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INSTITUTE OF FOOD TECHNOLOGISTS 1AUGUST 2004

This IFT Scientific Status Summary discusses Salmonella, Shigella,Campylobacter, Listeria, and Vibrio species, and Yersinia enterocolitica,Staphylococcus aureus, Clostridium perfringens, Clostridium botulinum,Bacillus cereus, and Enterobacter sakazakii.

Bacteria Associatedwith FoodborneDiseases

A

Scientific Status Summary

n estimated 76 million cases of foodborne illness occur each

year in the United States, costing between $6.5 and $34.9

billion in medical care and lost productivity (Buzby and

Roberts, 1997; Mead et al., 1999). In the United States, incidence of

foodborne illness is documented through FoodNet, a reporting system

used by public health agencies that captures foodborne illness in over

13% of the population. Of the 10 pathogens tracked by FoodNet,

Salmonella, Campylobacter, and Shigella are responsible for most cases

of foodborne illness. When both the estimated number of cases and

mortality rate are considered for bacterial, viral, and parasitic cases of

foodborne illness, Salmonella causes 31% of food related deaths,

followed by Listeria (28%), Campylobacter (5%), and Escherichia coli

O157:H7 (3%) (Mead et al., 1999).Data are only available for confirmed cases, and it is generally accepted

in the scientific community that the true incidence of foodborne disease isunderreported. Of the estimated 13.8 million cases of foodborne illnessdue to known agents, roughly 30% are due to bacteria. The remaining cas-es of known etiology are due to parasites in 3% of the cases and viruses in67% of the cases (Mead et al., 1999). Because parasites (Orlandi et al.,2002), E. coli O157:H7 (Buchanan and Doyle, 1997), and viruses (Cliver,1997) have been addressed by previously published Scientific Status Sum-maries, these pathogens are not discussed here. Enterobacter sakazakii, anemerging pathogen of concern in infant formula, is a new inclusion in thisrevised Scientific Status Summary.

This Scientific Status Summary is an updatedreview of the bacteria of primary significancein foodborne disease, their significance aspathogens, their association with foods, andrelated control measures. The original publica-tion appeared in the April 1988 edition of FoodTechnology.

2 INSTITUTE OF FOOD TECHNOLOGISTS AUGUST 2004

Table 1— Foodborne Disease in the United States, Including Estimated Annual Prevalence. Adapted from IFT (2000).

Bacteria Principal symptomsa Potential food contaminationa No. of illnessesb No. of deathsb

Bacillus cereus Diarrheal—watery diarrhea, abdominal Meats, milk, vegetables, fish 27,360 0cramps and painEmetic—nausea and vomiting Rice products, starchy foods n/a n/a

Campylobacter spp. Watery diarrhea, fever, abdominal Raw chicken, beef, pork, shellfish, raw milk 1,963,141 99pain, nausea, headache, muscle pain

Clostridium botulinum Weariness, weakness, vertigo, double Improperly canned or fermented goods 58 4vision, difficulty swallowing and speaking

Clostridium perfringens Intense abdominal cramps, diarrhea Meat, meat products, gravies 248,520 7

Enterobacter sakazakii c Meningitis, enteritis Powdered infant formula 1:100,000 infantsc 20-50%c

Listeria monocytogenes Nausea, vomiting, diarrhea; influenza-like Raw milk, cheeses (particularly soft ripened 2,493 499symptoms, meningitis, encephalitis; varieties), raw vegetables, raw meats, rawsepticemia in pregnant women, their and smoked fish, fermented sausagesfetuses, or newborns; intrauterine or cervicalinfection that may result in spontaneousabortion or stillbirth

Salmonella Typhi and Typhoid-like fever, malaise, headache, Raw meats, poultry, eggs, milk and 659 3 Salmonella Paratyphi abdominal pain, body aches, diarrhea, dairy products, fish, shrimp, yeast, coconut,

or constipation sauces, salad dressings (i.e., homemade itemscontaining unpasteurized eggs and no orinsufficient acidification for destroying pathogens)

Other Salmonella spp. Nausea, vomiting, abdominal cramps, fever, Raw meats, poultry, eggs, milk and 1,341,873 553headache; chronic symptoms (e.g., arthritis) dairy products, fish, shrimp, yeast, coconut,

sauces, salad dressings (i.e., homemade itemscontaining unpasteurized eggs and no orinsufficient acidification for destroying pathogens)

Shigella spp. Abdominal pain and cramps; diarrhea; Salads (potato, tuna, chicken, macaroni) 89,648 14fever; vomiting; blood, pus, or mucus raw vegetables, bakery products (e.g.,in stools, tenesmus cream-filled pastries), sandwich fillings,

milk and dairy products, poultry

Staphylococcus aureus Nausea, vomiting, retching, abdominal Meat and meat products, poultry, egg 185,060 2cramps, prostration products, salads (chicken, potato, macaroni),

cream-filled bakery products, milk anddairy products

Vibrio cholera Serogroup 01 Mild watery diarrhea, acute diarrhea, Raw or recontaminated oysters, clams, crabs 49 0rice-water stools

V. cholera Serogroup non-01 Diarrhea, abdominal cramps, fever, vomiting, Raw or recontaminated oysters, clams, crabs 49 0nausea; blood or mucus-containing stools

Vibrio vulnificus Fever, chills, nausea, septicemia in individuals Raw or recontaminated oysters, clams crabs 47 18with some underlying diseases or takingimmunosuppressive drugs or steroids

Other Vibrio spp. Diarrhea, abdominal cramps, nausea, Raw, improperly cooked or recontaminated n/a n/avomiting, headache, fever, chills shellfish or fish

Yersinia enterocolitica Fever, abdominal pain, diarrhea and/or vomiting Meats, oysters, fish, raw milk 86,731 2

aBased on FDA/CFSAN (2003b).bBased on Mead et al. (1999). n/a indicates that no estimate was provided for this pathogen.c From WHO (2004).


Bacteria are the causative agents offoodborne illness in 60% of cases re-quiring hospitalization (Mead et al.,1999). The international impact offoodborne illness is difficult to estimate.However, about 2.1 million children indeveloping countries die due to diarrhe-al-related illnesses annually. It is suspect-ed that food or water is the vehicle formany of these illnesses (WHO, 2002).

Because food is biological in natureand is capable of supplying consumerswith nutrients, it is equally capable ofsupporting the growth of contaminating

microorganisms. Three types of bacteri-al foodborne diseases are recognized:intoxications, infections, and toxicoin-fections. Foodborne bacterial intoxica-tion is caused by the ingestion of foodcontaining preformed bacterial toxin,such as the toxins produced by Staphy-lococcus aureus and Clostridium botuli-num, resulting from bacterial growth inthe food. Foodborne infection, on theother hand, is caused by ingestion offood containing viable bacteria such asSalmonella or Listeria which then growand establish themselves in the host, re-

sulting in illness. Foodborne toxicoin-fections result when bacteria present infood, such as Clostridium perfringens,are ingested and subsequently produce atoxin in the host.

Some pathogens reside in the intesti-nal tracts of normal, healthy animalsand, in some instances, humans. Certainmicroorganisms are ubiquitous in na-ture, occurring on soil and vegetation,in animal wastes, and on animal carcass-es. Human skin surfaces and nasal pas-sages harbor staphylococci. Water sup-plies may contain pathogens when con-


practices probably have the greatestchances for success in controlling food-borne illness. The need for continualeducation of consumers and all foodhandlers concerning the significant haz-ards associated with foodborne patho-genic microorganisms and proper foodhandling procedures is evident. Regula-tory control over food handling in thehome is not possible, but increased con-sumer education could have a beneficialeffect.

SalmonellaRussell S. Flowers

Salmonella is a generic name appliedto a group of nearly 2,000 biochemicallyrelated serotypes responsible for food-borne illness.

Significance as a PathogenThe total number of cases of human

salmonellosis have remained fairly con-stant between 1996 and 2002 (Figure 1),although the serotypes causing illnesshave changed (CDC, 2003a). Roughlytwo to four million cases of foodbornesalmonellosis occur annually in theUnited States, and the estimated 1.3million cases that occurred in 2000 cost$2.4 billion in medical costs and lostproductivity (USDA/ERS, 2003). Be-tween 1988 and 1995 there were be-tween 40,000 and 50,000 reported, con-firmed cases of salmonellosis annually;since 1997, that number has been below35,000 (CDC, 2004; FDA/CFSAN,2003b).

The disease is grossly underreportedbecause it is generally a self-limitinggastroenteritis which may be misdiag-nosed as intestinal influenza by the pa-tient or the physician. As a consequence,estimates of the true incidence of dis-ease are based on assumptions derivedfrom epidemiological evidence. Clearly,salmonellosis continues to be an impor-tant cause of foodborne disease world-wide.

Two clinical manifestations causedby Salmonella are recognized: enteric fe-ver (a severe, life-threatening illness)and the more common foodborne ill-ness syndrome. In both cases, the re-sponsible microorganisms enter thebody via the oral route.

Enteric fever, commonly referred toas typhoid fever, is primarily caused byone species, Salmonella Typhi, but othersalmonellae such as SalmonellaParatyphi are potentially capable of pro-

taminated with fecal matter. Coastal wa-ters may also naturally harbor thepathogen Vibrio vulnificus. It is thus ob-vious how difficult it is to prevent oneor more pathogens from contaminatingfoods.

The presence of potentially life-threatening pathogens in our environ-ment, the ability of some of them tosurvive and/or proliferate under refrig-eration and in reduced oxygen atmo-spheres, and, for some pathogens, thelow number necessary for causing dis-ease indicate the seriousness of the po-tential hazards with which we are faced.The food industry implements a varietyof effective control measures to limitpotential hazards. This generally beginson the farm with the implementation ofgood agricultural practices. Some rawproducts, such as poultry, are subject toperformance standards in spite of thefact that the consumer will presumablycook the product before consumption.While all food manufacturers utilizecontrol measures to ensure food safety,some food manufacturers are requiredto create and follow a Hazard Analysisand Critical Control Point (HACCP)plan. Critical control points identifiedthrough HACCP may include destruc-tion or inactivation of the relevant bac-teria or their spores through the use ofheat treatments (e.g., pasteurization,canning), dehydration, freezing, refrig-eration, specialized packaging, and/orapproved antimicrobial preservatives.Additionally, extensive quality controlprocedures are maintained to ensurethat these processes are effective.

It is impossible, however, to create arisk-free food supply. While food manu-facturers and distributors employ neces-sary control measures to ensure thesafety of food until it reaches the con-sumer, all food handlers and consumershave the responsibility upon purchaseof the food to maintain these controlmeasures until consumption. Whileoutbreaks associated with a particularcommercially processed food receivewidespread public attention, a muchgreater number of individual cases offoodborne illness occurring in restau-rants and in the home are not reported.The Centers for Disease Control andPrevention (CDC) reported that be-tween 1993 and 1997, approximately19% of foodborne illness outbreaks oc-curred in private residences. Numeroussurveys have highlighted inadequatehome food storage and handling prac-

tices as major contributors to foodborneillness (Altekruse et al., 1996; Meer andMisner, 2000). For most of the patho-gens discussed in this Scientific StatusSummary (Table 1), illness can be avoid-ed by heating and cooling foods to theappropriate temperatures, storing foodsat the appropriate conditions for therecommended period of time, hand-washing, and avoiding cross-contamina-tion. Delicatessens, cafeterias, and res-taurants were responsible for 33% ofoutbreaks (CDC, 2000b). Only one ofmore than 1,300 outbreaks, includingbacterial and viral, with known or un-known etiology, was attributed to a“commercial product” in 2000 (CDC,2000d). Proper handling, cooking, andstorage practices in foodservice opera-tions and in the home can prevent themajority of foodborne illnesses.

In the foodservice sector, educationof the food preparer and server, withemphasis on good personal hygiene, isthe best preventive measure. Unfortu-nately, the majority of foodserviceworkers are young (under 30 years ofage), inexperienced, and stay on the jobless than a year; thus, finding and edu-cating these food handlers while theyare actively working is difficult (Marth,1985; National Restaurant Association,2004).

However, training programs such asServsafe®, conducted through the Na-tional Restaurant Association EducationFoundation, and SuperSafeMark®, of-fered through the Food Marketing Insti-tute, have a positive impact on retailfood safety practices (Cotterchio et al.,1998; Lynch et al., 2003; Mathias et al.,1995).

Several studies and reviews havehighlighted the contribution of chang-ing demographics in the United Stateswith the increased risk of foodborne ill-ness (Knabel, 1995; Zink, 1997). The in-crease in the elderly population and in-dividuals with weakened immune sys-tems underscores the need for rigorousfood safety efforts, both on the part ofthe food manufacturer and the consum-er. Foodborne illnesses are more likelyto be life-threatening for the immunecompromised, the elderly, and individu-als debilitated by underlying healthproblems such as cirrhosis, hepatitis,hemachromatosis, etc.

Consumer education and increasedregulatory control of foodservice estab-lishments through inspection and strictenforcement of proper food handling


Figure 1. Most Common Salmonella Isolates from Human Sources (data from CDC PHLIS Surveillance Data:Salmonella Annual Summaries); left axis: (g) Heidelberg, (#) Enteritidis, (•) Javiana, (!) Newport, (F)Typhimurium; right axis: (-t—) total number of all Salmonella isolates.


ducing this syndrome. The illness iscommonly associated with foreign traveland affects an estimated 800 people an-nually (Mead et al., 1999). Although theroute of entry of the pathogen into thebody is primarily oral, the symptoms ofenteric fever are generally not elicitedthrough the intestinal tract. However, ashort episode of vomiting and diarrheasometimes occurs in the first day or twoin typhoid fever. The onset times varyconsiderably between typhoid andparatyphoid enteric fevers. Onset timefor typhoid is usually 8–15 days, seldomas short as five days but sometimes aslong as 30–35 days; while onset time forparatyphoid fever tends to be shorter,and may be so short as to suggest typicalfood poisoning (Parker, 1984). The usu-al symptoms of both are headache, mal-aise, anorexia, and congestion of themucous membranes, especially of theupper respiratory tract. Bacteremia gen-erally occurs in the first week of illness.In a study of 1,138 cases, Coleman andBuxton (1907) reported positive bloodcultures in 89% of patients during thefirst week, 73% in the second week, 60%in the third, 38% in the fourth, and 26%after the fourth week. Arthritic symp-toms may emerge three to four weeks af-ter infection in roughly 2% of the cases.The 10% mortality rate of SalmonellaTyphi and Salmonella Paratyphi is highcompared to other Salmonella species(FDA/CFSAN, 2003b).

Typically, common foodborne illness

resulting from Salmonella infection ischaracterized by a self-limiting acutegastroenteritis. Contaminated food orwater is the usual, but not the only, vehi-cle. The incubation period varies fromsix to 48 hr and generally falls within arange of 12–36 hr. Variation in the incu-bation time may be attributed to the sizeof the infecting dose, the virulence (de-gree of pathogenicity) of the microor-ganisms, the susceptibility of the host,and the physicochemical composition ofthe transmitting food. As few as 15 cellscan cause illness (FDA/CFSAN, 2003b).Symptoms include diarrhea, abdominalcramps, vomiting, and fever, which gen-erally last from one to seven days. How-ever, the microorganisms may be excret-ed in the feces for many weeks aftersymptoms subside. Although the illnessis usually self limiting, there is a 15%mortality rate in elderly who have devel-oped septicemia due to Salmonella dub-lin, and a 3.6% mortality rate in nursinghome cases of Salmonella Enteritidis(FDA/CFSAN, 2003b). Salmonellosismay be confused clinically with staphy-lococcal intoxication, but there are im-portant distinctions. Salmonella has alonger incubation period than staphylo-cocci (usually 12–36 hr vs. 2–4 hr) andis usually accompanied by fever, which isabsent in staphylococcal intoxication.And unlike Salmonella food poisoning,the acute symptoms of staphylococcalfood poisoning normally disappearwithin 24 hours.

In some cases, gastroenteritis may befollowed by extraintestinal invasion re-sulting in enteric fever, which is morelikely to occur in the very young, theaged, and debilitated patients.

Association with FoodsThere are three main ways Salmonel-

la can enter the food supply to cause ill-ness. Animals harbor Salmonella, mak-ing meats, poultry, eggs, and milk oftenimplicated vehicles. Salmonella, whichare introduced into the environment,possibly through manure and litter, maypersist and contaminate fruits and vege-tables on the farm. Cross-contaminationin the food service environment or thehome, often between raw poultry andready-to-eat (RTE) products, such asraw vegetables, can also cause salmonel-losis.

Some Salmonella are host-adapted(e.g., Salmonella pullorum in poultry,Salmonella dublin in cattle); however,most Salmonella are not. Although anySalmonella is a potential pathogen forhumans, most foodborne salmonellosisis caused by non-host-adapted sero-types.

Much human salmonellosis is direct-ly related to human association with an-imals, both wild and domestic. Foods ofanimal origin are vehicles for salmonel-losis. Salmonella was isolated in 19–54%of cattle carcasses, 1.9% of beef samplesat retail and 4.2% of retail chicken sam-ples (Beach et al., 2002; Zhao et al.,2001). In a review of Salmonella inmeats and poultry, Silliker and Gabis(1986) introduced the problem as fol-lows:

“The animal-to-man link is only onefactor in the epidemiology of humansalmonellosis. Contaminated red meatand poultry provide a nidus (source ofinfection) for Salmonella, which mannurtures through mishandling. Further-more, inedible parts of the animal areprocessed to yield important compo-nents of livestock feeds. As a result ofpoor manufacturing practices (post-processing contamination), these ren-dered animal by-products become re-contaminated with Salmonella, which,in turn, are carried into the feeds. Theconsumption of these feeds by livestock,followed by animal-to-animal transmis-sion, completes the Salmonella cycle.This is not to suggest, however, that con-taminated feeds are the only source ofSalmonella in livestock. Epidemiologicalevidence indicates that there is a direct


link between the presence of Salmonellain meat and poultry and human salmo-nellosis. Man induces salmonellosisthrough improper food handling prac-tices and perpetuates salmonellosisthrough recontamination of renderedanimal by-products, which are incorpo-rated into livestock feeds.”

Historically, egg products (dried, fro-zen or liquid eggs, with or without add-ed ingredients, as defined in the EggProducts Inspection Act; U.S. Congress,1970) were a significant source of hu-man salmonellosis in the United States.In 1990, the U.S. Dept. of Agriculture(USDA) required that eggs produced byflocks previously implicated in humandisease be tested for Salmonella (FDA/CFSAN, 2003b). Mandatory pasteuriza-tion of egg products was responsible forgreatly reducing eggs as a major cause ofsalmonellosis, and consumer awarenessof the need to refrigerate eggs also con-tributed in the decreased incidence ofdisease. However, since 1999, the inci-dence of Salmonella Enteritidis has re-mained stable. The microorganism is lo-calized inside eggs, making thoroughcooking imperative. The CDC estimatesthat 75% of Salmonella Enteritidis casesresult from the consumption of raw orundercooked grade A whole-shell eggs(FDA/CFSAN, 2003b). This serotypewas the second most commonly report-ed human serotype to the CDC in 2001.A portion of these illnesses occurred asa result of pooling raw shell eggs in in-stitutional or foodservice settings. Thepotential for temperature abuse or longholding times, combined with the possi-bility of inadequate cooking, resulted inthe recommendation that institutionsuse pasteurized egg products or pasteur-ized in-shell eggs instead of raw-shelleggs (CDC, 2003c).

Consumption of raw milk may alsocause human salmonellosis. In onestudy, Salmonella was isolated in 6.1%of bulk raw milk samples (Jayarao andHenning, 2001). Between 1972–2000, 16outbreaks of salmonellosis resultedfrom raw milk consumption (CDC,2003a). Milk-borne salmonellosis wasparticularly prevalent in Scotland,where, prior to 1983, the sale and con-sumption of raw milk was common(Sharp, 1986). Milk outbreaks in En-gland and Wales were primarily associ-ated with raw milk. The sale of raw milkis legal in 27 states, although many haverestrictions (CDC, 2003a; The WestonA. Price Foundation, 2003). Pasteuriza-

tion of milk destroys Salmonella andcurrently is the only effective means ofcontrol for milk. However, inadequatepasteurization or contact with raw milkafter pasteurization can result in con-taminated milk. In one pasteurized milkoutbreak in 1985, there were more than16,000 laboratory-confirmed cases ofsalmonellosis and several deaths (Anon-ymous, 1986).

Animal products are not the onlysources of human salmonellosis. Pro-duce can serve as a vehicle of Salmonellaas well, becoming contaminated eitheron the farm or through cross-contami-nation with contaminated products.Consumption of produce contaminatedwith Salmonella has recently caused sev-eral outbreaks. In 2000, 361 individualsin England and Wales acquired salmo-nellosis after eating lettuce, and al-though the lettuce appeared to comefrom one of three farms, investigatorscould not conclusively find the sourceof contamination (Horby et al., 2003).The strain, S. enterica subsp. enterica se-rotype Typhimurium DT 104, is resis-tant to several antibiotics, and the in-crease in its prevalence poses challengesin treatment of the infection. Rawsprouts, particularly alfalfa and mungbean, have been involved in 15 salmo-nellosis outbreaks since 1995 (Thomaset al., 2003). Seeds may be contaminat-ed, and the warm, moist conditions forsprouting allow growth of the pathogen.On-farm contamination of cantaloupesresulted in 24 cases of salmonellosis in1997 (Mohle-Boetani et al., 1999). Con-tamination of cantaloupes on one farmcaused mulitstate outbreaks of salmo-nellosis each spring between 2000 and2002 (CDC, 2002b). Since produce maybe eaten raw, different control measuresare necessary to prevent illness when thepathogen is introduced on the farm.

While Salmonella may survive incontaminated foods as a result of im-proper cooking, it is more common thatcross-contamination of foods aftercooking is the source of Salmonella.Foodservice workers or in-home foodpreparers may transfer salmonellae fromraw products to cooked or other uncon-taminated foods as a result of unsani-tary preparation practices (e.g., failureto wash hands) between handling ofthese foods. Salmonella can also betransferred from contaminated rawfoods to equipment surfaces, such asknives, cutting boards, and counter tops,and then from equipment to previously

uncontaminated foods. Once contami-nation occurs, the situation may be fur-ther complicated by improper storage ofthe product before serving (e.g., kept atroom temperature, improperly refriger-ated, or held in warmers within thegrowth range for Salmonella).

Although responsible for fewer out-breaks, contamination of foods by in-fected workers cannot be ignored as acause of foodborne salmonellosis. Someinfected individuals may excrete Salmo-nella for weeks, months, and, occasion-ally, years with little or no evidence ofdisease. Improper hygiene practices bythese individuals may lead to either con-tamination of foods or direct person-to-person contamination.

Control MeasuresDifferent control measures exist de-

pending on the mode of contaminationof the food. Reduction of the incidenceof Salmonella contamination of foodsrequires a number of approaches to theproblem, beginning at the farm and go-ing right through to the kitchen.

Several approaches have been takento reduce the carriage of Salmonella byanimals. Vaccinination of laying chick-ens significantly reduced the percent ofeggs positive for Salmonella Enteritidis(Woodward et al., 2002). Other researchhas examined the effect of probiotics onthe intestinal microflora of chickens. Inthis approach, the goal is to colonize thechicken with known microorganisms to“competitively exclude” Salmonella. Suc-cessful reduction in the percent of theflock positive for Salmonella wasachieved when the feed was supple-mented with yeast (Line et al., 1998) orwhen chickens were both sprayed with apatented mucosal starter culture and fedthe culture through water (Bailey et al.,2000).

In July 1996, USDA’s Food Safety andInspection Service (FSIS) publishedpathogen reduction performance stan-dards for Salmonella. The performancestandard (percent positive) was estab-lished by collecting the average baselinein the industry. Companies must useHACCP and other measures to ensurethat the percent of their product that ispositive for Salmonella falls below thisbaseline. The performance standard hasbeen expanded to include steers, cows,broilers, hogs, and several related prod-ucts (FSIS, 1998). Between 1997 and2003, FSIS reported a 66% decrease inthe presence of Salmonella in raw meat


and storing foods at the right tempera-tures.

ShigellaRussell S. Flowers

Shigellosis, or bacillary dysentery, asit is commonly known, is caused by bac-teria of the genus Shigella, which includeS. dysenteriae, S. flexneri, S. boydii, andS. sonnei (Figure 2) (Bryan, 1979). Thenormal habitat for Shigella is the intesti-nal tract of humans and other primates.

Significance as a PathogenData suggest that Shigella is responsi-

ble for less than 10% of reported food-borne illnesses per year, infecting ap-proximately 300,000–450,000 peopleannually (FDA/CFSAN 2003b; Mead etal., 1999). Although the primary modeof transmission appears to be person-to-person by the fecal-oral route (Feld-man and Riley, 1985), there have beensome documented cases of foodborneshigellosis. The main source of Shigellainvolved in outbreaks is asymptomaticcarriers or persons recovering from dis-ease. Shigella may persist in the intesti-nal tract for months (Bryan, 1978).

Symptoms of shigellosis include diar-rhea, abdominal pain, fever, and vomit-ing. The severity of the disease may varyfrom very mild to severe diarrhea withbloody stools, mucus secretia, and dehy-dration. Reactive arthritis, hemolyticuremic syndrome and Reiters disease aresequelae associated with shigellosis(FDA/CFSAN, 2003b). Generally, food-borne shigellosis is characterized by ahigh attack rate, common-source epide-miology, and short incubation periodsof 12–50 hr (FDA/CFSAN, 2003b).Symptoms usually persist 3–14 days. In-fection by some strains results in a 10–15% mortality rate. Frequently, an as-ymptomatic carrier state may developduring convalescence, lasting from a fewdays to several months. Human-volun-teer studies indicate that ingestion of asfew as 10–100 microorganisms can in-duce illness (Morris, 1986).

The disease is caused by invasion ofthe intestinal mucosa. Enterotoxins,classically referred to as Shiga toxins,may be produced by S. dysenteriae andpossibly by S. flexneri type 2A (Keuschet al., 1985). Shiga toxins block mRNAtranscription and induce apoptosis inendothelial cells (Erwert et al., 2003;Thorpe et al., 1999).

Secondary infections with Shigella


and poultry (USDA, 2003). Due to thedifficulty in completely eradicating Sal-monella from poultry, irradiation wasapproved in 1999 as a means to controlthe microorganism as part of a HACCPsystem (USDA, 1999).

The most common method of elimi-nating Salmonella from food products isheat processing. Salmonella is heat sensi-tive, and ordinary pasteurization orcooking conditions are generally suffi-cient to kill it in high-moisture foods. Aswith other microorganisms, the heat re-sistance of Salmonella is markedly in-creased as water activity (a

w) decreases.

Current U.S. standards for milk pasteur-ization result in a more than 1012-foldreduction of viable Salmonella found inmilk (Read et al., 1968). Minimum heatprocesses for pasteurization or cookingof most high-moisture products willeliminate Salmonella from these prod-ucts. Occurrence of Salmonella in suchprocessed products generally resultsfrom post-processing contamination.

In some cases, dry products may beheat processed for the purpose of elimi-nation of Salmonella. Ayres and Slos-berg (1949) and Banwart and Ayres(1956) demonstrated the effectivenessof dry-heat treatment on reducing Sal-monella in pan-dried egg white. Thiswork led to the eventual requirementfor dry-heat pasteurization of this prod-uct (U.S. Congress, 1970). Similar treat-ments have been applied to gelatin, ren-dered animal by-products, nonfat drymilk, and chocolate.

Other than heat, the major manufac-turing factors responsible for elimina-tion of Salmonella from food productsare acidification and reduction of a

w.

Goepfert and Chung (1970) investigatedthe fate of Salmonella during sausagefermentation and concluded that thecombination of acidity and sodiumchloride was the principal reason for thedemise of Salmonella during fermenta-tion. Similarly, Smittle (1977) reviewedthe literature on survival of Salmonellain mayonnaise and salad dressing andconcluded that the principal factor re-sponsible for the death of salmonellae inthese products is acidity, but that a sec-ond important factor is reduction of a

w

as affected by moisture, NaCl, and sugarconcentration. These same factors areresponsible for control of Salmonella ina variety of fermented dairy, meat, andvegetable products.

Salmonella may survive for extendedperiods in dehydrated foods. However,

some death occurs during dehydratedstorage, depending on the relative hu-midity (or a

w) and storage atmosphere.

The rate of death of Salmonella in foodpreserved by reduced a

w is increased at

higher aw levels, temperatures, and oxy-

gen levels (Genigeorgis and Riemann,1979). In low-moisture products such aspeanut butter and chocolate, Salmonellamay remain for years, with little loss ofviability.

Moist, perishable products are gener-ally distributed under refrigerated orfrozen conditions. Although freezingand frozen storage can have some lethaleffects on Salmonella, it is well recog-nized that Salmonella remain viable forlong periods of time in frozen foods andthat survival is enhanced as the storagetemperature decreases (Georgala andHurst, 1963).

The presence of Salmonella in certaintypes of produce seems to result fromintroduction of the pathogen on thefarm. Contamination in these products,which are typically not heated beforeconsumption, presents challenges to thefood industry (CDC, 2002b; Thomas etal., 2003). For sprouts, chemical disin-fection of the seed coat is not mandato-ry, but is recommended by the Food andDrug Administration (FDA). A surveyof sprout producers in California foundthat most alfalfa sprout growers fol-lowed FDA recommendations. Mungbean producers were less apt to followthe guidance (Thomas et al., 2003). Anoutbreak involving 87 cases of Salmo-nella mbandaka was linked back togrowers who did not disinfect seeds(Gill et al., 2003). The recommendedlevel of chlorine treatment, 20,000 ppm,still may not be sufficient to eliminateSalmonella, especially if they are local-ized inside the sprout tissue, as Gandhiet al. (2001) observed. Therefore, com-pliance on the part of the grower, whileworthwhile, does not guarantee a Sal-monella-free product since chemicalsanitizers may not be wholly sufficientto eliminate the presence of Salmonellain fruits and vegetables with naturalopenings and crevices (Beuchat andRyu, 1997).

When purchasing items likely to becontaminated with Salmonella, such aschicken, there are steps the food handlercan take to prevent salmonellosis fromoccurring in the home or food serviceenvironments. These general food safetypractices include avoiding cross-con-tamination, thoroughly cooking foods,


occur frequently. Shigellosis can be amajor problem in daycare centers, andcontrol is difficult due to the low infec-tious dose, unpredictable acquisition ofantimicrobial resistance, frequency ofmild infections and carrier status, andthe frequency with which young chil-dren are transferred from one daycarecenter to another (CDC, 1984). The dis-ease is disproportionately commonamong the urban poor, immigrants, Na-tive Americans, daycare centers, and in-stitutions (Bryan, 1978; Morris, 1986).FoodNet data from 2003 captured 3,875confirmed cases of shigellosis within the13% of the population monitored. Ex-trapolated, this suggests almost 30,000cases would be confirmed in the U.S.population (CDC, 2003b). In the early1900s, S. dysenteriae and S. flexneri pre-dominated (Bryan, 1978). The patternof isolations suggests that prior to 1945,direct fecal contamination of food andwater supplies may have been the pri-mary vector of disease. As socioeconom-ic and environmental sanitation (mu-nicipal sewage disposal, safe water sup-plies, and food hygiene) have improved,the relative prevalence of S. sonnei hasincreased. The explanation for this phe-nomenon is not clear, but it may resultfrom differences among the shigella intheir ability to survive and grow outsidetheir human and animal hosts.

Association with FoodsAn estimated 20% of the total num-

ber of cases of shigellosis involve food asthe vehicle of transmission (Mead et al.,1999). The principal foods involved inoutbreaks include a variety of saladsand seafoods (Morris, 1986) contami-nated during handling by infectedworkers (Bryan, 1978). Six separately re-ported outbreaks in the United Statesand Canada in 1998 involving S. sonneiwere traced back to parsley grown inMexico (Naimi et al., 2003). The pres-ence of S. sonnei in commercially pro-duced dip caused an outbreak on thePacific Coast in 2000. The product, mar-keted under several brand names, wasultimately recalled (CDC, 2000c). Im-proper refrigeration of the contaminat-ed product often contributes to illness.After the initial infection from a con-taminated food, the disease readilyspreads from person to person by the fe-cal-oral route of transmission.

Shigella usually are considered to berelatively fragile; i.e., they do not survivewell outside the host (Bryan, 1978). Like

most other members of the family En-terobacteriaceae, they are readily killedby most heat treatments employed inthe processing and preparation of foods,and do not survive well at pH below 4.5.However, studies on the survival of shi-gella suggest that under certain selectedconditions, they can survive for extend-ed periods in foods (Bryan, 1978). Forexample, S. sonnei and S. flexneri havebeen reported to survive at 25°C (77°F)in flour and in pasteurized whole milkfor more than 170 days; in eggs, clams,and shrimp for more than 50 days; inoysters for more than 30 days; and inegg whites for more than 20 days. Atlower temperatures, such as 20°C (-4°F)and 0.5°C (32.9°F), survival for longerperiods has been reported (Taylor andNakamura, 1964).

These laboratory data suggest thatshigella may survive for extended peri-ods in foods and may grow under select-ed conditions. However, in practice, Shi-gella is rarely isolated from processedproducts. Manufacturers do not rou-tinely test their products, raw materials,or processing environments for Shigella,and there is no evidence to suggest thatroutine testing is warranted. Most out-breaks result from contamination of rawor previously cooked foods during prep-aration in the home or in foodserviceestablishments. Generally, the source ofcontamination can be traced to a carrierwhose personal hygiene is poor (Bryan,1979). In underdeveloped countries,harvesting of seafood from fecally con-taminated waters and use of unsanitarywater in food preparation may besources of disease.

Control MeasuresShigellosis is a significant cause of

foodborne illness in both developed anddeveloping countries. Food microbiolo-gists should be aware that Shigella spe-cies can cause a rather severe form offoodborne illness (Smith, 1987) andthat relatively low numbers of the mi-croorganisms can cause disease. Prod-ucts suspected of foodborne illnessshould be analyzed for Shigella.

Further studies are needed to deter-mine the roles of the various toxins andvirulence factors in the disease process.The epidemiology of shigellosis shouldbe reevaluated, with particular emphasison the environmental factors associatedwith the increasing incidence of S. son-nei and S. flexneri. The waning incidenceof S. dysenteriae in U.S. epidemiologicaldata clearly indicate that the majorcause of shigellosis in developed coun-tries is infected food handlers. Thus,prevention and control of shigellosis re-quires either that infected humans notbe permitted to handle foods or thatthey practice good personal hygienesuch that, even if infected, the food doesnot become contaminated. Certainly,exclusion of infected individuals fromfood handling is desirable. However,routine testing of food workers for Shi-gella is neither practical nor necessary.

The principles necessary for controlof Shigella at the preparation and servicelevel are the same as those necessary forcontrol of other more widely recognizedpathogens such as Salmonella andCampylobacter. The increasing aware-ness of consumers and employees offoodservice establishments of the po-

Figure 2. Relative contribution of species of Shigella causing foodborne illness; (g) S. boydii,(#) S. dysenteriae, (•) S. flexneri, (!) S. sonnei.


tential for foodborne diseases should domuch to reduce the incidence ofshigellosis in the developed countries. Inthe developing countries, control ofshigellosis, as always, will depend onimproved hygiene and waste-handlingpractices.

Campylobacter jejuniand other speciesMichael P. Doyle

Originally considered a pathogenprincipally of veterinary significance,Campylobacter jejuni (formerly knownas Vibrio fetus) was known to causeabortion in sheep. Other enteric campy-lobacters that may be transmittedthrough food include C. coli, C. lari, andC. upsaliensis, although many other spe-cies have been identified.

Significance as a PathogenFollowing the development of proce-

dures for detecting campylobacters instool specimens, C. jejuni became recog-nized as a leading cause of acute bacteri-al gastroenteritis in humans. Recent evi-dence suggests that Campylobacter caus-es 2–4 million cases per year in theUnited States (FDA/CFSAN, 2003b;Mead et al., 1999). Common symptomsof Campylobacter enteritis include pro-fuse watery or sticky diarrhea (some-times containing blood), abdominalcramps, headaches, muscle pain, andnausea. The mortality rate is low (0.1%)and often occurs in susceptible popula-tions such as the elderly, young, or im-munocompromised (FDA/CFSAN,2003b). Roughly 25% of cases experi-ence relapses. Although sequelae such asGuillain-Barré syndrome, meningitis,recurrent colitis and acute cholecystitisare uncommon, campylobacter is asso-ciated with more than 30% of cases ofGuillain-Barré (FDA/CFSAN, 2003b;Mead et al., 1999). Human volunteerand retrospective studies of food-associ-ated outbreaks of Campylobacter enteri-tis revealed that ingesting relativelysmall numbers (only a few hundredcells) of C. jejuni can produce illness.Symptoms manifest after an incubationperiod of two to five days, and generallylast 7–10 days. Treatment with erythro-mycin can decrease an individual’s du-ration of shedding the microorganism(FDA/CFSAN, 2003b).


Association with FoodsCampylobacters are harmless inhab-

itants of the gastrointestinal tract of avariety of wild and domestic animals.Studies have revealed that as many as30–100% of poultry, 40–68% of cattle,and up to 76% of swine carry C. jejunior C. coli in their intestinal tracts (Beachet al., 2002; Harvey et al., 1999). For thisreason, the microorganism is often asso-ciated with unprocessed foods of animalorigin. Surveys of U.S. retail fresh redmeat and poultry show that 12–35% ofturkey, 64% of chicken, 2–5% of pork,0–5% of beef, 8% of lamb and 9% bulktank raw milk contained C. jejuni and/or C. coli (Jayarao and Henning, 2001;Logue et al., 2003; Stern et al., 1985;Zhao et al., 2001). It is estimated thatroughly half of all cases of Campylo-bacter entertitis are associated with un-dercooked chicken or cross-contamina-tion with raw chicken (FDA/CFSAN,2003b). In a 1996 outbreak, raw lettuceserved as the vehicle for Campylobacterinfections, presumably after contactwith raw chicken (CDC, 1998). Otherfoods implicated as vehicles of out-breaks include raw milk, raw beef,clams, and cake (likely contaminated bya C. jejuni-infected food handler). Con-sumption of C. jejuni-contaminated rawmilk from an organic dairy farm made75 Wisconsin residents ill in 2001(CDC, 2002c). Incompletely pasteurizedmilk was responsible for an outbreak ofCamplyobacter enteritis affecting 110people in the United Kingdom (Fahey,1995). Although C. jejuni is responsiblefor more foodborne illness than C. coli,the significance of the latter should notbe overlooked. Estimates from studies inEngland and Wales revealed that ap-proximately 25,000 cases of foodborneillness were caused by C. coli in the year2000 (Tam et al., 2003). Outbreaks ofCamplylobacter enteritis are generallysmall; however, untreated surface waterused in municipal water supplies hasbeen responsible for large outbreaks in-volving thousands of individuals.

The principal route by which C. jeju-ni contaminates food is through fecalcontamination by Campylobacter-infect-ed carriers. Raw meats and poultry be-come contaminated during processingwhen intestinal contents contact meatsurfaces. Raw milk may be contaminat-ed by contact with bovine feces or pos-

sibly by a mastitic infection of the bo-vine udder. Fresh produce can be con-taminated by fecal-laden irrigation wa-ter, infected harvesters with poor hy-gienic practices, or tainted processingwater. Although eggs have rarely beenlinked to outbreaks of Campylobacterenteritis, C. jejuni has been isolatedfrom the surface of eggs laid by Campy-lobacter-infected hens (Doyle, 1984;Finch and Blake, 1985).

C. jejuni and other campylobactersare not likely to grow in foods that re-main edible because the organisms: (1)will not grow below 30°C (86°F); (2) re-quire microaerophilic (low concentra-tions of oxygen, ideally 5–10%) condi-tions for growth; (3) grow slowly evenunder optimal conditions; and (4) arenot good competitors with other micro-flora. Additionally, C. jejuni is a veryfragile microorganism, being sensitive todrying, normal atmospheric concentra-tions of oxygen, storage at room tem-perature, acidic conditions, disinfec-tants, and heat. Because the microor-ganism is quite sensitive to stressful en-vironmental conditions when outside ofthe host’s digestive tract, campylobactersare not likely to be a problem in pro-cessed foods that receive a pasteuriza-tion treatment or are dehydrated. How-ever, the microorganism is often associ-ated with and transmitted by raw, refrig-erated foods of animal origin.

Control MeasuresC. jejuni infections can be prevented

by properly pasteurizing or cookingfoods (especially foods of animal origin)and avoiding cross-contamination ofcooked or RTE foods by utensils, equip-ment, or cutting surfaces that are notproperly cleaned and disinfected aftercontact with fresh, uncooked meats andfoods likely to be contaminated with themicroorganism. Pasteurization willeliminate viable campylobacters in milk.Use of potable irrigation and processingwater and safe food handling practicesby harvesters are important to the safetyof produce. Although C. jejuni does notsurvive well in foods, refrigeration pro-longs survival. Freezing food will sub-stantially reduce the initial Campylo-bacter population; however, a smallnumber of survivors may remain viablefor many months (Moorhead andDykes, 2002). Testing foods for the pres-


ence of Campylobacter is difficult sinceenrichment with antibiotics is necessaryand cells must be grown in an environ-ment of 5% O

2 and 10% CO

2.

Yersinia enterocoliticaMichael P. Doyle

Yersinia enterocolitica is not a fre-quent cause of human infection in theUnited States; illness is more commonin Northern Europe, Scandinavia, andJapan.

Significance as a PathogenAlthough CDC estimates that there

are 17,000 cases of yersiniosis, the infec-tion caused by Y. enterocolitica, per year,less than 7,500 total cases were reportedbetween 1990 and 1999 (Ackers et al.,2000; FDA/CFSAN, 2003b). When themicroorganism is involved in illness,which typically manifests 24–48 hr afteringestion, the symptoms can be quite se-vere; but fatalities are rare. The infectivedose is unknown. Yersiniosis occursmost commonly in the form of gastro-enteritis. Children are most severely af-fected with symptoms of intense ab-dominal pain (often mimicking appen-dicitis), diarrhea, fever, and vomiting.Symptoms of pseudoappendicitis haveresulted in many unnecessary appendec-tomies in children. The illness has alsobeen misdiagnosed as Crohn’s Disease(FDA/CFSAN, 2003b). Fatality fromgastroenteritis is rare, and recovery isgenerally complete within one or twodays. Arthritis has been identified as aninfrequent but significant sequela in 2–3% of Y. enterocolitica infection cases.Other syndromes may include mesen-teric lymphadenitis, terminal ileitis,erythema nodosum, septicemia, andmeningitis.

Association with FoodsY. enterocolitica is principally a

zoonotic bacterium that has been isolat-ed from a variety of animals. However,most animal isolates, with the exceptionof swine isolates, are not human patho-gens. Swine is an important and theprincipal reservoir for virulent strains ofY. enterocolitica, with such isolates oftenrecovered from the tonsils and tonguesof apparently healthy animals (Doyle etal., 1981). A study in Japan revealed thatrats in slaughterhouse areas may also be

pathogenic strains in foods may explainwhy there are relatively few reportedfood-associated outbreaks of yersiniosis.The microorganism is one of a fewfoodborne pathogens that can grow atrefrigeration temperatures. Studies inraw pork revealed that within 10 days at7°C (44.5°F) the microorganism couldgrow from a few hundred cells to mil-lions/g (Hanna et al., 1977). Hence, re-frigeration, which traditionally has beenan important means of controlling thegrowth of foodborne pathogens, is notan effective method for controlling Yers-inia.

Control MeasuresY. enterocolitica is sensitive to heat

(e.g., pasteurization at 71.8°C for 18sec), sodium chloride (5%), and highacidity (pH 4.0), and will generally beinactivated by environmental conditionsthat will kill Salmonella (Robins-Browne, 1997). However, since Yersiniawill grow at refrigeration temperatures,cold storage can be a means of selective-ly promoting growth of the microorgan-ism in foods. Therefore, it is importantto eliminate the microorganism fromfoods (especially pork, milk, and foodsthat may have direct or indirect contactwith porcine waste) by pasteurization orcooking. Care should be taken to avoidcross-contamination of processed, RTEfoods with pork and porcine wastes, aswell as animal and human fecal contam-ination.

Listeria monocytogenesand other Listeria speciesOriginally authored by Joseph Lovettand Robert M. Twedt. Reviewed byStephanie Doores.

Prior to the 1980s, listeriosis, the dis-ease caused predominantly by Listeriamonocytogenes, was primarily of veteri-nary concern, where it was associatedwith abortions and encephalitis in sheepand cattle. Interestingly, evidence indi-cates that veterinary listeriosis was fre-quently foodborne. As a result of itswide distribution in the environment,its ability to survive for long periods oftime under adverse conditions, and itsability to grow at temperatures as low as–0.4°C, Listeria has since become recog-nized as an important foodborne patho-gen (ICMSF, 1996).

carriers of virulent types of Yersinia(Zen-Yoji et al., 1974).

Not surprisingly, because swine arethe primary reservoir of virulent typesof Y. enterocolitica, illness has been asso-ciated with the consumption of pork in-testines. The product, known as chitter-lings, is traditionally eaten during thewinter holidays, generally by AfricanAmericans, and the incidence of yersini-osis increases in December for the con-suming population. Infants and chil-dren are most often affected, largely be-cause of caretakers who do not properlywash their hands between preparingchitterlings and feeding young children,or practice adequate kitchen hygiene(Jones et al., 2003; Lee et al., 1991). Forepidemiological purposes, these casesare not reported as foodborne illness,since the food is not ingested, but clearly

the porcine intestines served as thesource of the pathogen.

A study of bulk tank milk revealedthat 6.1% of samples were positive for Y.enterocolitica. Further testing deter-mined that all strains were virulent (Ja-yarao and Henning, 2001). Chocolatemilk, pasteurized milk, and tofu packedin unchlorinated spring water have allbeen vehicles of outbreaks of yersiniosis(Ackers et al., 2000). The precise modeof transmission was not determined,but in each situation there was thoughtto be a lack of sanitary practice or defi-ciencies in good manufacturing practic-es. Circumstantial evidence suggeststhat porcine waste may have been theoriginal source of Y. enterocolitica in-volved in some of the food-associatedoutbreaks (Toma and Deidrick, 1975;Wauters, 1979).

Y. enterocolitica is commonly presentin foods; however, with the exception ofpork, most isolates from foods are avir-ulent. The infrequent presence of

Y. enterocolitica is com-monly present in foods;however, with the excep-tion of pork, most isolatesfrom foods are avirulent.



Significance as a Pathogen Immunocompromised humans are

highly susceptible to virulent Listeria.The microorganism causes approxi-mately 1,600 cases of listeriosis, result-ing in approximately 415 deaths annual-ly (FDA/CFSAN, 2003b). Only thehemolytic species of Listeria (L. monocy-togenes, L. seeligeri, and L. ivanovii) areassociated with pathogenicity. Of these,only L. monocytogenes is consistentlypathogenic; L. ivanovii has rarely beenreported to be involved in human pa-thology, and L. seeligeri was reportedonly once to be the cause of meningitisin a non-immunocompromised adult.

Listeriosis is a very serious and oftenfatal infection primarily affecting theelderly and perinates. However, gas-trointestinal illness has recently beenrecognized as a possible manifestationof ingestion of Listeria. Unlike the moresevere form of listeriosis, gastrointesti-nal illness, which generally results morethan 12 hr after ingestion, primarily af-fects seemingly healthy adults. In someinstances, this type of illness is associatedwith the consumption of large doses of L.monocytogenes (Dalton et al., 1997).

In humans, ingestion of as few as1,000 cells of the bacteria, which theninvade macrophages, may be marked bya flu-like illness (malaise, diarrhea, andmild fever). This enteric phase, however,may be without symptoms, or so mildas to go unnoticed. A carrier state maydevelop. Between 1–10% of symptom-less persons may be excretors of L.monocytogenes (FDA/CFSAN, 2003b).Identification of Listeria as the causativeagent of foodborne illness is complicat-ed by the high carrier rate in humansand must be isolated from the food forconfirmation (FDA/CFSAN, 2003b).

Following invasion of macrophages,virulent strains of Listeria may thenmultiply, resulting in disruption of thesecells and septicemia. In this stage, occur-ring five days to three weeks after inges-tion, the microorganism has access to allbody areas and may involve the centralnervous system, the heart, the eyes, orother locations, including the fetuses ofpregnant women (FDA/CFSAN, 2003b).The stage of pregnancy upon exposureof the fetus to the microorganism deter-mines the outcome for the fetus (abor-tion, stillbirth, or neonatal sepsis). Theperinatal and neonatal mortality rate is80% (FDA/CFSAN, 2003b). Death israre in healthy adults; however, the mor-tality rate may approximate 50% in the

immunocompromised, newborn, orvery young (FDA/CFSAN, 2003b). Feveris a common symptom, and other com-plaints may vary from nonspecific fa-tigue and malaise to enteric symptoms.

Forms of listeriosis involving thecentral nervous system include meningi-tis, encephalitis, and abscesses. The clin-ical course for meningitis develops andprogresses suddenly, and the fatality ratecan be as high as 70%. Localized formsof listeriosis, which are rare, include en-docarditis, endophthalmitis, and osteo-myelitis, with the involvement of othersites such as skin, spleen, gall bladder,and lymph nodes. All forms of listeriosisare more likely to accompany immuno-

compromised states, whether natural (asin pregnancy, or in the elderly) or in-duced as a result of medical treatment(such as with corticosteroids). Individu-als using antacids or cimetidine mayalso be more susceptible to infection,because of changes in stomach acidity.Listeriosis is often treated with penicillinor ampicillin (FDA/CFSAN, 2003b).

Association with FoodsListeria is ubiquitous in nature, oc-

curring in soil, vegetation, and water(Beuchat and Ryu, 1997; Coyle et al.,1984; Pearson, 1970), and therefore isfrequently carried by humans and ani-mals. Listeria can survive for long peri-ods in both soil and plant materials. In-gestion of contaminated silage by rumi-nants has been linked to the occurrenceof Listeria in milk (Donnelly, 1987).

Biological oxidation during wastewa-ter treatment favors the growth of Liste-ria in sewage. Sludge provides a favor-able environment for survival andgrowth of L. monocytogenes for manymonths (Guenich and Muller, 1984). L.monocytogenes in concentrations of 103–104 cells/mL have been found associatedwith sewage plant effluents, while sew-age sludges have shown higher concen-trations. This makes the use of sewageplant effluent or waters receiving sewageplant effluent for irrigation of edible

crops dangerous. The use of sludge forfertilization of edible crops is equallyhazardous.

L. monocytogenes can grow in the pHrange of 4.39–9.4 in a good growth me-dium (ICMSF, 1996). Environmentswith pH values outside that range areunfavorable for survival. The microor-ganism has readily survived the pH 5environments of cottage cheese and rip-ening Cheddar cheese, and was detect-able in the latter product (pH 5.0–5.15)for more than one year (Ryser andMarth, 1987).

Listeria is salt tolerant, growing tolevels of visible turbidity in the labora-tory media Tryptic Soy Broth (pH 5.0)after 41 hr at 25°C in presence of 10%sodium chloride (ICMSF, 1996). How-ever, 12.5% sodium chloride inhibitedthe growth of L. monocytogenes at pH6.0 between 8 and 30°C (Razavilar andGenigeorgis, 1998). Listeria is reportedto survive for three months in dry fod-der and more than six months in drystraw. Dry feces have been reported tomaintain viability of the microorganismfor more than two years (Gray, 1963),and survival in soil for up to 295 dayshas been recorded. Listeria is relativelyresistant to drying, as demonstrated onglass beads and tile surfaces. Survival istemperature dependent, with lower tem-peratures enhancing survival (Dickgie-ber, 1980; Welshimer, 1960), an impor-tant factor in Listeria’s significance inthe food chain.

The first reported outbreak of food-borne listeriosis occurred betweenMarch and September 1981. Coleslawwas implicated as the cause of 34 casesof perinatal listeriosis and seven cases ofadult listeriosis in the Maritime Prov-ince of Nova Scotia. Perinatal cases werecharacterized by acute febrile illness inpregnant women, followed by spontane-ous abortion (five cases), stillbirth (fourcases), live birth of a seriously ill prema-ture or term infant (23 cases), or livebirth of a well infant (two cases). The fa-tality rate for infants born alive was27%. The outbreak strain was isolatedfrom two unopened packages of cole-slaw from the plant but was not cul-tured from the manufacturing plant en-vironment. The incriminated coleslawwas traced to a farm where the cabbagewas grown in fields fertilized with sheepmanure. Two of the sheep on the farmhad previously died from listeriosis(Schlech et al., 1983).

Although listeriosis is more com-

The first reported outbreakof foodborne listeriosisoccurred between Marchand September 1981.


raw and finished product, whether thesecross-connections be humans, equip-ment, water, air, or the piping arrange-ment within the plant. If a personnelpractice or movement within the plantprovides contamination potential, itshould be eliminated or modified. If apiping arrangement can, under somecircumstances, transport potentiallycontaminated material from the “dirty”area or process to the finished productarea, it should be eliminated, no matterhow many valves intervene. Not eventhe air in the potentially contaminatedarea should be shared with the finishedproduct area.

Thus, the sanitation regimen of theentire plant must be under constantscrutiny and evaluation. Even with strictsanitation practices, all open productprocesses should be considered poten-tial contamination points, and carefulscrutiny, including microbiologicalanalysis, should be a part of the qualitycontrol practice.

Concern cannot stop at the point ofpackaging; storage and distributionpractices must also be evaluated as partof an effective quality control program.The ability of L. monocytogenes to with-stand harsh environmental conditionssuch as pH extremes, heat, and freezingrequires special considerations whenplanning for storage and distribution.Surviving microorganisms can grow inproducts under conditions that wouldinactivate other microorganisms. Addi-tionally, L. monocytogenes is capable ofdoubling in number each 1.5 days whenrefrigerated at 4°C (39.2°F). This is ofparticular concern since more than halfthe home refrigerators surveyed in a1999 Audits International study exceed-ed 39°F (FDA/CFSAN and USDA/FSIS,2003).

Vibrio speciesJoseph M. Madden

The three Vibrio species of primarysignificance in foodborne illnesses are V.cholerae (serogroups O1, non-O1, andrecently, O139), V. parahaemolyticus,and V. vulnificus. Another species, V.mimicus sp. novo (Davis et al., 1981),formerly included in the species V. chol-erae, also is a recognized pathogen.

Significance as a PathogenAn estimated 10.1 Vibrio infections

per million adults who consume rawoysters occur annually in the United

monly reported as sporadic cases, ofwhich about 20% result from the con-sumption of unheated hot dogs and un-dercooked chicken, there have beenmany outbreaks involving a variety offoods (FDA/CFSAN and USDA/FSIS,2003). Hot dogs and deli meats contam-inated with L. monocytogenes serotype4b caused a 1998–1999 outbreak affect-ing 101 individuals in 22 states (FDA/CFSAN and USDA/FSIS, 2003). In 2002,63 cases of listeriosis, of which threewere perinatal, resulted from the con-sumption of sliced deli meat in eightNortheastern states. This type of foodproduct was also responsible for an out-break in 2000 affecting 29 people in 10states (CDC, 2000a).

The soft cheeses are now infamousfor their susceptibility to contaminationwith L. monocytogenes, presumably dueto the use of raw milk or because ofpost-pasteurization contamination. Notonly is the manufacturing process opento contamination, but the cheese canserve as a growth environment where L.monocytogenes can multiply during thestorage period, even under refrigerationat 4°C (39.2°F). Concentrations of Liste-ria as great as 106 cfu/g have been noted.Mexican-style cheese made with rawmilk was implicated in two separateoutbreaks, one in 1984 causing 142 ill-nesses, and one in 2000–2001 resultingin 12 cases. The mortality rate was33.8% and 41.7%, respectively (CDC,2001; Linnan et al., 1988).

While association of listeriosis withraw milk consumption is not rare, it isinfrequent and sporadic. The results ofsurveys of raw milk for L. monocytogenesshow that the presence of the microor-ganism in raw milk is common in theUnited States and Europe (DominguezRodriguez et al., 1985; Lovett et al.,1987). The 2003 risk assessment con-ducted jointly by FDA/CFSAN andUSDA/FSIS pooled the data of 45 stud-ies, which showed an overall positiverate of 4.1% (FDA/CFSAN and USDA/FSIS, 2003). The presence of L. monocy-togenes in pasteurized milk in the UnitedStates is rare, with an average percentpositive of 0.4% in more than 10,000samples tested in 30 studies conductedworldwide (FDA/CFSAN and USDA/FSIS, 2003).

Control MeasuresBecause of the severity of listeriosis

and the ability of the microorganism togrow at refrigeration temperatures, both

the FDA and USDA have a “zero toler-ance” (absence in 25 g) for L. monocyto-genes in RTE foods. The 2003 risk as-sessment prepared jointly by the agen-cies estimated the relative risk of listeri-osis in 23 categories of RTE foods. Theagencies report that the results of therisk assessment will be used to focuscontrol efforts on high-risk foods (FDA/CFSAN and USDA/FSIS, 2003). In termsof risk per serving, deli meats ranked ashaving the highest risk.

Because of the link between dairyfoods, particularly milk, and L. monocy-togenes, the adequacy of pasteurizationtemperatures and times to inactivate L.monocytogenes were intensively studied.Scientific consensus on this subjectholds that the risk of post-pasteuriza-tion contamination is a far more seriousthreat than survival of the microorgan-ism to pasteurization heat treatment(63°C for 30 min or 71.7°C for 15 sec).Two studies by Bunning et al. (1986,1988) failed to support the hypothesisthat an intracellular location of the bac-teria in polymorphonuclear leucocytesmay confer heat resistance and allowsurvival during pasteurization. Heatshock at 42 or 48°C for up to 60 min didnot confer significantly increased ther-motolerance at 52 or 57.8°C (Bunninget al., 1990).

The control of Listeria in food prod-ucts begins at the lowest level in theprocessing chain—at the raw productsource. The growing environmentshould be kept free of the opportunityfor potential contamination. Waters sus-pected of contamination by Listeria,sewage plant effluent, and sewage slud-ges should not be used to produce cropsthat will be eaten without cooking orthat will become an ingredient of aproduct that will be eaten raw. Contain-ers, cartons, boxes, tankers, trucks, orrail cars used to transport the productto the process plant should be frequent-ly cleaned.

At the plant, the raw product itselfbecomes a source of contamination forthe environment. Product flow must bedesigned to segregate Listeria-free foodfrom potentially contaminated environ-ments by control of the product pro-gression through the plant in a mannerthat separates “clean from dirty,” or pro-cessed from raw. Personnel, too, musthave their movements within the plantrestricted depending on their exposureto potential contamination. Thereshould be no cross-connections between


States (FDA/CFSAN, 2001). Between1981 and 1994, V. parahaemolyticus andV. cholerae non-O1 caused roughly thesame number of illnesses, but gastroen-teritis was the common symptom forthe former, whereas septicemia occurredmore often in cases of the latter (FDA/CFSAN, 2001). V. vulnificus was thestrain most often responsible for illness,which usually manifested as septicemia(FDA/CFSAN, 2001).

V. cholerae is the bacterium responsi-ble for cholera epidemics and outbreaks.Three groups of V. cholerae strains arerecognized: serogroups O1, O139, andnon-O1. V. cholerae O1 is the serogrouptraditionally associated with choleracases, involving severe, watery diarrheathrough the action of cholera toxin(Farmer et al., 1985). The serogroup isdivided into the Classical and El Torbiotypes (Peterson, 2002). A new sero-type of V. cholerae, O139, emerged in In-dia in 1992 and spread to neighboringcountries. V. cholerae O139 also causescholera, but many secondary infectionsare asymptomatic (Albert, 1996).

In volunteer feeding studies of sero-group O1, ingestion of 106 cells causedillness. The incubation period is typical-ly from six hours to five days (FDA/CF-SAN, 2003b). Symptoms of severe chol-era include massive diarrhea with largevolumes of “rice-water” stool (clear flu-id with sloughed-off dead intestinalcells). If left untreated, severe dehydra-tion and death can occur. The massivewater loss associated with V. cholerae in-fections has been attributed to the pro-duction of toxin by the microorganismsthat adhere to and colonize the small in-testine of infected individuals. The chol-era toxin causes massive fluid loss fromcells lining the intestinal tract, and thefamiliar rice-water stool. Diarrhea canlast six to seven days and is usually ac-companied by vomiting. Milder formsof diarrhea due to V. cholerae O1, lastingone to five days (usually less than three),are more difficult to distinguish fromother mild diarrheas (Farmer et al.,1985).

V. cholerae non-O1 can cause a lesssevere cholera-like illness, but it alsocauses a much wider spectrum of dis-eases (Morris et al., 1981), including ex-traintestinal infections (Hughes et al.,1978). While cholera and gastroenteritiscaused by V. cholerae O1 is rare in theUnited States, illnesses caused by thenon-O1 serogroup are common and di-arrhea usually occurs 48 hr after inges-

tion. V. cholerae non-O1 can cause sep-ticemia in susceptible individuals (thosewith other underlying diseases such ascirrhosis, diabetes, and hemachromato-sis, or those on immunosuppressivetherapy) via the gastrointestinal tract orwound infections. Septicemia may beaccompanied by nausea, but is usuallynot accompanied by diarrhea. Fever,chills, and nausea may result from thecellulitis occurring with the wound in-fections.

Only about 3% and 0.2–0.3% ofstrains of V. parahaemolyticus arepathogenic on the West Coast and GulfCoast, respectively (FDA/CFSAN,2001). Pathogenic strains of V. para-haemolyticus are responsible for out-breaks of acute gastroenteritis, withsymptoms of nausea, vomiting, abdom-inal cramps, low-grade fever, chills, and

diarrhea (usually watery, sometimesbloody) that begin 12–96 hr after inges-tion. The disease is generally mild andself-limiting, lasting from two hours to10 days, but progression to septicemiacan be fatal (Farmer et al., 1985; FDA/CFSAN, 2001).

In Japan, where ingestion of raw sea-food is common, V. parahaemolyticus isan extremely important diarrheal agent,responsible for 50–70% of enteritis cas-es (Sakazaki and Balows, 1981). Al-though outbreaks caused by V. para-haemolyticus do occur worldwide, theywere uncommon in the United Statesuntil four outbreaks involving morethan 700 people occurred in 1997–1998. This prompted the FDA to begina risk assessment of the public healthimpact of V. parahaemolyticus in rawoysters, the food with which they aremost often associated (FDA/CFSAN,2001). A level up to 10,000 viable V.parahaemolyticus cells/g was consideredacceptable until outbreaks showed thatsome oysters contained as few as 100cells/g (FDA/CFSAN, 2001). The FDA

risk assessment estimated that only 15%of illnesses are caused by the consump-tion of raw oysters containing >10,000viable V. parahaemolyticus cells.

V. vulnificus is primarily associatedwith serious wound infections and life-threatening primary septicemia (Blake etal., 1979). Primary septicemia is a veryserious disease with a 50% fatality rate.Most cases identified also had preexist-ing liver disease (Blake et al., 1979);however, others have been healthy indi-viduals (Tison and Kelly, 1984). The se-vere wound infections, which may re-quire amputation or result in death,usually occur after trauma and exposureto marine animals or the marine envi-ronment (Blake et al., 1979). The mor-tality rate associated with wound infec-tions, 7%, is much lower than that asso-ciated with septicemia.

V. mimicus is associated with a mildgastrointestinal illness similar to V. chol-era non-O1, usually occurring after theconsumption of raw seafood, particular-ly oysters (Shandera et al., 1983). Themicroorganism is probably distributedworldwide in countries situated on anocean, and its ecology may be similar tothat of V. cholerae non-O1 (Farmer et al.,1985).

Association with FoodsApproximately 10–20% of the popu-

lation consumes raw seafood annuallyand is at increased risk of infection byVibrio species (FDA/CFSAN, 2001).Roughly two-thirds of cases are associat-ed directly or indirectly with seafood;i.e., either by consumption of raw sea-food or by consumption of seafoodwhich has been recontaminated aftercooking. Oysters are commonly associ-ated with Vibrio contamination. In ayear-long survey of retail oysters fromvarious locations in North America,78% of samples from the North Atlantic,Pacific, and Canadian coasts containedless than 0.2 MPN/g of V. vulnificus; fre-quency and levels of V. parahaemolyticuswere higher. The highest levels of Vibriowere in the Gulf Coast (Cook et al.,2002). Some Vibrio species may be ubiq-uitous in the estuarian environment;however, in many cases the presence ofVibrio is due to contamination of waterby sewage.

Asiatic cholera, caused by V. choleraeO1, remains endemic to the Asian conti-nent, and it is suspected that it was rein-troduced to the United States in 1973,when the first case of cholera since 1911

Approximately 10–20% ofthe population consumesraw seafood annually andis at increased risk ofinfection by Vibrio species.



cus by 60% in 2007 compared to the av-erage level of illness in 1995–1999 inLouisiana, Texas, Florida, and California(NSSP, 2002).

The number of vibrios in estuarinewaters tends to increase as the ambientwater temperature rises, resulting intheir appearance in larger numbers inshellfish and other fishes in these watersduring the warmer months of the year(Cook et al., 2002). This may explain theincrease in V. vulnificus infections inFlorida between May and October com-pared to the rest of the year (CDC,1993). Special precautions should betaken during these months. The ModelOrdinance identifies specific controlmeasures for harvesting areas where twoor more cases of V. vulnificus have oc-curred. In those areas, the time betweenharvesting and refrigeration is halved,from 20 hr to 10 hr, in the summermonths. Cooling immediately uponharvesting and maintaining lower tem-peratures until consumption controlsthe growth of vibrios in raw seafood.According to the 2001 Risk Assessmentconducted by FDA/CFSAN, the annualnumber of illnesses due to V. para-haemolyticus in raw oysters could be re-duced from 3,000 to 240, if rapid cool-ing was employed, or to 15, if oysterswere frozen (FDA/CFSAN, 2001). Heat-ing rapidly (5 min, 50°C) kills 4.5–6 logsof all vibrios. The risk assessment esti-mated that there would be less than 10cases of V. parahaemolyticus infectioncaused by raw oysters if mild heatingwas used (FDA/CFSAN, 2001). Ade-quate cooking and avoidance of recon-tamination will ensure the safety of sea-food.

All of the disease-causing vibrios arenaturally occurring in the marine envi-ronment and are thus naturally occur-ring contaminants of seafood, making itimpossible to prevent the contamina-tion of these fresh products. The mostimportant control measures to preventhuman infections with these microor-ganisms are hygienic measures designedto prevent their multiplication in theseuncooked foods and prevent the recon-tamination of cooked seafood. Certainindividuals (e.g., those with diabetes,those on immunosuppressive drugs, al-coholics, and cirrhotics) should avoidthe consumption of any raw seafood atany time of the year. As long as individ-uals continue to consume raw or under-cooked seafood, there will continue tobe Vibrio infections in the United States.

was reported. Latin America recently ex-perienced an epidemic affecting nearlyone million people and causing morethan 9,000 deaths (Guthmann, 1995).There have been occasional outbreaks ofcholera in the United States, suggestingthat the microorganism has become en-demic on the U.S. Gulf Coast, with spo-radic cases expected to occasionally oc-cur. Between 1965 and 1991, there were136 cases of cholera reported to theCDC, of which 93 were acquired in theUnited States. Fifty-six of the 93 caseswere attributed to V. cholerae O1. Con-taminated crabs and other shellfish har-vested near the Gulf Coast were themain source of infection (Weber et al.,1994). Large outbreaks of illness havebeen eliminated by the establishment ofadequate sewage treatment facilities.However, in 1997 more than 90,000Rwandan refugees became infected withV. cholerae O1 (Anonymous, 1998).

V. parahaemolyticus and, because oftheir physiological similarities, the othervibrios, are found mainly in marine en-vironments throughout the world.However, in a few instances, these vibri-os have been isolated from fresh-wateror non-marine fish. In these cases, it isassumed that the sodium chloride con-tent of the water was elevated, or thevibrios were present because of pollu-tion of the waters. A correlation be-tween the isolation of these vibrios andwater temperatures also exists. Vibriosare isolated more frequently and inhigher numbers during the warmermonths of the year. Oysters from statesalong the West Coast and British Co-lumbia contaminated with V. para-haemolyticus caused an outbreak inNorth America in 1997, resulting in 209cases (FDA/CFSAN, 2003b). In 1998, V.parahaemolyticus in oysters and clamsharvested from the Long Island Soundcaused an outbreak in Connecticut,New Jersey, and New York, and a sepa-rate outbreak occurred in the GulfCoast. The implicated serotype (O3:K6)was recognized as pathogenic in Asia,but had not previously been isolated inthe United States. (FDA/CFSAN, 2001).

V. vulnificus is ubiquitous in coastalwaters from Florida to Maine. CDC re-ports that between 1988 and 1995, 302cases of V. vulnificus infections were re-ported in the Gulf Coast states (CDC,1996b). An outbreak affecting 16 indi-viduals in Los Angeles was associatedwith the consumption of raw oysters.The three case fatalities consumed raw

oysters harvested in different places—one in Louisiana and two from two dif-ferent harvesters in Texas. All three hadunderlying liver diseases.

Vibrios in general will survive attemperatures below 10°C (50°F), buttheir reproduction is slowed or arrestedat these lower temperatures. An excep-tion is V. parahaemolyticus, which diesunder refrigeration temperatures of 0–5°C (ICMSF, 1996). At elevated temper-atures, the growth of vibrios in contam-inated seafood is extremely rapid. Gen-eration times of 12–18 min are commonin various seafood held at 30–37°C (86–98.6°F). All of the vibrios are sensitive todrying, yet are able to grow in a widerange of sodium chloride levels, varyingfrom 0.5 to 10% with optimal growth at2.2% (FDA/CFSAN, 2001).

Control MeasuresThe distribution of V. cholerae and V.

parahaemolyticus in the environment isbetter understood than that of the otherhuman pathogenic members of the ge-nus. This is mainly due to the lack ofsuitable enrichment and primary isola-tion media for the other vibrios, as wellas their recent discovery and associationwith human illness. These vibrios are,however, thought to be distributed simi-lar to V. parahaemolyticus because oftheir salt tolerance and similar physio-logical makeup.

Presence in estuarine waters does notcorrelate with fecal coliform counts;therefore, presence of the bacteria is nei-ther restricted to nor related to poor-quality oyster beds. There is less infor-mation concerning the occurrence of V.mimicus in the environment and sewage.It has been suggested, however, that thismicroorganism may also be a normalpart of the marine estuarine environ-ment and may also increase in numbersduring the warmer months. Vibrio maybe distributed throughout differentparts of the world through ballast water.

The Interstate Shellfish SanitationConference created a Model Ordinance,contained in the Guide for the Controlof Molluscan Shellfish, through the Na-tional Shellfish Sanitation Program. Thedocument provides the standards forharvesting areas and specifies testingmethods for the waters. Because the sea-food industry has been required to im-plement HACCP since 1997, the ModelOrdinance also gives details on HACCPfor shellfish. The ultimate public healthgoal is to reduce illness due to V. vulnifi-


Staphylococcus aureusOriginally authored by Rosetta L.Newsome. Reviewed by Cynthia M.Stewart.

Given adequate time, temperature,pH, water activity (a

w), and atmosphere

for growth, contaminating S. aureusmay multiply, and many strains mayproduce enterotoxins when the popula-tion exceeds 105 cells/g. Ingestion of thethermostable enterotoxins, rather thanthe bacterium itself, is responsible forfoodborne illness.

Significance as a PathogenStaphylococcal food intoxication is

estimated to cause 185,000 cases offoodborne illness annually (Mead et al.,1999). The true incidence is unknownfor several reasons, including poor re-sponse by victims to interviews con-ducted by health officials; misdiagnosisdue to symptoms being similar to thoseof other types of food poisoning (suchas the vomiting caused by Bacillus cereusemetic toxin); inadequate collection ofsamples for laboratory analysis; and im-proper laboratory examination (Ben-nett, 1986).

Unlike other foodborne illnesses,which usually have longer incubationperiods, onset of staphylococcal food-borne illness may occur between 30 minand 8 hr following consumption of thetoxin-containing food (Bergdoll, 1979).Most illnesses, however, occur within 2–4 hr (Bergdoll, 1979). Staphylococcalenterotoxins (SE) cause severe gastroen-teritis (inflammation of the intestinaltract lining). Common symptoms ofstaphylococcal intoxication include nau-sea, vomiting, retching, abdominalcramping, sweating, chills, prostration,weak pulse, shock, shallow respiration,and subnormal body temperature. Re-covery from this intoxication (which israrely fatal) usually occurs uneventfullywithin 24–48 hr.

Growth of staphylococci to a popula-tion of a million or more cells per gramof food is considered necessary for suffi-cient toxin production to elicit symp-toms of food poisoning. Several anti-genically different protein enterotoxinsexist (Tatini et al., 1984). To date, SE A,B, C1, C2, C3, D, E, G, H, I, and J havebeen identified (Balaban and Rasooly,2000). Enterotoxin A is the most toxicand the one most commonly involved instaphylococcal food poisoning out-breaks. Data from outbreaks involving

along with Salmonella infantis, was iso-lated from turkey that caused a food-borne outbreak at a Florida prison in1990 (Meehan et al., 1992). Methicillin-resistant S. aureus (MRSA), commonlyassociated with hospital infections,caused a foodborne outbreak when adelicatessen employee prepared cole-slaw. Tests concluded that the employeecarried the outbreak strain of MRSA,which was presumably transferred froma nursing home that the employee fre-quently visited (Jones et al., 2002).

Although the acidity (low pH) ofmayonnaise is inhibitory to the growthof staphylococci, growth may occur insalads (such as egg or chicken), wherethe otherwise low pH of the mayonnaiseis raised or buffered by other salad in-gredients. Food systems that contain saltor sugar also provide a favorable envi-ronment for growth of S. aureus, be-cause growth of more sensitive microor-ganisms is inhibited, but growth of S.aureus is not. Staphylococci can tolerateup to 10–20% salt and 50–60% sucrose.S. aureus is also tolerant to nitrite andmay therefore multiply in curing solu-tions or cured meats if they are subject-ed to conditions favorable for growth.

S. aureus is a facultative anaerobewhich grows best under aerobic condi-tions but is also capable of growth whenoxygen is present at reduced concentra-tions. Some oxygen must be present,however, for toxin production (Bergdoll,1979; Niskanen, 1977; Tatini, 1973). Themicroorganism prefers temperatures of95–98.6°F (35–37°C) but can grow attemperatures as low as 44°F (67°C) or ashigh as 118°F (47.5°C) (Bergdoll, 1979).While heat processing and normalcooking temperatures are sufficient tokill the bacterial cells, the enterotoxinsare heat-stable and are not inactivatedby heat processing or cooking. The ab-sence of viable staphylococci, therefore,does not ensure that a food is safe.

Growth may occur at aw as low as

0.86 under aerobic conditions or at aw

0.90 under anaerobic conditions. Al-though staphylococci are fermentativeand proteolytic, they do not usuallyproduce off-odors in foods or makethem appear spoiled. Presence of thebacteria or their toxins in foods would,therefore, not be detectable by sensorymethods.

Control MeasuresThe ubiquity of the staphylococci in

the human and animal environment ne-

enterotoxin A indicate that less than 1µgof toxin can result in illness (Tatini etal., 1984).

Association with FoodsS. aureus is commonly found in the

nose and throat (and thus on the handsand fingertips) and on the hair and skinof more than 50% of healthy individuals(Bergdoll, 1979). Any food which re-quires handling in preparation maytherefore easily become contaminated.Infected wounds, lesions, and boils offood handlers may also be sources ofcontamination, as well as coughing andsneezing by individuals with respiratoryinfections. S. aureus also commonly oc-curs on the skin and hides of animals,and may thus contaminate foods fromthese animals as a result of cross-con-tamination during slaughter.

A variety of foods can support thegrowth of S. aureus. Foods which sup-port growth best include proteinaceousfoods, such as meat and meat products,poultry, fish and fish products, milk anddairy products, cream sauces, salads(ham, chicken, potato, etc.), puddings,custards, and cream-filled bakery prod-ucts. Staphylococci are usually outnum-bered by competitive harmless microor-ganisms in raw foods because they donot compete well with other bacteria,and it is probably for this reason thatraw foods are less frequently implicatedas the cause of staphylococcal food poi-soning. Cooking, however, eliminatesthe normal competitive bacteria of rawfoods; therefore, it is in prepared foodssuch as meat, potato, and macaroni sal-ads, custards, and cream-filled bakeryproducts that growth of contaminatingstaphylococci may be permitted.

S. aureus intoxications are often as-sociated with institutions such asschools and prisons where food is oftenprepared in mass quantities and helduntil consumption. Lack of sanitationby workers and improper time-temper-ature combinations can lead to contam-ination of the product and growth ofthe microorganism to levels at whichtoxin is produced. Accordingly, out-breaks can affect a large number of peo-ple. An outbreak in 1990 in two schoolsin Rhode Island resulted from ham pre-pared at a satellite location that was sub-sequently handled by an employee whoharbored S. aureus. The ham was storedat inappropriate temperatures for atleast 15 hr and was not adequately re-heated (Richards et al., 1993). S. aureus,



cessitates good sanitation in processingoperations and food handling, and strictcontrol measures for prevention of bac-terial growth and subsequent toxin pro-duction. For staphylococcal food poi-soning to occur, four things must hap-pen: (1) the food must be contaminatedwith enterotoxin-producing staphylo-cocci; (2) the food must be capable ofsupporting the growth of the contami-nant; (3) the food must be held at atemperature sufficiently high and for asufficient period of time to permit suffi-cient growth to result in the formationof an emetic (vomiting) level of entero-toxin; and (4) the food must be con-sumed. There is little that can be doneabout the first two items, because mostfoods are subject to contamination (forthe reasons already given) and are capa-ble of supporting growth. Control of thetemperature is the most effective routeto control staphylococcal food poison-ing. Indeed, the majority of staphylo-coccal food poisoning outbreaks occurbecause of inadequate cooling and re-frigeration of foods. Adequate heat pro-cessing and cooking and proper coolingand refrigeration are therefore impor-tant control measures.

Clostridium perfringensRonald G. Labbe

Clostridium perfringens is probablythe most extensively studied anaerobicbacterial human pathogen. During thelast 90 years, C. perfringens has beenmost closely associated with gas gan-grene. The first indication of an associa-tion with food poisoning, however,came in the 1940s from Knox and Mac-Donald in England and McClung in theUnited States (Hobbs, 1979).

Significance as a PathogenBecause of the degree of underre-

porting, estimates of the number of an-nual cases range from 10,000 to 250,000in the United States. (FDA/CFSAN,2003b; Mead et al., 1999). Strains of themicroorganism are divided into five tox-in types, A to E, based on the produc-tion of four lethal extracellular toxins(alpha, beta, epsilon, and iota). Sincevirtually all food poisoning outbreaksare caused by type A strains, it is notnecessary to determine the toxin type insuch outbreaks.

Illness due to C. perfringens usuallyoccurs 8–22 hr after ingestion of foodcontaining large numbers (106 or more)

of vegetative cells. The toxicoinfection iscaused by sporulation of the bacterialcells in the intestine, accompanied byproduction of an intracellular entero-toxin. Sporulation and enterotoxin pro-duction may also occur in foods, how-ever (Craven, 1980; Naik and Duncan,1977). Diarrhea and severe abdominalpain are the usual symptoms. Nausea isless common, and fever and vomitingare unusual. Death is uncommon, usu-ally occurring in debilitated or institu-tionalized individuals, especially the eld-erly. Illness generally lasts 24 hr, but maylast up to two weeks (FDA/CFSAN,2003).

Association with FoodsC. perfringens type A can be consid-

ered part of the microflora of soil. Vir-tually all soil samples examined have re-vealed type A strains at levels of 103–104/

g. Types B, C, D, and E are obligate para-sites, mostly of domestic animals, anddo not persist in soils. The microorgan-ism is also found in the intestinal con-tents of virtually every animal exam-ined, with wide variation in numberswithin and between species. For exam-ple, after chilling, carcasses were positivefor C. perfringens in 13 of 16 broilerflocks. In the affected flocks, 8–68% ofindividual carcasses harbored C. perfrin-gens, with a mean of 30% (Craven et al.,2001). In human infants, adult levels ofC. perfringens (103–105/g) are estab-lished by six months of age.

Meat and poultry products are by farthe most common C. perfringens vehi-cles. This is not surprising, given the in-cidence of C. perfringens in such foodsand the microorganism’s fastidious nu-trient requirements. When 197 commi-nuted meat samples were tested for the

presence of C. perfringens after cookingat 73.9°C, only two samples, bothground pork, had levels above the detec-tion limit of three spores/g (Kalinowskiet al., 2003). Nevertheless, in the UnitedStates, from 1988 to 1997, meat andpoultry items were involved in at least33% of C. perfringens outbreaks; asource was not identified nearly 20% ofthe time, and multiple vehicles, whichmay have included meat or poultry wereinvolved in 28% of outbreaks (CDC,1996a; CDC, 2000b). Fish are not com-monly involved in outbreaks, althoughthe body surface and the alimentary ca-nal of most fish of several species harborthe microorganism.

C. perfringens has been found, albeitat a much lower incidence, in virtuallyall types of processed foods examined.This is important because some of theseitems, such as spices and herbs, are add-ed to larger volumes of cooked foods.Although processed foods are rarely ve-hicles for this type of food poisoning,minestrone soup was the vehicle for a1990 outbreak in Michigan that made32 people ill (Roach and Sienko, 1992).The soup was prepared two days beforeconsumption and was not promptlycooled or adequately reheated.

Foodservice establishments are themost likely sites for acquiring the illness.This reflects the need for such facilitiesto cook large amounts of food well inadvance of serving. Because of the rela-tively mild nature of the illness, manyoutbreaks, especially those involvingfamilies, go unreported. The reportedincidence is undoubtedly only the tip ofthe iceberg. The incidence from properlyhandled commercially prepared food isvirtually zero.

Cured meat products are rarely vehi-cles for outbreaks of C. perfringens foodpoisoning. Two unrelated outbreaks in1993 were due to improper holding ofcorned beef. The outbreaks stemmedfrom the fact that in both instances, thecorned beef was prepared in advance inmass in anticipation of the large de-mand around St. Patrick’s Day. The loadof C. perfringens in corned beef samplesexceeded 105 CFU/g in each outbreak(CDC, 1994). In general, these productsare considered safe when properly han-dled due to the curing agents them-selves, the low initial spore load in suchproducts, the heat treatment adminis-tered, and the relative sensitivity to thecuring agents of the surviving, and per-haps injured, spore population.

Since C. perfringens willnot grow at refrigeratedtemperatures, the princi-pal cause of virtually alloutbreaks is failure toproperly refrigeratecooked foods, especiallylarge portions.


poultry. Thus, preventive measures de-pend to a large extent on knowledge ofproper food preparation and storagetechniques, especially temperature con-trol. In the case of C. perfringens foodpoisoning, it is clear that education andsupervision of food handlers remaincritical control points.

Clostridium botulinumMerle D. Pierson and N.R. Reddy

The popularity of botulinum neuro-toxin (BoNT), also known as “Botox,” toreduce facial wrinkles and the potentialto use C. botulinum and its neurotoxinas agents of intentional contaminationhave sparked renewed interest in thispathogen, historically associated withhome-canned or prepared products.The etiologic agent of botulism was firstisolated from inadequately cured hamin 1896 by E.P.M. van Ermengen. SinceC. botulinum spores are widely distrib-uted, they may find their way into pro-cessed foods through raw food materialsor by contamination of foods after pro-cessing and cause botulism in humans.Processors and consumers must takepreventive measures to eliminate C. bot-ulinum or to inhibit growth and toxinproduction by this microorganism toprevent outbreaks of botulism from oc-curring. Instances of botulism resultingfrom improper canning are primarilyassociated with home canning, not in-dustrial processes. This results in a lim-ited number of cases annually, but doesnot mean that attention should be di-verted from this pathogen.

Significance as a PathogenSeven types (A, B, C, D, E, F, and G)

of C. botulinum are recognized based onthe antigenic specificity of toxin. How-ever, all types of C. botulinum produceprotein neurotoxins with similar effectson an affected host. The types of C. bot-ulinum differ in their tolerance to saltand water activity, minimum growthtemperature, and heat resistance ofspores. All type A strains are proteolytic,and all type E strains are nonproteolytic.Types B and F both contain some pro-teolytic and nonproteolytic strains.Types A, B, E, and F are involved in hu-man botulism; type C causes botulismin birds, turtles, cattle, sheep, and hors-es; and type D is associated with foragepoisoning of cattle and sheep in Austra-lia and South Africa. No outbreaks oftype G have been reported; however,

Control MeasuresThe problem of C. perfringens food-

borne illness is one associated with thefoodservice industry and consumers.The two most important attributes of C.perfringens are its ability to grow rapidlyat elevated temperatures and its abilityto form spores. Control measures mustconsider both of these.

Since C. perfringens will not grow atrefrigerated temperatures, the principalcause of virtually all outbreaks is failureto properly refrigerate cooked foods, es-pecially large portions. An analysis ofoutbreaks occurring between 1973 and1987 showed that 97% were due to im-proper holding temperatures (Bean andGriffin, 1990). Spores on raw meat andpoultry can survive cooking and resumevegetative cell growth when the coolingproduct reaches a suitable temperature.Rapid, uniform cooling is therefore im-perative. Prior to 1996, USDA/FSIS pre-scribed specific time-temperature com-binations for the cooking and cooling ofRTE meats. In 1999, USDA issued a finalrule allowing processors some flexibilityin determining the process. With respectto C. perfringens, any cooling operationthat prevented one log of growth wouldmeet the standard. The process does notneed to be validated for uncured meatscooled from 54.4°C (130°F) to 26.7°C(80°F) in less than 1.5 hr, and furthercooled to 4.4°C (40°F) in less than 5 hr.(Anonymous, 1999). Cooking also re-duces the oxidation/reduction potentialand thereby promotes anaerobic condi-tions, a factor especially important inliquid foods and rolled meats, where thecontaminated outside surface is rolledinto the middle. Other factors includepreparation of food a day or more be-fore serving, inadequate hot-holding,and inadequate reheating of cooked,chilled foods. Cooked foods should bedivided into small portions so that re-frigeration temperature is reached with-in two hr. These foods should be re-heated to a minimum internal tempera-ture of 74°C (165°F) immediately beforeserving to destroy vegetative cells.Cooked meat should be kept above 60°C(140°F) or below 4°C (40°F).

It is not possible to prevent carriersof C. perfringens from handling food,since all people harbor the microorgan-ism in their intestinal tract. Similarly,the bacterium is present in a wide vari-ety of foods. However, food handlersplay a minimal role as a contaminationsource of C. perfringens in meat and

type G has been isolated in cases of sud-den and unexpected death in humans(Sonnabend et al., 1981) and infants(Sonnabend et al., 1985). C. botulinum isisolated from a variety of sources, suchas soil, marine and lake sediments, fecesand carcasses of birds and animals, hu-man autopsy specimens, rotting vegeta-tion, and foods. In addition to thesetypes, C. baratii and C. butyricum,which produce BoNT, have been isolat-ed from infant botulism cases (Hall etal., 1985; McCroskey et al., 1986; Suen etal., 1988).

C. botulinum produces a BoNT,which is a simple polypeptide that con-sists of a 100-kDa “heavy” chain joinedby a single disulfide bond to a 50-kDa“light” chain. Three-dimensional struc-ture of type A BoNT has been resolvedto 3.3Å resolution using data collectedfrom multiple crystals at 4°C (Lacy etal., 1998). BoNTs are toxic to humansand animals; on a weight basis, they arethe most lethal substances known, beingup to 100,000 times more toxic thansarin (Shapiro et al., 1998). The exact le-thal dose of BoNT for humans is notknown. However, by extrapolation, Ar-non and others (2001) estimated the le-thal dose of crystalline type A BoNT forhumans from primate data. The lethaldose of crystalline type A BoNT for a 70kg (154 lb) human would be approxi-mately 0.09–0.15µg via intravenous orintramuscular injection, 0.70-0.90µg viainhalation, and 70µg via oral ingestion(1µg/kg).

Type A BoNT, which causes roughlyhalf the annual cases of foodborne bot-ulism, is more lethal than types B and E,which are equally responsible for the re-maining cases (Shapiro et al., 1998). Thetoxin is a protein that can be inactivatedby heat (80°C for 10 min or 85°C for 5min). The toxin in solution is colorlessand odorless and can be absorbed intothe blood stream through the respirato-ry mucous membranes as well asthrough the wall of the stomach and in-testine. Type A BoNT is the first biologi-cal toxin licensed for treatment of hu-man diseases, namely for cervical torti-collis, strabismus, and blepharospasmassociated with dystonia, to reduce facialwrinkles and hemifacial spasm, and forseveral unlicensed treatments of otherhuman diseases (Arnon et al., 2001;Johnson, 1999; Schantz and Johnson,1992).

Botulism is currently classified intofour categories: (1) classical foodborne



botulism and intoxication caused by theingestion of preformed BoNT in con-taminated food; (2) wound botulism,the rarest form of botulism, which re-sults from the elaboration of BoNT invivo after growth of C. botulinum in aninfected wound; (3) infant botulism, inwhich botulinal toxin is elaborated invivo in the intestinal tract of an infantwho has been colonized with C. botuli-num; and (4) an “undetermined” classi-fication of botulism for those cases in-volving individuals older than 12months in which no food or woundsource is implicated (Anonymous, 1979;Rhodehamel et al., 1992).

Foodborne botulism results from theconsumption of food in which C. botuli-num has grown and produced toxin.The toxin is absorbed and irreversiblybinds to peripheral nerve endings. Signsand symptoms of botulism develop 12–72 hr after consumption of the toxin-containing food. The signs and symp-toms include nausea; vomiting; fatigue;dizziness; headache; dryness of skin;mouth; and throat; constipation; paraly-sis of muscles; double vision; and diffi-culty breathing. Duration of illnessranges anywhere from 1–10 days ormore, depending upon the host resis-tance, type and amount of toxin ingest-ed, and type of food. Treatment includesadministration of botulinal antitoxinand appropriate supportive care, partic-ularly respiratory assistance lasting be-tween two to eight weeks (Shapiro et al.,1998). Recovery may take several weeksto months. Death may result; however,the mortality rate is less than 10%.

Infant botulism, which affects infantsunder 14 months of age, was first recog-nized in California in 1976 (Midura andArnon, 1976). This type of botulism isthought to be caused by the ingestion ofC. botulinum spores that colonize andproduce toxin in the intestinal tract ofinfants (Paton et al., 1982; Wilcke et al.,1980). Honey has been implicated as apossible source of spores. Other non-sterilized foods, as well as nonfooditems in the infant’s environment, mayalso be sources of spores. Infant botu-lism is diagnosed by demonstrating bot-ulinal toxins and spores in the infant’sstools.

Clinical symptoms of infant botu-lism start with constipation that occursafter a period of normal development.This is followed by poor feeding, lethar-gy, weakness, pooled oral secretions,and weak or altered cry. Loss of head

control is striking. Recommended treat-ment is primarily supportive care. Re-cent clinical studies have shown that theuse of botulism immune globulin (BIG)reduces the time of hospitalization andthe need for supportive ventilation andtube feeding (AAP, 2000). Antimicrobialtherapy is not recommended (Sakagu-chi, 1979).

Due to the rarity of the illness, botu-lism is often misdiagnosed; however,rapid detection is necessary for thetimely administration of antitoxin. De-tection of the toxin, either in a suspectedfood vehicle or the serum or stool ofsuspected cases, is generally accom-plished using an Association of Analyti-cal Communities-approved mouse bio-assay. Amplified enzyme-linked immun-osorbent assay (ELISA) and amplified

ELISA/enzyme-linked coagulation assay(ELCA) show promise for more rapiddetection of toxin; however, these assayshave not yet replaced the mouse bioas-say in general use (Ferreira, 2001; Ferrei-ra et al., 2003; Roman et al., 1994).

Association with FoodsSpores of C. botulinum are widely

distributed in cultivated and forest soils,shore and bottom deposits of streams,lakes, and coastal waters; gills and vis-cera of crabs and other shellfish; and theintestinal tracts of fish and animals (Ey-les, 1986; ICMSF, 1980; Lynt et al., 1975;NFPA/CMI, 1984; Rhodehamel et al.,1992; Sakaguchi, 1979). Although thespores are widespread, only about 22–24cases of foodborne botulism are report-ed to the CDC annually (Shapiro et al.,1998; Townes et al., 1996). Type A oc-curs more frequently in soils of thewestern regions of the United States,and type B is found more frequently in

the eastern states and in Europe. Type Eis most often associated with water envi-ronments.

Since fruits and especially vegetablesare often in contact with soil, thesefoods can easily become contaminatedwith spores of C. botulinum. The micro-organism has been isolated from freshand processed meats, but the incidenceis generally low (Hauschild and Hilshe-imer, 1980; Rhodehamel et al., 1992).

C. botulinum spores have been de-tected in honey and corn syrup (Lynt etal., 1982). C. botulinum is a natural con-taminant of fish. The intestinal tract offish and its contents have been reportedto be the main reservoirs (Huss et al.,1974). Since many processes do not in-clude steps that are lethal to spores of C.botulinum, the microorganism may befound occasionally in some finishedproducts, such as smoked fish.

Proteolytic types A, B, and F strainsproduce heat-resistant spores that are amajor concern in the processing of low-acid canned foods. The nonproteolytictypes B, E, and F strains produce sporesof low heat resistance and cause prob-lems primarily in pasteurized or un-heated foods (Sperber, 1982). Proteina-ceous foods, such as meats, and nonpro-teinaceous foods, such as vegetables, canprovide sufficient nutrients for growthand toxin production by C. botulinum.The proteolytic types digest proteins infoods and produce foul odors, whichmay warn consumers. However, therehave been outbreaks where only a smallamount of food having an off-odor wasconsumed or food without evidence ofspoilage was consumed. The latter is es-pecially true for the nonproteolyticstrains (Lynt et al., 1975).

The proteolytic and nonproteolytictypes of C. botulinum differ in their tol-erance to salt, water activity, minimumgrowth temperature, and spore heat re-sistance. The optimum temperature forgrowth and toxin production by theproteolytic types is about 35°C (95°F),with very slow growth at both 12.5°C(55°F) and 50°C (122°F); however, atthe latter temperature, toxin may beslowly inactivated (Bonventre andKempe, 1959). The nonproteolytic typescan grow between 3.3°C (38°F) and45°C (113°F), with an optimum forgrowth and toxin production at about30°C (86°F). Refrigeration above 3.3°Cmay not be a complete safeguard againstbotulism for foods containing nonpro-teolytic strains (Eklund et al., 1967; Pa-tel et al., 1978).

Conditions that favor C.botulinum growth andtoxin production include arelatively high-moisture,low-salt, low-acid (pH >4.6) food that is devoid ofoxygen and stored withoutrefrigeration.


control measures, such as proper refrig-eration and frozen storage, must bemaintained from purchase until con-sumption. If a commercial product ismishandled, control measures are com-promised and botulism may result.

Bacillus cereusMichael P. Doyle

Bacillus cereus has been a recognizedcause of foodborne illness for almost 50years. Two types of illnesses, the emeticand diarrheal responses, are caused bytwo distinct enterotoxins produced bythis microorganism (Melling et al.,1976).

Significance as a PathogenAn estimated 27,000 cases of food-

borne illness due to B. cereus occur an-nually in the United States (Mead et al.,1999). A large molecular weight proteincauses the diarrheal response, whereascereulide, a small peptide stable for 20min at 121°C, causes the emetic re-sponse (Granum, 1997). The diarrhealresponse, which closely mimics symp-toms of the illness caused by C. perfrin-gens, generally produces mild symptomsthat usually develop between 6–15 hrafter ingestion. The emetic (or vomit-ing) response occurs within a few (oneto six) hr after ingestion, closely mim-icking symptoms produced by staphylo-coccal enterotoxin. Symptoms of the di-arrheal syndrome include diarrhea, ab-dominal cramps, and tenesmus, whereasnausea and vomiting are the principalsymptoms of the emetic syndrome. Re-covery is usually complete within 24 hrafter onset, and further complicationsdo not occur. The diarrheal syndromeresults following ingestion of largenumbers of the microorganism, whereasthe emetic syndrome is likely to resultfrom ingestion of preformed toxin inthe food.

Association with FoodsB. cereus is common in soil and on

vegetation and can be readily isolatedfrom a variety of foods, including dairyproducts, meats, spices and dried prod-ucts, and cereals (especially rice). Inview of its wide distribution in the envi-ronment, B. cereus is ingested from timeto time and inevitably is part of thetransitory human intestinal microflora.Studies have revealed that low numbersof B. cereus are present in the intestinal

Almost every type of food product(dairy products, vegetables, jarred pea-nuts, fishery products, meat products[beef, pork, and poultry], and condi-ments [chili sauce, chili peppers, tomatorelish, and salad dressing]) has been im-plicated in foodborne botulism out-breaks (Arnon et al., 2001; Aureli et al.,2000; Chou et al., 1988; Kalluri et al.,2003; Peck, 2002; Rhodehamel et al.,1992; Weber et al., 1993). The largestU.S. outbreak since 1978 occurred in1994 as the result of improper storage ofbaked potatoes later used to make dip.Of the 30 reported cases, four requiredmechanical ventilation (Angulo et al.,1998). Cheese and other dairy productswere implicated in less than 1% of re-ported cases of C. botulinum intoxica-tion since 1899 (Townes et al., 1996).Improper handling and storage of com-mercially canned cheese was responsiblefor a restaurant outbreak in 1993 affect-ing eight people and resulting in onedeath (Townes et al., 1996). In 1989,toxin production in garlic-in-oil causedthree cases of botulism. Since this wasnot the first time this type of product,which had a pH >4.6, was implicated,FDA began requiring that products ofthis nature be acidified or contain anti-microbial agents (Morse et al., 1990).FDA issued guidance for labeling offoods requiring refrigeration followingbotulism outbreaks caused by blackbean dip and clam chowder. Illnesscould have been prevented had theitems, which were not commerciallysterile, been properly refrigerated by theconsumer. Although the products weredisplayed in the refrigerated section ofthe grocery store, FDA felt that becauseof the packaging consumers did not fol-low the directions for refrigeration.Temperature abuse allowed the growthof C. botulinum to toxin-producing lev-els. To prevent future outbreaks, FDArecommended that foods that maypresent a safety risk if not refrigeratedbear the label, “IMPORTANT: Must BeKept Refrigerated” (FDA, 1997).

Control MeasuresConditions that favor C. botulinum

growth and toxin production include arelatively high-moisture, low-salt, low-acid (pH > 4.6) food that is devoid ofoxygen and stored without refrigeration(above 38°F or 3.3°C). The food indus-try uses a variety of physical and chemi-cal treatments to either destroy C. botu-

linum spores or control growth andsubsequent toxin production. Reviewsof specific treatments and hurdle con-cepts used for control of C. botulinumin a variety of food products have beenpublished (Hauschild, 1989; Rhode-hamel et al., 1992).

Typical methods used to preventbotulism include reduction of vegetativecell and/or spore contamination in thefood by heat treatment (e.g., pressurecooking for canning) to obtain “com-mercial sterility” and use of lower heattreatments (pasteurization) in combina-tion with other control measures. Ni-trite and salt (in a brine solution) arecontrol measures used in addition toheat treatment for low-acid cannedmeats (e.g., canned ham). In addition tothese controls, refrigeration is an impor-tant control for perishable vacuum-packaged meats. Acidification is used forother food products such as pickles,mayonnaise, and canned fruit products.Reduction of moisture level (drying) be-low an a

w of 0.93 is employed in certain

foods. Frozen storage is yet another sig-nificant factor in controlling pathogengrowth and subsequent toxin formation.

The excellent safety record of curedmeats relative to botulism outbreaks hasbeen largely attributed to the use of ni-trite as a curing ingredient. Many stud-ies have been published on the efficacyof sodium nitrite in inhibiting C. botuli-num growth and toxin production inperishable cured meats such as wieners,bacon, canned ham, luncheon meat, andcanned comminuted meat (Rhode-hamel et al., 1992). In perishable curedmeats, safety cannot be totally attribut-ed to nitrite alone, but a variety of fac-tors and their interaction with nitriteprovide protection against botulinalgrowth and toxin production. Factorssuch as heat treatment, acidity (pH),salt, and bacterial spore level influencethe effect of nitrite on C. botulinumgrowth and toxin production. Othercuring adjuncts, such as ascorbic acidand sodium erythorbate, may also influ-ence the efficacy of nitrite.

The greater incidence of botulismdue to improper home processing andstorage of foods, particularly home-canned foods, compared to commercial-ly processed foods reflects the food in-dustry’s greater awareness and controlof the key factors in inhibition of C. bot-ulinum growth and toxin production.Consumers must recognize that certain



tract of more than 10% of the healthyadult population (Ghosh, 1978; Shina-gawa et al., 1980).

Foods that have been implicated asvehicles in outbreaks of B. cereus diar-rheal-type food poisoning include cere-al dishes that contain corn and cornstarch, mashed potatoes, vegetables,meat products, puddings, soups, andsauces. B. cereus emetic-type food poi-soning is most frequently associatedwith fried or boiled rice dishes(Johnson, 1984). Chicken fried rice wasimplicated in a 1993 outbreak in Virgin-ia that affected 14 people (FDA/CFSAN,2003b). Other starchy foods such asmacaroni and cheese also have been in-criminated as vehicles of the emetic syn-drome.

B. cereus is a spore-forming microor-ganism whose spores will generally sur-vive cooking. Spores of the microorgan-ism normally occur on many foodsfrom harvest through primary process-ing. The microorganism is not consid-ered a hazard at the very low levels typi-cally present in food. However, prob-lems develop when food (especiallycooked foods which are devoid of mostheat-sensitive microbial competitors) isheld at 10–55°C (50–131°F) for a longperiod of time. Under these conditions,the microorganism grows to large num-bers, releasing toxin during growth inthe food and in the intestinal tract fol-lowing ingestion. Although ingestion ofmore than 105 cells/g is required to pro-duce illness (Hobbs and Gilbert, 1974),ingesting large numbers of B. cereusdoes not always produce illness (Dack etal., 1954). To date, all outbreaks of bothtypes of B. cereus illness have resultedbecause of improper holding tempera-tures or slow cooling of large amountsof food. Because these illnesses are in-toxications, they are not infections andare not transmitted. It does not appearthat processing of raw agricultural com-modities contributes to the incidence ofB. cereus illness.

Control MeasuresMeasures to reduce or eliminate B.

cereus food poisoning are well estab-lished. Most disease incidents resultfrom time-temperature abuse of food infood-handling establishments. This canbe prevented by holding foods at greaterthan 60°C (140°F) until served, or rap-idly cooling foods to below 10°C (50°F)for storage. Refrigerating foods in small

30% to 80% despite antibiotic treatment(Muytjens et al., 1983). There have beenconflicting reports regarding low birthweight as a risk factor for infection(FDA/CFSAN, 2002; Muytjens et al.,1983; van Acker, 2001); however, theContaminants and Natural ToxicantsSubcommittee of the FDA/CFSAN’sFood Advisory Committee reviewed riskfactors for E. sakazakii infections andconcluded that infants born at less than36 weeks gestational age were at risk un-til six weeks post-term. Other risk fac-tors identified were hospitalization inlevel-two or level-three neonatal inten-sive care units and immunocompro-mised health status (FDA/CFSAN,2003a).

Illness symptoms normally appear afew days after birth, and the health ofthe infant rapidly deteriorates(Muytjens et al., 1983). Infection mayresult in meningitis (58%), necrotizingenterocolitis (29%), or sepsis (17%). Inone outbreak in which 11 infants werepositive for E. sakazakii, one developedmeningitis and four others had clinicalsigns of severe sepsis, although the mi-croorganism could not be isolated fromthe blood (Arseni et al., 1987). Bactere-mia is often, but not necessarily, con-firmed (Muytjens et al., 1983).

The severe consequences of infectionin some cases may be linked to the pro-duction of enterotoxin by E. sakazakii.More than 20% of the 18 tested strainsproduced enterotoxin. High levels (108

cfu) of each strain were lethal to micewhen administered by intraperitonealinjection. Peroral administration of thesame levels were only lethal for twostrains (Pagotto et al., 2003).

When infection does not result indeath, the affected infant may have per-manent neurological or developmentaldeficiencies. In one case, mortality wasaverted by months of antibiotic treat-ment, but the patient was mentally re-tarded and paralyzed by age two (Bier-ing et al., 1989). In another study ofeight infected infants, the two survivorswere both retarded, suffered from hy-drocephalus, and eventually died(Muytjens et al., 1983). However, full re-covery is also possible (Simmons et al.,1989).

Infants may be colonized with E.sakazakii without developing symptoms(Arseni et al., 1987; FDA/CFSAN, 2002).Although colonization and fecal car-riage may last 8–18 weeks, secondary

quantities will facilitate rapid cooling.Ideally, foods should be cooled to below15°C (59°F) within two to three hr aftercooking. Regarding preparation of riceand fried rice, the following recommen-dations have been made: (1) preparequantities of rice as needed; (2) keepprepared rice hot (55–63°C, 131–146°F);(3) cool cooked rice quickly; and (4) re-heat cooked rice thoroughly before serv-ing (Bryan et al., 1981; Gilbert et al.,1974; Moreland, 1976; PHLS, 1976; Slyand Ross, 1982).

B. cereus spores in foods can be de-stroyed by retorting (pressure cooking)or applying an equivalent thermal pro-cess or by irradiation. However, suchtreatments may be too severe and,hence, unacceptable for many foods.

Enterobacter sakazakiiJennifer C. McEntire and Francis F.Busta. Reviewed by R. BruceTompkin

Advances in medicine have vastly im-proved the mortality rate for infants.Unfortunately, this population is partic-ularly susceptible to often-fatal infectionby Enterobacter sakazakii, introducedthrough reconstituted powdered infantformula.

Significance as a PathogenAs of 1989, 20 cases of neonatal in-

fection by E. sakazakii were reportedworldwide (Biering et al., 1989). By2003, 48 neonatal cases were recognized(FDA/CFSAN, 2003b). Within the lastseveral years, awareness of the problemassociated with this pathogen has in-creased; as a result, research and reportsof E. sakazakii infection in infants havealso risen.

In 1980, yellow-pigmented Entero-bacter cloacae was recognized as its ownspecies—E. sakazakii. Although infec-tion by this microorganism has been re-ported since the 1960s, the pathogenwas not recognized as a cause of food-borne illness because the ecology of themicroorganism was unknown.

Infection by E. sakazakii is an ex-tremely rare event. Six of the 58 report-ed cases of E. sakazakii infection world-wide involved individuals more thanfour years of age, and the median agewas 74 (FDA/CFSAN, 2003b). The vastmajority (83%) of cases have been re-ported in infants less than one year ofage, where the fatality rate ranged from


transfer is not known to occur (Arseniet al., 1987; Block et al., 2002).

Association with foodsAlthough powdered infant formula

was postulated as the vehicle for E. saka-zakii infection as early as 1983(Muytjens et al., 1983), the first conclu-sive evidence linking the infectiousmicroorganism with the product was in1989 (Biering et al., 1989). After severalcases of infant infection in Iceland, testsof the powdered milk showed that themicroorganism could be isolated fromall lot numbers examined. However, nei-ther every individual sample nor everypackage from a lot showed E. sakazakiicontamination after enrichment, indi-cating that the microorganism waspresent at low levels. Plasmid analysisand antibiograms of 22 out of 23 strainsisolated from the formula were identicalto the strains that caused illness (Bieringet al., 1989). These techniques, as well aschromosomal restriction endonucleaseanalysis, ribotyping, arbitrarily primedPCR, and multilocus enzyme electro-phoresis, have been used to conclusivelylink outbreak strains with strains isolat-ed from formula in several other out-breaks (Clark et al., 1990; Simmons etal., 1989; van Acker et al., 2001).

After a fatal case of E. sakazakii infec-tion was confirmed in a premature in-fant in Tennessee, 49 other infants in thehospital were screened for infection orcolonization over a 10-day period. Themicroorganism colonized in seven as-ymptomatic infants and caused eitherconfirmed or suspected infection inthree others. Pulsed-field gel electro-phoresis confirmed that the strain of E.sakazakii isolated from the cerebral spi-nal fluid of an infected infant was iden-tical to the strain cultured from openedand unopened packages of infant for-mula (FDA/CFSAN, 2002). As a result,the implicated lot was recalled. A sepa-rate, unrelated recall of powdered infantformula produced by a different manu-facturer was conducted in 2003, againdue to the presence of E. sakazakii.

A 2002 FDA field survey collected 22samples of finished infant formula fromeight different plants. Five samples(22.7%) were positive for E. sakazakii atthe lowest detectible level, 0.36 MPN/100 g. The positive samples includedfour out of 14 formulas for full-term in-fants and one of four made specificallyfor pre-term infants. The study showedthat the percent of positive samples was

similar regardless of whether the formu-la was manufactured through wet-mix-ing, spray drying, or dry blending, or ifthe formula was made with milk or soy(FDA/CFSAN, 2004). A survey of pow-dered infant formula from 35 countriesshowed that more than 14% of sampleswere positive for E. sakazakii. This in-cluded two of 12 samples from pow-dered infant formula purchased in theUnited States. The levels of E. sakazakiiwere low—0.36 and 0.92 cfu/100 g(Muytjens et al., 1988). Levels of thepathogen were less than 1 cfu/100 g in17 of 20 positive samples, and all tested

samples complied with Codex Alimen-tarius recommendations for coliformcounts. The Codex standard requiresfour of five samples to have no positivetubes for coliforms in a three-tubeMPN; the fifth sample may contain upto 20 cfu/g (FAO, 1994). The batch ofpowdered infant formula implicated ina Belgian outbreak also met the specifi-cations of Codex Alimentarius, but notthe more stringent Belgian law whichrequires all samples to have less than 1cfu/g. Levels of E. sakazakii were so lowthat the microorganism was not initiallydetected in the dry powder, and its con-tinued use resulted in another case (vanAcker et al., 2001).

Control measuresAlthough E. sakazakii contamination

of powdered infant formula occurs atvery low levels, generally less than 1 cfu/100 g, some infants with the associatedrisk factors are susceptible to infection.The current control measure used byhospitals to prevent infection is the use

of commercially sterile liquid formula inlieu of powdered infant formula for at-risk infants. The FDA/CFSAN AdvisoryCommittee subcommittee (2003a)charged with determining if there was alink between E. sakazakii infection andpowdered infant formula reiterated thisrecommendation.

To decrease the risk of infant infec-tion by E. sakazakii, FDA prepared guid-ance for the preparation and use ofpowdered infant formula in neonatalunits (FDA/CFSAN, 2002). Time andtemperature control are key. Initially,guidance proposed using boiling waterto reconstitute the powder and destroythe microorganism, but essential nutri-ents could potentially be destroyed aswell. The thermotolerance of themicroorganism seems to be strain-de-pendent with the more tolerant strainshaving D

58°C values up to 600 sec (Edel-

son-Mammel and Buchanan, 2004).While pasteurization should be suffi-cient to kill the pathogen, the microor-ganism may be more heat resistant thanother Enterobacteriaceae (Breeuwer etal., 2003; Nazarowec-White and Farber,1997). Edelson-Mammel and Buchanan(2004) showed that preparing formulawith hot, but not boiling, water (i.e.,70°C) is sufficient to reduce E. sakazakiiby more than 3.9 logs within about 15sec. Although changes in preparationand storage of formula may help pre-vent infections, guidance and recom-mendations do not address reducingcontamination at the production level.

The survival of E. sakazakii in pow-dered infant formula may be partiallydue to the ability of the microorganismto survive desiccation. Breeuwer et al.(2003) speculated that this is related totrehalose accumulation in stationaryphase cells. Although the microorgan-ism presumably does not survive pas-teurization, it may be able to survive theconditions of powdered milk if intro-duced during or after drying. This isconsistent with Muytjens et al. (1988),who speculated that the low levels of thepathogen indicate that E. sakazakii is in-troduced through post-process contam-ination.

An understanding of the environ-mental reservoir(s) would facilitate con-trol of the pathogen. Until recently, thiswas unknown. Insects recently havebeen suggested as carriers of E. sakaza-kii. For example, Mexican fruit flies wereshown to harbor the microorganism(Kuzina et al., 2001). Hamilton et al.

Although E. sakazakiicontamination of pow-dered infant formulaoccurs at very low levels,generally less than 1 cfu/100 g, some infants withthe associated risk fac-tors are susceptible toinfection.



(2003) reported the isolation of E. saka-zakii from the larvae of Stomoxys calci-trans, commonly known as the stable fly.Rats also have been identified as asource of the pathogen (Gakuya et al.,2001). These reports stress the impor-tance of appropriate pest control as ameans to reduce the presence of E. saka-zakii in production facilities. The pres-ence of the microorganism in five of 16homes and eight of nine manufacturingfacilities producing a variety of driedfoods suggests this opportunistic patho-gen is more ubiquitous than had beenpreviously assumed, making controlmore difficult (Kandhai et al., 2004).

The FDA/CFSAN Advisory Commit-tee subcommittee (2003a) encouragedmicrobiological sampling and testingfor E. sakazakii in powdered infant for-mula. Biochemical assays can be used toidentify E. sakazakii and distinguish itfrom related Enterobacteriaceae, and theFDA is developing a real time PCR-based system for rapid detection (Seo etal., 2003). Dupont Qualicon has alreadyincluded E. sakazakii in its BAX® PCR-based detection system (Dupont, 2003).The committee also recommended thatmanufacturers formulate products tomaximize microbiological safety, anddesign and operate plants under hygien-ic conditions. Currently, FDA does nothave GMPs for powdered infant formu-la. The committee noted, however, thatthese interventions can not completelyeliminate the risk of E. sakazakii infec-tion to infants fed reconstituted pow-dered formula.

The risk reduction strategies suggest-ed by a joint FAO/WHO committeemade similar recommendations, andconducted a risk assessment to show theeffects of possible interventions (WHO,2004). The group recommended thatCodex develop microbiological specifi-cations for E. sakazakii in powdered in-fant formula. An IFT authoritative re-port on performance standards outlinesthe questions that would need to be an-swered in order to develop such specifi-cations (IFT, 2004).

ConclusionWhen the first edition of this Scien-

tific Status Summary was published in1988, E. coli O157:H7 and L. monocyto-genes were emerging pathogens thatwere not yet “household names.”Campylobacteriosis was just being rec-ognized as the cause of illnesses previ-ously identified as salmonellosis, and al-

though one report isolated E. sakazakiifrom powdered milk, the cause of infec-tion in neonates was largely unknown.Strides have since been made to im-prove food safety, and fortunately, someof the pathogens discussed are currentlyrecognized as minimal contributors tofoodborne illness. The emergence ofnew pathogens continues, not unex-pectedly, as the cause of foodborne ill-ness is unknown in 81% of cases (Meadet al., 1999). This Scientific Status Sum-mary has focused on the bacteria of sig-nificance in foodborne disease. Thepathogenic (disease-causing) bacteriadiscussed in this summary cause mostof the known bacterial foodborne ill-ness in North America. Certain charac-teristics of these bacteria have been wellknown for decades, while others haveappeared more recently in somewhatunexpected situations.

The principal control measures forprevention of foodborne diseases con-tinue to be use of adequate time-tem-perature heat treatments or cookingprocedures; avoidance of cross-contam-ination of cooked or RTE foods byutensils, equipment, or cutting surfacesthat are not properly cleaned and disin-fected after contact with fresh, un-cooked raw foods; avoidance of con-sumption of undercooked or contami-nated raw foods; and avoidance of con-tamination of raw foods during han-dling by infected food handlers.

The majority of serious foodbornedisease outbreaks occur as the result ofimproper attention to detail in foodser-vice establishments and in the home.The need for education of all food han-dlers, including consumers, is readilyapparent.

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The Society for Food Science and Technology

The organizing editor of the original Scientific Status Summary, published in 1988, was James L.Oblinger, North Carolina State University, Raleigh. The principal authors of the original document were:Michael P. Doyle, Center for Food Safety, University of Georgia, Athens; Russell S. Flowers, SillikerLaboratories, Chicago Heights, Ill.; Ronald G. Labbe, Dept. of Food Science and Nutrition, University ofMassachusetts, Amherst; Joseph Lovett, formerly of Div. of Microbiology, Food and Drug Administration,Cincinnati, Ohio; Robert M. Twedt, formerly of Div. of Microbiology, Food and Drug Administration,Cincinnati, Ohio; Joseph M. Madden, Neogen, Lansing, Mich.; Rosetta L. Newsome, Office of Science,Communications, and Government Relations, Institute of Food Technologists, Chicago, Ill.; Merle D.Pierson, U.S. Dept. of Agriculture, Washington, D.C.; and N. Rukma Reddy, National Center for Food Safetyand Technology, Summit-Argo, Ill.

The document was updated by Jennifer McEntire, Office of Science, Communications, and GovernmentRelations, Institute of Food Technologists. The original authors reviewed the updated sections, with theexception of Staphylococcus aureus, reviewed by Cynthia M. Stewart, Food Microbiology Group, FoodScience Australia, CSIRO, Sydney, and Listeria monocytogenes, reviewed by Stephanie Doores, Dept. ofFood Science, The Pennsylvania State University. The new inclusion on Enterobacter sakazakii wasauthored by McEntire and Francis F. Busta, Department of Food Science, University of Minnesota, andwas reviewed by R. Bruce Tompkin.

This and other Scientific Status Summaries are prepared by the Institute of Food Technologists as onesource of accurate and objective scientific information suitable for many different audiences, includingIFT members. The Science Reports and Emerging Issues Committee of IFT oversees timely publication ofScientific Status Summaries, which are rigorously peer-reviewed by individuals with specific expertise inthe subject.

The Scientific Status Summaries may be reprinted without permission, provided that suitable credit isgiven.

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