Indicators of Bacterial Pathogens

IntroductionAll pathogenic microorganisms implicated in foodborne diseases are considered enteric pathogens, except S. aureus, B. cereus, C. botulinum. C. perfringens, and toxigenic molds.Enteric pathogens can survive and multiply or establish in the GIT of humans, food animals, and birds.A food contaminated directly or indirectly with fecal materials from these sources may theoretically contain one or more of these pathogens and thus can be potentially hazardous to consumers.

To ensure consumer safety, it is necessary to know that a food is either free of some enteric pathogens, such as Salmonella and E. coli 0157:H7.The procedures used to isolate and confirm a pathogen from a food involve several steps, take a relatively long time, and are costly. Instead, food samples are examined for a group or a species of bacteria that are of fecal origin, usually present in higher density than pathogens, but considered to be nonpathogenic.

Their presence is viewed as resulting from direct or indirect contamination of a food with fecal materials and indicates the possible presence of enteric pathogens in food. These bacterial groups or species are termed indicators of enteric pathogens.S. aureus, and clostridia are not considered as classical enteric pathogens because their presence in food is not normally considered to be of fecal origin and the indicators of enteric pathogens do not apply for them

Criteria For Ideal Indicators• The indicator should preferably contain a single

species or a few species with some common and identifiable biochemical characteristics.

• The indicator should be of enteric origin, that is, it should share the same habitat as the enteric pathogens.

* The indicator should be nonpathogenic so that its handling in the laboratory does not require safety precautions.

• The indicator should be present in the fecal matter in much higher numbers than the enteric pathogens so they can be easily detected,

• The indicator should be detected and identified within a short time.

• The indicator should have a growth and survival rate in a food as that of the enteric pathogen.

* The indicator should preferably be present when the pathogens are present in food; conversely, it should be absent when the enteric pathogens are absent.

It is apparent that no single bacterial group or species will be able to meet all the criteria of an ideal indicator. Several bacterial groups or species satisfy many of these criteria: Coliform Group A- coliforms Organisms and Sources: Escherichia, Enterobacter, Klebsiella, Citrobacter.

They are all Gram-negative, nonsporeforming rods, many are motile, facultative anaerobes, resistant to many surface active agents, ferment lactose to produce acid and gas within 48 h at 32 or 35C. Some species can grow at higher temperature(44.5C), whereas others can grow at 4-5C. All are able to grow in foods, all are killed by pasteurization.They can be present in feces of humans and warm-blooded animals and birds. Some can be present in the environment and contaminate food.

Occurrence and Significance in FoodColiforms are present in many raw foods and food ingredients of animal and plant origin.In some plant foods, they are present in very high numbers because of contamination from soil.Because they can grow in foods, some even at refrigerated temperature, a low initial number can reach a high level during storage. The occurrence of some coliforms of nonfecal origin and their ability to grow in many foods reduce the specificity of coliforms as an indicator.

In heat-processed foods, their specificity as an indicator is considered to be due to improper sanitation rather than fecal contamination.The presence of sublethally or stressed or injured cells can reduce the recovery in selective media.The presence of lysozyme(egg-based products) in a food, can further reduce the enumeration of stressed cells.

B- Fecal coliforms Organisms and Source: Fecal coliform bacteria also constitute a group of bacteria and include those coliforms whose specificity as fecal contaminants is much higher than that of coliforms. This group includes mostly E. coli, along with some klebsiella and Enterobacter SPP..Nonfecal coliforms are eliminated by using a high incubation temperature(44.5-45C) for 24 h in selective broths containing lactose. Lactose fermentation, with the production of gas, is considered a presumptive test for fecal coliforms.

Occurrence and Significance in FoodSome fecal coliforms are present in raw foods of animal origin.They can be present in plant foods from contaminated soil and water.High numbers can be due to either gross contamination or growth from a low initial level, probably because of improper storage temperature.Their presence in heat-processed (pasteurized) foods is probably because of improper sanitation after heat treatment.

In heated products and ready to eat products, the presence of fecal coliforms, especially above a certain level, is viewed cautiously for possible Fecal contamination and presence of enteric pathogens.A food can be accepted or rejected based on the numbers present.Fecal coliforms are extensively used as an indicator in foods of marine origin(shellfish), and in water and waste water.

C- Escherichia coli Organisms and Sources: E. coli includes only the Escherichia spp. Of the coliform and fecal coliform groups. E. Coli strains conform to the general characteristics described for coliform groups.Biochemically they are differentiated from other coliforms by the indole production from tryptone, methyl red reduction due to acid production, Voges Proskauer reaction (production of acetyl-methyl carbinol from glucose), and citrate utilization as a carbon source(IMViC).

E. Coli Type I and type II give IMViC reaction pattern, respectively, of + + - - and - + - - The IMViC tests are conducted with an isolate obtained after testing a food sample for coliform group or fecal coliform group. Initially, E. coli types were used as indicators of fecal contamination and possible presence of enteric pathogens(in food), with the consideration that they are nonpathogenic and occur normally in GIT of humans, animals and birds. However, it is now known that some variants and strains of E. coli are pathogenic(e.g., E. coli 0157:H7). There are specific tests to identify E. coli pathogenic strains.

Occurrence and Significance in Food

E. Coli is present in the lower intestinal tract of humans and warm-blooded animals and birds.Its presence in raw foods is considered an indication of direct or indirect fecal contamination.Direct fecal contamination occurs during the processing of raw foods of animal origin and because of poor personal hygiene of food handlers.Indirect contamination can occur through sewage and polluted water.32

In heat-processed (pasteurized) foods, its presence is viewed with great concern.Its value as an indicator of fecal contamination and the possible presence of enteric pathogens is much greater than that of coliform and fecal coliform groups. As the time to complete IMViC is relatively long, some direct plating methods have been developed that give an indication of E. coli in a shorter time.There are some problems with E. coli as an indicator(dying at faster rate in dried, frozen, and low-pH products than some enteric pathogens).

Enterococcus GroupCharacteristics and Habitat The genus Enterococcus includes many species that were previously grouped as fecal streptococci and other streptococci.They are Gram-positive, nonsporeforming, nonmotile cocci or coccobacilli, catalase negative, and facultative anaerobes. They can grow between 10-45C, some species can grow at 50C.Some require B vitamins and amino acids for growth.

Some can survive pasteurization temperature.In general, they are more resistant than most coliforms to refrigeration, freezing, drying, low pH, NaCl and water.They are found in the intestinal tracts of humans, and warm and cold-blooded animals, birds and insects.They include Enterococcus faecalis, E. faecium, E. durans, E. gallinarum, E. avium, and E. hirae.Many have been found in vegetation and processing equipment. They are found in sewage and water, can survive longer than coliforms, they can grow in foods.

Occurrence and Significance in FoodEnterococcus can get in different foods through fecal contamination or through water, vegetation, or equipment and processing environments, and may not be of fecal origin.In this respect, its value as an indicator of fecal contamination and possible presence of enteric pathogens is questionable.Also, the ability of some strains to survive pasteurization temperature(being thermodurics) reduces their value as an indicator.

On the other hand their better survivability in dried, frozen, refrigerated, and low pH foods and water can make them favorable indicators.Currently, their presence in high numbers, especially in heat-processed(pasteurized) foods, can be used to indicate their possible presence in high numbers in raw materials and improper sanitation of the processing equipment and environment.Some strains are associated with gastroenteritis.They have been used to determine the sanitary quality of water in shellfish beds and are considered to be better as indicators than coliforms for shellfish.