OR I G I N A L A R T I C L E

Survival of male-specific coliphage (MS2) as a surrogate forenteric viruses in the production process of traditional icecream

Sheila Ghadirzad | Masoud Yavarmanesh | Mohammad B. Habibi Najafi

Department of Food Science and

Technology, Faculty of Agriculture, Ferdowsi

University of Mashhad, Mashhad, Iran

Correspondence

Masoud Yavarmanesh, Department of Food

Science and Technology, Faculty of

Agriculture, Ferdowsi University of

Mashhad, Mashhad, Iran.

Email: yavarmanesh@um.ac.ir

AbstractThe objective of this study was to investigate the survival of coliphage MS2 as a surrogate for

enteric viruses under different process conditions in the production of traditional ice cream. The

results showed that heat treatment (73 8C, 10 min) decreased the recovery of coliphage MS2 to

1.33, 1.11, and 0.97 log10 (pfu/ml) in nonfrozen ice cream samples with 15%, 10%, and 5% fat,

respectively. Also the homogenization process (22,000 rpm in 1 min) decreased the recovery of

coliphage MS2 in nonfrozen ice cream samples with 15%, 10%, and 5% fat, to 2.33, 2.20, and 2.10

log10 (pfu/ml), respectively. All data in frozen ice cream samples was less than that of nonfrozen.

Pasteurization along with homogenization decreased the recovery of MS2 coliphage to 0.85 and

0.66 (pfu/ml) in nonfrozen and frozen ice cream samples, respectively. However, these processes

cannot totally eliminate all of the MS2 coliphages. During the storage time (6 weeks), ice cream

samples showed the lowest recovery of coliphage MS2 in the sixth week, and highest recovery of

coliphage MS2 on the first day. It was then concluded that pasteurization along with homogeniza-

tion has the most marked effect on the elimination of coliphage MS2 in frozen ice cream.

Practical applicationsEnteric viruses have been reported among outbreaks in which a pasteurized dairy product, such as

pasteurized milk, yogurt, and cheese, was involved. Since enteric viruses cause acute diseases and

are of major public health concern, together with the fact that overall global dairy production and

consumption have increased, a more comprehensive analysis of dairy products with a risk of con-

tamination with enteric viruses seems essential. Ice cream is a dairy product for which there is

considerable demand in the dairy market. However, there has been a paucity of research on the

survival of enteric viruses in ice cream. Therefore, we performed the current study to understand

the effects of different treatments on the survival of enteric viruses in traditional ice-cream.

1 | INTRODUCTION

Foodborne viruses are increasingly attracting a worldwide concern

about food safety as one of the most important causes of gastroenteri-

tis outbreaks. In contrast to most microbial agents, viruses are not only

unable to grow in food, but they also leave no visible sign of viral con-

tamination in foodstuffs. Therefore, the organoleptic characteristics of

food will not change because of such contamination (Zuber, Butot, &

Baert, 2013). Furthermore, only a few virus particles, apparently 10–

100 infectious viral particles are needed to produce an illness in most

cases which are much lower amounts compared to infected persons

who can shed virus via stool and vomitus (Teunis et al., 2008).

Some modes of virus transmission include the fecal–oral route,

direct person-to-person transmission, or consumption of contaminated

water and food (Bitler, Matthews, Dickey, Eisenberg, & Leon, 2013;

Kotwal & Cannon, 2014). Besides, food plays an important role in the

transmission of enteric viruses. For example, it is estimated that about

40% of norovirus infections are caused by the consumption of conta-

minated foodstuffs (Koopman & Duizer, 2004). Various food products

could cause HAV (Hepatitis A Virus) transmission such as shellfish,

salads, sandwiches, vegetables, fruit, reconstituted frozen orange juice,

ice cream, cheese, rice pudding, iced cake, custard, milk, bread, cookies,

and other raw or undercooked food (Cliver, 1997). Also, enteric viruses

could be transmitted through raw milk (Mortazavi, Habibi Najafi,

J Food Saf. 2018;e12450.https://doi.org/10.1111/jfs.12450

wileyonlinelibrary.com/journal/jfs VC 2018Wiley Periodicals, Inc. | 1 of 8

Received: 13 September 2017 | Revised: 27 December 2017 | Accepted: 1 January 2018

DOI: 10.1111/jfs.12450

Yavarmanesh, & Barouei, 2008; Raska, Helcl, Jezek, Kabelka, & Litiv,

1966) and dairy products such as pasteurized milk, cheese, and yoghurt

(Tiron, 1992). When foodstuffs are contaminated by foodborne viruses,

their survival depends on various parameters, such as the stability of

the virus, the method of food processing, and environmental conditions

(Koopmans, Bonsdorff, Vinj�e, Medici, & Monroe, 2002).

Generally, the composition of food matrices (especially pH, fat, and

sugar content) and the processes which are used for producing food-

stuffs influence virus survival (Zuber et al., 2013). Different food compo-

nents may provide protective effects on virus particles (Grove et al.,

2006). Also, the role of different milk components in the recovery of

viruses from raw milk has been reported (Yavarmanesh et al., 2010). For

example, inactivation of heat-treated HAV occurs at a higher tempera-

ture in products with a higher fat content (e.g., cream) compared to

products with lower fat (e.g., skim milk). Moreover, it has been indicated

that pasteurization temperatures are not sufficient to inactivate HAV in

dairy products (Bidawid, Farber, Sattar, & Hayward, 2000). Additionally,

the protective effect of milk in heat-treated poliovirus has been related

to milk fat, lactose, or milk proteins (Strazynski, Kramer, & Becker,

2002). Also, the heat resistance and survival of HAV in different compo-

sitions of food matrices such as shellfish, fruit-based and dairy products

have been described (Deboosere et al., 2010). Surrogates that are used

for pathogenic foodborne enteric viruses include HAV, feline calicivirus

(FCV), murine norovirus (MNV-1), bacteriophage MS2, Tulane virus,

porcine sapovirus, and poliovirus (Bozkurt, D’souza, & Davidson, 2015).

FCV was the first animal virus used as a surrogate for human norovi-

ruses but because of its high sensitivity to low pH (2.0 to 4.0), it may

not efficiently mimic the survival of human noroviruses in the environ-

ment or food. Despite the fact that MNV-1 causes an illness in mice

that is different from the human illness (Cannon et al., 2006), they are

similar to human noroviruses from immunological, biochemical, genetic,

and molecular points of view (Wobus, Thackray & Virgin, 2006).

However, the ideal surrogate should be similar to the intended

virus in structure and size as well as in survival and resistance charac-

teristics, cultivable in the laboratory, rather resistant to treatments,

nonpathogenic, and transmittable by the fecal-oral route (Busta et al.,

2003). Another potential human enteric virus surrogate is the bacterio-

phage MS2. It has been reported that bacteriophage MS2 survives in

fresh products and on environmental surfaces for prolonged periods.

Therefore, it was concluded that bacteriophage MS2 is an ideal surro-

gate for human norovirus and HAV (Dawson, Paish, Staffell, Seymour,

& Appleton, 2005).

The aim of this study was to evaluate the survival of bacteriophage

MS2 as an appropriate surrogate for enteric viruses in the process of

traditional ice cream production.

2 | MATERIALS AND METHODS

2.1 | Preparation of frozen host bacteria stock culture

(Escherichia coliFamp)

The host bacterial strain which is Escherichia coliFamp (ATCC#700891)

were obtained from the American Type Culture Collection was

prepared according to the EPA standard method No. 1601 (EPA,

2001). Frozen stock was streaked onto 1.5% Trypticase soy agar (TSA)

plates including appropriate streptomycin and ampicillin antibiotics to

obtain pure colonies of the bacterial host. All plates were then incu-

bated for 24 hr (overnight) at 36 61.0 8C. Subsequently, a pure colony

was inoculated into tryptic soy broth (TSB) containing appropriate

streptomycin and ampicillin antibiotics and grown to log phase. The

mixture of sterile glycerol and TSB with host bacteria propagated in log

phase in a ratio of 1:4 was kept frozen at270 8C.

2.2 | Preparation of overnight host bacteria stock

culture (Escherichia coliFamp)

Twenty-five milliliters of TSB containing streptomycin and ampicillin

was distributed into a sterile 125-ml flask. A loopful of E. coliFamp from

the frozen stock culture was then transferred to each flask and incu-

bated at 36 8C 61.0 8C while being shaken at 100 rpm for 18 to 20 hr

(overnight). Finally it was stored at 4 8C 61.0 8C to be used on the

same day (EPA, 2001).

2.3 | Preparation of log-phase host bacteria stock

culture

E. coliFamp host bacteria stock culture overnight activated of 0.1 to

1.0 ml was added to a 125 ml flask which contained 25 ml of TSB with

streptomycin and ampicillin. Then the flask was incubated at 36 8C

61.0 8C while being shaken at 100 to 150 rpm for approximately 4 hr

or when cultures were obviously turbid (cloudy), indicating log-phase

growth. For measuring absorption at 520 nm, 1 ml of culture from the

flask was removed aseptically and transferred into a cuvette. An

absorbance reading between 0.1 and 0.5 optical density (OD) units is a

sign of log-phase growth. Otherwise, cultures would be returned to a

shaker incubator until the proper OD was reached (EPA, 2001).

2.4 | Preparation of male-specific coliphage

For the preparation of coliphage MS2 (ATCC#15597-B1) were

obtained from the American Type Culture Collection, distilled water

was added to the vial containing coliphage MS2 stock. After that,

30 ml of TSB with Escherichia coliFamp was incubated at 36/5 8C 62 8C.

Besides, 1 ml of coliphage MS2 was added to culture media and further

incubated at 36/5 8C 6 2 �C for 4 hr. Finally, it was filtered and stored

in a sterile tube. The tube was labeled with source, date, and initials,

and stored at 4 8C6 1 �C (EPA, 2001).

2.5 | Preparation of MS2 stock dilutions

MS2 coliphage stock was diluted in buffer phosphate solution (pH 7.2)

to acquire the target concentrations 102, 104, and 106 pfu/ml. After

that, double agar layer (DAL) method, according to US-EPA Method

1601, was used to determine the mentioned concentrations (EPA,

2001).

2 of 8 | GHADIRZAD ET AL.

2.6 | Production of traditional ice cream

Raw milk was obtained from Ferdowsi University’s Dairy Farm. Pasteur-

ized and homogenized cream (30% milk fat) was obtained from Pegah

Dairy Industry Co, Mashhad, Iran. Sugar, Salep, Rosewater and Saffron

were purchased from a local confectionery market. Ice cream was pre-

pared according to a traditional recipe for Persian ice cream which con-

tains 77/7% milk and cream, 19% sugar, 3% rose water, and saffron, and

0.3% salep (Shaviklo, Thorkelsson, Sveinsdottir, & Rafipour, 2011).

According to the Pearson square, raw milk and cream were mixed

to acquire samples with 5, 10, and 15% fat. Then, the appropriate

amount of coliphage MS2 (102, 104, and 106 pfu/ml) was aseptically

added to the samples (fat: 5, 10, and 15%). After adding sugar, the

mixtures were heated to 40 8C before adding the other ingredients,

that is, saffron, stabilizer, and rose water. For the production of pas-

teurized and unpasteurized, also homogenized and nonhomogenized

traditional ice cream, the mixes were divided into four parts. The

processes of pasteurization and homogenization were done at 73 8C

for 10 min (Shaviklo et al., 2011) and at 22,000 rpm for 1 min, respec-

tively (Ika Homogenizer T-25 basic Ultra Turrax, Germany) (Bahram

Parvar, Mazaheri Tehrani, & Rayavi, 2013). Afterwards, the mixed

samples were cooled to 4 8C and stored for 4 hr for ageing. After the

end of the aging period, recovery of coliphage MS2 and pH changes

(pH meter model 691; Metrohm, Switzerland) in all mixed samples

were measured (as nonfrozen samples). After aging, the mix was fed

to an ice cream maker (Feller ice cream maker, Model IC Feller Tech-

nologic GmbH, Germany) and blended for 25–30 min. The ice cream

was packaged in polyethylene cups. Each cup, with a lid, was then put

into a plastic zip lock bag and kept frozen in a home freezer at

218 8C. Recovery of coliphage MS2 and pH changes were measured

in frozen and nonfrozen samples and during freezing time. All analy-

ses were performed after 1 day and at three week intervals during

the storage time (as frozen samples).

Weighing the ingredients+

Mixing raw milk and cream+

Adding coliphage MS2+

Adding sugar+

Heating (40 8C)+

Blending the ice cream mixture

(saffron, Salep and rose water)+

Dividing ice cream mixture into four parts+

Pasteurized (73 8C for 10min) and homogenized

(22000 rpm for 1 min) / unpasteurized and

Homogenized/ Pasteurized and Non- Homogenized/

unpasteurized and Non-Homogenized+

Cooling (4 8C)+

Ageing (4h)

+Measuring recovery of coliphage MS2 and pH

(as non-frozen samples)+

Freezing (batch ice-cream maker)+

Packaging (polyethylene cup)+

Storing at 218 8C

3 | STATISTICAL ANALYSIS

First, All data obtained was converted to logarithmic then a com-

pletely randomized factorial design was used for data analysis. Sta-

tistical analysis was conducted using Minitab version 14 (Minitab

Inc., State College, PA), and all figures were prepared using Slide

write software (plus. 2.0). Analysis of variance and LSD test were

used where applicable to determine statistically significant differen-

ces at p< .5.

4 | RESULTS

4.1 | Effect of pasteurization on the recovery of

coliphage MS2 in frozen and nonfrozen ice cream

Thermal processing (pasteurization at 73 8C, 10 min) of ice cream mixes

caused a significant decline in the recovery of coliphage MS2 (p< .05).

Also, the recovery of coliphage MS2 in frozen samples was less than in

nonfrozen ice cream (Table 1).

4.2 | Effect of homogenization on the recovery of

coliphage MS2 in frozen and nonfrozen ice cream

Results showed a significant decrease in the recovery of coliphage

MS2 in which the recovery of MS2 in frozen samples was less than in

nonfrozen ice cream (p< .05) (Table 2).

4.3 | Effect of storage time on the recovery of

coliphage MS2 and pH in frozen ice cream

The results demonstrated that by increasing storage time, the

recovery of MS2 decreased significantly. Also a significant

decrease in pH value occurred during storage time (p< .05)

(Table 3).

TABLE 1 Comparison of recovery for coliphage MS2 in frozen andnonfrozen ice cream

Ice cream mixes

Recovery of MS2in nonfrozensamples (pfu/ml) Std

Recovery ofMS2 in frozensamples (pfu/ml) Std

Pasteurized 1.15b .052 0.88b .046

Unpasteurized 3.70a .052 3.42a .046

p< .05.Different letters indicate significant differences.

GHADIRZAD ET AL. | 3 of 8

4.4 | Interaction effect of pasteurization andhomogenization on the recovery of coliphage MS2 in

frozen and nonfrozen ice cream

The interaction effect of pasteurization and homogenization on the recov-

ery of coliphage MS2 was significant (p< .05) (Table 4). It is estimated

that the highest coliophage recovery was achieved in nonfrozen ice cream

produced from unpasteurized and nonhomogenized mixes of ice cream.

4.5 | Interaction effect of pasteurization and fat onthe recovery of coliphage MS2 in frozen and

nonfrozen ice cream

The interaction effect of ice cream fat content and pasteurization was

significant (p< .05). The maximum amount of recovery was observed in

unpasteurized nonfrozen ice cream with 15% fat (Table 5).

4.6 | Interaction effect of homogenization and fatcontent on the recovery of coliphage MS2 in frozen

and nonfrozen ice cream

The investigation of the interaction effect of homogenization and fat

content on ice cream mixes showed a significant reduction in coliphage

recovery (p< .05). The highest recovery of coliphage MS2 was

observed in nonfrozen samples with 15% fat (Table 6).

4.7 | Interaction effect of spiking concentrations andfat content on recovery of coliphage MS2 in

nonfrozen ice cream

Based on the response surface curves in Figure 1a, by increasing

spiked concentrations and fat content, the recovery of coliphage

MS2 increased efficiently in pasteurized samples. Also, recovery in

unpasteurized samples increased more effectively (Figure 1b).

The recovery of coliphage MS2 decreased using the process of

homogenization (Figure 1c). However, it increased in nonhomogenized

samples (Figure 1d).

4.8 | Interaction effect of spiking concentrations and

fat content on the recovery of coliphage MS2 in

frozen ice cream

It is estimated that in the process of pasteurization, a rise in spik-

ing concentrations and fat content caused the recovery of coli-

phage MS2 to increase (Figure 2a), with an obvious increase in the

recovery of coliphage in unpasteurized samples was (Figure 2b).

Increase in spiking concentrations and fat content caused the

recovery of coliphage MS2 to increase in homogenized samples

(Figure 2c) and the increase was higher in nonhomogenized sam-

ples (Figure 2d).

4.9 | Interaction effect of storage time and fat

content on the recovery of coliphage MS2 in frozen

ice cream

An increase in storage time and a decrease in fat content in pasteurized

samples resulted in a decreasing trend in the recovery of coliphage

MS2. Frozen ice cream samples with 15% fat in the first week of stor-

age had the highest recovery of coliphage MS2 (Figure 3a,b). The

recovery of coliphage MS2 decreased by a reduction in fat content and

an increase in storage time for homogenized and nonhomogenized

samples (Figure 3c,d).

TABLE 2 Comparison of recovery for coliphage MS2 in frozen andnonfrozen ice cream

Ice cream mixes

Recovery of MS2in nonfrozensamples (pfu/ml) Std

Recovery of MS2in frozensamples (pfu/ml) Std

homogenized 2.22b .052 1.98b .046

Nonhomogenized 2.62a .052 2.32a .046

p< .05.Different letters indicate significant differences.

TABLE 3 Comparison of storage time and pH on recovery of coli-phage MS2 in frozen ice cream

Storage timeRecovery of MS2 infrozen samples (pfu/ml) Std pH

First day 2.25a .056 6.64a

Third week 2.17b .056 6.59b

Sixth week 2.04c .056 6.44c

p< .05.Different letters indicate significant differences.

TABLE 4 Comparison of interaction effect of pasteurization and homogenization on recovery of coliphage MS2 in frozen and nonfrozen icecream

Pasteurization HomogenizationRecovery of MS2 in nonfrozenice cream (pfu/ml)

Recovery of MS2 in frozenice cream (pfu/ml)

Unpasteurized Nonhomogenized 3.80a 3.55a

Unpasteurized Homogenized 3.60b 3.29b

Pasteurized Nonhomogenized 1.45c 1.11c

pasteurized Homogenized 0.85d 0.66d

p< .05.Different letters indicate significant differences.

4 of 8 | GHADIRZAD ET AL.

5 | DISCUSSION

The results showed that the recovery of coliphage MS2 can be affected

by various factors. One of the most important factors was thermal

processing (pasteurization) which is known to be very effective since

large volumes of liquid food samples can lead to insufficient heating in

the interior areas of the sample (Parry & Mortimer, 1984), Therefore,

one aspect of heat treatment is heat transfer rate. It was reported that

foodstuff size and the shape of the container affect heat transfer rate

(Chung, Wang, & Tang, 2007). Another aspect is the sort of thermal

systems which are used. It was suggested that using a water bath for

heating induces both conduction and convection heat transfer. Then,

heat transfer is quite equal in foodstuffs (Stringer, George, & Peck,

2000). The findings of some researchers asserted that Polioviruses in

milk, yogurt, and water were inactivated by long-time pasteurization,

high-temperature heating, and short-time pasteurization for 30 s,

although all of the polioviruses were not inactivated by short-time pas-

teurization for 15 s (Strazynski et al., 2002). Also, protective effects of

some components such as protein and fat on foodborne enteric viruses

during thermal processing were mentioned (Bidawid et al., 2000; Croci,

Suffredini, Di Pasquale, & Cozzi, 2012; Millard, Appleton, & Parry,

TABLE 5 Comparison of interaction effect of pasteurization and fat% on recovery of coliphage MS2 in frozen and nonfrozen ice cream

Pasteurization Fat %

Recovery of MS2in nonfrozen icecream (pfu/ml)

Recovery of MS2in frozen icecream (pfu/ml)

Unpasteurized 15 3.78a 3.51a

Unpasteurized 10 3.69b 3.42b

Unpasteurized 5 3.63c 3.33c

pasteurized 15 1.33d 1.02d

Pasteurized 10 1.11e 0.90e

Pasteurized 5 0.97f 0.74f

p< .05.Different letters indicate significant differences.

TABLE 6 Comparison of interaction effect of homogenization andfat % on recovery of coliphage MS2 in frozen and nonfrozen icecream

Homogenization Fat %

Recovery of MS2in nonfrozen icecream (pfu/ml)

Recovery of MS2in frozen icecream (pfu/ml)

Non homogenized 15 2.74a 2.42a

Non homogenized 10 2.63b 2.35b

Non homogenized 5 2.50c 2.21c

Homogenized 15 2.33d 2.11d

Homogenized 10 2.20e 1.97e

Homogenized 5 2.10f 1.85f

p< .05.Different letters indicate significant differences.

FIGURE 1 (a) Effect of spiking concentration and fat% on recovery of MS2 in pasteurized nonfrozen ice cream samples. (b) Effect ofspiking concentration and fat% on recovery of MS2 in unpasteurized nonfrozen ice cream samples. (c) Effect of spiking concentration andfat% on recovery of MS2 in homogenized nonfrozen ice cream samples. (d) Effect of spiking concentration and fat% on recovery of MS2 innonhomogenized nonfrozen ice cream samples

GHADIRZAD ET AL. | 5 of 8

FIGURE 2 (a) Effect of spiking concentration and fat% on recovery of MS2 in pasteurized frozen ice cream samples. (b) Effect of spikingconcentration and fat% on recovery of MS2 in unpasteurized frozen ice cream samples. (c) Effect of spiking concentration and fat% onrecovery of MS2 in homogenized frozen ice cream samples. (d) Effect of spiking concentration and fat% on recovery of MS2 innonhomogenized frozen ice cream samples

FIGURE 3 (a) Effect of storage time and fat% on recovery of MS2 in pasteurized frozen ice cream samples. (b) Effect of storage time and fat%on recovery of MS2 in unpasteurized frozen ice cream samples. (c) Effect of storage time and fat% on recovery of MS2 in homogenized frozenice cream samples. (d) Effect of storage time and fat% on recovery of MS2 in nonhomogenized frozen ice cream samples

6 of 8 | GHADIRZAD ET AL.

1987). Bidawid et al. showed the protective effect of fat on heat resist-

ance of HAV in milk, mainly in high concentrations. It was explained

that the presence of fats and proteins can lead to viral aggregation and

protection of cell receptors (Croci et al., 2012). Additionally, high fat

content increased heat stability of enteroviruses in ice-cream and ice-

cream products. Therefore, enteroviruses can survive during the pro-

cess of pasteurization at 71.7 8C for 30 s (Cliver & Salo, 1978). In

another study, sugar played a protective effect on the heat stability of

hepatitis A virus (HAV) (Deboosere, Legeay, Caudrelier, & Lange,

2004). Also, it was reported that heat stability of porcine parvovirus,

human parvovirus B19, HAV, and polioviruses during pasteurization at

60 8C can increase with high concentrations of sucrose (Ng & Dobkin,

1985; Nissen, Konig, Feinstone, & Pauli, 1996; Yunoki et al., 2003). It

can be assumed that a decrease in the water activity (aw) of Phosphate-

buffered saline (PBS) and a reduction in the solvability of the virus could

occur due to high sucrose concentrations. Therefore, virus aggregation

could increase heat stability. Another statement can be noted that the

high sugar content of the buffer causes the residual water in the viral

capsid to remove. The structural changes in capsid proteins afford a

higher resistance to heat (Jarke et al., 2013). Moreover, water activity

(aw) is known as an important factor on the behavior of coliphage MS2.

Sensitivity to thermal treatment in vegetables and herbs may be due to

high water activity (Bertrand et al., 2012). Also, it is estimated that lower

water activity in mashed strawberry with a high sugar content may lead

to higher heat resistance of HAV rather than in spinach (Bozkurt et al.,

2015). In traditional ice cream using salep as a carbohydrate and stabi-

lizer led to a decrease in water activity and it may indicate a protective

effect on the thermal resistance of coliphage MS2. Also, high contents

of fat and sugar in traditional ice cream can be a further reason for ther-

mal resistance of coliphage MS2.

Following heat processing, another treatment such as homogeniza-

tion can be effective on viral inactivation. Homogenized samples can

lead to a more effective heat transfer because of creating a uniform

food matrix (Stringer et al., 2000).

Another investigation showed that using homogenization along

with adding lecithin causes the viruses from food particles to release

and exposes them directly during the process of homogenization

(Johnson, Ellender, & Tsai, 1984). Also, lecithin showed extra effects on

reducing FCV-F9 at homogenizing pressures (Horm, Harte, & D’souza,

2012). It was concluded that adding salep as a stabilizer along with the

process of homogenization may have a cumulative effect on the

inactivation of viruses.

A gradual decrease in pH occurs in the storage time of ice cream

(Murtaza, Meenud Din, Huma, Asim Shabbier, & Shahid, 2004). It was

explained that this decrease could be due to the transformation of lac-

tose into lactic acid by some kinds of bacteria during storage period

(Khan, 1989). Also, poliovirus (PV) survival in oysters under frozen con-

ditions decreased about 1-log10 after 4 to 12 weeks while stored at

217.5 8C (DiGirolamo, Liston, & Matches, 1970). It was assumed that

this phenomenon could be as a result of the freezing and thawing proc-

esses, rather than being under frozen storage. Also, the titer of norovi-

ruses (NOV) declined approximately 10% for each turn of freezing and

thawing (Richards, McLeod, & Le Guyader, 2010).

The reduction in pH value could decrease the stability of coliphage

MS2 and lead to a reduction in the coliphage recovery rate (Guan,

Schulze-Makuch, Schaffer, & Pillai, 2003; Jo�nczyk, Kłak, MieRdzybrodzki,

& G�orski, 2011). Thus, a decrease in pH value and round of freezing

and thawing can possibly be a main reason for the inactivation of coli-

phage MS2 during storage time.

6 | CONCLUSION

The results of this study revealed that pasteurization along with

homogenization is the most effective process on the inactivation of

coliphage MS2 in the production of traditional ice cream, largely (espe-

cially) in low fat ice cream, although all coliphage MS2 cannot be elimi-

nated. It can be related to the complex matrix of traditional ice cream.

With regard to using unpasteurized milk in the production of traditional

ice cream, it seems that the exerted thermal process on the mix of tra-

ditional ice cream will be necessary to ensure consumers’ public health,

especially children.

ORCID

Masoud Yavarmanesh http://orcid.org/0000-0002-4771-5359

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How to cite this article: Ghadirzad S, Yavarmanesh M, Habibi

Najafi MB. Survival of male-specific coliphage (MS2) as a surro-

gate for enteric viruses in the production process of traditional

ice cream. J Food Saf. 2018;e12450. https://doi.org/10.1111/

jfs.12450

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