PREVALENCE OF STAPHYLOCCUS AND AEROMONAS IN SOME SALTED DAIRY PRODUCTS

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PREVALENCE OF STAPHYLOCCUS AND AEROMONAS IN SOME SALTED DAIRY PRODUCTS

AHMED HASSAN SAAD ¹; EHAB MOHAMED SALAMA ¹; HANAN A. EL DAHSHAN ²

and NOHA TALAAT ASSAF ¹

¹ Faculty of Veterinary Medicine, Suez Canal University, Egypt

² Food Hygiene Department, Port-Said Branch, Animal Health Research Institute

Received: 18 November 2018;     Accepted: 9 December 2018

INTRODUCTION

Pickled white soft cheese (domiata cheese) and Mish are well-known as local types of cheese in Egypt. Handling of milk during cheese manufacture considered the main source of microbial contamination of cheese which affecting on cheese quality and render it unfits for human consumption (Yousef et al., 2001).

Staphylococcus spp. are considering one of halophilic bacteria which its presence in milk or milk products can cause a public health hazard, Staphylococcus aureus is responsible for food poisoning (ICMSF, 1986) S. aureus is consider as one of important food-borne pathogen. It is a versatile pathogen of humans and animals; it causes a wide variety of diseases ranged in severity from slight skin infection to more severe diseases such as pneumonia and septicemia (Lowy, 1998). Staph.aureus is capable of producing several enterotoxin cause food poisoning in human with varying degree and generally characterized by nausea,  diarrhea,  abdominal   cramps   and   emesis

Corresponding author: HANAN A. EL DAHSHAN

E-mail address: mhananamen@yahoo.com

Present address: Food Hygiene Department, Port-Said Branch, Animal Health Research Institute

(Normanno et al., 2005) and (Brightwell et al., 2006). In the last few decades' Staphylococcal food poisoning (SFP) has been reported as third cause of food-borne illnesses in the world (Zhang et al., 1998).

Aeromonas spp are considered one of the most important halophilic bacteria which are widely distributed in nature. Moreover it is considered as food borne pathogen (Palumbo et al., 1992). Aeromonas spp. is pathogens that cause food-borne gastroenteritis, extra intestinal symptoms such as septicemia, meningitis, endocarditis and osteomyelitis with a high mortality rate in immune-compromised person (Gold and Salit, 1993). The mechanisms by which Aeromonas spp. cause diarrhea has been known that they produce enterotoxins, certain enzymes, they are able to adhere cell membranes and invade them (Kirov       et al., 1994). The presence of such bacteria in milk products may be due to direct contact with contaminated sources in dairy farm environment, excretion from udder of an infected animal or during processing of cheese.  (Oliver et al., 2005).

MATERIALS AND METHODS

1. Collection of samples

One hundred and twenty random samples of pickled white soft cheese and Mish (60 of each) were collected from different markets in Port Said city, and then taken aseptically to the laboratory immediately, where it is subjected to chemical and microbiological examinations.

2. Preparation of samples (A.P.H.A. 2004)

250 gm. of each sample were transferred aseptically in a sterilized polyethylene sac and thoroughly mashed in a sterile blender before being emulsified in the diluent solution under aseptic condition then divided into two subsamples for chemical examination and bacteriological examination.

3. Chemical Examination

3.1. Determination of sodium chloride content (A.O.A.C, 2000)

3 grams of prepared sample were weighted in 200 ml Erlyn – Meyer flask. 25 ml of N/10 silver nitrate solution, 10 ml of halogen free nitric acid and 50 ml of distilled water were added and the mixture was boiled. 15 ml of 5% potassium permanganate solution in 5 ml portion were added during boiling (till the solution become yellowish and clear). Then the solution was allowed to cool then filtered into 200 ml volumetric flask, the filter paper was washed thoroughly with distilled water at 20ºc, and the filtrate was made up to standard volume. The excess of silver nitrate in 100 ml of the clear solution was titrated against 0.1 N potassium thiocyanate solution (9.71 g / liter) using 2 ml of saturated solution of iron alum as indicator. The salt content was calculated according to the following equation:

Na CL % = 2 (25-R) X0.00584 X100

Weight (3g)

4.Microbiological Examination

4.1. Determination of total Halophilic bacterial counts (A.P.H.A.2004)

Preparation of serial dilution:

10gm of each sample was transferred in sterile stomacher bag with 90ml synthetic sea water solution (3% NaCL forSlight or10%NaCL for moderate Halophilic counts). Apply in a stomacher lab-blender (2000 rpm) used to homogenize the specimen for 2 minutes to make a1:10 dilution (wt/vol) then decimal dilution of 10^-2, 10^-3, 10^-4, 10^-5and 10^-6  were prepared.

0.1 ml from each dilution was inoculated in duplicated plates containing standard plate count agar following by spread plate technique and the plates were incubated at 37^oC for 48hours, then the total slight and moderate halophiles were counted and expressed as CFU/g.

4.2. Identification of isolated Staphylococci organisms according to (FDA, 2001).

Suspected colonies of Staphylococcus organismswere examined morphologically, microscopically according to (Ryan and Ray 2004)

4.3 Biochemical examination: according to (FDA, 2001)

The biochemical tests were Coagulase test, Catalase test, DNase test, Acetoin production, Oxidase test and D-mannitol fermentation.

4.4. Isolation of Aeronomas spp.

10gm of each sample were homogenized with 90 ml alkaline peptone water (with 3% or 10% NaCL) for 2 min, and then incubated for 24hr at 30ºC (Villari et al., 2000). A loopful from alkaline peptone water was subsequently plated on the surface of starch ampicillin agar plate and incubated for 48 hr. at 30ºc. Typical yellow colonies of Aeromonas species were purified on tryptone soya agar then stained by Gram^'s stain (A.P.H.A., 2004)and confirmed on the basis of the following test: Oxidase test, resistance to vibriostatic agent o/129, esculin hydrolysis, sugar fermentation and gas production, indole production and voges- proskaur test.

4.5. Differentiation of common motile Aeromonas isolated.

From clinical specimens according to Carnhan et al. (1991) as modified by Joseph and Carnahan (1994). All bacterial isolates would be short G -ve bacilli, oxidase positive that ferment glucose.

RESULTS

Sodium chloride content

Table 1: Statistical analytical results of salt conc. % in Pickled domiati cheese samples (n=60) and Mish samples (n=60) comparison with Egyptian Standards 2005.

Determination of total Slight & moderate Halophilic counts

Table 2: Statistical analytical results of Total Slight and moderateHalophilic count in examined samples.

Detection of Staphylococcal organisms at 3% and 10% NaCL:

Table 3:  prevalence of Staphylococci isolated from examined samples.

4. Isolation of Aeronomas spp. at 3% and 10% Na CL

Table 4: Incidence of Aeromonas spp. in examined samples at 10% NaCl.

DISCUSSION

Sodium chloride content

The use of salting is one of the classical methods of food preservation. Salting is used to extend the shelf-life of foods throughout civilization. In this study, it is evident in table (1) that salt percentage of examined pickled domiati samples ranged from 2.90 % to 5.30 %. While in mish samples ranged from 4.20 % to 8.30% all results obtained were within the normal range of (Egyptian Standards, 2005). Nearly similar results were reported by Ceylan et al. (2003), Hassan and Afify, (2007), Nawar, (2007), EL-Ansary et al. (2011), El Bakry, (2012), Yasser, (2015) and EL-Refaay, (2016), while higher result were obtained by Mohamed, (2004), Patrick et al., (2004), Hayaloglu and Kirbag, (2007) and EL Zahar, (2010). While lower result were obtained by Riad, (1996) and Sadek, (2009).

Determination of total Slight & moderate Halophilic counts

Slight, moderate, and extreme halophiles as those bacteria that grow best in media containing 2 to 5%, 5 to 20% and 20 to 30% salt respectively. (Kushner 1978). in table (2) Samples examination refer that maximum value of slight halophilic count in Pickled domiati cheese samples and Mish samples were 1x 10⁶ and 2.6 x10⁶ respectively while minimum value were 1x10³ and 1.6 x10³ respectively where salt conc. was 3%. This results are nearly similar to Freitas et al. (1993), andSaad and Moawad (1999). While higher results are obtained by Riad (1996)and Omer et al. (2007).

The maximum value of moderate halophilic count in Pickled domiati cheese samples and Mish were 3.2 x 10⁵ and 9. 0x10⁵cfu/g. respectively, while minimum value was 1.0 x10² and 5.0 x10² cfu/g.  Respectively with mean value 0.76 x 10⁵+ 0.14 x10⁵ in pickled domiati cheese samples. And with mean value 2.09 x10⁵+ 3.6 x10⁴ in Mish samples (Where NaCL concentration is 10%). These results are nearly similar to El-prince (1994) and Riad (1996),while lower results are obtained by Omar et al. (2007).

Detection of Staphylococcal organisms at 3% and 10% NaCL:

The presence of Staphylococcus is an index of contamination from operators or workers. It has a potential significance to public health due to its ability to produce enterotoxin leading to food poisoning. The results in Table (3) show that incidence of Staphylococcus in Pickled domiati cheese was 85% at 3% NaCL and 71.6% at 10% NaCL. While in Mish samples was 93% at 3% NaCL and 83.3% at 10% NaCL. These results are nearly similar to those obtained by Abou El-Makarem, (2009). Higher results are obtained by Sheleby(2008) and Elshafey (2011).

Isolation of Aeronomas spp. at 3% and 10% Na CL:

Aeromonas species are widely distributed in the aquatic environment, including raw and processed drinking water (Holmes et al., 1996), and have been frequently isolated from various food products such as fish and shellfish, raw meat, vegetables and raw milk Palumbo, (1996). It is associated with travellers' diarrhea (Hänninen et al., 1995 and Yamada et al., 1997). In this study, Aeromonas species was not isolated from any examined samples either Mish or Pickled domiati cheese using 3% salt concentration while at 10% Na CL, as shown in table (4) that the incidence of Aeromonas spp. was 25% A.hydrophila, 46.4% A.caviae, 21.4% A .trota and 7.1% A. schubertti in Mish cheese samples while the incidence of Aeromonas spp. was 25.7% A.hydrophila, 40% A.caviae, 20% A. trota and 14.3% A. schubertti in Pickled domiati cheese samples. This results are nearly similar to those obtained by to Freitas et al. (1993), Khalil (1997), Effat et al. (2000), Ahmed     et al. (2014) Higher results are obtained byYasser (2008), Nazem et al. (2010) and Alhazmi (2015).

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