PHYSICOCHEMICAL AND BACTERIOLOGICAL PROPERTIES OF SOME ALGERIAN TRADITIONAL DAIRY PRODUCTS ‘KLILA AND JBEN’ MARKETED IN SOUTHWEST OF ALGERIA AND THEIR IMPACT ON CONSUMERS HEALTH

Elhassan Benyagoub 1,3* , Bettache Guessas 1 , Mohammed Ayat 2 & Brahim Sanebaoui 3 .  1. Faculty of natural and life sciences, Department of Biology, Ahmed Ben bella University of Oran 1 (31000), Oran, Algeria. 2. Quality control laboratory, Oran (31000), Oran, Algeria. 3. Faculty of natural and life sciences, Department of Biology, Tahri Mohammed University of Bechar (08000), Bechar-Algeria. ...................................................................................................................... Manuscript Info Abstract ......................... ........................................................................ Manuscript History


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However, the Algerian traditional cheese varieties have not been studied thoroughly and are always confined at the family level. Some of these varieties have a good quality and have attractive properties in terms of flavor and texture. Therefore it's important to have knowledge on changing microbiological and biochemical properties during manufacture, as well as the parameters of their processes to produce a cheese with higher hygienic and technological quality.
Practically, few studies focused on Algerian cheeses in general and in particular Klila and Jben products, and there is not much data on their biochemical and microbiological characteristics and their transformation techniques.
This study aims to characterize the physicochemical and bacteriological properties of the traditional cheeses Klila and Jben, and their impact on public health in southwest of Algeria.

Manufacturing process of traditional cheeses:-
In Algeria, Klila and Jben are the most popular traditional cheeses and their traditional manufacturing methods are still in use until today (Hamama, 1989a). Nevertheless, there is an increased demand of this type of cheese, due to its pleasant taste and nutritional properties.
Although similar cheeses to Klila and Jben all over the world such as Jameed in Middle East and the Chhana in India are well characterized and produced on an industrial scale in continuous processes using new procedures such as atomization and lyophilization (Shaker et al.,1999;Dharam and Narender, 2007;Mazahreh et al., 2008;AL omari et al., 2008).
On the one hand, the Klila is a fermented cheese produced empirically in several regions of Algeria. It is manufactured by a relatively moderate heating (55-75°C) from Lben until the latter is curdled (10-15min), the curd is drained spontaneously or pressed by a stone, the resulting cheese is consumed as fresh or after drying. Klila is used as an ingredient after rehydration in traditional cooking. On the other hand, the Jben is a fresh cheese known and popular on the Algerian territory. It is produced by coagulation of fresh milk, then draining and molding the curd, and therefore the resulting cheese is ready for consumption.
In Europe, cheese valorization, like all traditional products, took place through various stages, whose main were to improve the production technology and to identify the factors affecting quality, as well as regulations for the protection and enhancement (eg: Protected appellation of origin, Protected geographical indication, guaranteed traditional specialty, etc.), that tend to preserve their peculiarities.
Even for Algerians dairy products, we headed towards the idea that we need to know what factors characterize them, in order to improve production technology and eliminate the hygienic risks (Galina et al., 2011). Therefore, a research should be made to keep the traditional skills, but at the same time, using modern knowledge; to have the right balance between quality, safety and convenience.

Samples:-
The levies were performed at markets of Bechar City, Ain Sefra and Beni Abbes between the period from February and April 2016, with a total of thirty samples divided between 25 samples of Klila and 05 samples of Jben. The samples were placed in a cooler and transported directly to the laboratory for analysis, where they are held at pedagogical laboratory of Bechar University, whose purpose is to assess the hygienic quality of traditional dairy products marketed in southwest of Algeria.
The pH was determined by pH meter Starter 2C Lab, Shanghai China. The titratable acidity was determined by measurement of lactic acid with sodium hydroxide in the presence of phenolphthalein at 1% as a color indicator according to (NF V04-206, 1994; ISO 11869, 1997).
The statistical treatment consisted of determining the average and standard deviation.
Microbiological Analysis:-Sampling was performed in concordance with the Algerian standardized sampling methods (ISO 707, 1994). Microbiological analysis included after preparing decimal dilutions according to (ISO 8261, 2001), where the stock solution was prepared after homogenization using a Stomacher 400C LabBlenders, the detection and enumeration of bacteria indicators of fecal contamination or sanitary quality defects, and the search for suspected pathogenic bacteria, including; Enumeration of total aerobic mesophilic flora incubated at 30°C for 72hours according to (NF V 04-016, 1985) on Plate Count Agar (PCA) (Fluka, Spain); Detection and enumeration of total and fecal coliform incubated at 37 and 44°C respectively for 24 to 48hours according to (ISO 4832, 1978)  Presumptive pathogenic strains isolated from selective media; Baird Parker, SS and Hecktoen have been tested to be identify by several biochemical tests 'Gram stain, catalase test, coagulase test, tests of classic gallery and miniaturized API 20E'.
Enumeration of Petri dishes having microorganisms is based on the standard set by legislation (AFNOR, 1980).

Interpretation of Results:-
The criteria for microbiological quality employed were in accordance with the publication on dairy products, Algerian standard (NA 35, 1998)

Hazard analysis 'HACCP system':-
The implementation of HACCP system "Hazard analysis and critical control points" system is performed in several successive stages, at first, it is to apply the prerequisite programs "PRP", hazards identification at each stage of production, processing or preparation of the product, evaluate the corresponding risks and to determine the stage where it is possible to act effectively (Faye and Loiseau, 2000).
The quality management by the analysis of risks or potential hazards associated with a product or process (HACCP) must be applied to the entire chain from the cow to the consumer (Leyral and Vierling, 2007). Hazard identification was performed using the Ishikawa diagram 'Diagram cause-effect' detailing 5 causes to consider "5M Method" (Medium, Workforce, materials, methods and equipment) to maintain a hygienic environment throughout the production (Notermans et al., 1994;Blanc, 2007), whereas the risk by approaching Zurich Hazard Analysis which involves assessing qualitatively and semi-quantitatively risk assessment, severity and frequency (Codex Alimentarius, 2003).
At each of the identified potential risks corresponding corrective actions pertinent and control plans, associated with critical limits, monitoring procedures and recording.

Results:-Physicochemical analysis:-
The average results of physicochemical parameters of Klila and Jben samples namely pH, titratable acidity, dry matter and relative humidity are represented in the table below (Table 1). However the average values found in titratable acidity of the samples analyzed are 3,3 and 10,1g/l with values range from 5 to 20,3 and from 2,6 to 4,6 g/l for Klila and Jben respectively.
The average levels of total dry matter of the samples analyzed were high, whose value obtained is 95,16%, with variations going from 89,4 to 97,8% for Klila, whereas samples of Jben recognized an average of 56,18% with minimum values of 25,6; and 85,2% as maximum values.

Microbiological analysis:-
The average results of bacteriological parameters of Klila and Jben are presented in the table below (Tab. 2). Enumeration of total aerobic mesophilic flora revealed an average estimated 5,1x10 7 and 2,88x10 6 cfu/g for samples of Klila and Jben. These results indicate that these products of lactic fermentation were too loaded with aerobic germs (Fig.1). Enumeration results of total coliforms show that the levels of these contaminants was important that the analyzed samples contained means of 3,89 x10 5 and 1,4 x10 4 ufc/g for the product of Klila and Jben respectively. However, we reported the complete absence of these germs for eight samples namely k11, k12, k13, k19, k20 for Klila and J2, J3, J5 for Jben.
After identification of isolates suspected as Salmonella sp or Shigella sp, the analyzed samples appear free from Salmonella sp and Shigella sp for different samples of Klila and Jben, while the isolated strains were identified as Enterobacter sakazakii, Escherichia coli and Serratia odorifera.
The sulphite-reducing clostridia spores were detected in Klila and Jben samples except the samples k15, J1, J4 and J5. Table 3 provides an illustration of a commodity chain approach to hazard identification, for industrial and semiindustrial soft and pressed cheese subsidiaries. Table 3:-Control measuring in the process production of raw milk, soft and pressed (hard and semi-hard) cheese.

Hazard analysis 'HACCP system ':-
Steps Control measuring The raw material: -Receipt of local raw milk -Check and control the health program and maintenance of premises and separation zones, personal hygiene "Uniforms of clothing + body and hand hygiene" -Managing the storage conditions of raw material, raw milk must be refrigerated no later than 2 hours after milking at 4°Cmaximum. The results of titratable acidity indicate that 52% of Klila samples were in the range of mean values given by (Hamama, 1989b;Kbibou, 1987), which the values obtained were 9,9 and 11,16g/l respectively.
The samples analyzed of Jben were characterized by relatively low titratable acidity and a slightly acidic pH reflecting the absence of an active lactic fermentation, may be due to the inhibitory activity of plant extracts used for the coagulation of milk (Lahsaoui, 2009).
The total solids is generally more important in the Klila product given its dry nature,while for Jben sample results were close to those found by (Hamama and Bayi, 1991) which were located in the range of 29,4 and 45,6%.
This difference is a consequence of salting fresh cheese and drainage time. This richness of dry matter of Jben product gives it a relatively firm consistence. According to (Hamama and Bayi, 1991), two major constituents of the dry matter are the fats and proteins which have an average of 16,5 and 15,8% respectively.

Bacteriological analysis:-
It should be noted that only fresh cheese "Jben" which is cited in the Algerian regulation, and there is no bacteriological standard for Klila product. Therefore, we have adopted the bacteriological parameters limits given by the Moroccan legislation (NM 624-04, 2004) to judge its hygienic quality.
The total coliforms results were correlated with the work of (Hamama, 1989a). In addition, we found that fresh cheese "Jben" samples are not contaminated with fecal coliforms. These results are in agreement with the work of (Mennane et al., 2007).

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However, fecal contamination is present in Klila samples with a large load close to the values given by (Hamama, 1989a;Rhiat et al., 2011) and that are 5,7x10 4 and 9x10 4 cfu/g respectively. The significant number of fecal contamination can be the consequence of a rapid and massive propagation of the fecal flora initially present in the raw milk used for cheese preparation.
Additional contamination of cheese by this flora can occur during the various manufacturing stages, views often unsanitary conditions wherein the preparation of the cheese is conducted (Hamama et al., 1996).
Isolation results of Clostridium showed a contamination of 96 and 40% of Klila and Jben samples respectively. These results were higher compared to those found by (Mennane et al., 2007) which 35,7% of Klila samples were contaminated by clostridia spores with absence of these contaminants in the fresh cheese "Jben". Most clostridia species are soil bacteria , so their presence indicates a telluric contamination.
Staphylococci enumeration revealed the presence of coagulase-negative staphylococci (CNS) with a percentage equal to 26 and 80% for Klila and Jben product respectively. One sample analyzed of Jben "J5" was contaminated by S. aureus. These results are consistent with the work of (Hamama et al., 1996) which reported a 73% contamination of fresh cheese samples examined by S. aureus with an average rate of 7x10 4 cfu/g, although only one sample has a population of pathogenic S. aureus greater than one million germs per gram.
The absence of pathogenic microorganisms namely Salmonella sp and Shigella sp, and that in all the analyzed samples is explained by the absence of contamination in milk and milk products at the source (Secke, 2007);  Original: milk from sick or carrier animals;  Direct: carriers or sick individuals;  Indirect: contacting the dairy product with a polluted environment during the preparation.

Hazards analysis:-
The prerequisite programs (PRP) represent the conditions and basic activities needed to maintain a hygienic environment for the production, storage and the provision of safe finished products throughout the process (Wallace and Williams, 2001). The prerequisites section, we have focused our efforts on education and personnel training on hygiene behavior, organized movement of raw materials, waste and personnel to avoid cross contamination.
However, the hazard analysis has been established only after the manufacturing diagram has been verified. It was carried out step by step the receipt of raw materials to the shipment of the finished product: First, we began by identifying hazards that are classified into three general types: biological (contaminants and pathogenic microorganisms), chemical (toxic substances) and physical (foreign bodies).
A 'HACCP' plan bringing together the key information of hazard identification revealed the existence of six critical points at different production stages of raw milk and the production process of dairy products, in which hazards are either biological 'microbial origin', which its parameters are determined in the regulations, or else physical and/or chemical. They are at: receipt and storage of raw materials 'raw milk'; Pasteurization/Sterilization; Inoculation and maturation; packaging; refrigeration and storage of finished product; cleaning and disinfecting, , it is imperative to be controlled by preventive and corrective measures that the industry must undertake to prevent the frequency of appearance of microbiological hazards probably the origin of food borne disease among consumers.
Awareness actions and information on potential risks are essential for consumers.

Conclusion:-
The results could highlight physicochemical and bacteriological properties of traditional cheeses namely Klila and Jben marketed at different towns in the southwest of Algeria compared to those reported in the regulations. However the absence of pathogenic organisms in substantially all of the samples analyzed, we recorded a high rate of fecal contaminants (total and fecal coliforms), which represent not only a factor in the deterioration of the quality of products but also may pose a risk to the health of consumers especially the presence of S. aureus recognized as the causative agent of food-borne infection 767 It is clear from this study, the importance of adopting a suitable manufacturing process that is vigilant and simple to prepare fresh cheese, fresh or dry white cheese, while respecting the measures of good hygienic practices and good manufacturing practices (GMP and GHP) to reduce the contamination level that can be produced and thus have a safe product and in compliance with the physicochemical and bacteriological standards established.