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Evaluation of the microbiological quality of broiler meat and antimicrobial susceptibility of isolates.

ABSTRACT

A total of 45 Enterobacteriaceae and staphylococci strains were isolated between February and May 2016. In this study, rates above the national microbiological standards for total aerobic mesophilic flora (FTAM), fecal coliforms, staphylococci and sulphite the gear boxes have been found. Salmonella was present in two samples on all four. The following types were present Serratia with (39.02%), and Escherichia Raoultella with (21.95%) for each genus Salmonella (12.2%), Enterobacter (2.44%) and Shigella (2.44%). Isolates doing on Baird Parker were identified as Staphylococcus spp. Staph are resistant to (100%) oxacillin and rifampin and sensitive (100%) to gentamicin and kanamycin. isolated enterobacteria are sensitive to the (100%) to amikacin, to (92.68%) to gentamicin, and (85.36%) to cephalexin and ciprofloxacin. Other sensitivities exist but, with lower rates. They are resistant to (80.48%) to Ticarcillin to (75.60%) to Tetracycline, and (73.17%) to Amoxicillin. They are resistant to other antibiotics, but with smaller proportions. This study showed that (46.34%) strains isolated enterobacteria are multi-resistant.

KEYWORDS: Broiler Meat, Enterobacteriaceae, Staphylococci, antibiotic, resistance.

INTRODUCTION

The production of poultry meat in 2007 accounted for over 26 percent of global meat supplies, has been more widely distributed between the developed and developing countries, given that it is China which recorded the highest growth rate [14].

The development of the poultry sector in Algeria has allowed a significant increase in broiler meat consumption. The latter increased from 0.82 kg/capita/year in 1972 to 9.18 kg/capita/year in 1986 and 9.70 kg/capita/year. Compared to other countries, Algeria remains in consumption, far behind the USA, Brazil, and the EU in 2003 which recorded respectively 51.8 kg/capita/year, 34.20 kg/capita/year and 22.9 kg/capita/year [16,28,2].

The development seems to have many causes including the very moderate price development that makes it very advantageous compared to red meat, its high protein and high effectiveness of its current production techniques.

In Algeria, the poultry sector "flesh" had know since 1980 a notable development, supported by an incentive policy. However, rearing and slaughter practices lag far behind the technological industrialized countries this resounding not only the productivity of poultry workshops, but especially on public health. Indeed, the problem of the poultry industry in terms of health still remains dependent on farming conditions in general, and especially the hygiene of buildings [24].

Livestock slaughter and workshops are exclusively private in Tebessa and do not meet the hygienic and sanitary requirements recommended by national and international legislation.

The sanitary barrier at the farm level and killings is so low, it is causing excessive mortality rates, of misuse of veterinary drugs and the spread of various diseases [24].

The richness of the water and protein in meat make it an essential food for a balanced diet. However, these same reasons make it a breeding ground for microbial growth. A big part of germs contaminating carcasses after the various stages of slaughter (skinned and eviscerated) are saprophytes. These bacteria, yeasts and molds. These are of spoilage microorganisms which cause putrefaction of meat. Moreover, the presence of pathogens responsible for foodborne illness is possible. It is often linked to health defects. These poisonings often caused by Salmonella spp, Escherichia coli, Staphylococcus aureus, Clostridium perfringens, etc...) can be quite severe [8].

Like all living things, animals are prone to diseases it is necessary to prevent or treat. The control of animal health guarantees not only the economic performance of a flock but also the welfare of animals. Only healthy animals may be slaughtered so that meat placed on the market pose no risk to consumer health.

In 2001, the World Health Organization (WHO) estimated that 50% of the antibiotics produced in the world were for farm and companion animals. Antibiotics are used in farming, for the treatment of infectious diseases of bacterial origin. But since the early sixties we see an increasing number of antibiotic-resistant bacteria and the emergence of new resistance.

Very few experimentally verifiable data exist on the microbiological quality of broiler meat and even less about the resistance of bacteria to antibiotics.

With this intention, our study is to investigate the alteration flora and contamination on broiler meat which aims to assess the frequency, identify its biochemical phenotype by analyzing a set of characters related enzyme activities and various metabolic pathways of these bacterial groups and study its resistance and antibiotic sensitivity.

MATERIALS AND METHODS

Sampling:

The biological material used for our study is represented by the meat broiler. The samples are taken at the butcher of the town of Tebessa. Samples are taken of the same compartment of the carcass, meat pectoral (chest). The choice of the chest muscle is based on the fact that it is the compartment of the richest carcass muscle tissue and the most requested by consumers, it is also the closest to the digestive system. The carcasses are selected randomly.

The work involves bacteriological analysis of 04 samples taken at chest level broiler meat bought at two outlets of the Tebessa region weekly. The samples are taken with a sterile knife. Being perishable, fresh meat therefore requires a transport accomplished in a refrigerant system. Indeed, the samples are held under cold in a cooling system (isothermal cooler) and quickly transferred to the laboratory. The samples were analyzed in the first hours after collection. The samples are rated as follows: E1, E2, E3, E4.

2[degrees]/Enumeration and isolation of bacteria:

Series of decimal dilutions of samples of broiler meat ([10.sup.-1] (Stock solution) to [10.sup.-4]) were performed on the tryptone salt broth and enumeration culture media and isolation were inoculated from dilutions: [10.sup.-1], [10.sup.-2], [10.sup.-3]et [10.sup.-4].

In Table 1 lists the culture media and incubation conditions for research of various germs. Research of Salmonella, we used two techniques depending on the analyzed portion

a) Chair:

We crushed in a mortar 25 g pectoral muscle meat in 225 ml trypton salt, after homogenization, we rested the homogenate for about twenty minutes for the revivification of microorganisms. The cutting operation was performed with a knife on a cutting board.

b) Skin:

The cut of 25 c[m.sup.2] of skin pectoral muscle meat chicken is made with a knife on a cutting board in a total asepsis. - Using a swab soaked in water buffered peptone we swabbed the entire surface of the cut skin; the swab is then placed in a test tube containing buffered peptone water. We mixed, remove the swab and close the tube. -Incube At 37[degrees] C for 24h. [22].

Isolation and conservation:

Isolation:

* The isolation was performed on agar VRBL, SS, Baird Parker previously poured and solidified in Petri dishes.

* From each box well suspect colonies isolated on which will be a Gram stain.

* The definite character bacteria are retained and transplanted until a pure culture. The purity and verified by macroscopic and microscopic observation by Gram stain, the end of the purification gram +/- characters must persist.

Preservation:

* It is performed on agar slant (GN), previously inoculated and incubated at 37 [degrees] C for 24 hours. Isolates will then be kept at 4 [degrees] C for a period of a few weeks.

Identification and preliminary analysis of isolates:

The identification of the isolates was performed by the application of conventional microbiology techniques, based on the research of a number of morphological, physiological and biochemical characteristics. All identification techniques have been described by Larpent [26], and Idoui Karam [19] and Gusils et al, [18].

The resulting colonies are observed under a dissecting microscope to determine their cultural characteristics (color, layout, shape and appearance).

After macroscopic examination of the purified strains, isolates were subjected to Gram staining, it allows to differentiate the Gram-positive bacteria from that of Gram-negative rods, shells, and the grouping mode.

* The catalytic activity has been demonstrated by the following technique: from a solid medium and aerobics, take a sufficient culture and suspended in a drop of [H.sub.2][O.sub.2] deposited on a slide. Presence of oxygen bubbles: catalase positive, no oxygen bubbles: catalase negative [10].

Highlighting oxidase is performed using oxidase discs, it consists of depositing on a blade of a disc oxidase, and imbibe a drop of sterile saline, collecting a colony to using a Pasteur pipette and spread it on the disc. If occurrence of a dark purple color, it is concluded that the bacterium is oxidase positive, and it has the cytochrome oxidase [10].

* The degradation of Mannitol was studied middle-Mannitol Mobility, mannitol fermentation causes the yellow in the middle of the turn, moving strains spread from the seeding line by creating an environment of disorder, while still strains only grow along the streak seeding [9].

* The determination of H2S production by the isolates is sought on middle Classic triple SugarIron agar (TSI) inoculated with the strain selected and incubated at 37 [degrees] C, producing results in blackening of the medium [4]. The TSI medium also allows to highlight the degradation of glucose, lactose, and sucrose, This medium is composed of a base and of a slope, and contains the methyl red as a pH indicator.

* The results are manifested as follows:

* Yellow Fitting, positive Glucose

* Slant turns yellow, lactose and/or sucrose positive

* Production of gas: the presence of gas bubbles in the pellet.

Specific identification of Staphylococcus aureus:

Pigmentation on GN:

* The medium used is nutrient agar. It is previously poured into the boxes because seeding is done at the surface.

* was inoculated 0.1 ml of each of the bacterial suspensions of the isolates, the dishes are incubated at 37 [degrees] C for 24h. Typical colonies are yellow to 1 to 2 mm in diameter.

Research nitrate reductase:

Nitrated is inoculated broth is incubated at 37 [degrees] C for 24h after incubation were added reagents NR I and NR II. If red/pink: + nitrate reductase, if colorless is added Zinc powder: If red: NR - if colorless + NR.

Research coagulase:

* are seeded broth heart-brain (BHIB) for 24 h at 37 [degrees] C.

* After incubation mixed in a test tube 0.5 ml human plasma + 0.5 ml of culture on BHIB.

Transfer to oven, the tube must be inclined. If Staphylococcus aureus, coagulation may take place after 30 minutes to 3 hours. if the plasma is not coagulated, species other than Staphylococcus aureus.

Clumping factor research (clotting factor):

Emulsify a little culture on GN in a plasma drop.

* If there is agglutination, the bacterium is Staphylococcus aureus.

Voges-Proskauer reaction (medium of Clark-Lubs):

Bacteria called positive Voges-Proskaeur (V[P.sup.+]) have a particular metabolic pathway in the fermentation of hexoses. A from pyruvic acid oxidation product of glucose, they can form the acetylmethylcarbinol (acetoin).

* were seeded the Clark-Lubs medium which is incubated for 24 h at 48 h at 37 [degrees] C. After incubation add 2 to 3 drops of each reagents VP I and VP II, tilt the tube and read after 10 min.

* If pink or red (VP +), so: the bacterium is Staphylococcus aureus.

The test of dihydrolase arginine (ADH):

The detection of this enzyme is very useful for the characterization of Staphylococcus aureus which are ADH +, this enzyme releases ammonia and citrulline from arginine. For it is inoculated with the test organism a tube without arginine base medium and supplemented with arginine tube, it covers the surface of the medium with 1 ml of sterile paraffin oil. The results are interpreted after 48 h incubation. The transformed control when the medium has a violet color. It is not yellow as if the [27].

Biochemical identification of enterobacteria:

Any isolate having Gram negative bacilli catalase positive, oxidase negative, was subjected to biochemical identification by API20E (BioMerieux R SA). It is a standardized system for the identification of Enterobacteriaceae and other Gram bacilli not tedious negative, with 20 miniaturized biochemical tests and a database.

Susceptibility:

Antibiograms has been made in quality control laboratory, Applied Biology Department, University Larbi Tbessi Tebessa, Algeria. The susceptibility testing is performed according to the agar diffusion method (discs methods). The technique is to deposit on the surface of Mueller Hinton agar previously seeded with a bacterial suspension, antibiotic discs. We used 13 antibiotic discs (BIOANALYSIS) for Enterobacteriaceae and 05 disks for staphylococci. The antibiotics tested are chosen according to the CA-SFM [6].

Each antibiotic diffuses within the agar. After 18 to 24 hours incubation at 37 [degrees] C, each disk is surrounded by a zone of inhibition of bacterial growth in the case of the sensitivity, the contrast and correct. The measurements of the diameters of inhibition zones are made using a ruler and the results are interpreted according to the criteria of the Committee of the antibiogram of the French Society for Microbiology (CA-SFM [7].

To determine the similarity of 41 Enterobacteriaceae strains to different antibiotics, we realized a dendrogram using the XLSTAT Version 2014 software.

Results:

I. Enumeration of the microbial flora of the broiler meat: I-1. Flora aerophilic total mesophilic (FTAM):

The listing of this flora demonstrated that there is significant contamination of meat for the four samples tested with a much higher value than fixed by the official newspaper of the Algerian Republic (JORA 035 of 27/05/1998) which is 5.[10.sup.5] CFU/g(Table 2).

I--2. Total and Fecal Coliforms:

The analysis showed a significant load of fecal coliform in the middle VRBL superior to Algerian standards that are [10.sup.3] CFU/g (JORA 035 of 27/05/1998). The samples are concerned the sample 2 with an average load of fecal coliform 2.8 [10.sup.3] CFU/g, sample 3 with an average value of 8.8.[10.sup.4] CFU/g and the sample 4 with a heavy load of 2.8.[10.sup.5] CFU/g. A significant burden of total coliforms was counted in all the samples studied. (Table 2).

I-3 Staphylococcus aureus:

The analysis of our samples showed high values for stapphylococcus spp on Baird Parker medium for samples 1 and 3 (1, 3.[10.sup.4] and 5.[10.sup.3]) respectively and a total absence for samples 2 and 4 (table 2).

I-4. Counting on SS medium:

Among four samples studied 3 (75%) had suspicious colonies to be salmonella. After identification of the isolates, samples 2 and 3 were contaminated with salmonella at the skin level for sample 2 and at the flesh for sample 3, which was a contamination rate (50%) (table 2).

I-5. sulphite-reducing:

The results show uncountable values of sulphite reducing agents in all the samples analyzed. The results obtained exceed the Algerian standard which is set at 30 CFU/g (JORA 035 of 27/05/1998).

II- Specific identification of Staphylococcus aureus:

The analysis of the results allowed us to see that the isolates were characterized by; the presence of catalase + except E3S2 isolate, an oxidase +, a H2S production except for E3S6, an DHA + for E3S6 and E3S7 isolates and a nitrate reductase + to all isolates, however, we noticed the absence of coagulase, mobility and pigmentations. Unfortunately these distinguishing characteristics do not allow us to link isolates identified to species references, reason for which our isolates were identified as Staphylococcus spp (Table 3).

IV-1 Resistance Staphylococcus spp to antibiotics:

This study showed 100% resistance to oxacillin and rifampcin. Against by a sensitivity of 100% was observed vis-a-vis of gentamicin and kanamycin. Any time we have seen a fusidic acid resistance and clindamycin of E3S2 strains and E3S6. In contrast, E3S3 and E3S7 strains are sensitive to these antibiotics. (Table 4).

III. Specific identification of Enterobacteriaceae:

In order to identify Enterobacteriaceae isolates, we have made a preliminary identification of forty one isolates by some conventional biochemical tests. Analysis of these results showed that all isolates are gramnegative, catalase positive and negative oxidase which is characteristic of Enterobacteriaceae. The isolates showed very similar characteristics and profile of enterobacteria. Nevertheless these characteristics are insufficient to give the species and type of these strains. Some serotypes of Salmonella are recognized by their H2S production and gas and others are negative and H2S gas. All isolates are mobility negative, positive nitrate reductase.

From the results of the API 20E, and using identification software, we could identify our isolates and bring the species presented in (Table 05).

III-1 Distribution function of genres strains and species:

The distribution of strains identified depending on gender and depending on the species are shown in the table (06). One notices a predominant gender Serratia, with a percentage (39.02%) particularly the species Serratia odorifera. The genera Escherichia and Raoultella occupy the second position with a percentage of (21.95%) by representing a single species for each genus Escherichia coli 1 for Escherichia Raoultella ornithinolytica for Raoultella, followed by the kind Salmonella third in is representative of a single species choleroessius spparizonae (12.2%), and finally there is Enterobactercloacae whose species and Shigella spp whose species with a rate of (2.44%) for each species.

Antibiotic susceptibility testing:

The 41 isolated Enterobacteriaceae strains were divided into three categories: resistant (R) sensitive (S) and intermediate (I). The percentages of resistant strains, S and I to different antibiotics are shown in Tables (07) and figure (01).

The results obtained allowed us to note that:

* The antibiotics used are active on strains isolated.

* The most active antibiotics by uncrossing order are: AN (100%), GM (92.68%), CL (85.36%), CAZ (78.04%), AMC and SXT (51.21%) PIP (46.34%), OFX (43.90%), AMX and TE (24.39%), NA (21.95%), ICT (19.51%), CIP (7.31%) and RCV (100%).

* * The largest resistance are; CIP (85.36%), ICT (80.48%), TE (75.60%), AMX (73.17%), NA (68.29%), SXT (48.78%), AMC (16.34%), PIP (36.58%), OFX (29.26%), CAZ (21.95%), CL (14.63%) and GM (4.87%).

V. multi-resistant Enterobacteriaceae:

* According to the recommendations of the committee susceptibility of the French Society of Microbiology (CA-SFM, 1999), were defined as multi-drug resistant, resistant strains (R + I) to at least one antibiotic in three different families of antibiotics among the following: [beta]-lactams, quinolons, aminoglycosides, sulfonamides (cotrimoxazole). Based on this definition, the detection rate of multidrug-resistant Enterobacteriaceae in our study was (46.34%). Nineteen (19) strains showed multi-resistance of 41 strains (table 8).

VI. Statistical study of the sensitivity of strains to antibiotics:

The reading of the susceptibility testing for isolates and determination of the similarity between these strains is based on the establishment of a dendrogram that pays the diameters of inhibition zones of each antibiotic tested on these strains. The results of the susceptibility of 41 isolates studied by HAC (Hierarchical Classification). We have made tables illustrating the diameters of inhibition zones on antibiotic strains studied. The rest of the calculations is performed using a XLSTAT software. The 41 isolates are noted in orderly, grouped according to their similarity, and x-axis taxonomic distances representing the similarity between strains (Figure 2).

An arbitrary cut-off of the tree includes the groups isolates. Determining the cutoff level gives the taxonomic distance (0.26) 26% as the most suitable to obtain homogeneous groups of isolates: CA'1, CA'2 and CA'3.

The CA'1 group (class 1) is composed of 9 strains. This group is quite homogeneous and strain sensitivity profiles to the tested antibiotics are almost identical. Strains of this group are characterized by sensitivity to CL, AMX, AMC and TC compared to strains of group and CA'2 CA'3.

The CA'2 group (class 2) brings together 29 strains. The strains of this group are characterized by resistance to nalidixic acid, gentamicin and ciprofloxacin.

The CA'3 group (Class 3) is composed of 03 strains, they are characterized by their resistance to AMC AMX, TC, PC and OFX.

Discussion:

The total mesophilic aerobic flora always informs us on the hygienic quality of broiler meat. It is considered as the factor determining the shelf life of meat. It is the most sought flora in microbiological analyzes. Sources of contamination of food by the total aerobic bacteria are very diverse: environment, animals, cross-contamination with other carcasses or food contamination manipulator. Their presence beyond the defined limits can mean a lack of hygiene in manufacturing processes or even, beyond [10.sup.7]CFU/g, a state of putrefaction [17]

This study showed a significant load of fecal coloformes exceeding the Algerian standards set in the Official Journal of the Algerian Republic (JORA 035 of 27/05/1998). In a study in Morocco [23] on fresh carcasses of broilers, fecal coliforms found the rate (1.48 [10.sup.6] CFU/g) was higher than the values found in our samples. These are witnesses coliform bacteria of fecal contamination. This microbial population is similar in practice to Escherichia coli. And enumeration of fecal coliforms can follow hygiene manipulators meat, in all its economic circuit [15]. Concerning the presence of fecal coloforms, Our results are similar to that found by [23] on fresh carcasses of chicken meat, which found (5.92 [10.sup.6] CFU/g)

The results on the baird parker medium revealed significant contamination of the E1 and E3 samples by Staphylococcus spp. These average values exceed the standard Algerian who is 5. [10.sup.2] CFU/g Staphylococcus aureus(JORA 035 of 27/05/1998). In Morocco an average of Staphylococcus spp 1.3 105 CFU/g was found on fresh carcasses of broiler meat [23]. Among the risk factors that may affect the number of staphylococcus present in the samples studied, the level of hygiene in the massacre, the cleanliness of the water used during the scalding, cleanliness fingers of pluckers, staff hygiene, the precautions taken at the time of evisceration and handling during the sale. Indeed, during the slaughter, phenomena of cross contamination occur, which induces proliferation of pathogens on initially healthy carcasses [1].

The presence of salmonella in broiler meat has been found in other studies. In a study in Constantine (Algeria), Salmonella contamination has affected (37%) of farms and (53%) of slaughterhouses [12].

According to our results, which showed a significant contamination of the samples by sulphite-reducers, we can explain this contamination of carcasses by these germs after slaughter, since they are of exogenous, often human, origin resulting from the handling of the meat by the staff.

About antibiotic resistance, the literature reports of resistance of Staphylococcus insulated food coagulase negative. Involved, according to this study the regular use of low-dose antibiotics in factory farming, making the animal an ideal breeding grounds for resistant bacteria. A phenomenon that produces bacteria resistant to 3, 4, 5 or 9 different antibiotics, leaving, in case of human infection, very few therapeutic option

From these results, we can conclude that there is a predominance of Serratia bacteria, Escherichia and Raoultella. We have no studies that have isolated Raoultella ornithinolytica in meat of poultry, we can confirm that this strain was isolated Tuna, and Sardine by [25]. In general, species of the genus Serratia are isolated from plants (vegetables, mushrooms, mosses), digestive rodents tube (40% of small wild mammals carry Serratia spp.), Insects, water and soil [13]. This is what can explain their presence in meat of poultry. Their transmission can occur through the water and soil.

The presence of salmonella and Escherchia coli in meat poultry was confirmed by other authors [11, 12, 5]. The reservoir of the genus Salmonella bacteria is primarily the gastrointestinal tract of vertebrates. Numerous animal species harbor these pathogens (poultry, cattle, pigs, fish, reptiles...). Enterica subspecies is more suited to warm-blooded animals and humans [31].

The collection of our strains is generally resistant to many antibiotics normally active on Gram-negative bacilli.

This rate of multi-resistant enterobacteriacae is close to the data found in Switzerland who noted that these multi-resistant bacteria can be detected in almost half of broiler flocks. Multi-resistant bacteria are now at the heart of public debate. The frequent presence of such bacteria in broiler poultry and in poultry meat is also regularly mentioned. In recent years, in Switzerland as in other countries, Enterobacteriaceae (intestinal bacteria) producing ESBL were also more frequently identified in farm animals healthy, especially in poultry. These animals are carriers of this kind of multi-resistant bacteria in their intestines [30].

The CA'1 group (class 1) is composed of 9 strains. This group is quite homogeneous and strain sensitivity profiles to the tested antibiotics are almost identical. Strains of this group are characterized by sensitivity to CL, AMX, AMC and TC compared to strains of group and CA'2 CA'3.

The CA'2 group (class 2) brings together 29 strains. The strains of this group are characterized by resistance to nalidixic acid, gentamicin and ciprofloxacin. According to CA-SFM [7], Salmonella spp strains resistant to nalidixic acid should be categorized resistant to fluoroquinolones. The categorized strains sensitive to nalidixic acid can be rendered susceptible to moxifloxacin, ciprofloxacin and ofloxacin. If resistance to nalidixic acid, fluoroquinolone sensitivity must be determined.

The CA'3 group (Class 3) is composed of 03 strains, they are characterized by their resistance to AMC AMX, TC, PC and OFX.

These results show that the meat of broiler sold in Tebessa does not offer a sufficient guarantee of safety especially with an important contamination as that found in this study. Efforts are still needed throughout the chain of production to sales through slaughter to present a final product quality ableto.the consumer confidence.

[FIGURE 1 OMITTED]

[FIGURE 2 OMITTED]

Conclusion:

* At the end of this work forty five (45) strains were isolated, purified and identified from four broiler meat samples marketed in Tebessa city of eastern Algeria.

* Identification of strains was carried out by the determination of morphological, physiological and biochemical.

* Analysis of the results obtained by specific identification of Staphylococcus aureus have we found that the isolates were identified as Staphylococcus spp.

The identification of Enterobacteriaceae isolates, we found that the 41 strains are divided into 6 types with predominant gender Serratia, with a percentage (39.02%), the genus Escherichia and Raoultella second with a percentage (21.95%), followed by the kind with Salmonella (12.2%) and Enterobacter and Shigella last position with (2.44%) for each of the two genres.

Antibiotic resistance shows a 100% resistance of Staphylococcus spp oxacillin and rifampin and a sensitivity of 100% to gentamicin and kanamycin.

For Enterobacteriaceae antibiotics used are active however stem.

However, the strains are:

100% sensitive to amikacin, 92.68% gentamicin, and 85.36% to cefalexin and ciprofloxacin. Other sensitivities exist but, with lower rates.

80. 48% resistant to ticarcillin, 75.60% to tetracycline, and 73.17% to amoxicillin. They are resistant to other antibiotics, but with smaller proportions.

The detection rate of multi-resistant Enterobacteriaceae shows that 19 strains (46.34%) are multi-resistant strains of the 41.

The study of the degree of similarity of isolates yielded three distinct classes of enterobacteria. These classes are characterized by their resistance to antibiotics as a result:

The CA'1 group: 9 strains groups that are characterized by their resistance to cifladizime, amoxicillin + clavulanic amoxicillin and ticarcillin from strains of group and CA'2 CA'3.

The group CA'2 has 29 strains which are characterized by their resistance to amoxicillin, piperacillin, ticarcillin, nalidixic acid, ofloxacin and tetracycline.

The group CA'3 is composed of 03 strains which are characterized by their resistance to the following antibiotics: Amoxicillin + clavulanic acid, amoxicillin, ticarcillin, ciprofloxacin and ofloxacin.

These results show that the meat of broiler sold in Tebessa does not offer a sufficient guarantee of safety especially with an important contamination as that found in this study. Efforts are still needed throughout the chain of production to sales through slaughter to present a final product quality ableto the consumer confidence.

ACKNOWLEDGMENTS

The authors of this article would like to thank everyone who participate to the realization of this work.

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(1,2,3) Salima Taleb, (1) Manel Debebza, (1) Yasmine Yousfi Y, (1) Nadia Rezkellah

(1) Faculty of exact sciences and sciences of nature and life. Applied Biology Department. University Larbi Tbessi 12000. Tebessa Algeria

(2) Laboratory of Nutrition and Food Technology. 1. University of Constantine Algeria

(3) Water and Environment Laboratory. University Larbi Tbessi 12000. Tebessa Algeria

Address For Correspondence:

TALEB Salima, Faculty of exact sciences and sciences of nature and life. Applied Biology Department. University Larbi Tbessi 12000. Tebessa Algeria

This work is licensed under the Creative Commons Attribution International License (CC BY).http://creativecommons.org/licenses/by/4.0/

[ILLUSTRATION OMITTED]

[ILLUSTRATION OMITTED]

Received 12 January 2016; Accepted 10 March 2017; Available online 26 March 2017
Table 1: Culture Media and incubation conditions

Germs sought       Culture medium used          Type seeding

FTAM               Nutrient Agar                Depth
Total coliforms    Lactose Agar biliee crystal  Depth
                   violet and neutral red
                   (VRBL)
Fecal coliform     Lactose Agar biliee crystal  Depth
                   violet and neutral red
                   (VRBL)
Staphylococcus     Baird Parker                 Area
Salmonella         --Water Buffered peptone     Inoculation
                   -Bouillon selenite cystine
                   (SFB)                        Inoculation
                   -Agar SS
                                                Area
Sulphite-reducing  Liver meat                   Inoculation
Germs sought       Incubation condition
                   Temperature   Time
FTAM               30[degrees]C  72h
Total coliforms    37[degrees]C  24h
Fecal coliform     44[degrees]C  24 a 48h
Staphylococcus     37[degrees]C  24h
Salmonella         37[degrees]C  24h
                   37[degrees]C  24h
                   37[degrees]C  24h
Sulphite-reducing  37[degrees]C  24h

Table 2: Prevalence of microflora

Samples
Microbial flora            E n[degrees] 01  E n[degrees]2
FTAM                       8.9.[10.sup.6]   Uncountable
Total Coliforms            3,6.[10.sup.5]   5,8.[10.sup.6]
(CFU/g)
Fecal coliforms (CFU/g)    8. [10.sup.2]    2.8.[10.sup.3]
Staphylococcus             1, 3.[10.sup.4]  Absence
spp(UFC/g)
Salmonella                 Flesh  Skin      Flesh  Skin
                           -      -         -      +
Sulphite-reducing          Uncountable      Uncountable
Samples
Microbial flora            E n[degrees]3    E n[degrees]4
FTAM                       5.2. [10.sup.7]  Uncountable
Total Coliforms            4,5.[10.sup.5]   1,2.[10.sup.7]
(CFU/g)
Fecal coliforms (CFU/g)    8.8 [10.sup.4]   2,8.[10.sup.5]
Staphylococcus             5.[10.sup.3]     Absence
spp(UFC/g)
Salmonella                 Flesh  Skin      Flesh  Skin
                            +     -         -      -
Sulphite-reducing          Uncountable      Uncountable

(-) = Absence, (+) = Presence

Table 3: Identification Test of Staphylococcus aureus

Physicochemical tests     Isolates
                          E3 S2             E3 S3     E3 S6     E3 S7
Catalase                  +                 +         +         +
Oxidase                   +                 +         +         +
Pigmentation on nutrient  -                 -         -         -
agar
Mannitol fermentation     Manitol+          Manitol+  Manitol+  Manitol+
Nitrate reductase         NR+               NR+       NR+       NR+
Coagulase                 -                 -         -         -
Clumping factor           No agglutination
Mobility ADH              -                 -         -        -
                          +                 +         -        -
     bottom               +                 +         +        +
TSI  Slope                +                 +         +        +
     H2S                  +                 +         +        +
     Gas                  +                 +         -        +
VP                        -                 -         -        -
Methyl Red (MR)           +                 +         +        +

E = sample, S = strain

Table 4: Results of staphylococci sensitivity test to antibiotics

ATB        GM  K  FA  OX  RA  DA
Stem code
E3 S2      S   S  R   R   R   R
E3S3       S   S  S   R   R   S
E3 S6      S   S  R   R   R   R
E3 S7      S   S  S   R   R   S

GM = Gentamicin, K = Kanamycin, FA = fusidic acid, OX = oxacillin,
RA = rifampicin, DA = clindamycin
S = Sensitive, R = resistant, E = sample, S = strain

Table 5: Results of the biochemical identification of different
insulation

Codes strains             Identification

STRAINS ISOLATED ON VRBL
E1S1CF                    Escherchia coli 1
E1S3CF                    Escherchia coli 1
E1S4CF                    Raoultella ornithinolytica
E1S5CF                    Raoultella ornithinolytica
E1S6CF                    Raoultella ornithinolytica
E1S7CF                    Raoultella ornithinolytica
E3S1CF                    Escherchia coli 1
E3S2CF                    Raoultella ornithinolytica
E3S5CF                    Enterobacter cloacae
E3S6CF                    Raoultella ornithinolytica
E3S7CF                    Raoultella ornithinolytica
E3S8CF                    Raoultella ornithinolytica
E4S7CF                    Escherchia coli 1
E4S8CF                    Escherchia coli 1
E2S1CT                    Escherchia coli 1
E2S4CT                    Serratia oderifera
E3S1CT                    Escherchia coli 1
E3S4CT                    Raoultella ornithinolytica
STRAINS ISOLATED ON SS
SSE2P1                    Salmonella choleroessius SPP arizonae
SSE2P2                    Salmonella choleroessius SPP arizonae
SSE2P3                    Salmonella choleroessius SPP arizonae
SSE2P4                    Serratia oderifera
SSE2P7                    Salmonella choleroessius SPP arizonae
SSE2P80                   Serratia oderifera
SS E1 S1                  Serratia oderifera
SS E1 S2                  Serratia oderifera
SS E1 S3                  Serratia oderifera
SS E1 S4                  Serratia oderifera
SS E1 S5                  Serratia oderifera
SS E1 S6                  Escherchia coli 1
SS E1 S7                  Escherchia coli 1
SS E2 S1                  Serratia oderifera
SS E2 S2                  Serratia oderifera
SS E2 S3                  Serratia oderifera
SS E2 S4                  Serratia oderifera
SS E3 S1                  Serratia oderifera
SS E3 S2                  Serratia oderifera
SS E3 S3                  Serratia oderifera
SS E3 S4                  Serratia oderifera
SS E3 S5                  Salmonella choleroessius SPP arizonae
SS E3 S6                  Shigella spp

E = sample, S = strain, CF = Fecal coliform, CT = Total coliform,
SS = ss agar

Table 6: Distribution of strains depending on gender

Kind          Bacterial species           Effective  Percentage

Escherichia   Escherichia coli            9          21,95%
Serratia      Serratia odorifera          16         39,02%
Enterobacter  Enterobacter cloacae        1          2,44%
Salmonella    Salmonella choleroessius    5          12,2%
              SPP arizonae
Raoultella    Raoultella ornithinolytica  9          21,95%
Shigella      Shigella spp                1          2,44%

Table 7: Distribution of strains according their sensitivity to
antibiotics

Antibiotics       Initials  Number of strains (percentage of strains)
                            Resistant    Sensitive    Intermediate

Amoxicillin +     AMC       19 (16,34%)  21 (51,21%)  1 (2,43%)
Clavulanic acid
Amoxicillin       AMX       30 (73,17%)  10 (24,39%)  1 (2,43%)
Ceftazidim        CAZ       9 (21,95%)   32 (78,04%)  0
Ticarcillin       TIC       33 (80,48%)  8 (19,51%)   0
Cefalexin         CL        6 (14,63%)   35 (85,36%)  0
Piperacillin      PIP       15 (36,58%)  19 (46,34%)  7 (17,07%)
Amikacin          AN        0            41 (100%)    0
Gentamicin        GM        2 (4,87%)    38 (92,68%)  1 (2,43%)
Tetracyclin       TE        31 (75,60%)  10 (24,39%)  0
Nalidixic acid    NA        28 (68,29%)  9 (21,95%)   4 (9,75%)
Ofloxacin         OFX       12 (29,26%)  18 (43,90%)  11 (26,82%)
Ciprofloxacin     CIP       3 (7,31%)    35 (85,36%)  3 (7,31%)
Sulfametoxazole   SXT       20 (48,78%)  21 (51,21%)  0
+, Trimethoprim

Table 8: Distribution of multi-resistant strains as antibiotics to
which they are resistant.

Code      Strains

CFE1S3    Escherchia coli 1
CFE1S7    Raoultella ornithinolytica
CF E3 S1  Escherchia coli 1
CF E3 S2  Raoultella ornithinolytica
CF E3 S5  Enterobacter cloacae
CF E3 S7  Raoultella ornithinolytica
CF E3 S8  Raoultella ornithinolytica
E2 SS P4  Serratia oderifera
E2 SS P7  Salmonella choleroessius SPP arizonae
SS E1 S4  Serratia oderifera
SS E2 S1  Serratia oderifera
SS E2 S2  Serratia oderifera
SS E2 S3  Serratia oderifera
SS E3 S1  Serratia oderifera
SS E3 S2  Serratia oderifera
SS E3 S3  Serratia oderifera
SS E3 S4  Serratia oderifera
SS E3 S5  Salmonella choleroessius SPP arizonae
SS E3 S6  Shigella spp
Code      PROFILE
CFE1S3    AMX, TIC, PIP, NA, OFX, SXT
CFE1S7    TIC, PIP, NA, OFX, SXT
CF E3 S1  AMX, CAZ, TIC, PIP, NA, OFX, SXT
CF E3 S2  AMC, AMX, CAZ, TIC, PIP, SXT, OFX
CF E3 S5  AMC, AMX, TIC, NA, SXT
CF E3 S7  AMX, TIC, PIP, NA, SXT
CF E3 S8  AMX, TIC, PIP, NA, OFX, SXT
E2 SS P4  AMC, AMX, TIC, PIP, NA, SXT
E2 SS P7  AMC, AMX, CAZ, TIC, PIP, NA, OFX, SXT
SS E1 S4  AMC, AMX, TIC, PIP, NA, OFX, GN, AK, SXT
SS E2 S1  TIC, NA, OFX, SXT
SS E2 S2  AMC, AMX, TIC, CL, NA, OFX, SXT
SS E2 S3  AMX, TIC, PIP, OFX, GN, SXT
SS E3 S1  AMC, AMX, CAZ, TIC, CL, PIP, NA, OFX, CIP, GN, SXT
SS E3 S2  AMC, AMX, TIC, NA, OFX, SXT
SS E3 S3  AMC, AMX, TIC, CL, PIP, NA, OFX, SXT
SS E3 S4  AMC, AMX, TIC, PIP, NA, OFX, SXT
SS E3 S5  AMC, TIC, PIP, NA, SXT
SS E3 S6  AMC, TIC, NA, SXT

E = sample, S = strain, CF = Fecal coliform, SS = ss agar, p = (Skin).
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Author:Taleb, Salima; Debebza, Manel; Yousfi Y, Yasmine; Rezkellah, Nadia
Publication:Advances in Environmental Biology
Article Type:Report
Geographic Code:6ALGE
Date:Mar 1, 2017
Words:6735
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