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Molecular Characterization and Virulence Gene Profile of Entero- pathogenic and Shiga-Toxin Producing Escherichia coli from Food of Animal Origin and Environmental Sources
(SKUAST Kashmir, 2023) Shah, Iqra Hussain; Syed, Akram Hussain
The present study describes occurrence, molecular characterization virulence gene profile and antibiogram of entero-pathogenic and shiga-toxin producing Escherichia coli (E.coli) from food of animal origin and environmental Sources. Of the 456 samples screened for the presence of E. coli, comprising of animals (156), food of animal origin (60), environment (240), 243 (53.28%) were positive for E.coli. Among all samples highest occurrence of E. coli was recorded from animals (85.28%) followed by environment (38.75%) and food of animal origin (28.33%). A polymerase chain reaction (PCR) was used to confirm the E. coli spp. to the genus level (16SrRNA). Molecular characterisation using multiplex PCR revealed a total of 29 (11.93%) isolates positive for eae gene, 58 (23.86%) for Stx1 gene, 43 (17.69%) for Stx2 gene, 20 (8.23%) for both Stx1& Stx2 gene, 20 (8.23%) for hlyA gene and 10 (4.11%) for ehx gene. None of the recovered isolates were positive for bfp gene. The occurrence of intimin (eae), Stx1, Stx2, hylA gene was recorded highest (15.03%, 27.81%, 24.81%, 14.81 and 11.27% respectively) in E. coli isolated from animals followed by environment (8.33%, 21.05%, 8.06%, 5.29% respectively) and food of animal origin (5.88%, 5.88%, 11.76%, 5.88%, 0.00% respectively). Out of 121 Shiga-toxin producing E. coli (STEC) isolates recovered, Stx1 gene was found associated with 44 (36.36%) isolates, 32 (26.44%) as Stx2, 20 (16.52%) as both stx1/stx2, 08 (6.61) as eae/stx1, 07 (5.78%) as eae/stx2, 6 (4.95%) as Stx1/ehx, and 4 (3.30%) as Stx2/ehx. Among all the samples, atypical STEC was observed to be more prevalent 111 (91.73%) than typical STEC 10(8.26%). The overall occurrence of entero-pathogenic E. coli (EPEC) noted from different samples was 14 (3.07%). Among the different samples, EPEC was observed more in animals 11(7.05%) compared to environment 03 (1.25%). None of the sample from food of animal origin was detected positive for EPEC. All the EPEC isolates were identified as atypical. Of the 243 E. coli isolated, the eae gene was identified in 29 (11.93%) isolates. Out of 29 eae positive isolates, 11 (37.93%) were typed as Int-β and 15 (51.72%) isolates were typed as Int-γ. Three (10.34%) eae positive isolates couldn’t be typed using the available set of primers. Upon subtyping of eae positive samples, Int- β was observed in 3 (37.50%) of isolates from environment, 7 (35.00%) of isolates from animals and 1 (100%) of isolates from food of animal origin. While as Int- γ was noted in 11 (55.00%) isolates from animals, 4 (50.00%) isolates from environment. A high level of resistance was observed against Ampicillin (100%) which was followed by tetracycline (60.59%), Co-trimoxazole (44.85%), streptomycin (43.62%), chloramphenicol (32.94%). However, the results showed that isolates recovered from different sources were completely (100%) sensitive towards Cefuroxim and Cefepime. The present study thus confirmed the presence of coliform organisms including STEC and EPEC in animals, food of animal origin and environment indicating a potential hazard. The wider prevalence of multidrug resistant E. coli poses a threat of spread of antibiotic resistance among micro-organisms, therefore proper and judicious use of antimicrobial agents is the need of hour to prevent spread of drug resistance.
Open Access
Molecular Epidemiology and Virulence Gene Profile of Aeromonas species from Milk and Environment
(SKUAST Kashmir, 2021) Maliha Gulzar; Rather, Mudasir Ali
The present study describes the occurrence, molecular epidemiology, virulence gene profile and antibiogram of Aeromonads from milk, water and environmental samples. Of the 810 samples screened for the presence of Aeromonas species, comprising of milk (238), environment (386), water (186), 330 (40.74%) were positive. The occurrence of Aeromonas spp. in milk, environment, and water samples was 29.8, 49.48 and 36.55%, respectively. A polymerase chain reaction (PCR) was used to confirm the Aeromonas to the genus level (16SrRNA) and species level (rpoB/gyrB) as well. The molecular identification revealed that of the 330 samples which were positive for isolation of Aeromonads, a total of 482 isolates were identified as Aeromonas spp. based on 16SrRNA amplification. The speciation based on rpoB/gyrB gene amplification revealed that A. hydrophila(40.29%) was the most predominant species recovered followed by A. caviae(29.12%) and A. veronii (3.64%). There were Aeromonas isolates which could be identified upto genus level only (26.94%). The molecular epidemiology of mesophilic Aeromonads was studied on the basis of sequence analysis of gyrA, gyrBandrpoD genes. The sequencing of gyrAand gyrBgenes revealed mixed clusters formed between the milk and environmental isolates and separate clusters formed by the water isolates. However, on rpoD gene analysis no separate cluster for water isolates was present and formed mixed clusters with isolates recovered from milk and environment. Comparison of study isolates housekeeping gene sequences with the sequences downloaded from patric database (GenBank) revealed specific spatial distribution of study isolates. A multiplex PCR was used for detection of 2 enterotoxin genes viz. cytotonic heat labile enterotoxin (ahl) and cytotoxic heat labile enterotoxin(ahc) and uniplex PCR for cytotonic heat stable enterotoxin (ast) and lateral flagellar gene (fla) of mesophilic Aeromonas species. The ahl, ahc, astand flagenes were detected in 147 (48.83%), 153 (50.83%), 53 (17.60%) and 282 (93.68%) of isolated Aeromonads, respectively irrespective of the targeted species. Twelve enterotoxin gene patterns viz. ahc/ahl/ast/fla, ahc/ast/fla, ast/ahl/fla, ahc/ahl/fla, ahl/fla, ahc/fla, ahc/ahl, ast/ahl, ast/fla, ahc, flaandnone were identified. The ahc/fla (25.24%) was the most predominant gene pattern.A high degree of resistance was observed against ampicillin (all isolates were ampicillin resistant as isolated on Ampicillin Dextrin Agar), ceftazidime, trimethoprim, tetracycline, ceftriaxone, imipenem in order with the high number of isolates showing minimum inhibitory concentration above 16 µg/ml for antibiotics ceftazidime and tetracycline. Results of this study revealed the isolates were found highly sensitive to gentamicin, amikacin, levofloxacin, ciprofloxacin in order with the high number of isolates having a minimum inhibitory concentration less than 1µg/ml for antibiotic levofloxacin, ciprofloxacin and gentamicin. A. hydrophila and A. caviae isolates were highly susceptible to gentamicin and A. veroniiwas highly susceptible tocefepime. Irrespective of species, the highest number of isolates were resistant to ceftazidime. All 301 mesophilic Aeromonas spp. were subjected to ESBL-AmpC Co-existence detection and it was found that 288 isolates were AmpC positive and 93 were ESBL positive. ESBL-AmpC was co-detected in 18 isolates which were ampicillin resistant and ceftriaxone susceptible, 4 isolates which were resistant to ampicillin and showed intermediate resistance against ceftriaxone, 8 isolates which were resistant against ampicillin and ceftriaxone. The high occurrence of virulent Aeromonads in milk, water and environmental samples is a public health concern and thereby preventive measures are to be adopted for its spread among the human population. The sequence analysis of gyrAand gyrB genes can be a useful molecular epidemiological tool to determine the source of contamination in foods and water. The wider prevalence of multidrug resistant Aeromonas species poses a threat of spread of antibiotic resistance among micro-organisms, therefore proper and judicious use of antimicrobial agents is need of hour to prevent spread of drug resistance.
Open Access
A Study on Efficacy of Lactobacillus Spp. as A Biocontrol Agent Against Some Important Foodborne Bacterial Pathogens
(SKUAST Kashmir, 2019) Peerzada Rouf Ahmad; Mudasir Ali
The present investigation was undertaken to study the efficiency of Lactobacillus plantarum as a biocontrol agent to inhibit the In-vitro growth of three important food-borne pathogens viz. Staphylococcus aureus, Escherichia coli and Bacillus cereus. The extent of inhibitory effect of cell free extract of L. plantarum against these pathogens and change in In-vitro virulence characteristics of these food-borne pathogens was also studied. The pathogens to be tested in the experiment were recovered from various food sources and clinical samples. For isolation of S. aureus, 20 samples comprising of raw milk (5), mastitis milk (5), raw chicken (5) and sheep faecal samples (5) were taken and 6 samples turned out positive making an overall occurrence of 30%. Among all the categories of the samples processed, the highest occurrence of S. aureus was found in mastitis milk (40%). Of the 20 samples comprising of mastitic milk (5), raw milk (5), raw chicken (5) and sheep faecal samples (5) screened for isolation of E. coli, 7 (35%) samples were found positive. The highest occurrence of E. coli was found in sheep faecal samples (60%). For the isolation of B. cereus, 20 samples comprising 5 samples each of mastitic milk, raw milk, meat (chicken) and sheep faecal samples were taken, of which 5 samples turned positive, with an overall occurrence of 25%. The percent occurrence was higher in raw milk (40%) and raw chicken (40%). The ploymerse Chain reaction was carried out to amplify important virulent genes of the pathogens to determine their potential pathogenic nature. In S. aureus delta haemolysin gene (hld) was targeted and of the 6 isolates, 4 were found to carry the virulent genes. A virulent gene, hemolysin F (hlyf), of E. coli was targeted and was detected in 5 out of 7 isolates. In B. cereus the simultaneous presence of all three genes of hbl complex (hblC, hblD and hblA) was detected in all the 5 isolates. The efficiency of L. plantarum in inhibiting In-vitro growth of selected pathogens (S. aureus, B. cereus and E. coli) was observed. For the experiment varying concentrations of L. plantarum (104 to 108/ml) were grown along with standard concentration (108/ml) of each of the pathogen. Evaluation of L. plantarum against S. aureus was carried out by growing S. aureus (108/ml) alone and along with different concentrations of L. plantarum (104 to 108/ml) designated thereby as treatment groups (A to E) and counts were evaluated after three different incubation periods (24, 36 and 48 hrs). After 24 hrs of incubation at 37oC the mean colony forming unit count (CFU) of S. aureus in the control group (without L. plantarum in it) was found to be 622.66 x108 CFU/ml. However, after 24 hr of incubation at 37oC, the mean colony forming units of S. aureus in the treatment groups treated with 108, 107, 106, 105 and 104/ml of L. plantarum were 2.56 x108 CFU/ml, 2.78 x108 CFU/ml, 2.90 x108 CFU/ml, 3.03 x108 CFU/ml and 39.19 x108 CFU/ml, respectively. Similar trends were observed for S. aureus after 36 and 48 hrs of incubation. Adapting similar procedure for E. coli it was found that mean colony forming units of E. coli in the control group after 24 hours of incubation was found out to be highest (446.66 x108 CFU/ml.) as compared to all the treatment groups (A-E). The mean colony forming counts of E. coli were 0.81 x108 CFU/ml, 1.09 x108 CFU/ml, 1.36 x108 CFU/ml, 1.85 x108 CFU/ml and 24.62 x108 CFU/ml grown with different concentration of L. plantarum viz, 108, 107, 106, 105 and 104/ml, respectively. Similar results were for E. coli observed after 36 and 48 hrs of incubation periods. In B. cereus after 24 hrs of incubation the mean colony forming counts in the control group was 83.60x108 CFU/ml and in the treatment groups the counts were 0.31x108 CFU/ml, 0.43x108 CFU/ml, 0.60x108 CFU/ml, 0.69x108/ CFU/ml and 7.12x108 CFU/ml, when grown with L. plantarum with concentrations of 108, 107, 106, 105 and 104/ml, respectively. Henceforth, the mean colony forming counts of all the three pathogens increased when the concentration of L. plantarum decreased. The extent of inhibitory effect of cell lysates of L. plantarum was also evaluated on the pattern of disc diffusion assay and it was seen that the cell free lysate of L. plantarum (absorbed on filter paper discs was effective in inhibiting the In-vitro growth of all the three pathogens by forming zones of inhibition. The virulence characteristics (virulence Gene profiling, lecithinase activity and haemolytic activity) of these pathogens were analysed before and after growing with L. plantarum and it was found that there was no change in the In-vitro virulence properties of the isolates pre and post inoculation with L. plantarum. Therefore, it can be concluded that L. plantarum is effective in inhibiting growth of all the three pathogens as counts in the control groups were higher than all the treatments groups across different incubation periods. Moreover, mean counts of all the three pathogens decreased with the increase in concentration of L. plantarum, suggesting higher concentrations of L. plantarum are more effective in inhibiting growth of these pathogens. Therefore, L. plantarum can be used as a biocontrol agent for inhibition of growth of pathogens in the foods.
Open Access
Detection of enterotoxin genes and Antibiogram of Bacillus cereus from Raw Mutton and its slaughtering environment in Srinagar City
(SKUAST Kashmir, 2019) Zehgeer, Mohammad Mateen; Munshi, Ziaul Hassan
The present study was envisaged with the aim to study the occurrence, enterotoxin gene profile and antibiogram of B. cereus from raw mutton and its slaughtering environment in Srinagar city. Of the 200 samples comprising sticking area, carcass hanging hooks, water sample, meat(mutton), chopping board, meat shop floor, weighing pan and knife screened for presence of B. cereus, 78 samples turned out to be positive resulting in an overall occurrence of 39%. Of the 78 isolates, 16 (64%), 10 (40%), 4 (16%), 8 (32%), 6 (24%), 11 (44%),11 (44%) and 12 (48%) were isolated from, sticking area, carcass hanging hooks, water sample, meat(mutton), chopping board, meat shop floor, weighing pan and knife respectively. All the B. cereus isolates (78) were screened by multiplex PCR for detection of four enterotoxigenic genes (hblCDA and cytK). The isolates were divided into six different groups based on the presence or absence of enteroxigenic genes. Among the total isolates (78), Group I was occupied by 48 (61.53%), Group II by 8 (10.25%), Group III by 6 (7.69%), Group IV by 10 (12.82%), Group V by 3 (3.84%) and Group VI by 3 (3.84%) of isolates, respectively. The complete hbl complex and cytK was present in 54 (69.23%) and 59 (75.64%) isolates, respectively. The occurrence of various enterotoxigenic genes recorded for hblD, hblA, hblC and cytK were 73.07, 73.07, 73.07 and 75.64 per cent respectively. In-vitro antibiotic sensitivity of isolates revealed a high sensitivity towards gentamicin (96.15%), chloramphenicol (93.58%), ciprofloxacin (93.50%), erythromycin (87.17%), amikacin (79.48%), levofloxacin (75.64%), tetracycline (67.94%) and azithromycin (48.71%). Quite the opposite, highest resistance against Penicillin-G (100%), metronidazole (93.58%), ampicillin (88.46%), amoxicillin (91.02%) and cotrimoxazole (88.40%) was recorded. Among all the collected samples, the sticking area showed the highest occurrence of enterotoxigenic B. cereus isolates revealing the importance of maintaining hygienic practices at the sticking area which may otherwise lead to increased contamination of meat thereby creating an alarming situation of public health concern.
Open Access
Occurence and Virulence gene profile of Salmonellae from raw chicken and its slaughtering environment
(SKUAST Kashmir, 2019) Iyman Binti Fayaz; Munshi, Ziaul Hassan
The present study was carried out with the aim to determine the Occurrence and Virulence gene profile of Salmonellae from raw chicken and its slaughtering ekdijgpoultry meat, chopping board, weighing pan, knife and butchers’ hands and were screened for the presence of Salmonellae spp. Nine (09) samples identified as positive for the presence of Salmonellae presenting an overall occurrence of 4.28%. From the 210 isolates, 1(4%), 2 (8%), 3 (5%), 2 (8%), 0 (0%), 1(4%) and 0 (0%) of Salmonellae were isolated from, sticking area, cages, poultry meat, chopping board, weighing pan, knife and butchers’ hands respectively. All the 09 Salmonellae isolates were confirmed and were screened for three virulence genes of Inv A, Stn, Sef A by employing PCR. All the positive isolates were found to be positive carrier for genus specific 16S rRNA along with Inv A gene and stn gene whereas sef gene was found in 4(44.44%) isolates only. The antibiogram of the isolates as against 14 antibiotics, showed that all the isolates were sensitive towards ciprofloxacin and nitrofurantoin whereas showed highest resistance towards chloramphenicol, trimethoprim and levofloxacin. The present study revealed that Salmonellae contamination in raw chicken meat poses highest risk in transferring Salmonella infection to human beings. However hygienic practices at the poultry slaughtering units during the slaughtering of birds, use of clean water and frequent changing of water during washing of meat and other equipments including knives, chopping boards can decrease the contamination of Salmonella spp. up to a greater extent.
Open Access
Molecular Epidemiology of Methicillin Resistant Staphylococcus aureus(MRSA) from Humans and Bovines
(SKUAST Kashmir, 2019) Fajar Farooq; Rather, Mudasir Ali
The present study was undertaken to determine the occurrence, virulence gene profile and molecular epidemiology of Methicillin resistant Staphylococcus aureus (MRSA) from humans and bovines. Of the 200 samples, comprising those from healthy individuals (50), human clinical cases (50), clinically diseased animals (50) and healthy animals (50) screened for the presence of Staphylococcusaureus,149turned positive. The presumptive S. aureusisolates identified biochemically were further confirmed by targeting species specific thermostable nuclease (nuc) gene employing PCR and 112 isolates were confirmed as S. aureus (56%). The highest occurrence of S. aureuswas found in human clinical cases (76%) followed by clinically diseased animals (62%), healthy individuals (48%) and least in healthy animals (38%). The virulence gene profiling involved detection of delta hemolysin (hld), alpha hemolysin gene (hla) and panton-valentine leukocidin gene (pvl) genes of S. aureus by PCR. Of the 112 isolates, hldgene was detected in 49 (43.7%) isolates, hla gene in 13 (11.6%) andpvlgene in 7 (6.2%) isolates. The In-vitro antibiotic sensitivity profile revealed that, all the S. aureus isolates were highly sensitive to Gentamicin (95.5 %), Vancomycin (89.2%), Tetracycline (78.5%) and Oxacillin (54.6%). The highest resistance was noted against Methicillin (100%), Penicillin (94.6%) and Amoxycillin (92.8%). The study revealed 10.7% isolates to be multi drug resistant, majority among which were resistant against beta-lactam, Macrolide and Tetracycline class of antibiotics. The organism was analyzed for antibiotic resistance gene (mecA) and mecA was detected in 40 (35.7%) isolates. A higher proportion of isolates with mecA gene were recovered from human clinical cases (63.1%) and clinically diseased animals (38.7%), whereas the lesser occurrence of mecA gene was found in isolates from healthy humans (12.5%) and animals (5.2%). MRSA isolates were further subjected to SCC mectyping. Out of 24 mecApositive human clinical isolates 19 (79%) isolates were typablewhere as 5 (20.3%) were untypable; most encountered types were SCCmectype-I (33.3%) followed by, SCCmectype-V (29.1%), SCCmectype IVa (16.6%), however, none of the isolates harboured SCCmectype-II and SCCmectype-III. Similarly out of 12mecApositive isolates from clinically diseased animals 8 (66.6%) isolates were typablewhere as 4 (33.3%) isolates were untypable; highest occurrence was of SCCmectype V (41.6%) followed by, SCCmectype IV (16.6%), SCCmectype I (8.3%) and none of the isolates carried SCCmectype II and SCCmectype III. ThemecApositive isolates from healthy individuals and healthy animal were untypable.These non-typeable strains pose an important challenge in clinical settings emphasizing the molecular evolution of the SCC mecelement of emerging MRSA strains to adapt to environmental changes. Moreover, similar SCCmectypes in both clinical diseased animals and human clinical cases suggests co-circulation of MRSA isolates between human and animal population which was indication of possible lateral gene transfer between the Staphylococcal isolates in this region.
Open Access
Virulence and Antibiotic Resistance Gene Profile of Staphylococcus aureus from Milk, Milk Products and Environmental Samples of Kashmir Valley
(SKUAST Kashmir, 2018) Qazi, Irfan Ul Hassan; Rather, Mudasir Ali
The present study was undertaken to determine the prevalence, virulence gene profile, antibiogram and antibiotic resistance gene profile ofStaphylococcus aureus from milk, milk products and environmental samples from Kashmir Valley. Of the 210 samples, comprising of mastitis milk (50), milk products (40), raw milk (60) and environmental samples (60) screened for the presence of Staphylococcus aureus,48 samples were positive summing up a prevalence of 22.9. The presumptive S. aureusisolates identified biochemically were further confirmed by targeting species specific thermostable nuclease (nuc) gene employing PCR and all the 48 isolates were confirmed asS. aureus. The highest prevalenceof S. aureuswas found in the mastitis milk (34%), followed by milk products (22.5%), while both the environmental samples and raw milk showed the same percentage prevalence of 18.3. Molecular characterization of virulence factors encompassed aPCR involving detection of delta hemolysin (hld), toxic shock syndrome toxin (tsst-1) and enterotoxin (sea) genes of S. aureus. Of the 48 isolates, hldgene was detected in 46 (95.8%) isolates, tsst-1 gene in 8 (16.7%) isolates while as sea gene was not detected in any of the isolates. The highest occurrence of hld gene was found in isolates recovered from milk products (100%) and environmental samples (100%), followed by mastitis milk (94.1%), whereas the lowest occurrence was seen in raw milk (90.9%) samples. The highest occurrence of tsst-1 gene was seen in isolates recovered from milk products (77.8%), followed by raw milk (9.1%), whereas none of the isolates from mastitis milk and environmental samples tested positive for tsst-1 gene. In-vitro antibiotic sensitivity profile revealed that all the S. aureus isolates were highly sensitive to gentamicin (95.8%), vancomycin (83.3%), tetracycline (79.1%) and oxacillin (68.7%). The highest resistance was noted against methicillin (100%), penicillin (93.7%) and amoxicillin (93.7%). The study revealed 10.4% isolates to be multi drug resistant and majority among which were resistant against beta-lactam, macrolide and tetracycline class of antibiotics. The S. aureus isolates were tested for three antibiotic resistance genes, tet-M, blaZ and mecA responsible for resistance against tetracycline, penicillin and methicillin antibiotics, respectively. tet-M was detected in 25 (52.1%) isolates, blaZ in 11 (22.9%) and mecA in 8 (16.7%) isolates. The study also detected 5 (10.4%) isolates to be oxacillin susceptible - MRSA (OS-MRSA), which probably is the first reported case in Jammu & Kashmir state. The study revealed a high occurrence of S. aureus in milk and milk products posing a high risk to the consumers. The occurrence of virulence and antibiotic resistance genes in the isolates and the multiple antibiotic resistance along with MRSA and OS-MRSA isolates may aggravate the problem further, which could be alarming andmay pose a significant public health threat.
Open Access
Studies on Prevalence of Coliform Bacteria with Special Reference to Shiga-toxigenic Escherichia coli (STEC) in Market Milk and Milk Products
(SKUAST Kashmir, 2019) Malik, Faisal Nazir; Syed Akram Hussain
The present study was carried out to determine hygienic quality of milk and milk products, with special reference to coliform and E. coli count. Prevalence of E. coli including STEC was also determined by detecting virulent Shiga toxigenic (stx1 and stx2) genes in the isolated strain. Antibiotic resistance pattern of the STEC isolates were studied against a panel of antibiotics. The mean coliform counts for raw milk, packaged, milk, paneer, dahi, kulfi and rasmlie were 4.680±0.164 log10 cfu /ml, 2.403±0.299 log10 cfu/ml, 2.771±0.230 log10 cfu/g, 3.614±0.247 log10 cfu/g, 3.615±0.247 log10 cfu/g and 3.911±0.215 log10 cfu/g. In present study, 80% raw milk, 26% packaged milk, 44% paneer, 36% dahi, 50% kulfi and 54% of rasmalaie samples had coliform count above permissible limits. The mean E. coli count of raw milk, packaged milk, paneer, dahi, kulfi and rasmalie were 3.48±0.15 log10 cfu/ml, 2.19±0.34 log10 cfu/ml, 2.05±0.32 log10 cfu/g , 2.43±0.37 log10 cfu/g, 2.69±0.25 log10 cfu/g and 3.10±0.23 log10 cfu/g respectively. The overall prevalence of E. coli was found to be 24.3% with highest (62.0 %) in raw milk and lowest (10.0%) in dahi. The overall prevalence of STEC from milk and milk products was found to be 3.6%. A total of eight (08) isolates were found positive for stx1 gene, two (02) for stx2 gene and one (01) for both stx1and stx2 genes. However, isolates recovered from packaged milk and dahi did not carry either stx1 or stx2 genes. All the E. coli isolates including STEC were found complete (100%) resistant to ampicillin and sensitive (100%) to cefuroxime. The present study thus confirmed the presence of coliform organisms including STEC in raw milk and their products indicating a potential hazard. Emergence of multi drug resistant STEC strains as observed in current study also possess threat to public health.
Open Access
Occurrence, Molecular Detection and Antibiogram of Staphylococcus aureus from Table Eggs
(SKUAST Kashmir, 2016) Handoo, Sabah; Munshi, Ziaul Hassan
The present study was conducted to determine the Occurrence, Molecular Detection and Antibiogram of Staphylococcus aureus from Table Eggs in organised and unorganised sectors. A total of 240 samples collected from local organized sector (60), local unorganized sector (90) and organized sector imported from outside the state- (90) were screened for the presence of Staphylococcus spp., 115 samples were positive summing up to an overall percentage occurrence of 47.91. The highest Staphylococcal load was recorded in the eggs of organized sector imported from outside the state (57.77%) followed by local unorganized sector (47.77%) and the lowest contamination was seen in the eggs of local organized sector (33.33%). Out of 115, Staphylococcus spp. isolates, 84 (35%) isolates were recovered from egg shells and 31 (12.91%) from egg contents. Staphylococcus isolates those tested positive for biochemical tests including coagulase production, were presumptively considered as Staphylococcus aureus, were further confirmed by targeting species specific thermostable nuclease (nuc) gene employing PCR thereby confirming 48 isolates as S. aureus, indicating an percentage occurrence of 20. S. aureus showed the highest occurrence (28.88 %) in the eggs imported from outside the state, followed by eggs produced locally in organized sector (16.6%), while as the lowest occurrence was found in the eggs produced locally in unorganized sector (13.33%). Among the 48 S. aureus isolates, 20 (22.22%) were isolated from the samples of district Srinagar, 17 (18.88%) from Anantnag and 11 (18.33%) from Ganderbal indicating the highest contamination in eggs from Srinagar followed by Anantnag and Ganderbal, respectively. Of the 48 isolates of S. aureus, 37 (77.08%) were isolated from the egg shells and 11 (22.91%) from the egg contents. Molecular characterization of virulence factors encompassed a PCR involving detection of enterotoxin gene seb and tsst-1 of Staphylococcus aureus isolates. The enterotoxin type B gene (seb) was detected in 4 (8.33%) and toxic shock syndrome toxin-1 gene (tsst-1) was detected in 5 isolates (10.41%) In-vitro antibiotic sensitivity profile revealed that all the Staphylococcus aureus isolates were highly sensitive to levofloxacin (87.50%), chloramphenicol (85.41%) ofloxacin (68.75%), azithromycin (66.66%), and cefotaxime (60.41%) and tetracycline (60.41%). The highest resistance was noted against ceftazidime (100%), penicillin-G (87.5%), erythromycin (72.91%), ampicillin (81.25%) and oxytetracycline (66.66%). Five (5) isolates of Staphylococcus aureus were phenotypically shown to be resistant towards methicillin. All the phenotypically resistant isolates against methicillin showed the presence of methicillin resistant gene (mecA) using PCR. The study revealed a high occurrence of Staphylococcus aureus in eggs posing a high risk to the handlers and consumers. The occurrence of enterotoxin genes in the isolates and the multiple antibiotic resistant along with Methicillin resistant isolates aggravates the problem further which otherwise will pose a high public health threat.

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