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ThesisItem Open Access OCCURRENCE OF ENROFLOXACIN AND TETRACYCLINE RESISTANT KLEBSIELLA PNEUMONIAE AND SALMONELLA SPP. IN BEEF AND COW MILK(COLLEGE OF VETERINARY AND ANIMAL SCIENCES MANNUTHY,THRISSUR, KERALA VETERINARY AND ANIMAL SCIENCES UNIVERSITY, 2023-03-22) JISNA JAMAL P. P; Dr. C. LathaAntimicrobial resistance (AMR) is a global public health threat thatchallenges the scope and ability of modern medicine to effectively treat common bacterial infections. The present study was conducted for the isolation and identification of enrofloxacin and tetracycline resistant K. pneumoniae and Salmonella spp. from beef and raw cow milk samples. A total of 200 beef and 200 raw cow milk samples were collected from different retail outlets and milk societies of Thrissur and Palakkad districts. The samples were collected during a span of 10 months, from January 2022 to October 2022. The samples were subjected to conventional culture techniques followed by biochemical tests for the preliminary identification of the organisms. On analysis of milk samples from both districts, highest number of K. pneumoniae was obtained from Palakkad (86 per cent) district, followed by Thrissur (69 per cent) district. In the case of beef samples, 45 per cent and 30 per cent revealed the presence of K. pneumoniae from Thrissur and Palakkad districts, respectively. On statistical analysis, it was revealed that, the difference in occurrence of K. pneumoniae in milk samples from Thrissur and Palakkad districts were significant (p<0.01). Similarly, there was significant difference (p<0.05) between the occurrence of K. pneumoniae in beef samples from Thrissur and Palakkad districts. Among the 200 milk samples analysed, seven per cent of the samples from Palakkad district and five per cent of samples from Thrissur district were contaminated with Salmonella spp. A higher amount of Salmonella spp. was detected in beef samples from Thrissur (17 per cent) and Palakkad (12 per cent) districts. Statistical analysis revealed that there was no significant difference (p>0.05) in occurrence of Salmonella spp. in both milk and beef samples from Thrissur and Palakkad districts. On molecular confirmation, 91.03 per cent K. pneumoniae and all theSalmonella spp. isolates generated amplicons specific for rpoB and invA genes, respectively by PCR. None of the Salmonella spp. isolates in this study amplified virulence specific spvC gene on PCR. The confirmed isolates were subjected to phenotypic confirmation of antibiotic resistance by Kirby-Bauer disc diffusion test against 15 commonly used antibiotics. Multiple antimicrobial resistance (MAR) index was calculated for both K. pneumoniae and Salmonella spp. The MAR index for K. pneumoniae isolates analysed in this study ranged from 0.06 to 0.6, and that of Salmonella spp. showed a range of 0.06 to 0.33. The phenotypically enrofloxacin resistant isolates were subjected to Mismatch Amplification Mutation Assay (MAMA) PCR, to detect point mutations in gyrA and parC genes. The gyrAmutations were detected in 13 isolates of K. pneumoniae and three isolates of Salmonella spp. isolates. None of the isolates in this study showed parC mutations on MAMA PCR. The enrofloxacin resistant isolates were also screened for the presence of plasmid mediated quinolone resistant, qnrS gene. A total of 31 isolates, comprising 23 isolates of K. pneumoniae and eight isolates of Salmonella spp. amplified qnrS gene on PCR. Genotypic confirmations of tetracycline resistance in the isolates were done by targeting the tetracycline resistant genes tetA, tetB, tetO and tetX on PCR. The presence of tetA gene was detected in 58.7 per cent of K. pneumoniae isolates and 80 per cent of Salmonella spp. isolates. The gene tetB was detected in 43.48 per cent and 50 per cent of K. pneumoniae and Salmonella spp. organisms respectively. None of the isolates amplified tetO and tetX genes on PCR. The representative amplicons of the resistant genes were sequenced and submitted to National Center for Biotechnology Information (NCBI), and accession numbers were obtained. The results of the study revealed that beef and milk acts as an important reservoir of enrofloxacin and tetracycline resistant K. pneumoniae and Salmonella spp. This highlights an urgent need for the implementation of control strategies in management of food animals and handling of food products. In order to effectively control the emergence and spread of the resistance, inter-sectoral collaboration and a One Health Approach incorporating agriculture, animal husbandry, veterinary medicine, human medicine and environment science should be adopted. The use of antimicrobials in human and animals should be monitored and specific control policies which acknowledge the use of antimicrobials in food producing animals should be developed.ThesisItem Open Access IN VIVO EFFICACY OF GARLIC AND CHITOSAN FOR CONTROL OF CAMPYLOBACTER SPP. IN MEAT TYPE CHICKEN(COLLEGE OF VETERINARY AND ANIMAL SCIENCES MANNUTHY, THRISSUR, KERALA VETERINARY AND ANIMAL SCIENCES UNIVERSITY, 2023-03-17) KAVYA A; Dr. Deepa JollyCampylobacteriosis, a major foodborne gastroenteritis in humans, is acquired primarily through the consumption of contaminated poultry meat. Campylobacter jejuni (C. jejuni) and Campylobacter coli (C. coli) are the most frequently reported thermophilic Campylobacter spp. The present study was undertaken to assess the occurrence of Campylobacter spp. in meat type breeds by conventional and molecular techniques and to assess the in vivo anticampylobacter effect of garlic and chitosan in meat type chicken. A total of 120 cloacal swabs of different meat type breeds from the University Poultry and Duck Farm (UPDF), Mannuthy were subjected to isolation and identification of Campylobacter spp. byconventional culture techniques. Isolates were confirmed by multiplex Polymerase Chain Reaction for genus and species identification. The overall occurrence of Campylobacter spp. in the farm was 31.67 per cent with no significant difference in the occurrence in different meat type breeds. Of the isolates, 71.05 per cent were identified as C. jejuni and 28.95 per cent as C. coli. Two natural compounds viz., garlic [Fresh Garlic Extract (FGE) and Garlic Powder (GP)] and chitosan were analysed for in vitro anticampylobacter activity by determining the Minimum Inhibitory Concentration (MIC).The Standard strain C. jejuni NCTC 11168 and one of the confirmed C. jejuni isolate (wild strain) were found to be more susceptible to chitosan than garlic. In vivo study was undertaken in UPDF, Mannuthy with 78 day-old meat type chicks. Oral challenge was performed on the third day with the wild isolate of C. jejuni and six birds were randomly examined for Campylobacter colonisation, which revealed cent per cent colonisation on 20th day. Four treatments (groups of 12 birds) which included, T1-feed+ground fresh garlic, T2-feed+GP, T3-feed+chitosan, T4-feed+chitosan+GP, alongwith positive and negative controls, were initiated on 21st day. After first and second weeks of treatment, statistical analysis revealed a significant reduction in Campylobacter count in all treatment groups with the lowest reduction in T3 group. None of the treatments could completely eliminate the organism. Histopathological examination showed degeneration and shortening of villi in the caeca and inflammatory changes in the liver. Antibiotics resistance profile of the isolates against 14 commonly usedantibiotics revealed the presence of multidrug-resistant Campylobacter and all the isolates showed complete resistance to cefotaxime. The Study highlighted the importance of preharvest strategies in farms along with strict biosecurity and hygienic measures to reduce the Campylobacter load in poultry.ThesisItem Open Access DETECTION OF COXIELLA BURNETII (C. BURNETII) IN MILK AND VAGINAL SWABS OF GOATS WITH REPRODUCTIVE DISORDERS(COLLEGE OF VETERINARY AND ANIMAL SCIENCES MANNUTHY, THRISSUR, KERALA VETERINARY AND ANIMAL SCIENCES UNIVERSITY, 2023-03-23) M. BINDU PUSHPA; Dr. C. SethulekshmiThe present study was undertaken to detect Coxiella burnetii in milk and vaginal swabs of goats with reproductive disorders. A total of 450 samples which included milk (150), vaginal swabs (150) and blood (150) were collected from Thrissur, Palakkad and Malappuram districts. The samples were collected randomly from households, organised and unorganised farms of Thrissur, Palakkad and Malappuram districts. Milk and vaginal swab samples collected were subjected to extraction of DNA. The DNA was extracted from goat milk samples using phenol chloroform method. Extraction of DNA from vaginal swabs was carried out using Qiagen blood and tissue kit. The molecular confirmation of C. burnetii was done by targeting Com1 and IS1111 gene. The targeted Com1 and IS1111 genes could not be detected in any of the collected goat milk samples using PCR. The C. burnetii could not be detected in any of goat vaginal swab samples using PCR assay targeting Com1 and IS1111 genes. Blood samples were collected from goats of Thrissur, Palakkad and Malappuram districts were subjected to extraction of serum. Serum of goats were tested for antibodies against C. burnetii using commercial ELISA kit (Bio X Diagnostics). Out of 150 goat sera samples, 12 sera samples showed seropositivity for C. burnetti. Overall seroprevalence reported was 7.99 per cent, while highest was reported in Malappuram (four per cent) followed by Palakkad (2.66 per cent) and Thrissur (1.33 per cent) districts. A survey was conducted using well designed questionnaire among the households, organised and unorganised farms of Thrissur, Palakkad and Malappuram districts. Information regarding the occurrence of various reproductive disorders like abortions, stillbirths, repeat breeding in goats was collected. Economic loss occurred due to abortions, stillbirths and repeat breeding was calculated. From the survey it was noticed that elevated type of flooring was most preferred as the state is known for rains and it is labor friendly. Malabari and Attappady were two common goat breeds reared in all the three districts. Losses due to occurrence of abortions and stillbirths in organised and unorganised farms is more compared to households. Occurrence of various reproductive disorders in goats is due to both infectious (bacteria, virus, parasite) and non- infectious causes (environmental stress, nutritional deficiencies).One of the infectious causes for reproductive disorders in goats is C. burnetii. Inhealthy animal it is usually asymptomatic, but in pregnant animals it either causes abortions, stillbirths, placentitis or sheds in various excretions like milk, vaginal mucus, faeces urine and thereby spreading the disease to entire flock. In order to prevent C. burnetii in goats detailed knowledge of risk factors is necessary and implementation of one health approach in milk industry can ensure food safety.ThesisItem Open Access PHYTOCONTROL STRATEGIES USING SILVER NANOPARTICLES AND ACTIVATED CARBON AGAINST COLIFORMS IN WATER(COLLEGE OF VETERINARY AND ANIMAL SCIENCES MANNUTHY, THRISSUR, KERALA VETERINARY AND ANIMAL SCIENCES UNIVERSITY, 2023-03-23) MIYADA YOOSUF; Dr. Deepa JollyWater quality is one of the most important factors for a sustainable andhealthy ecosystem. The present study was undertaken to evaluate phytocontrol strategies using silver nanoparticles and activated carbon against coliforms in water. A total of 100 water samples were evaluated for coliforms and coliphages and its physico-chemical parameters. Samples included 85 well water samples received at the NABL accredited laboratory of the Department of Veterinary Public Health, CVAS, Mannuthy and 15 river water samples collected from various parts of Thrissur district. The samples were evaluated for total coliform count using Most Probable Number (MPN) method, which revealed 88 positive samples exceeding the WHO recommended limit of 10 MPN index/100 mL. These positive samples were subjected to isolation and identification of E. coli of which 57 E. coli positive samples were further subjected to isolation and identification of coliphages in water using Double Agar Layer Method. A total of five samples positive for coliphages were obtained and of these, the concentration of coliphages were more in river water samples (60 per cent) compared to well water (40 per cent). From the water samples positive for coliforms, six samples were selected at random for finding the efficacy of a bioadsorbent filter column which was prepared in the laboratory. This consisted of a layer of activated carbon sandwiched between sterilised sand layers within a Poly vinyl chloride (PVC) pipe. Two filters of two different diameters (20 mm and 25 mm) were assembled. The activated carbon used in the filter was made from the stems and leaves of cassava plant. All the water samples (unfiltered and filtered) were assessed for physico-chemical parameters and coliform count. The filter column of 20 mm diameter did not reduce the coliform count significantly but the filter with 25 mm diameter reduced the coliform count of the water. The layer thickness and flow rate of the filtered water were also estimated. The qualitative phytochemical screening and antibacterial effect of silver nanoparticles prepared from the hot aqueous extract of fresh neem leaves was also determined. The silver nanoparticles were characterised using X ray diffraction analysis which revealed a good yield of silver nanoparticles. The in vitro antibacterial activity of the synthesised silver nanoparticles revealed that silver nanoparticles have antibacterial activity and showed a zone of inhibition ranging from 6 mm to 16 mm against Gram negative standard culture of E. coli (MTCC 13706) and no activity against three PCR confirmed isolates. This study suggested that efficient control measures should be advocated to control the quality of our water resources.ThesisItem Open Access EPIDEMIOLOGICAL INVESTIGATION OF RISK FACTORS ASSOCIATED WITH OCCURRENCE OF LEPTOSPIROSIS IN DAIRY CATTLE(COLLEGE OF VETERINARY AND ANIMAL SCIENCES MANNUTHY, THRISSUR, KERALA VETERINARY AND ANIMAL SCIENCES UNIVERSITY, 2022-10-19) SRIJI A. P.; Dr. K. Vrinda MenonLeptospirosis is a neglected bacterial zoonotic disease with worldwide distribution posing significant threat to humans and animals. The present study was conducted to identify the risk factors associated with bovine leptospirosis in Pananchery panchayat and Irinjalakuda municipality in Thrissur district, Kerala. Blood and urine (102 each) samples from dairy cattle and water (50) samples were collected from both the study areas in pre and post-monsoon seasons. A survey was conducted at the time of sample collection in order to identify risk factors. Blood, urine and water samples were examined by Dark Field Microscopy (DFM) and were found to be negative. Microscopic Agglutination Test (MAT) of serum samples revealed seroprevalence of 25 per cent in pre-monsoon and 36.54 per cent in post-monsoon season from samples collected from Pananchery. In Irinjalakuda, 22 per cent and 28 per cent seroprevalence was observed in pre and post-monsoon seasons, respectively. The predominant serovars detected in both seasons were Hardjo (30 per cent in pre-monsoon and 28.95 per cent postmonsoon) and Grippotyphosa (16.67 per cent in pre-monsoon and 23.68 per cent in post-monsoon). Urine and water samples were subjected to PCR targeting 16S rRNA, lipl 21, lipl 32 and lipl 41 genes. On molecular analysis of urine by PCR, 15.38 per cent samples from Pananchery and 10 per cent samples from Irinjalakuda were found positive for leptospires in pre-monsoon. In postmonsoon, 23.08 per cent urine samples from Pananchery and 18 per cent from Irinjalakuda were detected by PCR. Pathogenic leptospires were detected in 5.88 per cent and 8.82 per cent of the urine samples in pre and post-monsoon, respectively. Analysis of water samples by PCR revealed that two per cent samples in pre-monsoon, 10 per cent of samples in post-monsoon were found to be positive without any significant difference between two study areas and two seasons. On molecular analysis, only six per cent water samples in post-monsoon revealed pathogenic leptospires. Initially, the risk factors were analysed by subjecting the data to univariate analysis (Chi-Square) for identifying risk factors associated with infection in both pre and post-monsoon seasons. The factors which were identified as significantly associated with leptospirosis in univariate analysis were then subjected to multivariate analysis. Using multivariate logistic regression analysis, possibility of contamination of grazing land with animal urine (OR=3.605; p<0.01) was identified as risk factor in pre-monsoon. In post monsoon, semi intensive and extensive system of rearing of animals (OR=6.493; p<0.01) and animals that used water sources shared by human and cattle for drinking (OR=2.487; p<0.05) were significantly associated with the presence of leptospirosis. The research revealed that apparently healthy cattle and water sources may lead to endemicity of disease in the area. Identification of risk factors will help to adopt proper intervention strategies to minimise the disease occurrence in endemic areas.ThesisItem Open Access ISOLATION AND DETECTION OF ANTIMICROBIAL RESISTANCE OF THERMOPHILIC CAMPYLOBACTER FROM LAYER FARMS AND CHICKEN PROCESSING LINES(COLLEGE OF VETERINARY AND ANIMAL SCIENCES, MANNUTHY, THRISSUR, KERALA VETERINARY AND ANIMAL SCIENCES UNIVERSITY, 2022-10-19) PRAVITHA C. P; Dr. Deepa JollyCampylobacteriosis is the leading cause of bacterial gastroenteritis worldwide. Chicken and other poultry species are the major reservoir for thermophilic Campylobacter spp. like C. jejuni and C. coli. The present study was undertaken to determine the occurrence of Campylobacter spp. in layer farms, chicken processing lines, molecular confirmation and antibiotic resistance profile studies of isolates, investigation of genotypic resistance pattern of isolates against tetracycline and ampicillin and evaluation of antibacterial efficacy of chitosan against Campylobacter spp. on chicken skin. A total of 260 samples, comprising of 120 cloacal swabs and 140 environmental samples were collected from two organised layer farms (F1 and F2), one each in Thrissur and Ernakulam districts. In order to establish the important contamination points of Campylobacter during slaughter, a total of 324 samples were analysed from various points of two poultry processing lines, SL1 and SL2. All the samples were subjected to isolation and identification of Campylobacter spp. by conventional culture technique. Genus confirmation and species level identification of the isolates was done by polymerase chain reaction. The overall occurrence of Campylobacter spp. in layer farms F1 and F2 was 20 per cent and 15.4 per cent, respectively. Upon comparing the contamination points from poultry processing lines, contamination in SL1 (37.65 per cent) was found to be higher than SL2 (15.4 per cent). In both slaughterlines, Campylobacter positive status of the birds brought for slaughter was the prime source for C. jejuni contamination. The presence of the organism which increased during scalding, defeathering and evisceration steps, decreased after final washing and chilling of the carcass. Antibiotic resistance profiling revealed all isolates to be resistant to cefipime, cefixime, ceftazidime and cefuroxime. The genes conferring tetracycline and ampicillin resistance, tetO and blaOXA-61 were found among 73.3 per cent and 42.8 per cent of the isolates. Upon statistical analysis, no significant reduction of Campylobacter counts on chicken skin was observed after treatments with different concentrations of chitosan (0.5 per cent, 1 per cent or 2 per cent). The results of present study signify the prevalence of C. jejuni, a zoonotic pathogen, in the layer farms and poultry processing environment of central Kerala. Appropriate biosecurity measures and processing interventions along the poultry production chain is essential for the effective control of this pathogen.ThesisItem Open Access EFFICACY OF CAMPYLOPHAGES AND SELECTED PHYTOCHEMICALS IN THE CONTROL OF CAMPYLOBACTER BIOFILMS(COLLEGE OF VETERINARY AND ANIMAL SCIENCES MANNUTHY, THRISSUR, KERALA VETERINARY AND ANIMAL SCIENCES UNIVERSITY, 2023-03-23) VIVEKANANDHAN R; Dr. B. SunilCampylobacteriosis is one of the most common causes of bacterial foodborne disease. Campylobacter spp. are found as commensals in the intestines of a wide variety of animals, particularly pigs utilised for food production. The present study was undertaken to analyse the efficacy of campylophages and selected phytochemicals in the control of campylobacter biofilms. A total of 130 samples including 40 samples each of faecal and rectal swabs and sewage (n=50) were collected from the two unorganised pig farms. An overall occurrence of 26.15 per cent with higher isolation from rectal swabs (57.5 per cent) than faecal and sewage samples (25 per cent and two per cent) were observed. In both the farms, no effective biosecurity measures were followed. In pork processing line, a total of 80 samples including 20 each of carcass swabs, cutting board swabs, knife swabs and sewage samples were collected. An overall occurrence of 12.5 per cent was observed. All the isolates were subjected to biofilm forming ability at three different temperatures 42℃ (10 per cent CO2 condition), 37℃ & 25℃ (aerobic condition) and compared with two media (Dulbecco’s Modified Eagle’s Medium (DMEM) & Muller Hinton Broth (MHB)). Strong biofilm formers were predominant in aerobic condition at 37℃& 25℃ incubated in MHB. Weak biofilm formers were more on stainless steel and nylon fibre. Carvacrol, trans-cinnamaldehyde, eugenol, ethanolic extract of citrus by-product and bleaching powder had minimum biofilm inhibition concentration of 0.078 mg/mL, 0.156 mg/mL, 0.156 mg/mL, 0.321 mg/mL and 0.625 mg/mL, respectively. Trans-cinnamaldehyde significantly (p<0.001) inhibited 99.39 per cent biofilm formation by the campylobacter isolates on polystyrene surface at MBIC of 0.156 mg/mL. Citrus by-product ethanolic extract significantly (p<0.001) inhibited 99.43 per cent biofilm formation by the campylobacter isolates on stainless steel surface at MBIC of 0.312 mg/mL.Carvacrol significantly (p<0.001) inhibited 97.80 per cent biofilm formation by the campylobacter isolates on nylon fibre surfaces at MBIC of 0.078 mg/mL. Carvacrol, trans-cinnamaldehyde, eugenol, ethanolic extract of citrus by-product and bleaching powder had minimum biofilm inactivation concentration of 0.312mg/mL, 2.5 mg/mL, 1.25 mg/mL, 2.5 mg/mL and 5 mg/mL, respectively. Trans cinnamaldehyde inactivated 21.49, 57.89 and 95.92 per cent pre-formed biofilms by campylobacter isolates after 2, 5, 10 min of exposure, respectively at MBIAC of 2.5 mg/mL on polystyrene surface. Eugenol significantly (p<0.001) inactivated 53.79, 70.87 and 98.90 per cent pre-formed biofilms by campylobacter isolates after 2, 5, 10 min of exposure, respectively on stainless steel surface at MBIAC of 1.25 mg/mL. Trans-cinnamaldehyde significantly (p<0.001) inactivated 87.71, 93.9 and 99.15 per cent pre-formed biofilms by campylobacter isolates after 2, 5, 10 min of exposure, respectively on nylon fibre surface at MBIAC of 2.5 mg/mL. Three campylophages were isolated from sewage samples of farm B. Phage 2 and 3 were characterised as Siphoviridae and phage 5 was as Myoviridae by transmission electron microscope. Phage 5 inactivated 30.3 per cent, 30.55 per cent and 29.60 per cent pre-formed biofilms of campylobacter isolates on polystyrene, stainless steel and nylon fibre surface after 24 h of exposure, respectively. Campylophages were effective in inactivating pre-formed biofilms by the isolates which were lysed by the phages. Campylophages were not effective in inactivating biofilms formed by the campylobacter isolates which did not show lytic activity. Cost analysis showed citrus by-product extract as cost effective (Rs 0.41/L) than other phytochemicals.ThesisItem Open Access DETECTION OF COLISTIN AND QUINOLONE RESISTANT SALMONELLA ENTERICA FROM POULTRY PROCESSING LINE(COLLEGE OF VETERINARY AND ANIMAL SCIENCES MANNUTHY, THRISSUR, KERALA VETERINARY AND ANIMAL SCIENCES UNIVERSITY, 2022-02-10) RADHIKA A. N.; Dr. Binsy MathewThe present study was undertaken to detect colistin and quinolone resistant Salmonella enterica from poultry processing lines. A total of 450 samples were collected which included 75 samples each of cloacal swabs, caecal contents and carcass rinsates from two processing plants viz., Plant A, Thrissur and Plant B, Ernakulam. The samples were subjected to isolation of Salmonella by conventional culture technique and Salmonella were detected in 14.22 per cent of the samples. There was significant difference (p < 0.05) in the occurrence of Salmonella spp. in cloacal swabs and carcass rinsates of two processing plants. The molecular confirmation of positive isolates targeting invA gene for Salmonella genus and iroB gene for Salmonella enterica species was detected in 67.19and81.4per cent of isolates, respectively. The Salmonella enterica confirmed isolates were subjected to antibiotic sensitivity against quinolones by disc diffusion method and colistin agar test for colistin; wherein, highest resistance was observed to enrofloxacin and levofloxacin, whereas 8.57 per cent of the isolates were resistant to colistin. For molecular confirmation of quinolone resistance mismatch amplification mutation assay (MAMA) PCR was employed to detect mutations in gyrA and parC genes, 15.15 per cent of isolates had mutations in gyrA gene but none of the isolates had mutations in parC gene. The qnrS gene was present in 18.18 per cent of the isolates. However, none of the isolates harboured oqxAB gene. For molecular confirmation of colistin resistance,mutations in pmrA and pmrB genes were analysed by sequencing; out of three resistant isolates, two isolates viz., pmrA1 and pmrA2 had mutations in pmrA gene at 345th and 346th position, respectively. Further, two pmrB harbouring isolates showed mutation in 79th, 244th, 319th, 388th and 577th position of the sequence. All the representative amplicons of the genes were sequenced and submitted to the NCBI. The results of the study revealed the occurrence of antibiotic resistant Salmonella enterica throughout the poultry or chickenprocessing lines. In order to prevent antibiotic resistant Salmonella contamination in the processing line elucidating risk factors is necessary and implementation of one health approach in the meat production so as to ensure food safety to the consumers.ThesisItem Open Access ASSESSMENT OF POST HARVEST CONTAMINATION OF SEAFOOD WITH DRUG RESISTANT PATHOGENS OF PUBLIC HEALTH SIGNIFICANCE IN WAYANAD DISTRICT(COLLEGE OF VETERINARY AND ANIMAL SCIENCS, POOKODE, WAYANAD, KERALA VETERINARY AND ANIMAL SCIENCES UNIVERSITY, 2023-01-13) ANJUSHA.K.M; Dr. PrejitAntimicrobial resistance is a global public health emergency that causes existing antibiotics to fail therapeutically and raises healthcare costs among humans, animals, and aquaculture sectors. The presence of antimicrobial-resistant pathogens such as Escherichia coli, Salmonella spp., and Staphylococcus aureus in retail fish outlets and their environments acts as a potential source of transmission of AMR genes to humans. Hence, the present study aims to determine the occurrence of the above pathogens from different fish markets of the Wayanad district and to assess the prevalence of carbapenem-resistant and Extended- spectrum beta-lactamase (ESBL) -producing E. coli, Salmonella spp, and methicillin-resistant S. aureus (MRSA). A total of 216 samples, comprising finfishes (60), crustaceans (60), and molluscans (60) and 36 market environmental (hand wash, surface swab, and ice) samples, were analyzed for the presence of E. coli, Salmonella spp., and S. aureus by conventional microbiological and molecular methods. The prevalence was observed to be 79.63, 12.03, and 67.59 per cent, respectively. Furthermore, the characterization of isolates for ESBL production through the phenotypic disc diffusion method confirmed 66.27 per cent of E. coli isolates and 65.38 per cent of Salmonella spp. isolates as ESBL producers, and by PCR-based genotypic characterization of ESBL revealed 66.27 per cent of E. coli and 65.38 per cent of Salmonella spp. isolates harbouring at least one of the selected ESBL genes, i.e., blaCTX-M, blaTEM, or blaSHV. The ascending order of carbapenem sensitivity of the isolates upon phenotypic assay was imipenem, ertapenem, doripenem, and meropenem for E. coli, whereas the order was imipenem, ertapenem, meropenem and doripenem in the case of Salmonella spp. Carbapenemase genotypic characterization using PCR revealed that E. coli was positive for genes viz., blaOXA (14.53%), blaNDM (6.97%), and Salmonella spp. was positive for blaOXA (3.84 %), blaNDM (3.84%), with none of the isolates positive for the blaIMP gene. The characterization of isolates for MRSA production by the phenotypic disc diffusion method showed more resistance towards oxacillin (11.64%), followed by cefoxitin (5.48 %), and a genotypic assay revealed that 5.48 percent of isolates were found to harbor the mecA gene. The present study confirmed the prevalence of drug-resistant E. coli, Salmonella spp., and S. aureus in retail fish outlets and their environment. Hence, the study recommends retailers adopt stringent hygiene practices to prevent post-harvest contamination of fish.
