Veterinary Research Forum

Abstract This study was important to improve proper biosecurity measures and controlling the spread of Aeromonas to prevent future outbreaks. This research sought to determine whether virulent Aeromans species were present in morbid rainbow trout, their resistance and their genetic relatedness. A total number of 542 tissue lesion specimens were collected from gill, liver, heart and kidneys in morbid domesticated fish in Duhok province, Iraq. The gyrB DNA sequence analysis was used to determine the species classification. Drug susceptibility testing was conducted for all isolated strains using disc diffusion technique. The genotyping analysis was carried out using enterobacterial repetitive intergenic consensus-polymerase chain reaction. Thirty-four isolates were found and they were classified into three species (Aeromonas veronii, Aeromonas sorbia, and Aeromonas allosaccharophila), where A. veronii stand as one of the most prevalent species. The most frequently affected organ by Aeromonas was the gills among four different organs. The detection frequencies of the virulence genes aerolysin, outer membrane protein, glycerophospholipid-cholesterol acyltransferase, elastase, flagella, serine protease, cytotonic heat-labile, and hemolysin were 100%, 100%, 79.41%, 64.70%, 76.47%, 67.64%, 70.58%, and 41.17, respectively. None of the strains possessed all of the virulence markers. All isolates were completely resistant to ceftazidime, amoxicillin and doxycycline. All isolates were found to be multi-drug-resistant. Regardless of the nearest geographic source area of samples and the same Aeromonas species, there was a high genetic diversity. The results of this study could help farmers and researchers make informed decisions about measures of biosecurity and proper therapeutic drugs to apply to prevent current outbreaks and prevent them from recurring again.

Abstract The most serious problem in public health is salmonellosis, a common disease in horse. The aim of this study was to investigate the shedding of Salmonella serotypes in healthy Caspian pony. We examined 143 pony's fecal samples collected from the north of Iran belonging to different ages and sexes. Samples were cultured, then identification of isolates were performed by common bacteriological methods and polymerase chain reaction (PCR). The PCR was also used to explore the presence of fimA and salmonella secreted effector L (SseL) genes as virulence factors in the isolates and all were assigned to antibiotic susceptibility test via disc diffusion method. Results showed two fecal samples (1.39%) contaminated with Salmonella and further examination demonstrated the isolates belonging to S. enterica serotype typhimurium. Both serotypes were isolated from female and ˂6 years of age group of ponies and we detected fimA and SseL genes in the isolates. Observing multiple drug resistance and virulence genes in isolates is of utmost importance from both clinical and public health perspectives. It is highly likely that we face instances of salmonellosis in animals or humans that lead to severe infections and fail to respond to treatment in future. This study revealed that the occurrence of Salmonella was low in ponies, however, regarding the presence of virulence factors with multidrug resistant trend in this zoonotic bacterium, establishment of good hygienic measurement to prevent the transmission of bacteria between animal and human is necessary. 

Abstract Staphylococcus aureus is gaining worldwide attention because of its substantial impact on public health. The current study aimed to characterize S. aureus strains isolated from wild birds in the Kasur district of Punjab, Pakistan from 2021 to 2022. A total of one hundred samples were collected from five wild bird species. The samples were enriched, inoculated on selective agars and cultured for 24 hr at 37.00 ˚C. All isolates were verified by matrix-assisted laser desorption ionization–time-of-flight mass spectrometry (MALDI-TOF MS) and polymerase chain reaction (PCR) after Gram staining. Positive isolates were screened for phenotypic (Kirby-Bauer disk diffusion and minimum inhibitory concentration s), genotypic antibiotic resistance, and virulence genes. These samples yielded 30 (30.00%) S. aureus isolates, confirmed by polymerase chain reaction utilizing the 16S rRNA gene. Staphylococcus aureus was more prevalent in cloacal samples (16.00%) than oral samples (14.00%). Various S. aureus isolates showed varying degrees of resistance to three different antibiotics. Oxacillin (56.66%; n = 17) and tetracycline (33.33%; n = 10) showed the highest resistance rates with the lowest susceptibility (43.33%; n = 13). In contrast, vancomycin, rifampicin, linezolid, and daptomycin were 100% susceptible. Further disc diffusion study revealed resistance to tetracycline (33.33%), erythromycin (16.66%), and gentamicin (10.00%). The tetK gene was found in 33.33% of wild bird samples, while the ermA gene was found in 16.66% of samples. The aacA-D gene was only found in three (10.00%) isolates. None of the isolates tested positive for virulence genes. In conclusion, S. aureus is carried by wild birds in this area, posing a potentail threat to both humans and animals.

Abstract Russian sturgeon (Acipenser gueldenstaedtii) is an endangered fish species and also an important resource for the sturgeon aquaculture industry in Turkiye. Recently, a fatal and persistent bacterial disease occurred in the reared sturgeon kept in a trout farm in Turkiye. The disease outbreak has been with notable external signs including petechial hemorrhages and systemic anemia. This outbreak lasted for six weeks, and cumulative mortality reached around 35.00 - 40.00%. In this study, no parasitic and viral agents were observed in the sturgeons. Citrobacter gillenii was isolated from the diseased fish and identified by biochemical and molecular methods including API 20E and 20NE and 16S rRNA gene region sequencing, respectively. As a result, C. gillenii was identified for the first time in Russian sturgeon in Turkiye. The sequence was also deposited under the Genbank with MW057770 accession number. According to the result of disc diffusion method, bacteria were sensitive to enrofloxacin, streptomycin, amoxicillin and oxytetracycline and resistant to penicillin, trimethoprim/sulfamethoxazole, florfenicol and erythromycin. Also, ampC, sul1 and floR resistance genes were detected in the isolated bacteria. The results of this study provide important information for the diagnosis and treatment of this newly emerged disease of Russian sturgeon.

Abstract Campylobacter jejuni and C. coli are the main causes of gastrointestinal diseases in humans even in industrialized countries affecting public health. The aim of the current study was to evaluate the occurrence and antibiotic resistance of C. jejuni and C. coli in chicken meat, beef, mutton and water buffalo meat slaughtered in Ahvaz city, Iran. A total of 380 samples including chicken meat from industrial abattoirs (n = 150), chicken meat from traditional abattoirs (n = 50), fresh packed chicken meat from local markets (n = 30) and beef, mutton and water buffalo meat from industrial abattoirs (50 samples for each meat) in Ahvaz,were collected and tested for the presence of Campylobacter spp. The procedure was one-step enrichment in Preston enrichment broth followed by plating on supplemented blood agar for 24 hr under microaerophilic conditions at 42 ˚C. Suspected colonies were tested by polymerase chain reaction assay and susceptibility of the confirmed isolates to various antibiotics was investigated by the Kirby-Bauer disk diffusion method. Overall, 32 samples (8.40%) were contaminated with Campylobacter spp. Mutton was the most contaminated meat (24%), while fresh packed chicken meat were not contaminated. Among the 32 isolates, 40.60%, 34.40%, 21.90%, and 15.60% were resistant to tetracycline, ciprofloxacin, ampicillin, and streptomycin, respectively. Moreover, a high number of multi-antibiotic resistant Campylobacter spp. was determined. Since foods of animal origin are the most sources of Campylobacter infection, the presence of resistant strains to antibiotics is a potential risk to public health.

Abstract Althoughpoultry meat is considered as the main source for human Campylobacter infections,there is limited information about non-poultry sources. The present study was aimed to investigate the prevalence and the antibiotic resistance of thermophilic Campylobacter spp. in fecal samples of the cattle and sheep in Shiraz, Iran. A total of 302fecal samples were obtained from clinically healthy, slaughtered cattle and sheep from Shiraz slaughterhouse. The animals were clinically healthy before being slaughtered. The samples were cultured according to the specific cultivation method under thermophilic conditions. The susceptibility of Campylobacter isolates were determined for 13 antimicrobial agents. All enriched samples and cultured isolates were targeted for polymerase chain reaction (PCR) detection of 16S rRNA and multiplex PCR for determining their species. Among 302 fecal samples, 65 (21.5%) and 205 (67.8%) samples were positive for the presence of Campylobacter species with the cultivation and PCR techniques, respectively. All 65 distinct isolates were susceptible to neomycin and colistin and the isolates showed high resistance to cephalotin (83.0%) and ciprofloxacin (67.7%). After the multiplex PCR, 78.5% of total positive samples showed the simultaneous presence of Campylobacter jejuni and Campylobacter coli. In conclusion, the results emphasized that non-poultry farms are important as a possible source of Campylobacter infections.

Abstract Staphylococcus aureus is generally regarded as a leading cause of mastitis in dairy cattle. The aim of this study was to investigate the pattern of agr groups and any possible relationship between agr groups and antibiotic resistance among S. aureus strains isolated from bovine mastitis in Northeast of Iran. For this purpose, a total of 300 bovine mastitic milk samples were taken from dairy industry farms of Khorasan Razavi Province, Iran. S. aureus were isolated and identified according to the standard methods. Antibiotic susceptibility testing was conducted by disk diffusion method. In this study a total of 31 isolates of S. aureus were evaluated for agrD gene polymorphism by specific primers. Most of the isolates belonged to agr group I (54.8%), followed by agr group III (25.8%) and agr group II (19.4%). There was not any isolates belonging to group IV. Resistance to methicillin in agr group I isolates was more than other groups. Agr groups II and III were quite susceptible to methicillin. Due to high prevalent of S. aureus isolates and high antibiotic resistance rate in bovine mastitic isolates, it is important to verify the characteristics of S. aureus strains in Iran.

Abstract The aim of this study was to determine the occurrence of enterotoxigenic and enteroaggregative Escherichia coli strains and antibiotic resistance of the isolates in raw milk and unpasteurized cheese. Out of 200 samples of raw milk and 50 samples of unpasteurized cheeses, 96 and 24 strains of E. coli were isolated, respectively. Polymerase chain reaction (PCR) was used to detect the genes encoding heat-stable enterotoxin a (STa), heat-stable enterotoxin b (STb), heat labile toxin (LT) and enteroaggregative heat-stable toxin1 (EAST1). Twelve out of 120 (10.00%) isolates harbored the gene for EAST1, 2(1.66%) isolates were detected as producing STb and LT  toxins and 12 (10.00%) strains contained STb and EAST1 genes.  None of the strains contain the STa gene. All of the strains were tested for antibiotic resistance by disk diffusion method. Disks included: ciprofloxacin (CFN), trimetoprim-sulfamethoxazole (TSX), oxytetracycline (OTC), gentamicin (GMN), cephalexin (CPN), nalidixic acid (NDA) and nitrofurantoin (NFN), ampicillin (AMP), neomycin (NEO) and streptomycin (STM).  Among 120 isolated strains of E. coli, the resistance to each antibiotics were as follows: OTC100%, CPN 86.00%, NDA 56.00%, NFN 42.00%, GMN 30.00%, TSX 28.00%, CFN 20%, AM 23.40% and STM 4.25%. None of the isolates were resistant to NEO. The present data indicate that different resistant E. coli pathogens may be found in raw milk and unpasteurized cheese. It poses an infection risk for human and transferring the resistant factors to microflora of the consumers gut.