Molecular Characterization of Multidrug-Resistant and Extended-Spectrum ß-Lactamases-Producing Salmonella enterica Serovars Enteritidis and Typhimurium Isolated from Raw Meat in Retail Markets.

In the present study, a total of 720 samples were collected from retail raw meat from 13 upazilas in Sylhet District, Bangladesh, of which 225 samples were from cattle meat, 210 samples were from goat meat, and 285 samples were from chicken meat. Salmonella enterica serovars Typhimurium and Enteritidis were screened for extended-spectrum ß-lactamase (ESBL) genes using multiplex PCR. Among the 720 samples, Salmonella spp. was detected in 28.06% (202 out of 720) of the samples, with S. Enteritidis and S. Typhimurium were identified in 11.53% (83 out of 720) and 12.22% (88 out of 720) of the samples, respectively. It was found that all Salmonella enterica serovars isolated from cattle meat displayed multidrug resistance (MDR) based on antimicrobial susceptibility testing. Notably, a significant proportion of S. Enteritidis isolates and all S. Typhimurium isolates from goat meat demonstrated complete resistance to multiple drugs (ampicillin, cefuroxime, and ceftazidime). Regarding chicken meat, out of 89 isolates encompassing both S. Typhimurium and S. Enteritidis, 57 isolates (64.04%) exhibited MDR. Additionally, blaCTX-M-1 exhibited the highest occurrence at 15.69% for S. Typhimurium and 7.89% for S. Enteritidis in chicken meat. Moreover, blaCTX-M-9 was only detected at 3.92% for S. Enteritidis in chicken meat. Furthermore, blaOXA had the highest prevalence rate of 19.04% for S. Enteritidis and 25.80% for S. Typhimurium in cattle meat, followed by chicken meat. These findings highlight the urgency for monitoring ESBL-producing Salmonella in retail raw meat and the need for strict measure to manage antibiotic use to prevent the spread of multidrug-resistant and ESBL-producing Salmonella strains, thereby protecting humans and reducing public health risks.

Exploring of spectrum beta lactamase producing multidrug-resistant Salmonella enterica serovars in goat meat markets of Bangladesh.

The emergence of Salmonella enterica serovars that produce extended-spectrum beta-lactamase and exhibit multi-drug resistance (MDR) poses a substantial global threat, contributing to widespread foodborne illnesses and presenting an alarming issue for public health. This study specifically concentrated on the isolation and identification of ESBL-resistant genes (bla TEM, bla SHV, bla CTX-M1, bla CTX-M2, bla CTX-M9, MultiCase ACC, MultiCase MOX, MultiCase DHA, bla OXA) and the antibiogram profiling of Salmonella enterica serovars found in goat meat samples procured from retail outlets in Bangladesh. During the research in the Sylhet district of Bangladesh, researchers gathered a total of 210 samples of goat meat from 13 different Upazilas. Primarily, cultural and biochemical methods were used for isolation of bacteria from the selected samples. Salmonella enterica serovars Typhimurium and Enteritidis, along with three ESBL-resistant genes, were identified through polymerase chain reactions (PCRs). The disk diffusion test was used to determine antimicrobial susceptibilities. Out of 210 samples analysed, Salmonella spp. was detected in 18.10 % (38 out of 210), with S. Enteritidis and S. Typhimurium found in 9.05 % (19 out of 210) and 5.24 % (11 out of 210) of the samples, respectively. A total of 72.73 % (8/11) of S. Enteritidis and 100 % (19/19) of S. Typhimurium isolates were positive by Multidrug-resistant patterns. The positive outcomes were found of S. Typhimurium tested 63.16 % (12 out of 19) for the bla TEM gene and 21.05 % (4/19) for the bla SHV, gene. The study proposes that the retail goat meat market channel could be a prominent transmission way of ESBL-producing MDR Salmonella enterica serovars, representing a significant public health hazard.

Investigating the infection dynamics and molecular detection of Cryptosporidium in Buffaloes in Sylhet, Bangladesh.

Cryptosporidiosis, a zoonotic infection impacting both livestock and humans, is inadequately understood in terms of its prevalence and transmission dynamics involving buffaloes in Bangladesh. This research, conducted in the Sylhet division, aimed to explore the prevalence and potential risk factors influencing Cryptosporidium spp. in the faecal samples of 392 buffaloes. Detection of the parasite utilized modified Ziehl-Neelsen staining, with molecular identification achieved through nested PCR (nPCR). The comprehensive analysis revealed 9.18% (36/392) prevalence at the individual animal level and 40.48% (17/42) at the herd level. Age-based analysis revealed fluctuating infection rates of Cryptosporidium spp. in buffaloes across distinct age brackets, with rates of 22.61% in those aged 0-6 months, 5.00% in those aged 6-12 months, and 1.03% in those aged 12-18 months. Diarrheic buffaloes showed a significantly (p < 0.001) higher infection rate (26.67%; 28/105) compared to non-diarrheic buffaloes (2.79%; 8/287). In risk factor analysis, binary logistic regression revealed that buffaloes aged 0-6 months were experiencing a likelihood that is 14.84 times higher to be affected by Cryptosporidium in contrast to their older counterparts (OR = 14.85; p = 0.02). Additionally, diarrhoeic buffaloes were found to be more susceptible to Cryptosporidium compared to healthy buffaloes (OR = 17.50; p < 0.001). A higher stocking density was associated with an increased likelihood of infection in buffaloes (OR = 11.20; p = 0.01). The results of this study emphasize the necessity for targeted interventions, considering factors like diarrheic condition and stocking density, to effectively manage and control cryptosporidiosis in Bangladesh.

Antibacterial potential of lactic acid bacteria isolated from raw cow milk in Sylhet district, Bangladesh: A molecular approach.

BACKGROUND: The most prevalent probiotic bacterium employed in the food industry is Lactobacillus because it can produce metabolites with antibacterial capabilities and exhibits hostility towards infections and microorganisms that cause spoilage. AIM: This study set out to identify naturally occurring Lactobacillus and plantaricin (pln EF) coding genes in raw cow milk and to assess the antibacterial potency of isolated Lactobacillus isolates. METHODS: Following enrichment in De Man, Rogosa and Sharpe (MRS) broth, single colonies were isolated, and pure colonies were obtained by streaking on MRS agar. The 16S rRNA gene was amplified using polymerase chain reaction (PCR) to confirm the cultural positivity of all isolates. Additionally, the presence of plantaricin was verified by targeting the pln EF gene through PCR. OUTCOME: Out of the 166 raw milk specimens acquired from cows, 153 (91.17%; CI: 86.98-95.76) were identified as positive for Lactobacillus through both culture and biochemical screening. Subsequently, 121 (72.89%; CI: 65.46-79.49) of the isolates were affirmed to harbour Lactobacillus through PCR analysis. Within this subset, 6 isolates (4.96%; CI: 1.84-10.48) were found to possess the plnEF gene. When exposed to Lactobacillus isolates, Salmonella Typhimurium and Salmonella enterica displayed an average maximum zone of inhibition with a diameter measuring 24 mm. In contrast, Escherichia coli exhibited an average minimum zone of inhibition, featuring a diameter of 11 mm. Additionally, the Lactobacillus isolates demonstrated inhibitory zones against Staphylococcus aureus, Klebsiella pneumoniae and Klebsiella oxytoca, measuring 14, 22 and 19 mm, respectively. CLINICAL SIGNIFICANCE: Lactic acid bacteria, particularly Lactobacilli, are plentiful in cow milk and possess broad-spectrum antibacterial properties.

Molecular Detection of Colistin Resistance mcr-1 Gene in Multidrug-Resistant Escherichia coli Isolated from Chicken.

Zoonotic and antimicrobial-resistant Escherichia coli (hereafter, E. coli) is a global public health threat which can lead to detrimental effects on human health. Here, we aim to investigate the antimicrobial resistance and the presence of mcr-1 gene in E. coli isolated from chicken feces. Ninety-four E. coli isolates were obtained from samples collected from different locations in Bangladesh, and the isolates were identified using conventional microbiological tests. Phenotypic disk diffusion tests using 20 antimicrobial agents were performed according to CLSI-EUCAST guidelines, and minimum inhibitory concentrations (MICs) were determined for a subset of samples. E. coli isolates showed high resistance to colistin (88.30%), ciprofloxacin (77.66%), trimethoprim/sulfamethoxazole (76.60%), tigecycline (75.53%), and enrofloxacin (71.28%). Additionally, the pathotype eaeA gene was confirmed in ten randomly selected E. coli isolates using primer-specific polymerase chain reaction (PCR). The presence of mcr-1 gene was confirmed using PCR and sequencing analysis in six out of ten E. coli isolates. Furthermore, sequencing and phylogenetic analyses revealed a similarity between the catalytic domain of Neisseria meningitidis lipooligosaccharide phosphoethanolamine transferase A (LptA) and MCR proteins, indicating that the six tested isolates were colistin resistant. Finally, the findings of the present study showed that E. coli isolated from chicken harbored mcr-1 gene, and multidrug and colistin resistance. These findings accentuate the need to implement strict measures to limit the imprudent use of antibiotics, particularly colistin, in agriculture and poultry farms.

In Silico Investigation of Conserved miRNAs and Their Targets From the Expressed Sequence Tags in Neospora Caninum Genome.

Neospora caninum is a protozoan parasite, the etiologic agent of Neosporosis-a common cause of abortion in cattle worldwide. Herd level prevalence of Neosporosis could be as high as 90%. However, there is no approved treatment and vaccines available for Neosporosis. MicroRNA (miRNA) based prophylaxis and therapeutics could be options for Neosporosis in cattle and other animals. The current study aimed to investigate the genome of Neospora caninum to identify and characterize the conserved miRNAs through Expressed Sequence Tags (ESTs) dependent homology search. A total of 1,041 mature miRNAs of reference organisms were employed against 336 non-redundant ESTs available in the genome of Neospora caninum. The study predicted one putative miRNA "nca-miR-9388-5p" of 19 nucleotides with MFEI value -1.51 kcal/mol and (A + U) content% 72.94% corresponding with its pre-miRNA. A comprehensive search for specific gene targets was performed and discovered 16 potential genes associated with different protozoal physiological functions. Significantly, the gene "Protein phosphatase" was found responsible for the virulence of Neospora caninum. The other genes were accounted for gene expression, vesicular transport, cell signaling, cell proliferation, DNA repair mechanism, and different developmental stages of the protozoon. Therefore, this study finding will provide pivotal information to future aspirants upon Bovine Neosporosis. It will also serve as the baseline information for further studies of the bioinformatics approach to identify other protozoal miRNAs.

Molecular detection and phylogenetic analysis of Canine Parvovirus (CPV) in diarrhoeic pet dogs in Bangladesh.

Since its emergence, Canine Parvovirus type 2 (CPV-2) has been considered as a deadly pathogen in dogs with high mortality in puppies for its clinical gastroenteritis and severe haemorrhagic diarrhoea. Although several studies on CPV-2 were conducted in Bangladesh, molecular investigation is poorly understood. The aim of the study was molecular detection and phylogenetic analysis of CPV in diarrhoeic pet dogs. During Jan-July 2019, anal swabs were collected from 96 unvaccinated pet dogs with suspected CPV infection from Sylhet region of Bangladesh. The CPV infection was initially screened through rapid Immunochromatographic (IC) strip test, and then CPV-2 (VP2 gene) were detected by conventional PCR assay. Then the nucleotide sequence of amplified VP2 gene was compared with other CPV strains from GeneBank. Of the total samples, 17.7% (17/96) found positive in IC strip test, and 15.62% (15/96) were found positive in PCR assay by using primer pair P2 that detect original CPV-2 type. The IC test showed 100% sensitivity and 97.5% specificity with PCR. In sequence analysis, our isolates showed the highest 90.40% homology with the isolates of China and the USA. Our strains had also an evolutionary relationship with the other strains of CPV from China and India. This study demonstrates the presence of CPV-2 in Bangladesh and futher sequence analysis of more VP2 gene will help in details insight of the molecular and genetic evolution of CPV in Bangladesh.

In Silico Identification and Functional Characterization of Conserved miRNAs in the Genome of Cryptosporidium parvum.

Cryptosporidium parvum, a predominant causal agent of a fatal zoonotic protozoan diarrhoeal disease called cryptosporidiosis, bears a worldwide public health concern for childhood mortality and poses a key threat to the dairy and water industries. MicroRNAs (miRNAs), small but powerful posttranscriptional gene silencing RNA molecules, regulate a variety of molecular, biological, and cellular processes in animals and plants. As to the present date, there is a paucity of information regarding miRNAs of C. parvum; hence, this study was used to identify miRNAs in the organism using a comprehensible expressed sequence tag-based homology search approach consisting of a series of computational screening process from the identification of putative miRNA candidates to the functional annotation of the important gene targets in C. parvum. The results revealed a conserved miRNA that targeted 487 genes in the model organism (Drosophila melanogaster) and 85 genes in C. parvum, of which 11 genes had direct involvements in several crucial virulence factors such as environmental oocyst protection, excystation, locomotion, adhesion, invasion, stress protection, intracellular growth, and survival. Besides, 20 genes showed their association with various major pathways dedicated for the ribosomal biosynthesis, DNA repair, transportation, protein production, gene expression, cell cycle, cell proliferation, development, immune response, differentiation, and nutrient metabolism of the organism in the host. Thus, this study provides a strong evidence of great impact of identified miRNA on the biology, virulence, and pathogenesis of C. parvum. Furthermore, the study suggests that the detected miRNA could be a potential epigenomic tool for controlling the protozoon through silencing those virulent and pathway-related target genes.

First molecular characterization and phylogenetic analysis of the VP2 gene of feline panleukopenia virus in Bangladesh.

Feline panleukopenia virus (FPV) is a highly contagious infectious pathogen of cats globally. However, there is no information on the molecular identification and characterization of FPV in Bangladesh. Here, 8.16% (8/98) and 18.37% (18/98) of diarrheic cats tested positive for FPV by an immunochromatography (IC) test and PCR, respectively. The IC test showed 44.44% sensitivity and 100% specificity in comparison with PCR. Our newly sequenced Bangladeshi FPV strain (MN826076) showed the highest (99.71%) sequence identity to strains from the United Arab Emirates (UAE). Strain MN826076 contained two characteristic amino acid variations in VP2 identifying it as an FPV strain: valine at position 103 and aspartic acid at position 323. Phylogenetically, the VP2 of strain MN826076 was found to be closely related to 19 FPV strains, sharing the same clade.

Multidrug Antimicrobial Resistance and Molecular Detection of mcr-1 Gene in Salmonella Species Isolated from Chicken.

Colistin (polymyxin E) is widely used in animal and human medicine and is increasingly used as one of the last-resort antibiotics against Gram-negative bacilli. Due to the increased use of colistin in treating infections caused by multidrug-resistant Gram-negative bacteria, resistance to this antibiotic ought to be monitored. The study was undertaken to elucidate the molecular mechanisms, genetic relationships and phenotype correlations of colistin-resistant isolates. Here, we report the detection of the mcr-1 gene in chicken-associated Salmonella isolates in Bangladesh and its in-silico functional analysis. Out of 100 samples, 82 Salmonella spp. were isolated from chicken specimens (liver, intestine). Phenotypic disc diffusion and minimum inhibitory concentration (MIC) assay using different antimicrobial agents were performed. Salmonella isolates were characterized using PCR methods targeting genus-specific invA and mcr-1 genes with validation for the functional analysis. The majority of the tested Salmonella isolates were found resistant to colistin (92.68%), ciprofloxacin (73.17%), tigecycline (62.20%) and trimethoprim/sulfamethoxazole (60.98%). When screened using PCR, five out of ten Salmonella isolates were found to carry the mcr-1 gene. One isolate was confirmed for Salmonella enterica subsp. enterica serovar Enteritidis, and other four isolates were confirmed for Salmonella enterica subsp. enterica serovar Typhimurium. Sequencing and phylogenetic analysis revealed a divergent evolutionary relationship between the catalytic domain of Neisseria meningitidis lipooligosaccharide phosphoethanolamine transferase A (LptA) and MCR proteins, rendering them resistant to colistin. Three-dimensional homology structural analysis of MCR-1 proteins and molecular docking interactions suggested that MCR-1 and LptA share a similar substrate binding cavity, which could be validated for the functional analysis. The comprehensive molecular and in-silico analyses of the colistin resistance mcr-1 gene of Salmonella spp. of chicken origin in the present study highlight the importance of continued monitoring and surveillance for antimicrobial resistance among pathogens in food chain animals.

Data on prevalence, distribution and risk factors for Foot and Mouth Disease in grazing cattle in haor areas of Bangladesh.

Foot and mouth disease (FMD) is a highly contagious and devastating viral disease among all cloven-footed animals. In Bangladesh, the disease is endemic, with outbreaks occurring throughout the year in the haor regions. Thus, the FMD outbreaks impact livelihoods in the haor area and are of great concern. Therefore, a cross-sectional study was undertaken to evaluate the prevalence, distribution, and risk factors for clinical FMD in some selected areas of haor in Sylhet division of Bangladesh. We examined 1,388 cattle, of which 343 were clinically affected with FMD (prevalence 24.71%, CI95% = 22.44 - 26.98) during the period from July 2017 through June 2018. Though production loss was observed, no mortality was recorded in the infected animals. The data article shows the spatial distribution of FMD prevalence. The temporal pattern indicates a higher number of FMD cases in June (47.01%, CI95% = 38.97 - 55.07). The gender was found associated (OR = 2.98; p < 0.001) with the potential risk of FMD occurrence through univariate analysis. Besides, indigenous breeds of cattle (OR = 2.83; p < 0.001) are found to be more susceptible to FMD compared to exotic and crossbreeds. The risk factors identified in this article will serve as a baseline for the development of risk based FMD control program in future.