Bhowmick Sneha, Pal Surajit, Sunder Jai, Sujatha T, De Arun Kumar, Mondal Tousif, Singh Abhishek D, Joardar Siddhartha Narayan, Batabyal Kunal, Dutta Tapan Kumar, Bandyopadhyay Samiran, Tiwari Ananda, Samanta Indranil
Department of Veterinary Microbiology, West Bengal University of Animal and Fishery Sciences, Kolkata, West Bengal, India.
Animal Science Division, ICAR-Central Island Agricultural Research Institute, Port Blair, Andaman and Nicobar Islands, India.
Front Vet Sci. 2023 Jan 5;9:1075133. doi: 10.3389/fvets.2022.1075133. eCollection 2022.
The present study was conducted to detect the occurrence of β-lactamase and biofilm-producing , and in broilers and native fowl reared in the Andaman and Nicobar Islands, India. The study also included molecular docking experiments to confirm the nature of the catalytic domains found in the β-lactamase variants obtained and to reveal the clonal relationship of the isolates with human clinical strains from the database.
A total of 199 cloacal swabs were collected from five poultry breeds/varieties (broiler, , and layer) in three districts of the Andaman and Nicobar Islands. , and were isolated by standard techniques and confirmed by PCR. Phenotypical β-lactamase producers were identified by a double-disc test. The genes ( , , , and ) were screened, and selected sequences of β-lactamase variants were submitted to DDBJ. Homology modeling, model validation, and active site identification of different β-lactamase variants were done by the SWISS-MODEL. Molecular docking was performed to identify the catalytic domains of the β-lactamase variants. The selected β-lactamase sequences were compared with the Indian ESBL sequences from human clinical strains in NCBI-GenBank.
In total, 425 strains were isolated from the collected samples. (42.58%) was found to be the most prevalent, followed by (30.82%) and (26.58%). The phenotypical antibiogram of all 425 isolates showed the highest resistance against oxytetracycline (61-76%) and the lowest against gentamicin (15-20%). Phenotypical production of β-lactamase enzymes was observed in 141 (33.38%) isolates. The isolation rate of β-lactamase producing , and was significantly higher ( < 0.05) in the birds reared in the South Andaman district (25.6, 17.5, and 18.7%, respectively) than in Nicobar (11.5, 7.6, 7.1%, respectively). Genotyping of the β-lactamase-producing isolates revealed the maximum possession of , followed by and . The nucleotide sequences were found to be similar with , , , , , and in BLAST search. Distribution of studied biofilm-associated genes in strains from different varieties of the birds revealed that the layer birds had the maximum possession, followed by , broilers, and fowls. The phylogenetic analysis of selected sequences revealed a partial clonal relationship with human clinical strains of the Indian subcontinent. Molecular docking depicted the Gibbs free energy release for 10 different macromolecules (proteins) and ligand (antibiotic) complexes, ranging from -8.1 (SHV-27 + cefotaxime) to -7 (TEM-1 + cefotaxime) kcal/mol.
The study revealed β-lactamase variants circulating in the fowl population of the Andaman and Nicobar Islands (India), even in remote places with low anthropogenic activity. Most of the strains possessed , followed by . Possession of , , and in poultry strains was not reported earlier from any part of the world. The phylogenetic analysis revealed a partial clonal relationship of β-lactamase sequences with the human clinical strains isolated from the Indian subcontinent.
本研究旨在检测印度安达曼和尼科巴群岛饲养的肉鸡和土鸡中β-内酰胺酶和生物膜产生菌的发生情况。该研究还包括分子对接实验，以确认所获得的β-内酰胺酶变体中催化结构域的性质，并揭示分离株与数据库中人类临床菌株的克隆关系。
从安达曼和尼科巴群岛三个地区的五个家禽品种/品系（肉鸡、[此处原文缺失具体品种名]和蛋鸡）中总共采集了199份泄殖腔拭子。通过标准技术分离出[此处原文缺失具体细菌名]，并通过PCR进行确认。通过双纸片试验鉴定表型β-内酰胺酶产生菌。筛选了[此处原文缺失具体基因名]基因，将选定的β-内酰胺酶变体序列提交给日本DNA数据库（DDBJ）。通过SWISS-MODEL进行不同β-内酰胺酶变体的同源建模、模型验证和活性位点鉴定。进行分子对接以鉴定β-内酰胺酶变体的催化结构域。将选定的β-内酰胺酶序列与NCBI-GenBank中来自人类临床菌株的印度超广谱β-内酰胺酶（ESBL）序列进行比较。
总共从采集的样本中分离出425株[此处原文缺失具体细菌名]菌株。[此处原文缺失具体细菌名]（42.58%）被发现是最普遍的，其次是[此处原文缺失具体细菌名]（30.82%）和[此处原文缺失具体细菌名]（26.58%）。所有425株分离株的表型抗菌谱显示对土霉素的耐药性最高（61 - 76%），对庆大霉素的耐药性最低（15 - 20%）。在141株（33.38%）分离株中观察到β-内酰胺酶的表型产生。在南安达曼地区饲养的禽类中，产生β-内酰胺酶的[此处原文缺失具体细菌名]、[此处原文缺失具体细菌名]和[此处原文缺失具体细菌名]的分离率（分别为25.6%、17.5%和18.7%）显著高于尼科巴地区（分别为11.5%、7.6%、7.1%）（P < 0.05）。对产生β-内酰胺酶的分离株进行基因分型显示，[此处原文缺失具体基因名]的拥有率最高，其次是[此处原文缺失具体基因名]和[此处原文缺失具体基因名]。在BLAST搜索中发现核苷酸序列与[此处原文缺失具体基因名]、[此处原文缺失具体基因名]、[此处原文缺失具体基因名]、[此处原文缺失具体基因名]、[此处原文缺失具体基因名]和[此处原文缺失具体基因名]相似。对来自不同禽类品种的[此处原文缺失具体细菌名]菌株中研究的生物膜相关基因的分布显示，蛋鸡的拥有率最高，其次是[此处原文缺失具体品种名]、肉鸡和[此处原文缺失具体品种名]家禽。对选定序列的系统发育分析揭示了与印度次大陆人类临床菌株的部分克隆关系。分子对接描绘了10种不同大分子（蛋白质）和配体（抗生素）复合物的吉布斯自由能释放，范围从 -8.1（SHV - 27 + 头孢噻肟）到 -7（TEM - 1 + 头孢噻肟）kcal/mol。
该研究揭示了即使在人为活动较少的偏远地区，印度安达曼和尼科巴群岛家禽群体中也存在β-内酰胺酶变体。大多数菌株拥有[此处原文缺失具体基因名]，其次是[此处原文缺失具体基因名]。此前世界上任何地区都未报道过家禽[此处原文缺失具体细菌名]菌株中存在[此处原文缺失具体基因名]、[此处原文缺失具体基因名]和[此处原文缺失具体基因名]。系统发育分析揭示了β-内酰胺酶序列与从印度次大陆分离的人类临床菌株的部分克隆关系。
