Department of Exercise and Medical Science, Graduate School, Dankook University, Cheonan, Republic of Korea.
Department of Microbiology, College of Science & Technology, Dankook University, Cheonan, Republic of Korea.
PLoS One. 2024 Mar 27;19(3):e0298745. doi: 10.1371/journal.pone.0298745. eCollection 2024.
Aeromonas spp. are the opportunistic pathogens that infect both aquatic and terrestrial homeotherms. They were commonly present in aquatic environments, including effluent, tap water, marine, river, and lake, where they are often isolated from aquatic animals, including fish, molluscs, and crustaceans. The Aeromonas infections can cause sepsis, ulcer, and other symptoms, resulting in the death of massive aquatic animals. Therefore, the prevention and control of Aeromonas is of great significance for the healthy development of aquaculture. In this study, we used modern molecular methods to enhance disease control of Aeromonas isolates from freshwater fish in Hebei Province. A total of 130 Aeromonas spp. isolates were isolated from freshwater fish farms in Hengshui, Handan, and Shijiazhuang and all 130 Aeromonas spp. isolates were sequenced for species identification. Of the 130 Aeromonas spp. isolates, 104 isolates were successfully sequenced, and BLAST analysis showed that Aeromonas veronii was predominant in freshwater fish farms in Hebei Province. In addition, 26 antibiotic resistance profiles were obtained from 102 fully cultured isolates among the 104 Aeromonas spp. isolates whose species was primarily identified, and 44 multidrug-resistant bacteria among the 102 isolates were identified using an antibiotic susceptibility test. Using the Multilocus Sequence Typing (MLST) method, 33 out of 44 multidrug-resistant isolates with 14 non-Aeromonas reference strains were selected for phylogenetic and MLST analysis, and all 33 multidrug-resistant isolates were A. veronii. A total of 30 new Sequence Types (STs) were obtained by comparing concatenated sequences (gyrB-groL-gltA-metG-ppsA-recA) on PubMLST website. Furthermore, recombination event analysis detected using RDP5 and ClonalFrameML software 42 and 49 recombination events, respectively, and 22 recombination events were validated by four or more algorithms. Since mutation and recombination events increase clonal diversity and single housekeeping gene sequence alignments are limited for identifying species, we propose the use of multiple concatenated sequence loci to increase discriminatory power. In addition, we propose that the MLST method is an appropriate technique to study and develop the resistance mechanisms of multidrug-resistant Aeromonas and to identify Aeromonas systematically in complex samples obtained from the environment.
气单胞菌属是一种机会性病原体，可感染水生和陆生恒温动物。它们通常存在于水生环境中，包括污水、自来水、海洋、河流和湖泊，经常从水生动物（包括鱼类、软体动物和甲壳类动物）中分离出来。气单胞菌感染可引起败血症、溃疡等症状，导致大量水生动物死亡。因此，预防和控制气单胞菌对水产养殖业的健康发展具有重要意义。在这项研究中，我们使用现代分子方法增强了对河北省淡水鱼类气单胞菌分离株的疾病控制。从衡水、邯郸和石家庄的淡水养殖场共分离出 130 株气单胞菌，对 130 株气单胞菌进行了测序以进行种属鉴定。在 130 株气单胞菌分离株中，有 104 株分离株成功测序，BLAST 分析显示，河北省淡水养殖场中气单胞菌 veronii 占优势。此外，在对 104 株气单胞菌分离株中的 102 株完全培养分离株进行种属初步鉴定后，获得了 26 种抗生素耐药谱，通过抗生素敏感性试验鉴定出 102 株分离株中有 44 株多药耐药菌。使用多位点序列分型（MLST）方法，从 44 株多药耐药菌中选择了 33 株与 14 株非气单胞菌参考菌株一起进行了系统发育和 MLST 分析，所有 33 株多药耐药菌均为气单胞菌 veronii。通过在 PubMLST 网站上比较串联序列（gyrB-groL-gltA-metG-ppsA-recA），共获得 30 种新的序列类型（ST）。此外，使用 RDP5 和 ClonalFrameML 软件分别检测到重组事件分析 42 次和 49 次，通过四种或更多算法验证了 22 次重组事件。由于突变和重组事件增加了克隆多样性，并且单一管家基因序列比对对于鉴定物种的能力有限，因此我们建议使用多个串联序列位点来提高鉴别力。此外，我们建议 MLST 方法是研究和开发多药耐药气单胞菌耐药机制以及在从环境中获得的复杂样本中系统鉴定气单胞菌的合适技术。
