野生水鸟感染霍乱弧菌和沙门菌等病原菌的分离和鉴定
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摘要
目的从死亡的野生水鸟标本中分离、鉴定病原菌,评估迁徙鸟类传播人兽共患病原菌的风险。方法采集的组织标本涂布营养琼脂平板,分别在需氧和厌氧的条件下培养。分离的菌株使用Phoenix100全自动微生物鉴定仪和16s rRNA测序的方法鉴定细菌种属,随后通过PCR的方法检测病原菌携带主要毒力基因的情况。结果从骨顶鸡、鸬鹚和红嘴鸥等鸟类的组织标本中分离鉴定出霍乱弧菌、沙门菌和产气荚膜梭菌等多种肠道感染相关的病原细菌。霍乱弧菌经PCR鉴定不属于O1和O139血清群,无霍乱毒素和毒素相关菌毛的编码基因,但是有溶血素的编码基因。结论检测的死亡鸟类个体存在多病原细菌的感染。
The pathogenetic bacteria were isolated from wild water birds following sudden death in Inner mongolia autonomous region of China, under both aerobic and anaerobic conditions. Those strains were identified as Vibrio cholerae, Salmonella and Clostridium perfringens etc. by Phoenix100 automatic microorganism identification instrument or 16s rRNA sequencing. The mainly virulence genes, serotype or toxinotype of different strains were detected by specific PCR assays. All the C. perfringens isolates were determined as toxinotype A. All the V. cholerae isolates were determined as Non-O1/O139 sero-group, which contain only the hemolysin gene but not the cholera toxin and toxin-coregulated pili genes. Since those zoonotic bacteria can be isolated from wild migratory birds, it should deserve more attention for the biosafety and public health issues.
The pathogenetic bacteria were isolated from wild water birds following sudden death in Inner mongolia autonomous region of China, under both aerobic and anaerobic conditions. Those strains were identified as Vibrio cholerae, Salmonella and Clostridium perfringens etc. by Phoenix100 automatic microorganism identification instrument or 16s rRNA sequencing. The mainly virulence genes, serotype or toxinotype of different strains were detected by specific PCR assays. All the C. perfringens isolates were determined as toxinotype A. All the V. cholerae isolates were determined as Non-O1/O139 sero-group, which contain only the hemolysin gene but not the cholera toxin and toxin-coregulated pili genes. Since those zoonotic bacteria can be isolated from wild migratory birds, it should deserve more attention for the biosafety and public health issues.
引文
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[14] Doran JL, Collinson SK, Clouthier SC, et al. Diagnostic potential of sefA DNA probes to Salmonella Enteritidis and certain other O-serogroup D1 Salmonella serovars[J]. Mol Cell Probes, 1996, 10(4): 233-246.
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[3] Benskin CM, Wilson K, Jones K, et al. Bacterial pathogens in wild birds: a review of the frequency and effects of infection[J]. Biol Rev Camb Philos Soc, 2009, 84(3): 349-373. DOI: 10.1111/j.1469-185X.2008.00076.x
[4] Greig J, Rajic A, Young I, et al. A scoping review of the role of wildlife in the transmission of bacterial pathogens and antimicrobial resistance to the food chain[J]. Zoonoses Public Health, 2015, 62(4): 269-284. DOI: 10.1111/zph.12147
[5] Nandi B, Nandy RK, Mukhopadhyay S, et al. Rapid method for species-specific identification of Vibrio cholerae using primers targeted to the gene of outer membrane protein OmpW[J]. J Clin Microbiol, 2000, 38(11): 4145-4151.
[6] Li F, Du P, Li B, et al. Distribution of virulence-associated genes and genetic relationships in non-O1/O139 Vibrio cholerae aquatic isolates from China[J]. Appl Environ Microbiol, 2014, 80(16): 4987-4992. DOI: 10.1128/AEM.01021-14
[7] Alam M, Sultana M, Nair GB, et al. Toxigenic Vibrio cholerae in the aquatic environment of Mathbaria, Bangladesh[J]. Appl Environ Microbiol, 2006, 72(4): 2849-2855. DOI: 10.1128/AEM.72.4.2849-2855.2006
[8] Awasthi SP, Asakura M, Neogi SB, et al. Development of a PCR-restriction fragment length polymorphism assay for detection and subtyping of cholix toxin variant genes of Vibrio cholerae[J]. J Med Microbiol, 2014, 63(Pt 5): 667-673. DOI: 10.1099/jmm.0.070797-0
[9] Oliveira SD, Rodenbusch CR, Cé MC, et al. Evaluation of selective and non-selective enrichment PCR procedures for Salmonella detection[J]. Lett Appl Microbiol, 2003, 36(4): 217-221. DOI: org/10.1046/j.1472-765X.2003.01294.x
[10] Malorny B, Paccassoni E, Fach P, et al. Diagnostic real-time PCR for detection of Salmonella in food[J]. Appl Environ Microbiol, 2004, 70(12): 7046-7052. DOI: 10.1128/AEM.70.12.7046-7052.2004
[11] Yoo HS, Lee SU, Park KY, et al. Molecular typing and epidemiological survey of prevalence of Clostridium perfringens types by multiplex PCR[J]. J Clin Microbiol, 1997, 35(1): 228-232.
[12] Zhu L, Zhou W, Wang T, et al. Isolation of Clostridium perfringens type A from wild bharals (Pseudois nayaur) following sudden death in Tibet, China[J]. Anaerobe, 2017, 44: 20-22. DOI: 10.1016/j.anaerobe.2017.01.004
[13] Purdy AE, Balch D, Lizárraga-Partida ML, et al. Diversity and distribution of cholix toxin, a novel ADP-ribosylating factor from Vibrio cholerae[J]. Environ Microbiol Rep, 2010, 2(1): 198-207. DOI: 10.1111/j.1758-2229.2010.00139.x
[14] Doran JL, Collinson SK, Clouthier SC, et al. Diagnostic potential of sefA DNA probes to Salmonella Enteritidis and certain other O-serogroup D1 Salmonella serovars[J]. Mol Cell Probes, 1996, 10(4): 233-246.