[目的]建立检测新型阿卡斑病毒(AKAV)的SYBR Green I荧光定量RT-PCR,并结合序列测定开展广西边境地区蚊携带新型AKAV调查,了解其流行趋势及遗传进化特征,为建立重要蚊媒病毒病预警机制提供技术支持.[方法]分别针对新型AKAV的S基因和M基因主要变异位点设计引物,建立SYBR Green I荧光定量RT-PCR并通过Ct值、标准曲线斜率、相关系数、扩增效率及扩增准确性等指标确定最佳反应体系及扩增程序;以优化的SYBR Green I荧光定量RT-PCR检测2019年6—10月在广西边境地区采集的139份蚊样品,获得的新型AKAV经测序后采用ClustalW构建遗传进化树.[结果]SYBR Green I荧光定量RT-PCR最佳反应体系:SYBR Premix Ex TaqⅡ10.0μL,上、下游引物(4μmol/L)各1.0μL,标准品或反转录产物2.0μL,双蒸水补齐至20.0μL.扩增程序:95℃预变性30 s;95℃30 s,64℃(S基因)/66℃(M基因)30 s,72℃30 s,进行40个循环.SYBR Green I荧光定量RT-PCR检测新型AKAV S基因和M基因的极限为1.0×101 copies/μL和1.0×102 copies/μL,对应的扩增效率分别为98.61％和105.81％,但不能扩增蓝舌病病毒、鹿流行性出血热病毒、竹鼠圆环病毒和布鲁氏杆菌;检测S基因和M基因的批内重复试验、批间重复试验变异系数(CV)均低于5.00％.采用建立的SYBR Green I荧光定量RT-PCR成功从广西边境地区防城港市、东兴市、宁明县、大新县及北海市的阿蚊和库蚊中检出新型AKAV,且均属于基因Ia型,尤其与广东和湖南蠓虫、中华按蚊、致倦库蚊分离毒株具有较高的相似性.[结论]针对新型AKAV S基因和M基因建立的SYBR Green I荧光定量RT-PCR具有特异性强、灵敏度高、重复性好的特点,更适用于检测病毒拷贝数低的样品.此外,从广西边境地区蚊样品中检出的AKAV毒株均为新型AKAV,属于基因Ia型,与广东和湖南蠓虫、中华按蚊、致倦库蚊分离毒株具有较高的相似性,推测新型AKAV已在广西周边省份流行.

The causes of death of pigs in a pig farm were confirmed and the pathogenicity and drug resist-ance of isolates were investigated.The bacteria were isolated by conventional methods,and the culture char-acteristics,1 6 S rDNA and pathogenicity,virulence genes and drug resistance were detected.The results showed that 9 strains of Streptococcus suis were isolated from dead pigs and 9 9.9% homology with Strep-tococcus suis GZ0565 in GenBank.Virulence factor test results showed for cps9H+orf2+gdh+fdps+virulent type,and mouse pathogenicity test identified as virulent strains.The results showed that the strain had high sensitivity to nitrofurantoin,ciprofloxacin and cephalosporin,and had complete resistance to oxacillin,az-treonam,kanamycin,cotrimoxazole,polymyxin B,lincomycin,presented resistant genes of tetracyclines, aminoglycosides,sulfonamides,quinolones.This study showed that the outbreak is caused by type 9 patho-genic Streptococcus suis.

为了解广西牛片形吸虫病的流行情况,在广西8个地级市不同规模牛场及散养户共采集了1 571份牛粪,采用水洗沉淀法进行检测,同时屠宰时随机抽查50头牛,观察组织及病理变化.调查结果表明,广西牛片形吸虫感染较为严重,平均感染率达69.3％(1 089/1 571),其中南宁71.9％ (41/57)、柳州46.3％ (139/300)、梧州72.3％ (88/122)、北海37.5％ (6/16)、崇左87.8％ (652/742)、河池69.1％ (65/94)、百色39.6％(87/220)和贵港55％(11/20),各地感染情况均有不同.牛感染片形吸虫主要病变器官为肝脏、胆囊、淋巴结,而胆囊是片形吸虫病寄生的主要地方.本调查为指导广西防治牛片形吸虫病,促进肉牛产业发展提供参考依据.

To identify the causative agent infecting Cavia porcellus on a farm in Wuxu town of Nanning,suspect bacteria were isolated from the eye discharge,neck cyst and lungs of diseased Caviaporcellus.Their morphological characteristics,physiological and biochemical characteristics and the results of 16s rRNA gene sequence analysis proved that the pathogenic bacteria were Streptococcus equi subsp,zooepidemicus.In a pathogenic test,the isolate was lethal to adult mice.Their 16S rRNA was closely related to Streptococcus equi subsp.Zooepidemicus(AB104843) with homologies of 99.8％.Phylogenetic analysis indicated that the isolate was closely related to Streptococcus equi subsp,zooepidemicus(AB104843) and Streptococcus equi subsp,equi(JQ726495 and JQ726496),positioning the same branch.The results of antibiotic susceptibility testing demonstrated that the Streptococcus was highly sensitive to ampicillin,aztreonam,cefradine,cefazolin,cephalexin,cefuroxim,streptomycin,kanamycin etc..but resistant to penicillin,imipenem,tetracycline,minocycline etc.These findings suggest that Streptococcus equi subsp.Zooepidemicus 16-58 is the pathogen of the dead Cavia porcellus in Wuxu town of Nanning.

This study aimed to determine the pathogen infecting the bamboo rats with abscessing pneumonia in Jinling town of Nanning.One bacterial strain was isolated from the lungs of diseased bamboo rats.The analysis by morphological characteristic,physiological and biochemical assays,and 16S rRNA gene sequence demonstrated that the pathogenic bacterium was Escherichia coli.The virulence assay showed that the isolate was highly pathogenic to the adult mice.The 16S rRNA gene sequence closely related to Escherichia coli (DQ337503,FJ418574,KR189581) with homologies of 100％.The susceptibility test to antibiotic demonstrated that the bacterial strain was highly sensitive to cefixime,kanamycin,spectinomycin,netilmicin,florfenicol and ninocycline,but resistant to ampicillin,amoxicillin,imipenem,cefazolin,cephalexin,tetracycline,erythromycin and clarithromycin.The study provides a basis for disease control of the bamboo rats in Jinling town of Nanning.