食源性沙门氏菌的PCR检测

该试验根据GenBank数据库沙门氏菌的特有基因invA(AE008832)设计扩增长度为285 bp的引物,研究PCR技术在检测冷冻肉中沙门氏菌的检测限度.在生理盐水中食源性沙门氏菌的检测限度为103 cfu/mL;在人为污染肉汤中,普通FCR的检出限为1.98×103 cfu/mL;人为污染后的样品肉汤中沙门氏菌浓度为100,101,102 cfu/mL时,分别培养7.6.6h即可检测到.     

鸭疫里默氏杆菌的共感染情况调查

为了调查山东省鸭疫里默氏杆菌病与其他疾病的混合感染情况,对从山东省6个肉鸭养殖较多县(市)收集到的118份样品进行了大肠杆菌、沙门氏菌和鸭病毒性肝炎的分离鉴定。结果显示,鸭疫里默氏杆菌和大肠杆菌单独感染分别占到样品总数的21.18%和11.86%,鸭疫里默氏杆菌与大肠杆菌和鸭病毒性肝炎的双重感染最为普遍,分别占到样品总数的38.14%和18.64%,鸭疫里默氏杆菌、鸭大肠杆菌、鸭病毒性肝炎的三重感染占到样品总数的6.78%。其中,鸭大肠杆菌的优势血清型为O₇₈和O₉₂,分别占总分离株数的16.90%和15.49%,其次是血清型O₉₃和O₁₄₂,分别占到总分离株的11.27%和8.45%。     

猪链球菌2型湖南分离株多重耐药性及相关耐药基因研究

基因突变和基因转移是细菌耐药性产生和存在的重要内因,检测与抗生素耐药性相关的基因具有重要的意义.试验选取25份猪链球菌2型湖南分离株对16种抗生素进行药敏试验,检测菌株耐药性;选出了23株有红霉素抗性的菌株,用PCR检测其erm(B)基因.结果显示,猪链球菌湖南分离株对红霉素、四环素、万古霉素和克林霉素具有高耐药性,在23株红霉素抗性菌株中有18株存在erm(B)基因,由erm(B)基因产生的红霉素耐药菌株占到其中的78.3%.因此,erm(B)基因是链球菌2型湖南分离株对红霉素耐药的主要抗性决定基因.     

禽流感病毒H9N2亚型三重PCR检测方法的建立

为建立简便快速检测禽流感病毒(avian influenza virus,AIV)并同时区分出H9、N2亚型的方法,本试验根据基因库中H9亚型AIV的HA基因、N2亚型AIV的NA基因及AIV的M基因序列,分别设计了3对针对这3种基因保守序列的引物,建立了AIV H9N2亚型的三重PCR检测方法。应用该方法对H9N2亚型AIV模板进行PCR扩增,可得到3条与试验设计相符的目的条带,分别为313 bp (HA基因)、451 bp (NA基因)和667 bp(M基因);对非H9亚型的N2亚型AIV模板进行扩增,出现2条特异性扩增条带,即451 bp (NA基因)和667 bp(M基因);对非H9、N2亚型AIV模板进行扩增则只出现一条目的条带,即667 bp(M基因);对其他禽呼吸道病原体进行PCR扩增,结果均为阴性。敏感性试验结果显示此三重PCR方法最低检出限为10^-2 ng/μL。应用所建立的三重PCR方法对120份临床病料进行检测的结果与病毒分离鉴定结果一致。各项试验结果均表明,该方法对于禽流感病毒尤其是H9、N2亚型禽流感病毒的检测具有快捷、特异、灵敏的特点。     

Identification and quantification of benzimidazole resistance polymorphisms in Haemonchus contortus isolated in Northeastern Brazil

Haemonchus contortus is the most prevalent nematode in Brazil. The objective of this study was to select 6 populations of H. contortus of known or suspected benzimidazole resistance status and characterize these using quantitative real-time polymerase chain reaction (qPCR) for single nucleotide polymorphisms (SNPs) F200Y, F167Y and E198A in the β-tubulin isotype 1 gene. qPCR was performed using DNA from a pool of 10 adult male H. contortus from a single animal per farm. Faecal egg count reduction test (FECRT) and egg hatch test (EHT) were used to determine the resistance status. Samples were obtained from 6 farms located in 5 counties in the Ceará State: Tauá, Boa Viagem, Quixadá, Santa Quitéria and Solonópole. The inbred-susceptible-Edinburgh (ISE) isolate was used as reference for comparative purposes in the qPCR. Benzimidazole resistance was detected by FECRT on all farms with efficacy values ranging from 0 to 51%. EC50 values as determined by EHT were all above 1.49 μg/ml. High frequencies of the resistant SNPs F200Y and F167Y alleles were detected but no resistance was detected at SNP E198A. Our results suggest that the SNPs F167Y and F200Y are both important for benzimidazole resistance in the studied populations.     

荧光定量PCR法比较研究健康和子宫内膜炎奶牛阴道菌群结构

为揭示奶牛阴道菌群结构与子宫内膜炎发生关系,本实验选择健康奶牛和患有子宫内膜炎奶牛各5头,采集其阴道粘液样品,提取样品总DNA,利用SYBR Green Ⅰ荧光定量PCR方法对这两类奶牛阴道菌群结构的差异进行了检测.检测结果显示,健康奶牛阴道内乳杆菌属(p＜0.05)、芽孢杆菌属(p＜0.05)和魏斯氏菌属(p＜0.01)的细菌数量显著或极显著高于子宫内膜炎奶牛,而子宫内膜炎奶牛阴道内大肠杆菌数量显著高于健康奶牛(p＜0.05).研究结果表明,阴道菌群结构失衡可能是奶牛子宫内膜炎发生的原因.     

Prevalence and molecular characterization of Hepatozoon canis in dogs from urban and rural areas in Southeast Brazil

The objective of this survey was to investigate the prevalence of Hepatozoon infection in dogs in the rural and urban areas of Uberlândia, Brazil by PCR and molecular characterization. DNA was obtained from blood samples collected from 346 local dogs from both genders and various ages. Seventeen PCR products from positive blood samples of urban dogs and 13 from the rural dogs were sequenced. Partial sequences of the 18S rRNA gene indicated that all 30 dogs were infected with Hepatozoon canis similar in sequence to H. canis from southern Europe. Four local dog sequences were submitted to GenBank (accessions JN835188; KF692038; KF692039; KF692040). This study indicates that H. canis is the cause of canine hepatozoonosis in Uberlândia and that infection is similarly widespread in rural and urban dogs.     

奶牛无浆体病PCR诊断方法的建立及应用

利用边缘无浆体(Anaplasma marginale)高度保守的msp5基因,建立了奶牛无浆体病PCR诊断方法。特异性试验表明与牛巴贝斯虫、双芽巴贝斯虫、温氏附红细胞体、大肠杆菌、金黄色葡萄球菌和牛白细胞DNA无交叉反应;敏感性试验表明可检测到约200个感染红细胞。利用该方法对黑龙江省西部多个养牛场送检的140份血样进行检测,其中曾出现过高热、气喘、流涎和贫血等临床症状的"无名高热"奶牛血样95份、无临床症状奶牛血样45份,结果有上述临床症状奶牛PCR血样阳性率为44.2%,无症状奶牛血样PCR阳性率为13.3%。从而证实无浆体是引起奶牛"无名高热"的病原之一。     

3种优势牧草在不同生育期对干旱胁迫的响应

选择松嫩草地上的3种优势牧草羊草(Leymus chinensis)、碱蒿(Artemisia anethifolia)和全叶马兰(Kalimeris integrifolia) ,通过野外干旱胁迫试验、定点实时观测和隶属函数法,研究3种优势牧草在不同生育期内对不同等级干旱胁迫的响应,探究不同等级干旱胁迫对牧草在不同生育期内形态特性的影响及其抗旱能力。结果表明：在牧草同一生育期内,随干旱胁迫程度加剧,株高、叶长、叶绿素含量和产量受到的影响越大,在特旱程度下,3种牧草生长受干旱胁迫影响最严重;同一干旱胁迫处理下,随着生育期推移,株高、叶长、叶绿素含量和产量受干旱胁迫的影响总体呈减少趋势。3种牧草在其生育期内的抗旱性呈现出一定的规律性,羊草在开花期抗旱能力最强,其次是在返青期;全叶马兰在开花期抗旱能力最强,其次是在返青期,在抽穗期最差;碱蒿在开花期抗旱能力最强,其次是在抽穗期,在返青期最差。本研究结果对松嫩草地3种优势草种在不同生育期内遭受不同程度干旱胁迫时采取不同应急方案具有指导作用,也为培育优良耐旱草种及草地生态系统可持续发展提供了一定的参考。