Investigations of milk quality from teats with milk flow disorders

The objective of this study was to investigate the quality of milk from teats with milk flow disorders. Somatic cell count, pathogens, and signs of mastitis (>100,000 cells/ml and pathogens detected) were determined in the milk from all teats of the udder before treatment of the affected teat, as well as 1 and 6 mo later. Teats with milk flow disorders were compared to all of the other teats from the same udder. Before treatment, the SCC from affected teats was 4.3 million higher, the odds of detecting pathogens 6 times higher, and the odds of mastitis 11 times higher than in control teats (when adjusted for other significant explanatory variables). SCC and the risk of mastitis decreased after surgical treatment of the affected teats, whereas the chance of detecting pathogens was not affected. Six months after treatment, the SCC was 1.3 million higher, and the odds of mastitis 6.5 times higher than in control teats. Throughout the study period neither SCC, the odds of detecting pathogens, nor mastitis changed significantly in control teats. It may be concluded from this study that milk quality from teats with milk flow disorders is decreased before treatment and does not reach the milk quality from unaffected teats within 6 mo after treatment.     

Effects of pathogen-specific clinical mastitis on probability of conception in Holstein dairy cows

The objective of this study was to estimate the effects of pathogen-specific clinical mastitis (CM), occurring in different weekly intervals before or after artificial insemination (AI), on the probability of conception in Holstein cows. Clinical mastitis occurring in weekly intervals from 6 wk before until 6 wk after AI was modeled. The first 4 AI in a cow’s lactation were included. The following categories of pathogens were studied: Streptococcus spp. (comprising Streptococcus dysgalactiae, Streptococcus uberis, and other Streptococcus spp.); Staphylococcus aureus; coagulase-negative staphylococci (CNS); Escherichia coli; Klebsiella spp.; cases with CM signs but no bacterial growth (above the level that can be detected from our microbiological procedures) observed in the culture sample and cases with contamination (≥3 pathogens in the sample); and other pathogens [including Citrobacter, yeasts, Trueperella pyogenes, gram-negative bacilli (i.e., gram-negative organisms other than E. coli, Klebsiella spp., Enterobacter, and Citrobacter), Corynebacterium bovis, Corynebacterium spp., Pasteurella, Enterococcus, Pseudomonas, Mycoplasma, Prototheca, and others]. Other factors included in the model were parity (1, 2, 3, 4 and higher), season of AI (winter, spring, summer, autumn), day in lactation of first AI, farm, and other non-CM diseases (retained placenta, metritis, ketosis, displaced abomasum). Data from 90,271 AI in 39,361 lactations in 20,328 cows collected from 2003/2004 to 2011 from 5 New York State dairy farms were analyzed in a generalized linear mixed model with a Poisson distribution. The largest reductions in probability of conception were associated with CM occurring in the week before AI or in the 2 wk following AI. Escherichia coli and Klebsiella spp. had the greatest adverse effects on probability of conception. The probability of conception for a cow with any combination of characteristics may be calculated based on the parameter estimates. These findings may be helpful to farmers in assessing reproduction in their dairy cows for more effective cow management.     

Quarter- and cow-level risk factors for intramammary infection with coagulase-negative staphylococci species in Swiss dairy cows

Bacteriological status, evaluation of udder symmetry, udder hygiene, and teat end scores of 92 dairy cows were assessed on 3 Swiss dairy farms in a longitudinal 1-yr study to determine risk factors for intramammary infection (IMI) with coagulase-negative staphylococci (CNS) species. Farm visits were performed monthly including sterile quarter milk sampling and udder evaluation of all lactating cows. Milk samples were evaluated for the presence of staphylococci using selective agar plates. Species identification was performed using MALDI-TOF mass spectrometry. Intramammary infection was defined as milk samples having ≥100 cfu per mL of milk according to culture results. Overall, 3,151 quarter samples were included in the statistical analysis. Staphylococcus chromogenes, Staphylococcus haemolyticus, Staphylococcus xylosus, and a Staphylococcus warneri-like species were the 4 most prevalent CNS species found. Hierarchical multivariable logistic regression models were built to evaluate risk factors for species-specific CNS IMI. Risk factors for Staph. chromogenes IMI were presence in herd B, the period from June 2014 to August 2014 and December 2014 to February 2015, and presence of udder edema. For Staph. haemolyticus, the relevant risk factor included coinfection with Staph. xylosus coinfection with other than the above-mentioned CNS species (“others”) and the period from June 2014 to November 2014. Coinfection with Staph. haemolyticus and “others,” the periods from June 2014 to August 2014 and December 2014 to February 2015, early phase of lactation (1–60 d in milk), and belonging to herd B were significantly associated with Staph. xylosus IMI. Mid and late lactation, coinfection with Staph. xylosus, and the period September 2014 to May 2015 were identified as significant risk factors for Staph. warneri-like IMI. For Staph. chromogenes, 60.6 and 26% of the variance was observed at the quarter and cow level, respectively, whereas for the other investigated species the highest variance was observed at the sample level. The predominant species within herds differed and was most pronounced for the Staph. warneri-like species.     

Pathogen group-specific risk factors for intramammary infection in treated and untreated dairy heifers participating in a prepartum antimicrobial treatment trial

Heifer mastitis is a well-known problem, with several pathogens being involved. Several generic risk factors associated with the likelihood of intramammary infections (IMI) in fresh dairy heifers have been identified before. Yet, a need exists to identify pathogen group-specific factors, as the effect of (groups of) pathogens on udder health and milk yield is different. The aim of the present study was to identify pathogen group-specific risk factors for IMI in heifers participating in a prepartum antimicrobial treatment trial, allowing us to test the hypothesis that different factors are of importance between treated and untreated control heifers as well. Data from a clinical trial in which end-term heifers were treated systemically (over 3 consecutive days) 2 wk before calving with penethamate hydriodide (n = 76) or remained untreated (n = 73), were available. Several potential risk factors at the herd, heifer, and quarter level were recorded in the first 3 d in milk. Quarters from untreated heifers supplemented with ≥4 mg of selenium/d prepartum were significantly less likely to be infected with coagulase-negative staphylococci (CNS), whereas quarters were more likely to be infected with CNS when assistance during calving was needed. Udder edema before calving significantly decreased the odds of IMI with major pathogens. In treated heifers, no factors were detected that were associated with the likelihood of CNS IMI, whereas quarters from heifers were significantly more likely to be infected with major pathogens when they were housed in the calving pen more than 1 d and when they had been in contact with the lactating cows before calving. The risk factors for IMI that were identified in treated heifers were different than those in untreated heifers, independent of the pathogen group that was considered. It looks as if prepartum treatment not only changed the likelihood of infection, but also the factors that were associated with infection. However, except for treated heifers with an IMI with major pathogens, only a small proportion of the variation could be explained in the final models. Therefore, factors other than those that were studied could explain the likelihood of infection.     

黄瓜褐斑病病原菌鉴定及生物学特性研究

对辽宁省保护地发生的黄瓜褐斑病进行病原菌的分离、鉴定 ,明确该病的病原菌为多主棒孢霉[Corynesporacassiicola(B.&C.)Wei]。生物学特性研究表明 ,该菌孢子萌发的最适温度范围为15～35℃ ,最适pH值为5～8,水滴是孢子萌发的必要条件。病菌营养生长的最适温度为15～35℃ ,最适 pH值为6～8 ,在PSA、PDA等常规培养基上生长良好 ,光照对菌丝生长速度影响不显著。病菌产孢的最适温度为25～30℃,最适pH值为5～8,光照有助于孢子形成,在PSA、PDA等常规培养基上能大量产孢。病菌分生孢子在室外难以安全越冬,在保护地内能够安全越夏。该菌寄主范围广,能寄生黄瓜和番茄等20余种作物     

玉米纹枯病病菌侵染过程研究

研究了纹枯病菌侵入玉米叶鞘的显微和超微过程,结果表明：在接种病菌后１２～２４ｈ,病菌通过表皮、乳孔和自然孔口３种途径侵入寄主,其中以表皮直接侵入为主。侵染垫是病菌主要侵入结构。接种１２ｈ,可在叶鞘表面形成少量近圆形的侵染垫。接种２４ｈ,则有大量侵染垫形成,侵染垫下可见侵入钉直接侵入寄主表皮。接种２４ｈ后叶鞘组织细胞内可见大量侵入的菌丝。     

我国食品微生物定量风险评估的研究进展

近10余年来,我国政府重视食品安全并逐渐加强风险分析体系的构建和实施。根据历年来国家卫生与计生委的统计数据,表明由致病微生物导致的食物中毒发病率一直高于其他危害。食源性致病菌引起的食品安全风险是全球性问题,发展中国家面临的情况更为严峻,因此加强我国微生物定量风险评估以减少与发达国家的差距,从国家层面上势在必行。特别是2009年我国颁布实施《食品安全法》和2011年成立国家食品安全风险评估中心（ChinaNational Center for Food Safety Risk Assessment,CFSA）以来,已有不少针对国内具体食品致病菌情况开展的食品微生物定量风险评估（quantitative microbial risk assessment,QMRA）研究。本文对2000年至今我国已开展的QMRA研究,包括涉及的食品、致病菌、微生物预测模型、剂量-效应模型等进行详细综述。同时指出我国开展QMRA面临的技术性难题及解决方法,并对采用QMRA结果用于构建危害分析与关键控制点（Hazard Analysis and CriticalControl Point,HACCP）和食品安全目标（ food safety objective,FSO）以及执行目标（performance objective,PO）的应用前景进行探讨。建议加强风险评估与风险管理的互动交流、进一步完善风险监测、微生物限量制定和国际合作,在完善实施指南的基础上针对我国具体国情开展更多具有科学性和系统性的QMRA研究。     

基于tdh1基因变异的大流行副溶血性弧菌鉴别方法的建立

目的基于副溶血性弧菌tdh1 DNA序列中特异性变异位点,建立新的大流行株鉴别方法。方法通过对多种型别菌株的tdh序列进行比对,查找大流行株特异性变异位点,并基于该位点设计位点特异性PCR体系和检测方法。以GS-PCR和tdh基因联合检测为参照方法,用已知大流行株、非大流行株和2014年收集的1 067株副溶血性弧菌对新方法进行验证。结果 3株菌株的6条tdh全序列存在26个多态性位点,其中tdh1基因的第368位碱基的变异[G/A]能用于鉴别大流行株,本研究基于此位点建立了tdh1_368位点特异性PCR。该变异能将1996年后的大流行O3∶K6型菌株与1996前的进行区分,也能将大流行株其他血清型变种与非流行菌株进行区分。对2014年的1 067株菌株的检测结果显示,tdh1_368位点特异性PCR与参考方法检测结果完全一致。结论本研究以副溶血性弧菌tdh基因的多态性位点建立检测大流行菌型的试验方法,从对1 067株菌株的实测结果来看,tdh1_368位点特异性PCR与既往报道的方法具有高度的一致性,且指标更为直接。     

基于内参的副溶血性弧菌实时荧光定量PCR快速检测方法的建立

目的建立基于内参的副溶血性弧菌实时荧光定量PCR方法,快速检测样品中的副溶血性弧菌。方法根据Gen Bank已公布的副溶血性弧菌基因组序列,筛选特异性靶基因,设计特异性引物探针,优化反应体系,并在体系中加入内参(IAC),通过标记不同荧光基团的Taq Man探针来监测IAC,进而实时监控整个PCR反应。按照5~50 cfu/25 g的细菌量人工污染样品,以评价所建立反应的体系。结果以副溶血性弧菌基因组DNA为模板,最低检测限为1 pg/μl;以10倍梯度稀释的菌液经水煮法提取的DNA为模板,最低检测限为4×102cfu/ml;以含有gyr B的质粒为模板,最低检测极限可以达到100 copies/μl;建立gyr B和gyr B-IAC标准曲线,Ct值与模板拷贝数均呈良好线性关系(r2=0.999);人工污染初始菌量为7 cfu/25 g时,样品中副溶血性弧菌增菌6 h即可检出。结论本研究所建立的gyr B-IAC实时荧光定量PCR方法,既能有效检测食品中副溶血性弧菌,又能实时监测PCR反应过程,有效防止\"假阴性\"的发生,结果可靠,有利于实现海产品中副溶血性弧菌实时荧光定量PCR检测方法的标准化。