利用博达法测试奶牛隐性乳房炎试验报告

利用美国近年研究并应用于实践的博达R牛奶测试技术对太原某牛场奶牛隐性乳房炎进行监测,试验显示,博达R牛奶测试技术方法简便快捷、测定数据准确,具有较大推广应用价值;牛场奶牛隐性乳房炎头阳性率为55.5%(111/200),乳区阳性率为26.34%(196/744);3胎及以上奶牛阳性率达到70.18%(40/57)并显著高于1-2胎35.42%(17/48)(P<0.01);奶牛个体混合乳样体细胞与个体乳区乳样体细胞平均数呈直线相关,差异极显著(r=0.7418,t>t0.01,P<0.01)。     

乳房炎牛乳中少动鞘氨醇单胞菌的分离鉴定和药敏试验

为了对陕西省某奶牛养殖场出现乳房炎的病例进行确诊,无菌采集患病奶牛的乳汁对其进行细菌分离、培养特性观察、生化试验、16S rRNA基因测序、动物致病性试验以及药敏试验。结果表明,分离到的细菌,其培养特性和生化试验结果与少动鞘氨醇单胞菌相吻合;16S rRNA基因测序结果与少动鞘氨醇单胞菌的同源性为98%;小鼠致病性试验表明该菌株有致病性;药敏试验结果表明,该菌对环丙沙星、诺氟沙星、复方新诺明等耐药,对多黏菌素、阿米卡星、亚胺培南、庆大霉素、头孢曲松等敏感。     

奶牛乳腺炎病原菌的分离及其药敏试验

在临床调查的基础上,采用普通琼脂培养分离临床型乳腺炎奶牛的病原菌,并用平板纸片法进行药物敏感试验,以确定农场奶牛临床型乳腺炎的致病菌种类及致病菌对不同抗生素的敏感性.结果发现,患临床型乳腺炎奶牛的主要致病菌是葡萄球菌和链球菌,有部分大肠杆菌;泰诺、氨苄青霉素钠、先锋9号和盐酸环丙沙星的抑菌效果较好.结合临床治疗效果观察发现,盐酸环丙沙星对乳腺炎的治疗效果最佳.     

Evaluation of a portable test system for assessing endotoxin activity in raw milk

The aim of the present study was to compare endotoxin activities detected in raw milk samples obtained from cattle by a commercially available portable test system (PTS) and traditional microplate limulus amebocyte lysate (LAL)-based assay, which determined activities using a kinetic turbidimetric (KT) assay. Raw milk samples were obtained from 53 and 12 dairy cattle without and with clinical mastitis, respectively. Comparison between the KT and PTS was performed by the Friedman test. The Pearson product moment correlation coefficients were calculated to evaluate associations between any two continuous variables. Linear regression model analysis was also performed to obtain the equation describing the relationship between PTS and KT assay. The endotoxin activities detected in 200- or 400-fold diluted milk samples were similar between PTS and KT assay, whereas a significant difference was observed in 100-fold diluted milk (P<0.001). The results obtained from 200- (r²=0.778, P<0.001) and 400-fold diluted milk samples (r²=0.945, P<0.001) using PTS correlated with those using KT assay. The median milk endotoxin activities in Gram-positive and Gram-negative clinical mastitis cows were 0.655 and 11,523.5 EU/ml, respectively. The results of the present study suggest that PTS as a simple and easy test to assess endotoxin activity in raw milk is efficient, simple and reproducible.     

隐性乳腺炎患牛乳汁电导率变化与体细胞数相关关系的研究

以随机采集的奶牛乳样为研究对象,对乳样的电导率与体细胞数进行了测定与分析。结果显示,体细胞在20-50万/mL区间时,奶牛乳汁电导率值在0.40-0.55mho/m之间;体细胞在50万/mL以上时,奶牛乳汁电导率0.6mho/m以上。结果表明,牛乳电导率的变化与体细胞的含量呈正相关,可确定利用牛乳体细胞数与电导率变化的相关关系来准确判断奶牛隐性乳房炎。     

Staphylococcal Nuclease in Udder Secretions of Cows with Acute Mastitis

High concentrations of nuclease produced by Staphylococcus aureus were demonstrated within a few hours by direct examination of udder secretions from cows with a severe staphylococcal mastitis. A positive correlation between the nuclease concentration and the severity of the mastitis was found. The cows with high nuclease concentrations were generally young individuals and/or in the first 2 weeks of the lactation period. Most had a low titre of antibodies against staphylococcal nuclease. Two-thirds of the cows in which high nuclease concentrations were demonstrated were culled or died because of the mastitis attack.The role which nuclease plays for staphylococcal virulence is discussed, and it is concluded that nuclease contributes to the pathogenicity of S. aureus.     

快速鉴别奶牛乳腺炎细菌或真菌的双重PCR方法

综合多种细菌、真菌DNA模板的提取方法,采用β-巯基乙醇裂解细胞壁,利用蜗牛酶和溶菌酶消化细胞壁,再利用石英砂机械破壁,能彻底破坏细菌、真菌细胞壁,提取高质量的DNA,摸索出一套从奶样中同时提取细菌、真菌核酸的新方法。建立双重PCR方法,同时完成对细菌16SrRNA保守区特异性基因片段和真菌18SrRNA保守区特异性基因片段的扩增,快速诊断奶牛乳房炎是细菌感染还是真菌感染或是混合感染。对采自临床型乳腺炎和隐性乳腺炎病例的共计84个乳样分别用传统细菌学培养法和双重PCR方法对比鉴定。结果表明,双重PCR检测乳样细菌的最小浓度为10^2CFU／mL,检测乳样真菌的最小浓度为10^3CFU／mL。双重PCR方法对细菌的检测与平板培养检测方法比较差异不显著（P〉0．05）,对真菌的检测与平板培养检测方法比较具有更高的检出率（P〈0．01）。     

奶牛急性乳房炎病原菌的分离与鉴定

[目的]为了掌握奶牛急性乳房炎致病菌的种类.[方法]通过采集病牛乳汁样品3份,分别对其病原茵进行分离与鉴定.检测[结果]表明:检出4种共13株细菌,其中致病性金黄色葡萄球菌6株;停乳链球菌5株;大肠埃希茵1株,乳房链球茵1株.[结论]该场奶牛的急性乳房炎是由致病性金黄色葡萄球菌、停乳链球菌、乳房链球菌和大肠杆菌共同引起...     

Mastitis Whey — a Good Medium for Bacteria?

Growth of mastitis pathogenic bacteria was measured in bovine whey samples by a turbidometric microtechnique. Whey from mastitis cows supported growth as compared with whey prepared from normal milk. Blood proteins leak into milk during mastitis. A study was undertaken to analyze which molecules from blood would promote bacterial growth in whey Fractions containing hemoglobin showed a distinct growth-promoting effect. An inadequate iron supply is one of the restricting growth factors for bacteria in milk. By utilizing heme-compounds the pathogens can by-pass the effect of antimicrobial iron-binding present in milk in the form of lactoferrin.     

山东潍坊市部分奶牛场奶牛乳房炎病原菌的分离鉴定及耐药性检测

为了解潍坊市当前奶牛乳房炎的发病以及病原菌的种类和耐药性情况,为该地区奶牛乳房炎的防治提供参考,采集潍坊市6个奶牛场临床型和隐性型乳房炎奶样各100份,进行了细菌分离和鉴定。结果细菌检出率为91.50%（183/200）,细菌混合感染奶样占检出细菌奶样的84.70%（155/183）;共分离到265株细菌,其中链球菌（无乳链球菌、停乳链球菌、乳房链球菌）占38.11%,金黄色葡萄球菌占34.34%,大肠杆菌占16.98%,这些是引起奶牛乳房炎的主要病原菌。将这些病原菌进行药物敏感试验,结果显示,敏感率高的药物依次是头孢喹肟（80.59%）、恩诺沙星（68.35%）、头孢噻呋（68.35%）和氟苯尼考（67.51%）,耐药率高的药物依次是青霉素（37.97%）、链霉素（26.16%）和四环素（10.55%）。     

奶牛隐性乳房炎病原菌的分离鉴定及药敏试验

为了解贵阳市某乳业有限公司奶牛场隐性乳房炎流行情况及其主要病原菌,采用CMT方法检出隐性乳房炎乳样后,对阳性乳样进行细菌分离与鉴定,同时对其主要病原菌进行了体外药敏试验。结果表明,该公司奶牛场隐性乳房炎的感染率为40％,主要病原菌为葡萄球菌属和链球菌属细菌,其敏感的抗菌药物有先锋V霉素、利福平、丙氟哌酸、氟哌酸、新霉素、卡那霉素、庆大霉素等,经临床用药观察,有效地控制了该公司奶牛场隐性乳房炎。     

奶牛乳房炎病原菌人工诱发奶山羊急性乳房炎试验

用从临床型乳房炎奶牛的乳样中分离的金黄色葡萄球菌、无乳链球菌、停乳链球菌 3种菌 ,经乳室注入不同的菌数感染泌乳奶山羊分别诱导急性乳房炎试验。结果表明 ,3种菌经奶山羊乳头注入均可引起程度不等的乳房炎症状 ,出现乳腺肿胀湿热、触之硬实或有痛感 ,乳汁颜色发生变化 ,奶中氯化物及细胞数均升高 ,产奶量下降 ,并伴有体温升高 ,心率增加等全身症状。这些病理变化的严重程度与攻菌菌株的毒力及攻菌量有一定的关系 ,其中以金黄色葡萄球菌攻菌后引起的乳房炎病理变化最为明显 ,无乳链球菌次之 ,停乳链球菌较轻。通过本试验初步测得了 3种菌人工感染奶山羊引起急性乳房炎的最小攻菌量 ,金黄色葡萄球菌为1 .52× 1 0 6个菌 ,无乳链球菌为 7.7× 1 0 6个菌 ,停乳链球菌为 3 .0× 1 0 5个菌 ,同时初步建立了人工感染泌乳奶山羊的乳房炎实验动物病理模型 ,为进一步研究乳房炎菌苗、病理学、抗菌药物药动学及药效学提供了实验基础     

Chelex-100法直接提取乳样中奶牛乳房炎主要致病菌DNA方法的建立

为建立一种直接从乳样中快速提取细菌DNA的方法,试验通过人工制备8个倍比稀释细菌的乳样,检测了Chelex-100法提取3种奶牛乳房炎主要致病菌(金黄色葡萄球菌、大肠杆菌和无乳链球菌)DNA进行PCR扩增的敏感性,并与苯酚—氯仿法进行了比较分析。结果显示,以Chelex-100法提取乳样中细菌DNA进行PCR扩增具有较高的敏感性,所检测金黄色葡萄球菌、大肠杆菌和无乳链球菌的最小浓度分别为10³、10²、10² CFU/mL;而使用苯酚—氯仿法提取乳样中各细菌DNA的PCR敏感性均为10⁴ CFU/mL。综上所述,Chelex-100法提取乳样细菌DNA的PCR敏感性可以满足临床检测奶牛乳房炎的需要,显现了简单快速、经济、无污染的优点,为从乳样中直接提取细菌DNA提供了新的思路,对PCR快速检测乳房炎致病菌具有重要意义。     

Association of California Mastitis Test Scores with Intramammary Infection Status in Lactating Dairy Cows Admitted to a Veterinary Teaching Hospital

Background

Subclinical mastitis is of concern in veterinary hospitals because contagious mastitis pathogens might be unknowingly transmitted to susceptible cows and then back to their farm of origin.

Objectives

To evaluate the California mastitis test (CMT) as an indicator of intramammary infection (IMI) in lactating dairy cows admitted to a veterinary hospital.

Animals

A total of 139 admissions of 128 lactating dairy cows admitted to the University of Illinois Veterinary Teaching Hospital over a 2‐year period.

Methods

A retrospective study with a convenience sample was conducted. Medical records of cows with CMT results and milk culture results for the day of admission were reviewed. Breed, age, season, maximum CMT score for the 4 quarters, maximum CMT score difference, and clinical diagnosis were evaluated as predictors of IMI by the chi‐square test and stepwise logistic regression.

Results

An IMI was identified in 51% of quarters. For cows admitted without evidence of clinical mastitis, the sensitivity of a CMT score ≥trace in predicting an IMI on a quarter or cow basis was 0.45 and 0.68, respectively. The distributions of maximal quarter CMT score and the maximum difference in quarter CMT score for cows without evidence of clinical mastitis did not differ (P = 0.28, P = 0.84, respectively) for cows with and without IMI. Stepwise logistic regression did not identify significant predictors of IMI in cows without clinical mastitis.

Conclusions

Lactating dairy cattle admitted to a veterinary hospital should be managed as if they have an IMI, even in the absence of clinical mastitis.     

陕西地区临床型奶牛乳房炎病原菌的分离鉴定及耐药性分析

为了分析陕西省奶牛乳房炎的主要致病性病原菌及其耐药性,为奶牛乳房炎的治疗提供依据。对陕西铜川、杨凌和宝鸡等地奶牛场患临床型奶牛乳房炎的48头奶牛进行奶样采集,并进行病原菌的分离鉴定及耐药性分析,根据培养特性、形态特征及生化试验鉴定结果,将分离得到的54株病原菌分为9种,分别为链球菌(15/54)、大肠埃希菌(12/54)、金黄色葡萄球菌(10/54)、绿脓杆菌(8/54)和志贺氏菌(5/54),药敏试验结果表明,分离的54株病原菌对13种常用抗菌素均具有一定的耐药性。     

Aquagrams of Raw Milk for Oestrus Detection in Dairy Cows

The purpose of this research was to develop rapid and cost‐effective method for oestrus detection in dairy cows by means of near infrared spectroscopy and aquaphotomics, using raw milk from individual cows. We found that aquaphotomics approach showed consistent specific water spectral pattern of milk at the oestrus periods of the investigated Holstein cows. Characteristic changes were detected especially in foremilk collected at morning milking. They were reflected in calculated aquagrams of milk spectra where distinctive spectral pattern of oestrus showed increased light absorbance of strongly hydrogen‐bonded water. Results showed that monitoring of raw milk near infrared spectra provides an opportunity for analysing hormone levels indirectly, through the changes of water spectral pattern caused by complex physiological changes related to fertile periods.     

内蒙古地区乳房炎奶样与环境中细菌的分离鉴定及耐药性分析

【目的】研究内蒙古地区部分奶牛养殖场乳房炎生鲜乳样本及环境样本致病菌种类和耐药性,为乳房炎的防控提供临床用药指导和参考。【方法】采用平板划线法对内蒙古地区4个规模化养殖场468份(乳房炎生鲜乳样本199份,乳房炎奶牛饲养环境样本269份)样本进行细菌分离培养,采用形态学观察、革兰氏染色镜检以及16S rDNA测序对分离出的细菌进行鉴定,且对引发乳房炎的主要致病菌进行药敏性试验。【结果】高盐甘露醇培养基上呈现出小、白且偏黄色的菌落,经16S rDNA测序比对,与金黄色葡萄球菌高度相似的菌株共56株,分离率为11.97%;在伊红-美蓝培养基上呈现出黑紫色泛有金属光泽的菌落,经16S rDNA测序比对,与大肠杆菌高度相似的菌株共44株,分离率为9.40%。在199份生鲜乳样本中,金黄色葡萄球菌和大肠杆菌的分离率最高,分别为21.11%和17.09%;在269份环境样本中,金黄色葡萄球菌和大肠杆菌的分离率分别为5.20%和3.72%。药敏试验结果显示,金黄色葡萄球菌对青霉素、磺胺异噁唑、氨苄西林、阿莫西林/克拉维酸的耐药性相对较高,耐药率分别为90.63%、78.13%、75.00%、68.75%;对庆大霉素、环丙沙星、复方新诺明的敏感性较高。大肠杆菌对氨苄西林、头孢噻吩、磺胺异噁唑存在明显耐药性,耐药率分别为100%、94.29%、45.71%;对链霉素、卡那霉素、庆大霉素、多粘菌素E、美罗培南、阿莫西林/克拉维酸以及利福昔明表现出较高敏感性。【结论】内蒙古地区引发奶牛乳房炎的主要致病菌为金黄色葡萄球菌、大肠杆菌;金黄色葡萄球菌对青霉素、磺胺异噁唑、氨苄西林、阿莫西林/克拉维酸的耐药性相对较高;大肠杆菌对氨苄西林、头孢噻吩、磺胺异噁唑存在明显的耐药性。     

青海省某奶牛场顽固性乳房炎主要病原菌的分离鉴定与耐药性研究

为了明确青海省某奶牛场顽固性乳房炎的病原菌,并制定有效的防控措施,本试验采用细菌分离、16SrDNA序列分析、细菌基因分型等方法对病牛乳样中的病原菌进行分离鉴定,并进行17种药物的敏感性试验。试验从15份乳样中分离到10株金黄色葡萄球菌、1株表皮葡萄球菌和2株化脓隐秘杆菌,其中10株金黄色葡萄球菌均为荚膜多糖5型分离株;所分离金黄色葡萄球菌对青霉素G、红霉素、头孢氨苄和呋喃妥因耐药率分别为100%、90%、10%和10%;表皮葡萄球菌对青霉素G、复方新诺明和甲氧苄氨嘧啶耐药,而对其他14种抗菌药物敏感;化脓隐秘杆菌对四环素和甲氧苄氨嘧啶耐药,对强力霉素和复方新诺明中度敏感,对其他13种抗菌药物敏感。该牛场顽固性乳房炎是多种病原菌混合感染所致,但以荚膜多糖5型金黄色葡萄球菌为主。不同的病原菌的药物敏感性不同,故而治疗困难,建议使用恩诺沙星治疗病牛并加强隐性乳房炎的筛查与防治,病情得到控制。     

鉴别显色培养基在奶牛乳房炎病原菌快速鉴定中的应用

为研发用于奶牛乳房炎常见病原菌分离鉴定的鉴别显色培养基,并检验其与普通分离鉴定培养基辅以生化鉴定和16S rRNA核酸序列测定两种传统方法鉴定结果的一致性,笔者以生物信息学方法筛出各种属细菌的特异性酶、可作为唯一碳源发酵产酸的碳源,然后合成酶显色底物,连同碳源、酸碱指示剂制备显色培养基,将各种属标准菌株、野生型菌株涂布培养,评估菌落形态、菌落颜色及培养基基质颜色变化。采集隐性乳房炎(CMT法检测)及临床型乳房炎病例的牛奶样品(絮状物、凝块、清亮状或血乳)共计482份,用鉴别显色培养基和两种传统方法进行病原菌的分离、鉴定。鉴别显色培养基上菌落纯化后提取DNA用于16S rRNA序列扩增,产物送去测序(n=194)。以两种传统方法为参考,判定鉴别显色培养基鉴定病原菌的可靠性,用SAS 9.4的FREQ程序计算鉴别显色培养基的简单科恩κ系数。鉴别显色培养基上常见乳房炎病原菌不同种属的菌落形态和培养基基质颜色明显不同,肉眼即可辨别。鉴别显色培养基与两种传统方法鉴定结果的一致性参数分别为κ=0.70、κ=0.96。鉴别显色培养基能够作为常见奶牛乳房炎病原菌的标准鉴定方法。     

宁夏地区奶牛乳房炎源性放射根瘤菌的分离鉴定与药敏试验

从宁夏地区部分奶牛场采集164份患有乳房炎奶牛的乳样,采用 VITEK-2全自动微生物鉴定系统进行病原菌的分离鉴定,共分离鉴定出放射根瘤菌30株,分离率高于奶牛乳房炎常见的病原菌溶血葡萄球菌、巴氏杆菌、金黄色葡萄球菌等;15种抗菌药物的药敏试验结果表明,放射根瘤菌对阿莫西林、林可霉素、先锋霉素Ⅴ、头孢曲松已产生不同程度的耐药性,而其他磺胺类、氨基糖苷类、喹诺酮类、头孢类中的11种药物则表现出较好的抑菌效果。