奶牛乳汁体细胞数的快速检测及其临床意义

本文用新型便携式体细胞计数仪C-Reader对奶牛乳汁体细胞数(SCC)进行快速、精确地检测,同时对乳样中的病原菌进行分离鉴定,旨在指导奶牛乳房炎的诊断、预防及治疗。选用北京及河北地区10个奶牛场的111头奶牛共428份奶样,进行SCC计数。SCC20万/mL的奶样占63.6%,SCC介于20万/mL～50万/mL的奶样占13.1%,SCC50万/mL占23.3%。检测的111头奶牛中,60头奶牛患有隐性乳房炎,头发病率54.1%,乳区发病率23.3%。隐性乳房炎SCC变化范围50.6万/mL～984.3万/mL。对隐性乳房炎乳样进行致病菌分离培养,其阳性率高达82.0%。共鉴定出100株致病菌,其中金黄色葡萄球菌64株,表皮葡萄球菌11株,大肠杆菌8株,链球菌10株,其他菌7株。结果表明,C-Reader计数仪能够快速准确地检测北京及河北地区的奶牛隐性乳房炎,其病原菌以金黄色葡萄球菌为主。     

乳房炎对中国荷斯坦牛测定日泌乳性能及体细胞数变化的影响

旨在研究乳房炎及病原菌组成对中国荷斯坦牛测定日泌乳性能及体细胞数变化的影响.本研究于2009年9月对江苏某奶牛场140头成年中国荷斯坦泌乳牛进行乳房炎检测和DHI测定,同时对隐性乳房炎阳性乳区进行病原菌分离鉴定,并分析了不同乳房炎类型和隐性乳房炎病原菌组成对测定日泌乳性能和乳中SCC变化的影响.结果表明,该牛场临床乳房炎、隐性乳房炎和正常奶牛比例分别为10.00％、51.10％和37.90％,其中隐性乳房炎以大肠杆菌和链球菌混合感染最多(19头,占26.00％),其次为链球菌单独感染(17头,占23.30％).由1种细菌单独感染的比例为26头(占36.11％),2种或2种以上细菌混合感染为44头(占61.11％).乳房炎类型对测定日产奶量、乳糖含量、体细胞数和体细胞评分4个指标影响达到极显著水平(P＜0.01),临床乳房炎和隐性乳房炎奶牛日产奶量和乳糖含量极显著低于正常奶,同时体细胞数和体细胞评分极显著高于正常奶,而临床乳房炎奶和隐性乳房炎奶在所有检测指标上均无显著差异,但临床乳房炎奶比隐性乳房炎奶体细胞数高出约100万·mL-1.不同病原菌组成类型对乳中体细胞数和体细胞评分有极显著影响(P＜0.01),而对其它性状无显著影响(P＞0.05).混合感染奶牛测定日产奶量极显著低于单独感染奶牛(P＜0.01),同时乳中体细胞数显著高于单独感染奶牛(P＜0.01).乳房炎类型对乳中SCC的变化趋势有显著影响(P＜0.01),正常奶牛维持较低SCC的能力较强,隐性乳房炎奶牛在下一泌乳月体细胞增加的可能性较大.不同隐性乳房炎病原菌组成类型对乳中SCC的变化无显著影响.奶牛生产中隐性乳房炎发病率高,危害性也最大,不同病原菌混合感染引起的隐性乳房炎对测定日产奶量和乳中体细胞数影响最大.该结果为奶牛场采取合理措施,降低隐性乳房炎发生率提供了参考.     

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%）。     

牛乳体细胞数的控制研究及实践

为了检验"牛博士Ⅲ"体细胞数检测仪的可信度,控制奶牛隐性乳房炎的指标SCC(体细胞数),同时采集两份牛奶样品,其中一份送达湖南省奶牛生产性能测定中心进行检测;分析乳腺炎的成因,提出了控制SCC的措施。结果表明,"牛博士Ⅲ"与丹麦福斯公司FossomaticTMFC体细胞仪检测结果的相对误差小于10%,相对标准偏差小于5%,处于可接受范围。牛奶中的菌落总数小于5×104CFU/mL,SCC控制在30万个/mL左右。     

牦牛隐性乳房炎的调查及致病菌的分离鉴定

为调查阿坝州红原县牦牛隐性乳房炎的发病情况,无菌采取68份无临床乳房炎牦牛奶样,采用体细胞计数、细菌分离培养、革兰染色与PCR相结合的方法分离鉴定细菌种属,并统计分析体细胞数与细菌分离情况的相关性。结果显示,牦牛奶样体细胞数从4 000个/mL～240 000个/mL不等;各种细菌阳性奶样数分别为金黄色葡萄球菌1份,溶血性葡萄球菌30份,非溶血性葡萄球17份,大肠埃希菌2份,乳酸乳球菌18份和藤黄微球菌2份;未分离到无乳、停乳、乳房链球菌。结果表明,分离到溶血性葡萄球菌的牦牛奶样体细胞平均数显著高于未分离到溶血性葡萄球菌的牦牛奶样体细胞的平均数(P0.05),当牦牛奶中体细胞数大于90 000个/mL时,溶血性葡萄球菌的阳性率大于80%(10/12)。研究结果提示溶血性葡萄球菌是引起牦牛隐性乳房炎的病原菌,为牦牛隐性乳房炎的防控提供了依据。     

新疆褐牛与荷斯坦牛隐性乳房炎与体细胞数、病原菌种类的比较

本试验以新疆乌鲁木齐市地区一大中型奶牛场的新疆褐牛、荷斯坦牛奶样为研究对象,对奶样进行了病原菌分离培养,同时对不同品种牛体细胞数与菌落形成单位（colony-forming unit, CFU）、体细胞数与不同年龄进行了统计分析。结果表明,2个品种牛感染病原菌种类稍有差别且新疆褐牛感染葡萄球菌的机率明显高于荷斯坦牛;荷斯坦牛比新疆褐牛更容易患乳房炎;新疆褐牛、荷斯坦牛体细胞在小于20万/mL时均与体细胞大于50万/mL时的CFU值呈显著差异（P＜0.05）,而这2个品种牛的体细胞在20～50万/mL时分别与体细胞小于20万/mL、大于50万/mL时的CFU值均无显著差异（P＞0.05）,初步推测体细胞数和细菌总数之间没有必然联系,还受其它因素影响;此外,随着年龄增长,褐牛和荷斯坦牛的体细胞评分（somatic cell score,SCS）都呈上升趋势,且不同品种间的SCS差异显著。     

一个高体细胞数牧场奶牛隐性乳腺炎病原菌的调查

对安徽某规模牧场随机采集的314份奶样进行细菌分离鉴定、牛奶体细胞计数(SCC),并对分离比例较高的凝固酶阴性葡萄球菌(CNS)进行亚种鉴定及药敏试验。结果显示120个低体细胞数奶样(SCC25×104/mL)的细菌检出率只有8.3%,分离细菌为CNS;而194个高体细胞数奶样(SCC≥25×104/mL)细菌检出率高达72.2%,分离出的细菌主要为金黄色葡萄球菌、CNS、大肠杆菌、停乳链球菌等。其中CNS感染比例最高、为28.8%,主要包括松鼠葡萄球菌、表皮葡萄球菌、人型葡萄球菌等。乳腺内细菌感染可引起患病乳区SCC显著升高(P0.05),松鼠葡萄球菌和表皮葡萄球菌感染也可引起SCC显著升高(P0.05),不同亚种的CNS对临床常用抗菌药物青霉素、庆大霉素、红霉素有较高的耐药率,分别为71.2%、53.8%和82.7%。     

山东不同地区乳房炎和非乳房炎牛奶中主要病原菌的调查

通过对13个地区35个牧场的985份非乳房炎奶样和284份乳房炎奶样进行常见病原菌分离鉴定,以期了解山东地区牛奶中主要病原菌的流行情况。采用常规方法与生化鉴定、PCR技术对样品进行细菌分离鉴定,利用系统进化群试验对大肠杆菌致病性进行分析。结果显示:(1)获得金黄色葡萄球菌150株(11.82%),大肠杆菌248株(19.54%),无乳链球菌35株(2.76%),停乳链球菌18株(1.42%),乳房链球菌49株(3.86%);乳房炎样品细菌检出率(51.41%)明显高于非乳房炎样品(35.94%)(P<0.01);(2)青岛地区样品检出率(67.91%),高于其他地区(P<0.05),金黄色葡萄球菌在青岛检出率(50.0%)最高,大肠杆菌在东营的检出率(40.0%)最高;乳房链球菌在日照的检出率(31.25%)最高,无乳链球菌和停乳链球菌在各地的检出率无明显差异;(3)6月份样品中细菌检出率(67.91%)最高,且明显高于其他采样时间(P<0.01);(4)248株大肠杆菌中,共生群A群213株(85.89%),致病群35株(14.11%),乳房炎样品致病群检出率(22.39%);明显高于非乳房炎样品(11.05%)(P=0.023)。表明山东省牛奶中主要致病菌为金黄色葡萄球菌,乳房炎样品细菌和致病群检出率均高于非乳房炎样品,不同地区致病菌检出率存在差异,且夏季最高,秋冬季节低。     

不同饲喂方式下西南地区中国荷斯坦牛乳中体细胞数变化规律研究

为研究TMR日粮和精粗料分别饲喂方式下,中国荷斯坦奶牛乳中体细胞数(SCC)的变化规律.用SPSS13.0-般线性模型(GLM)对西南地区某奶业集团中国荷斯坦牛27 528次奶牛群改良计划(DHI)测定日记录进行了分析.结果表明:以TMR日粮饲喂方式下,体细胞数正常奶牛头次数(SCC<50万个/mL)占78.7%,患严重乳房炎的奶牛头次数(SCC>200万个/mL)占4.3%.乳中体细胞数(SCC)平均值为41.2±67.9万个/mL;而以精粗料分别饲喂方式下,SCC正常的奶牛头次数占77.3%,患严重乳房炎的奶牛头次数占4.5%,SCC平均值为42.4±68.9万个/mL.两种饲喂方式下二者SCC差异不显著(P>0.05).不同月份、产犊季节、泌乳月和胎次对两种饲喂方式乳中SCC的影响都达到极显著水平(P<0.01).两种饲喂方式下SCC都随着泌乳月和胎次的增加有升高的趋势,夏季SCC显著高于冬季.     

牦牛隐性乳房炎病原菌的分离及鉴定

牦牛隐性乳房炎被认为是高原藏族地区牦牛奶产量低下的主要原因之一。探索和研究牦牛隐性乳房炎致病菌特性及流行病学特征,可为牦牛隐性乳房炎防治提供一定的依据。本试验采集了红原地区不同牧群牦牛的80份奶样,由体细胞数、p H值测定初步筛选出了30份疑似牦牛乳房炎奶样,然后对其进行培养观察及生化检测,分离并鉴定出了牦牛隐性乳房炎病原菌。结果显示:80份奶样中有28份为牦牛隐性乳房炎奶样,检出率为35%;细菌学检测表明,28份奶样中金黄色葡萄球菌占10.71%,无乳链球菌占25%,乳房链球菌占32.14%,停乳链球菌占17.86%,大肠杆菌占14.29%。     

我国规模牛场奶牛个体生鲜乳体细胞数水平及影响因素探析

本研究旨在探析我国规模奶牛场奶牛个体生鲜乳体细胞数（SCC）的水平、分布和影响因素。利用SAS9.0的GLM模型,统计分析覆盖16个省（市、区）,33个规模奶牛场,23 351头中国荷斯坦牛,从2007年至2009年连续3年的225 775条奶牛个体生鲜乳SCC检测记录。结果表明：奶牛个体生鲜乳SCC的总体均值为48×104cell/mL,标准差为117×104cell/mL,个体生鲜乳SCC波动范围较大;其中,77.9%的奶牛个体,其生鲜乳SCC小于50×104cell/mL,对群体混合样生鲜乳SCC的影响系数为0.26;11.3%的奶牛个体,其生鲜乳SCC介于50×104～100×104cell/mL之间,乳房处于隐性感染状态,对群体混合样生鲜乳SCC的影响系数为0.16;10.8%的奶牛个体,生鲜乳SCC大于100×104cell/mL,理论上乳房处于临床感染状态,对群体混合样生鲜乳SCC的影响系数为0.58。奶牛个体生鲜乳体细胞数评分（SCS）呈正态分布,与奶牛个体因素（产奶量、乳脂率、乳蛋白含量、胎次和泌乳阶段）和环境因素（泌乳月份、泌乳季节）差异极显著（P〈0.01）,与奶牛养殖区域、奶牛场差异不显著（P〉0.05）。我国规模奶牛场奶牛个体生鲜乳SCC主要在50×104cell/mL以下,依据影响参数能实现对生鲜乳SCC水平的调控。     

奶水牛乳中体细胞数与乳成分的相关分析

乳汁体细胞数(somatic cell count,SCC)是检查奶牛乳房炎和牛奶质量的指标,为了解南宁某规模化奶牛场奶水牛SCC情况及SCC与其他乳成分的关系,试验随机选取该场152份泌乳早期乳样,测定其SCC、乳汁比重、蛋白质、脂肪、总固形物、非脂固形物及乳糖的含量,并对其进行相关性分析。结果表明,SCC小于50万/mL的乳样占被检奶样总数的88.16%,大于100万/mL的占1.97%;乳汁比重、乳蛋白、乳糖和非脂固形物含量均随着SCC的升高而下降,而乳脂含量随着SCC的升高而呈上升趋势;SCC与乳糖及非脂固形物含量呈极显著负相关(P<0.01)。因此,奶水牛乳中SCC不仅与乳糖和非脂固形物含量密切相关,也提示其与奶水牛产后营养代谢状况有关。     

Preliminary studies on the prevalence of Mycoplasma bovis mastitis in dairy in cattle in Australia

A highly sensitive and specific PCR (MB-PCR) was used in preliminary studies to detect M. bovis in milk samples to investigate its association with high somatic cell count (SCC), an indicator of subclinical mastitis and one of the factors in down grading the quality of milk. A total of 186 and 167 herds were tested with 43% and 62% of herds positive for M. bovis in Victoria and North Queensland, respectively. The quarter milks from 52 cows with persistently high SCC were tested by MB-PCR and culture to investigate the association of M. bovis with major mastitis pathogens (MMP). M. Bovis was detected in 77% of cows of which 19% alone had M. bovis without any other bacteria, 17% had M. bovis in combination with major mastitis pathogens and 40% had M. bovis in combination with non-major mastitis pathogens. We believe that M. bovis is widespread in dairy cattle and has the potential to produce disease alone or to predispose the udder to disease caused by major mastitis and environmental pathogens. These studies have revealed a hitherto unrecognised high prevalence of M. bovis in dairy cattle in North Queensland and Victoria in Australia. These initial studies also give a clear association between M. bovis and elevated somatic cell counts.     

牛乳电导率值与体细胞数相关性的研究

试验以随机采集的奶牛乳样为研究对象，对乳样的电导率值与体细胞数进行了测定与分析。结果显示，体细胞数在50万个/ml以下时，电导率值在4．40～4．75ms／cm之间；体细胞数在50万～500万个／ml时，电导率值在5．20～9．00ms／cm之间；体细胞数在500万／个ml时，电导率值在9．00ms／cm以上。结果表明，牛乳电导率的变化与体细胞数呈正相关，可利用牛乳体细胞数与电导率变化的相关关系判断奶牛隐性乳房炎发病程度。     

Between-herd prevalence of Mycoplasma bovis in bulk milk in Flanders, Belgium

Mycoplasma bovis (M. bovis) is a highly infectious pathogen of cattle causing pneumonia, polyarthritis, otitis, and less frequently, subcutaneous abscesses, abortions and meningitis. Ineffective drugs treatments, culling of infected cows and loss of milk production can lead to significant economic loss on dairy farms. The early detection of cows excreting M. bovis bacteria to prevent mastitis outbreaks is warranted. Reports suggest that the risk of M. bovis mastitis is higher in larger dairy herds. The objective of this study is to estimate the herd-level prevalence of M. bovis in Flanders, Belgium by culturing bulk tank milk samples taken from dairy farms. Three bulk tank milk samples per dairy herd were taken over four weeks, with collection intervals of two weeks. Culturing was done after pre-incubation using modified Hayflicks media to increase the chances of recovery of bacteria. For the identification of M. bovis, tDNA intergenic spacer PCR was used. In three herds (1.5%) of the 200 herds sampled, M. bovis was isolated from one of the three consecutive bulk tank milk samples. We conclude that in Flanders in 2009 at least 1.5% of the dairy herds had one or more cows excreting M. bovis in the milk. The frequent monitoring of bulk tank milk to detect the presence of M. bovis, especially in expanding herds on farms that often purchase replacement animals, should be encouraged in order to detect the presence of M. bovis and to monitor the success of control procedures following an outbreak of mycoplasmal mastitis in the herd.     

牛支原体、巴氏杆菌A型和化脓隐秘杆菌多重PCR快速检测方法的建立

本试验旨在建立一种可同时鉴别牛支原体、巴氏杆菌A型和化脓隐秘杆菌的多重PCR方法。分别针对多杀性巴氏杆菌A型特异的hyac-hvaD基因区段、化脓隐秘杆菌的16SrRNA基因上保守区段和牛支原体的UvrC基因设计特异性引物,多重PCR的最佳扩增条件确定为：95℃ 10min预变性;95℃ 1min,56℃ 50s,72℃ 1min,循环30次;72℃ 210min延伸。结果表明,该多重PCR方法可同时扩增出以上三种致病菌的特异性片段,不能扩增出其他病原菌的相关片段;对多杀性巴氏杆菌A型、化脓隐秘杆菌和牛支原体的最低检测浓度分别为8×10^5CFU／mL、8×10^5CFU／mL和4×10^6CFU／mL。同时用该方法检测了牛支原体肺炎患牛的鼻拭子与肺组织,发现12h预增菌后,肺组织检测与牛支原体培养的阳性符合率为92％。对临床样本进行牛支原体分离培养需要3-4d时间,而采用多重PCR方法检测12h预增菌则能在24h内出结果。该多重PCR方法显著加快了临床诊断速度,具有推广应用价值。     

Dynamics of lingual antimicrobial peptide,lactoferrin concentrations and lactoperoxidase activity in the milk of cows treated for clinical mastitis

The aim of the present study was to examine changes in innate immune factors in the milk of mastitic dairy cows treated with antibiotics. Cows in the antibiotics group (n = 13) were infused into the mammary gland with cefazolin on the sixth day after mastitis was diagnosed (the day of the mastitis diagnosis = day ?6). The control group (n = 12) was not treated. Milk samples were collected once every 2 days from days ?6 to 12 and somatic cell count (SCC), lingual antimicrobial peptide (LAP), and lactoferrin (LF) concentrations and lactoperoxidase (LPO) activity were measured. SCC and LF concentrations in the antibiotics group markedly decreased after the antibiotic treatment. When cows in the antibiotics group were divided according to SCC on day 0, LAP concentrations and LPO activity in cows with a lower SCC on day 0 (<5 × 10⁶ cell/mL) were significantly higher and lower than those in cows with a higher SCC, respectively. These results suggest that LF concentration decreased with decrease in SCC after treatment and that LAP concentration and LPO activity differed depending on the severity of mastitis. This is the first report to reveal the dynamics of innate immune factor in milk of cows treated for clinical mastitis.     

Reliability in somatic cell count measurement of clinical mastitis milk using DeLaval cell counter

Somatic cell counts (SCC) measurements are typically performed using quantitative methods, such as the Breed method (Breed) and the Fossomatic method (FSCC). The DeLaval cell counter (DCC) developed recently is a quantitative somatic cell counter with a low initial cost and superior portability. However, since the DCC was specifically developed for measuring SCC of ≤ 4 × 10⁶ cells/mL milk from bulk tanks or individual cows, its reliability for estimating SCC that exceed this concentration has not yet been clarified. This study therefore examined whether it is possible to accurately measure SCC by diluting milk samples with initial SCC of 4 × 10⁶ cells/mL, as seen in clinical mastitis milk. We collected milk samples from 99 quarters of 99 Holstein cows with clinical mastitis. These milk samples were diluted 10‐fold with saline and thoroughly mixed before performing SCC measurement with the DCC. The correlation coefficients of SCC measured by the FSCC, Breed and DCC methods indicated strong correlations between each pair of methods. The findings showed that DCC can be used to identify bovine clinical mastitis milk and is useful as a quantitative SCC measurement device on farm sites.     

Experimental infection of lactating bovine mammary glands with Streptococcus uberis in quarters colonized by Corynebacterium bovis

Twenty-seven quarters of 18 lactating dairy cows were inoculated intramammarily with 3.6 X 10(4) colony-forming units (CFU) of a strain of Streptococcus uberis isolated from a cow with clinical mastitis. Before quarters were inoculated, 22 were considered as naturally colonized with Corynebacterium bovis, and 5 were considered bacteriologically negative. Streptococcus uberis was isolated from all quarters within 2 days after inoculation, and all quarters developed clinical mastitis by 3 days after inoculation. Mastitis was acute, and most cows had increased rectal temperatures. The number of somatic cells increased significantly (P less than 0.05), and milk production decreased significantly. In many cows, rectal temperatures remained increased, and Str uberis was isolated from infected glands after intramammary and systemic antimicrobial treatments were given. A decreased number (110 CFU) of the same strain of Str uberis caused equally severe mastitis in 3 quarters colonized with C bovis and in 1 bacteriologically negative quarter in 2 cows. Streptococcus uberis was isolated from all inoculated quarters, and all quarters developed clinical mastitis by 2 days after inoculation. Two quarters colonized with C bovis and 2 bacteriologically negative quarters were inoculated once with 25 CFU and once with 240 CFU of a different strain of Str uberis (ATCC 27958). Streptococcus uberis was never isolated from inoculated quarters, and changes in milk yield or number of somatic cells were not observed.