Mastitis Pathogens Isolated from Samples of Milk in Dairy Cows and Their Resistance against Antimicrobial Agents

The objective of this study was to evaluate the effectiveness of different antibiotics against mastitis causing microorganisms during first phase of lactation in herd of 320 Holstein cows in east of Slovakia. Milk samples from quarters were cultured and identified bacteria were subjected to antimicrobial susceptibility test by disc diffusion method to a large number of antibiotics. The results revealed higher sensitivity against tetradelta (100% of Strep. Agalactiae and Strep. uberis, Coagulase Negative Staphylococci), (94.4% of Stap. aureus) with highest number of bacterial isolates, followed by cefalexin + kanamycin amoxicillin plus clavulanat acid and ceftiotur (100% of Strep. agalactiae and Strep. uberis). Resistance was observed against penicillin (22.2% of Stap. aureus) amoxicillin (22.2% of Stap. aureus and 10.5% of Strep. uberis) and streptomycin (22.2% of Stap. aureus and 52.7% of Strep. uberis).

Rumen microbiome structure and metabolites activity in dairy cows with clinical and subclinical mastitis

Background:Due to the high prevalence and complex etiology,bovine mastitis(BM)is one of the most important diseases to compromise dairy cow health and milk quality.The shift in milk compositions has been widely investigated during mastitis,but recent studies suggested that gastrointestinal microorganism also has a crucial effect on the inflammation of other peripheral tissues and organs,including the mammary gland.However,research focused on the variation of rumen inner-environment during mastitis is still limited.Therefore,the ruminal microbial profiles,metabolites,and milk compositions in cows with different udder health conditions were compared in the present study.Furthermore,the correlations between udder health status and ruminal conditions were investigated.Based on the somatic cell counts(SCC),California mastitis test(CMT)parameters and clinical symptoms of mastitis,60 lactating Holstein dairy cows with similar body conditions(excepted for the udder health condition)were randomly divided into 3 groups(n=20 per group)including the healthy(H)group,the subclinical mastitis(SM)group and the clinical mastitis(CM)group.Lactation performance and rumen fermentation parameters were recorded.And rumen microbiota and metabolites were also analyzed via 16S rRNA amplicon sequencing and untargeted metabolomics,respectively.Results:As the degree of mastitis increased,rumen lactic acid(LA)(P<0.01),acetate,propionate,butyrate,valerate(P<0.001),and total volatile fatty acids(TVFAs)(P<0.01)concentrations were significantly decreased.In the rumen of CM cows,the significantly increased bacteria related to intestinal and oral inflammation,such as Lachnospiraceae(FDR-adjusted P=0.039),Moraxella(FDR-adjusted P=0.011)and Neisseriaceae(FDR-adjusted P=0.036),etc.,were accompanied by a significant increase in 12-oxo-20-dihydroxy-leukotriene B4(FDR-adjusted P=5.97×10^(−9))and 10beta-hydroxy-6beta-isobutyrylfuranoeremophilane(FDR-adjusted P=3.88×10^(−10)).Meanwhile,in the rumen of SM cows,the Ruminiclostridium_9(FDR-adjusted P=0.042)and Enterorhabdus(FDR-adjusted P=0.043)were increased along with increasing methenamine(FDR-adjusted P=6.95×10^(−6)),5-hydroxymethyl-2-furancarboxaldehyde(5-HMF)(FDR-adjusted P=2.02×10^(−6))and 6-methoxymellein(FDR-adjusted P=2.57×10^(−5)).The short-chain fatty acids(SCFAs)-producing bacteria and probiotics in rumen,including Prevoterotoella_1(FDRadjusted P=0.045)and Bifidobacterium(FDR-adjusted P=0.035),etc.,were significantly reduced,with decreasing 2-phenylbutyric acid(2-PBA)(FDR-adjusted P=4.37×10^(−6)).Conclusion:The results indicated that there was a significant shift in the ruminal microflora and metabolites associated with inflammation and immune responses during CM.Moreover,in the rumen of cows affected by SM,the relative abundance of several opportunistic pathogens and the level of metabolites which could produce antibacterial compounds or had a competitive inhibitory effect were all increased.

Month-Wise Prevalence of Subclinical Mastitis in Dairy Cows in Guangdong Province,China

To evaluate the month-wise prevalence of subclinical mastitis （SM） and its relations with climatic temperature and humidity, and to provide references for control and prevention of mastitis in dairy cows in Guangdong, China, California mastitis test method was used to determine the monthly prevalence of SM （MPSM, %） of lactating Holstein cows from 11 dairy farms in Guangdong Province, China, including about 516 cows per monthly examination. The average MPSM on a cow and a quarter basis were 30 and 13% respectively, and there was a positive linear correlation between monthly mean air temperature （MT, °C） and MPSM on a cow basis （r=0.763, P=0.004） or a quarter basis （r=0.577, P=0.049）, but there were no correlations between MT and MPSM on a cow or a quarter basis （P〉0.05）. There was a shift in MPSM trend that the highest MPSM （38.4%） was not for the hottest July with MT at 30.8°C, but for the later September at 27.9°C. The farms need to develop new housing conditions to control cowshed temperatures in the hot season to reduce the mastitis prevalences in the post-hot months.

Experimental Intramammary Infection with a Strain of <i>Escherichia coli</i>Isolated from a Cow with Persistent <i>E. coli</i>Mastitis

Transient E. coli intramammary infections (IMI) are usually associated with rapid onset of clinical signs including mammary gland swelling and abnormal milk with rapid clearance of bacteria from milk. Conversely, reports have described strains of E. coli showing very different clinical trends. Persistent E. coli IMI are associated with mild clinical symptoms that disappear shortly after the onset of infection, possibly flaring-up intermittently during lactation. In the present study, we evaluated a strain of E. coli isolated from a cow with persistent mastitis to determine if the experimental infection model mimics naturally occurring persistent E. coli IMI. Uninfected mammary quarters of 7 Holstein heifers were infused within 10 days of calving with 50 colony-forming units of a persistent E. coli strain. Six of 7 heifers developed mild clinical mastitis with elevated rectal temperatures within 9 to 36 h after infusion. The challenge strain was isolated intermittently in milk from all infected mammary quarters during the first two weeks after infusion and 3 animals continued to shed E. coli periodically during the sampling period. One animal shed E. coli intermittently in milk for 172 d after challenge and developed clinical mastitis four times during this period. The isolated strain had an identical pulsed-field gel electrophoresis profile as the E. coli strain used to infuse mammary glands. The experimental IMI model described here mimics very closely naturally occurring persistent E. coli IMI, thus providing an excellent in vivo model to better understand pathogenesis and to facilitate development of control strategies for this important mastitis pathogen.

Feeding a Saccharomyces cerevisiae fermentation product improves udder health and immune response to a Streptococcus uberis mastitis challenge in mid-lactation dairy cows

Background:We aimed to characterize the protective effects and the molecular mechanisms of action of a Saccharomyces cerevisiae fermentation product(NTK)in response to a mastitis challenge.Eighteen mid-lactation multiparous Holstein cows(n=9/group)were fed the control diet(CON)or CON supplemented with 19 g/d NTK for 45 d(phase 1,P1)and then infected in the right rear quarter with 2500 CFU of Streptococcus uberis(phase 2,P2).After 36-h,mammary gland and liver biopsies were collected and antibiotic treatment started until the end of P2(9 d post challenge).Cows were then followed until day 75(phase 3,P3).Milk yield(MY)and dry matter intake(DMI)were recorded daily.Milk samples for somatic cell score were collected,and rectal and udder temperature,heart and respiration rate were recorded during the challenge period(P2)together with blood samples for metabolite and immune function analyses.Data were analyzed by phase using the PROC MIXED procedure in SAS.Biopsies were used for transcriptomic analysis via RNA-sequencing,followed by pathway analysis.Results:DMI and MY were not affected by diet in P1,but an interaction with time was recorded in P2 indicating a better recovery from the challenge in NTK compared with CON.NTK reduced rectal temperature,somatic cell score,and temperature of the infected quarter during the challenge.Transcriptome data supported these findings,as NTK supplementation upregulated mammary genes related to immune cell antibacterial function(e.g.,CATHL4,NOS2),epithelial tissue protection(e.g.IL17C),and anti-inflammatory activity(e.g.,ATF3,BAG3,IER3,G-CSF,GRO1,ZFAND2A).Pathway analysis indicated upregulation of tumor necrosis factorα,heat shock protein response,and p21 related pathways in the response to mastitis in NTK cows.Other pathways for detoxification and cytoprotection functions along with the tight junction pathway were also upregulated in NTK-fed cows.Conclusions:Overall,results highlighted molecular networks involved in the protective effect of NTK prophylactic supplementation on udder health during a subclinical mastitic event.

Identification of novel molecular markers of mastitis caused by Staphylococcus aureus using gene expression profiling in two consecutive generations of Chinese Holstein dairy cattle

Background: Mastitis in dairy cows caused by Staphylococcus aureus is a major problem hindering economic growth in dairy farms worldwide. It is difficult to prevent or eliminate due to its asymptomatic nature and long persistence of infection. Although transcriptomic responses of bovine mammary gland cells to pathogens that cause mastitis have been studied, the common responses of peripheral blood leukocytes to S. aureus infection across two consecutive generations of dairy cattle have not been investigated.Methods: In the current study, RNA-Seq was used to profile the transcriptomes of peripheral blood leukocytes sampled from S. aureus-infected mothers and their S. aureus-infected daughters, and also healthy non-infected mothers and their healthy daughters. Differential gene expression was evaluated as follows: 1) S. aureus-infected cows versus healthy non-infected cows(S vs. H, which include all the mothers and daughters), 2) S. aureus-infected mothers versus healthy non-infected mothers(SM vs. HM), and 3) S. aureus-infected daughters versus healthy noninfected daughters(SMD vs. HMD).Results: Analysis of all identified expressed genes in the four groups(SM, SMD, HM, and HMD) showed that EPOR,IL9, IFNL3, CCL26, IL26 were exclusively expressed in both the HM and HMD groups, and that they were significantly(P < 0.05) enriched for the cytokine-cytokine receptor interaction pathway. A total of 17, 13 and 10 differentially expressed genes(DEGs)(FDR Padj. < 0.1 and |FC| > 1.2) were detected in the three comparisons, respectively. DEGs with P < 0.05 and |FC| > 2 were used for functional enrichment analyses. For the S vs. H comparison, DEGs detected included CCL20, IL13 and MMP3, which are associated with the IL-17 signaling pathway. In the SM vs. HM and SMD vs. HMD comparisons, five(BLA-DQB, C1 R, C2, FCGR1 A, and KRT10) and six(BLA-DQB, C3 AR1, CFI, FCAR, FCGR3 A, and LOC10498484) genes, respectively, were involved in the S. aureus infection pathway.Conclusions: Our study provides insights into the transcriptomic responses of bovine peripheral blood leukocytes across two generations of cattle naturally infected with S. aureus. The genes highlighted in this study could serve as expression biomarkers for mastitis and may also contain sequence variation that can be used for genetic improvement of dairy cattle for resilience to mastitis.

Molecular characteristics,antibiogram and prevalence of multi-drug resistant Staphylococcus aureus (MDRSA) isolated from milk obtained from culled dairy cows and from cows with acute clinical mastitis

Objectives: To study the molecular characteristics, antibiogram and prevalence of multidrug resistant Staphylococcus aureus(S. aureus)(MDRSA) isolated from milk obtained from culled dairy cows and from cows with acute clinical mastitis.Methods: Bacteria were cultured from 188 quarter milk samples obtained from cows before culling(n=139) and from cows affected with acute mastitis(n=49) belonging to 10 dairy farms. The bacteria were identified using colony moiphology, Gram staining and biochemical characteristics. S. aureus isolates were then subjected to molecular characterization using PCR targeting 16 S rRNA and mecA gene to identify Methicillin resistant S. aureus(MRSA). The antibiogram of all isolates was performed using the Kirby-Bauer disk diffusion method against 10 commonly used antibiotics in dairy farms.Results: S. aureus was isolated from 19(13.7%) samples obtained from culled cows and 11(22.4%) samples obtained from cows with acute mastitis. In both culled cows and cows with acute mastitis, in vitro antibiogram revealed that 100% of S. aureus isolates were resistant to erythromycin, penicillin G, streptomycin, doxycyclin, and trimethoprim/sulpha. The prevalence of MRSA in milk of culled cows and cows with acute mastitis was 26.3% and 18.2%, respectively, with an overall prevalence of 3.7% among all samples. All MRSA isolates were completely resistant to all tested antibiotics. All MRSA isolates were positive for the presence of the mecA gene.Conclusions: MRSA carrying the mecA gene were isolated from mastitic milk from dairy cows in Jordan for the first time. MRSA may pose a potential health risk to the public, farm workers and veterinarians.

Effects of Xiaoruling on Subclinical Mastitis in Dairy Cows

[Objective] TO investigate the effects of traditional Chinese medicine （TCM） preparation ＂Xiaoruling＂ on the subclinical mastitis in dairy cows. [Method] A total of 28 dairy cows suffering from subclinical mastitis were randomly divided into experimental group （n =20） and control group （ n =8）. The dairy cows in the experimental group were treated with Xiaoruling for 2 weeks. [ Rsults] The cure rate of subclinical mastitis in dairy cows was up to 89.47% and milk yield increased greatly （ P 〈 0.05）. Meanwhile, the milk somatic cell count （SCC） was greatly decreased （ P 〈 0.01 ）. SOD and GSH-Px activities in plasma were increased greatly （ P 〈 0.01 ）. [ Conclusion] Xiaoruling has an effective treatment on subclinical mastitis in dairy cows.

Multiple PCR for Detection of Subclinical Mastitis in Dairy Cow

[ Objective] To develop a rapid, sensitive, specific and accurate diagnostic method for subclinical mastitis in dairy cow. [ Method] A total of 70 samples of suspected clinical mastitis cows were detected in gene level by multiple PCR. [Result] The 58 samples were positive with the developed multiple PCR method and the positive rate was 82.5% （58/70）. The 53 samples were positive with the traditional biochemical method and the positive rate was 75.5% （53/70）. As a result, the coincidence rate of both methods was 92%. [ Conclusion] The multiple PCR method is rapid and specific for the detection of pathogenic subclinical mastitis in dairy cow.

Differences in severity of mastitis and the pathogens causing various oxidative product levels

Differences in severity of mastitis and their causing pathogens might be associated with oxidative product levels occurring during the inflammatory process in infected udders. The objectives of this study were to determine the relationship of oxidative product levels, using malondialdehyde (MDA) as a marker on both mastitis severity and its causing pathogens. Quarter milk samples of all lactating cows in the selected farms were primarily tested for mastitis severity levels including clinical and subclinical mastitis. All quarter milk samples from mastitis cows were separately collected for MDA measurement and bacteriological analyses. Results showed that MDA in clinical mastitis quarters was highest in comparison to sub-clinical mastitis and healthy quarters. MDA in milk samples with Strep. uberis and coagulase negative staphylococci was higher than in milk samples without any pathogens (p < 0.05). In conclusion, differences in both severity of mastitis and mastitis pathogens were associated with differences of oxidative products in infected udders.

Comparative Proteomic Analysis of Plasma from Clinical Healthy Cows and Mastitic Cows

The current research presents the protein changes in plasma from healthy dairy cows and clinical mastitic cows using two- dimensional gel electrophoresis （2-DE）. After staining with silver nitrate and Coomassie Blue, differential expression proteins were detected by PDQuest 7.4 software, and then subjected to ion trap mass spectrometer equipped with a Surveyor HPLC System, differential spots of protein were identified. Three protein spots that originated from preparation gels were identified to be two proteins. Overall, haptoglobin precursor was up-regulated in cows infected with clinical mastitis and could be a mastitis-associated diagnostic marker, whereas SCGB 2A1 （secretoglobin, family 2A, member 1） was down-regulated protein. Plasma protein expression patterns were changed when cows were infected with mammary gland inflammation; it suggests that analysis of differential expression protein might be useful to clarify the mechanisms involved in the pathophysiology, and find new diagnostic markers of mastitis and potential protein targets for treatment.

基于SourceTracker分析牧场环境对乳房炎乳菌群的影响

乳房炎是奶牛常见的疾病之一,通常是由牧场环境中的病原微生物引起,因此,开展乳房炎病原微生物溯源研究对乳房炎防治具有重要意义。本研究从中国天津某牧场收集了乳房炎乳、牧场环境(空气、饮用水、饲料、粪便、垫料、新垫料及喷淋水)和挤奶厅(前药浴液、后药浴液、前药浴杯、后药浴杯、奶杯、乳头皮肤)样品共计74份,提取可培养微生物DNA和总DNA,进行16S rDNA扩增测序和SourceTracker分析。在采集的74份样品中,共检测到50个不同科水平上的细菌。Beta多样性和ANOSIM分析表明,样品间细菌群落相似性低。乳房炎乳和乳头皮肤及卧床样品的细菌群落结构相似。此外,SourceTracker结果发现,乳头皮肤是奶牛乳房炎的主要污染源,其次是空气、奶杯和粪便,这表明牧场管理会影响奶牛感染乳房炎的情况。因此,牧场应加强管理控制措施,减少或消除牧场中病原微生物的传播风险。

Preparation of Polyclonal Antisera of Dairy Cow S100A12 Protein

[Objective] This study aimed to prepare dairy cow anti-S100A12 antisem and develop a highly effective and sensitive immunological detection reagent for further investigation of the functions of dairy cow S100A12. [Method] Purified S100A12 protein was respectively emulsified with Freund＇s complete adjuvant and Freund＇s incomplete adjuvant as the antigen for immunizing New Zealand white rabbits to prepare the polyclonal antisera. The titer was detected using agar double diffusion assay and indirect enzyme-linked immunoserbent assay （ELISA） and the specificity was determined with Western Blot. [ Result ] The titer of anti- S100A12 antisera was 1： 8 as determined by agar double diffusion assay and over 1：409 600 by ELISA. Western Blot result showed that the polyclonal antisera could be specifically combined with S100A12 protein. [ Conclusion] The results indicated that anti-S100A12 polyclonal antibody with high fiter and high specificity was successfully obtained, which provided a novel tool for further investigation of the functions of S100A12 gene.

融合数据增强与改进ResNet34的奶牛热红外图像乳腺炎检测

乳腺炎是奶牛生产养殖中最为严重的疾病之一,奶牛乳腺炎的早期检测可以为后续治疗提供依据,从而提高疾病治疗效率,降低养殖风险。为了对自然行走的奶牛实现快速、高精度的“一步式”乳腺炎疾病检测,提出了一种基于热红外图像,融合数据增强与改进ResNet34的奶牛乳腺炎疾病检测方法。相对于现有的“多步式”奶牛红外图像乳腺炎检测方法,该方法无需奶牛关键部分如乳房和眼睛的定位以及温度提取等,可有效避免“多步式”造成的误差累计,从而实现更高效的乳腺炎检测。首先,将包含奶牛关键部位的局部图片水平拼接成信息完整的整体图片,结合RandAugment数据增强方法扩增训练样本;其次,采用ResNet34残差网络作为实验的基础网络,并根据热红外图像特性对模型进行如下改进:(1)精简网络内部冗余层使得模型更轻量化;(2)中间层添加辅助分类器弥补由于模型精简带来的特征损失;(3)将改进的多融合池化层代替原有单一池化层,使得特征提取内容更丰富。随机选取3298张热红外图像(66头奶牛)作为实验对象,并设置多组对比实验,结果表明:与传统ResNet34相比改进后ResNet34模型分类准确率提高3.4%,基于改进ResNet34并融合迁移学习和数据增强的模型验证准确率达到90.3%,测试准确率为88.4%,分类时间仅需3.39×10^(-3)s。为了保证实验数据集的样本独立性,进一步将奶牛个体数量按照3∶1∶1划分为训练集、验证集和测试集,测得模型测试准确率达到80.3%,证明所提出模型具有很好的鲁棒性。根据测试结果,计算出模型查准率为91.2%、查全率为91.6%、F1分数为91.4%,与前人所做实验相比准确率提高了5.1%,特异度提升5.3%。该研究方法可以为初期奶牛乳腺疾病筛选和医学诊断提供辅助和参考。

基于CiteSpace对中草药防治奶牛乳房炎的研究趋势及热点的可视化分析

试验旨在探究中草药防治奶牛乳房炎的研究热点及趋势。试验运用CiteSpace软件对中草药防治奶牛乳房炎的相关文献从年发文量、文献来源及关键词进行可视化分析。结果显示,中草药防治奶牛乳房炎文献主要来源于黑龙江畜牧兽医、中国奶牛、动物医学进展等期刊,研究热点涉及抑菌试验、致病菌、药敏试验、耐药性分析、分离鉴定等领域,研究方法包括综述、系统评价、试验研究、临床治疗及安全性评价等。研究表明,中草药防治奶牛乳房炎的研究主要集中在畜牧科学、兽医科学及饲料与添加剂等领域,其中致病菌的分离鉴定与耐药性评价始终是研究的热点。

奶牛微生物培养物BLS的研究

用鉴别培养基从土壤、新疆传统奶酪和马奶酒中分离出地衣芽孢杆菌、米酒乳杆菌和酿酒酵母。经鉴定和安全试验 ,上述三株菌混合后的联合培养物 ,接种生料发酵制成微生态制剂BLS ,饲喂泌乳牛。试验组平均每日奶产量显著高于对照组。

国外奶牛乳房炎研究进展——基于Web of Science核心数据库

随着人类社会工业化现代化进程的加快和经济的高速发展,人类对牛奶的需求量逐渐加大,高产奶牛的选育和替代人工的挤奶设备应用开始进入发展的快车道,与此同时也带来了奶牛乳房炎的“爆发式”增长,成为了当代奶牛养殖业的三大最主要疾病之一。国内外的专家学者也在不断地寻求有效方法,以最大限度地减少奶牛乳房炎的发生。国外在乳房炎方面的研究走在了国内的前面,他们的先进技术理念、手段和方法非常值得国内的借鉴。本文基于web of science核心数据库,用CiteSpace软件对近3年乳房炎相关的文献进行检索,并将其中经典文献进行总结归纳,以展示国外最新的乳房炎研究进展。

VITEK2.Compact对奶牛乳源金黄色葡萄球菌的条件摸索及MRSA检测结果准确性的评价

为探究金黄色葡萄球菌全自动微生物检测平台(VITEK2 Compact)检测条件,及该方法在金黄色葡萄球菌耐药性筛查的准确性及其兽医临床应用价值,以金黄色葡萄球菌ATCC 29213为质控菌株,选取ASTGP79药敏板,参考美国临床实验室标准研究所(CLSI/NCCLS)的《抗生素敏感试验操作标准》(S100-17和M31-A2)中兽医临床抗生素敏感性试验选药原则,通过摸索不同稀释浓度菌液,对照药敏标准,摸索全自动微生物生长分析仪的高通量筛选平台的适用检测条件。将该耐药性筛查方法应用于在奶牛乳汁中分离获得的39株金黄色葡萄球菌,与经典K-B法的耐药性进行比较,以此验证VITEK2 Compact检测金黄色葡萄球菌药敏结果准确性。结果显示,综合考虑检测结果、透光度、重复性等因素,确定了质控菌株的菌液比例为1×10^(8)CFU/mL,且证实了VITEK2 Compact药敏检测方法的结果符合临床折点的判断标准;对39株乳源金黄色葡萄球菌的耐药性检测与K-B法比较,标准符合率(CA):β-内酰胺类为92.30%、氨基糖苷类为93.32%、四环素类为92.30%、磺胺类为94.87%、喹诺酮类为92.30%、大环内酯类为90.59%、林可酰胺类为92.30%、氯霉素类为94.87%;两种检测方法对耐甲氧西林金黄色葡萄球菌(meticillin-resistant Staphylociccus aureus,MRSA)的检出阳性率无统计学差异(P>0.05)。表明了本研究探讨的金黄色葡萄球菌全自动微生物检测平台的药敏检测方法的准确性与可靠性,为该方法应用于金黄色葡萄球菌引起的奶牛乳房炎防治提供了理论依据。

乳房炎奶牛乳汁体细胞数与乳汁HP、SAA和AGP水平关系的研究

对8头正常奶牛和22头自发性乳房炎奶牛的混合乳汁中血清淀粉样蛋白(SAA)、结合珠蛋白(HP)、α1-酸性糖蛋白(AGP)水平和乳汁体细胞数(SCC)的关系进行了研究.结果表明:乳房炎奶牛与正常奶牛乳汁中HP水平差异显著(P<0.05),与乳汁SCC存在极显著正相关关系(P=0.000,r=0.406);乳房炎奶牛与正常奶牛乳汁中SAA和AGP的差异不明显(P>0.05),而且与乳汁SCC的相关性也均不显著,说明乳汁HP水平能反映奶牛乳腺感染的严重程度.

Isolation and Identification of Pathogenic Bacteria Causing Bovine Subclinical Mastitis in Eastern Hebei Province

In this study, the milk samples of 1 021 cows in eight dairy farms in Eastern Hebei Province were collected and detected with LMT reagent and somatic cell count for subclinical mastitis. Pathogenic bacteria in subclinical mastitis positive milk samples were isolated and identified.The results showed that 60.63%（619/1 021） of the sampled cows were diagnosed with subclinical mastitis, and mixed infections accounted for 88.21%（546/619） of the cases. In addition, 82 strains of 14 species were isolated from the subclinical mastitis positive milk samples, including 36 strains of Staphylococcus（43.90%）, 33 strains of Streptococcus（40.24%）, 8 strains of Enterobacteriaceae（9.76%） and 5 strains of Corynebacterium（6.10%）, respectively. The results proved that Staphylococcus aureus and Streptococcus agalactiae are the main pathogenic bacteria causing bovine subclinical mastitis in Eastern Hebei Province.