Invasive potential of biofilm-forming Staphylococci bovine subclinical mastitis isolates

Staphylococcus (S.) aureus is a common infectious agent of bovine chronic mastitis, a disease that is difficult to eradicate. The abilities of Staphylococci to be internalized and form a biofilm can contribute to host immunological defence evasion that subsequently impairs antimicrobial therapy. The invasive capability of six S. aureus field isolates with different biofilm-forming profiles was compared in vitro using a bovine mammary epithelial cell line. This was further confirmed in primary cell cultures using fluorescent rRNA probes against S. aureus. The results suggest that S. aureus invasion levels are not related to biofilm formation.     

An evaluation of a hand-held electrical resistance meter for the diagnosis of bovine subclinical mastitis in late lactation under Australian conditions

OBJECTIVE: To assess the ability of a hand-held device to differentiate between infected and noninfected bovine mammary glands according to the electrical resistance of milk, under Australian conditions. DESIGN: A cross-sectional study. PROCEDURE: Milk samples were collected from 236 quarters of 60 cows selected from a commercial dairy herd with a high prevalence of mastitis. The true infection status of these quarters was determined using bacteriology. Various methods were used in an attempt to relate the electrical resistance of milk from each quarter to the presence or absence of infection in that quarter. RESULTS: Although the electrical resistance of milk from infected quarters was generally lower than that of noninfected quarters, the overlap of readings between the two populations limited the ability of this device to indicate accurately whether a quarter was infected. The use of methods comparing the readings from each of the four quarters of a single cow did not allow the reliable detection of infected cows. CONCLUSION: Although this device may have some practical advantages in comparison with some other methods of diagnosing subclinical mastitis, the predictive value of the method was generally poor.     

冀东地区奶牛隐性乳房炎病原菌的分离鉴定

通过LMT法、乳汁pH检验法和体细胞直接计数等方法相结合,对冀东地区3个大型奶牛场、8个奶牛养殖户选取的1 021头奶牛进行隐性乳房炎流行病学调查与病菌分离鉴定。结果表明,隐性乳房炎发病率为60.63%(619/1021),乳房炎阳性乳样品中细菌分离率达88.21%(546/619)。从543头隐性乳房炎患牛的阳性乳区乳样中分得细菌共4类14种菌82株,其中葡萄球菌36株,占43.90%;链球菌33株,占40.24%;肠杆菌类8株,占9.76%;棒状杆菌5株,占6.10%。冀东地区奶牛隐性乳房炎主要病原菌为金黄色葡萄球菌和无乳链球菌。     

川渝地区奶牛隐性乳房炎调查和主要病原菌分离鉴定及药敏试验

用LMT法对川渝地区134头泌乳奶牛的491个乳区进行隐性乳房炎发病情况调查,并对阳性乳样进行主要病原菌的分离鉴定及药敏试验。结果显示,被检奶牛患隐性乳房炎头阳性率为70.90%,乳区阳性率为35.64%,175份阳性乳样的细菌检出率为90.86%,病原菌单独感染率为10.69%,混合感染率为89.31%;共分离到金黄色葡萄球菌、链球菌和大肠杆菌3种主要病原菌134株,其中金黄色葡萄球菌93株,占分离菌株的69.40%,链球菌34株,占25.37%,大肠杆菌7株,占5.23%;主要病原菌对呋喃妥因、新生霉素、头孢菌素类和喹诺酮类药物敏感性较好,对大多数抗菌药物产生不同程度的耐药性,对青霉素和链霉素则完全不敏感,耐药程度接近100%。     

头孢噻呋治疗隐性乳房炎对乳汁中主要致病菌区系的影响

采用细菌分离鉴定和高通量测序技术,比较分析了头孢噻呋治疗隐性乳房炎对乳汁中主要致病菌区系的影响。结果:经5～10 d的乳头灌注头孢噻呋治疗,有效率为68. 75%(11/16),治愈率仅为18. 75%(3/16)。无乳链球菌、酵母菌和金黄色葡萄球菌是治疗前乳汁中的主要致病菌,分别占分离菌株总数的33. 33%(6/18)、27. 78 (5/18)%和16. 67%(3/18);治疗后乳汁中主要致病菌是酵母菌,比例为58. 33%(7/12),其次是大肠杆菌(16. 67%)和其他链球菌(16. 67%)。治疗前后乳汁中主要真菌组成差异不显著,且假丝酵母属(Candida,63. 01%)、Meyerozyma属(25. 15%)和德巴利酵母属(Debaryomyces,5. 73%)是主要的3种真菌属。因此,在该奶牛场使用头孢噻呋治疗慢性隐性乳房炎效果并不理想,原因可能是抗菌治疗后真菌特别是酵母菌的大量繁殖。     

PCR-based detection of genes encoding virulence determinants in Staphylococcus aureus from bovine subclinical mastitis cases

The present study was carried out to genotypically characterize Staphylococcus aureus (S. aureus) isolated from bovine mastitis cases. A total of 37 strains of S. aureus were isolated during processing of 552 milk samples from 140 cows. The S. aureus strains were characterized phenotypically, and were further characterized genotypically by polymerase chain reaction using oligonucleotide primers that amplified genes encoding coagulase (coa), clumping factor (clfA), thermonuclease (nuc), enterotoxin A (entA), and the gene segments encoding the immunoglobulin G binding region and the X region of protein A gene spa. All of the isolates yielded an amplicon with a size of approximately 1,042 bp of the clfA gene. The amplification of the polymorphic spa gene segment encoding the immunoglobulin G binding region was observed in 34 isolates and X-region binding was detected in 26 isolates. Amplification of the coa gene yielded three different products in 20, 10, and 7 isolates. The amplification of the thermonuclease gene, nuc, was observed in 36 out of 37 isolates. All of the samples were negative for the entA gene. The phenotypic and genotypic findings of the present strategies might provide an understanding of the distribution of the prevalent S. aureus clones among bovine mastitis isolates, and might aid in the development of steps to control S. aureus infections in dairy herds.     

Genetic associations between somatic cell score and pathogen‐specific subclinical mastitis in Norwegian Red cows

The aim of this study was to estimate heritabilities of and genetic correlations between pathogen‐specific subclinical mastitis (SCM) traits and lactation mean somatic cell score (LSCS) in Norwegian Red cattle. Based on data from 130 733 first‐lactation cows four binary pathogen‐specific SCM traits, Staphylococcus aureus, Streptococcus dysgalactiae, Streptococcus uberis and coagulase‐negative staphylococci SCM, were analysed together with unspecific SCM and LSCS using a multivariate sire model with threshold models for binary traits and a linear model for LSCS. Posterior means (SD) of heritabilities were 0.17 (0.01) for LSCS, 0.11 (0.01) for liability to unspecific SCM and ranged from 0.04 (Staph. aureus) to 0.14 (Strep. dysgalactiae) for liability to pathogen‐specific SCM. Genetic correlations were positive and moderate to high, ranging from 0.37 to 0.98. All genetic correlations except the one between LSCS and unspecific SCM were lower than 1, indicating that SCM caused by different pathogens can be considered as partly different traits.     

隐性乳房炎患牛乳汁pH值变化与体细胞数关系的研究

采集奶牛乳样604份, 测定乳汁pH值与体细胞的含量, 确定奶牛隐性乳房炎与牛乳pH值的变化关系。结果表明, 牛乳pH值的变化与体细胞的含量呈正相关。正常乳汁的pH值在6 4~6 6之间; 体细胞在20万~50万/mL区间时, 奶牛乳汁pH值在6 6~6 8之间; 体细胞在50~150万/mL区间时, 奶牛乳汁pH值在6 8~7 0之间; 体细胞在150万~500万/mL区间时, 奶牛乳汁pH值在7 0~7 2之间; 体细胞大于500万/mL时, 奶牛乳汁pH值在7 2以上。     

Difference in virulence between Staphylococcus aureus isolates causing gangrenous mastitis versus subclinical mastitis in a dairy sheep flock

Staphylococcus aureus mastitis in dairy sheep ranges from subclinical mastitis to lethal gangrenous mastitis. Neither the S. aureus virulence factors nor the host-factors or the epidemiological events contributing to the different outcomes are known. In a field study in a dairy sheep farm over 21 months, 16 natural isolates of S. aureus were collected from six subclinical mastitis cases, one lethal gangrenous mastitis case, nasal carriage from eight ewes and one isolate from ambient air in the milking room. A genomic comparison of two strains, one responsible for subclinical mastitis and one for lethal gangrenous mastitis, was performed using multi-strain DNA microarrays. Multiple typing techniques (pulsed-field-gel-electrophoresis, multiple-locus variable-number, single-nucleotide polymorphisms, randomly amplified polymorphic DNA, spa typing and sas typing) were used to characterise the remaining isolates and to follow the persistence of the gangrenous isolate in ewes’ nares. Our results showed that the two strains were genetically closely related and they shared 3 615 identical predicted open reading frames. However, the gangrenous mastitis isolate carried variant versions of several genes (sdrD, clfA-B, sasA, sasB, sasD, sasI and splE) and was missing fibrinogen binding protein B (fnbB) and a prophage. The typing results showed that this gangrenous strain emerged after the initial subclinical mastitis screening, but then persisted in the flock in the nares of four ewes. Although we cannot dismiss the role of host susceptibility in the clinical events in this flock, our data support the hypothesis that S. aureus populations had evolved in the sheep flock and that S. aureus genetic variations could have contributed to enhanced virulence.     

乳房淋巴结内注射胸腺五肽治疗奶牛隐性乳房炎的研究

将患隐性乳房炎的奶牛30头随机分为5组,3个试验组奶牛分别在乳房淋巴结内注入低、中、高剂量的胸腺五肽（TP-5）;阳性药物对照组奶牛在乳房内灌注＂乳炎＂（主要含林可霉素和新霉素）;空白对照组,不作任何处理。治疗前和治疗后1、2、3周采集奶样进行细菌学检查和体细胞计数（SCC）。停药后4d之内每天采样进行牛奶抗生素残留检测。结果表明,和空白对照组比较,经药物治疗后各组感染乳区有不同程度减少,牛奶SCC减少的幅度依次为阳性药物对照组〉TP-5高剂量组〉TP-5中剂量组〉TP-5低剂量组。＂乳炎＂治疗组在停药后3d之内可以在牛奶中检测到抗生素残留,而TP-5则没有。因此,TP-5对奶牛隐性乳房炎具有治疗作用,并且不会在牛奶中造成有害的残留。     

An outbreak of Streptococcus canis mastitis in a dairy herd in Israel

CASE HISTORY: An increase in the bulk somatic cell count (BSCC) of up to 1,000 × 10³ cells/ml occurred in a dairy herd in Israel at the end of 2001 and beginning of 2002.

CLINICAL FINDINGS: Bacteriological examination of milk from 69 cows revealed a high prevalence of Streptococcus group G bacteria, identified as S. canis, affecting 38% of cows and 20% of all quarters. Isolates were sensitive to cephalothin and moderately sensitive to penicillin G. Infected cows were separated from the herd, treated with intramammary antibiotics, milked last, and strict hygiene practices were introduced to the milking routine. The pathogen was cleared from the herd and BSCC decreased to 250-350 × 103 cells/ml after 6 months.

DIAGNOSIS: Streptococcus canis mastitis.

CLINICAL RELEVANCE: Streptococcus canis infection may cause subclinical mastitis and high bulk SCC in dairy herds and be resolved by treatment with intramammary antibiotics and the introduction of strict hygiene practices.     

Acute phase proteins as indicators of calf herd health

The potential for using acute phase proteins (APPs) in the assessment of herd health was studied by examining the levels of serum haptoglobin, serum amyloid A (SAA) and plasma fibrinogen in relation to clinical findings and leukocyte counts in calves. Two groups of calves from conventional dairy farms were studied. The animals were examined 10 times during the first six weeks after introduction into a new environment. Haptoglobin, SAA and fibrinogen were analysed and weight gain, disease symptoms and treatments were recorded. Analysis of antibodies against viral infections was performed. An acute phase reaction (APR) score was established at each sampling by combining the APP results and total leukocyte counts. The health status differed between the two groups, although no manipulation of health had been performed, except that the group with a higher incidence of disease had a concurrent experimental infection with lungworm as part of another study. In the group with a higher incidence of disease, the mean weight gain was significantly lower, and the number of sampling days with elevated serum concentrations of APPs, and the mean maximum concentrations of haptoglobin and fibrinogen were significantly higher compared to the healthier group. The APR score was significantly higher at days 4 and 8 of the study in the group with a higher incidence of disease. The results indicate that measurement of APPs could be a useful tool for evaluation of health in calf herds.     

Acute phase proteins as indicators of stress in Baladi goats from Jordan

This study aimed to evaluate haptoglobin (Hp), serum amyloid A (SAA), and C-reactive protein (CRP) in goats following exposure to heat and transportation stressors. In the heat stress (HS) experiment (4-week period), six goats were exposed to direct solar radiation vs. six goats exposed to shade regimen. In the transportation stress experiment, six goats were transported for 2?h in a vehicle vs. a control group (n?=?6). Blood samples were collected and serum concentrations of acute phase proteins (APPs) were measured. Rectal temperature, respiratory rate, heart rate, and body weight were recorded. All statistical analyses were performed using the proc General Linear Model (SPSS 19.0). Results indicated that Hp and SAA concentrations were significantly affected by HS (P?P?相似文献   

丹参酮乳房注入剂中有效成分含量测定

采用高效液相色谱测定丹参酮乳房注入剂有效成分隐丹参酮和丹参酮ⅡA的含量。将供试品用正己烷萃取后挥干,甲醇溶解后检测;采用反相高效液相色谱,ZORBAX SB-C18键合硅胶柱(4.6×250 mm,5μm);柱温30℃;流动相为乙腈(A)-0.026%磷酸水溶液(B),梯度洗脱(0~20 min:V(A)60→90,V(B)40→10;20~30 min:V(A)∶V(B)=90∶10);流速1.2 m L/min;检测波长270 nm。丹参酮乳房注入剂中隐丹参酮含量在500~1750μg/g(r=0.9999)范围内线性关系良好,平均加样回收率为101.15%,RSD为1.55%;丹参酮ⅡA的含量在1125~3937.5μg/g(r=0.9999)范围内线性关系良好,平均加样回收率为100.89%,RSD为1.64%。结果显示,该方法简单、准确、灵敏,精密度、重复性好,可作为丹参酮乳房注入剂中有效成分含量测定的参考方法。     

Acute phase proteins in naturally occurring respiratory disease of feedlot cattle

The aim of this study was to evaluate three acute phase proteins (APP) [haptoglobin (HPT), lipopolysaccharide binding protein (LBP) and transferrin (Tf)] in feedlot cattle with naturally occurring respiratory disease diagnosed by a calf health scoring chart (CHSC). Seventy-seven beef calves were observed for signs of Bovine Respiratory Disease (BRD) during the first 28 days after arrival at the feedlot. Fourteen cases and pen matched controls were selected based on the CHSC. BRD cases were defined as a score of ≥5, while controls were defined as a score ≤4. The mean CHSC score in cases was 6.9 which was significantly greater than the controls 2.8 (P < 0.01). Mean plasma LBP and HPT concentrations were significantly greater in cases than controls (P < 0.01). Our study results show that measurement of HPT and LBP could be useful in detecting respiratory disease in feedlot conditions. Transferrin concentrations between the two groups were not statistically different.     

Efficacy of antimicrobial therapy for bovine acute Klebsiella pneumoniae mastitis

The purpose of this retrospective study was to assess the efficacy of antimicrobial therapy for bovine acute Klebsiella pneumoniae mastitis. We evaluated data from cattle in Ehime, Japan, with naturally occurring acute mastitis due to K. pneumoniae (n=208) or Escherichia coli (n=201). Survival was significantly shorter in cattle with acute K. pneumoniae mastitis (median, 76 days) compared with the disease caused by E. coli (median 464 days). In 2004–2008, because both species were highly susceptible to cefazolin, cases of K. pneumoniae and E. coli mastitis were treated solely with cefazolin, yielding clinical cure rates of 52.8% for K. pneumoniae and 86.0% for E. coli. However, since 2009, the efficacy of treatment of K. pneumoniae mastitis with cefazolin alone has decreased. When cefazolin administered on the first disease day led to clinical improvement, treatment with cefazolin was continued. However, when cefazolin administered on the first disease day failed to yield clinical improvement, the antibiotic was switched to a fluoroquinolone on the second day, resulting in cure rates of 76.7% for K. pneumoniae and 80.0% for E. coli. These findings suggest that, when the first-line drug (e.g., cefazolin) is ineffective, promptly changing to a second-line drug (e.g., a fluoroquinolone) increases the cure rate for bovine K. pneumoniae mastitis.     

Acute phase proteins in cattle after exposure to complex stress

Stressors such as weaning, mixing and transportation have been shown to lead to increased blood concentrations of acute phase proteins (APP), including serum amyloid A (SAA) and haptoglobin, in calves. This study was therefore undertaken to assess whether SAA and haptoglobin levels in blood mirror stress in adult cattle. Six clinically healthy Holstein cows and two Holstein heifers were transported for four to six hours to a research facility, where each animal was housed in solitary tie stalls. Blood samples for evaluation of leukocyte counts and serum SAA and haptoglobin concentrations were obtained before (0-sample) and at 8, 24 and 48 hours after the start of transportation. Upon arrival the animals gave the impression of being anxious, and they appeared to have difficulty coping with isolation and with being tied on the slippery floors of the research stable. Serum concentrations of SAA and haptoglobin increased significantly in response to the stressors (P < 0.01 and 0.05 at 48 hours, respectively). Additionally, the animals had transient neutrophilia at 8 and 24 hours (P < 0.05). In conclusion, the results of the study suggest that SAA and haptoglobin may serve as markers of stress in adult cattle.     

我国部分地区奶牛乳房炎病例中金黄色葡萄球菌血清型及毒力调查

为了解我国奶牛乳房炎病例中金黄色葡萄球菌(S.aureus)流行血清型和不同血清型菌株的毒力情况,本研究采用玻板凝集和双重PCR方法分别对北京、山西、内蒙古、山东、浙江和新疆奶牛乳房炎乳样中分离的191株S.aureus进行血清分型并从各血清型中随机选取100株菌以小白鼠进行毒力测定.结果表明:这些地区牛源S.aureus中,336PS型为流行血清型,占60.2％(115/191),CP8型占19.4％(37/191),CP5型占11.5％(22/191),未分型菌株占8.9％(17/191).毒力测定显示:336PS、CP8和CP5型菌株中强毒力菌株分别占65.1％、62.9％及61.9％,未分型菌株中强毒力菌株为22.2％.该调查为我国奶牛乳房炎S.aureus疫苗菌株的筛选提供了依据.     

奶牛乳房炎金黄色葡萄球菌基因多态性分型试验

利用随机引物AP-7,建立引物随机多态性扩增(RAPD)体系对71株引起内蒙古和贵州地区奶牛乳房炎的金黄色葡萄球菌分离株进行基因分型研究。结果表明,71株金黄色葡萄球菌均得到清晰的RAPD指纹图谱,扩增产物为2～9条带,产物大小为240～4 500bp。菌株共分为6个基因型,其中Ⅰ型17株(占23.9%)、Ⅱ型3株(占4.2%)、Ⅲ型33株(占46.5%)、Ⅳ型15株(占21.1%)、Ⅴ型2株(占2.8%)、Ⅵ型2株(占2.8%)。Ⅰ型为内蒙古地区的流行优势菌群,Ⅲ型为贵州地区的流行优势菌群。两地区各基因型菌株比例有明显差异,这可能与奶牛养殖业水平和环境差异有关。     

The role of acute phase proteins in diagnosis and management of steroid-responsive meningitis arteritis in dogs

Acute phase proteins (APPs) have become an important tool in the diagnosis, management and prognosis of inflammatory diseases in humans and are developing a similar utility in domestic species. Steroid-responsive meningitis arteritis (SRMA) is a well-recognised inflammatory disease of the dog, the diagnosis of which remains unsatisfactory based on clinical criteria and non-specific laboratory investigations. In this prospective pilot study the authors examined the acute phase response throughout the course of SRMA in serum and cerebrospinal fluid (CSF) by evaluating three key stages in disease management: presentation, treatment response and putative relapse.Serum APPs were found to be of value in supporting the diagnosis of SRMA and monitoring its treatment. C-reactive protein (CRP), serum amyloid-A (SAA), alpha-1-acid glycoprotein (AGP) and haptoglobin (Hp) all exhibited an increase above our laboratory reference range in nine patients at initial presentation. During treatment APPs decreased significantly compared to presentation except Hp which increased (Wilcoxon–Signed–Rank-test: CRP, SAA and AGP P < 0.05). Serum CRP and SAA were also found to be of clinical value in the identification of putative relapse (seven cases), particularly in the light of unperturbed CSF parameters where APP concentrations were elevated. CSF APPs were found to be less reliable markers in the management of this disease. The results indicate that SRMA causes a significant APP response in dogs, which although not disease specific, is of value in supporting the diagnosis of SRMA.