Acute Clinical Mastitis Research Articles

Identification of potential key circular RNAs associated with Escherichia coli-infected bovine mastitis using RNA-sequencing: preliminary study results.

Escherichia coli (E. coli) is commonly found in dairy farms and can invade mammary gland tissue, often causing acute clinical mastitis. Mammary infections with E. coli have shown a wide range of clinical signs, causing abnormal appearance of the milk, udder inflammation and systemic signs of illness. Circular RNA is a class of endogenous non-coding RNA that plays an important role in the occurrence and development of various inflammatory diseases. However, there is little information on the circRNA associated with bovine mastitis. In this study investigated the involvement of circRNAs in bovine mastitis through the construction of an E. coli-infected bovine mastitis model by injecting of E. coli into the mammary gland of dairy cows, using healthy gland mammary tissue as a control (M_C). High-throughput RNA-seq was performed on the E. coli-infected mammary gland tissue (M_E) and differentially expressed circRNAs between theM_C and M_E groups, followed by an analysis of their potential functions using bioinformatics methods. A total of 164 differentially expressed circRNAs were identified, including 92 downregulated circRNAs and 72 upregulated circRNAs. As shown by Gene Ontology enrichment analysis these DE circRNAs were mostly enriched in ras protein signal transduction, cytoplasmic vesicle parts, and enzyme binding, and Kyoto Encyclopedia of Genes and genome singal pathway enrichment analysis indicated significant associations with phagosome signal pathway. Additionally, the expression of bovine mastitis-related circRNAs, including novel_circRNA_0000128, novel_circRNA_0011103, novel_circRNA_0012656, novel_circRNA_0015099, novel_circRNA_005648, novel_circRNA_000074, and novel_circRNA_0011796 were verified via quantitative reverse-transcription polymerase chain reaction (RT-qPCR). These results provide a new direction for further investigation of the molecular mechanisms underlying bovine mastitis.

Effect of 2% meloxicam injection in Holstein dairy cows on acute clinical mastitis without systemic symptoms.

This study aimed the efficacy of meloxicam (MX) in treating acute clinical mastitis (ACM) without systemic symptoms in Holstein cows by studying improvement in udder pain, changes in prostaglandin E2(PGE2) and bradykinin (BK) levels in the milk, and milk yield (MY) after healing. Forty-two cows with ACM were randomly assigned to the MX treatment group (T group; n=21) and the control group (C group; n=21). At onset of illness (day 0), the T group received a 0.5 mg/kg subcutaneous (SC) injection of MX whereas the C group received 15 mL SC of saline solution as a placebo. Udder tenderness (UT) was measured, and milk samples were collected on days 0-3. There was little change in the MY of the T group before and after healing, whereas MY in the C group was significantly lower than after healing. UT on day 3 in the T group was significantly lower than that in the C group. PGE2 levels significantly decreased from day 0 to day 3 in both groups. A significant negative correlation between PGE2 and linear score was observed on day 1 in the T group, but not in the C group. In ACM without systemic symptoms, the administration MX may be useful for restoring MY and reducing udder pain after healing.

Outbreak of Acute Clinical Mastitis in Primigravidae Ewes in the Immediately Pre-Partum Period

An outbreak of acute mastitis was investigated in a dairy sheep farm in Central Greece with a prevalence of 8.2%. Two animals were examined with symptoms of hyperacute inflammation in one of the two mammary glands. Affected glands presented swollen to grossly enlarged, warm and painful. In both animals, haematological examination revealed mild anaemia, increased leucocyte counts with mature neutropaenia and left shift, and marked thrombocytosis. Bacteriological examinations of mammary gland secretion yielded heavy growth of Streptococcus dysgalactiae and Escherichia coli from each of the two animals, respectively. Serological examination revealed a high percentage of positive samples for small ruminant lentivirus infection in both animals. The somatic cell count in the bulk-tank milk was 0.65 × 106 cells/mL. The housing conditions were inadequate with high levels of ammonia, intense stocking and muddy bedding. This pathological condition in nulliparous ewes was attributed to the extensive oedema of mammary gland and teat, the relaxation of immunity during the peri-parturient period and the heavily contaminated environment. These findings are in general agreement with corresponding incidents in heifers. Following our instructions for improving housing conditions, no other similar incidents were recorded.

β-defensin-4 as an endogenous biomarker in cows with mastitis.

Defensins are peptides with antimicrobial and immunomodulatory effects. Their concentration could be altered during infections and thus provide information on the prognosis and course of the disease. The aim of the present study was to investigate the defensin concentration in cows with mastitis in order to find correlations between clinical expression and course of the disease and the defensin concentration in milk and blood. A total of 85 dairy cows were examined. Of these, 30 animals suffered from acute clinical mastitis, 25 animals were diagnosed with subclinical mastitis and 30 animals were considered a healthy comparison group. Beta-Defensin-4 (DEFB-4) was determined by a species-specific enzyme-linked immunosorbent assay (ELISA) (Bovine Defensin Beta 4 ELISA Kit, MyBioSource). The highest concentrations of DEFB-4 were detected in the animals with acute clinical mastitis. Values of 0 to 895 pg/mL (median: 115 pg/mL) were measured in milk and 40-1,016 pg/mL (median: 245 pg/mL) in serum. The concentrations of this group differed significantly from those of the animals with subclinical mastitis (p < 0.0001 serum; p = 0.015 milk). In this group, concentrations of 15-211 pg/mL (median: 46 pg/mL) were recorded in milk and 20-271 pg/mL (median: 85 pg/mL) in serum. Our results also show that in cases of acute mastitis after 12 days of treatment there is still an active inflammatory process in the tissue, because no significant reduction of somatic cells and defensin could be found after re-examination. Since the DEFB-4 concentrations of animals with clinical mastitis that had to be treated with antibiotics differed significantly from those of animals with subclinical mastitis that did not require antibiotic treatment, it can be assumed that bovine DEFB-4 is an important endogenous parameter for the defense against bacterial infections of the udder.

Prediction of Acute and Chronic Mastitis in Dairy Cows Based on Somatic Cell Score and Mid-Infrared Spectroscopy of Milk.

Simple SummaryMid-infrared (MIR) spectroscopy is the method of choice to determine milk components like fat, protein and urea. We examined the potential of MIR spectra analyses for the prediction of clinical mastitis events of dairy cows additionally, or alternatively, to somatic cell count, which is routinely used as an indicator for mastitis monitoring. Prediction models based on MIR spectra and a somatic cell count-derived score (SCS) were developed and compared. A model based on MIR spectra and SCS proved more accurate at predicting mastitis than models based on either indicator alone. Consequently, MIR spectra analyses add extra value in the prediction of clinical mastitis, making them potentially useful for dairy farm management and as an auxiliary trait for the genetic evaluation of udder health.Monitoring for mastitis on dairy farms is of particular importance, as it is one of the most prevalent bovine diseases. A commonly used indicator for mastitis monitoring is somatic cell count. A supplementary tool to predict mastitis risk may be mid-infrared (MIR) spectroscopy of milk. Because bovine health status can affect milk composition, this technique is already routinely used to determine standard milk components. The aim of the present study was to compare the performance of models to predict clinical mastitis based on MIR spectral data and/or somatic cell count score (SCS), and to explore differences of prediction accuracies for acute and chronic clinical mastitis diagnoses. Test-day data of the routine Austrian milk recording system and diagnosis data of its health monitoring, from 59,002 cows of the breeds Fleckvieh (dual purpose Simmental), Holstein Friesian and Brown Swiss, were used. Test-day records within 21 days before and 21 days after a mastitis diagnosis were defined as mastitis cases. Three different models (MIR, SCS, MIR + SCS) were compared, applying Partial Least Squares Discriminant Analysis. Results of external validation in the overall time window (−/+21 days) showed area under receiver operating characteristic curves (AUC) of 0.70 when based only on MIR, 0.72 when based only on SCS, and 0.76 when based on both. Considering as mastitis cases only the test-day records within 7 days after mastitis diagnosis, the corresponding areas under the curve were 0.77, 0.83 and 0.85. Hence, the model combining MIR spectral data and SCS was performing best. Mastitis probabilities derived from the prediction models are potentially valuable for routine mastitis monitoring for farmers, as well as for the genetic evaluation of the trait udder health.

Escherichia coli Mastitis in Dairy Cattle: Etiology, Diagnosis, and Treatment Challenges.

Bovine mastitis is an inflammation of the udder tissue parenchyma that causes pathological changes in the glandular tissue and abnormalities in milk leading to significant economic losses to the dairy industry across the world. Mammary pathogenic Escherichia (E.) coli (MPEC) is one of the main etiologic agents of acute clinical mastitis in dairy cattle. MPEC strains have virulence attributes to resist the host innate defenses and thrive in the mammary gland environment. The association between specific virulence factors of MPEC with the severity of mastitis in cattle is not fully understood. Furthermore, the indiscriminate use of antibiotics to treat mastitis has resulted in antimicrobial resistance to all major antibiotic classes in MPEC. A thorough understanding of MPEC’s pathogenesis and antimicrobial susceptibility pattern is required to develop better interventions to reduce mastitis incidence and prevalence in cattle and the environment. This review compiles important information on mastitis caused by MPEC (e.g., types of mastitis, host immune response, diagnosis, treatment, and control of the disease) as well as the current knowledge on MPEC virulence factors, antimicrobial resistance, and the dilemma of MPEC as a new pathotype. The information provided in this review is critical to identifying gaps in knowledge that will guide future studies to better design diagnostic, prevent, and develop therapeutic interventions for this significant dairy disease.

The Link Between 15-F2t-Isoprostane Activity and Acute Bovine Endothelial Inflammation Remains Elusive.

Modern dairy cattle suffer from increased incidence and severity of mastitis during major physiological transitions of the lactation cycle. Oxidative stress, a condition resulting from inadequate antioxidant defense against reactive oxygen and nitrogen species, is a major underlying component of mastitis pathophysiology. Isoprostanes (IsoP) are molecules derived from cellular lipid membranes upon non-enzymatic interaction with reactive species during inflammation, and are regarded as highly sensitive and specific biomarkers of oxidative stress. Changes in IsoP concentrations have been noted during major physiological transitions and diseases such as coliform mastitis in dairy cattle. However, the biological role of IsoP during oxidative stress in dairy cows has not been well-elucidated. Therefore, this study aimed to characterize the impacts of IsoP on oxidative stress outcomes in a bovine model of acute endothelial inflammation. Bovine aortic endothelial cells (BAEC; n = 4) were stimulated with 2,2'-azobis (2-amidinopropane) dihydrochloride (AAPH) or lipopolysaccharide (LPS) with or without 15-F2t-IsoP to determine how IsoP influence oxidative stress outcomes. Our endothelial inflammation model showed relatively decreased reactive metabolites and increased barrier integrity in cells treated with both the agonist and IsoP compared to agonist treatment alone. However, IsoP do not appear to affect oxidative stress outcomes during acute inflammation. Understanding the effect of IsoP on BAEC is an early step in elucidating how IsoP impact dairy cows during times of oxidative stress in the context of acute clinical mastitis. Future studies should define the optimal dosing and treatment timing of IsoP to maximize their cytoprotective potential during acute inflammation.

Sensitivity and Resistance of the Microbiota of Reproductive Organs and Mammary Gland of Cows to Anti-microbial Agents in Cases of Inflammation

This paper contains the results of complex microbiological studies (culture method, real-time PCR) of biological material from cows affected by inflammatory diseases of reproductive organs and mammary gland. Milk microbiota with underlying subclinical mastitis was represented by pathogenic (S. aureus, enteropathogenic E. coli), opportunistic bacteria (Staphylococcus spp., E. coli, E. faecium, Streptococcus spp., S. agalactiae, P. aeruginosa) and yeast-like fungi of Candida spp. in association. Combined infectious and inflammatory diseases of genital tract and mammary gland in cows resulted in the same types of microorganisms in microflora composition, which confirms information obtained by other researchers on associated microbiota during inflammatory processes. S. aureus, P. aeruginosa, E. coli, E. faecium, E. faecalis, S. epidermidis, S. saprophyticus, Bacillus spp., and yeast-like fungi of Candida spp. were obtained from milk samples from cows with acute clinical mastitis and in vaginal washes from cows with acute postpartum endometritis. In the course of defining the sensitivity to antibiotics and identifying genes of antimicrobial resistance, multidrug resistance was established in most part of obtained strains, which should be taken into account when planning treatment measures.

Antimicrobial susceptibility of nine udder pathogens recovered from bovine clinical mastitis milk in Europe 2015–2016: VetPath results

VetPath is an ongoing pan-European antimicrobial susceptibility monitoring programme collecting pathogens from diseased cattle, pigs and poultry not recently treated with antibiotics. Non-duplicate isolates (n = 1244) were obtained from cows with acute clinical mastitis in eight countries during 2015–2016 for centrally antimicrobial susceptibility testing according CLSI standards.Among Escherichia coli (n = 225), resistance was high to ampicillin and tetracycline, moderate to kanamycin and low to amoxicillin/clavulanic acid and cefazolin. The MIC50/90 of danofloxacin, enrofloxacin and marbofloxacin were 0.03 and 0.06 μg/mL. For Klebsiella spp. (n = 70), similar results were noted, except for ampicillin and kanamycin. We detected 3.7 % (11/295) Enterobacteriaceae isolates carrying an ESBL/AmpC gene. Staphylococcus aureus (n = 247) and coagulase-negative staphylococci (CoNS; n = 189) isolates were susceptible to most antimicrobials tested except to penicillin (25.1 and 29.1 % resistance). Two S. aureus and thirteen CoNS isolates harboured mecA gene. Streptococcus uberis isolates (n = 208) were susceptible to β-lactam antibiotics (87.1–94.7 % susceptibility), 23.9 % were resistant to erythromycin and 37.5 % to tetracycline. Resistance to pirlimycin was moderate. For Streptococcus dysgalactiae (n = 132) the latter figures were 10.6 and 43.2 %; pirlimycin resistance was low. MIC values for Streptococcus agalactiae, Trueperella pyogenes and Corynebacterium spp. were generally low.This current VetPath study shows that mastitis pathogens were susceptible to most antimicrobials with exceptions of staphylococci against penicillin and streptococci against erythromycin or tetracycline. For most antimicrobials, the percentage resistance and MIC50/90 values among the major pathogens were comparable to that of the preceeding VetPath surveys. This work highlights the high need to set additional clinical breakpoints for antimicrobials frequently used to treat mastitis.

Antimicrobial profile of multidrug-resistant Streptococcus spp. isolated from dairy cows with clinical mastitis.

Objective:The current investigation was designed to point out the prevalence of multidrug-resistant Streptococcus spp. causing acute clinical mastitis and their pattern of antibiotic resistance in dairy cows.Materials and methods:Milk was sampled from 128 dairy cows with 191 infected quarters during the period from August 2017 to December 2018. Bacterial species were isolated from the milk samples and identified based on colony morphology and biochemical tests. Multiplex PCR was done for confirmatory detection of the Streptococcus spp. isolates.Results:The chief isolation percentages, from the sampled milk, were Escherichia coli (26%), then Staphylococcus aureus (23%), and Streptococcus dysagalactiae (23%), then Streptococcus agalactiae (20.1%), and finally coagulase-negative Staphylococci (7.7%). In confirmed PCR streptococci isolates, the antibiotic resistance genes have been detected, including macrolides antibiotic resistance genes (ermB and mefA genes), lincosamides antibiotic resistance genes (linB gene), and tetracycline resistance genes (tetM and tetO genes). Age, parity number, cleaning of bedding materials, cleaning of milking facilities, and utensils and udder cleaning practice were significant risk factors for multidrug-resistant streptococcal mastitis in dairy cows.Conclusion:The results of this study explored the phenotypic and genotypic traits of Streptococcus spp. which constitute a usual cause of acute clinical mastitis in dairy cows. The ermB, mefA, tetM, and tetO antibiotic-resistant genes were identified in streptococci isolates from dairy cows’ milk with acute clinical mastitis, indicating a public health hazard. Thus, veterinary clinical breakpoints are needed to improve surveillance data, improve the hygiene regimen on the farms, and promote the wise use of antimicrobials.

Intramammary honey preparation for treatment of subclinical bovine mastitis: a preliminary study

Honey has been used in medicine against bacterial infections for thousands of years. The objective of this study was to evaluate the efficacy of intramammary honey administered during lactating period to eliminate Staphylococcus aureus and Pseudomonas aeruginosa infection. Three infected cows were investigated. Five milliliters of multifloral undiluted honey was administered aseptically in quarter by intramammary infusion during two consecutive days (day 1 and day 2) of experiment. Cows’ milk samples were collected for bacteriological analysis during interval of time: 0, 7, 14, 21, 52, and 60 days. The cure following administration of honey was two cows at 7 and 14 days; three at 21, 52, and 60 days for S. aureus infection and two cows at days 21, 52, and 60 for P. aeruginosa. In this preliminary study, the efficacy of honey has demonstrated as one of the new alternative therapeutic product against P. aeruginosa and S. aureus subclinical mastitis. Further studies may elucidate the susceptibility of pathogens microorganisms implicated in acute clinical and subclinical bovine mastitis, the dynamic of honey in mammary gland, and appropriate intramammary infusion.

Outbreak of mastitis in ewes caused by Streptococcus agalactiae

Many cases of acute clinical mastitis, in seven neighboring sheep flocks of Larissa Prefecture, were investigated. The causative agent was identified by biochemical and serological tests as Streptococcus agalactiae. Further investigation of the isolates by pulsed-field gel electrophoresis (PFGE) revealed identical profiles. The results indicate that S. agalactiae should be considered as a significant pathogen that induces mastitis in ewes; the disease is characterized by high morbidity, easy transmission between ewes and rapid reduction in milk production. The antibiotic therapy proved to be ineffective, even though antibiotics administered after the sensitivity test, however, the administration of an autovaccine was promising.

Monitoring of antimicrobial susceptibility of udder pathogens recovered from cases of clinical mastitis in dairy cows across Europe: VetPath results

VetPath is an ongoing pan-European antimicrobial susceptibility monitoring programme collecting pathogens from diseased cattle, pigs and poultry not recently treated with antibiotics. Non-duplicate milk samples were collected from cows with acute clinical mastitis in nine countries and 934 isolates were obtained during 2009–2012 for subsequent antimicrobial susceptibility testing in a central laboratory. CLSI broth microdilution methodology was used, and where available, MICs were interpreted using CLSI approved veterinary-specific (ceftiofur) otherwise human clinical breakpoints.Among Escherichia coli (n=207) and Klebsiella spp., (n=87), resistance was moderate to tetracycline and high to cephapirin (E. coli only) whereas resistance to other β-lactam antibiotics was very low (ceftiofur) to low (amoxicillin/clavulanic acid, cephalexin, cephalonium). The MIC90 of enrofloxacin and marbofloxacin was 0.03 and 0.06μg/mL respectively (E. coli) with 0.5% strains displaying higher MICs. Staphylococcus aureus (n=192) and coagulase-negative staphylococci (CNS; n=165) strains were susceptible to most antibiotics tested except to penicillin (25.0 and 29.1% resistance), respectively. Three S. aureus and seven CNS strains were oxacillin-resistant and harboured mecA. Streptococcus uberis strains (n=188) were susceptible to the β-lactam antibiotics although 35.6% were penicillin intermediately susceptible, and 20.2% were resistant to erythromycin, 36.7% to tetracycline. For Streptococcus dysgalactiae (n=95) the latter figures were 13.7 and 56.8%, respectively.For most antibiotics, the percentage resistance among E. coli, S. aureus and S. uberis was comparable to that of the VetPath 2002–2006 survey. This current, expanded VetPath study shows that mastitis pathogens were susceptible to most antibiotics with exceptions of staphylococci tested against penicillin and streptococci against erythromycin or tetracycline. This work highlights the high need to set additional clinical breakpoints for antibiotics frequently used to treat mastitis.

Immunosensing system for rapid multiplex detection of mastitis-causing pathogens in milk

Mastitis, an inflammation of the mammary gland and udder tissue, is the major endemic disease of dairy cattle. In addition to causing health problems to the animals, mastitis leads to the reduction of milk production and quality, representing a significant economic burden for farmers. To enable timely treatment of infected animals with pathogen-specific antibiotics, the development of automated analytical methods for rapid on-site identification and quantification of mastitis-causing pathogens in milk is particularly important.An immunosensing system for multiplex detection of the two most common mastitis-causing pathogens Staphylococcus aureus and Escherichia coli is proposed in the present study. This immunosensor combines Bead Injection Analysis (BIA), attachment of pathogens onto renewable micro-column, biorecognition of bound pathogens by specific antibodies, conjugated with different fluorescence markers and the measurement of fluorescence signals. The analysis takes 20min and exhibits detection limits of < 50 CFU mL−1 for E. coli and 100 CFU mL−1 for S. aureus in milk. The applicability of the immunosensor was demonstrated by analyzing milk samples from cows, who were suffering from acute clinical 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

ObjectivesTo study the molecular characteristics, antibiogram and prevalence of multi-drug resistant Staphylococcus aureus (S. aureus) (MDRSA) isolated from milk obtained from culled dairy cows and from cows with acute clinical mastitis. MethodsBacteria 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 morphology, Gram staining and biochemical characteristics. S. aureus isolates were then subjected to molecular characterization using PCR targeting 16S 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. ResultsS. 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. ConclusionsMRSA 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.

Bacteriological etiology and treatment of mastitis in Finnish dairy herds

BackgroundThe Finnish dairy herd recording system maintains production and health records of cows and herds. Veterinarians and farmers register veterinary treatments in the system. Milk samples for microbiological analysis are routinely taken from mastitic cows. The laboratory of the largest dairy company in Finland, Valio Ltd., analyzes most samples using real-time PCR. This study addressed pathogen-specific microbiological data and treatment and culling records, in combination with cow and herd characteristics, from the Finnish dairy herd recording system during 2010–2012.ResultsThe data derived from 240,067 quarter milk samples from 93,529 dairy cows with mastitis; 238,235 cows from the same herds served as the control group. No target pathogen DNA was detected in 12% of the samples. In 49% of the positive samples, only one target species and in 19%, two species with one dominant species were present. The most common species in the samples with a single species only were coagulase-negative staphylococci (CNS) (43%), followed by Staphylococcus aureus (21%), Streptococcus uberis (9%), Streptococcus dysgalactiae (8%), Corynebacterium bovis (7%), and Escherichia coli (5%). On average, 36% of the study cows and 6% of the control cows had recorded mastitis treatments during lactation. The corresponding proportions were 16 and 6% at drying-off. For more than 75% of the treatments during lactation, diagnosis was acute clinical mastitis. In the milk samples from cows with a recorded mastitis treatment during lactation, CNS and S. aureus were most common, followed by streptococci. Altogether, 48% of the cows were culled during the study. Mastitis was reported as the most common reason to cull; 49% of study cows and 18% of control cows were culled because of mastitis. Culling was most likely if S. aureus was detected in the milk sample submitted during the culling year.ConclusionsThe PCR test has proven to be an applicable method also for large-scale use in bacterial diagnostics. In the present study, microbiological diagnosis was unequivocal in the great majority of samples where a single species or two species with one dominating were detected. Coagulase-negative staphylococci and S. aureus were the most common species. S. aureus was also the most common pathogen among the culled cows, which emphasizes the importance of preventive measures.

Genotyping of Staphylococcus aureus in bovine mastitis and correlation to phenotypic characteristics

Reducing the prevalence of mastitis caused by Staphylococcus aureus (S. aureus) is essential to improve animal health and reduce economic losses for farmers. The clinical outcome of acute mastitis and risk of progression to persistent mastitis can, at least to some extent, be related to genetic variants of the strain causing the infection. In the present study we have used microarrays to investigate the presence of virulence genes in S. aureus isolates from dairy cows with acute clinical mastitis (n=70) and correlated the findings to other genotypic and phenotypic characteristics. Among the most commonly found virulence factors were genes encoding several hemolysin types, leukocidins D and lukM/lukF-P83, clumping factors A and B, fibrinogen binding protein and fibronectin-binding protein A. Some virulence factors e.g. fibronectin-binding protein B and Staphylococcus aureus surface protein G were less common. Genes coding for several staphylococcal enterotoxins and toxic shock syndrome toxin-1 (TSST-1) were commonly found, especially in one major pulsotype. No beta-lactamase genes were found in any common pulsotype, while present in some rare pulsotypes, indicated to be of human origin. Production of TSST-1, enterotoxins, hemolysins and beta-lactamase could all be positively correlated to presence of the corresponding genes. This study reveals a number of genotypic differences and similarities among common and rare pulsotypes of S. aureus from cases of mastitis in Sweden. The results could help the design of diagnostic tools to guide on-farm interventions according to the expected impact on udder health from a specific S. aureus genotype.

Antimicrobial susceptibility monitoring of mastitis pathogens isolated from acute cases of clinical mastitis in dairy cows across Europe: VetPath results

VetPath is an ongoing pan-European antimicrobial susceptibility monitoring programme collecting pathogens from diseased cattle, pigs and poultry not recently treated with antibiotics. Non-replicate milk samples were collected from cows with acute clinical mastitis in eight countries. Escherichia coli, Staphylococcus aureus and Streptococcus uberis were isolated by standardised methods. Antimicrobial susceptibility was determined in a central laboratory by CLSI broth microdilution methodology; results were interpreted using clinical breakpoints where available. Among E. coli (n=280), resistance to tetracycline (14.3%) and cefapirin (11.1%) were most common. Resistance to other β-lactam antibiotics was absent (ceftiofur) or very low (cefalexin, amoxicillin/clavulanic acid). The MIC90 of enrofloxacin and marbofloxacin was 0.03 and 0.06μg/mL, respectively, with 0.7% of strains displaying a deviating high MIC. Staphylococcus aureus (n=250) were susceptible to most antibiotics tested, although 36.0% were resistant to penicillin G. For other β-lactam antibiotics where a CLSI breakpoint was available, no resistance was detected. Tetracycline resistance was low (5.2%). Streptococcus uberis (n=282) were susceptible to all β-lactam antibiotics, although 29.8% were intermediately susceptible to penicillin G; 18.8% of strains were resistant to erythromycin and 28.7% to tetracycline. This European study shows that bacteria associated with acute clinical mastitis are susceptible to most antibiotics with the exception of penicillin G against S. aureus, and erythromycin and tetracycline against S. uberis. The results of this study should serve as a reference baseline. This work also highlights the urgent need to set additional clinical breakpoints for antibiotics frequently used to treat mastitis.

Efficacy of enrofloxacin for the treatment of acute clinical mastitis caused by Escherichia coli in dairy cows

Evidence for the efficacy of antimicrobial treatments in Escherichia coli mastitis is limited. The aim of this double-blinded field trial was to investigate the efficacy of enrofloxacin compared with placebo,...

Cytokine Response and Oxidative Stress Status in Dairy Cows with Acute Clinical Mastitis

Reactive oxygen species are natural end products of the intensive metabolism in cells located in organism high-yielding dairy cows. When the disturbance of homeostasis occurs, oxidative processes lead to oxidative stress causing inflammation of the mammary gland (mastitis) in high-yielding dairy cows. This study was performed to evaluate blood antioxidant profile and pro-inflammatory cytokines in dairy cows with acute clinical mastitis. For this purpose, venous blood samples were obtained from 50 randomly selected clinically healthy lactating cows (control group) and 50 dairy cows with acute clinical mastitis based on detailed clinical examination. In cows with acute clinical mastitis, there was a significant (p<0.05) increase in the clinical index score compared with control group. Biochemically, there were a significant (p<0.05) decrease in the total antioxidant capacity, activity of reduced glutathione and catalase as well as in the levels of zinc and iron. However, there was a significant (p<0.05) increase in the levels of malondialdehyde, nitric oxide, interleukin-6 and tumor necrosis factor-?, activity of superoxide dismutase and the oxidative stress index. In dairy cows with acute clinical mastitis, there was a negative correlation between malondialdehyde and superoxide dismutase (r=?0.653), malondialdehyde and interleukin-6 (r=?0.860), and glutathione reductase and interleukin-6 (r=?0.928). The results of the current study indicate that the body antioxidant defense system is compromised in dairy cows with acute clinical mastitis creating a state of oxidative stress. Likewise, alteration of antioxidant trace element levels and pro-inflammatory cytokines is considered the most reliable index of increased oxidative stress and tissue damage in dairy cows with acute clinical mastitis.