Intramammary Infections In Dairy Cows Research Articles

Efficacy of an internal teat seal associated with a dry cow intramammary antibiotic for prevention of intramammary infections in dairy cows during the dry and early lactation periods

ABSTRACT: The present study aimed to evaluate the use of an internal dry period teat seal containing bismuth subnitrate (Teatseal®, Zoetis®, Florham Park, Nova Jersey, USA) associated with a long-acting cloxacilin preparation (Orbenin® Extra dry cow, Zoetis®, Florham Park, Nova Jersey, USA), in preventing new infections during the dry-off and early postpartum period. A total of 150 Holstein cows (average production of 9,000 kg of milk per lactation), with four functional udder quarters without clinical mastitis was included in the study. All animals were dried-off 60 days before the expected calving date. Two teats positioned diagonal-contralaterally received only dry cow antibiotic, control group C (n=300) and the other two teats, treatment group T (n=300) received dry cow antibiotic and infusion with an internal teat seal. Data from SCC variable were transformed by log base-10 transformation. Duncan’s test was used accepting 5% as the level of statistical significance. The occurrence of intramammary infection (IMI) and chronicity rate, and frequency of microorganisms isolated at drying and immediately postpartum in teats of group C and group T were evaluated using a non-parametric Chi-square Test, accepting 10% as the statistical significance level. There was a decrease in the occurrence of new infections in the early postpartum in cows which the sealant was used (C=19.6%, T=11.4%). In the postpartum period, Gram-negative bacteria were isolated from 16 teats in C and seven in T. The greatest reduction was observed for Escherichia coli (8 vs 1) in group T. There was no effect using the internal sealant on the frequency of isolation of environmental Streptococus. The use of sealant reduced the prevalence of subclinical mastitis cows between drying-off and the early postpartum period (C=51% versus T=42%) and resulted in a lower somatic cell count (SCC) in the treatment group when compared with the control group (T=1,073x103, C=1,793x103). The use of the internal teat seal combined with dry cow antibiotic is effective in the prevention of IMI during the dry period and early lactation and results in the reduction of SCC in immediate postpartum period. The treatment is effective in reducing infection between dry-off and the immediate postpartum caused by major and minor pathogens. However, no effect on infections caused by contagious pathogens was observed.

Sensitivity and specificity of PCR analysis and bacteriological culture of milk samples for identification of intramammary infections in dairy cows using latent class analysis

Real-time PCR analysis of milk samples is a fast method to identify intramammary infections (IMI) in dairy cows, and has the potential to be used for routine analysis of test milking composite milk samples. However, the results of the PCR analysis can be difficult to interpret. The objective of this study was to compare the sensitivity (Se) and specificity (Sp) of PCR analysis of composite milk samples, and conventional bacteriological culturing (BC) of quarter milk samples, when used to identify cows with IMI. The comparisons were performed for IMI with four common udder pathogens; Staphylococcus aureus (S aureus), Streptococcus dysgalactiae (Str dysgalactiae), Str uberis and coagulase negative staphylococci (CoNS). The Se and Sp of real-time PCR (SePCR; SpPCR) and BC (SeBC; SpBC) was simultaneously estimated using latent class analysis (LCA), studying one pathogen at the time. Milk samples from 970 dairy cows from 25 herds were included. Aseptically collected quarter milk samples taken at the day before test milking (TM), at the day of TM, and at the day after TM, were analyzed using BC. Non-aseptically collected composite milk samples taken at the day of TM were analyzed using PCR. Moreover, the composite milk somatic cell count (SCC) was recorded and summarized by diagnostic test and bacterial finding. LCA was first performed using only test results from samples taken at the day of TM, but in a second analysis BC results from the three consecutive samplings, interpreted in parallel, were included. The SePCR was significantly higher than the SeBC for S aureus, Str dysgalactiae and CoNS in the first analysis, but only for CoNS in the second analysis. The SpPCR was significantly lower than the SpBC for Str dysgalactiae and CoNS. In conclusion, using PCR analysis of composite milk samples, as a diagnostic tool for identifying cows with IMI increased the Se for all the pathogens investigated (although not always significantly), while Sp in general remained on a similar level, compared to BC of quarter milk samples. The use of repeated quarter milk sampling improved the SeBC, making the results of PCR analysis and BC more similar. However, the SCC of cows with IMI according to BC was higher than for cows with IMI according to PCR, suggesting that some of the cows with IMI according to PCR did not have an active inflammation. Hence, extra caution is needed when decisions about treatment of IMI are based on PCR.

Factors associated with intramammary infection in dairy cows caused by coagulase-negative staphylococci, Staphylococcus aureus, Streptococcus uberis, Streptococcus dysgalactiae, Corynebacterium bovis, or Escherichia coli

The aim of this study was to determine risk factors for bovine intramammary infection (IMI) associated with the most common bacterial species in Finland. Large databases of the Finnish milk-recording system and results of microbiological analyses of mastitic milk samples from Valio Ltd. (Helsinki, Finland) were analyzed. The study group comprised 29,969 cows with IMI from 4,173 dairy herds. A cow with a quarter milk sample in which DNA of target species was detected in the PathoProof Mastitis PCR Assay (Thermo Fisher Scientific, Waltham, MA) was determined to have IMI. Only cows with IMI caused by the 6 most common pathogens or groups of pathogens, coagulase-negative staphylococci (CNS), Staphylococcus aureus, Streptococcus uberis, Streptococcus dysgalactiae, Corynebacterium bovis, and Escherichia coli, were included. The control group comprised 160,176 IMI-free cows from the same herds as the study group. A multilevel logistic regression model was used to study herd- and cow-specific risk factors for incidence of IMI. Pathogen-specific results confirmed those of earlier studies, specifically that increasing parity increases prevalence of IMI regardless of causative pathogen. Holsteins were more susceptible to IMI than Nordic Reds except when the causative pathogen was CNS. Occurrence of IMI caused by C. bovis was not related to milk yield, in contrast to IMI caused by all other pathogens investigated. Organic milk production was associated with IMI only when the causative pathogen of IMI was Staph. aureus; Staph. aureus IMI was more likely to occur in conventional than in organic production. Cows in older freestall barns with parlor milking had an increased probability of contracting an IMI compared with cows in tiestall barns or in new freestall barns with automatic milking. This was the case for all IMI, except those caused by CNS, the prevalence of which was not associated with the milking system, and IMI caused by Staph. aureus, which was most common in cows housed in tiestall barns. A better breeding index for milk somatic cell count was associated with decreased occurrence of IMI, indicating that breeding for improved udder health has been successful in reducing the incidence of IMI caused by the most common pathogens in Finland. In the Finnish dairy sector, the importance of other measures to control IMI will increase as the Holstein breed progressively takes the place of the Nordic Red breed. Attention should be paid to hygiene and cleanliness, especially in old freestall barns. Based on our results, the increasing prevalence of automatic milking is not a reason for special concern.

Susceptibility to cephalosporins of bacteria causing intramammary infections in dairy cows with a high somatic cell count in Germany

The objective of this cross-sectional study was to determine the minimal inhibitory concentrations of cephalosporins of the first (cefalonium and cefapirin) and fourth generation (cefquinome) against bacteria isolated from intramammary infections in dairy cows with elevated somatic cell counts in Germany. Additionally, possible regional differences of the minimal inhibitory concentrations within Germany should be evaluated. In total, 6936 quarter milk samples from cows with a somatic cell count >200,000cells/ml were taken in 43 herds.The concentrations of the first generation cephalosporins inhibiting at least 90% of the isolates of a pathogen (MIC90) were ≥64μg/ml against Gram-negative bacteria and enterococci whereas the respective MIC90 against the other Gram-positive bacteria were ≤4μg/ml. The MIC90 of cefquinome were ≥16μg/ml against Gram-negative bacteria, bacilli and enterococci, and ≤2μg/ml against the other Gram-positive bacteria.Only the minimal inhibitory concentrations against coagulase-negative staphylococci differed significantly between regions in parametric survival models with shared frailties for the herds. However, the minimal inhibitory concentrations of cefquinome against staphylococci were higher than the minimal inhibitory concentrations of the tested cephalosporins of the first generation. Therefore, cefquinome should not be the first choice to treat staphylococcal mastitis in dairy cows.

Influences of season, parity, lactation, udder area, milk yield, and clinical symptoms on intramammary infection in dairy cows

The aim of this study was to evaluate the influences of season, parity, lactation, udder area, milk yield, and clinical symptoms on bacterial intramammary infection (IMI) in dairy cows. A total of 2,106 mastitis pathogens in 12 species were isolated from 125 dairy farms distributed in 30 different cities in China, and the information about these factors was recorded at the same time. Mastitis pathogens were isolated from 63.43% of the milk samples, whereas Streptococcus agalactiae accounted for 38.61% of all pathogens, followed by Str. dysgalactiae (28.16%), Staphylococcus aureus (19.10%), Escherichia coli (6.90%), and other pathogens (7.23%). According to our investigation, IMI was more common in spring with the isolation rate of pathogens at 81.04%, and lowest in winter (52.34%). Cows were more likely to be infected by environmental pathogens (E. coli or Str. uberis) in summer, in rear quarters and in cows with higher daily milk yield or lower somatic cell count. In addition, Str. dysgalactiae exhibited a higher prevalence with increased parity. Different clinical symptoms of quarters with bacterial IMI were seen in this study, and mastitis pathogens were isolated from healthy quarters.

Short communication: Subtyping of Staphylococcus haemolyticus isolates from milk and corresponding teat apices to verify the potential teat-skin origin of intramammary infections in dairy cows

Coagulase-negative staphylococci (CNS) are a major cause of intramammary infections (IMI) in dairy cows and they colonize the teat skin. Staphylococcus haemolyticus, one of the more common CNS, has been identified as a highly versatile opportunistic species. The aim of the present study was to gain better insight into the adaptation of S. haemolyticus subtypes to the udder ecosystem with respect to IMI development. During a longitudinal observational study conducted over 13 mo on 6 Flemish dairy herds, S. haemolyticus isolates were recovered from milk and teat apices. A total of 44 S. haemolyticus isolates originating from milk (24 isolates) and teat apices (20 isolates) of 6 selected udder quarters were singled out and analyzed using a combined methodology of (GTG)5-PCR and amplified fragment length polymorphism (AFLP) fingerprinting to determine intraspecies differences. Combining both fingerprinting methods, 4 S. haemolyticus subtypes were obtained (I to IV). Subtypes I, II, and IV were recovered from both milk and teat apex samples and were found to be associated with persisting IMI. Subtype III, not apparently related to IMI, was isolated solely from teat apices and not from milk. In general, S. haemolyticus subtypes found in milk from infected quarters could be recovered from the corresponding teat apices, although the latter could be colonized with up to 3 different subtypes. Comparing subtypes from milk and teat apices indicates that the IMI-causing agent likely originates from the teat skin.

Genomic characteristics of Staphylococcus aureus strains associated with high within-herd prevalence of intramammary infections in dairy cows

Staphylococcus aureus is one of the most important causes of mastitis in dairy cattle. Based on previous research, Staph. aureus genotypes with different pathogenic and contagious properties can cause intramammary infection (IMI) and coexist in the same herd. Our study aimed to compare Staph. aureus strains from herds that differed in IMI prevalence using different molecular approaches such as ribosomal spacer (RS)-PCR, multilocus sequence typing (MLST), spa typing, ribotyping, pulsed-field gel electrophoresis (PFGE), and multiplex PCR. For this purpose, 31 dairy herds with Staph. aureus IMI were selected, and 16 of these were chosen for a comparison study: the 8 high-prevalence (HP) herds had Staph. aureus IMI prevalence >28% and the 8 low-prevalence (LP) herds had an IMI prevalence <4%. A total of 650 isolates of Staph. aureus from mammary quarters of all positive cows were genotyped with RS-PCR, a technique based on amplification of a portion of the intergenic spacer 16S-23S rRNA, and a subset of 54 strains was also analyzed by multiplex PCR, ribotyping, PFGE, MLST, and spa typing. The RS-PCR analysis revealed 12 different profiles. Staphylococcus aureus strains isolated from 5 out of 8 HP herds showed a profile identical to the genotype B (GTB), described in previous studies as being strongly associated with high within-herd prevalence of Staph. aureus mastitis and the presence of the genes coding for enterotoxins sea, sed, and sej, a long x-region of spa gene, and 3 lukE fragments. Moreover, all strains isolated in the HP herds possessed genes coding for staphylococcal enterotoxins. In LP herds, a limited number of strains of 6 genotypes, different from those isolated in HP herds, were identified and GTB was not found. Within these genotypes, 4 strains were positive for the mecA gene. Preliminary results and comparison with other genotyping methods confirmed that genotyping by RS-PCR is an accurate, rapid, and inexpensive tool for future field studies on Staph. aureus mastitis strains and generates clinically relevant results.

Methicillin-resistant Staphylococcus aureus (MRSA) is associated with low within-herd prevalence of intra-mammary infections in dairy cows: Genotyping of isolates

Staphylococcus aureus is one of the most common mastitis-causing pathogens worldwide. In the last decade, livestock-associated methicillin-resistant S. aureus (LA-MRSA) infections have been described in several species, included the bovines. Hence, this paper investigates the diffusion of MRSA within Italian dairy herds; the strains were further characterized using a DNA microarray, which detects 330 different sequences, including the methicillin-resistance genes mecA and mecC and SCCmec typing. The analysis of overall patterns allows the assignment to Clonal Complexes (CC). Overall 163 S. aureus isolates, collected from quarter milk samples in 61 herds, were tested. MRSA strains were further processed using spa typing. Fifteen strains (9.2%), isolated in 9 herds (14.75%), carried mecA, but none harboured mecC. MRSA detection was significantly associated (P<0.011) with a within-herd prevalence of S. aureus intra-mammary infections (IMI) ≤5%. Ten MRSA strains were assigned to CC398, the remaining ones to CC97 (n=2), CC1 (n=2) or CC8 (n=1). In 3 herds, MRSA and MSSA co-existed: CC97-MRSA with CC398-MSSA, CC1-MRSA with CC8-MSSA and CC398-MRSA with CC126-MSSA. The results of spa typing showed an overall similar profile of the strains belonging to the same CC: t127-CC1, t1730-CC97, t899 in 8 out of 10 CC398. In the remaining 2 isolates a new spa type, t14644, was identified. The single CC8 was a t3092. The SCCmec cassettes were classified as type IV, type V or type IV/V composite. All or most strains harboured the genes encoding the β-lactamase operon and the tetracycline resistance. Streptogramin resistance gene was related to CC398. Enterotoxin and leukocidin genes were carried only by CC1, CC8 and CC97-MRSA. The persistence of MRSA clones characterized by broader host range, in epidemiologically unrelated areas and in dairy herds with low prevalence of S. aureus IMI, might enhance the risk for adaptation to human species.

Characterization of persistent and transient Escherichia coli isolates recovered from clinical mastitis episodes in dairy cows

Escherichia coli usually cause transient intramammary infections in dairy cows, but persistent intramammary infections have been observed. The objective of the study was to compare antimicrobial resistance and virulence genes found in persistent and transient E. coli isolated from clinical mastitis cases in a cohort of 91 Canadian dairy herds monitored over a 2-year period. Antimicrobial susceptibility was determined by broth microdilution and the presence of 27 virulence genes associated with extra-intestinal E. coli infections was determined by colony hybridization. Proportion of resistance in persistent E. coli ranged from 0.0% (enrofloxacin) to 27.8% (ampicillin and tetracycline). Proportion of resistance in transient E. coli ranged from 0.0% (enrofloxacin) to 16.8% (tetracycline). Odds of being classified as a persistent isolate increased by a factor of 1.6 (95% CI: 1.1, 2.4) for each aditional resistance observed (e.g. isolates resistant to four antimicrobial agents had 1.6 times higher odds of belonging to the persistent groups compared to isolates demonstrating resistance to three agents). Persistency was associated with higher odds of resistance to ampicillin (OR: 9.8, P<0.01) or cephalothin (OR: 7.6, P=0.02). Persistent isolates had 5.4 times higher odds (95% CI: 1.2, 24.0) of harboring virulence gene iroN. Similarly, persistent isolates had 8.6 times higher odds (95% CI: 2.8, 27.1) of possessing the virulence gene sitA. In conclusion, this study confirmed that persistency of intramammary E. coli isolates is associated with certain traits. Findings concerning iron-acquisition shed new light on the mechanisms of intramammary survival.

Longitudinal study of the effects of teat condition on the risk of new intramammary infections in dairy cows

Machine milking–induced alterations of teat tissue may impair local defense mechanisms and increase the risk of new intramammary infections. The objective of the current study was to assess the influence of short-term and long-term alterations of teat tissue and infectious status of the udder quarter on the risk of naturally occurring new intramammary infections, inflammatory responses, and mastitis. Short-term and long-term changes in teat condition of right udder quarters of 135 cows of a commercial dairy farm in Saxony-Anhalt, Germany, were recorded monthly for 10 mo using simple classification schemes. Quarter milk samples were collected from all examined quarters at each farm visit. Bacteriological culture results and somatic cell counts of quarter milk samples were used to determine new inflammatory responses (increase from ≤100,000 cells/mL to >100,000 cells/mL between 2 samples), new infections (detection of a pathogen from a quarter that was free of the same pathogen at the preceding sampling), and new mastitis (combination of new inflammatory response and new infection). Separate Poisson mixed models for new inflammatory responses, new infections, and new mastitis caused by specific pathogens or groups of pathogens (contagious, environmental, major, minor, or any) were used to estimate risk ratios and 95% confidence intervals. Data preparation and parameter estimation were performed using the open source statistical analysis software R. We observed no effect of any variable describing teat condition on the risk of new intramammary infections, inflammatory responses, or mastitis. Intramammary infections of the same udder quarter in the preceding month did not affect risk either.

A Streptococcus uberis transposon mutant screen reveals a negative role for LiaR homologue in biofilm formation

The environmental pathogen Streptococcus uberis causes intramammary infections in dairy cows. Because biofilm growth might contribute to Strep.uberis mastitis, we conducted a biological screen to identify genes potentially involved in the regulation of biofilm growth. By screening a transposon mutant library of Strep.uberis, we determined that the disruption of 13 genes (including hasA, coaC, clpP, miaA, nox and uidA) led to increased biofilm formation. One of the genes (SUB1382) encoded a homologue of the LiaR response regulator (RR) of the Bacillus subtilis two-component signalling system (TCS). Electrophoretic mobility shift assays revealed that DNA binding by LiaR was greatly enhanced by phosphorylation. Two-dimensional differential in-gel electrophoresis analyses of the liaR mutant and the parental Strep.uberis strain revealed five differentially produced proteins with at least a 1·5-fold change in relative abundance (P<0·05). The DNA-binding protein LiaR is a potential regulator of biofilm formation by Strep.uberis. Several molecular primary and downstream targets involved in biofilm formation by Strep.uberis were identified. This provides a solid foundation for further studies on the regulation of biofilm formation in this important pathogen.

Associations of udder-health indicators with cow factors and with intramammary infection in dairy cows

The objective of this study was to investigate if and how cow factors and intramammary infection (IMI) are associated with 4 different udder-health indicators in dairy cows as a first step in investigating whether the diagnostic performance of these indicators can be improved. The investigated indicators were somatic cell count (SCC), lactate dehydrogenase (LDH), N-acetyl-β-d-glucosaminidase (NAGase), and alkaline phosphatase (AP) measured in milk. In this cross-sectional study, approximately 1,000 cows from 25 dairy herds were sampled for bacteriology (quarter milk samples) during 3 consecutive days: the day before test milking, at the day of test milking, and at the day after test milking. The whole-udder test milking sample was analyzed for milk composition, SCC, LDH, NAGase, and AP. Cow data (parity, breed, milk yield, percentage of milk fat and protein, milk urea concentration, and days in milk from the sampled test milking) were collected from the Swedish milk-recording scheme. Of the sampled cows 485 were considered IMI negative and were used in multivariable mixed-effect linear regression models to investigate associations between cow factors and the udder-health indicators. A second modeling including all cows, both IMI negative and IMI positive (256 cows), was also performed. The results showed that all udder-health indicators were affected by cow factors but that different cow factors were associated with different indicators. Intramammary-infection status was significantly associated with all udder-health indicators except AP. Parity and milk urea concentration were the only cow factors associated with all indicators in all models. The significant cow factors explained 23% of the variation in SCC and >30% of the variation in LDH, NAGase, and AP in IMI-negative cows, showing that LDH, NAGase, and AP are more affected than SCC by cow factors. The IMI status explained 23% of the variation in SCC in the model with all cows but only 7% of the variation in LDH and 2% of the variation in NAGase, indicating that SCC has the best potential as a diagnostic tool in finding cows with IMI. However, further studies are needed to investigate whether the diagnostic properties of these udder-health indicators will improve with adjustment according to their associations with different cow factors when used as a diagnostic tool for finding cows with IMI.

Randomised controlled trials demonstrate efficacy of a novel internal teat sealant to prevent new intramammary infections in dairy cows and heifers

AIMS: The main aim was to investigate the effectiveness of a novel internal teat sealant (ITS) formulation containing bismuth subnitrate and the antiseptic chlorhexidine, in adult cows (Trial A) and nulliparous heifers (Trial B), to reduce the incidence of new intramammary infection (IMI) between treatment and calving and reduce the prevalence of IMI within 6 days following calving.METHODS: In Trial A dairy cows (n=326) without history of clinical mastitis and with a maximum composite milk somatic cell count <200,000 cells/mL during the current lactation were enrolled from three spring-calving mainly pasture-based herds immediately following the final milking. In Trial B all heifers (n=166) were enrolled from two herds approximately 3 weeks before the start of calving. Glands were randomly assigned to controls or treatment with ITS, and mammary secretion samples were taken from all glands prior to treatment, and on two occasions 0–4 and 3–6 days post-calving to determine the incidence of new IMI and the prevalence of IMI immediately post-calving. Additionally, cows in Trial A were monitored for cases of clinical mastitis (CM) in the non-lactating period and animals in both trials were monitored for cases of CM within 30 days of calving.RESULTS: The ITS reduced the estimated incidence of new IMI with both major and any pathogens compared to control glands in cows (6.2 vs. 14.2% and 14.7 vs. 32.8%, respectively) and in heifers (3.0 vs. 13.1% and 13.4 vs. 25.3%, respectively) (p<0.001). The ITS reduced the estimated prevalence of a major or any pathogen compared to untreated controls at both 0–4 and 3–6 days post-calving, in both cows and heifers. Additionally, the ITS reduced the incidence of CM in the non-lactating period in cows (−1.0%, p=0.01) and in the 30 days following calving in heifers (−2.0%, p=0.06), and tended to reduce the estimated incidence of CM in cows post-calving (−1.7%, p=0.10).CONCLUSIONS: The novel ITS reduced the incidence of new IMI, and the prevalence of IMI at calving in both adult cows and heifers, and reduced the incidence of CM in the non-lactating period of cows and in the first 30 days after calving in heifers.CLINICAL RELEVANCE: The results of these trials demonstrate the efficacy of a novel ITS for the control of mastitis in likely uninfected dairy cows over the non-lactating period and in heifers in late pregnancy.

Accuracy of the composite somatic cell count to detect intra-mammary infection in dairy cows using latent class analysis

The somatic cell count (SCC) is considered an important indicator of intra-mammary infection (IMI). The purpose of this study was to determine the accuracy of both SCC and culture to detect IMI and their conditional dependence by means of latent class methods. This study involved 175 dairy cows from 2 herds with different udder infection prevalences. Quarter and composite milk samples were collected for SCC and bacteriological culture. Latent-class models using Bayesian methods were used to estimate test sensitivity (Se) and specificity (Sp) and population prevalence. The models ran involved only major mastitis pathogens and composite SCC (CSCC). Five thresholds between 100,000 and 300,000cells/mL were evaluated and the receiver operating characteristics (ROC) curve analysis was performed. Fifty-five percent of the cows had CSCC ≥200,000cells/mL and 95.4% of the cows had at least one infected quarter either with minor or major pathogens. Considering a threshold of 150,000cells/mL, the estimated Se and Sp for the CSCC were, 0.80 (95% CrI 0.71–0.88) and 0.57 (95% CrI 0.44–0.71), respectively. The estimated culture Se and Sp were 0.83 (95% CrI 0.73–0.93) and 0.89 (95% CrI 0.74–0.98), respectively. There was no evidence of dependence between CSCC and culture. The area under curve for CSCC was 0.72. To the best of our knowledge, this is the first report of the CSCC accuracy to detect IMI for major pathogens considering the effect of culture misclassification. The estimates provided here could help to examine the performance of sampling schemes based on CSCC to manage udder health.

Associations of herd- and cow-level factors, cow lying behavior, and risk of elevated somatic cell count in free-stall housed lactating dairy cows

Recent evidence suggests that the risk of intramammary infection in dairy cows is related to lying patterns. The objectives of this study were to quantify the standing and lying behavior of dairy cows milked 3×/d, determine the cow- and herd-level factors associated with these behaviors, and relate these findings to the risk of an elevated somatic cell count (SCC). Five commercial free-stall dairy herds in Eastern Ontario, milking 3×/d, were enrolled in a longitudinal study. Forty Holstein-Friesian cows/herd were randomly selected as focal animals based on days in milk (<200d) and SCC (<100,000cells/mL). Farms were followed for 4, 5-week periods. Individual-cow SCC was recorded at the beginning of each period and end of the final period. Elevated SCC (eSCC) was used as an indicator of subclinical mastitis. A new incident eSCC was defined as an individual cow that started the period with a SCC <100,000cells/mL but whose next SCC exceeded 200,000cells/mL. Lying behavior was recorded 5d after each milk sampling using data loggers. For these 5d, individual milking times and feeding times were also recorded. On d1 of each recording period 2 trained observers scored focal cows for hygiene and lameness. Throughout the course of the study, cows averaged 11.2h/d of lying time, split into 8.6 lying bouts/d that were on average 84.6min in length. Later lactation cows had longer daily lying times that were split into fewer lying bouts of longer duration than cows earlier in lactation. Lame cows had longer daily lying times and lying bout durations than non-lame cows. Cows with greater milk yield had lower lying times than lower producing cows. Average post-milking standing time across the study herds was 103min. Manipulation of feed (feed delivery or push-up) by the stockperson, in the hour before milking or shortly thereafter, resulted in the longest post-milking standing times. Over the study period, 48 new eSCC were detected, resulting in a mean herd incidence rate of 0.91 eSCC/cow-year at risk for all study herds. A non-linear relationship between post-milking standing time and eSCC incidence was found; compared to those cows that lie down <90min after milking, cows that lie down for the first time >90min after milking had a lower risk of acquiring a new eSCC. The risk of experiencing an eSCC was also increased in multiparous cows, and in those cows with a higher SCC at the beginning of the study. These results indicate that management practices that promote post-milking standing time, such as the manipulation of feed delivery around milking times, should be encouraged to reduce the risk of cows experiencing new eSCC.

Efficacy Evaluation of Herbal Teat Dip “Mastidip Liquid” in Sub-Clinical Mastitis in Crossbred Cows

The present investigation was conducted to evaluate the efficacy of herbal teat dip Mastidip Liquid (supplied by m/s Ayurvet Ltd, Baddi, India) in subclinical mastitic cows on the basis of restoration of altered milk pH and somatic cells count (SCC). Twenty crossbred cows screened positive for subclinical mastitis were selected and divided into two equal groups. Herbal teat dip Mastidip Liquidwas applied post milking in 1:1 dilution (one part concentrated solution + one part clean water) twice daily immediately after milking for a period of 30 days in one group (II) and for 45 days in other group (III). One additional group of 10 intact cows were taken as normal control group (I). Milk samples were collected for SCC and pH examination before initiation of teat dipping and on 7,14,21,28, 35 and 49th day post application of teat dip. The pH and SCC was normalized and milk yield improved after treatment with Mastidip Liquid. In conclusion, post milking teat dipping with Mastidip Liquidis found to be effective teat sanitizer in prevention of new intramammary infection in dairy cows and reducing the incidence of subclinical mastitis.

Latent class analysis of the diagnostic characteristics of PCR and conventional bacteriological culture in diagnosing intramammary infections caused by Staphylococcus aureus in dairy cows at dry off

BackgroundStaphylococcus aureus is one of the most common causes of intramammary infections in dairy cows at dry off. Reliable identification is important for disease management on herd level and for antimicrobial treatment of infected animals. Our objective was to evaluate the test characteristics of PathoProof ™ Mastitis PCR Assay and bacteriological culture (BC) in diagnosing bovine intramammary infections caused by S. aureus at dry off at different PCR cycle threshold (Ct)-value cut-offs.MethodsSterile quarter samples and non-sterile composite samples from 140 animals in seven herds were collected in connection with the dairy herd improvement (DHI) milk recording. All quarter samples were analyzed using BC whereas all composite samples were analyzed with PathoProof ™ Mastitis PCR Assay. Latent class analysis was used to estimate test properties for PCR and BC in the absence of a perfect reference test. The population was divided into two geographically divided subpopulations and the Hui-Walter 2-test 2-populations model applied to estimate Se, Sp for the two tests, and prevalence for the two subpopulations.ResultsThe Se for PCR increased with increasing Ct-value cut-off, accompanied by a small decrease in Sp. For BC the Se decreased and Sp increased with increasing Ct-value cut-off. Most optimal test estimates for the real-time PCR assay were at a Ct-value cut-off of 37; 0.93 [95% posterior probability interval (PPI) 0.60-0.99] for Se and 0.95 [95% PPI 0.95-0.99] for Sp. At the same Ct-value cut-off, Se and Sp for BC were 0.83 [95% PPI 0.66-0.99] and 0.97 [95% PPI 0.91-0.99] respectively. Depending on the chosen PCR Ct-value cut-off, the prevalence in the subpopulations varied; the prevalence increased with increasing PCR Ct-value cut-offs.ConclusionNeither BC nor real-time PCR is a perfect test in detecting IMI in dairy cows at dry off. The changes in sensitivity and prevalence at different Ct-value cut-offs for both PCR and BC may indicate a change in the underlying disease definition. At low PCR Ct-value cut-offs the underlying disease definition may be a truly/heavily infected cow, whereas at higher PCR Ct-value cut-offs the disease definition may be a S. aureus positive cow.

Manageable risk factors associated with the lactational incidence, elimination, and prevalence of Staphylococcus aureus intramammary infections in dairy cows

Staphylococcus aureus intramammary infections (IMI) are a major cause of mastitis on farms worldwide. Incidence and elimination rates are the key determinants of prevalence of Staph. aureus, and risk factors associated with these rates must be identified, prioritized, and controlled to obtain long-term reduction in prevalence. The objectives of this study were to identify manageable risk factors associated with the lactational incidence, elimination, and prevalence of Staph. aureus IMI. A cohort of 90 Canadian dairy farms was recruited and followed in 2007 and 2008. Quarter milk samples were collected repeatedly from a selection of cows, and bacteriological culture was realized to assess incidence, elimination, and prevalence of Staph. aureus IMI. Practices used on farms were measured using direct observations and a validated questionnaire. A linear regression model was used to explore the relationship between herd IMI prevalence and incidence and elimination rates. Multilevel logistic regression models were used to compute measures of associations between practices used on farms and IMI incidence, elimination, and prevalence. The herd incidence rate was the most important predictor of herd IMI prevalence: a reduction of the incidence rate equivalent to its interquartile range (0.011 new IMI/quarter-month) was associated with a prevalence reduction of 2.2 percentage points; in comparison, an equivalent increase of the elimination rate by its interquartile range (0.36 eliminated IMI/quarter-month) resulted in a prevalence reduction of 0.4 percentage points. Postmilking teat disinfection and blanket dry-cow therapy were already implemented by most herds. Most of the practices associated with Staph. aureus IMI incidence were related to milking procedures. Among these, wearing gloves during milking showed desirable associations with IMI incidence, elimination, and prevalence. Similarly, adequate teat-end condition and use of premilking teat disinfection were associated with lower IMI incidence and prevalence. The initial herd prevalence of Staph. aureus IMI was positively associated with subsequent IMI incidence. This indicates that, in some situations, an initial reduction of the pool of infected quarters could be justified. Some housing practices were associated with IMI incidence, elimination, or prevalence. The effects of these latter practices, however, were often influenced by specific cow characteristics such as parity or days in milk. These results highlight the importance of good milking practices to prevent Staph. aureus IMI acquisition and, therefore, reduce their prevalence.

Host-response patterns of intramammary infections in dairy cows

Many different bacterial species have the ability to cause an infection of the bovine mammary gland and the host response to these infections is what we recognize as mastitis. In this review we evaluate the pathogen specific response to the three main bacterial species causing bovine mastitis: Escherichia coli, Streptococcus uberis and Staphylococcus aureus. In this paper we will review the bacterial growth patterns, host immune response and clinical response that results from the intramammary infections. Clear differences in bacterial growth pattern are shown between bacterial species. The dominant pattern in E. coli infections is a short duration high bacteria count infection, in S. aureus this is more commonly a persistent infection with relative low bacteria counts and in S. uberis a long duration high bacteria count infection is often observed. The host immune response differs significantly depending on the invading bacterial species. The underlying reasons for the differences and the resulting host response are described. Finally we discuss the clinical response pattern for each of the three bacterial species. The largest contrast is between E. coli and S. aureus where a larger proportion of E. coli infections cause potentially severe clinical symptoms, whereas the majority of S. aureus infections go clinically unnoticed. The relevance of fully understanding the bovine host response to intramammary infection is discussed, some major gaps in our knowledge are highlighted and directions for future research are indicated.

Short communication: Epidemiology and genotyping of Candida rugosa strains responsible for persistent intramammary infections in dairy cows

The present study was undertaken during an outbreak of clinical and subclinical mastitis in 14 dairy cows caused by Candida rugosa, in which high somatic cell counts were seen and cases did not respond to antibiotic treatment. Intramammary infection cured spontaneously in 10 cows, whereas 4 cows were culled as a result of persistent infections. Repeated sampling of these cows and biomolecular analysis of the isolates showed that the infections were caused by the same genotype, even over a period of 2 lactations. Random amplification of the genome of C. rugosa milk isolates gave 3 different DNA banding patterns (genotypes G1, G2, and G3). Viable cells of C. rugosa were also isolated from various environmental sources and were present in high concentrations in total mixed ration samples, which could be considered the primary source of diffusion of viable yeast cells in the environment, as demonstrated by genotyping. The proven capacity of these microorganisms to survive in the environment of the cow, such as the total mixed ration, bedding, water, and cow skin, and to cause persistent intramammary infections highlights the importance of mycotic spread in dairy herds.