K. Persson Waller

Microbial Aetiology of Acute Clinical Mastitis and Agent-Specific Risk Factors

A nation wide study on the microbial aetiology of cases of acute clinical mastitis in Swedish dairy cows was conducted with the aim to investigate changes in the microbial panorama compared to a previous study performed 1994-1995. Another aim was to investigate some agent-specific environmental and individual risk factors. Milk samples were collected from 987 udder quarter cases from 829 cows during six 2-month periods from May 2002 to April 2003, and data on risk factors and demography were collected at sampling by means of a questionnaire. In total, 1056 bacteriological diagnoses were made. The most frequently isolated bacterial species was Staphylococcus aureus constituting 21.3% of the diagnoses, followed by Escherichia coli (15.9%), Streptococcus dysgalactiae (15.6%), Streptococcus uberis (11.1%), coagulase-negative staphylococci (6.2%), Arcanobacterium pyogenes (6.1%) and Klebsiella spp. (4.2%). Samples with no growth or contamination constituted 10.6% and 4.5% of the diagnoses, respectively. A major shift in the panorama of udder pathogens was not observed compared to the survey in 1994-1995. Isolation of Klebsiella spp. was strongly associated with the use of sawdust as bedding material. On the other hand, using sawdust as bedding reduced the risk of isolating S. uberis relative to using straw or peat. The risk of isolating E. coli increased with increasing milk yield and was higher in loose housing systems than in tie stalls. Isolation of S. aureus was associated with tie stalls, and A. pyogenes with low yielding cows and teat lesions. S. dysgalactiae infections were also associated with teat lesions.

Bovine subclinical mastitis caused by different types of coagulase-negative staphylococci

Subclinical mastitis caused by intramammary infections (IMI) with coagulase-negative staphylococci (CNS) is common in dairy cows and may cause herd problems. Control of CNS mastitis is complicated by the fact that CNS contain a large number of different species. The aim of the study was to investigate the epidemiology of different CNS species in dairy herds with problems caused by subclinical CNS mastitis. In 11 herds, udder quarter samples were taken twice 1 mo apart, and CNS isolates were identified to the species level by biochemical methods. The ability of different CNS species to induce a persistent infection, and their associations with milk production, cow milk somatic cell count, lactation number, and month of lactation in cows with subclinical mastitis were studied. Persistent IMI were common in quarters infected with Staphylococcus chromogenes, Staphylococcus epidermidis, and Staphylococcus simulans. The results did not indicate differences between these CNS species in their association with daily milk production, cow milk somatic cell count, and month of lactation in cows with subclinical mastitis. In cows with subclinical mastitis, S. epidermidis IMI were mainly found in multiparous cows, whereas S. chromogenes IMI were mainly found in primiparous cows.

Cytokines in mammary lymph and milk during endotoxin-induced bovine mastitis.

Cytokine kinetics were examined in milk and in afferent and efferent lymph of the supramammary lymph node after intramammary infusion of endotoxin from Escherichia coli. Cows were sampled 0, 2 and 4h after infusion (p.i.). Neutrophils appeared in afferent lymph 2h p.i., and in efferent lymph and milk 4h p.i. The milk contained high concentrations of interleukin-8 (IL-8) at 2 and 4h p.i. IL-8 was also found in lymph, but at lower concentrations. The tumor necrosis factor-alpha (TNF-alpha) concentration tended to increase in afferent lymph at 2h p.i., and increased in milk at 4h p.i. The level of IL-1beta increased at 4h p.i. in milk, but was not detected in lymph. Interferon-gamma was not detected in any sample, at any time. The results indicate a primary role for IL-8 in the recruitment of neutrophils into the gland, and suggest that IL-1beta and TNF-alpha are not necessary for IL-8 production and release in response to endotoxin.

Adjuvant effect of ginseng extracts on the immune responses to immunisation against Staphylococcus aureus in dairy cattle.

A crude ginseng extract (GS) and the purified ginsenoside R(b1) (R(b1)) were evaluated for their adjuvant effects in dairy cattle at immunisation with ovalbumin (OVA) and/or a Staphylococcus aureus bacterin used for prevention of bovine mastitis. To evaluate a suitable dose of GS as an adjuvant, 36 lactating cows were randomly divided into six groups. The cows were inoculated twice intramuscularly with a 2-week interval, with saline solution, OVA in saline, or OVA in combination with 4, 16 or 64 mg GS, or Al(OH)(3). The level of specific antibodies to OVA in serum and milk whey was measured before immunisations and 1-5 weeks after the second immunisation. The antibody response in serum was significantly higher in animals immunised with OVA and GS than in animals immunised with OVA alone. A significant increase in milk antibody titres compared with OVA only was only found 2 weeks after the second immunisation in the group immunised with OVA and 4 mg GS. In the second part of the study, 18 heifers were randomly divided into three groups and were immunised twice intramuscularly with a two week interval, with the S. aureus bacterin (control), or with the bacterin in combination with 4 mg GS or 1mg R(b1). The specific antibody response to S. aureus and the lymphocyte proliferation after stimulation with PWM, concanavalin A (Con A) or a specific S. aureus antigen was evaluated in blood samples taken before and after immunisations as specified above. Addition of R(b1) resulted both in significantly higher antibody production and lymphocyte proliferation in response to PWM, Con A and S. aureus antigens than in the control group. Addition of GS induced a significantly higher lymphocyte proliferation in response to PWM and Con A than the control, but had no additional effect on the antibody production. In conclusion, both GS and R(b1) were safe adjuvants, and R(b1) had the strongest adjuvant effects, when used for immunisation against S. aureus in dairy cattle. Field trials are warranted to test the ability of GS and R(b1) to enhance the efficacy of mastitis vaccines in protection against intramammary infections.

Changes in some Blood Micronutrients, Leukocytes and Neutrophil Expression of Adhesion Molecules in Periparturient Dairy Cows

Dairy cows are highly susceptible to infectious diseases, like mastitis, during the period around calving. Although factors contributing to increased susceptibility to infection have not been fully elucidated, impaired neutrophil recruitment to the site of infection and changes in the concentrations of some micronutrients related with the function of the immune defence has been implicated. Most of the current information is based on studies outside the Nordic countries where the conditions for dairy cows are different. Therefore, the aim of the study was to evaluate changes in blood concentrations of the vitamins A and E, the minerals calcium (Ca), phosphorous (P), and magnesium (Mg), the electrolytes potassium (K) and sodium (Na) and the trace elements selenium (Se), copper (Cu) and zinc (Zn), as well as changes in total and differential white blood cell counts (WBC) and expression of the adhesion molecules CD62L and CD18 on blood neutrophils in Swedish dairy cows during the period around calving. Blood samples were taken from 10 cows one month before expected calving, at calving and one month after calving. The results were mainly in line with reports from other countries. The concentrations of vitamins A and E, and of Zn, Ca and P decreased significantly at calving, while Se, Cu, and Na increased. Leukocytosis was detected at calving, mainly explained by neutrophilia, but also by monocytosis. The numbers of lymphocytes tended to decrease at the same time. The mean fluorescent intensity (MFI) of CD62L and CD18 molecules on blood neutrophils remained constant over time. The proportion of CD62L+ neutrophils decreased significantly at calving. The animals were fed according to, or above, their requirements. Therefore, changes in blood levels of vitamins, minerals and trace elements were mainly in response to colostrum formation, changes in dry matter intake, and ruminal metabolism around calving. Decreased levels of vitamins A and E, and of Zn at calving might have negative implications for the functions of the immune defence. The lower proportion of CD62L+ neutrophils at calving may result in less migration of blood neutrophils into the tissues, and might contribute to the increased susceptibility to infections at this time.SammanfattningMjölkkor är mycket känsliga för infektionssjukdomar,som mastit, under perioden runt kalvningen. Det är inte helt klarlagt vilka faktorer som bidrar till den ökade infektionskänsligheten men försämrad rekrytering av neutrofiler till infektionsplatsen och förändringar i koncentrationen av vissa mikroelement har troligen betydelse. De flesta befintliga uppgifter baseras på studier gjorda utanför Norden där förhållandena för mjölkkor är annorlunda. Syftet med denna studie var därför att undersöka förändringar i blodkoncentrationen av vitaminerna A och E, mineralerna kalcium (Ca), fosfor (P) och magnesium (Mg), elektrolyterna kalium (K) och natrium (Na) samt spårämnena selen (Se), koppar (Cu) och zink (Zn) hos svenska mjölkkor under perioden runt kalvning. Dessutom undersöktes förändringar i antalet vita blodkroppar och uttrycket av adhesionsmolekylerna CD62L och CD18 på blodneutrofiler. Resultaten stämde huvudsakligen med tidigare rapporterfrån andra länder. Koncentrationen av vitaminerna A och E, Zn, Ca och P minskade signifikant vid kalvning medan Se, Cu och Na ökade. Leukocytos sågs vid kalvning huvudsakligen orsakat av neutrofili men också till viss del monocytos. Antalet lymfocyter tenderade att minska vid samma tidpunkt. Medelvärdet av fluorescensintensiteten för CD62Loch CD18-molekylerna på blodneutrofiler var konstant över tiden. Andelen CD62L+ neutrofiler minskade dock signifikant vid kalvningen. Djuren utfodrades i enlighet med eller över deras behov varför förändringarna i blodnivåerna av vitaminer, mineraler och spårämnen huvudsakligen var ett resultat avkolostrumproduktion, samt förändringar i torrsubstansintag och våmmetabolism runt kalvningen.Minskade nivåer av vitaminerna A och E och av Zn vid kalvning kan ha negativ effekt på immunförsvarets funktion. Den lägre andelen CD62L+ neutrofiler vid kalvning kan resultera i en försämrad mi-.

Metabolites and Immune Variables Associated with Somatic Cell Counts of Primiparous Dairy Cows

The main objective of this study was to investigate associations between serum concentrations of several blood variables related to metabolic and immunological status around calving, and udder health measured as milk somatic cell counts (SCC), Box-Cox transformed to bcSCC, at first test-milking in 287 primiparous cows in 20 Swedish dairy herds. Possible systematic effects of breed and age at calving on blood profiles were also investigated. Ordinary linear regression models, with robust standard errors and adjusting for clustering within herds, were used to investigate associations between blood variables and bcSCC. Hierarchical linear regression models, with herd as random factor, were used to investigate systematic effects on blood variables. The results showed that greater concentrations of beta-hydroxybutyrate (BHBA) and glucose before calving were associated with lesser bcSCC at first test-milking, whereas greater concentrations of nonesterified fatty acids (NEFA) before calving and greater delta NEFA (describing the difference in concentrations before and after calving) were associated with greater bcSCC at first test-milking. In addition, greater alpha-tocopherol concentrations in the period -5 to +5 d relative to calving were associated with lesser bcSCC at first test-milking, whereas greater concentrations of collectin of 43 kDa (CL-43) postpartum (1 to 21 d after calving) were associated with greater bcSCC. Postpartum concentrations of conglutinin and haptoglobin were also associated with bcSCC, but not independently of each other. Moreover, significant breed differences were observed for insulin, urea nitrogen, conglutinin, cholesterol, NEFA, and CL-43, the latter 3 as an interaction with period. Overall, cows of the Swedish Red breed had greater concentrations of insulin, cholesterol, urea nitrogen, and conglutinin, and lesser concentrations of NEFA and CL-43 than cows of the Swedish Holstein breed. Age at calving as main effect was significantly associated with BHBA, glucose, insulin, NEFA, urea nitrogen, and conglutinin. Heifers calving at >27 mo of age had greater BHBA and NEFA values, and lesser glucose, insulin, and urea nitrogen values compared with heifers calving at <27 mo. Heifers calving at an age <25 mo had greater conglutinin and urea nitrogen values, and lesser NEFA values compared with heifers calving at >25 mo. The results show that there are several associations among metabolites, immunological variables, and udder health of primiparous cows, but also that these variables vary between breeds and between cows of different age at first calving.

Comparison of a commercialized phenotyping system, antimicrobial susceptibility testing, and tuf gene sequence-based genotyping for species-level identification of coagulase-negative staphylococci isolated from cases of bovine mastitis.

In order to evaluate the usefulness of some phenotypic and genotypic methods for species identification of coagulase-negative staphylococci (CNS), isolates were obtained from bovine cases of clinical and sub-clinical mastitis from different geographical areas in Sweden. By using the Staph-Zym test, antimicrobial susceptibility testing, and sequencing of part of the CNS tuf gene and, when needed, part of the 16S rRNA gene we characterized 82 clinical isolates and 24 reference strains of 18 different species of staphylococci. The genotypic methods identified nine different species of CNS among the 82 milk isolates. A comparison with results obtained by tuf gene sequencing showed that Staph-Zym correctly identified CNS reference strains to species level more often than bovine milk CNS isolates (83% and 61%, respectively). In addition, tests supplementary to the Staph-Zym were frequently needed in both groups of isolates (50% of reference strains and 33% of milk isolates) to obtain an identification of the strain. It is notable that Staph-Zym judged two isolates as CNS, although they belonged to other species, could not give a species name in 11% of the bovine CNS isolates, and gave 28% of the isolates an incorrect species name. The present study indicates that the studied phenotypic methods are unreliable for identification of CNS from bovine intra-mammary infections.

Management practices associated with udder health of first-parity dairy cows in early lactation.

This study aimed at investigating associations between management routines including feeding, housing, and milking around calving, and udder health of first-parity cows in early lactation in Swedish large, high producing, low bulk-milk somatic cell count (SCC) dairy herds housed in free stalls. Seventy-two dairy herds participated and data concerning 1189 first-parity cows calving during the study period (October 2005-January 2006) was collected. Multivariable regression analysis were performed with three different outcomes; within-herd number of first-parity cows veterinary treated for clinical mastitis at days -10 to 60 after calving, within-herd number of first-parity cows with a SCC> or =200,000cells/mL at first test-day, and SCC of first-parity cows at first test-day. Cow factors significantly associated with good udder health of first-parity cows (few cases of clinical mastitis and or low SCC) were being of the Swedish Red breed, having a high milk yield at first test-day, and a milk-urea > or =5mmol/L at first test-day. Herd factors significantly associated with good udder health were having mattresses as flooring in the cubicles in the lactating cow housing, and to house the first-parity cows in tie stalls 1 month before calving. Cow factors significantly associated with poor udder health of first-parity cows were having a milk-urea <4mmol/L at first test-day. Herd factors significantly associated with poor udder health of first-parity cows were feeding first-parity cows sugar-beet pulp or corn silage, and to give silage from a different batch to pregnant heifers than to lactating cows. Moreover, to have sawdust or shavings in the calving pen, to be moved from the calving pen > or =2 days after calving, to milk first-parity cows at the calving site instead of in the parlor, and to rinse, clean or disinfect milking units before a first-parity cow was milked were also significantly associated with poor udder health of first-parity cows. The results indicate that different control measures must be taken depending on the nature of the udder health problem.

Prediction of Vitamin A, Vitamin E, Selenium and Zinc Status of Periparturient Dairy Cows Using Blood Sampling During the Mid Dry Period

AbstractVitamins A and E, and the trace elements selenium (Se) and zinc (Zn) are essential for the health and performance of dairy cows. Their concentrations often decrease around calving and extra supplementation is sometimes recommended at that time. However, the need for this varies, for example depending on quantity and quality of feedstuffs in the diet. The aim of this study was to measure the concentrations of serum vitamin A (S-vit A) and vitamin E (S-vit E), plasma Se (P-Se) and serum Zn (S-Zn) in blood samples taken at several time points from one month before to one month after calving, and to evaluate if a blood sample taken during the mid dry period can accurately predict the blood concentration at calving and early lactation. Dairy cows on 3 different feeding regimens during the dry period were included in the study. A significant decrease in the concentrations of S-vit A and S-vit E, and S-Zn, was observed at calving, and P-Se was significantly lower during the dry period and at calving than in early lactation. The blood concentrations of S-vit E and P-Se in the mid dry period significantly predicted the occurrence of values considered marginal or deficient at the time of calving. The data indicate that a mid dry period concentration of ≥5.4 mg/l of S-vit E and ≥0.09 mg/l of P-Se will result in a 90% chance that the cow stays above marginal levels at calving given that a feed of the same quality is offered.SammanfattninBedömning av vitamin A-, vitamin E-, selen- och zinkstatus hos kor runt kalvning med hjälp av blodprov tagna i mitten av sinperioden Vitamin A och E, och spårelementen selen (Se) och zink (Zn) är viktiga för mjölkkornas hälsa och produktion. Koncentrationen av dessa ämnen minskar runt kalvning och extra tillskott rekommenderas ibland under denna tid. Behovet av detta varierar dock till exempel beroende på fodrets kvantitet och kvalitet. Syftet med denna studie var att mäta koncentrationen av vitamin A (S-vit A), vitamin E (S-vit E), Se (P-Se) och Zn (S-Zn) i blodprov tagna vid flera tidpunkter från en månad före till en månad efter kalvning, och att utvärdera om ett blodprov taget mitt i sinperioden kan förutsäga blodkoncentrationen vid kalvning och tidig laktation. Mjölkkor som utfodrades med 3 olika mängder av samma foderstat under sinperioden ingick i studien. En markant minskning i koncentrationen av S-vit A och S-vit E, och till en mindre del i koncentrationen av P-Se och S-Zn, sågs runt kalvning. Blodkoncentrationen av S-vit E och PSemitt i sintiden kunde signifikant förutsäga förekomst av värden vid kalvning som indikerar marginell nivå eller brist. Data tyder på att om blodprovet mitt i sintiden innehåller ≥5.4 mg/l S-vit E och ≥0.09 mg/l P-Se har kon 90% chans att nå högre nivåer vid kalvning än vad som anses marginellt om ett foder avsamma kvalitet ges.

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.