Short communication: Drying-off practices and use of dry cow therapy in Finnish dairy herds.

The objective of this study was to survey drying-off practices and use of dry cow therapy (DCT) in Finland through an online questionnaire. The questionnaire was accessible to all dairy farmers of the Finnish dairy herd recording system in 2016 (approximately 5,400 farms). In total, 715 dairy producers across the country, representative of the Finnish dairy industry, participated in the survey. Cows were dried off gradually in most of the farms. Most farms (78%) reported using selective DCT, whereas 9% of farms did not use any DCT, and 13% of farms applied blanket DCT. A significant trend was observed with increasing herd size and proportion of farms using blanket DCT. Percentage of farms using blanket DCT was also higher in farms with automatic milking system. Farmer's own experience was the most commonly reported reason for choosing a particular approach to DCT. Microbiological testing of milk samples at dry-off was the preferred method of selecting cows for DCT; 82 and 64% of farms using selective and blanket DCT approach, respectively, reported testing milk samples before treatment. The second most common criteria for using antibiotic DCT were clinical mastitis history and high somatic cell count. A high number of farms using selective DCT reported treating only up to one-fourth of their cows at dry-off. Information acquired on drying-off practices in Finland allows for future monitoring of prudent antimicrobial usage at dry-off.

Short communication: Lameness impairs feeding behavior of dairy cows.

The automated, reliable, and early detection of lameness is an important aim for the future development of modern dairy operations. One promising indicator of lameness is a change in the feeding behavior of a cow. In this study, the associations between feeding behavior and lameness were evaluated. A herd of 50 cows was investigated during the winter season in a freestall barn. Feeding behavior, feed intake, milk yield, and body weight were monitored using electronic feeding troughs and an automated milking system. Gait scoring every second week was used as a measure of lameness. To analyze the effect of lameness on feeding behavior and milk yield, linear mixed models were used. Cows with more severe lameness spent less time feeding per day (104 ± 4, 101 ± 4, and 91 ± 4 min/d for lameness scores 2, 3, and 4, respectively). An interaction between parity and lameness score was detected, with severely lame primiparous cows spending the least time feeding. Severely lame cows fed faster; however, their body weights were lower than for less-lame cows. Increase in lactation stage was associated with longer daily feeding time, longer duration of feeding bouts, and lower feeding rate. Worsening of gait was associated with lower silage intake and less time spent feeding even before severe lameness was scored. The results indicate that lameness is associated with changes in feeding behavior and that such changes could be considered in the future development of remote monitoring systems. It should also be noted that impaired feeding behavior along with lameness can put the welfare of especially early lactating primiparous cows at risk.

Efficacy of 5-day parenteral versus intramammary benzylpenicillin for treatment of clinical mastitis caused by gram-positive bacteria susceptible to penicillin in vitro.

The efficacy of parenteral (intramuscular) or intramammary (IMM) benzylpenicillin treatment for clinical mastitis caused by gram-positive bacteria susceptible to penicillin in vitro was investigated. Cows with clinical mastitis in 1 udder quarter were randomly placed into 2 treatment groups. The preliminary bacteriological diagnosis of intramammary infection (IMI) was based on on-farm culturing, and the bacteriological diagnoses were later confirmed by a quantitative PCR assay. Clinical mastitis caused by gram-positive bacteria susceptible to benzylpenicillin was treated with penicillin via either the parenteral route (20mg/kg) or IMM route (600mg) once per day for 5d. The outcome of the treatment was evaluated 3 to 4wk after the onset of the treatment. The affected quarter was examined to assess the clinical cure, and milk samples were collected from the affected quarter to determine the bacteriological cure and milk N-acetyl-ß-d-glucosaminidase activity. The survival and the composite milk somatic cell counts of the treated cows were followed up for 6 and 3mo after treatment, respectively. A total of 140 cows with clinical mastitis were included in the study, 61 being treated with benzylpenicillin parenterally and 79 via the IMM route. From all quarters treated, 108 of 140 (77.1%) were cured clinically and 77 of 140 (55.0%) were cured bacteriologically. The route of treatment did not significantly affect the outcome of the treatment; 80.3% of the quarters with parenteral treatment and 74.7% of the quarters with IMM treatment showed a clinical cure, and 54.1 and 55.7% a bacteriological cure, respectively. The milk N-acetyl-ß-d-glucosaminidase activity was significantly lower in the quarters with a clinical or bacteriological cure than in the quarters with no cure. The 6-mo survival and the proportion of cows with composite milk somatic cell counts <200,000/mL among the treated cows during the 3-mo follow-up period did not significantly differ between the treatment groups. In conclusion, the outcome of either parenteral or IMM benzylpenicillin treatment of clinical mastitis caused by penicillin-susceptible bacteria was similar.

Milk haptoglobin, milk amyloid A, and N-acetyl-ß-D-glucosaminidase activity in bovines with naturally occurring clinical mastitis diagnosed with a quantitative PCR test.

The associations between quantitative bacteriological results from a real-time PCR test and concentrations of acute-phase proteins (APP) and N-acetyl-ß-d-glucosaminidase (NAGase) activity in milk in naturally occurring clinical mastitis were investigated. Milk APP concentrations and NAGase activity in clinical mastitis caused by different udder pathogens were studied. The associations between the severity of the clinical signs and concentrations of APP and NAGase activity were estimated. Milk samples from 281 cases of clinical mastitis were collected from 3 Estonian dairy farms and analyzed by PCR to identify pathogens. Twenty-seven samples out of 281 (9.6%) were PCR negative. Milk samples containing 4 or more bacterial species (n=28) were considered possibly contaminated and excluded from all further analyses. In total, 443 bacterial identifications were made from the remaining 226 milk samples. A single bacterial species was detected in 68 samples (30.1%), 2 species were detected in 99 samples (43.8%), and 3 species were detected in 59 (26.1%) samples. To determine the inflammatory response in the udder, the concentrations of milk amyloid A (MAA) and haptoglobin (Hp) and NAGase activity in the milk were analyzed. A significant positive association was found between the severity of the clinical signs and inflammatory markers in the milk. Milk amyloid A and Hp concentrations and NAGase activity were significantly higher in samples with large quantities of bacterial DNA from Escherichia coli or Streptococcus dysgalactiae compared with milk samples not containing those species. Large quantities of bacterial DNA from Trueperella pyogenes or Streptococcus uberis in the milk were associated with elevated concentrations of Hp and high NAGase activity, but not with increased MAA concentrations. Milk samples containing Corynebacterium bovis and coagulase-negative staphylococci had significantly lower concentrations of MAA and Hp and lower NAGase activity compared with samples where these species were not detected. It can be concluded that concentrations of APP and NAGase activity in the milk were associated with the quantity of bacterial DNA in the milk samples.

Dry cow therapy and early lactation udder health problems-Associations and risk factors.

Mastitis remains the most expensive disease of dairy cows, and antibiotic dry cow therapy (DCT) at dry-off is an important part of mastitis control. Regardless of the infection status, blanket DCT is administered to all quarters of all cows, which is controversial due to the worldwide problem of antimicrobial resistance. Even though selective DCT of only infected cows is a more sustainable approach, choosing animals for treatment is not always straightforward. Our aim was to evaluate whether the herd-level DCT approach is associated with early lactation udder health problems, taking into account the cow characteristics. The information source was 2015-2017 Dairy Herd Improvement data with 7461 multiparous cows from 241 Finnish dairy herds. Information on the herd-level DCT approach was obtained from farmers' questionnaire responses in 2017, and the three different approaches were selective DCT, blanket DCT, and no DCT. The statistical tool for the data analysis was a generalized linear mixed model with a random herd effect for binary outcomes and a linear mixed model with a random herd effect for a continuous outcome. The two binary outcomes were the odds of having high milk somatic cell count (SCC ≥ 200 000 cells/mL) on the first test-day within 5-45 days in milk (DIM) and the odds of mastitis treatment in early lactation up to 45 DIM. The third outcome was the mean milk lnSCC (× 1000 cells/mL) within 120 DIM. Selective DCT was the prevailing treatment practice in our data. Blanket DCT was associated with lower SCC after calving. Cows more likely to have high SCC after calving were older cows, cows with high average SCC during the previous lactation, and cows with high milk yield near dry-off. A mastitis treatment in the early lactation was more likely if, during the previous lactation, the cow had high average SCC, high peak milk production, or high milk yield near dry-off. Our findings indicate that DCT is still effective in mastitis control. Cows with high milk yield, especially near dry-off, and cows with persistently high SCC require attention when considering next lactation udder health.

Efficacy of enrofloxacin in the treatment of naturally occurring acute clinical Escherichia coli mastitis.

The efficacy of the combination of systemic enrofloxacin (5mg/kg twice with a 24-h interval, first dose i.v., second dose s.c.) and the nonsteroidal antiinflammatory agent ketoprofen (3mg/kg i.m. or 4 mg/kg per os daily for 1 to 3 d) treatment was compared with antiinflammatory treatment only in dairy cows with naturally occurring acute clinical Escherichia coli mastitis. A total of 132 cows with acute clinical mastitis and with confirmed growth of E. coli in a pretreatment milk sample were randomly allocated to 1 of 2 treatment groups. Response to treatment was evaluated clinically and by bacteriological culturing and determination of N-acetyl-beta-d-glucosaminidase (NAGase) activity on d 2 and 21 posttreatment. Enrofloxacin treatment did not increase bacteriological (90.5% of treated vs. 86.8% of nontreated cured) or clinical cure (46.7% of treated vs. 57.1% of nontreated cured), cow survival (95.3% of treated vs. 92.7% of nontreated), or quarter milk production assessed 21 d posttreatment (21.8 vs. 29.3% return to preinfection level for nontreated cows), nor did it decrease mammary gland tissue damage estimated using determination of milk NAGase activity (24.0+/-0.3 vs. 18.3+/-1.3 pmol of 4-methylumbelliferone per min per microL for nontreated cows). Treatment did not influence the number of study cows remaining in the herd after 6 mo (71.9% of treated vs. 80.6% of nontreated). The only significant effects of enrofloxacin were enhancing the bacteriological cure (odds ratio=3.32 for treated cows) and decreasing the clinical cure (odds ratio=0.05 for treated cows) on d 2 posttreatment. Our results did not support the use of enrofloxacin to treat acute clinical E. coli mastitis.

Antibiotic dry cow therapy, somatic cell count, and milk production: Retrospective analysis of the associations in dairy herd recording data using multilevel growth models.

Antibiotic dry cow therapy (DCT) is an important part of most mastitis control programs. Updating DCT recommendations is an ongoing topic due to the global problem of antimicrobial resistance. Finland, along with other Nordic countries, has implemented selective DCT for decades. Our study analyzed Dairy Herd Improvement (DHI) information from 241 Finnish farmers who participated in a survey about their drying-off practices. The aim was to evaluate herd-level associations between milk somatic cell count (SCC), milk production, and various antimicrobial DCT approaches both cross-sectionally in 2016 and longitudinally in 2012-2016. The three DCT approaches in the study were selective, blanket, and no DCT use. An additional aim was to evaluate whether dynamic changes occurred in herd-average SCC and annual milk production over five years, and whether these potential changes differed between different DCT approaches. The method for the longitudinal analyses was growth modeling with random coefficient models. Differences in SCC and milk production between farms with different DCT approaches were minor. Regardless of the farm's DCT approach, annual milk production increased over the years, while average SCC was reasonably constant. The variability in SCC and milk production across all DCT groups was low between years, and most of the variability was between farms. Compared to other milking systems, farms with automatic milking system (AMS) had higher SCC, and in 2016 higher milk production. The results of this study suggest that it is possible to maintain low herd-average SCC and good milk production when using selective DCT and following the guidelines for prudent antimicrobial use. Average SCC and milk production varied across the herds, suggesting that advice on DCT practices should be herd-specific. The methodology of growth modeling using random coefficient models was applicable in analyzing longitudinal data, in which the time frame was relatively short and the number of herds was limited.

Host response in bovine mastitis experimentally induced with Staphylococcus chromogenes.

An experimental infection model was developed to study host response to intramammary infection in cows caused by Staphylococcus chromogenes. CNS intramammary infections have become very common in modern dairy herds, and they can remain persistent in the mammary gland. More information would be needed about the pathophysiology of CNS mastitis, and an experimental mastitis model is a means for this research. Six primiparous Holstein-Friesian cows were challenged with S. chromogenes 4 weeks after calving. One udder quarter of each cow was inoculated with 2.1 x 10(6)cfu of S. chromogenes. All cows became infected and clinical signs were mild. Milk production of the challenged quarter decreased on average by 16.3% during 7 days post-challenge. Cows eliminated bacteria in a few days, except for one cow which developed persistent mastitis. Milk indicators of inflammation, SCC and N-acetyl-beta-D-glucosaminidase (NAGase) returned to normal within a week. Milk NAGase activity increased moderately, which reflects minor tissue damage in the udder. Concentrations of serum amyloid A (SAA) and milk amyloid A (MAA) were both elevated at 12h PC. MAA was affected by the milking times, and was at its highest before the morning milking. In our experimental model, systemic acute phase protein response with SAA occurred as an on-off type reaction. In conclusion, this experimental model could be used to study host response in CNS mastitis caused by the main CNS species and also for comparison of the host response in a mild intramammary infection and in more severe mastitis models.

Real-time polymerase chain reaction-based identification of bacteria in milk samples from bovine clinical mastitis with no growth in conventional culturing.

In more than 30% of milk samples from clinical and subclinical bovine mastitis, bacteria fail to grow even after 48 h of conventional culture. The "no-growth" samples are problematic for mastitis laboratories, veterinarians, and dairy producers. This study provides the first investigation of the bacteriological etiology of such samples, using a real-time PCR-based commercial reagent kit. The assay targets the DNA of the 11 most common bacterial species or groups in mastitis and the staphylococcal blaZ gene (responsible for penicillin resistance) and can identify and quantify bacterial cells even if dead or growth-inhibited. A study was made of 79 mastitic milk samples with no-growth bacteria in conventional culture, originating from cows with clinical mastitis. Of the 79 samples, 34 (43%) were positive for 1 (32 samples) or 2 (2 samples) of the target bacteria. The positive findings included 11 Staphylococcus spp. (staphylococci other than Staphylococcus aureus), 10 Streptococcus uberis, 2 Streptococcus dysgalactiae, 6 Corynebacterium bovis, 3 Staph. aureus, 1 Escherichia coli, 1 Enterococcus, and 1 Arcanobacterium pyogenes. The positive samples contained as many as 10(3) to 10(7) bacterial genome copies per milliliter of milk. This study demonstrates that in nearly half of the clinical mastitis cases in which conventional culture failed to detect bacteria, mastitis pathogens were still present, often in substantial quantities. The clearly elevated N-acetyl-beta-d-glucosaminidase activity values of the milk samples, together with clinical signs of the infected cows and quarters, confirmed the diagnosis of clinical mastitis and indicated that real-time, PCR-based bacterial findings are able to reveal bacteriological etiology. We conclude that all common mastitis bacteria can occur in large quantities in clinical mastitis samples that exhibit no growth in conventional culture, and that the real-time PCR assay is a useful tool for bacteriological diagnosis of such milk samples. Low bacterial concentration is commonly speculated to explain the no-growth milk samples. This hypothesis is not supported by the results of the current study.

Comparison of Finnish meat inspection records and average daily gain for cattle herds differing in Mycoplasma bovis test-status.

Detecting Mycoplasma bovis on cattle farms represents a challenge in the absence of an outbreak or cases of M. bovis mastitis, yet identification of an infection is essential to control the spread of the disease successfully. The objectives of this study were: (1) to determine whether meat inspection records can aid identification of cattle farms supporting M. bovis infection, and (2) to compare the average daily weight gain estimated from carcass weight for cattle originating from farms differing in M. bovis test-status. Meat inspection records were collected from two abattoirs in 2015; 80 677 animals in total. All the dairy and mixed breed cows and bulls used for meat production were categorized according to known M. bovis infection status of the farms from which the cattle were derived; positive, contact or control farms. The associations between animals from different M. bovis categories and lung lesions of bulls and cows (pneumonia and pleuritis), identified during meat inspection, and estimated average daily gain (ADG) of bulls, were investigated. The odds ratios for lung lesions, especially pleuritis, were higher in M. bovis test-positive or contact farms compared with control farms. Additionally, odds ratios for pleuritis were higher among animals from M. bovis test-positive farms and animals from contact slaughtering farms originating from M. bovis-free rearing farms. Bulls originating from M. bovis test-positive farms had higher estimated average daily gain than cattle from control farms. Meat inspection records can be used alongside other methods to detect M. bovis-positive farms where M. bovis causes lung lesions.

Characterisation of the course of Mycoplasma bovis infection in naturally infected dairy herds.

Mycoplasma bovis causes bovine respiratory disease, mastitis, arthritis and otitis. The importance of M. bovis has escalated because of recent outbreaks and introductions into countries previously free of M. bovis. We characterized the course of M. bovis infection on 19 recently infected dairy farms over 24 months. Our objective was to identify diagnostic tools to assess the efficacy of control measures to assess low risk infection status on M. bovis infected farms. PCR assays and culture were used to detect M. bovis, and in-house and BioX ELISAs were used to follow antibody responses. Cows and young stock were sampled on four separate occasions, and clinical cases were sampled when they arose. On 17 farms, a few cases of clinical mastitis were detected, mostly within the first eight weeks after the index case. Antibodies detected by in-house ELISA persisted in the serum of cows at least for 1.5 years on all farms, regardless of the M. bovis infection status or signs of clinical disease or subclinical mastitis on the farm. Six out of 19 farms became low risk as the infection was resolved. Our results suggest that, for biosecurity purposes, regular monitoring should be conducted on herds by screening for M. bovis in samples from cows with clinical mastitis and calves with pneumonia, in conjunction with testing young stock by screening longitudinally collected nasal swabs for M. bovis and sequential serum samples for antibody against recombinant antigen.

The effect of a cannula milk sampling technique on the microbiological diagnosis of bovine mastitis.

Two methods of collecting milk samples from mastitic bovine mammary quarters were compared. Samples were taken in a consistent order in which standard aseptic technique sampling was done first, followed by insertion of a sterile cannula through the teat canal and collection of a second sample. Microbiological results of those two sampling techniques were compared. Milk samples were analysed using multiplex real-time polymerase chain reaction (PCR). The cannula technique produced a reduced number of microbial species or groups of species per sample compared with conventional sampling. Staphylococcus spp. were the most common species identified and were detected more often during conventional sampling than with cannula sampling. Staphylococcus spp. identified in milk samples could also have originated from the teat canal without being present in the milk. The number of samples positive for Trueperella pyogenes or yeasts in the conventional samples was twice as high as in the cannula samples, indicating that the presence of Trueperella pyogenes and yeast species should not necessarily be interpreted as being the causative agents of bovine intra-mammary infections (IMI).

Is the biofilm formation and slime producing ability of coagulase-negative staphylococci associated with the persistence and severity of intramammary infection?

Biofilm and slime formation assists bacteria in avoiding the host immune defence and antimicrobial therapy. It is suspected to affect the severity or persistence of mastitis caused by coagulase-negative staphylococci (CNS), which are a common cause of bovine mastitis. The phenotypic biofilm formation ability of 244 CNS isolates (199 isolates from bovine mastitis and 52 type and reference strains) was investigated with a tissue culture plate (TCP) assay and fluorescent in situ hybridization (FISH). Slime production of the strains was assessed using Congo red agar (CRA) plates. Additionally, genes encoding the adhesion proteins MSCRAMM (microbial surface components recognizing adhesive matrix molecules) and biofilm-associated proteins (bap) were detected. The severity of intramammary infection (IMI) in mastitis from which the isolates originated was measured with milk N-acetyl-ß-D-glucosaminidase (NAGase) activity. One-third of isolates from mastitis produced biofilm when analysed with TCP or FISH. The kappa test value, measuring the agreement between two tests, differed between CNS species. Slime production was less frequent for isolates of the common mastitis species Staphylococcus chromogenes (0.2% of isolates produced slime) and Staphylococcus simulans (3.5%) compared to Staphylococcus epidermidis (40%). No association was found between the phenotypic ability to form biofilm and the persistence of IMI or severity of mastitis. Slime production was rare in isolates originating from IMI. Only 12.7% of isolates from persistent IMI and 1.8% of isolates from spontaneously eliminated IMI produced slime. The eno gene encoding laminin-binding protein was most frequently detected among the isolates from mastitis, 75% of them having this gene. Only a few other MSCRAMM genes were detected.

Acute phase proteins in milk in naturally acquired bovine mastitis caused by different pathogens.

The concentrations of haptoglobin (Hp) and serum amyloid A (SAA) and the activity of N-acetyl-ß-D-glucosaminidase (NAGase) in milk from 234 cows with spontaneous mastitis caused by different pathogens were measured to assess whether they corresponded with the clinical signs of mastitis and whether there were any differences between pathogens. Ninety-eight of the cows had clinical mastitis and 136 had subclinical mastitis. There were statistically significant positive correlations between the concentrations of SAA and Hp and the activity of NAGase. Significant differences in the concentrations of acute phase proteins and NAGase activity were found in milk from cows with mastitis caused by different pathogens. The highest concentrations of Hp and NAGase were found in cases of mastitis caused by Escherichia coli and Arcanobacterium pyogenes, and the lowest concentrations were from cases of mastitis caused by coagulase-negative staphylococci. Very low SAA concentrations were found in milk from the cases caused by A pyogenes, in contrast to cases caused by other major mastitis pathogens. The median concentration of SAA was over 10 times higher in cases of mastitis caused by E coli than in mastitis caused by other pathogens. There were significant differences in the mean Hp concentration and NAGase activity between clinical and subclinical mastitis. In approximately one-third of the samples, the Hp concentration was below the detection limit, potentially compromising the use of Hp as a mastitis marker.