Changes in mammary infection status in dairy cows during the dry period using dry cow therapy approaches on three farms.

This case study evaluated the mammary infection status of dairy cows during the dry periods and explored the associated problems in their quarters with dry cow therapy (DCT). This study assessed intramammary infections, antibiotic efficacy, and antimicrobial resistance of pathogens in 464-quarter milk samples from 59 dairy cows during the dry periods after applying blanket DCT, non-DCT, and selective DCT approaches on three farms. The recovery rates of intramammary infections were 95% (19/20 quarters) with blanket DCT on farm A, 70% (14/20) with non-DCT on farm B, and 19% (4/21) with selective DCT on farm C. Analysis of mammary infections in cows with DCT revealed that mammary infections were controlled by blanket DCT, well controlled by non-DCT, and substantial problems remained in selective DCT. Lower intramammary infection prevalence in the quarters at postpartum appeared to be associated with higher recovery of mammary infections, fewer new infections, and lower uncured mammary infections within the herds. Antibacterial resistance in 14 coagulase-negative staphylococci isolated to six antimicrobial drugs was suggested to be linked to antibiotic use on the farm. Follow-up studies on the quarter-based infection status with DCT will assist in improving mastitis control in cows during the dry period.

The relationships between environmental parameters in livestock pen and physiological parameters of Holstein dairy cows.

There has been an increase in temperature and the incidence of extreme weather events, such as heat wave, due to global warming, which has promoted the incidence of livestock diseases. Therefore, it is important to examine the effect of changes in environmental parameters on livestock performance. The aim of this study was to examine the relationship between ambient environmental conditions in livestock pen and the physiological parameters of Holstein dairy cows. The results showed that there was a decrease in the red blood cell counts, hemoglobin concentrations, and mean corpuscular hemoglobin concentration of the cows with increasing pen temperature, wet bulb globe temperature (WBGT), and temperature humidity index (THI). Additionally, high daily variation in temperature caused a decrease in the serum albumin levels of the cows. Moreover, the lowest serum calcium, inorganic phosphorus, and magnesium concentrations were observed in November, and were negatively correlated with the 24-hr temperature, WBGT, and THI range of the pen prior to sampling. Multiple regression analysis showed a positive correlation between serum cortisol concentration and 24-hr WBGT range of the pen prior to samplings and packed cell volume. However, serum cortisol and total protein concentrations were negatively correlated. Overall, the findings of the study suggest that large variation in temperature induced stress in the cows, which could be overcome by increased water consumption and improved protein digestion and absorption by the animals, and the addition of minerals, such as calcium to the diet.

A novel polymerase chain reaction assay for the detection of seven Mycoplasma species of cattle origin.

The study aimed to develop a pair of polymerase chain reaction primers for detecting ruminant mycoplasma pathogens. We designed a set of primers based on the most similar sequences within 16 S rRNA regions of seven Mycoplasma spp. These primers have high sensitivity for detecting Mycoplasma dispar, M. arginine, M. canadense, M. bovis, M. alkalescens, M. californicum, and M. bovigenitalium within the annealing temperature range of 46 to 48 °C. The minimum amount of DNA that can be detected using the protocol is 250 ng, which is equivalent to 2,000 colony-forming units per mL. The primers can detect mycoplasma from DNA extracted directly from milk samples. The common bovine mastitis pathogens of Staphylococcus aureus coagulase-negative staphylococci, Escherichia coli, Streptococcus uberis, Klebsiella pneumonia, and Kocuria rosea were not detected by the primers. We believe the high sensitivity and specificity of these primers make them useful for detecting infection with seven Mycoplasma species in ruminants, allowing the primers to be used in clinical settings.

Evaluation of the health status of mammary glands and compositional changes in udder-half milk obtained from dairy goats for milk quality management.

This study evaluated the health status of the mammary glands and milk composition of dairy goats. The California mastitis test (CMT) score, somatic cell counts (SCCs), somatic cell type, electrical conductivity (EC), N-acetyl-ß-d-glucosaminidase (NAGase) activity, mastitis-causing pathogens, and milk composition in 121 udder-half milk samples from 62 crossbreed goats (1-3 years old) at 23-45 days postpartum were compared in four categories with SCCs of <200 × 103 , <300 × 103 , 301-1000 × 103 , and >1010 × 103 cells/ml. The SCC, CMT score, EC, and NAGase activity were significantly lower (p < 0.05) in udder-half milk with SCCs of <200 × 103 and <300 × 103 cells/ml than those in milk with SCCs of 301-1000 × 103 and >1010 × 103 cells/ml. The protein, lactose/ash, and solids-not-fat (SNF) in milk with an SCC of <300 × 103 cells/ml were similar to the previously reported values. The proportion of polymorphonuclear neutrophils was significantly higher (p < 0.05) in milk with SCCs of 301-1000 and >1010 × 103 cells/ml than in milk with an SCC of <300 × 103 cells/ml. Assessing mammary gland health and milk composition based on categorization of udder-half milk by SCC may be useful for milk quality control on goat farms.

Somatic cell and innate immune responses in mammary glands of lactating cows to intramammary infusion of Bifidobacterium breve at pre-drying off period.

Intramammary infusion of Bifidobacterium breve (B. breve)-induced somatic cell (SC) counts, chemiluminescent response (CL), lactoferrin (LF) concentrations and mastitis-causing pathogens from quarters with subclinical mastitis were measured to evaluate innate immune response of mammary glands in dairy cows at 3 to 4 weeks before drying off. SC counts in 7 quarters of 7 control cows and 5 quarters of 6 cows with mastitis increased markedly on day 1 and SC values in control cows were significantly (P<0.05) increased and returned to pre-infusion levels on day 5 after B. breve-infusion. CL values in both groups increased markedly on day 1 and then decreased after B. breve-infusion; however, CL values in cows with mastitis did not return to normal levels on day 5 and at postpartum. The CL values were highly correlated with their SC counts in milk from both groups. LF concentrations increased toward day 3 after B. breve-infusion and were higher in cows with mastitis. B. breve-infusion eliminated 16.6% (1/6) of pathogens from 6 quarters with chronic subclinical mastitis. B. breve-induced SC responses in quarters from 3 cows with mastitis showed characteristic patterns of recovery, persistent and new infections. B. breve-induced SC counts in quarters from the cows in the pre-drying off were lower (25.7-70.6%) than those of the cows in mid-lactation. The intrinsic innate immune response in cows on pre-drying off may be decreased and appears to be insufficient to eliminate pathogens from mammary gland in the pre-drying off.

Innate immune response in bovine neutrophils stimulated with Mycoplasma bovis.

Mycoplasma bovis (M. bovis) is a significant worldwide pathogen of cattle. Neutrophils have an important role in the innate immune response during infection with M. bovis. However, even though neutrophils accumulate in M. bovis infection, the interaction of M. bovis and neutrophils has not been fully elucidated. We attempted to elucidate the innate immune response of neutrophils stimulated with M. bovis and evaluate the transcriptome and functional analysis of bovine neutrophils stimulated with M. bovis. Proinflammatory cytokines, such as inducible nitric oxide (iNOS), which was the most increased gene in transcriptome analysis, were increased in quantitative polymerase chain reaction analysis of bovine neutrophils stimulated with live or heat-killed M. bovis. Nitric oxide and intracellular reactive oxygen species production of neutrophils stimulated with M. bovis was significantly increased. Neutrophils stimulated with M. bovis showed an increased ratio of nonapoptotic cell death compared to unstimulated controls. We demonstrated that neutrophil extracellular traps (NETs) formation was not recognized in neutrophils stimulated with live M. bovis. However, heat-killed M. bovis induced NETs formation. We also showed the interaction with M. bovis and bovine neutrophils regarding proinflammatory cytokine gene expression and functional expression related to NETs formation. Live and killed M. bovis induced innate immune responses in neutrophils and had the potential to induce NETs formation, but live M. bovis escaped NETs.

Forestomach developmental failure in an 11-month-old Japanese Black steer with severely retarded growth and chronic ruminal tympany.

This study reports findings from the pathological examination of the forestomach of an 11-month-old Japanese Black steer with severely retarded growth (41% of expected weight) and chronic ruminal tympany. The ruminal papillae were weakly formed (0.3-0.5 cm long) and unevenly distributed. The cellulae and cristae reticuli were underdeveloped; the cristae were 0.4-0.7 cm in height and milky white. The keratinized layer in the stratified squamous epithelium was thickened. Ruminal pH was 5.25, and ruminal volatile fatty acid concentration was 11.7 mM. The steer's severely retarded growth was considered to be caused by malnutrition due to developmental and functional failure of the forestomach.

Transcriptome analysis of Mycoplasma bovis stimulated bovine peripheral blood mononuclear cells.

Mycoplasma bovis is a pathogenic bacterium in bovines that causes huge global economic losses. Numerous factors play important roles in M. bovis pathogenesis; however, the host immune response involved in M. bovis infection has not been fully elucidated. We aimed to determine the characteristics of the host immune response to Mycoplasma infection. We evaluated the responsiveness of bovine peripheral blood mononuclear cells (PBMCs) stimulated with M. bovis via microarray analysis. The transcriptional abundance of innate immune-related genes IL-36A, IL-27, IFN-γ, and IL-17 in PBMCs increased after M. bovis exposure. Upon M. bovis infection, there was increased expression of the lymphocyte activated genes basic leucine zipper transcription factor (BATF) and signaling lymphocytic activation molecule family members 1 and 7 (SLAMF 1 and SLAMF 7) in PBMCs compared with that in unstimulated cells. The study revealed that the transcriptional abundance of innate immunity genes in PBMCs increased during M. bovis infection. This induced the activation of PBMCs, giving rise to an immune response, which is followed by the development of the inflammatory response. The results from this study could be used as the basis for the development of novel vaccine candidates against M. bovis.

Innate immune response of mammary gland induced by intramammary infusion of Bifidobacterium breve in lactating dairy cows.

This study aimed to evaluate innate immune responses of mammary glands induced by intramammary infusion of Bifidobacterium breve in dairy cows. Somatic cell counts in quarters of cows showed a marked increase following B. breve infusion on days 1 and 2. Opsonized-stimulated chemiluminescence response in quarter milk was significantly (P<0.05) increased by B. breve infusion on days 1 to 3 compared to that of pre-infusion. Lactoferrin concentrations in B. breve-infused quarter milk increased significantly (P<0.05) on days 2 to 4 and 6 compared to those of pre-infusion. IgG and IgA concentrations in B. breve-infused quarters significantly (P<0.05) increased on days 2 to 4 for IgG and days 3, 4, 6 and 8 for IgA compared to those of pre-infusion. Interleukin (IL)-1ß and IL-8 mRNA levels in somatic cells from B. breve-infused quarters were significantly (P<0.05) upregulated on day 1 compared to those on days 0 and 14. Conversely, IL-6 mRNA levels in somatic cells from B. breve-infused quarters on days 0, 1 and 14 and NF-κB mRNA levels on day 0 were significantly (P<0.05) down-regulated compared to those of control. IL-1ß, tumor necrosis factor (TNF)-α and IL-6 concentrations increased on days 1, 3 and 7 after B. breve infusion in quarters. Intramammary infusion of B. breve (3 × 109 cfu) induces a massive influx of leukocytes and enhances innate immune response in mammary glands. This event may contribute to the enhancing host defense in the mammary gland.

Effects of intramammary infusion of Bifidobacterium breve on mastitis pathogens and somatic cell response in quarters from dairy cows with chronic subclinical mastitis.

The present study assessed the effects of intramammary infusion of Bifidobacterium breve (B. breve) on mastitis-causing pathogens and on the somatic cell counts (SCC) in lactating cows with chronic subclinical mastitis. The bacteriological cure rates of 42 quarters from 42 cows infected with Staphylococcus aureus, Corynebacterium bovis, coagulase-negative staphylococci, and environmental streptococci were 18.2% (2/11), 14.3% (1/7), 58.8% (10/17), and 28.6% (2/7), respectively, on day 14 after B. breve infusion. In a second trial, B. breve was infused into 18 quarters from 18 cows with chronic subclinical mastitis from which pathogens had not been isolated; the rates of quarters showing SCC > 50 × 104 cells/ml prior to B. breve infusion that decreased to < 30 × 104 cells/ml after infusion were significantly (p < .01) increased to 61.1% (11/18) on day 14 compared to that prior to infusion (0/18). The intramammary infusion of B. breve appears to be a non-antibiotic approach for elimination of minor pathogens and decreasing SCC in quarters with chronic subclinical mastitis in dairy cows.

Prevalence of sub-clinical mastitis and its association with milking practices in an intensive dairy production region of Uganda.

A cross-sectional study was conducted to investigate the risk factors for sub-clinical mastitis (SCM) in Mbarara District, an intensive dairy production region of Uganda where hand-milking is dominant. In 30 farms, herd-level milking practices and SCM prevalence were studied. The SCM prevalences were 68.6% (417/608, 95% confidence interval (CI): 64.9-72.2%) and 39.2% (946/2,411, 37.3-41.2%) at the cow- and quarter-levels, respectively. A preventive factor for SCM was cow calmness at the end of milking (OR: 0.20, 95%CI: 0.05-0.79, P=0.021); a risk factor was rough teat-end (OR: 1.75, 95%CI: 1.14-2.68, P=0.011). Good cow hygiene was negatively associated with environmental mastitis (P=0.002). Appropriate hand-milking practices that avoid teat damage are expected to reduce SCM in Uganda.

Immunosuppression in Cows following Intramammary Infusion of Mycoplasma bovis.

Mycoplasma bovis is a destructive pathogen that causes large economic losses in rearing cattle for beef and dairy worldwide. M. bovis causes suppression of and evades the host immune response; however, the mechanisms of host immune function involved in M. bovis mastitis have not been elucidated. The purpose of this study was to elucidate the characteristics of the bovine immune response to mycoplasmal mastitis. We evaluated the responsiveness of the bovine mammary gland following infusion of M. bovis Somatic cell counts and bacterial counts in milk from the infected quarter were increased. However, the proliferation of peripheral blood mononuclear cells (blood MNCs) and mononuclear cells isolated from M. bovis-stimulated mammary lymph nodes (lymph node MNCs) did not differ from that in the unstimulated cells. Transcriptome analysis revealed that the mRNA levels of innate immune system-related genes in blood MNCs, complement factor D (CFD), ficolin 1 (FCN1), and tumor necrosis factor superfamily member 13 (TNFSF13) decreased following intramammary infusion of M. bovis The mRNA levels of immune exhaustion-related genes, programmed cell death 1 (PD-1), programmed cell death-ligand 1 (PD-L1), lymphocyte activation gene 3 (LAG3), and cytotoxic T-lymphocyte-associated protein 4 (CTLA4) of milk mononuclear cells (milk MNCs) in the infected quarter were increased compared with those before infusion. Increase in immune exhaustion-related gene expression and decrease in innate immune response-related genes of MNCs in quarters from cows were newly characterized by M. bovis-induced mastitis. These results suggested that M. bovis-induced mastitis affected the immune function of bovine MNCs, which is associated with prolonged duration of infection with M. bovis.

Effect of Mycoplasma bovis on expression of inflammatory cytokines and matrix metalloproteinases mRNA in bovine synovial cells.

Mycoplasma bovis causes chronic arthritis in calves. Mycoplasma arthritis shows severe inflammatory reactions in joints that is commonly treated with antibiotics and results in significant economic losses in the calf industry. A previous study showed that inflammatory cytokines and matrix metalloproteinases (MMPs) produced by synovial cells promote progression of the pathophysiology of bacterial arthritis. However, the mechanism underlying the pathogenesis of bovine Mycoplasma arthritis has not been fully clarified. In this study, we examined the immunologic response of bovine synovial tissue to M. bovis. We observed significant increases in expression of interleukin (IL)-1ß, IL-6, IL-8, MMP-1, and MMP-3 mRNA in synovial tissue from Mycoplasma arthritis calves compared with tissues from normal calves. Expression of IL-6, IL-8, and MMP-1 mRNA was also induced in cultured synovial cells stimulated with M. bovis, but not expression of IL-1ß and MMP-3 mRNA. In contrast, the culture supernatant of peripheral blood mononuclear cells stimulated with M. bovis induced marked increases in the expression of IL-1ß, IL-6, IL-8, MMP-1, and MMP-3 mRNA in synovial cells. Our results indicate that inflammatory cytokines and MMPs produced by synovial cells play a key role in the pathogenesis of Mycoplasma arthritis. We suggest that interactions between synovial cells and mononuclear cells in the presence of M. bovis induce expression of these cytokines and MMPs in synovial cells, resulting in severe inflammatory reactions in the joints.

Innate immune response of bovine mammary epithelial cells to Mycoplasma bovis.

Mycoplasma spp. are contagious bacteria, and mycoplasmal mastitis is a serious productivity problem on dairy farms. Bovine mammary epithelial cells (bMECs) have an important role in the elimination of pathogens, but the effect of Mycoplasma bovis on bMECs has not been fully described. To elucidate the immune response against intramammary infection by M. bovis, we undertook microarray analysis to examine and profile mRNA expression in bMECs after stimulation with M. bovis. We also compared the effects of M. bovis, Staphylococcus aureus, and Escherichia coli on immune-related mRNA expression in bMECs. Transcriptome analysis indicated a significant decrease in the level of mRNA-encoding lysine-specific demethylase 4D, suggesting that the immune response is suppressed by a decrease in histone demethylase activity. Interleukin (IL)-1ß, IL-6, tumor necrosis factor alpha, toll-like receptor (TLR) 2, and TLR4 mRNA expression levels were significantly increased in bMECs stimulated with heat-killed M. bovis, but the expression levels were lower than those following stimulation by heat-killed S. aureus or E. coli. Our results suggest that M. bovis weakly affects mRNA expression in bMECs compared to the effects of E. coli or S. aureus. Moreover, live M. bovis may induce suppression of the immune response in bMECs.

Bacteriological evaluation of composted manure solids prepared from anaerobic digested slurry for hygienic recycled bedding materials for dairy cows.

Changes in mastitis-causing pathogens, pH and water content in composted manure solids (CMS) prepared from digested slurry were evaluated during turning at 2-day intervals for 8 days (C1-C4). The numbers of streptococci, coagulase-negative staphylococci and coliforms were 2.6 × 101 , 1.7 × 102 and 1.0 × 101  colony-forming units (cfu)/g in CMS (C4) (summer), and these counts were markedly lower (P < 0.05) than those in CMS (C0 and C1). The bacterial counts ranged from 101 to 1.7 × 102  cfu/g in CMS (C4) (summer) and were within approved levels, <1 × 106  cfu/g, indicating a minimal mastitis risk. The temperatures in CMS (C1-C4) increased to 63°C-74°C in summer and 67°C-70°C in winter. The mean pH values in CMS (C0-C4) were 9.2 in summer and 8.7 in winter, and water contents ranged from 61.7% to 69.6% in summer and 73.2% to 66.2% in winter. The significant decrease of pathogenic bacteria in CMS appears to be closely related to temperature >63°C for 8 days, pH 8.7-9.2, and water content 62% to 73%. This study demonstrates that prepared CMS has value as a recycled material with the potential to alleviate udder health issues in dairy cows.

Control of Pseudomonas mastitis on a large dairy farm by using slightly acidic electrolyzed water.

The disinfection effect of slightly acidic electrolyzed water (SAEW) use in a farm where Pseudomonas mastitis has spread was evaluated. Despite the application of antibiotic therapy and complete cessation of milking infected quarters, numerous new and recurrent Pseudomonas aeruginosa clinical mastitis infections (5.8-7.1% of clinical mastitis cases) occurred on the farm from 2003 to 2005. Procedural changes and equipment modifications did not improve environmental contamination or the incidence of Pseudomonas mastitis. To more thoroughly decontaminate the milking parlor, an SAEW system was installed in 2006. All milking equipment and the parlor environment were sterilized with SAEW (pH 5-6.5, available chlorine 12 parts per million) before and during milking time. After adopting the SAEW system, the incidence of clinical and subclinical Pseudomonas mastitis cases decreased significantly (P < 0.0001) and disappeared. These findings suggest that SAEW effectively reduced the incidence of mastitis in a herd contaminated by Pseudomonas species. This is the first report to demonstrate the effectiveness of disinfection by SAEW against mastitis pathogens in the environment.

Mycoplasma bovis escapes bovine neutrophil extracellular traps.

Mycoplasma bovis is a significant pathogen in bovine infections including mastitis, pneumonia, arthritis and otitis media, and is the cause of large economic losses in beef and dairy farms. During infection with M. bovis, recruited neutrophils are not sufficient to eradicate M. bovis from the infection site. The release of neutrophil extracellular traps (NETs) is one of the innate immune responses of neutrophils but the effect of M. bovis on NET formation by bovine neutrophils has not yet been clarified. The objective of our research was to examine the effect of M. bovis on NET formation and the killing activity of bovine neutrophils. We showed that NETs were not detected following stimulation of neutrophils by M. bovis alone or with Phorbol 12-myristate 13-acetat (PMA). Reactive oxygen species production is essential for NET formation but the levels in neutrophils stimulated with M. bovis at multiplicity of infections of 10, 100, and 1000 were similar to those of unstimulated cells. NET formation induced by PMA stimulated neutrophils disappeared following the addition of M. bovis but this phenomenon was not observed when ethylenediaminetetraacetic acid (EDTA) was added. M. bovis colony forming units were significantly decreased by the addition of EDTA in the presence of NETs. Our results suggested that M. bovis infection alone did not induce NETs and that M. bovis nucleases, as hypothesis-based, contributed to resistance against the killing activity of NETs.

An unusual case of thoracic ectopia cordis in a Toggenburg Goat and its three-dimensional images constructed with X-ray computed tomography.

A two-day-old female Toggenburg goat with thoracic ectopia cordis (EC) was diagnosed via radiography and computed tomography. The goat was born with EC, defects of the sternum and a supra-umbilical abdominal wall, but without the presence of Cantrell's syndrome. Necropsy and histopathological findings indicated the affected kid had malformation of the heart with an enlarged left ventricle. The findings showed the heart (9 x 5 x 5 cm) stayed outside the thorax, and was covered by a semitransparent membrane. This report is the first to describe a case of thoracic EC in a goat whose sternum was not developed fully and was not connected to the ribs. It is also the first paper to describe three-dimensional images of this condition constructed from computed tomography scans.

Phage Therapy Is Effective in a Mouse Model of Bacterial Equine Keratitis.

UNLABELLED: Bacterial keratitis of the horse is mainly caused by staphylococci, streptococci, and pseudomonads. Of these bacteria, Pseudomonas aeruginosa sometimes causes rapid corneal corruption and, in some cases, blindness. Antimicrobial resistance can make treatment very difficult. Therefore, new strategies to control bacterial infection are required. A bacteriophage (phage) is a virus that specifically infects and kills bacteria. Since phage often can lyse antibiotic-resistant bacteria because the killing mechanism is different, we examined the use of phage to treat horse bacterial keratitis. We isolated Myoviridae or Podoviridae phages, which together have a broad host range. They adsorb efficiently to host bacteria; more than 80% of the ΦR18 phage were adsorbed to host cells after 30 s. In our keratitis mouse model, the administration of phage within 3 h also could kill bacteria and suppress keratitis. A phage multiplicity of infection of 100 times the host bacterial number could kill host bacteria effectively. A cocktail of two phages suppressed bacteria in the keratitis model mouse. These data demonstrated that the phages in this study could completely prevent the keratitis caused by P. aeruginosa in a keratitis mouse model. Furthermore, these results suggest that phage may be a more effective prophylaxis for horse keratitis than the current preventive use of antibiotics. Such treatment may reduce the use of antibiotics and therefore antibiotic resistance. Further studies are required to assess phage therapy as a candidate for treatment of horse keratitis. IMPORTANCE: Antibiotic-resistant bacteria are emerging all over the world. Bacteriophages have great potential for resolution of this problem. A bacteriophage, or phage, is a virus that infects bacteria specifically. As a novel therapeutic strategy against racehorse keratitis caused by Pseudomonas aeruginosa, we propose the application of phages for treatment. Phages isolated in this work had in vitro effectiveness for a broad range of P. aeruginosa strains. Indeed, a great reduction of bacterial proliferation was shown in phage therapy for mouse models of P. aeruginosa keratitis. Therefore, to reduce antibiotic usage, phage therapy should be investigated and developed further.

Predictors of fatal outcomes resulting from acute Escherichia coli mastitis in dairy cows.

To evaluate the prognostic criteria for identifying cows at an increased risk of a fatal outcome from acute Escherichia coli mastitis, the potential cut-off values for five diagnostic parameters associated with a high mortality were determined by receiver operator characteristic curve analysis. These criteria were hematocrit value >32%, blood non-esterified fatty acid concentration >0.4 mEq/l, antithrombin activity <120%, platelet count <15 × 10(4)/ml and presence of dysstasia. Exceeding the cut-off values for at least three parameters on day 2 after onset predicted fatality (predictive value 87.5). When these prognostic criteria were applied to 34 clinical cases, cows that met three criteria were seven times more likely to die than cows that met fewer than three criteria.