Milk as diagnostic fluid for udder health management.

BACKGROUND: Mastitis is the major disease affecting milk production of dairy cattle, and milk is an obvious substrate for the detection of both the inflammation and its causative infectious agents at quarter, cow, or herd levels. In this review, we examine the use of milk to detect inflammation based on somatic cell count (SCC) and other biomarkers, and for the detection of mastitis pathogens through culture-based and culture-free methods. FINDINGS: The use of SCC at a cow or bulk milk level to guide udder health management in lactation is well-established, and SCC is increasingly used to guide selective dry cow treatment. Other markers of inflammation include electrical conductivity, which is used commercially, and markers of disease severity such as acute phase proteins but are not pathogen-specific. Some pathogen-specific markers based on humoral immune responses are available, but their value in udder health management is largely untested. Commercial pathogen detection is based on culture or polymerase chain reaction, with other tests, for example, loop-mediated isothermal amplification or 16S microbiome analysis still at the research or development stage. Matrix-assisted laser desorption ionisation time of flight (MALDI-ToF) is increasingly used for the identification of cultured organisms whilst application directly to milk needs further development. Details of test sensitivity, specificity, and use of the various technologies may differ between quarter, cow, and bulk milk applications. CONCLUSIONS: There is a growing array of diagnostic assays that can be used to detect markers of inflammation or infection in milk. The value of some of these methods in on-farm udder health improvement programs is yet to be demonstrated whilst methods with proven value may be underutilised.

Wooden hoof blocks: are we using the right wood?

AIMS: To investigate the association between the density of wooden hoof blocks and resistance to wear in pasture-based dairy herds, and to assess the density of commercially available wooden hoof blocks. METHODS: Three types of wooden hoof blocks with different densities (low, medium and high) were attached to 36 lactating dairy cows with parity ≤2 and sound locomotion (score ≤2 on a scale of 1-4). The height of wooden blocks was measured in three different regions, front, abaxial and caudal on Days 7, 11, 14, 18, 21, 25 and 28 after application. Due to the loss of low-density wooden blocks, the data for these blocks were analysed for only two measurements on Days 7 and 11. The data for medium and high-density wooden blocks were analysed from Days 7-25. A linear mixed model with repeated measures was used to analyse the repeated observations. Height, density and surface area of commercially available hoof blocks (n = 19) were measured and compared to the blocks used in this study. RESULTS: The magnitude of wear, in the front and the abaxial point of the blocks were greater in blocks made of low-density wood compared to those made of medium and high-density wood (p < 0.001). The amount of wear increased over time for all groups (p < 0.001). Wood density was negatively associated with wear and loss. Measurements of commercial wooden blocks revealed that the 13/19 (63%) had lower density and 12/19 (68%) less surface area than the wooden blocks with medium density used in this study. CONCLUSION: In this study, the density of the wood was significantly associated with the longevity of hoof blocks when applied to hooves of pasture-based dairy cows. CLINICAL RELEVANCE: The longevity of the wooden hoof blocks applied to treat lame cows plays a significant role in the healing of the claw horn lesions. The density of a wooden hoof block affects the rate of wear of the block, and this should be considered by manufacturers and those treating lame cows.

The effect of participation in a regular reproductive herd health program on 80-day submission rate and 100-day in-calf rate in year-round calving dairy farms.

Herd health management programs are commonly implemented on dairy farms to improve farm reproductive performance. In year-round calving systems, herd health programs can be supported by extension programs such as InCalf, which focus on improving the 100-day in-calf rate (100DICR) as a key reproductive performance indicator. The effect of implementing a herd health program was evaluated by analysing the 80-day submission rate (80DSR) and 100DICR during the initial 18-month period following commencement of regular herd health visits at 11 year-round calving dairy farms. The mean 80DSR and 100DICR was calculated for each of six consecutive three-month intervals following the initial herd health visit at 11 dairy farms distributed across the north and south coasts of New South Wales (NSW). On the 11 study farms 80DSR increased from 36.9% (95% confidence interval [CI] 36.7-37.2) at 0-3 months to 61.9% (95% CI 61.4-62.4) and 100DICR increased from 19.1% (95% CI 18.9-19.3) at 0-3 months to 28.5% (95% CI 28.0-29.0) at 15-18 months. 80DSR increased at similar rates on north coast farms compared to south coast farms; however, north coast farms had a lower 100DICR at 0-3 months which increased at a greater rate compared to south coast farms. 100DICR on north coast farms increased from 12.8% (95% CI 12.5-13.2) at 0-3 months to 27.6% (95% CI 26.8-28.4) at 15-18 months, whereas 100DICR on south coast farms increased from 26.7% (95% CI 26.3-27.0) at 0-3 months to 29.6% (95% CI 28.8-30.5) at 15-18 months. Participation in a regular herd health program resulted in an increased 80DSR and 100DICR.

Mycoplasma species in vaginas of dairy cows before and after exposure to bulls and their association with conception.

Mycoplasma species can colonize the urogenital tract of dairy cattle. However, interrelationships between Mycoplasma spp. and reproductive performance in dairy herds are unclear. In this study, we measured apparent prevalences of Mycoplasma spp. in the vaginas of dairy cows (n = 629) pre- and post-bull exposure in dairy herds with and without Mycoplasma bovis clinical disease (n = 5 herds), and assessed associations between variables describing reproductive performance and consequent Mycoplasma spp. isolation. Mycoplasma spp. were infrequently isolated from the vagina pre- (1.9%; 12/629) and post-bull (3.2%; 20/629) exposure. Of the mycoplasmas isolated, Mycoplasma bovigenitalium was isolated most frequently (87.5%; 28/32), followed by Mycoplasma californicum (9.3%; 3/32). Mycoplasma bovis was only isolated from one cow. We were unable to provide any evidence of venereal transmission of M. bovis in cows in M. bovis-infected herds that use natural service bulls. There was an insufficient number of cows with Mycoplasma spp. in the vagina pre-bull exposure to assess effects on subsequent reproductive performance. Cows that had not conceived before post-bull exposure sampling had much greater odds (odds ratio 14.8; 95% confidence interval 4.2 to 52.3) of having a Mycoplasma sp. isolated from the vagina at this time compared with those that had conceived. Also, within those that had conceived, delayed conception increased the odds of having a Mycoplasma spp. isolated from the vagina at the post-bull exposure sampling by a factor of 1.62 for every additional week not pregnant. The likely cause of these findings is that cows that remain not pregnant for longer are more likely to be served by a bull (likely repeatedly) and subsequently become colonized with a Mycoplasma sp. (mostly M. bovigenitalium) through venereal transmission. In dairy herds that use bulls, there is a greater chance of isolating a Mycoplasma sp. (mostly M. bovigenitalium) after a period of bull breedings from the vaginas of cows that have remained nonpregnant for longer during the bull breeding period.

Mycoplasma bovis and other Mollicutes in replacement dairy heifers from Mycoplasma bovis-infected and uninfected herds: A 2-year longitudinal study.

Replacement dairy heifers exposed to Mycoplasma bovis as calves may be at risk of future clinical disease and pathogen transmission, both within and between herds; however, little information is available about these risks. We conducted a 2-yr longitudinal (panel) study starting with 450 heifer calves reared to weaning in 8 herds (7 M. bovis infected with clinical disease, 1 uninfected) under the same ownership. After weaning, heifers were commingled and managed with non-study heifers at a single heifer rearing facility. Nose, conjunctival, and vaginal swabs were collected along with a blood sample at weaning, prebreeding, precalving, and approximately 1 mo postcalving. Additionally, a colostrum sample was collected upon calving and a composite milk sample was collected 1 mo postcalving. The swabs, colostrum, and milk samples were cultured for Mycoplasma spp., and serum from the blood was evaluated for serological evidence of exposure to M. bovis using an ELISA. Despite a high M. bovis ELISA seroprevalence at weaning in the heifers from the 7 M. bovis-infected herds with clinical disease [72% (289/400); range by herd: 28-98%], M. bovis was isolated from only 4% (16/400) of the same heifers at the same time. In heifers from the uninfected herd at weaning, M. bovis seroprevalence was 2% (1/50) and M. bovis was not detected by culture. Mycoplasma bovis was isolated from 0.5% (2/414) of heifers at prebreeding, 0% (0/374) of heifers at precalving, and 0.3% (1/356) of heifers 1 mo postcalving. The nose was the predominant anatomical site of M. bovis colonization (74%; 14/19 culture positives). A single heifer (from an M. bovis-infected herd with clinical disease) was repeatedly detected with M. bovis in its nose at weaning, prebreeding, and postcalving samplings. This demonstrates the possibility, albeit rare, of a long-term M. bovis carrier state in replacement heifers exposed to M. bovis as calves, up to at least 1 mo after entry into the milking herd. No M. bovis clinical disease was detected in any heifer from weaning through to the end of the study (approximately 1 mo after calving). Acholeplasma spp. were commonly isolated throughout the study. Mycoplasma bovigenitalium, Mycoplasma bovoculi, and Mycoplasma bovirhinis were isolated infrequently. Mycoplasma bovis seroprevalences at prebreeding, precalving, and postcalving samplings were 27% (112/414), 12% (46/374), and 18% (65/356), respectively. Overall, the results show that replacement heifers from groups exposed to M. bovis preweaning can become colonized with M. bovis and that colonization can, uncommonly, be present after their first calving. For groups of 50 or more heifers exposed to M. bovis preweaning, there is at least a nontrivial probability that the group will contain at least 1 shedding heifer postcalving.

Whole dairy herd sampling to detect subclinical intramammary Mycoplasma bovis infection after clinical mastitis outbreaks.

After clinical Mycoplasma bovis mastitis outbreaks in dairy herds, M. bovis can persist as subclinical intramammary infections. Identification and culling of sub-clinically infected cows may be warranted to reduce future pathogen transmission and disease. In this study, apparent cow-level prevalences of M. bovis intramammary infection within 4 milking herds immediately following outbreaks of clinical disease due to M. bovis were determined utilising PCR and culture. All clinically affected M. bovis cows had been culled from the herds prior to herd sampling. Composite milk samples were collected once from each cow (n = 2,258) using a routine milk recording sampling technique. These samples were pooled for PCR screening; positive pools were analysed in different sized pools as needed from large to small, until individual PCR-positive animals could be identified. Despite M. bovis seroprevalences of 76% (herd 1), 40% (herd 2), 20% (herd 3) and 16% (herd 4), apparent prevalences of intramammary infection in the main milking group based on PCR in herds 1 to 4 were 0.2% (1/497), 0.0% (0/475), 0.1% (1/816) and 0.0% (0/444), respectively. Due to the low apparent prevalences of subclinical intramammary mycoplasma infections in these herds and the high expense associated with milk sample collection and testing, the return on diagnostic investment was very limited, particularly considering that additional cows are likely to have been colonised with mycoplasma in other anatomical sites. The results of this study suggest that pursuing identification of cows with subclinical intramammary mycoplasma infections following resolution of clinical M. bovis disease outbreaks in dairy herds may be of minimal benefit in programs designed to control or eradicate M. bovis.

Farmers' perceptions and approaches to detection, treatment and prevention of lameness in pasture-based dairy herds in New South Wales, Australia.

AIMS: The objective of this study was to investigate farmers' perception of lameness in comparison to the estimated prevalence of lameness in NSW pasture-based dairies to evaluate farmers' perceptions and approaches to detection, treatment and prevention of lameness. METHODS: Across-sectional study was conducted on 62 pasture-based dairy farms across NSW, Australia. The prevalence of lameness in these farms was estimated using locomotion scoring (1-4 scale). A survey was also conducted, using a questionnaire and face-to-face interview, to explore farmers' perceived prevalence of lameness and approaches to treatment and prevention. RESULTS: The prevalence of lameness estimated by farmers was 3.7 times less (mean: 5%; range 0% to 26%) than that determined by locomotion scoring (mean: 19.1%; range 5.0%-44.5%). Approaches to treatment included antimicrobial therapy, hoof inspection with or without application of wooden blocks. In 28% of the farms, the lame cows were managed by farmers or farm staff with no official training in treatment of lame cows. The mean interval from detection of lameness to examination of the affected hoof was almost 55 hours (range 2-720 hours). A very low percentage of farms kept lameness records or implemented lameness preventive strategies such as footbaths and prophylactic foot trimming. CONCLUSIONS: Farmers and farm managers were found to underestimate the prevalence of lameness which could be due to the low level of awareness and can contribute to subsequent lack of implementation of prophylactic procedures and preventive management strategies for lameness. These findings accentuate the need to improve farmers' ability to detect lame cows and to emphasise the importance of recording in order to facilitate the management of lameness in dairy herds.

Isolation of Mycoplasma spp. and serological responses in bulls prior to and following their introduction into Mycoplasma bovis-infected dairy herds.

With the common use of bulls for breeding following a period of artificial insemination in seasonally bred dairy herds, it is important to consider the potential role of the bull in transmission of Mycoplasma spp. within and between herds. This study aimed to assess the prevalence of Mycoplasma spp. in a population of bulls before and after use in Mycoplasma bovis-infected herds. The frequency of subclinical infection was also measured serologically postbreeding, and the association of Mycoplasma spp. on semen quality was evaluated. Mycoplasma bovis was isolated from 4 of 118 bulls after use in 4 herds infected with M. bovis. In the bulls, M. bovis seroprevalence increased from 9% prebreeding to 46% postbreeding with a total seroconversion rate of 44% across the 4 herds, with no evidence of clinical disease. There was no association of Mycoplasma spp. in the bulls' semen and abnormal palpation characteristics (enlarged or nodular) of seminal vesicular glands or poor semen quality attributes such as semen mass activity, sperm motility, and morphology. These results demonstrate a high degree of subclinical exposure of the bulls to M. bovis in infected herds and highlight the potential for bulls to be mycoplasma carriers within and between herds. Herd biosecurity protocols and control programs should take into account the potential role of bulls in the introduction and spread of Mycoplasma spp.

Short communication: Shedding of Mycoplasma bovis and antibody responses in cows recently diagnosed with clinical infection.

Mycoplasma bovis can have significant consequences when introduced into immunologically naïve dairy herds. Subclinically infected carrier animals are the most common way that M. bovis is introduced into herds. Although M. bovis udder infections can be detected by milk sampling lactating animals before their introduction, currently, no definitive way of identifying M. bovis carrier animals that are nonlactating (i.e., calves, heifers, dry cows, or bulls) is available. Understanding the prevalence of M. bovis shedding from various body sites in clinically infected animals could inform strategies for the detection of subclinical infection in nonlactating stock. The mucosal surfaces of the nose, eye, and vagina of 16 cows with recent clinical mastitis caused by M. bovis were examined for the presence of M. bovis shedding. Blood was collected for serological evaluation by a commercially available ELISA. Mycoplasma bovis was isolated from the vagina of only 3 (18.8%) of the cows and was not detected from the noses or eyes of any of the cows. Fifteen of the 16 (93.8%) cows were seropositive to the ELISA. With such low prevalence of detection of M. bovis from the vagina and no detections from the noses or eyes of recently clinically infected animals, it is very likely that sampling these sites would be ineffective for detecting subclinical infection in cattle. Serology using the ELISA may have some use when screening animals for biosecurity risk assessment. However, more information regarding time to seroconversion, antibody longevity, and test diagnostic sensitivity and specificity are required to define the appropriate use of this ELISA for biosecurity purposes.

Bulk tank milk antibody ELISA as a biosecurity tool for detecting dairy herds with past exposure to Mycoplasma bovis.

In Australia, one of the biosecurity recommendations to help prevent the introduction of Mycoplasma bovis into a dairy herd is to use a PCR assay on bulk tank milk (BTM) samples to evaluate the M. bovis infection status of potential source herds. An alternative approach is to assess the immunological status of the herd with respect to previous exposure to M. bovis via the use of an ELISA that is commercially available for use on cattle milk and serum. The objectives of this study were to (1) evaluate factors potentially associated with variation in the ELISA BTM optical density coefficient (ODC%) in previously exposed herds, (2) evaluate the association between the proportion of cows that are ELISA positive and the BTM ELISA ODC%, (3) assess agreement between the BTM ELISA and PCR and culture, and (4) compare BTM ELISA ODC% between the "hospital" herd and the main lactating herd on the same farm. Bulk tank milk samples (n = 192) were collected from 19 dairy herds with a history of clinical M. bovis disease and from 6 control herds (herds with no known clinical cases of mycoplasmosis). For 28 of the BTM samples collected, blood was also collected from 50 lactating cows contributing to that bulk tank sample. From 1 herd, concurrent paired BTM samples were collected from the main herd and the hospital herd on 16 occasions. All BTM samples were analyzed by ELISA (Bio-X Bio K 302, Bio-X Diagnostics, Rochefort, Belgium), PCR, and culture. The BTM ELISA ODC% was associated with time since initial M. bovis outbreak and time since the start of the herd's calving period. Following an initial outbreak of M. bovis, the BTM ELISA ODC% was highest in the first 8 mo. In split- and seasonal-calving herds, significantly higher BTM ELISA ODC% results were observed 5 to 8 wk after the commencement of the calving period. A significant association was observed between the within-herd seroprevalence for the lactating herd and BTM ELISA ODC%, but within-herd seroprevalence explained little of the variation in BTM ELISA ODC%. When comparing the BTM ELISA with a multiplex probe PCR and culture followed by 16S to 23S rRNA sequencing, there was virtually no agreement above that expected by chance; prevalence-adjusted bias-adjusted kappa values were 0.22 and 0.25 for ELISA category versus PCR category and culture, respectively. Finally, the hospital herd BTM ELISA ODC% mirrored that for the main herd BTM but was significantly higher. This study demonstrates that this commercially available ELISA used on BTM samples may complement the use of BTM PCR or culture in identifying herds from which purchase of animals may pose a higher biosecurity risk for introduction of M. bovis into noninfected herds.

Effect of insemination site and diameter of the pre-ovulatory follicle on the odds of pregnancy in heifers using sexed or non-sexed semen.

OBJECTIVE: To determine if insemination site or pre-ovulatory follicle diameter at fixed-time artificial insemination (FTAI) affects the odds of pregnancy when heifers are inseminated with sexed semen. METHODS: The study was conducted in 422 Holstein heifers enrolled into 531 inseminations. Inseminations were randomly allocated to 1 of 16 treatment combinations involving three variables: semen type (sexed vs non-sexed), insemination site (uterine horn vs uterine body) and one of four sires. Ovaries were examined by transrectal ultrasound prior to FTAI to determine the follicle diameter and location. AI technician, times bred, age, weight and temperature-humidity index were also recorded. Pregnancy diagnosis was conducted 29 days post-insemination. Follicle diameter and body weight were categorised according to arbitrary cut-points. Each variable was analysed by logistic regression to determine the effect on pregnancy per AI and compare between sexed and non-sexed inseminations. RESULTS: Insemination site did not affect pregnancy per AI for either sexed (P = 0.528) or non-sexed (P = 0.886) inseminations. Heifers with an 18-22 mm follicle had better odds of pregnancy than heifers that did not (odds ratio (OR) 1.45, 95% confidence interval (CI) 1.004-2.09), although no effect was detected for only sexed or only non-sexed inseminations. Heifers weighing 310-370 kg had a higher pregnancy per AI than heifers weighing > 370 kg for non-sexed inseminations (P = 0.004) and sexed semen from sire 4 caused lower odds of pregnancy than semen from sire 1 (OR 0.40, 95% CI 0.18-0.89). CONCLUSION: Insemination site did not affect pregnancy per AI, but heifers with an 18-22 mm pre-ovulatory follicle at insemination had better odds of pregnancy for both sexed and non-sexed inseminations.

Demographics, farm and reproductive management strategies used in Australian automatic milking systems compared with regionally proximal conventional milking systems.

OBJECTIVE: To determine the management practices utilised in automatic milking systems (AMS) that affect reproductive management and performance and how these compare with the management practices used in regionally proximal conventional milking systems (CMS). METHODS: This study examined demographic and management data from AMS and CMS dairy farms through a survey, with a specific focus on reproductive management procedures. RESULTS: Overall, responses from AMS and CMS dairy farms showed little difference in terms of respondent demographics, farm size, herd structure and most farm management strategies. AMS dairies were more likely to use activity meters or other electronic oestrus detection aids than CMS dairies (P < 0.001) and were also more likely to have changed to electronic recording systems (P = 0.007). Although many respondents indicated that they used key monitoring parameters to assess reproductive performance (e.g. days in milk, conception vs pregnancy rate etc.), the format of responses varied significantly, indicating a relatively widespread (among the respondents) lack of knowledge regarding the meaning and usage of some of these common parameters/terminology. CONCLUSIONS: Ultimately, reproductive management practices of AMS dairies were largely similar to those of CMS dairies, indicating that such practices can be implemented in a practical sense, even though the resultant reproductive performance is not yet understood. Understanding that the key reproductive management strategies do not need to change vastly is important to ensure that new adoptees are well informed. Further work is needed to objectively measure AMS performance to increase the knowledge base and generate the confidence that will facilitate further adoption of this innovation.

Effects of ovarian structures identified at Ovsynch™ enrolment, disease history and lactation variables on odds of pregnancy to a fixed-time artificial insemination after Ovsynch™.

OBJECTIVE: To determine the effects of ovarian structures present at Ovsynch™ enrolment, disease history, lactation variables and times bred on pregnancy per fixed-time artificial insemination (FTAI) following fixed-time insemination after Ovsynch and to determine the relationship between the size of ovarian follicular structures and concurrent presence of a corpus luteum (CL). METHODS: The study was conducted in a 3000-cow Holstein dairy herd. Over a 6-month period, 886 non-pregnant cows were examined by transrectal ultrasound and enrolled into 1132 Ovsynch events. Enrolled cows were synchronised using an Ovsynch-56 protocol. At enrolment, both ovaries were scanned by transrectal ultrasound to identify and record the presence and size of ovarian structures. Cows that did not return to oestrus were pregnancy tested 32-38 days post-insemination. Univariable and multivariable generalised linear mixed models (GLMMs) were used to determine the effect of each variable on pregnancy per FTAI. A univariable GLMM was also used to analyse the relationship between size of the dominant follicular structure and concurrent presence of a CL. RESULTS: CL size (P = 0.039) and presence of a luteal cyst (P = 0.002) at Ovsynch enrolment significantly increased the odds of pregnancy. Occurrence of lameness (P = 0.035) or mastitis (P = 0.008) between calving and enrolment significantly decreased the odds of pregnancy. Neither the presence nor size of a follicular structure significantly affected the odds of pregnancy, although cows with larger follicular structures were less likely to have a concurrent CL (P < 0.001). CONCLUSION: Presence of luteal tissue at Ovsynch enrolment and recent disease events affected pregnancy per FTAI in cows inseminated after Ovsynch.

Evaluation of a cow-side milk progesterone assay and assessment of the positive predictive value of oestrus diagnosis by dairy farmers in New South Wales.

OBJECTIVES: The three objectives of this study were to determine the positive predictive value (PPV) of oestrus diagnosis (heat detection accuracy) by dairy farmers, calculate the diagnostic sensitivity and specificity of the P4 Rapid milk progesterone assay for detecting a corpus luteum and evaluate the economics of using a cow-side milk progesterone assay designed to aid oestrus diagnosis. METHODS: Milk samples were collected from 752 cows diagnosed in oestrus by farm personnel on 14 dairy farms. Samples were tested using the P4 Rapid milk progesterone assay to estimate the PPV of oestrus diagnosis at each farm and a crude pooled mean of PPV of oestrus diagnosis across all farms. A further 156 milk samples were collected from cows with luteal tissue status determined by transrectal ultrasound and tested by the P4 Rapid assay to enable calculation of the sensitivity and specificity of the P4 Rapid assay. RESULTS: For pooled farm samples, the PPV was 97.0%, with a range between farms of 88.9-100%. Sensitivity of the P4 Rapid milk progesterone assay for detecting a corpus luteum was 90.1% and specificity was 98.7%. Misclassification of oestrus in cows previously identified as pregnant was the most common cause of false-positive oestrus diagnoses by farm personnel. CONCLUSION: Routine testing of milk progesterone in all cows diagnosed in oestrus is not economically justified and may even slightly reduce submission rates; conversely, strategic use of cow-side milk progesterone assays can improve herd reproductive performance by facilitating decisions on whether to rebreed cows previously diagnosed as pregnant.

Milk acidification to control the growth of Mycoplasma bovis and Salmonella Dublin in contaminated milk.

Bacterial contamination of milk fed to calves compromises calf health. Several bacterial pathogens that infect cows, including Mycoplasma bovis and Salmonella enterica ssp. enterica serovar Dublin, are shed in milk, providing a possible route of transmission to calves. Milk acidification lowers the milk pH so that it is unsuitable for bacterial growth and survival. The objectives of this study were to (1) determine the growth of M. bovis and Salmonella Dublin in milk, and (2) evaluate the efficacy of milk acidification using a commercially available acidification agent (Salstop, Impextraco, Heist-op-den-Berg, Belgium) to control M. bovis and Salmonella Dublin survival in milk. For the first objective, 3 treatments and a positive control were prepared in 10 mL of milk and broth, respectively, and inoculated with M. bovis or Salmonella Dublin to an approximate concentration of 104 cfu/mL. Each treatment was retained at 5, 23, or 37°C with the positive control at 37°C. Aliquots were taken at 4, 8, 24, 28, 32, 48, 52, and 56 h after inoculation and transferred onto agar medium in triplicate following a 10-fold dilution series in sterile phosphate-buffered saline. All plates were incubated and colonies counted. For the second objective, 4 treatments and a positive control were prepared with 100 mL of milk and inoculated with M. bovis or Salmonella Dublin to an approximate concentration of 106 cfu/mL. With the use of Salstop, treatments were adjusted to an approximate pH of 6, 5, 4, or 3.5. The positive control was left untreated. At 1, 2, 4, 6, 8, and 24 h after treatment, triplicate aliquots were taken, the pH measured, and then the aliquots were transferred onto agar medium and into broth for enrichment. Following incubation, agar colonies were counted, while broths were plated and incubated prior to colonies being counted. All trials were repeated. Mycoplasma bovis did not grow in milk, but Salmonella Dublin proliferated. The pH of all acidification treatments remained stable for 24 h. No viable M. bovis organisms were detected at 1 h of exposure to pH 3.5 and 4 or at 8 h of exposure to pH 5. Following 24 h of exposure to pH 6 M. bovis remained viable. No viable Salmonella Dublin organisms were detected at 2 and 6 h of exposure to pH 3.5 and 4, respectively. Salmonella Dublin remained viable following 24 h of exposure to pH 5 and 6. These results demonstrate that milk acidification using Salstop is effective at eliminating viable M. bovis and Salmonella Dublin organisms in milk if the appropriate pH and exposure time are maintained.

Identifying risk factors associated with lameness in pasture-based dairy herds.

Lameness is a significant welfare concern for dairy farmers and a major contributing economic loss to the dairy industry. Information is limited on environmental and managerial risk factors associated with lameness in Australian dairy herds. The objective of this study was to explore and quantify the environmental and management risk factors associated with lameness in pasture-based dairy herds. A cross-sectional study was conducted in 63 pasture-based dairy herds between 2011 and 2014, where all lactating cows were locomotion scored (scale 1-4) during a single visit. Environmental and management variables, such as length of main track and animal handling practices, were recorded during the visit. The prevalence of lameness was measured for each farm and associated risk factors were analyzed using a Generalized Linear Model, where farm was the unit of analysis. Estimated average prevalence of lameness was 18.9% (range 5 to 44.5%). The prevalence of lameness was associated with the amount of rainfall during the 30 d before the farm assessment, smoothness of concrete surface and available space per cow in the holding yard, and length of feed-pad available per cow. Inappropriate handling of cows on the track (e.g., causing sideways pushing among cows) was also a contributing risk factor to high prevalence of lameness in these dairy herds. The findings of this study suggest that by managing several environmental and farming practices, producers can reduce the prevalence of lameness, leading to improved productivity of their herds.

Artificial insemination field data on the use of sexed and conventional semen in nulliparous Holstein heifers.

This study investigated conception rates and other reproductive outcomes achieved with artificial insemination (AI) of nulliparous Holstein heifers using sexed and conventional semen in a commercial Australian dairy herd in central western New South Wales from January 2004 to April 2009. Retrospective data from on-farm records of 9,870 inseminations of 4,456 heifers were analyzed using several mixed models to assess the effect of temperature and humidity surrounding breeding, insemination sire, artificial insemination technician, service number, and heifer weight and age at breeding on reproductive traits (conception rates, sex ratios, gestation length, and abortion and stillbirth rates). Semen was used from 15 sexed sires and 41 unsexed sires. Sexed semen was primarily used at first and second service. Empirical conception rates of 31.6 and 39.6% were achieved for sexed and unsexed semen respectively, whereas model-based predictions were lower, at 21.3 and 32.1%. Conception rates were significantly affected by insemination sire, sex-sorting, heifer age at breeding, temperature and humidity surrounding breeding, service number, and AI technician. Sexed semen yielded 86% heifers, compared with 48% for conventional semen. Significant predictors of calf sex included semen sexing, gestation length, and insemination sire. Twinning rate was high, at 3.6% for both semen types, and gestation length and heifer weight at breeding were significant predictors of twinning. Abortion rates for sexed and unsexed conceptions were similar at 6.1 and 6.5%, respectively, and were affected by heifer age at breeding. Stillbirth rate was affected by calf sex, twinning, gestation length, and AI technician; semen sorting, age at breeding, and temperature and humidity were marginally significant predictors. No abnormalities were observed in the development of offspring, except for a marginally higher stillbirth rate for sexed calves, a finding that needs further investigation. Many variables influence the breeding outcomes associated with the application of sex-sorted sperm on commercial dairy farms. Recognition and management of these variables will increase the economic return from the investment in sex-sorted sperm.

Comparison of three diagnostic techniques for detection of rotavirus and coronavirus in calf faeces in Australia.

OBJECTIVE: Compare real-time reverse transcription polymerase chain reaction (qRT-PCR), a commercially available enzyme-linked immunosorbent assay (ELISA) and lateral flow immunochromatography assay (LAT) for the detection of rotavirus and coronavirus in faecal samples collected from diarrhoeic calves. DESIGN: Prospective survey. METHOD: Samples were tested at two separate facilities using a commercial ELISA and an in-house qRT-PCR. Simple logistic regression was performed to examine the relationship between the two tests. A subset of samples was screened using qRT-PCR, ELISA and a commercial LAT dipstick (132 faecal samples were tested for coronavirus and 122 samples for rotavirus). RESULTS: Of the 586 samples tested, 131 (22.39%) and 468 (79.86%) were positive for coronavirus and group A rotavirus, respectively, using qRT-PCR. The number of samples positive on ELISA for coronavirus and rotavirus was 73 (12.46%) and 225 (38.40%), respectively. Using LAT, 30 (22.73%) and 43 (35.35%) samples were positive for coronavirus and rotavirus, respectively. Simple linear regression revealed a statistically significant (P < 0.05) but weak (r(2) =-0.07 and -0.40) correlation between the rotavirus/coronavirus qRT-PCR and ELISA, respectively. There was also poor agreement between the LAT and qRT-PCR assays. CONCLUSION: The sensitivity and specificity of the commercial ELISA and LAT assays evaluated in this study were low compared with qRT-PCR. The low positive and negative predictive values of the assays suggests that they were of limited diagnostic benefit in the population sampled.

Development of a novel in-water vaccination protocol for DNA adenine methylase deficient Salmonella enterica serovar Typhimurium vaccine in adult sheep.

Intensive livestock production is associated with an increased incidence of salmonellosis. The risk of infection and the subsequent public health concern is attributed to increased pathogen exposure and disease susceptibility due to multiple stressors experienced by livestock from farm to feedlot. Traditional parenteral vaccine methods can further stress susceptible populations and cause carcass damage, adverse reactions, and resultant increased production costs. As a potential means to address these issues, in-water delivery of live attenuated vaccines affords a low cost, low-stress method for immunization of livestock populations that is not associated with the adverse handling stressors and injection reactions associated with parenteral administration. We have previously established that in-water administration of a Salmonella enterica serovar Typhimurium dam vaccine conferred significant protection in livestock. While these experimental trials hold significant promise, the ultimate measure of the vaccine will not be established until it has undergone clinical testing in the field wherein environmental and sanitary conditions are variable. Here we show that in-water administration of a S. Typhimurium dam attenuated vaccine was safe, stable, and well-tolerated in adult sheep. The dam vaccine did not alter water consumption or vaccine dosing; remained viable under a wide range of temperatures (21-37°C); did not proliferate within fecal-contaminated trough water; and was associated with minimal fecal shedding and clinical disease as a consequence of vaccination. The capacity of Salmonella dam attenuated vaccines to be delivered in drinking water to protect livestock from virulent Salmonella challenge offers an effective, economical, stressor-free Salmonella prophylaxis for intensive livestock production systems.