Machine learning modeling to predict causes of infectious abortions and perinatal mortalities in cattle.

A plethora of infectious and non-infectious causes of bovine abortions and perinatal mortalities (APM) have been reported in literature. However, due to financial limitations or a potential zoonotic impact, many laboratories only offer a standard analytical panel, limited to a preestablished number of pathogens. To improve the cost-efficiency of laboratory diagnostics, it could be beneficial to design a targeted analytical approach for APM cases, based on maternal and environmental characteristics associated with the prevalence of specific abortifacient pathogens. The objective of this retrospective observational study was to implement a machine learning pipeline (MLP) to predict maternal and environmental factors associated with infectious APM. Our MLP based on a greedy ensemble approach incorporated a standard tuning grid of four models, applied on a dataset of 1590 APM cases with a positive diagnosis that was achieved by analyzing an extensive set of abortifacient pathogens. Production type (dairy/beef), gestation length, and season were successfully predicted by the greedy ensemble, with a modest prediction capacity which ranged between 63 and 73 %. Besides the predictive accuracy of individual variables, our MLP hierarchically identified predictor importance causes of associated environmental/maternal characteristics of APM. For instance, in APM cases that happened in beef cows, season at APM (spring/summer) was the most important predictor with a relative importance of 24 %. Furthermore, at the last trimester of gestation Trueperella pyogenes and Neospora caninum were the most important predictors of APM with a relative importance of 22 and 17 %, respectively. Interestingly, herd size came out as the most relevant predictor for APM in multiparous dams, with a relative importance of 12 %. Based on these and other mix of predicted environmental/maternal and pathogenic potential causes, it could be concluded that implementing our MLP may be beneficial to design a more cost-effective, case-specific diagnostic approach for bovine APM cases at the diagnostic laboratory level.

Milk yield residuals and their link with the metabolic status of dairy cows in the transition period.

The transition period is one of the most challenging periods in the lactation cycle of high-yielding dairy cows. It is commonly known to be associated with diminished animal welfare and economic performance of dairy farms. The development of data-driven health monitoring tools based on on-farm available milk yield development has shown potential in identifying health-perturbing events. As proof of principle, we explored the association of these milk yield residuals with the metabolic status of cows during the transition period. Over 2 yr, 117 transition periods from 99 multiparous Holstein-Friesian cows were monitored intensively. Pre- and postpartum dry matter intake was measured and blood samples were taken at regular intervals to determine ß-hydroxybutyrate, nonesterified fatty acids (NEFA), insulin, glucose, fructosamine, and IGF1 concentrations. The expected milk yield in the current transition period was predicted with 2 previously developed models (nextMILK and SLMYP) using low-frequency test-day (TD) data and high-frequency milk meter (MM) data from the animal's previous lactation, respectively. The expected milk yield was subtracted from the actual production to calculate the milk yield residuals in the transition period (MRT) for both TD and MM data, yielding MRTTD and MRTMM. When the MRT is negative, the realized milk yield is lower than the predicted milk yield, in contrast, when positive, the realized milk yield exceeded the predicted milk yield. First, blood plasma analytes, dry matter intake, and MRT were compared between clinically diseased and nonclinically diseased transitions. MRTTD and MRTMM, postpartum dry matter intake and IGF1 were significantly lower for clinically diseased versus nonclinically diseased transitions, whereas ß-hydroxybutyrate and NEFA concentrations were significantly higher. Next, linear models were used to link the MRTTD and MRTMM of the nonclinically diseased cows with the dry matter intake measurements and blood plasma analytes. After variable selection, a final model was constructed for MRTTD and MRTMM, resulting in an adjusted R2 of 0.47 and 0.73, respectively. While both final models were not identical the retained variables were similar and yielded comparable importance and direction. In summary, the most informative variables in these linear models were the dry matter intake postpartum and the lactation number. Moreover, in both models, lower and thus also more negative MRT were linked with lower dry matter intake and increasing lactation number. In the case of an increasing dry matter intake, MRTTD was positively associated with NEFA concentrations. Furthermore, IGF1, glucose, and insulin explained a significant part of the MRT. Results of the present study suggest that milk yield residuals at the start of a new lactation are indicative of the health and metabolic status of transitioning dairy cows in support of the development of a health monitoring tool. Future field studies including a higher number of cows from multiple herds are needed to validate these findings.

Enhancing bovine abortion surveillance: A learning experience.

Abortions and perinatal mortalities (APM) substantially affect cattle industry efficiency. Various infectious and noninfectious factors have been associated with bovine APM worldwide. Infections are often considered pivotal due to their abortifacient potential, leading laboratories to primarily investigate relevant infectious agents for APM cases. Some infectious causes, such as Brucella abortus, have also a zoonotic impact, necessitating monitoring for both animal and human health. However, underreporting of bovine APM is a global issue, affecting early detection of infectious and zoonotic causes. Previous studies identified factors influencing case submission, but regional characteristics may affect results. In Belgium, farmers are obliged to report cases of APM within the context of a national brucellosis monitoring program. The inclusion criteria for this monitoring program cover abortions (gestation length of 42-260 d) and perinatal mortalities of (pre)mature calves following a gestation length of more than 260 d, which were stillborn or died within 48 h after birth. The objective of the present study was to describe the evolution in submission of APM cases within a mandatory abortion monitoring program in relation to subsidized initiatives in the northern part of Belgium. Based on the proportion of APM submissions versus the proportion of bovine reproductive females, an APM proportion (APMPR) was calculated, and factors at both animal and herd level that may influence this APMPR were explored by using linear models. This evaluation revealed that the APMPR increased with the introduction of an extensive analytical panel of abortifacient agents and a free on-farm sample collection from 0.44% to 0.94%. Additionally, an increase of the APMPR was associated with an outbreak of an emerging abortifacient pathogen (Schmallenberg virus; 1.23%), and the introduction of a mandatory eradication program for bovine viral diarrhea virus (BVDv; 1.20%). The APMPR was higher in beef compared with dairy cattle, and it was higher in winter compared with fall, spring, and summer. Smaller herds categorized in the first quartile had a higher APMPR compared with larger herds. Herds that submitted an APM in the previous year had a higher APMPR in the next year compared with herds without an APM submission. Finally, herds for which there was evidence of the presence of BVDv had a higher APMPR compared with herds without evidence of the presence of BVDv. In conclusion, the number of APM submissions increased after the introduction of a free on-farm sample collection and an extensive pathogen screening panel. Production type (beef), season (winter), smaller herd size, previous APM, and presence of BVDv seemed to have a positive effect on APMPR. However, even under mandatory circumstances, APM still seems to be underreported, since the APMPR was lower than the expected minimal rate of 2%. Therefore, further research is necessary to identify the drivers that convince farmers to submit APM cases to improve submission rates and ensure an efficient monitoring program for APM and eventually associated zoonotic pathogens.

An in-depth investigation of the microbiota and its virulence factors associated with severe udder cleft dermatitis lesions.

Udder cleft dermatitis (UCD) is a skin condition affecting the anterior parts of the udder in dairy cattle. In the present study, we aimed to shed light on the microbiota in severe UCD lesions versus healthy udder skin by putting forward a taxonomic and functional profile based on a virulence factor analysis. Through shotgun metagenomic sequencing, we found a high proportion of bacteria in addition to a low abundance of archaea. A distinct clustering of healthy udder skin versus UCD lesion samples was shown by applying principal component analysis and (sparse) partial least squares analysis on the metagenomic data. Proteobacteria, Bacillota, and Actinomycetota were among the most abundant phyla in healthy udder skin samples. In UCD samples, Bacteroidota was the most abundant phylum. At genus level, Bifidobacterium spp. had the highest relative abundance in healthy skin samples, whereas Porphyromonas spp. and Corynebacterium spp. had the highest relative abundance in UCD samples. In the differential abundance analysis, Porphyromonas spp. and Bacteroides spp. were significantly differentially abundant in UCD samples, whereas Bifidobacterium spp., Staphylococcus sp. AntiMn-1, and Staphylococcus equorum were more commonly found in healthy samples. Moreover, the abundance of several treponeme phylotypes was significantly higher in lesion samples. The streptococcal cysteine protease speB was among the most abundant virulence factors present in severe UCD lesions, while a plethora of virulence factors such as the antitoxin relB were downregulated, possibly contributing to creating the ideal wound climate for the dysbiotic community. Network analysis showed healthy lesion samples had a large network ofpositive, correlations between the abundances of beneficial species such as Aerococcus urinaeequi and Bifidobacterium angulatum, indicating that the healthy skin microbiome forms an active protective bacterial network, which is disrupted in case of UCD. In UCD samples, a smaller microbial network mainly consisting of positive correlations between the abundances of Bacteroides fragilis and anaerobic Bacteroidota was exposed. Moreover, a high correlation between the taxonomic data and virulence factors was revealed, concurrently with 2 separate networks of microbes and virulence factors. One network, matching with the taxonomic findings in the healthy udder skin samples, showcased a community of harmless or beneficial bacteria, such as Bifidobacterium spp. and Butyrivibrio proteoclasticus, associated with hcnB, hcnC, relB, glyoxalase, and cupin 2. The other network, corresponding to UCD samples, consisted of pathogenic or facultative pathogenic and mainly anaerobic bacteria such as Treponema spp., Mycoplasmopsis spp., and bovine gammaherpesvirus 4, that correlated with virulence factors SpvB, fhaB, and haemagglutination activity domain-associated factor. Our results point toward a dysbiotic community with a notable decrease in diversity and evenness, with a loss of normal skin inhabitants and innocuous or useful species making way for predominantly anaerobic, facultative pathogens. The shift in the abundance of virulence factors such as fhaB and SpvB could play a role in the manifestation of a local micro-environment favorable to the microbiome associated with udder skin lesions. Lastly, the presence of specific networks between microbial species, and between microbes and virulence factors was shown.

The effect of maternal supply of rumen-protected protein to Holstein Friesian cows during the dry period on the transfer of passive immunity and colostral microbial composition.

The objective of this study was to analyze if maternal supply of rumen-protected protein during the dry period can affect the IgG concentration and microbial composition of colostrum and the IgG absorption and fecal microbial composition in the calf. Seventy-four multiparous Holstein Friesian (HF) dairy cows were stratified per parity and randomly assigned to one of 2 different dry period diets, a diet with a low crude protein (CP) level (LP) and a diet with a high CP level (HP) by addition of rumen-undegraded protein (RUP; formaldehyde-treated soybean meal, Mervobest, Nuscience, Drongen, Belgium). Colostrum was collected within 1 h after calving and IgG concentration was quantified by radial immunodiffusion analysis. Forty-nine calves (23 female and 26 male) were enrolled in the trial with a 2 × 2 factorial design, with prenatal and postnatal treatment as the 2 independent variables. This led to 4 experimental groups: LPLP, LPHP, HPLP, and HPHP, in which the first 2 letters refer to the prenatal treatment (diet of the dam) and the last 2 refer to the postnatal treatment (diet of the colostrum-producing cow). Calves received 3× 2 L of colostrum within 2, 6, and 24 h after birth. Meconium and feces were collected solely from female calves (n = 18) by digital palpation of the rectum, immediately after birth and before colostrum administration and at d 3 of age. Microbial DNA was extracted from meconium (n = 9), feces (n = 15), and colostrum (n = 49). Amplicon sequencing of the bacterial V3-V4 region of the 16S rRNA gene was performed for characterization of the bacterial communities. Colostrum IgG concentration was higher in cows that were supplemented with RUP, especially in cows entering their second lactation (LSM ± SEM 61.3 ± 2.3 vs. 55.2 ± 2.8 g of IgG/L). Calves born out of LP cows that received colostrum from HP cows (LPHP) had a lower serum IgG level compared with HPHP and LPLP calves (LSM ± SEM 14.2 ± 1.3 vs. 18.8 ± 1.2 and 20.9 ± 1.3 g of IgG/L in HPHP and LPLP, respectively). The most abundant phyla in colostrum were Proteobacteria (48.2%), Firmicutes (24.8%), Bacteroidetes (9.5%), and Actinobacteria (5.0%). The most abundant phyla in calf meconium and feces were Firmicutes (42.5 and 47.5%), Proteobacteria (21.7% and 33.7%), Bacteroidetes (16.8% and 15.7%), and Actinobacteria (2.9% and 3.1%). There was no difference in the overall microbial communities between colostrum from HP and LP cows. However, 2 genera (both members of the family Lachnospiraceae) were more abundant in colostrum from HP cows compared with LP cows. The microbial composition of meconium, feces and colostrum differed from each other. Fecal samples were more similar to each other and are characterized by a lower intersample diversity compared with colostrum and meconium samples. To conclude, increasing the CP level by addition of RUP in the dry period diet affected the colostrum IgG concentration and the transfer of passive immunity, but did not change the overall microbial composition of colostrum nor of meconium and feces in the calf.

Severe udder cleft dermatitis lesion transcriptomics points to an impaired skin barrier, defective wound repair and a dysregulated inflammatory response as key elements in the pathogenesis.

This study is the first to investigate the transcriptomic changes occurring in severe udder cleft dermatitis lesions (UCD) in Holstein-Friesian cows. An examination of the gene expression levels in natural UCD lesions and healthy udder skin through RNA Seq-Technology provided a deeper insight into the inflammatory pathways associated with this disease. A clear distinction between the gene expression patterns of UCD lesions and healthy skin was shown in the principal component analysis. Genes coding for inflammatory molecules were upregulated such as the chemokines C-X-C motif ligand 2 (CXCL2), 5 (CXCL5) and 8 (CXCL8), and C-C motif ligand 11 (CCL11). Moreover, the genes coding for the multifunctional molecules ADAM12 and SLPI were amongst the highest upregulated ones, whereas the most downregulated genes included the ones coding for keratins and keratin-associated molecules. Predominantly inflammatory pathways such as the chemokine signaling, cytokine receptor interaction and IL-17 signaling pathway were significantly upregulated in the pathway analysis. These results point towards a fulminant, dysregulated inflammatory response concomitant with a disruption of the skin barrier integrity and a hampered wound repair mechanism in severe UCD lesions.

Prediction of first test day milk yield using historical records in dairy cows.

The transition between two lactations remains one of the most critical periods during the productive life of dairy cows. In this study, we aimed to develop a model that predicts the milk yield of dairy cows from test day milk yield data collected in the previous lactation. In the past, data routinely collected in the context of herd improvement programmes on dairy farms have been used to provide insights in the health status of animals or for genetic evaluations. Typically, only data from the current lactation is used, comparing expected (i.e., unperturbed) with realised milk yields. This approach cannot be used to monitor the transition period due to the lack of unperturbed milk yields at the start of a lactation. For multiparous cows, an opportunity lies in the use of data from the previous lactation to predict the expected production of the next one. We developed a methodology to predict the first test day milk yield after calving using information from the previous lactation. To this end, three random forest models (nextMILKFULL, nextMILKPH, and nextMILKP) were trained with three different feature sets to forecast the milk yield on the first test day of the next lactation. To evaluate the added value of using a machine-learning approach against simple models based on contemporary animals or production in the previous lactation, we compared the nextMILK models with four benchmark models. The nextMILK models had an RMSE ranging from 6.08 to 6.24 kg of milk. In conclusion, the nextMILK models had a better prediction performance compared to the benchmark models. Application-wise, the proposed methodology could be part of a monitoring tool tailored towards the transition period. Future research should focus on validation of the developed methodology within such tool.

Unaffected semen quality parameters in Neospora caninum seropositive Belgian Blue bulls.

Neospora caninum is a protozoan parasite that causes abortion, perinatal mortality, and subfertility in cattle worldwide. Despite the presence of the DNA of the parasite in semen of infected bulls, the effect on semen quality has not been extensively studied. This study aimed to investigate the effect of a natural Neospora caninum infection on fresh and frozen semen quality parameters in Belgian Blue bulls. Two hundred and fourteen bulls were serologically screened with an indirect ELISA-test specific for anti-Neospora caninum antibodies, every two months during one year. In addition to serological screening, semen was collected twice weekly using an artificial vagina. The following semen quality parameters were assessed: ejaculate volume, concentration, total motility of fresh semen samples, as well as morphology, total and progressive motility for frozen/thawed semen samples. Bulls were semen sampled throughout the whole year, but only semen samples of bulls that had six consecutive positive or negative ELISA-test results were included in our dataset (n = 98). Generalized linear and binomial mixed models were used for statistical analysis of each outcome variable. In these models the explanatory variables were defined as: age, barn location, mean Temperature Humidity Index (THI) during sperm production (14-42 days before sampling), maximum daily THI at collection, season of sperm production, season at collection and the Neospora caninum antibody test results. Initially, individual explanatory variables were tested in univariable models for each outcome variable. Akaike information criterion (AIC) values were used to select explanatory variables to build a multivariable model, where the Neospora caninum test result was forced in all models. The present study reveals an overall apparent seroprevalence of Neospora caninum of 9,2% in the study population. No significant associations were detected between natural neosporosis, substantiated by ELISA-antibody levels, and any of our tested outcome variables on fresh and frozen/thawed semen samples. Based on the results of the present study, we conclude that Neospora caninum seropositive bulls do not have lower semen quality parameters compared with seronegative bulls.

Exploring the microbial composition of Holstein Friesian and Belgian Blue colostrum in relation to the transfer of passive immunity.

For centuries, multicellular organisms have lived in symbiosis with microorganisms. The interaction with microorganisms has been shown to be very beneficial for humans and animals. During a natural birth, the initial inoculation with bacteria occurs when the neonate passes through the birth canal. Colostrum and milk intake are associated with the acquisition of a healthy gut flora. However, little is known about the microbial composition of bovine colostrum and the possible beneficial effects for the neonatal calf. In this prospective cohort study, the microbial composition of first-milking colostrum was analyzed in 62 Holstein Friesian (HF) and 46 Belgian Blue (BB) cows by performing amplicon sequencing of the bacterial V3-V4 region of the 16S rRNA gene. Calves received, 3 times, 2 L of their dam's colostrum within 24 h after birth. Associations between colostral microbial composition and its IgG concentration, as well as each calf's serum IgG levels, were analyzed. Colostrum samples were dominated by the phyla Proteobacteria, Firmicutes, Bacteroidetes, and Actinobacteria. The 10 most abundant genera in the complete data set were Acinetobacter (16.2%), Pseudomonas (15.1%), a genus belonging to the Enterobacteriaceae family (4.9%), Lactococcus (4.0%), Chryseobacterium (3.9%), Staphylococcus (3.6%), Proteus (1.9%), Streptococcus (1.8%), Enterococcus (1.7%), and Enhydrobacter (1.5%). The remaining genera (other than these top 10) accounted for 36.5% of the counts, and another 8.7% were unidentified. Bacterial diversity differed significantly between HF and BB samples. Within each breed, several genera were found to be differentially abundant between colostrum of different quality. Moreover, in HF, the bacterial composition of colostrum leading to low serum IgG levels in the calf differed from that of colostrum leading to high serum IgG levels. Results of the present study indicate that the microbes present in colostrum are associated with transfer of passive immunity in neonatal calves.

Effect of lycopene supplementation to bovine oocytes exposed to heat shock during in vitro maturation.

We investigated the effect of the antioxidant lycopene supplemented into the in vitro maturation medium (TCM-199 with 20 ng/mL epidermal growth factor and 50 mg/mL gentamycin) in a heat shock (HS) model to mimic in vivo heat stress conditions. Bovine cumulus-oocyte complexes were supplemented with 0.2 µM lycopene (or not supplemented; control) under HS (40.5 °C) and non-HS (NHS; 38.5 °C) during maturation. After 22 h of maturation, we evaluated the nuclear status of the oocytes, the level of reactive oxygen species (ROS) production, and the respective blastocyst development and quality (via differential staining). Data were fitted in logistic and linear regression models, and the replicates were set as a random effect. The nuclear maturation was higher in NHS (84.0 ± 3.2%; least square mean ± standard error) than HS control (60.4 ± 4.3%; P < 0.001). Remarkably, the nuclear maturation in HS lycopene (71.7 ± 4.1%) was similar to NHS control (P = 0.7). Under HS conditions lycopene reduced ROS production (27.4 ± 4.8; relative fluorescence units (RFU)) in comparison to HS control (33.8 ± 1.8 RFU; P = 0.009). However, the ROS production in NHS lycopene (18.9 ± 2.0 RFU) was similar to NHS control (18.7 ± 1.8 RFU; P = 0.9). The cleavage rate in HS lycopene (76.1 ± 3.3%) was not lower than NHS lycopene (83.3 ± 2.5%; P > 0.1). On the day 8 of embryo development, the blastocyst rate was higher for NHS lycopene (55.2 ± 4.7%) versus NHS control (44.5 ± 4.7%; P = 0.04), but under HS the day 8 blastocyst rate was similar between control (29.9 ± 4.2%) and lycopene (32.3 ± 4.2%; P = 0.9). Lycopene supplementation increased the cell number of the embryos (total cell, trophectoderm, and inner cell mass numbers) under NHS conditions (P > 0.03). The apoptotic cell ratio was lower in lycopene (NHS and HS) versus control (NHS and HS) (P > 0.04). Lycopene has the ability to scavenge oocyte ROS and improved the cleavage rate of embryos under HS conditions. However, this could not be translated to a higher blastocyst development, which remained lower under HS. Results of our study indicate that antioxidant supplementation like lycopene during the maturation of bovine cumulus-oocyte complexes may be routinely used to improve blastocyst rate and quality under standard maturation conditions.

Researching 100 t cows: An innovative approach to identify intrinsic cows factors associated with a high lifetime milk production.

Longevity is an important trait both from an economic and social perspective. Modern dairy cows are criticized for their short productive lifespan: only a minority of animals survives to a fourth lactation, implying that most cows are culled before reaching their maximal potential. In contrast, the population of 100 t cows (HT), reaching the threshold of 100,000 kg lifetime milk yield, is growing rapidly. As these cows combine a long lifespan with high functionality, a better understanding of their intrinsic characteristics might help us to improve the overall lifespan and lifetime production in dairy cows. The aim of the present research was to compare HT with their less-producing herd mates in order to identify intrinsic cow factors associated with longevity and high lifetime production. Therefore, we matched 26,248 HT with 691,597 herd mates, born in the same year in the same herd. Data were provided by Coöperatie rundveeverbetering (CRV) and contained birth dates, calving dates, milk yield and dam information. In addition, scores for conformation traits based on classifications in the first lactation and breeding values (for milk yield, fertility, udder health and claw health) were provided. Multivariable conditional logistic regression models were built to identify factors associated with reaching a lifetime milk yield of 100,000 kg. Results revealed cows born in September and born out of heifers to have the highest odds to become a HT. When cows received a score ≥ 83 (population average 80) for udder and feet & legs conformation, they had higher odds of reaching the 100,000 kg threshold. While a greater body condition and larger rump angle increased the odds of becoming a HT, this was decreased in cows with a large body depth. Finally, breeding values for milk yield, fertility, udder health and claw health were positively associated with the likelihood of reaching a lifetime milk yield of 100,000 kg. In conclusion, to increase lifetime milk yield in dairy herds, farmers should select heifers with high scores for conformation traits like udder and feet & legs and high breeding values for milk yield, fertility and udder health. Furthermore, our data suggest that being born in September out of a heifer potentially contributes to reaching a high lifetime milk yield.

Effect of overconditioning on the hepatic global gene expression pattern of dairy cows at the end of pregnancy.

Overconditioning is a risk factor for upregulated pre- and postpartum fat mobilization. Therefore, we hypothesized that overconditioning at the end of pregnancy leads to the accumulation of lipids in the liver and modifications of the hepatic gene expression pattern. The aim of this study was to evaluate the effect of normal- versus overconditioning on the hepatic transcriptomic profile of dairy cows at the end of pregnancy. Ten dry multiparous Holstein cows were killed 2 wk before expected calving. Body condition score (BCS) and backfat thickness (BFT) were evaluated, and blood samples for nonesterified fatty acids (NEFA) were taken before cows were killed. After cows were killed, liver biopsy samples were collected for further assessment of total lipids and RNA sequencing. Five cows were classified as normal-conditioned (median BCS = 3, range 2.75-3.5) and 5 as overconditioned (median BCS = 4, range 4-5). Regression models confirmed that normal-conditioned cows had lower BFT (1.29 ± 0.29 cm; least squares means ± standard error) and serum NEFA (0.16 ± 0.04 mmol/L) in comparison to overconditioned cows (3.14 ± 0.43 cm and 0.38 ± 0.07 mmol/L for BFT and NEFA, respectively). Total liver lipid percentage tended to be lower in normal- versus overconditioned cows (4.63 ± 0.40% and 6.06 ± 0.44%, respectively). In comparison to the mean liver lipid percentage of the normal- and overconditioned cows, 1 overconditioned cow had a relatively low (5.21%) and 1 normal-conditioned cow had a relatively high (6.07%) liver lipid percentage. Differentially expressed genes analysis (edgeR quasi-likelihood method) showed that normal-conditioned cows presented 11 upregulated and 12 downregulated genes in comparison to overconditioned cows. Linear discriminant analysis effects size revealed 133 differentially expressed genes between normal- versus overconditioned cows. Notably, the liver of normal-conditioned cows had upregulated genes associated with liver functionality (ALB, SELENOP, IGF1, and IGF2). On the other hand, overconditioned cows had upregulated genes associated with the acute-phase response (C3, HPX, and, LBP). High basal lipolysis in overconditioned cows at the end of pregnancy increased liver lipid content, and this may alter the hepatic gene expression pattern to a pro-inflammatory state.

Retrospective study of factors associated with bovine infectious abortion and perinatal mortality.

Abortion and perinatal mortality, leading causes of economic loss in cattle industry, are the consequence of both non-infectious and a wide range of infectious causes. However, the relative contribution of pathogens to bovine abortion and perinatal mortality is poorly documented, since available studies involved only a limited number of pathogens. Therefore, the objectives of the present monitoring study were to determine the prevalence of infectious agents associated with bovine abortion and perinatal mortality, and to identify differences in production type, gestation length, parity and seasonality by using mixed effect models (logistic regression). A pre-established sampling protocol based on the collection of the aborted fetus/calf and a corresponding maternal blood sample, involving diagnostic testing for 10 pathogens, was performed. At least one potential causal agent of the abortion or perinatal mortality was detected in 39 % of cases. In these diagnosed cases, Neospora caninum was the most detected pathogen, followed by Trueperella pyogenes, BVDv, Escherichia coli, and Aspergillus fumigatus. Neospora caninum [odds ratio (OR): 0.4; 95 % confidence interval (CI): 0.3-0.7] and Aspergillus fumigatus (OR: 0.1; 95 % CI: 0.1-0.3) were detected less in late versus early gestation. Aspergillus fumigatus was less common in dairy in comparison to beef abortion cases (OR: 0.2; 95 % CI: 0.1-0.6). Winter was associated with a lower positivity for Neospora caninum and BVDv in comparison to warmer seasons. Despite extensive diagnostic testing, an etiological diagnosis was not reached in 61 % of cases, highlighting the need for even more extensive (non-)infectious disease testing or more accurate tests.

Low microbial biomass within the reproductive tract of mid-lactation dairy cows: A study approach.

The microbiome from the reproductive tract is being investigated for its putative effect on fertility, embryo development, and health status of the human or animal host postpartum. Besides the presence of a vaginal microbiome, recent studies have claimed the existence and putative role of the uterine microbiome. Yet, the extremely low bacterial numbers and high eukaryotic/prokaryotic DNA ratio make this a highly challenging environment to study with next-generation sequencing (NGS) techniques. Here, we describe the methodological challenges that are typically encountered when performing an accurate analysis of low microbial biomass samples, illustrated by data of our own observational study. In terms of the research question, we compared the microbial composition throughout different parts of the reproductive tract of clinically healthy, mid-lactation Holstein-Friesian cows. Samples were collected from 5 dairy cows immediately after killing. Swabs were taken from the vagina, and from 4 pre-established locations of the uterine endometrium. In addition to the conventional DNA extraction blank controls, sterile swabs rubbed over disinfected disposable gloves and the disinfected surface of the uterus (tunica serosa) before incision were taken as sampling controls. The DNA extraction, DNA quantification, quantitative PCR of the 16S rRNA genes, and 16S rRNA gene sequencing were performed. In terms of NGS data analysis, we performed prevalence-based filtering of putative contaminant operational taxonomic units (OTU) using the decontam R package. Although the bacterial composition differed between the vagina and uterus, no differences in bacterial community structure (α and ß diversity) were found among the different locations in the uterus. At phylum level, uterine samples had a greater relative abundance of Proteobacteria, and a lesser relative abundance of Firmicutes than vaginal samples. The number of shared OTU between vagina and uterus was limited, suggesting the existence of bacterial transmission routes other than the transcervical one to the uterus. The mid-lactation bovine genital tract is a low microbial biomass environment, which makes it difficult to distinguish between its constitutive versus contaminant microbiome. The integration of key controls is therefore strictly necessary to decrease the effect of accidentally introduced contaminant sequences and improve the reliability of results in samples with low microbial biomass.

A large-scale study on the effect of age at first calving, dam parity, and birth and calving month on first-lactation milk yield in Holstein Friesian dairy cattle.

Milk yield during first lactation is an important economical trait. Age at first calving (AFC) is considered an important predictor of subsequent milk yield. In addition, both season of birth, as well as season of calving, have been shown to influence milk production, with conflicting results. Finally, higher parity of the dam has been associated with a lower performance of the offspring. The aim of the present study was to assess the effect of the above-mentioned factors based on a large-scale study and to rank the most important determinants for first-lactation milk yield. Data on 3,810,678 Holstein Friesian heifers, born in Belgium and the Netherlands between 2000 and 2015, were provided by Cooperative CRV and CRV BV (Arnhem, the Netherlands) and consisted of birth dates, calving dates, and first-lactation productions. In addition, herd, sire, and dam information was provided. Linear regression models were built with herd-calving year and sire as random effects and 305-d energy-corrected milk (ECM) yield during first lactation as outcome variable. Birth month, calving month, parity of the dam, and AFC were included as fixed effects in the model and a dominance analysis was performed to rank the associated factors according to importance. Results revealed AFC to be the most important factor (R2 = 0.047), with an increase in ECM up to an age of 33 mo. Calving month was a more important predictor than birth month (R2 = 0.010 vs. R2 = 0.002, respectively), with the highest first-lactation production in heifers calving in October to December, and the lowest in heifers calving in June and July. Birth month had a limited effect on first-lactation milk yield (R2 = 0.002), potentially masked by rearing strategies during early life. Finally, parity of the dam ≥3 was associated with a reduced ECM of the offspring (R2 = 0.002). In conclusion, our results show AFC to be an important determinant of milk yield during first lactation. In addition, seasonal patterns in milk production are seen, which should be further explored to identify the underlying mechanism.

Genome-wide association for metabolic clusters in early-lactation Holstein dairy cows.

The aim of this study was to detect the genomic region or regions associated with metabolic clusters in early-lactation Holstein cows. This study was carried out in 2 experiments. In experiment I, which was carried out on 105 multiparous Holstein cows, animals were classified through k-means clustering on log-transformed and standardized concentrations of blood glucose, insulin-like growth factor I, free fatty acids, and ß-hydroxybutyrate at 14 and 35 d in milk (DIM), into metabolic clusters, either balanced (BAL) or other (OTR). Forty percent of the animals were categorized in the BAL group, and the remainder were categorized as OTR. The cows were genotyped for a total of 777,962 SNP. A genome-wide association study was performed, using a case-control approach through the GEMMA software, accounting for population structure. We found 8 SNP (BTA11, BTA23, and BTAX) associated with the predicted metabolic clusters. In experiment II, carried out on 4,267 second-parity Holstein cows, milk samples collected starting from the first week until 50 DIM were used to determine Fourier-transform mid-infrared (FT-MIR) spectra and subsequently to classify the animals into the same metabolic clusters (BAL vs. OTR). Twenty-eight percent of the animals were categorized in the BAL group, and the remainder were classified in the OTR category. Although daily milk yield was lower in BAL cows, we found no difference in daily fat- and protein-corrected milk yield in cows from the BAL metabolic cluster compared with those in the OTR metabolic cluster. In the next step, a single-step genomic BLUP was used to identify the genomic region(s) associated with the predicted metabolic clusters. The results revealed that prediction of metabolic clusters is a highly polygenic trait regulated by many small-sized effects. The region of 36,258 to 36,295 kb on BTA27 was the highly associated region for the predicted metabolic clusters, with the closest genes to this region (ANK1 and miR-486) being related to hematopoiesis, erythropoiesis, and mammary gland development. The heritability for metabolic clustering was 0.17 (SD 0.03), indicating that the use of FT-MIR spectra in milk to predict metabolic clusters in early-lactation across a large number of cows has satisfactory potential to be included in genetic selection programs for modern dairy cows.

The potential interaction between body condition score at calving and dietary starch content on productive and reproductive performance of early-lactating dairy cows.

Improving reproductive performance is one of the most important factors affecting the profitability of dairy herds. This study investigated the effect of feeding a high starch (HS) diet and body condition score (BCS) at calving on blood metabolites, fertility and ovarian function and milk production in Holstein dairy cows. One hundred seventy-four multiparous cows were fed common close-up and early lactation diets during the first 15 days in milk (DIM). Cows were randomly assigned to 1 of 2 experimental diets from 16 until 50 DIM (n = 87 per group); normal starch (228 g/kg diet DM; NS) or HS (270 g/kg diet DM; HS) diets. Each treatment group was further subdivided based on BCS at calving as normal BCS (BCS ⩽ 3.5; normal BCS (NBCS); n = 45) or high BCS (HBCS) (BCS ⩾ 3.75; HBCS; n = 42). A significant difference was detected for increased milk production (47.24 v. 44.55 kg/day) and decreased milk fat (33.93 v. 36.33 g/kg) in cows fed HS or NS, respectively. Plasma glucose and insulin concentrations were significantly higher in cows fed the HS compared to the NS diet. Diets significantly affected DIM at first artificial insemination (AI, 79.51 ± 3.83 v. 90.40 ± 3.83 days for cows fed HS and NS diets, respectively). High BCS groups had greater milk fat content and elevated plasma nonesterified fatty acids (NEFA), ß hydroxybutyrate (BHB) and bilirubin concentrations. In general, feeding higher starch diets to normal BCS cows during the first 50 DIM improved productive and reproductive performance of early-lactating dairy cows.

Assessment of associations between transition diseases and reproductive performance of dairy cows using survival analysis and decision tree algorithms.

This study aimed to evaluate the associations between transition cow conditions and diseases TD with fertility in Holstein cows, and to compare analytic methods for doing so. Kaplan-Meier, Cox proportional hazard, and decision tree models were used to analyze the associations of TD with the pregnancy risk at 120 and 210 DIM from a 1-year cohort with 1946 calvings from one farm. The association between TD and fertility was evaluated as follows: 1 cows with TD whether complicated with another TD or not TD-all, versus healthy cows, and 2 cows with uncomplicated TD TD-single, versus cows with multiple TD TD+; complicated cases, versus healthy cows. The occurrence of twins, milk fever, retained placenta, metritis, ketosis, displaced abomasum, and clinical mastitis were recorded. Using Kaplan-Meier models, in primiparous cows the 120 DIM pregnancy risk was 62% (95% CI: 57-67 %) for healthy animals. This was not significantly different for TD-single (58%; 95% CI: 51-66 %) but was reduced for TD+ (45%; 95% CI: 33-60 %). Among healthy primiparous cows, 80% (95% CI: 75-84 %) were pregnant by 210 DIM, but pregnancy risk at that time was reduced for primiparous cows with TD-single (72%; 95% CI: 65-79 %) and TD+ (62%; 95% CI: 49-75 %). In healthy multiparous cows, the 120 DIM pregnancy risk was 53% (95% CI: 49-56 %), which was reduced for TD-single (36%; 95% CI: 31-42 %) and TD+ (30%; 95% CI: 24-38 %). The 210 DIM pregnancy risk for healthy multiparous cows was 70% (95% CI: 67-72 %), being higher than the 210 DIM pregnancy risk for multiparous cows with TD-single (47%; 95% CI: 42-53 %) or TD+ (46%; 95% CI: 38-54 %). Cows with TD-all presented similar pregnancy risk estimates as for TD + . Cox proportional hazards regressions provided similar magnitudes of effects as the Kaplan-Meier estimates. Survival analysis and decision tree models identified parity as the most influential variable affecting fertility. Both modeling techniques concurred that TD + had a greater effect than TD-single on the probability of pregnancy at 120 and 210 DIM. Decision trees for individual TD identified that displaced abomasum affected fertility at 120 DIM in primiparous while metritis was the most influential TD at 120 and 210 DIM for multiparous cows. The data were too sparse to assess multiple interactions in multivariable Cox proportional hazard models for individual TD. Machine learning helped to explore interactions between individual TD to study their hierarchical effect on fertility, identifying conditional relationships that merit further investigation.