Investigating the uses of machine learning algorithms to inform risk factor analyses: The example of avian infectious bronchitis virus (IBV) in broiler chickens.

Infectious bronchitis virus (IBV) is a contagious coronavirus causing respiratory and urogenital disease in chickens and is responsible for significant economic losses for both the broiler and table egg layer industries. Despite IBV being regularly monitored using standard epidemiologic surveillance practices, knowledge and evidence of risk factors associated with IBV transmission remain limited. The study objective was to compare risk factor modeling outcomes between a traditional stepwise variable selection approach and a machine learning-based random forest Boruta algorithm using routinely collected IBV antibody titer data from broiler flocks. IBV antibody sampling events (n = 1111) from 166 broiler sites between 2016 and 2021 were accessed. Ninety-two geospatial-related and poultry-density variables were obtained using a geographic information system and data sets from publicly available sources. Seventeen and 27 candidate variables were screened to potentially have an association with elevated IBV antibody titers according to the manual selection and machine learning algorithm, respectively. Selected variables from both methods were further investigated by construction of multivariable generalized mixed logistic regression models. Six variables were shortlisted by both screening methods, which included year, distance to urban areas, main roads, landcover, density of layer sites and year, however, final models for both approaches only shared year as an important predictor. Despite limited significance of clinical outcomes, this work showcases the potential of a novel explorative modeling approach in combination with often unutilized resources such as publicly available geospatial data, surveillance health data and machine learning as potential supplementary tools to investigate risk factors related to infectious diseases.

Contrasting PRRSV temporal lineage patterns at the individual farm, production system, and regional levels in Ohio and neighboring states from 2017 to 2021.

Porcine reproductive and respiratory virus (PRRSV), one of the most significant viruses in the swine industry, has been challenging to control due to its high mutation and recombination rates and complexity. This retrospective study aimed to describe and compare the distribution of PRRSV lineages obtained at the individual farm, production system, and regional levels. PRRSV-2 (type 2) sequences (n = 482) identified between 2017 - 2021 were provided by a regional state laboratory (Ohio Department of Agriculture, Animal Disease Diagnostic Center (ODA-ADDL)) collected from swine farms in Ohio and neighboring states, including Indiana, Michigan, Pennsylvania, and West Virginia. Additional sequences (n = 138) were provided by one collaborating swine production system. The MUSCLE algorithm on Geneious Prime® was used to align the ORF5 region of PRRSV-2 sequences along with PRRSV live attenuated vaccine strains (n = 6) and lineage anchors (n = 169). Sequenced PRRSV-2 were assigned to the most identical lineage anchors/vaccine strains. Among all sequences (n = 620), 29.8% (185/620) were ≥ 98.0% identity with the vaccine strains, where 93.5% (173/185) and 6.5% (12/185) were identical with the L5 Ingelvac PRRS® MLV and L8 Fostera® PRRS vaccine strains, respectively, and excluded from the analysis. At the regional level across five years, the top five most identified lineages included L1A, L5, L1H, L1C, and L8. Among non-vaccine sequences with production system known, L1A sequences were mostly identified (64.3% - 100.0%) in five systems, followed by L1H (0.0% - 28.6%), L1C (0.0% - 10.5%), L5 (0.0% - 14.4%), L8 (0.0% - 1.3%), and L1F (0.0% - 0.5%). Furthermore, among non-vaccine sequences with the premise identification available (n = 262), the majority of sequences from five individual farms were either classified into L1A or L5. L1A and L5 sequences coexisted in three farms, while samples submitted by one farm contained L1A, L1H, and L5 sequences. Additionally, the lineage classification results of non-vaccine sequences were associated with their restriction fragment length polymorphism (RFLP) patterns (Fisher's exact test, p < 0.05). Overall, our results show that individual farm and production system-level PRRSV-2 lineage patterns do not necessarily correspond to regional-level patterns, highlighting the influence of individual farms and systems in shaping PRRSV occurrence within those levels, and highlighting the crucial goal of within-farm and system monitoring and early detection for accurate knowledge on PRRSV-2 lineage occurrence and emergence.

Dairy producers' colostrum practices, marketing, and attitudes toward male dairy calves.

Optimal early-life care of surplus calves born on dairy farms is critical for health and welfare. This cross-sectional study aimed to describe the marketing practices of male dairy calves, differences in the colostrum management between male and female calves on dairy farms, and the relationship between discrepant colostrum practices and dairy producers' attitudes toward male calf care. United States dairy producers (n = 1,000) in the states of Florida, Michigan, Ohio, Vermont, and Wisconsin were selected using stratified random sampling. A questionnaire containing questions about farm demographics, colostrum management (including quantity and timeliness of colostrum delivery), and producers' attitudes toward male calf care was mailed in February 2021. Attitudes toward male calf care were assessed using 5-point Likert scales for 10 statements regarding perceptions of the value of male calves and barriers in providing optimal care. Producer responses to questions about the quantity and timeliness of colostrum delivery between male and female calves were compared using a nonparametric Wilcoxon rank-sum test. Multivariable logistic regression models were used to investigate the association between differences in colostrum management and producers' attitudes. By May 2021, 953 surveys were delivered and 315 (33.1%) were returned with complete responses. Most producers (>90%) reported feeding equal volumes of colostrum and performing the first postbirth feeding within a similar time frame for male and female calves. However, compared with females, male calves had a longer delay to the first colostrum feeding. Approximately 40% of producers marketed their male calves through auctions, and over half (54.6%) of farms sold the male calves between 3 and 10 d of age. Large farms (≥500 lactating cows) were found to market male calves at a younger age (≤3 d of age). Most producers (>78%) believed their male calves were receiving optimal care and did not consider the workload and financial costs as obstacles to providing good care to male calves. However, those who viewed the workload as an obstacle to good care and produced organic products were marginally more likely to feed a lower volume of colostrum to male calves in the first feeding after birth. These findings suggest that interventions to improve colostrum practices should be more broadly targeted to all calves born on dairy farms, and that age at transport is markedly different between large and small farms.

Exploring the effects of transport duration on the fecal microbial communities of surplus dairy calves.

Transportation significantly affects the health and welfare of surplus dairy calves, largely due to the various stressors and pathogen exposures encountered during the process. Concurrently, an animal's microbiome is known to correlate with its health status, with stress-induced alterations in the microbiota potentially precipitating various diseases. This study aimed to compare the effects of transportation durations of 6, 12, or 16 h on the fecal microbiota in young surplus dairy calves. We used a randomized controlled design in which surplus dairy calves aged 1 to 19 d from 5 commercial dairy farms in Ontario were allocated into 1 of 3 transportation groups (6, 12, and 16 h of continuous transportation). Health assessments were conducted before, immediately after, and for 2 wk following transportation. Fecal samples were collected before, immediately after, and at 24 and 72 h after transportation and subjected to 16S rRNA sequencing. Alpha diversity metrics showed no significant differences between the 3 transportation groups at any of the sampling time points. Although ß diversity metrics revealed no clustering by transportation groups, they indicated significant differences across sampling time points within each group. The overall analysis revealed a total of 22 phyla and 353 genera, with Firmicutes, Bacteroidetes, Proteobacteria, Actinobacteria, and Fusobacteria being the most abundant phyla. Bacteroides, Escherichia/Shigella, Lactobacillus, Collinsella, and Bifidobacterium were the most abundant genera. The reduction in Fusobacteria abundance before and after transport was significantly larger in the 16-h transportation group when compared with the 6-h transportation group. We also identified several genus-level and amplicon sequence variation-level taxa that displayed significant differences in their abundances across various transportation groups, observed at all sampling time points investigated. This research identifies microbiota changes due to varying transportation durations in surplus dairy calves, providing a broad understanding of the microbial shifts in surplus dairy calves after transportation across varying durations. Although these variations may not directly correlate with overall calf health or indicate dysbiosis, these results emphasize the importance of further investigating transportation practices to enhance calf health and well-being. Further studies are warranted to elucidate the relationship between microbiota and calf health.

Normalizing real-time PCR results in routine testing.

Normalization, the process of controlling for normal variation in sampling and testing, can be achieved in real-time PCR assays by converting sample quantification cycles (Cqs) to "efficiency standardized Cqs" (ECqs). We calculated ECqs as E-ΔCq, where E is amplification efficiency and ΔCq is the difference between sample and reference standard Cqs. To apply this approach to a commercial porcine reproductive and respiratory syndrome virus (PRRSV) RT-qPCR assay, we created reference standards by rehydrating and then diluting (1 × 10-4) a PRRSV modified-live vaccine (PRRS MLV; Ingelvac) with serum or oral fluid (OF) to match the sample matrix to be tested. Sample ECqs were calculated using the mean E and reference standard Cq calculated from the 4 reference standards on each plate. Serum (n = 132) and OF (n = 130) samples were collected from each of 12 pigs vaccinated with a PRRSV MLV from -7 to 42 d post-vaccination, tested, and sample Cqs converted to ECqs. Mean plate Es were 1.75-2.6 for serum and 1.7-2.3 for OF. Mean plate reference standard Cqs were 29.1-31.3 for serum and 29.2-31.5 for OFs. Receiver operating characteristic analysis calculated the area under the curve for serum and OF sample ECqs as 0.999 (95% CI: 0.997, 1.000) and 0.947 (0.890, 1.000), respectively. For serum, diagnostic sensitivity and specificity of the commercial PRRSV RT-qPCR assay were estimated as 97.9% and 100% at an ECq cutoff ≥ 0.20, and for OF, 82.6% and 100%, respectively, at an ECq cutoff ≥ 0.45.

A survey of antimicrobial use practices on veal farms in the United States.

Neonatal veal calves are highly susceptible to bacterial diseases. Occasional sub-optimal early-life care and long-distance transportation result in high disease burden and antimicrobial treatment incidence. Nonetheless, judicious use of antimicrobials is necessary to mitigate the animal and human health impacts of antimicrobial resistance. The objective of this cross-sectional study was to use a clinical vignette-based survey approach to evaluate the potential for reduced group or individual therapeutic antimicrobial use. The survey included items probing the adherence to veterinary-written protocols, antimicrobial use (AMU) at the calf- and group-level, and the treatment actions to case vignettes of calf diarrhoea, pneumonia, and navel infection. The survey was mailed to all veal calf producers within two U.S. production companies (n = 32 producers in Company 1; n = 70 producers in Company 2) in February and December 2019, respectively. The overall survey response rate was 36% (21/59). Although 95% of producers reported having veterinary-written treatment protocols for diarrhoea, pneumonia, and navel infection, veal producers infrequently (<50% of the time) referenced these protocols. Veal producers were primarily trained for disease identification and treatment by observing other personnel "on-the-job" (81%). Veal producers reported a high incidence of calf diarrhoea relative to pneumonia and naval infection, a lower percentage (≤40%) of diarrhoea cases being treated individually with antimicrobials. Using clinical vignettes, our results suggest that AMU decisions among veal producers depend on the severity of clinical signs for diarrhoea and pneumonia, while navel infections are often treated with antimicrobials regardless of sign severity. Nearly two-thirds of veal producers reported treating uncomplicated cases of diarrhoea (watery stool with normal body temperature, activity, and appetite), more than previously reported from dairy producers (37.1%). Findings from this survey suggest that calf producer-focused training to guide antimicrobial treatment decisions and improve producer adherence to veterinary-written treatment protocols may have important impacts on judicious antimicrobial use.

Performance of a Differentiation of Infected from Vaccinated Animals (DIVA) Classical Swine Fever Virus (CSFV) Serum and Oral Fluid Erns Antibody AlphaLISA Assay.

Classical swine fever virus (CSFV) is an OIE-listed disease that requires effective surveillance tools for its detection and control. The aim of this study was to develop and evaluate the diagnostic performance of a novel CSFV Erns IgG AlphaLISA for both serum and oral fluid specimens that would likewise be compatible with the use of CSFV E2 DIVA vaccines. Test performance was evaluated using a panel of well-characterized serum (n = 760) and individual (n = 528) or pen-based (n = 30) oral fluid samples from four groups of animals: (1) negative controls (n = 60 pigs); (2) inoculated with ALD strain wild-type CSFV (n = 30 pigs); (3) vaccinated with LOM strain live CSFV vaccine (n = 30 pigs); and (4) vaccinated with live CSFV marker vaccine on commercial farms (n = 120 pigs). At a cutoff of S/P ≥ 0.7, the aggregate estimated diagnostic sensitivities and specificities of the assay were, respectively, 97.4% (95% CI 95.9%, 98.3%) and 100% for serum and 95.4% (95% CI 92.9%, 97.0%) and 100% for oral fluid. The Erns IgG antibody AlphaLISA combined DIVA capability with solid diagnostic performance, rapid turnaround, ease of use, and compatibility with both serum and oral fluid specimens.

Swine industry stakeholders' perception on the use of water-based foam as an emergency mass depopulation method.

The U.S. pork supply chain is vulnerable to various internal and external threats and in need of prompt, comprehensive response plans. Under urgent circumstances, for example in the case of foreign disease incursions, swine farms will have to perform on-farm animal depopulation to prevent disease spread. Several animal depopulation methods including water-based foam (WBF) have been proposed and are under evaluation for feasibility in the field. However, the psychological/emotional impacts of applying depopulation methods for personnel managing and carrying on the tasks are not currently well understood. Thus, this study aimed to investigate WBF as an alternative for depopulation compared to existing methods approved by the American Veterinary Medical Association. Swine industry stakeholders were invited to voluntarily observe a WBF depopulation trial and to provide their self-reported perspectives before and after the observation. A survey was designed to explore key areas on expected and perceived method effectiveness, efficiency, and animal welfare considerations, as well as to evaluate short-term post-observation psychological impacts. Among 42 recruited stakeholders, 31.0% (13/42) were educators/researchers followed by animal health officials (26.2%, 11/42) and veterinarians (23.8%, 10/42), with an average of 11.7 ± 12.6 (n = 39) years of work experience. After the trial, respondents' positive perception of WBF depopulation increased specifically regarding the animal loading process being less stressful than restrained in-barn depopulation options (P = 0.003) and by the observation of fewer swine escape attempts and vocalizations than expected (P < 0.001). Respondents' positive perception of WBF also increased regarding to the time required to fill the trailer with foam, to stop hearing animal vocalization, and stop hearing animal movement, as the observed trial times were faster than their pre-observation estimates (P < 0.001). Additionally, 79.5% (31/39) of respondents agreed that the rapid destruction of animal populations had priority over animal welfare under urgent scenarios. Minor post-traumatic stress disorder-like (PTSD-like) symptoms from the observed trials were reported (26.7%, 4/15 respondents) one month after the observation. This study showed that the WBF depopulation process was perceived positively by swine stakeholders and may have limited short-term psychological impacts on personnel involved in animal depopulation.

REST-repressed lncRNA LINC01801 induces neuroendocrine differentiation in prostate cancer via transcriptional activation of autophagy.

The association between REST reduction and the development of neuroendocrine prostate cancer (NEPC), a novel drug-resistant and lethal variant of castration-resistant prostate cancer (CRPC), is well established. To better understand the mechanisms underlying this process, we aimed to identify REST-repressed long noncoding RNAs (lncRNAs) that promote neuroendocrine differentiation (NED), thus facilitating targeted therapy-induced resistance. In this study, we used data from REST knockdown RNA sequencing combined with siRNA screening to determine that LINC01801 was upregulated and played a crucial role in NED in prostate cancer (PCa). Using The Cancer Genome Atlas (TCGA) prostate adenocarcinoma database and CRPC samples collected in our laboratory, we demonstrated that LINC01801 expression is upregulated in NEPC. Functional experiments revealed that overexpression of LINC01801 had a slight stimulatory effect on the NED of LNCaP cells, while downregulation of LINC01801 significantly inhibited the induction of NED. Mechanistically, LINC01801 is transcriptionally repressed by REST, and transcriptomic analysis revealed that LINC01801 preferentially affects the autophagy pathway. LINC01801 was found to function as a competing endogenous RNA (ceRNA) to regulate the expression of autophagy-related genes by sponging hsa-miR-6889-3p in prostate cancer cells. In conclusion, our data expand the current knowledge of REST-induced NED and highlight the contribution of the REST-LINC01801-hsa-miR-6889-3p axis to autophagic induction, which may provide promising avenues for therapeutic opportunities.

Correction: Hsu et al. Development of Folic Acid-Conjugated and Methylene Blue-Adsorbed Au@TNA Nanoparticles for Enhanced Photodynamic Therapy of Bladder Cancer Cells. Nanomaterials 2020, 10, 1351.

Within the publication of this article in Nanomaterials 2020, 10, 1351 [...].

Refinement of water-based foam depopulation procedures for finisher pigs during field conditions: Welfare implications and logistical aspects.

Water-based foam (WBF) depopulation is currently being researched as an alternative for rapid destruction of swine populations under emergency circumstances. Appropriate guidelines are needed to maintain method reliability and depopulation efficacy while minimizing animal distress under field conditions. Finisher pigs were depopulated using WBF with a 7.5-minute dwell time in two trials to evaluate the effect of; trial 1) foam fill level (1.5, 1.75, or 2.0 times the pig's head height) and trial 2) foam fill rate (slow, medium, or fast) on aversive pig responses (surface breaks, vocalization, and escape attempts) and time to cessation of cardiac activity. Activity and cardiac activity were recorded using subcutaneous bio-loggers for swine in trial 2. The average time to cessation of movement (COM) from the start of foam filling was compared for the foam fill rate groups using a generalized linear mixed effect model under Poisson distribution. Foam rate group was used as an independent variable, and replicates as a random effect. For trial 1, the average (mm:ss ± SD) time to fill completion was 01:18 ± 00:00, 00:47 ± 00:05, and 00:54 ± 00:05, for 1.5, 1.75, and 2.0 times the pig's head height, respectively. For trial 2, the average time to fill completion was 03:57 ± 00:32, 01:14 ± 00:23 and 00:44 ± 00:03, and the average time (mm:ss ± SE) to COM was 05:22 ± 00:21, 03:32 ± 00:14, and 03:11 ± 00:13 for slow, medium, and fast fill rate groups, respectively. A higher number of aversive pig responses were observed for the lowest foam fill level and slowest foam fill rate compared to increased fill levels and faster fill rates. For trial 2 the median (mm:ss ± IQR) time to fatal arrhythmia was 09:53 ± 02:48, 11:19 ± 04:04, and 10:57 ± 00:47 post-foam initiation for fast, medium, and slow foam rate groups, respectively. Time to cessation of cardiac activity was significantly shorter for the fast foam rate group compared to medium and slow foam rates groups (P = 0.04). For both trials, vocalizations were absent, and all pigs were unconscious following the 7.5-minute dwell time and no pigs needed a secondary euthanasia method. This WBF study showed that slower fill rates and low foam fill levels may extend the time until cessation of cardiac activity in swine during depopulation. A conservative recommendation with consideration of swine welfare during an emergency scenario would be a minimum foam fill level twice the pig's head height and a foam fill rate capable of covering pigs in foam within 60 s to minimize aversive responses and expedite cessation of cardiac activity.

Water-based medium-expansion foam depopulation of adult cattle.

Current options for depopulation of adult cattle are limited, have logistic constraints, and may not be practical on a large scale. Aspirated water-based foam (WBF) has been shown to be successful in depopulating poultry and swine but has yet to be tested in cattle. WBF is advantageous because necessary equipment can be readily available, easy to use, and presents minimal personnel risk. With the use of a modified rendering trailer in a field setting, we evaluated the efficacy of aspirated WBF for depopulation of adult cattle. Water-based medium-expansion foam was added to the trailer holding cattle to a depth of approximately 50 cm greater than head height. The study was conducted as a gated design and the initial trial was conducted using six anesthetized and six conscious animals for verification of the process and followed by four replicates each containing 18 conscious cattle. A total of 84 cattle were used, with a subset (n = 52) implanted with subcutaneous bio-loggers that recorded activity and electrocardiograms. Cattle were loaded onto the trailer and three gasoline-powered water pumps delivered foam into the trailer followed by a 15-min foam dwell period. Average (± SD) time to completely fill the trailer with foam was 84.8 ±â€…11.0 s. No animal vocalizations were heard during foam application or the dwell period, and all cattle were confirmed dead upon removal from the trailer after 15 min of immersion. Necropsies of a subset of cattle revealed foam extending to at least the tracheal bifurcation in all cattle and distal to this level in 67% (8/12) animals. Time to cessation of movement, which served as a proxy for loss of consciousness, was 2.5 ±â€…1.3 min and time to cardiac death was 8.5 ±â€…2.5 min as determined by data from animals carrying subcutaneous bio-loggers. The results of this study indicate that WBF is a rapid and effective method for depopulation of adult cattle with potential advantages in speed and carcass handling and disposal over current methods.

Effect of extrinsic factors on the detection of PRRSV and a porcine-specific internal sample control in serum, oral fluid, and fecal specimens tested by RT-rtPCR.

We characterized the effect of 1) temperature × time, 2) freeze-thaw cycles, and 3) high porcine reproductive and respiratory syndrome virus (PRRSV) RNA concentrations on the detection of PRRSV and a porcine-specific internal sample control (ISC) in serum, oral fluid, and fecal samples using a commercial PRRSV RT-rtPCR assay (Idexx). In study 1, the effect of temperature × time on PRRSV and ISC detection was shown to be specimen dependent. In serum stored at 4, 10, or 20°C, PRRSV detection was consistent for up to 168 h, but storage at 30°C reduced detectable PRRSV RNA. ISC RNA was stable in serum held at 4 and 10°C, but not at 20 and 30°C. In contrast, PRRSV and ISC RNAs in oral fluid and fecal samples continuously decreased at all temperature × time treatments. Based on these data, serum samples should be stored at ≤ 20°C to optimize PRRSV RNA detection. Oral fluid and fecal samples should be frozen in a non-self-defrosting freezer until tested. In study 2, freeze-thaw cycles had little impact on PRRSV and ISC detection, but more so in oral fluids than serum or fecal samples. Thus, freeze-thaw cycles in oral fluids should be minimized before RT-rtPCR testing. In study 3, the ISC was not affected by high concentrations of PRRSV RNA in serum, oral fluid, or fecal samples. It should not be assumed that data from our PRRSV study are applicable to other pathogens; additional pathogen-specific studies are required.

Detection of Mannheimia haemolytica-Specific IgG, IgM and IgA in Sera and Their Relationship to Respiratory Disease in Cattle.

Mannheimia haemolytica is one of the major causes of bovine respiratory disease in cattle. The organism is the primary bacterium isolated from calves and young cattle affected with enzootic pneumonia. Novel indirect ELISAs were developed and evaluated to enable quantification of antibody responses to whole cell antigens using M. haemolytica A1 strain P1148. In this study, the ELISAs were initially developed using sera from both M. haemolytica-culture-free and clinically infected cattle, then the final prototypes were tested in the validation phase using a larger set of known-status M. haemolytica sera (n = 145) collected from feedlot cattle. The test showed good inter-assay and intra-assay repeatability. Diagnostic sensitivity and specificity were estimated at 91% and 87% for IgG at a cutoff of S/P ≥ 0.8. IgM diagnostic sensitivity and specificity were 91% and 81% at a cutoff of sample to positive (S/P) ratio ≥ 0.8. IgA diagnostic sensitivity was 89% whereas specificity was 78% at a cutoff of S/P ≥ 0.2. ELISA results of all isotypes were related to the diagnosis of respiratory disease and isolation of M. haemolytica (p-value < 0.05). These data suggest that M. haemolytica ELISAs can be adapted to the detection and quantification of antibody in serum specimens and support the use of these tests for the disease surveillance and disease prevention research in feedlot cattle.

Comparison of Two Diagnostic Assays for the Detection of Serum Neutralizing Antibody to Porcine Epidemic Diarrhea Virus.

Lactogenic immunity is important for the protection of piglets against many pathogens including porcine epidemic diarrhea virus. Circulating neutralizing antibodies levels in sow sera may help determine if a detectable immune response could confer protection to piglets. Neutralizing antibodies can be detected through various diagnostic assays. This study evaluated the diagnostic characteristics of two neutralizing antibody assays for porcine epidemic diarrhea virus neutralizing antibodies in serum of challenged gilts. Four treatment groups, control, non-vaccinated, vaccinated prior to challenge, and vaccinated following challenge, were comprised of 20 gilts. Serum sample were collected from each gilt prior to and following challenge with porcine epidemic diarrhea virus. Samples were evaluated for the presence of neutralizing antibodies via a fluorescent focus neutralization assay and a high-throughput neutralization assay. Diagnostic sensitivity and specificity for the fluorescent focus neutralization and high-throughput neutralization assays for this study were optimized at a cutoff of a dilution of 80 and 80% fluorescent reduction respectively and demonstrated moderate agreement based off the kappa statistic. The focus fluorescent neutralization and high-throughput neutralization assays can be used to monitor the status of neutralizing antibodies within animals or a population of animals. The high-throughput assay has advantages over the focus fluorescent assay in that it has a higher specificity at the indicated cut-off and the nature of the results allows for more discrimination between individual results.

A survey of male and female dairy calf care practices and opportunities for change.

The primary objective of this study was to compare male and female dairy calf management practices and evaluate risk factors associated with differences in care. Secondary objectives were to understand surplus calf transportation and marketing practices and investigate incentives to motivate calf care improvements. An online survey was distributed to all dairy producers in Ontario (n = 3,367) from November 2020 to March 2021 and Atlantic Canada (n = 557) from April to June 2021. Dairy producers were identified through provincial dairy associations and contacted via e-mail and social media. Descriptive statistics were computed, and a logistic regression model was created to evaluate factors associated with using discrepant feeding practices (i.e., fed less colostrum, fed colostrum later, or fed raw, unsalable milk) for male calves compared with females. The survey had a 7.4% response rate (n = 289/3,924) and was primarily filled out by farm owners (76%). Although colostrum and milk feeding practices were similar between male and female calves, male calves received less milk while still on the dairy farm of origin compared with females. Male calves were also more likely to be fed a higher proportion of raw, unsalable milk. Female producers and those that kept their male calves beyond 10 d of age had lower odds of using poorer feeding practices for male calves. Male calves were mostly sold between 1 and 10 d (64%), primarily through direct sales to a calf-rearing facility (45%), with auctions being the next most common method (35%). A small but notable proportion of producers (18%) agreed that euthanizing male calves is a reasonable alternative when their sale price is very low; however, few producers (13%) reported that financial costs limited their male calf care. The largest proportion (43%) of producers reported that a price premium for more vigorous calves would motivate them to take better care of their male calves. Conversely, only 28% of producers reported that a price discount for calves in poor condition would be motivating. Producers placed importance on the opinion of their calf buyer, their herd veterinarian, and the Canadian Code of Practice for the Care and Handling of Dairy Cattle when considering their calf care practices, and they highly valued practices that promote calf health. Respondents to this survey reported a lower proportion of tiestall barn use and higher milk productivity compared with typical dairy herds in the region, suggesting selection bias for more progressive dairy producers. Nevertheless, our results suggest that dairy producers provide similar care between male and female calves, but some male calves experience challenges due to milk feeding and marketing practices. Feedback from calf buyers along with continued support and guidance from herd veterinarians and the Code of Practice may motivate dairy producers to improve male calf care.

What Should I do and Who's to blame? A cross-national study on youth's attitudes and beliefs in times of COVID-19.

The COVID-19 crisis has had a major impact on youth. This study examined factors associated with youth's attitudes towards their government's response to the pandemic and their blaming of individuals from certain risk groups, ethnic backgrounds, and countries or regions. In a sample of 5,682 young adults (Mage = 22) from 14 countries, lower perceived burden due to COVID-19, more collectivistic and less individualistic values, and more empathy were associated with more positive attitudes towards the government and less blaming of individuals of certain groups. Youth's social identification with others in the pandemic mediated these associations in the same direction, apart from the COVID-19 burden on attitudes, which had a positive indirect effect. No evidence of country-level moderation was found.

Internal reference genes with the potential for normalizing quantitative PCR results for oral fluid specimens.

In basic research, testing of oral fluid specimens by real-time quantitative polymerase chain reaction (qPCR) has been used to evaluate changes in gene expression levels following experimental treatments. In diagnostic medicine, qPCR has been used to detect DNA/RNA transcripts indicative of bacterial or viral infections. Normalization of qPCR using endogenous and exogenous reference genes is a well-established strategy for ensuring result comparability by controlling sample-to-sample variation introduced during sampling, storage, and qPCR testing. In this review, the majority of recent publications in human (n = 136) and veterinary (n = 179) medicine did not describe the use of internal reference genes in qPCRs for oral fluid specimens (52.9% animal studies; 57.0% human studies). However, the use of endogenous reference genes has not been fully explored or validated for oral fluid specimens. The lack of valid internal reference genes inherent to the oral fluid matrix will continue to hamper the reliability, reproducibility, and generalizability of oral fluid qPCR assays until this issue is addressed.

Description of electroencephalographic data gathered using water-based medium-expansion foam as a depopulation method for nursery pigs.

The United States' swine industry is under constant threat of foreign animal diseases, which may emerge without warning due to the globalized transportation networks moving people, animals, and products. Therefore, having disease control and elimination protocols in place prior to pathogen introduction is paramount for business continuity and economic recovery. During extraordinary circumstances, it may become necessary to depopulate large populations of animals, including swine, as a disease containment measure. Currently approved depopulation methods for swine present significant logistical challenges when scaled to large populations or performed in field conditions. In the United States, water-based foam is currently approved for poultry depopulation, and recent field studies demonstrate water-based foam is an effective depopulation alternative for swine. While effective, the speed at which water-based foam induces loss of consciousness prior to death, a major welfare consideration, has not been adequately investigated. In this study, 12 nursery pigs were terminated using water-based medium-expansion foam to quantify the time to induce loss of consciousness and ultimately brain death. Each pig was implanted with subdermal electrodes to capture electroencephalographic data, placed in a body sling, and suspended in a plastic bulk container that was subsequently filled with water-based foam. Electroencephalographic data was recorded for 15 min, during which the pigs remained immersed in the water-based foam. Conservatively, average (± SD) time to unconsciousness and brain death was 1 min, 53 s ± 36 s and 3 min, 3 s ± 56 s, respectively. The relatively rapid loss of consciousness compared to other methods limits the amount of distress and is overall a positive finding for the welfare of the pigs that might be depopulated with water-based foam. The findings of this study add additional evidence supporting the use of water-based medium-expansion foam for an emergency depopulation of swine.