Inter-laboratory comparison of eleven quantitative or digital PCR assays for detection of proviral bovine leukemia virus in blood samples.

Bovine leukemia virus (BLV) is the etiological agent of enzootic bovine leukosis and causes a persistent infection that can leave cattle with no symptoms. Many countries have been able to successfully eradicate BLV through improved detection and management methods. However, with the increasing novel molecular detection methods there have been few efforts to standardize these results at global scale. This study aimed to determine the interlaboratory accuracy and agreement of 11 molecular tests in detecting BLV. Each qPCR/ddPCR method varied by target gene, primer design, DNA input and chemistries. DNA samples were extracted from blood of BLV-seropositive cattle and lyophilized to grant a better preservation during shipping to all participants around the globe. Twenty nine out of 44 samples were correctly identified by the 11 labs and all methods exhibited a diagnostic sensitivity between 74 and 100%. Agreement amongst different assays was linked to BLV copy numbers present in samples and the characteristics of each assay (i.e., BLV target sequence). Finally, the mean correlation value for all assays was within the range of strong correlation. This study highlights the importance of continuous need for standardization and harmonization amongst assays and the different participants. The results underscore the need of an international calibrator to estimate the efficiency (standard curve) of the different assays and improve quantitation accuracy. Additionally, this will inform future participants about the variability associated with emerging chemistries, methods, and technologies used to study BLV. Altogether, by improving tests performance worldwide it will positively aid in the eradication efforts.

Paediatric nail consultation in an academic centre in Belgium: a 10-year retrospective study.

BACKGROUND: Very few studies have been conducted to establish the nature and prevalence of nail disorders in children. OBJECTIVES: To determine the frequency of various nail conditions in the paediatric setting and to report their management and follow-up. METHODS: This was a retrospective study between 2007 and 2017 of children under 18. All the patients were evaluated in our paediatric nail clinic at the dermatology department of Queen Fabiola Children's University Hospital. The data were synthesized from information obtained through medical records as well as from photographs taken during consultation. Follow-up was completed by phone interview. RESULTS: Three hundred and one patients were included. The majority of nail abnormalities involved the toenails (57.6%). The most common clinical signs were, in descending order, Beau's lines, pachyonychia, subungual hyperkeratosis and onycholysis. The most frequent diagnoses were fever-related Beau's lines or onychomadesis (9.7%), trachyonychia (8.4%), longitudinal melanonychia (8.1%) and congenital malalignment of the great toenail (8.1%). The main diagnoses by age group were as follows: congenital hypertrophy of the lateral nail folds (21.4%) [0-2 years old]; fever-related Beau's lines or onychomadesis (21%) [2-6 years old]; trachyonychia (22%) [6-12 years old]; and juvenile ingrown nail (21.4%) [12-18 years old]. Management included clinical observation for 119 patients and specific advices for 108 patients. A treatment was prescribed for 134 patients, topical in 76.5% of cases. Follow-up demonstrated complete healing in 50.6% of patients and improvement in 19.7%. CONCLUSION: The most frequent nail disorders are benign, and their distribution varies with age. Management mainly involves conservative care, and the prognosis is favourable in the majority.

Analysis of tRNA halves (tsRNAs) in serum from cattle challenged with bovine viral diarrhea virus.

Acute infections of bovine viral diarrhea virus (BVDV) lead to a range of clinical presentations. Laboratory tests for detection depend on collection of samples during a short viremia. Acutely infected animals remain largely undiagnosed. Transfer RNA halves (tsRNAs) are hypothesized to function like microRNAs to regulate gene expression during an immune response. The objective of this study was to identify tsRNAs in cattle that had been challenged with a non-cytopathic field strain of BVDV. Colostrum-deprived neonatal Holstein calves were either challenged with BVDV (n=5) or mock challenged (n=4). Sera was collected prior to challenge and days 4, 9, and 16 post challenge. RNA was extracted and read counts of small non-coding RNAs were assessed using next-generation sequencing. A total of 87,838,207 reads identified 41 different tsRNAs. Two 5' tsRNAs, tsRNAProAGG and tsRNAValAAC, differed across time. Two 5' tsRNAs, tsRNAGlyCCC and tsRNAGlyGCC, differed between treatment groups across time. Four days post challenge, 5' tsRNAGlyCCC and tsRNAGlyGCC were significantly lower in the challenged group than the control group. Further studies are needed to identify the importance and function of 5' tsRNAGlyCCC and tsRNAGlyGCC in serum samples of cattle challenged with BVDV.

Perceptions of general and postpresidential election discrimination are associated with loss of control eating among racially/ethnically diverse young men.

OBJECTIVE: The purpose of this study was to examine the association between young men's perceived experiences with discrimination, both general and following the 2016 presidential election, and their loss of control (LOC) eating. The degree to which men identified with their ethnic identity was evaluated as a moderator. METHOD: The sample included 798 men (18-30 years; M = 24.0 ± 3.6) who identified as African American (n = 261), Asian/Asian American (n = 266), or Hispanic/Latino (n = 271). Participants completed an online survey of items assessing demographic characteristics; perceived discrimination; perceptions of race-related discrimination following the 2016 U.S. presidential election; ethnic identity; and LOC eating. RESULTS: After adjusting for income, education, generational status and body mass index, perceived discrimination was positively associated with LOC eating frequency in African American and Hispanic/Latino men (ps < .01). Ethnic identity was inversely associated with LOC eating frequency in Hispanic/Latino men (p < .001). In Asian/Asian American men, perceived discrimination was only associated with more LOC eating among those with a low ethnic identity (p < .001). Higher levels of perceived discrimination following the presidential election were uniquely associated with more frequent LOC eating (p < .01) only among Asian/Asian American men who were not born in the United States or whose parents were not born in the United States. DISCUSSION: LOC eating may partially explain known associations between discrimination and heightened risk for obesity and chronic diseases among African American and Hispanic/Latino men. Asian/Asian American men's LOC eating may be linked to postpresidential election and general experiences with racial discrimination, particularly if they report a low sense of belonging to their ethnic group.

Current Demographics and Roles of Florida Community Health Workers: Implications for Future Recruitment and Training.

The high prevalence of health disparity diseases (e.g., obesity, Type 2 diabetes) among underserved populations in the United States suggests the need for increased resources to prevent these diseases and to improve health care access and quality in underserved communities. Community health workers are valuable resources and facilitators of health care access and quality treatment. The purpose of the present study is to provide descriptive information about community health workers in Florida and to provide recommendations for improved training and expansion of community health workers' roles in research and intervention. The study participants were 396 adults (85.1% Female, 75.5% Community health workers) who completed the 2015 Florida Community Health Workers Census. Participants were recruited by the Health Council of South Florida through emails and phone calls to members of the Florida Community Health Worker Coalition and various organizations. It was found that several groups disproportionately affected by health disparities were underrepresented among the community health workers who participated in the census and among the communities served by these community health workers. Actions are needed to improve and increase the recruitment and training of community health workers in Florida.

Structural organization of membrane-inserted hexamers formed by Helicobacter pylori VacA toxin.

Helicobacter pylori colonizes the human stomach and is a potential cause of peptic ulceration or gastric adenocarcinoma. H. pylori secretes a pore-forming toxin known as vacuolating cytotoxin A (VacA). The 88 kDa secreted VacA protein, composed of an N-terminal p33 domain and a C-terminal p55 domain, assembles into water-soluble oligomers. The structural organization of membrane-bound VacA has not been characterized in any detail and the role(s) of specific VacA domains in membrane binding and insertion are unclear. We show that membrane-bound VacA organizes into hexameric oligomers. Comparison of the two-dimensional averages of membrane-bound and soluble VacA hexamers generated using single particle electron microscopy reveals a structural difference in the central region of the oligomers (corresponding to the p33 domain), suggesting that membrane association triggers a structural change in the p33 domain. Analyses of the isolated p55 domain and VacA variants demonstrate that while the p55 domain can bind membranes, the p33 domain is required for membrane insertion. Surprisingly, neither VacA oligomerization nor the presence of putative transmembrane GXXXG repeats in the p33 domain is required for membrane insertion. These findings provide new insights into the process by which VacA binds and inserts into the lipid bilayer to form membrane channels.

The players may change but the game remains: network analyses of ruminal microbiomes suggest taxonomic differences mask functional similarity.

By mapping translated metagenomic reads to a microbial metabolic network, we show that ruminal ecosystems that are rather dissimilar in their taxonomy can be considerably more similar at the metabolic network level. Using a new network bi-partition approach for linking the microbial network to a bovine metabolic network, we observe that these ruminal metabolic networks exhibit properties consistent with distinct metabolic communities producing similar outputs from common inputs. For instance, the closer in network space that a microbial reaction is to a reaction found in the host, the lower will be the variability of its enzyme copy number across hosts. Similarly, these microbial enzymes that are nearby to host nodes are also higher in copy number than are more distant enzymes. Collectively, these results demonstrate a widely expected pattern that, to our knowledge, has not been explicitly demonstrated in microbial communities: namely that there can exist different community metabolic networks that have the same metabolic inputs and outputs but differ in their internal structure.

Frequency of visits to a health care provider, health promoting behaviors, and perceived health status among African American women.

The primary purpose of this study was to examine whether the self-reported number of health care visits over a 1-year period was associated with engagement in health promoting behaviors (i.e., healthy eating and physical activity) and perceived health status among a cross-sectional sample of African American women who were pre-hypertensive/hypertensive and/or overweight or obese (N = 180). The study participants were recruited in predominantly African American churches and had their data collected in April and May of 2009. Age, income, and education were also examined as moderators in the aforementioned relationships. Results revealed that the self-reported number of health care visits was significantly positively associated with healthy eating and perceived health status. Income moderated the relationship between self-reported number of health care visits and engagement in healthy eating. These results provide support for health promotion programs for African American women with program components that explain the relationships among routine care from a health care provider, engagement in health promoting behaviors, and prevention of chronic health conditions.

A Nonoligomerizing Mutant Form of Helicobacter pylori VacA Allows Structural Analysis of the p33 Domain.

Helicobacter pylori secretes a pore-forming VacA toxin that has structural features and activities substantially different from those of other known bacterial toxins. VacA can assemble into multiple types of water-soluble flower-shaped oligomeric structures, and most VacA activities are dependent on its capacity to oligomerize. The 88-kDa secreted VacA protein can undergo limited proteolysis to yield two domains, designated p33 and p55. The p33 domain is required for membrane channel formation and intracellular toxic activities, and the p55 domain has an important role in mediating VacA binding to cells. Previous studies showed that the p55 domain has a predominantly ß-helical structure, but no structural data are available for the p33 domain. We report here the purification and analysis of a nonoligomerizing mutant form of VacA secreted by H. pylori The nonoligomerizing 88-kDa mutant protein retains the capacity to enter host cells but lacks detectable toxic activity. Analysis of crystals formed by the monomeric protein reveals that the ß-helical structure of the p55 domain extends into the C-terminal portion of p33. Fitting the p88 structural model into an electron microscopy map of hexamers formed by wild-type VacA (predicted to be structurally similar to VacA membrane channels) reveals that p55 and the ß-helical segment of p33 localize to peripheral arms but do not occupy the central region of the hexamers. We propose that the amino-terminal portion of p33 is unstructured when VacA is in a monomeric form and that it undergoes a conformational change during oligomer assembly.

A structural model for binding of the serine-rich repeat adhesin GspB to host carbohydrate receptors.

GspB is a serine-rich repeat (SRR) adhesin of Streptococcus gordonii that mediates binding of this organism to human platelets via its interaction with sialyl-T antigen on the receptor GPIbα. This interaction appears to be a major virulence determinant in the pathogenesis of infective endocarditis. To address the mechanism by which GspB recognizes its carbohydrate ligand, we determined the high-resolution x-ray crystal structure of the GspB binding region (GspB(BR)), both alone and in complex with a disaccharide precursor to sialyl-T antigen. Analysis of the GspB(BR) structure revealed that it is comprised of three independently folded subdomains or modules: 1) an Ig-fold resembling a CnaA domain from prokaryotic pathogens; 2) a second Ig-fold resembling the binding region of mammalian Siglecs; 3) a subdomain of unique fold. The disaccharide was found to bind in a pocket within the Siglec subdomain, but at a site distinct from that observed in mammalian Siglecs. Confirming the biological relevance of this binding pocket, we produced three isogenic variants of S. gordonii, each containing a single point mutation of a residue lining this binding pocket. These variants have reduced binding to carbohydrates of GPIbα. Further examination of purified GspB(BR)-R484E showed reduced binding to sialyl-T antigen while S. gordonii harboring this mutation did not efficiently bind platelets and showed a significant reduction in virulence, as measured by an animal model of endocarditis. Analysis of other SRR proteins revealed that the predicted binding regions of these adhesins also had a modular organization, with those known to bind carbohydrate receptors having modules homologous to the Siglec and Unique subdomains of GspB(BR). This suggests that the binding specificity of the SRR family of adhesins is determined by the type and organization of discrete modules within the binding domains, which may affect the tropism of organisms for different tissues.

Influence of Maternal BLV Infection on miRNA and tRF Expression in Calves.

Small non-coding RNAs, such as microRNAs (miRNA) and tRNA-derived fragments (tRF), are known to be involved in post-transcriptional gene regulation. Research has provided evidence that small RNAs may influence immune development in calves. Bovine leukosis is a disease in cattle caused by Bovine Leukemia Virus (BLV) that leads to increased susceptibility to opportunistic pathogens. No research has addressed the potential influence that a maternal BLV infection may have on gene regulation through the differential expression of miRNAs or tRFs in progeny. Blood samples from 14-day old Holstein calves born to BLV-infected dams were collected. Antibodies for BLV were assessed using ELISA and levels of BLV provirus were assessed using qPCR. Total RNA was extracted from whole blood samples for small RNA sequencing. Five miRNAs (bta-miR-1, bta-miR-206, bta-miR-133a, bta-miR-133b, and bta-miR-2450d) and five tRFs (tRF-36-8JZ8RN58X2NF79E, tRF-20-0PF05B2I, tRF-27-W4R951KHZKK, tRF-22-S3M8309NF, and tRF-26-M87SFR2W9J0) were dysregulated in calves born to BLV-infected dams. The miRNAs appear to be involved in the gene regulation of immunological responses and muscle development. The tRF subtypes and parental tRNA profiles in calves born to infected dams appear to be consistent with previous publications in adult cattle with BLV infection. These findings offer insight into how maternal BLV infection status may impact the development of offspring.

No evidence for a negative association between bovine leukemia virus status and fertility in Kansas beef herds: a cross-sectional study.

OBJECTIVE: Determine the association between bovine leukemia virus (BLV) status and fertility in beef cows. BLV-status was defined using 3 different testing strategies (ELISA-, quantitative polymerase chain reaction- [qPCR], and high proviral load [PVL]-status). Fertility was defined as the overall probability of pregnancy as well as the probability of becoming pregnant in the first 21 days of the breeding season. ANIMALS: Convenience sample of 2,820 cows from 43 beef herds. PROCEDURES: The association of BLV-status with the probability of becoming pregnant was evaluated with a multivariable logistic regression analysis that used pregnancy status as a binary outcome, herd nested within ranch as a random effect, and BLV-status (ELISA-, qPCR-, and PVL-status as separate models) and potential covariates (eg, age, Body Condition Score [BCS] category, and interactions) as fixed effects. RESULTS: Raw data revealed that 55% (1,552/2,820) of cows were classified as BLV-positive by ELISA, and 95.3% (41/43) of herds had a least 1 ELISA-positive cow. Classification as BLV ELISA-positive was positively associated with the probability of being pregnant; however, when qPCR or PVL were used to classify BLV-status, there was no association with the probability of being pregnant. None of the methods of classifying BLV-status were associated with the probability of becoming pregnant in the first 21 days of the breeding season. CLINICAL RELEVANCE: This study did not find evidence that testing beef cows for BLV-status using ELISA, qPCR, or a cut-off of 0.9 PVL and removing test-positive cows will improve cowherd fertility as described by the probability of becoming pregnant during the breeding season or becoming pregnant during the first 21 days of the breeding season.

Red cell distribution width is a predictor of all-cause mortality in hospitalized dogs.

OBJECTIVE: To determine whether RBC distribution width (RDW) is associated with an increased odds of mortality in hospitalized dogs and cats. DESIGN: Retrospective, single-center study; data collected from 2007 to 2017. SETTING: University teaching hospital. ANIMALS: Six thousand six hundred and sixty-one animals (5183 dogs and 1478 cats). INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: Medical records were identified from animals presented to the emergency service and admitted to the ICU with a CBC and serum biochemistry performed on admission. Patients were stratified into quintiles based upon presenting RDW, and logistic regression modeling was performed to evaluate the relationship between RDW and in-hospital mortality. Canine patients with a presenting RDW in the upper fourth and fifth quintiles had an increased odds of all-cause in-hospital mortality (p < 0.0001). Specifically, dogs in the upper fifth quintile had a 2.1-fold greater odds of death compared to dogs in the first quintile, and dogs in the upper fourth quintile had a 1.9-fold greater odds of death compared to dogs in the first quintile. This relationship remained significant after adjusting for age, sex, key laboratory values excluding HCT, medical versus surgical disease, and diagnosis category. This relationship was no longer significant with the inclusion of HCT. No significant association was identified between presenting RDW and in-hospital mortality in cats. CONCLUSIONS: Hospitalized dogs with higher RDW on presentation to the emergency service have greater odds of all-cause in-hospital mortality compared to dogs with a lower RDW. A similar association between RDW and mortality was not found in cats. Further studies are warranted to assess the usefulness of this biomarker for specific diseases in dogs and to better understand the mechanisms driving the association between increased RDW and mortality in canine patients.

Identification of BoLA Alleles Associated with BLV Proviral Load in US Beef Cows.

Bovine leukemia virus (BLV) causes enzootic bovine leukosis, the most common neoplastic disease in cattle. Previous work estimates that 78% of US beef operations and 38% of US beef cattle are seropositive for BLV. Infection by BLV in a herd is an economic concern for producers as evidence suggests that it causes an increase in cost and a subsequent decrease in profit to producers. Studies investigating BLV in dairy cattle have noted disease resistance or susceptibility, measured by a proviral load (PVL) associated with specific alleles of the bovine leukocyte antigen (BoLA) DRB3 gene. This study aims to investigate the associations between BoLA DRB3 alleles and BLV PVL in beef cattle. Samples were collected from 157 Midwest beef cows. BoLA DRB3 alleles were identified and compared with BLV PVL. One BoLA DRB3 allele, *026:01, was found to be associated with high PVL in relation to the average of the sampled population. In contrast, two alleles, *033:01 and *002:01, were found to be associated with low PVL. This study provides evidence of a relationship between BoLA DRB3 alleles and BLV PVL in US beef cows.

Cross-sectional study to describe bovine leukemia virus herd and within-herd ELISA prevalence and bovine leukemia virus proviral load of convenience-sampled Kansas beef cow-calf herds.

OBJECTIVE: Determine bovine leukemia virus (BLV) seroprevalence of adult female cattle in Eastern Kansas beef herds and the proviral load (PVL) of those cattle found to be ELISA positive. ANIMALS: Convenience sample of 2,845 cows from 44 beef herds. PROCEDURES: BLV serostatus was determined using an ELISA antibody test (gp-51; IDEXX). BLV quantitative PCR (qPCR) status and PVL were determined utilizing a qPCR test (SS1 qPCR test; CentralStar Laboratories). The association of age, herd size, and body condition score (BCS) category on the probability of being BLV positive was evaluated with a multiple variable logistic regression analysis that used BLV status as a binary outcome, herd nested within ranch as a random effect, and BCS, herd size, and age category as fixed effects. RESULTS: Forty-two of 44 herds had at least 1 BLV ELISA-positive cow (95.5% herd seroprevalence). Overall, 1,564 of the 2,845 cows were BLV ELISA positive (55.0% individual animal prevalence). No association between BLV ELISA status and herd size or BCS was identified. When evaluated by age, the model-adjusted probability of being BLV ELISA positive was lowest for heifers (1 year of age, first parity) and increased until 5 to 6 years of age. Of the 1,564 ELISA-positive animals, 838 were qPCR positive (53.6%). The model-adjusted probability of being qPCR positive was not associated with age, herd size, or BCS category. CLINICAL RELEVANCE: This study indicated that BLV-seropositive status both as a herd classification and individual animal classification was very common in this population. Because the percentage of BLV-seropositive cows varied between herds and by age, this study provides evidence that it is essential for investigators to control for herd and age in any analysis of the association of BLV serostatus and health and production outcomes of interest. Some BLV ELSIA-seropositive cows were classified as BLV negative by qPCR, and risk factors may differ between classification status by ELISA and qPCR.

Phenotypic Selection of Dairy Cattle Infected with Bovine Leukemia Virus Demonstrates Immunogenetic Resilience through NGS-Based Genotyping of BoLA MHC Class II Genes.

Characterization of the bovine leukocyte antigen (BoLA) DRB3 gene has shown that specific alleles associate with susceptibility or resilience to the progression of bovine leukemia virus (BLV), measured by proviral load (PVL). Through surveillance of multi-farm BLV eradication field trials, we observed differential phenotypes within seropositive cows that persist from months to years. We sought to develop a multiplex next-generation sequencing workflow (NGS-SBT) capable of genotyping 384 samples per run to assess the relationship between BLV phenotype and two BoLA genes. We utilized longitudinal results from milk ELISA screening and subsequent blood collections on seropositive cows for PVL determination using a novel BLV proviral load multiplex qPCR assay to phenotype the cows. Repeated diagnostic observations defined two distinct phenotypes in our study population, ELISA-positive cows that do not harbor detectable levels of provirus and those who do have persistent proviral loads. In total, 565 cows from nine Midwest dairy farms were selected for NGS-SBT, with 558 cows: 168 BLV susceptible (ELISA-positive/PVL-positive) and 390 BLV resilient (ELISA-positive/PVL-negative) successfully genotyped. Three BoLA-DRB3 alleles, including one novel allele, were shown to associate with disease resilience, *009:02, *044:01, and *048:02 were found at rates of 97.5%, 86.5%, and 90.3%, respectively, within the phenotypically resilient population. Alternatively, DRB3*015:01 and *027:03, both known to associate with disease progression, were found at rates of 81.1% and 92.3%, respectively, within the susceptible population. This study helps solidify the immunogenetic relationship between BoLA-DRB3 alleles and BLV infection status of these two phenotypic groupings of US dairy cattle.

Accuracies of genomic breeding values in American Angus beef cattle using K-means clustering for cross-validation.

BACKGROUND: Genomic selection is a recently developed technology that is beginning to revolutionize animal breeding. The objective of this study was to estimate marker effects to derive prediction equations for direct genomic values for 16 routinely recorded traits of American Angus beef cattle and quantify corresponding accuracies of prediction. METHODS: Deregressed estimated breeding values were used as observations in a weighted analysis to derive direct genomic values for 3570 sires genotyped using the Illumina BovineSNP50 BeadChip. These bulls were clustered into five groups using K-means clustering on pedigree estimates of additive genetic relationships between animals, with the aim of increasing within-group and decreasing between-group relationships. All five combinations of four groups were used for model training, with cross-validation performed in the group not used in training. Bivariate animal models were used for each trait to estimate the genetic correlation between deregressed estimated breeding values and direct genomic values. RESULTS: Accuracies of direct genomic values ranged from 0.22 to 0.69 for the studied traits, with an average of 0.44. Predictions were more accurate when animals within the validation group were more closely related to animals in the training set. When training and validation sets were formed by random allocation, the accuracies of direct genomic values ranged from 0.38 to 0.85, with an average of 0.65, reflecting the greater relationship between animals in training and validation. The accuracies of direct genomic values obtained from training on older animals and validating in younger animals were intermediate to the accuracies obtained from K-means clustering and random clustering for most traits. The genetic correlation between deregressed estimated breeding values and direct genomic values ranged from 0.15 to 0.80 for the traits studied. CONCLUSIONS: These results suggest that genomic estimates of genetic merit can be produced in beef cattle at a young age but the recurrent inclusion of genotyped sires in retraining analyses will be necessary to routinely produce for the industry the direct genomic values with the highest accuracy.

Tracing Viral Transmission and Evolution of Bovine Leukemia Virus through Long Read Oxford Nanopore Sequencing of the Proviral Genome.

Bovine leukemia virus (BLV) causes Enzootic Bovine Leukosis (EBL), a persistent life-long disease resulting in immune dysfunction and shortened lifespan in infected cattle, severely impacting the profitability of the US dairy industry. Our group has found that 94% of dairy farms in the United States are infected with BLV with an average in-herd prevalence of 46%. This is partly due to the lack of clinical presentation during the early stages of primary infection and the elusive nature of BLV transmission. This study sought to validate a near-complete genomic sequencing approach for reliability and accuracy before determining its efficacy in characterizing the sequence identity of BLV proviral genomes collected from a pilot study made up of 14 animals from one commercial dairy herd. These BLV-infected animals were comprised of seven adult dam/daughter pairs that tested positive by ELISA and qPCR. The results demonstrate sequence identity or divergence of the BLV genome from the same samples tested in two independent laboratories, suggesting both vertical and horizontal transmission in this dairy herd. This study supports the use of Oxford Nanopore sequencing for the identification of viral SNPs that can be used for retrospective genetic contact tracing of BLV transmission.

Purification, crystallization and preliminary X-ray diffraction analysis of the carbohydrate-binding region of the Streptococcus gordonii adhesin GspB.

The carbohydrate-binding region of the bacterial adhesin GspB from Streptococcus gordonii strain M99 (GspB(BR)) was expressed in Escherichia coli and purified using affinity and size-exclusion chromatography. Separate sparse-matrix screening of GspB(BR) buffered in either 20 mM Tris pH 7.4 or 20 mM HEPES pH 7.5 resulted in different crystallographic behavior such that different precipitants, salts and additives supported crystallization of GspB(BR) in each buffer. While both sets of conditions supported crystal growth in space group P2(1)2(1)2(1), the crystals had distinct unit-cell parameters of a = 33.3, b = 86.7, c = 117.9 Šfor crystal form 1 and a = 34.6, b = 98.3, c = 99.0 Šfor crystal form 2. Additive screening improved the crystals grown in both conditions such that diffraction extended to beyond 2 Šresolution. A complete data set has been collected to 1.3 Šresolution with an overall R(merge) value of 0.04 and an R(merge) value of 0.33 in the highest resolution shell.

Current Developments in the Epidemiology and Control of Enzootic Bovine Leukosis as Caused by Bovine Leukemia Virus.

Enzootic Bovine Leukosis (EBL) caused by the bovine leukemia virus (BLV) has been eradicated in over 20 countries. In contrast, the U.S. and many other nations are experiencing increasing prevalence in the absence of efforts to control transmission. Recent studies have shown that BLV infection in dairy cattle has a greater impact beyond the long-recognized lymphoma development that occurs in <5% of infected cattle. Like other retroviruses, BLV appears to cause multiple immune system disruptions, affecting both cellular and humoral immunity, which are likely responsible for increasingly documented associations with decreased dairy production and decreased productive lifespan. Realization of these economic losses has increased interest in controlling BLV using technology that was unavailable decades ago, when many nations eradicated BLV via traditional antibody testing and slaughter methods. This traditional control is not economically feasible for many nations where the average herd antibody prevalence is rapidly approaching 50%. The ELISA screening of cattle with follow-up testing via qPCR for proviral load helps prioritize the most infectious cattle for segregation or culling. The efficacy of this approach has been demonstrated in at least four herds. Breeding cattle for resistance to BLV disease progression also appears to hold promise, and several laboratories are working on BLV vaccines. There are many research priorities for a wide variety of disciplines, especially including the need to investigate the reports linking BLV and human breast cancer.