Retrospective cohort study investigating the relationship between diarrhea during the preweaning period and subsequent survival, health and production in dairy cows.

The primary objective of this study was to evaluate the association between preweaning diarrhea and measures of survival, health and production. The measures of interest included survival (mortality before first calving and time to removal from the herd), health (retained fetal membranes, metritis, mastitis and somatic cell count), and production (305d mature equivalent milk yield and time to first calving). A secondary objective was to investigate if these associations varied according to the age of the calf when the case of diarrhea occurred. Herd records from a farm located in Southern Australia were used to conduct a retrospective cohort study where subjects (calves) with diarrhea were enrolled at the time of their first case along with 2 subjects without diarrhea, matched for age (±3 d) and date of birth (±15 d), amounting to 9833 calves in the data set. Survival analysis was conducted to determine if preweaning diarrhea was associated with death in the short term (first 20 d after enrollment), medium term (21 to 100 d after enrollment) and long-term (101 d after enrollment to first calving). Crude incidence rate ratios, Kaplan-Meier curves and hazard ratios (HR, Cox regression) were derived for each event-based outcome (e.g., death, calving, mastitis). Multivariable linear models were used for continuous outcomes. Calves with preweaning diarrhea had greater mortality rates in the short term (hazard ratio, HR = 2.48, 95% CI: 1.87-3.29) and medium term (HR = 1.89, 95% CI: 1.41-2.55) but not in the long term (HR = 0.98, 95% CI: 0.79-1.22). A small negative association between preweaning diarrhea and time to first calving was found, with calves with diarrhea calving 4 d later than calves without diarrhea (HR = 0.95, 95% CI: 0.91-1.00). Rates of post-calving removal from the herd (death or culling) were higher in calves with a history of preweaning diarrhea (HR = 1.13, 95% CI: 1.01 to 1.26), as were peak lactation log somatic cell count (+0.08, 95% CI: 0.02 to 0.14). Further research is needed to replicate these novel findings as they are based on exploratory analyses and could be spurious findings. No substantial associations were observed for the other measures of interest. Our study findings support existing research demonstrating the significance of preweaning diarrhea as a significant cause of calf mortality, and raises new hypotheses about other potential impacts during lactation.

Etiology and epidemiology of digital dermatitis in Australian dairy herds.

Bovine digital dermatitis (BDD) is an important cause of lameness in dairy cows worldwide. However, very little is known about this disease in Australian herds, which are predominantly managed on pasture. The primary objectives of this cross-sectional study were to describe the presence and prevalence of BDD in Australian dairy herds and to characterize the microbiota of healthy skin and M4 lesions of BDD-affected, pasture-managed cows. Cows from 71 dairy herds were examined at milking time to identify the presence of BDD lesions. True prevalence was estimated using Bayesian methods with informative priors for sensitivity and specificity. Biopsy samples (n = 60) were collected from cows with and without BDD lesions in 7 pasture-based herds. The microbiota in the superficial and deep strata of each tissue biopsy were characterized via sequencing of the V3-V4 region of the bacterial ribosomal RNA gene. Lesions were detected in 1,817 (11.5%) of 15,813 cows and in 68 of 71 (95.8%) herds. The median herd-level apparent and true prevalences of BDD were 8.5% and 18.1%, respectively, but prevalences varied considerably between farms. On farms with BDD, M4 lesions accounted for 70% to 100% of all lesions (interquartile range = 95.1%-100%, median = 100%); M2 lesions (i.e., large ulcerative lesions) were observed at low prevalence (<2.2%) in the few herds (7/71, 9.9%) where they were found. There was a significant difference in the composition of the microbiota between healthy skin and M4 lesions but not between superficial and deep tissue layers. Several gut- and effluent-associated bacterial taxa, including Lentimicrobium and Porphyromonas, which have previously been associated with BDD, were abundant in BDD lesions but not in control biopsies. Our study supports the idea that such taxa are involved in, although possibly not essential to, lesion development and persistence in pasture-managed cows in Australia. Our results also suggest that Dichelobacter may contribute to the disease process. We conclude that BDD is likely to occur in most Australian dairy farms, but that further studies are needed to identify its effect on cow welfare and productivity. Further investigation of the etiology of BDD in Australian dairy herds is also necessary to inform prevention and control strategies.

Evaluation of point-of-care tests for identification of pathogens to inform clinical mastitis treatment decisions in pasture- and confinement-managed dairy cows in Australia.

To support antimicrobial stewardship in livestock production, there is a growing array of point-of-care diagnostics to guide antimicrobial treatment. The primary objective of this observational study was to evaluate the diagnostic performance of 5 point-of-care tests currently available in Australia for guiding lactational treatment of nonsevere clinical mastitis. A secondary objective was to describe the pathogen profiles of mastitis-causing organisms in cows managed in barns ("intensive") and on pasture ("nonintensive"). Foremilk samples (n = 641) were collected by farm staff in dairy herds in Australia (n = 30) and tested at a university laboratory using a reference test and 5 index tests. The reference test was aerobic culture on trypticase soy agar with 5% sheep blood followed by MALDI-TOF for identification of isolates. The following point-of-care tests were evaluated as index tests: Accumast, biplate, Check-Up, Mastatest (with AUP3-2022 product), and 3M Petrifilm. We found that 23% of samples were contaminated, with the median herd contamination prevalence being 22%. After excluding contaminated samples, the most common diagnoses (according to the reference test) in intensive herds were no growth (31.7%), Klebsiella spp. (28.1%), Escherichia coli (15.0%), and Streptococcus uberis (8.4%). The most common diagnoses in noncontaminated samples from cows in nonintensive herds were Strep. uberis (35.0%), no growth (26.9%), and E. coli (13.3%). After 24 h of incubation, all index tests demonstrated limited diagnostic sensitivity for identification of pathogens of interest (range: 0.06-0.63). Diagnostic performance was better at the group-level, with sensitivity and specificity for identification of noncontaminated gram-positive growths (i.e., cases that are widely considered to be candidates for antimicrobial treatment) being 0.84 and 0.75, respectively (biplate), 0.76 and 0.90 (Accumast), 0.89 and 0.79 (Check-Up), 0.67 and 0.83 (Petrifilm), and 0.55 and 0.81 (Mastatest). In intensive herds, 22.7% to 40% of cases were classified as antimicrobial treatment candidates by index tests, which was less than for cows in nonintensive herds (41.3%-61.0%). Despite limited diagnostic reliability at the genus level and species level, and the need to ensure samples are collected aseptically, our findings indicate that implementation of selective treatment protocols using the tests evaluated in this study would likely reduce antimicrobial usage in Australian herds.

Selective dry-cow therapy can be implemented successfully in cows of all milk production levels.

Antibiotic stewardship on dairy farms can be heightened through the implementation of selective dry-cow therapy (SDCT). However, some producers are concerned that this practice may be related to poor udder health outcomes in cows with high milk production at the time of dry-off. The objective of this study was to evaluate if the effect of culture-guided SDCT (Cult-SDCT) and algorithm-guided SDCT (Alg-SDCT) on dry-period intramammary infection (IMI) dynamics and postcalving udder health and performance [when compared with blanket dry-cow therapy (BDCT)] varied according to milk production level before dry-off. Data were compiled from clinical trials conducted in the United States and Canada that compared Cult-SDCT and Alg-SDCT to a positive control, i.e., BDCT. In those trials, cows were enrolled 1-2 d before dry-off, randomized to their dry-cow therapy strategy and followed until 120 d in milk of the subsequent lactation. The number of cows and quarters in the final data set were 1,485 and 5,097, respectively. Measured outcomes included quarter-level antibiotic use at dry-off, quarter-level IMI prevalence after calving, quarter-level dry-period IMI cure risk, quarter-level dry-period new IMI risk, cow-level clinical mastitis and removal from the herd during 1-120 d in milk, and somatic cell count and milk yield during 1-120 DIM. The primary objective of analysis was to investigate if the effect of Cult-SDCT and Alg-SDCT on these outcomes, when compared with BDCT, varied according to milk production level before dry-off. To do this, each cow was classified as having low, mid or high production, based on her milk yield tertile group at the most recent herd test before enrollment (low: <23.7 kg/d, mid: 23.7 to 30.4 kg/d, and high >30.4 kg/d). Multivariable generalized estimating equations were used to estimate risk differences and differences in means, and Cox regression was used to estimate hazard ratios. For Cult-SDCT, the proportion of quarters treated with dry-cow antibiotics within each milk production level were 40.7% (low), 41.7% (mid) and 47.2% (high). For Alg-SDCT, the proportions were 60.6% (low), 38.7% (mid), and 35.1% (high). Measures of udder health were not markedly different when comparing Cult-SDCT to BDCT and Alg-SDCT to BDCT. This was consistently observed in low, mid and high producing cows. In conclusion, the findings from this study indicate that Cult-SDCT and Alg-SDCT can be successfully implemented in cows of all milk production levels.

Negatively controlled trial investigating the effects of dry cow therapy on clinical mastitis and culling in multiparous cows.

Blanket dry cow therapy (DCT) is a major contributor to overall antibiotic usage on dairy farms in the United States. With low prevalence of intramammary infections at dry-off in US herds today, alternative DCT approaches have been the focus of much research. We hypothesized that complete cessation of DCT [i.e., use of internal teat sealants (ITS) only at dry-off] could be a practical alternative to blanket DCT in well-managed herds. The objective of this negatively controlled clinical trial was to determine the effects of DCT on clinical mastitis (CM) and removal from the herd during the dry period and the first 200 d of the subsequent lactation in multiparous dairy cows treated with only ITS at dry-off. As a secondary objective, we conducted exploratory analysis to identify subpopulations in the herd (based on parity, previous CM history, and dry-period length) where DCT would not affect postcalving udder health, to generate hypotheses about potential alternative selective DCT programs. The study was conducted in a commercial dairy herd in South Dakota from June 2020 to January 2021. Dry-off sessions (n = 43) were scheduled such that all cows at a given session were dried off using ITS alone (ITS only, n = 20 sessions, n = 1,108 cows) or an intramammary DCT product containing 500 mg of cloxacillin (Dry-Clox, Boehringer Ingelheim) followed by ITS (ITS+ABX, n = 23 sessions, n = 1,331 cows). Culling and CM events were recorded by farm workers who were blinded to the treatment status of cows. Hazard ratios (HR) for the effects of the treatment group on CM and removal from the herd were estimated using multivariable Cox proportional hazards, adjusting for the clustered treatment allocation strategy. Risk of removal from the herd during the dry period was lower in ITS+ABX than ITS-only cows (1.1 vs. 2.7%; HR = 0.45; 95% CI: 0.25 to 0.81). Risk of removal from the herd during the first 200 d of lactation was similar in ITS+ABX and ITS-only cows (17.3 vs. 18.0%; HR = 0.98; 95% CI: 0.82 to 1.18). Risk of CM during the first 200 d of lactation was lower in ITS+ABX cows (6.9%; HR = 0.56; 95% CI: 0.41 to 0.76) compared with ITS-only cows (13.4%). The beneficial effects of DCT on CM and removal from the herd were consistently observed across strata of parity, previous CM history, and dry-period length, indicating that no subpopulations could be identified to withhold DCT. The findings from this study indicate that the omission of DCT from the dry-off procedure, when udder health is not taken into consideration, in multiparous cows can have a negative effect on cow health and welfare. Findings from previous research suggest that culture- or algorithm-guided selective dry cow therapy are likely to be safer approaches to improving antibiotic stewardship.

Correction: The multifaceted nature of antimicrobial peptides: current synthetic chemistry approaches and future directions.

Correction for 'The multifaceted nature of antimicrobial peptides: current synthetic chemistry approaches and future directions' by Bee Ha Gan et al., Chem. Soc. Rev., 2021, 50, 7820-7880, DOI: 10.1039/D0CS00729C.

Invited review: Selective use of antimicrobials in dairy cattle at drying-off.

Administering intramammary antimicrobials to all mammary quarters of dairy cows at drying-off [i.e., blanket dry cow therapy (BDCT)] has been a mainstay of mastitis prevention and control. However, as udder health has considerably improved over recent decades with reductions in intramammary infection prevalence at drying-off and the introduction of teat sealants, BDCT may no longer be necessary on all dairy farms, thereby supporting antimicrobial stewardship efforts. This narrative review summarizes available literature regarding current dry cow therapy practices and associated impacts of selective dry cow therapy (SDCT) on udder health, milk production, economics, antimicrobial use, and antimicrobial resistance. Various methods to identify infections at drying-off that could benefit from antimicrobial treatment are described for selecting cows or mammary quarters for treatment, including utilizing somatic cell count thresholds, pathogen identification, previous clinical mastitis history, or a combination of criteria. Selection methods may be enacted at the herd, cow, or quarter levels. Producers' and veterinarians' motivations for antimicrobial use are discussed. Based on review findings, SDCT can be adopted without negative consequences for udder health and milk production, and concurrent teat sealant use is recommended, especially in udder quarters receiving no intramammary antimicrobials. Furthermore, herd selection should be considered for SDCT implementation in addition to cow or quarter selection, as BDCT may still be temporarily necessary in some herds for optimal mastitis control. Costs and benefits of SDCT vary among herds, whereas impacts on antimicrobial resistance remain unclear. In summary, SDCT is a viable management option for maintaining udder health and milk production while improving antimicrobial stewardship in the dairy industry.

The role of chemical synthesis in developing RiPP antibiotics.

The growing antimicrobial resistance crisis necessitates the discovery and development of novel classes of antibiotics if a 'postantibiotic era' is to be avoided. Ribosomally synthesised and post-translationally modified peptides, or RiPPs, are becoming increasingly recognised as a potential source of antimicrobial drugs. This is due to a combination of their potent antimicrobial activity and their high stability relative to unmodified linear peptides. However, as peptide drugs, their clinical development is often perturbed by issues such as low solubility and poor bioavailability. Chemical synthesis has the potential to overcome some of these challenges. Furthermore, the structural complexity of RiPPs makes them interesting synthetic targets in their own right, with the total synthesis of some structural classes having only been recently realised. This review focusses on the use of RiPPs as antimicrobial agents and will highlight various strategies that have been employed to chemically synthesise three major classes of RiPPs: lasso peptides, cyclotides, and lanthipeptides.

The multifaceted nature of antimicrobial peptides: current synthetic chemistry approaches and future directions.

Bacterial infections caused by 'superbugs' are increasing globally, and conventional antibiotics are becoming less effective against these bacteria, such that we risk entering a post-antibiotic era. In recent years, antimicrobial peptides (AMPs) have gained significant attention for their clinical potential as a new class of antibiotics to combat antimicrobial resistance. In this review, we discuss several facets of AMPs including their diversity, physicochemical properties, mechanisms of action, and effects of environmental factors on these features. This review outlines various chemical synthetic strategies that have been applied to develop novel AMPs, including chemical modifications of existing peptides, semi-synthesis, and computer-aided design. We will also highlight novel AMP structures, including hybrids, antimicrobial dendrimers and polypeptides, peptidomimetics, and AMP-drug conjugates and consider recent developments in their chemical synthesis.

Longitudinal study of herd udder hygiene and its association with clinical mastitis in pasture-based dairy cows.

The objectives of this exploratory study were to (1) describe the association between herd-level udder hygiene and clinical mastitis and (2) investigate how sample size and milking stage affect the accuracy and precision of herd udder hygiene assessments made at milking time. A prospective longitudinal study was conducted in a dairy herd in Northern Australia as part of a previously published clinical trial of premilking teat disinfection. Video footage from 35 afternoon milkings was used to conduct 12,544 udder hygiene scores from 504 cows during an 89-d period and measure udder hygiene of the herd (proportion of cows with udder hygiene ≥3 out of 4). Linear interpolation was used to estimate herd udder hygiene on the days that were not scored, such that a herd-level udder hygiene measure was available for all cow-days in the study. Clinical mastitis events occurring during the study period were detected and recorded by farm staff according to a standardized definition. The relationship between herd udder hygiene on each of 1, 2, and 3 d before each study day (d -1, -2, and -3, respectively) and clinical mastitis at the cow level on each study day (each in turn being set as d 0) was determined using multivariable generalized estimating equations (family = Poisson, link = log), with the unit of analysis being the cow-day, adjusting for potential confounders and the clustering within the data. In addition, sampling strategies were evaluated by simulating herd udder hygiene assessments using a subset of cows in the herd. Herd udder hygiene from d -1, -2, and -3 was positively associated with clinical mastitis on d 0 (incidence rate ratio = 1.4 per 10-point increase in the percentage of cows with poor udder hygiene). Sampling strategy simulation found that at least 80 cows needed to be scored to achieve sufficiently precise estimations of herd udder hygiene. Furthermore, cows scored later during milking were slightly more likely to have poor udder hygiene than those scored earlier (risk ratio = 1.02 for cows that were 10% later in the milking order). More research is needed to evaluate risk factors for poor udder hygiene and potential interventions in pasture-based dairy cows.

Quantum dot interactions with and toxicity to Shewanella oneidensis MR-1.

Combining abiotic photosensitisers such as quantum dots (QDs) with non-photosynthetic bacteria presents an intriguing concept into the design of artificial photosynthetic organisms and solar-driven fuel production. Shewanella oneidensis MR-1 (MR-1) is a versatile bacterium concerning respiration, metabolism and biocatalysis, and is a promising organism for artificial photosynthesis as the bacterium's synthetic and catalytic ability provides a potential system for bacterial biohydrogen production. MR-1's hydrogenases are present in the periplasmatic space. It follows that for photoenergised electrons to reach these enzymes, QDs will need to be able to enter the periplasm, or electrons need to enter the periplasm via the Mtr pathway that is responsible for MR-1's extracellular electron transfer ability. As a step towards this goal, various QDs were tested for their photo-reducing potential, nanotoxicology and further for their interaction with MR-1. CdTe/CdS/TGA, CdTe/CdS/Cysteamine, a commercial, negatively charged CdTe and CuInS2/ZnS/PMAL QDs were examined. The photoreduction potential of the QDs was confirmed by measuring their ability to photoreduce methyl viologen with different sacrificial electron donors. The commercial CdTe and CuInS2/ZnS/PMAL QDs showed no toxicity towards MR-1 as evaluated by a colony-forming units method and a fluorescence viability assay. Only the commercial negatively charged CdTe QDs showed good interaction with MR-1. With transmission electron microscopy, QDs were observed both in the cytoplasm and periplasm. These results inform on the possibilities and bottlenecks when developing bionanotechnological systems for the photosynthetic production of biohydrogen by MR-1.

Electron Accepting Units of the Diheme Cytochrome c TsdA, a Bifunctional Thiosulfate Dehydrogenase/Tetrathionate Reductase.

The enzymes of the thiosulfate dehydrogenase (TsdA) family are wide-spread diheme c-type cytochromes. Here, redox carriers were studied mediating the flow of electrons arising from thiosulfate oxidation into respiratory or photosynthetic electron chains. In a number of organisms, including Thiomonas intermedia and Sideroxydans lithotrophicus, the tsdA gene is immediately preceded by tsdB encoding for another diheme cytochrome. Spectrophotometric experiments in combination with enzymatic assays in solution showed that TsdB acts as an effective electron acceptor of TsdA in vitro when TsdA and TsdB originate from the same source organism. Although TsdA covers a range from -300 to +150 mV, TsdB is redox active between -100 and +300 mV, thus enabling electron transfer between these hemoproteins. The three-dimensional structure of the TsdB-TsdA fusion protein from the purple sulfur bacterium Marichromatium purpuratum was solved by X-ray crystallography to 2.75 Å resolution providing insights into internal electron transfer. In the oxidized state, this tetraheme cytochrome c contains three hemes with axial His/Met ligation, whereas heme 3 exhibits the His/Cys coordination typical for TsdA active sites. Interestingly, thiosulfate is covalently bound to Cys330 on heme 3. In several bacteria, including Allochromatium vinosum, TsdB is not present, precluding a general and essential role for electron flow. Both AvTsdA and the MpTsdBA fusion react efficiently in vitro with high potential iron-sulfur protein from A. vinosum (Em +350 mV). High potential iron-sulfur protein not only acts as direct electron donor to the reaction center in anoxygenic phototrophs but can also be involved in aerobic respiratory chains.

Asymmetric Syntheses of (+)-Preussin B, the C(2)-Epimer of (-)-Preussin B, and 3-Deoxy-(+)-preussin B.

Efficient de novo asymmetric syntheses of (+)-preussin B, the C(2)-epimer of (-)-preussin B, and 3-deoxy-(+)-preussin B have been developed, using the diastereoselective conjugate addition of lithium (S)-N-benzyl-N-(α-methylbenzyl)amide to tert-butyl 4-phenylbut-2-enoate and diastereoselective reductive cyclizations of γ-amino ketones as the key steps to set the stereochemistry. Conjugate addition followed by enolate protonation generated the corresponding ß-amino ester. Homologation using the ester functionality as a synthetic handle gave the corresponding γ-amino ketone. Hydrogenolytic N-debenzylation was accompanied by diastereoselective reductive cyclization in situ; reductive N-methylation then gave 3-deoxy-(+)-preussin B as the major diastereoisomeric product. Meanwhile, the same conjugate addition but followed by enolate oxidation with (+)-camphorsulfonyloxaziridine gave the corresponding anti-α-hydroxy-ß-amino ester. α-Epimerization by oxidation and diastereoselective reduction then gave access to the corresponding syn-α-hydroxy-ß-amino ester. Homologation of both of these diastereoisomeric α-hydroxy-ß-amino esters gave the corresponding ß-hydroxy-γ-amino ketones. N-Debenzylation and concomitant diastereoselective reductive cyclization, followed by reductive N-methylation, provided the C(2)-epimer of (-)-preussin B and (+)-preussin B as the major diastereoisomeric products, respectively. The overall yields (from phenylacetaldehyde) were 19% for 3-deoxy-(+)-preussin B over seven steps, 8% for the C(2)-epimer of (-)-preussin B over nine steps, and 7% for (+)-preussin B over eleven steps.

Synthesis of Cyclopropanes via Hydrogen-Borrowing Catalysis.

Cyclopropanes are highly useful motifs that are often incorporated into drug candidates to improve potency, metabolic stability, or pharmacokinetic properties. An expedient method for the α-cyclopropanation of ketones using hydrogen borrowing (HB) catalysis is described. The transformation occurs via HB alkylation of a hindered ketone with subsequent intramolecular displacement of a pendant leaving group affording the cyclopropanated product. The leaving group can be installed in either the ketone or alcohol component of the HB system, giving access to α-cyclopropyl ketones via two complementary approaches. Conversion to the corresponding carboxylic acids can be achieved in a simple two-step sequence to afford synthetically useful 1,1-substituted spirocyclopropyl acid building blocks.

Can apps be used to formulate fluid therapy plans in veterinary medicine?

BACKGROUND: Formulating sophisticated fluid therapy plans can be complicated and time consuming. Consequently, veterinarians in the field who lack experience, time, or confidence may formulate suboptimal fluid therapy plans. OBJECTIVES: Compare conventional and app-guided fluid therapy plans for simulated cases of neonatal calf diarrhea. PARTICIPANTS: Third and fourth year veterinary students (n = 55) from The University of Sydney. METHODS: We developed a web app to assist fluid therapy formulation (http://calfaid.com) that was evaluated in a randomized case simulation trial. Participants were instructed to perform fluid therapy calculations and formulate an integrated fluid therapy plan for case scenarios using conventional methods and using the fluid therapy app. Responses were scored by a blinded study investigator using an a priori scoring guide and groups (conventional vs. app-guided) were compared using linear mixed models. RESULTS: On average, total scores for app-guided fluid therapy calculations were 20.6% points higher (95% confidence interval [CI], 14.1-27.1) than calculations completed using the conventional method (88.2% vs. 67.5%, respectively). On average, total scores for app-guided integrated fluid therapy plans were 14.2% points higher (95% CI, 6.3-22.2; 65.8% vs. 51.2%). Eighty percent of respondents indicated they would prefer to use the app-guided method over the conventional method. CONCLUSION AND CLINICAL IMPORTANCE: Our findings suggest that fluid therapy plans can be improved using apps.

The microbiome of common bedding materials before and after use on commercial dairy farms.

BACKGROUND: Bovine mastitis is one of the most economically important diseases affecting dairy cows. The choice of bedding material has been identified as an important risk factor contributing to the development of mastitis. However, few reports examine both the culturable and nonculturable microbial composition of commonly used bedding materials, i.e., the microbiome. Given the prevalence of nonculturable microbes in most environments, this information could be an important step to understanding whether and how the bedding microbiome acts as a risk factor for mastitis. Therefore, our objective was to characterize the microbiome composition and diversity of bedding material microbiomes, before and after use. METHODS: We collected 88 bedding samples from 44 dairy farms in the U.S. Unused (from storage pile) and used (out of stalls) bedding materials were collected from four bedding types: new sand (NSA), recycled manure solids (RMS), organic non-manure (ON) and recycled sand (RSA). Samples were analyzed using 16S rRNA sequencing of the V3-V4 region. RESULTS: The overall composition as well as the counts of several microbial taxa differed between bedding types, with Proteobacteria, Actinobacteria, Bacteroidetes and Firmicutes dominating across all types. Used bedding contained a significantly different microbial composition than unused bedding, but the magnitude of this difference varied by bedding type, with RMS bedding exhibiting the smallest difference. In addition, positive correlations were observed between 16S rRNA sequence counts of potential mastitis pathogens (bacterial genera) and corresponding bedding bacterial culture data. CONCLUSION: Our results strengthen the role of bedding as a potential source of mastitis pathogens. The consistent shift in the microbiome of all bedding types that occurred during use by dairy cows deserves further investigation to understand whether this shift promotes pathogen colonization and/or persistence, or whether it can differentially impact udder health outcomes. Future studies of bedding and udder health may be strengthened by including a microbiome component to the study design.

Evaluation of three immunological assays to mitigate the risk of transboundary spread of Coxiella burnetii by alpacas.

Coxiella burnetii causes coxiellosis in animals and Q fever in humans, a potentially debilitating zoonotic disease commonly transmitted through domestic ruminants. To prevent transboundary spread of C. burnetii, animals may be tested prior to export. In alpacas, this process is complicated by the lack of scientific evidence for C. burnetii infection in the species, and the unique composition of camelid antibodies, which may cause false-positive results in assays developed for ruminants. We evaluated a complement fixation test (CFT; currently recommended for alpacas in New Zealand), an enzyme-linked immunosorbent assay (ELISA) and an immunofluorescence assay (IFA). Positive analytical control samples were generated through vaccination of alpacas with a human Q fever vaccine, whereas negative analytical control samples were sourced from New Zealand (deemed free of C. burnetii). Immunological assays were conducted on 131 alpaca sera submitted for export testing. Test characteristics (sensitivity, specificity, positive and negative predictive values) for CFT, ELISA and IFA were determined using Bayesian latent class analysis. Due to anticomplementary activity, 37 (28.2%) of the CFT results were inconclusive, making CFT unsuitable for routine use. Of the remaining 94 samples, 10.6%, 0% and 7.4% were positive for C. burnetii antibodies based on CFT, ELISA and IFA, respectively, yielding estimated sensitivities of 58%, 26% and 78%, and estimated specificities of 95%, 98% and 95%, with the estimates for sensitivity being imprecise, as evidenced by wide 95% credible intervals. Positive predictive values were similar across assays, albeit very low at the estimated seroprevalence of 5%. Our results indicate that, of the tests available, IFA appears to be the most appropriate for use in alpacas. Higher sensitivity of antibody detection, use of antigen detection assays and availability of samples from individuals with evidence of infection could provide additional insight into the risk of transboundary spread of C. burnetii by alpacas.

Diagnosing Intramammary Infection: Meta-Analysis and Mapping Review on Frequency and Udder Health Relevance of Microorganism Species Isolated from Bovine Milk Samples.

Matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry provides accurate species-level identification of many, microorganisms retrieved from bovine milk samples. However, not all those microorganisms are pathogenic. Our study aimed to: (1) determine the species-specific prevalence of microorganisms identified in bovine milk of apparently healthy lactating quarters vs. quarters with clinical mastitis (CM); and (2) map current information and knowledge gaps on udder health relevance of microorganisms retrieved from bovine milk samples. A mixed study design (meta-analysis and mapping review) was chosen. We gathered several large Canadian, US and Brazilian data sets of MALDI-TOF results for organisms cultured from quarter milk samples. For meta-analysis, two datasets (apparently healthy quarters vs. CM samples) were organized. A series of meta-analyses was conducted to determine microorganisms' prevalence. Then, each species reported was searched through PubMed to investigate whether inflammation (increased somatic cell count (SCC) or signs of CM) was associated with microorganism's recovery from milk. A total of 294 different species of microorganisms recovered from milk samples were identified. Among 50,429 quarter-milk samples from apparently healthy quarters, the 5 most frequent species were Staphylococcus chromogenes (6.7%, 95% CI 4.5-9.2%), Aerococcus viridans (1.6%, 95% CI 0.4-3.5%), Staphylococcus aureus (1.5%, 95% CI 0.5-2.8%), Staphylococcus haemolyticus (0.9%, 95% CI 0.4-1.5%), and Staphylococcus epidermidis (0.7%, 95% CI 0.2-1.6%). Among the 43,924 quarter-milk CM samples, the 5 most frequent species were Escherichia coli (11%, 95% CI 8.1-14.3%), Streptococcus uberis (8.5%, 95% CI 5.3-12.2%), Streptococcus dysgalactiae (7.8%, 95% CI 4.9-11.5%), Staphylococcus aureus (7.8%, 95% CI 4.4-11.9%), and Klebsiella pneumoniae (5.6%, 95% CI 3.4-8.2%). When conducting the PubMed literature search, there were 206 species identified by MALDI-TOF for which we were not able to find any information regarding their association with CM or SCC. Some of them, however, were frequently isolated in our multi-country dataset from the milk of quarters with CM (e.g., Citrobacter koseri, Enterococcus saccharolyticus, Streptococcus gallolyticus). Our study provides guidance to veterinarians for interpretation of milk bacteriology results obtained using MALDI-TOF and identifies knowledge gaps for future research.

Micronutrients and respiratory infections: the biological rationale and current state of clinical evaluation.

A range of nutrients has been studied or proposed for use in preventing respiratory tract infections and reducing their severity. This article gives a narrative review of the existing literature, biological rationales and current state of clinical evaluation for micronutrient therapies. The importance of vitamin A, the B vitamins, vitamin C, vitamin D, eicosapentaenoic acid, vitamin E, selenium, zinc and a range of combination therapies are discussed, looking at their effects on reducing rates of infection, reducing severity of infection and improved recovery from infection. Further discussion regarding the level of evidence required for nutritional interventions is included.