Detection and diversity of a putative novel heterogeneous polymorphic proline-glycine repeat (Pgr) protein in the footrot pathogen Dichelobacter nodosus

Dichelobacter nodosus, a Gram-negative anaerobic bacterium, is the essential causative agent of footrot in sheep. Currently, depending on the clinical presentation in the field, footrot is described as benign or virulent; D. nodosus strains have also been classified as benign or virulent, but this designation is not always consistent with clinical disease. The aim of this study was to determine the diversity of the pgr gene, which encodes a putative proline-glycine repeat protein (Pgr). The pgr gene was present in all 100 isolates of D. nodosus that were examined and, based on sequence analysis had two variants, pgrA and pgrB. In pgrA, there were two coding tandem repeat regions, R1 and R2: different strains had variable numbers of repeats within these regions. The R1 and R2 were absent from pgrB. Both variants were present in strains from Australia, Sweden and the UK, however, only pgrB was detected in isolates from Western Australia. The pgrA gene was detected in D. nodosus from tissue samples from two flocks in the UK with virulent footrot and only pgrB from a flock with no virulent or benign footrot for >10 years. Bioinformatic analysis of the putative PgrA protein indicated that it contained a collagen-like cell surface anchor motif. These results suggest that the pgr gene may be a useful molecular marker for epidemiological studies

Multiple locus VNTR analysis highlights that geographical clustering and distribution of Dichelobacter nodosus, the causal agent of footrot in sheep, correlates with inter-country movements

Dichelobacter nodosus is a Gram-negative, anaerobic bacterium and the causal agent of footrot in sheep. Multiple locus variable number tandem repeat (VNTR) analysis (MLVA) is a portable technique that involves the identification and enumeration of polymorphic tandem repeats across the genome. The aims of this study were to develop an MLVA scheme for D. nodosus suitable for use as a molecular typing tool, and to apply it to a global collection of isolates. Seventy-seven isolates selected from regions with a long history of footrot (GB, Australia) and regions where footrot has recently been reported (India, Scandinavia), were characterised. From an initial 61 potential VNTR regions, four loci were identified as usable and in combination had the attributes required of a typing method for use in bacterial epidemiology: high discriminatory power (D > 0.95), typeability and reproducibility. Results from the analysis indicate that D. nodosus appears to have evolved via recombinational exchanges and clonal diversification. This has resulted in some clonal complexes that contain isolates from multiple countries and continents; and others that contain isolates from a single geographic location (country or region). The distribution of alleles between countries matches historical accounts of sheep movements, suggesting that the MLVA technique is sufficiently specific and sensitive for an epidemiological investigation of the global distribution of D. nodosus

Ovine pedomics : the first study of the ovine foot 16S rRNA-based microbiome

We report the first study of the bacterial microbiome of ovine interdigital skin based on 16S rRNA by pyrosequencing and conventional cloning with Sanger-sequencing. Three flocks were selected, one a flock with no signs of footrot or interdigital dermatitis, a second flock with interdigital dermatitis alone and a third flock with both interdigital dermatitis and footrot. The sheep were classified as having either healthy interdigital skin (H), interdigital dermatitis (ID) or virulent footrot (VFR). The ovine interdigital skin bacterial community varied significantly by flock and clinical condition. The diversity and richness of operational taxonomic units was greater in tissue from sheep with ID than H or VFR affected sheep. Actinobacteria, Bacteriodetes, Firmicutes and Proteobacteria were the most abundant phyla comprising 25 genera. Peptostreptococcus, Corynebacterium and Staphylococcus were associated with H, ID and VFR respectively. Sequences of Dichelobacter nodosus, the causal agent of ovine footrot, were not amplified due to mismatches in the 16S rRNA universal forward primer (27F). A specific real time PCR assay was used to demonstrate the presence of D. nodosus which was detected in all samples including the flock with no signs of ID or VFR. Sheep with ID had significantly higher numbers of D. nodosus (104-109 cells/g tissue) than those with H or VFR feet

Biological and clinical characteristics of gene carriers far from predicted onset in the Huntington's disease Young Adult Study (HD-YAS): a cross-sectional analysis.

BACKGROUND: Disease-modifying treatments are in development for Huntington's disease; crucial to their success is to identify a timepoint in a patient's life when there is a measurable biomarker of early neurodegeneration while clinical function is still intact. We aimed to identify this timepoint in a novel cohort of young adult premanifest Huntington's disease gene carriers (preHD) far from predicted clinical symptom onset. METHODS: We did the Huntington's disease Young Adult Study (HD-YAS) in the UK. We recruited young adults with preHD and controls matched for age, education, and sex to ensure each group had at least 60 participants with imaging data, accounting for scan fails. Controls either had a family history of Huntington's disease but a negative genetic test, or no known family history of Huntington's disease. All participants underwent detailed neuropsychiatric and cognitive assessments, including tests from the Cambridge Neuropsychological Test Automated Battery and a battery assessing emotion, motivation, impulsivity and social cognition (EMOTICOM). Imaging (done for all participants without contraindications) included volumetric MRI, diffusion imaging, and multiparametric mapping. Biofluid markers of neuronal health were examined using blood and CSF collection. We did a cross-sectional analysis using general least-squares linear models to assess group differences and associations with age and CAG length, relating to predicted years to clinical onset. Results were corrected for multiple comparisons using the false discovery rate (FDR), with FDR 0·16). CSF neurofilament light protein (NfL), plasma NfL, and CSF YKL-40 were elevated in this far-from-onset preHD cohort compared with controls (FDR<0·0001, =0·01, and =0·03, respectively). CSF NfL elevations were more likely in individuals closer to expected clinical onset (FDR <0·0001). INTERPRETATION: We report normal brain function yet a rise in sensitive measures of neurodegeneration in a preHD cohort approximately 24 years from predicted clinical onset. CSF NfL appears to be a more sensitive measure than plasma NfL to monitor disease progression. This preHD cohort is one of the earliest yet studied, and our findings could be used to inform decisions about when to initiate a potential future intervention to delay or prevent further neurodegeneration while function is intact. FUNDING: Wellcome Trust, CHDI Foundation

Pf7: an open dataset of Plasmodium falciparum genome variation in 20,000 worldwide samples

We describe the MalariaGEN Pf7 data resource, the seventh release of Plasmodium falciparum genome variation data from the MalariaGEN network.  It comprises over 20,000 samples from 82 partner studies in 33 countries, including several malaria endemic regions that were previously underrepresented.  For the first time we include dried blood spot samples that were sequenced after selective whole genome amplification, necessitating new methods to genotype copy number variations.  We identify a large number of newly emerging crt mutations in parts of Southeast Asia, and show examples of heterogeneities in patterns of drug resistance within Africa and within the Indian subcontinent.  We describe the profile of variations in the C-terminal of the csp gene and relate this to the sequence used in the RTS,S and R21 malaria vaccines.  Pf7 provides high-quality data on genotype calls for 6 million SNPs and short indels, analysis of large deletions that cause failure of rapid diagnostic tests, and systematic characterisation of six major drug resistance loci, all of which can be freely downloaded from the MalariaGEN website

The role of the environment in transmission of Dichelobacter nodosus between ewes and their lambs

Dichelobacter nodosus (D. nodosus) is the essential causative agent of footrot in sheep. The current study investigated when D. nodosus was detectable on newborn lambs and possible routes of transmission. Specific qPCR was used to detect and quantify the load of D. nodosus in foot swabs of lambs at birth and 5–13 h post-partum, and their mothers 5–13 h post-partum; and in samples of bedding, pasture, soil and faeces. D. nodosus was not detected on the feet of newborn lambs swabbed at birth, but was detected 5–13 h after birth, once they had stood on bedding containing naturally occurring D. nodosus. Multiple genotypes identified by cloning and sequencing a marker gene, pgrA, and by multi locus variable number tandem repeat analysis (MLVA) of community DNA from swabs on individual feet indicated a mixed population of D. nodosus was present on the feet of both ewes and lambs. There was high variation in pgrA tandem repeat number (between 3 and 21 repeats), and multiple MLVA types. The overall similarity index between the populations on ewes and lambs was 0.45, indicating moderate overlap. Mother offspring pairs shared some alleles but not all, suggesting lambs were infected from sources(s) other than just their mother's feet. We hypothesise that D. nodosus is transferred to the feet of lambs via bedding containing naturally occurring populations of D. nodosus, probably as a result of transfer from the feet of the group of housed ewes. The results support the hypothesis that the environment plays a key role in the transmission of D. nodosus between ewes and lambs

First study of pathogen load and localisation of ovine footrot using fluorescence in situ hybridisation (FISH)

Analysis of bacterial populations in situ provides insights into pathogen population dynamics and potential reservoirs for disease. Here we report a culture-independent study of ovine footrot (FR); a debilitating bacterial disease that has significant economic impact on sheep farming worldwide. Disease begins as an interdigital dermatitis (ID), which may then progress to separation of the hoof horn from the underlying epidermis causing severe footrot (SFR). Dichelobacter nodosus is the causative agent of ovine FR, however, the role of Fusobacterium necrophorum and other bacteria present in the environment and on the feet of sheep is less clear. The objective of this study was to use fluorescence in situ hybridisation (FISH) to detect, localise and quantify D. nodosus, F. necrophorum and the domain Bacteria from interdigital skin biopsies of healthy, ID- and SFR-affected feet. D. nodosus and F. necrophorum populations were restricted primarily to the epidermis, but both were detected more frequently in feet with ID or SFR than in healthy feet. D. nodosus cell counts were significantly higher in feet with ID and SFR (p < 0.05) than healthy feet, whereas F. necrophorum cell counts were significantly higher only in feet with SFR (p < 0.05) than healthy feet. These results, together with other published data, indicate that D. nodosus likely drives pathogenesis of footrot from initiation of ID to SFR; with D. nodosus cell counts increasing prior to onset of ID and SFR. In contrast, F. necrophorum cell counts increase after SFR onset, which may suggest an accessory role in disease pathogenesis, possibly contributing to the severity and duration of SFR

Latitude, depth and environmental variables influence deepwater fish assemblages off Western Australia

Continental shelf and upper slope fishes are important global resources which have significant economic and biodiversity values. Understanding the drivers of their abundance and distribution are fundamental to guide sustainable fisheries management arrangements. Deepwater demersal fish assemblages were sampled between depths of 91–570 m at six locations along the Western Australian coastline spanning 17° of latitude (i.e. 17–34°S at the Rowley Shoals, Montebello Islands, Ningaloo Reef, Abrolhos Islands, Perth Canyon and South West Capes) using Baited Remote Underwater Stereo-Video systems (stereo-BRUVs, n = 417 deployments). A total of 5965 individual fishes were identified belonging to 252 species and 92 families. Assemblage composition data were modelled against in-situ habitat data, temperature, salinity, current speed, and current direction. Multivariate statistical analyses revealed that fish assemblage composition was predominantly influenced by latitude (9.5% of variation), depth (7.3%) and salinity (7.2%). Three distinct assemblages were defined in response to latitude; a tropical, a sub-tropical and a southern temperate bioregion. Species richness declined with increasing latitude, and average biomass was highest in the southern bioregion. A mid-depth peak in biomass and lengths indicated the dominance of meso-predator species in the 300–399 m depth range. The information provided by this study on the factors influencing the abundance and distribution of fish assemblages over an exceptionally broad depth and latitudinal range along a continuous coastline will be useful for predicting the effects of future climate shifts on this and similar teleost species occupying tropical and temperate oceans elsewhere

Exploring perceptions of low risk behaviour and drivers to test for HIV among South African youth.

Human Immunodeficiency Virus (HIV) prevalence among South African youth is high, yet HIV testing remains suboptimal. We explored how perceptions of HIV risk and behaviours informed decisions to test for HIV. This study was conducted from April 2018 to March 2019 in Ekurhuleni district, Gauteng Province with males and females aged between 15-24 years. Twenty-five youth with unknown HIV status participated in in-depth interviews (IDIs); while four focus group discussions (FGDs) were conducted with those that previously tested for HIV. Probes used in the guides included types of incentives that youth would value when testing for HIV or receiving treatment; barriers and motivators to HIV testing; enablers and challenges to using cellphone technology and preferences on type of social media that could be used to create awareness about HIV testing services. IDIs and FGDs were audio-recorded, transcribed, and translated. QSR NVIVO 10 was used for the analysis. The majority of the youth perceived that their risk of HIV infection was low due to factors such as being young, lacking physical signs of HIV, being sexually inactive and parents not being HIV positive. However, youth identified high risk behaviours such as unprotected sex, multiple sexual partners, excessive drinking of alcohol, being victims of sexual abuse, road accidents and violent behaviour as increasing their vulnerability to HIV. Most youth highlighted cues to action that would motivate them to test for HIV such as support of parents, receiving incentives, improved confidentiality during HIV testing and receiving information about HIV via social media (Facebook, Twitter and Whatsapp). Despite perceptions of low risk to HIV, youth remain vulnerable to HIV. Disseminating HIV information via digital platforms; giving youth options to choose between testing locations that they consider to be private; providing incentives and equipping parents/guardians to encourage youth to test could optimise HIV testing