Genomic heterogeneity of Dichelobacter nodosus within and between UK sheep flocks and between age groups within a flock.

BACKGROUND: Footrot and interdigital dermatitis are endemic infectious diseases in all sheep farming regions, impairing welfare and production. The development of efficacious vaccines against the primary causative pathogen has been hampered by the extensive antigenic diversity of Dichelobacter nodosus. Understanding the heterogeneity of the pathogen within and between flocks is essential if the feasibility of bespoke vaccine production is to be assessed for use in the U.K. RESULTS: In this study 56 ewe and lamb isolates from 9 flocks were compared by D. nodosus serogroup and Multi Locus Sequence Type which provides significantly enhanced discriminatory power for molecular epidemiology. Serogroup heterogeneity between flocks ranged from two to five unique serogroups per flock. Three flocks contained isolates of two serogroups, two flocks contained isolates of three serogroups and one flock included isolates of five serogroups. Analysis of 25 isolates from one flock with high prevalence of lameness, identified that serogroup and sequence type was significantly correlated with age. Significantly higher proportion of lambs were infected with serogroup B (principally ST85) as opposed to serogroup H (principally ST86), which predominated amongst adult sheep. CONCLUSIONS: Genomic heterogeneity of the pathogen was significantly lower within flock compared to heterogenicity observed between flocks. Furthermore, this study indicates that within a flock, the host-pathogen dynamics and susceptibility to particular D. nodosus strains may be age dependent.

The Influence of Food Intake Specificity in Children with Autism on Gut Microbiota.

Autism spectrum disorder (ASD) is a complex of neurodevelopmental conditions with increasing incidence. The microbiota of children with ASD is distinct from neurotypical children, their food habits are also different, and it is known that nutrient intake influences microbiota in a specific way. Thus, this study investigates the food habits of children with ASD and their association with the gut microbiota. Children with ASD had their dietary energy intakes similar to controls, but they more often demonstrated food selectivity, which seemed to result in deficiency of micronutrients such as vitamins K, B6, C, iron, cooper, docosahexaenoic and docosapentanoic acid. Using high-throughput sequencing, a DNA library of intestinal microbiota was performed. Core microbiota was similar in children with and without ASD, but Dichelobacter, Nitriliruptor and Constrictibacter were found to be putative markers of ASD. The changes in gut microbiota that we observed in connection to food selectivity, intake of fats and omega-3 in particular, fermented milk products and animal/plant protein consumption had similar character, independent of diagnosis. However, high fibre intake was connected with a decreased α-diversity only in children with ASD. High carbohydrate and fibre intake influenced ß-diversity, changing the abundance of Bacteroides and other genera, many of them members of the Clostidiaceae. Modulating food habits of ASD children can influence their gut microbiota composition.

The prevalence of Dichelobacter nodosus in clinically footrot-free sheep flocks: a comparative field study on elimination strategies.

BACKGROUND: Ovine footrot caused by Dichelobacter nodosus (D nodosus) is an infectious disease affecting sheep worldwide. Switzerland plans a nationwide footrot eradication program, based on PCR-testing of interdigital swab samples. The aim of this study was to test for the presence of D nodosus in clinically footrot-free sheep flocks which had been subjected to different treatment strategies, to assess whether they were feasible for the eradication process, especially focussing on antimicrobial flock treatments. Clinical scoring and PCR-results were compared. Ten farms had used hoof bathing and hoof trimming without causing bleeding, ten had used individual treatments and flock vaccines to gain the free status and ten had become free through whole-flock systemic macrolide treatment. For every farm, three risk-based collected pool samples were analysed for the occurrence of virulent and benign D nodosus by PCR detection of aprV2/aprB2. RESULTS: Six flocks from any treatment group tested positive for aprB2 in all pools. Clinical signs were absent at the time of sampling, but some flocks had experienced non-progressive interdigital inflammation previously. Two flocks tested aprV2-positive in the high-risk pool. One of them underwent a progressive footrot outbreak shortly after sampling. Individual retesting indicated, that virulent D nodosus most likely was reintroduced by a recently purchased ram. In the second flock, a ram was tested positive and treated before clinical signs occurred. CONCLUSIONS: All treatment strategies eliminated the causative agent and were found to be suitable for implementation in the PCR-based eradication process. PCR-testing proved to be more sensitive than visual scoring, as it also detected clinically healthy carriers. It will be of benefit as a diagnostic tool in elimination and surveillance programs.

Dichelobacter nodosus in sheep, cattle, goats and South American camelids in Switzerland-Assessing prevalence in potential hosts in order to design targeted disease control measures.

Footrot is a contagious foot disease caused by the bacterium Dichelobacter nodosus (D. nodosus) that affects sheep worldwide. Due to substantial economic and welfare impact, various countries have developed control programs against footrot. The aim of this cross-sectional study was to estimate the national prevalence of virulent and benign D. nodosus in Switzerland in the four domestic ruminant species sheep, cattle, goats and South American camelids (SAC) to detect potential host populations and to propose targeted disease control measures. Risk factors for infection with the virulent strain of D. nodosus, based on a survey carried out among farmers, were investigated on animal and herd level. Overall, 613 farms and 2920 animals were investigated during 2017-18 applying a two-stage cluster sampling strategy. A Real-Time PCR method for simultaneous detection of virulent and benign strains of D. nodosus was used for the first time in such a large study. On animal level, the true prevalence (TP) of virulent D. nodosus in sheep was estimated at 16.9% (95% confidence interval (CI95%): 9.5-24.3%). In cattle and goats no virulent D. nodosus was detected and in SAC an apparent prevalence (AP) of 0.2% (CI95%: 0.0-0.4%) was observed. On farm level, a TP of virulent D. nodosus of 16.2% (CI95%: 8.4-25.2%) for sheep and an AP of 1.5% (CI95%: 0.3-5.2%) for SAC herds was estimated. Since the Swiss control program only targets the virulent strains of D. nodosus, it was concluded that cattle, goats and SAC do not play a role in footrot epidemiology in Switzerland. Adult sheep were at higher risk of infection for virulent D. nodosus compared to lambs and yearlings. On herd level, risk factors for infection with virulent D. nodosus in sheep were earlier occurence of footrot, winter compared to summer and autumn, and goat contact on pasture. Liming pastures had a protective effect on D. nodosus infection. For benign D. nodosus, the TP in sheep was 6.3% (CI95%: 1.6-11.0%) and in cattle 88.4% (CI95%: 83.8-93.0%). The TP for benign D. nodosus in sheep farms was 2.8% (CI95%: 0.0-10.5%) and in cattle farms 95.9% (CI95%: 91.7-98.1%). In goat and SAC farms, the AP was 6.6% (CI95%: 3.4-11.5%) and 7.4% (CI95%: 3.8-13.1%), respectively. These findings could be relevant for wild ruminants such as Alpine ibex (Capra ibex ibex), which can develop clinical footrot after infection with benign D. nodosus. The findings of this study are crucial for assessing targeted disease control measures in Switzerland.

Potential transmission routes of Dichelobacter nodosus.

Footrot caused by Dichelobacter nodosus is a highly contagious bacterial disease affecting the claw of sheep and the main cause of lameness in these animals. It is not only an economic burden but also a serious animal welfare issue. More information about the transmission of D. nodosus is needed for effective footrot control programs. We therefore determined the prevalence of D. nodosus in sheep presented at shows and markets where commingling of animals occurs. Furthermore, possible transmission vectors during foot trimming were investigated and trimming knife decontamination protocols evaluated. Sheep at six markets and four shows were sampled and tested for the presence of D. nodosus by real-time PCR. Different vectors, such as trimming knives were tested by real-time PCR and for viable D. nodosus by culture. The prevalence of virulent D. nodosus in sheep presented at shows and markets ranged from 1.7% to 100%. Regions with an ongoing control program showed significantly lower prevalence. After trimming, positive real-time PCR and culture results were obtained from the knives, the hands of the claw trimmers as well as removed claw horn material whereas boots were only positive by real-time PCR. In conclusion, markets and shows pose a risk for transmission of D. nodosus. The risk of transmission is particularly high during claw trimming and recommended measures to limit this risk include wiping the knife with a disinfection towel, wearing and changing gloves after every sheep, as well as proper disposal of trimmed and infectious horn.

Pathology and bacteria related to digital dermatitis in dairy cattle in all year round grazing system in Brazil.

Digital dermatitis (DD) is one of the main causes of lameness in dairy cattle worldwide, and it is frequently reported in high-yielding, free stall dairy herds from regions with a temperate climate. However, DD is also observed with high prevalence in grazing cattle with a low milk yield in tropical regions. To clarify whether these differences have an impact on the etiology of the disease, we studied DD lesions from all year round grazing cattle of mixed breed in Brazil using high-throughput 16S rRNA gene sequencing and fluorescent in situ hybridization. The study included samples from 66 skin lesions and 5 healthy skins collected from five farms. Both techniques showed Treponema spp. to be the most abundant bacteria, present in all but one of the samples with minimal epidermal alterations. We identified eleven different Treponema strains belonging to the six major phylotypes of Treponema which have all previously been identified in DD lesions. Furthermore, we identify Dichelobacter nodosus in DD lesions by gene sequencing and also by fluorescent in situ hybridization in almost half of biopsy specimens in areas with mild epithelial damage and together with Treponema. The present data support the hypothesis that Treponema constitutes the main pathogen responsible for DD, independent of the environment and region where cows are kept, and it further suggests D. nodosus as another potentially important pathogen.

Detection and Serogrouping of Dichelobacter nodosus Infection by Use of Direct PCR from Lesion Swabs To Support Outbreak-Specific Vaccination for Virulent Footrot in Sheep.

Virulent footrot is an economically significant disease in most sheep-rearing countries. The disease can be controlled with vaccine targeting the fimbriae of virulent strains of the essential causative agent, Dichelobacter nodosus However, the bacterium is immunologically heterogeneous, and 10 distinct fimbrial serogroups have been identified. Ideally, in each outbreak the infecting strains would be cultured and serogrouped so that the appropriate serogroup-specific mono- or bivalent vaccine could be administered, because multivalent vaccines lack efficacy due to antigenic competition. If clinical disease expression is suspected to be incomplete, culture-based virulence tests are required to confirm the diagnosis, because control of benign footrot is economically unjustifiable. Both diagnosis and vaccination are conducted at the flock level. The aims of this study were to develop a PCR-based procedure for detecting and serogrouping D. nodosus directly from foot swabs and to determine whether this could be done accurately from the same cultured swab. A total of 269 swabs from the active margins of foot lesions of 261 sheep in 12 Merino sheep flocks in southeastern Australia were evaluated. DNA extracts taken from putative pure cultures of D. nodosus and directly from the swabs were evaluated in PCR assays for the 16S rRNA and fimA genes of D. nodosus Pure cultures were tested also by the slide agglutination test. Direct PCR using extracts from swabs was more sensitive than culture for detecting and serogrouping D. nodosus strains. Using the most sensitive sample collection method of the use of swabs in lysis buffer, D. nodosus was more likely to be detected by PCR in active than in inactive lesions, and in lesions with low levels of fecal contamination, but lesion score was not a significant factor. PCR conducted on extracts from swabs in modified Stuart's transport medium that had already been used to inoculate culture plates had lower sensitivity. Therefore, if culture is required to enable virulence tests to be conducted, it is recommended that duplicate swabs be collected from each foot lesion, one in transport medium for culture and the other in lysis buffer for PCR.

Pooling of interdigital swab samples for PCR detection of virulent Dichelobacter nodosus.

Virulent ovine foot rot is a contagious foot disease. Given the development and validation of a real-time PCR to detect Dichelobacter nodosus isolates that contain the virulence-associated protease genes aprV2 and aprB2, the diagnosis of foot rot has made considerable progress. We evaluated pooling methods to reduce the number of samples during a foot rot control program. Samples of individual feet were compared to a 4-feet sample of the same sheep. All further analyses based on 4-feet samples (pools-of-5 and pools-of-10 4-feet samples) were compared to samples of individual sheep, and a risk-based herd sampling was evaluated and compared to the whole flock. The sensitivity and specificity of the 4-feet samples for detection of aprV2-positive strains was 93.8% (CI: 87.6-97.5%) and 98.3% (CI: 96.5-99.3%), respectively. The sensitivity and specificity of the pools-of-10 was 86.7% (CI: 78.4-92.7%) and 100.0% (CI: 97.4-100%), respectively. Pools-of-5 were not significantly more sensitive than pools-of-10. The pooling of 4 individual foot samples into one 4-feet sample is an adequate method to reduce the number of samples of individual sheep. The sensitivity of pools-of-5 and pools-of-10 is too imprecise for a control program. Risk-based sampling allowed for a substantial reduction of samples to be tested, had a sensitivity of 95.8% (CI: 78.9-99.9%) and specificity of 100.0% (CI: 88.1-100.0%) when determining the foot rot flock status, and represents an adequate methodology to predict within-flock freedom from infection.

A financial cost-benefit analysis of eradicating virulent footrot.

In 2008, virulent footrot was detected in sheep in south-west Norway. Footrot is caused by Dichelobacter nodosus, and the outbreak was linked to live sheep imported from Denmark in 2005. A large-scale program for eradicating the disease was implemented as a joint industry and governmental driven eradication project in the years 2008-2014, and continued with surveillance and control measures by the Norwegian Food Safety Authority from 2015. The cost of the eradication program including surveillance and control measures until 2032 was assumed to reach approximately €10.8 million (NOK 90 million). A financial cost-benefit analysis, comparing costs in the eradication program with costs in two simulated scenarios, was carried out. In the scenarios, designated ModerateSpread (baseline) and SlowSpread, it was assumed that the sheep farmers would undertake some voluntary measures on their own that would slow the spread of the disease. The program obtained a positive NPV after approximately 12 years. In a stochastic analysis, the probabilities of a positive NPV were estimated to 1.000 and to 0.648 after 15 years and to 0.378 and 0.016 after ten years, for the ModerateSpread and SlowSpread scenarios respectively. A rapid start-up of the program soon after the detection of the disease was considered crucial for the economic success as the disease would have become more widespread and probably raised the costs considerably at a later start-up.

Evaluation of Genotypic and Phenotypic Protease Virulence Tests for Dichelobacter nodosus Infection in Sheep.

Dichelobacter nodosus is a fastidious, strictly anaerobic bacterium, an obligate parasite of the ruminant hoof, and the essential causative agent of virulent ovine footrot. The clinical disease results from a complex interplay between the pathogen, the environment, and the host. Sheep flocks diagnosed with virulent but not benign footrot in Australia may be quarantined and required to undergo a compulsory eradication program, with costs met by the farmer. Virulence of D. nodosus at least partially depends on the elaboration of a protease encoded by aprV2 and manifests as elastase activity. Laboratory virulence tests are used to assist diagnosis because clinical differentiation of virulent and benign footrot can be challenging during the early stages of disease or when the disease is not fully expressed due to unfavorable pasture conditions. Using samples collected from foot lesions from 960 sheep from 40 flocks in four different geographic regions, we evaluated the analytical characteristics of qPCR tests for the protease gene alleles aprV2 and aprB2, and compared these with results from phenotypic protease (elastase and gelatin gel) tests. There was a low level of agreement between clinical diagnosis and quantitative PCR (qPCR) test outcomes at both the flock and sample levels and poor agreement between qPCR test outcomes and the results of phenotypic virulence tests. The diagnostic specificity of the qPCR test was low at both the flock and individual swab levels (31.3% and 18.8%, respectively). By contrast, agreement between the elastase test and clinical diagnosis was high at both the flock level (diagnostic sensitivity [DSe], 100%; diagnostic specificity [DSp], 78.6%) and the isolate level (DSe, 69.5%; DSp, 80.5%).

Identification and characterization of Dichelobacter nodosus serogroup H from ovine footrot in India.

A total of 56 foot swabs were collected from inter digital spaces of sheep with footrot lesions were screened for 16 rRNA of Dichelobacter nodosus by PCR. Out of the 56 samples, 38(67.85%) were found to be positive. All the positive samples were subjected to multiplex PCR targeting fimA gene for identification of serogroups of D. nodosus. Serogroup H was found along with serogroup B in 12 (55.26%) samples and with serogroup I in 8 (22.2%) samples. The serogroup H was identified for the first time from the Indian subcontinent. The phylogenetic analysis of the present sequence with the available serogroup H sequences of GenBank revealed to be in close association with the serotype H1.

Ovine footrot: new insights into bacterial colonisation.

Ovine footrot is characterised by interdigital dermatitis (ID) and by the separation of the skin and hoof horn (under-running footrot). Dichelobacter nodosus is the essential pathogen causing footrot; the role of other microorganisms in this disease remains unclear. The aims of this study were (i) to investigate the colonisation of D nodosus, Fusobacterium necrophorum and Treponema species in biopsies from the ovine interdigital skin of healthy, ID and footrot-affected feet and (ii) to characterise the virulence of D nodosus strains in those biopsies. Postslaughter biopsy samples (n=241) were collected and analysed by real-time PCR to determine prevalence and load of the different bacterial species. The highest prevalence and load of D nodosus were found on feet with ID. The vast majority of samples contained virulent D nodosus and some samples contained both virulent and benign D nodosus Notably, the more pathogenic subspecies of F necrophorum was found in samples from UK sheep. Our findings provide further insights into the role bacterial colonisation may play in the early stage of ID and in the progression towards footrot.

Survival of the ovine footrot pathogen Dichelobacter nodosus in different soils.

Dichelobacter nodosus (D. nodosus) is the causative agent of footrot in sheep; one of the most important health and welfare issues of sheep worldwide. For control programmes to be effective, it is essential that the transmission cycle of D. nodosus is understood and bacterial reservoirs in the environment are better defined. This study evaluated the survival of D. nodosus in different soils using soil microcosms. Cultivation independent and dependent methods were used to detect D. nodosus over 40 days from seeding in soil. A D. nodosus specific probe was used for quantification by qPCR and viability was assessed by cell permeability to an intercalating dye, PMA, and by culture. Survival varied dramatically depending on soil type, matric potential (MP) and temperature. Our findings indicate that D. nodosus survival was higher at 5 °C compared with 25 °C in all soils and significantly longer at both temperatures in clay soil (>44% clay) compared with other soil types. Survival under all conditions was longer than 30 days for both culture independent and dependent methods, this is substantially longer than previous studies and, if this is an infectious dose, longer than the current recommendation of resting a field for 14 days to prevent onward infection.

The detection of Dichelobacter nodosus and Fusobacterium necrophorum from ovine footrot in Kashmir, India.

In a study conducted, a total of 450 swab samples from footrot lesions of naturally infected sheep were collected in all the ten districts of the Kashmir valley and were examined for the presence of Dichelobacter nodosus (D. nodosus) and Fusobacterium necrophorum (F. necrophorum), in order to determine if F. necrophorum was associated with ovine footrot. The detection of F. necrophorum and D. nodosus was carried out by polymerase chain reaction targeting the leukotoxin (lktA) and 16S rRNA genes, respectively. In this study, only less than 50% of positive samples contained both the bacteria, so it is not possible to conclude with certainty that both bacteria are together required for the disease manifestation.

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.

Characterisation of Dichelobacter nodosus and detection of Fusobacterium necrophorum and Treponema spp. in sheep with different clinical manifestations of footrot.

The aim of this study was to determine the proportion of Dichelobacter nodosus, Fusobacterium necrophorum and Treponema spp. in sheep with different clinical manifestations of footrot compared to healthy sheep both at flock and individual level. The second aim was to characterise D. nodosus with respect to virulence, presence of intA gene and the serogroups. Swab samples (n=1000) from footrot-affected (n=10) and healthy flocks (n=10) were analysed for the presence of D. nodosus, F. necrophorum and Treponema spp. by real-time PCR and culturing (D. nodosus only). Dichelobacter nodosus isolates (n=78) and positive swabs (n=474) were analysed by real-time PCR for the aprV2/B2 and the intA genes and by PCR for the fimA gene (isolates only). D. nodosus was more commonly found in flocks affected with footrot than in clinically healthy flocks. A significant association was found between feet with severe footrot lesions and the aprV2 gene and between feet with moderate or no lesions and the aprB2 gene, respectively. F. necrophorum was more commonly found in flocks with footrot lesions than in flocks without lesions. No significant association was found between sheep flocks affected with footrot and findings of Treponema spp. or the intA gene. Benign D. nodosus of six different serogroups was detected in twelve flocks and virulent D. nodosus of serogroup G in one. In conclusion, D. nodosus and F. necrophorum were more commonly found in feet with footrot than in healthy feet. The majority of D. nodosus detected was benign, while virulent D. nodosus was only detected in a single flock.

High-level association of bovine digital dermatitis Treponema spp. with contagious ovine digital dermatitis lesions and presence of Fusobacterium necrophorum and Dichelobacter nodosus.

Contagious ovine digital dermatitis (CODD) is an important foot disease in sheep, with significant animal welfare and economic implications. It is thought that CODD emerged from bovine digital dermatitis (BDD) via treponemal bacteria. With wildlife species such as elk now suffering a CODD-like disease, it is imperative to clarify these disease etiologies. A large investigation into treponemal association with CODD is warranted. CODD lesions (n = 58) and healthy sheep foot tissues (n = 56) were analyzed by PCR for the three BDD-associated Treponema phylogroups and two other lameness-associated bacteria, Dichelobacter nodosus and Fusobacterium necrophorum. Spirochete culture was also attempted on CODD lesions. "Treponema medium/Treponema vincentii-like," "Treponema phagedenis-like," and Treponema pedis spirochetes were identified in 39/58 (67%), 49/58 (85%), and 41/58 (71%) of CODD lesions, respectively. One or more BDD-associated Treponema phylogroups were detected in 100% of CODD lesions. Healthy foot tissues did not amplify BDD-associated Treponema phylogroup DNA. D. nodosus and F. necrophorum were present in 34/58 (59%) and 41/58 (71%) of CODD lesions and 22/56 (39%) and 5/56 (9%) of healthy foot tissues, respectively. Thirty-two spirochetes were isolated from CODD lesions, with representatives clustering with, and indistinguishable from, each of the three BDD-associated Treponema phylogroups based on 16S rRNA gene comparisons. This study for the first time demonstrates a high-level association for BDD treponeme phylogroups in CODD and their absence from healthy tissues, supporting the hypothesis that BDD treponemes play a primary causative role in CODD and confirming that the specific PCR assays are an effective differential diagnostic tool for CODD.

Outbreak of severe foot rot associated with benign Dichelobacter nodosus in an Alpine ibex colony in the Swiss Prealps.

An outbreak of foot rot occurred in the ibex colony "Vanil Noir" in Switzerland from May to December 2014. This article describes field observations and the analyses carried out on the limbs of 3 animals submitted for postmortem examination. Disease signs observed in the field included lameness, poor body condition and overgrown hooves. Macroscopic examination of selected limbs revealed severe lesions in all of them, including interdigital inflammation with ulceration and malodorous exudation. Histological changes were consistent with chronic laminitis with bone resorption, which was not detected at radiographical examination. Grocott-positive organisms compatible with Dichelobacternodosus were detected in the lesions. Samples collected from the lesions were positive by polymerase chain reaction for benign D. nodosus, which is typically associated with only mild lesions in domestic sheep. Whether D. nodosus is endemic in the colony or had previously been transmitted from sympatric domestic livestock is unclear. The unusual warm and humid weather conditions in 2014 may well have contributed to the outbreak.

A recently introduced Dichelobacter nodosus strain caused an outbreak of footrot in Norway.

BACKGROUND: In 2008, an outbreak of ovine footrot occurred in Norway. Dichelobacter nodosus isolates collected between 2008 and 2011 have been characterised. Isolates defined as virulent by the gelatin gel test (GG-test) were only found in sheep in Rogaland County, where the severe cases of footrot were registered. The majority (96%) of the virulent isolates belonged to serogroup A. It is suspected that they represent a newly introduced strain, and the aim of the present study was to investigate whether they are genetically similar. Sixty-one virulent isolates from sheep and 116 benign isolates from sheep, cattle and goats were included. Four GG-test virulent isolates from Danish sheep were also included. All isolates were genotyped by pulsed-field gel electrophoresis (PFGE) and by PCR for pgr variant determination. RESULTS: The Norwegian virulent isolates were assigned to 8 pulsotypes (PTs), while the benign isolates were assigned to 66 PTs. Thirty-seven (68.5%) of the 54, virulent, serogroup A isolates belonged to the same PT, and included isolates from 2008 through 2011. Isolates belonging to this PT were defined as the outbreak strain. The remaining virulent serogroup A isolates belonged to 4 PTs differing by ≤3 bands from the outbreak strain. Two virulent, Danish, serogroup A isolates differed by 2 bands from the Norwegian outbreak strain. All but 3 (95%) of the virulent isolates had the pgrA variant while 85% of the benign isolates had the pgrB variant. CONCLUSION: This study provides evidence that the footrot outbreak in Norway in 2008 most likely was caused by new introduction and local spread of one virulent D. nodosus strain.

A longitudinal study of the role of Dichelobacter nodosus and Fusobacterium necrophorum load in initiation and severity of footrot in sheep.

Footrot is an infectious bacterial disease of sheep that causes lameness. The causal agent is Dichelobacter nodosus. There is debate regarding the role of Fusobacterium necrophorum in disease initiation. This research used an observational longitudinal study of footrot, together with quantitative PCR (qPCR) of bacterial load of D. nodosus and F. necrophorum, to elucidate the roles of each species in the development of disease. All feet of 18 a priori selected sheep were monitored for five weeks assessing disease severity (healthy, interdigital dermatitis (ID) and severe footrot (SFR)) and bacterial load. A multinomial model was used to analyse these data. Key unadjusted results were that D. nodosus was detected more frequently on feet with ID, whereas F. necrophorum was detected more frequently on feet with SFR. In the multinomial model, ID was associated with increasing log10 load of D. nodosus the week of observation (OR=1.28 (95% CI=1.08-1.53)) and the week prior to development of ID (OR=1.20 (95% CI=1.01-1.42). There was no association between log10 load(2) of F. necrophorum and presence of ID (OR=0.99 (95% CI=0.96-1.02))). SFR was associated with increasing log10 load of D. nodosus the week before disease onset (OR=1.42 (95% CI=1.02-1.96)) but not once SFR had occurred. SFR was positively associated with log10 load(2) of F. necrophorum once disease was present (OR=1.06 (95% CI=1.01-1.11)). In summary, there was an increased risk of increasing D. nodosus load the week prior to development of ID and SFR and during an episode of ID. In contrast, F. necrophorum load was not associated with ID before or during an episode, and was only associated with SFR once present. These results contribute to our understanding of the epidemiology of footrot and highlight that D. nodosus load plays the primary role in disease initiation and progression, with F. necrophorum load playing a secondary role. Further studies in more flocks and climates would be useful to confirm these findings. This study identifies that D. nodosus load is highest during ID. This supports previous epidemiological findings, which demonstrate that controlling ID is the most effective management strategy to prevent new cases of ID and SFR.