Search

Zebu cattle (Bos indicus) is reported to be more resistant towards harmful environmental factors than taurine cattle (Bos taurus). A few hundred zebu cattle are kept in Switzerland and in contrast to the Swiss indigenous breeds, infectious hoof disease in zebu is not observed. Therefore, we compared the prevalence of three ruminant hoof pathogens in zebu and taurine cattle. These included Treponema spp., Fusobacterium necrophorum and Dichelobacter nodosus which are associated with bovine digital dermatitis (BDD), different bovine hoof diseases and ovine footrot, respectively. Interdigital swabs and punch biopsies from hind feet of slaughter animals were tested for the three pathogens by PCR. Sixty zebu from eight farms were compared to a convenience sample of 20 taurine cattle from 17 farms. Treponema spp. associated with BDD were not detected in zebu while 23 % of animals and 50 % of farms were positive for benign D. nodosus, with results indicating environmental contamination rather than colonization. Taurine cattle showed 35 % of animals and 41 % of farms positive for T. phagedenis while 90 % of animals and 94 % of farms were colonized by D. nodosus as indicated by a 500-fold higher bacterial load than in zebu. The difference in prevalence of the two pathogens between zebu and taurine cattle was highly significant. F. necrophorum was as well only detected in taurine cattle with values of 15 % of animals and 17.7 % of farms, being significantly different at the animal level. Furthermore, genetic analysis of Swiss zebu indicates high genomic diversity and clear separation from taurine cattle. This is the first evidence that zebu show resistance towards colonization by bacterial hoof pathogens in contrast to taurine cattle., Competing Interests: Declaration of Competing Interest None, (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Fusobacterium necrophorum is a Gram-negative anaerobic bacterium responsible for localized infections of the oropharynx that can evolve into bacteremia and/or septic thrombophlebitis of the jugular vein or peritonsillar vein, called Lemierre's syndrome. To identify microbial genetic determinants associated with the severity of this life-threatening disease, 70 F. necrophorum strains were collected and grouped into two categories according to the clinical presentation: (i) localized infection, (ii) bacteremia with/without Lemierre's syndrome. Comparative genomic analyses revealed two clades with distinct genetic content, one clade being significantly enriched with isolates from subjects with bacteremia. To identify genetic determinants contributing to F. necrophorum pathogenicity, genomic islands and virulence factor orthogroups (OVFs) were predicted. The presence/absence profiles of OVFs did not group isolates according to their clinical category, but rather according to their phylogeny. However, a variant of lktA, a key virulence factor, with a frameshift deletion that results in two open reading frames, was associated with bacteremia. Moreover, a genome-wide association study identified three orthogroups associated with bacteremic strains: (i) cas8a1, (ii) a sodium/solute symporter, and (iii) a POP1 domain-containing protein. Further studies must be performed to assess the functional impact of lktA mutation and of these orthogroups on the physiopathological mechanisms of F. necrophorum infections., (© 2024. The Author(s).)

We described a case of a 24-year-old man with multiple organ failure caused by Fusobacterium necrophorum subsp. funduliforme F1260. This is the first described case of Lemierre's syndrome with multiple organ failure due to F. necrophorum subsp. funduliforme F1260 in an adult in China. Our study highlights that there may be a risk of misdiagnosis based solely on typical manifestations of internal jugular vein thrombophlebitis, metastatic lesions, and F. necrophorum isolated from blood cultures or normally sterile sites. Clinicians should be cognizant of the potential utility of metagenomic next-generation sequencing in facilitating early pathogen detection in severe infections, thus enabling timely and appropriate administration of antibiotics to reduce mortality rates and improve prognosis., Competing Interests: Declaration of competing interest No potential conflict of interest was reported by the authors., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Background: Legionella pneumonia is one of the most severe types of atypical pneumonia, impairing multiple organ systems, posing a threat to life. Diagnosing Legionella pneumonia is challenging due to difficulties in culturing the bacteria and limitations in immunoassay sensitivity and specificity., Case Presentation: This paper reports a rare case of sepsis caused by combined infection with Legionella pneumophila and Fusobacterium necrophorum, leading to respiratory failure, acute kidney injury, acute liver injury, myocardial damage, and electrolyte disorders. In addition, we systematically reviewed literature on patients with combined Legionella infections, analyzing their clinical features, laboratory results and diagnosis., Conclusions: For pathogens that require prolonged incubation periods and are less sensitive to conventional culturing methods, metagenomic next-generation sequencing (mNGS) can be a powerful supplement to pathogen screening and plays a significant role in the auxiliary diagnosis of complex infectious diseases., (© 2024. The Author(s).)

Tonsillar Fusobacterium necrophorum PCR Ct-values were higher in participants with asymptomatic tonsillar carriage than patients with pharyngeal infections. However, Ct-values were not associated with severity of disease or predictive of development of complications and hence lacked clinical usefulness. The reporting of F. necrophorum Ct-values in clinical samples is not recommended., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Background: Peritonsillar abscess (PTA) is a severe deep neck space infection with an insufficiently characterized bacterial etiology. We aimed to reveal the bacteria associated with PTA applying next generation sequencing (NGS). Tonsil biopsies and pus samples of 91 PTA patients were analysed applying NGS method., Results: Over 400 genera and 800 species belonging to 34 phyla were revealed. The most abundant species in both sample types were Streptococcus pyogenes, Fusobacterium necrophorum and Fusobacterium nucleatum. When present, S. pyogenes was normally a predominant species, although it was recovered as a minor population in some samples dominated by F. nucleatum and occasionally F. necrophorum. S. pyogenes and F. necrophorum were the predominant species (> 10% in a community) in 28 (31%) pus samples, while F. nucleatum in 21 (23%) and S. anginosus in 8 (9%) pus samples. We observed no substantial differences between the microbial findings in pus and tonsil biopsies., Conclusions: The most probable causative agents of PTA according to our NGS-study include Streptococcus pyogenes, Fusobacterium necrophorum and Fusobacterium nucleatum. Some other streptococci (S. anginosus) and anaerobes (Prevotella, Porphyromonas) may contribute to the infection as well. Pus of the peritonsillar abscess is more representative specimen for microbiological examination than the tonsillar tissue. Our results are important in the context of optimizing the handling of the PTA patients., (© 2023. The Author(s).)

Objective: Fusobacterium necrophorum causes bovine hepatic abscess, foot rot, mastitis, and endometritis. The 43 kDa outer membrane protein (43 K OMP) of F. necrophorum is a porin protein that plays an important role in infections by this bacterium, but the biological function and the pathogenesis of this protein are largely unknown., Methods: In this study, we investigated the role of the 43 K OMP in bacterial infection of bovine mammary epithelial cells (MAC-T cells) by Tandem Mass Tag proteomic analysis. The RAW264.7 cells were incubated with recombinant 43 K OMP (12.5 μg/mL) for 2 h, 4 h, 6 h, and 12 h, and then the inflammatory related protein and inflammatory cytokine production were measured by Western blot analysis and ELISA, the mRNA expression levels of inflammatory cytokine were measured by Real-Time PCR., Results: Proteomic analysis results demonstrated there were 224 differentially expressed proteins in the MAC-T cells stimulated with the 43 K OMP compared with control, and 118 proteins were upregulated and 106 proteins were downregulated. These differentially expressed proteins were mainly involved in NF-kappa B signaling, bacterial invasion of epithelial cells, cell adhesion, complement and coagulation cascades. The top six differentially expressed proteins were; MMP9, PLAU, STOM, PSMD13, PLAUR, and ITGAV, which were involved in a protein-protein interaction network. Furthermore, TLR/MyD88/NF-κB pathway related proteins and inflammatory cytokines (IL-6, TNF-α, and IL-1β) were assessed by Western blot analysis and ELISA. Results showed the 43 K OMP to enhance the expression of TLR4 protein at 2 h (P < 0.01) and the MyD88 protein at 4 h (P < 0.05) post-stimulation, and to decrease IκBα expression at 4 h, 6 h and 12 h (P < 0.05) post-infection, as well as induce phosphorylation at Ser536 (P < 0.01). Levels of IL-6, IL-1β, and TNF-α in the supernatants of mouse macrophages were increased (P < 0.05), as were mRNA expression levels of IL-6, IL-1β, and TNF-α (P < 0.05), while IL-4 mRNA expression was decreased (P < 0.05)., Conclusions: Taken together, these results suggested the important role for 43 K OMP in F. necrophorum infection, promoting the production of pro-inflammatory cytokines (IL-6 and TNF-α) by activation of the TLR/MyD88/NF-κB pathway. These findings provided a theoretical basis for a better understanding of the pathogenesis of F. necrophorum infection., Competing Interests: Declaration of competing interest The authors declare that there are no conflicts of interest with respect to this manuscript., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Fusobacterium necrophorum is a Gram-negative, filamentous anaerobe prevalent in the mucosal flora of animals and humans. It causes necrotic infections in cattle, resulting in a substantial economic impact on the cattle industry. Although infection severity and management differ within F. necrophorum species, little is known about F. necrophorum speciation and the genetic virulence determinants between strains. To characterize the clinical isolates, we performed whole-genome sequencing of four bovine isolates (8L1, 212, B17, and SM1216) and one human isolate (MK12). To determine the phylogenetic relationship and evolution pattern and investigate the presence of antimicrobial resistance genes (ARGs) and potential virulence genes of F. necrophorum, we also performed comparative genomics with publicly available Fusobacterium genomes. Using up-to-date bacterial core gene (UBCG) set analysis, we uncovered distinct Fusobacterium species and F. necrophorum subspecies clades. Pangenome analyses revealed a high level of diversity among Fusobacterium strains down to species levels. The output also identified 14 and 26 genes specific to F. necrophorum subsp. necrophorum and F. necrophorum subsp. funduliforme , respectively, which could be essential for bacterial survival under different environmental conditions. ClonalFrameML-based recombination analysis suggested that extensive recombination among accessory genes led to species divergence. Furthermore, the only strain of F. necrophorum with ARGs was F. necrophorum subsp. funduliforme B35, with acquired macrolide and tetracycline resistance genes. Our custom search revealed common virulence genes, including toxins, adhesion proteins, outer membrane proteins, cell envelope, type IV secretion system, ABC (ATP-binding cassette) transporters, and transporter proteins. A focused study on these genes could help identify major virulence genes and inform effective vaccination strategies against fusobacterial infections. IMPORTANCE Fusobacterium necrophorum is an anaerobic bacterium that causes liver abscesses in cattle with an annual incidence rate of 10% to 20%, resulting in a substantial economic impact on the cattle industry. The lack of definite biochemical tests makes it difficult to distinguish F. necrophorum subspecies phenotypically, where genomic characterization plays a significant role. However, due to the lack of a good reference genome for comparison, F. necrophorum subspecies-level identification represents a significant challenge. To overcome this challenge, we used comparative genomics to validate clinical test strains for subspecies-level identification. The findings of our study help predict specific clades of previously uncharacterized strains of F. necrophorum. Our study identifies both general and subspecies-specific virulence genes through a custom search-based analysis. The virulence genes identified in this study can be the focus of future studies aimed at evaluating their potential as vaccine targets to prevent fusobacterial infections in cattle.

Fusobacterium necrophorum causes a range of mild to life threatening infections and there is uncertainty in terms of diagnosis and treatment due to the lack of knowledge on their pathogenic mechanisms. This study characterised genomes of F. necrophorum to compare their virulence factors and investigate potential infection markers. 27 isolates of F. necrophorum from patients with pharyngotonsillitis were subjected to whole genome sequencing and compared with 42 genomes published in the NCBI database. Phylogenomics, pangemome, pan-GWAS and virulome were analysed to study strain variations with reference to virulence factors. Core genome based phylogenomic tree exhibited three clades of which Clade A belonged to F. necrophorum subsp necrophorum, clades B and C were F. necrophorum subsp funduliforme. Pan-GWAS and Pan-Virulome suggest some marker genes associated with clinical sources of isolation that needs further validation. Our study highlights some interesting features of the pathogenesis of F. necrophorum infections. Although the animal isolate genomes had some marker genes, the genomes of human isolates did not exhibit clear correlation to their clinical sources of isolation. This prompts to think of other mechanisms such as co-infections or host factors that can be involved in the pathogenesis., (Copyright © 2022 The Author(s). Published by Elsevier B.V. All rights reserved.)

Fusobacterium necrophorum can cause liver abscess, foot rot in ruminants, and Lemire syndrome in humans, Also, its virulence factors can induce the apoptosis of macrophages and neutrophils. However, the detailed mechanism has not been fully clarified. This study investigated the mechanisms of apoptosis and inflammatory factor production in F. necrophorum -induced neutrophils and macrophages (RAW246.7). After infection of macrophages with F. necrophorum , 5-ethynyl-2'-deoxyuridine labeling assays indicated that F. necrophorum inhibited macrophage proliferation in a time- and dose-dependent manner. Hoechst staining and DNA ladder assays showed significant condensation of the nucleus and fragmentation of genomic DNA in F. necrophorum -infected macrophages, Annexin V (FITC) and propidium iodide (PI) assay confirmed the emergence of apoptosis in the macrophages and sheep neutrophils with F. necrophorum compared with the control. The group with significant apoptosis was subjected to RNA sequencing (RNA-Seq), and the sequencing results revealed 2581 up- and 2907 downregulated genes. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analysis of the differentially expressed genes showed that F. necrophorum drove apoptosis and production of inflammatory factors by activating genes related to the Nuclear Factor-κB (NF-κB) and death receptor pathways. Meanwhile, quantitative reverse transcription PCR and Western blot validation results were consistent with the results of transcriptome sequencing analysis. In conclusion, F. necrophorum induced apoptosis and production of pro-inflammatory factors through the NF-κB and death receptor signaling pathway, providing a theoretical basis for further mechanistic studies on the prevention and control of F. necrophorum disease treatment., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Wang, Zhao, He, Jiang, Wang, Xiao, Sun and Guo.)

The Mediterranean climate region of Alentejo in the Southern of Portugal is an important sheep production centre but little is known about the presence and characteristics of Dichelobacter nodosus in association with Fusobacterium necrophorum in the different footrot lesion scores. DNA from 261 interdigital biopsy samples, taken from 14 footrot affected flocks and from three non-affected flocks, were analysed for the presence of D. nodosus and F. necrophorum by real-time PCR. Both virulence and serogroup were determined for 132 and 53 D. nodosus positive biopsy samples, respectively. The co-infection with both bacteria was the commonest epidemiological finding associated with a greater disease severity. There was a statistically significant association (p = 0.002) between footrot-affected flocks and the presence of D. nodosus. Most D. nodosus positive samples were virulent (96.2 %) and belonged to serogroup B (90 %)., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Fusobacterium necrophorum, a Gram-negative anaerobe, is an important bovine pathogen that causes hepatic abscesses, foot rot, mastitis and endometritis. We have previously shown that the 43 kDa outer membrane protein (43 K OMP) of F. necrophorum is a porin protein that plays an important role in bacterial infections; however, the molecular mechanisms by which this protein mediates adhesion remain unclear. In this study, we investigated the role of 43 K OMP in F. necrophorum adhesion to bovine epithelial cells using 43 K OMP-deficient mutants, and identified the protein that interacts with 43 K OMP by immunoprecipitation-mass spectrometry. Our results indicated that the native 43 K OMP and recombinant 43 K OMP could bind to the cell membrane of MAC-T or bovine endometrial epithelial cells (BEECs). When F. necrophorum was preincubated with antibodies against the recombinant 43 K OMP or bovine epithelial cells were preincubated with 43 K OMP, the adhesion of F. necrophorum to MAC-T or BEECs decreased significantly (P<0.01). We successfully constructed a 43 K OMP-deficient strain (A25Δ43 K OMP) and bacterial attachment to MAC-T or BEECs was significantly higher with the F. necrophorum A25 strain than with mutant strain A25Δ43 K OMP (P<0.01). The deficiency of 43 K OMP reduced the binding of F. necrophorum to bovine epithelial cells by 90.5 %-94.9 %. Among the 39 potential differential proteins, fibronectin, collagen and myosin were selected as the target proteins, and direct interaction between 43 K OMP of F. necrophorum and fibronectin was demonstrated. Taken together, these results suggest that 43 K OMP plays a key role in adhesion of F. necrophorum to bovine epithelial cells through its interaction with fibronectin. These findings provide a theoretical basis for the pathogenic mechanism of F. necrophorum., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Fusobacterium necrophorum, a gram-negative anaerobe, causes pharyngotonsillitis primarily in adolescents and young adults (approximately 15-30 years old). The same age group has the highest incidence of peritonsillar abscess and the Lemierre syndrome. The same organism, F. necrophorum, is the most common cause of peritonsillar abscess in this age group and causes at least 80% of Lemierre syndrome cases. We outline the case for empiric antibiotic treatment of some patient in this age group who have a significant probability that F. necrophorum is the cause of their pharyngotonsillitis., Competing Interests: Declaration of competing interest This research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors., (Copyright © 2021 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Fusobacterium necrophorum, a Gram-negative anaerobe, is the primary etiologic agent of liver abscesses of beef cattle. The bacterium, a member of the microbial community of the rumen, travels to the liver via portal circulation to cause abscesses. The severity of liver abscesses vary from mild with one or two small abscesses to severe with medium to large multiple abscesses. Leukotoxin, a secreted protein, is the critical virulence factor involved in the infection. Our objective was to compare leukotoxin production between strains of F. necrophorum isolated from mild and severe liver abscesses collected from slaughtered cattle. The quantification of leukotoxin was based on assays to measure cytotoxicity and protein antigen concentration. One-hundred strains, 50 from mild and 50 from severe abscesses, were utilized in the study. Cell-free supernatants were prepared from cultures grown in anaerobic broth at 9 and 24 h incubations. The leukotoxic activity was quantified by measuring cytotoxicity based on the release of lactic dehydrogenase from bovine lymphocyte cells, BL3, treated with the culture supernatant. Leukotoxin protein concentration was quantified by a sandwich ELISA assay with a leukotoxin-specific monoclonal antibody as the capture antibody. The leukotoxin activity and concentration were highly variable among the strains within each severity of liver abscesses. Although the leukotoxic activity was unaffected by incubation time, leukotoxin protein concentration was consistently higher at 24 h compared to 9 h incubation. Strains from severe liver abscesses had significantly higher leukotoxic activity and higher protein concentration compared to strains from mild liver abscesses (P < 0.0001) at both 9 and 24 h culture supernatants. Across all strains, the correlation coefficients between leukotoxic activity and leukotoxin concentration at 9 and 24 h were 0.14 (P = 0.17) and 0.47 (P < 0.0001), respectively. In conclusion, strains isolated from severe liver abscesses had significantly higher leukotoxic activities and leukotoxin protein concentrations compared to strains isolated from mild liver abscesses., Competing Interests: Declaration of competing interest The authors declare no conflicts of interest., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

A 1-mo-old reticulated giraffe had progressive anorexia and died at the Ordos Zoo. Autopsy revealed necrotic stomatitis with severe bilateral necroulcerative lesions at the base of the tongue and of the cheeks near the commissures of the mouth. There was also severe bilateral confluent bronchopneumonia with a pronounced bronchial pattern and multifocal fibrinous pleuritis. Histologically, there was serofibrinous-suppurative bronchopneumonia with necrosuppurative bronchiolitis and necrotic arteritis. Filamentous bacteria with morphology consistent with Fusobacterium necrophorum were observed at the advancing edge of the necrotic tissue in the tongue and cheeks, as well as in the affected alveolar spaces and bronchioles. Aggregates of slender, gram-negative, rod-like or filamentous bacteria were identified in the lung impression smear. PCR results of 16S rDNA of the tongue and lung lesions had 100% homology to the F. necrophorum subsp. funduliforme B35 sequence (EF447425.1). The gross, histologic, Gram stain, and PCR product sequencing features in our case were consistent with oral and pulmonary necrobacillosis in ruminants, a rare disease of giraffes.

Objectives: We aimed to evaluate the nationwide incidence and a potential increase in invasive infections with Fusobacterium necrophorum. Secondly, we aimed to describe epidemiology, clinical characteristics and outcomes for the different presentations: Lemierre's syndrome (LS), invasive head and neck-infection without LS and invasive non-head and neck-infection., Methods: A retrospective multicentric population-based study of all invasive infections with F. necrophorum diagnosed in Sweden from 2010 to 2017 with 6 months of follow-up was performed through reviews of medical records. Invasive infections were defined and identified by a positive blood culture or sequencing of 16S rDNA, targeted PCR or culture from normally sterile sites. Incidence calculations were performed, including comparisons between 2010-13 and 2014-17, age groups and clinical presentations. Patient and infection characteristics, treatment and clinical outcomes were analysed., Results: Invasive infections with F. necrophorum were diagnosed in 300 cases in Sweden 2010-17. The incidence increased from 2.9 to 5.0 cases/million/year from 2010-13 to 2014-17 (p 0.001). A total of 104/300 (35%) patients developed LS, 102/300 (34%) invasive head and neck infection without LS and 94/300 (31%) invasive non-head and neck infection. The median age was 20, 25 and 64 years, respectively. Among patients with LS 72/96 (75%) had thrombocytopenia on admission, 86/104 (83%) had sepsis, 19/104 (18%) developed septic shock and 45/104 (43%) needed intensive care. 30-day mortality in LS was 2/104 (2%)., Conclusion: We describe an increased incidence of invasive infections with F. necrophorum in Sweden and highlight its full spectrum of invasive clinical presentations. LS, in particular, causes considerable morbidity in young and previously healthy patients., (Copyright © 2019 European Society of Clinical Microbiology and Infectious Diseases. Published by Elsevier Ltd. All rights reserved.)

Here we report an outbreak of an atypical, ulcerative dermatitis in North Country mule lambs, located in South Gloucestershire, UK. The lesions, which appeared to be contagious, occured between the coronary band and the carpal joint as a focal, well demarcated, circular, ulcerative dermatitis. Histopathological examination of the lesion biopsies revealed areas of ulceration, epidermal hyperplasia, suppurative dermatitis and granulation tissue. Clumped keratohyalin granules and intracellular keratinocyte oedema (ballooning degeneration) were evident within lesion biopsies, consistent with an underlying viral aetiology. A PCR-based microbiological investigation failed to detect bovine digital dermatitis-associated treponeme phylogroups, Dichelobacter nodosus, Staphylococcus aureus, Dermatophilus congolensis or Chordopoxvirinae virus DNA. However, 3 of the 10 (30 %) and 6 of 10 (60 %) lesion samples were positive for Fusobacterium necrophorum and Streptococcus dysgalactiae DNA, respectively. Contralateral limb swabs were negative by all standard PCR assays. To better define the involvement of F. necrophorum in the aetiology of these lesions, a qPCR targeting the rpoB gene was employed and confirmed the presence of F. necrophorum DNA in both the control and lesions swab samples, although the mean F. necrophorum genome copy number detected in the lesion swab samples was ∼19-fold higher than detected in the contralateral control swab samples (245 versus 4752 genome copies/μl, respectively; P < 0.001). Although we have not been able to conclusively define an aetiological agent, the presence of both F. necrophorum and S. dysgalactiae in the majority of lesions assayed supports their role in the aetiopathogenesis of these lesions., (Copyright © 2020 The Authors. Published by Elsevier B.V. All rights reserved.)

Background: Non-healing bovine foot lesions, including non-healing white line disease, non-healing sole ulcer and toe necrosis, are an increasingly important cause of chronic lameness that are poorly responsive to treatment. Recent studies have demonstrated a high-level association between these non-healing lesions and the Treponema phylogroups implicated in bovine digital dermatitis (BDD). However, a polymicrobial aetiology involving other gram-stain-negative anaerobes is suspected., Methods: A PCR-based bacteriological survey of uncomplicated BDD lesions (n=10) and non-healing bovine foot lesions (n=10) targeting Fusobacterium necrophorum , Porphyromonas endodontalis, Dichelobacter nodosus and Treponema pallidum/T. paraluiscuniculi was performed., Results: P. endodontalis DNA was detected in 80.0% of the non-healing lesion biopsies (p=<0.001) but was entirely absent from uncomplicated BDD lesion biopsies. When compared to the BDD lesions, F. necrophorum was detected at a higher frequency in the non-healing lesions (33.3% vs 70.0%, respectively), whereas D. nodosus was detected at a lower frequency (55.5% vs 20.0%, respectively). Conversely, T. pallidum/T. paraluiscuniculi DNA was not detected in either lesion type., Conclusion: The data from this pilot study suggest that P. endodontalis and F. necrophorum should be further investigated as potential aetiological agents of non-healing bovine foot lesions. A failure to detect syphilis treponemes in either lesion type is reassuring given the potential public health implications such an infection would present., Competing Interests: Competing interests: None declared., (© British Veterinary Association 2020. Re-use permitted under CC BY. Published by BMJ.)

Fusobacterium species are components of the normal microbiota of the oral cavity, gastrointestinal tract, and female genital tract. They are increasingly recognized as causative agents of oral, laryngeal, and tonsillar infections. Several fusobacterial species are involved in infections, with F. necrophorum and F. nucleatum being the most commonly cultured subtypes. In this study, we aimed to investigate clinical and prognostic differences in terms of mortality and association with malignancy between F. necrophorum and F. nucleatum detected by culture and 16S rRNA gene sequencing. This is a systematic, comparative, retrospective, non-interventional study. Data were extracted from the Department of Clinical Microbiology, Region Zealand, Denmark: all patients with F. necrophorum or F. nucleatum detected by culture or 16S rRNA gene sequencing from 1st of January 2010 to 30th of June 2015 were included. In total, F. necrophorum was detected in samples from 75 patients, and F. nucleatum in samples from 68 patients (total: n = 143). Thirteen patients had a current cancer diagnosis at the time of fusobacterial sampling. Multivariate analyses revealed a significant association of "current cancer" with 30-day mortality. Fusobacterial subtype was not associated with mortality neither in overall nor in subgroups with or without current cancer. Despite differences in clinical disease pattern between F. necrophorum and F. nucleatum, mortality was unaffected by fusobacterial subtype. Mortality was significantly related to comorbidity, especially a current diagnosis of cancer. Our data highlights the current debate whether fusobacterial involvement in cancer may have disease-altering properties, rather than being opportunistic pathogens secondary to cancer disease.

Purpose: Recent data suggest an association between Fusobacterium necrophorum infection and pharyngotonsillitis among adolescents and adults. However, existing reports are only from North America and Europe. We aimed to identify and compare the prevalence of F. necrophorum among patients with pharyngitis and asymptomatic controls in Japan and clarify the epidemiological characteristics of pharyngitis., Methods: Patients aged ≥16 years with pharyngitis and asymptomatic controls were prospectively included. F. necrophorum was detected by using both conventional culture methods and real-time F. necrophorum-specific PCR targeting the rpoB gene. The prevalence of β-hemolytic streptococci was also identified and compared between groups., Results: Forty-four pharyngitis patients and 31 asymptomatic controls were included. F. necrophorum was identified using PCR in 6 (13.6%) pharyngitis cases and 2 (6.5%) controls, with no significant difference (p = 0.457). The median bacterial load of F. necrophorum identified with real-time PCR was significantly higher in pharyngitis cases than in controls (p = 0.046). Patients with a high Centor Score tended to have a higher bacterial load than those with a low Centor Score and controls. In cases of pharyngitis, the prevalence of F. necrophorum was similar to that of Streptococcus pyogenes (F. necrophorum-positive: 6 [13.6%] vs. S. pyogenes-positive: 5 [11.4%], p = 0.99)., Conclusion: F. necrophorum was similarly prevalent among pharyngitis cases as S. pyogenes in Japan. The association of higher F. necrophorum bacterial load with symptomatic pharyngitis in accordance with the previous findings from a different geographical region suggests that F. necrophorum is an important causative agent of bacterial pharyngitis., (Copyright © 2018 Japanese Society of Chemotherapy and The Japanese Association for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.)

Fusobacterium necrophorum has recently been suggested to be associated with tonsillopharyngitis, peritonsillar abscess, and recurrent tonsillitis. Between the 2 subspecies of F. necrophorum, subsp. funduliforme is known to be a major human pathogen. To better understand the epidemiology of F. necrophorum subsp. funduliforme (FNSF), we studied the prevalence of FNSF in the tonsils of patients undergoing elective tonsillectomy (TE) for different indications. Adult patients who underwent elective TE from October 2014 to November 2015 were included. The tonsils were sent for aerobic and anaerobic tissue culture within 30 min of excision; the presence of FNSF was detected using PCR with gyrB primers and 16S rRNA. A total of 32 patients were enrolled. The prevalence of FNSF identified by either culture or gyrB PCR did not significantly differ between infectious and noninfectious TE indications. The constant presence of FNSF might not be associated with recurrent pharyngotonsillitis.

The objective of this study was to determine the prevalence and identification of leukotoxin gene, lktA, variant strains of Fusobacterium necrophorum in the footrot lesions of sheep. The detection of F. necrophorum was carried out by PCR targeting the lktA gene fragment and identification of lktA variant strains was done by PCR-single stranded conformational polymorphism (PCR-SSCP) and gene sequencing. Of the 450 swabs collected from footrot lesions of sheep, 117 were lktA-positive for F. necrophorum. Of the 50 swabs collected from apparently asymptomatic sheep, only one was lktA-positive for F. necrophorum. The overall prevalence of F. necrophorum in footrot affected sheep in Kashmir valley was 26%, and ranged from 20 to 34.8%, respectively. PCR-SSCP of lktA gene fragment analysis revealed three lktA variants, designated as JKS-F1/F2/F3, while two samples (1.7%) showed multiple lktA variant strains of F. necrophorum in a single footrot-affected sheep hoof. This appears to be the first report on the presence of more than one lktA variant of F. necrophorum in a footrot lesion of sheep. The JKS-F3 lktA variant was the most frequent (75.4%), followed by JKS-F2 (14.4%) and JKS-F1 (8.4%), respectively. Among the three lktA variants identified, JKS-F3 was detected in 74 (86.0%) samples from severe footrot affected sheep with a lesion score of 4. The data suggest that JKS-F3 is the predominant lktA variant of F. necrophorum and is associated with severe footrot in sheep. Hence, JKS-F3 may be a significant variant contributing to the severity and duration of the disease in sheep., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Fusobacterium necrophorum (Fn), a gram-negative anaerobe, is increasingly implicated as an etiologic agent in older adolescents and young adults with sore throat. Inadequately treated Fn pharyngitis may result in suppurative complications such as peritonsillar abscess and Lemierre's syndrome. Data from the literature suggest that the incidence of life-threating complications in these age groups from Fn pharyngitis (Lemierre's syndrome) in the United States exceeds those associated with group A beta-hemolytic streptococcal (GAS) pharyngitis (acute rheumatic fever). Using real-time PCR, we previously reported about a 10% prevalence of Fn in asymptomatic medical students and about 20% in students complaining of sore throat at a university student health clinic (p = 0.009). In this study, a comprehensive microbiome analysis of the same study samples confirms that Fn pharyngitis was more common than GAS pharyngitis. Eighteen patients were found to have Fn OTU values exceeding an arbitrary cutoff value of 0.1, i.e. greater than 10% of total sequences, with five subjects reaching values above 0.7. By contrast only 9 patients had GAS OTU values greater than 0.1 and none exceeded 0.6. When the data were analyzed using five separate assessments of alpha diversity, in each case for Fn there were statistically significant differences between Fn positive&#95;high (OTU abundance > 0.1) vs control, Fn positive&#95;high vs Fn negative (OTU abundance = 0), Fn positive&#95;high vs Fn positive&#95;low (OTU abundance > 0 and < 0.1). When the data were analyzed using three beta diversity indexes (Bray-Curtis, weighted unifrac, and unweighted unifrac), there were statistically significant differences between Fn positive&#95;high (OTU abundance ≥ 0.1) vs control for all three. Statistically significant differences remained if we chose somewhat different OTU abundance cutoffs of 0.05 or 0.15. We conclude that Fn appears to play a dominant role in bacterial pharyngitis in the older adolescent and young adult age groups and that the development of a productive mucosal infection with Fn is linked to a significant decrease in the diversity of the associated tonsillar microbiome.

Fusobacterium necrophorum is associated with various diseases in humans and animals. Reservoirs (sites where the pathogen persists in the absence of disease) of F. necrophorum are believed to be present in healthy individuals e.g. tonsillar epithelium, or their environment e.g. soil, but for most diseases the reservoir sites are unknown. Strain typing of F. necrophorum would facilitate linking specific reservoirs with a specific disease. The aim of this study was to develop multiple locus variable number tandem repeat analysis (MLVA) as a strain typing technique for F. necrophorum, and to test the use of this scheme to analyse both isolates and mixed communities of bacteria. Seventy-three tandem repeat regions were identified in the F. necrophorum genome; three of these loci were suitable and developed as a MLVA scheme. The MLVA scheme was sensitive, specific, and discriminatory for both isolates and communities of F. necrophorum. The MLVA scheme strain typed 46/52F. necrophorum isolates including isolates of both subspecies and from different countries, host species and sample sites within host. There were 12 unique MLVA strain types that clustered by subspecies. The MLVA scheme characterised the F. necrophorum community in DNA from 32/49 foot- and 28/33 mouth swabs from sheep. There were 17 community types in total. In 31/32 foot swabs, single strains of F. necrophorum were detected while in the 28 mouth swabs there were up to a maximum of 8 strains of F. necrophorum detected. The results demonstrate the potential for this method to elucidate reservoirs of F. necrophorum., (Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.)

Leukotoxin is a well-known virulence factor of animal isolates of Fusobacterium necrophorum subspecies necrophorum, and is also expressed by animal isolates of subspecies funduliforme, whereas its presence in isolates from humans has not been fully established. In this study we found that the leukotoxin gene was present in all tested F. necrophorum isolates from humans. Three sequence variants were found, two of which have not been described previously. The sequence types correlated to source of infection. Further studies are needed to examine the role of the leukotoxin in human infections., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Pyometra is a common disease of cattle that causes infertility and thereby financial losses to the cattle industry. Bacteria involved in the development and progression of pyometra have been investigated by microbial culture but their tissue invading abilities, which is an important aspect of bacterial pathogenicity and development of lesions, have not been investigated. Bacterial invasion of the uterus and oviducts was studied in 21 cows diagnosed with pyometra at the time of slaughter by applying fluorescence in situ hybridization using probes targeting 16S ribosomal RNA of Fusobacterium necrophorum, Porphyromonas levii, Trueperella pyogenes and the overall bacterial domain Bacteria. Fusobacterium necrophorum and P. levii were found to invade the endometrium, especially if the endometrium was ulcerated, and penetrated deep into the lamina propria. These species co-localized within the tissue thus indicating a synergism. Trueperella pyogenes did not invade the uterine tissue. In addition to endometrial lesions, most cows with pyometra also had salpingitis but without significant bacterial invasion of the oviductal wall., (Copyright © 2017 Elsevier Inc. All rights reserved.)

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., (Copyright © 2015 The Authors. Published by Elsevier B.V. All rights reserved.)

Bovine digital dermatitis (BDD) is an infective foot disease commonly reported in dairy cattle where Treponema are considered as the primary causative infectious agents. There still remains little definitive information on the etiology of BDD in beef cattle suggesting further investigations are warranted. Beef BDD lesions (n=34) and healthy beef foot tissues (n=38) were analysed by PCR for three BDD-associated Treponema phylogroups and also for Dichelobacter nodosus and Fusobacterium necrophorum. Spirochete culture was attempted on all BDD lesion samples. One or more BDD-associated Treponema phylogroups were detected in 100% of beef BDD lesions. "Treponema medium/Treponema vincentii-like", "Treponema phagedenis-like" and Treponema pedis spirochetes were identified in 27/34 (79%), 31/34 (91%) and 24/34 (71%) of BDD lesions, respectively. No BDD-associated treponeme DNA was amplified from beef healthy foot tissues. D. nodosus and F. necrophorum were present in 24/34 (71%) and 15/34 (44%) of lesions and 10/38 (26%) and 12/38 (32%) of healthy foot tissues, respectively. Twenty spirochetes were isolated from beef BDD lesions; 19 were representatives of the three BDD-associated Treponema phylogroups. One spirochete isolate shared less than 97% 16S rRNA gene similarity to the three cultivable BDD-associated Treponema phylogroups and therefore may represent a novel taxa of Treponema. Upon comparison, sheep contagious ovine digital dermatitis (CODD), dairy cattle and beef cattle BDD lesions appear to have extremely similar bacteriological data and therefore provides evidence of a shared etiopathogenesis posing concerns for cross-species transmission., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Fusobacterium necrophorum is a Gram-negative obligate anaerobe associated with several diseases in humans and animals. Despite its increasing clinical significance, there is little or no data on the relationship between its metabolism and virulence. Previous studies have shown that bacteria grown under iron-limitation express immunogenic antigens similar to those generated in vivo. Thus, this paper describes the relationship between F. necrophorum subsp. necrophorum (Fnn) metabolism and the expression of the encoded putative virulence factors under iron-restricted conditions. At the midlog phase, iron limitation reduced Fnn growth but the cell density was dependent on the size of the inoculum. Preferential utilization of glucose-1-phosphate, d-mannitol and l-phenylalanine; production of 2-hydroxycaproic acid and termination of dimethyl sulphide production were major Fnn response-factors to iron limitation. Ultimately, iron restriction resulted in an increased ability of Fnn to metabolize diverse carbon sources and in the expression of stress-specific virulence factors. Iron starvation in low Fnn cell density was associated with the up-regulation of haemagglutinin (HA) and leukotoxin (lktA) genes (2.49 and 3.72 fold change respectively). However, Fnn encoded Haemolysin (Hly), yebN homologue (febN) and tonB homologue, were down-regulated (0.15, 0.79 and 0.33, fold changes respectively). Interestingly, cell density appeared to play a regulatory role in the final bacteria cell biomass, induction of a metabolic gene expression and the expression pattern virulence factors in Fnn suggesting the role of a cell density-associated regulatory factor. This report suggest that future studies on differential expression of bacterial genes under altered environmental condition(s) should consider testing the effect of cell concentrations as this is often neglected in such studies. In conclusion, iron restriction induces preferential utilization of carbon sources and altered metabolism in Fnn with associated changes in the expression pattern of the virulence factors., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Fusobacterium necrophorum, a Gram-negative anaerobe, is the primary etiologic agent of liver abscesses in cattle. There are two subspecies; subsp. necrophorum and subsp. funduliforme, which differ in morphological, biochemical, molecular characteristics, and virulence. The subsp. necrophorum, which is more virulent, occurs more frequently in liver abscesses than the subsp. funduliforme. Bacterial adhesion to the host cell surface is critical to the pathogenesis of several bacterial infections, and in F. necrophorum, outer membrane proteins (OMP) have been shown to mediate adhesion to bovine endothelial cells. The objective of this study was to identify potential adhesins that are involved in adhesion of F. necrophorum subsp. necrophorum to the host cells. An OMP of 42.4 kDa, which binds with high affinity to the bovine endothelial cells and is recognized by the sera from cattle with liver abscesses, was identified. N-terminal sequencing of the protein showed 96% homology to the FomA protein of F. nucleatum. The PCR analysis showed that this fomA gene was present in several strains of subsp. necrophorum, subsp. funduliforme of bovine and subsp. funduliforme of human origin. The purified native and recombinantly expressed protein when preincubated with the endothelial cells, prevented the attachment of subsp. necrophorum significantly. In addition, the polyclonal antibody produced against the protein prevented the binding of subsp. necrophorum to bovine endothelial cells., (Copyright © 2014 Elsevier B.V. All rights reserved.)

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., (Copyright © 2014. Published by Elsevier B.V.)

Oral necrobacillosis (ON) is a model polymicrobial disease that affects macropods in captivity and livestock. Several studies in humans and animals have focused mainly on the bacterial etiology of this disease with little or no information on the role/association of fungi with ON. Using a Polymerase Chain Reaction-Denaturing Gradient Gel Electrophoresis (PCR-DGGE) assay and statistical analysis of the fungal community structure in healthy and disease groups, a reduction in the species diversity and drastic reduction (>1000 fold) in the fungal population in wallabies with ON was observed. Furthermore, an in vitro assay revealed a potential anaerobic-bacteria antibiosis mechanism in the observed decrease in fungal population in ON and a synergistic bacterial-fungal interaction in wallabies with healthy oral status. This study contributes to our knowledge of the fungal community structure associated with ON and forms the basis for an investigation at an epidemiological scale in order to exploit the clinical potentials of these findings., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

We report the case of a 71-year-old woman who presented a primary spinal epidural abscess caused by Fusobacterium necrophorum. This is the second report in the medical literature to associate this organism with a primary spinal epidural abscess without spondylodiscitis. After treatment with emergency laminectomy followed by 8 weeks of antibiotic treatment the patient was cured. Oral metronidazole (500 mg every 8 h) was the definitive choice of treatment. F. necrophorum spinal epidural abscess is rare, although samples for anaerobic culture should be collected in order to improve detection of anaerobic spinal infections. PCR amplification and sequencing of the 16S rRNA permits early diagnosis in anaerobic infections., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

A pair of primers was designed in an attempt to amplify outer membrane protein (OMP) gene of Fusobacterium necrophorum based on nucleotide sequence of the OMP of Fusobacterium nucleatum. Further analysis was performed to characterize its molecular properties and phylogeny in the genus Fusobacterium. We identified a predicated 43kDa outer membrane protein (43K OMP) in F. necrophorum, which showed the same properties as other pore-forming proteins of Gram-negative anaerobic bacteria according to analysis of signal peptide, AT-rich, membrane-spanning region and conserved motifs. The predicated 43K OMP exhibited 70.22%, 62.04%, 56.75%, 58.72%, 51.59%, 31.49% and 50.26% amino acid identity with the OMPs of F. nucleatum, Fusobacterium varium, Fusobacterium ucerans, Fusobacterium periodonticum, Fusobacterium mortiferum, Fusobacterium gonidiaformans and F. necrophorum (hypothetical protein), respectively. 11 common conserved domains and 10 common variable domains were found among the 45 aligned OMPs of Fusobacterium species. Distributions of the conserved and variable domains were highly associated with predicted membrane-spanning regions, cell surface exposed regions and B-cell epitope regions. Phylogenetic analysis revealed the predicated 43K OMP of F. necrophorum was closely related with the OMPs from F. nucleatum and F. periodonticum. These data will increase understanding of pathogenesis and genetic evolution of F. necrophorum., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Fusobacterium necrophorum, a Gram-negative, anaerobic bacterium, is an opportunistic animal and human pathogen that causes a variety of infections termed necrobacillosis. There are 2 subspecies of F. necrophorum (subsp. necrophorum and subsp. funduliforme) that differ morphologically and biochemically and in virulence. Leukotoxin, a secreted protein, is considered to be the major virulence factor. In camelids, F. necrophorum causes a variety of infections, generally involving the lips, tongue, pharynx, interdigital spaces, foot pad, larynx, mandible, or maxillary bones. The objective of the current study was to characterize the presumptive Fusobacterium isolates from a variety of necrotic infections in llama (Lama glama) and alpaca (Vicugna pacos) and determine whether the strains possess leukotoxin activities. A total of 7 isolates from alpaca and 2 isolates from llama were characterized. Based on growth characteristics in broth culture, and biochemical and polymerase chain reaction analyses, all 9 isolates belonged to subsp. necrophorum and possessed the putative hemagglutinin gene. Western blot analysis with antileukotoxin antibodies raised in rabbit showed the presence of leukotoxin protein in the culture supernatant of all isolates. Furthermore, flow cytometry of the culture supernatants demonstrated cytotoxicity to bovine and alpaca polymorphonuclear leukocytes (PMNs). The extent of cytotoxicity to either alpaca or bovine PMNs differed among camelid strains. The cytotoxicity of many of the camelid strains was higher (P < 0.05) toward alpaca PMNs compared to bovine PMNs. Fusobacterium necrophorum isolates from llama and alpaca are similar to bovine isolates, and leukotoxin may be a major virulence factor.

Periodontal diseases (PD) are diseases of polymicrobial aetiology and constitute major health problems in captive macropods. Increasing knowledge of the causal pathogens is therefore crucial for effective management and prevention of these diseases. PCR survey and sequence analyses of potential periodontopathogens in captive wallaby populations revealed a co-incidence of the diseases with the detection of Fusobacterium necrophorum subsp. necrophorum (Fnn) and its encoded leukotoxin (lktA) gene. Sequence analyses showed that the outer membrane protein of Fnn in the GenBank database shared significant homology (99%) with the Fnn encoded haemagglutinin-related-protein gene fragment identified in this study. In addition, this report suggests the existence of a variant of Fnn with no detectable lktA gene and thus warrants further studies. In contrast to reports associating Porphyromonas gingivalis and F. nucleatum with PD, this study revealed that PD in macropods are associated with Porphyromonas gulae and Fnn and raises the question: is there a possible host pathogen co-evolution in the pathogenesis of PD in animals and humans? These findings contribute to the understanding of the aetiology of periodontal disease in macropods as well as opening up a new direction of research into the microbial interactions involved in the pathogenesis of PD in macropods., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Fusobacterium necrophorum, a Gram-negative anaerobe, is frequently associated with suppurative and necrotic infections of animals and humans. The organism is a major bovine pathogen, and in cattle, the common fusobacterial infections are hepatic abscesses, foot rot, and necrotic laryngitis. The species comprises two subspecies: F. necrophorum subsp. necrophorum and F. necrophorum subsp. funduliforme. Bacterial adhesion to the host cell surface is a critical initial step in the pathogenesis, and outer membrane proteins (OMP) play an important role in adhesion and establishment of certain Gram-negative bacterial infections. The means by which F. necrophorum attaches to epithelial or endothelial cells has not been determined. We evaluated whether OMP of F. necrophorum, isolated from a liver abscess, mediated adhesion to bovine endothelial cells (adrenal gland capillary endothelial cell line). The extent of binding of subsp. necrophorum to the endothelial cells was higher than that of F. necrophorum subsp. funduliforme. Trypsin treatment of bacterial cells decreased their binding to endothelial cells indicating the protein nature of adhesins. Preincubation of endothelial cells with OMP extracted from F. necrophorum decreased the binding of bacterial cells. In addition, binding of each subspecies to endothelial cells was inhibited by polyclonal antibodies raised against respective OMP and the antibody-mediated inhibition was subspecies specific. The western blot analysis of OMP bound to endothelial cells with anti-OMP antibodies showed four OMP of 17, 24, 40 and 74 kDa. We conclude that OMP of F. necrophorum play a role in adhesion of bacterial cells to the endothelial cells., (Copyright © 2012 Elsevier B.V. All rights reserved.)

The aim of this study was to determine which of the two species, Fusobacterium necrophorum or Dichelobacter nodosus, are associated with hoof thrush in horses. Fourteen hoof samples, collected from eight horses with thrush and 14 samples collected from eight horses with healthy hooves, were examined for the presence of F. necrophorum, Fusobacterium equinum and D. nodosus. Only isolates with phenotypic characteristics representing Fusobacterium could be cultured. Total DNA extracted from the 28 hoof samples was amplified by using DNA primers designed from gene lktA, present in F. necrophorum subsp. necrophorum, F. necrophorum subsp. funduliforme and F. equinum, and gene fimA, present in D. nodosus. The lktA gene was amplified from five of the 14 infected hoof samples and from one hoof sample without thrush. The DNA sequence of the amplified ltkA gene was identical to the lktA gene of the type strain of F. necrophorum (GenBank accession number AF312861). The isolates were phenotypically differentiated from F. equinum. No DNA was amplified using the fimA primer set, suggesting that F. necrophorum, and not D. nodosus, is associated with equine hoof thrush. Hoof thrush in horses is thus caused by F. necrophorum in the absence D. nodosus. This is different from footrot in sheep, goats, cattle and pigs, which is caused by the synergistic action of F. necrophorum and D. nodosus., (Copyright © 2012 Elsevier B.V. All rights reserved.)

The objective of this study was to evaluate the relationship between bacterial species-specific virulence factors (VFs) present in the uterus at 3 different stages of lactation (1-3, 8-10, and 34-36 days in milk (DIM)) and the incidence of metritis and clinical endometritis in dairy cows. The following VF genes were investigated: plo (pyolysin), cbpA (collagen-binding protein), and fimA (fimbriae expression) which are Arcanobacterium pyogenes specific; fimH (a type 1 pilus component), Escherichia coli specific; and lktA (leukotoxin), Fusobacterium necrophorum specific. Uterine swabs were collected from 111 postpartum dairy cows. PCR was used to detect the presence of plo, cbpA, fimA, fimH, and lktA genes. A. pyogenes cbpA was detected in only 5 samples and therefore was not subjected to further analysis. E. coli (fimH) was significantly associated with metritis and endometritis when detected at 1-3 DIM; F. necrophorum (lktA) was significantly associated with metritis when detected at 1-3 and 8-12 DIM and with endometritis when detected at 34-36 DIM; and A. pyogenes (fimA and plo) was associated with metritis (fimA) when detected at 1-3 DIM and endometritis (fimA and plo) when detected at 8-10 and 34-36 DIM., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Footrot is a bacterial disease that has substantial economic and welfare impacts in sheep and can be difficult to manage. Research is focussed on reducing the impact that footrot has on farmers and their flocks and better understanding the aetiology of the disease. Key areas of current research include, developing better vaccines, deploying tailored vaccines in a specific and targeted fashion on individual farms, analysing and developing better farm management practices to suit specific sheep farming environments, elucidating the virulence genes and bacterial population dynamics that drive footrot and using genetic testing in combination with selective breeding to produce stock that are more resilient to disease., (Copyright © 2010 Elsevier B.V. All rights reserved.)

Human and animal bites may lead to serious infection. The organisms involved tend to originate from the oral cavity of the offending biter, as well as the environment where the injury occurred. A variety of aerobic as well as anaerobic organisms have been isolated from bite wounds, with infection ranging from localized cellulitis to systemic dissemination, leading to severe disease ranging from abscess to bone and joint infection, to endocarditis and brain abscess. Immediate wound management, including recognition of the most commonly associated infectious pathogens, and judicious use of empiric antibiotics are crucial in providing the best care after a bite. Here, we discuss the common animal bite associated infections, and provide the most up to date information regarding their management.

Photonic crystals (PCs) are periodic dielectric structures that have a band-gap that forbids propagation of a certain range of wavelengths of light. This property enables control of light with remarkable facility by modification of the band-gaps and produce effects that are impossible with conventional optics. Using chemically functionalized PCs, where the chemical functional group consists of amine and carboxyl group, in conjunction with a biomolecular probe material, the detection of pathogens and viral disease is possible, indicated by the shift in wavelength signal. Moreover, this system using the bioinspired PCs allows specific target detection in biosensor chip fields through control of the PCs. In this study, we demonstrated that two bacterial pathogens (Fusobacterium necrophorum and Acinetobacter baumannii) causing sepsis were detected by DNA-probe hybridization and a severe acute respiratory syndrome coronavirus was detected by antigen-antibody interaction using the functional PCs. Optical readout with the integrated sensor detecting the signals from PCs, allows for low cost and robust readout of resonance peak shift. This biosensor system using the functional PCs on the photonic crystal-fabricated chip can efficiently and effectively detect various targets, and be easily prepared with high productivity and economic property.

Within the last decade, Fusobacterium necrophorum subsp. funduliforme has been considered a clinically important pathogen causing pharyngitis especially in adolescents and young adults. F. necrophorum pharyngitis can progress into Lemierre's syndrome, which is a severe and life-threatening infection. However, throat swabs are not cultured anaerobically in the routine and even if cultured anaerobically, it can be difficult to identify F. necrophorum from the normal flora of the throat. F. necrophorum is therefore often overlooked as the cause of pharyngitis. In our laboratory, a F. necrophorum selective agar has been developed containing vancomycin and nalidixin, which inhibit the growth of most Gram-positive and many Gram-negative bacteria, respectively. β-haemolysis of horse blood can be detected, which further facilitates the detection and identification of F. necrophorum. The F. necrophorum selective agar was evaluated against a quantitative real-time polymerase chain reaction assay and shown to have a significantly higher sensitivity for detecting F. necrophorum than the anaerobic agar commonly used in Denmark. Furthermore, the F. necrophorum selective agar does not require experienced laboratory technicians, require fewer subcultures, is probably less expensive and is faster to perform than other culture methods., (© 2010 The Authors. APMIS © 2010 APMIS.)

Fusobacterium necrophorum is the main pathogen that causes numerous necrobacilloses. Hemolysin is one of the major virulence factors involved in fusobacterial infections. In order to investigate the genetic basis of hemolytic activity and the regulation mechanism of the hemolysin expression, a genomic library was constructed from F. necrophorum DNA by ligating DNA fragments generated by partial HindIII digestion with pUC18 vector. The screening of the genomic library with polymerase chain reaction, DNA sequencing and sequence assembly led to a 7.45 kb sequence which includes the putative hly gene and upstream sequence. Clustered putative genes encoding short chain acyl-CoA dehydrogenase (Scad) and electron transfer flavoprotein (Etf) alpha and beta subunits locate upstream of hly. A 535 bp non-coding sequence, possibly with some cis-regulatory elements involved in the regulation of the hemolysin expression in F. necrophorum, locates between etf-beta and hly. The nucleotide sequence of the hly gene indicates it encodes hemolysin. It is the first characterized hemolysin coding gene in F. necrophorum., (Copyright (c) 2010 Elsevier Ltd. All rights reserved.)

Outbreaks of digital necrobacillosis in Norwegian wild tundra reindeer (Rangifer tarandus tarandus) are described. The outbreaks occurred in late summer and autumn 2007 and 2008, subsequent to periods with an unusually high number of days with precipitation and high air temperature. Lesions were generally restricted to one foot and the disease incidence was highest in calves. Single limbs from 20 animals and six whole carcasses were submitted for laboratory examination. Gross lesions were characterized by swelling of the fetlock to coronary band area and cutaneous sinus tracts with sparse discharge of pus. Subcutaneous tissue was inflamed and oedematous with focal necrosis. Tendons, tendon sheaths, joints and periosteum of the digital bones were often affected. Animals shot during winter showed severe chronic periostitis and osteomyelitis and necrotizing deforming arthritis. Microscopically, skin lesions were characterized by deep ulcers with centrally located necrotic tissue, bordered by a zone of oedema and intense inflammation with granulation tissue and fibrosis. Necrosis, suppurative inflammation and oedema were found in the synovial membranes, tendons and tendon sheaths. Digital bone lesions were characterized by necrosis, fibrosis and extensive bone proliferation. Vasculitis and thrombosis were common in all lesions. Elongate filamentous gram-negative bacteria in necrotic lesions from all animals were identified as Fusobacterium necrophorum by fluorescence in-situ hybridization. F. necrophorum was cultured from the foot lesions of six animals. Five of these isolates were examined by 16S rRNA sequencing. The sequences were identical and differed from all other strains listed in GenBank. These results are consistent with circulation of a reindeer-adapted pathogenic strain of F. necrophorum in the wild reindeer population, causing outbreaks of digital necrobacillosis following warm and humid summers., ((c) 2010 Elsevier Ltd. All rights reserved.)