Emayella augustorita, New Member of Pasteurellaceae, Isolated from Blood Cultures of Septic Patient.

We report discovery of a new bacterial genus and species of the family Pasteurellaceae by using phylogenetic and metabolic analysis. The bacterium, Emayella augustorita, was isolated from blood cultures of a patient in France diagnosed with an adenocarcinoma of the intestines and who was treated with a biliary prosthesis placement.

Aggregatibacter actinomycetemcomitans infection in children: two case reports and a review of the literature.

Aggregatibacter actinomycetemcomitans (Aa), a Gram-negative coccobacillus commonly associated with endocarditis, poses a rare diagnostic challenge in pediatric cases. The presentation of two pediatric cases-myositis and chest mass-highlights novel aspects, including unusual symptom presentations in children which can be mistaken for malignancy. The limited sensitivity of standard blood tests complicates diagnosis, leading to delayed diagnosis and treatment. Representative samples must be taken, especially if blood cultures are negative. Despite advances in detection methods, diagnosing Aa infection remains difficult due to its rarity in children and variable clinical presentation. In conclusion, a comprehensive understanding of Aa infection in children is essential for early and effective diagnostic and therapeutic management.

Atypical post-infectious glomerulonephritis with c-ANCA positivity followed by endocarditis.

Post-infectious glomerulonephritis (PIGN), an uncommon variety of glomerulonephritis (GN), is characterized by emergence of nephritic syndrome within a few weeks following an infectious event. PIGN typically presents as a mild condition and tends to resolve by the time of diagnosis for GN. Aggregatibacter actinomycetemcomitans belongs to the HACEK group of bacteria, which constitutes less than 3% of bacteria responsible for community-acquired infective endocarditis. We present a case of 29-year-old man suspected of lymphoma with B-symptoms along with severe splenomegaly and nephromegaly. Shortly after, he developed an episode of nephritic syndrome accompanied by acute kidney injury (AKI) and high titers of cytoplasmic ANCA (c-ANCA)-positivity. Kidney biopsy revealed PIGN with tubulointerstitial nephritis. Despite treatment with antibiotics and corticosteroid, he visited the emergency room due to worsening dyspnea and multi-organ failure. An echocardiogram showed a bicuspid aortic valve with vegetation unseen on previous echocardiogram. He underwent aortic valve replacement immediately without adverse events. Four months after valve replacement, his renal function and cardiac performance have remained stable. We report a case of PIGN with AKI and high titers of c-ANCA appearing later as an infective endocarditis due to Aggregatibacter actinomycetemcomitans. With careful clinical observation and appropriate and timely management, satisfactory outcomes for patient health are possible.

Mannheimia haemolytica-associated fibrinonecrotizing abomasitis in lambs.

Mannheimia haemolytica-associated abomasitis has been clinically described as a cause of sudden death in lambs, but it is poorly characterized. We describe the pathological features of a severe fibrinonecrotizing abomasitis in 3 lambs that died suddenly. All 3 abomasums had a thickened submucosa due to edema and necrotic areas delimited by bands of degenerate neutrophils with slender nuclei (oat cells) and angiocentric distributions. The overlying mucosa was congested. Myriads of gram-negative coccobacilli were observed within the oat cell bands. M. haemolytica was isolated from the abomasum in all 3 animals and was serotyped as A2 in one of them. Pericarditis and pleuritis were observed in 2 of the lambs. Clostridium spp. were isolated in 1 lamb and detected by immunohistochemistry in the 3 animals, suggesting clostridial co-infection. M. haemolytica should be considered among the differential diagnoses of necrotizing abomasitis in lambs.

Gallibacterium anatis infection in poultry: a comprehensive review.

Gallibacterium anatis (G. anatis), a member of the Pasteurellaceae family, normally inhabits the upper respiratory and lower genital tracts of poultry. However, under certain circumstances of immunosuppression, co-infection (especially with Escherichia coli or Mycoplasma), or various stressors, G. anatis caused respiratory, reproductive, and systemic diseases. Infection with G. anatis has emerged in different countries worldwide. The bacterium affects mainly chickens; however, other species of domestic and wild birds may get infected. Horizontal, vertical, and venereal routes of G. anatis infection have been reported. The pathogenicity of G. anatis is principally related to the presence of some essential virulence factors such as Gallibacterium toxin A, fimbriae, haemagglutinin, outer membrane vesicles, capsule, biofilms, and protease. The clinical picture of G. anatis infection is mainly represented as tracheitis, oophoritis, salpingitis, and peritonitis, while other lesions may be noted in cases of concomitant infection. Control of such infection depends mainly on applying biosecurity measures and vaccination. The antimicrobial sensitivity test is necessary for the correct treatment of G. anatis. However, the development of multiple drug resistance is common. This review article sheds light on G. anatis regarding history, susceptibility, dissemination, virulence factors, pathogenesis, clinical picture, diagnosis, and control measures.

Comparative study of antibacterial activity and stability of D-enantiomeric and L-enantiomeric bovine NK-lysin peptide NK2A.

L-enantiomers of antimicrobial peptides (AMPs) are sensitive to proteolytic degradation; however, D-enantiomers of AMPs are expected to provide improved proteolytic resistance. The present study aimed to comparatively investigate the in vitro antibacterial activity, trypsin and serum stability, toxicity, and in vivo antibacterial activity of L-enantiomeric bovine NK2A (L-NK2A) and its D-enantiomeric NK2A (D-NK2A). Circular dichroism spectroscopy of D-NK2A and L-NK2A in anionic liposomes showed α-helical structures and the α-helical conformation of D-NK2A was a mirror image of L-NK2A. Both D-NK2A and L-NK2A displayed minimal in vitro and in vivo toxicities. RP-HPLC and mass spectrometry analyses revealed that D-NK2A, but not L-NK2A, was resistant to trypsin digestion. D-NK2A and L-NK2A showed similar in vitro bacterial killing activities against Histophilus somni. Slightly reduced antibacterial activity was observed when D-NK2A and L-NK2A were pre-incubated with serum. Confocal and transmission electron microscopic findings confirmed that both peptides induced disruption of bacterial inner- and outer-membranes. Improved survivals with D-NK2A treatment were observed when compared to L-NK2A in a murine model of acute H. somni septicemia. We conclude that antibacterial activity and mode of action of NK2A are not chiral specific. With further optimization, D-NK2A may be a viable AMP candidate to combat bacterial infections.

Host Chromatin Regulators Required for Aggregatibacter actinomycetemcomitans Cytolethal Distending Toxin Activity in Saccharomyces cerevisiae Model.

Cytolethal distending toxin (CDT) is a bacterial genotoxin that causes host cell cycle arrest and death. We previously employed a Saccharomyces cerevisiae model with inducible expression of the CDT catalytic subunit from Aggregatibacter actinomycetemcomitans, AaCdtB, and showed that a wide variety of host factors play a role in facilitating the activity of CdtB. Our observation that a yeast H2B mutant defective in chromatin condensation was partially resistant to CdtB implies that chromatin structure may affect CDT function. In this study, we identified host chromatin regulatory genes required for CdtB cytotoxicity. We found that the deletion of HTZ1 or certain subunits of SWR, INO80, and SIR complexes increased cellular resistance to CdtB. We hypothesized that CdtB may interact with Htz1 or the chromatin, but immunoprecipitation experiments failed to detect physical interaction between CdtB and Htz1 or the chromatin. However, we observed reduced nuclear localization of CdtB in several mutants, suggesting that impaired nuclear translocation may, at least partly, explain the mechanisms of CdtB resistance. In addition, mutations in chromatin regulatory genes induce changes in the global gene expression profile, and these may indirectly affect CdtB toxicity. Our results suggest that decreased expression of endoplasmic reticulum (ER)-Golgi transport-related genes that may be involved in CdtB transport and/or increased expression of DNA repair genes may contribute to CdtB resistance. These results suggest that the functions of chromatin regulators may contribute to the activity of CDT in host cells.

Glaesserella parasuis serotype 4 HPS4-YC disrupts the integrity of the swine tracheal epithelial barrier and facilitates bacterial translocation.

Glaesserella parasuis (G. parasuis) is a commensal bacterium in the upper respiratory tract of pigs that can also cause the swine Glässer disease, which induces an intensive inflammatory response and results in significant economic losses to the swine industry worldwide. G. parasuis can cause disease through infection of the respiratory tract, resulting in systemic infection, but the mechanism is largely unknown. Recently we showed that Glaesserella parasuis serotype 4 (GPS4) increased swine tracheal epithelial barrier permeability, resulting in easier bacterial translocation. Tight junction proteins (TJ) play a crucial role in maintaining the integrity and impermeability of the epithelial barrier. GPS4 decreased the expression of the TJ ZO-1 and occludin in swine tracheal epithelial cells (STEC). Furthermore, the proinflammatory cytokines IL-6, IL-8 and TNF-α were significantly upregulated in GPS4-infected STEC, and both the MAPK and NF-κB signaling pathways were activated and contributed to the expression of TNF-α. We demonstrate that the production of proinflammatory cytokines, especially TNF-α, during GPS4 infection was involved in barrier dysfunction. Additionally, animal challenge experiments confirmed that GPS4 infection downregulated TJ in the lungs of piglets and induced a severe inflammatory response. In general, G. parasuis infection downregulated the expression of TJ and induced massive secretion of proinflammatory cytokines, resulting in epithelial barrier disruption and favoring bacterial infection. This study allowed us to better understand the mechanism by which G. parasuis crosses the respiratory tract of pigs.

Clonal spread of multi-resistant Gallibacterium anatis isolates among Iranian broilers and layers.

Gallibacterium anatis is a common cause of reproductive tract infection in chickens, which leads to reduced egg production and increased mortality. This study was undertaken to investigate prevalence of G. anatis in 12 poultry flocks originating from Iranian provinces with leading chicken production and to determine genetic diversity, antimicrobial resistance, and the presence of major antigens of the isolates investigated. Out of the 120 chicken tracheal samples collected and tested, 84 (70%) were positive for G. anatis. Genotyping by Pulse Field Gel Electrophoresis and genome sequencing revealed a total of 24 pulsotypes for 71 strains (at a 87% similarity level) and seven genome clusters comprising 21 strains (97% similarity level), respectively. The combination of the two typing methods confirmed the presence of several genotypes originating from a common ancestor affecting poultry yet also suggested that identical clones were shared among chickens within farms and between different farms. The latter finding is to our knowledge the first example of clonal presence of G. anatis in epidemiologically unrelated farms. The 21 sequenced strains were characterized against a panel of commonly used antibiotics and showed lowered sensitivity to tetracycline (76.2%) and enrofloxacin (90.5%). The widespread presence of multiresistant G. anatis isolates calls for non-antibiotic prophylactics. Three major immunogen genes, gtxA, Gab_1309 and Gab_2312 were detected in the isolates indicating these antigens likely represent effective vaccine targets. A conserved sequence of the gtxA gene across a range of epidemiologically independent strains suggests the use of GtxA for future vaccine development purposes.

On-farm colorimetric detection of Pasteurella multocida, Mannheimia haemolytica, and Histophilus somni in crude bovine nasal samples.

This work modifies a loop-mediated isothermal amplification (LAMP) assay to detect the bovine respiratory disease (BRD) bacterial pathogens Pasteurella multocida, Mannheimia haemolytica, and Histophilus somni in a colorimetric format on a farm. BRD causes a significant health and economic burden worldwide that partially stems from the challenges involved in determining the pathogens causing the disease. Methods such as polymerase chain reaction (PCR) have the potential to identify the causative pathogens but require lab equipment and extensive sample processing making the process lengthy and expensive. To combat this limitation, LAMP allows accurate pathogen detection in unprocessed samples by the naked eye allowing for potentially faster and more precise diagnostics on the farm. The assay developed here offers 66.7-100% analytical sensitivity, and 100% analytical specificity (using contrived samples) while providing 60-100% concordance with PCR results when tested on five steers in a feedlot. The use of a consumer-grade water bath enabled on-farm execution by collecting a nasal swab from cattle and provided a colorimetric result within 60 min. Such an assay holds the potential to provide rapid pen-side diagnostics to cattle producers and veterinarians.

Protective Effects of Baicalin on Peritoneal Tight Junctions in Piglets Challenged with Glaesserella parasuis.

Glaesserella parasuis (G. parasuis) causes inflammation and damage to piglets. Whether polyserositis caused by G. parasuis is due to tight junctions damage and the protective effect of baicalin on it have not been examined. Therefore, this study aims to investigate the effects of baicalin on peritoneal tight junctions of piglets challenged with G. parasuis and its underlying molecular mechanisms. Piglets were challenged with G. parasuis and treated with or without baicalin. RT-PCR was performed to examine the expression of peritoneal tight junctions genes. Immunofluorescence was carried out to detect the distribution patterns of tight junctions proteins. Western blot assays were carried out to determine the involved signaling pathways. Our data showed that G. parasuis infection can down-regulate the tight junctions expression and disrupt the distribution of tight junctions proteins. Baicalin can alleviate the down-regulation of tight junctions mRNA in peritoneum, prevent the abnormalities and maintain the continuous organization of tight junctions. Our results provide novel evidence to support that baicalin has the capacity to protect peritoneal tight junctions from G. parasuis-induced inflammation. The protective mechanisms of baicalin could be associated with inhibition of the activation of PKC and MLCK/MLC signaling pathway. Taken together, these data demonstrated that baicalin is a promising natural agent for the prevention and treatment of G. parasuis infection.

MOLECULAR IDENTIFICATION OF MEMBERS OF THE FAMILY PASTEURELLACEAE FROM THE ORAL CAVITY OF KOALAS (PHASCOLARCTOS CINEREUS) AND THEIR RELATIONSHIP WITH ISOLATES FROM KOALA BITE WOUNDS IN HUMANS.

A total of 22 Pasteurellaceae isolates obtained from the oral cavity of koalas (Phascolarctos cinereus) at different wildlife centers in Australia were investigated using amplification and sequencing of two housekeeping genes, rpoA and recN. The available sequences from the Lonepinella koalarum type strain (ACM3666T) and the recent isolates of Lonepinella-like bacteria obtained from human infected wounds associated with koala bites were also included. Phylogenetic analysis was performed on the concatenated rpoA-recN genes and genome relatedness was calculated based on the recN sequences. The oral cavity isolates, the koala bite wound isolates, and L. koalarum ACM3666T resulted in four clusters (Clusters 1-4). Clusters 1-3 were clearly not members of the genus Lonepinella. Cluster 1 was closely related to the genus Fredericksenia, and Clusters 2 and 3 appeared to be novel genera. Cluster 4 consisted of three subclusters: Cluster 4a with one koala bite wound isolate and L. koalarum ACM3666T, Cluster 4b with three oral cavity isolates and two Lonepinella-like wound isolates, and Cluster 4c with three nearly identical oral cavity isolates that may represent a different species within the genus Lonepinella. The rich Pasteurellaceae population, including potential novel taxa in the oral cavity of koalas supports an important role of these highly adapted microorganisms in the physiology of koalas. Moreover, the pathogenic potential of Lonepinella-like species is an important consideration when investigating infected koala bites in humans.

Transferrin Binding Protein B and Transferrin Binding Protein A2 Expand the Transferrin Recognition Range of Histophilus somni.

The bacterial bipartite transferrin receptor is an iron acquisition system that several important human and animal pathogens require for survival. It consists of the TonB-dependent transporter transferrin binding protein A (TbpA) and the surface lipoprotein transferrin binding protein B (TbpB). Curiously, the Tbps are only found in host-specific pathogens and are themselves host specific, meaning that they will bind to the transferrin of their host species but not to the transferrins of other animal species. While this phenomenon has long been established, neither the steps in the evolutionary process that led to this exquisite adaptation for the host nor the steps that could alter it are known. We sought to gain insight into these processes by studying Tbp specificity in Histophilus somni, an economically important pathogen of cattle. A past study showed that whole cells of H. somni specifically bind bovine transferrin but not transferrin from sheep and goats, two bovids whose transferrins share 93% amino acid sequence identity with bovine transferrin. To our surprise, we found that H. somni can use sheep and goat transferrins as iron sources for growth and that HsTbpB, but not HsTbpA, has detectable affinity for sheep and goat transferrins. Furthermore, a third transferrin binding protein found in H. somni, HsTbpA2, also showed affinity for sheep and goat transferrins. Our results suggest that H. somni TbpB and TbpA2 may contribute to broadening the host transferrin recognition range of H. somniIMPORTANCE Host-restricted pathogens infect a single host species or a narrow range of host species. Histophilus somni, a pathogen that incurs severe economic losses for the cattle industry, infects cattle, sheep, and goats but not other mammals. The transferrin binding proteins, TbpA and TbpB, are thought to be a key iron acquisition system in H. somni; however, despite their importance, H. somni TbpA and TbpB were previously shown to be cattle transferrin specific. In our study, we find that H. somni TbpB and another little-studied Tbp, TbpA2, bind sheep and goat transferrins, as well as bovine transferrin. Our results suggest that TbpB and TbpA2 may allow for host range expansion and provide a mechanism for how host specificity in Tbp-encoding pathogens can be altered.

Oral Lactobacillus strains reduce cytotoxicity and cytokine release from peripheral blood mononuclear cells exposed to Aggregatibacter actinomycetemcomitans subtypes in vitro.

BACKGROUND: This study evaluated the effect of oral lactobacilli on the cytotoxicity and cytokine release from peripheral blood mononuclear cells (PBMCs) when exposed to Aggregatibacter actinomycetemcomitans subtypes in vitro. The supernatants and cell wall extracts (CWEs) of eight A. actinomycetemcomitans strains, representing different subtypes, and three Lactobacillus strains were used. The PBMCs from six blood donors were exposed to supernatants and CWEs of A. actinomycetemcomitans or Lactobacillus strains alone or combinations and untreated cells as control. The cytotoxicity was determined by trypan blue exclusion method and IL-1ß secretion by ELISA. TNF-α, IL-6, and IL-8 secretions were measured using Bioplex Multiplex Immunoassay. RESULTS: Supernatants or CWEs from all bacterial strains showed cytotoxicity and IL-1ß secretion and the subtypes of A. actinomycetemcomitans showed generally a significantly higher effect on PBMCs than that of the Lactobacillus strains. Two highly toxic A. actinomycetemcomitans strains (JP2 and JP2-like) induced a higher response than all other strains. When combined, Lactobacillus significantly reduced the toxicity and the IL-1ß secretion induced by A. acinomycetemcomitans. The effect varied between the subtypes and the reduction was highest for the JP2 and JP2-like strains. The Lactobacillus paracasei strain SD1 had a higher reducing effect than the other Lactobacillus strains. This strain had a consistent reducing effect on all subtypes of A. actinomycetemcomitans cytotoxicity, and release of IL-1ß, IL-6, IL-8, and TNF-α from PBMCs of the blood donors. A strong and significant variation in cytokine release between the six blood donors was noticed. CONCLUSIONS: Lactobacillus spp. and L. paracasei SD1 in particular, showed a limited but statistically significant reducing interaction with A. actinomycetemcomitans toxicity and release of cytokines in vitro.

GtxA is a virulence factor that promotes a Th2-like response during Gallibacterium anatis infection in laying hens.

GtxA, a leukotoxic RTX-toxin, has been proposed a main virulence factor of Gallibacterium anatis. To evaluate the impact of GtxA during infection, we experimentally infected laying hens with a G. anatis wild-type (WT) strain and its isogenic gtxA deletion mutant (ΔgtxA), respectively, and monitored the birds during a 6 day period. Birds inoculated with ΔgtxA had significantly reduced gross lesions and microscopic changes compared to the birds inoculated with the WT strain. To assess the host response further, we quantified the expression of pro-inflammatory cytokines and apoptosis genes by RT-qPCR. In the ovarian tissue, the expression levels of IL-4 and TNF-α were significantly lower in the ΔgtxA group compared to the WT group, while IL-6 and IL-10 levels appeared similar in the two groups. In the spleen tissue of ΔgtxA infected chickens, IL-4 expression was also lower compared to the WT infected chickens. The results indicated that GtxA plays a key role in an acute cytokine-mediated Th2-like response against G. anatis infection in the ovary tissue. The pro-inflammatory response in the ovary tissue of birds inoculated with ΔgtxA mutant was thus significantly lower than the wild-type response. This was, at least partly, supported by the apoptosis gene expression levels, which were significantly higher in the ΔgtxA mutant compared to the wild-type infected chickens. In conclusion, GtxA clearly plays an important role in the pathogenesis of G. anatis infections in laying hens. Further investigations into the specific factors regulating the host response is however needed to provide a more complete understanding of the bacteria-host interaction.

Genotypic divergence of Avibacterium paragallinarum isolates with different growth requirements for nicotinamide adenine dinucleotide.

In 2017, for the first time in Asia, we reported the isolation of variants of Avibacterium paragallinarum with atypical NAD dependency. The present study was conducted to characterize the genotypes of 24 isolates of Av. paragallinarum in Korea, including the four variants reported previously. Most of the typical isolates (19/20) showed a unique ERIC-PCR pattern with no ERIC-PCR patterns in common between the typical isolates and the variants. Furthermore, the variants shared no ERIC-PCR patterns among themselves. All the typical NAD-dependent isolates belonged to the same phylogenetic group based on both 16S rRNA and hagA gene sequences. The four variants were placed in several groups distinct from the typical isolates. In the 16S rRNA phylogenetic analysis, two of the variants were not closely aligned to any other Av. paragallinarum, isolate although they were clearly members of the genus Avibacterium. The other variants were clustered together with NAD atypical isolates from geographically diverse global locations. Compared with the Modesto reference strain AY498870, all the variants lacked a TTTTT stretch at positions 182-186 in the 16S rRNA gene and the same deletion was shown in most of the reported variants. The typical isolates and variants shared 97.3-98.2% and 95.2-97.2% nucleotide sequence similarity, for 16S rRNA and hagA, respectively. In addition, the similarities among variants were within 98.3-100% and 96.5-98.4% for the two genes, respectively. Our results indicate that the Av. paragallinarum variants with altered NAD growth requirements were genetically different and highly divergent from the typical NAD-dependent isolates.RESEARCH HIGHLIGHTS NAD variant Korean Av. paragallinarum isolates show genetic diversity, whereas typical Korean Av. paragallinarum isolates do not.The Korean variants were not closely aligned to all other Av. paragallinarum in the 16S rRNA phylogeny.NAD atypical isolates from geographically diverse global locations clustered together.Almost all variants, including all Korean variants of Av. paragallinarum, lack a specific fragment of the 16S rRNA gene.

High phylogenetic diversity of Gallibacterium anatis is correlated with low biosecurity measures and management practices on poultry farms.

Gallibacterium anatis is considered one of the most common bacterial causative agents of reproductive tract disorders in poultry. In this study, phylogenetic analysis of partial rpoB sequences and biotyping using MALDI-TOF MS was done in order to investigate the genetic diversity of Gallibacterium isolates from 13 farms with different biosecurity measures and management practices. Sampling was done as a part of regular monitoring, except for Farms 9-13 that were included in the study to represent extensive production systems with lowest biosecurity levels. Pharyngeal and cloacal swabs were taken from live birds, while swabs from trachea, liver, peritoneum and oviduct were taken during necropsies. After cultivation and identification, strains from each farm were randomly selected for sequencing and biotyping. Both results showed high level of heterogeneity among the isolates originating from farms with low biosecurity levels, unlike isolates from farms with higher biosecurity levels and proper management that were more closely related and clustered together. Such correlation was statistically significant. Low biosecurity levels enable horizontal transmission of the pathogens, as well as gene transfer. The results confirm the importance of adequate biosecurity measures and management on poultry farms as they greatly affect the genetic diversity of the pathogens. Therefore, implementation of basic biosecurity measures could help control the heterogeneity of Gallibacterium strains, which would alleviate control of the infection prevalence on farms through immunoprophylaxis, and consequently improve poultry production. Also, the genetic diversity of G. anatis on poultry farms could be a good bioindicator of management practices and biosecurity measures used. RESEARCH HIGHLIGHTS High correlation between low biosecurity and high diversity of Gallibacterium anatis. Diversity of Gallibacterium is a good bioindicator of management practices on farms.

Prevalence and temporal trends in antimicrobial resistance of bovine respiratory disease pathogen isolates submitted to the Wisconsin Veterinary Diagnostic Laboratory: 2008-2017.

The objective of this study was to describe the prevalence and trends in antimicrobial resistance for bacterial pathogens associated with bovine respiratory disease (BRD) isolated from samples submitted to the Wisconsin Veterinary Diagnostic Laboratory (WVDL). Data were retrospectively collected from bovine respiratory isolates including Pasteurella multocida, Mannheimia haemolytica, Histophilus somni, and Bibersteinia trehalosi identified at the WVDL between January 2008 and December 2017. Antimicrobial susceptibility testing data were queried from antimicrobial resistance databases at the WVDL. A total of 4,261 isolates were identified. Pasteurella multocida was most frequently identified, accounting for 2,094 isolates (49% of total) over the study period. Mannheimia haemolytica was the second most frequently isolated bacterial respiratory pathogen (n = 1,267, 30%) followed by H. somni (n = 749, 18%) and B. trehalosi (n = 151, 4%). Over the 10-yr period, B. trehalosi had the highest median percentage of isolates that were resistant to at least one antibiotic at 33% (interquartile range: 24, 47) followed by M. haemolytica (13%; 8, 23). For P. multocida, 10% (4, 26) of isolates were classified as resistant to at least one antibiotic, whereas H. somni had the fewest resistant isolates (9%; 3, 15). When comparing 2013-2017 to 2008-2012, the overall percentage of resistant isolates for P. multocida and B. trehalosi decreased, whereas the percentage of resistant isolates for M. haemolytica and H. somni increased. Increased resistance against florfenicol, fluoroquinolones, gentamicin, tilmicosin, and tulathromycin was observed for M. haemolytica. These data show that antimicrobial susceptibility for BRD bacterial pathogens has changed in the population served by the WVDL over this 10-yr period. For P. multocida, resistance is relatively low and has either improved or at least remained constant for the majority of drugs labeled for treatment of respiratory disease in dairy cattle. Veterinarians and producers should be aware of the bacterial pathogens most commonly associated with BRD and work toward early disease detection, proper antibiotic administration, and monitoring lung lesions to ensure that their treatment protocols improve lung health.

Occurrence of Pasteurellaceae and Neisseriaceae bacteria in the pharyngeal and respiratory tract of dogs and cats - Short communication.

The occurrence of members of the Pasteurellaceae and Neisseriaceae families was studied in dogs and cats. A total of 110 nasal and pharyngeal swab samples from 47 dogs and 8 cats were collected. Most of the strains were identified by 16S rDNA sequencing, except Frederiksenia canicola and Pasteurella multocida where species-specific polymerase chain reactions were applied. The most frequently isolated species was F. canicola, which occurred only in dogs, mainly in the pharyngeal cavity. The second commonest bacterium, P. multocida was found in both types of samples and in both hosts. Other species from the family Pasteurellaceae, such as Haemophilus haemoglobinophilus, Pasteurella canis and P. dagmatis, were detected only in dogs. All isolated species belonging to the family Neisseriaceae, mainly representing Neisseria weaveri, were found only in the pharyngeal cavity. Neisseria weaveri and N. zoodegmatis could be detected in both hosts. Neisseria dumasiana and N. canis were isolated from dogs, while N. shayeganii only from a cat. For phylogenetic analysis, rpoB gene sequencing was performed, where the strains were on monophyletic branches and clearly separated from each other. In this study, recently described species such as F. canicola, N. shayeganii and N. dumasiana were detected that had never been isolated in Hungary before.

Interactions between the Trimeric Autotransporter Adhesin EmaA and Collagen Revealed by Three-Dimensional Electron Tomography.

Bacterial adhesion to host tissues is considered the first and critical step of microbial infection. The extracellular matrix protein adhesin A (EmaA) is a collagen-binding adhesin of the periodontal pathogen Aggregatibacter actinomycetemcomitans Three 202-kDa EmaA monomers form antenna-like structures on the bacterial surface with the functional domain located at the apical end. The structure of the 30-nm functional domain has been determined by three-dimensional (3D) electron tomography and subvolume averaging. The region exhibits a complex architecture composed of three subdomains (SI to SIII) and a linker between subdomains SII and SIII. However, the molecular interaction between the adhesin receptor complexes has yet to be revealed. This study provides the first detailed 3D structure of reconstituted EmaA/collagen complexes obtained using 3D electron tomography and image processing techniques. The observed interactions of EmaA with collagen were not to whole, intact fibrils, but rather to individual collagen triple helices dissociated from the fibrils. The majority of the contacts with the EmaA functional domain encompassed subdomains SII and SIII and in some cases the tip of the apical domain, involving SI. These data suggest a multipronged mechanism for the interaction of Gram-negative bacteria with collagen.IMPORTANCE Bacterial adhesion is a crucial step for bacterial colonization and infection. In recent years, the number of antibiotic-resistant strains has dramatically increased; therefore, there is a need to search for novel antimicrobial agents. Thus, great efforts are being devoted to develop a clear understanding of the bacterial adhesion mechanism for preventing infections. In host/pathogen interactions, once repulsive forces are overcome, adhesins recognize and tightly bind to specific receptors on the host cell or tissue components. Here, we present the first 3D structure of the interaction between the collagen-binding adhesin EmaA and collagen, which is critical for the development of endocarditis in humans.