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.

Clinical-pathological findings induced by Histophilus somni isolated in subacute cardiac death in feedlot cattle.

The purpose of this report is to provide information about the different presentations of cardiac and extra-cardiac histophilosis and, to assess the antimicrobial (ATM) susceptibility of Histophilus somni isolated from these cardiac lesions to different ATM agents commonly used for treating bovine bacterial respiratory pathogens. Eight feedlot calves, which died after suffering from food rejection, apathy, hyperthermia, cough and nasal mucous discharge, and lack of response to ATM therapy, were studied. Cardiac lesions observed at necropsy included valvular/mural endocarditis, myocardial infarction, and necrotizing myocarditis, miliar non-suppurative myocarditis, myocardic necrotic sequestrum, and/or pericarditis. Histopathological, bacteriological and molecular studies confirmed the presence of a fastidious microorganism in the affected organs. H. somni showed no resistance to most ATM tested (ceftiofur, gamithromycin, enrofloxacin, florfenicol, tilmicosin). The results obtained in this study confirmed that H. somni was the main cause of the subacute cardiac lesions associated with hyperthermia, apathy and respiratory signs observed in cattle examined in this research. These presentations must be considered by veterinary practitioners in order to establish a rational therapeutic.

A Conserved Histophilus somni 23S Intervening Sequence Yields Functional, Fragmented 23S rRNA.

Histophilus somni is a Gram-negative bacterial organism that acts as an opportunistic pathogen and is a fastidious member of the Pasteurellaceae family associated with diseases of respiratory, reproductive, cardiac, and other tissues of ruminants. We identified an intervening sequence (IVS) embedded in all five copies of the 23S rRNA gene in the closed genome sequence of the H. somni isolate USDA-ARS-USMARC-63250 that may play an important role in affecting the biology of the organism. Sequencing the RNA from this isolate shows that most of the IVS is cleaved from the transcript, resulting in independent fragments of this structural rRNA that remain functional within the bacterial ribosome. The IVS lies between positions 1170 and 1278 bp of the 3,017-bp gene and exhibits self-complementarity between its 5' and 3' ends that predicts a stem-loop structure interrupting helix-45 in the transcribed 23S rRNA. Excision removes a 94-nucleotide (nt) stem-loop structure that displays an unusual 1-nt 3' end overhang instead of the more typical 2-nt overhang commonly observed at the ends of other excised IVS stem-loops. A comparison with genomes of other H. somni isolates indicates that this IVS is highly conserved, with 31 of 32 complete genomes having similar interruptions of canonical 23S rRNA genes. The potential biological effects of either the released IVS or the fragmentation of the functional 23S rRNA are unknown, but fragmentation may enhance rRNA degradation in ways that contribute to the regulation of gene expression. IMPORTANCE The genome biology underlying H. somni virulence, pathogenicity, environmental adaptability, and broad tissue tropism is understood poorly. We identified a novel H. somni 109-nt IVS stem-loop structure, of which the central portion is excised from the 23S rRNA transcript, resulting in the fragmentation of this rRNA in the H. somni isolate USDA-ARS-USMARC-63250 and the release of a 94-nt structured RNA of unknown function. We determined that this peculiar rRNA biology is widespread among sequenced H. somni isolates, suggesting it has importance to organism biology. The fragmented 23S rRNA molecules remain functional in the ribosome, given that the isolate grows in culture. The structured excised portion of the IVS, presumably due to the action of the endoribonuclease III, has an unusual 1-nt 3' end overhang. This newly discovered H. somni 23S rRNA fragmentation may enhance rRNA degradation providing a previously unrecognized avenue for regulating H. somni biological processes.

An improved multiplex PCR for Actinobacillus pleuropneumoniae, Glaesserella australis and Pasteurella multocida.

Glaesserella australis, a newly described bacterial species, has been isolated from pig lungs that displayed lesions very similar to those caused by Actinobacillus pleuropneumoniae, prompting the need for a validated diagnostic tool. In this work, we have altered a multiplex PCR used for the identification of cultures of G. australis, A. pleuropneumoniae and Pasteurella multocida to be more sensitive and then evaluated the use of the altered diagnostic tool on cultures and directly on tissues. The altered multiplex PCR was validated using 47 related species, both type/reference strains and field isolates. The sensitivity was assessed by serial dilutions and used a mixture of target bacteria in different concentrations. Further, 166 lung samples from 54 farms from four Australian States were used to validate the ability of the multiplex PCR to detect bacteria in lung swabs. The multiplex PCR was specific for the three target species. The assay could detect a minimum of 40 colony forming units (CFU) of G. australis, 786 CFU of A. pleuropneumoniae and 238 CFU of P. multocida. The multiplex PCR yielded more positives than coventional bacteriological examination. From a total of 166 lung samples, 51.9%, 51.9% and 5.6% of farms were PCR positive for P. multocida, A. pleuropneumoniae and G. australis, respectively. The results suggested that the new multiplex PCR was specific, sensitive and out performed traditional culture. The prevalence of G. australis was not very high, but it was the dominant pathogen in infected pigs.

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.

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.

Identification, HPG2 Sequence Analysis, and Antimicrobial Susceptibility of Avibacterium paragallinarum Isolates Obtained from Outbreaks of Infectious Coryza in Commercial Layers in Sonora State, Mexico.

This is the first extensive report on the identification and characterization of Avibacterium paragallinarum (AVP) isolates obtained from outbreaks of infectious coryza (IC) in IC-vaccinated layer flocks from Sonora State in Mexico. Isolates obtained from IC outbreaks during the years 2007, 2014, 2015, 2017, and 2019 were identified by conventional PCR test and 16S rRNA gene analysis, serotyped by Page serotyping and genotyped by the recently described partial sequence analysis of the HPG2 region. Furthermore, antimicrobial susceptibility profiles were determined by a recently improved minimal inhibitory concentration (MIC) test. The conventional PCR test and the 16S rRNA analyses confirmed the isolates as AVP. Serotyping results showed the involvement of isolates belonging to serotypes A, B, and C in the IC outbreaks. Genotyping of the HPG2 region revealed the presence of sequence type (ST)1, ST4, and ST11, of which the latter has also been identified in Europe. The MIC susceptibility test showed that all tested isolates were susceptible for the majority of tested antimicrobials, including erythromycin and tetracycline, which are important antibiotics for the treatment of IC. The IC situation in Sonora State, Mexico, is complex because of the presence of serotypes A, B, and C. This finding emphasizes the importance of biosecurity in combination with the application of the most optimal vaccination programs in the control of IC in Sonora State, Mexico.

Nota de investigación­Análisis de secuencias de la región HPG2 y susceptibilidad antimicrobiana de aislamientos de Avibacterium paragallinarum obtenidos de brotes de coriza infecciosa en aves de postura comerciales en el estado de Sonora, México. Este es el primer informe extenso sobre la identificación y caracterización de aislamientos de Avibacterium paragallinarum (AVP) obtenidos de brotes de coriza infecciosa (IC) de parvadas de ponedoras vacunadas con coriza infecciosa en el estado de Sonora en México. Los aislamientos obtenidos de los brotes de coriza infecciosa durante los años 2007, 2014, 2015, 2017 y 2019 se identificaron mediante una prueba de PCR convencional y el análisis del gene de ARNr 16S, se serotipificaron mediante el método de Page y se genotipificaron mediante el análisis parcial de secuencias descrito recientemente de la región HPG2. Además, se determinaron los perfiles de susceptibilidad a los antimicrobianos mediante la prueba de concentración mínima inhibitoria (MIC) que ha sido mejorada recientemente. La prueba de PCR convencional y los análisis de secuencias del gene ARNr 16S confirmaron que los aislados eran A. paragallinarum. Los resultados de la serotipificación mostraron la participación de aislamientos pertenecientes a los serotipos A, B y C en los brotes de coriza infecciosa. La genotipificación de la región HPG2 reveló la presencia de secuencias del tipo (ST) 1, ST4 y ST11, de los cuales este último también ha sido identificada en Europa. La prueba de susceptibilidad por concentración mínima inhibitoria mostró que todos los aislados analizados eran susceptibles a la mayoría de los antimicrobianos analizados, incluida la eritromicina y la tetraciclina, que son antibióticos importantes para el tratamiento contra la coriza infecciosa. La situación de coriza infecciosa en el estado de Sonora, México, es compleja por la presencia de los serotipos A, B y C. Este hallazgo enfatiza la importancia de la bioseguridad en combinación con la aplicación de los programas de vacunación óptimos en el control de la coriza infecciosa en el estado de Sonora, México.

Effect of Baicalin on Transcriptome Changes in Piglet Vascular Endothelial Cells Induced by a Combination of Glaesserella parasuis and Lipopolysaccharide.

Glaesserella parasuis causes porcine Glässer's disease and lipopolysaccharide (LPS) induces acute inflammation and pathological damage. Baicalin has antioxidant, antimicrobial, and anti-inflammatory functions. Long noncoding RNAs (lncRNAs) play key regulatory functions during bacterial infection. However, the role of lncRNAs in the vascular dysfunction induced by a combination of G. parasuis and LPS during systemic inflammation and the effect of baicalin on lncRNA expression induced in porcine aortic vascular endothelial cells (PAVECs) by a combination of G. parasuis and LPS have not been investigated. In this study, we investigated the changes in lncRNA and mRNA expression induced in PAVECs by G. parasuis, LPS, or a combination of G. parasuis and LPS, and the action of baicalin on lncRNA expression induced in PAVECs by the combination of G. parasuis and LPS. Our results showed 133 lncRNAs and 602 genes were differentially expressed when PAVECs were stimulated with the combination of G. parasuis and LPS, whereas 107 lncRNAs and 936 genes were differentially expressed when PAVECs were stimulated with the combination of G. parasuis and LPS after pretreatment with baicalin. The Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis showed the dominant signaling pathways triggered by the combination of G. parasuis and LPS were the tumor necrosis factor signaling pathway, phosphatidylinositol signaling system, and inositol phosphate metabolism. Protein-protein interaction network analysis showed the differentially expressed target genes of the differentially expressed lncRNAs (DELs) were related to each other. A coexpression analysis indicated the expression levels of the DELs were co-regulated with those of their differentially expressed target genes. This is the first study to systematically compare the changes in lncRNAs and mRNAs in PAVECs stimulated with a combination of G. parasuis and LPS. Our data clarified the mechanisms underlying the vascular inflammation and damage triggered by G. parasuis and LPS, and it may provide novel targets for the treatment of LPS-induced systemic inflammation.

Severe pneumonia in a street rat (Rattus norvegicus) caused by Rodentibacter rarus strain RMC2.

Background: Rodents are one of the most dangerous reservoirs and carriers of infectious diseases. Gradually, rats have become predominant in cities, sometimes staying in close vicinity to humans, pets, and other animals. Consequently, they tend to increase the transmission risk of pathogens. Case Description: Here, we report an original case of bacterial pneumonia in a street rat (Rattus norvegicus). The rat was found dead on a street in the chief town of Marseille (France) after being run over by a car. The necropsy of the corpse revealed generalized granulomatous pneumonia in almost all the pulmonary lobes. Lung lesions and predominantly multiple fibro-inflammatory areas are presumably the witness of an infectious etiology. Bacterial isolation was carried out from lung tissues. Colonies were identified by MALDI-TOF MS and confirmed by 16S rRNA sequencing. The following bacteria were identified: Staphylococcus cohnii, Bordetella bronchiseptica, Bordetella parapertussi, Corynebacterium glucuronolyticum, Pelistega suis and Rodentibacter rarus. Based on the histopathological diagnosis and the avoidance approach, the most likely etiological agent of pneumonia is therefore R. rarus, a little-known Pasteurellales bacterium that is closely related to Rodentibacter pneumotropicus. Conclusion: These data emphasize the severity of R. rarus infection in rodents. Thus, pointing out a potential risk for other animals (dogs, cats, and birds), as well as humans. The health monitoring program for rodents and rabbits pasteurellosis should now include R. rarus. Therefore, the pathological effect of the Rodentibacterspecies and/or strains needs to be better explored.

Mortality in Common (Sterna hirundo) and Sandwich (Thalasseus sandvicensis) Terns Associated with Bisgaard Taxon 40 Infection on Marco Island, Florida, USA.

Widely distributed aquatic species such as terns are highly dependent on, and can serve as indicators of, the global health of marine and other aquatic environments. Documented mass mortality events in terns have been associated with anthropogenic, weather-related and, less commonly, infectious causes. This study describes a multispecies mortality event associated with brevetoxicosis and Bisgaard taxon 40-induced sepsis involving common (Sterna hirundo) and sandwich (Thalasseus sandvicensis) terns off the southwest coast of Florida, USA, in November and December 2018. During an approximately 6-8-week period, a large number of birds were found dead or displayed weakness, ataxia or other neurological signs. Many were admitted to a wildlife hospital for evaluation, but most died or were euthanized due to poor prognosis. Necropsy of 12 birds revealed minimal or non-specific gross lesions. Initial toxicology screening of tissues for brevetoxins revealed levels that could be consistent with brevetoxicosis. However, histology revealed multiorgan inflammation and necrosis associated with a gram-negative bacillus. A bacterium isolated on aerobic culture of liver and heart tissues was unidentifiable in the MALDI-TOF database. Subsequently, 16 S rRNA gene sequencing revealed that the isolate shared 99.33% homology with Bisgaard taxon 40 from the Pasteurellaceae family. While the source of the bacterium and potential association with brevetoxin exposure are unclear, histopathology suggests that the bacterium was the proximate cause of clinical signs and mortality in all birds examined as well as the scale of the mortality event. This report highlights the need to conduct detailed investigations into wildlife mortality events and expands on the current, limited knowledge of the effects of novel Pasteurellaceae bacteria on avian health.

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.

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.

Sialidase of Glaesserella parasuis Augments Inflammatory Response via Desialylation and Abrogation of Negative Regulation of Siglec-5.

Siglecs are sialic acid-binding immunoglobulin-like lectins that play an important role in tissue homeostasis, immune response, and pathogen infection. Bacterial sialidases act on natural ligands of Siglecs, interfering with the Siglec-mediated immune response. Glaesserella parasuis is a porcine bacterial pathogen that secretes sialidase. However, little is known about the sialidase of G. parasuis and its impact on immune regulation. Here, we used wild-type G. parasuis, a sialidase-deficient mutant, and complementary strains to investigate the role of sialidase in porcine alveolar macrophage infection. Sialidase induced the release of proinflammatory cytokines, such as interleukin-1α (IL-1α), IL-6, and tumor necrosis factor alpha, from porcine alveolar macrophages. Moreover, sialidase desialylated the surface of porcine alveolar macrophages and altered the expression of Siglecs (the expression of Siglec-5 was reduced). Furthermore, sialidase led to a reduction in endogenous SH2 domain-containing protein tyrosine phosphatase (SHP-2) recruitment to Siglec-5 and simultaneously activated the inflammatory response via the mitogen-activated protein kinase and nuclear factor kappa light chain enhancer of activated B cell signaling pathways. This desialylation occurred before the release of proinflammatory cytokines, suggesting that the sialidase-induced inflammatory response was followed by reduced recruitment of SHP-2 to Siglec-5. Thus, this study is the first to demonstrate the role of sialidase in the inflammatory response of G. parasuis. This role resulted from the abrogation of negative regulation of Siglec-5 on proinflammatory cytokine release. This study helps to understand the molecular mechanism underlying the inflammatory response induced by sialidase secreted by G. parasuis and the acute inflammation caused by G. parasuis.

Differentiation among the most important Rodentibacter species by multiplex PCR assays targeting the ITSile+ala sequences of the rRNA operons.

Screening for the Rodentibacter species is part of the microbiologic quality assurance programs of laboratory rodents all over the world. Nevertheless, currently there are no PCR amplification techniques available for the diagnostic of R. ratti, R. heidelbergensis and of a Rodentibacter related ß-haemolytic taxon. The aim of this study was to utilize the differences in the sequence of the Internal Transcribed Spacer (ITS) regions of R. pneumotropicus, R. heylii, R. ratti, R. heidelbergensis and of the ß-haemolytic Rodentibacter taxon for the design of specific PCR assays for these species. The ITSile+ala sequence variations allowed the design of specific forward and reverse primers for each species included, that could be combined in different multiplex assays. The performance characteristics specificity and sensitivity registered for each primer pair against a diverse collection of Pasteurellaceae isolated from rats and mice and of further non-Pasteurellaceae strains was 100% for all five Rodentibacter species included. In addition, the PCR assays displayed high limits of detection and could be successfully used for detection of Rodentibacter spp. DNA in clinical swabs of laboratory mice and rats. Overall, the assays described here represent the first PCRs able to diagnose R. ratti, R. heidelbergensis and the ß-haemolytic Rodentibacter taxon, whose diagnostic to species level could further facilitate better understanding of their geographic distribution, prevalence, and biology in the future.

Acute fibrinous pleuropneumonia and septicaemia caused by Bibersteinia trehalosi in neonatal calves in New Zealand.

Case history: In July and August 2019, 15/40, ≤48-hour-old calves became acutely ill. The calves were all born on-farm, transferred to pens soon after birth, and fed with "gold" colostrum. The hygiene, biosecurity and ventilation in the pens were poor. Of the 15 calves, 11 died or were euthanised and four calves, ≤48-hour-old, that became acutely ill later in the outbreak were treated with cefquinome, a fourth-generation cephalosporin, and recovered. Clinical findings: The affected calves presented with acute recumbency, lethargy, tachypnoea, tachycardia, increased lung sounds, inability to stand or feed, and dehydration without pyrexia. Pathological findings: Gross findings in a calf that died naturally included fibrinous pleuropneumonia, marked oedematous expansion of the interlobular septa, especially in the ventral lung lobes, fibrinous polyserositis and fibrinous polyarthritis. A second calf that was euthanised had strikingly similar lung lesions. Histologically, the pulmonary interlobular septa of both calves were prominently expanded by oedema, dilated lymphatics and the infiltration of numerous neutrophils and macrophages interspersed with small Gram-negative rod bacteria. Likewise, the visceral pleura showed fibrinopurulent inflammation with numerous small Gram-negative rods. Microbiological findings: Microbial culture and matrix-assisted laser desorption ionisation time-of-flight (MALDI-TOF) mass spectrometry identified Bibersteinia trehalosi in the lung, stifle joint and peritoneal cavity of the first calf and lung of the second. Diagnosis: B. trehalosi acute fibrinous pleuropneumonia and septicaemia. Clinical relevance: This is the first report of the clinical findings and histological lesions of B. trehalosi pleuropneumonia and septicaemia in calves in New Zealand. The pathogen is isolated with increasing frequency from cases of bovine respiratory disease in dairy cows, feedlot cattle and calves in the United Kingdom and North America. The importance of microbial culture in cases such as this with unusual lung lesions in calves <48 hours of age, cannot be over emphasised. Cefquinome was administered to all remaining heifer calves and four calves that became ill later in the outbreak recovered after cefquinome treatment.

First report of pulmonary disease associated with Nicoletella semolina in a horse in New Zealand.

Case history: A 9-year-old warmblood gelding with a history of chronic intermittent tachypnoea and dyspnoea was presented for evaluation and removal of a mass on the left side of the neck. A fibrous mass adherent to the left jugular vein developed and was removed surgically 6 weeks later, at which time the owner requested an evaluation of the cause of the persistent respiratory signs first noted on primary admission. Clinical findings and treatment: Clinical findings included coarse lung sounds on thoracic auscultation, tracheal wheeze, and an abnormal trans-tracheal aspirate. These findings, in addition to the results of ultrasonographic imaging of the thorax and transtracheal cytology, were suggestive of bacterial bronchopneumonia. Initial antimicrobial therapy included I/M 22 mg/kg procaine penicillin every 12 hours and I/V 6.6 mg/kg gentamicin sulphate every 24 hours. The horse's clinical signs improved within 36 hours. It was discharged after 6 days, and at the owner's request antimicrobial therapy was changed to 25 mg/kg trimethoprim/sulphadimidine to be given orally every 12 hours for 10 days. One month later, the horse had recovered and there were no further complications reported by the owner except for an occasional cough while grazing Laboratory findings: Bacterial culture of transtracheal wash fluid resulted in the isolation of Nicoletella semolina as the sole organism, later confirmed by genotyping. Attempts to subculture the organism for antimicrobial susceptibility testing were unsuccessful. Diagnosis: Infectious bronchopneumonia associated with Nicoletella semolina Clinical relevance: Further work is required to determine whether N. semolina is acting as an opportunistic commensal of the equine respiratory tract or a primary pathogen. However, this article reports the first instance in New Zealand of an association between the presence of this organism and respiratory disease in a horse.

Identification of a large repetitive RTX immunogen in a highly virulent Rodentibacter heylii strain.

Rodentibacter (R.) heylii is frequently detected in laboratory rodents. Repeats in toxin (RTX) toxins are considered important virulence factors of this major murine pathogen. We evaluated the virulence of a R.heylii strain negative for all known RTX toxin genes and Muribacter (M.) muris, a commensal in mice, in experimental infections of C57BL/6 and BALB/c mice. Experimental intranasal infection with 108 CFU of the pnxI-, pnxII- and pnxIII- R. heylii strain resulted in 75% and 100% mortality in C57BL/6 and BALB/c mice, respectively. In early losses, multiple internal organs were infected and purulent bronchopneumonia was the main pathology. Intranasal application of M. muris did not result in mortality or severe weight loss. Immunoproteomics led to the identification of a surface-associated and specific immunogen, which was designated as R. heylii immunogen A (RhiA) and which was exclusively recognised by sera obtained from mice infected with this R. heylii pathotype. RhiA is a 262.6 kDa large protein containing long imperfect tandem repeats and C-terminal RTX consensus sequences. Immunohistochemical analysis confirmed that this R.heylii pathotype expresses RhiA in the lower respiratory tract. In summary, this study describes a specific immunogen in a virulent R. heylii, strain which is an excellent antigen for pathotype-specific serological screenings and which might carry out RTX-related functions.

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.

Prevalence and antimicrobial resistance of opportunistic pathogens associated with bovine respiratory disease isolated from nasopharyngeal swabs of veal calves in Switzerland.

The composition of the bacterial flora in the calf nasopharynx might influence the risk of bovine respiratory disease (BRD). The aims of the present study were, firstly, to investigate the prevalence of bacteria potentially involved in BRD in the nasopharynx of veal calves and to identify associated risk factors for their presence, and, secondly, to provide data on antimicrobial resistance levels in these bacteria. Deep nasopharyngeal swabs were collected from veal calves on 12 Swiss farms over a period of one year by non-random, but systematic sampling for isolation of Pasteurellaceae and Mycoplasma (M.) bovis and dispar. Associations of potential risk factors with occurrence of these bacteria were tested in multivariable mixed logistic regression analyses, based on information gained from extensive questionnaires completed with the farmers. Antimicrobial susceptibility testing was performed for Pasteurellaceae by broth microdilution method to obtain minimal inhibitory concentrations (MIC). Pasteurellaceae, including Pasteurella (P.) multocida, Mannheimia (M.) haemolytica, Bisgaard Taxon 39 and Histophilus (H.) somni, were almost twice as prevalent as M. bovis and dispar in this study. Continuous stocking was a risk factor for the presence of Pasteurellaceae, especially when calves originated from more than six suppliers. In young calves (≤ 91 days), feeding of California Mastitis Test (CMT) positive milk was an additional risk factor for the presence of Pasteurellaceae whereas transport of calves by farmers and livestock traders (as opposed to transport only by farmers) increased the risk in older calves (> 91 days). Risk factors for the presence of M. bovis/dispar were higher number of calves per drinking nipple in young calves, and no access to an outside pen and feeding of CMT positive milk in older calves, respectively. While further research will have to investigate the observed associations in more detail, this suggests that management can play an important role in the prevalence of nasopharyngeal bacteria with a potential subsequent involvement in BRD. Antimicrobial resistance differed between the three bacterial species tested in this study and was highest to oxytetracycline and spectinomycin in P. multocida, oxytetracycline and penicillin in M. haemolytica, and ampicillin and penicillin in H. somni. Only two European VetCAST breakpoints (for florfenicol in P. multocida and M. haemolytica) have been published to date, matching the MIC distribution of the present isolate populations well, in contrast to certain commonly applied American Clinical and Laboratory Institute interpretive criteria. This highlights the potential for further refinement of clinical breakpoints in veterinary medicine.