Histophilus Ovis Research Articles

Taxonomy of Histophilus somni.

Histophilus somni was proposed in 2003 as a common name for bacteria that earlier had been called "Haemophilus somni", "Haemophilus agni", "Haemophilus somnifer", and "Histophilus ovis". The species is clearly separated from other species and genera within the family Pasteurellaceae. The species is phenotypically variable, but highly uniform regarding the 16S rDNA sequence. Whole-genome sequencing has revealed distinct genetic differences between a commensal and a pathogenic strain, particularly in regard to putative virulence factors. However, broad generalizations regarding the genetics of H. somni cannot be applied to the entire species until the genomes of additional strains are sequenced.

Histophilus somni

Haemophilus somnus has recently been renamed Histophilus somni and it has been proposed this name also includes Haemophilus agni and Histophilus ovis. The organism occurs in all management systems of the cattle industry, but the most serious losses occur in beef feedlots where co-mingling of cattle from numerous farms occurs. The disease in feedlots most often manifests as the septicemic form and is known as the Histophilus somni disease complex (HSDC), but pleuritis and bronchopneumonia are two other very common causes of disease or death. Despite effective vaccines and antibiotics, sudden losses from 30 to 90 days after arrival at feedlots continue to occur in significant numbers, especially in certain geographical areas ofNorth America and Europe. There undoubtedly remains much to be learned about predisposing factors to histophilosis and how this organism avoids natural body defense mechanisms.

Transferrin-dependent expression of TbpA by Histophilus ovis involves a poly G tract within tbpA

A poly G tract in tbpA of Histophilus ovis strain 3384Y was suspected of being responsible for the transferrin (Tf)-dependent expression of TbpA. The region encompassing the poly G tract was amplified using DNA from H. ovis strains 9L and 3384Y grown under iron-replete conditions and under iron-restricted conditions in the presence of bovine Tf. Sequence analysis of the amplification products revealed that regardless of the growth conditions, the poly G tract in strain 9L contained eight Gs, a situation that maintains the correct reading frame of the gene. Similarly, the poly G tract in strain 3384Y contained eight Gs when the organisms were grown under iron-restricted conditions in the presence of bovine Tf but when grown under iron-replete conditions, the poly G tract contained nine Gs resulting in a frame shift and the introduction of a premature stop codon. It is concluded that the Tf-dependent expression of TbpA in H. ovis strain 3384Y is due to a form of phase variation.

Production of transferrin receptors by Histophilus ovis: three of five strains require two signals.

Five strains of Histophilus ovis (9L, 642A, 714, 5688T, and 3384Y) were investigated with respect to iron acquisition. All strains used ovine, bovine, and goat transferrins (Tfs), but not porcine or human Tfs, as iron sources for growth. In solid phase binding assays, total membranes from only two (9L and 642A) of the five strains, grown under iron-restricted conditions, were able to bind Tfs (ovine, bovine, and goat, but not porcine or human). However, when the organisms were grown under iron-restricted conditions in the presence of bovine transferrin (Tf), total membranes from all strains exhibited Tf binding (as above); competition experiments demonstrated that all three Tfs (ovine, bovine, and goat) were bound by the same receptor(s). Membranes from organisms grown under iron-replete conditions in the presence or absence of bovine Tf failed to bind any of the test Tfs. An affinity-isolation procedure allowed the isolation of two putative Tf-binding polypeptides (78 and 66 kDa) from total membranes of strains 9L and 642A grown under iron-restricted conditions, and from membranes of all strains if the growth medium also contained Tf. It is concluded that all strains tested acquire Tf-bound iron by means of siderophore-independent mechanisms involving surface receptors analogous to the Tf-binding proteins (TbpA and TbpB) found in comparable organisms; although iron restriction alone is sufficient to promote the expression of these proteins by strains 9L and 642A, their production by strains 714, 5688T, and 3384Y appears to require two signals, iron restriction and the presence of Tf.

Production of transferrin receptors by <i>Histophilus ovis</i>: three of five strains require two signals

Production of transferrin receptors by <i>Histophilus ovis</i>: three of five strains require two signals

Specific PCR primers from the 16S-23S rRNA spacer region for the rapid detection and identification of Actinobacillus seminis.

Actinobacillus seminis is a common cause of ovine epididymitis and ram infertility. The ability to detect and identify this organism promptly is important commercially for the quality control of ram semen samples. Actinobacillus seminis is a fastidious and slow-growing bacterium and primary isolation and presumptive identification can be difficult and time-consuming. In this study, two ribosomal operons, termed rrnA and rrnB, have been characterized in the A. seminis genome, and these contain one and two tRNAs, respectively, in the spacer region between the 16S and 23S rRNA genes. Species-specific primers for A. seminis were developed from the sequence of the spacer region of rrnB for the identification and detection of A. seminis by PCR. The PCR assay was specific for A. seminis and gave no amplification products with phenotypically similar organisms such as Histophilus ovis. Storage solution used to preserve semen for long-term storage was found to inhibit the PCR. Therefore, for diagnostic purposes, the assay would best be performed after primary isolation or perhaps on fresh semen prior to storage if obvious contamination is indicated.

PCR methods and plasmid profile analysis for characterisation of Histophilus ovis strains.

The value of polymerase chain reaction (PCR)-based DNA fingerprinting and plasmid profile analysis for differentiation of Histophilus ovis isolates was assessed. Nineteen isolates of H. ovis were typed by PCR-ribotyping, repetitive extragenic palindromic element (REP)-PCR and enterobacterial repetitive intergenic consensus (ERIC)-PCR. These methods distinguished five types by PCR-ribotyping, 11 types by REP-PCR and seven types by ERIC-PCR. The ribotyping method produced a relatively simple pattern and a small number of distinct types and was useful for differentiation of H. ovis from the phenotypically similar organism, Haemophilus somnus. REP- and ERIC-PCR both produced complex banding patterns, but increased the discrimination between strains. Plasmids were found in 12 of the 19 isolates and there were four different plasmid profiles. A combination of the PCR methods and plasmid profile analysis provided a high resolution typing method for H. ovis.

Identification and differentiation of isolates of Haemophilus somnus, Histophilus ovis and Actinobacillus seminis by PCR methods

Identification and differentiation of isolates of Haemophilus somnus, Histophilus ovis and Actinobacillus seminis by PCR methods

Thrombotic meningoencephalitis associated with Histophilus ovis infection in lambs in Europe

Thrombotic meningoencephalitis is an acute, usually fatal disease of cattle caused by Haemophilus somnus. Although it has been widely recognised in cattle there have been only two previous reports of...

Antigenic cross-reactions between the causative agent of ovine footrot, Dichelobacter nodosus, and other bacteria

Lack of test specificity for the causative agent of ovine footrot, Dichelobacter nodosus, currently precludes routine application of serological tests. To investigate this problem antigenic cross-reactivity between D. nodosus and other bacteria was studied using double immunodiffusion and enzyme-linked immunosorbent assay (ELISA). Sera from sheep and rabbits immunised with D. nodosus reacted with other taxa but particularly Yersinia pseudotuberculosis, Y. enterocolitica, Cardiobacterium hominis, Suttonella indologenes, Corynebacterium pseudotuberculosis, Brucella ovis, Proteus vulgaris and Histophilus ovis. Sheep sera were then adsorbed with bacterial antigens and reacted in ELISA with cell envelope antigens of D. nodosus to determine whether non-specific reactivity could be reduced. C. pseudotuberculosis and the antigenically related organisms Nocardia asteroides and Mycobacterium phlei, as well as Bacteroides ruminicola and several other taxa reduced the signal in the ELISA. Antibodies reactive with many taxa increased with age of sheep. Sheep sera were more cross reactive than rabbit sera and distinguished relatively poorly between the antigens of different bacteria. Unfortunately the simple expedient of immunoadsorption was not effective in reducing non-specific reactions to the extent required for practical application of a serological test for ovine footrot.

A case of ovine abortion associated with Histophilus ovis infection.

A case of ovine abortion associated with Histophilus ovis infection.

Characterization of an immunoreactive 17.5-kilodalton outer membrane protein of Haemophilus somnus by using a monoclonal antibody.

A single outer membrane protein (OMP) of Haemophilus somnus, with an apparent molecular mass of 17.5 kDa, was identified in the sodium dodecyl sulfate (SDS)-insoluble fraction after extraction with 1% SDS-0.5 M NaCl-0.1% beta-mercaptoethanol. A hybridoma derived from mice immunized with H. somnus OMP fractions produced a monoclonal antibody (MAb), designated 20-3-5, that bound to the 17.5-kDa OMP of H. somnus. The MAb 20-3-5 epitope was present on 45 of 45 strains of H. somnus tested. MAb 20-3-5 cross-reacted with Haemophilus agni, Histophilus ovis, and Haemophilus haemoglobinophilus but not with 13 other species and subspecies of gram-negative bacteria. Immunoelectron-microscopic and antibody absorption studies revealed that the MAb 20-3-5 epitope is exposed on the surface of bacteria. In an immunoblot analysis, convalescent-phase sera obtained from calves with experimental H. somnus pneumonia contained antibodies to the 17.5-kDa OMP of H. somnus. Future studies will be directed toward examining the role of the 17.5-kDa OMP in immunity to H. somnus infections.

Genomic fingerprinting of "Haemophilus somnus" isolates by using a random-amplified polymorphic DNA assay

The random-amplified polymorphic DNA (RAPD) assay was used to generate DNA fingerprints for 16 isolates of "Haemophilus somnus," and one isolate each of "Haemophilus agni," "Histophilus ovis," "Actinobacillus seminis," Pasteurella haemolytica, and Escherichia coli. The RAPD assay differentiated among "H. somnus" isolates, which shared similarity coefficients of 0.46 to 1.00 on the basis of pairwise comparisons of RAPD markers produced with nine random decamer primers. Three virulent encephalitic "H. somnus" isolates exhibited identical banding patterns, suggesting a common clonal ancestry. The RAPD assay clearly distinguished between the "H. somnus"-"H. agni"-"H. ovis" group and the other bacterial species tested. The results of the present study suggest that DNA fingerprinting of "H. somnus" isolates by the RAPD assay could be valuable in revealing subspecific divisions within this largely unexplored species.

Meningoencephalitis and other conditions associated with Histophilus ovis infection in sheep

Histophilus ovis was isolated from 29 sheep in 20 flocks and 2 artificial insemination (AI) centres in southern New South Wales from 1984 to 1990. The clinical and pathological findings were consistent with previous reports and included polyarthritis (7 flocks), epididymo-orchitis (5), meningoencephalitis (3), pneumonia (3), septicaemia (2), mastitis (1) and metritis (1). Six sheep had meningoencephalitis, a syndrome not previously associated with H ovis infection in sheep, which was similar pathologically to thromboembolic meningoencephalitis in cattle, caused by the related organism, Haemophilus somnus. H ovis was isolated from the semen of 12-month-old rams in a flock that had polyarthritis due to H ovis, in 4-month-old ram lambs and from the uterus of a ewe in a flock that had sporadic cases of H ovis septicaemia.

Isolation of Actinobacillus seminis from rams in the United Kingdom

Actinobacillus seminis was isolated from the semen of five rams on four farms. Four of the rams had abnormal semen and three were also infertile. The isolates of A seminis showed similar phenotypic profiles and electrophoretic protein patterns to the type strain of A seminis but were distinct from Histophilus ovis previously isolated from rams with epididymitis in Scotland. The infection appeared to be subclinical but two of the five rams had palpable abnormalities of their testes. Three rams were treated with antibiotics but the infection persisted. No gross lesions were found in the genitalia of two of three rams examined post mortem but one had necrotic abscesses in the testes and epididymis. A seminis was isolated from the seminal vesicles and epididymis of one ram without gross lesions but not from the genitalia of the other two. On one farm the infection in a recently purchased ram led to the detection of another case as a result of the bacteriological screening of 11 stock rams not in contact with the initial case. These five subclinical cases, which included a supposedly healthy stock ram, suggest that A seminis infection may be widespread and should be considered in cases of infertility.

Inter- and Intrafamilial Similarities of rRNA Cistrons of the Pasteurellaceae

We performed hybridizations between labeled rRNAs from seven representative members of the family Pasteurellaceae and from three other taxa on the one hand and DNAs from 53 strains known or presumed to belong to the Pasteurellaceae on the other hand. The members of the Pasteurellaceae are most closely related to members of the Enterobacteriaceae, the Vibrionaceae, the Aeromonadaceae, and the genus Alteromonas. The family Pasteurellaceae is very heterogeneous. There are at least seven rRNA branches. Several organisms with the same genus name are dispersed over the entire dendrogram. The "Histophilus ovis," [Haemophilus] ducreyi, [Actinobacillus] actinomycetemcomitans, and [Haemophilus] aphrophilus rRNA branches are separate and quite remote from the three authentic genera in this family; this might justify eventual later separate generic status. DNA-rRNA hybridization with suitable, labeled rRNA probes is an excellent method to establish whether an organism belongs in the Pasteurellaceae; e.g., some strains of Bisgaard's taxa 7, 13, and 16 and of the gas-producing "SP" group certainly belong in this family, whereas three bovine lymphangitis organisms (strains NCTC 10547, NCTC 10549, and NCTC 10553), [Haemophilus] piscium ATCC 10801T (T = type strain), and [Pasteurella] piscicida ATCC 17911 belong in the Enterobacteriaceae, the Aeromonadaceae, and the Vibrionaceae, respectively.

Trials with an Enzyme‐Linked Immunosorbent Assay (ELISA) for the Diagnosis of Subclinical Genital Infections in Rams Caused by Histophilus ovis and Actinobacillus seminis

The performance of an enzyme-linked immunosorbent assay (ELISA) was evaluated in the serological diagnosis of subclinical genital infection in 6 naturally infected ram flocks and 2 experimentally infected ram hoggets. The test employs lipopolysaccharide (LPS) antigen prepared by autoclaving from Actinobacillus seminis and Histophilus ovis. A total of 193 sheep (118 unmated virgin rams and 75 mature breeding rams) were examined clinically, serologically (by ELISA) and bacteriologically (semen bacteriology) at the same time. Serum samples from all animals were also tested by an ELISA employing LPS antigen prepared from Brucella ovis in the same way. Shedding of A. seminis and H. ovis did not show close correlation with serological positivity (Table 1), as only 9 (15.0%) out of the 60 A. seminis shedders were ELISA seropositive at the same time. As regards H. ovis only 10 (19.2%) out of the 52 H. ovis shedders were ELISA seropositive at the same time. The results indicate that, when used alone, the ELISA employing LPS antigen is unsuitable for diagnosing subclinical genital infection caused by H. ovis and A. seminis in rams. The authors discussed shortly the employing fields of this ELISA test in the diagnostic work.

Evidence of Host-Specific Subgroups among "Histophilus ovis" Isolates

An investigation of 20 bovine and 24 ovine isolates of organisms variously described as “Histophilus ovis,” “Haemophilus agni,” and “Haemophilus somnus” revealed no consistent host-related differences in 47 biochemical characteristics; however, agarose gel electrophoresis of predominantly plasmid-derived deoxyribonucleic acid demonstrated the presence of variable numbers of plasmids of divergent size in each of the ovine strains but in none of the bovine isolates. Restriction enzyme analysis of deoxyribonucleic acids from an ovine culture and a bovine culture also suggested chromosomal differences. Our observations support the concept of one species, Histophilus ovis, comprising ovine and bovine isolates, but point to the existence of subspecific divisions related to host predilection.

Evaluation of three LPS-specific monoclonal antibodies for the immunodiagnosis of bovine brucellosis

Bovine sera collected during the Australian brucellosis eradication campaign were used to assess the value of three monoclonal antibodies (MAb Bruce 1, 4 and 7) for the immunodiagnosis of bovine brucellosis in a competitive enzyme immunoassay (CEIA). Each MAb reacted to a different epitope of lipopolysaccharide molecules on the cell surface of Brucella abortus. When the sensitivity of the CEIA was set at 100 per cent so that all infected animals were identified, the specificity of the test using MAb Bruce 1 and Bruce 7 was 69 per cent and 52 per cent, respectively. On the other hand, a quarter of the sera from infected cattle did not inhibit the binding of MAb Bruce 4 to the antigen. With a maximum sensitivity of 75 per cent, the specificity of the CEIA using MAb Bruce 4 was 94 per cent. However, all three MAb cross reacted with sera from sheep infected with Bovis, Histophilus ovis and Actinobacillus seminis.

Isolation and characterization of Actinobacillus seminis strains from ovine semen samples and epididymitis.

SummaryEighteen strains of Actinobacillus seminis isolated from ovine semen samples and epididymitis were characterized according to their cultural and biochemical behaviour. The semen samples were collected from young, unmated and mature breeding rams suffering from epididymitis or subclinical genital infection.All A. seminis strains were non‐motile, non spore‐forming, uncapsulated, Gram‐negative pleomorphic rods, approximately 1–8 μm × 0.8‐1 μm. On bovine blood agar they formed low convex, greyish‐white, non‐adherent and non‐haemolytic colonies, which grew under aerobic conditions, but more luxuriantly when cultures were incubated in an atmosphere containing 10% CO2 at 37°C.All strains were catalase, oxidase, nitrate and ornithine decarboxylase positive, but indole, urease, phosphatase and beta‐galactosidase (ONPG) negative. They were fermentative in the O/F test and usually produced acid within 1 to 3 days from glucose, arabinose, fructose, maltose and xylose, but not from lactose, mannose and sucrose.In the agar gel diffusion test, antigenic differences were demonstrated among the isolates. On the basis of the cultural, morphological and biochemical characteristics, A. seminis can be differentiated from other Gram‐negative bacteria (e.g. Brucella ovis, Histophilus ovis etc.) involved in genital infections in rams.