Ovine Haemophilus somnus: experimental intracisternal infection and antigenic comparison with bovine Haemophilus somnus.

Experimental infection was produced by two of four isolates of ovine Haemophilus somnus given by intracisternal inoculation into two to three-month-old lambs. Isolate 2041 (originally obtained from a septicemic lamb in Alberta) caused lethal infection in eight of nine lambs, isolate 67p from the prepuce of a normal lamb produced less acute disease in four of nine lambs, and the other two isolates (93p and 1190) caused no detectable disease. Significant lesions were limited to the brain and spinal cord. Purulent meningitis was characteristic but vasculitis or septicemia were not detected, perhaps due to the route of inoculation. Since a difference in virulence was noted among strains, we analyzed surface proteins thought to be virulence factors of bovine H. somnus. Protein profiles of bovine and ovine H. somnus done by sodium dodecyl sulphate-polyacrylamide gel electrophoresis showed similar patterns for virulent bovine isolates and ovine septicemic isolates. Preputial isolates showed a lower molecular mass major outer membrane protein than septicemic isolates. Antigenic analysis revealed that outer membrane proteins p270, p78, p76, p40, and p39 were detected in both ovine and bovine isolates except for 1190, which was probably not a true H. somnus isolate. Thus the preputial and septicemic isolates of ovine H. somnus were similar to bovine H. somnus in pathogenicity and in surface antigens.     

Hemophilus somnus in the reproductive tracts of slaughtered cows: location and frequency of isolations and lesions

One hundred bovine female reproductive tracts were examined for the presence of Hemophilus somnus, and fifty were examined for gross and microscopic lesions. H. somnus was isolated from eight of 100 tracts. The isolation was made from the vagina in seven cows. Of 24 tracts in which the major vestibular gland was sampled, three were positive at this site, and two of these three also had the organism isolated from the vagina. Further investigations into the role of the major vestibular gland as a source for continuing infections of the reproductive tract of cattle were indicated. In this investigation, H. somnus was as likely to be found in a region without lesions as one with lesions.     

Haemophilus somnus (Histophilus somni) in bighorn sheep.

Respiratory disease and poor lamb recruitment have been identified as limiting factors for bighorn-sheep populations. Haemophilus somnus (recently reclassified as Histophilus somni) is associated with respiratory disease in American bison, domestic sheep, and cattle. It is also harbored in their reproductive tracts and has been associated with reproductive failure in domestic sheep and cattle. Therefore, reproductive tract and lung samples from bighorn sheep were evaluated for the presence of this organism. Organisms identified as H. somnus were isolated from 6 of 62 vaginal but none of 12 preputial swab samples. Antigen specific to H. somnus was detected by immunohistochemical study in 4 of 12 formalin-fixed lung tissue samples of bighorn sheep that died with evidence of pneumonia. Notably, H. somnus was found in alveolar debris in areas of inflammation. The 6 vaginal isolates and 2 H. somnus isolates previously cultured from pneumonic lungs of bighorn sheep were compared with 3 representative isolates from domestic sheep and 2 from cattle. The profiles of major outer membrane proteins and antigens for all of the isolates were predominantly similar, although differences that may be associated with the host-parasite relationship and virulence were detected. The DNA restriction fragment length profiles of the bighorn-sheep isolates had similarities not shared with the other isolates, suggesting distinct phylogenetic lines. All of the isolates had similar antimicrobial profiles, but the isolates from the bighorn sheep produced less pigment than those from the domestic livestock, and growth of the former was not enhanced by CO2. Wildlife biologists and diagnosticians should be aware of the potential of these organisms to cause disease in bighorn sheep and of growth characteristics that may hinder laboratory detection.     

Reproductive difficulties in cattle with antibody titers to Haemophilus somnus

A herd of Holstein cows was examined because of suspected embryonic death. Four cows had embryonic loss, and 2 cows had aborted. Paired serum samples were tested for antibodies to infectious bovine rhinotracheitis virus, bovine viral diarrhea virus, and Haemophilus somnus. Of 16 cows, 8 had antibody titers to H somnus greater than 1:1,024, and 3 had greater than or equal to four-fold changes in antibody titers to H somnus. Haemophilus somnus infection was active in this herd and may have been responsible for the herd's reproductive problems.     

Haemophilus somnus and reproductive disease in the cow: A review

Haemophilus somnus causes inflammatory disease in the genital tract of cows as reported in several field surveys and experimental trials. This organism can also innocuously colonize the healthy genital mucosa of the cow, which indicates its dual relationship with the host, that of pathogen and commensal. Experimental data indicate embryocidal capability of this pathogen suggesting a possible role in early embryonic death. Haemophilus somnus also causes sporadic abortions after a bacteremia in the dam. Retrograde infection of the pregnant uterus from the lower genital tract appears unlikely; however, this process can account for post-parturient endometritis. Detection of high homologous IgG₂ serum antibody titers using an ELISA test may be useful for the diagnosis of current or recent genital inflammation. Experimental laboratory data indicate that a proportion of genital strains of H. somnus are pathogenic and capable of causing thrombotic meningoencephalitis and perhaps pneumonia. In vivo testing of the pathogenicity of genital strains remains to be conducted.     

Atypical nervous lesion in Haemophilus somnus infection of cattle

Two cows affected with Haemophilus somnus were examined pathologically. Although manifested lesions were composed of fibrinopurulent meningitis without the usual parenchymatous lesions in the central nervous system, Haemophilus somnus was isolated in pure culture from the cerebral cortex of both cases. Presence of this atypical lesion in the nervous system should be kept in mind for the differential diagnosis of the disease.     

Female genital tract abnormalities of Zebu cattle slaughtered at Bahir-Dar Town, north-west Ethiopia

A study was conducted to assess the type and prevalence of abnormalities occurring in the female reproductive tracts of 201 Zebu cattle of Fogera type (161 cows and 40 heifers) slaughtered at Bahir-Dar town, north-west Ethiopia. Out of the 201 female genital tracts collected and examined, abnormalities were recorded in 74 (36.8%). The most common abnormalities encountered were ovariobursal adhesion (5.5%), endometritis (3.9%) and cystic ovaries (3.5%). Other abnormalities recorded were ovarian hypoplasia, vaginitis, cervicitis, tortuous cervical canal, mucometra, vaginal cyst, parovarian cyst, hypoplastic cervical rings, cervical cyst, freemartins, closed external cervical os, uterine and oviducts adhesion, cystic uterine tube, remnant of retained fetal membrane and cyst in the uterine wall. The prevalence of the abnormalities was significantly (p < 0.05) higher in parous than in nulliparous cows. Moreover, evidence of ovarian cyclicity was found in 51.6% and 30% of non-pregnant parous and nulliparous cows examined, respectively. This study revealed that reproductive tract abnormalities seem to be an important problem with possible subsequent infertility in Fogera-type Zebu cows in the study area.     

Chlamydia abortus in Cows Oviducts,Occasional Event or Causal Connection?

Fifty‐seven genital tracts of regularly slaughtered culled Piedmontese cows, aged 7.4 ± 4.3 years (mean ± SD), range: 2.6–15.6 years, were grossly and microscopically examined. DNA extracted from oviducts was subjected to PCR to evaluate the presence of Chlamydia spp. The 15 PCR‐positive oviducts were subjected to Sanger sequencing and showed the presence of Chamydia abortus, with an identity range between 99 and 100%. Nine of the PCR‐positive samples belonged to the 24 animals with a normal macroscopic appearance of the whole genital tract (percentage of positive oviducts in normal genital tracts 9/24 = 37.5%), while six belonged to the 33 genital tracts with lesions in one or more organs (percentage of positive oviducts in pathological genital tracts 6/33 = 18.1%); of these, a single animal had salpingitis. The detection of C. abortus in bovine oviducts is of particular interest because it has never been previously investigated or reported.     

Development of a defined medium for Haemophilus somnus isolated from cattle

Nutritional factors that influence the growth of Haemophilus somnus were examined, and a defined medium was developed. Optimal growth of H somnus in broth occurred under conditions of maximum aeration. Nutritional components required for or enhanced growth of most H somnus isolates in the defined medium included uracil, D-glucose, isotonic NaCl, Na2HPO4, nicotinamide, flavin mononucleotide, pantothenic acid, pyridoxine, and a variety of salts and amino acids. The defined medium supported optimum growth of 18 of 21 isolates of H somnus from cattle.     

Characterisation of Histophilus ovis and related organisms by restriction endonuclease analysis

The banding profiles generated by Bam H1 restriction endonuclease cleavage of bacterial DNA from clinical and reference isolates of Histophilus ovis, Haemophilus somnus and related bacteria were compared. H. ovis, H. somnus and Haemophilus agni isolates were found to have distinct similarities in banding profiles characterised by 10 common bands between 2.0 and 9.6 kilobases (kb). The close taxonomic relationship of these isolates was reinforced by these findings. The reference isolates examined in this study--Actinobacillus lignieresii, Actinobacillus seminis, H. agni, H. somnus, H. ovis, Haemophilus influenzae, Haemophilus parainfluenzae, Haemophilus parahaemolyticus--could be distinguished from each other on the basis of their characteristic banding profiles. Actinobacillus sp were observed to have more bands between 2 and 23 kb compared with the H. ovis and Haemophilus sp isolates studied. Analysis of isolates from an experimental infection trial illustrated the potential of restriction endonuclease analysis in molecular epidemiological applications. It was possible to demonstrate by this means that the post-challenge isolates had identical banding profiles to the challenge (or infecting) isolate which had a distinctly different banding profile from that of pre-challenge H. ovis isolates. Furthermore, restriction endonuclease analysis of H. ovis isolates obtained from follow-up investigations of a recurrent problem of epididymitis in unmated rams, indicated that the H. ovis isolates implicated in epididymitis, were present as a single strain in a number of sheep over a period of time. This suggested that the mechanism of transmission was by perinatal perputial contamination.     

An evaluation of the API ZYM system as a means of identifying Haemophilus somnus and related taxa.

The commercially available API ZYM microbiological identification system was evaluated for the rapid identification of Haemophilus somnus. Eighty-seven isolates of the organism had API ZYM profiles which were characteristic. The API ZYM profiles demonstrate clear differences between H. somnus and other genera but suggest a close association to three related organisms. Enzyme activity of H. somnus isolates were similar to organisms identified as Histophilus ovis, Haemophilus agni and strains UQV of Actinobacillus actinoides and Actinobacillus seminis but was clearly different from isolates of Pasteurella haemolytica, Pasteurella multocida, Bordetella bronchiseptica and group EF4. The API ZYM system allowed more rapid identification of H. somnus than conventional biochemical tests and may be a useful adjunct to conventional methods used for identification of H. somnus isolates. The test did not reveal obvious differences between isolates from various anatomic locations.     

Differentiation of pathogenic and nonpathogenic Escherichia coli isolated from poultry

Twenty-one isolates of Escherichia coli recovered from chickens and turkeys were evaluated for pathogenicity in 1-week-old chicks. Fifteen produced coli-septicemia (pathogenic) and six were innocuous (nonpathogenic). Both pathogenic and nonpathogenic E. coli were tested for their ability to selectively absorb Congo red (CR) dye incorporated into agar medium. Eight of 15 pathogenic E. coli (somatic antigen types O1, O78, O11, O88, and OX9) absorbed the dye and produced red colonies (CR+) between 48 to 72 hours of incubation. All serotypes of E. coli with homologous somatic antigen O78 were CR+, while those of O2 antigen were CR- (white colonies). Five of six nonpathogenic E. coli also were CR+. In contrast to pathogenic E. coli, however, nonpathogenic isolates absorbed CR early, between 18 to 24 hours of incubation. Although CR dye binding did not correlate well with pathogenicity, it may be an identifiable property of some serotypes of E. coli.     

Gross abnormalities, bacteriology and histological lesions of uteri of dairy cows failing to conceive or maintain pregnancy

AIM: To describe the gross pathology, bacteriology and histopathology of the reproductive tracts of dairy cows that failed to conceive or maintain pregnancy. METHODS: The reproductive tracts of 105 cows that were not pregnant at the end of the seasonal breeding programme were retrieved following slaughter. The tracts were examined grossly, both horns of the uterine lumen were swabbed for bacteriology, and tissue was collected from each horn and the body of the uterus for histopathology. Grossly enlarged uteri and tracts containing purulent vaginal content were excluded. Histopathology was performed on three sections of uterine tissue from each of three cows in which no gross pathological changes were detected and from which no bacteria were isolated, from three cows in which no gross pathology was detected but from which bacteria were isolated, and from three cows in which gross pathological changes were detected but from which no bacteria were isolated. RESULTS: Thirty-six (34%) cows had one or more gross lesions which involved the ovary, uterine tube, uterus or vagina. Bacteria were isolated from the uteri of 22 (21%) cows. Isolates included Arcanobacterium pyogenes (n = 1), Escherichia coli (n = 1), Fusobacterium spp (n = 1), Haemophilus somnus (n = 5), Streptococcus acidominimus (n = 12), S. bovis (n = 2), S. uberis (n = 1) and S. salivarious (n = 1). In only five cows were both gross pathology and bacteria detected. There was no relationship between the isolation of bacteria and the diagnosis of gross pathology of the uterus. There were no differences in the degree of histopathological changes in the uteri from the three groups of cows examined, and lesions present were minor. CONCLUSIONS: Gross pathological changes and intra-uterine bacteria were found in 34% and 20% of cows, respectively, but the correlation between the two was poor. Histopathological changes were unremarkable, suggesting the bacteriological findings were coincidental, that causative agents of infertility were not present at the time of examination, or that unrelated causes such as nutritional anoestrus may have been responsible for the failure of some cows to conceive.     

Comparative pathogenicity of selected bovine viral diarrhea virus isolates in gnotobiotic lambs

The infectivity and pathogenicity of selected bovine viral diarrhea virus (BVDV) isolates were determined in gnotobiotic, colostrum-deprived neonatal lambs. Five-day-old cesarean-derived gnotobiotic lambs were exposed to 1 of 10 BVDV isolates via aerosol suspension. These isolates were from tissues or secretions of calves or lambs affected with respiratory tract disease, weak neonatal calves, aborted bovine fetuses, or reference Singer or Draper BVDV. The pathogenicity of each isolate, relative to the others, was evaluated in lambs by measurement of the neutralizing antibody response, virus isolation from nasal secretions or tissues, and postmortem lesions. The BVDV isolates varied in their infectivity and pathogenicity. Singer, the cytopathic reference strain, was the most lymphotrophic isolate and stimulated the greatest neutralizing antibody response. Encephalitis was the most consistent lesion observed and was used as the final determinant of relative pathogenicity of the viruses. The most neuropathogenic isolates were the 2 viruses originating from lambs affected with respiratory tract disease, the 2 weak neonatal calf isolates, and 1 isolate from an aborted bovine fetus. The least pathogenic isolates were the 2 reference isolates, Draper and Singer; the 2 mucosal disease isolates; and 1 isolate originating from an aborted bovine fetus.     

In ovo pathogenicity of Mycoplasma gallisepticum strains in the presence and absence of maternal antibody

Mycoplasma gallisepticum (MG) strains showing marked variation in pathogenicity were examined for virulence in ovo. No correlation was found between in ovo pathogenicity and other in vivo or in vitro methods for pathogenicity evaluation. For certain highly pathogenic strains, there was a clear relationship between the titer of MG inoculated and the embryo mortality and time of death; an LD50 for these strains could be calculated by yolk-sac inoculation. However, not every strain that caused lesions in the respiratory tract in vivo caused embryo mortality. Less pathogenic strains that grow well and colonize the respiratory tract usually caused embryo mortality during the later stages of incubation, and there was no strict correlation between titer of inoculum and embryo mortality. It appeared that embryo death in these cases may have resulted from generalized stress due to mass multiplication of the MG. Embryo mortality due to virulent MG was completely blocked in eggs containing maternal antibody to MG, although the mycoplasma could be reisolated from the yolk-sac membrane of the live embryonated egg after 17 days of incubation. Attempts to mimic the effect of maternal antibody by injecting exogenous MG antiserum were not successful.     

A comparative study of bovine and ovine Haemophilus somnus isolates.

Bacterial isolates (including 17 Haemophilus somnus isolates and an H. somnus-like isolate) from asymptomatic or diseased cattle and sheep, were evaluated for markers associated with virulence and host predilection. The isolates were separated into 6 distinct biovariants, 3 for sheep and 3 for cattle, based on reactions in a battery of 21 test media. Three bovine isolates associated with disease caused hemolysis of bovine blood. The rest of the isolates did not hemolyze either bovine or ovine erythrocytes. Protein profiles of all H. somnus isolates were similar with the exception of the major outer membrane proteins (MOMPs). The MOMPs of isolates associated with disease in cattle had a relative molecular weight of approximately 41 kDa compared with 33 kDa for the MOMPs of isolates from asymptomatic cattle. The MOMPs from sheep isolates were either slightly higher or lower than the 41 kDa MOMPs of bovine isolates. Major antigens detected by Western blotting were similar in all isolates except the H. somnus-like isolate. An immunodominant 40 kDa antigen was conserved in all H. somnus isolates. Antibodies to this antigen have previously been found to be protective in cattle and may also be protective for sheep. Marked differences between cattle and sheep isolates were revealed by use of restriction enzyme analysis, which separated the isolates into 12 ribotypes and 15 unique DNA profiles. Thus, cattle and sheep isolates in this collection had distinctive differences in biochemical reactions, MOMP profiles, and DNA analyses. Such differences have potential value for epidemiological studies and may also be used to evaluate host specificity of H. somnus isolates.     

A Sero-Epidemiological Study of Haemophilus Somnus Infection in Dairy Cattle

Akhtar, S., Farver, T.B. and Riemann, H.P., 1997. A sero-epidemiological study of Haemophilus somnus infection in dairy cattle. Veterinary Research Communications, 21 (4), 229-239Five repeated cross-sectional serological surveys of 790 dairy cattle in 4 dairy herds between December 1985 and February 1987 provided an opportunity to study the changes in the seroprevalence of Haemophilus somnus across the 5 surveys and with respect to some demographic and disease variables. The demographic variables included were age (heifers or cows) and farm, representing two groups of herds (two herds in each group, located in the Central and Northern Valleys of California). The serological status of cattle as either negative or positive against H. somnus, Campylobacter fetus and Leptospira hardjo were determined with enzyme linked-immunosorbent assays. Logistic regression analysis was used to compute maximum likelihood estimates of adjusted odds ratios and their respective 95% confidence intervals.The baseline risk of being H. somnus seropositive in the cattle observed at first sampling did not vary significantly during the study period after adjustment for the effects of covariates. Only at first sampling were cows about twice as likely to be H. somnus seropositive than heifers. At samplings 1 and 3, but not at 2 and 5, being in the herds of the Central Valley appeared protective. In contrast, at sampling 4 the cattle in herds in the Central Valley were about 7 times more likely to be H. somnus seropositive. C. fetus-positive cattle were about 3 times more likely to be H. somnus seropositive at sampling 1 only. The relationship between H. somnus status and L. hardjo was not significant during the study period.     

Further characterization of the agent causing coryza in turkeys

A total of 128 isolates of Alcaligenes faecalis, from the respiratory tract of turkeys and chickens, were identified and divided into two types designated type I and type II. Type I isolates were pathogenic in poults, hemagglutinated guinea pig red blood cells (RBCs), and did not grow on minimal essential medium (MEM) agar, and most did not grow in 6.5% NaCl broth. Type II isolates were nonpathogenic and nonhemagglutinating and grew on MEM agar, and most grew in 6.5% NaCl broth. Hemagglutination of guinea pig RBCs was a reliable characteristic for distinguishing type I from type II isolates, and it correlated with pathogenicity. In serological studies using 62 type I and 21 type II isolates, cross-reactions were observed when type I but not type II antigens were used to test antisera in the microagglutination test. Eleven bacterial isolates, different from type I and type II isolates, were urease-positive. Although frequently isolated from turkeys with coryza, these isolates were nonpathogenic and were always found in association with type I A. faecalis. Urease-positive isolates and type I and type II A. faecalis isolates were stable following 50 in vitro passages. Bordetella avium sp. nov. (the nomenclature suggested in Europe for A. faecalis) was pathogenic in poults. The colonial morphology, biochemical characteristics, and hemagglutinating activity of B. avium sp. nov. were the same as those of type I A. faecalis isolates. Based on the results of these studies, it was concluded that type I A. faecalis is the etiologic agent of turkey coryza.     

Aerosol challenge of calves with Haemophilus somnus and Mycoplasma dispar

The aim of the study was to examine the ability of Haemophilus somnus and Mycoplasma dispar to induce pneumonia in healthy calves under conditions closely resembling the supposed natural way of infection, viz. by inhalation of aerosol droplets containing the microorganisms. The infections were investigated by recording clinical data, cytokine expression of peripheral blood cells and pathology. Twelve calves were included in the study: Three animals were exposed to H. somnus only, and two to M. dispar only, whereas five were challenged to M. dispar followed by exposure to H. somnus 11-14 days later. Also, one calf was exposed to M. dispar followed by exposure to a sterile saline solution 11 days later, and one calf was only exposed to a sterile saline solution. Just one animal, only challenged with H. somnus, developed a focal necrotizing pneumonia, from which H. somnus was isolated. Thus, the ability of H. somnus and M. dispar to act as primary pathogens under these conditions were minimal and inconsistent.However, a transient rise in body temperature, a marked granulocytosis and increased levels of interleukin-8 in peripheral blood after inoculation with H. somnus indicated a clear systemic response, probably as a consequence of the natural non-specific local and systemic defence mechanisms acting in healthy calves.     

Adherence of Haemophilus somnus to tumor necrosis factor-alpha-stimulated bovine endothelial cells in culture.

Vascular thrombosis and tissue infarction is a principal lesion in Haemophilus somnus septicemia known also as thrombotic meningoencephalitis. This study was undertaken to examine whether tumor necrosis factor-alpha (TNF-alpha) can influence the adherence of H. somnus to cultured bovine aortic endothelial cells (BAEC). Confluent BAEC were exposed to 0-100 nM of human recombinant TNF-alpha for 12-48 h. Suspensions of different strains of H. somnus (approximately 1.5-3 x 10(8) labelled with [methyl-3H]-thymidine, were added to BAEC and incubated for 1.5 h. Initial studies with one pathogenic (P) strain and one non-pathogenic (NP) strain revealed that both strains adhered to normal endothelial cells but minimally to subendothelial matrix remaining after removal of BAEC. Adherence to BAEC was reduced by an excess of unlabelled H. somnus of the same strain. Adherence was enhanced for both strains by exposure of BAEC to TNF-alpha in a manner that increased with TNF-alpha concentration and with duration of exposure to TNF-alpha prior to addition of bacteria. A survey of adherence of six live P strains and six NP strains demonstrated considerable variation but no difference in adherence between P and NP strains to normal or to TNF-alpha-stimulated BAEC. However, TNF-alpha consistently increased adhesion of each strain to BAEC. Both P and NP strains caused more severe cytotoxic changes in TNF-alpha-treated BAEC. Tumor necrosis factor-alpha also increased adhesion of formalin-killed bacteria of P and NP strains.(ABSTRACT TRUNCATED AT 250 WORDS)