Toxicity of Alpholate to Cattle

Although Apholate is used as a sexual sterilant of both sexes of houseflies (Musca domestica L.) it can not be used for `systemic' chemosterilization of the prehypodermic larvae of the warble flies Hypoderma bovis (L.) and H. lineatum (De Vill.) because of its high toxicity to 4- to 5-month-old calves.

An intramuscular dose of 2.5 mg./Kg. killed the calves in 5 to 7 days. The pathognomonic clinical signs were impaction of the rumen, anorexia, depression, and general weakness. The hematopoietic system was affected. There was marked leukocytopenia characterized by lymphocytopenia within 24 hours of Apholate injections.

     

Serological diagnosis of human hypodermosis: A preliminary report

In France, farm workers are occasionally infested with the larvae of Hypoderma lineatum and H. bovis, which are primirily parasites of cattle. A specific serological test for diagnosing human cases is described here.The immunological responses in cattle are directed mainly against the proteolytic secretion of the first instar larvae. This secretion was used as an antigen for the diagnostic immuno-electrophoretic test in man. The antigen was highly specific and produced negative results to human sera from 108 patients with other forms of parasitoses and 23 non- parasitized controls.We report here on 13 cases of Hypoderma infestation in man. Ten of these were positive and three negative to the test. In the negative cases, the serum was collected 1–2 months after the emergence of the larvae. The immunological response disappeared rapidly after larval emergence; in one case within 9 days.     

An Intradermal Test to Detect Latent Warble (Hypoderma spp.) Infection in Cattle

An intradermal test was developed to screen cattle for infection with the first-instar larvae of the warble flies Hypoderma lineatum and H. bovis. The diagnostic antigen, prepared from the first-instar larvae of H. lineatum, produced a distinct dermal reaction in cattle infected with the first-instar larvae of either species, but not in cattle in which the infection could not be confirmed later either on necropsy or by the appearance of warbles in the back. The reaction to the antigen was unpredictable in cattle with warbles in their backs. The diagnostic property of the antigen was also demonstrated in rabbits and guinea pigs artificially sensitized to the antigen.     

Induction of immunity in calves to mycoplasma bovis infection of the respiratory tract

Immunity to colonization of the respiratory tract by Mycoplasma bovis (formerly Mycoplasma agalactiae subsp. bovis was induced in calves by inoculation of formalin inactivated organisms. Animals inoculated intramuscularly and then intratracheally with inactivated mycoplasmas had significantly fewer M. bovis in their lungs, compared with non-vaccinated animals, 3 weeks after intratracheal challenge with viable organisms. In contrast, there was no significant difference in the numbers of M. bovis isolated from the lungs of control animals and of calves given two intramuscular inoculations of inactivated organisms. These results indicate that stimulation of the local immune system is important in the development of resistance to M. bovis respiratory infection following vaccination with inactivated organisms.     

Histopathological findings,phenotyping of inflammatory cells,and expression of markers of nitritative injury in joint tissue samples from calves after vaccination and intraarticular challenge with Mycoplasma bovis strain 1067

Background

The pathogenesis of caseonecrotic lesions developing in lungs and joints of calves infected with Mycoplasma bovis is not clear and attempts to prevent M. bovis-induced disease by vaccines have been largely unsuccessful. In this investigation, joint samples from 4 calves, i.e. 2 vaccinated and 2 non-vaccinated, of a vaccination experiment with intraarticular challenge were examined. The aim was to characterize the histopathological findings, the phenotypes of inflammatory cells, the expression of class II major histocompatibility complex (MHC class II) molecules, and the expression of markers for nitritative stress, i.e. inducible nitric oxide synthase (iNOS) and nitrotyrosine (NT), in synovial membrane samples from these calves. Furthermore, the samples were examined for M. bovis antigens including variable surface protein (Vsp) antigens and M. bovis organisms by cultivation techniques.

Results

The inoculated joints of all 4 calves had caseonecrotic and inflammatory lesions. Necrotic foci were demarcated by phagocytic cells, i.e. macrophages and neutrophilic granulocytes, and by T and B lymphocytes. The presence of M. bovis antigens in necrotic tissue lesions was associated with expression of iNOS and NT by macrophages. Only single macrophages demarcating the necrotic foci were positive for MHC class II. Microbiological results revealed that M. bovis had spread to approximately 27% of the non-inoculated joints. Differences in extent or severity between the lesions in samples from vaccinated and non-vaccinated animals were not seen.

Conclusions

The results suggest that nitritative injury, as in pneumonic lung tissue of M. bovis-infected calves, is involved in the development of caseonecrotic joint lesions. Only single macrophages were positive for MHC class II indicating down-regulation of antigen-presenting mechanisms possibly caused by local production of iNOS and NO by infiltrating macrophages.     

Bovine babesiosis: The immunization of cattle with fractions of erythrocytes infected with Babesia bovis (syn B. argentina)

Soluble antigen which protected susceptible cattle against challenge with $\text{Babesia bovis}$ was extracted from $\text{B. bovis}$-infected erythrocytes by sonic disintegration and separation of the soluble from the insoluble matter by ultracentrifugation. The material was then fractionated by the precipitation of fibrinogen-like proteins. The precipitate contained the babesial antigens that were located on the stroma of the infected erythrocytes. Antigen originally located on the parasite remained in solution. Both fractions conferred protection on splenectomized calves against challenge with $\text{B. bovis}$. However, the fraction containing the parasite antigens appeared to have more potential for development as a killed vaccine because it was not heavily contaminated with antigenic material from bovine erythrocytes.     

Expression of Mycoplasma bovis variable surface membrane proteins in the respiratory tract of calves after experimental infection with a clonal variant of Mycoplasma bovis type strain PG45

The pathomorphological findings and the expression and distribution of variable surface protein antigens (Vsp) of Mycoplasma (M.) bovis were characterised immunohistochemically in lungs of eight calves following inoculation with a Vsp A-expressing clonal variant of M. bovis type strain PG45. Within 48 h post inoculation (p.i.) an innate immune response dominated by macrophages and neutrophils develops. The monoclonal antibodies (mAbs) 1A1 and 1E5 detected M. bovis Vsp antigens in paraffin tissue sections of seven calves. Vsp antigens were widely distributed and were already present at day two p.i. within macrophages and other lung compartments. Taken together, the results demonstrate that the bovine is unable to eliminate M. bovis during the time period examined. Based on the different immunohistochemical labelling patterns obtained with the mAbs, the results also support the speculation that the in vivo variability of Vsps together with immunological factors may contribute to the chronicity of pulmonary disease.     

Immunization of Crossbred Cattle (Bos indicus×Bos taurus) with Fractionated Midgut Antigens against Hyalomma anatolicum anatolicum

Crossbred calves (Bos indicus×Bos taurus) were immunized with a fractionated midgut supernate antigen (GS-F Ag from Hyalomma anatolicum anatolicum). The first inoculation on day 0 was given intramuscularly after emulsification with Freund's complete adjuvant; the second was given subcutaneously on day 14 in incomplete Freund's adjuvant; and the third on day 35 was given subcutaneously without adjuvant. Each injection comprised 1 mg of antigen protein. Ten days after the last inoculation, the immunized calves were challenged simultaneously with 1000 larvae and 20 pairs (20 males and 20 females) of adult H. a. anatolicum on one ear and a similar number of Hyalomma dromedarii ticks on the other ear. There was a significant decrease in the percentage larval engorgement and larval rejection of up to 34% on the immunized calves. A significant increase in the engorgement and preoviposition periods and a significant decrease in the engorged weight, egg mass weight and reproductive index were observed for adult female ticks when fed on the immunized calves. The GS-F Ag also induced a considerable degree of cross-protection in calves against H. dromedarii larval ticks.     

FAILURE OF VACCINE STRAINS OF BABESIA BOVIS TO REGAIN INFECTIVITY FOR TICKS DURING LONG-STANDING INFECTIONS IN CATTLE

SUMMARY Two strains of Babesia bovis that were known to have lost infectivity for the normal tick vector, Boophilus microplus, due to repeated blood passaging in cattle, were studied to determine whether the strains would regain infectivity for ticks during longstanding infections. Parasitaemlas were monitored in 4 chronically infected calves that were regularly infested with ticks. Two strains of ticks known to be susceptible to infection with unmodified strains of B. bovis were used. Adult female ticks that dropped from the calves on days that a parasitaemia was evident were tested for B. bovis infection. Sixty-six batches of ticks collected up to 279 days after infection of the calves produced 14 pools of larvae, none of which transmitted infection. Primary infections established from the chronic infections by subinoculation at 200, 259 and 333 days after infection of the calves were also not transmitted by ticks.     

Hypodermosis in cattle slaughtered in Nigde province,Turkey<Superscript>*</Superscript>

This study was carried out to investigate the prevalence of hypodermosis in cattle between January and June 2005 in Nigde province, which is located in the middle of Turkey. A total of 1336 cattle, which were slaughtered in Nigde, were investigated for Hypoderma larvae. The 68 out of 1336 cattle (%5.08) were found positive for Hypoderma larvae. A total of 536 Hypoderma larvae were found in the skin and subcutaneous tissue of the back of infested cattle. The 489 out of 536 larvae (%91.23) were identified as Hypoderma bovis and 47 out of 536 (%8.77) as H. lineatum. Number of Hypoderma larvae counted on single infested cattle varied between 1–45 and the mean number of Hypoderma larvae per cattle was 7.88 (536/68). Hypodermosis was recorded for the first time in cattle from Nigde. ^*This study was supported by the Scientific Research Projects Unit of Nigde University (FEB 2004/07).     

Colostral transfer of antibodies specific for Hypoderma lineatum proteins

Antibodies to Hypoderma lineatum were transferred to calves via colostrum. The antibodies transferred in the colostrum demonstrated specificity to all the H. lineatum first-instar proteins which were resolved by non-denaturing polyacrylamide gel electrophoresis and were capable of mediating a Type I hypersensitivity reaction. The kinetic decline of colostrum-acquired antibodies, the effect upon development of an autologous humoral response to H. lineatum and the host or parasite protective role of these antibodies are discussed.     

The use of tick transmission by Boophilus microplus to isolate pure strains of Babesia bovis,Babesia bigemina and Anaplasma marginale from cattle with mixed infections

Pure strains of Babesia bovi, Babesia bigemina and Anaplasma marginale were isolated from cattle infected with all 3 species as well as a Theileria sp. and Eperythrozoon teganodes, using only transmission by the tick, Boophilus microplus. Unengorged adult ticks transferred to susceptible cattle transmitted A. marginale, but not Babesia. Engorged adults gave rise to progeny that transmitted Babesia, B. bovis by larvae and B. bigemina by male ticks. The Theileria and E. teganodes were not transmitted by the ticks and thus did not appear in calves used for isolating the pure strains of Babesia and A. marginale.     

Infectious bovine keratoconjunctivitis: Bacteriologic,immunologic, and clinical responses of cattle to experimental exposure with Moraxella bovis

The bacteriologic, immunologic, and clinical responses of 3- to 4-month old Holstein-Friesian calves to experimental exposure with Moraxella bovis type 10900 has been investigated. After u.v. radiation and intraconjunctival exposure with 1.9 × 10⁷ microorganisms, each eye of 16 calves exhibited signs of blepharospasm, photophobia, and increased lacrimation. Bacteria were recovered from exposed eyes for 2–7 consecutive weeks before maximal clinical response occurred. The severity of the cases varied from eyes that exhibited mild signs to severe clinical cases with profuse lacrimation, conjunctival swelling, corneal opacity, and ulceration. By 70 days after exposure, M. bovis could not be recovered from any conjunctival swabs, and clinical signs were not observed. Four non-exposed control animals did not develop clinical signs nor was M. bovis recovered from conjunctival swabs.Lacrimal secretions collected at the time of and 1 week after maximal clinical response had significantly elevated levels of total protein as compared to those collected 3, 2, and 1 week before, and 2 and 3 weeks after maximal clinical response. A passive hemagglutination test, using tanned formalized sheep erythrocytes sensitized with M. bovis sonicate antigen, detected antibody in lacrimal secretions from 22 of 32 eyes. The appearance of specific antibody in lacrimal secretions correlated with the amelioration of clinical signs and the decline in numbers of M. bovis microorganisms recovered from conjunctival swabs.     

Detection of antibodies to Cooperia ssp. and Ostertagia ssp. in calves with the enzyme linked immunosorbent assay (ELISA)

The ELISA technique was found to be very useful for the detection and monitoring of anti-Cooperia and anti-Ostertagia antibodies in calves. Six differens antigens were used; saline extracts from third- and fourth-stage larvae and from adult worms of both genera.Some degree of genus specificity was found when using L₄ or adult antigens but not when L₃ antigens were used. Stage-specificity could be observed for Cooperia L₄ antigen for a limited period after primary single infection. These findings were supported by the results of immuno-electrophoresis.     

Prevalence of respiratory bacterial pathogens and associated management factors in dairy calves in Taiwan

This study aimed to investigate the prevalence at both farm-level and calf-level and to identify the risk factors of respiratory bacterial pathogens in dairy calves in Taiwan. The status of bovine respiratory disease (BRD) was evaluated by using the Wisconsin scoring system from a total of 400 pre-weaned calves from 32 different farms in Taiwan, then the nasopharyngeal swabs were collected. The prevalence of respiratory pathogens was 84.37% at farm-level and 45.50% at calf-level, and Pasteurella multocida (P. multocida) was the most prevalent pathogen. The presence of Mycoplasma bovis (M. bovis), P. multocida, Mannheimia haemolytica (M. haemolytica) and Histophilus somni (H. somni) were all higher in BRD positive calves than BRD negative calves, but only in H. somni was significant (P<0.001). Then nine farm management risk factors were analyzed by using multivariate logistic regression models to determine the risk factors of respiratory bacterial pathogens (farm and calf-level). In the result at farm-level, only unheated colostrum was significantly associated with pathogen positive farms (Odds Ratio (OR)=11.43). At calf-level, the predominant risk factor for each pathogen, M. bovis, P. multocida, M. haemolytica and H. somni, was late first colostrum feeding (OR=272.82), unheated colostrum (OR=3.41), waste milk feeding (OR=6.59) and high pneumonia treatment cost (OR=2.52), respectively. For effective preventive measures, farmer education on milk and colostrum feeding are urgently warranted.     

First report of the use of mucosal swabs of the palatine tonsillar crypt for detection of Mycoplasma bovis in naturally infected calves

ABSTRACT

Aims: To compare detection by real-time PCR of DNA from Mycoplasma bovis on mucosal swabs taken from the palatine tonsillar crypt and the mainstem bronchi of clinically asymptomatic calves after slaughter.

Methods: We compared the sensitivity of mucosal swabs taken from two sites: the palatine tonsillar crypt and the mainstem bronchi. Paired samples were taken post-mortem at slaughter from 55 clinically well calves from an infected herd and were tested by real-time PCR for the presence of M. bovis-specific DNA.

Results: Mycoplasma bovis DNA was detected in 51 palatine tonsillar crypt swabs (92.7 (95% CI?=?82.4–98.0)%) and seven mainstem bronchial swabs (12.7 (95% CI?=?5.3–24.5)%). All seven calves with positive mainstem bronchial swabs also had positive palatine tonsillar crypt swabs.

Conclusions: When compared to mucosal swabs of the mainstem bronchi, mucosal swabs of the palatine tonsillar crypt were seven times more sensitive for the post-mortem detection of M. bovis DNA. The viability of detected M. bovis was not assessed, because any cattle carrying viable or non-viable M. bovis DNA were determined to be a potential risk to eradication. Palatine tonsillar crypt mucosa may be a useful anatomical site for real-time PCR detection of M. bovis DNA in naturally infected calves. More work is needed to define the persistence and viability of M. bovis at this anatomical site.

Clinical relevance: The results of this study helped form the basis of surveillance tools used in M. bovis control and eradication efforts. Familiarity with these results may help veterinarians better communicate with their clients about the science behind the eradication efforts.     

Immune response of calves following the inoculation of Mycoplasma dispar and Mycoplasma bovis

A small but significant reduction in the number of Mycoplasma dispar colonising the respiratory tract after intratracheal challenge was observed in gnotobiotic-calves previously inoculated subcutaneously three times with formalin-killed organisms and oil adjuvant. Injection of M. dispar by the intramuscular route, with oil adjuvant, and 2 weeks later by the intratracheal route, without adjuvant, failed to induce immunity to subsequent intratracheal challenge.Following the subcutaneous injection of killed M. dispar, the amount of antibody detected by single radial haemolysis (SRH) increased markedly with increasing age in groups of calves with average ages of 16 to 155 days when first injected. Most calves aged less than 40 days failed to produce an antibody response to a singel injection of M. dispar. With M. bovis a smaller difference was observed between antibody levels generated in calves of different ages; also, calves less than 40 days old produced a detectable SRH antibody response following a single injection of killed M. bovis.IgG1 and IgG2 antibody to M. dispar and M. bovis were measured by ELISA. IgG1 appeared before IgG2 antibody and this was particularly pronounced in younger calves. Also, for both mycoplasmas IgG2 antibody levels were lower in younger than older calves. The IgG1 response to M. dispar was compared in three groups of calves with average ages of 16, 55 and 155 days and was greatest in the oldest and least in the youngest animals. In contrast, the IgG1 response to M. bovis varied little in calves of different ages. It therefore appears that the immune response of young calves to M. dispar is impaired or defective.     

Establishment of an antibody avidity test to differentiate vaccinated cattle from those naturally infected with Mycoplasma bovis

Mycoplasma bovis is a major pathogen of bovine respiratory disease (BRD) in China and a live attenuated vaccine has recently been developed. This study aimed to establish an IgG avidity test to differentiate between naturally infected and vaccinated animals. An indirect ELISA (iELISA) was first established in the laboratory to detect antibodies specific to M. bovis using whole cell proteins as coating antigens and serum samples from experimentally infected cattle. The specificity and sensitivity of the iELISA was confirmed using a commercial ELISA kit as a reference standard. Both tests showed substantial agreement as indicated by a κ value of 0.78 (95% confidence interval, CI, 0.62, 0.93), and an overall 92.0% (80/87) agreement between the two tests. Based on the laboratory iELISA, a sodium thiocyanate (NaSCN) competitive iELISA was then developed for the detection of IgG avidity, expressed as relative avidity index (AI).Two-hundred and one experimentally immunised and naturally infected animals were used. These comprised 36 immunised calves, 38 negative control calves, 37 naturally infected calves, 87 calves of unknown status, and an additional three immunised calves that were used for a time trial. By testing true positive and negative antisera from either naturally infected or immunised calves, the AI cut-off value was defined as 70.4%. The diagnostic accuracy of the in-house NaSCN competitive iELISA was determined using serum samples collected from the experimental animals. The IgG avidity test demonstrated 96.0% sensitivity (95% CI 80.5%, 99.3%) and 95.8% specificity (95% CI 79.8%, 99.3%), and was successfully established as a valuable first test for differentiating vaccinated animals from those infected with M. bovis. This test may be a useful tool for clarifying the magnitude of M. bovis infection and in assessing the efficacy of vaccination in exposed animal populations.     

Bacteriological, serological, pathological and immunohistochemical studies of Mycoplasma bovis respiratory infection in veal calves and adult cattle at slaughter

Mycoplasma bovis is an important cause of calf pneumonia worldwide. In this study, we examined 140 cattle at slaughter comprising 70 veal calves and 70 beef cattle; 115 animals with pneumonic lesions and 25 without. Lung samples were submitted for bacteriological, histological, and M. bovis-immunohistochemical analyses. Serology for M. bovis was positive in 76% of beef cattle and 100% of veal calves. M. bovis was isolated only from veal calves in 16 out of 64 pneumonic cases. M. bovis was detected by immunohistochemistry in seven bacteriologically positive cases. M. bovis antigen was associated with bronchogenic necrosuppurative or fibrinonecrotizing lesions. Bacteriologically positive and immunohistochemical negative cases were associated with catarrhal bronchointerstitial pneumonia. Results suggest that M. bovis infection may develop into a severe necrosuppurative bronchopneumonia or fibrinonecrotizing pneumonia when associated with a high number of intralesional organisms or, conversely, into a mild catarrhal bronchointerstitial pneumonia when associated with a low number of organisms.     

Immunoprophylaxis of experimental Mycoplasma bovis arthritis in calves. Protective efficacy of live organisms and formalinized vaccines

Live Mycoplasma bovis (M. bovis) organisms given subcutaneously or intraperitoneally protected nine of ten calves and eight of nine calves, respectively, from clinical arthritis, while the formalinized vaccine given subcutaneously protected eight of ten calves. In contrast, clinical arthritis was induced in all non-vaccinated calves that were challenged intravenously. The arthritic lesion was more severe in non-vaccinated calves than in the few vaccinated calves that developed clinical arthritis. Unlike formalinized vaccine, live M. bovis culture given subcutaneously provoked a local reaction at the site of injection in most calves in the form of oedematous plaques of about 7–8 cm in diameter. Results suggest that the formalinized vaccine may offer a practical approach to the control of Mycoplasma bovis arthritis in calves.