N. F. Friis

Mycoplasma hyopneumoniae infection in pigs: Duration of the disease and evaluation of four diagnostic assays

200 SPF pigs were infected by aerosol with Mycoplasma hyopneumoniae and the development of clinical signs, serological and pathological reactions were studied. Mean time to onset of coughing was 13 days. A mean delay of 9 days was observed from onset of coughing until seroconversion against M. hyopneumoniae as measured by ELISA. At an individual level, the sensitivity for this ELISA was estimated to 98-100% and the specificity to 93-100%. Pasteurella multocida was isolated from the majority of the lungs 4 weeks post inoculation with M. hyopneumoniae and the lung lesions in pigs were significantly larger when P. multocida was present as compared to pigs with M. hyopneumoniae alone. An evaluation of cultivation, immunofluorescence, ELISA and polymerase chain reaction for demonstration of M. hyopneumoniae in lungs showed that all four methods have a high sensitivity in the acute stages of pneumonia. In the later stages the sensitivity of cultivation was superior to the other methods. No differences in specificity were observed between the methods. The antigen-ELISA OD values and the immunofluorescence scores revealed a strong positive correlation. Nasal swabs were additionally used for demonstration of M. hyopneumoniae and the polymerase chain reaction was found superior to the other methods.

A monoclonal blocking ELISA detecting serum antibodies to Mycoplasma hyopneumoniae.

A monoclonal blocking enzyme-linked immunosorbent assay (ELISA) for detection of antibodies to Mycoplasma hyopneumoniae in porcine serum has been developed. The monoclonal antibody (mAb) reacts with an M. hyopneumoniae specific epitope on a molecule of approximately 74 kDa. Only sera from M. hyopneumoniae infected pigs were able to block the binding of the mAb although antibodies from M. flocculare infected pigs also recognized a 74 kDa molecule. Sera from experimentally infected pigs as well as field samples were compared by the ELISA and by an indirect hemagglutination assay (IHA). In experimental pigs, the earliest detectable antibody response was found to be almost identical for both assays, but for some of the pigs the time of detection was significantly earlier by blocking ELISA than by IHA. In naturally infected herds more samples were found to be positive by ELISA than by IHA. Furthermore, the results indicate that sera from naturally M. flocculare infected pigs may give rise to cross-reactions in the IHA. The blocking ELISA appears to be a valuable and reproducible tool in the surveillance and serodiagnosis of M. hyopneumoniae infections in pigs.

Mycoplasma canis and urogenital disease in dogs in Norway

Mycoplasmas identified as Mycoplasma canis were isolated from nine dogs with clinical signs of urogenital disease in Norway over a period of 20 months. Some of the dogs had been treated unsuccessfully with antibiotics, and three were euthanased as a result of severe persistent disease. Seven of the dogs had a urinary tract infection, one had chronic purulent epididymitis and one had chronic prostatitis. Overt haematuria was frequently observed among the dogs with cystitis. M canis was isolated in pure culture from seven of the dogs and in mixed culture from the other two. In three cases the mycoplasma was cultivated only from urinary sediment, and it was typically obtained in smaller numbers than would be considered indicative of a urinary tract infection. In contrast with most mycoplasmas, the M canis isolated from all the dogs grew on ordinary blood agar plates used for routine bacteriological cultivation. Specific mycoplasma media were not used and the presence of other Mycoplasma or Ureaplasma species cannot be excluded.

Molecular analysis of field strains of Mycoplasma capricolum subspecies capripneumoniae and Mycoplasma mycoides subspecies mycoides, small colony type isolated from goats in Tanzania

A molecular analysis of strains of Mycoplasma capricolum subsp. capripneumoniae (M. capripneumoniae) and Mycoplasma mycoides subsp. mycoides, small colony type (M. mycoides SC) isolated from goats was performed using the amplified fragment length polymorphism (AFLP) and pulsed-field gel electrophoresis (PFGE) fingerprinting techniques. Among the 11 field strains of M. capripneumoniae from Tanzanian goats, two AFLP patterns were demonstrated, with 10 of the strains showing indistinguishable patterns. Five Kenyan strains of M. capripneumoniae produced three AFLP patterns, with two of them being indistinguishable from the 10 identical Tanzanian and one Ugandan strain (M74/93) isolated from sheep. The AFLP pattern of the type strain (F38(T)) was identical to two Kenyan strains (Baringo and G183/82). On PFGE analysis, all the examined M. capripneumoniae strains exhibited identical PFGE profiles.Five field strains of M. mycoides SC isolated from goats displayed identical AFLP patterns except for one strain which differed from others at only one position. The AFLP pattern of the type strain of M. mycoides SC (PG1(T)) was different from the field strains. The five field strains of M. mycoides SC produced identical PFGE profiles, which were, however, different from the type strain. The AFLP and PFGE profiles of M. mycoides SC strains from goats were identical to those of six strains isolated from cattle affected with contagious bovine pleuropneumonia (CBPP) in the same areas. The results of this study suggest a close epidemiological linkage between strains of M. capripneumoniae and between M. mycoides SC type, respectively, isolated from goats in Tanzania.

Genomic diversity among Danish field strains of Mycoplasma hyosynoviae assessed by amplified fragment length polymorphism analysis

Genomic diversity among strains of Mycoplasma hyosynoviae isolated in Denmark was assessed by using amplified fragment length polymorphism (AFLP) analysis. Ninety-six strains, obtained from different specimens and geographical locations during 30 years and the type strain of M. hyosynoviae S16(T) were concurrently examined for variance in BglII-MfeI and EcoRI-Csp6I-A AFLP markers. A total of 56 different genomic fingerprints having an overall similarity between 77 and 96% were detected. No correlation between AFLP variability and period of isolation or anatomical site of isolation could be demonstrated. An examination of the clonal appearance of M. hyosynoviae isolates recovered from seven herds affected with arthritis revealed presence of several genotypically distinct variants of the organism in a single herd, indicating that different strains may simultaneously be involved in an outbreak of the disease.

Absence of strictly age related resistance to Mycoplasma hyosynoviae infection in 6-week-old pigs

Mycoplasma hyosynoviae has never been reported to cause arthritis in pigs younger than 10 weeks of age. In order to investigate whether a strict age-related resistance exists, four 6-week-old pigs and four 13-week-old pigs, all immunologically naïve with respect to M. hyosynoviae, were inoculated intranasally with the agent and autopsied at day 11 or 13 after infection. One uninoculated pig per age group was included as a negative control. Just as the 13-week-old pigs, the 6-week-old piglets were susceptible to blood, joint and tonsillar infection with M. hyosynoviae and developed clinical arthritis following inoculation with the agent. Thus, we found no evidence of an age-related resistance to the infection.

Mycoplasma alkalescens demonstrated in bronchoalveolar lavage of cattle in Denmark

Mycoplasma alkalescens is an arginine-metabolizing mycoplasma, which has been found in association with mastitis and arthritis in cattle. Routine bacteriological examination of 17 bronchoalveolar lavage samples from calves with pneumonia in a single herd in Denmark, identified M. alkalescens in eight samples. The organism was found as a sole bacterilogical findings in five of the samples as well as in combination with Mannheimia haemolytica, Haemophilus somni and Salmonella Dublin. This is the first report of isolation of M. alkalescens in Denmark.

Mycoplasma hyopharyngis Isolation From Swine

Mycoplasma hyopharyngis is a seldom isolated porcine species. It was found, adequately described, and named by [3]. In that study 7 strains were isolated from the upper respiratory tract of pigs in 2 different herds, one of which was an institutional herd. All the strains fulfilled the usual criteria put up for characterization of mycoplasmas. They could degrade the amino acid arginine, but pathogenic capabilities were not reported. They were found antigenically distinct from all other mycoplasmas, inclusive the recognised porcine species. A later report on the isolation of M. hyopharyngis was done by [2] who in a single swine herd on 2 occasions found the microorganism in inflamed joints and adjacent subcutaneous abcesses of some animals. In a later phylogenetic study [6] found that the mycoplasma belongs to the Mycoplasma lipophilum cluster within the Mollicutes.

Transfer of maternal immunity to piglets is involved in early protection against Mycoplasma hyosynoviae infection

Mycoplasma hyosynoviae causes arthritis in pigs older than 12 weeks. The role of colostrum in protection of piglets against M. hyosynoviae infection is not clear. Our objective was therefore to investigate whether transfer of maternal immunity to piglets was involved in early protection against the infection. Experimental infections were carried out in three groups of weaners receiving different levels of M. hyosynoviae-specific colostrum components; Group NC derived from Mycoplasma free sows and possessed no specific immunity to M. hyosynoviae. Group CAb pigs, siblings of the NC group, received colostrum with M. hyosynoviae-specific antibodies immediately after birth. Group CCE pigs were born and raised by infected sows and presumably had the full set of colostrally transferred factors, including specific antibodies. When 4½ weeks old, all pigs were inoculated intranasally with M. hyosynoviae. The course of infection was measured through clinical observations of lameness, cultivation of M. hyosynoviae from tonsils, blood and synovial fluid and observation for gross pathological lesions in selected joints. Specific immune status in the pigs was evaluated through detection of antibodies by immunoblotting and measurement of M. hyosynoviae-specific T-cell proliferation. The latter analysis may possibly indicate that M. hyosynoviae infection induces a T-cell response. The CCE piglets were significantly protected against development of lameness and pathology, as well as infection with M. hyosynoviae in tonsils, blood and joints, when compared to the two other groups. Raising the CCE pigs in an infected environment until weaning, with carrier sows as mothers, apparently made them resistant to M. hyosynoviae-arthritis when challenge-infected at 4½ weeks of age. More pigs in group NC had M. hyosynoviae related pathological lesions than in group CAb, a difference that was significant for cubital joints when analysed on joint type level. This finding indicates a partially protective effect of passively transferred M. hyosynoviae-specific colostral antibodies upon development of M. hyosynoviae related pathology. Thus, the level of passive immunity transferred from sow to piglet seems to provide, at least partial, protection against development of arthritis. It cannot be ruled out that the CCE pigs, by growing up in an infected environment, have had the chance to establish an active anti-M. hyosynoviae immune response that complements the maternally transferred immune factors. Evident from this study is that the general absence of M. hyosynoviae arthritis in piglets can be ascribed mainly to their immunological status.