Serological comparison of type strains of porcine, bovine, and ovine mycoplasmas with atypical colony morphology

The type strains of Mycoplasma hyopneumoniae, M. flocculare, M. dispar, and M. ovipneumoniae, all characterized by nipple-less colonies on solid media, were compared serologically. By indirect hemagglutination and by complement fixation tests they were found to constitute a related group. By crossed immunoelectrophoresis a sharing of common antigens was demonstrated, whereas no cross reactivity was noted by the metabolism inhibition test.The type strains of Mycoplasma hyorhinis and Mycoplasma bovirhinis were included in the study for comparison. Although some cross reaction was noted, they appeared just moderately related to the nipple-less group as well as to each other.     

Pathogenicity of some Mycoplasma and Acholeplasma species in the lungs of gnotobiotic calves

Cloned cultures of 16 strains, representing nine different species of Mycoplasma and Acholeplasma, were inoculated intratracheally into gnotobiotic calves. Strains of M bovirhinis, M canadense, M verecundum, A axanthum and A modicum did not produce visible pneumonic lesions and were not reisolated from the lungs. Strains of M alkalescens and M arginini colonised the lower respiratory tract but failed to produce visible pneumonia. M bovigenitalium (strain M991/70) and M dispar (strain Gri226) both colonised the respiratory tract and induced pneumonic lesions estimated to involve up to 8 per cent (M bovigenitalium) and 17 per cent (M dispar) of the lung. Histologically M bovigenitalium produced a cuffing pneumonia and M dispar produced a interstitial alveolitis.     

Studies of bovine mycoplasma mastitis. 1. Review of literature on the occurrence of bovine mastitis with Mycoplasma involvement]

The clinical pattern as well as the pathologico-anatomic or histological changes due to mycoplasma mastitis are neither specific nor pathognomic. Mastitis pathogens so far described included M. bovis, M. bovigenitalium, A. laidlawii, A. axanthum, M. alkalescens, M. canadense, M. dispar, M. bovirhinis, strains of Group 7 according to Leach, strain ST 6, and ureaplasma strains. In the GDR, enzootic mastitis has been confined to A. laidlawii and A. axanthum.     

An improved method for the identification of Mycoplasma dispar

An indirect immunoperoxidase test is described for easy and reliable identification of Mycoplasma dispar. Cultures suspected of being M. dispar were grown on nitrocellulose filter papers and could be identified without having produced centre-forming colonies. These develop only after several passages in vitro and are required for standard identification procedures such as immunofluorescence or growth inhibition. It is to be expected that the method reported here could also be useful for the identification of other mycoplasma species forming centre-less colonies.     

Colonisation pattern of the respiratory tract of calves by Mycoplasma dispar

Eight conventionally reared, 1- to 11-week old Ayrshire calves were naturally infected by a strain of Mycoplasma dispar (M. dispar). The colonisation was quantitatively followed by nasal swab samples, transtracheal aspiration samples and by the examination of the whole of the respiratory tract for mycoplasmas at slaughter after a follow-up period of 7–10 months.The fairly uniform pattern of the colonisation by M. dispar was revealed: A high degree of colonisation, measuring 10⁵–10⁸ colour change units (ccu) per nasal sample, lasted for a period of 2–5 months and was followed by a slow decrease in titres. Seven of the calves still harboured M. dispar in their respiratory tracts at slaughter. Intermittently obtained transtracheal aspiration samples were all positive for M. dispar and the titres were regularly higher than those for the simultaneously taken nasal samples indicating a high ability of M. dispar to continuously colonize the more distal parts of the respiratory tract. It was demonstrated that the sensitivity of nasal swabbing in the detection of M. dispar infection largely depended on the phase of colonisation : The method was good for the detection of a fairly recent infection of M. dispar, but inadequate for detection of low grade carriers.In various phases, the calves also became infected by Mycoplasma bovirhinis and Acholeplasma laidlawii. Their ability to colonize the whole respiratory tract was lower than that of M. dispar.     

Cross-reactive proteins among eight bovine mycoplasmas detected by monoclonal antibodies

Whole cell proteins of eight bovine mycoplasmas (M. bovoculi, M. bovis, M. dispar, M. bovirhinis, M. arginini, M. verecundum, M. canadense, M. alkalescens) were separated by SDS-PAGE and transferred to nitrocellulose paper. Rabbit anti-M. bovoculi serum was found to react with immunoblots of all mycoplasma species tested. These cross-reactive proteins were in the range of 35,000-100,000 molecular weight. Monoclonal antibody MA25.5 developed against a M. bovoculi 94 kDa surface protein cross-reacted with a band of 62 kDa from M. dispar and three bands of 89, 85 and 74 kDa from M. arginini only while MA18.13 that recognized a band of 57 kDa from M. bovoculi did not react with the other species. The role of MA25.5 monoclonal antibody in inhibiting the growth of M. bovoculi, M. dispar and M. arginini was tested using the metabolic-inhibition (MI) test. Monoclonal antibody MA25.5 inhibited the growth of M. bovoculi and also inhibited M. dispar growth but at lower MI titers, while it showed no effect on the growth of M. arginini.     

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.     

The prevalence and level of colonisation by Mycoplasma dispar and other mycoplasmas on calf rearing farms

The prevalence and level of colonisation by respiratory mycoplasmas, especially by Mycoplasma dispar (M. dispar), in calves on 24 Finnish calf rearing farms were studied by using nasal swabs. A minimum of 5 calves from each farm was examined and 91.3 % of the 206 calves examined were 1 to 5 months old. Nine herds were selected on account a diagnosed respiratory disease problem, the others without anamnestic knowledge of their health condition.All 24 farms and 96.1 % of the calves examined were found to harbour one or more species of mycoplasma. M. dispar, M. bovirhinis and Acholeplasma laidlawii were isolated from 23, 19 and 3 farms or from 91.7, 51.9 and 3.4 % of the calves investigated, respectively.The prevalence for M. dispar was 97.8 % and the geometric mean titer 4.9 log₁₀ color changing units (ccu) among 1- to 5-month old calves belonging to the infected farms. The prevalence in the age-group of 6- to 12-month old calves was significantly lower (40 %). No significant difference was found in the level of colonisation by M. dispar among the one-month age-groups of the 1- to 5-month old calves. A high degree of colonisation among 1- to 2-month old calves was indicative of the high ability of M. dispar to spread and colonise the respiratory pathway of young calves in the conditions described. The same state still prevailing in calves aged 4 to 5 months supported the concept of a relatively long duration of a heavy colonisation by this mycoplasma. The prevalence of M. bovirhinis among 1-to 5-month old calves was lower than that of M. dispar. Any decrease in the level of colonisation of M. bovirhinis could not be demonstrated in older calves. The titer levels of both M. dispar and M. bovirhinis as well as the prevalence of M. bovirhinis were significantly higher in herds having a current respiratory disease problem than in healthy or mildly affected herds. The same relationship applied to the husbandry method of common pens as compared with individual pens. A causal relationship of increased colonisation level, either as cause or effect, to the respiratory disease is suggested, as well as a probably more indirect one to the common pen husbandry type.     

四川地区鸡毒支原体的分离鉴定及血清学定型

在成都、重庆等１０个市、地、县，从患有慢性呼吸道疾病的鸡场采集１０８份样品，分离的２７个菌株，经Ｌ型细菌检验，理化特性鉴定，符合支原体的生物学特性。通过血清学鉴定（生长抑制试验），证实分离的支原体与鸡毒支原体国际代表株Ｓ６为同一血清型。人工复制试验结果，表明分离的鸡毒支原体ＳＭＧ－１和ＳＭＧ－２具有高致病性，感染鸡出现典型的慢性呼吸道病症状     

Prevalence of mycoplasmas in the respiratory tracts of pneumonic calves.

The prevalence of mycoplasmas in the respiratory tracts of 148 pneumonic calves originating from 25 herds in the Netherlands is reported. Four types of culture media were used to isolate mycoplasmas: solid modified Edward medium, 2 types of Friis media, and A7B differential agar medium. Mycoplasmas were isolated both from nasal swab specimens and lung lavage fluids collected from live calves and from nasal mucosa and lung tissue specimens collected post mortem. All of the mycoplasma strains isolated could be identified as either Ureaplasma diversum (isolated from 80% of 25 herds), Mycoplasma dispar (92%), M. bovirhinis (88%), M. bovis (20%), M. bovigenitalium (4%), M. arginini (16%), or M. canis (12%). Isolation rates of M. dispar and U. diversum were considerably higher from lung lavage fluids than from nasal swab specimens. M. bovis was detected only in fattening herds and not in dairy herds. The respiratory tracts of 75% of the calves examined contained at least 2 mycoplasma species. In total, 25 different combinations of mycoplasma species were detected in specimens collected from noses and lungs. The pathogenic species U. diversum and M. dispar had not been isolated before in the Netherlands.     

牦牛肺炎病原多杀性巴氏杆菌和殊异支原体的分子鉴定与分析

青海省刚察县某牦牛产业基地牦牛发生咳嗽,鼻孔有分泌物流出,呼吸困难等症状的疾病。为快速确诊发病牛的致病原因,及时防控和治疗,从发病和病死的牦牛中,采集鼻腔拭子和肺脏、胸腔积液,利用PCR方法进行分子鉴定与分析。结果显示:此牛场中引发肺炎的病原是A型多杀性巴氏杆菌和殊异支原体两种病原体混合感染,测序结果显示5个有效样品中鉴定的为完全相同的病原,P.multocida-GC1的Kmt基因与参考序列中国株CP031554/3/2/1同源性在99%以上,M.dispar-GC1与参考序列的16S rRNA基因同源性在97%以上。同时在遗传进化角度再次分析确认了鉴定的P.multocida与中国和俄罗斯的A型多杀性巴氏杆菌聚成一大支;鉴定的M.dispar与殊异支原体聚为一支。结果表明,青海刚察地区牦牛存在感染多杀性巴氏杆菌和殊异支原体的潜在风险,该研究为牦牛呼吸道疾病与肺炎的病原学调查和病原的流行病学调查提供了参考数据。     

Isolation and characterization of a new porcine mycoplasma

In a study on the distribution of Mycoplasma sui- (hyo-) pneumoniae (M. suip.) among Danish swine it was found that most isolates possessing the characteristic colonial morphology of M. suip. would be inhibited significantly in the growth inhibition (g.i.) and metabolic inhibition (m.i.) tests by antiserum for a type strain* of this species. However, a few isolates were found to be completely unaffected by this antiserum. Five such strains have been recovered, viz. 4 from cases of catarrhal pneumonia in bacon pigs, 1 from the nasal cavity of a 40-kg pig. The pigs in question originated from 5 different herds. The recovery of 1 of the strains has been reported (Friis 1971b).     

Comparison of Mycoplasma hyopneumoniae strains by serologic methods

Six field strains of Mycoplasma hyopneumoniae isolated from pneumonic lungs of pigs, reference strains 11 and J of M hyopneumoniae, Ms 42 strain of Mycoplasma flocculare, and BTS 7 strain of Mycoplasma hyorhinis were compared serologically, using hyperimmune antisera produced in rabbits. All strains of M hyopneumoniae were closely related as determined with the disk growth-inhibition test; however, differences in zone sizes indicated that some antigenic heterogeneity existed. Cross-reactions were not detected between M hyopneumoniae, M flocculare, and M hyorhinis with the growth-inhibition test. The metabolic-inhibition test was more useful for detection of intraspecies antigenic difference than was the growth-inhibition test, since antigenic diversity was clearly detected among M hyopneumoniae strains. Slight cross-reactions were observed between M hyopneumoniae and M flocculare. Using 2-dimensional immunoelectrophoresis, antigenic differences were observed among M hyopneumoniae strains, although many common components also were detected in electropherograms. Mycoplasma flocculare possessed a close antigenic relationship to M hyopneumoniae, as determined by two-dimensional immunoelectrophoresis, whereas both organisms were less related to M hyorhinis. Evidence obtained in this study indicated that strains of mycoplasmas tentatively identified as M hyopneumoniae were similar antigenically, but evidence was obtained also of some diversity in antigenic structure among these strains.     

Biochemical Characterization of Mycoplasma bovirhinis,Mycoplasma dispar and Recent Bovine Isolates of Mycoplasma canis

The pattern and kinetics of substrate utilization by the type strains of Mycoplasma canis, M. bovirhinis and M. dispar and ten recent M. canis isolates from cattle were determined. Metabolism of a range of sugars and organic acids by M. dispar was detectable by measurement of oxygen uptake. Organic acids were not utilized by M. bovirhinis or M. canis, and there was no oxygen uptake during metabolism of glucose or other sugars, as monitored by a pH-change method. The M. canis strains varied in their ability to metabolize sugars; seven of the isolates from cattle had the distinctive ability to metabolize sucrose, and one isolate, plus the type strain (from a dog), metabolized N-acetylglucosamine. The M. bovirhinis strain metabolized maltose. However, all the test strains oxidized glycerol at high rates and with a high affinity. Oxidation of glycerol has been reported for other mycoplasmas from the bovine respiratory tract and leads to the production of hydrogen peroxide, a potential virulence factor.     

Evaluation of Some Serological Techniques for The Identification of Mycoplasma Hyorhinis and Mycoplasma Suipneumoniae

Eighteen strains of Mycoplasma hyorhinis and a strain of Mycoplasma suipneumoniae were tested in 4 serological tests, i. e., disc growth inhibition, metabolic inhibition, indirect haemagglutination and indirect epi-immunofluorescence. Only with immunofluorescence could all tested strains of M. hyorhinis be shown; no cross-reactions between M. hyorhinis and M. suipneumoniae could be detected. The other tests failed in many cases to identify strains of the same species, and they gave cross-reactions between M. hyorhinis and M. suipneumoniae.     

FP1株番鸭源禽Ⅰ型副粘病毒抗原性分析

为明确FP1株番鸭源禽Ⅰ型副粘病毒强毒与鸡新城疫强毒标准株F48E9之间的抗原性差异,本研究通过交互血凝抑制试验、血清交叉中和试验及雏番鸭免疫攻毒保护试验比较了FP1株番鸭源禽I型副粘病毒强毒与鸡新城疫强毒标准株F48E9之间的抗原性.结果显示同源阳性血清对病毒的血凝抑制效价比异源阳性血清对病毒的血凝抑制效价高2.66～4个滴度;两者之间的抗原相关值R为0.35;对于5日龄经肌注途径免疫接种1羽份量La Sota弱毒疫苗14d后的雏番鸭,以FP1株番鸭源禽Ⅰ型副粘病毒强毒、鸡新城疫强毒标准株F48E9分别攻击后,其保护指数分别为54.5、92.9.以上结果表明,FP1株番鸭源禽Ⅰ型副粘病毒与鸡新城疫强毒标准株F48E9存在较明显的抗原性差异,为鸭副粘病毒病疫苗的研制和该病的防控提供了依据.     

Characterization of Mycoplasma hyosynoviae strains by amplified fragment length polymorphism analysis,pulsed-field gel electrophoresis and 16S ribosomal DNA sequencing

Mycoplasma hyospnoviae strains from Denmark, Germany, Japan, Sweden, the Netherlands and the UK were examined for variations in the genomic DNA and within the 16S ribosomal RNA (rRNA) gene. Variations in the chromosomal DNA among 57 isolates recovered from the respiratory tract and joints of pigs, were investigated by analysis of amplified fragment length polymorphisms of the Bg/II and MfeI restriction sites and by pulsed-field gel electrophoresis of a BssHII digest of chromosomal DNA. Both methods allowed unambiguous differentiation of the analysed strains and showed similar discriminatory potential for the differentiation of M. hyosynoviae isolates. Concordant results obtained with the two whole-genome fingerprinting techniques evidence the considerable intraspecies genetic heterogeneity of M. hyosynoviae. Sixteen field strains of M. hyosynoviae and the type strain S16(T) were further examined for variation within the 16S rRNA gene. Ten field strains possessed the 16S rDNA sequences identical to the type strain, while the remaining six strains had sequences that differed by one to two nucleotides from that obtained from the type strain.     

Use of a polymerase chain reaction for detection of Mycoplasma dispar in the nasal mucus of calves.

A polymerase chain reaction (PCR) assay was developed for the detection of Mycoplasma dispar in nasal mucus samples collected from calves. The target DNA sequence was the 16S rRNA gene, and the fragment was selected within a region of high polymorphism. The specificity and detection limit of the method were determined. This method was then used for the detection of M. dispar in nasal swabs collected from 301 calves, including 155 clinical samples from animals showing signs of respiratory disease and 146 samples from healthy animals. PCR with generic primers was applied to the detection of Mollicutes, followed by the detection of M. dispar. Mollicutes were detected in 52.05% of clinical samples from healthy animals and in 90.96% of samples from sick animals. Mycoplasma dispar was detected in 6.16% of healthy animals and in 34.84% of sick animals. The PCR assay was useful in verifying the presence of M. dispar in calves and may be a useful tool in monitoring this mycoplasma in cattle herds.     

Serological Characterization of 8 Haemophilus Pleuropneumoniae Strains and Proposal of a New Serotype: Serotype 8

Eight strains of Haemophilus pleuropneumoniae isolated from 8 herd outbreaks of pleuropneumonia in pigs were studied by means of the slide agglutination test, the tube agglutination test, the IHA test and by gel diffusion.The 8 strains were antigenically homogeneous and serologically distinct from serotypes 1 through 7. It is therefore proposed to refer these strains to a new serotype: serotype 8, with strain 405 as the type strain.In addition to the serotype-specific capsular antigens, capsular antigen of serotype 3 (strain 1421) and serotype 6 (strain Femø) could be demonstrated in the 8 strains by means of the IHA test and by gel diffusion analyses.     

The association between serological evidence of mycoplasma infection and respiratory disease in feedlot calves.

Calves from five Ontario feedlots were bled on arrival and approximately 28 days later. Calves treated during this interval for undifferentiated respiratory disease were classified as cases and untreated calves were classified as controls. Serum was titrated blindly for antibodies to Mycoplasma bovis and Mycoplasma dispar. Indirect hemagglutination titers of 1:20 or more were assumed to reflect recent or current exposure, whereas 1:10 or less were not. The titers to M. bovis increased in all feedlots indicating active infection. The initial titers to M. dispar were higher than the titers against M. bovis, yet they increased in all feedlots except one, suggesting widespread infection with this organism. There was an increased risk (although not statistically significant) of being treated if the titer against M. bovis rose during the period. Calves with low M. dispar titers on arrival were at increased risk of being treated and titer increases were strongly associated with treatment (statistically significant). Thus, the serological results indicate high prevalence of M. bovis and M. dispar in the feedlot calves and that calves with increasing titers in particular to M. dispar are at increased risk of being treated for respiratory disease.