Studies of Mycoplasma mastitis in cattle. 7. Mycoplasma mastitis in 3 dairy cattle herds

Reported in this paper is the occurrence of enzootic mastitis in three dairy cattle stocks. The outbreaks had been caused by Mycoplasma bovis, Acholeplasma laidlawii, Acholeplasma axanthum as well as by one unidentified strain of the family of mycoplasmataceae. All animals with positive response to mycoplasma tests were identified an selected by repetitive testing of cultures in milk samples which had been taken from all lactating and dry cows and heifers as well as by evaluation of organ samples obtained from slaughtered cows. Regular cleaning and disinfection of stands in cowsheds, cattle tracks, and milk parlours as well as disinfection of udders and milking cups worked extremely well throughout the action in control of those cases of enzootic mycoplasma mastitis.     

Studies of bovine Mycoplasma mastitis. 4. Serological classification of Mycoplasma strains isolated from 3 stocks

Thirty-two isolates from altered milk samples taken from cows with mastitis of two herds were serologically tested by the WHT against rabbit hyperimmune sera of 16 reference and type strains. The strains tested were associated and grouped in the following way: 16 M. bovis, nine A. laidlawii, and seven A. axanthum. The classification of on M. bovis strain was confirmed by SHT. Five isolates from another stock, with biochemical properties related to the family of mycoplasmataceae, were not serologically identified.     

Studies of bovine Mycoplasma mastitis. 3. Testing of biochemical properties of Mycoplasma strains isolated from 3 stocks

Thirty-two isolates of altered milk samples taken from cows with mastitis on three industrialised dairy cattle units were biochemically tested following five passes through no-inhibitor media, assessment of filtratability through 450-nm-membrane filter, and clone assay. Something between five and ten clones of each of the strains involved were tested for their growth properties both at 22 degrees C and in no-serum media, sensitivity to digitonin and capability of aesculin hydrolysis. TTC reduction (triphenyltetrazolium-chloride), as well as for decomposition of glucose, arginine, and urea. The properties found were characteristic of M. bovis, A. laidlawii, and A. axanthum, and of the family of mycoplasmataceaea in the strains of one of the stocks.     

Incidence and transmission of Mycoplasma bovis mastitis in Holstein dairy cows in a hospital pen: A case study

The objective was to determine the incidence and transmission of mycoplasma mastitis in the hospital pen in a dairy herd of 650 lactating cows after a hospital pen was established following an outbreak of this disease. Mycoplasma mastitis status was monitored for 3 months through repeated collection of milk samples from cows with clinical mastitis (CM) and from bulk tank milk. During the outbreak 13 cows were diagnosed with Mycoplasma bovis CM, 1 cow with Mycoplasma sp. mastitis and 8 cows showed signs of arthritis, 3 of which were confirmed as having M. bovis arthritis. M. bovis isolates from cows with CM, arthritis and bulk tank milk had indistinguishable chromosomal digest pattern fingerprints. Incidence rates of M. bovis CM cases in the milking and hospital pens were 0.01 and 1.7 cases per 100 cow-days at risk. Approximately 70% of cows with M. bovis CM became infected within 12 days of entering the hospital pen. Transmission of M. bovis in the hospital pen occurred as 3 episodes. Each episode corresponded to the introduction of a cow with M. bovis CM from a milking pen. Evidence indicates that cows with M. bovis CM from milking pens were the source of transmission of the disease in the hospital pen and thus their presence in the hospital pen appeared to be a risk factor for transmission of M. bovis mastitis in this single case study herd.     

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.     

Preliminary studies on the prevalence of Mycoplasma bovis mastitis in dairy in cattle in Australia

A highly sensitive and specific PCR (MB-PCR) was used in preliminary studies to detect M. bovis in milk samples to investigate its association with high somatic cell count (SCC), an indicator of subclinical mastitis and one of the factors in down grading the quality of milk. A total of 186 and 167 herds were tested with 43% and 62% of herds positive for M. bovis in Victoria and North Queensland, respectively. The quarter milks from 52 cows with persistently high SCC were tested by MB-PCR and culture to investigate the association of M. bovis with major mastitis pathogens (MMP). M. Bovis was detected in 77% of cows of which 19% alone had M. bovis without any other bacteria, 17% had M. bovis in combination with major mastitis pathogens and 40% had M. bovis in combination with non-major mastitis pathogens. We believe that M. bovis is widespread in dairy cattle and has the potential to produce disease alone or to predispose the udder to disease caused by major mastitis and environmental pathogens. These studies have revealed a hitherto unrecognised high prevalence of M. bovis in dairy cattle in North Queensland and Victoria in Australia. These initial studies also give a clear association between M. bovis and elevated somatic cell counts.     

The course of mastitides in cows infected into the mammary gland with strains of Mycoplasma bovigenitalium and M. bovis]

Six cows were inoculated into the mammary gland with eight mycoplasma strains isolated from the genital tract of bulls and two type strains. The milk of cows infected with Mycoplasma bovigenitalium strains isolated from the genital tracts of bulls showed a change in the appearance and contained large quantities of mycoplasmas and specific antibodies. The mastitis was most intense in about 9 days and began to subside in 17 days infection. The type strain of M. bovigenitalium PG11 failed to produce mastitis. On the other hand, the type strain of M. bovis PG45 produced severe mastitis after a 14-day latency period, with the infection spreading to the uninoculated quarters, causing atrophy of the mammary gland, and persisting till slaughter. The sera of all cows that developed mastitis after experimental infection contained high titres of specific antibodies. The two infecting mycoplasma species were recovered from the inner organs and mammary glands of these cows after slaughter.     

牛支原体引起奶牛乳房炎的诊断

牛支原体（Mycoplasma bovis）是引起成年牛乳房炎和犊牛肺炎、关节炎的主要病原之一。本文从有乳房炎临床症状的奶牛中采集牛奶样本,经细菌培养和支原体培养、特异性PCR与环介导等温扩增（LAMP扩增）和16S rRNA测序等病原学检测证实为牛支原体感染,同时还有其他细菌的混合感染。     

Experience in identification and control of mycoplasma in dairy cattle stock under routine laboratory conditions

Exacting demands have to be met by milk testing routine laboratories where mycoplasma species are to be cultured from milk samples. Cleanliness of sampling, immediate cold storage, and no-delay transport of the milk samples to the testing centre are of greatest importance to the informative value of all mycoplasma tests. In the summer season, it is recommended to freeze the milk samples immediately after sampling. Mycoplasma broth should not be inoculated to animals of the herd from which the samples had been taken. A modified mycoplasma culturing medium is described in this paper. It will enable wider introduction of mycoplasma diagnosis by more Regional Institutes of Veterinary Medicine. Also reported in this paper are diagnosis and successful control of enzootic mastitis caused by Acholeplasma laidlawii and Acheloplasma axanthum. Repeated bacteriological testing of milk and secretion of all cows in the herd helped in picking out all bacteriologically positive animals and isolating them from the negative individuals. All animals that had produced positive responses to bacteriological testing were killed, notwithstanding clinical udder and general milk findings. Definite success of any control action undertaken against mycoplasmic mastitis will depend strongly on no-delay bacteriological testing of milk and secretion samples from all cows of the given herd, as early as in the acute phase of mastitis caused by mycoplasma.     

Between-herd prevalence of Mycoplasma bovis in bulk milk in Flanders, Belgium

Mycoplasma bovis (M. bovis) is a highly infectious pathogen of cattle causing pneumonia, polyarthritis, otitis, and less frequently, subcutaneous abscesses, abortions and meningitis. Ineffective drugs treatments, culling of infected cows and loss of milk production can lead to significant economic loss on dairy farms. The early detection of cows excreting M. bovis bacteria to prevent mastitis outbreaks is warranted. Reports suggest that the risk of M. bovis mastitis is higher in larger dairy herds. The objective of this study is to estimate the herd-level prevalence of M. bovis in Flanders, Belgium by culturing bulk tank milk samples taken from dairy farms. Three bulk tank milk samples per dairy herd were taken over four weeks, with collection intervals of two weeks. Culturing was done after pre-incubation using modified Hayflicks media to increase the chances of recovery of bacteria. For the identification of M. bovis, tDNA intergenic spacer PCR was used. In three herds (1.5%) of the 200 herds sampled, M. bovis was isolated from one of the three consecutive bulk tank milk samples. We conclude that in Flanders in 2009 at least 1.5% of the dairy herds had one or more cows excreting M. bovis in the milk. The frequent monitoring of bulk tank milk to detect the presence of M. bovis, especially in expanding herds on farms that often purchase replacement animals, should be encouraged in order to detect the presence of M. bovis and to monitor the success of control procedures following an outbreak of mycoplasmal mastitis in the herd.     

Application of an indirect ELISA to milk samples to identify cows with Mycoplasma bovis mastitis

An indirect ELISA was used to detect antibodies to Mycoplasma bovis in milk samples collected from a herd with M bovis mastitis. Antibodies were detected in samples from nine cows which had developed clinical M bovis mastitis. Milk from only three consistently antigen-negative cows tested positive for M bovis antibodies. These results indicate the potential value of the indirect ELISA for the detection of cows which have recently developed M bovis mastitis during the early stages of an outbreak.     

Examination of cattle with respiratory diseases for Mycoplasma and bacterial bronchopneumonia agents

A total of 247 mycoplasma strains was isolated from 435 lungs, tracheobronchial secretions and nasal swabs originating from cattle with symptoms of bronchopneumonia. Mycoplasma (M.) bovis was found 89 times (36%) and was the most common mycoplasma species in the lungs. M. bovirhinis, M. bovigenitalium, M. spec. and Acholeplasma (A.) laidlawii were isolated 158 times (64%). Among these mycoplasmas M. bovirhinis was the most widespread species (114 isolations). In 55 cases (62%) M. bovis was associated with Pasteurella or Actinomyces (A.) pyogenes. The other mycoplasma species were found in 67 cases (42%) together with these bacteria. Without mycoplasmas Pasteurella and A. pyogenes occurred in 33 of the probes investigated (21%). Beside mycoplasmas Haemophilus (H.) somnus was isolated from 16 of 162 tracheobronchial secretions investigated. The results confirm earlier suppositions that in most of the cases bronchopneumonia of cattle is a multifactorial event, frequently associated with mycoplasmas--especially M. bovis.     

A field study of Mycoplasma bovis infection in cattle

After an outbreak of mastitis in cattle caused by Mycoplasma bovis a study was made in 5 herds with recent cases (principal herds) and in 4 control herds. In the principal herds, M. bovis was isolated from milk samples, nasal swabs, and from one vaginal swab. M. bovis was also isolated from nasal swabs of calves in 2 of the 4 control herds, whereas all milk samples and vaginal swabs from the control herds were negative. Evaluation of serum antibody titres to M. bovis among non-mastitic animals of 3 principal herds and 1 control herd showed no difference in distribution of the titre values, which generally were low. However, cows excreting M. bovis in the milk had high antibody titres. The way of introduction to the herds and the spread of the infection within the herds could not be established by the study, which was supplemented by a DNA restriction fragment analysis of a number of M. bovis isolates.     

Bovine mastitis in Ontario due to Mycoplasma agalactiae subsp. bovis.

Bovine mastitis caused by Mycoplasma agalactiae subsp. bovis was first diagnosed in 16 of 55 cows in an Ontario herd in Feburary 1972. A total of 182 of 598 (30.4%) cows from 33 of 64 (51.5%) farms in widely separated areas of the province were culturally positive. Herd incidence varied from 15 to 40% with one closed herd having an incidence of 61%. Four herds were investigated culturally and serologically by the growth inhibition test for 15 months. In the acute phase the organism was present in the milk in extremely high numbers and could still be isolated from a few cows after eight to 12 months. The sera from 89.5% of the animals with clinical mycoplasma mastitis produced a zone of surface "film" and/or colony inhibition and some cows remained positive for six to 12 months. The disease was experimentally reproduced with a pure culture of the organism isolated from the milk of a cow from one of the herds.     

Detection of mycoplasma in mastitic milk by PCR analysis and culture method.

Mycoplasma alkalescens, M. bovigenitalium, M. bovirhinis and M. bovis were directly detected from milk specimens by a polymerase chain reaction (PCR) when milk specimens were centrifuged and treated with mycoplasmal lysis buffer. The sensitivity of this PCR method was 110 to 1,400 colony forming units (CFU). This method was useful for the detection of mycoplasmas in milk specimens from cows at an early stage of mycoplasmal mastitis since a small amount of mycoplasma could be detect in milk without culture. The results were available within 12 hr, which is faster than conventional culture techniques. M. bovirhinis was detected in more than 70% of mastitic milk specimens when mycoplasmas were detected in milk specimens from 30 cows with mastitis by this PCR method.     

Persistence of Mycoplasma bovis infection in the mammary glands of lactating cows inoculated experimentally

To study the course of clinical mycoplasma mastitis and investigate its potential for persistence, 10(8) colony-forming units (cfu) of an Irish isolate of Mycoplasma bovis was inoculated aseptically into the right fore teat canal of three lactating cows. M bovis rapidly colonised the infected quarters and grew exponentially to more than 10(10) cfu/ml within the first three days, and spread to other quarters of each of the three cows within five to 10 days. After periods of between 24 and 72 hours the infected quarters became distended and sensitive to touch, and their secretions changed from containing visible particles, to a seropurulent exudate, to an aqueous suspension of fine particles which formed a sediment after a sample was collected. M bovis-specific antibody levels increased to varying degrees in all three cows. Subsequently, the concentrations of mycoplasma decreased to less than 10(7) cfu/ml in two of the cows, but remained at more than 10(8) cfu/ml to the end of the lactation of the other cow. Apparently normal milk was secreted by one of the cows within a month of the challenge, and by the other two cows at the start of their next lactation. However, in two of the cows subclinical M bovis infection persisted through the dry periods and into their next lactations.     

Studies of Mycoplasma mastitis in cattle. 5. Studies of udder pathogenicity of Mycoplasma and Acholeplasma strains of different origins

Isolated acholeplasma laidlawii strains exhibited highly differentiated behaviours regarding their udder pathogenicity. Twelve of 16 tested strains were pathogenic to udder. Symptoms of acute udder inflammation were caused by all ten A. laidlawii strains isolated from differentiated material of calf, but by only two of six strains isolated from differentiated material of cattle. Intracisternal instillation of both strains from milk and one strain each from udder skin or cervical mucus caused merely temporary disorders of secretion. Ultrasonic extracts of A. laidlawii strains, some of them additionally heated, were intracisternally applied, as well. Udder irritation was caused only by those acholeplasma strains which were udder-patha was assumed to be attributable to a toxin of the polysaccharide type. Pathogenicity to udder was recorded also from one M. alkalescens strain isolated from a nose swab taken of cattle as well as from two A. granularum strains isolated from calf lungs.     

Mycoplasmal mastitis in dairy cows in the Moghan region of Ardabil State, Iran

Mycoplasmas are an important and economically significant cause of mastitis in dairy cows in various parts of the world. The organisms are highly contagious, with the main reservoir of infection originating from cows with subclinical mastitis. In 1998 the 1st cases of bovine mastitis due to Mycoplasma bovis were diagnosed in Ardabil State, Iran. An investigation was carried out with the aim of establishing the extent of mycoplasma infections in dairy cows in Ardabil State. Milk samples obtained from 80 cows with clinical mastitis were cultured in the laboratory for the presence of mycoplasmas. Similarly, 48 bulk-tank milk samples were examined for the presence of mycoplasmas. A modified Hayflick broth was used to isolate the mycoplasmas and an immunoperoxidase test used for the species identification of the isolates. Mycoplasma bovis was isolated from 39 (48.75%) of the clinical mastitis samples and from 48 of the bulk-tank milk samples tested. This indicated that mycoplasma udder infections were more prevalent in dairy cows in Ardabil State than previously thought.     

Isolation of Mycoplasma bovis from bovine clinical mastitis cases in Northern Greece

Mycoplasma bovis was detected in 18/219 (8.2%) quarter milk samples collected from cases of bovine clinical mastitis in Northern Greece between November 1997 and March 1999. The cases occurred in 2/37 (5.4%) of the herds examined. The micro-organism was isolated from bulk milk samples (BTS) from the two positive herds but was not isolated from 111 composite milk samples collected from clinically healthy cows from all 37 herds. Isolates were identified as M. bovis by polymerase chain reaction (PCR) assay. Other micro-organisms were also isolated from the M. bovis positive samples. The M. bovis-positive cows had all been imported into Greece from other European countries.     

Prevalence of mycoplasmal bovine mastitis in California

Seven species of mycoplasma plus one or more unknown species were found to cause bovine mastitis in California. Both the frequency of cases and number of species of mycoplasma in samples received at the laboratory have increased from 1976 to 1978. By survey, nearly 4% of samples of bulk tank milk from dairy farms were found to contain mycoplasma of potential pathogenic significance. Acholeplasma laidlawii was frequently isolated from samples both from cows and from farm bulk tanks during wet, rainy weather in the spring of 1978, apparently as contaminants only. The prevalence of positive bulk tank milk samples in an area appeared to parallel the prevalence of clinical mycoplasmal mastitis problem herds.