Environmental temperature and relative humidity may predispose to respiratory disease by enhancing pathogen survival, lowering host resistance to infection, ...
Br. vel . J. (1990) . 146, 4 1 9
EFFECTS OF AGE, ENVIRONMENTAL TEMPERATURE AND RELATIVE HUMIDITY ON THE COLONIZATION OF THE NOSE AND TRACHEA OF CALVES BY MYCOPLASMA SPP .
Z . WOLDEHIWET, B . MAMACHE and T . G . ROWAN* University of Liverpool, Departments of Veterinary Pathology and *Animal Husbandry, Veterinary Field Station, Leahurst, Neston, Wirral L64 7TE, UK
SUMMARY Nasal and tracheal swabs sequentially collected from three groups of eight calves between the ages of 1 and 98 days indicated that the nose and trachea were colonized by Mycoplasma spp . during the first weeks of life . Over 92% of all calves harboured Mycoplasma s pp . i n their noses when they were 2 weeks old, the rate of recovery falling gradually thereafter . The peak period of recovering mycoplasmas from the noses and tracheas of calves was at 6 weeks old . M. bovirhinis, M. arginini and Acholeplasma laidlawii predominated in the nose while M. dispar and M. bovirhinis predominated in the trachea . There was no association between rates of isolation and clinical signs of respiratory disease . There were no significant differences between the frequencies of isolation of Mycoplasma spp . from groups of calves kept under different environmental temperatures and relative humidities .
INTRODUCTION Several mycoplasmas which are regarded as pathogens of the lung are often present in the respiratory tract of healthy calves but the rates of isolation vary according to the health status and the age of calves (Thomas & Smith, 1972 ; Bennett & Jasper, 1977 ; Tanskanen, 1987) . More than 12 species of mycoplasma have been isolated from the respiratory tract of cattle but only a few, M. mycoides subsp . mycoides, M. bovis, M. dispar and Ureaplasma, are known to be pathogenic (Gourlay & Howard, 1979 ; Howard et al., 1987). It seems, therefore, that some mycoplasmas are normal commensals while others may cause disease either as primary or secondary agents (Gourlay & Howard, 1979) . It is generally believed that stress factors such as transportation, climatic changes or concurrent infection with other viral or bacterial agents may be necessary for these mycoplasmas to express their full pathogenicity (Gourlay & Howard, 1979) . Environmental temperature and relative humidity may predispose to respiratory disease by enhancing pathogen survival, lowering host resistance to infection, influencing the replication rate of the pathogen or precipitating disease in animals already harbouring the pathogen (Dennis, 1986) .
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The aim of the present study was to investigate the effects of age, environmental temperature, relative humidity and vaccination with a live attenuated viral vaccine on the colonization of the nose and trachea of calves by mycoplasmas .
MATERIALS AND METHODS Calves Three groups of eight Friesian male calves were purchased directly from farms with no recent problems of respiratory disease . The calves were maintained in a controlled environment chamber and fed as previously described (Woldehiwet et at., 1990) . One group of eight calves (group A) was exposed to an air temperature of 5 ° C and the other group (group B) was kept at 16 °C . For both groups, the relative humidity (RH) was approximately 58% . In a third experiment (group C) eight calves were kept at 17 °C and 92% RH from day 1 until they had a mean age of 45 days . Thereafter, the environment was changed abruptly to 5 °C and 58% RH . The environment was maintained at this level for 11 days before it was changed to 17 °C and 85% RH . All calves were vaccinated with a live attenuated thermosensitive vaccine against infectious bovine rhinotracheitis (IBR) (RLB 106ts strain, Tracherine, Smith Kline Animal Health Ltd, Stevenage, England) when they were 12 weeks old . Twice daily, the calves were clinically examined by a veterinary surgeon . Any coughing was recorded as mild, moderate or severe . Isolation and identification of Mycoplasma spp. Nasal and tracheal swabs were collected within the first day of life and every 14 days thereafter in groups A and B . In group C, samples were collected when the calves were less than 30 h old, at a mean age of 2, 4 and 6 weeks and at intervals of 1-3 days thereafter until they were 10 weeks old . Tracheal mucus was collected by introducing protected prenasal swabs as described by Jones (1987) with some modifications (Woldehiwet et al., 1990). Mycoplasma were isolated using the dilution method, followed by subinoculation onto solid media for identification purposes, as described by Gourlay & Howard (1983) . Numbers of mycoplasma were measured as colour changing units (CCU) (Rodwell & Whitcombe, 1983) . Analysis of data The xz test was used to analyse the differences in the frequencies of isolation of Mycoplasma spp . between nasal and tracheal swabs within groups of calves and between groups of calves (Snedecor & Cochran, 1980) . The significance of the differences of frequencies of isolation in nasal and tracheal swabs from swabs collected at different ages were analysed by the Mann-Whitney U-test .
RESULTS Initial samples were collected from 24 calves . One calf in group A was withdrawn from the experiment in the third week, because it was pyrexic and had to be treated . Subsequent samples were taken from 23 calves . Of the swabs tested, 93 of 169 nasal
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swabs and 85 of 169 tracheal swabs yielded growth of mycoplasmas which were positively identified to species level . Five species were identified from the nose and from the trachea (Tables I and II) . Mycoplasma bovirhinis and M. arginini were recovered from nasal swabs more frequently than from the tracheal swabs (x 2=5 . 95 ; P 0 . 05) . Effects of abrupt changes of environmental temperature and RH The abrupt changes in environmental temperature and relative humidity in group C did not have a significant effect in the number and type of mycoplasma isolated from
MYCOPLASMAS IN CALVES
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tracheal or nasal swabs, but the change from warm humid (17 ° C and 92% RH) to cold dry (5 ° C and 58% RH) increased the rates of isolation of M. bovis from 12% to 38% . Effects of vaccination with live IBR The vaccination of calves with live thermosensitive IBR vaccine did not significantly affect the number of CCU or the type of mycoplasmas isolated (Tables I, II, and III) (P> 0 . 05) . Clinical signs All the calves in group A and some calves in groups B and C had mild to moderate coughing which lasted for 2-21 days but there was no apparent association between coughing and isolation of Mycoplasma spp . from the nose or trachea . One calf in group A was withdrawn from the experiment because it became pyrexic and was treated . None of the other calves showed pyrexia or required treatment . DISCUSSION Respiratory disease is a major problem in housed calves . Of the many infectious agents which have been associated with calf pneumonia, respiratory syncytial virus, parainfluenza type 3 (PI-3) virus, Pasteurella haemolytica, P . multocida, Mycoplasma bovis and M. dispar are probably the most important (Stalheim, 1983 ; Stott et al., 1987) . Because most of these agents are also found in the respiratory tract of normal calves, it has been generally accepted that environmental factors such as ambient temperature and relative humidity may be necessary to precipitate disease . Several surveys on respiratory disease in cattle claim to show correlation between peaks of incidence and climatic changes . Roy (1980), for example, stated that the incidence of respiratory disease in the UK was greater in winter than in summer. Other workers presented experimental data showing that changes in environmental temperature and relative humidity may be associated with changes in bacterial flora and pneumonia (Jones & Webster, 1984) . Whittlestone (1976) reported that climatic changes may affect the incidence of enzootic pneumonia in pigs by influencing the survival of M. hypopneumoniae . Most Mycoplasma spp ., including those reported to cause respiratory disease under experimental conditions, are often found in the respiratory tract of healthy calves . Gourlay et al. (1970), for example, isolated Mycoplasma dispar from 60% and M. bovirhinis from 20% of lungs from apparently healthy calves of between 2 and 4 months of age . Thomas & Smith (1972) isolated M. bovirhinis from 91% of nasal swabs obtained from apparently healthy calves between the ages of 3 and 4 months and M. dispar from the tracheas and lungs of 55% of calves in the same age group . The same workers found that 20% of calves which were less than 2 days old harboured Acholeplasma laidlawii, and 10% had M. dispar in their noses and tracheas . Calves which were more than 10 months old had significantly fewer mycoplasma in their noses and tracheas . The main finding of the present study was that the number and type of Mycoplasma spp . that colonize the nose and trachea of calves was significantly influenced by the age of calves and not by the environmental temperature or humidity . By sampling the same calves sequentially until they were 3 months old, we have shown in this study that Mycoplasma spp . start to colonize the upper respiratory tract as soon as the calves were born . The peak rates of isolating mycoplasmas from the nose were obtained when calves were 2-6 weeks old and from the trachea when calves were 6-8 weeks old, with gradual
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reductions thereafter. The calves were weaned at 5 weeks of age . M. bovirhinis, M. arginini and A . laidlawii dominated the nasal isolates while M. dispar and M. bovirhinis dominated the tracheal isolates . Several workers had also reported that M. bovirhinis predominates in the upper part of the respiratory tract while M. dispar was found mainly on the distal end of the respiratory tract (Thomas & Smith, 1972 ; Gourlay & Howard, 1979 ; Stalheim, 1983) . M. dispar affects the ciliated epithelium of the respiratory tract while M. bovis affects lung parenchyma without apparently affecting the epithelium (Thomas et al., 1987) . No evidence was found to associate rates of isolation with clinical signs of respiratory disease but it is perhaps significant that individual and mean titres of Mycoplasma spp . isolated in the present study were relatively low . Several workers who isolated Mycoplasma spp . following outbreaks of respiratory disease reported higher titres of mycoplasma (Thomas et al., 1982 ; Tanskanen, 1987) . Tanskanen (1987) reported that the recovery rates and titres of M. dispar and M. bovirhinis were significantly higher in calves from farms with respiratory diseases than in calves from farms with no respiratory disease .
ACKNOWLEDGEMENT We are grateful to Dr J . Bradbury for her advice and help in the identification of Mycoplasma spp . and to Mr T . Houghton for technical assistance .
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