Mycoplasma bovis in cattle : evaluation of risk factors and control measures for M. bovis infections

During recent years Mycoplasma bovis has spread to many countries that had previously recorded no M. bovis infections. M. bovis has also caused more severe outbreaks in some countries where it was recorded. In Finland, the first case of M. bovis was detected in November 2012 from a calf-rearing unit. The first dairy farm that registered a positive infection occurred one month later. Different interest groups working with cattle had a mutual objective to attempt to control M. bovis in Finland. A three-year research project was established (2014–2017) and this thesis comprises three published articles on topics pursued during the project. Locating all M. bovis positive farms and controlling the infection are important factors in improving animal health, lowering the usage of antibiotics and reducing economic costs associated with M. bovis. The general objective of this thesis was to find the most effective tools to prevent and control M. bovis in Finland. We evaluated the risk factors for a country with recently recorded M. bovis infections and studied if control measures implemented in dairy farms might lower the risk for a farm to be recorded as M. bovis positive. In addition, we wanted to see if meat inspection data could be used to help to detect M. bovis positive farms and if the average daily gain of bulls at slaughter is affected by M. bovis. The following statistical and laboratory methods were used. In article I, meat inspection data were analysed using a mixed effects logistic regression model (lung lesions) and a linear regression model (average daily gain). In article II, we analysed more closely two dairy farms where typical initial sources of M. bovis were not found. Bacteriological culture and real time PCR targeting the oppD gene were used for analysing semen samples. Nasal smears of calves were also analysed using real time PCR. Whole genome sequencing of isolated M. bovis bacteria was used to verify epidemiological results. In article III, risk factors for M. bovis were evaluated with a mixed effects logistic regression model. Most nasal smears were analysed with the same real time PCR targeting the oppD gene as used with semen in article II. Milk samples were analysed with real time PCR in commercial laboratories (Pathoproof® Complete 16-kit, Thermo Fisher Scientific, Finland). From our results, we concluded that meat inspection data can be used with other methods to locate M. bovis positive farms. Lung lesions, especially pleuritis, were associated with animals from M. bovis positive farms. Animals from M. bovis positive farms did not grow less than animals from control farms, but it has to be taken into account that we did not evaluate the increased use of antibiotics or welfare issues and mortality. We also showed that two naive dairy farms got M. bovis through artificial insemination (AI) of M. bovis positive semen. In both farms, the first detected M. bovis mastitic cows had been inseminated with semen from an M. bovis positive lot. In our analysis, farms using an M. bovis positive bull in insemination showed only a trend in the risk analysis because only few lots were positive. With the univariable analysis, contaminated semen lots were a significant risk factor. Control measures implemented against M. bovis in dairy farms helped the farms to maintain a low risk level. The control measures mainly included raising calves separately from older animals, culling all M. bovis mastitis cases and analysing bacteriological mastitis samples for M. bovis routinely with PCR. Out of nineteen follow-up farms, thirteen reached a low risk level, meaning that M. bovis was not detected in three consecutive nasal swabs taken from calves approximately every six months and was not detected in routine bacteriological clinical quarter milk samples or periodical bulk tank milk samples using PCR to detect M. bovis. Our studies furnish new information on the risk factors associated with M. bovis and more knowledge on the effects of control measures in dairy farms. Contaminated semen has been considered a risk but our studies present an actual natural transmission to cows on two dairy farms and we were able to include the use of contaminated semen in our risk analysis. It would be beneficial to have M. bovis free semen available for countries that do not have M. bovis and for those trying to control M. bovis. Control measures appeared to be effective in dairy farms included in our study. Further research is needed to determine a more precise length of time needed to isolate the calves from older animals on M. bovis positive farms.

Nautojen Mycoplasma bovis-bakteeri aiheuttaa huolta leviten uusiin maihin ja aiheuttaen joissain maissa vakavampia taudinpurkauksia kuin aiemmin. Suomessa ensimmäinen tapaus todettiin marraskuussa 2012 vasikkakasvattamon hengitystietulehdusnäytteessä. Kuukausi tämän jälkeen löydettiin ensimmäinen positiivinen lypsykarjatila. Tämän väitöskirjan julkaisut liittyvät vuonna 2014 aloitetun kolmivuotisen tutkimushankkeen tuloksiin. Väitöskirjan tavoitteena oli löytää keinoja ehkäistä ja hallita M. bovis tartuntoja Suomessa. Selvitimme tartunnan leviämisen riskitekijöitä ja tutkimme, auttoivatko sovitut toimenpiteet tiloilla hallitsemaan tartuntaa. Lisäksi tutkimme lihantarkastustuloksien hyödyntämistä M. bovis -positiivisten tilojen löytämisessä ja M. bovis –tartunnan vaikutusta teurastettujen sonnien kasvuun. Tutkimuksen tuloksien perusteella lihantarkastustietoja voidaan käyttää muiden keinojen tukena M. bovis -positiivisten tilojen kartoittamisessa. Keuhkomuutoksia, erityisesti keuhkokalvontulehduksia havaittiin enemmän naudoilla, jotka olivat peräisin M. bovis positiivisilta tiloilta. M. bovis -positiivisilta tiloilta peräisin olevat eläimet eivät kasvaneet huonommin kuin kontrollitiloilta peräisin olevat, mutta tuloksissa pitää ottaa huomioon, että emme selvittäneet mahdollista antibioottien lisääntynyttä käyttöä sekä eläinten hyvinvointiin liittyviä asioita ja kuolleisuutta. Tutkimuksemme avulla saimme uutta tietoa M. bovis -tartunnan riskitekijöistä ja ymmärrystä vastustustoimenpiteiden tehosta lypsykarjatiloilla. Pystyimme toteamaan, että ainakin kaksi lypsykarjatilaa sai M. bovis -tartunnan keinosiemennyksessä käytetyn M. bovis -positiivisen siemennesteen välityksellä. Molemmilla tiloilla ensimmäiset todetut M. bovis -positiiviset lehmät olivat keinosiemennetty M. bovis -positiivisella siemenerällä. Lypsykarjatiloilla tehdyt vastustustoimenpiteet näyttivät auttavan tartunnan hallinnassa. Toimenpiteisiin kuuluivat vasikoiden kasvattaminen erikseen vanhemmista eläimistä, M. bovis -positiivisten utaretulehduslehmien poisto ja maitonäytteiden tutkiminen rutiinisti M. boviksen varalta PCR -menetelmällä. Seuranta-ajalla kolmetoista tilaa yhdeksästätoista saavuttivat pienen riskin tason eli M. bovista ei havaittu kolmessa peräkkäisessä noin kuuden kuukauden välein vasikoista otetuissa sierainlimanäytteissä eikä maitonäytteissä (yksilö tai tankkimaito).