Haemolytic anaemia is not common in cattle [11, 12] although parasitic anaemia occurs more commonly, particularly in tropical regions. Babesia, Anaplasma, Theileria and Mycoplasma species have all been reported to cause anaemia in cattle [13, 14]. Haemotropic Mycoplasma species have also been reported to cause haemolytic anaemia in small ruminants [15, 16] and pigs .
Initially haemorrhage secondary to significant calving trauma was suspected to be the most likely cause of anaemia in this case due to the history of vulval/vaginal bleeding soon after calving. This however was ruled out based on clinical examination and the history form the farmer reporting that the cow calved unassisted. Further support of the belief that acute haemorrhage was an unlikely cause of anaemia in this case was the stable clinical appearance of the animal in spite of a very low PCV which would suggest a more chronic, insidious disease progression (allowing the animal to establish compensatory mechanisms). Post-parturient haemoglobinuria was also considered as a potential differential diagnosis but phosphorus levels were within the reference range. Abomasal haemorrhage was ruled out based on the absence of melena and bright, appetant demeanour of the animal with no apparent abdominal pain. Post-mortem examination further confirmed that neither abomasal haemorrhage nor ulceration were evident in this animal.
Haematology and serum biochemistry indicated that the anaemia in this case was haemolytic and other potential causes of haemolytic anaemia were investigated. Heavy metal toxicities and adverse drug reactions have been reported to cause haemolytic anaemia in cattle [11, 17, 18] but could be ruled out in this case by the histories from the referring veterinary surgeon and the farmer. Similarly, haemolytic anaemias associated with onion, brassica and bracken toxicities were ruled out by history and laboratory findings.
Other parasitic and infectious causes of haemolytic anaemia such as babesiosis, anaplasmosis and leptospirosis were also considered as potential underlying differential diagnoses. Leptospirosis was considered to be very unlikely as this animal (and the herd from which it was referred) was vaccinated with a leptospirosis vaccine and there was no evidence in the history to suggest that this had been ineffectively instigated. This animal presented in November, which in Scotland makes tick borne infections such as anaplasmosis and babesiosis unlikely due to the environmental conditions at that time of year; in addition this cow was housed all year round therefore tick exposure was much less likely in this case.
Haemotropic mycoplasmas have been reported in a number of domestic species, as well as humans and are known to cause both chronic and acute disease . Haemotropic Mycoplasma species adhere to the erythrocyte cell wall and cause haemolysis, resulting in anaemia. The exact mechanism resulting in haemolytic anaemia is not yet fully understood, however several mechanisms have been proposed including direct damage to the erythrocyte cell wall and the development of autoantibodies resulting in an immune-mediated anaemia . In sheep, infection with M.ovis has been shown to increase the fragility of erythrocytes compared to PCR-negative control animals . Autoreactive actin antibodies have been identified in pigs infected with M.suis and it is likely that these are involved in lysis of infected erythrocytes . Autoantibodies have also been reported in haemotropic Mycoplasma infections in other species including cats, dogs, mice and rats [21–23]. To our knowledge the presence of autoantibodies has not yet been identified in cattle infected with M.wenyonii however it has been speculated that the hind limb oedema more commonly seen associated with M.wenyonii infections in cattle could result from the formation of local immune-complexes . In cats the association between Mycoplasma haemofelis and positive direct Coombs’ test has been well documented [1, 7]; however, to our knowledge there are no reports in the literature of this association occurring in cattle, indeed there are relatively few cases of immune-mediated haemolytic anaemia of any cause reported in cattle [11, 12].
Disease caused by Mycoplasma wenyonii typically presents in individual animals as reduced milk yield and pitting oedema of the hind limbs and udder or scrotum. In the animal described in this case study, the classically described oedema was not reported to be present at initial presentation by the referring veterinary surgeon and was not seen whilst the animal was at the SCPAHFS. This case highlights the fact that whilst hind limb and udder/scrotal oedema occur in typical cases of M.wenyonii infection, the absence of oedema cannot rule out infection, as atypical infections do occur. M.wenyonii should be considered a differential diagnosis in any cow presenting with anaemia, particularly if there has been a recent reduction in milk yield. If bacteraemia is at a sufficient level, it can be economically diagnosed by taking an EDTA-anticoagulated blood sample and preparing a fresh smear for submission to your local reference laboratory. Blood smears can be examined microscopically for erythrocyte inclusion bodies by a haematologist or clinical pathologist.
Anaemia is an inconsistent finding in previous reports of cattle infected with M.wenyonii; when reported, anaemia is most commonly mild to moderate and associated with other clinical signs such as pyrexia, malaise and/or oedema of the hind limbs and udder or scrotum [3, 9, 10, 24, 25]. The case reported here presented with a severe anaemia in the absence of associated clinical signs such as those previously mentioned, which to our knowledge has not yet been reported in a mature animal. Severe anaemia is more commonly reported in splenectomised calves experimentally infected with M.wenyonii parasites [26, 27], however there is one recent report of severe anaemia occurring in a naturally infected mature cow, which also presented with hind limb oedema . There are reports of adult cattle co-infected with M.wenyonii and other blood parasites such as Anaplasma species, Babesia species and Theileria species presenting with severe haemolytic anaemia  and in sheep concurrently infected with M.ovis and Anaplasma ovis severe haemolytic anaemia has been reported . There was no evidence of concurrent Anaplasma or Babesia infection in the case reported and Theileria species are not present in the United Kingdom.
The case reported was direct Coombs’ test positive, indicating an immune mediated response resulting in haemolysis which to our knowledge has not been reported in cattle infected with M.wenyonii before. However, as previously mentioned, an immune-mediated haemolytic process has been implicated in the underlying pathogenesis resulting in anaemia in other species infected with haemotropic Mycoplasma species and it is probable that this also occurs in cattle, but until now has not been identified.
Since haemotropic mycoplasmas are uncultivable [19, 20, 29] Mycoplasma infection has traditionally been diagnosed solely on the presence of inclusion bodies on blood smears as described previously . Specificity of this test is high however sensitivity can be low  due to the lack of a specific stain for M.wenyonii on blood smears. In addition, blood smear examination cannot be used to determine which Mycoplasma species is involved. Recent advances have enabled the use of 16S rDNA PCR and denaturing gradient gel electrophoresis (DGGE) to more accurately detect and speciate mycoplasmas, including haemotropic mycoplasmas. The primer set used in this case report are Mycoplasma genus-specific primers [4, 5] and are more commonly used in our laboratory to detect non-haemotropic mycoplasmas.
A volume of 200 μL whole blood in EDTA was sufficient to detect Mycoplasma in the first un-treated sample. However, Mycoplasma was not detected in 200 μL blood sampled during treatment and immediately prior to euthanasia; positive and negative template controls performed as expected. Mycoplasma was however detected in the latter sample when a much larger volume of blood was processed (7 mL). The limits of detection for Mycoplasma in bovine blood using this PCR are not known, but 200 μL may be inadequate to detect low Mycoplasma loads. Further evaluation of this assay with positive clinical material is needed.
How M.wenyonii spreads between cattle is currently unknown but arthropod vectors have been suggested as a possible mode of transmission. Arthropod transmission has been implicated in the spread of M.haemofelis between cats [7, 30]. In addition, transmission of Mycoplasma ovis by Culex annulirostris mosquitoes has been demonstrated in Australia . Extrapolating from this evidence, arthropod vectors could be a possible route of infection in cattle and this is supported by limited Animal Health and Plant Agency (AHPA) (formerly Animal Health and Veterinary Laboratories Agency (AHVLA)) data suggesting the presence of a seasonal incidence in M.wenyonii infection in cattle in the United Kingdom . It is also possible that management interventions that result in exposure of uninfected animals to blood from infected animals (e.g. vaccination, ear-tagging, dehorning, castrating etc.) may result in transmission of M.wenyonii, as has been suggested in other species . However, neither of these modes of transmission has been proven in cattle. Further work is warranted to determine the mode of transmission of Mycoplasma wenyonii in cattle and how this may affect the herd as a whole.
The recommended treatments for M.wenyonii infection are tetracyclines, macrolides or fluoroquinolones [2, 29], all of which have readily accessible licensed formulations in the United Kingdom. The use of tetracyclines has been reported in cattle to result in short-term improvement in clinical signs, however treatment is unlikely to shorten the duration of disease [3, 9, 10]. Oxytetracycline and doxycycline have both been used in cats to successfully treat M.haemofelis  and Lester et al.  report a case of successful treatment of a dog infected with M.haemocanis (formerly Haemobartonella canis) with tetracycline .
It is possible that prompt treatment with antibiotics may prevent further production losses and spread to other animals in the herd; however, there are no reports of complete cure being achieved in cattle infected with M.wenyonii and it is likely that Mycoplasma wenyonii -infected cows, in common with other species infected with mycoplasmas, remain chronic carriers for the rest of their lives [3, 19]. There is little information available on the effect of chronic haemotropic Mycoplasma infection on production  and further study is warranted to determine whether a chronic carrier status affects the lifetime production of the infected animal.