From Haemobartonella to hemoplasma: molecular methods provide new insights

Hemotropic mycoplasmas (aka hemoplasmas) are the causative agents of infectious anemia in numerous mammalian species. Originally known as Haemobartonella and Eperythrozoon species, these organisms have been reclassified within the genus Mycoplasma. The development of new molecular assays has expanded our knowledge of this heterogeneous group of agents and allowed us to study their epidemiology and pathogenesis. The present review summarizes recently gained insights into feline hemotropic mycoplasmas, formerly known as Haemobartonella felis. Besides the two initially identified feline hemoplasma species, Mycoplasma haemofelis and Candidatus Mycoplasma haemominutum, we discovered a third novel hemoplasma in a Swiss pet cat; preliminary results suggest that the pathogenic potential of the latter agent depends on cofactors. In applying PCR-based assays, feline hemoplasma infections have been documented in domestic cats and wild felids worldwide. Differences between the three hemoplasmas in regard to response to antibiotic treatment and establishment of a carrier status have been reported. Additionally, besides an ostensible vector-borne transmission, direct transmission by aggressive interaction of cats or interspecies transmission might play a role in the epidemiology of these organisms. Based on a potential vector-borne and interspecies transmission, a zoonotic potential of hemoplasmas should be further investigated.     

Molecular identification of `Candidatus Mycoplasma haemovis' in sheep with hemolytic anemia

We examined the presence of hemoplasmas, hemotropic mycoplasmas, among 11 sheep (Ovis aries) with regenerative and hemolytic anemia and found six of them were positive by real-time PCR. The positive samples were then subjected to conventional PCR for direct sequencing of the 16S rRNA gene. Nucleotide sequences of all the positive samples were identified as the 16S rRNA gene of `Candidatus Mycoplasma haemovis' by phylogenetic analysis, demonstrating the infections with this particular hemoplasma species in Japan.     

Development and application of a real-time TaqMan(®) qPCR assay for detection and quantification of 'Candidatus Mycoplasma haemolamae' in South American camelids

Two alpacas from a herd in southwest Switzerland died for unknown reasons. Necropsy revealed chronic weight loss and pale mucous membranes. Infection with hemotropic mycoplasmas was suspected and subsequently confirmed by molecular methods. In order to investigate the epidemiological situation in this herd, a real-time TaqMan(?) qPCR assay for the specific detection and quantification of hemoplasma infection in South American camelids was developed. This assay was based on the 16S rRNA gene and amplified 'Candidatus Mycoplasma haemolamae' DNA, but not DNA from other hemoplasmas or non-hemotropic mycoplasma species. The lower detection limit was one copy/PCR, and the amplification efficiency was 97.4%. In 11 out of 24 clinically healthy herd mates of the two infected alpacas, 'Candidatus M. haemolamae' infection was confirmed. No correlation was found between bacterial load and clinical signs or anemia. The assay described herein enables to detect and quantify 'Candidatus M. haemolamae' and may be used in future studies to investigate the prevalence, pathogenesis and treatment follow-up of hemoplasma infections in South American camelids.     

Real-time PCR-based prevalence study, infection follow-up and molecular characterization of canine hemotropic mycoplasmas

Hemotropic mycoplasmas (hemoplasmas) have been reported in several mammalian species including dogs. Infections may lead to hemolytic anemia, but investigations in the dog had been hampered by the lack of adequate diagnostic methods. Only recently sensitive PCR-based assays were reported for the two canine hemoplasmas, Mycoplasma haemocanis and Candidatus Mycoplasma haematoparvum. By applying these assays, 15.4% of 460 dogs from the south of France tested hemoplasma positive. It was hypothesized that this high prevalence may be associated with the presence of Rhipicephalus sanguineus, a proposed vector for canine hemoplasmas. To address this hypothesis and expand the PCR-based knowledge on canine hemoplasmosis, we investigated dogs in a climatic zone that does not allow for the permanent establishment of R. sanguineus. Blood samples were collected throughout a year from 889 dogs in Switzerland: 1.2% of the dogs tested real-time PCR positive. The infection status was not significantly associated with anemia, age or gender. Phylogenetic analyses of Candidatus M. haematoparvum and M. haemocanis isolates revealed > or =99.8% identity to published sequences. All samples collected from three infected dogs throughout a follow-up period of < or =13 months tested PCR positive. Interestingly, the majority of the infected dogs either had been imported from or had visited regions where R. sanguineus is indigenous. Thus, canine hemoplasma prevalence was found to be low in a country with a climate incompatible with frequent occurrence of R. sanguineus. Nonetheless, veterinarians may expect hemoplasma infections in dogs with a travel history and/or after potential tick vector exposure.     

Occurrence of hemotropic mycoplasmas in non-human primates (Alouatta caraya,Sapajus nigritus and Callithrix jacchus) of southern Brazil

Hemoplasmas, the erythrocyte-associated mycoplasmas, have been detected in several primates, causing mostly subclinical infection. This study aimed to determine the prevalence of hemoplasma infection in captive and free-ranging monkeys from southern Brazil, as well as factors and hematological abnormalities associated with infection. Blood samples from 40 non-human primates (NHP) were tested for hemoplasmas and coinfections. An overall of 10/40 (25.0%) NHP tested positive for hemoplasmas using PCR-based assays, including 9/14 (64.3%) black howler monkeys (Alouatta caraya) and 1/24 (4.2%) black-horned capuchin (Sapajus nigritus). Infection was not statistically associated with anemia, but wild-born monkeys and male black howler monkeys were more likely to be positive when compared with captive-born animals and female black howler monkeys, respectively. The sequences from the black howler monkey hemoplasma were similar (94% identity) to the squirrel monkey hemoplasma (“Candidatus Mycoplasma kahanei”) and were phylogenetically located in a different cluster when compared to the human hemoplasma (“Candidatus Mycoplasma haemohominis”).     

Feline hemotropic mycoplasmosis (feline hemobartonellosis).

Hemotropic mycoplasmas represent an important cause of anemia in cats worldwide. Previously known as Haemobartonella species, sequencing of the 16S rRNA genes of these organisms has led to their reclassification as mycoplasmas. Two species have been identified in cats, M haemofelis and "Candidatus M haemominutum." The latter organism alone has not been associated with disease in naturally infected cats but may cause anemia in FeLV-infected cats and accelerate development of FeLV-induced myeloproliferative disease. The mode of transmission of these organisms remains enigmatic. Nevertheless, development of sensitive DNA-based tests for these unculturable organisms has improved the understanding of the epidemiology and pathogenesis of FHM. Cats with clinical signs and laboratory abnormalities consistent with FHM should be treated with doxycycline; enrofloxacin may represent an effective alternative. Transfusion with packed red blood cells after cross-matching may be required for severely anemia cats, and addition of prednisone may be required if the diagnosis of FHM is uncertain, or response to antimicrobials alone is insufficient. Affected cats should be tested for FeLV, the most common concurrent infection in cats with FHM.     

PCR检测新疆南疆部分地方品种绵羊的嗜血支原体和泰勒虫感染情况

为了解新疆南疆部分地方品种绵羊嗜血支原体和泰勒虫的感染情况,采用PCR法检测新疆南疆5个地方品种绵羊共100份血液DNA样本。结果表明:新疆南疆部分地方品种绵羊泰勒虫和嗜血支原体感染率分别为35.0%(35/100)和3.0%(3/100)。仅和田羊和多浪羊上检测到嗜血支原体感染,感染率分别为8.0%(2/25)和5.0%(1/20),不同品种羊的嗜血支原体感染率无统计学差异(P>0.05);泰勒虫感染率多浪羊最高,为60.0%(12/20),卡拉库尔羊感染率最低,为15.0%(3/20),不同品种羊泰勒虫感染率差异显著(P<0.05)。散养绵羊嗜血支原体感染率6.0%(3/50),圈养绵羊未发现感染,两者间无统计学差异(P>0.05);散养和圈养绵羊的泰勒虫感染率分别为26.0%(13/50)和44.0%(22/50),两者之间无统计学差异(P>0.05)。将PCR产物进行双向测序,对序列进行比对分析,发现3个嗜血支原体阳性样本均被鉴定为绵羊支原体,35个泰勒虫阳性样本均被签定为绵羊泰勒虫。说明新疆南疆部分地方品种绵羊的泰勒虫感染均较为普遍,而嗜血支原体的感染情况有待于进一步调查。     

Mycoplasms of the swine--A review]

The mycoplasmas constitute a group of microorganisms placed between bacteria and virus. The name, Mycoplasma, is derived from the mycelial morphology of the organisms. The minimal reproductive unit, the elementary body, measures 0.2-0.5 mum. Unlike bacteria, mycoplasmas are not confined by a rigid cell wall, but just by a thin membrane. For their cultivation, though common bacteriological technique is adequate, especially enriched media are required. Antibiotics, as a rule penicillins, are added to the medium for inhibition of bacteria. Up to the present, 5 porcine species of mycoplasma are known: Mycoplasma suipneumoniae, Mycoplasma hyorhinis, Mycoplasma hyosynoviae, Mycoplasma flocculare, and Acholeplasma granularum. The 4 species first mentioned are very common among swine in Denmark. A. granularum has not been demonstrated so far. Occasionally, other species of mycoplasma are found in swine. M. suipneumoniae is by far the most important porcine mycoplasma, being to-day regarded as the primary etiologic agent in porcine enzootic pneumonia. A pure mycoplasma infection usually results in only weak clinical signs of pneumonia, but the disease may be aggravated by secondary factors as bacteria, parasites, and bad housing conditions. Enzootic pneumonia is usually prevalent only in fattening units, where it tends to persist indefinitely. The mycoplasma infection is practically incurable. Control of the disease is attempted by the SPF-program launched by the Danish Meat Research Institute, Roskilde. In this connexion the high sensitivity of mycoplasmas to physico-chemical influence is of advantage, because it results in a low rate of survival of the organisms outside the host. A further advantage is afforded by the fast that M. suipneumoniae is a definitely swine-specific organism. The rest of the porcine mycoplasmas are of far lesser importance. Yet, M. hyorhinis may produce a sero-fibrinous inflammation of serous cavities and joints in pigs less than 10 weeks old, and M. hyosynoviae may produce arthritis in fattening pigs.     

Occurrence of 'Candidatus Mycoplasma turicensis' infection in domestic cats in Japan

We have examined the presence of hemoplasmas, hemotropic mycoplasmas, among domestic cats (Felis catus) in Japan by using a species specific PCR, and found 'Candidatus Mycoplasma turicensis', a recently recognized hemoplasma species. A total of 60 feline blood samples collected in 2004 and 2005 were subjected to PCR amplification for the detection of Mycoplasma haemofelis, 'Candidatus Mycoplasma haemominutum' and 'Candidatus Mycoplasma turicensis'. Six blood samples collected from domestic cats were found infected with the 'Candidatus M. turicensis'. All of them are also infected with other species of hemoplasmas, M. haemofelis and/or 'Candidatus M. haemominutum'. This is the first to demonstrate 'Candidatus M. turicensis' infections among cat population in Japan.