Molecular detection of Bartonella coopersplainsensis and B. henselae in rats from New Zealand.

AIM To identify Bartonella spp. in rats from New Zealand using molecular methods. METHODS DNA was extracted from the spleens of 143 black rats (Rattus rattus) captured in the Tongariro National Park, New Zealand. PCR was performed using Bartonella genus-specific primers amplifying segments of the 16S-23S rRNA internal transcribed spacer and citrate synthase (gltA) and beta subunit of the RNA polymerase (rpoB) genes. PCR products were sequenced and compared online with sequences stored in the database of the National Center for Biotechnology Information of the United States of America. RESULTS DNA sequences matching Bartonella coopersplainsensis and B. henselae were detected in samples from 22/143 (15.4%) and 3/143 (2.1%) rats, respectively. Co-occurrence of B. coopersplainsensis and B. henselae sequences was observed in the sample from one rat. CONCLUSIONS AND CLINICAL RELEVANCE Gram-negative fastidious bacteria belonging to the genus Bartonella are associated with a range of human diseases. Rodents play an important role as reservoirs of a broad range of Bartonella species. To our knowledge, this is the first report of a molecular detection of Bartonella spp. DNA in rodents from New Zealand, and the first identification of B. henselae DNA in rats, worldwide. Whereas the public health significance of B. coopersplainsensis remains undefined, B. henselae is the agent of cat scratch disease, and the presence of this bacterium in rats may have public health implications. Our results are preliminary and additional analyses of larger samples, preferably by bacterial culture, would provide more information on the prevalence and diversity of Bartonella spp., in particular B. henselae, in rats.

"Candidatus Neoehrlichia chilensis" sp. nov.: Molecular detection and characterization of a novel Anaplasmataceae in wild rodents from Valdivia, southern Chile.

This study aimed to screen wild rodents from southern Chile, for the presence of Anaplasmatacea. Spleen samples from 33 wild rodents trapped in Valdivia Province were screened by conventional PCR (cPCR), targeting the Anaplasmataceae 16S rRNA gene (16S). Positive samples were further evaluated, targeting a larger 16S fragment, groEL operon, and gltA gene, followed by sequencing and phylogenetic analysis. Anaplasmataceae DNA was detected in 15% (five of 33) of the tested rodents (Abrothrix sp. [four of five] and Mus musculus [one of five]). Analysis of sequenced products based on the 16S gene revealed high similarity with "Ca. Neoehrlichia mikurensis," "Ca. Neoehrlichia lotoris" and "Ca. Neoehrlichia arcana" (97.8%-98.6%). A lower similarity was observed with Candidatus Neoehrlichia groEL (89.7%-92%) and gltA (79.5%-79.9%) loci. According to the 16SrRNA, groEL and gltA phylogenetic analyses, two closely related genotypes of "Candidatus Neoehrlichia" spp. from Chile were observed, which clustered together in a separate clade from other species in this genus. This study suggests the presence of two genotypes of a novel species of "Candidatus Neoehrlichia," proposed as "Candidatus Neoehrlichia chilensis," circulating in rodents from Chile. This is the first report of "Ca. Neoehrlichia" species in rodents from America.

Morphological and molecular identification of Rhipicephalus (Boophilus) microplus in Nigeria, West Africa: a threat to livestock health.

The cattle tick Rhipicephalus (Boophilus) microplus was first reported in West Africa in Ivory Coast, in 2007. Since then it has made an aggressive eastward advancement having been reported in four other West African countries: Mali, Burkina Faso, Togo and Benin. We herein report the first molecular identification of this tick species in Nigeria, West Africa, and highlight the threat it poses to livestock health. A nation-wide tick survey was conducted in 12 out of 36 states across the various agro ecological zones of Nigeria over a 1 year period (April 2014-March 2015). In total 1498 ticks belonging to three genera collected from cattle were morphologically identified. Overall, Amblyomma species constituted the highest percentage of sampled ticks, 50.2% (752/1498), followed by Rhipicephalus (including the subgenus Boophilus) species, 29.4% (440/1498) and Hyalomma species, 20.4% (306/1498). The presence of Rh. (B.) microplus was identified morphologically from four out of the 12 states. This finding was confirmed for the first time in Nigeria using a molecular method targeting the ITS-2 region of the ticks in three of the 12 states. This study ascertained the presence of Rh. (B.) microplus in Nigeria in addition to a broad variety of cattle tick species, most of which are of veterinary importance. The implication of this finding is that there may be additional economic burden to livestock farmers due to increased cost of tick control occasioned by the acaricide resistance by this tick species widely reported from different climes. Additionally, there may be a potential upsurge in incidence of hemoparasitic infections in cattle leading to increased morbidity, cost of treatment and mortalities.

Molecular detection of zoonotic bartonellae (B. henselae, B. elizabethae and B. rochalimae) in fleas collected from dogs in Israel.

Fleas represent an acknowledged burden on dogs worldwide. The characterization of flea species infesting kennel dogs from two localities in Israel (Rehovot and Jerusalem) and their molecular screening for Bartonella species (Rhizobiales: Bartonellaceae) was investigated. A total of 355 fleas were collected from 107 dogs. The fleas were morphologically classified and molecularly screened targeting the Bartonella 16S-23S internal transcribed spacer (ITS). Of the 107 dogs examined, 80 (74.8%) were infested with Ctenocephalides canis (Siphonaptera: Pulicidae), 68 (63.6%) with Ctenocephalides felis, 15 (14.0%) with Pulex irritans (Siphonaptera: Pulicidae) and one (0.9%) with Xenopsylla cheopis (Siphonaptera: Pulicidae). Fleas were grouped into 166 pools (one to nine fleas per pool) according to species and host. Thirteen of the 166 flea pools (7.8%) were found to be positive for Bartonella DNA. Detected ITS sequences were 99-100% similar to those of four Bartonella species: Bartonella henselae (six pools); Bartonella elizabethae (five pools); Bartonella rochalimae (one pool), and Bartonella bovis (one pool). The present study indicates the occurrence of a variety of flea species in dogs in Israel; these flea species are, in turn, carriers of several zoonotic Bartonella species. Physicians, veterinarians and public health workers should be aware of the presence of these pathogens in dog fleas in Israel and preventive measures should be implemented.

Molecular detection of Rickettsia aeschlimannii in Hyalomma spp. ticks from camels (Camelus dromedarius) in Nigeria, West Africa.

Several species of the spotted fever group rickettsiae have been identified as emerging pathogens throughout the world, including in Africa. In this study, 197 Hyalomma ticks (Ixodida: Ixodidae) collected from 51 camels (Camelus dromedarius) in Kano, northern Nigeria, were screened by amplification and sequencing of the citrate synthase (gltA), outer membrane protein A (ompA) and 17-kDa antigen gene fragments. Rickettsia sp. gltA fragments were detected in 43.3% (42/97) of the tick pools tested. Rickettsial ompA gene fragments (189 bp and 630 bp) were detected in 64.3% (n = 27) and 23.8% (n = 10) of the gltA-positive tick pools by real-time and conventional polymerase chain reaction (PCR), respectively. The amplicons were 99-100% identical to Rickettsia aeschlimannii TR/Orkun-H and R. aeschlimannii strain EgyRickHimp-El-Arish in GenBank. Furthermore, 17-kDa antigen gene fragments of 214 bp and 265 bp were detected in 59.5% (n = 25) and 38.1% (n = 16), respectively, of tick pools, and sequences were identical to one another and 99-100% identical to those of the R. aeschlimannii strain Ibadan A1 in GenBank. None of the Hyalomma impressum ticks collected were positive for Rickettsia sp. DNA. Rickettsia sp. gltA fragments (133 bp) were detected in 18.8% of camel blood samples, but all samples were negative for the other genes targeted. This is the first report to describe the molecular detection of R. aeschlimannii in Hyalomma spp. ticks from camels in Nigeria.

16S rRNA gene mutations associated with decreased susceptibility to tetracycline in Mycoplasma bovis.

Mycoplasma bovis isolates with decreased susceptibilities to tetracyclines are increasingly reported worldwide. The acquired molecular mechanisms associated with this phenomenon were investigated in 70 clinical isolates of M. bovis. Sequence analysis of the two 16S rRNA-encoding genes (rrs3 and rrs4 alleles) containing the primary binding pocket for tetracycline (Tet-1 site) was performed on isolates with tetracycline hydrochloride MICs of 0.125 to 16 µg/ml. Mutations at positions A965T, A967T/C (Escherichia coli numbering) of helix 31, U1199C of helix 34, and G1058A/C were identified. Decreased susceptibilities to tetracycline (MICs, ≥2 µg/ml) were associated with mutations present at two (A965 and A967) or three positions (A965, A967, and G1058) of the two rrs alleles. No tet(M), tet(O), or tet(L) determinants were found in the genome of any of the 70 M. bovis isolates. The data presented correlate (P<0.0001) the mutations identified in the Tet-1 site of clinical isolates of M. bovis with decreased susceptibility to tetracycline.

Bartonella species in fleas from Palestinian territories: prevalence and genetic diversity.

Bartonellosis is an infectious bacterial disease. The prevalence and genetic characteristics of Bartonella spp. in fleas of wild and domestic animals from Palestinian territories are described. Flea samples (n=289) were collected from 121 cats, 135 dogs, 26 hyraxes and seven rats from northern (n=165), central (n=113), and southern Palestinian territories (n=11). The prevalent flea species were: Ctenocephalides felis (n=119/289; 41.2%), Ctenocephalides canis (n=159/289; 55%), and Xenopsylla sp. (n=7/289; 2.4%). Targeting the Intergenic Transcribed Spacer (ITS) locus, DNA of Bartonella was detected in 22% (64/289) of all fleas. Fifty percent of the C. felis and 57% of the Xenopsylla sp. contained Bartonella DNA. DNA sequencing showed the presence of Bartonella clarridgeiae (50%), Bartonella henselae (27%), and Bartonella koehlerae (3%) in C. felis. Xenopsylla sp. collected from Rattus rattus rats were infected with Bartonella tribocorum, Bartonella elizabethae, and Bartonella rochalimae. Phylogenetic sequence analysis using the 16S ribosomal RNA gene obtained four genetic clusters, B. henselae and B. koehlerae as subcluster 1, B. clarridgeiae as cluster 2, while the rat Bartonella species (B. tribocorum and B. elizabethae) were an outgroup cluster. These findings showed the important role of cat and rat fleas as vectors of zoonotic Bartonella species in Palestinian territories. It is hoped that this publication will raise awareness among physicians, veterinarians, and other health workers of the high prevalence of Bartonella spp. in fleas in Palestinian territories and the potential risk of these pathogens to humans and animals in this region.

Serum acute phase proteins as clinical phase indicators and outcome predictors in naturally occurring canine monocytic ehrlichiosis.

BACKGROUND: Canine monocytic ehrlichiosis (CME), caused by Ehrlichia canis, is an important tick-borne disease of global importance. Currently, limited information is available on the diagnostic and prognostic value of acute phase proteins (APPs) in dogs naturally infected with E. canis. HYPOTHESIS: APPs may be useful indicators of the clinical phase of CME and predictive of the clinical outcome (death or survival). ANIMALS: Fifty-six dogs naturally infected with E. canis and 7 clinically healthy control dogs. METHODS: C-reactive protein (CRP), serum amyloid A (SAA), haptoglobin (Hp), and albumin concentrations determined on admission were retrospectively compared among 27 dogs with nonmyelosuppressive CME, 29 dogs with myelosuppressive CME and 7 healthy dogs. Diagnosis of CME was based on clinical and clinicopathological findings, seropositivity to E. canis, polymerase chain reaction amplification of E. canis-specific 16S rDNA, microscopic observation of Ehrlichia sp. morulae in blood monocytes or some combination of these. RESULTS: Mean concentrations of CRP, SAA, and Hp were significantly higher in the myelosuppressed dogs compared with the other groups, but no significant differences were found in the concentration of albumin. Survival analysis of the affected animals indicated that APP concentrations were not associated with clinical outcome; the latter was strongly associated with pancytopenia (odds ratio for death 22.7) and neutropenia (odds ratio for death 7.7). CONCLUSIONS AND CLINICAL IMPORTANCE: CRP, SAA, and Hp serum concentrations on admission are useful indicators of the clinical phase of CME, but are not useful predictors of clinical outcome.

Combined MLST and AFLP typing of Bartonella henselae isolated from cats reveals new sequence types and suggests clonal evolution.

Bartonella species are Gram-negative, fastidious bacteria. Bartonella henselae is found in cats and transmitted to humans via cat scratches or bites causing cat-scratch disease, characterized by clinical symptoms with varying severity. The prevalence of bartonellosis among humans in Germany appears to be high, and severe clinical cases have been described. However, epidemiological data of B. henselae in cats are rare. In this study we determined the detection rates of Bartonella ssp. in cats by culture and real-time PCR. Furthermore, B. henselae isolates were genetically characterized by highly discriminatory amplified fragment length polymorphism (AFLP) and multilocus sequence typing (MLST). Bartonella spp. were isolated by culture from 11 (2.2%) of 507 blood samples. Out of 169 blood samples additionally analyzed by PCR, 28 (16.6%) were found positive for Bartonella spp., illustrating the advantage of PCR in Bartonella spp. detection. PCR-REA identified B. henselae in 27 cats and Bartonella clarridgeiae in one cat. B. henselae isolates from different geographical regions in Germany were genetically characterized by AFLP and MLST. Both methods confirmed genetic diversity of B. henselae on the strain level. MLST identified 11 new sequence types, all of them assigned to three clonal complexes as determined by eBURST. AFLP typing revealed genetic relation among the B. henselae isolates from the same geographical region. Combining AFLP typing and MLST/eBURST analyses revealed that B. henselae of the same AFLP subcluster belonged to the same clonal complex. Altogether these results indicate that B. henselae may evolve clonally.

Molecular detection of Rickettsia massiliae, Rickettsia sibirica mongolitimonae and Rickettsia conorii israelensis in ticks from Israel.

Rickettsioses are recognized as important emerging vector-borne infections of humans worldwide. Previous reports documented the presence of two spotted fever group rickettsiae in Israel, Rickettsia conorii israelensis and Rickettsia felis. The aim of this study was to characterize the diversity of rickettsiae in ticks collected from vegetation and the ground, from different parts of Israel. Non-engorged questing adult ticks were collected from 13 localities. A total of 131 tick pools, 83 of Rhipicephalus turanicus and 48 of Rhipicephalus sanguineus (each with 2-10 ticks per pool), were included in this study. In addition, 13 Hyalomma sp. ticks were collected. The ticks were molecularly screened for rickettsiae, targeting the citrate synthase (gltA) and the outer membrane protein A (ompA) gene loci. Rickettsia massiliae ompA DNA (100% sequence identity; 180 bp) was detected in 32 Rh. turanicus and 12 Rh. sanguineus tick pools. R. conorii israelensis was detected in three Rh. sanguineus pools. Rickettsia sibirica mongolitimonae ompA DNA (100% sequence identity; 182 bp) was found in one Hyalomma tick. This study reports the first detection of R. massiliae and R. sibirica mongolitimonae in ticks from Israel. This is the first report describing the presence of these human pathogens in the Middle East.

Molecular detection of Ehrlichia canis, Anaplasma bovis, Anaplasma platys, Candidatus Midichloria mitochondrii and Babesia canis vogeli in ticks from Israel.

: Ticks are vectors of important pathogens of human and animals. Therefore, their microbial carriage capacity is constantly being investigated. The aim of this study was to characterize the diversity of domestic animal pathogens in ticks collected from vegetation and the ground, from different parts of Israel. Non-engorged questing adult ticks were collected from 13 localities. A total of 1196 ticks in 131 pools-83 pools of Rhipicephalus turanicus and 48 of Rhipicephalus sanguineus (with two to ten ticks per pool)-were included in this study. In addition, 13 single free-roaming Hyalomma spp. ticks were collected. Screening by molecular techniques revealed the presence of Ehrlichia canis, Anaplasma platys, Anaplasma bovis and Babesia canis vogeli DNA in R. turanicus ticks. E. canis, A. bovis, B. canis vogeli and Candidatus Midichloria mitochondrii DNA sequences were detected in R. sanguineus ticks. Candidatus Midichloria mitochondrii DNA was also detected in Hyalomma spp. ticks. Neither Hepatozoon spp. nor Bartonella spp. DNA was detected in any of the ticks examined. This study describes the first detection of E. canis in the tick R. turanicus, which may serve as a vector of this canine pathogen; E. canis was the most common pathogen detected in the collected questing ticks. It also describes the first detection of A. bovis and Candidatus Midichloria mitochondrii in Israel. To the best of the author's knowledge, this is the first report describing the detection of DNA of the latter two pathogens in R. sanguineus, and of A. bovis in R. turanicus.

Prognostic indicators for canine monocytic ehrlichiosis.

In order to identify prognostic factors for survival in canine monocytic ehrlichiosis (CME), clinical records of 40 cases of CME were retrospectively studied. The dogs were assigned as survivors (n=21) and non-survivors (n=19), and their signalment, anamnesis, clinical and clinicopathological signs, and treatment protocols were compared. Pale mucous membranes, bleeding tendencies and weakness were more prevalent in the non-survivors compared to the survivors. Dogs in the non-survivor group had significantly lower white blood cell (WBC), hematocrit (HCT), and platelet (PLT) counts. Pronounced pancytopenia (WBC < 4 x 10(3) microL(-1); HCT < 25%; PLT < 50 x 10(3) microL(-1)) was found as a risk factor for mortality. In this study, severe leucopenia (WBC < 0.93 x 10(3) microL(-1)), severe anemia (PCV < 11.5%), prolonged activated partial thromboplastin time (APTT>18.25s) and hypokalemia (K<3.65 mmol/L) were each found to predict mortality with a probability of 100%. In contrast, WBC counts above 5.18 x 10(3) microL(-1), platelet counts above 89.5 x 10(3) microL(-1), PCV > 33.5%, APTT < 14.5s and serum potassium concentration above 4.75 mmol/L, each provided 100% prediction for survival. These prognostic indicators can be easily obtained at presentation, are inexpensive, and may be useful aids when treatment and prognosis are being considered.

Seroprevalence of Anaplasma phagocytophilum among healthy dogs and horses in Israel.

The presence of reacting antibodies to Anaplasma phagocytophilum has previously been demonstrated in Israel, both in humans and the golden jackal (Canis aureus syriacus). This study was undertaken to determine the seroprevalence of A. phagocytophilum antibodies in two additional potential hosts, domestic dogs and horses in order to investigate the possibility of exposure to the organism in Israel. Of 195 dogs tested, 9% were seroreactive with A. phagocytophilum antigen and 30% were seroreactive to Ehrlichia canis. Twenty-nine percent of the dogs seropositive for E. canis were also reactive to A. phagocytophilum. Two dogs had immunofluorescence antibody (IFA) antibody titres for A. phagocytophilum greater than E. canis. The equine serological survey (n = 300) revealed no seroreactive horses. The results presented in this study suggest that dogs in Israel could have been accidentally exposed to A. phagocytophilum, for example by ticks carried on migrating birds, however, the possibility of cross-reaction with E. canis should also be considered. In spite of the high prevalence of ticks on horses in Israel during the summer months, no evidence for exposure to A. phagocytophilum was apparent.

Drivers for the emergence and re-emergence of vector-borne protozoal and bacterial diseases.

In recent years, vector-borne parasitic and bacterial diseases have emerged or re-emerged in many geographical regions causing global health and economic problems that involve humans, livestock, companion animals and wild life. The ecology and epidemiology of vector-borne diseases are affected by the interrelations between three major factors comprising the pathogen, the host (human, animal or vector) and the environment. Important drivers for the emergence and spread of vector-borne parasites include habitat changes, alterations in water storage and irrigation habits, atmospheric and climate changes, immunosuppression by HIV, pollution, development of insecticide and drug resistance, globalization and the significant increase in international trade, tourism and travel. War and civil unrest, and governmental or global management failure are also major contributors to the spread of infectious diseases. The improvement of epidemic understanding and planning together with the development of new diagnostic molecular techniques in the last few decades have allowed researchers to better diagnose and trace pathogens, their origin and routes of infection, and to develop preventive public health and intervention programs. Health care workers, physicians, veterinarians and biosecurity officers should play a key role in future prevention of vector-borne diseases. A coordinated global approach for the prevention of vector-borne diseases should be implemented by international organizations and governmental agencies in collaboration with research institutions.

Fatal Vipera xanthina palestinae envenomation in 16 dogs.

Sixteen fatal dog envenomations by the snake Vipera palaestinae over a 14-y period are described. Most envenomations occurred during the late night hours in the warm months, and 8/16 dogs were bitten on the limbs. The most frequent clinical signs upon admission were soft tissue swelling and edema, local pain, depression, bleeding, lameness, dyspnea, and 6 dogs were in shock. Thrombocytopenia was present in 14/16 cases and increased hematocrit (13/16) and hemoglobin (9/16) concentration were the most common hematological abnormalities upon admission. Biochemical abnormalities included increased activities of muscle enzymes and alkaline phosphatase, hypocalcemia, and hypocholesterolemia. Creatine kinase activity was markedly increased in 2 dogs. During hospitalization serious complications in many dogs were disseminated intravascular coagulation, acute renal failure, seizures, cardiac arrhythmias, acute necrotizing pancreatitis and severe laryngeal edema; these required intensive and expensive therapies. Specific antivenin (10 ml) administered to 8/16 dogs did not prevent death. Glucocorticosteroids were given in 8 cases; however, their use was associated with complications. Four dogs suffered sudden death, 2 of which died 1-2 d after discharge. Necropsy performed on 3/16 dogs found soft tissue swelling and local bleeding at the envenomation sites as well as bleeding in several distal body organs and tissues.

Vipera palaestinae envenomation in 327 dogs: a retrospective cohort study and analysis of risk factors for mortality.

Vipera palaestinae (Vp), formerly a subspecies of the near east viper Vipera xanthina, is the most common poisonous snake in Israel and neighbouring countries (Jordan, Lebanon and Syria), and is responsible for most envenomations in humans and domestic animals. Hospital records were retrospectively reviewed for confirmed cases of Vp envenomations in dogs over a 13-year period and 327 cases were included in the study. Most envenomations occurred between May and October, and between 02:00 and 10:00 PM. The most frequent clinical signs included: local swelling and oedema (99.6%), viper teeth penetration marks (51%), tachypnoea (50%), panting (44%), increased body temperature (19.2%), tachycardia (>160/min, 19%), salivation (18%) and lameness (15.6%). Common haematological findings included: increased haematocrit (47%), increased haemoglobin concentration (45%), leucocytosis (39%), and thrombocytopenia (30%). The prothrombin time and activated partial thromboplastin time were prolonged in 68 and 21% of the dogs, respectively. Blood biochemistry abnormalities included increased activities of muscle enzymes, hyperglycaemia, hyperbilirubinaemia, hyperglobulinaemia and hypocholesterolaemia. The mortality rate was 4% (13 dogs). The following variables were significantly (p < 0.05) associated with mortality: body weight below 15 kg (p = 0.01), limb envenomation (0.008), envenomation at night (p = 0.025), severe lethargy (P < 0.001), hypothermia (p = 0.04), systemic bleeding (p = 0.001), shock (p = 0.007), dyspnoea (p = 0.002), tachycardia (p = 0.002), thrombocytopenia (p = 0.02), and glucocorticosteroid therapy (p = 0.002). Dogs younger than 4 years had a lower death risk (p = 0.01). The association of steroid therapy with increased mortality suggests that the use of steroids in Vp envenomations may be harmful. Specific antivenom therapy (10 ml/dog) was not associated with a higher survival rate, thus its use, dose and timing of administration should be further investigated.

Spirocerca lupi in dogs: prophylactic effect of doramectin.

Spirocerca lupi is primarily a parasite of dogs, which typically causes oesophageal nodules, aortic aneurysms, and spondylitis. This study investigated the efficacy of doramectin as a prophylactic agent for canine spirocercosis. Five beagle dogs were injected subcutaneously with doramectin (400 microg/kg on 3 occasions 30 days apart q30d), while 5 other beagle dogs served as untreated controls. All dogs were inoculated with 40 infectious S. lupi larvae (L3) one month after the last doramectin treatment. All control dogs and 4/5 treated dogs became infected. Two control dogs died of ruptured aortic aneurysms, while no deaths occurred in treated dogs. Oesophageal nodules appeared 40-103 day later in treated as compared to control dogs, and eggs appeared in the faeces 49-106 day later in treated as compared to control dogs. The mean faecal egg count on day 223 in the treatment group was reduced by 99.77%. All control dogs had thoracic radiographic changes during the study, while only 2/5 study dogs showed radiographic changes. This study shows that although doramectin did not entirely prevent canine spirocercosis it reduced the clinical signs associated with infection and delayed and reduced egg output.

Phylogenetic analysis of hemoplasma species: an international study.

Nearly complete 16S rRNA gene sequences for feline and canine hemoplasma isolates from Europe, Australia, Africa, and Asia showed almost 100% identity to those previously reported for United States isolates. Partial sequences of the RNA subunit of the RNase P gene were also determined, and RNase P-based phylogenetic analysis showed that the hemoplasmas are most closely related to the members of the Mycoplasma pneumoniae group.