Investigation of possible rickettsial infection in patients with malaria.

Rickettsioses are a common health problem in many geographical areas, including rural areas in Southeast Asia. Co-infection of rickettsioses and malaria has been reported in Africa, where common reservoir and vectors are available. In this study, blood samples of Malaysian patients microscopically positive (n=148) and negative (n=88) for malaria parasites (Plasmodium knowlesi, Plasmodium malariae, Plasmodium falciparum, and Plasmodium vivax) were screened for the presence of rickettsial DNA, using PCR assays targeting specific genes. A partial fragment of rickettsial ompB gene was successfully amplified and sequenced from a patient microscopically positive for Plasmodium spp. and PCR-positive for P. vivax. BLAST analysis of the ompB sequence demonstrated the highest sequence similarity (99.7% similarity, 408/409nt) with Rickettsia sp. RF2125 (Genbank accession no. JX183538) and 91.4% (374/409 nt) similarity with Rickettsia felis URRWXCal2 (Genbank accession no. CP000053). This study reports rickettsial infection in a malaria patient for the first time in the Southeast Asia region.

Investigation of possible rickettsial infection in patients with malaria

@#Rickettsioses are a common health problem in many geographical areas, including rural areas in Southeast Asia. Co-infection of rickettsioses and malaria has been reported in Africa, where common reservoir and vectors are available. In this study, blood samples of Malaysian patients microscopically positive (n=148) and negative (n=88) for malaria parasites (Plasmodium knowlesi, Plasmodium malariae, Plasmodium falciparum, and Plasmodium vivax) were screened for the presence of rickettsial DNA, using PCR assays targeting specific genes. A partial fragment of rickettsial ompB gene was successfully amplified and sequenced from a patient microscopically positive for Plasmodium spp. and PCR-positive for P. vivax. BLAST analysis of the ompB sequence demonstrated the highest sequence similarity (99.7% similarity, 408/409nt) with Rickettsia sp. RF2125 (Genbank accession no. JX183538) and 91.4% (374/409 nt) similarity with Rickettsia felis URRWXCal2 (Genbank accession no. CP000053). This study reports rickettsial infection in a malaria patient for the first time in the Southeast Asia region.

Molecular evidence of hemoplasmas in Malaysian cattle and ticks.

Hemotrophic mycoplasma (hemoplasmas) is a cell wall-less bacterium causing infectious anemia in animals. As data on hemoplasmas infecting cattle in Malaysia is scarce, specific polymerase chain reaction assays were used for detection of hemoplasmas from blood samples of cattle and ticks in this study. Hemoplasma DNA was detected in 69 (69.0%) of 100 cattle blood samples obtained from different breeds. A total of 50.0% of the cattle in this study were infected with only Mycoplasma wenyonii, while 2.0% were infected with only Candidatus Mycoplasma haemobos and 17% were infected with both species. Based on sequence analysis of the partial or nearly full length sequences of hemoplasma 16S rRNA gene, the presence of M. wenyonii and Candidatus M. haemobos was confirmed. Hemoplasmapositive cattle of less than three years appeared to have higher infection rate compared to other age groups. M. wenyonii was identified for the first time in approximately 30% of cattle ticks (Rhipicephalus microplus and Haemaphysalis sp.) in this study. This study presents the first molecular evidence of hemoplasmas in Malaysian cattle and ticks.

Molecular Detection of Coxiella burnetii from Farm Animals and Ticks in Malaysia.

Coxiella burnetii, the causative agent of Q fever, is an intracellular bacterium of medical and veterinary importance. The reservoirs of C. burnetii are extensive which include mammals and arthropods, particularly ticks. As the organism is difficult to culture, this study was aimed to detect C. burnetii DNA in animal (mainly blood and vaginal samples of cattle, goats and sheep) and tick samples obtained from farm animals, wild rodents and vegetation. Two polymerase chain reaction (PCR) assays targeting IS1111 transposon-like gene (TransPCR) and com1 gene (OMP-PCR) were used for C. burnetii detection. Sequence determination of the amplified fragments and a real-time PCR assay were used to confirm PCR findings. C. burnetii DNA was detected from 9.1% of cattle blood and 4.2% vaginal samples, respectively. A small percentage (5.8%) of ticks (including Amblyomma, Dermacentor, Rhipicephalus and Haemaphysalis spp.) haboring C. burnetii were identified in this study. This study provides molecular evidence on the presence of C. burnetii in cattle and ticks. The possible zoonotic transmission of C. burnetii is yet to be investigated.

Molecular evidence of hemoplasmas in Malaysian cattle and ticks

Hemotrophic mycoplasma (hemoplasmas) is a cell wall-less bacterium causing infectious anemia in animals. As data on hemoplasmas infecting cattle in Malaysia is scarce, specific polymerase chain reaction assays were used for detection of hemoplasmas from blood samples of cattle and ticks in this study. Hemoplasma DNA was detected in 69 (69.0%) of 100 cattle blood samples obtained from different breeds. A total of 50.0% of the cattle in this study were infected with only Mycoplasma wenyonii, while 2.0% were infected with only Candidatus Mycoplasma haemobos and 17% were infected with both species. Based on sequence analysis of the partial or nearly full length sequences of hemoplasma 16S rRNA gene, the presence of M. wenyonii and Candidatus M. haemobos was confirmed. Hemoplasmapositive cattle of less than three years appeared to have higher infection rate compared to other age groups. M. wenyonii was identified for the first time in approximately 30% of cattle ticks (Rhipicephalus microplus and Haemaphysalis sp.) in this study. This study presents the first molecular evidence of hemoplasmas in Malaysian cattle and ticks.

Whole-genome sequence analysis and exploration of the zoonotic potential of a rat-borne Bartonella elizabethae.

Bartonella elizabethae has been known to cause endocarditis and neuroretinitis in humans. The genomic features and virulence profiles of a B. elizabethae strain (designated as BeUM) isolated from the spleen of a wild rat in Kuala Lumpur, Malaysia are described in this study. The BeUM strain has a genome size of 1,932,479bp and GC content of 38.3%. There is a high degree of conservation between the genomes of strain BeUM with B. elizabethae type strains (ATCC 49927 and F9251) and a rat-borne strain, Re6043vi. Of 2137 gene clusters identified from B. elizabethae strains, 2064 (96.6%) are indicated as the core gene clusters. Comparative genome analysis of B. elizabethae strains reveals virulence genes which are known in other pathogenic Bartonella species, including VirB2-11, vbhB2-B11, VirD4, trw, vapA2-5, hbpA-E, bepA-F, bepH, badA/vomp/brp, ialB, omp43/89 and korA-B. A putative intact prophage has been identified in the strain BeUM, in addition to a 8kb pathogenicity island. The whole genome analysis supports the zoonotic potential of the rodent-borne B. elizabethae, and provides basis for future functional and pathogenicity studies of B. elizabethae.

Identification of rickettsiae from wild rats and cat fleas in Malaysia.

Rickettsioses are emerging zoonotic diseases reported worldwide. In spite of the serological evidence of spotted fever group rickettsioses in febrile patients in Malaysia, limited studies have been conducted to identify the animal reservoirs and vectors of rickettsioses. This study investigated the presence of rickettsiae in the tissue homogenates of 95 wild rats and 589 animal ectoparasites. Using PCR assays targeting the citrate synthase gene (gltA), rickettsial DNA was detected in the tissue homogenates of 13 (13.7%) wild rats. Sequence analysis of the gltA amplicons showed 98.6-100% similarity with those of Rickettsia honei/R. conorii/R. raoultii (Rickettsiales: Rickettsiaceae). Sequence analysis of outer membrane protein A gene (ompA) identified Rickettsia sp. TCM1 strain from two rats. No rickettsia was detected from Laelaps mites, Rhipicephalus sanguineus and Haemaphysalis bispinosa ticks, and Felicola subrostratus lice in this study. R. felis was identified from 32.2% of 177 Ctenocephalides felis fleas. Sequence analysis of the gltA amplicons revealed two genotypes of R. felis (Rf31 and RF2125) in the fleas. As wild rats and cat fleas play an important role in the enzoonotic maintenance of rickettsiae, control of rodent and flea populations may be able to reduce transmission of rickettsioses in the local setting.

Molecular survey and sequence analysis of Anaplasma spp. in cattle and ticks in a Malaysian farm.

This study was conducted to determine the occurrence of Anaplasma spp. in the blood samples of cattle, goats, deer and ticks in a Malaysian farm. Using polymerase chain reaction (PCR) and sequencing approach, Anaplasma spp. was detected from 81(84.4%) of 96 cattle blood samples. All blood samples from 23 goats and 22 deer tested were negative. Based on the analysis of the Anaplasma partial 16S ribosomal RNA gene, four sequence types (genotypes 1 to 4) were identified in this study. Genotypes 1-3 showed high sequence similarity to those of Anaplasma platys/ Anaplasma phagocytophilum, whilst genotype 4 was identical to those of Anaplasma marginale/ Anaplasma centrale/ Anaplasma ovis. Anaplasma DNA was detected from six (5.5%) of 109 ticks which were identified as Rhipicephalus (formely known as Boophilus) microplus ticks collected from the cattle. This study reported for the first time the detection of four Anaplasma sequence types circulating in the cattle population in a farm in Malaysia. The detection of Anaplasma DNA in R. microplus ticks in this study provides evidence that the ticks are one of the potential vectors for transmission of anaplasmosis in the cattle.

Molecular survey and sequence analysis of Anaplasma spp. in cattle and ticks in a Malaysian farm

This study was conducted to determine the occurrence of Anaplasma spp. in the blood samples of cattle, goats, deer and ticks in a Malaysian farm. Using polymerase chain reaction (PCR) and sequencing approach, Anaplasma spp. was detected from 81(84.4%) of 96 cattle blood samples. All blood samples from 23 goats and 22 deer tested were negative. Based on the analysis of the Anaplasma partial 16S ribosomal RNA gene, four sequence types (genotypes 1 to 4) were identified in this study. Genotypes 1-3 showed high sequence similarity to those of Anaplasma platys/ Anaplasma phagocytophilum, whilst genotype 4 was identical to those of Anaplasma marginale/ Anaplasma centrale/ Anaplasma ovis. Anaplasma DNA was detected from six (5.5%) of 109 ticks which were identified as Rhipicephalus (formely known as Boophilus) microplus ticks collected from the cattle. This study reported for the first time the detection of four Anaplasma sequence types circulating in the cattle population in a farm in Malaysia. The detection of Anaplasma DNA in R. microplus ticks in this study provides evidence that the ticks are one of the potential vectors for transmission of anaplasmosis in the cattle.