Dataset Information

Molecular Detection and Genetic Characterization of Ehrlichia ruminantium Harbored by Amblyomma hebraeum Ticks of Domestic Ruminants in North West Province, South Africa.

ABSTRACT: Ehrlichia ruminantium (E. ruminantium) is the causative agent of heartwater disease and it is mainly transmitted to livestock by Amblyomma hebraeum (A. hebraeum) tick in South Africa. This study investigated the occurrence of E. ruminantium and its genetic diversity in ticks within Ngaka Modiri Molema district of North West Province in South Africa. Genomic DNA was extracted from whole A. hebraeum ticks totaling 876 and resulted in a total of 292 pooled DNA samples. Firstly, conventional PCR was used to detect Ehrlichia spp. targeting the dsbA gene, followed by nested PCR targeting the Map1 gene performed on DNA pool samples that were positive from the first PCR. One hundred and six tick DNA pool samples were positive by dsbA gene PCR for the presence of Ehrlichia spp. with minimum infection rate (MIR) of 121, while 13/106 were positive by Map1 PCR with MIR of 15. Different E. ruminantium Map1 genotypes (NWUe1, NWUe2, NWUe3, NWUe5, and NWUe6) were detected from tick samples and were closely related to more than 13 gene sequences of E. ruminantium from the NCBI GenBank database. These findings suggest that there is a significant diversity of E. ruminantium infecting ticks in the study area.

SUBMITTER: Mnisi SS

PROVIDER: S-EPMC9559644 | biostudies-literature | 2022 Sep

REPOSITORIES: biostudies-literature

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Molecular Detection and Genetic Characterization of Ehrlichia ruminantium Harbored by Amblyomma hebraeum Ticks of Domestic Ruminants in North West Province, South Africa.

Mnisi Sifiso S SS Mphuthi Malekoba B N MBN Ramatla Tsepo T Mofokeng Lehlohonolo S LS Thekisoe Oriel O Syakalima Michelo M

Animals : an open access journal from MDPI 20220921 19

Ehrlichia ruminantium (E. ruminantium) is the causative agent of heartwater disease and it is mainly transmitted to livestock by Amblyomma hebraeum (A. hebraeum) tick in South Africa. This study investigated the occurrence of E. ruminantium and its genetic diversity in ticks within Ngaka Modiri Molema district of North West Province in South Africa. Genomic DNA was extracted from whole A. hebraeum ticks totaling 876 and resulted in a total of 292 pooled ...[more]

PMID: 36230251

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Project description:BackgroundEhrlichia ruminantium is the causal agent of heartwater, a fatal tropical disease affecting ruminants with important economic impacts. This bacterium is transmitted by Amblyomma ticks and is present in sub-Saharan Africa, islands in the Indian Ocean and the Caribbean, where it represents a threat to the American mainland.MethodsAn automated DNA extraction method was adapted for Amblyomma ticks and a new qPCR targeting the pCS20 region was developed to improve E. ruminantium screening capacity and diagnosis. The first step in the preparation of tick samples, before extraction, was not automated but was considerably improved by using a Tissue Lyser. The new pCS20 Sol1 qPCR and a previously published pCS20 Cow qPCR were evaluated with the OIE standard pCS20 nested PCR.ResultspCS20 Sol1 qPCR was found to be more specific than the nested PCR, with a 5-fold increase in sensitivity (3 copies/reaction vs 15 copies/reaction), was less prone to contamination and less time-consuming. As pCS20 Sol1 qPCR did not detect Rickettsia, Anasplasma and Babesia species or closely related species such as Panola Mountain Ehrlichia, E. chaffeensis and E. canis, its specificity was also better than Cow qPCR. In parallel, a tick 16S qPCR was developed for the quality control of DNA extraction that confirmed the good reproducibility of the automated extraction. The whole method, including the automated DNA extraction and pCS20 Sol1 qPCR, was shown to be sensitive, specific and highly reproducible with the same limit of detection as the combined manual DNA extraction and nested PCR, i.e. 6 copies/reaction. Finally, 96 samples can be tested in one day compared to the four days required for manual DNA extraction and nested PCR.ConclusionsThe adaptation of an automated DNA extraction using a DNA/RNA viral extraction kit for tick samples and the development of a new qPCR increased the accuracy of E. ruminantium epidemiological studies, as well as the diagnostic capabilities and turn-over time for surveillance of heartwater. This new method paves the way for large-scale screening of other bacteria and viruses in ticks as well as genetic characterization of ticks and tick-pathogen coevolution studies.

Project description:Rhipicephalus microplus ticks are vectors for multiple pathogens infecting animals and humans. Although the medical importance of R. microplus has been well-recognized and studied in most areas of China, the occurrence of tick-borne Rickettsiales has seldom been investigated in Guizhou Province, Southwest China. In this study, we collected 276 R. microplus ticks from cattle (209 ticks) and goats (67 ticks) in three locations of Guizhou Province. The Rickettsia, Anaplasma, and Ehrlichia were detected by targeting the 16S rRNA gene and were further characterized by amplifying the key genes. One Rickettsia (Ca. Rickettsia jingxinensis), three Ehrlichia (E. canis, E. minasensis, Ehrlichia sp.), and four Anaplasma (A. capra, A. ovis, A. marginale, Ca. Anaplasma boleense) species were detected, and their gltA and groEL genes were recovered. Candidatus Rickettsia jingxinensis, a spotted fever group of Rickettsia, was detected in a high proportion of the tested ticks (88.89%, 100%, and 100% in ticks from the three locations, respectively), suggesting the possibility that animals may be exposed to this type of Rickettsia. All the 16S, gltA, groEL, and ompA sequences of these strains are 100% identical to strains reported in Ngawa, Sichuan Province. E. minasensis, A. marginale, and Candidatus Anaplasma boleense are known to infect livestock such as cattle. The potential effects on local husbandry should be considered. Notably, E. canis, A. ovis, and A. capra have been reported to infect humans. The relatively high positive rates in Qianxinan (20.99%, 9.88%, and 4.94%, respectively) may indicate the potential risk to local populations. Furthermore, the genetic analysis indicated that the E. minasensis strains in this study may represent a variant or recombinant. Our results indicated the extensive diversity of Rickettsiales in R. microplus ticks from Guizhou Province. The possible occurrence of rickettsiosis, ehrlichiosis, and anaplasmosis in humans and domestic animals in this area should be further considered and investigated.

Project description:The Lone Star tick, Amblyomma americanum, transmits several bacterial pathogens including species of Anaplasma and Ehrlichia. Amblyomma americanum also hosts a number of non-pathogenic bacterial endosymbionts. Recent studies of other arthropod and insect vectors have documented that commensal microflora can influence transmission of vector-borne pathogens; however, little is known about tick microbiomes and their possible influence on tick-borne diseases. Our objective was to compare bacterial communities associated with A. americanum, comparing Anaplasma/Ehrlichia -infected and uninfected ticks. Field-collected questing specimens (n = 50) were used in the analyses, of which 17 were identified as Anaplasma/Ehrlichia infected based on PCR amplification and sequencing of groEL genes. Bacterial communities from each specimen were characterized using Illumina sequencing of 16S rRNA gene amplicon libraries. There was a broad range in diversity between samples, with inverse Simpson's Diversity indices ranging from 1.28-89.5. There were no statistical differences in the overall microbial community structure between PCR diagnosed Anaplasma/Ehrlichia-positive and negative ticks, but there were differences based on collection method (P < 0.05), collection site (P < 0.05), and sex (P < 0.1) suggesting that environmental factors may structure A. americanum microbiomes. Interestingly, there was not always agreement between Illumina sequencing and PCR diagnostics: Ehrlichia was identified in 16S rRNA gene libraries from three PCR-negative specimens; conversely, Ehrlichia was not found in libraries of six PCR-positive ticks. Illumina sequencing also helped identify co-infections, for example, one specimen had both Ehrlichia and Anaplasma. Other taxa of interest in these specimens included Coxiella, Borrelia, and Rickettsia. Identification of bacterial community differences between specimens of a single tick species from a single geographical site indicates that intra-species differences in microbiomes were not due solely to pathogen presence/absence, but may be also driven by vector life history factors, including environment, life stage, population structure, and host choice.

Dataset Information

Molecular Detection and Genetic Characterization of Ehrlichia ruminantium Harbored by Amblyomma hebraeum Ticks of Domestic Ruminants in North West Province, South Africa.

Publications

Molecular Detection and Genetic Characterization of <i>Ehrlichia ruminantium</i> Harbored by <i>Amblyomma hebraeum</i> Ticks of Domestic Ruminants in North West Province, South Africa.

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