Development of twenty-four microsatellite markers for Afrotropical Ornithodoros ticks
Data files
Feb 02, 2024 version files 6.53 KB
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Genotyping.zip
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README.md
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Abstract
Background: Soft ticks of the genus Ornithodoros are responsible for the maintenance and transmission of the African swine fever (ASF) virus in the sylvatic and domestic viral cycles in Southern Africa. They are also the main vectors of Borrelia species causing relapsing fevers. Currently, no genetic markers are available for Afrotropical Ornithodoros ticks. As ASF spreads globally, such markers are needed to assess the role of ticks in the emergence of new outbreaks. The aim of this study was to design microsatellite markers that could be used for ticks of the Ornithodoros moubata complex, particularly Ornithodoros phacochoerus, to assess population structure and tick movements in ASF endemic areas.
Methods: One hundred and fifty-one markers were designed using the O. moubata and O. porcinus genomes after elimination of repeated sequences in the genomes. All designed markers were tested on O. phacochoerus and O. porcinus DNA to select the best markers.
Results: Twenty-four microsatellite markers were genotyped on two populations of O. phacochoerus and on few individuals from four other Ornithodoros species. Nineteen markers were selected to be as robust as possible for population genetic studies on O. phacochoerus.
Conclusions: The microsatellite markers developed here represent the first genetic tool to study nidicolous populations of Afrotropical Ornithodoros. This dataset contains the genotyping results obtained for all twenty-four markers tested.
Summary
The three datasheets made available here result from the genotyping of twenty-four microsatellite loci on five Ornithodoros species.\
These datasheets can be transformed (using CREATE for instance) to be used in any population genetics software (Fstat, Genepop, Micro-Checker…).
This dataset was used to test the robustness of twenty-four microsatellite loci for the species Ornithodoros phacochoerus. The microsatellite loci were also tested on four other Ornithodoros species (O. moubata, O. porcinus, O. maritimus, and O. erraticus) to assess specificity.
Description of the data and file structure
This dataset contains three different datasheets of genotyping results. Two of the datasheets present genotyping data from the species O. phacochoerus. Ornithodoros phacochoerus ticks came from two different sites: Coutada9 Game Reserve (pop 69) and Gorongosa National Park (pop G3) in Mozambique.
The file names indicate which data are contained in the datasheet:
- “Plex1-2-3_pop69etG3_19markers” contains the genotyping results for O. phacochoerus species (pop69etG3) from the 19 microsatellite loci that were selected to be as robust as possible. (Number of ticks tested n=58, 29 individuals per site)
- “Plex1-2-3_pop69etG3_5markersDiscarded” contains the genotyping results for O. phacochoerus species (pop69etG3) from the 5 microsatellite loci that were discarded due to lack of robustness in this species. (Number of ticks tested n=58, 29 individuals per site)
- “Plex1-2-3_FourOtherSpecies_ 24markers” contains the genotyping results for O. moubata, O. porcinus, O. maritimus, and O. erraticus species from all the 24 microsatellite loci. (Number of ticks tested n=8, 2 individual per species tested)
Here is the detail of how the information were collected/registered/transformed for each variable:
“Pop“: Site number for O. phacochoerus ticks (69 or G3). Was replaced by the species name for the four other species.
“SampleName“: Individual unique identifier for each of the ticks.
“NumPop“: Number associated with the “Pop” column for easier use in population genetics softwares.
“Parc“: Name of the game reserve or national park from which O. phacochoerus ticks were collected. Was replaced by “AutreEspece” (OtherSpecies) for the four other species.
“ms35“, “ms48”, “ms59” etc…: Names of microsatellite loci that were tested. For each microsatellite locus, there is two columns, one for each allele detected in the tick. Alleles are named according to their size in base pair (for instance, individual 69_1 presents two alleles for microsatellite locus ms35, one allele which is 67 base pair long and one which is 71 base pair long)
Empty cells: Empty cells were not filled with “null,” or “n/a,” due to interference with population genetics software.
Empty cells occur either after no data was obtained for the genotyping of a tick at a microsatellite locus, or if the data were of low quality and did not allow correct reading of the peaks.
Data files are in .txt format with tabulations as separators, they can be visualized using any software capable of handling .txt files (Excel, LibreOffice Calc, TextPad etc…).
Sharing/Access information
This dataset is associated with a paper entitled “Development of microsatellite markers for Afrotropical Ornithodoros ticks” submitted for peer-reviewing (no DOI available yet, DOI will be provided as soon as possible).
Software
The authors used this dataset by converting it to Fstat and Micro-Checker format using CREATE (Coombs, Letcher, and Nislow 2008). Fstat v 2.9.4. was used to calculate Linkage disequilibrium p-values, observed heterozygosity, expected heterozygosity, Fis, and Fst. Micro-Checker (Van Oosterhout, Hutchinson, Wills, and Shipley 2004) was used to test for presence of null alleles, stuttering and short allele dominance.
This dataset results from genotyping of twenty-four microsatellite loci on Ornithodoros phacochoerus ticks and on four other Ornithodoros species.
Fluorescent-labeled forward primers (FAM, VIC, NED or PET) were used for the 24 microsatellite loci. Touchdown PCRs were performed in six multiplexes of four microsatellite loci each.
The amplification mix consisted in 2 μL of DNA template, 10 μL of 2x Type-it Microsatellite PCR Kit (Qiagen, Courtaboeuf, France), adjusted volume of fluorescence-labeled forward primer, and reverse primer for microsatellite loci, in a final volume of 20 μL. The touchdown PCR program was set as follows: 95°C for 3 min, then 10 cycles of 95°C for 20 sec, 60°C -0.5°C/cycle, for 30 sec, and 72°C for 1 min, then 30 cycles of 95°C for 20 sec, 55°C for 30 sec, and 72°C for 1 min, followed by a final extension step at 72°C for 7 min.
Formamide for denaturation and GeneScan-600 (LIZ) Size Standard Kit for ladder were added to the PCR products before genotyping by capillary electrophoresis at the GPTR laboratory (Great Regional Technical Platform of genotyping, AGAP Institut/CIRAD, Montpellier, France) with an ABI 3500xL Genetic Analyzer (Applied Biosystems, Foster City, CA, USA).
Genotypes were read using GeneMapper® v.6 software (Applied Biosystems, Waltham, MA, USA). Allele bins were set manually after a review of all samples. Allele scoring was performed automatically according to the bin set designed for the marker, then manually checked by two different experimenters. Alleles were named according to their length in base pairs.