Computational protocol: The Genotypic Structure of a Multi-Host Bumblebee Parasite Suggests a Role for Ecological Niche Overlap

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Protocol publication

[…] We first analyzed our data for the set of singly infected bees alone. In a second step, all infections were included, i.e. including the reconstructed strains retrieved from multiple infections as identified by the above algorithm. The overall population genetic structure was calculated using the software Genepop . Pairwise FST–values and their significance levels were calculated using Arlequin 3.1 . Each directly identified or reconstructed multi-locus genotype was entered as one independent data point; populations with sample sizes smaller than 5 were not included in each of these analyses, and those with sample sizes lower than 10 are marked in italics. Locus 1.B6 was not included in the final analysis at all due to ambiguities in genotyping on the ABI Prism Fragment Analyzer (scattered ranges instead of clear peaks). We also tested for linkage disequilibrium according to the method of Black and Kraftsur implemented in the software GENETIX .In a further step, C. bombi populations for each site where bees had been sampled for consecutive years were pooled together into one data set for that given site. The resulting three parasite populations in the region Basler Jura (Röschenz, Movelier, Soyhières) and four parasite populations in the Lower Engadin (La Munt, Buffalora, Lavin, Stabelchod), respectively, were analyzed with respect to their phylo-geography using allele frequencies. Relationships were based on Cavalli-Sforza's and Edwards' chord distance DC . According to Takezaki and Nei , this distance is superior to other measures because it explicitly considers the stepwise nature of microsatellite mutation. Allele frequencies were bootstrapped 1'000 times, using the subprogram SEQBOOT within the program package PHYLIP 3.1. ; the consensus tree was assembled in NEIGBOR and CONSENSUS, and finally visualized in TREEVIEW .At last, G∶N-ratios (number of different multi-locus genotypes, G, over sample size, N) were calculated as a measure of genetic diversity and clonality of C. bombi according to Ivey and Richards . Calculations were done for all successfully typed multi-locus (4 loci) genotypes. Note that diversity at these four loci was high enough for the resolution to be appropriate for our analyses. […]

Pipeline specifications

Software tools Genepop, Arlequin, PHYLIP, TreeViewX
Applications Phylogenetics, Population genetic analysis
Organisms Toxoplasma gondii
Diseases Infection