In the replicates for which the pre-bottleneck laboratory-cultured populations did not significantly depart from a mutation/drift equilibrium, an important assumption of the program Bottleneck, only a portion of the bottleneck events were detected.
The program Bottleneck reported a significant excess of heterozygosity in only one (Pinta; P = 0.027) of the seven populations; however, sample sizes may have been too small to provide statistical resolution.
The program Bottleneck showed a heterozygosity excess in one of the Chilean populations (R. Bonito, P = 0.025), characterized by negative FIS values (FIS = −0.015), while population expansion was only detected in one Falkland population (Sarnys Creek, P = 0.022).
While several methods are designed to estimate the effective size of a population or its fluctuation over time, the program Bottleneck is specifically aimed at detecting a recent bottleneck (within the past 2Ne-4Ne generations).
To see if we could detect signatures of a population bottleneck, we used the program BOTTLENECK (Piry et al. 1999), which tests for heterozygote excess as compared with that expected under mutation drift equilibrium.
For each generated data set, using the program Bottleneck version 1.2.02, we performed a Wilcoxon sign-rank test to determine whether a significant number of loci featured a heterozygosity excess, which is indicative of a recent botteneck event, assuming a two-phase mutation model (TPM; model of microsatellite mutation; [20]).
The tests in program bottleneck are based on the expectation that a population bottleneck will lead to a rapid loss of rare alleles producing an excess of heterozygotes and a shift in allele frequency proportions [ 63].
The data were also analyzed with the program Bottleneck 33 to determine whether there was a significant excess of heterozygotes, which is typical of populations growing in size after a reduction in their numbers.
To investigate whether populations bear the signature of a recent fluctuation in population size, we investigated heterozygosity deficiency and heterozygosity excess with respect to gene diversity using the program BOTTLENECK v. 1.2.02 with 1,000 replications of a two-phase mutation model [ 59].
A recent review by Peery and others 46 suggests that the program Bottleneck can give spurious results when sample sizes are small, and that results can be affected by the duration and time since the bottleneck event.
We tested for a signal of a genetic bottleneck in each of the three time periods using the program bottleneck[ 62].
