Development, characterization, and cross-amplification of polymorphic microsatellite markers for North American Trachymyrmex and Mycetomoellerius ants

Abstract

Objective: The objective of this study is to develop and identify polymorphic microsatellite markers for fungus-gardening (attine) ants in the genus Trachymyrmex sensu lato. These ants are important ecosystem engineers and have been a model group for understanding complex symbiotic systems, but very little is understood about the intraspecific genetic patterns across most North American attine species. These microsatellite markers will help to better study intraspecific population genetic structure, gene flow, mating habits, and phylogeographic patterns in these species and potentially other congeners.Results: Using next-generation sequencing techniques, we identified 17 and 12 polymorphic microsatellite markers from T. septentrionalis and Mycetomoellerius (formerly Trachymyrmex) turrifex, respectively, and assessed the genetic diversity of each marker. We also analyzed the cross-amplification success of the T. septentrionalis markers in two other closely related Trachymyrmex species, and identified 10 and 12 polymorphic markers for T. arizonensis and T. pomonae, respectively.

Male‐biased dispersal in a fungus‐gardening ant symbiosis

“…Two primary advantages of mtDNA sequences are that (a) sequences can be readily obtained and (b) the problems associated with mtDNA are understood and can be easily examined, whereas the problems associated with nuclear markers are more uncertain (Bowen et al., 2014; Moreau, 2009). Moreover, a preliminary examination of the microsatellite markers of T. septentrionalis used in this study supported one of the main findings of mtDNA‐based studies in this species: pronounced genetic differentiation across the Mississippi River Valley (Matthews et al., 2020), thus making it unlikely that the COI sequences analyzed were numts, which usually lack variation because of purifying selection in nuclear genomes (Martins et al., 2007). Therefore, mtDNA sequences in this species are likely robust genetic tools (Matthews et al., 2020; Mikheyev et al., 2008; Seal et al., 2015).…”

“…For example, haplotypes are represented by phylogroups east and west of the Mississippi River, with western populations consisting of one clade whereas those found in the east are more diverse, consisting of at least three clades (Mikheyev et al., 2008; Seal et al., 2015). A preliminary analysis of nuclear microsatellites supports this overall pattern—higher diversity in populations east of the Mississippi River relative to the west (Matthews et al., 2020). Trachymyrmex septentrionalis is one of the most common and abundant ant species in the longleaf pine forests of Florida whose populations appear to respond rapidly to annual variations in climate (especially rainfall) and local conditions (e.g., relief) and often move significant amounts of soil in the process (Seal & Tschinkel, 2006, 2008, 2010; Tschinkel & Seal, 2016).…”

“…We extracted genomic DNA for microsatellite genotyping from 64 individual T. septentrionalis workers using a QIAamp DNA Micro Kit (QIAGEN). We implemented the M13‐tail polymerase chain reaction (PCR) method (Schuelke, 2000), which involves three primers in the PCR: forward primers (Matthews et al., 2020) with an M13‐tail at the 5’ end, an unlabeled reverse primer, and a universal M13 primer labeled with 6‐FAM (6‐carboxy‐fluorescine) fluorescent dye. These were used to amplify nine microsatellite loci (Ts21, Ts25, Ts32, Ts33, Ts39, Ts41, Ts43, Ts46, Ts5) (Matthews et al., 2020).…”

“…Surprisingly, there have been very few studies that examine sex‐biased dispersal and gene flow in fungus‐gardening ants, even though the molecular markers necessary to do so have been developed. For instance, microsatellite markers have been used to examine polyandry, polygyny, and population structure (Bekkevold et al., 1999; Fjerdingstad & Boomsma, 2000; Helmkampf et al., 2008; Kellner et al., 2013; Matthews et al., 2020; Murakami et al., 2000; Rabeling et al., 2011, 2013, 2014) and mitochondrial DNA (mtDNA) markers have been used to examine phylogeographic structure (i.e., female dispersal patterns) in several attine species (Cardoso et al., 2015; Seal et al., 2015; Solomon et al., 2008). While at least one study reported comparable dispersal abilities in both male and female Atta colombica , only a small portion of its known range was surveyed (Helmkampf et al., 2008).…”

“…In this study, we employ mtDNA markers and diploid microsatellite markers that were developed using whole genome sequencing (Matthews et al., 2020) to examine population structure, gene flow, and signatures of sex‐biased dispersal in a fungus‐gardening ant species, Trachymyrmex septentrionalis (McCook, 1881). Trachymyrmex septentrionalis is the only fungus‐gardening ant found entirely within the United States and is distributed from Texas to Florida to Long Island, New York, with a center of genetic diversity found in northern Florida (Matthews et al., 2020; Rabeling et al., 2007; Seal et al., 2015; Senula et al., 2019). Although northern Florida is near the southern edge of the distribution of T. septentrionalis , the population does not appear to exhibit traits of a peripheral population (Mueller et al., 2011; Sexton et al., 2009).…”