Environmental DNA analysis does not detect the endangered Rio Concepcion topminnow, Poeciliopsis jackschultzi.

Authors: Alanna Fulkerson, Alejandro Varela-Romero, Alexsandre Gutierrez-Barragán, Doug Duncan, Mariana Mateos

Year: 2025

Abstract

Like their counterparts in other regions of the arid Northwestern America, freshwater fishes of the US-Mexico borderland rivers Rio De La Concepcion and Rio Santa Cruz (a tributary of the Gila River) are imperiled, due to habitat loss/degradation and invasive species. The primary goal of this study was to evaluate the status of the Rio Concepcion Topminnow, Poeciliopsis jackschultzi (Conway, Mateos, Vrijenhoek 2019), an endangered cienega-specialist and microendemic that historically occupied ca. 30 km stretch of the Rio De La Concepcion, just south of the Arizona-Sonora border. Survey efforts are hampered by the inability to use external morphology to distinguish P. jackschultzi from its co-occurring congeners: the Gila Topminnow, Poeciliopsis occidentalis; and asexual hybrids whose maternal progenitor (and thus mitochondrial genome) is the Headwater topminnow, Poeciliopsis monacha. We surveyed the condition of freshwater habitats, and used environmental (e)DNA analyses (metabarcoding with the “MiFish” primer set, which target a 163-185 bp region of the mitochondrial 12S rRNA gene of teleosts) to document the fish taxa present (native and non-native), within the known range of P. jackschultzi, and in nearby localities and tributaries, over two collection periods (May 2022 and 2023). A secondary goal was to document fish taxa present in the Mexican side of the Rio Santa Cruz, upstream from where it re-enters the US (east of Nogales) and from where the Poeciliopsis asexual hybrid (considered non-native) was recently recorded. Our analyses, based on >500 million Illumina sequence reads, recovered 2925 unique sequences (i.e. features), of which 2774 were assigned to teleosts. Unfortunately, we did not detect a single feature assigned to P. jackschultzi, suggesting that it was absent or at an abundance below our detection ability. Multiple sites that used to harbor cienega and other freshwater fish habitat were completely dry during our surveys. We recommend future surveys during several seasons (including periods with more water) to continue searching for extant P. jackschultzi. Across all sites surveyed, the most frequent eDNA features were assigned to the native leuciscids Longfin dace, Agosia chrysogaster and Sonoran Chub, Gila ditaenia. Features assigned to P. monacha mitochondrion (i.e. asexual hybrid) and P. occidentalis had the fifth and sixth highest frequency. The following previously documented non-native taxa were also detected with eDNA: Green sunfish, Lepomis cyanellus (third highest eDNA frequency); Tilapia (fourth highest frequency); Black Bullhead, Ameiurus melas; Western mosquitofish, Gambusia affinis; Bluegill, Lepomis macrochirus; and Largemouth Bass, Micropterus salmoides. The Green Swordtail, Xiphophorous helleri, appears to reflect a more recent (< ~20 years) introduction around the town of Imuris, Sonora. We document the likely extirpation of Poeciliopsis spp. from most of the Cocospera tributary (i.e. upstream of the dam El Comaquito). We discuss the strengths and weaknesses of the eDNA metabarcoding method, including a small number of false negatives (i.e., failure to detect taxa whose presence we visually detected), and false positive detections. At the single site sampled on the Mexican side of the Santa Cruz (Parque Mascareñas), we confirmed the presence of the natives Gila Topminnow and Longfin Dace, and of the putative non-native Poeciliopsis asexual hybrid, which likely disperses downstream into US sites during high precipitation periods (e.g. during Monsoon season). In addition, we detected eDNA from a sucker and a chub. Surprisingly, rather than matching the sequences of the respective native Santa Cruz taxa (i.e., Catostomus clarkii, Catostomus insignis, and the Gila robusta complex), the sucker and chub eDNA features, respectively, perfectly matched reference sequences from taxa native to the Rio Yaqui: Yaqui Sucker, Catostomus bernardini and Mexican Roundtail Chub, Gila minacae. We hypothesize that human introductions from the Yaqui to the Santa Cruz have occurred.