Cryptobenthic Coral Reef Fishes: Resolving Critical Gaps in Species Delimitation, Sampling Techniques, and Phylogenetic relationships

Abstract

Cryptobenthic reef fishes (CRF) are the smallest vertebrates on coral reefs but represent about 40% of the fish species and about 50% of fish abundance in coral reef ecosystems. Their diversity can be explained by their extremely limited dispersal abilities and short generation times (promoting allopatric speciation) coupled with their ability to partition microhabitats at a very fine scale. One of their main contributions to coral reef ecosystems is as food for larger piscivorous fishes and invertebrates. Despite their importance, CRF are often overlooked, as their cryptic nature and very small size make them difficult to assess using visual methods. The prevailing method for quantifying communities of CRF is to use enclosures and anesthetics to collect fishes from within the reef. Environmental DNA (eDNA) sampling of water around reefs may be an alternative or supplemental way to quantify CRF communities. In Chapter 1, I compared the ability to characterize CRF communities using eDNA sequences from water samples collected from within the interstices of the reef, with corresponding anesthetic stations from the same microhabitat. A total of 676 CRF from 33 species were collected from anesthetic stations, whereas only four species of CRF were detected from the eDNA samples taken at the same locations. Main factors contributing to these results may have been: (1) low standing biomass of CRF, thus low abundance of detectable DNA in the eDNA water samples; (2) large number of non-targeted metazoan reads that “drowned” the detections of specific CRF sequences; and (3) lack of sequences available in public databases that represent the high level of endemism present in CRF and a need for more complete taxonomic inventory. For some groups of CRF, their small size, cryptic nature, and conserved morphology have resulted in many undetected cryptic species, which may require a genome-wide species delimitation approach to discern how many species are present. One of the most species-rich groups of CRF, the genus Eviota, has 132 species described to date, is widely distributed from the Red Sea to Hawaii and French Polynesia, and is known to comprise numerous cryptic species. In Chapter 2, I focused on the Eviota sigillata complex which is represented by two nominal species described by morphological characters, yet preliminary genetic data suggest the presence of multiple cryptic lineages. I use molecular data from mitochondrial DNA and genome-wide SNP data generated via double digest restriction site associated sequencing (ddRADseq), in combination with morphological data to infer the number of species in the E. sigillata complex. Specifically, I constructed phylogenetic trees and conducted several types of single-locus and multilocus species delimitation analyses and compared these to groupings based on morphology, as well as their geographic distribution. Overall, I found evidence for the presence of 9-13 lineages within the E. sigillata species complex, with genetic lineages corresponding well with the biogeography of the group. I further confirmed that the original morphological diagnostic characters used for the separation of the two nominal species were not useful for distinguishing each of the nine clades in the complex but may be helpful in diagnosing groups of species. Overall, Chapter 2 shed light onto the patterns of speciation within CRF and provided a glimpse of the tremendous hidden diversity that remains in coral reef fishes. The goby genera Eviota and Sueviota (family Gobiidae) are commonly known as dwarfgobies, and collectively, the two genera are among the most abundant and diverse groups of fishes on coral reefs. Despite the diversity, abundance, and ecological importance of this group, and the large and growing number of species described to date (132 Eviota, 9 Sueviota), there is a lack of understanding of the phylogenetic relationships among the major clades of Eviota, poor knowledge of the relationships between dwarfgobies and other Gobiidae species, and no information on the placement of Sueviota. In addition, as is the case with most small reef fishes, a clear understanding of taxonomically informative phenotypic characters is also lacking. In Chapter 3, to resolve the evolutionary history of dwarfgobies, I inferred a time-calibrated phylogeny of the group using genome-wide data from 440 ddRADseq loci captured across 98 Eviota and Sueviota specimens, plus 66 specimens of other related gobies. I also assessed the distribution of 14 external and osteological morphological characters across the tree to assess which may be useful for diagnosing clades. Results from my Chapter 3 robustly established the non-monophyly of the genus Eviota, which was resolved into two separate clades, both of which were resolved within a lineage of other coral associated genera (Gobiodon, Paragobiodon, Pleurosicya, Minisicya and Bryaninops). One of the two clades is herein elevated to its own genus, Eviotops, a name which was previously considered synonymous with Eviota. Additionally, I established that the genus Sueviota is deeply nested within one of the Eviota clades and is herein synonymized with Eviota. Furthermore, I found strong phylogenetic signals for 12 out of the 14 phenotypic traits examined, providing strong complementary support for the two recovered clades and establishing the validity of phenotypic traits that strongly correspond with genetic groupings that should aid in future taxonomic studies for this group.