Response and resilience of karst subterranean estuary communities to precipitation impacts.

The impact of meteorological phenomena on ecosystem communities of karst subterranean estuaries (KSEs) remains unknown. KSEs are characterized by vertically stratified groundwater separated by a halocline and host endemic aquatic cave-adapted fauna (stygobionts). In October 2015, 8 days of heavy precipitation caused the first recorded mortality event in the KSE. This event was marked by a halocline shift 5 m deeper. The present study aimed to provide insights into resilience of KSEs faunal communities to temporal shifts in temperature and precipitation. Cave water temperature decreased on average 0.0068°C per mm of accumulated precipitation over 4 days, which can add up to, and surpass, the interannual temperature variation in cases of heavy precipitations. Biological surveys (2012-2021) conducted within cave systems El Aerolito and La Quebrada, in Cozumel, indicated that change in community structure was not detected and stygobionts were resilient; however, marine species inhabiting the caves were impacted. Overall, the faunal community at KSEs remains resilient within short-term meteorological phenomena despite shifts of non-stygobionts.

El impacto de fenómenos meteorológicos en las comunidades de los ecosistemas de estuarios subterráneos kársticos (KSE, por sus siglas en inglés) sigue siendo desconocido. Los KSE se caracterizan por aguas subterráneas estratificadas verticalmente separadas por una haloclina, y albergan fauna acuática endémica adaptada a la vida en cuevas (estigobiontes). En octubre de 2015, ocho días de fuertes precipitaciones causaron el primer evento de mortandad registrado en un KSE. Este evento estuvo marcado por un desplazamiento de la haloclina, resultando en una haloclina 5 m más profunda. El presente estudio tuvo como objetivo proveer información sobre la resiliencia de las comunidades faunísticas de los KSE a cambios temporales en la temperatura y la precipitación. La temperatura del agua de la cueva disminuyó en promedio 0.0068°C por mm de precipitación acumulado durante cuatro días, lo cual puede alcanzar, e incluso superar, la variación interanual de la temperatura debido a fuertes precipitaciones. Los estudios biológicos (2012­2021) realizados dentro de los sistemas de cuevas El Aerolito y La Quebrada, en Cozumel, indicaron que no se detectó un cambio en la estructura de la comunidad y que los estigobiontes fueron resilientes, sin embargo, las especies marinas que habitan las cuevas se vieron afectadas. En general, la comunidad de fauna de KSEs se mantiene resiliente a los fenómenos meteorológicos a corto plazo a pesar de los cambios en la fauna no estigobia.

An integrative re-evaluation of Typhlatya shrimp within the karst aquifer of the Yucatán Peninsula, Mexico.

The Yucatán Peninsula, Mexico is a carbonate platform well-known for extensive karst networks of densely stratified aquifer ecosystems. This aquifer supports diverse anchialine fauna, including species of the globally distributed anchialine shrimp genus Typhlatya (Atyidae). Four species (T. campecheae, T. pearsei, T. dzilamensis and T. mitchelli) are endemic to the Peninsula, of which three are federally listed in Mexico. This first integrative evaluation (i.e., molecular, morphological, broad geographic and type locality sampling, and environmental data) of Yucatán Typhlatya reveals considerable species identity conflict in prior phylogenetic assessments, broad species ranges, syntopy within cave systems and five genetic lineages (of which two are new to science). Despite sampling from the type locality of endangered T. campecheae, specimens (and molecular data) were indistinguishable from vulnerable T. pearsei. Ancestral/divergence reconstructions support convergent evolution of a low-salinity ancestor for a post-Paleogene arc Yucatán + Cuba Typhlatya clade within the anchialine Atyidae clade. A secondary adaptation for the coastal-restricted euryhaline (2-37 psu), Typhlatya dzilamensis (unknown conservation status) was identified, while remaining species lineages were low-salinity (< 5 psu) adapted and found within the meteoric lens of inland and coastal caves. This study demonstrates the need for integrative/interdisciplinary approaches when conducting biodiversity assessments in complex and poorly studied aquifers.

Ideating iDNA: Lessons and limitations from leeches in legacy collections.

Indirect methods for conducting faunal inventories present great promise, and genomic inventories derived from environmental sources (eDNA) are improving. Invertebrate ingested DNA (iDNA) from terrestrial leeches in the family Haemadipsidae has shown potential for surveying vertebrates and biodiversity monitoring in protected areas. Here we present an initial, and critical, evaluation of the limitations and biases of current iDNA protocols for biodiversity monitoring using both standard and NGS barcoding approaches. Key findings include the need for taxon relevant multi-locus markers and reference databases. In particular, the limitations of available reference databases have profound potential to mislead and bias eDNA and iDNA results if not critically interpreted. Nevertheless, there is great potential for recovery of amplifiable DNA from gut contents of invertebrate museum specimens which may reveal both temporal patterns and cryptic diversity in protected areas with increased efficiency. Our analyses of ingested DNA (iDNA) from both freshly stored and previously collected (legacy) samples of terrestrial leeches successfully identified vertebrates from Myanmar, Australia and Madagascar and indicate the potential to characterize microbial communities, pathogen diversity and interactions at low cost.

Phylogeny and systematics of Aphroditiformia.

Aphroditiformia represents one of the most successful radiations of annelids, and is therefore an interesting model to understand morphological and functional evolution. Previous phylogenetic analyses yielded most families as monophyletic but excluded anchialine and interstitial species while failing to recover relationships within Sigalionidae. Here we address these shortcomings through the analysis of four molecular markers and 87 morphological characters sampled across 127 species under the assumptions of parsimony and model-based methods. Of the 34 newly sequenced taxa, five anchialine and 24 interstitial species were included, with increased representation of Sigalionidae. An additional 28 elusive Sigalionidae taxa were included, represented only by morphological partitions. Molecular and morphological partitions were evaluated under exhaustive sensitivity analyses, testing the effects of alignment algorithms and optimization criteria on tree topologies. Our trees congruently recovered six clades corresponding to the families within Aphroditiformia: Acoetidae, Aphroditidae, Eulepethidae, Iphionidae, Polynoidae and Sigalionidae, respectively. An anchialine polynoid lineage was nested among strictly deep sea species, and interstitial pisionids and pholoids formed two independent clades nested within Sigalionidae. Additionally, Sigalionidae resulted in four clades, defined by combinations of apomorphies, and hereby we propose the subfamilies Pelogeniinae, Pholoinae, Pisioninae, Sthenelanellinae, as well as the provisionally included polyphyletic Sigalioninae.

Genetic spatial structure of an anchialine cave annelid indicates connectivity within - but not between - islands of the Great Bahama Bank.

Land-locked anchialine blue holes are karstic sinkholes and caves with tidally influenced, vertically stratified water bodies that harbor endemic fauna exhibiting variable troglomorphic features. These habitats represent island-like systems, which can serve to elucidate evolutionary and biogeographic processes at local scales. We investigated whether the 'continuous spelean corridor' hypothesis may elucidate the biogeographical distributions of the stygobitic annelid Pelagomacellicephala iliffei (Polynoidae) collected from the Great Bahama and Caicos Banks of the Bahamas Archipelago. Phylogenetic reconstructions were performed using Bayesian Inference on individual and combined datasets of three molecular markers (16S rDNA, COI, 18S rDNA) and species delimitation employed three widely accepted methods in DNA taxonomy, namely GMYC, bPTP, and ABGD. Mantel tests were used to test the effect of geography on genetic structure. Using these analyses, we recovered five independently evolving entities of the focal species across four islands of the Great Bahama Bank including Cat, Eleuthera, Exumas, and Long. Genetic data yielded strong correlations between islands and phylogenetic entities, signifying independent evolutionary histories within anchialine caves across the platform. The island of Eleuthera showed intra-island gene flow and dispersal capabilities between blue holes separated by 115km, providing evidence of a crevicular spelean corridor within the island. However, no evidence of inter-island dispersal is present in the analyzed system. Consistent with previous biogeographic studies of cave crustaceans, the major barriers shaping the cave biota of the Bahamas Archipelago appears to be the deep trenches and channels separating the Bahamian banks.

Cryptic species of Archinome (Annelida: Amphinomida) from vents and seeps.

Since its description from the Galapagos Rift in the mid-1980s, Archinome rosacea has been recorded at hydrothermal vents in the Pacific, Atlantic and Indian Oceans. Only recently was a second species described from the Pacific Antarctic Ridge. We inferred the identities and evolutionary relationships of Archinome representatives sampled from across the hydrothermal vent range of the genus, which is now extended to cold methane seeps. Species delimitation using mitochondrial cytochrome c oxidase subunit I (COI) recovered up to six lineages, whereas concatenated datasets (COI, 16S, 28S and ITS1) supported only four or five of these as clades. Morphological approaches alone were inconclusive to verify the identities of species owing to the lack of discrete diagnostic characters. We recognize five Archinome species, with three that are new to science. The new species, designated based on molecular evidence alone, include: Archinome levinae n. sp., which occurs at both vents and seeps in the east Pacific, Archinome tethyana n. sp., which inhabits Atlantic vents and Archinome jasoni n. sp., also present in the Atlantic, and whose distribution extends to the Indian and southwest Pacific Oceans. Biogeographic connections between vents and seeps are highlighted, as are potential evolutionary links among populations from vent fields located in the east Pacific and Atlantic Oceans, and Atlantic and Indian Oceans; the latter presented for the first time.

The curious case of Hermodice carunculata (Annelida: Amphinomidae): evidence for genetic homogeneity throughout the Atlantic Ocean and adjacent basins.

Over the last few decades, advances in molecular techniques have led to the detection of strong geographic population structure and cryptic speciation in many benthic marine taxa, even those with long-lived pelagic larval stages. Polychaete annelids, in particular, generally show a high degree of population divergence, especially in mitochondrial genes. Rarely have molecular studies confirmed the presence of 'cosmopolitan' species. The amphinomid polychaete Hermodice carunculata was long considered the sole species within its genus, with a reported distribution throughout the Atlantic and adjacent basins. However, recent studies have indicated morphological differences, primarily in the number of branchial filaments, between the East and West Atlantic populations; these differences were invoked to re-instate Hermodice nigrolineata, formerly considered a junior synonym of H. carunculata. We utilized sequence data from two mitochondrial (cytochrome c oxidase subunit I, 16S rDNA) markers and one nuclear (internal transcribed spacer) marker to examine the genetic diversity of Hermodice throughout its distribution range in the Atlantic Ocean, including the Mediterranean Sea, the Caribbean Sea, the Gulf of Mexico and the Gulf of Guinea. Our analyses revealed generally low genetic divergences among collecting localities and between the East and West Atlantic, although phylogenetic trees based on mitochondrial data indicate the presence of a private lineage in the Mediterranean Sea. A re-evaluation of the number of branchial filaments confirmed differences between East and West Atlantic populations; however, the differences were not diagnostic and did not reflect the observed genetic population structure. Rather, we suspect that the number of branchial filaments is a function of oxygen saturation in the environment. Our results do not support the distinction between H. carunculata in the West Atlantic and H. nigrolineata in the East Atlantic. Instead, they re-affirm the older notion that H. carunculata is a cohesive species with a broad distribution across the Atlantic Ocean.

Flavour preferences conditioned by protein solutions in post-weaning pigs.

Prior studies report in various mammalian species that a neutral flavour can become strongly preferred after being associated with a positive consequence of its consumption. Two experiments were performed to study flavour preferences conditioned by a protein source in weaned pigs. In experiment 1, pigs were trained to drink (30 min/day) one flavour (CS+) mixed into a 2% protein solution (Soybean Protein Concentrate; SPC or Porcine Digestible Peptides; PDP) and another flavour (CS-) mixed into water during 6 alternate sessions. The pigs in the SPC and PDP groups consumed more CS+ than CS- in the two-choice tests with both flavours presented in water (552 vs. 409 mL, 571 vs. 414 mL, respectively). In the last choice SPC and PDP animals preferred the CS+ over the CS- when both flavours were present in feed rather than water (650 vs. 536 g and 678 vs. 513 g, respectively). No differences were observed between the conditioning effects of the two proteins. In experiment 2, pigs were trained (30 min/day) with a garlic flavour (CS) mixed with 4% PDP in sessions 1, 3, 5 and 7 unflavoured tap water in sessions 2, 4, 6 and 8 (Conditioned group) or with garlic flavour in water in sessions 1, 3, 5 and 7 and 4% PDP without added flavour in sessions 2, 4, 6 and 8 (control group). In subsequent choice tests conditioned pigs consumed more PDP+Garlic than PDP in Tests 1 (550 mL vs. 372 ml, P<0.05) and 3 (763 mL vs. 503 mL, P<0.05). In addition, pigs in the Conditioned group made significantly more first contacts (FC, number of piglets at a pan during the first 15s) with the PDP+Garlic solution than PDP solution in Tests 1 and 2 but not in Test 3. In contrast, the control group did not differ in their intakes of or first contacts to the two PDP solutions. The present results indicate that piglets can acquire preferences for a cue flavour added to protein products (PDP and SCP). The conditioned flavour preference also enhanced the attraction to the palatable protein (PDP) when the flavour and protein were combined.

On the classification, evolution and biogeography of terrestrial haemadipsoid leeches (Hirudinida: Arhynchobdellida: Hirudiniformes).

A scourge of tropical and subtropical jungles, bloodfeeding terrestrial leeches of Haemadipsidae have long confused systematists and defied sensible biogeographic interpretation. The family Haemadipsidae usually includes problematic taxa that neither fit the typical IndoPacific distribution of the group, nor properly match diagnostic characters used to define the family. Historically, four additional families-Xerobdellidae, Diestecostomatidae Mesobdellidae and Nesophilaemonidae-have occasionally been recognized for New World and European representatives, though agreement on the composition of those families has not been consistent. Here, we expand the phylogenetic sampling of non-IndoPacific (among other) genera to include Meso American Diestecostoma species and Nesophilaemon skottsbergii from the Juan Fernandez Archipelago in order to critically assess prior hypotheses in a molecular phylogenetic analysis of arhynchobdellid leeches. The result, based on nuclear 18S rDNA and 28S rDNA and mitochondrial COI indicates that there are two distantly related lineages of bloodfeeding terrestrial leeches. The otherwise monophyletic family Haemadipsidae is found to exclude species of Xerobdella, Mesobdella and Diestecostoma. Xerobdellidae is formally resurrected to accommodate species of those three genera. Morphological characteristics corroborate the distinction of Haemadipsidae and Xerobdellidae on the basis of sexual and nephridial characters. Idiobdella seychellensis belongs in Haemadipsidae notwithstanding its lacking respiratory auricles. Nesophilaemon skottsbergii too is in Haemadipsidae notwithstanding its geographic proximity to the xerobdellid Mesobdella gemmata. The characters used to define haemadipsoid families are reevaluated. Feeding preferences and biogeographic patterns are also examined.

Arhynchobdellida (Annelida: Oligochaeta: Hirudinida): phylogenetic relationships and evolution.

A remarkable diversity of life history strategies, geographic distributions, and morphological characters provide a rich substrate for investigating the evolutionary relationships of arhynchobdellid leeches. The phylogenetic relationships, using parsimony analysis, of the order Arhynchobdellida were investigated using nuclear 18S and 28S rDNA, mitochondrial 12S rDNA, and cytochrome c oxidase subunit I sequence data, as well as 24 morphological characters. Thirty-nine arhynchobdellid species were selected to represent the seven currently recognized families. Sixteen rhynchobdellid leeches from the families Glossiphoniidae and Piscicolidae were included as outgroup taxa. Analysis of all available data resolved a single most-parsimonious tree. The cladogram conflicted with most of the traditional classification schemes of the Arhynchobdellida. Monophyly of the Erpobdelliformes and Hirudiniformes was supported, whereas the families Haemadipsidae, Haemopidae, and Hirudinidae, as well as the genera Hirudo or Aliolimnatis, were found not to be monophyletic. The results provide insight on the phylogenetic positions for the taxonomically problematic families Americobdellidae and Cylicobdellidae, the genera Semiscolex, Patagoniobdella, and Mesobdella, as well as genera traditionally classified under Hirudinidae. The evolution of dietary and habitat preferences is examined.