First extraction of eDNA from tree hole water to detect tree frogs: a simple field method piloted in Madagascar

Environmental DNA (eDNA) is becoming an increasingly used tool for monitoring cryptic species within terrestrial and aquatic systems. We present the first method for extracting water from tree holes for eDNA studies of tree-dwelling frogs, and the first use of eDNA for amphibian monitoring in Madagascar. This pilot study expands on a previously developed method and aims to provide a simple field protocol for DNA extraction from very small water samples, using a relatively inexpensive kit compared to other collection methods. We collected 20 ml of water from tree holes in Ambohitantely Special Reserve in Madagascar, with the aim to survey for the Critically Endangered tree frog Anodonthyla vallani, and we developed species specific cytochrome c oxidase 1 primers for this species. While our two samples did not detect A. vallani, we successfully extracted up to 16.6 ng/µl of eDNA from the samples and using 16S rRNA primers barcoded the tree frog Plethodontohyla mihanika in one of the samples. Despite just two samples being collected, we highlight the future potential of eDNA from tree holes for investigating cryptic habitat specialist amphibians given we extracted frog eDNA from just 20 ml of water. The method provides a rapid, simple, and cost-effective method which can assist cryptic species monitoring in challenging and time-consuming field conditions and should be developed further for frog surveying in Madagascar and beyond. The newly developed primers can be used for further work using this eDNA method to survey threatened Anodonthyla frog species

The complete mitochondrial genome of rare and critically endangered Anilany helenae (Microhylidae) of Madagascar

Anilany helenae is a Critically Endangered frog native to the central highlands of Madagascar. Due to ongoing habitat loss of its known range, this species’ population is considered declining, while little is known about its ecology, behavior, and taxonomy. Within the context of developing tools that can aid the conservation of Madagascar’s amphibian fauna, and add to the continued understanding of their taxonomy, we assembled its complete mitochondrial genome (Genbank Accession number MZ751042). This contributes the first complete mitochondrial genome of a microhylid from Madagascar, despite there being over 100 species in the Cophylinae subfamily alone. Anilany helenae’s circular mitochondrial genome is 17,519 bp long, contains 37 genes, and exhibits differences in gene arrangement compared with other microhylids, including the placement of protein coding genes nad1 and nad2. A phylogeny of the 13 protein coding genes of the few Madagascan anuran mitogenomes available, along with species from Africa and East Asia, places A. helenae along with the New Guinean Mantophryne lateralis in a basal position with respect to the other microhylids in the tree

Methanotrophic bacterial symbionts fuel dense populations of deep-sea feather duster worms (Sabellida, Annelida) and extend the spatial influence of methane seepage

Deep-sea cold seeps are dynamic sources of methane release and unique habitats supporting ocean biodiversity and productivity. Here, we describe newly discovered animal-bacterial symbioses fueled by methane, between two species of annelid (a serpulid Laminatubus and sabellid Bispira) and distinct aerobic methane-oxidizing bacteria belonging to the Methylococcales, localized to the host respiratory crown. Worm tissue δ¹³C of −44 to −58‰ are consistent with methane-fueled nutrition for both species, and shipboard stable isotope labeling experiments revealed active assimilation of ¹³C-labeled methane into animal biomass, which occurs via the engulfment of methanotrophic bacteria across the crown epidermal surface. These worms represent a new addition to the few animals known to intimately associate with methane-oxidizing bacteria and may further explain their enigmatic mass occurrence at 150–million year–old fossil seeps. High-resolution seafloor surveys document significant coverage by these symbioses, beyond typical obligate seep fauna. These findings uncover novel consumers of methane in the deep sea and, by expanding the known spatial extent of methane seeps, may have important implications for deep-sea conservation

Methanotrophic bacterial symbionts fuel dense populations of deep-sea feather duster worms (Sabellida, Annelida) and extend the spatial influence of methane seepage

Deep-sea cold seeps are dynamic sources of methane release and unique habitats supporting ocean biodiversity and productivity. Here, we describe newly discovered animal-bacterial symbioses fueled by methane, between two species of annelid (a serpulid Laminatubus and sabellid Bispira) and distinct aerobic methane-oxidizing bacteria belonging to the Methylococcales, localized to the host respiratory crown. Worm tissue δ¹³C of −44 to −58‰ are consistent with methane-fueled nutrition for both species, and shipboard stable isotope labeling experiments revealed active assimilation of ¹³C-labeled methane into animal biomass, which occurs via the engulfment of methanotrophic bacteria across the crown epidermal surface. These worms represent a new addition to the few animals known to intimately associate with methane-oxidizing bacteria and may further explain their enigmatic mass occurrence at 150–million year–old fossil seeps. High-resolution seafloor surveys document significant coverage by these symbioses, beyond typical obligate seep fauna. These findings uncover novel consumers of methane in the deep sea and, by expanding the known spatial extent of methane seeps, may have important implications for deep-sea conservation

An unexpected new red-bellied Stumpffia (Microhylidae) from forest fragments in central Madagascar highlights remaining cryptic diversity

The Madagascan endemic subfamily Cophylinae in the family Microhylidae, is an example of a taxonomic group for which much is still to be discovered. Indeed, the cophyline frogs present a large portion of Madagascar’s cryptic and microendemic amphibian diversity, yet they remain understudied. A new red-bellied species of the microhylid frog genus Stumpffia is described from the central plateau of Madagascar. Visual encounter surveys in Ambohitantely and Anjozorobe in 2019 and 2020 identified this previously unknown Stumpffia species, which closely resembles Stumpffia kibomena known from Andasibe in the east. Stumpffia lynnae sp. nov. adds another species to the red-bellied species complex, differing from S. kibomena by genetic differentiation in the mitochondrial 16S rRNA gene (3.6–3.9%) and distinct nuclear RAG1 haplotypes, as well as strongly by its advertisement call. The new species is known from across Ambohitantely Special Reserve and Anjozorobe Angavo protected area, but is known only from one complete specimen and eight individual tissue samples. Based on the rarity of the species, the small number of locations in which it has been found, and its disappearing forest habitat, its IUCN Red List classification is suggested as “Endangered”. This species is the first Stumpffia described from Madagascar’s central plateau, highlighting the importance of conserving the remnant forest fragments in this area and the ongoing need to survey and protect this threatened habitat type

The gut of the finch: uniqueness of the gut microbiome of the Galápagos vampire finch.

BACKGROUND: Darwin's finches are a clade of 19 species of passerine birds native to the Galápagos Islands, whose biogeography, specialized beak morphologies, and dietary choices-ranging from seeds to blood-make them a classic example of adaptive radiation. While these iconic birds have been intensely studied, the composition of their gut microbiome and the factors influencing it, including host species, diet, and biogeography, has not yet been explored. RESULTS: We characterized the microbial community associated with 12 species of Darwin's finches using high-throughput 16S rRNA sequencing of fecal samples from 114 individuals across nine islands, including the unusual blood-feeding vampire finch (Geospiza septentrionalis) from Darwin and Wolf Islands. The phylum-level core gut microbiome for Darwin's finches included the Firmicutes, Gammaproteobacteria, and Actinobacteria, with members of the Bacteroidetes at conspicuously low abundance. The gut microbiome was surprisingly well conserved across the diversity of finch species, with one exception-the vampire finch-which harbored bacteria that were either absent or extremely rare in other finches, including Fusobacterium, Cetobacterium, Ureaplasma, Mucispirillum, Campylobacter, and various members of the Clostridia-bacteria known from the guts of carnivorous birds and reptiles. Complementary stable isotope analysis of feathers revealed exceptionally high δ15N isotope values in the vampire finch, resembling top marine predators. The Galápagos archipelago is also known for extreme wet and dry seasons, and we observed a significant seasonal shift in the gut microbial community of five additional finch species sampled during both seasons. CONCLUSIONS: This study demonstrates the overall conservatism of the finch gut microbiome over short (< 1 Ma) divergence timescales, except in the most extreme case of dietary specialization, and elevates the evolutionary importance of seasonal shifts in driving not only species adaptation, but also gut microbiome composition

The gut of the finch: uniqueness of the gut microbiome of the Galápagos vampire finch

Background: Darwin’s finches are a clade of 19 species of passerine birds native to the Galápagos Islands, whose biogeography, specialized beak morphologies, and dietary choices—ranging from seeds to blood—make them a classic example of adaptive radiation. While these iconic birds have been intensely studied, the composition of their gut microbiome and the factors influencing it, including host species, diet, and biogeography, has not yet been explored. Results: We characterized the microbial community associated with 12 species of Darwin’s finches using high-throughput 16S rRNA sequencing of fecal samples from 114 individuals across nine islands, including the unusual blood-feeding vampire finch (Geospiza septentrionalis) from Darwin and Wolf Islands. The phylum-level core gut microbiome for Darwin’s finches included the Firmicutes, Gammaproteobacteria, and Actinobacteria, with members of the Bacteroidetes at conspicuously low abundance. The gut microbiome was surprisingly well conserved across the diversity of finch species, with one exception—the vampire finch—which harbored bacteria that were either absent or extremely rare in other finches, including Fusobacterium, Cetobacterium, Ureaplasma, Mucispirillum, Campylobacter, and various members of the Clostridia—bacteria known from the guts of carnivorous birds and reptiles. Complementary stable isotope analysis of feathers revealed exceptionally high δ15N isotope values in the vampire finch, resembling top marine predators. The Galápagos archipelago is also known for extreme wet and dry seasons, and we observed a significant seasonal shift in the gut microbial community of five additional finch species sampled during both seasons. Conclusions: This study demonstrates the overall conservatism of the finch gut microbiome over short (< 1 Ma) divergence timescales, except in the most extreme case of dietary specialization, and elevates the evolutionary importance of seasonal shifts in driving not only species adaptation, but also gut microbiome composition

Collaborative International Research in Clinical and Longitudinal Experience Study in NMOSD.

Objective: To develop a resource of systematically collected, longitudinal clinical data and biospecimens for assisting in the investigation into neuromyelitis optica spectrum disorder (NMOSD) epidemiology, pathogenesis, and treatment. Methods: To illustrate its research-enabling purpose, epidemiologic patterns and disease phenotypes were assessed among enrolled subjects, including age at disease onset, annualized relapse rate (ARR), and time between the first and second attacks. Results: As of December 2017, the Collaborative International Research in Clinical and Longitudinal Experience Study (CIRCLES) had enrolled more than 1,000 participants, of whom 77.5% of the NMOSD cases and 71.7% of the controls continue in active follow-up. Consanguineous relatives of patients with NMOSD represented 43.6% of the control cohort. Of the 599 active cases with complete data, 84% were female, and 76% were anti-AQP4 seropositive. The majority were white/Caucasian (52.6%), whereas blacks/African Americans accounted for 23.5%, Hispanics/Latinos 12.4%, and Asians accounted for 9.0%. The median age at disease onset was 38.4 years, with a median ARR of 0.5. Seropositive cases were older at disease onset, more likely to be black/African American or Hispanic/Latino, and more likely to be female. Conclusions: Collectively, the CIRCLES experience to date demonstrates this study to be a useful and readily accessible resource to facilitate accelerating solutions for patients with NMOSD