Proteus anguinus
is a neotenic cave salamander, endemic to the Dinaric Karst and a symbol of world natural heritage. It is classified as “vulnerable” by the International Union for Conservation of ...Nature (IUCN) and is one of the EU priority species in need of strict protection. Due to inaccessibility of their natural underground habitat, scientific studies of the olm have been conducted mainly in captivity, where the amphibians are particularly susceptible to opportunistic microbial infections. In this report, we focused on the diversity of cultivable commensal fungi isolated from the skin of asymptomatic and symptomatic animals obtained from nature (20 specimens) and captivity (22 specimens), as well as from underground water of two karstic caves by direct water filtration and by exposure of keratin-based microbial baits and subsequent isolation from them. In total 244 fungal isolates were recovered from the animals and additional 153 isolates were obtained from water samples. Together, these isolates represented 87 genera and 166 species. Symptomatic animals were colonized by a variety of fungal species, most of them represented by a single isolate, including genera known for their involvement in chromomycosis, phaeohyphomycosis and zygomycosis in amphibians:
Acremonium
,
Aspergillus
,
Cladosporium
,
Exophiala
,
Fusarium
,
Mucor
,
Ochroconis
,
Phialophora
and
Penicillium
. One symptomatic specimen sampled from nature was infected by the oomycete
Saprolegnia parasitica
, the known causative agent of saprolegniosis. This is the first comprehensive report on cultivable skin mycobiome of this unique amphibian in nature and in captivity, with an emphasis on potentially pathogenic fungi and oomycetes.
1. Various groundwater habitats have exceptionally high levels of endemism caused by strong hydrographical isolation and low dispersal abilities of their inhabitants. More than 10% of ...macro-stygobiotic species nevertheless occupy relatively large ranges, measuring from some hundred to over 2000 km in length. These species represent a challenge because their distributions disregard hydrographical boundaries, and their means to disperse and maintain long-term gene flow are unknown. 2. Based on mitochondrial and nuclear gene sequences, we examined the phylogeographic structure of six formally recognised stygobiotic species (Niphargus virei, N. rhenorhodanensis, Troglocaris anophthalmus, T. hercegovinensis, Spelaeocaris pretneri, Proteus anguinus) and searched for cryptic lineage diversity in a genus-wide phylogeny of Niphargus. Using tree-based criteria as well as comparative divergence measures, we identified cryptic lineages, which may tentatively be equated with cryptic species. 3. Fourteen analysed nominal stygobiotic species with large ranges emerged as highly diversified, splitting into 51 tentative cryptic lineages. The degree of divergence was within the range or larger than the divergence of other related pairs of sister species. A substantial part (94%) of the cryptic lineages had ranges <200 km in length. One half of them were recorded at single sites only. The largest range recorded was that of a cryptic N. virei lineage (700 km), while none of the very large traditional ranges (e.g. Niphargus aquilex- 2300 km, N. tauri- 1900 km) could be corroborated. 4. These data suggest that small ranges of macro-stygobionts are the rule, and ranges over 200 km are extremely rare. 5. The implications of this result for groundwater biodiversity assessment and conservation include a considerable increase in overall diversity at the regional and continental scale and a strong decrease in faunal similarities among regions, coupled with greater endemism.
Microplastic pollution in karst systems is still poorly studied, despite the presence of protected species and habitats, and important water reserves. Vulnerable key species hosted in these habitats ...could consume or assimilate microplastics, which can irreversibly damage management efforts, and thus ecosystems functionality. This can be particularly true for subterranean water habitats where microplastic pollution effects on wildlife management programs are not considered. The aim of this study is to provide a case study from the Classical Karst Region, which hosts peculiar habitats and key species protected at European level, such as the olm Proteus anguinus. As this area has been deeply exploited and modified over time, and is adjacent to highways, roads and railways, which could contribute to pollution within the karst system, threatening the ecosystems, it provides a perfect model system.
In this study we collected and investigated water and sediment samples from aquatic environments of surface and subterranean habitats hosting several subterranean environment-adapted organisms. Examined particles were counted and characterized by size, color and shape via visual identification under a microscope, with and without UV light. Furthermore, spectroscopic analyses were carried out in order to identify microplastics typology. Microplastics were found in all examined habitats. In water, microplastics concentration ranged from 37 to 86 items/L, in sediments from 776 to 2064 items/kg. Fibre-shape was the main present, followed by fragments and beads, suggesting multiple sources of pollution, especially textile products. Most of the particles were fluorescent under UV light and were mainly transparent, while not-fluorescent ones were especially black, blue or brown. Samples contained especially polyesters and copolymers. These results highlight intense MP pollution in karst areas, with significant impacts on water quality, and potential effects on subterranean environment-dwelling species. We stress the importance of monitoring pollution in these critical environments for biodiversity and habitat conservation: monitoring in karst areas must become a priority for habitat and species protection, and water resources management, improving analyses on a larger number of aquatic surface and subterranean habitats.
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•Karst surface and subterranean waters are polluted by microplastics.•Protected habitats and stygobiont species are threatened by microplastics.•Small size and fibre shape microplastics dominate the samples.•Samples contained especially polyesters and copolymers.•Monitoring aquatic environments is fundamental for their conservation.
In comparison to biodiversity on Earth's surface, subterranean biodiversity has largely remained concealed. The olm (Proteus anguinus) is one of the most enigmatic extant cave inhabitants, and until ...now little was known regarding its genetic structure and evolutionary history. Olms inhabit subterranean waters throughout the Dinaric Karst of the western Balkans, with a seemingly uniform phenotypic appearance of cave‐specialized traits: an elongate body, snout and limbs, degenerated eyes and loss of pigmentation (“white olm”). Only a single small region in southeastern Slovenia harbours olms with a phenotype typical of surface animals: pigmented skin, eyes, a blunt snout and short limbs (“black olm”). We used a combination of mitochondrial DNA and genome‐wide single nucleotide polymorphism data to investigate the molecular diversity, evolutionary history and biogeography of olms along the Dinaric Karst. We found nine deeply divergent species‐level lineages that separated between 17 and 4 million years ago, while molecular diversity within lineages was low. We detected no signal of recent admixture between lineages and only limited historical gene flow. Biogeographically, the contemporaneous distribution of lineages mostly mirrors hydrologically separated subterranean environments, while the historical separation of olm lineages follows microtectonic and climatic changes in the area. The reconstructed phylogeny suggests at least four independent transitions to the cave phenotype. Two of the species‐level lineages have miniscule ranges and may represent Europe's rarest amphibians. Their rarity and the decline in other lineages call for protection of their subterranean habitats.
Most research on mechanisms of aging is being conducted in a very limited number of classical model species, i.e., laboratory mouse (
Mus musculus
), rat (
Rattus norvegicus domestica
), the common ...fruit fly (
Drosophila melanogaster
) and roundworm (
Caenorhabditis elegans
). The obvious advantages of using these models are access to resources such as strains with known genetic properties, high-quality genomic and transcriptomic sequencing data, versatile experimental manipulation capabilities including well-established genome editing tools, as well as extensive experience in husbandry. However, this approach may introduce interpretation biases due to the specific characteristics of the investigated species, which may lead to inappropriate, or even false, generalization. For example, it is still unclear to what extent knowledge of aging mechanisms gained in short-lived model organisms is transferable to long-lived species such as humans. In addition, other specific adaptations favoring a long and healthy life from the immense evolutionary toolbox may be entirely missed. In this review, we summarize the specific characteristics of emerging animal models that have attracted the attention of gerontologists, we provide an overview of the available data and resources related to these models, and we summarize important insights gained from them in recent years. The models presented include short-lived ones such as killifish (
Nothobranchius furzeri
), long-lived ones such as primates (
Callithrix jacchus, Cebus imitator, Macaca mulatta
), bathyergid mole-rats (
Heterocephalus glaber, Fukomys spp.
), bats (
Myotis spp.
), birds, olms (
Proteus anguinus
), turtles, greenland sharks, bivalves
(Arctica islandica
), and potentially non-aging ones such as
Hydra
and
Planaria
.
In surveillance of subterranean fauna, especially in the case of rare or elusive aquatic species, traditional techniques used for epigean species are often not feasible. We developed a non-invasive ...survey method based on environmental DNA (eDNA) to detect the presence of the red-listed cave-dwelling amphibian, Proteus anguinus, in the caves of the Dinaric Karst. We tested the method in fifteen caves in Croatia, from which the species was previously recorded or expected to occur. We successfully confirmed the presence of P. anguinus from ten caves and detected the species for the first time in five others. Using a hierarchical occupancy model we compared the availability and detection probability of eDNA of two water sampling methods, filtration and precipitation. The statistical analysis showed that both availability and detection probability depended on the method and estimates for both probabilities were higher using filter samples than for precipitation samples. Combining reliable field and laboratory methods with robust statistical modeling will give the best estimates of species occurrence.
The biology of cave‐dwelling species is a fascinating area, yet it is understudied due to logistic constraints, especially in regard to aquatic organisms. The olm (Proteus anguinus) is the largest ...known troglobiont vertebrate, showing extreme life‐history characteristics such as a possible lifespan of over 100 years and a reproductive cycle of over 12 years. However, most studies carried out on the species to date are based on laboratory studies, resulting in a severe lack of ecological data from natural populations studied in their original habitat. We applied a capture–mark–recapture approach on an Eastern Herzegovinian population for eight years to reveal its spatial strategy and general movement patterns. We found that P. anguinus is sedentary, can often be found within a few square metres over several years, and their moving distance is uncorrelated with the time elapsed between recaptures. Previous laboratory studies constrained to confined space and artificial environments have already suggested site fidelity and our results from a natural population confirm this. The low reproductive activity of the species together with the reported extreme site fidelity makes this top predator of aquatic cave communities highly vulnerable and a sensitive bio‐indicator of habitat‐changing human activities.
This is the first capture–mark–recapture study on a natural population of the obligate aquatic cave‐dwelling olm (Proteus anguinus), known for its extreme life‐history characteristics (lifespan over 100 years, reproductive cycle over 10 years). We observed extreme site fidelity; individuals moved typically less than 15 m over years.
Vitellogenin (Vtg) is a precursor protein of egg yolk proteins in oviparous and ovoviviparous vertebrates. Except in a case of exposure to estrogenic endocrine disruptors, Vtg is a female-specific ...protein and could be used as a molecular marker for sex identification. This would be especially useful in the case of the endangered European cave salamander Proteus anguinus in which sexes are indistinguishable according to external morphology, which hinders the establishment of a successful captive breeding program. Here we describe the identification, partial characterization, and purification of Vtg from P. anguinus. Vtg was identified in the plasma of a vitellogenic proteus female with visible oocytes. The identification of this protein was accomplished by mass spectrometry analysis. Two-dimensional gel electrophoresis revealed proteus Vtg as a mix of 190 kDa isoforms with isoelectric points in the pH range 5.3–6.0. Vtg was purified from proteus blood by gel filtration followed by anion-exchange chromatography. Using specific staining of SDS-PAGE gels, the Vtg was found to be phosphorylated and lipidated. Unlike the case in some other aquatic vertebrates, in P. anguinus, Vtg was not present in detectable amounts in cutaneous mucus. Degradation of oocytes in the captive vitellogenic female was accompanied by simultaneous decrease of Vtg concentration. Over a period of 10 months, the concentration of Vtg dropped from maximal to sub-detectable. Our results show that Vtg is a promising molecular marker for sex identification and ovary maturation in P. anguinus, which could contribute to the development of a viable program for captive reproduction of this unique species.
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•In Proteus anguinus, sex detemination is indistinguishable by outer morphology, sex -specific loci or sex chromosomes.•Vitellogenin, a female-specific protein, was identified in the plasma of Proteus anguinus.•Vitellogenin is a 190 kDa phospholipoprotein with an isoelectric point of 5.3–6.0.•In a captive female, the concentration of vitellogenin dropped from maximum to sub-detectable over a period of 10 months.
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