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The mucous skin of amphibians provides a habitat for microorganisms which may interact with their hosts and thereby affect their condition and health. Cultivation-independent analyses of the bacterial communities based on the detection of PCR-amplified bacterial 16S rRNA genes provides a direct approach to characterize their diversity. In the present pilot study we utilized this approach in combination with a high-throughput DNA sequencing technology (454 pyrosequencing), to characterize the bacterial community structure of the skin of three newt species (Lissotriton vulgaris, Ichthyosaura alpestris, Triturus cristatus), collected near Braunschweig, Germany. 16S rDNA sequences were obtained from 19 unique samples. On average, 6113 amplicon sequences were obtained per sample and these could phylogenetically be assigned to a total of 1615 different operational taxonomic units (OTUs). Altogether, most samples were rather similar in their dominant bacterial taxa. Most represented were Betaproteobacteria (46%; mostly Janthinobacterium), Gammaproteobacteria (28%; mostly Pseudomonas), Flavobacteria (phylum Bacteroidetes: 19%, mostly Flavobacterium), and Sphingobacteria (Bacteroidetes: 5%, mostly Pedobacter). We found no significant differences between the three newt species, or between hemi-nested vs. non-nested PCR, but a strong difference among sampling dates (15 and 17 April 2013) which might be explained by the ongoing transition of the newts from their terrestrial to aquatic phase which coincided with this period, or by differences between sexes as these were unevenly sampled on the two dates. 16S rRNA gene sequences retrieved in this study in several cases were identical or very similar to those previously found on the skin of North American salamanders.

Due to their extraordinary coloration, mountain brook newts of the genus Neurergus found in the Near East have fascinated herpetologists since their initial description more than 150 years ago. Although the monophyly of Neurergus newts within the Salamandridae has been unambiguously shown for mitochondrial genes, and recent comprehensive molecular phylogenies placed Neurergus as a sister taxon of banded newts (genus Ommatotriton), we know almost nothing about the structure and relatedness of populations at the intraspecific level. In this study, we therefore analysed sequence variation of a mitochondrial DNA segment (covering a partial region of the control region and the 12S ribosomal RNA) from more than 100 individuals and of two nuclear genes (KIAA and SACS) for a representative subset of individuals originating from nine distinct populations, representing N. strauchii, N. crocatus and N. microspilotus. We also studied individuals of N. derjugini, a taxon that has long been synonymized as N. crocatus, and of which individuals have not been accessible to the scientific community since its original description in 1916. Our results suggest high genetic diversity of populations within species for the mitochondrial DNA marker, while the resolution of applied nuclear genes did not go beyond the level of species. For N. strauchii and N. crocatus, two species that inhabit the largest geographic ranges within the genus, we found a high proportion of diversity both within and between populations for the mitochondrial control region. Individuals of N. microspilotus and N. derjugini only displayed considerable genetic differentiation for one nuclear gene (SACS), while only very little or none genetic differentiation could be found for the mitochondrial control region and the KIAA gene, respectively. As both taxa are also morphologically not well differentiated, we suggest on the basis of the current dataset to taxonomically synonymize N. microspilotus due to priority reasons as N. derjugini. It can be therefore concluded that the most accurate taxonomy of the genus Neurergus should consider N. crocatus, N. strauchii, N. kaiseri and N. derjugini as valid taxonomic units at the species level.

In many species of the Salamandridae family, females provide parental care by carefully wrapping plant material around their eggs. As this behaviour has been shown to have a large effect on offspring survival, variation in this trait is expected to be low. Detailed investigations are, however, lacking. In the present study, we analyzed the consistency of egg-wrapping behaviour in two time periods within a breeding season in female smooth newts (Lissotriton vulgaris). We found a surprisingly low proportion of wrapped eggs of around 47% during the first period (when males were present) and an almost doubled ratio of around 92% in the second period (when males were absent). Also, the variation between individuals was significantly lower in the second period than in the first one. Furthermore, the bigger the females were, the more the proportion of wrapped eggs increased day by day within the first period; however, this relationship did not fully explain the observed difference between the two periods. Our results suggest that parental care in smooth newts can be influenced by the presence of mating partners and body size, and provide the first empirical evidence for within-individual variation in egg-wrapping in a salamandrid species.

Finding a potential mate, that is an individual of the opposite sex, is a fundamental step for sexual reproduction in animal species. Signals involved in the context of mate attraction are mediated by acoustic, visual, and/or chemical signals. For amphibians in general, and especially for many newt and salamander species, chemical cues are known to play important roles in inter- and intraspecific communication. We therefore investigated the use of olfactory cues for sex recognition in terrestrial fire salamanders (Salamandra salamandra) that belong to the group of true salamanders within the Salamandridae. Specifically, we performed odor preference tests with adult salamanders of both sexes and tested whether substrate-borne chemical cues provide sex-specific signals. We found an overall preference for chemical cues of the opposite sex, i.e. males and females differed significantly in their preference for a specific sex. Females spent significantly more time in the compartment with the chemical cues of a male, whereas males did not show a significant preference, but in general more males preferred the compartment with the chemical cues of a female. Our results suggest that fire salamanders are capable to discriminate the sex of conspecific individuals based on pure chemical cues. We discuss our results in the light of the presence of dorsal glands in the males' cloaca of true salamander species, from which pheromones can be directly released onto the substrate.

In the present study, we applied statistical methods to quantitative image analysis of the persistent and individual ventral colour pattern of Salamandrina salamanders, in order to discriminate between individuals of the two species belonging to this genus. Pictures of 238 individuals from three populations of S. perspicillata and pictures of 95 S. terdigitata from two populations were analysed. Partial least squares discriminant analysis (PLSDA) classified 98.78% of individuals into the correct species. PLSDA reaches lower percentages of correct classification when applied to discriminate individuals from different populations of the same species (74.14% for S. perspicillata, 78.26% for S. terdigitata). An ANOVA analysis of colour abundances in different body sectors reveals significant differences between species. The results show that colour pattern has a specific basis, the most discriminant areas being the head and the pectoral girdle. We discuss these results in the light of the proposed evolutionary scenarios of the species, and suggest that ventral colour patterns were driven by founder effect.

The potential of the seventh intron of the β-subunit of the fibrinogen gene (β-fibint 7) for phylogenetic analysis within the Salamandridae family was explored, comparing the topologies of trees based on this marker to those based on mitochondrial 12S rRNA gene previously published. Using primers designed specifically for amphibians, we amplified 25 sequences of β-fibint 7 corresponding to 15 species of salamandrids and one plethodontid species. There was considerable length variation among the β-fibint 7 sequences examined, ranging from 1123 bp in S. atra to 400 bp in P. waltl. Many aspects of the phylogenetic relationships estimated by the two independent loci were congruent and corroborate current taxonomic hypothesis. Although the number of taxa analysed is small, the data obtained in this work suggested that β-fibint 7 is a useful marker for assessing phylogenetic relationships within the Salamandridae family, and is probably appropriate for phylogenetic studies among closely related salamanders that have diverged over the last 20 Myr.

Five new locations of N. strauchii are reported from Turkey, closing the gap between the two subspecies N. s. barani and N. s. strauchii. A molecular analysis based on 829 base pairs from two mitochondrial ribosomal genes (12S and 16S rRNA), together with geographical data from the area concerned, indicate all new populations found are very closely related to the mitochondrial sequences from specimens of N. s. strauchii from its type locality and suggest the river Euphrates might have acted as a natural barrier separating the only populations of N. s. barani known to date from all other populations of N. s. strauchii. A morphological analysis of all N. strauchii populations sampled for this study indicates that belly patterns are not a good diagnostic character to differentiate between N. s. barani and N. s. strauchii, suggesting the only reliable morphological taxonomic character that allows their identification is the different number of yellow spots in adult specimens, which is significantly higher in N. s. strauchii. The analyses also indicate that the number of spots is similar between sub-adult and adult N. s. barani but significantly different between sub-adult and adult specimens of N. s. strauchii, suggesting there is also a difference between both subspecies in increase in the number of spots during maturation.Listed by IUCN as vulnerable and protected by the Bern Convention (1979), our findings indicate N. s. strauchii is more widely distributed than previously thought, while N. s. barani would be restricted to just a few localities. Disturbance of its prime habitat, mountain brooks, and uncontrolled illegal collection, especially of N. s. barani, are still the main threats to the Anatolia newt throughout its known distribution range.

The cloacal anatomy of the palmate newt, Triturus helveticus, was studied in both sexes using light and scanning electron microscopes. The female cloaca is characterized by the occurrence of rugae and folds. Two folds are especially prominent in the region where the spermathecal tubules open into the cloaca. In addition to sperm storage structures, vent glands are obvious in the connective tissue around the cloacal orifice. Their pores occur externally to the cloacal borders, thus suggesting that these glands are a possible source of mating pheromones. Typical features of the male cloaca are: a ciliated epithelium; a cloacal tube dorsal and separate from the anterior cloacal chamber; a broad pseudopenis and wide lateral recesses in the cloacal chamber. Four main types of cloacal glands are recognized in males (dorsal, pelvic, Kingsbury's, and ventral gland). Quite unusual is the occurrence of two distinct types of dorsal glands, possibly related to the production of different pheromonal substances. These glands in the male, together with the occurrence of vent glands in the female, are proposed as cloacal specializations for the production of unequivocal species-specific sex attractants.

The reproductive cycle of the golden-striped salamander, Chioglossa lusitanica, was studied in the vicinity of Porto (northwestern Portugal), in a population that breeds in a mine gallery. Salamanders (67 females and 64 males) were collected between January and November of 1998. Both sexes showed a seasonal reproductive cycle. Spermatogenesis and oogenesis took place between winter and late spring and the presence of spermatozoa and mature oocytes were observed mainly between early summer and late autumn. Gonad, liver and tail weights varied seasonally in both sexes and appear related to different phases of the sexual cycle. Average potential clutch size was 17.8 (sx = 0.8; range = 9-34). Clutch size was correlated with snout-vent length and apparently not influenced by tail-length or age.

Spatial behaviour of Salamandra atra aurorae Trevisan, 1982, was investigated by a capture-recapture program during a whole season of activity. Salamanders changed their behaviour with growth: while juveniles usually dispersed on the ground, adults showed a strong fidelity to a limited area, a few square metres wide, also from year to year. Each individual frequently changed its diurnal refuge over the course of the whole activity season, choosing among a large variety of potential shelters. The distribution pattern of the home ranges was slightly aggregated, irrespective of age and sex. Under the assumptions of a closed population and an equal catchability among individuals, a density of 475 (±184; 95% confidence interval) salamanders per hectare was estimated in a mature and quite open wood, whereas a density of 97 (±23; 95% confidence interval) salamanders per hectare was estimated in a thicker wood.

A molecular phylogeny of the salamandrid genus Neurergus was reconstructed based on two sections of the 12S and 16S mitochondrial ribosomal genes (810 bp), 19 allozyme and three plasma protein loci. When representative species of all closely related salamandrid groups were included, mitochondrial data provided evidence for monophyly of Neurergus within the Salamandridae. Mitochondrial and allozyme data showed homogenous intrageneric tree topologies, but different estimates of times of separation. We calibrated the evolutionary rate to 0.46% pairwise sequence divergence per million years. Accordingly Neurergus diverged 18 million years ago (mya) from a lineage that comprised Euproctus asper and large bodied newts of the genus Triturus. A split around 11 mya produced two major clades within Neurergus. Further separation within the southern 'N. crocatus-clade' (comprising N. crocatus, N. microspilotus and N. kaiseri) occurred ca. 5 mya. The northern 'N. strauchii-clade' separated into N. s. strauchii and N. s. barani ca. 3 mya. Our molecular phylogeny indicates that stream-reproduction is the ancestral state for Neurergus, adaptations to pond-reproduction in N. kaiseri are the result of an evolutionary switch of reproductive biology within the 'N. crocatus-clade'.

The Chinhai Salamander, Echinotriton chinhaiensis, mates on land. The male of a courting pair marks a trail by a mucous excretion from his cloaca. The pair follows an ellipsoid track during which manoeuvre a spermatophore is deposited. Partners align their bodies such that the female is led over the spermatophore. Clasping was not observed. The female traces the spermatophore with her cloaca through shifting movements with the pelvic region. A male may deposit several spermatophores on one evening. The courtship is compared to that in the genus Tylototriton.

We studied age structure and growth in two populations of the golden-striped salamander, Chioglossa lusitanica, in northern Portugal by cohort analysis and skeletochronology. Lines of Arrested Growth (LAG) deposited during the larval phase could be distinguished from LAG deposited after metamorphosis. One or two LAG were found in larvae, with counts corresponding to age in years as predicted from larval size distributions. Post-metamorphic modal age was 5 to 6 years and longevity was 8 years. Von Bertalanffy growth curves for males and females from both populations were different from one another. Sexual maturity was reached 4 to 5 years after metamorphosis and corresponded with a snout-vent length of 43-44 mm in both sexes. A tendency was observed for females to be older than males. Mature females were on average larger than mature males. Larval growth was higher in spring than in winter and differed between populations and years. The population in which larvae grew relatively slowly was characterized by large young adults and vice versa, perhaps reflecting contrasting aquatic and terrestrial feeding conditions. The knowledge here presented is important for the better understanding of the population dynamics and ecological and conservation requirements of the golden-striped salamander.

The feeding ecology of 28 amphibian species with complete life cycles has been studied from the last pre-metamorphic stages to metamorphosed juveniles. The widespread view that feeding ceases completely during metamorphosis is not confirmed. Generally, however, amphibian feeding rate decreases at metamorphosis. Foraging in Caudata either does not cease (Hynobiidae, rheophilous Salamandridae) or ceases only before the end of transformation, which takes less than one metamorphic stage. The cessation of foraging in Anura coincides with the transformation of the mouth and digestive tract at the beginning of the metamorphic climax. Foraging on small animals starts just after the change from a larval to a post-metamorphic mouth, i.e., before the end of metamorphosis.

Oviposition site selection in Triturus helveticus, T. alpestris and T. cristatus was studied both in natural and laboratory situations. In a natural breeding site, eggs of the three species were not laid equally on several plants: T. cristatus laid practically all their egges on only one plant (Nasturtium officinale) while T. helveticus eggs were collected on four plant species. In laboratory experiments, plastic and cotton fabric supports made it possible to test ovipositing female selectivity according to flexibility and shape of support and egg-laying distance below the surface. In all three species, a high proportion of eggs were laid on supports of linear shape (as opposed to arborescent shape) and egg frequency decreased as a function of depth (from 0 to 40 cm below the surface). Selection according to flexibility was also observed, the smallest species (T. helveticus) laying more eggs on the thinner supports. The role of support selectivity during oviposition in Triturus is discussed, in term of proximate (i.e. female protection against predators, breathing necessity) and ultimate (i.e. egg survival) factors, and related to new ecology.

In the spring of 1984 an ecological investigation on the endemic salamanders Hydromantes genei flavus and Euproctus platycefihalus was carried out in eastern Sardinia. The investigation was focussed on natural population sizes and densities. The Sardinian cave salamander was found to occur in two clearly distinct habitats, viz. humid holm oak forests and limestone caves. The density found in the forests was 300/ha and 98 ± 7 individuals were found in one cave. The Sardinian brook salamander was found in a habitat not reported before, viz. a stagnant mountain lake. There the animals were rather robust, and a neotenic specimen was captured.

In its general pattern the sexual behaviour of Triturus montandoni most resembles that of Triturus helveticus. The courtship consists of three phases: orientation, static display and retreat display, followed by a spermatophore transfer phase. During display the male performs three tail movements: the fan, the whip, and the wave. The relative frequencies of tail movements and the duration of fanning bouts are the main difference between T montandoni, T. helveticus, and T. vulgaris. The phylogenetic relationships between these three species are discussed.

An electrophoretic survey of proteins from 27 populations examines the level of genetic differentiation among four subspecies of T. vulgaris in Yugoslavia. Populations of T. v. vulgaris, T. v. meridionalis and T. v. graecus are genetically the most similar to each other, while T. v. dalmaticus is substantially different from these subspecies. This pattern , which is not in accordance with the pattern of morphological similarities, is discussed with special reference to Pleistocene glaciation periods.

Abstract Mertensiella caucasica specimens, collected from various parts of northeastern Anatolia, are compared statistically and from the point of view of their pattern and coloration. While no significant taxonomic differences were detected between the samples of different populations, the range diagrams, constructed from several body ratios, show a marked difference between the adult males, adult females, sub-adults, juveniles and larvae of the same population.

Abstract The karyotype of Neurergus strauchii shows pericentric and subterminal C-bands heterochromatin and nucleolar organizers on chromosome IX. This site is revealed by both a C-band and a chromomycin A3 fluorescent band; the GC-specific fluorochrome also highlights additional sites of fluorescence. The lampbrush chromosomes have a general morpho-structural feature known for Triturus lampbrush chromosomes, but giant loops are lacking. Nucleolar organizer and sphere sites are the main cytogenetic markers in the lampbrush karyotype of Neurergus. The implication of these results to our understanding of cytotaxonomy and evolutionary biology of Palearctic Salamandrids is discussed.

A field study has been made in the relatively narrow sympatric zone of Triturus cristatus and T. marmoratus, in the department Mayenne (France). Ecological data have been analysed. In the sympatric area specific preferences appear to exist: ecological isolation has developed in some areas. Typical T. marmoratus areas are hilly and wooded, provided with many terrestrial hiding-places, while forest pools and springs especially serve as spawning sites. T. cristatus areas are flat and open. Comparison with distribution data of 24 years ago shows expansion of T. cristatus at the expense of T. marmoratus, which presumably is true for the whole sympatric zone. The preferences of the species in combination with changes in environment by human intervention explain the detected changes in distribution. Ecological isolation as a phenomenon in the speciation of both species is discussed.

Chromosome number, karyotype and chromosomal banding pattern of Mertensiella caucasica were studied in differentially stained mitotic preparations. The main feature of the C-banded appearance concerns the centromere regions, which are heavily stained by the Giemsa and are outlined by proximal C-bands. This morpho-structural character seems to be peculiar of the genome of M. caucasica, when compared with the C-banding pattern shown by the chromosomes of Salamandra, considered the most closely related form. Therefore, a preliminary cytotaxonomic survey seems to support the opinion that the Caucasian salamander can be referred to a well defined genus Mertensiella rather than a subgenus of Salamandra.

During 1977 and in October 1978 fieldwork was carried out on the Golden-striped Salamander (Chioglossa lusitanica) in the surroundings of Porto (Portugal). The occurrence of C. lusitanica appears to be limited to the northwestern part of the Iberian Peninsula; its distribution is well defined by the region with a precipitation of more than 1000 millimeters per annum and not exceeding 1500 meters altitude. Here it leads a nocturnal and secretive life along small streams in hilly and mountainous terrain. In autumn, the adults meet on their mating-sites, observations on the mating process conform with data already available. Oögenesis takes place in summer. A positive relation was found between the length of the female and its fecundity. C. lusitanica can autotomize its tail, which can regenerate completely. It was found that oögenesis proceeds at the expense of tail-regeneration. Egg-deposition takes place in September, October and November, the larvae hatch after six to nine weeks. Metamorphosis takes place from the beginning of summer onward. Some larvae postpone metamorphosis until next season, having spent two winters in the water. Adults spend the summer in hidden places and remain largely inactive. It was found that with the drying up of the brooks a part of some populations migrated to summer-refuges such as dams and deserted mine galleries, thereby sometimes covering long distances. These refuges may also serve as a place for copulation and for egg-deposition. Migration-patterns in the populations under study were markedly different, largely depending on the availability of resorts and summer-refuges. Juveniles lead a live less nocturnal than the adults. Furthermore they show a stronger tendency to stay close to the brook and to stay there in summer. The population-density along one of the brooks investigated was estimated at four to five adult specimens per section of one meter in length. Water withdrawal presents a serious danger to certain populations.

In its general pattern, courtship of Paramesotriton caudopunctatus is similar to that of Triturus species. The male courts a female by making fanning movements with his tail. When the female's response is positive, she follows the male. The male deposits a spermmass, which remains sticking to the cloaca of the female, when she creeps over it. P. caudopunctatus displays only one type of tailmovement: the'fanning'. The tip of the male's tail, which is marked with a bright spot, probably plays a role in courtship as a visual stimulator. At the time of oviposition the female's cloaca is swollen. The eggs are mounted in clefts and hollows in or between stones.