Proteomics of Galápagos Marine Iguanas Links Function of Femoral Gland Proteins to the Immune System

Communication between individuals via molecules, termed chemosignaling, is widespread among animal and plant species. However, we lack knowledge on the specific functions of the substances involved for most systems. The femoral gland is an organ that secretes a waxy substance involved in chemical communication in lizards. Although the lipids and volatile substances secreted by the femoral glands have been investigated in several biochemical studies, the protein composition and functions of secretions remain completely unknown. Applying a proteomic approach, we provide the first attempt to comprehensively characterize the protein composition of femoral gland secretions from the Galápagos marine iguana. Using samples from several organs, the marine iguana proteome was assembled by next-generation sequencing and MS, resulting in 7513 proteins. Of these, 4305 proteins were present in the femoral gland, including keratins, small serum proteins, and fatty acid-binding proteins. Surprisingly, no proteins with discernible roles in partner recognition or inter-species communication could be identified. However, we did find several proteins with direct associations to the innate immune system, including lysozyme C, antileukoproteinase (ALP), pulmonary surfactant protein (SFTPD), and galectin (LGALS1) suggesting that the femoral glands function as an important barrier to infection. Furthermore, we report several novel anti-microbial peptides from the femoral glands that show similar action against Escherichia coli and Bacillus subtilis such as oncocin, a peptide known for its effectiveness against Gram-negative pathogens. This proteomics data set is a valuable resource for future functional protein analysis and demonstrates that femoral gland secretions also perform functions of the innate immune system.

Diversity of compounds in femoral secretions of Galápagos iguanas (genera:AmblyrhynchusandConolophus), and their potential role in sexual communication in lek-mating marine iguanas (Amblyrhynchus cristatus)

BackgroundChemical signals are widely used in the animal kingdom, enabling communication in various social contexts, including mate selection and the establishment of dominance. Femoral glands, which produce and release waxy secretions into the environment, are organs of central importance in lizard chemical communication. The Galápagos marine iguana (Amblyrhynchus cristatus) is a squamate reptile with a lek-mating system. Although the lekking behaviour of marine iguanas has been well-studied, their potential for sexual communication via chemical cues has not yet been investigated. Here we describe the diversity of the lipophilic fraction of males’ femoral gland secretions among 11 island populations of marine iguanas, and compare it with the composition of its sister species, the Galápagos land iguana (Conolophus subcristatus). We also conducted behavioural observations in marine iguana territorial males in order to explore the possible function of these substances in the context of male dominance in leks.MethodsFemoral secretions were analysed by gas chromatography coupled to mass spectrometry (GC–MS), and chromatography with a flame ionisation detector (GC-FID) in order to characterise the lipophilic composition. To understand the potential role of femoral secretions in marine iguana intraspecific communication, territorial males were sampled for their femoral glands and monitored to record their head bob rate—a territorial display behaviour in males—as well as the number of females present in their leks.ResultsWe found that the gland secretions were composed of ten saturated and unsaturated carboxylic acids ranging in chain length between C16and C24, as well as three sterols. Cholesterol was the main compound found. Intriguingly, land iguanas have a higher diversity of lipophilic compounds, with structural group of lipids (i.e. aldehydes) entirely absent in marine iguanas; overall the chemical signals of both species were strongly differentiated. Lipid profiles also differed among populations of marine iguanas from different islands, with some islands demonstrating a high diversity of lipophilic compounds (i.e. full spectra of compounds), and others lacking one or more compounds. Among the compounds most frequently found missing were 11- and 13-eicosenoic acids. Gland secretions of males with a better body condition and with a higher dominance status (i.e. those accompanied by females and with higher head bob display) were proportionately richer in C20-unsaturated fatty acids (11-eicosenoic acid).DiscussionLand and marine iguanas strongly diverged in their chemical composition of the femoral glands likely due to ecological differences between both species. Despite that marine iguana populations varied in their femoral gland composition that was not related to their genetic structure. Our results indicated that 11-eicosenoic acid may play an important role in intraspecific chemical communication in marine iguanas.

Generation glands of cordylid lizards: mechanism of secretion transfer to the environment

Cordylid lizards possess two types of holocrine epidermal glands involved in chemical communication, femoral glands and generation glands. We investigated how the glandular products of generation glands are dispersed to the environment, as it is assumed in earlier studies that glandular material of cordylid generation glands is transferred to the substrate via abrasion, similar to the situation in femoral glands. By means of scanning electron microscopy, we established that the outer gland surfaces of adult Cordylus cordylus, Hemicordylus capensis and Pseudocordylus microlepidotus show only localized signs of wear and tear, indicative of a very low abrasion rate. Using standard histological techniques and light microscopy, we studied the appearance of multi-layer generation glands in a series of individuals of different ages in Cordylus macropholis. In this species, generation glands are already conspicuous in neonates. The significant relationship between SVL and the number of generation layers that we recorded for C. macropholis suggests that abrasion is not the routine method of dispersal of glandular material. We did record significant differences in cross-sectional area and height between the outer and inner layers of multi-layer generation glands of Cordylus macropholis, but this is more likely attributable to ‘desiccation’ of the outer layer, rather than abrasion. Although we found signs of abrasion in cordylid generation glands, these appear minor. Evidence seems to suggest that a substantial part of the mature glandular generations remains intact and that probably only a liquid/semi-liquid component is dispersed to the environment. More research is, however, needed to confirm this.

A new microendemic frog species of the genus Blommersia (Anura: Mantellidae) from the east coast of Madagascar

Field surveys in northeastern Madagascar have revealed the existence of a new Blommersia frog species (Anura: Mantellidae), populations of which were recorded within a 30 km radius of the town of Maroantsetra. We combined morphological, bioacoustic and molecular techniques and show that it is indeed a distinct evolutionary lineage which we describe as Blommersia variabilis sp. nov. from the type locality Ambodivoahangy near Maroantsetra. This new species is morphologically most similar to B. wittei by the presence of vomerine teeth and relatively small, well-delimited femoral glands, but differs from that species by advertisement call consisting of only 2–6 notes of comparatively longer duration, and a wider separation of femoral glands in males. It is associated with dense secondary vegetation fringing lentic water bodies. Anecdotal evidence suggests its life history is similar to other congeners. The restricted range of this species implies that it is microendemic, being possibly confined to the Antainambalana watershed. Mitochondrial and nuclear DNA variation show that its closest known relatives are Blommersia galani and B. dejongi, both of which also have restricted ranges on the east coast. A putative hybrid between a B. galani female and B. dejongi male with intermediate morphology was identified based on nuclear and mitochondrial DNA variation.

Review of the genus Petropedetes Reichenow, 1874 in Central Africa with the description of three new species (Amphibia: Anura: Petropedetidae)

We review and summarize the present knowledge on Central African Petropedetes, discuss their taxonomy and describe three new species. We synonymise one name: P. newtonii is a junior synonym of P. johnstoni. Frogs from the African mainland, namely Cameroon and eastern Nigeria, assigned to “P. newtonii” in more recent literature, were misidentified. They are herein described anew as Petropedetes vulpiae sp. nov. Two further new taxa are morphologically similar to other described species. Petropedetes euskircheni sp. nov. is morphologically close to P. parkeri but differs foremost in the smaller size of the femoral glands in adult males. P. juliawurstnerae sp. nov. is morphologically similar to P. perreti and P. cameronensis. It can be distinguished from P. perreti by its less developed webbing and from P. cameronensis by the size of its tympanum and the occurrence of a tympanic papilla in males. A key to adult males of the currently known Central African Petropedetes species is given and an outlook on potential further new taxa is provided.

A new species of Nyctibatrachus Boulenger (Amphibia: Anura: Nyctibatrachidae) from the surroundings of Bhadra Wildlife Sanctuary, Western Ghats, India

A new species of Indian nyctibatrachid frog, Nyctibatrachus dattatreyaensis sp. nov. (Amphibia: Anura), is described from the montane Shola forests of Dattatreya Peeta, Bhadra Wildlife sanctuary, Karnataka, southwestern India, on the basis of unique combination of characters: head wider than long; skin on the dorsum highly wrinkled with transverse corrugated folds; three discontinuous longitudinal folds, one dorsolaterally and two laterally; webbing on toes medium (3/ 4 th ); two yellowish bands on the dorsolateral area, prominent from sub-adult to adult stage; femoral glands present. The largest specimen in the type series has a snout-vent length of 41.0 mm. A revised key to the species of Nyctibatrachus is provided.