The phylogeography of Middle Eastern tree frogs in Israel.

Western Palearctic treefrogs of the genus Hyla provide an example of a morphologically and ecologically cryptic group. Up to three distinct Hyla species have been proposed as resident in Israel and this number has consistently been subject to taxonomical debates. Here, we analyzed 16S rRNA and COI gene fragments of 658 individuals sampled at 47 pools in nine regions across Israel and the West Bank in order to resolve the taxonomic status of Hyla frogs. We generated both Bayesian and Maximum Likelihood phylogenies, and constructed time-calibrated trees to provide an evolutionary and historical context of sequence variations. We further applied SAMOVA as well as Monmonier's maximum-difference algorithm to study the genetic structure among populations and to identify potential zones acting as barriers to gene flow across locations. Our results revealed two distinct haplogroups for each gene fragment, with 95% CI divergence times dated from 8.9-17.1 Mya (16S) and 7.1-23.6 Mya (COI), respectively. SAMOVA and barrier analyses partitioned the populations into three groups. Our results highlight that, while there are probably only two Hyla species in Israel, one population of one of the species might qualify as a separate evolutionarily significant unit. Our findings elucidate the taxonomic status of Hyla frogs in Israel and provide the basis for determining appropriate management and conservation priorities.

Genetic and behavioural factors affecting interpopulation colour pattern variation in two congeneric chameleon species.

We conducted a study on interpopulation variation of colour patterns in two congeneric chameleon species, which have an analogous life history. Both species are able to rapidly change colour pattern, and their context-dependent colour patterns often vary across a wide geographical range. Specifically, we tested four hypotheses that can explain the observed interpopulation variation of colour patterns by a series of behavioural field trials where the colour patterns of individuals were recorded and later analysed by a deep neural network algorithm. We used redundancy analysis to relate genetic, spectral and behavioural predictors to interpopulation colour pattern distance. Our results showed that both isolation by distance (IBD) and alternative mating tactics were significant predictors for interpopulation colour pattern variation in Chamaeleo chamaeleon males. By contrast, in Chamaeleo dilepis, the interpopulation colour pattern variation was largely explained by IBD, and evidence for alternative mating tactics was absent. In both chameleon species, the environmental colours showed no evidence of influencing chameleon interpopulation colour pattern variation, regardless of sex or behavioural context. This contrasting finding suggests that interpopulation context-dependent colour pattern variations in each species are maintained under a different set of selective pressures or circumstances.

The interaction between cortisol and testosterone predicts leadership within rock hyrax social groups.

Group movement leadership is associated with higher risks for those in the front. Leaders are the first to explore new areas and may be exposed to predation. Individual differences in risk-taking behavior may be related to hormonal differences. In challenging circumstances, such as risk-taking leadership that may pose a cost to the leader, cortisol is secreted both to increase the likelihood of survival by restoring homeostasis, and to mediate cooperative behavior. Testosterone too has a well-established role in risk-taking behavior, and the dual-hormone hypothesis posits that the interaction of testosterone and cortisol can predict social behavior. Based on the dual-hormone hypothesis, we investigated here whether the interaction between testosterone and cortisol can predict risk-taking leadership behavior in wild rock hyraxes (Procavia capensis). We used proximity loggers, observations, and playback trials to characterize hyrax leaders in three different leadership contexts that varied in their risk levels. In support of the dual-hormone hypothesis, we found that cortisol and testosterone interactions predict leadership that involves risk. Across different circumstances that involved low or high levels of risk, testosterone was positively related to leadership, but only in individuals (both males and females) with low levels of cortisol. We also found an interaction between these hormone levels and age at the low-risk scenarios. We suggest that the close social interactions and affiliative behavior among hyrax females within small egalitarian groups may make female leadership less risky, and therefore less stressful, and allow female leaders to influence group activities.

Sub-zero temperatures and large-scale weather patterns induce tooth damage in Icelandic arctic foxes.

Tooth damage in carnivores can reflect shifts in both diet and feeding habits, and in large carnivores, it is associated with increased bone consumption. Variation in tooth condition in Icelandic arctic foxes, a mesocarnivore, was recorded from 854 individual foxes spanning 29 years. We hypothesized that annual climatic variations, which can influence food abundance and accessibility, will influence tooth condition by causing dietary shifts toward less edible prey. We examined tooth condition in relation to four climatic predictors: mean annual winter temperature, indices of both the El Niño anomaly and North Atlantic subpolar gyre (SPG), and the number of rain-on-snow days (ROS). We found unequivocal evidence for a strong effect of annual climate on tooth condition. Teeth of Icelandic foxes were in better condition when winter temperatures were higher, when the SPG was more positive, and when the number of ROS was low. We also found a substantial subregional effect with foxes from northeastern Iceland having lower tooth damage than those from two western sites. Contradicting our original hypothesis that foxes from northeastern Iceland, where foxes are known to scavenge on large mammal remains (e.g., sheep and horses), would show the highest tooth damage, we suggest that western coastal sites exhibited greater tooth damage because cold winter temperatures lowered the availability of seabirds, causing a shift in diet toward abrasive marine subsidies (e.g., bivalves) and frozen beach wrack. Our study shows that monitoring tooth breakage and wear can be a useful tool for evaluating the impact of climate on carnivore populations and that climate change may influence the condition and fitness of carnivores in complex and potentially conflicting ways.

Testing the morphological constraint hypothesis of tail length in the sexually dimorphic Cerastes vipera and new perspectives.

The morphological constraint hypothesis (MCH) states that, in snakes, males typically have relatively longer tails than females to accommodate the hemipenes and retractor muscles. To date, most studies testing the MCH have been interspecific and results have been equivocal. We tested the MCH intraspecifically on Cerastes vipera, a species with a relatively short tail and suitable for testing the MCH. The relative tail length and length of the hemipenes pocket in Cerastes vipera were measured in preserved museum-maintained males (n = 35) and in free-ranging males and females (n = 277). Males exhibited relatively longer tails than females, which was explained fully by the length of the hemipenes pocket. The relatively short tail of C. vipera presents a constraint to the reproductive structures in males, as the length of the hemipenes pocket occupies a greater proportion in shorter- than longer-tailed individuals. This is the first report presenting these intraspecific findings in support of the MCH. Whether these relations are widespread among snake families, within Viperidae, or specifically within C. vipera warrants further studies.

Both selection and drift drive the spatial pattern of adaptive genetic variation in a wild mammal.

The major histocompatibility complex (MHC) has been intensively studied for the relative effects of different evolutionary forces in recent decades. Pathogen-mediated balancing selection is generally thought to explain the high polymorphism observed in MHC genes, but it is still unclear to what extent MHC diversity is shaped by selection relative to neutral drift. In this study, we genotyped MHC class II DRB genes and 15 neutral microsatellite loci across 26 geographic populations of European badgers (Meles meles) covering most of their geographic range. By comparing variation of microsatellite and diversity of MHC at different levels, we demonstrate that both balancing selection and drift have shaped the evolution of MHC genes. When only MHC allelic identity was investigated, the spatial pattern of MHC variation was similar to that of microsatellites. By contrast, when functional aspects of the MHC diversity (e.g., immunological supertypes) were considered, balancing selection appears to decrease genetic structuring across populations. Our comprehensive sampling and analytical approach enable us to conclude that the likely mechanisms of selection are heterozygote advantage and/or rare-allele advantage. This study is a clear demonstration of how both balancing selection and genetic drift simultaneously affect the evolution of MHC genes in a widely distributed wild mammal.

Male rock hyraxes that maintain an isochronous song rhythm achieve higher reproductive success.

Rhythmic stability (nonrandom temporal structure) is required for many neural and physiological functions, whereas rhythmic irregularities can indicate genetic or developmental deficiencies. Therefore, rhythmic courtship or contest signals are widespread in nature as honest advertisement displays. Examination of bird songs revealed the pervasiveness of categorical rhythmic patterns that can be described as small integer ratios between sequential inter-call intervals. As similar rhythmic profiles are prevalent in human music, it was suggested that a shared functionality could drive both animal songs and human musical rhythms, facilitating synchrony between signallers and enabling easy identification of performance errors. Here we examined whether the rhythmic structure and the rhythmic stability of vocal displays are related to reproductive success in male rock hyraxes (Procavia capensis), which presents an unusual case of a terrestrial singing mammal. We combined long-term parentage analysis of 13 male hyraxes (22 male/years) with an analysis of an audio library of 105 hyrax songs. Male annual reproductive success was determined by the number of offspring that survived to the age of 1 year. The frequency of singing events was used to determine the seasonal singing effort for each male. Songs were analysed for rhythmic structure, focusing on the presence of categorical rhythms and the contribution of rhythmic stability to annual reproductive success. We found that male hyraxes that sing more frequently tend to have more surviving offspring and that the rhythmic profile of hyrax songs is predominantly isochronous with sequential vocal element pairs nearly equally spaced. The ratio of isochronous vocal element transitions (on-integer) to element transitions that deviate from an isochronous pattern (off-integer) in hyrax songs is positively correlated with male reproductive success. Our findings support the notion that isochronous rhythmic stability can serve as an indication of quality in sexually selected signals and is not necessarily driven by the need for multiple caller synchronization. The relative scarcity of nonisochronous rhythmic categories in individually performed hyrax songs raises the question of whether such rhythmic categories could be a product of collective, coordinated signalling, while being selected against in individual performance.

Sex-associated and context-dependent leadership in the rock hyrax.

In many mammalian species, both sexes may take leadership role, but different traits may play a role in determining variation within species. Here we examine the effect of sex on leadership. We present three complementary datasets derived from a well-studied population of wild rock hyrax (Procavia capensis). The findings demonstrated that male and female rock hyraxes take on different leadership positions, depending on the context. When risk is moderate, more likely to lead are younger resident males, which experience high cortisol and lower testosterone levels. However, during acute predation scenarios, more likely to lead are males with lower centrality status. We suggest that hyrax males exhibit risky behaviors that may reflect their need for self-advertisement. In contrast, leadership among group females is more equally distributed. Females have little to gain from risky actions due to the lack of competition among them, but nonetheless take leadership positions.

Annual climatic fluctuations and short-term genetic variation in the eastern spadefoot toad.

In addition to variations on the spatial scale, short- and long-term temporal variations, too, can impose intense selection on the overall genetic diversity and composition of a population. We hypothesized that the allelic composition in populations of the eastern spadefoot toad (Pelobates syriacus) would change among successive years in accordance with the short-term changes in environmental conditions. Surprisingly, the effect of short-term climate fluctuations on genetic composition have rarely been addressed in the literature, and to our knowledge the effect of annual climatic fluctuations have not been considered meaningful. Our findings show that climatic variation among successive years, primarily the amount of rainfall and rainy days, can significantly alter both microsatellite allelic composition and diversity. We suggest that environmental (i.e. fluctuating) selection is differential across the globe, and that its intensity is expected to be greatest in regions where short-term climatic conditions are least stable.

The "Law of Brevity" in animal communication: Sex-specific signaling optimization is determined by call amplitude rather than duration.

The efficiency of informational transfer is one of the key aspects of any communication system. The informational coding economy of human languages is often demonstrated by their almost universal fit to Zipf's "Law of Brevity," expressing negative relationship between word length and its usage frequency. Animal vocal systems, however, provided mixed results in their adherence to this relationship, potentially due to conflicting evolutionary pressures related to differences in signaling range and communicational needs. To examine this potential parallel between human and animal vocal communication, and also to explore how divergent, sex-specific, communicational settings affect signaling efficiency within a species, we examined the complete vocal repertoire of rock hyraxes (Procavia capensis). As male and female hyraxes differ in their sociality levels and male hyraxes vocal repertoire is dominated by sexual advertisement songs, we hypothesized that sex-specific vocal repertoires could be subjected to different signaling optimization pressures. Our results show that the sexes differ in repertoire size, call usage, and adherence to coding efficiency principles. Interestingly, the classic call length/call usage relationship is not consistently found in rock hyraxes. Rather, a negative relationship between call amplitude and call usage is found, suggesting that the efficiency of the vocal repertoire is driven by call amplitude rather than duration. We hypothesize that, in contrast to human speech that is mainly intended for short distance, the need for frequent long-range signaling shapes an animal's vocal repertoire efficiency according to the cost of call amplitude rather than call length. However, call duration may be a secondary factor affecting signaling efficiency, in cases where amplitude is under specific selection pressures, such as sexual selection.

Phylogeography of a cryptic speciation continuum in Eurasian spadefoot toads (Pelobates).

Cryptic phylogeographic diversifications provide unique models to examine the role of phylogenetic divergence on the evolution of reproductive isolation, without extrinsic factors such as ecological and behavioural differentiation. Yet, to date very few comparative studies have been attempted within such radiations. Here, we characterize a new speciation continuum in a group of widespread Eurasian amphibians, the Pelobates spadefoot toads, by conducting multilocus (restriction site associated DNA sequencing and mitochondrial DNA) phylogenetic, phylogeographic and hybrid zone analyses. Within the P. syriacus complex, we discovered species-level cryptic divergences (>5 million years ago [My]) between populations distributed in the Near-East (hereafter P. syriacus sensu stricto [s.s.]) and southeastern Europe (hereafter P. balcanicus), each featuring deep intraspecific lineages. Altogether, we could scale hybridizability to divergence time along six different stages, spanning from sympatry without gene flow (P. fuscus and P. balcanicus, >10 My), parapatry with highly restricted hybridization (P. balcanicus and P. syriacus s.s., >5 My), narrow hybrid zones (~15 km) consistent with partial reproductive isolation (P. fuscus and P. vespertinus, ~3 My), to extensive admixture between Pleistocene and refugial lineages (≤2 My). This full spectrum empirically supports a gradual build up of reproductive barriers through time, reversible up until a threshold that we estimate at ~3 My. Hence, cryptic phylogeographic lineages may fade away or become reproductively isolated species simply depending on the time they persist in allopatry, and without definite ecomorphological divergence.

Social context mediates testosterone's effect on snort acoustics in male hyrax songs.

Testosterone affects physical and motivational states, both of which may strongly influence vocalization structure and acoustics. The loud complex calls (i.e., songs) of male rock hyraxes (Procavia capensis) are used as honest signals for advertising physical and social states. The snort, a low frequency, noisy element of the song, encodes information on the singer's age and social rank via harshness, as measured by jitter (i.e., acoustic frequency stability) and duration; suggesting that the snort concomitantly advertises both vocal stability and aggression. Our past findings revealed that testosterone levels are related to both vocal elements and social status of male hyraxes, suggesting that hormonal mechanisms mediate the motivation for aggressive and courtship behaviors. Here we examined whether long-term androgen levels are related to snort acoustics and song structure by comparing levels of testosterone in hair with acoustic and structural parameters. We found that songs performed by individuals with higher testosterone levels include more singing bouts and longer, smoother snorts, but only in those songs induced by external triggers. It is possible that hyraxes with higher levels of testosterone possess the ability to perform higher-quality singing, but only invest in situations of high social arousal and potential benefit. Surprisingly, in spontaneous songs, hyraxes with high testosterone were found to snort more harshly than low-testosterone males. The context dependent effects of high testosterone on snort acoustics suggest that the aggressive emotional arousal associated with testosterone is naturally reflected in the jittery hyrax snort, but that it can be masked by high-quality performance.

Population genetic analysis of the recently rediscovered Hula painted frog (Latonia nigriventer) reveals high genetic diversity and low inbreeding.

After its recent rediscovery, the Hula painted frog (Latonia nigriventer) has remained one of the world's rarest and least understood amphibian species. Together with its apparently low dispersal capability and highly disturbed niche, the low abundance of this living fossil calls for urgent conservation measures. We used 18 newly developed microsatellite loci and four different models to calculate the effective population size (Ne) of a total of 125 Hula painted frog individuals sampled at a single location. We compare the Ne estimates to the estimates of potentially reproducing adults in this population (Nad) determined through a capture-recapture study on 118 adult Hula painted frogs captured at the same site. Surprisingly, our data suggests that, despite Nad estimates of only ~234-244 and Ne estimates of ~16.6-35.8, the species appears to maintain a very high genetic diversity (HO = 0.771) and low inbreeding coefficient (FIS = -0.018). This puzzling outcome could perhaps be explained by the hypotheses of either genetic rescue from one or more unknown Hula painted frog populations nearby or by recent admixture of genetically divergent subpopulations. Independent of which scenario is correct, the original locations of these populations still remain to be determined.

The contextual separation of lateral white line patterns in chameleons.

While many animals display different colour patterns that signal different messages, some species use various tactics to separate between colour and pattern displays. The common chameleon (Chamaeleo chamaeleon) is capable of rapidly changing and separating among displays of colour patterns and ornaments. We used chameleons to study the contextual role of separation among colour and pattern displays. Specifically, we studied the predominant white badge, which is composed of multiple parts, during different seasons and in different social contexts. We hypothesized that the badge contains important information about the sender and, therefore, would be present during important social contexts. We carried out a series of trials to document the presence/absence of the badge and found that the badge is individually specific and reflects body size. We also revealed that the badge remained fixed throughout other body colour changes, but was replaced by other colour patterns during mating behaviour. During social encounters, additional dark patches delineating the badge appeared, presumably amplifying its signal. Thus, we suggest that the badge constitutes an important feature in intraspecific communication, and is possibly employed to display quality. However, the replacement of the badge by other displays during courtship suggests that during important social events like mating, chameleons transmit exclusive information that is not broadcast by the badge. Our findings demonstrate the importance of separation between colour patterns, and the alternative use of intraspecific colour patterns for specific social contexts in chameleons.

Living quarters of a living fossil-Uncovering the current distribution pattern of the rediscovered Hula painted frog (Latonia nigriventer) using environmental DNA.

One of the greatest challenges of effective conservation measures is the correct identification of sites where rare and elusive organisms reside. The recently rediscovered Hula painted frog (Latonia nigriventer) has not been seen for many decades and was therefore categorized extinct. Since its rediscovery in 2011, individuals from the critically endangered species have been found, with great effort, only in four restricted sites. We applied the environmental DNA (eDNA) approach to search for new populations of the Hula painted frog in suitable aquatic habitats. We further used the eDNA data to classify the landscape factors associated with the species distribution and to predict its suitable habitats. We sampled 52 aquatic sites in the Hula Valley during the spring of 2015 and 2016 and amplified the samples with a species-specific qPCR assay. DNA of the Hula painted frog was detected in 22 of the sites, all of which clustered within three main areas. A boosting classification model showed that soil type, vegetation cover and the current and former habitats are all key predictors of the frog's current distribution. Intriguingly, the habitat suitability models reveal a high affinity of the species to its long-lost habitat of the historical wetlands. Our findings encourage a series of informed searches for new populations of this threatened frog and provide guidance for future conservation management programmes. In the era of global conservation crisis of amphibians, developing the eDNA approach, a reliable detection method for many critically endangered and elusive amphibians, is of particular importance.

The progression pattern of male hyrax songs and the role of climactic ending.

The study of animal vocal signals can either focus on the properties of distinct vocal elements or address the signal as a whole. Although some attention has been given to the continuous progression patterns of bird songs, such patterns in mammalian vocalisations have been largely overlooked. We examined temporal changes in structural and acoustic parameters in male rock hyrax songs. We found a gradual increase in call frequency and amplitude towards the song ending, as well as an abrupt increase in bout syntactic complexity, peaking in the last quintile of a song. In musical terms, such a pattern can be described as a crescendo (amplitude increase) with a terminal climax. In Western music, crescendos are used to maintain attention and direct the listeners towards a memorable highpoint of the musical piece. This structure may have an analogous function in animal communication, recruiting audience attention towards the climactic and potentially most informative part of the signal. Our playback experiments revealed that hyrax males tend to reply more to songs with a climactic ending, indicating that this progression pattern is important for hyrax communication. We suggest that animal vocal communication research can benefit from adding musical concepts to the analysis toolbox.

The plant component of an Acheulian diet at Gesher Benot Ya'aqov, Israel.

Diet is central for understanding hominin evolution, adaptation, and environmental exploitation, but Paleolithic plant remains are scarce. A unique macrobotanical assemblage of 55 food plant taxa from the Acheulian site of Gesher Benot Ya'aqov, Israel includes seeds, fruits, nuts, vegetables, and plants producing underground storage organs. The food plant remains were part of a diet that also included aquatic and terrestrial fauna. This diverse assemblage, 780,000 y old, reflects a varied plant diet, staple plant foods, environmental knowledge, seasonality, and the use of fire in food processing. It provides insight into the wide spectrum of the diet of mid-Pleistocene hominins, enhancing our understanding of their adaptation from the perspective of subsistence. Our results shed light on hominin abilities to adjust to new environments, facilitating population diffusion and colonization beyond Africa. We reconstruct the major vegetal foodstuffs, while considering the possibility of some detoxification by fire. The site, located in the Levantine Corridor through which several hominin waves dispersed out of Africa, provides a unique opportunity to study mid-Pleistocene vegetal diet and is crucial for understanding subsistence aspects of hominin dispersal and the transition from an African-based to a Eurasian diet.

Molecular assessment of the phylogeny and biogeography of a recently diversified endemic group of South American canids (Mammalia: Carnivora: Canidae).

To investigate the evolution and biogeography of an endemic group of South American foxes, we examined mitochondrial DNA control region sequences for 118 individuals belonging to all six extant species of the genus Lycalopex. Phylogenetic and molecular dating analyses supported the inference that this genus has undergone a very recent and rapid radiation, stemming from a common ancestor that lived ca. 1 million years ago. The Brazilian endemic L. vetulus was supported as the most basal species in this genus, whereas the most internal group is comprised by the recently diverged (ca. 350,000 years ago) Andean/Patagonian species L. griseus and L. culpaeus. We discuss the inferred phylogenetic relationships and divergence times in the context of the current geographic distributions of these species, and the likely effects of Pleistocene climatic changes on the biogeography of this group. Furthermore, a remarkable finding was the identification of multiple individuals classified as L. gymnocercus bearing mtDNA haplotypes clearly belonging to L. griseus, sampled in regions where the latter is not known to occur. At a minimum, this result implies the need to clarify the present-day geographic distribution of each of these fox species, while it may also indicate an ongoing hybridization process between them. Future testing of this hypothesis with in-depth analyses of these populations is thus a priority for understanding the history, evolutionary dynamics and present-day composition of this endemic Neotropical genus.

Alternative Mating Tactics in Male Chameleons (Chamaeleo chamaeleon) Are Evident in Both Long-Term Body Color and Short-Term Courtship Pattern.

Alternative mating tactics in males of various taxa are associated with body color, body size, and social status. Chameleons are known for their ability to change body color following immediate environmental or social stimuli. In this study, we examined whether the differential appearance of male common chameleon during the breeding season is indeed an expression of alternative mating tactics. We documented body color of males and used computer vision techniques to classify images of individuals into discrete color patterns associated with seasons, individual characteristics, and social contexts. Our findings revealed no differences in body color and color patterns among males during the non-breeding season. However, during the breeding season males appeared in several color displays, which reflected body size, social status, and behavioral patterns. Furthermore, smaller and younger males resembled the appearance of small females. Consequently, we suggest that long-term color change in males during the breeding season reflects male alternative mating tactics. Upon encounter with a receptive female, males rapidly alter their appearance to that of a specific brief courtship display, which reflects their social status. The females, however, copulated indiscriminately in respect to male color patterns. Thus, we suggest that the differential color patterns displayed by males during the breeding season are largely aimed at inter-male signaling.

Effects of parasite pressure on parasite mortality and reproductive output in a rodent-flea system: inferring host defense trade-offs.

Evaluating host resistance via parasite fitness helps place host-parasite relationships within evolutionary and ecological contexts; however, few studies consider both these processes simultaneously. We investigated how different levels of parasite pressure affect parasite mortality and reproductive success in relationship to host defense efforts, using the rodent Gerbillus nanus and the flea Xenopsylla conformis as a host-parasite system. Fifteen immune-naïve male rodents were infested with 20, 50, or 100 fleas for four weeks. During this time number of new imagoes produced per adult flea (our flea reproductive output metric), flea mortality, and change in circulating anti-flea immunoglobulin G (our measure of adaptive immune defense) were monitored. Three hypotheses guided this work: (1) increasing parasite pressure would heighten host defenses; (2) parasite mortality would increase and parasite reproductive output would decrease with increasing investment in host defense; and (3) hosts under high parasite pressure could invest in behavioral and/or immune responses. We predicted that at high infestation levels (a) parasite mortality would increase; (b) flea reproductive output per individual would decrease; and (c) host circulating anti-flea antibody levels would increase. The hypotheses were partially supported. Flea mortality significantly increased and flea reproductive output significantly decreased as flea pressure increased. Host adaptive immune defense did not significantly change with increasing flea pressure. Therefore, we inferred that investment in host behavioral defense, either alone or in combination with density-dependent effects, may be more efficient at increasing flea mortality and decreasing flea reproductive output than antibody production during initial infestation in this system.