Population genetics and phylogeographic history of the insular lizard Podarcis lilfordi (Gunther, 1874) from the Balearic Islands based on genome-wide polymorphic data.

Islands provide a great system to explore the processes that maintain genetic diversity and promote local adaptation. We explored the genomic diversity of the Balearic lizard Podarcis lilfordi, an endemic species characterized by numerous small insular populations with large phenotypic diversity. Using the newly available genome for this species, we characterized more than 300,000 SNPs, merging genotyping-by-sequencing (GBS) data with previously published restriction site-associated DNA sequencing (RAD-Seq) data, providing a dataset of 16 island populations (191 individuals) across the range of species distribution (Menorca, Mallorca, and Cabrera). Results indicate that each islet hosts a well-differentiated population (F ST = 0.247 ± 0.09), with no recent immigration/translocation events. Contrary to expectations, most populations harbor a considerable genetic diversity (mean nucleotide diversity, P i = 0.144 ± 0.021), characterized by overall low inbreeding values (F IS < 0.1). While the genetic diversity significantly decreased with decreasing islet surface, maintenance of substantial genetic diversity even in tiny islets suggests variable selection or other mechanisms that buffer genetic drift. Maximum-likelihood tree based on concatenated SNP data confirmed the existence of the two major independent lineages of Menorca and Mallorca/Cabrera. Multiple lines of evidence, including admixture and root testing, robustly placed the origin of the species in the Mallorca Island, rather than in Menorca. Outlier analysis mainly retrieved a strong signature of genome differentiation between the two major archipelagos, especially in the sexual chromosome Z. A set of proteins were target of multiple outliers and primarily associated with binding and catalytic activity, providing interesting candidates for future selection studies. This study provides the framework to explore crucial aspects of the genetic basis of phenotypic divergence and insular adaptation.

Joint estimation of survival and dispersal effectively corrects the permanent emigration bias in mark-recapture analyses.

Robust and reliable estimates of demographic parameters are essential to understand population dynamics. Natal dispersal is a common process in monitored populations and can cause underestimations of survival and dispersal due to permanent emigration. Here, we present a multistate Bayesian capture-mark-recapture approach based on a joint estimation of natal dispersal kernel and detection probabilities to address biases in survival, dispersal, and related demographic parameters when dispersal information is limited. We implement this approach to long-term data of a threatened population: the Bonelli's eagle in Catalonia (SW Europe). To assess the method's performance, we compare demographic estimates structured by sex, age, and breeding status in cases of limited versus large data scales, with those of classical models where dispersal and detection probabilities are estimated separately. Results show substantial corrections of demographic estimates. Natal dispersal and permanent emigration probabilities were larger in females, and consequently, female non-breeder survival showed larger differences between separate and joint estimation models. Moreover, our results suggest that estimates are sensitive to the choice of the dispersal kernel, fat-tailed kernels providing larger values in cases of data limitation. This study provides a general multistate framework to model demographic parameters while correcting permanent emigration biases caused by natal dispersal.

Insular holobionts: persistence and seasonal plasticity of the Balearic wall lizard (Podarcis lilfordi) gut microbiota.

Background: Integrative studies of animals and associated microbial assemblages (i.e., the holobiont) are rapidly changing our perspectives on organismal ecology and evolution. Insular vertebrates provide ideal natural systems to understand patterns of host-gut microbiota coevolution, the resilience and plasticity these microbial communities over temporal and spatial scales, and ultimately their role in the host ecological adaptation. Methods: Here we used the endemic Balearic wall lizard Podarcis lilfordi to dissect the drivers of the microbial diversity within and across host allopatric populations/islets. By focusing on three extensively studied populations/islets of Mallorca (Spain) and fecal sampling from individually identified lizards along two years (both in spring and autumn), we sorted out the effect of islet, sex, life stage, year and season on the microbiota composition. We further related microbiota diversity to host genetics, trophic ecology and expected annual metabolic changes. Results: All the three populations showed a remarkable conservation of the major microbial taxonomic profile, while carrying their unique microbial signature at finer level of taxonomic resolution (Amplicon Sequence Variants (ASVs)). Microbiota distances across populations were compatible with both host genetics (based on microsatellites) and trophic niche distances (based on stable isotopes and fecal content). Within populations, a large proportion of ASVs (30-50%) were recurrently found along the four sampling dates. The microbial diversity was strongly marked by seasonality, with no sex effect and a marginal life stage and annual effect. The microbiota showed seasonal fluctuations along the two sampled years, primarily due to changes in the relative abundances of fermentative bacteria (mostly families Lachnospiraceae and Ruminococcaceae), without any major compositional turnover. Conclusions: These results support a large resilience of the major compositional aspects of the P. lilfordi gut microbiota over the short-term evolutionary divergence of their host allopatric populations (<10,000 years), but also indicate an undergoing process of parallel diversification of the both host and associated gut microbes. Predictable seasonal dynamics in microbiota diversity suggests a role of microbiota plasticity in the lizards' metabolic adaptation to their resource-constrained insular environments. Overall, our study supports the need for longitudinal and integrative studies of host and associated microbes in natural systems.

Seafloor litter sorting in different domains of Cap de Creus continental shelf and submarine canyon (NW Mediterranean Sea).

We analyzed litter occurrence in 68 underwater video transects performed on the middle/outer continental shelf and submarine canyon off Cap de Creus (NW Mediterranean), an area recently declared Site of Community Importance (SCI). Low densities of urban litter were registered on the shelf (7.2 items ha-1), increasing in abundance towards the deepest part of the submarine canyon, with 188 items ha-1 below 1000 m depth. We hypothesize that the strong bottom currents that recurrently affect this area efficiently move litter objects from the shelf towards the deep. Of all litter items, approximately 50% had a fishing-related origin, mostly longlines entangled on rocks in the canyon head and discarded trawl nets in deeper areas. Over 10% of cold-water colonies observed had longlines entangled, indicating the harmful effects of such practices over benthic habitats. These results should be considered when designing mitigation measures to reduce litter pollution in Cap de Creus SCI.

Microbial co-occurrence networks of gut microbiota reveal community conservation and diet-associated shifts in cichlid fishes.

BACKGROUND: The extent to which deterministic rather than stochastic processes guide gut bacteria co-existence and ultimately their assembling into a community remains largely unknown. Co-occurrence networks of bacterial associations offer a powerful approach to begin exploring gut microbial community structure, maintenance and dynamics, beyond compositional aspects alone. Here we used an iconic model system, the cichlid fishes, with their multiple lake assemblages and extraordinary ecological diversity, to investigate a) patterns of microbial associations that were robust to major phylogeographical variables, and b) changes in microbial network structure along dietary shifts. We tackled these objectives using the large gut microbiota sequencing dataset available (nine lakes from Africa and America), building geographical and diet-specific networks and performing comparative network analyses. RESULTS: Major findings indicated that lake and continental microbial networks were highly resembling in global topology and node taxonomic composition, despite the heterogeneity of the samples. A small fraction of the observed co-occurrences among operational taxonomic units (OTUs) was conserved across all lake assemblages. These were all positive associations and involved OTUs within the genera Cetobacterium and Turicibacter and several OTUs belonging to the families of Peptostreptococcaceae and Clostridiaceae (order Clostridiales). Mapping of diet contribution on the African Lake Tanganyika network (therefore excluding the geographic variable) revealed a clear community change from carnivores (C) to omnivores (O) to herbivores (H). Node abundances and effect size for pairwise comparisons between diets supported a strong contrasting pattern between C and H. Moreover, diet-associated nodes in H formed complex modules of positive interactions among taxonomically diverse bacteria (mostly Verrucomicrobia and Proteobacteria). CONCLUSIONS: Conservation of microbial network topologies and specific bacterial associations across distinct lake assemblages point to a major host-associated effect and potential deterministic processes shaping the cichlid gut microbiota. While the origin and biological relevance of these common associations remain unclear, their persistence suggests an important functional role in the cichlid gut. Among the very diverse cichlids of L. Tanganyika, diet nonetheless represents a major driver of microbial community changes. By intersecting results from predictive network inferences and experimental trials, future studies will be directed to explore the strength of these associations, predict the outcome of community alterations driven by diet and ultimately help understanding the role of gut microbiota in cichlid trophic diversification.

Phylogeography and Ecological Niche Shape the Cichlid Fish Gut Microbiota in Central American and African Lakes.

Cichlid fishes, with their repeated colonization of lakes and subsequent radiations at different scales of phylogenetic and ecological diversification, offer an excellent model system to understand the factors shaping the host-gut microbiota association in nature. Here, we characterized the gut microbiota of the Amphilophus species complex from Central America (known as the Midas cichlid complex), encompassing 158 wild specimens (13 species) collected from seven Nicaraguan lakes, and combined these data with previously published data from two African lakes (spanning 29 species). Our aim was to comprehensively explore trends in microbiota variation and persistence along the large spatial and temporal scales of cichlid diversification (from the oldest radiation in L. Tanganyika, 9-12 My old, to young ones in Nicaraguan crater lakes, <0.5 My old), in allopatry and sympatry (within and across lakes), and across the range of dietary niches (from highly specialized to generalist feeders). Despite their extraordinary diversity, cichlids shared a remarkably conserved microbial taxonomic profile, which argues for a primary role of the host genetics in the assembly and maintenance of these microbial communities. Within this partly constrained microbiota profile, geographic isolation (continent and lake) represented the first level of discrimination. For the Midas cichlid, a partial congruency was found between host microbiota and genetic distances, suggesting that microbial communities have partly diversified along their cichlid phylogeographic history of crater lake colonization. In sympatry (within lakes), the young and poorly ecologically diversified cichlid assemblages of Central American lakes display largely unresolved gut microbiotas (in terms of both alpha and beta diversities), whereas the phylogenetically and ecologically diverse species found in African lakes showed greater microbial interspecific diversity. This pattern largely points to the level of habitat segregation, trophic niche overlap, and reproductive barriers as major modulators of the gut microbiota connectivity among sympatric species.

The optimal sampling design for littoral habitats modelling: A case study from the north-western Mediterranean.

Species distribution models (SDMs) have been used to predict potential distributions of habitats and to model the effects of environmental changes. Despite their usefulness, currently there is no standardized sampling strategy that provides suitable and sufficiently representative predictive models for littoral marine benthic habitats. Here we aim to establish the best performing and most cost-effective sample design to predict the distribution of littoral habitats in unexplored areas. We also study how environmental variability, sample size, and habitat prevalence may influence the accuracy and performance of spatial predictions. For first time, a large database of littoral habitats (16,098 points over 562,895 km of coastline) is used to build up, evaluate, and validate logistic predictive models according to a variety of sampling strategies. A regularly interspaced strategy with a sample of 20% of the coastline provided the best compromise between usefulness (in terms of sampling cost and effort) and accuracy. However, model performance was strongly depen upon habitat characteristics. The proposed sampling strategy may help to predict the presence or absence of target species or habitats thus improving extensive cartographies, detect high biodiversity areas, and, lastly, develop (the best) environmental management plans, especially in littoral environments.

Effective dispersal and density-dependence in mesophotic macroalgal forests: Insights from the Mediterranean species Cystoseira zosteroides.

Dispersal and recruitment are fundamental processes for population recovery following disturbances in sessile species. While both processes are well understood for many terrestrial species, they still remain poorly resolved for some macroalgal species. Here we experimentally investigated the effective dispersal and recruit survival of a mesophotic Mediterranean fucoid, Cystoseira zosteroides. In three isolated populations, four sets of settlement collectors were placed at increasing distances (from 0 to 10 m) and different orientations (North, South, East and West). We observed that effective dispersal was restricted to populations' vicinity, with an average of 6.43 m and not further than 13.33 m, following a Weibull distribution. During their first year of life, survival was up to 50%, but it was lower underneath the adult canopy, suggesting a negative density-dependence. To put our results in a broader context we compared the effective dispersal of other fucoid and kelp species reported in the literature, which confirmed the low dispersal ability of brown algae, in particular for fucoids, with an effective dispersal of few meters. Given the importance of recruitment for the persistence and recovery of populations after disturbances, these results underline the vulnerability of C. zosteroides and other fucoid species to escalating threats.

Processes shaping gut microbiota diversity in allopatric populations of the endemic lizard Podarcis lilfordi from Menorcan islets (Balearic Islands).

Compositional variation of the gut microbiota across host allopatric populations can reflect both adaptation and stochasticity since the time of separation. Major factors shaping this variation include the host phylogeographic and demographic history, the microbiota inheritance, environmental inputs and dispersal of bacteria. Here we explored the impact of these factors in driving gut community diversity in seven allopatric populations of the omnivorous lizard Podarcis lilfordi from the Menorcan coastal islets, all descending from an ancestral mainland population. Using 16S rRNA Illumina sequencing, we showed that 'islet' and 'age' (time since islet separation from mainland) were the only significant variables in microbial community clustering, suggesting a partial islet-restricted diversification following these lizards phylogeography. Despite a significant variation, islets/populations were characterized by a remarkably low bacterial uniqueness (2.4% of total OTUs) and a minor differential enrichment of taxa, indicating a negligible impact of local inputs and important host common constraints. Overall, the extant pattern of similarity/dissimilarity among islets is compatible with partial retention of the ancestral mainland microbial pool, with differences among islets potentially explained by a differential loss of bacteria following population fragmentation and bottlenecks (i.e. ecological drift). While more quantitative data are needed to validate this hypothesis, this study unveils the importance of considering both neutral and niche-driven processes in driving contemporary patterns of gut metacommunity diversity.

Microeukaryote community in a partial nitritation reactor prior to anammox and an insight into the potential of ciliates as performance bioindicators.

An in-depth, long-term, multidisciplinary study was conducted in order to study the microeukaryote community in a partial nitritation (PN) reactor prior to anammox. The PN reactor operated with moving bed biofilm reactor (MBBR) technology, using plastic supports (carriers) for biofilm development. The microeukaryote community from the biofilm (BF) and the surrounding media (mixed liquor or ML) were analysed separately. Despite the physicochemical conditions under which the PN-MBBR operated (an average of 305.9±117mg TAN l-1 and 328.4±131.9mg N-NO2- l-1), up to 24 microeukaryotic taxa were observed by microscope. Microeukaryote species showed an uneven distribution in the PN-MBBR, thus suggesting the existence of two habitats: the BF, preferred by species with specific structures for adhering to a substrate, such as the stalked Peritrichia, and the ML, preferred by free-swimming or non-substrate dependent species. The results indicated that most ciliate population dynamics mainly responded to the nitrous acid and free ammonia concentrations and, to a lesser extent, to sCOD values. In the BF, variations in the population of Epistylis camprubii and Opercularia coarctata suggest the existence of competition between these species due to niche overlap. A V4 18S rDNA molecular survey (Illumina) was carried out for some samples with the aim of obtaining maximum coverage of the main eukaryote species that were microscopically detected throughout the study. The diversity and abundance data provided by both detection methods were compared. The study helped identify broader tolerance ranges of the microeukaryote taxa to the physicochemical parameters analysed.

Convergence of gut microbiotas in the adaptive radiations of African cichlid fishes.

Ecoevolutionary dynamics of the gut microbiota at the macroscale level, that is, in across-species comparisons, are largely driven by ecological variables and host genotype. The repeated explosive radiations of African cichlid fishes in distinct lakes, following a dietary diversification in a context of reduced genetic diversity, provide a natural setup to explore convergence, divergence and repeatability in patterns of microbiota dynamics as a function of the host diet, phylogeny and environment. Here we characterized by 16S rRNA amplicon sequencing the gut microbiota of 29 cichlid species from two distinct lakes/radiations (Tanganyika and Barombi Mbo) and across a broad dietary and phylogenetic range. Within each lake, a significant deviation between a carnivorous and herbivorous lifestyle was found. Herbivore species were characterized by an increased bacterial taxonomic and functional diversity and converged in key compositional and functional community aspects. Despite a significant lake effect on the microbiota structure, this process has occurred with remarkable parallels in the two lakes. A metabolic signature most likely explains this trend, as indicated by a significant enrichment in herbivores/omnivores of bacterial taxa and functions associated with fiber degradation and detoxification of plant chemical compounds. Overall, compositional and functional aspects of the gut microbiota individually and altogether validate and predict main cichlid dietary habits, suggesting a fundamental role of gut bacteria in cichlid niche expansion and adaptation.

Gut Microbiota Dynamics during Dietary Shift in Eastern African Cichlid Fishes.

The gut microbiota structure reflects both a host phylogenetic history and a signature of adaptation to the host ecological, mainly trophic niches. African cichlid fishes, with their array of closely related species that underwent a rapid dietary niche radiation, offer a particularly interesting system to explore the relative contribution of these two factors in nature. Here we surveyed the host intra- and interspecific natural variation of the gut microbiota of five cichlid species from the monophyletic tribe Perissodini of lake Tanganyika, whose members transitioned from being zooplanktivorous to feeding primarily on fish scales. The outgroup riverine species Astatotilapia burtoni, largely omnivorous, was also included in the study. Fusobacteria, Firmicutes and Proteobacteria represented the dominant components in the gut microbiota of all 30 specimens analysed according to two distinct 16S rRNA markers. All members of the Perissodini tribe showed a homogenous pattern of microbial alpha and beta diversities, with no significant qualitative differences, despite changes in diet. The recent diet shift between zooplantkon- and scale-eaters simply reflects on a significant enrichment of Clostridium taxa in scale-eaters where they might be involved in the scale metabolism. Comparison with the omnivorous species A. burtoni suggests that, with increased host phylogenetic distance and/or increasing herbivory, the gut microbiota begins differentiating also at qualitative level. The cichlids show presence of a large conserved core of taxa and a small set of core OTUs (average 13-15%), remarkably stable also in captivity, and putatively favoured by both restricted microbial transmission among related hosts (putatively enhanced by mouthbrooding behavior) and common host constraints. This study sets the basis for a future large-scale investigation of the gut microbiota of cichlids and its adaptation in the process of the host adaptive radiation.

Temporal variability of nitrogen stable isotopes in primary uptake compartments in four streams differing in human impacts.

Understanding the variability of the natural abundance in nitrogen stable isotopes (expressed as δ(15)N) of primary uptake compartments (PUCs; e.g., epilithon or macrophytes) is important due to the multiple applications of stable isotopes in freshwater research and can give insights into environmental and anthropogenic factors controlling N dynamics in streams. While previous research has shown how δ(15)N of PUCs varies with δ(15)N of dissolved inorganic N (DIN) among streams, less is known about how δ(15)N of PUCs varies over time. Here, we examined monthly variation of δ(15)N of PUCs and of DIN species (nitrate and ammonium) over a year, and compared it among streams with contrasting human impacts and PUC types. Our results showed no evidence of isotopic seasonal patterns. Temporal variability in δ(15)N-PUCs increased with human impact, being the highest in the urban stream, probably influenced by the high variability of δ(15)N-DIN. Among compartments, in-stream PUCs characterized by fast turnover rates, such as filamentous algae, showed the highest temporal variability in δ(15)N values (from -3.6 to 23.2 ‰). Our study elucidates some of the environmental and biological controls of temporal variability of δ(15)N in streams, which should be taken into account when using stable isotopes as an ecological tool.

Nitrogen stable isotopes in primary uptake compartments across streams differing in nutrient availability.

High variability in the natural abundance of nitrogen stable isotopes (δ(15)N) has been reported for primary uptake compartments (PUCs; e.g., epilithon, filamentous algae, bryophytes, macrophytes) in human-impacted aquatic ecosystems, but the origin of this variability is not yet well understood. We examined how δ(15)N of different PUC types relate to δ(15)N of dissolved inorganic nitrogen (DIN) species (nitrate and ammonium) and to the stream nutrient concentrations in which they grow. We selected 25 reaches located across the fluvial network of La Tordera catchment (NE Spain, 868.5 km(2)), encompassing a gradient of human pressures from headwaters to the river valley. δ(15)N-PUC variability was mostly explained by location within the fluvial network and was strongly related to the δ(15)N of DIN species, especially of ammonium. Models were stronger for PUCs growing within the stream channel and thus using streamwater as their main source of nutrients. Regression models including nutrient concentrations improved the prediction power for δ(15)N-PUCs, suggesting that nutrient concentrations and stoichiometry cannot be ignored in explaining the natural abundance of nitrogen isotopes in PUCs. These results provide insights into what controls variability in δ(15)N of PUCs within a stream network, with implications for the application of stables isotopes as an ecological tool.