eDNA metabarcoding reveals the role of habitat specialization and spatial and environmental variability in shaping diversity patterns of fish metacommunities.

Information is scarce on how environmental and dispersal processes interact with biological features of the organisms, such as their habitat affinity, to influence patterns in biodiversity. We examined the role of habitat specialist vs. generalist species, and the spatial configuration, connectivity, and different environmental characteristics of river-floodplain habitats to get a more mechanistic understanding of alpha and beta diversity of fish metacommunities. We used environmental DNA metabarcoding to characterize species (taxa) richness and composition in two separate floodplains of the river Danube (Austria and Hungary) during two different hydrological conditions. Results showed that differences in the number of generalist and specialist species and their responses to connectivity and environmental gradients influenced patterns in alpha and beta diversity. Of the components of beta diversity, richness difference (nestedness) showed consistently higher values than replacement (turnover), mainly due to the decrease of specialist species along the connectivity gradient (i.e., from the mainstem to the most isolated oxbows). Variance in both alpha and beta diversity could be well predicted by a set of local and regional variables, despite high environmental variability, which characterizes river-floodplain ecosystems. Of these, the joint or shared variance fractions proved to be the most important, which indicates that the effects of local and regional processes cannot be unambiguously separated in these river-floodplain systems. Local scale environmental variables were more important determinants of both alpha and beta diversity in the low water period than in the high water period. These results indicate the differential role of local and regional processes in community organization during different hydrological conditions. Maintenance of both local and regional scale processes are thus important in the preservation of alpha and beta diversity of floodplain fish metacommunities, which should be considered by environmental management.

Small-scale metapopulation structure of a limnophilic fish species in a natural river system investigated using microsatellite genotyping by amplicon sequencing (SSR-GBAS).

Habitat niches of fish species can exert a strong influence on population structure, even on a small geographical scale. In this scope, Pelasgus thesproticus is a great model species to study connectivity in riverine environments owing to its naturally patchy habitat distribution. Furthermore, it is important to conduct such studies in near-natural systems to avoid the impact of human disturbances on the river, such as fragmentation, morphological changes and habitat degradation. In this sense, the Vjosa in Albania is an excellent study area. A total of 204 individuals were sampled from five locations in the lower Vjosa and two tributaries and genotyped with 33 newly designed microsatellites loci using high throughput sequencing. The application of microsatellite genotyping by sequencing revealed genetic structure and some differentiation, even at a small spatial scale (< 65 river km). A total of 500 alleles were found with an average of 0.93 private alleles among sites with rather low FST values (< 0.04). The extent of admixture observed in some populations indicate that the genetic structure is mainly influenced by upstream populations, either from the main river itself or from tributaries. In addition, the connection between a tributary and the other sites is disrupted by the flow regime, which is reflected in a high degree of divergence from the other populations. Our results indicate that hydrological conditions of the flowing river present strong barriers to gene flow, particularly in the upstream direction, but at the same time act as dispersal corridors in the downstream direction and exhibit source-sink dynamics in which upstream populations contribute disproportionately to downstream populations for this habitat specialist along the river. It is suggested that processes of colonization and reinforcement may play an important role in shaping the genetic structure of patchily distributed fish species in natural river systems. Future studies should increase the knowledge of dispersal factors, habitat heterogeneity, consequence of source-sink dynamics, and gene flow within the system, which will help to understand and maintain important processes related to metapopulation theory and the potential evolutionary consequences of habitat loss and fragmentation.

Quantitative monitoring of diverse fish communities on a large scale combining eDNA metabarcoding and qPCR.

Environmental DNA (eDNA) metabarcoding is an effective method for studying fish communities but allows only an estimation of relative species abundance (density/biomass). Here, we combine metabarcoding with an estimation of the total abundance of eDNA amplified by our universal marker (teleo) using a quantitative (q)PCR approach to infer the absolute abundance of fish species. We carried out a 2850-km eDNA survey within the Danube catchment using a spatial integrative sampling protocol coupled with traditional electrofishing for fish biomass and density estimation. Total fish eDNA concentrations and total fish abundance were highly correlated. The correlation between eDNA concentrations per taxon and absolute specific abundance was of comparable strength when all sites were pooled and remained significant when the sites were considered separately. Furthermore, a nonlinear mixed model showed that species richness was underestimated when the amount of teleo-DNA extracted from a sample was below a threshold of 0.65 × 106 copies of eDNA. This result, combined with the decrease in teleo-DNA concentration by several orders of magnitude with river size, highlights the need to increase sampling effort in large rivers. Our results provide a comprehensive description of longitudinal changes in fish communities and underline our combined metabarcoding/qPCR approach for biomonitoring and bioassessment surveys when a rough estimate of absolute species abundance is sufficient.

Identifying geographically differentiated features of Ethopian Nile tilapia (Oreochromis niloticus) morphology with machine learning.

Visual characteristics are among the most important features for characterizing the phenotype of biological organisms. Color and geometric properties define population phenotype and allow assessing diversity and adaptation to environmental conditions. To analyze geometric properties classical morphometrics relies on biologically relevant landmarks which are manually assigned to digital images. Assigning landmarks is tedious and error prone. Predefined landmarks may in addition miss out on information which is not obvious to the human eye. The machine learning (ML) community has recently proposed new data analysis methods which by uncovering subtle features in images obtain excellent predictive accuracy. Scientific credibility demands however that results are interpretable and hence to mitigate the black-box nature of ML methods. To overcome the black-box nature of ML we apply complementary methods and investigate internal representations with saliency maps to reliably identify location specific characteristics in images of Nile tilapia populations. Analyzing fish images which were sampled from six Ethiopian lakes reveals that deep learning improves on a conventional morphometric analysis in predictive performance. A critical assessment of established saliency maps with a novel significance test reveals however that the improvement is aided by artifacts which have no biological interpretation. More interpretable results are obtained by a Bayesian approach which allows us to identify genuine Nile tilapia body features which differ in dependence of the animals habitat. We find that automatically inferred Nile tilapia body features corroborate and expand the results of a landmark based analysis that the anterior dorsum, the fish belly, the posterior dorsal region and the caudal fin show signs of adaptation to the fish habitat. We may thus conclude that Nile tilapia show habitat specific morphotypes and that a ML analysis allows inferring novel biological knowledge in a reproducible manner.

Genetic diversity of Nile tilapia (Oreochromis niloticus) populations in Ethiopia: insights from nuclear DNA microsatellites and implications for conservation.

BACKGROUND: Nile tilapia, Oreochromis niloticus (Linnaeus, 1758) is among the economically most important freshwater fish species in East Africa, and a major source of protein for local consumption. Human induced translocations of non-native stocks for aquaculture and fisheries have been found as a potential threat to the genetic diversity and integrity of local populations. In the present study, we investigate the genetic structure of O. niloticus from 16 waterbodies across Ethiopia using 37 microsatellite loci with SSR-GBAS techniques. RESULTS: The samples are structured into three main clusters shaped either by biogeographic factors or stocking activities. High FST values (Global FST = 0.438) between populations indicate a high level of genetic differentiation and may suggest long term isolation even within the same drainage systems. Natural populations of the Omo-Turkana system and the lakes in the Southern Main Ethiopian Rift showed the highest genetic variability while low variability was found in stocked populations of lakes Hora, Hashenge and Hayq. CONCLUSIONS: The results presented herein, may provide an essential basis for the management and conservation of the unique genetic resources in northern East Africa, and advance our understanding of biodiversity, phylogeny, evolution and development towards phylogenetically more accurate taxonomic classifications.

New data on Garra makiensis (Cyprinidae, Labeoinae) from the Awash River (Ethiopia) with remarks on its relationships to congeners on the Arabian Peninsula.

On the African continent, the genus Garra consists of several species often insufficiently separated from each other by diagnostic characters. Herein, a detailed morphological redescription of Garra makiensis from the Awash River drainage is presented, together with additional data on the type specimens of G. makiensis and G. rothschildi. Mitochondrial CO1 sequence data are also provided, including the historic paralectotype of G. makiensis, with a comparison to Garra species from Africa and the Middle East. Based on these sequences, G. makiensis clusters outside the group of African congeners and is a sister lineage to species from the south-east of the Arabian Peninsula. Although morphologically variable, G. makiensis is characterised by having a single unbranched pectoral-fin ray, a short distance between vent and anal-fin origin (7.3-19.7 % of pelvic - anal distance), chest and belly covered with scales, and a prominent axillary scale at base of pelvic fin (18.8-35.5 % of pelvic-fin length).