The black piranha (Serrasalmus rhombeus), a widely spread species in the rivers of the Amazon basin, plays a vital role as both key predator and important prey. Despite its essential contribution to ecosystem stability, there is a lack of information regarding its genetic diversity and population dynamics in the central Amazon region. As the Amazon continues to undergo environmental changes in the context of growing anthropogenic threats, such knowledge is fundamental for assist in the conservation of this species. This study is the first to analyze the genetic diversity and population structure of S. rhombeus in the central Amazon region using high-resolution genomic data. We employed a Genotyping-by-Sequencing approach with 248 samples across 14 study sites from various tributaries, encompassing diverse water types (black, white and clear water) and characterized by 34 physiochemical parameters. The data reveals low diversity accompanied by pronounced signs of inbreeding in half of the sites and robust genetic differentiation and variation among sites and within-sites. Surprisingly, we also found evidence of higher dispersal capacity than previously recognized. Our analysis exposed a complex and high population structure with genetic groups exclusive to some sites. Gene flow was low and some groups presented ambiguous genealogical divergence index (gdi) signals, suggesting the occurrence of potential cryptic species. Moreover, our results suggest that the the population structure of black piranha appears more influenced by historical events than contemporary factors. These results underscore the need to give greater attention to this keystone species, for which no regulatory framework or conservation strategies is presently in effect.

The study of the eukaryotic fraction of the microbiota using a metabarcoding approach is usually hindered by the high host to eukaryotic microbiota DNA ratio in samples. Indeed, the 18S rRNA gene is very similar for both the host and its eukaryotic communities, leading to a preferential amplification of the predominant host DNA when using universal primers. Multiple approaches have been developed to reduce host DNA amplification. One method is based on elongation arrest blocking primers, oligonucleotides modified with a C3 Spacer that stops the advancement of the DNA polymerase at non-conserved regions of a target gene. In this paper, we successfully developed and tested species-specific elongation arrest blocking primers to block the Flag cichlid, Mesonauta festivus, 18S rRNA SSU. Our elongation arrest blocking primers significantly reduced the amount of host DNA amplicons by 66 %. In addition to reducing the amount of sequencing wasted, the blocking primers increased the detectability of potentially dangerous parasitic taxa in fish gut, highlighting the potential of the method for parasitic screening. For instance, we discovered a case of infection by the parasitic ciliate Nyctotherus sp. and detected the presence of a parasitic Trematode and an Amoebae. While our data support the possibility of achieving a complete inhibition of host DNA amplification using elongation arrest blocking primers, more research is still required. Still, there is a need for the development and additional testing of protocols to study the eukaryotic diversity present in fish gut, a slow-growing field of study in comparison to its prokaryotic counterpart.

Lowland central Amazonia is characterized by heterogeneous riverscapes dominated by two chemically divergent water types: black (ion-poor, rich in dissolved organic carbonate and acidic) and white (rich in nutrient and turbid) waters. Recent phylogeographic and genomic studies have associated the ecotone formed by these environments to an ecologically driven genetic divergence between fish present in both water types. With the objective of better understanding the evolutionary forces behind the central Amazonian Teleostean diversification, we sampled 240 Mesonauta festivus from 12 sites on a wide area of the Amazonian basin. These sites included three confluences of black and white water environments to seek for repeated evidences of ecological divergence at the junction of these ecotones. Results obtained through our genetic assessment based on 41,268 SNPs contrast with previous findings and support a low influence of diverging water physicochemical characteristics on the genetic structuration of M. festivus populations. Conversely, we detected patterns of isolation by downstream water current and evidence of past events of vicariance potentially linked to the Amazon River formation. Using a combination of population genetics, phylogeographic analysis and environmental association models, we decomposed the spatial and environmental genetic variances to assess which evolutionary forces shaped inter-population differences in M. festivus’ genome. Our sampling design, comprising three confluences of black and white water rivers, supports the main role of evolution by allopatry. While an ecologically driven evolution admittedly played a role in Amazonian fish diversification, we argue that neutral evolutionary processes explain most of the divergence between M. festivus populations.