Conventional morphology-based identification is commonly used for routine assessment of freshwater ecosystems. However, cost and time efficient techniques such as high-throughput sequencing (HTS) based approaches may resolve the constraints encountered in conducting morphology-based surveys. Here, we characterized stream macroinvertebrate species diversity and community composition via metabarcoding and morphological analysis from environmental samples collected from the Shigenobu River Basin in Ehime Prefecture, Japan. We compared diversity metrics and assessed both approaches' ability to evaluate the relationship between macroinvertebrate community and environmental variables. In total, we morphologically identified 45 taxa (3 families, six subfamilies, 31 genera, and five species) from 8,276 collected individuals from ten study sites. We detected 44 species by metabarcoding, with 35 species collapsed into 11 groups matching the morphologically identified taxa. A significant positive correlation between logged depth (number of HTS reads) and abundance of morphological taxa was observed, which implied that quantitative data can be used for subsequent analyses. Relatively higher estimates of alpha diversity were calculated from the metabarcoding data in comparison to morphology-based data. However, beta diversity estimates between metabarcoding and morphology data based on both incidence and abundance-based matrices were correlated proving that community differences between sampling sites were preserved in the molecular data.Also, both models were significant, but metabarcoding data (93%) explained a relatively higher percentage of variation in the relationship between community composition and the environmental variables than morphological data (91%). Overall, we present both the feasibility and limitations of HTS-driven estimations of taxonomic richness, community composition, and diversity metrics, and that metabarcoding was proven comparable and more sensitive against morphology-based analysis for stream macroinvertebrate biodiversity assessment and environmental monitoring.

Zwicknia Murányi, gen. n. is erected for the Capnia bifrons species group sensu Zhiltzova, 2001 with the description of three new species based on morphology, mating call, and the mitochondrial DNA marker cytochrome c oxidase I: Z. acuta Murányi & Orci, sp. n., Z. kovacsi Murányi & Gamboa, sp. n. and Z. rupprechti Murányi, Orci & Gamboa, sp. n.. Zwicknia bifrons (Newman, 1838) comb. n. is selected as the type species and redescribed. The other three species placed into Zwicknia, gen. n., Z. sevanica (Zhiltzova, 1964) comb. n., Z. tuberculata (Zhiltzova, 1964) comb. n., and Z. turkestanica (Kimmins, 1950) comb. n. are redescribed based only on morphological characters. Comparative morphological studies and newly discovered characters of the genitalia has allowed for the first time a synopsis of the adults of the West Palaearctic and Nearctic genera of Capniidae. Arsapnia Banks, 1897 (type species: A. decepta Banks, 1897 comb. rev.) is removed from synonymy with Capnia Pictet, 1841 with new combinations, Arsapnia arapahoe (Nelson & Kondratieff, 1988) comb. n., A. coyote (Nelson & Baumann, 1987) comb. n., A. pileata (Jewett, 1966) comb. n., A. sequoia (Nelson & Baumann, 1987) comb. n., A. teresa (Claassen, 1924) comb. n., A. tumida (Claassen, 1924) comb. n., and A. utahensis (Gaufin & Jewett, 1962) comb. n. A new sensu stricto diagnosis of Capnia is proposed with comments on the taxa retained in Capnia sensu lato.

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10Conventional morphology-based identification is commonly used for routine assessment of freshwater 11 ecosystems. However, cost and time efficient techniques such as high-throughput sequencing (HTS) based 12 approaches may resolve the constraints encountered in conducting morphology-based surveys. Here, we 13 characterized stream macroinvertebrate species diversity and community composition via metabarcoding and 14 morphological analysis from environmental samples collected from the Shigenobu River Basin in Ehime 15 Prefecture, Japan. We compared diversity metrics and assessed both approaches' ability to evaluate the 16 relationship between macroinvertebrate community and environmental variables. In total, we morphologically 17 identified 45 taxa (3 families, six subfamilies, 31 genera, and five species) from 8,276 collected individuals 18 from ten study sites. We detected 44 species by metabarcoding, with 35 species collapsed into 11 groups 19 matching the morphologically identified taxa. A significant positive correlation between logged depth (number 20 of HTS reads) and abundance of morphological taxa was observed, which implied that quantitative data can 21 be used for subsequent analyses. Relatively higher estimates of alpha diversity were calculated from the 22 metabarcoding data in comparison to morphology-based data. However, beta diversity estimates between 23 metabarcoding and morphology data based on both incidence and abundance-based matrices were 24 correlated proving that community differences between sampling sites were preserved in the molecular data. 25 Also, both models were significant, but metabarcoding data (93%) explained a relatively higher percentage of 26 variation in the relationship between community composition and the environmental variables than 27 morphological data (91%). Overall, we present both the feasibility and limitations of HTS-driven estimations 28 of taxonomic richness, community composition, and diversity metrics, and that metabarcoding was proven 29 comparable and more sensitive against morphology-based analysis for stream macroinvertebrate biodiversity 30 assessment and environmental monitoring. 31 32 community composition 33 42 al., 2016). 43 Conventional morphological analysis is most commonly used in routine monitoring programs evaluating 44 environmental quality changes. However, this is not only time consuming but has serious issues with accuracy, 45 and consistency in the level of taxonomic identification that highly depends on taxonomic expertise (Hajibabaei 46 et al., 2011). Specifically, small organisms such as the larval stages of stream macroinvertebrates frequently 47 used for river biomonitoring are often difficult or impossible to identify at finer taxonomic resolution (e.g., 48 species level) (Sweeny et al., 2011). A promising alternative approach is DNA metabarcoding -a combination 49 of amplicon-based high-throughput sequencing (HTS) analysis and DNA taxonomy (Hebert et al., 2003). High-50 throughput amplicon sequencing can process large number ...

Non-alcoholic fatty liver disease (NAFLD) and non-alcoholic steatohepatitis (NASH) are emerging worldwide epidemics, projected to become the leading cause of liver transplants. The strongest genetic risk factor for NAFLD/NASH susceptibility and progression is a single-nucleotide polymorphism (SNP) in the patatin-like phospholipase domain-containing 3 gene (PNPLA3), rs738409, encoding the missense mutation I148M. This aminoacidic substitution interferes with the normal remodeling of lipid droplets in hepatocytes. It is also thought to play a key role in promoting liver fibrosis by inhibiting the release of retinol from hepatic stellate cells. Reducing PNPLA3 levels in individuals homozygous for 148M may be an effective treatment for the entire spectrum of NAFLD, based on gene dosage analysis in the human population, as well as the protective effect of another naturally occurring SNP (rs2294918) in PNPLA3 which, when co-inherited, reduces PNPLA3 mRNA levels to 50% and counteracts disease risk. By screening a clinical compound library targeting specific signaling pathways active in primary human hepatocytes, we identified momelotinib, a drug evaluated in clinical trials to treat myelofibrosis, as a potent down-regulator of PNPLA3 expression, across all genotypes. We found that momelotinib treatment yielded >80% reduction in PNPLA3 mRNA in human primary hepatocytes and stellate cells, as well as in vivo via acute and chronic treatment of WT mice. Using a human multilineage 3D spheroid model of NASH homozygous for the PNPLA3 mutant protein, we additionally show that it decreases PNPLA3 mRNA as well as intracellular lipid content. Furthermore, we show that the effects on PNPLA3 coincide with changes in chromatin accessibility within regulatory regions of the PNPLA3 locus, consistent with inhibition occurring at the level of transcription. In addition to its primary reported targets, the JAK kinases, momelotinib inhibits several non-JAK kinases, including ACVR1. Using a combination of targeted siRNA knockdowns and signaling pathway perturbations, we show that momelotinib reduces the expression of the PNPLA3 gene largely through the inhibition of BMP signaling rather than the JAK/STAT pathway. Overall, our work identified momelotinib as a potential NASH therapeutic and uncovered previously unrecognized connections between signaling pathways and PNPLA3. These pathways may be exploited by drug modalities to “tune down” the level of gene expression, and therefore offer a potential therapeutic benefit to a high at-risk subset of NAFLD/NASH patients.

The generation of the high species diversity of insects in Japan was profoundly influenced by the formation of the Japanese Archipelago. We explored the species diversification and biogeographical history of the Nemouridae Billberg, 1820 family in the Japanese Archipelago using mitochondrial DNA and nuclear DNA markers. We collected 49 species among four genera: Indonemoura Baumann, 1975; Protonemura Kempny, 1898; Amphinemura, Ris 1902 and Nemoura Latreille, 1796 in Japan, China, South Korea and North America. We estimated their divergence times—based on three molecular clock node calibrations—using Bayesian phylogeography approaches. Our results suggested that Japanese Archipelago formation events resulted in diversification events in the middle of the Cretaceous (<120 Ma), speciation in the Paleogene (<50 Ma) and intra-species diversification segregated into eastern and western Japan of the Fossa Magna region at late Neogene (20 Ma). The Indonemoura samples were genetically separated into two clades—that of Mainland China and that of Japan. The Japanese clade clustered with the Nemouridae species from North America, suggesting the possibility of a colonisation event prior to the formation of the Japanese Archipelago. We believe that our results enhanced the understanding both of the origin of the species and of local species distribution in the Japanese Archipelago.

Background Vector control measures are critical for the prevention and reduction of dengue virus (DENV) transmission. Effective vector control is reliant not only on knowledge of mosquito abundance, but also on the timely and accurate detection of mosquito-borne infection. Mosquito-based virus surveillance programs typically rely on pool-based mosquito testing, although whether individual-based mosquito testing is a feasible alternative to this has not been widely studied. Applying an individual-based mosquito testing approach, we conducted a 1-month surveillance study of DENV in adult Aedes aegypti mosquitoes in homes of suspected dengue patients during the 2015 peak dengue season in Tarlac City, Philippines to more accurately assess the mosquito infection rate and identify the DENV serotypes and genotypes concurrently co-circulating in mosquitoes and patients there. Methods We performed a one-step multiplex real-time reverse transcription-polymerase chain reaction (RT-PCR) assay for the simultaneous detection and serotyping of DENV in patients and individual female Ae. aegypti mosquitoes. Additionally, we performed sequencing and phylogenetic analyses to further characterize the detected DENV serotypes in mosquitoes and patients at the genotype level. Results We collected a total of 583 adult Ae. aegypti mosquitoes, of which we individually tested 359 female mosquitoes for the presence of DENV. Ten (2.8%) of the 359 female mosquitoes were positive for the presence of DENV. We detected DENV-1, DENV-2, and DENV-4 in the field-collected mosquitoes, which was consistent with the serotypes concurrently found in infected patients. Sequencing and phylogenetic analyses of the detected DENV serotypes based on the partial sequence of the evelope (E) gene revealed three genotypes concurrently present in the sampled mosquitoes and patients during the study period, namely DENV-1 genotype IV, DENV-2 Cosmopolitan genotype, and DENV-4 genotype II. Conclusions We demonstrated the utility of a one-step multiplex real-time RT-PCR assay for the individual-based DENV surveillance of mosquitoes. Our findings reinforce the importance of detecting and monitoring virus activity in local mosquito populations, which are critical for dengue prevention and control.

BackgroundEnvironmental heterogeneity continuously produces a selective pressure that results in genomic variation among organisms; understanding this relationship remains a challenge in evolutionary biology. Here, we evaluated the degree of genome-environmental association of seven stonefly species across a wide geographic area in Japan and additionally identified putative environmental drivers and their effect on co-existing multiple stonefly species. Double-digest restriction-associated DNA (ddRAD) libraries were independently sequenced for 219 individuals from 23 sites across four geographical regions along a nationwide latitudinal gradient in Japan.ResultsA total of 4251 candidate single nucleotide polymorphisms (SNPs) strongly associated with local adaptation were discovered using Latent mixed models; of these, 294 SNPs showed strong correlation with environmental variables, specifically precipitation and altitude, using distance-based redundancy analysis. Genome–genome comparison among the seven species revealed a high sequence similarity of candidate SNPs within a geographical region, suggesting the occurrence of a parallel evolution process.ConclusionsOur results revealed genomic signatures of local adaptation and their influence on multiple, co-occurring species. These results can be potentially applied for future studies on river management and climatic stressor impacts.Electronic supplementary materialThe online version of this article (10.1186/s12864-019-5453-3) contains supplementary material, which is available to authorized users.