Significant Effects of Associated Microorganisms on the Community of Photosynthetic Picoeukaryotes.

Photosynthetic picoeukaryotes (PPEs) form associations with other microorganisms. However, whether and how the associated microbes affect PPE communities remain unknown. We used flow cytometric cell sorting combined with parallel high-throughput sequencing of the 18S and 16S rRNA genes to simultaneously investigate PPEs and their associated microbial communities in the Yangtze-connected Lake Dongting. The lake harbors a great diversity of PPEs. PPE communities exhibited significant temporal rather than spatial variations. Two distinct PPE taxa affiliated with Discostella nipponica and Poterioochromonas malhamensis were dominant during winter/spring and summer, respectively. Parallel high-throughput sequencing revealed a great diversity of associated bacteria and non-pigmented eukaryotes (NPEs) in PPEs sorts. Proteobacteria, Actinobacteria, Bacteroidetes, and Cyanobacteria among the associated bacteria and fungi among the associated NPEs were dominant. PPEs were more apparently associated with bacteria than with NPEs. The co-occurrence network of PPEs and associated microbes formed five major modules, which exhibited distinct temporal patterns, being specific to a certain period. Variations in PPEs communities were significantly correlated with both environmental factors and associated microbial communities. In variation partitioning analysis, the associated bacteria explained the greatest variations in PPE communities, and associated bacteria and NPEs co-explained a large portion of environmental effects on PPE communities. Our results highlight the significance of associated microbes in shaping PPE communities.

Pleistocene climate and geomorphology drive the evolution and phylogeographic pattern of Triplophysa robusta (Kessler, 1876).

Montane systems provide excellent opportunities to study the rapid radiation influenced by geological and climatic processes. We assessed the role of Pleistocene climatic oscillations and mountain building on the evolution history of Triplophysa robusta, a cold-adapted species restricted to high elevations in China. We found seven differentiated sublineages of T. robusta, which were established during the Mid Pleistocene 0.87-0.61 Mya. The species distribution modeling (SDM) showed an expansion of T. robusta during the Last Glacial Maximum (LGM) and a considerable retraction during the Last Interglacial (LIG). The deep divergence between Clade I distributed in Qinling Mountains and Clade II in Northeastern Qinghai-Tibet Plateau (QTP) was mainly the result of a vicariance event caused by the rapid uplifting of Qinling Mountains during the Early Pleistocene. While the middling to high level of historical gene flow among different sublineages could be attributed to the dispersal events connected to the repetition of the glacial period during the Pleistocene. Our findings suggested that frequent range expansions and regressions due to Pleistocene glaciers likely have been crucial for driving the phylogeographic pattern of T. robusta. Finally, we urge a burning question in future conservation projection on the vulnerable cold-adapted species endemic to high elevations, as they would be negatively impacted by the recent rapid climate warming.

Spatiotemporal characteristics of the pharyngeal teeth in interspecific distant hybrids of cyprinid fish: Phylogeny and expression of the initiation marker genes.

As an important feeding organ and taxonomical characteristic, the pharyngeal teeth of cyprinid fish have very high morphological diversity and exhibit species-specific numbers and arrangements. Many genes have been verified to regulate the pharyngeal teeth development and act as the initiation marker for teeth. Six initiation marker genes for pharyngeal teeth were used as RNA probes to investigate the expression pattern, and these genes were further used to construct a phylogenetic tree for cyprinid fish including some distant hybrids. The results from in situ hybridization showed that similarities and differences existed in the expression of dlx2b, dlx4b, dlx5a, pitx2, fth1b, and scpp5 in the pharyngeal region of the hybrids (BT) by the crosses of blunt snout bream (BSB, ♀) × topmouth culter (TC, ♂). Particularly, we found a high specificity marker gene scpp5 for the early development of pharyngeal teeth. The Scpp5 expression pattern established a clear graphic representation on the spatiotemporal characteristics of the early morphogenesis of pharyngeal teeth in BT and BSB. Our results suggested that the scpp5 expression in 4V1, 3V1, and 5V1 in BT occurred earlier than that in BSB, while the replacement rate of pharyngeal teeth (4V2, 3V2, and 5V2) was faster in BSB. Phylogenetic analysis revealed that the six marker genes were highly conserved and could be used as the molecular marker for identifying the parents of the distant hybrids in cyprinid fish. The expression patterns of the scpp5 gene was examined in various tissues, including the brain, gill, heart, liver, muscle, skin, fins, gonad, eye, and kidney, showing that the scpp5 gene was ubiquitously expressed, indicating its important role in cyprinid fish.

Immunoglobulin heavy-chain loci in ancient allotetraploid goldfish.

As an ancient allotetraploid species, goldfish (Carassius auratus) have two sets of subgenomes. In this study, immunoglobulin heavy-chain (IGH) genes were cloned from the red crucian carp (Carassius auratus red var.), and the corresponding loci were identified in the gynogenetic diploid red crucian carp (GRCC) genome as well as the genomes of three other goldfish strains (Wakin, G-12, and CaTCV-1). Examination showed that each goldfish strain possessed two sets of parallel IGH loci: a complete IGHA locus and a degenerated IGHB locus that was nearly 40 × smaller. In the IGHA locus, multiple τ chain loci were arranged in tandem between the µ&δ chain locus and the variable genes, but no τ-like genes were found in the IGHB locus.

Phylogeographic relationships and the evolutionary history of the Carassius auratus complex with a newly born homodiploid raw fish (2nNCRC).

BACKGROUND: An important aspect of studying evolution is to understand how new species are formed and their uniqueness is maintained. Hybridization can lead to the formation of new species through reorganization of the adaptive system and significant changes in phenotype. Interestingly, eight stable strains of 2nNCRC derived from interspecies hybridization have been established in our laboratory. To examine the phylogeographical pattern of the widely distributed genus Carassius across Eurasia and investigate the possible homoploid hybrid origin of the Carassius auratus complex lineage in light of past climatic events, the mitochondrial genome (mtDNA) and one nuclear DNA were used to reconstruct the phylogenetic relationship between the C. auratus complex and 2nNCRC and to assess how demographic history, dispersal and barriers to gene flow have led to the current distribution of the C. auratus complex. RESULTS: As expected, 2nNCRC had a very close relationship with the C. auratus complex and similar morphological characteristics to those of the C. auratus complex, which is genetically distinct from the other three species of Carassius. The estimation of divergence time and ancestral state demonstrated that the C. auratus complex possibly originated from the Yangtze River basin in China. There were seven sublineages of the C. auratus complex across Eurasia and at least four mtDNA lineages endemic to particular geographical regions in China. The primary colonization route from China to Mongolia and the Far East (Russia) occurred during the Late Pliocene, and the diversification of other sublineages of the C. auratus complex specifically coincided with the interglacial stage during the Early and Mid-Pleistocene in China. CONCLUSION: Our results support the origin of the C. auratus complex in China, and its wide distribution across Eurasia was mainly due to natural Pleistocene dispersal and recent anthropogenic translocation. The sympatric distribution of the ancestral area for both parents of 2nNCRC and the C. auratus complex, as well as the significant changes in the structure of pharyngeal teeth and morphological characteristics between 2nNCRC and its parents, imply that homoploid hybrid speciation (HHS) for C. auratus could likely have occurred in nature. The diversification pattern indicated an independent evolutionary history of the C. auratus complex, which was not separated from the most recent common ancestor of C. carassius or C. cuvieri. Considering that the paleoclimate oscillation and the development of an eastward-flowing drainage system during the Pliocene and Pleistocene in China provided an opportunity for hybridization between divergent lineages, the formation of 2nNCRC in our laboratory could be a good candidate for explaining the HHS of C. auratus in nature.

Population Genomic Evidence for the Diversification of Bellamya aeruginosa in Different River Systems in China.

Clarifying the genetic structure can facilitate the understanding of a species evolution history. It is crucial for the management of germplasm resources and providing useful guidance for effective selective breeding. Bellamya is an economically and ecologically important freshwater snail for fish, birds and even humans. Population genetic structures of the Bellamya species, however, were unknown in previous studies. Population genomics approaches with tens to hundreds of thousands of single nucleotide polymorphisms (SNPs) make it possible to detect previously unidentified structures. The population genomic study of seven populations of B. aeruginosa across three river systems (Yellow River, Yangtze River and Pearl River) in China was conducted by SLAF-seq. SLAF-seq obtained a total of 4737 polymorphisms SLAF-tags and 25,999 high-consistency genome-wide SNPs. The population genetic structure showed a clear division among populations from the Yellow River basin (YH and WL) and the Pearl River basin (QSH and LB), as well as population YC from the Yangtze River basin using the SNPs data. However, there existed no distinct population structure using the mitochondrial DNA (mtDNA). The anthropogenic translocation from the Yangtze River basin to the Pearl River basin and the passive dispersion from the Yangtze River basin to the Yellow River basin by flooding have weakened the phylogeographic pattern of B. aeruginosa. The divergence of B. aeruginosa in the three river systems suggests that the anthropogenic dispersal for aquaculture and breeding requires serious consideration of the population structure for the preservation of genetic diversity and effective utilization of germplasm resources.

Integrated proteomic and transcriptomic analysis reveals that polymorphic shell colors vary with melanin synthesis in Bellamya purificata snail.

The snail Bellamya purificata is an ecologically and economically important freshwater gastropod species. However, limited genomic resources are available for this snail. In this study, the transcriptome of mantle tissues and proteome of shells of B. purificata with two shell colors (namely light-cyan line (LC) and light-purple line (LP)) were deeply sequenced and characterized. A total of 5.72 million contigs were assembled into 157,015 unigenes, 21,455 (13.66%) of these unigenes were significantly matched to NR, Swiss-Prot, KOG, GO and KEGG database. 1807 differentially expressed genes (DEGs) were identified between the two different shell color lines. These DEGs were significantly enriched in five KEGG pathways including tyrosine metabolism, tryptophan metabolism, phenylalanine, tyrosine and tryptophan biosynthesis, phenylalanine metabolism, and histidine metabolism, which suggested that the shell color polymorphism in B. purificata was a result of melanin synthesis variation. A total of 1521 proteins were identified in B. purificata here as well. The differentially expressed protein analysis showed that the tyrosinase content in LP was significantly decreased in comparison to LC, which agreed with the transcriptome analysis results. This study provides valuable genomic resources of B. purificata and improves our understanding of molecular mechanisms of biomineralization and shell color polymorphism in snail.

Sox Gene Family Revealed Genetic Variations in Autotetraploid Carassius auratus.

The Sox gene family encoded transcription factors that played key roles in developmental processes in vertebrates. To further understand the evolutionary fate of the Sox gene family in teleosts, the Sox genes were comprehensively characterized in fish of different ploidy levels, including blunt snout bream (2n = 48, Megalobrama amblycephala, BSB), goldfish (2n = 100, Carassius auratus red var., 2nRCC), and autotetraploid C. auratus (4n = 200, 4nRCC). The 4nRCC, which derived from the whole genome duplication (WGD) of 2nRCC, were obtained through the distant hybridization of 2nRCC (♀) × BSB (♂). Compared with the 26 Sox genes in zebrafish (2n = 50, Danio rerio), 26, 47, and 92 putative Sox genes were identified in the BSB, 2nRCC, and 4nRCC genomes, respectively, and classified into seven subfamilies (B1, B2, C, D, E, F, and K). Comparative analyses showed that 89.36% (42/47) of Sox genes were duplicated in 2nRCC compared with those in BSB, while 97.83% (90/92) of Sox genes were duplicated in 4nRCC compared with those in 2nRCC, meaning the Sox gene family had undergone an expansion in BSB, 2nRCC, and 4nRCC, respectively, following polyploidization events. In addition, potential gene loss, genetic variations, and paternal parent SNP locus insertion occurred during the polyploidization events. Our data provided new insights into the evolution of the Sox gene family in polyploid vertebrates after several rounds of WGD events.

Blood biochemistry profile of Qihe crucian carp Carassius auratus in different aquaponic systems.

Blood biochemical profile can be used to evaluate the health status of fish and ambient environmental conditions. However, it is not well known in the aquaponic systems, let alone their differences between hydroponic and aquatic plants. A 5-month trial was conducted to investigate the changes in the growth performance and blood index of Qihe crucian carp Carassius auratus in eight aquaponic systems (control, Ipomoea aquatica, Lactuca sativa, Lemna minor, Amaranthus tricolor, Ceratophyllum demersum, Vallisneria spiralis, and C. demersum-net, indicated by CK, Ia, Ls, Lm, At, Cd, Vn, and Cd-ns). Results showed that weight gain rate and specific growth rate did not significantly differ among the eight groups. However, most blood parameters significantly differed among the eight groups. The glucose level and activities of alkaline phosphatase, alanine aminotransferase, and aspartate aminotransferase were generally larger in Cd, Lm, Vn, and CK groups, and smaller in Ia, Ls, or At groups. Additionally, the triglyceride, cholesterol, albumin, and albumin/globulin were mostly elevated in Ia and At groups. Moreover, the creatinine, total proteins, and globulin levels peaked in Vn group. These findings showed that the blood biochemical profile was more sensitive than the growth performances and that hydroponic plants can induce a higher production and more healthy status of Qihe crucian carp C. auratus when compared with aquatic plants or no plants.

Variations in the Mitochondrial Genome of a Goldfish-Like Hybrid [Koi Carp (♀) × Blunt Snout Bream (♂)] Indicate Paternal Leakage.

Previously, a homodiploid goldfish-like fish (2n = 100; GF-L) was spontaneously generated by self-crossing a homodiploid red crucian carp-like fish (2n = 100; RCC-L), which was in turn produced via the distant hybridization of female koi carp (Cyprinus carpio haematopterus, KOC, 2n = 100) and male blunt snout bream (Megalobrama amblycephala, BSB, 2n = 48). The phenotypes and genotypes of RCC-L and GF-L differed from those of the parental species but were similar to diploid red crucian carp (2n = 100; RCC) and goldfish (2n = 100; GF), respectively. We sequenced the complete mitochondrial DNAs (mtDNAs) of the KOC, BSB, RCC-L, GF-L, and subsequent generations produced by self-crossing [the self-mating offspring of RCC-L (RCC-L-F2) to the self-mating offspring of RCC-L-F2 (RCC-L-F3) and the self-mating offspring of GF-L (GF-L-F2)]. Paternal mtDNA fragments were stably embedded in the mtDNAs of both lineages, forming chimeric DNA fragments. In addition to these chimeras, several nucleotide positions in the RCC-L and GF-L lineages differed from the parental bases, and were instead identical with RCC and GF, respectively. Moreover, RCC-L and GF-L mtDNA organization and nucleotide composition were more similar to those of RCC and GF, respectively, compared to parental mtDNA. Finally, phylogenetic analyses indicated that RCC-L and GF-L clustered with RCC and GF, not with the parental species. The molecular dating time shows that the divergence time of KOC and GF was about 21.26 Mya [95% highest posterior density (HPD): 24.41-16.67 Mya], which fell within the period of recent. The heritable chimeric DNA fragments and mutant loci identified in the mtDNA of the RCC-L and GF-L lineages provided important evidence that hybridizations might lead to changes in the mtDNA and the subsequent generation of new lineages. Our findings also demonstrated for the first time that the paternal mtDNA was transmitted into the mtDNA of homodiploid lineages (RCC-L and GF-L), which provided evidence that paternal DNA plays a role in inherited mtDNA. These evolutionary analyses in mtDNA suggest that GF might have diverged from RCC after RCC diverged from koi carp.

Rapid genomic DNA variation in newly hybridized carp lineages derived from Cyprinus carpio (♀) × Megalobrama amblycephala (♂).

BACKGROUND: Distant hybridization can generate changes in phenotypes and genotypes that lead to the formation of new hybrid lineages with genetic variation. In this study, the establishment of two bisexual fertile carp lineages, including the improved diploid common carp (IDC) lineage and the improved diploid scattered mirror carp (IDMC) lineage, from the interspecific hybridization of common carp (Cyprinus carpio, 2n = 100) (♀) × blunt snout bream (Megalobrama amblycephala, 2n = 48) (♂), provided a good platform to investigate the genetic relationship between the parents and their hybrid progenies. RESULT: In this study, we investigated the genetic variation of 12 Hox genes in the two types of improved carp lineages derived from common carp (♀) × blunt snout bream (♂). Hox gene clusters were abundant in the first generation of IDC, but most were not stably inherited in the second generation. In contrast, we did not find obvious mutations in Hox genes in the first generation of IDMC, and almost all the Hox gene clusters were stably inherited from the first generation to the second generation of IDMC. Interestingly, we found obvious recombinant clusters of Hox genes in both improved carp lineages, and partially recombinant clusters of Hox genes were stably inherited from the first generation to the second generation in both types of improved carp lineages. On the other hand, some Hox genes were gradually becoming pseudogenes, and some genes were completely pseudogenised in IDC or IDMC. CONCLUSIONS: Our results provided important evidence that distant hybridization produces rapid genomic DNA changes that may or may not be stably inherited, providing novel insights into the function of hybridization in the establishment of improved lineages used as new fish resources for aquaculture.

Analysis of genetic diversity and population structure of Bellamya quadrata from lakes of middle and lower Yangtze River.

As an endemic species of freshwater gastropods in China, Bellamya quadrata plays an important role in ecosystem service provision and commercial importance. However, the species is overharvested and its natural habitats are under severe threat due to fragmentation and loss. To estimate the genetic diversity and population structure of B. quadrata, 285 individuals from eight lake populations across middle and lower Yangtze River were sampled. Seven microsatellite loci were genotyped. Our results showed that (i) the genetic diversity of B. quadrata was high in most of the studied populations, yet effective population sizes appear to be rather small in some populations; (ii) low levels of genetic differentiation exists among populations but gene flow was generally high; (iii) no clear geographic or genetic structure was observed in the studied region, implying mechanisms (zoochoric dispersal and anthropogenic translocations) that enhance dispersal and gene flow have promoted population connectivity. However, the comparatively high genetic diversity of B. quadrata could be attributed to a lag phase, suggesting that the genetic diversity of this species may be lost in the future and the priorities for conservation of B. quadrata are necessary.

[Macrobenthic community structure and bioassessment for water quality of Banqiao Reservoir in Huaihe River basin].

In order to demonstrate macrobenthic community structure dynamics and conduct a biological evaluation of water quality in Banqiao Reservoir, we collected seasonal samples from January to November 2014 across 15 sampling sites. A total of 17 species belonging to five families, four classes, and 3 phyla were identified, including 8 chironomidae, 4 oligochaeta and 5 mollusc. Bellamya aeruginosa, Corbicula fluminea, Branchiura sowerbyi, Pelopia sp. and Glyptotendipes sp. were the important species in Banqiao Reservoir. The total density, biomass and biodiversity of marobenthos showed significant spatial and temporal differences. ANOSIM analysis indicated that the macrobenthic community structure also differed significantly among regions and seasons, and the main contributing species were Glyptotendipes sp., Pelopia sp. and B. sowerbyi. The abundance-biomass comparison curves (ABC curves) indicated that the current macrobenthic community in Banqiao Reservoir was stable. Combined with the biodiversity index, biological pollution index (BPI) and Hilsenhoff biotic index (BI), it was suggested that Banqiao Reservoir suffered slight pollution.