Mitochondrial Genetic Diversity of the Freshwater Snail Melanoides tuberculata.

According to geological history, Peninsular Malaysia and Borneo formed at different times and were once connected during Quaternary glaciations. To determine how this history has influenced phylogeography, our study examined the population genetic structure of the tropical freshwater gastropod Melanoides tuberculata across Peninsular Malaysia and Borneo using the sequences from mitochondrial DNA 16S rRNA and cytochrome oxidase subunit I genes (1168 bp). In total, 104 specimens were collected from seventeen populations. All mtDNA haplotypes were identified as belonging to two highly divergent lineages, and these lineages were almost allopatric in their distributions. Our study found that the freshwater fauna in Malaysia might be divided into four regions: northeast Peninsular Malaysia, northwest Peninsular Malaysia, south Peninsular Malaysia, and Borneo. The phylogeography of M. tuberculata in Malaysia was shaped by the landforms of Peninsular Malaysia and by the paleo-river systems in the Sunda continental shelf. In addition, our study found that these two lineages in Malaysia have invaded the globe. These results suggest that Malaysia is located in important shipping lanes throughout the world, and the populations of M. tuberculate might be widely distributed throughout the world by shipping.

Bioactive Constituents from the Termite Nest-Derived Medicinal Fungus Xylaria nigripes.

Six new eremophilane-type sesquiterpenes, namely, nigriterpenes A-F (1-6), and one new phenolic compound, named 2-hydroxymethyl-3-pentylphenol (7), along with fomannoxin alcohol, 3-butyl-7-hydroxyphthalide, scytalone, and fomannoxin were isolated from the ethyl acetate extracts of the fermented broths of termite nest-derived Xylaria nigripes, which has long been used as a traditional Chinese medicine for treating insomnia and depression. Their structures were elucidated on the basis of spectroscopic data analysis and compared with the literature. All the pure isolates were evaluated against lipopolysaccharide-induced inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2) expression, and NO production in murine brain microglial BV-2 cells. Of the compounds tested, nigriterpene C (3) and fomannoxin alcohol exerted significant inhibitory effects on two induced enzymes and NO production without any significant cellular toxicity. The most potent compound, 3, exhibited concentration-dependent inhibition on NO production and iNOS and COX-2 expression with IC50 values of 21.7 ± 4.9, 8.1 ± 2.3, and 16.6 ± 5.5 µM, respectively. These results indicated that the potential anti-inflammatory effects of nigriterpene C (3) and fomannoxin alcohol on murine brain microglial BV-2 cells may provide a rationale for the traditional medical uses of X. nigripes for treating insomnia and depression.

Aliphatic phenolic ethers from Trichobotrys effusa.

Four novel aliphatic phenolic ethers, namely, trichoethers A-D (1-4), possessing a unique C11-O-C10 skeleton, together with coriloxin, zythiostromic acid A, radicicol, and 3,5-dihydroxytoluene were isolated from the ethyl acetate extracts of the fermented broths of Trichobotrys effusa YMJ1179. The structures of all the compounds were determined based on spectroscopic data analysis. The configurations of 1-4 were established by J values and NOESY and compared with published data. Compounds 1-4 and radicicol exhibited growth-inhibitory activities against the A549 non-small-cell lung cancer cell line with GI50 values of 25.61, 19.32, 16.19, 24.31, and 1.43 µM, respectively, in comparison with 5-fluorouracil (GI50 = 4.55 µM).

Multilocus Phylogeography and Population Genetic Analyses of Opsariichthys hainanensis Reveal Pleistocene Isolation Followed by High Gene Flow around the Gulf of Tonkin.

The ichthyofauna of continental islands is characterized by immigration through a land bridge due to fluctuating sea levels. Hainan Island is adjacent to the southern margin of mainland China and provides opportunities for understanding the origin and diversification of freshwater fishes. The aim of our study was to evaluate the level of genetic variation and phylogeographic structure of Opsariichthys hainanensis on Hainan Island and mainland China, using mtDNA cyt b gene (1140 bp) and D-loop (926 bp), nuclear RAG1 gene (1506 bp), and 12 microsatellite loci. Mitochondrial phylogenetic analysis identified five major lineages according to the geographical distribution from different populations. We suggested that two dispersal events occurred: the population in the Changhua River migrated to the Red River (Lineage B), and the populations in the South Hainan region moved northwards to the North Hainan region. However, populations in Northwest Hainan Island dispersed to the populations around the Gulf of Tonkin (Lineage A1) and populations in Northeast Hainan Island dispersed to the populations in mainland China (Lineage A2). Our results indicated that the populations of O. hainanensis suffered a bottleneck event followed by a recent population expansion supported by the ABC analysis. We suggest that O. hainanensis populations were found mostly in the lowlands and a lack of suitable freshwater habitat in southern mainland China and Hainan during the Last Interglacial period, and then expansion occurred during the Last Glacial Maximum.

Population genetic diversity and structure of Rhinogobius candidianus (Gobiidae) in Taiwan: Translocation and release.

Rhinogobius candidianus is a freshwater goby distributed in north, northwest, west, and south Taiwan, but this species has been introduced to east Taiwan and became dominant. To investigate its native population genetic diversity and structure and evaluate the sources and diversity of translocated populations, the mitochondrial DNA control region and cytochrome b gene (1981 bp) from 220 specimens were analyzed. These results indicated that (1) the east populations originated from two sources in west Taiwan; (2) translocated populations exist in east Taiwan and south Taiwan; (3) many populations have likely been moved secondarily by human intervention; (4) the effective size of the populations had declined greatly; (5) within the native populations, the ancestral populations colonized Taiwan during the land bridge phase in the Pleistocene through north Taiwan; (6) the landform changes in Taiwan shaped the population structure; and (7) the landforms of the coastline during glaciation also shaped the native range. The low-level genetic diversity, high population differentiation, and population decline greatly suggest the need for resource management and conservation interventions. Four clades (α-δ) should be managed as four distinct evolutionarily significant units, while the translocated populations should be managed as separate management units. Moreover, the translocated populations in east Taiwan should be evaluated and monitored carefully.

Sesquiterpene glycosides from Cosmospora joca.

Five novel sesquiterpene glycosides, namely, cosmosporasides A-E (1-5), were isolated from the ethyl acetate extract of the fermented broth of Cosmospora joca. The structures of 1-5 were elucidated on the basis of spectroscopic data analyses, monosaccharide composition analyses, and monosaccharide chirality analyses. The relative configuration of the sesquiterpene moiety of 3-5 was determined by J-based configuration analyses and supported by NOESY assignments. The inhibitory effects of 1-5 on the nitric oxide (NO) production in lipopolysaccharide (LPS)-activated murine macrophage RAW264.7 cells were evaluated; all except 3 inhibited NO production at 100 µM. 4 was the most potent, with an average maximum inhibition and a median inhibitory concentration value of 71.70 ± 1.23% and 65.98 ± 0.53 µM, respectively.

Polymorphic microsatellite loci isolated from the Squalidus argentatus using PCR-Based Isolation of Microsatellite Arrays (PIMA).

Squalidus argentatus (Sauvage and Dabry de Thiersant 1874) is a small-sized freshwater fish which is distributed in Mainland China, Hainan Island and Taiwan. The populations of S. argentatus have dropped sharply probably due to overharvesting and water pollution recently. Eleven polymorphic microsatellite markers were developed for the cyprinid fish S. argentatus. These new markers were tested on 43 individuals collected from Yangtze River and Qiantang River. The number of alleles, observed and expected heterozygosity per locus, in two populations ranged from 3 to 14, from 0.333 to 0.954 and from 0.480 to 0.928, respectively. Only two loci are significantly deviated from Hardy-Weinberg expectations due to the heterozygote deficiency. No significant linkage disequilibrium was detected between the pairwise comparisons of these loci. These polymorphic microsatellite loci will enable us to study the genetic variation, population structure, and conservation genetics of this species in the future.

Genetic diversity of Rhinogobius delicatus (Perciformes: Gobiidae): origins of the freshwater fish in East Taiwan.

Mitochondrial DNA cytochrome b and d-loop sequences (1,984 bp) from 92 specimens of the freshwater goby Rhinogobius delicatus from seven drainages in East Taiwan were identified as two major lineages exhibiting a southern or northern distribution. The existence of low genetic diversity, a pattern of population decline and high population differentiation (FST=0.711) support the need for the development of management strategies for the conservation of localized populations. The results of a statistical dispersal-vicariance analysis suggested that the ancestral populations of R. delicatus were widely distributed in East Taiwan. Compared with the phylogeographic patterns of the other endemic eastern Taiwan freshwater fishes, Onychostoma alticorpus, Aphyocypris kikuckii and Hemimyzon taitungensis, our study suggests that the freshwater fishes colonized East Taiwan through northeastern and southwestern Taiwan, although the ancestral populations colonized the island before it reached its present shape.

Characterization of the mitochondrial genome of Favonigobius reichei (Perciformes, Gobiidae).

Favonigobius reichei, belongs to the family Gobiidae, which is widely distributed in estuarine and marine waters of the coasts of the Indian and the western Pacific Oceans. The entire mitochondrial genome of F. reichei is 16,415 base pairs (bp) in length and contained 13 protein-coding genes (PCGs), 2 ribosomal RNA (rRNA) genes, and 22 transfer RNA (tRNA) genes. The overall base composition is 27.71% A, 26.78% T, 28.02% C, and 17.49% G, showing AT-rich feature (54.49%). Phylogenetic analysis based on 13 PCGs shows the F. reichei has the closest evolutionary relationship with Myersina macrostoma. This work provides a valuable mitogenome resource for better understanding of the conservation genetics, environmental DNA, and population studies in the family Gobiidae.

Genetic diversity and structure of the Chinese lake gudgeon (Sarcocheilichthys sinensis).

Mitochondrial DNA cytochrome b and d-loop sequences (2,137 bp) in 65 specimens of Sarcocheilichthys sinensis from five populations were identified as two lineages (I and II). The pairwise genetic distance between lineages I and II was 1.94%. SAMOVA analyses suggested that the best grouping occurred at three groups, Yangtze, Qiantang and Minjiang Rivers. High haplotype diversity (0.949) and low nucleotide diversity (θπ = 1.067%) were detected. The results of the neutrality tests, mismatch distribution and approximate Bayesian computation (ABC) did not support demographic expansions. The results of phylogenetic analysis, statistical dispersal-vicariance analysis (S-DIVA), ABC, MIGRATE-N and the time to the most recent common ancestor (TMRCA) indicated two colonization routes. First, before the Wuyi Mountains lifted, S. sinensis dispersed from the Yangtze River to the Minjiang River. Second, during glaciation, the continental shelf was exposed, which contributed to the dispersion of populations from the Yangtze River.

Complete mitochondrial genome and the phylogenetic position of the pelagic octopus Tremoctopus violaceus (Mollusca: Tremoctopodidae).

Tremoctopus violaceus is a small species of pelagic octopods which inhabit in subtropical and tropical open oceans. The mitogenome of T. violaceus is 16,015 base pairs (bp) in length and contained 13 protein-coding genes, two rRNA genes, and 22 tRNA genes. Sequence analysis showed that the overall base composition is 31.5% for A, 40.47% for T, 7.79% for C, and 20.24% for G. The newly characterized complete mitochondrial genome of T. violaceus will provide essential data for further studies of this endangered species.

Mitochondrial diversity and phylogeography of Acrossocheilus paradoxus (Teleostei: Cyprinidae).

Mitochondrial DNA cytochrome b sequences (1141 bp) in 229 specimens of Acrossocheilus paradoxus from 26 populations were identified as four lineages. The pairwise genetic distances among these four lineages ranged from 1.57 to 2.37% (mean= 2.00%). Statistical dispersal-vicariance analysis suggests that the ancestral populations were distributed over mainland China and Northern and Western Taiwan. Approximate Bayesian computation approaches show that the three lineages in Taiwan originated from the lineage in mainland China through three colonization routes during two glaciations. The results indicated that during the glaciation and inter-glacial periods, the Taiwan Strait was exposed and sank, which contributed to the dispersion and differentiation of populations. Furthermore, the populations of A. paradoxus colonized Taiwan through a land bridge to the north of the Formosa Bank, and the Miaoli Plateau in Taiwan was an important barrier that limited gene exchange between populations on both the sides.

The complete mitochondrial genome of Montipora aequituberculata (Scleractinia, Acroporidae).

The complete mitogenome of the hexacorallia, Montipora aequituberculata has been amplified and sequenced. The mitogenome consists of 17,886 bp, with 13 protein-coding genes, 2 ribosomal RNA genes, 2 transfer RNA genes and a control region. It has been observed that ND5 gene is split into two parts by a large fragment of genes, which commonly presented in scleractinian coral. The overall base composition of the H-strand was A, 24.91%; G, 24.1%; C, 14.2%; and T, 36.8%, with a slight AT bias of 61.7%. The control region was 627 bp in length and located between 12S rRNA and COIII gene. Based on the neighbour-joining (NJ) tree, M. aequituberculata was grouped with M. cactus, Anacropora matthai and Acropora tenuis, and formed a clade of Acroporidae. In conclusion, the complete mitogenome of M. aequituberculata data may provide more informative for phylogenetic approach for corals phylogeny.

Complete mitochondrial genome of Junceella fragilis (Gorgonacea, Ellisellidae).

In this article, the complete mitogenome of the Octocorallia, zooxanthellate, Junceella fragilis has been amplified and sequenced. This mitochondrial genome consists of 18,724 bp, with 14 protein-coding genes, 2 ribosomal RNA genes, 1 transfer RNA genes, no intron was observed. It has been observed that a mitochondrial mismatch repair (mtMutS) gene was present in all octocorals. The overall base composition of the heavy strand was A, 29.1%; G, 20.4%; C, 33.0%; and T, 17.5%, with a slight AT bias of 62.1%. The complete mitogenomic data may provide more informative for phylogenetic approach for soft corals phylogeny especially for octocorallian species.

Complete mitochondrial genome of the amphidromous, red-tailed goby Sicyopterus lagocephalus (Pallas) (Teleostei, Gobiidae).

In this article, the complete mitogenome of the amphidromous, red-tailed goby, Sicyopterus lagocephalus has been amplified and sequenced by long polymerase chain reaction. This mitochondrial genome consists of 16,500 bp, with 13 protein-coding genes, 2 ribosomal RNA genes, 22 transfer RNA genes, and a non-coding control region (CR), and its gene arrangement is identical to those of most vertebrates. The CR (841 bp) is located between tRNA(Pro) and tRNA(Phe). The overall base composition of the heavy strand is A, 28.9%; G, 16.4%; C, 28.3%; and T, 26.4%, with a slight AT bias of 55.3%. The complete mitogenomic data may provide more informative for phylogenetic approach for gobioid phylogeny especially for Sicydiine gobies.

SLDP: a novel protein related to caleosin is associated with the endosymbiotic Symbiodinium lipid droplets from Euphyllia glabrescens.

Intracellular lipid droplets (LDs) have been proposed to play a key role in the mutualistic endosymbiosis between reef-building corals and the dinoflagellate endosymbiont Symbiodinium spp. This study investigates and identifies LD proteins in Symbiodinium from Euphyllia glabrescens. Discontinuous Percoll gradient centrifugation was used to separate Symbiodinium cells from E. glabrescens tentacles. Furthermore, staining with a fluorescent probe, Nile red, indicated that lipids accumulated in that freshly isolated Symbiodinium cells and lipid analyses further showed polyunsaturated fatty acids (PUFA) was abundant. The stable LDs were purified from endosymbiotic Symbiodinium cells. The structural integrity of the Symbiodinium LDs was maintained via electronegative repulsion and steric hindrance possibly provided by their surface proteins. Protein extracts from the purified LDs revealed a major protein band with a molecular weight of 20 kDa, which was termed Symbiodinium lipid droplet protein (SLDP). Interestingly, immunological cross-recognition analysis revealed that SLDP was detected strongly by the anti-sesame and anti-cycad caleosin antibodies. It was suggested that the stable Symbiodinium LDs were sheltered by this unique structural protein and was suggested that SLDP might be homologous to caleosin to a certain extent.

Complete mitochondrial genome of Sicyopterus japonicus (Perciformes, Gobiidae).

In this study, we sequenced the complete mitochondrial genome of Sicyopterus japonicus (Perciformes, Gobiidae). This mitochondrial genome consists of 16,514 base pairs (bp), with 13 protein-coding genes, 2 ribosomal RNA genes, 22 transfer RNA genes, and 1 non-coding control region (CR), also with the gene synteny identical to that of typical vertebrates. CR, of 843 bp in length, is located between tRNA(Pro) and tRNA(Phe). The overall base composition of the heavy strand shows T 26.1%, C 28.7%, A 28.3%, and G 16.91%, with a slight AT bias of 54.4%.

Isolation and characterization of sesquiterpenes from Arecophila saccharicola YMJ96022401 with NO production inhibitory activity.

Sesquiterpenes, arecoic acids A-F and arecolactone, were isolated from the ethyl acetate extracts of the fermented broth of Arecophila saccharicola YMJ96022401 along with two known analogues 1,7α,10α-trihydroxyeremophil-11(13)-en-12,8-olide and 1,10α,13-trihydroxyeremophil-7(11)-en-12,8-olide. Their structures were elucidated on the basis of spectroscopic data analyses. The inhibitory effects of all of these compounds on nitric oxide (NO) production in lipopolysaccharide (LPS, 200 µg/mL)-activated murine macrophage RAW264.7 cells were also evaluated. Among these compounds, 1,7α,10α-trihydroxyeremophil-11(13)-en-12,8-olide significantly inhibited NO production without any cytotoxicity, and its average maximum inhibition (E(max)) at 100 µM and median inhibitory concentration (IC50) were 85.7%±0.8% and 16.5±1.0 µM, respectively. Arecolactone was the most potent, with the E(max) at 12.5 µM and IC50 being 94.7%±0.8% and 1.32±0.1 µM, respectively, but displayed cytotoxicity at considerable higher concentrations than 25 µM. Analyses of Western blotting indicated that arecolactone (0.8-12.5 µM) inhibited induction of inducible NO synthase (iNOS) by LPS, which involved suppression of NF-κB activation and the phosphorylation of extracellular signal-regulated kinase (ERK), c-Jun NH2-terminal kinase (JNK) and p38 mitogen-activated protein kinases (MAPKs) in activated RAW 264.7 cells. In addition, arecolactone concentration-dependently prevented the vascular hyporeactivity to phenylephrine induced by LPS (300 ng/mL) through iNOS pathway in isolated rat thoracic aortic rings. These results indicated that both of these naturally occurring iNOS inhibitors may provide a rationale for the potential anti-inflammatory effect of A. saccharicola YMJ96022401.