Genetic diversity of the regionally endangered Chinese ricefish (Oryzias sinensis) in Taiwan, with comments on its conservation status.

The Chinese ricefish (Oryzias sinensis) is a freshwater fish that is regionally endangered in Taiwan. To evaluate its conservation status and further formulate conservation plans for this species in Taiwan, this study analyzed the fish collected from all known localities, including a newly discovered population. The phylogenetic trees based on D-loop and COI divided the Chinese ricefish into two major clades: Taiyangpi-Shuanglienpi and Taiping-Gongliao. This study showed that both the Shuanglienpi and Taiyangpi wild populations had significantly higher Hd and π values, while the Shuanglienpi restored population only had one haplotype from the wild. However, their genetic diversity may have decreased over the past few decades. The Gongliao and Taiping populations shared the same haplotype as a population from a previous study. We propose that the Gongliao and Taiping populations were founded by a few individuals and might have originated from a single introduction event from southeastern mainland China. Low genetic diversity was clearly observed in the Taiping, Gongliao and Shuanglienpi restored populations. Based on the present investigation and previous studies, the invasive poeciliid fish (such as mosquitofish and green swordtail) contribute the most to the decline of the Chinese ricefish population and genetic diversity in Taiwan.

Chub movement is attracted by the collision sounds associated with spawning activities.

Cyprinids (carps, chubs and minnows) possess well-developed hearing and high sensitivity to sound pressure. The sensitive hearing may assist cyprinids with searching for food, territory defense, and mating behavior. Many paired fishes violently shake in sand and gravel while spawning in rivers. However, no study has examined the ecological importance of the collision sound made by the behavior. This study examined whether cohabitated chubs (Opsariichthys evolans and Zacco platypus) use the collision sound as a signal to locate spawning events so they can be a male satellite or egg eater. Three types of sounds (i.e., collision sound, music noise and ambient noise) were played with or without jerkbaits at the midstream of the Keelung River, Taiwan during the spawning season in 2018 and 2019. Generalized linear mixed models were then built to examine the effects of the sound types and the presence of jerkbaits on the number of individuals that the two chubs attracted. Results showed significantly different levels of attractiveness among the three sound types, with the collision sound attracting most fishes, including both females and males, followed by music noise and ambient noise. The presence of jerkbaits increased the number of fishes attracted, but the effect was only statistically marginally significant. These results suggest that the collision sound as an acoustic signal may be more important than a visual signal for the chubs to locate spawning events of other mating pairs, probably because of the longer transmission distance of the former. The present study demonstrates the ecological meanings of the collision sounds made in association with spawning activities of the chubs and implies that the native chub's spawning activities may be affected by the introduced Z. platypus. More studies on the interactions between these cohabitated chubs will benefit the conservation of native chubs.

Stomach content analysis of young Russell's oarfish (Regalecus russelii) from Taiwan, and a report on an unusual case of predation.

In this study, we analyzed the stomach content of two individuals of rare, young Russell's oarfish, Regalecus russelii, from Taiwan. One of them was captured alive along with larval fish and crustaceans by a stow net from the shallow waters (15-18m deep) of the Tamsui River mouth. A total of 38 individuals of larval fish belonging to three species (86.4% of all prey) and 6 individuals of crustaceans belonging to three species were found in the Russell's oarfish specimen's stomach. Among the prey, the Japanese anchovy (Engraulis japonicus) was the most dominant fish species, accounting for 44.7% of all fishes; and luciferids were the most dominant crustaceans, accounting for 66.7% of all crustaceans. All the prey fish were sandy or a mixture of sandy and muddy sediment dwellers. The luciferids and Japanese anchovies are characterized by having diel vertical migration behavior. Based on the sampling information and biological characteristics of the fish larvae and crustaceans, we conjectured that the abundance of these prey when they moved up to the upper layer of the Tamsui River mouth at nighttime had attracted the young Russell's oarfish to hunt there. In addition, we speculated that it might be easier for the vertical-swimming Russell's oarfish to hunt a large number of luciferids assembled in the upper layer of the water by its particular posture or angle of view.

Interspecific Mating Behavior Between Introduced Zacco platypus and Native Opsariichthys evolans in Taiwan.

Introduced freshwater fishes considerably influence the ecology and populations of native species. Previous research has revealed that introduced Zacco platypus may hybridize with chubs that are sister but distinct genera. However, we have little knowledge of Z. platypus' mate choice or its impact on Taiwanese chubs. Therefore, this study identified the interspecific mating behavior between introduced Z. platypus and native Opsariichthys evolans and evaluated the former's invasive impact on cohabitants. Our observations showed that interspecific mating pairs do occur between Z. platypus male(s) and O. evolans female(s). Fifty-three percent of spawning events were interspecific mating and only 43% were between native O. evolans mating pairs. This study showed that Z. platypus male satellites might prefer to engage with Z. platypus, while O. evolans might engage by chance. However, introduced males of Z. platypus may be unable to recognize conspecific females. Meanwhile, introduced females of Z. platypus also have a mate choice preference for males of Z. platypus. Therefore, Z. platypus male hybridization might significantly reduce the successful mating ratio of O. evolans, leading to a dramatic reduction in native O. evolans offspring in the future.

A slender symbiotic goby hiding in burrows of mud shrimp Austinogebia edulis in western Taiwan.

The present study recorded the population of the goby fish (Perciformes: Gobiidae), Eutaeniichthys cf. gilli Jordan & Snyder, 1901, from the tunnel burrowed by the mud shrimp Austinogebia edulis Ngo-Ho and Chan, 1992 in a mudflat in Shengang and Wangong of Changhua County, western Taiwan. This finding is not only a new record of the genus in Taiwan, it is also the first record of this species in a mudflat near an industrial park. In total, 56 individuals of E. cf. gilli were collected from June 2016 to September 2018. Morphological traits of males and females were measured. The resin casting method trapped bodies of E. cf. gilli that were present in the tunnel burrow and proved that the fish inhabits burrows of the mud shrimp A. edulis. In addition, a species of snapping shrimp was also found in the same tunnel. Symbiotic interaction may occur between E. cf. gilli, A. eduli and the snapping shrimp. The China Coastal Current (CCC) runs along the coastlines of Japan, Korea, China, and reaches western Taiwan during the northeast monsoon period. The CCC, therefore, might play an important role in the biogeographic distribution of E. cf. gilli in the western Pacific Ocean. Since E. cf. gilli is listed in the Red List as an endangered species of Japan for many years, Taiwan waters may provide a refuge for this fish species warranting a broader investigation. Since Taiwan is some distance away from the previously recorded locations in Japan, Korea, the Yellow Sea, and the Bohai Sea, a phylogenic analysis is warranted for population and species differentiation in the future.

Using DNA Barcodes to Aid the Identification of Larval Fishes in Tropical Estuarine Waters (Malacca Straits, Malaysia).

Larval descriptions of tropical marine and coastal fishes are very few, and this taxonomic problem is further exacerbated by the high diversity of fish species in these waters. Nonetheless, accurate larval identification in ecological and early life history studies of larval fishes is crucial for fishery management and habitat protection. The present study aimed to evaluate the usefulness of DNA barcodes to support larval fish identification since conventional dichotomous keys based on morphological traits are not efficient due to the lack of larval traits and the rapid morphological changes during ontogeny. Our molecular analysis uncovered a total of 48 taxa (21 families) from the larval samples collected from the Klang Strait waters encompassing both spawning and nursery grounds of marine and estuarine fishes. Thirty-two (67%) of the larval taxa were identified at the species level, two taxa (4%) at the genus level, and 14 taxa (29%) at family level. The relatively low rate of species-level identification is not necessarily due to the DNA barcoding method per se, but a general lack of reference sequences for speciose and non- commercial fish families such as Gobiidae, Blenniidae, and Callionymidae. Larval morphology remains important in species diagnoses when molecular matches are ambiguous. A lower ethanol percentage (50%) for larva preservation is also useful to keep the body of larvae intact for morphological identification, and to preserve DNA for subsequent molecular analyses. The 10% Chelex resin used to extract DNA is also cost- effective for long term monitoring of larval fishes. Hence, the DNA barcoding method is an effective and easy way to aid the identification of estuarine larval fishes at the species level.

The freshwater snake eel genus Lamnostoma (Anguilliformes: Ophichthidae) in Taiwan, with description of a new species.

A review of the freshwater ophichthid eel genus Lamnostoma from Taiwan is provided with three species recognized. Lamnostoma mindora has been recorded in previous literature and its presence is confirmed. Two specimens of Lamnostoma polyophthalmum represent the first record in Taiwan. A new species, Lamnostoma taiwanense sp. nov., is described on the basis of two specimens collected from northeastern Taiwan. It can be distinguished by having 3 preopercular pores, higher number of teeth on lower jaw, and a series of white spots with hill-shaped papillae along the head. A key to all known species of Lamnostoma is provided.

Description of A New Species of the Gudgeon Genus Microphysogobio Mori 1934 (Cypriniformes: Cyprinidae) from Guangdong Province, Southern China.

Shih-Pin Huang, I-Shiung Chen, Yahui Zhao, and Kwang-Tsao Shao (2018) Microphysogobio luhensis n. sp., a new cyprinid species, is described from the Rongjiang River in eastern Guangdong Province, China. Morphological characters and molecular evidence based on mitochondrial DNA Cytochrome b (Cyt b) and Cytochrome oxidase subunit I (COI) sequences were used to compare this new species with other related species from mainland China, Vietnam and Taiwan. The present molecular evidences revealed that this new species is closely related to M. kachekensis and M. yunnanensis. However, these three species can be well distinguished based on the number of small pearl-like papillae on their inside papillae, lip papillae shape, barbel length, barbel width and color pattern. Furthermore, the morphometric comparison between M. kachekensis and the poorly known species M. yunnanensis is also discussed in this study for the first time. In addition, a diagnostic key to all 14 valid species of Microphysogobio from southern mainland China, Hainan Island and Taiwan is also provided.

A New Species of Microphysogobio (Cypriniformes: Cyprinidae) from Guangxi Province, Southern China.

Shih-Pin Huang, Yahui Zhao, I-Shiung Chen, and Kwang-Tsao Shao (2017) Microphysogobio zhangi n. sp., a new cyprinid species is described from Guangxi Province, China. Morphological and molecular evidence based on mitochondrial DNA Cytochrome b (Cyt b) sequence were used for comparing this new species and other related species. The phylogenetic tree topology revealed that this new species is closely related to M. elongatus and M. fukiensis. We also observed the existence of a peculiar trans-river gene ow in the Pearl River and the Yangtze River populations, and speculated that it was caused by an ancient arti cial canal, the Lingqu Canal, which forming a pathway directly connecting these two rivers.

Molecular Phylogeny of the Opsariichthys Group (Teleostei: Cypriniformes) Based On Complete Mitochondrial Genomes.

Shih-Pin Huang, Feng-Yu Wang, and Tzi-Yuan Wang (2017) The complete mitochondrial genomes of 76 species from 43 genera under Cyprinidae sensu lato were collected to reassess the molecular phylogeny of Opsariichthyinae sensu Liao et al. 2011. The mitogenomes of three species, Candidia barbata, Opsariichthys evolans, and Opsariichthys pachycephalus, were newly sequenced. Phylogenetic trees were reconstructed based on 13 concatenated multiple protein-coding genes with two ribosomal RNA genes. The concatenated dataset provided a new perspective on systematics and relationships. Tree topologies show that a monophyletic group containing Parazacco, Candidia, Nipponocypris, Zacco, and Opsariichthys should belong to the Opsariichthys group. In addition, the present results also strongly support that Candidia and Nipponocypris should be regarded as distinct genera within the Opsariichthys group. Aphyocypris, Yaoshanicus, Nicholsicypris, and Pararasbora form a monophyletic group within Xenocyprididae, distinct from the Opsariichthys group. Furthermore, Hemigrammocypris is nested with four species of Metzia, a genus of ex-Cultrinae in Xenocyprididae. In addition, two major types of distinct stripes - longitudinal and vertical - were observed among species of the Opsariichthys group and were highly correlated with molecular phylogenetic relationships. Such types of vertical and longitudinal stripes presented in the Opsariichthys group might have originated in an ancestor species, after which distinct vertical stripes might have been lost among these cyprinids but retained in the Opsariichthys group.

A data-mining framework for exploring the multi-relation between fish species and water quality through self-organizing map.

The steep slopes of rivers can easily lead to large variations in river water quality during typhoon seasons in Taiwan, which may poses significant impacts on riverine eco-hydrological environments. This study aims to investigate the relationship between fish communities and water quality by using artificial neural networks (ANNs) for comprehending the upstream eco-hydrological system in northern Taiwan. We collected a total of 276 heterogeneous datasets with 8 water quality parameters and 25 fish species from 10 sampling sites. The self-organizing feature map (SOM) was used to cluster, analyze and visualize the heterogeneous datasets. Furthermore, the structuring index (SI) was adopted to determine the relative importance of each input variable of the SOM and identify the indicator factors. The clustering results showed that the SOM could suitably reflect the spatial characteristics of fishery sampling sites. Besides, the patterns of water quality parameters and fish species could be distinguishably (visually) classified into three eco-water quality groups: 1) typical upstream freshwater fishes that depended the most on dissolved oxygen (DO); 2) typical middle-lower reach riverine freshwater fishes that depended the most on total phosphorus (TP) and ammonia nitrogen; and 3) low lands or pond (reservoirs) freshwater fishes that depended the most on water temperature, suspended solids and chemical oxygen demand. According to the results of the SI, the representative indicators of water quality parameters and fish species consisted of DO, TP and Onychostoma barbatulum. This grouping result suggested that the methodology can be used as a guiding reference to comprehensively relate ecology to water quality. Our methods offer a cost-effective alternative to more traditional methods for identifying key water quality factors relating to fish species. In addition, visualizing the constructed topological maps of the SOM could produce detailed inter-relation between water quality and the fish species of stream habitat units.

Complete Mitochondrial Reveals a New Phylogenetic Perspective on the Brackish Water Goby Mugilogobius Group (Teleostei: Gobiidae: Gobionellinae).

The Mugilogobius group consists of brackish water gobionellines widely distributed in the Indo-West Pacific region. Complete mitochondrial genome and morphological evidence was collected to estimate their phylogenetic relationship and taxonomic status. A total of 11 genera were sampled, including Brachygobius, Calamiana, Hemigobius, Mugilogobius, Pandaka, Pseudogobiopsis, Pseudogobius, Redigobius, Rhinogobius, Stigmatogobius, and Wuhanlinigobius, five of which were sequenced for the first time. A morphological phylogenetic tree was also reconstructed based on 35 characters. The molecular phylogenetic trees reveal that the Mugilogobius group contains four major clades. The present study also reveals that the adult male mouth size and forked sensory papillae row d can be considered as synapomorphies, and that the head pores on inter-orbital, anterior oculoscapular, and preopercular regions can be regarded as derived features among the Mugilogobius group. Furthermore, the absence of posterior oculoscapular pores may provide a clue for understanding the evolutionary history of the Mugilogobius group.

The complete mitochondrial genome of Temminck's minnow Nipponocypris temminckii (Cypriniformes, Cyprinidae).

The complete mitochondrial genome of Temminck's minnow Nipponocypris temminckii (Temminck & Schlegel, 1846) which is endemic to Japan and Korean Peninsula has been amplified and sequenced in this study. The complete mitochondrial genome is 16,618 base pairs (bp) in total length, had the typical vertebrate mitochondrial gene arrangement, including 13 protein-coding genes, 2 rRNAs, 22 tRNAs, 1 control region (CR) and 1 light-strand replication origin (OL). The overall base composition of N. temminckii is 30.5% for A, 26.5% for T, 26.4% for C, 16.6% for G, with higher AT content 57.0%. This study will be rather beneficial for understanding the phylogenetic approach of Nipponocypris species and other related genera of cyprinid fishes.

The complete mitochondrial genome of the redigoby Redigobius bikolanus (Perciformes, Gobiidae).

The complete mitochondrial genome of the Biko's redigoby Redigobius bikolanus (Herre, 1927) has been amplified in this study. R. bikolanus contributes in Taiwan, as well as Southeast Asia. The complete mitochondrial genome is 16,464 base pairs (bp) in total length, had the typical vertebrate mitochondrial gene arrangement, including 13 protein-coding genes, 2 rRNAs, 22 tRNAs, 1 control region (CR) and 1 light-strand replication origin (OL). The overall base composition of R. bikolanus is 28.9% for A, 26.7% for T, 28.5% for C, 15.9% for G, with higher AT content 55.6%. This study will contribute and be rather beneficial for understanding the phylogenetic approach of genus Redigobius.

The complete mitochondrial genome of Hoeven's mullet-goby Hemigobius hoevenii (Bleeker) (Teleostei, Gobiidae).

In this study, the complete mitochondrial genome of Hoeven's mullet-goby Hemigobius hoevenii (Bleeker, 1851) collected from Malay Peninsula has been amplified and sequenced. The complete mitochondrial genome is 16,469 base pairs (bp) in total length, had the typical vertebrate mitochondrial gene arrangement, including 13 protein-coding genes, 2 rRNAs, 22 tRNAs, 1 control region (CR) and 1 light-strand replication origin (OL). The overall base composition of H. hoevenii is 27.1% for A, 26.6% for T, 29.0% for C, 17.3% for G, with higher AT content 53.7%. This study will contribute for understanding the phylogenetic approach of Hemigobius species as well as the related genera of gobiid fishes.

The complete mitochondrial genome of the Korean minnow Nipponocypris koreanus (Cypriniformes, Cyprinidae).

The complete mitochondrial genome of the Korean minnow Nipponocypris koreanus (Kim et al., 2005) which is endemic to Korean Peninsula has been amplified and sequenced in this study. The complete mitochondrial genome is 16,615 base pairs (bp) in total length, had the typical vertebrate mitochondrial gene arrangement, including 13 protein-coding genes, 2 rRNAs, 22 tRNAs, 1 control region (CR) and 1 light-strand replication origin (OL). The overall base composition of N. koreanus is 30.0% for A, 26.8% for T, 26.1% for C, 17.1% for G, with higher AT content 56.8%. This study will be rather beneficial for understanding the phylogenetic approach of Nipponocypris species and other related genera of cyprinid fishes.

The complete mitochondrial genome of the small-scaled Wu's goby Wuhanlinigobius polylepis (Perciformes, Gobiidae).

In this study, the complete mitochondrial genome of the small-scaled Wu's goby Wuhanlinigobius polylepis (Wu & Ni, 1985) found in eastern and southern China has been amplified and sequenced. The complete mitochondrial genome is 16,496 base pairs (bp) in total length, had the typical vertebrate mitochondrial gene arrangement, including 13 protein-coding genes, 2 rRNAs, 22 tRNAs, 1 control region (CR) and 1 light-strand replication origin (OL). The overall base composition of Wuhanlinigobius polylepis is 28.6% for A, 29.0% for T, 26.7% for C, 15.7% for G, with higher AT content 57.6%. This study will contribute for further understanding the molecular phylogenetic anaylsis in Hemigobius generic group of gobiid fishes.

The Recognition and Molecular Phylogeny of Mugilogobius mertoni Complex (Teleostei: Gobiidae), with Description of a New Cryptic Species of M. flavomaculatus from Taiwan.

Shih-Pin Huang, I-Shiung Chen, Mana M. N. Yung, and Kwang-Tsao Shao (2016) Mugilogobius mertoni (Weber, 1911) is considered as a widely distributed species around the Indo- West Pacific region, and several nominal species are considered as junior synonyms of M. mertoni. However, in our recent study, several different morphological types of M. mertoni were observed, they were collected from Taiwan, Palau and Phuket Island. This study aimed to investigate the taxonomic status of those M. mertoni-like individuals, we also attempted to assess their phylogenetic relationship base on combined mitochondrial DNA ND5, Cyt b and D-loop sequences. The present morphological and molecular evidences suggested that the current M. mertoni could be regarded as a species complex, and several cryptic species might be included in M. mertoni complex. One of these which collected from Taiwan is described as a new species, Mugilogobius flavomaculatus n. sp. based on both morphological and molecular evidence in this study. The phylogenetic tree also revealed that M. flavomaculatus n. sp. is the closest to M. mertoni. M. flavomaculatus n. sp. and its sister species M. mertoni are found to have different niches in the same estuary. Moreover, stable morphological characters and nuclear gene RAG2 also clearly show that no hybridization is detected in between M. flavomaculatus n. sp. and M. mertoni. Except the present new species, taxonomic status of all junior synonyms refers to M. mertoni are also discussed.

Differentiation and apoptosis induction by lovastatin and γ-tocotrienol in HL-60 cells via Ras/ERK/NF-κB and Ras/Akt/NF-κB signaling dependent down-regulation of glyoxalase 1 and HMG-CoA reductase.

Glyoxalase 1 (GLO1) and HMG-CoA reductase (HMGCR) are highly expressed in most tumor cells and little in normal cells. In this study, treatment of HL-60 cells with lovastatin induced characteristic apoptosis in a dose-dependent manner. We demonstrated that lovastatin treatment inhibited Ras and Raf protein translocation to cell membrane and eliminated the phosphorylation of the downstream effectors Akt and ERK, and the subsequent NF-κB translocation into nucleus. Specific inhibitors and γ-tocotrienol confirmed the Ras/Raf/ERK/NF-κB/GLO1 and Ras/Akt/NF-κB/GLO1 pathways. Data revealed that lovastatin induced HL-60 cell death was attenuated by mevalonate treatment. We demonstrated also that γ-tocotrienol showed its apoptotic effect on the HL-60 cell through the same pathway. γ-Tocotrienol enhanced the apoptotic effect of lovastatin through the down-regulation of GLO1 and HMGCR resulting in an increase of methylglyoxal and a decrease of cholesterol and led to the apoptosis of HL-60 cells. Data also revealed that both lovastatin and gamma-tocotrienol induced significant HL-60 cell differentiation. These results suggest that both lovastatin and gamma-tocotrienol could induce differentiation and followed by apoptosis.