Hsiao-Wei Kao

Evolution of infectious bronchitis virus in Taiwan: Characterisation of RNA recombination in the nucleocapsid gene

Abstract Avian infectious bronchitis virus (IBV) belongs to the Coronaviridae family and causes significant economic loss in Taiwan (TW), even in flocks that have been extensively immunised with Massachusetts (Mass)-serotype vaccines. Phylogenetic analysis of all non-structural and most structural genes shows that TW IBV is genetically distinct from the US strain and more similar to Chinese (CH) IBV. In contrast, the nucleocapsid (N) gene of TW IBV presents phylogenetic incongruence. RNA recombination at the 5′ end of the N gene between TW and US IBV is shown to be responsible for this discordance. Surprisingly, the recombinant N gene is found in all of tested TW IBV isolates, suggesting that a recombination event gave origin to a founder lineage. Our data indicate that RNA recombination in the recombinant 5′ end of the N gene may have caused the emergence of the current IBV population in Taiwan.

Genetic Diversity and Differentiation of Gray Mullet (Mugil cephalus) in the Coastal Waters of Taiwan

From mid-December to late January, schools of mature gray mullet (Mugil cephalus) migrate southward along the coastal waters of China to Taiwan for spawning. It has been proposed that there is no genetic differentiation of gray mullet in the coastal waters of Taiwan. To test this hypothesis, complete cytochrome b (cyt b) DNA sequences of 98 individuals of gray mullet, two individuals of Liza macrolepis, and three individuals of L. affinis were amplified by polymerase chain reaction and sequenced. Phylogenetic trees reconstructed with the Bayesian, maximum likelihood, maximum parsimony, and neighbor-joining methods all support the existence of three monophyletic groups (denoted as groups 1, 2, and 3), with net evolutionary divergences (p-distances) between groups ranging from 5.1% to 6.6%. To estimate the relative abundance of each group, a PCR-RFLP method was developed to examine 600 juveniles collected from October 2006 to March 2007. Groups 1, 2, and 3 comprised 85%, 3%, and 12% of the samples, respectively. Juveniles of groups 1 and 3 could be found as early as November, but juveniles of group 2 were not found until February. Based on the dates of specimen collection and phylogenetic analyses, we propose that groups 1 and 2 are migratory populations from China and Japan, respectively, whereas group 3 is a resident population in Taiwan.

Evolution of infectious bronchitis virus in Taiwan: positively selected sites in the nucleocapsid protein and their effects on RNA-binding activity.

Abstract RNA recombination has been shown to underlie the sporadic emergence of new variants of coronavirus, including the infectious bronchitis virus (IBV), a highly contagious avian pathogen. We have demonstrated that RNA recombination can give rise to a new viral population, supported by the finding that most isolated Taiwanese (TW) IBVs, similar to Chinese (CH) IBVs, exhibit a genetic rearrangement with the American (US) IBV at the 5’ end of the nucleocapsid (N) gene. Here, we further show that positive selection has occurred at two sites within the putative crossover region of the N-terminal domain (NTD) of the TW IBV N protein. Based on the crystal structure of the NTD, the stereographic positions of both predicted selected sites do not fall close to the RNA-binding groove. Surprisingly, converting either of the two residues to the amino acid present in most CH IBVs resulted in significantly reduced affinity of the N protein for the synthetic RNA repeats of the viral transcriptional regulatory sequence. These results suggest that modulating the amino acid residue at either selected site may alter the conformation of the N protein and affect the viral RNA–N interaction. This study illustrates that the N protein of the current TW IBV variant has been shaped by both RNA recombination and positive selection and that the latter may promote viral survival and fitness, potentially by increasing the RNA-binding capacity of the N protein.

Differential expression of zebrafish gpia and gpib during development

Glucose 6-phosphate isomerase (GPI), alternatively named phosphoglucose isomerase (PGI), autocrine motility factor (AMF) or neuroleukin (NLK), is a sugar metabolic enzyme catalyzing the interconversion between glucose-6-phosphate and fructose-6-phosphate. When secreted out of the cell, it can induce cellular activities of neighboring cells. As a prerequisite to study the function of gpi during development, we analyzed the sequences and expression patterns of zebrafish gpia and gpib. Phylogenetic analyses indicate that gpia and gpib are two paralogs resulting from the teleost fish-specific whole genome duplication. In adult zebrafish, gpia is widely expressed in many tissues, whereas gpib is only expressed in heart, muscle, and at lower abundance in eye. During embryonic development, gpia mRNA is maternally deposited in cleavage and blastula embryos, but gpib mRNA is not. Zygotic expression of gpia mRNA initiates in the primitive gut of segmentation embryos, and its expression increases in eye, cerebellum, hindbrain, heart, pharyngeal arches, pectoral fin buds and gut of hatching embryos at 60hpf (hours post fertilization). At 72hpf, gpia mRNA is strongly expressed in the retina of eye, tectum, hindbrain, and gut derivatives such as liver, intestine and circular smooth muscle layer of the swim bladder. By contrast, gpib is expressed in the yolk syncytial nuclei of late blastula, gastrula and segmentation embryos. Somitic gpib expression initiates in late pharyngula embryos, and it is most prominent in the muscle of hatching embryos around 48-60hpf. Transcript of gpib is also expressed in the pectoral fins and portions of the pharyngeal arches in the hatching period. The differential expression of gpia and gpib in zebrafish suggests partition or divergence of gpi functions between the two duplicates.

Erratum to: Mycena kentingensis, a new species of luminous mushroom in Taiwan, with reference to its culture method

Mycena kentingensis, a new luminous mushroom, was discovered in the tropical forest of Kenting National Park in Taiwan. The pileus of M. kentingensis is 3–8 mm in diameter and emits green light in the dark. The dorsal surface of the pileus is covered with short and white spines. The lamellae are nearly free. The stipe is puberulous with a white basal disc about 2 mm broad. Cheilocystidia are clavate to obpyriform, 16–40 × 4–8 μm, and have dense excrescences on the upper half or two-thirds of the cell. The pileus lacks cherocyte, and the stipe lacks caulocystidium. To further identify the species by the DNA barcode method, the internal transcribed spacer (ITS) DNA sequence of M. kentingensis was amplified by polymerase chain reaction, sequenced, and searched for in the GenBank. The hit with the maximum score was an uncultured fungus clone KFRI2121 (accession number HQ662846) with 88 % sequence identity. For those ITS sequences associated with Latin binominal names in the GenBank, the hit with the maximum score was Mycena stylobates with 87 % sequence identity. Phylogenetic analyses by Bayesian and maximum likelihood methods showed that M. kentingensis is close to M. stylobates and M. adscendens. To develop its culture method, mycelia of M. kentingensis were isolated. The optimal temperature for mycelial growth on potato-dextrose agar (PDA) medium was 24 °C. Commercial compost with the addition of 20 % rice bran and 50 % water was appropriate for fruit-body formation. The new species of Mycena kentingensis brings the total of luminous fungi in the world to 74 species.

Anadeninus Parvipenis (Pulmonata: Anadenidae), a Newly Recorded Species from Taiwanfica

Anadeninus parvipenis Wiktor, Chen et Wu (Anadenidae), an endangered species in China Species Red List, is recorded for the first time in Lalashan Forest Reserve, Taiwan. The spermatheca of this species is elongated, penis is small and epiphallus is long and thick. In this study, a description of this newly recorded species is given and the colored photographs of specimens, SEM images of radula and jaw and dissection of genitalia are also provided for aid of identification of this species.