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Anthropogenic acidification of water is an on-going environmental disaster for freshwater fishes. Fishes rely on ammonia excretion to eliminate the excess acid and mitigate the harmful effects; however, it remains largely unknown how ammoniagenesis occurs and is coordinated with ammonia excretion upon acidic stress. Medaka (Oryzias latipes) was used to examine the effects of acidic stress on ammonia production and excretion. We reveal an undiscovered ammonia-producing cell type that is rich in glutaminase (GLS) and located adjacent to the ammonia-excreting ionocytes, Na

We analyze 214 freshly laid eggs belonging to 16 species across three orders of Class Reptilia. Using mechanical compression tests, we measure each egg’s absolute stiffness (K, unit: N m−1) and relative stiffness (C number). The effective Young’s modulus, E, was obtained by combining experimental and numerical methods. The mineral (CaCO3) content was measured by acid–base titration, the microstructures by scanning electron microscopy (SEM), and the crystallography by electron backscatter diffraction (EBSD). We find that the C number of reptilian eggs is, on average, higher than that of bird eggs, indicating that reptilian eggs are stiffer with respect to the egg mass than birds. However, Young’s moduli of the reptilian eggshells (32.85 ± 3.48 GPa) are similar to those of avian eggshells (32.07 ± 5.95 GPa), even though those eggshells have different crystal forms, microstructures, and crystallography. Titration measurement shows that the reptilian eggshells are highly mineralized (>89% for nine Testudines species and 96% for Caiman crocodilus). Comparing the species with aragonite and calcite crystals, we find that calcite shells, including those of the Kwangsi gecko (inner part) and spectacled caiman (outer part), generally have larger grains than the aragonite ones. However, the grain size is not correlated to the effective Young’s modulus. Also, as measured by the C number, the aragonite shells are, on average, stiffer than the calcite ones (except for the Kwangsi gecko), primarily due to their thicker shells.

Gut microbiota are involved in many physiological functions such as metabolism, brain development, and neurodegenerative diseases. Many microbes in the digestive tract do not maintain a constant level of their relative abundance but show daily oscillations under normal conditions. Recent evidence indicates that chronic jetlag, constant darkness, or deletion of the circadian core gene can alter the composition of gut microbiota and dampen the daily oscillation of gut microbes. However, the neuronal circuit responsible for modulating gut microbiota remained unclear. Using genetic mouse models and 16s rRNA metagenomic analysis, we find that light‐dark cycle information transmitted by the intrinsically photosensitive retinal ganglion cells (ipRGCs) is essential for daily oscillations of gut microbes under temporal restricted high‐fat diet conditions. Furthermore, aberrant light exposure such as dim light at night (dLAN) can alter the composition, relative abundance, and daily oscillations of gut microbiota. Together, our results indicate that external light‐dark cycle information can modulate gut microbiota in the direction from the brain to the gut via the sensory system.

Finite element analysis (FEA) was used to conduct mechanical analyses on eggshells of giant birds, and relate this to the evolution and reproductive behavior of avian species. We aim to (1) investigate mechanical characteristics of eggshell structures of various ratite species, enabling comparisons between species with or without reversed sexual size dimorphism (RSSD), (2) quantify the safety margin provided by RSSD, (3) determine whether the Williams’ egg can have been incubated by an extinct giant bird Genyornis newtoni, (4) determine the theoretical maximum body mass for contact incubation. We use a dimensionless number C to quantify relative shell stiffness with respect to the egg size, allowing for comparison across wide body masses. We find that RSSD in moas significantly increases the safety margin of contact incubation by the lighter males. However, their safety margins are still smaller than those of the moa species without RSSD. Two different strategies were adopted by giant birds—one is RSSD and thinner shells, represented by some moa species, the other is no RSSD and regular shells, represented by the giant elephant bird. Finally, we predicted that the upper limit of body mass for contact incubation was 2000 kg.

Gut microbiota has been shown to involve in many physiological functions such as metabolism, brain development, and neuron degeneration disease. Intriguingly, many microbes in the digestive tract do not maintain a constant level of their relative abundance but show daily oscillations under normal conditions. Recent evidence indicates that chronic jetlag, constant darkness, or deletion of the circadian core gene can alter the composition of gut microbiota and dampen the daily oscillation of gut microbes. These studies suggest that the interaction between the host circadian clock and the light-dark cycle plays an important role in gut homeostasis and microbiota. However, how or whether environmental factors such as the light-dark cycle could modulate gut microbiota is still poorly understood. Using genetic mouse models and 16s rRNA metagenomic analysis, we found that light-dark cycle information transmitted by the ipRGC-sympathetic circuit was essential for daily oscillations of gut microbes under temporal restricted high fat diet condition. Furthermore, aberrant light exposure such as dim light at night (dLAN), acting through intrinsically photosensitive retinal ganglion cells (ipRGCs), could alter the composition, relative abundance, and daily oscillations of gut microbiota. Together, our results indicate that external stimulation, such as light-dark cycle information, through the sensory system can modulate gut microbiota in the direction from the brain to the gut.

ObjectiveThe gut microbiota-derived metabolite, trimethylamine N-oxide (TMAO) plays an important role in cardiovascular disease (CVD). The fasting plasma TMAO was shown as a prognostic indicator of CVD incident in patients and raised the interest of intervention targeting gut microbiota. Here we develop a clinically applicable method called oral carnitine challenge test (OCCT) for TMAO-related therapeutic drug efforts assessment and personalising dietary guidance.DesignA pharmacokinetic study was performed to verify the design of OCCT protocol. The OCCT was conducted in 23 vegetarians and 34 omnivores to validate gut microbiota TMAO production capacity. The OCCT survey was integrated with gut microbiome, host genotypes, dietary records and serum biochemistry. A humanised gnotobiotic mice study was performed for translational validation.ResultsThe OCCT showed better efficacy than fasting plasma TMAO to identify TMAO producer phenotype. The omnivores exhibited a 10-fold higher OR to be high TMAO producer than vegetarians. The TMAO-associated taxa found by OCCT in this study were consistent with previous animal studies. The TMAO producer phenotypes were also reproduced in humanised gnotobiotic mice model. Besides, we found the faecal CntA gene was not associated with TMAO production; therefore, other key relevant microbial genes might be involved. Finally, we demonstrated the urine TMAO exhibited a strong positive correlation with plasma TMAO (r=0.92, pConclusionThe OCCT can be used to identify TMAO-producer phenotype of gut microbiota and may serve as a personal guidance in CVD prevention and treatment.Trial registration numberNCT02838732; Results.

Using two advanced sequencing approaches, Illumina and PacBio, we derive the entire Dscam gene from an M2 assembly of the complete Penaeus monodon genome. The P. monodon Dscam (PmDscam) gene is ~266 kbp, with a total of 44 exons, 5 of which are subject to alternative splicing. PmDscam has a conserved architectural structure consisting of an extracellular region with hypervariable Ig domains, a transmembrane domain, and a cytoplasmic tail. We show that, contrary to a previous report, there are in fact 26, 81 and 26 alternative exons in N-terminal Ig2, N-terminal Ig3 and the entirety of Ig7, respectively. We also identified two alternatively spliced exons in the cytoplasmic tail, with transmembrane domains in exon variants 32.1 and 32.2, and stop codons in exon variants 44.1 and 44.2. This means that alternative splicing is involved in the selection of the stop codon. There are also 7 non-constitutive cytoplasmic tail exons that can either be included or skipped. Alternative splicing and the non-constitutive exons together produce more than 21 million isoform combinations from one PmDscam locus in the P. monodon gene. A public-facing database that allows BLAST searches of all 175 exons in the PmDscam gene has been established at http://pmdscam.dbbs.ncku.edu.tw/.

Background In pancrustaceans, the Down syndrome cell adhesion molecule (Dscam) is an extraordinarily complex, single-locus gene, with the potential for generating thousands of isoforms by combining alternative splicing exons. In the present study, we used two advanced sequencing approaches, Illumina and PacBio, with hybrid assembly to analyze the entire Dscam genomic structure in Penaeus monodon. Results The P. monodon Dscam (PmDscam) genome was ~250 kbp, with a total of 175 constitutive and alternative splicing exons. Analysis of PmDscam cDNA and genomics revealed a conserved architectural structure consisting of an extracellular region with hypervariable Ig domains, a transmembrane domain, and a cytoplasmic tail. While the number of splicing exon variants in N-terminal Ig2, N-terminal Ig3 and the entirety of Ig7 were previously reported to be 28, 43 and 19, we now show that there are in fact 26, 81 and 26 alternative exons in these regions, respectively. We also identified two alternative variants of two exons in the cytoplasmic tail, as well as 7 cytoplasmic tail elements that can either be included or skipped. The presence of three stop codon sites in the cytoplasmic tail region means that alternative splicing is involved in the selection of the stop codon. Conclusions In total, alternative splicing provides for 54,756 potential combinations in the extracellular region, plus 512 potential combinations in the cytoplasmic tail, all derived from one PmDscam genome locus. We have also established a public-facing PmDscam genome database (http://pmdscam.dbbs.ncku.edu.tw/) to facilitate future research on characterizing the involvement of Dscam in pancrustacean immunity.

Thermal homeostasis of mammals is constrained by body-size scaling. Consequently, small mammals require considerable energy to maintain a high mass-specific metabolic rate (MSMR) and sustain target body temperature. In association with gut microbiota, mammalian hosts acquire absorbable molecules and fulfill their metabolic requirements. Our objective was to characterize gut microbes in wild mammals and relate those findings to host body-size scaling. Two large (Petaurista philippensis grandis and P. alborufus lena), one medium (Trogopterus xanthipes) and one small (Pteromys volans orii) species of flying squirrels (FS) were studied. Using 16S rRNA genes, 1,104 OTUs were detected from four FS, with 1.99% of OTUs shared among all FS. Although all FS gut microbiota were dominated by Firmicutes, they were constituted by different bacterial families. Moreover, Bacteroidetes accounted for up to 19% of gut microbiota in small FS, but was absent in large FS. Finally, based on metagenome predictions, carbohydrate and amino acid metabolism genes were enriched in small body-size FS. In conclusion, gut microbiota compositions and predictive metabolic functions were characteristic of body-size in FS, consistent with their adaptations to folivorous dietary niches.

To determine the genetic diversity and geographic distribution of Cao Bang virus (CBNV) and to ascertain the existence of CBNV-related hantaviruses, natural history collections of archival tissues from Chinese mole shrews (Anourosorex squamipes) and Taiwanese mole shrews (Anourosorex yamashinai), captured in Guizho Province, People’s Republic of China, and in Nantou County, Taiwan, in 2006 and 1989, respectively, were analyzed for hantavirus RNA by RT-PCR. Pair-wise alignment and comparison of the S-, M- and L-segment sequences indicated CBNV in two of five Chinese mole shrews and a previously unrecognized hantavirus, named Xinyi virus (XYIV), in seven of 15 Taiwanese mole shrews. XYIV was closely related to CBNV in Vietnam and China, as well as to Lianghe virus (LHEV), recently reported as a distinct hantavirus species in Chinese mole shrews from Yunnan Province in China. Phylogenetic analyses, using maximum-likelihood and Bayesian methods, showed that XYIV shared a common ancestry with CBNV and LHEV, in keeping with the evolutionary relationship between Anourosorex mole shrews. Until such time that tissue culture isolates of CBNV, LHEV and XYIV can be fully analyzed, XYIV and LHEV should be regarded as genetic variants, or genotypes, of CBNV.

Gut microbial communities of animals are influenced by diet and seasonal weather changes. Since foraging strategies of wild animals are affected by phenological changes, gut microbial communities would differ among seasons. However, interactions of plant-animal-microbiota with seasonal changes have not been well characterized. Here, we surveyed gut microbial diversity of Siberian flying squirrels (Pteromys volans orii) from a natural forest in Hokkaido during spring and summer of 2013 and 2014. Additionally, we compared microbial diversity to temperature changes and normalized difference vegetation index (NDVI). Changes in both seasonal temperature and phenology were significantly associated with alterations in gut microbiota. There were two clusters of OTUs, below and above 20 °C that were significantly correlated with low and high temperatures, respectively. Low-temperature cluster OTUs belonged to various phyla, whereas the high-temperature cluster was only constituted by Firmicutes. In conclusion, gut microbiota of Siberian flying squirrels varied with environmental changes on an ecological scale.

Mammalian herbivores rely on microbial activities in an expanded gut chamber to convert plant biomass into absorbable nutrients. Distinct from ruminants, small herbivores typically have a simple stomach but an enlarged cecum to harbor symbiotic microbes; however, knowledge of this specialized gut structure and characteristics of its microbial contents is limited. Here, we used leaf-eating flying squirrels as a model to explore functional characteristics of the cecal microbiota adapted to a high-fiber, toxin-rich diet. Specifically, environmental conditions across gut regions were evaluated by measuring mass, pH, feed particle size, and metabolomes. Then, parallel metagenomes and metatranscriptomes were used to detect microbial functions corresponding to the cecal environment. Based on metabolomic profiles, >600 phytochemical compounds were detected, although many were present only in the foregut and probably degraded or transformed by gut microbes in the hindgut. Based on metagenomic (DNA) and metatranscriptomic (RNA) profiles, taxonomic compositions of the cecal microbiota were dominated by bacteria of the Firmicutes taxa; they contained major gene functions related to degradation and fermentation of leaf-derived compounds. Based on functional compositions, genes related to multidrug exporters were rich in microbial genomes, whereas genes involved in nutrient importers were rich in microbial transcriptomes. In addition, genes encoding chemotaxis-associated components and glycoside hydrolases specific for plant beta-glycosidic linkages were abundant in both DNA and RNA. This exploratory study provides findings which may help to form molecular-based hypotheses regarding functional contributions of symbiotic gut microbiota in small herbivores with folivorous dietary habits.

Taiwan has a fast-aging population, and it is expected to see an associated increase in the number of older adults with disabilities. We aimed to understand whether Taiwanese older adults’ disability trajectories mediated the association between disablement process factors and later long-term care (LTC) service use. Data were from the nationally representative Taiwan Longitudinal Study on Aging Survey, 1996–2007 (N = 3,429). Trajectories of disability in activities of daily living and instrumental activities of daily living were identified by using growth mixture modeling. Structural equation modeling was applied to examine the effect of disability trajectory as mediator on disablement process factors, such as demographics, number of comorbidities, depression, and leisure activities (LAs), and on later LTC service use. Three distinct disability trajectories were identified among the group of surveyed older adults: healthy (N = 2,998, 87.43 %), progressive disability (N = 363, 10.59 %), and maintained disability (N = 68, 1.98 %). These disability trajectories, either progressive disability or maintained disability trajectory, were found to have a positive mediating effect on the association between disablement process factors (older, lower educational attainment, larger number of comorbidities, and depressive symptoms) and greater use of LTC services. However, encouraging older adults to engage in LAs reduced later use of LTC services through both total and significant mediating effect of postponing development of disability trajectory in older Taiwanese adults. These findings suggest that an active lifestyle is important for older adults’ physical health and to decrease the societal burden of LTC service use.

Sequential polygyny is a reproductive strategy that allows males to continue to mate and compensates for the loss of future breeding opportunities incurred by parental care (i.e. egg attendance). Using the frog Kurixalus eiffengeri, we tested predictions that (1) attending males fathered two, overlapping clutches; and (2) that double clutching leads to improved offspring numbers. Using five microsatellite DNA markers, we genotyped 15 pairs of overlapping clutches, which differed slightly in developmental stage at a single egg-laying site. Parentage analyses showed at least 12 of 15 pairs of overlapping egg clutches were sired by the attending male mated with different females, providing the first genetic evidence to support an earlier prediction that attending males sired both egg clutches. Field surveys found a low incidence of overlapping clutches (4.9% of 263 egg-occupied stumps), suggesting sequential polygyny is uncommon. Stumps with multiple clutches contained significantly more eggs than stumps with single clutches but hatched similar number of tadpoles. Results suggest that continuous calling that attracts females during egg attendance is a reproductive tactic that maximizes mating opportunities. However, adoption of the sequential polygyny tactic may only result in marginal fitness gains for males that are traded off against average higher egg mortality in larger egg clutches.

Parthenogenesis has been known in a variety of animal taxa and is commonly found in oligochaetes. Many pheretimoid earthworms in Megascolecidae have been known to be or suspected to be parthenogenetic, but there has not been any genetic investigation on them. Amynthas catenus and Amynthas hohuanmontis are earthworms belonging to the Pheretima complex of the family Megascolecidae. Both are endemic to Mt. Hohuan at an elevation of about 3000 m in central Taiwan, and both have spermathecae from three pairs in segments VI–VIII (sexthecal) to absence (athecal) and reproductive organs in different degrees of degeneration. Here we present the genetic evidence revealed by amplified fragment length polymorphism (AFLP) markers to confirm the parthenogenetic mode of reproduction in A. catenus and A. hohuanmontis . Sixty-two selective primer combinations were used to generate a total of 4593 and 4812 bands for A. catenus and A. hohuanmontis , respectively. We found that the two earthworms are automictic (meiotic) parthenogens with offsprings showing slightly reduced number of AFLP markers compared with their parents. There was no sperm in any of the spermathecae examined. Selection and reproductive success among different genotypes of the two earthworms will be an important topic for future investigation.

The objective was to apply a novel modification of a genome-wide, comparative cytogenetic technique (comparative genomic hybridization, comparative genomic hybridization (CGH)), to study species belonging to the myrmecophagous (ant/termite eating) mammalian orders/superorders (Pholidota, Tubulidentata, Carnivora, and Xenarthra), as a model for other applications in mammalian systematics and conservation biology. In this study, CGH was applied to high-quality metaphase spreads of pangolin (Pholidota), using probes of sloth and canine (Xenarthra and Carnivora, respectively) genomic DNA labeled with different fluorophores, thereby facilitating analysis of the visible color spectrum on pangolin karyotypes. Our results posited that pholidotes are closer to carnivores than to xenarthrans, which confirmed the current consensus that myrmecophagy in these mammalian lineages was more likely because of homoplasy (convergent evolution) than being an ancestral character. Since the modified CGH technique used is genome-wide, has chromosome-level resolution, and does not need full genome sequencing, it has considerable potential in systematics and other fields.

The worldwide distributed house mouse, Mus musculus, is subdivided into at least three lineages, Mus musculus musculus, Mus musculus domesticus, and Mus musculus castaneus. The subspecies occur parapatrically in a region considered to be the cradle of the species in Southern Asia (‘central region’), as well as in the rest of the world (‘peripheral region’). The morphological evolution of this species in a phylogeographical context is studied using a landmark-based approach on mandible morphology of different populations of the three lineages. The morphological variation increases from central to peripheral regions at the population and subspecific levels, confirming a centrifugal sub-speciation within this species. Furthermore, the outgroup comparison with sister species suggests that M. musculus musculus and populations of all subspecies inhabiting the Iranian plateau have retained a more ancestral mandible morphology, suggesting that this region may represent one of the relevant places of the origin of the species. Mus musculus castaneus, both from central and peripheral regions, is morphologically the most variable and divergent subspecies. Finally, the results obtained in the present study suggest that the independent evolution to commensalism in the three lineages is not accompanied by a convergence detectable on jaw morphology. © 2012 The Linnean Society of London, Biological Journal of the Linnean Society, 2012, 105, 635–647.

Background Microbial diversity and community structures in acidic hot springs have been characterized by 16S rRNA gene-based diversity surveys. However, our understanding regarding the interactions among microbes, or between microbes and environmental factors, remains limited. Results In the present study, a metagenomic approach, followed by bioinformatics analyses, were used to predict interactions within the microbial ecosystem in Shi-Huang-Ping (SHP), an acidic hot spring in northern Taiwan. Characterizing environmental parameters and potential metabolic pathways highlighted the importance of carbon assimilatory pathways. Four distinct carbon assimilatory pathways were identified in five dominant genera of bacteria. Of those dominant carbon fixers, Hydrogenobaculum bacteria outcompeted other carbon assimilators and dominated the SHP, presumably due to their ability to metabolize hydrogen and to withstand an anaerobic environment with fluctuating temperatures. Furthermore, most dominant microbes were capable of metabolizing inorganic sulfur-related compounds (abundant in SHP). However, Acidithiobacillus ferrooxidans was the only species among key rare microbes with the capability to fix nitrogen, suggesting a key role in nitrogen cycling. In addition to potential metabolic interactions, based on the 16S rRNAs gene sequence of Nanoarchaeum-related and its potential host Ignicoccus-related archaea, as well as sequences of viruses and CRISPR arrays, we inferred that there were complex microbe-microbe interactions. Conclusions Our study provided evidence that there were numerous microbe-microbe and microbe-environment interactions within the microbial community in an acidic hot spring. We proposed that Hydrogenobaculum bacteria were the dominant microbial genus, as they were able to metabolize hydrogen, assimilate carbon and live in an anaerobic environment with fluctuating temperatures. Electronic supplementary material The online version of this article (doi:10.1186/s12864-015-2230-9) contains supplementary material, which is available to authorized users.

Despite the fact that some educational researchers believe that laboratory courses promote outcomes in cognitive and affective domains in science learning, others have argued that laboratory courses are costly in relation to their value. Moreover, effective measurement of student learning in the laboratory is an area requiring further investigation. The present study set out to examine learning outcomes by measuring students' academic performance and their skill in writing research proposals and investigated students' views about laboratory courses through interviews. Comparisons were made between students taking lectures only (control group) and students enrolled in both lecture and laboratory courses (experimental group). A total of 78 undergraduate students participated in the study. Stepwise discriminant analysis, analysis of covariance, and multidimensional scaling were used to analyse the data. Results showed that students in the experimental group did not perform as well as students in the control ...

The rhacophorid frog, Kurixalus eiffingeri, is one of only a few frog species that exhibits polyandry and paternal care of eggs. Previous studies predicted that multiple paternity within an egg clutch could influence the degree of paternal care and reproductive strategies. We used microsatellite DNA markers to assess the prevalence of multiple paternity within egg clutches and the relationship between male paternal care and the percent of male’s genetic contribution to the clutch, i.e., paternal share. We conducted field observations of paternal care and collected tissues from both male frogs and tadpoles for parentage analyses. Our results showed that at least five out of 31 egg clutches had multiple paternity. Attending males were always the genetic fathers of some, if not all of the eggs in the clutch they guarded. All egg clutches except one were attended by one male frog but the attending male did not necessarily sire the majority of offspring. Multiple paternity in all cases consisted of two fathers and one mother and most likely resulted from synchronous polyandry. Paternal care effort correlated significantly with the male’s genetic contribution to the clutch, suggesting that male frogs adjust the effort expended in care in response to paternal share. In addition, our results suggest that externally fertilizing species with parental care and multiple paternity may develop novel reproductive and behavioral strategies to safeguard their parental investment and overcome sperm competition.

Here we provide a genome-wide, high-resolution map of the phylogenetic origin of the genome of most extant laboratory mouse inbred strains. Our analysis is based on the genotypes of wild-caught mice from three subspecies of Mus musculus. We show that classical laboratory strains are derived from a few fancy mice with limited haplotype diversity. Their genomes are overwhelmingly Mus musculus domesticus in origin, and the remainder is mostly of Japanese origin. We generated genome-wide haplotype maps based on identity by descent from fancy mice and show that classical inbred strains have limited and non-randomly distributed genetic diversity. In contrast, wild-derived laboratory strains represent a broad sampling of diversity within M. musculus. Intersubspecific introgression is pervasive in these strains, and contamination by laboratory stocks has played a role in this process. The subspecific origin, haplotype diversity and identity by descent maps can be visualized using the Mouse Phylogeny Viewer (see URLs).

The systematic status of Pholidota has been a matter of debate, particularly regarding the apparent inconsistency between morphological and molecular studies. The Sry gene, a master regulator of male sex determination in eutherian mammals, has not yet been used for phylogenetic analyses of extant mammals. The objective of the present study was to clone and characterize the complete gene (1300 base pairs; bp) and amino acid sequences (229 residues) of Sry from the Formosan pangolin (Manis pentadactyla pentadactyla), a member of Pholidota. The Sry amino acid identity between pangolin and other reported species ranged from 42.5% (mouse, Mus musculus) to 84.1% (European hare, Lepus europaeus). Sequence conservation was primarily in the high motility group (HMG) box (234 bp), whereas homology outside the HMG box was low. The cloned Sry was mapped to the pangolin Y chromosome by fluorescence in situ hybridization (FISH); this was confirmed to be the first Y-borne molecular marker identified in Pholidota. Based on Bayesian phylogenetic analysis for Sry HMG sequences from 36 representative taxa, including the Formosan pangolin, Pholidota was more closely related to Carnivora than to Xenarthra, consistent with the emerging molecular tree inferred from markers not located on the Y chromosome. In conclusion, this study characterized the gene structure of Sry of the Formosan pangolin and provided insights into the phylogenetic position of Pholidota.

The black tiger shrimp (Penaeus monodon) is an ecologically and economically important penaeid species and is widely distributed in the Indo-Pacific region. Here we investigated the genetic diversity of P. monodon (n = 355) from eight geographical regions by genotyping at 10 microsatellite loci. The average observed heterozygosity at various loci ranged from 0.638 to 0.743, indicating a high level of genetic variability in this region. Significant departures from Hardy-Weinberg equilibrium caused by heterozygote deficiency were recorded for most loci and populations. Pairwise F(ST) and R(ST) values revealed genetic differentiation among the populations. Evidence from the assignment test showed that the populations in the West Indian Ocean were unique, whereas other populations examined were partially admixed. In addition, the non-metric multidimensional scaling analysis indicated the presence of three geographic groups in the Indo-Pacific region, i.e. the African populations, a population from western Thailand and the remaining populations as a whole. We also sequenced and analysed the mitochondrial control region (mtCR) in these shrimp stocks to determine whether the nuclear and mitochondrial genomes show a similar pattern of genetic differentiation. A total of 262 haplotypes were identified, and nucleotide divergence among haplotypes ranged from 0.2% to 16.3%. Haplotype diversity was high in all populations, with a range from 0.969 to 1. Phylogenetic analysis using the mtCR data revealed that the West Indian Ocean populations were genetically differentiated from the West Pacific populations, consistent with the microsatellite data. These results should have implications for aquaculture management and conservation of aquatic diversity.

Insulin genes in mouse and rat compose a two-gene system in which Ins1 was retroposed from the partially processed mRNA of Ins2. When Ins1 originated and how it was retained in genomes still remain interesting problems. In this study, we used genomic approaches to detect insulin gene copy number variation in rodent species and investigated evolutionary forces acting on both Ins1 and Ins2. We characterized the phylogenetic distribution of the new insulin gene (Ins1) by Southern analyses and confirmed by sequencing insulin genes in the rodent genomes. The results demonstrate that Ins1 originated right before the mouse–rat split (∼20 MYA), and both Ins1 and Ins2 are under strong functional constraints in these murine species. Interestingly, by examining a range of nucleotide polymorphisms, we detected positive selection acting on both Ins2 and Ins1 gene regions in the Mus musculus domesticus populations. Furthermore, three amino acid sites were also identified as having evolved under positive selection in two insulin peptides: two are in the signal peptide and one is in the C-peptide. Our data suggest an adaptive divergence in the mouse insulin two-gene system, which may result from the response to environmental change caused by the rise of agricultural civilization, as proposed by the thrifty-genotype hypothesis.

Coat color variation due to melanin pigment synthesis in house mice Mus musculus in Asia is described and found to be consistent with Gloger's rule, which states that individuals of endothermic animals are darker in humid habitats than those in drier habitats. Three properties of coat color (hue, value and chroma) were measured, and a lightness variable was derived from a principal components analysis using 428 skin specimens representing three subspecies from 85 localities. Dorsal coat color ranged from yellow through brown to black, whereas ventral coat color ranged from white to black. Dorsal coat color varied less than the ventral color. In our samples, the variation in coat color in natural populations was far less than that observed in the laboratory. We found a significant correlation between the lightness variable of dorsal coat color and precipitation. Dark coat color was observed in more humid and closed habitats (darker background color), and pale coat color in drier, more open habitats (lighter background color). This result might imply the role of concealment as a selective force affecting dorsal coat color that was observed in house mice. We also discussed other selective forces that could affect the coat color variation in house mice, such as resistance to bacterial degradation and thermoregulation. In addition, the color spectra of the dorsal pelage among the three subspecies were different, the major distinction being the environmental background color of the habitats in which they are distributed.

Supplimentary material on metagemic analysis. (DOCX 312 kb)

The presence of the pale chub Zacco platypus (Japanese name, Oikawa) in Taiwan has been suggested to be a result of its inadvertent introduction from Lake Biwa in Japan in the 1980s in conjunction with the Japanese Ayu, Plecoglossus altivelis altivelis, which was released several times into the Tamsui River to restock the extinct Ayu population of Taiwan. However, it is also possible that Z. platypus is native to Taiwan and has not been previously described for reasons of its narrow range. Knowledge of the colonizing history of Z. platypus is of considerable importance because it provides insight into the evolutionary process and, hence, impacts management decisions regarding this species in Taiwan. A portion of the mitochondrial D-loop was sequenced for 77 specimens from five populations of Z. platypus from Japan and Taiwan. A total of 22 haplotypes were identified, and nucleotide divergence among haplotypes ranged from 0.20% to 2.82%. Haplotype diversity was high in all populations examined, with a range from 0.718 in the Tagiri River population to 0.909 in the Lake Biwa population. Phylogenetic and statistical parsimony analyses of the molecular data revealed a close genetic relationship between Taiwanese and Japanese Z. platypus and supported the previous report that the Taiwanese Z. platypus originated in Lake Biwa in Japan.

Ma, G.-C., Tsao, H.-S., Lu, H.-P. & Yu, H.-T. (2006). AFLPs congruent with morphological differentiation of Asian common minnow Zacco (Pisces: Cyprinidae) in Taiwan. — Zoologica Scripta, 35 , 341–351. Morphological and amplified fragment length polymorphism (AFLP) analyses were used to examine the evolutionary relationships among Asian common minnows ( Zacco spp.) in Taiwan, where four morphotypes were recently reported. Congruent results from the two sets of analyses indicate that the four morphotypes represent four separate morphometric clades with distinct AFLPs. Consequently, we conclude that four species exist in Taiwan and the systematics of Zacco should be revised accordingly. We also discuss conservation implications and offer a key to the four proposed Zacco species.

Most members of the large family of rhodopsin-like G-protein-coupled receptors possess an evolutionarily conserved Asp-Arg-Tyr (DRY) motif in the C-terminal region of the third transmembrane domain. Mutations of residues within this motif usually abolish receptor function and, when they occur naturally, can even cause human diseases. By analyzing over 100 mammalian orthologs of the chemoattractant receptor GPR33 we identified several polymorphic and fixed sequence variations within the DRY motif. Unexpectedly, the naturally occurring mutation of Arg(3.50) to His in mouse GPR33 showed no difference from the wild-type receptor in several functional tests. Sequence analysis of GPR33 from Asian house mice revealed the polymorphic existence of Arg(3.50) and His(3.50) alleles in wild-trapped populations, further supporting the functional equivalence of both allelic variants. In contrast, the Arg(3.50) to Gly mutation found in hamster GPR33 inactivates the receptor and may have contributed to pseudogenization of this gene in this species. Functional data with GPR33 variants indicate different receptor- and context-specific consequences of DRY mutations. Our study also reveals GPR33 as a new example illustrating missense mutations as a first step in the pseudogenization process.

The genus Crocidura is one of the largest genera among mammals and widely distributed in Eurasia, Africa, and Southeast Asia. We review the recent advances in taxonomic and phylogenetic studies of this genus in East and Southeast Asia. Sympatric distributions of two or more species are known in several localities. Karyotypes of East and Southeast Asian species are divided into several groups. From karyological comparison and molecular phylogeny, those groups are suggested to have formed by geographical isolation in islands and in the continent in East and Southeast Asia. To clarify evolutionary history of East and Southeast Asian species, comprehensive studies with the West Asian and European species from karyological and molecular approaches are desired.

The identification of different clones is fundamental to the study of population structure among organisms with mixed reproductive modes such as cnidarians. However, due to the low genetic variation of coral mtDNA and contamination by zooxanthellate DNA, very few molecular markers are available for studying the clonal structure of cnidarians. Herein we used four polymorphic loci of microsatellite DNA isolated from a zooxanthellae-free octocoral, Junceella juncea, to study its clonal structure in seven populations collected from three localities in Taiwan. In total, 40 multilocus genotypes were found among 152 colonies, and the number of genotypes (clones) identified in the seven populations ranged from 2 to 16. Each of the 40 multilocus genotypes was restricted to a single population, even where adjacent populations were only 100 m distant. The ratio of observed to expected genotypic diversity (Go:Ge) ranged from 0.217 to 0.650, and Go showed a significant departure from Ge (p

Distinct Taura syndrome virus (TSV) isolates were found in Metapenaeus ensis (isolate Tw2KMeTSV), Penaeus monodon (isolate Tw2KPmTSV) and Litopenaeus vannamei (isolate Tw02LvTSV). Nucleotide sequence analysis of these three isolates revealed differences in the TSV structural protein (capsid protein precursor) gene orf2. TSV ORF2 amino acid sequence comparison and phylogenetic analysis suggested a comparatively close relationship between these three Taiwanese isolates and the Hawaiian isolate HI94TSV. In P. monodon specimens that were naturally and experimentally infected with the Tw2KPmTSV isolate, the virus was contained and shrimps showed no clinical signs of infection. However, when P. monodon was challenged with the Tw2KMeTSV isolate, the virus replicated freely. The ORF2 amino acid sequence of the Tw2KMeTSV isolate differed from that of isolate Tw2KPmTSV in four positions and these differences may account for their phenotypic differences, at least in terms of their ability to replicate in specific hosts.

Major histocompatibility complex (MHC) genes are the most polymorphic loci known for vertebrates. Here we employed five microsatellite loci closely linked to the MHC region in an attempt to study the amount of genetic variation in 19 populations of the southeast Asian house mouse (Mus musculus castaneus) in Taiwan. The overall polymorphism at the five loci was high (He = 0.713), and the level of polymorphism varied from locus to locus. Furthermore, in order to investigate if selection is operating on MHC genes in natural mouse populations, we compared the extent and pattern of genetic variation for the MHC-linked microsatellite loci (the MHC loci) with those for the microsatellite loci located outside the MHC region (the non-MHC loci). The number of alleles and the logarithm of variance in repeat number were significantly higher for the MHC loci than for the non-MHC loci, presumably reflecting linkage to a locus under balancing selection. Although three statistical tests used do not provide support for selection, their lack of support may be due to low statistical power of the tests, to weakness of selection, or to a profound effect of genetic drift reducing the signature of balancing selection. Our results also suggested that the populations in the central and the southwestern regions of Taiwan might be one part of a metapopulation structure.

We analyzed the population structure of the Japanese anchovy (Engraulis japonicus), a small pelagic fish, using 6 microsatellite DNA loci. The anchovy is known to have 2 separate spawning populations, one near northeastern Taiwan in the Pacific Ocean and the other near southwestern Taiwan in the Taiwan Strait. The planktonic larvae then drifted north to the feeding grounds in the East China Sea to advance in their life history. Three populations of the anchovy were analyzed, including 2 temporal population from the northeastern spawning ground (I-Lan 1999 and I-Lan 2000) and one population from the southwestern spawning ground (Peng-Hu 2000). The genetic variability of the 6 loci was high for all the populations. The average numbers of alleles per population ranged from 25.5 to 32.3, and the average observed heterozygosity ranged from 0.559 to 0.650. A significant population differentiation was found between geographic populations but not between the temporal populations. However, the level of geographic differentiation was weak, average FST 0.0088. The significant geographic population structure indicated that the populations of 2 spawning grounds belonged to separate stocks. Moreover, 16 of the 18 population-locus cases showed significant departure from Hardy-Weinberg equilibrium, implying that each spawning population in turn consisted of mixed native stocks. Finally, we posed 3 population models to be evaluated against the genetic data disclosed with the microsatellite markers.

We analyzed population subdivision and gene flow of the Southeast Asian house mouse (Mus musculus castaneus) in Taiwan by using six microsatellite DNA markers. Seven populations of the house mouse (187 individuals), including one from Fukien Province in southeastern China, which is separated from Taiwan by the Taiwan Strait, were analyzed in this study. The overall polymorphic level at the six loci was high (He = 0.76) although individual populations varied in their levels of heterozygosity (He = 0.35-0.83). For the populations within Taiwan, there was no evidence of isolation by distance and the level of gene flow was not (inversely) correlated to geographic distances. Gene flow was estimated to be higher across the Taiwan Strait than within the island of Taiwan. These observations of gene flow cannot be understood unless in the context of the historical human settlements and agricultural expansion, and the commensal habits of the species. We also discussed the causes of population subdivision and genetic variation among populations in terms of ecological characteristics of the house mouse in Taiwan.

The aims of this study were to investigate (1) whether and (2) the extent to which the trajectory of Taiwanese older adults’ leisure activities (LA) mediated the potential association between their sociodemographic factors and their functional disability trajectory. Longitudinal data from four waves of the Taiwan Longitudinal Study on Aging (TLSA), collected between 1996 and 2007, were used for analysis (N = 3,429). A parallel-process latent growth curve modeling was adopted to evaluate the mediational process of LA between sociodemographic factors (age, gender, education, self-rated health, comorbidities, and depression) and the outcome process of functional disabilities.

We analyzed bacterial communities of six distinct gut sites (the food bolus and mucus layer of the proximal small intestine, cecum and distal large intestine), using wild folivorous flying squirrels. We found significant spatial heterogeneity in composition, diversity and species abundance distributions (SADs) of gut microbiota, corresponding to physicochemical conditions. High diversity was detected in the mucus layer of small intestine and the food bolus of cecum, followed by the food bolus of large intestine and the mucus layer of cecum and relatively low diversity in the food bolus of small intestine and the mucus layer of large intestine, likely due to disturbance and resource partitioning. The SADs showed succession-like patterns in the food bolus communities from the proximal to distal gut. Notably, each mucus layer community had a unique pattern different from the food bolus community of the same compartment, with distinct relative abundances of dominant species. In combination with data from other mammalian fecal samples, we concluded that gut microbiota were apparently dynamic in community structure, from low species richness with unequal abundances to high species richness with equal abundances; these findings were interpreted as strong habitat effects on bacterial communities.

An open reading frame (ORF) that encodes a 715-amino-acid polypeptide was found in an 8421-bp Eco RI fragment of the shrimp white spot syndrome virus (WSSV) genome. The polypeptide shows significant homology to eukaryotic serine/threonine protein kinase (PK) and contains the major conserved subdomains for eukaryotic protein kinases. Coupled in vitro transcription and translation generated a protein having an apparent molecular mass of about 87 kDa according to sodium dodecyl sulfate–polyacrylamide gel electrophoresis. For transcriptional analysis of the pk gene, total RNA was isolated from WSSV-infected shrimp at different times after infection. Northern blot analysis with pk -specific riboprobe found a major and a minor transcript of 2.7 and 5.7 kb, respectively. Rapid amplification of the 5′ cDNA ends of the major 2.7-kb pk transcript showed that there were two transcriptional initiation sites located at nucleotide residues -38(G) and -39(G) relative to the ATG translational start codon. Temporal expression analysis by RT-PCR indicated that the transcription of the pk gene started 2 h after infection and continued for at least 60 h. Phylogenetic analysis showed that WSSV protein kinase does not have any close relatives and does not fall into any of the major protein kinase groups.

t-haplotypes are a meiotic drive system found on the 17th chromosome of the house mouse (Mus musculus). They can be found in wild populations of all four genetically differentiated subspecies. The drive phenomenon is male-specific, such that heterozygous males (+/t) show non-Mendelian transmission and transmit the t-chromosome to > 90% of their offspring. So far the most comprehensive studies on the frequencies of t-haplotypes in natural populations have been on just one of the subspecies (M. musculus domesticus). We applied molecular methods to accurately screen t-haplotypes in a large number of populations of a second subspecies (M. musculus castaneus) distributed in Taiwan. We found that the overall t-haplotype frequency is low in M. m. castaneus (0.108), and the chromosomes are patchily distributed among its populations, closely resembling the situation found in M. m. domesticus. Further, we found the frequencies of t-haplotypes in our sample did not differ in relation to the sex or age of mice. This resemblance in the frequency and distribution among populations of the two distinct subspecies suggests that similar general mechanisms might be responsible for the low frequencies in both subspecies.

From previously constructed genomic libraries of a Taiwan WSSV isolate, a putative WSSV tk-tmk gene was identified. Uniquely, the open reading frame (ORF) of this gene was predicted to encode a novel chimeric protein of 388 amino acids with significant homology to two proteins: thymidine kinase (TK) and thymidylate kinase (TMK). Northern blot analysis with a WSSV tk-tmk-specific riboprobe detected a major transcript of 1.6 kb. When healthy adult Penaeus monodon shrimp were inoculated with WSSV, the tk-tmk gene transcript was first detected by RT-PCR analysis at 4 h postinfection and transcription levels continued to increase over the first 18 h. The gene's major in vitro transcription and translation product, equivalent to the predicted size (43 kDa), is a single chimeric protein that includes both the TK and TMK functional motifs. Evidence for phylogenetic analysis and sequence alignment suggested that the gene may have resulted from the fusion of a cellular-type TK gene and a cellular-type TMK gene. Its unique arrangement may also provide a valuable gene marker for WSSV.

Population dynamics of the spiny country rat Niviventer coxingi were studied by mark–recapture methods from January 1982 to May 1983 in a subtropical montane forest at Chitou, central Taiwan. The population density was < 8 rats/ha. The annual pattern of population fluctuation was characterized by summer increase, autumn peak and spring decline. Adult recruits occurred mostly in summer and autumn 1982, whereas young rats joined the population as three cohorts: winter 1981, summer 1982 and winter 1982. Growth patterns differed between winter and summer cohorts, which was attributed mainly to the slow body weight increase in summer cohort males. Males, either adult or juvenile, survived the winter better than females, resulting in the shorter lifespans and the lower number of females in the population. Life-history traits in female N. coxingi, i.e. short reproductive lifespan, few litters per life time and prolonged maternal care, along with the low breeding intensity, rendered the number of females the limiting factor for population increase. However, females may improve the survival and maturation of their offspring by prolonged maternal care.

To determine the genetic relationship between different colour morphs (orange and blackmorphs) of Clark’s anemonefish ( Amphiprion clarkii ) in Taiwan, we isolated eight poly-morphic microsatellite loci. A large number of alleles (range, 6–30), and high levels ofobserved heterozygosity (range, 0.1231–0.8358) were resolved in 71 individuals from twopopulations, indicating that these markers should be useful in assessing the relationshipbetween the two colour morphs of A. clarkii . Keywords : Amphiprion clarkii , anemonefish, microsatellites Received 21 January 2007; revision accepted 4 April 2007 The colouration pattern of tropical reef fish has often beenused for taxonomic purposes. Nonetheless, the roles ofsexual selection and colour patterns on genetic different-iation and speciation are not completely understoodamong reef fishes. Only a few studies have focused on thephylogenetic relationship between closely related specieswith different colour morphs (McMillan et al . 1999; Bernardi

We define the species boundaries of white-toothed shrews (genus Crocidura) in Taiwan using karyological and morphological characteristics. Ninety-nine animals were obtained from all over Taiwan at capture rates usually less than 10%. Three species are recognized by distinct cytotypes: Crociduru uttenuata tanakae 2n = 40, FN = 56; Crocidura suaveolens hoslefti 2n = 40, FN = 50; Crocidura kurodai 2n = 40, FN = 54. A suite of six morphological characters diagnose the three species: shape of skull, position of incisive foramina, shape of fourth upper premolar, shape of pinna, tail vibrissae, and foot pads. A species key and notes on the life history of each species are provided. Finally, we discuss chromosomal evolution and biogeography of Crociduru in East and South East Asia.

Chirixalus eiffingeri is an arboreal breeding rhacophorid frog with unique parental care behaviours. We developed 11 polymorphic microsatellites as genetic makers for parentage analysis to resolve the ecology of parental care. The numbers of alleles per locus ranged from two to 17. The observed and expected heterozygosities averaged 0.433 and 0.656, respectively. Total exclusionary probability of these loci is 0.984 when no parental genotypes are known, and is 0.999 when one of the parental genotypes is known. The results indicate that the 11 markers should provide sufficient resolution for inferring genetic parentage in C. eiffingeri .