Two lineages of stream toads in the genus Ansonia from Malaysian Borneo have long been suspected to be specifically distinct on the basis of molecular data. We assessed the taxonomic status of these lineages using morphological and additional genetic data. In mtDNA phylogeny, each lineage-one from Bario, Kelabit Highlands of Sarawak, the other from Mt. Mulu of Sarawak and the Crocker Range of Sabah-is separated from other congeners by large genetic distances, comparable with those observed between heterospecific species in the genus. These lineages are also morphologically distinguishable from other species, and are considered to represent valid, independently evolving species. We therefore describe them as A. kelabitensis sp. nov. and A. kanak sp. nov.
Three new megophryid species, Leptolalax melanoleucus, L. fuliginosus, and L. solus, are described from southwestern and southern Thailand on the bases of acoustic and morphological characteristics. Leptolalax melanoleucus and L. fuliginosus are similar to L. pelodytoides from northern Thailand, but differ from it completely in advertisement call characteristics and ventral color. Leptolalax solus is similar to L. heteropus from peninsular Malaysia, but differs from it by advertisement call, as well as by some body proportions. The distributional pattern of Leptolalax within Thailand is discussed.
We establish a new interstitial polyclad species, Theama japonica sp. nov., based on specimens collected from coarse-sandy habitats in three Japanese main islands (Hokkaido, Honshu, and Shikoku) along the coasts of the Pacific Ocean and the Sea of Japan. Theama japonica is characterized by i) two pairs of cerebral eyespots and four to six precerebral eyespots; ii) eosinophilic secretion glands distributed in the distal half of the inner ventral part of the prostatic vesicle; iii) a conical penis papilla, bent up dorsally, with a sclerotized inner wall; iv) the prostatic sheath with an inner angular fold on the dorso-distal side; and v) the external cilia longer dorsally than ventrally. Partial sequences of the cytochrome c oxidase subunit I (COI) gene from 20 specimens collected at eight localities along Japanese coasts represented 19 haplotypes. The uncorrected p-distances among these COI haplotypes fell within intraspecific variations observed in other polyclads. A network analysis based on these COI haplotypes suggested a geographically non-cohesive genetic structure of the species, possibly indicating the species' high dispersibility. Molecular phylogenetic analyses based on a concatenated dataset of 18S and 28S rDNA sequences showed T. japonica formed a clade with other Theama species. The resulting tree also indicates that our new species is more closely related to Theama sp. from Colombia than species from Panama and Croatia.
Although the crab-eating frog Fejervarya cancrivora is one of the most widely distributed species in Asian region, taxonomic relationships among different populations remain unclarified. In this study, we attempted to elucidate the taxonomic status of F. cancrivora from Indonesian and other Asian populations. Five populations of F. cancrivora from Selangor (Malaysia), Cianjur (Java, Indonesia), Trat (Thailand), Khulna (Bangladesh), and Makassar (Sulawesi, Indonesia) were morphologically observed and subjected to crossing experiments. Principal component and clustering analyses revealed that these five populations could be organized into three groups corresponding to three observed morphological types: a Selangor and Cianjur group (large-type), a Trat and Khulna group (mangrove-type), and a Makassar group (Sulawesi-type). The limited crossing experiments revealed that hybrids between Selangor females and Cianjur and Trat males developed normally, whereas hybrids between Selangor females and Khulna males showed incomplete gametic isolation. Histological observations of the testes of mature males revealed the presence of pycnotic nuclei in the hybrids between Selangor females and Khulna males in addition to normal bundles of spermatozoa. In contrast, no pycnotic nuclei were observed in the Selangor controls. Although meiotic metaphases in the controls were normal, those in hybrids showed several abnormalities, such as the appearance of univalents and an increase in rod-shaped bivalents. Based on our findings from the morphological observations and crossing experiments, we conclude that each of three identified types represents a distinct species. We propose that the large-type is F. cancrivora, the mangrove-type is F. moodiei, and the Sulawesi-type represents an undescribed species.
Examination of types and recently collected specimens revealed that Ansonia anotis Inger, Tan, and Yambun, 2001 and Pedostibes maculatus (Mocquard, 1890), both described from Kinabalu, Sabah, Malaysia, are hardly differentiated morphologically. Analyses of a total of 2,427 bp of the 12S rRNA, tRNA(val), and 16S mitochondrial rRNA genes revealed that the two species are very close genetically. Thus A. anotis is regarded as conspecific and is synonymized with P. maculatus. Genetically, this species proved to form a lineage distinct from other bufonids from Southeast Asia, including species of Ansonia and Pedostibes. Because the species has also some unique morphological traits different from known bufonid genera, we propose to establish a new genus for Nectophryne maculata Mocquard, 1890.
The red-bellied form of Calliophis intestinalis (Laurenti, 1768) sensu lato was originally reported from Pahang, west Malaysia. To determine the taxonomic status of this form, we examined the type specimens of Elaps sumatranus Lidth De Jeude, 1890, Calliophis intestinalis everetti (Boulenger, 1896), and Callophis furcatus var. nigrotaeniatus Peters, 1863. The results indicated that the red-bellied form of C. intestinalis should be named as Calliophis nigrotaeniatus comb. nov., whose valid species status was based on morphological and molecular analyses. We designate a lectotype and redescribe the species, which is genetically close to Calliophis bilineatus (Peters, 1881) from the Philippines, and is clearly distinguishable from other congeners by possessing a pair of gray or dark blue lateral stripes and by being bright red on the ventrum. Elaps sumatranus and C. i. everetti are relegated to subjective junior synonyms of C. nigrotaeniatus.
This study was conducted to clarify the development of free neuromasts with growth of the barramundi, Lates calcarifer. A pair of free neuromasts was observed behind the unpigmented eyes in newly hatched eleutheroembryos with a mean total length of 1.93 mm, and two-hour-old eleuthero-embryos could respond to an approaching pipette. At 2 days after hatching, the egg yolk sac was mostly consumed, the eyes were pigmented, and the larvae commenced feeding on rotifers. Free neuromasts increased in number with growth and commenced developing into canal neuromasts in barramundi 15 days old with a mean total length of 8.07 mm. The average length of the major axis of the trunk free neuromasts attained approximately 12.9-15.5 microm, and the number of sensory cells was 15.4-17.5 at 15-20 days old. Developed cupulae of free neuromasts were observed in 1-day-old eleutheroembryos. The direction of maximum sensitivity of free neuromasts, determined from the polarity of the sensory cells, coincided with the minor axis of the lozenge-shaped outline of the apical surface of the free neuromasts. The polarity of trunk neuromasts was usually oriented along the antero-posterior axis of the fish body, but a few had a dorso-ventral direction. On the head, free neuromasts were oriented on lines tangential to concentric circles around the eye.
Nest predation is the main cause of hatching failure for many turtle populations. For green turtles (Chelonia mydas) nesting at Chagar Hutang in Redang Island, Malaysia, Asian water monitors (Varanus salvator) are a potential nest predator. However, no studies have documented the space use of this species in coastal habitat adjacent to a sea turtle nesting beach to assess its potential impact on turtle nests. Here, we used Global Positioning System (GPS) data loggers to quantify space use of Asian water monitors in order to establish the extent to which they use sea turtle nesting areas. Asian water monitors had a diurnal activity pattern and spent most of their time in rain forest habitat behind the sea turtle nesting beach. The home range occupied by Asian water monitors varied between 0.015 and 0.198 km2 calculated by the Kernel Brownian Bridge method. The space use patterns of individual Asian water monitors varied between individuals. Two males had relatively small home ranges, whereas one male and the female had a relatively large home range. Because tracked Asian water monitors in this study rarely visited the sea turtle nesting areas, it is probable that only a few individuals are responsible for opening nests.
Two forms of Staurois that are differentiated by body size occur parapatrically in the Crocker Range, Sabah, Borneo. Analyses of a total of 1,499 bp of the mitochondrial cytochrome b, 12S rRNA, and 16S rRNA genes revealed that the two forms could be completely split genetically. The two forms could be also clearly differentiated morphologically, not only by snout-vent length but also by the relative sizes of snout, eye, and finger disk. Comparisons of the two forms with all known species of the genus revealed the large and small forms to be S. tuberilinguis and S. parvus, respectively. The latter species has long been synonymized with the former, but we here consider them to represent different species.
In general, African catfish shows higher survival rates in the dark conditions than in the light conditions. In this study, larval behavior of African catfish was observed under 0, 0.01, 0.1, 1, 10, and 100 lx using a CCD camera to investigate the reason why African catfish larvae show higher survival rates in dark conditions. The larvae showed significantly higher swimming activity under 0, 0.01, and 0.1 lx than that under 10 and 100 lx. The larvae also showed significantly increased aggressive behavior under 10 and 100 lx; the swimming larvae attacked resting individuals more frequently under 10 and 100 lx than under 0, 0.01, and 0.1 lx. The aggressive behavior and sharp teeth of the attacking larvae appeared to induce skin surface lesions on injured larvae. Chemical substances were then generated from the injured skin surface, and these chemical stimuli triggered cannibalistic behavior in other fish near the injured fish. The results of this study demonstrate that the higher survival rates of African catfish larvae under dark conditions are a result of inactivity and subsequent increase in chemical releasing stimuli concentrations around inactive individuals that triggers feeding behavior in nearby active catfish. Therefore, we recommend larval rearing of African catfish in dark or dim conditions, as it improves catfish survival rates.
This study examined 396 base pairs of the mitochondrial cytochrome b gene from 110 individuals belonging to the genus Hampala, a group of freshwater cyprinids that inhabit Southeast Asia. The samples were taken from various locations throughout Sarawak, Sabah, and peninsular Malaysia. The nucleotide sequences were subjected to phylogenetic analyses by using the neighbor-joining, maximum parsimony, and maximum likelihood methods. All three methods revealed the reciprocally monophyletic relationship of Hampala macrolepidota to the other Hampala forms, thus strongly supporting its status as a distinct species. Phylogenetic analysis also discovered the existence of two H. bimaculata lineages endemic to Borneo: (1) a newly identified species from the southern and central part of Sarawak assigned as H. bimaculata Type A and (2) the previously described H. bimaculata from northern Sarawak and the west coast of Sabah assigned as H. bimaculata Type B. However, the status of H. sabana and an intermediate form were not elucidated. The results suggest that the intermediate form from the Tawau population is actually a subpopulation of H. sabana, while the highly divergent intermediate form from Kalabakan could represent a cryptic species. The sharing of H. macrolepidota haplotypes in the southern peninsular Malaysia and southern and central Sarawak samples (Hm1 and Hm2) reflected the recent disconnection of the two regions, during the late Pleistocene. Overall, the partial sequence of the mitochondrial cytochrome b gene was useful for resolving the phylogenetic relationships among Hampala fishes in Malaysia.
In order to elucidate the taxonomic status of the Fejervarya limnocharis complex relative to Malaysia and Japan populations, morphological observations and molecular phylogenetic analysis were carried out using three populations from Indonesia (type locality), Malaysia, and Japan. In addition, we conducted histological and spermatogenic observations using hybrids among these populations. Principal component and cluster analyses demonstrated that these populations could be clearly separated from one another. Abnormal testes were found in the hybrids between the Japan and Indonesia populations and between the Japan and Malaysia populations, but testes of the controls and hybrids between the Malaysia and Indonesia populations were quite normal. The mean number of univalents per cell was 5.42, 4.58, and 0.20 in hybrids between the Indonesia and Japan populations, Malaysia and Japan populations, and Indonesia and Malaysia populations, respectively. Sequence divergences in 16S rRNA and Cyt b genes were 0-0.4% (xbar=0.2%) and 0.3-1.5% (xbar=1.0%), respectively, between the Malaysia and Indonesia populations, and 2.4-2.6% (xbar=2.5%) and 11.0-12.0% (xbar=11.5%) between the Japan population and F. limnocharis complex, including the Malaysia and Indonesia populations and F. multistriata from China. This study indicated that the Malaysia population and F. multistriata from China should be designated as a subspecies of topotypic F. limnocharis, and that the Japan population should be regarded as a distinct species.
Rats (Rattus species) are the most notorious vertebrate pests in Malaysian oil palm plantations. Although many studies have been conducted on Asian rats, little attention has been paid to their species composition and phylogenetic relationships in oil palm plantations in Peninsular Malaysia. We determined the mitochondrial cytochrome oxidase subunit I (COI) gene sequence (708 bp) for 216 individual rats collected from five oil palm plantations in Peninsular Malaysia. Phylogenetic analysis in conjunction with comparison with sequences from the nucleotide sequence database revealed five distinct lineages in the Malaysian oil plantations: Rattus tiomanicus, Rattus argentiventer, Rattus exulans, Rattus tanezumi, and a taxon corresponding to the Malayan house rat, which was most frequently observed (∼50%). The last taxon has traditionally been classified as a synonym of Rattus rattus (Rattus rattus diardii) or Rattus tanezumi, but our phylogenetic analysis placed it as an independent lineage, which is not particularly closely related to R. rattus or R. tanezumi, and which we refer to as Rattus diardii. The construction of the network showed that there is considerable genetic variation within the lineages of R. diardii and R tiomanicus, suggesting that these two species are native to the Malay Peninsula.
Bats play crucial ecological and economic roles. However, this group of mammals is largely threatened due to anthropogenic activities inside or around their caves. In the present study, we investigate the effects of cave microclimate and physiological parameters on bat roost preference in Fairy Cave Nature Reserve (Fairy Cave NR). The microclimate and physiological parameters including temperature (°C), relative humidity (RH), light intensity (lux), air flow, passage dimension (m), roost height (m) and distance of roost from nearest entrance (m) were measured. Results showed that Emballonura monticola, Hipposideros diadema, Hipposideros larvatus and Penthetor lucasi roost in Fairy Cave NR. These bats can be distinguished by their colony size, roost posture, spatial arrangement and position of their roost. Penthetor lucasi makes up the largest colony in the bright zone, whereas E. monticola has the smallest colony, occupying the twilight zone throughout the cave. Members from the family Hipposideridae roost in the dark portion of the cave with zero light intensity. Emballonura monticola utilizes the hottest roost in the cave compared to the other microbats, whereas H. diadema inhabits the coolest roost. Physiological parameters such as light intensity, passage dimension, and distance from nearest entrance are significant parameters in roosting preferences. These parameters should be monitored to ensure the sustainability of not only the aforementioned species, but also other species that roost in nearby caves of Bau limestone (Wind Cave Nature Reserve: with 13 recorded species).
Mesohabitat selection in fluvial fishes was studied in a small tropical stream of the Malay Peninsula. A total of 681 individuals representing 24 species were sampled at 45 stations within heterogeneous stream (ca. 1 km in length), in which water depth, water velocity, substrate size, and riparian canopy cover were measured as environmental variables. A canonical correspondence analysis (CCA) yielded a diagram that shows a specific mesohabitat selection of the fish assemblage, in which the species were plotted widely on the CCA1-CCA2 biplot. Generalized linear model also revealed a significant pattern of the mesohabitat selection of several species. Water velocity and substrate size mainly separated on CCA1, indicating variation of pool (deep, slow-flow section) and riffle (shallow, fast-flow section) structures is a primary factor of mesohabitat selection in the fluvial fish assemblage. The mean body weight of species significantly correlated with CCA1; larger species tended to inhabit pools, while small ones occupied riffles. The riparian canopy cover separated on CCA2. The trophic level of species significantly correlated with CCA2; herbivorous species (low trophic level) selected open sites without riparian cover, whereas omnivorous/carnivorous (middle-high trophic level) species preferred highly covered sites. In conclusion, our results suggest that mesohabitat selection is closely related to the species feeding habit, which is consistent with the results of previous studies.
To elucidate genetic divergence and evolutionary relationship in Fejervarya cancrivora from Indonesia and other Asian countries, allozyme and molecular analyses were carried out using 131 frogs collected from 24 populations in Indonesia, Thailand, Bangladesh, Malaysia, and the Philippines. In the allozymic survey, seventeen enzymatic loci were examined for 92 frogs from eight representative localities. The results showed that F. cancrivora is subdivided into two main groups, the mangrove type and the large- plus Pelabuhan ratu types. The average Nel's genetic distance between the two groups was 0.535. Molecular phylogenetic trees based on nucleotide sequences of the 16S rRNA and Cyt b genes and constructed with the ML, MP, NJ, and BI methods also showed that the individuals of F. cancrivora analyzed comprised two clades, the mangrove type and the large plus Pelabuhan ratu / Sulawesi types, the latter further split into two subclades, the large type and the Pelabuhan ratu / Sulawesi type. The geographical distribution of individuals of the three F. cancrivora types was examined. Ten Individuals from Bangladesh, Thailand, and the Philippines represented the mangrove type; 34 Individuals from Malaysia and Indonesia represented the large type; and 11 individuals from Indonesia represented the Pelabuhan ratu / Sulawesi type. Average sequence divergences among the three types were 5.78-10.22% for the 16S and 12.88-16.38% for Cyt b. Our results suggest that each of the three types can be regarded as a distinct species.
The present study was conducted to elucidate the genetic divergence and the phylogenetic relationships in the F. limnocharis complex from Bangladesh and other Asian countries such as Sri Lanka, Thailand, Malaysia, Taiwan and Japan by allozyme analyses. We used a total of 95 frogs of the F. limnocharis complex from these countries and F. cancrivora from the Philippines as an outgroup. Based on body size, the F. limnocharis complex from Bangladesh was divided into three distinct groups: large, medium and small types. Allozyme analyses were carried out with 28 loci encoding 20 enzymes and two blood proteins by horizontal starch-gel electrophoresis. When genetic distance was calculated, distinct divergence was found among the three types: mean genetic distance was 0.782 between the small and medium types, 1.458 between the large and medium types, and 1.520 between the large and small types. Phylogenetic trees based on genetic distance showed that all populations of Bangladesh small type strongly formed a cluster and were found to be most closely related to the Sri Lanka population; that all populations of Bangladesh large type formed a very strong cluster and were grouped with several populations from Thailand, Malaysia, Japan, and Taiwan; and that the medium type was segregated from all other groups. This may imply that each of the three types is a different species, and that the medium type is possibly an undescribed taxon.
We record a tree frog of the genus Chiromantis for the first time from outside the Southeast Asian continent and describe it as a new species, Chiromantis inexpectatus. The new species from the Malaysian state of Sabah, Borneo, is a small-sized Chiromantis (male snout-vent length ca. 22 mm), and is distinguished from all other members of the genus by the combination of the following morphological characteristics: dark stripes absent, but dark spots present on dorsum; a dark-brown lateral band present from snout tip to half of body, bordered ventrally by white stripe; third and fourth fingers less than half webbed; third finger disk wider than tympanum diameter; and inner metatarsal tubercle present. Significance of findings of this species from Borneo Island, as well as phylogeny and breeding habit of the genus Chiromantis, are briefly discussed.
The two-pore domain potassium ion (K + ) channel-related K + (TREK) channel and melatonin receptors play roles in the regulation of reproduction in zebrafish. Since reproduction is regulated by diurnal rhythms, the TREK family and melatonin receptors may exhibit diurnal rhythms in expression. In this study, we aimed to investigate diurnal variations of the gene expressions of TREK family and melatonin receptors and their associations with kisspeptin and gonadotrophin-releasing hormone (GnRH). Diurnal variations of trek1b, trek2a, trek2b, mt1, mt2, mel1a, kiss2 and gnrh3 expressions were examined by real-time PCR. For reproduction-related genes, kiss2 and gnrh3 exhibited diurnal rhythms. trek2a revealed a diurnal rhythm in the TREK family. mt2 and mel1c exhibited diurnal rhythms in the melatonin receptors. Since Trek2a regulates gnrh3 expression, the diurnal rhythm of gnrh3 expression suggests to be regulated by the diurnal rhythm of trek2a expression.
We examined allozyme variation in two camaenid tree snails, Amphidromus atricallosus and A. inversus, across two principal regions of Thailand and from Singapore, plus for A. inversus, one site in peninsular Malaysia. Using horizontal starch gel electrophoresis, 13 allozyme loci (11 polymorphic) were screened for A. atricallosus and 18 (5 polymorphic) for A. inversus. Heterozygosity was higher in A. atricallosus (Hexp=0.018-0.201, mean=0.085) than in A. inversus (Hexp=0-0.023, mean= 0.002). Genetic heterogeneity among samples was higher in A. inversus (Fst=0.965) than in A. atricallosus (Fst=0.781). Within A. atricallosus, populations were more differentiated in southern Thailand (Fst=0.551) than in eastern Thailand (Fst=0.144). The high Fst and low Hexp in populations of A. inversus suggest that this species is likely to have experienced a series of strong bottlenecks, perhaps occurring chiefly on offshore continental-shelf islands. The low Fst values of A. atricallosus in eastern Thailand suggest frequent gene flows among populations in this region. The southern and eastern samples of A. atricallosus exhibited fixed allele differences at four loci and great genetic distance (Nei's D=0.485-0.946), suggesting that these two samples may actually represent, or else be evolving into, separate species.