Little is known about the classification and phylogenetic relationships of the leaf monkeys (Presbytis). We analyzed mitochondrial DNA sequences of cytochrome b (Cyt b) and 12S rRNA to determine the phylogenetic relationships of the genus Presbytis. Gene fragments of 388 and 371 bp of Cyt b and 12S rRNA, respectively, were sequenced from samples of Presbytis melalophos (subspecies femoralis, siamensis, robinsoni, and chrysomelas), P. rubicunda and P. hosei. The genus Trachypithecus (Cercopithecidae) was used as an outgroup. The Cyt b NJ and MP phylogeny trees showed P. m. chrysomelas to be the most primitive, followed by P. hosei, whereas 12S rRNA tree topology only indicated that these two species have close relationships with the other members of the genus. In our analysis, chrysomelas, previously classified as a subspecies of P. melalophos, was not included in either the P. m. femoralis clade or the P. m. siamensis clade. Whether or not there should be a separation at the species level remains to be clarified. The tree topologies also showed that P. m. siamensis is paraphyletic with P. m. robinsoni, and P. m. femoralis with P. rubicunda, in two different clades. Cyt b and 12S rRNA are good gene candidates for the study of phylogenetic relationships at the species level. However, the systematic relationships of some subspecies in this genus remain unclear.
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 aim of this study was to evaluate the genotoxic effects of a crude extract of khat (Catha edulis, Forsk) leaves in rats. Two groups were fed khat crude extract, 1000 and 2000mg/kg body weight, for 90 days and were compared with a control group. The alkaline (pH>13) version of comet assay was used in this study. However, no previous published work has been undertaken and showed the effect of khat on DNA migration in the comet assay. To compare the comet assay results with another genetic endpoint, blood samples were analyzed for chromosomal aberrations. These results showed no DNA damage detected using comet assay in both the khat treated groups, while the results of chromosomal aberrations assay showed a significant increase (P<0.05) in the 2000mg/kg body weight treated group compared to the control group.
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.
We classified diversity in eight new complete mitochondrial genome sequences and 41 partial sequences from living Aboriginal Australians into five haplogroups. Haplogroup AuB belongs to global lineage M, and AuA, AuC, AuD, and AuE to N. Within N, we recognize subdivisions, assigning AuA to haplogroup S, AuD to haplogroup O, AuC to P4, and AuE to P8. On available evidence, (S)AuA and (M)AuB are widespread in Australia. (P4)AuC is found in the Riverine region of western New South Wales, and was identified by others in northern Australia. (O)AuD and (P8)AuE were clearly identified only from central Australia. Our eight Australian full mt genome sequences, combined with 20 others (Ingman and Gyllensten 2003 Genome Res. 13:1600-1606) and compared with full mt genome sequences from regions to the north that include Papua New Guinea, Malaya, and Andaman and Nicobar Islands, show that ancestral connections between regions are deep and limited to clustering at the level of the N and M macrohaplogroups. The Australian-specific distribution of the five haplogroups identified indicates genetic isolation over a long period. Ancestral connections within Australia are deeper than those reflected by known linguistic or culturally based affinities. Applying a coalescence analysis to a gene tree for the coding regions of the eight genomic sequences, we made estimates of time depth that support a continuity of presence for the descendants of a founding population already established by 40,000 years ago.
DNA barcodes, typically focusing on the cytochrome oxidase I gene (COI) in many animals, have been used widely as a species-identification tool. The ability of DNA barcoding to distinguish species from a range of taxa and to reveal cryptic species has been well documented. Despite the wealth of DNA barcode data for fish from many temperate regions, there are relatively few available from the Southeast Asian region. Here, we target the marine fish Family Carangidae, one of the most commercially-important families from the Indo-Malay Archipelago (IMA), to produce an initial reference DNA barcode library.
Christmas Island is a remote Australian territory located close to the main Indonesian island of Java. Y-chromosome and mitochondrial DNA (mtDNA) markers were used to investigate the genetic structure of the population, which comprises communities of mixed ethnic origin. Analysis of 12 Y-chromosome biallelic polymorphisms revealed a high level of gene diversity and haplotype frequencies that were consistent with source populations in southern China and Southeast Asia. mtDNA hypervariable segment I (HVS-I) sequences displayed high levels of haplotype diversity and nucleotide diversity that were comparable to various Asian populations. Genetic distances revealed extremely low mtDNA differentiation among Christmas Islanders and Asian populations. This was supported by the relatively high proportion of sequence types shared among these populations. The most common mtDNA haplogroups were M* and B, followed by D and F, which are prevalent in East/Southeast Asia. Christmas Islanders of European descent were characterized by the Eurasian haplogroup R*, and a limited degree of admixture was observed. In general, analysis of the genetic data indicated population affinities to southern Chinese (in particular from the Yunnan Province) and Southeast Asia (Thailand, Malaysia, and Cambodia), which was consistent with historical records of settlement. The combined use of these different marker systems provides a useful and appropriate model for the study of contemporary populations derived from different ethnic origins.
Metabarcoding, the coupling of DNA-based species identification and high-throughput sequencing, offers enormous promise for arthropod biodiversity studies but factors such as cost, speed and ease-of-use of bioinformatic pipelines, crucial for making the leapt from demonstration studies to a real-world application, have not yet been adequately addressed. Here, four published and one newly designed primer sets were tested across a diverse set of 80 arthropod species, representing 11 orders, to establish optimal protocols for Illumina-based metabarcoding of tropical Malaise trap samples. Two primer sets which showed the highest amplification success with individual specimen polymerase chain reaction (PCR, 98%) were used for bulk PCR and Illumina MiSeq sequencing. The sequencing outputs were subjected to both manual and simple metagenomics quality control and filtering pipelines. We obtained acceptable detection rates after bulk PCR and high-throughput sequencing (80-90% of input species) but analyses were complicated by putative heteroplasmic sequences and contamination. The manual pipeline produced similar or better outputs to the simple metagenomics pipeline (1.4 compared with 0.5 expected:unexpected Operational Taxonomic Units). Our study suggests that metabarcoding is slowly becoming as cheap, fast and easy as conventional DNA barcoding, and that Malaise trap metabarcoding may soon fulfill its potential, providing a thermometer for biodiversity.
Molecular detection has overcome limitations of microscopic examination by providing greater sensitivity and specificity in Plasmodium species detection. The objective of the present study was to develop a quantitative real-time polymerase chain reaction coupled with high-resolution melting (qRT-PCR-HRM) assay for rapid, accurate and simultaneous detection of all five human Plasmodium spp. A pair of primers targeted the 18S SSU rRNA gene of the Plasmodium spp. was designed for qRT-PCR-HRM assay development. Analytical sensitivity and specificity of the assay were evaluated. Samples collected from 229 malaria suspected patients recruited from Sabah, Malaysia were screened using the assay and results were compared with data obtained using PlasmoNex(TM), a hexaplex PCR system. The qRT-PCR-HRM assay was able to detect and discriminate the five Plasmodium spp. with lowest detection limits of 1-100 copy numbers without nonspecific amplifications. The detection of Plasmodium spp. in clinical samples using this assay also achieved 100% concordance with that obtained using PlasmoNex(TM). This indicated that the diagnostic sensitivity and specificity of this assay in Plasmodium spp. detection is comparable with those of PlasmoNex(TM). The qRT-PCR-HRM assay is simple, produces results in two hours and enables high-throughput screening. Thus, it is an alternative method for rapid and accurate malaria diagnosis.
Matched MeSH terms: DNA Primers; DNA Copy Number Variations
Studying the effect of a magnetic field on oligonucleotide DNA can provide a novel DNA manipulation technique for potential application in bioengineering and medicine. In this work, the optical and electrochemical response of a 100 bases oligonucleotides DNA, cytosine-guanine (CG100), is investigated via exposure to different magnetic fields (250, 500, 750, and 1000 mT). As a result of the optical response of CG100 to the magnetic field, the ultra-violet-visible spectrum indicated a slight variation in the band gap of CG100 of about 0.3 eV. Raman spectroscopy showed a significant deviation in hydrogen and phosphate bonds' vibration after exposure to the magnetic field. Oligonucleotide DNA mobility was investigated in the external electric field using the gel electrophoresis technique, which revealed a small decrease in the migration of CG100 after exposure to the magnetic field.
In their initial stages of discovery, prokaryotic toxin-antitoxin (TA) systems were confined to bacterial plasmids where they function to mediate the maintenance and stability of usually low- to medium-copy number plasmids through the post-segregational killing of any plasmid-free daughter cells that developed. Their eventual discovery as nearly ubiquitous and repetitive elements in bacterial chromosomes led to a wealth of knowledge and scientific debate as to their diversity and functionality in the prokaryotic lifestyle. Currently categorized into six different types designated types I-VI, type II TA systems are the best characterized. These generally comprised of two genes encoding a proteic toxin and its corresponding proteic antitoxin, respectively. Under normal growth conditions, the stable toxin is prevented from exerting its lethal effect through tight binding with the less stable antitoxin partner, forming a non-lethal TA protein complex. Besides binding with its cognate toxin, the antitoxin also plays a role in regulating the expression of the type II TA operon by binding to the operator site, thereby repressing transcription from the TA promoter. In most cases, full repression is observed in the presence of the TA complex as binding of the toxin enhances the DNA binding capability of the antitoxin. TA systems have been implicated in a gamut of prokaryotic cellular functions such as being mediators of programmed cell death as well as persistence or dormancy, biofilm formation, as defensive weapons against bacteriophage infections and as virulence factors in pathogenic bacteria. It is thus apparent that these antitoxins, as DNA-binding proteins, play an essential role in modulating the prokaryotic lifestyle whilst at the same time preventing the lethal action of the toxins under normal growth conditions, i.e., keeping the proverbial wolves at bay. In this review, we will cover the diversity and characteristics of various type II TA antitoxins. We shall also look into some interesting deviations from the canonical type II TA systems such as tripartite TA systems where the regulatory role is played by a third party protein and not the antitoxin, and a unique TA system encoding a single protein with both toxin as well as antitoxin domains.
Matched MeSH terms: DNA-Binding Proteins; DNA Copy Number Variations
DNA methylation, a major regulator of epigenetic modifications has been shown to alter the expression of genes that are involved in aspects of glucose metabolism such as glucose intolerance, insulin resistance, β-cell dysfunction and other conditions, and it ultimately leads to the pathogenesis of type 2 diabetes mellitus (T2DM). Current evidences indicate an association of DNA methylation with T2DM. This review provides an overview of how various factors play crucial roles in T2DM pathogenesis and how DNA methylation interacts with these factors. Additionally, an update on current techniques of DNA methylation analysis with their pros and cons is provided as a basis for the adoption of suitable techniques in future DNA methylation research towards better management of T2DM. To elucidate the mechanistic relationship between vital environmental factors and the development of T2DM, a better understanding of the changes in gene expression associated with DNA methylation at the molecular level is still needed.
To date, three species of the genus Glischropus are recognized from the Indomalayan zoogeographic region-G. bucephalus from the Indochinese subregion, G. tylopus from the Sundaic subregion (Peninsular Thailand and Malaysia, Borneo, Sumatra, Moluccas) and G. javanus, restricted to Java. The investigation of the holotype and three topotype specimens of G. batjanus supported the view that the name was previously correctly regarded as the junior subjective synonym of G. tylopus. During review of material recently collected in southwestern Sumatra, Indonesia, one specimen of a yet undescribed species of Thick-thumbed bat was identified. G. aquilus n. sp. markedly differs from its congeners by its dark brown pelage, nearly black ear and tragus, and in skull proportions. The phylogenetic analysis based on cytb sequences also supports the specific distinctness of G. aquilus n. sp. Its discovery brings the count to 88 species of bats known from Sumatra.
Chinese knotweed (Persicaria chinensis) is of ecological and economic importance as a high-risk invasive species and a traditional medicinal herb. However, the insects associated with P. chinensis pollination have received scant attention. As a widespread invasive plant we would expect P. chinensis to be associated with a diverse group of insect pollinators, but lack of taxonomic identification capacity is an impediment to confirm this expectation. In the present study we aimed to elucidate the insect pollinators of P. chinensis in peninsular Malaysia using DNA barcoding. Forty flower visitors, representing the range of morphological diversity observed, were captured at flowers at Ulu Kali, Pahang, Malaysia. Using Automated Barcode Gap Discovery, 17 morphospecies were assigned to 23 species representing at least ten families and four orders. Using the DNA barcode library (BOLD) 30% of the species could be assigned a species name, and 70% could be assigned a genus name. The insects visiting P. chinensis were broadly similar to those previously reported as visiting Persicaria japonica, including honey bees (Apis), droneflies (Eristalis), blowflies (Lucilia) and potter wasps (Eumedes), but also included thrips and ants.
Phylogeographic patterns and population structure of the pelagic Indian mackerel, Rastrelliger kanagurta were examined in 23 populations collected from the Indonesian-Malaysian Archipelago (IMA) and the West Indian Ocean (WIO). Despite the vast expanse of the IMA and neighbouring seas, no evidence for geographical structure was evident. An indication that R. kanagurta populations across this region are essentially panmictic. This study also revealed that historical isolation was insufficient for R. kanagurta to attain migration drift equilibrium. Two distinct subpopulations were detected between the WIO and the IMA (and adjacent populations); interpopulation genetic variation was high. A plausible explanation for the genetic differentiation observed between the IMA and WIO regions suggest historical isolation as a result of fluctuations in sea levels during the late Pleistocene. This occurrence resulted in the evolution of a phylogeographic break for this species to the north of the Andaman Sea.
Fruit type in the genus Lithocarpus (Fagaceae) includes both classic oak acorns and novel modifications. Bornean taxa with modified fruits can be separated into two sections (Synaedrys and Lithocarpus) based on subtle shape differences. By following strict criteria for homology and representation, this variation in shape can be captured and the sections distinguished by using elliptic Fourier or eigenshape analysis. Phenograms of fruit shape, constructed by using restricted maximum likelihood techniques and these morphometric descriptors, were incorporated into combined and comparative analyses with molecular sequence data from the internal transcribed spacer (ITS) region of the nuclear rDNA, using branch-weighted matrix representation. The combined analysis strongly suggested independent derivation of the novel fruit type in the two sections from different acornlike ancestors, while the comparative analysis indicated frequent decoupling between the molecular and morphological changes as inferred at well-supported nodes. The acorn fruit type has undergone little modification between ingroup and outgroup, despite large molecular distance. Greater morphological than molecular change was inferred at critical transitions between acorn and novel fruit types, particularly for section Lithocarpus. The combination of these two different types of data improved our understanding of the macroevolution of fruit type in this difficult group, and the comparative analysis highlighted the significant incongruities in evolutionary pattern between the two datasets.
Methicillin-resistant Staphylococcus aureus (MRSA) has been prevalent in our hospital over the last three years. Differentiation among MRSA strains by DNA typing in addition to antibiotic resistance pattern surveillance is crucial in order to implement infection control measures. The aim of this study was to characterize MRSA isolates from patients admitted to Hospital Universiti Kebangsaan Malaysia (HUKM) by phenotypic (analyses of antibiotic susceptibility pattern) and genotypic (PFGE) techniques to determine the genetic relatedness of the MRSA involved and to identify endemic clonal profiles of MRSA circulating in HUKM. Seventy one MRSA strains collected between January to March 2000 from patients from various wards in HUKM were tested for antimicrobial resistance and typed by pulsed-field gel electrophoresis (PFGE). Four major types of PFGE patterns were identified (A, B, C and D) among MRSA strains. Two predominant PFGE types were recognised, Type A (59.2%) and Type B (33.8%). Most of these strains were isolated from ICU, Surgical wards and Medical wards. MRSA strains with different PFGE patterns appeared to be widespread among wards. Strains with the same antibiotype could be of different PFGE types. Most of isolates were resistant to ciprofloxacin, erythromycin, gentamicin and penicillin. One isolate with a unique PFGE pattern Type D and susceptible to gentamicin was identified as a different clone. Some isolates obtained from the same patient showed different PFGE subtypes suggesting that these patients were infected/colonized with multiple MRSA strains. PFGE analysis suggests that MRSA strains with different PFGE types was propagated within our hospital. The relationship between antibiotic susceptibility and PFGE patterns was independent. The ability of PFGE technique in differentiating our MRSA strains make it a method of choice for investigating the source, transmission and spread of nosocomial MRSA infection, and thus an appropriate control programme can be implemented to prevent the spread of MRSA infection.
The advent of recombinant DNA technology has already made a significant impact on various aspects related to the basic understanding of pathogenic mechanisms in infectious diseases, as well as practical applications related to diagnostics and prevention. The present paper discusses recent technological innovations and increased analytical capabilities which promise to have an even more significant impact on the control of viral and bacterial diseases.