We report here the first novel HIV-1 circulating recombinant form (CRF) 54_01B (CRF54_01B) isolated from three epidemiologically unlinked subjects of different risk groups in Malaysia. These recently sampled recombinants showed a complex genome organization composed of parental subtype B' and CRF01_AE, with identical recombination breakpoints observed in the gag, pol, and vif genes. Such a discovery highlights the ongoing active generation and spread of intersubtype recombinants involving the subtype B' and CRF01_AE lineages and indicates the potential of the new CRF in bridging HIV-1 transmission among different risk groups in Southeast Asia.
Three novel actinobacteria, strains 39(T), 40 and 41, were isolated from soil collected from Barrientos Island in the Antarctic. The taxonomic status of these strains was determined using a polyphasic approach. Comparison of 16S rRNA gene sequences revealed that strain 39(T) represented a novel lineage within the family Dermacoccaceae and was most closely related to members of the genera Demetria (96.9 % 16S rRNA gene sequence similarity), Branchiibius (95.7 %), Dermacoccus (94.4-95.3 %), Calidifontibacter (94.6 %), Luteipulveratus (94.3 %), Yimella (94.2 %) and Kytococcus (93.1 %). Cells were irregular cocci and short rods. The peptidoglycan type was A4α with an L-Lys-L-Ser-D-Asp interpeptide bridge. The cell-wall sugars were galactose and glucose. The major menaquinone was MK-8(H(4)). The polar lipids were diphosphatidylglycerol, phosphatidylglycerol, phosphatidylinositol, phosphoglycolipid, two glycolipids and one unknown phospholipid. The acyl type of the cell-wall polysaccharide was N-acetyl. The major cellular fatty acids were anteiso-C(17 : 0) (41.97 %), anteiso-C(17 : 1)ω9c (32.16 %) and iso-C(16 : 0) (7.68 %). The DNA G+C content of strain 39(T) was 68.4 mol%. On the basis of phylogenetic and phenotypic differences from other genera of the family Dermacoccaceae, a novel genus and species, Barrientosiimonas humi gen. nov., sp. nov., is proposed; the type strain of the type species is 39(T) (=CGMCC 4.6864(T) = DSM 24617(T)).
Malaria is still a public health problem in Malaysia with chloroquine (CQ) being the first-line drug in the treatment policy of uncomplicated malaria. There is a scarcity in information about the magnitude of Plasmodium falciparum CQ resistance. This study aims to investigate the presence of single point mutations in the P. falciparum chloroquine-resistance transporter gene (pfcrt) at codons 76, 271, 326, 356 and 371 and in P. falciparum multi-drug resistance-1 gene (pfmdr1) at codons 86 and 1246, as molecular markers of CQ resistance.
The present study aimed to isolate actinobacteria from soil samples and characterized them using molecular tools and screened their secondary metabolites for antimicrobial activities. Thirty-nine strains from four different location of Barrientos Island, Antarctica using 12 types of isolation media was isolated. The isolates were preceded to screening of secondary metabolites for antimicrobial and antifungal activities. Using high-throughput screening methods, 38% (15/39) of isolates produced bioactive metabolites. Approximately 18% (7/39), 18% (7/39), 10% (4/39) and 2.5% (1/39) of isolates inhibited growth of Candida albicans ATCC 10231(T), Staphylococcus aurues ATCC 51650(T), methicillin-resistant Staphylococcus aurues (MRSA) ATCC BAA-44(T) and Pseudomonas aeruginosa ATCC 10145(T), respectively. Molecular characterization techniques like 16S rRNA analysis, Enterobacterial repetitive intergenic consensus-polymerase chain reaction (ERIC-PCR), Random amplified polymorphic DNA (RAPD) and composite analyses were used to characterize the actinobacteria strains. Analysis of 16S rRNA sequences is still one of the most powerful methods to determine higher taxonomic relationships of Actinobacteria. Both RAPD and ERIC-PCR fingerprinting have shown good discriminatory capability but RAPD proved to be better in discriminatory power than ERIC-PCR. Our results demonstrated that composite analysis of both fingerprinting generally increased the discrimination ability and generated best clustering for actinobacteria strains in this study.
The feasibility of langsat empty fruit bunch waste for preparation of activated carbon (EFBLAC) by microwave-induced activation was explored. Activation with NaOH at the IR ratio of 1.25, microwave power of 600 W for 6 min produced EFBLAC with a carbon yield of 81.31% and adsorption uptake for MB of 302.48 mg/g. Pore structural analysis, scanning electron microscopy and Fourier transform infrared spectroscopy demonstrated the physical and chemical characteristics of EFBLAC. Equilibrium data were best described by the Langmuir isotherm, with a monolayer adsorption capacity of 402.06 mg/g, and the adsorption kinetics was well fitted to the pseudo-second-order equation. The findings revealed the potential to prepare high quality activated carbon from langsat empty fruit bunch waste by microwave irradiation.
Epidemic foci of Plasmodium knowlesi malaria have been identified during the past ten years in Malaysia, in particular in the States of Sarawak and Sabah (Malaysia Borneo), and in the Pahang region (peninsular Malaysia). Based on a review of the available recent international literature, the authors underline the importance of molecular biology examinations, polymerase chain reactions (PCR), performed with primers specific for P. knowlesi, since the current microscopic examination (haemoscope) may fail to distinguish P. knowlesi from Plasmodium malariae, due to the very similar appearance of the two parasites. P. knowlesi has been described as the causal agent of life-threatening and lethal forms of malaria: its clinical picture is more severe when compared with that of P. malariae, since the disease is characterized by greater parasitaemia, as opposed to that documented in the course of P. malariae disease. The most effective carrier is Anopheles leucosphyrus: this mosquito is attracted by both humans and monkeys. Among primates, the natural hosts of P. knowlesi are Macaca fascicularis and Macaca nemestina, while Saimiri scirea and Macaca mulatta, which cannot become infected in nature, may be useful in experimental models. When underlining the potentially severe evolution, we note the key role played by prompt disease recognition, which is expected to be more straightforward in patients monitored in endemic countries at high risk, but should be carefully implemented for subjects being admitted to hospital in Western countries suffering from the typical signs and symptoms of malaria, after travelling in South-East Asia where they were engaged in excursions in the tropical forest (trekking, and similar outdoor activities). In these cases, the diagnosis should be prompt, and suitable treatment should follow. According to data in the literature, in non-severe cases chloroquine proves very effective against P. knowlesi, achieving the disappearance of signs and symptoms in 96% of cases after only 24 hours after treatment start. In the light of the emerging epidemiological data, P. knowlesi should be added to Plasmodium vivax, Plasmodium ovale, P. malariae, and Plasmodium falciparum, as the fifth aetiological agent of malaria. During the next few years, it will become mandatory to plan an appropriate surveillance program of the epidemiological evolution, paying also great attention to the clinical features of patients affected by P. knowlesi malaria, which are expected to worsen according to the time elapsed; some studies seem to point out greater severity according to increased parasitaemia, paralleling the increased interhuman infectious passages of the plasmodium.
The viability and activity of Bifidobacterium pseudocatenulatum G4, B. longum BB 536 and yoghurt cultures (Lactobacillus delbrueckii ssp. bulgaricus and Streptococcus thermophilus) were studied in yoghurt containing 0.75% Mangefira pajang fibrous polysaccharides (MPFP) and inulin. Growth of probiotic organisms, their proteolytic activities, the production of short chain fatty acids (lactic, acetic and propionic) and the pH of the yoghurt samples were determined during refrigerated storage at 4 °C for 28 d. B. pseudocatenulatum G4 and B. longum BB 536 showed better growth and activity in the presence of MPFP and inulin, which significantly increased the production of short chain fatty acids as well as the proteolytic activity of these organisms.
In the field of diagnostics, molecular amplification targeting unique genetic signature sequences has been widely used for rapid identification of infectious agents, which significantly aids physicians in determining the choice of treatment as well as providing important epidemiological data for surveillance and disease control assessment. We report the development of a rapid nucleic acid lateral flow biosensor (NALFB) in a dry-reagent strip format for the sequence-specific detection of single-stranded polymerase chain reaction (PCR) amplicons at ambient temperature (22-25°C). The NALFB was developed in combination with a linear-after-the-exponential PCR assay and the applicability of this biosensor was demonstrated through detection of the cholera toxin gene from diarrheal-causing toxigenic Vibrio cholerae. Amplification using the advanced asymmetric PCR boosts the production of fluorescein-labeled single-stranded amplicons, allowing capture probes immobilized on the NALFB to hybridize specifically with complementary targets in situ on the strip. Subsequent visual formation of red lines is achieved through the binding of conjugated gold nanoparticles to the fluorescein label of the captured amplicons. The visual detection limit observed with synthetic target DNA was 0.3 ng and 1 pg with pure genomic DNA. Evaluation of the NALFB with 164 strains of V. cholerae and non-V. cholerae bacteria recorded 100% for both sensitivity and specificity. The whole procedure of the low-cost NALFB, which is performed at ambient temperature, eliminates the need for preheated buffers or additional equipment, greatly simplifying the protocol for sequence-specific PCR amplicon analysis.
The current in vitro study was designed to investigate the anti-inflammatory, cytotoxic and antioxidant activities of boesenbergin A (BA), a chalcone derivative of known structure isolated from Boesenbergia rotunda. Human hepatocellular carcinoma (HepG2), colon adenocarcinoma (HT-29), non-small cell lung cancer (A549), prostate adenocarcinoma (PC3), and normal hepatic cells (WRL-68) were used to evaluate the cytotoxicity of BA using the MTT assay. The antioxidant activity of BA was assessed by the ORAC assay and compared to quercetin as a standard reference antioxidant. ORAC results are reported as the equivalent concentration of Trolox that produces the same level of antioxidant activity as the sample tested at 20 µg/mL. The toxic effect of BA on different cell types, reported as IC50, yielded 20.22 ± 3.15, 10.69 ± 2.64, 20.31 ± 1.34, 94.10 ± 1.19, and 9.324 ± 0.24 µg/mL for A549, PC3, HepG2, HT-29, and WRL-68, respectively. BA displayed considerable antioxidant activity, when the results of ORAC assay were reported as Trolox equivalents. BA (20 µg/mL) and quercetin (5 µg/mL) were equivalent to a Trolox concentration of 11.91 ± 0.23 and 160.32 ± 2.75 µM, respectively. Moreover, the anti-inflammatory activity of BA was significant at 12.5 to 50 µM and without any significant cytotoxicity for the murine macrophage cell line RAW 264.7 at 50 µM. The significant biological activities observed in this study indicated that BA may be one of the agents responsible for the reported biological activities of B. rotunda crude extract.
In this study, a total of 426 human faecal samples were examined for the presence of Entamoeba histolytica, Entamoeba dispar, Entamoeba moshkovskii infection via a combination of microscopic examination and nested polymerase chain reaction (PCR) targeting 16S ribosomal RNA of Entamoeba species.
Tor tambroides, a common and appreciated cyprinid fish of the Tasik Kenyir water reservoir in Malaysia, is one of the species selected for propagation. This fish was first successfully propagated in Malaysia by the Department of Agriculture, Sarawak, Malaysia, and the breeding program continued throughout the country. The gills were frequently infected by a Myxobolus species to be described as Myxobolus tambroides sp. n. The small, 50 to 70 μm, round plasmodia of this species is located intralamellarly. Plasmodia were filled with pyriform myxospores, 9.9 and 7.4 μm wide. In sutural view, the caudal end of the myxospores had a distinctive valvular groove, parallel with the suture. Plasmodia caused deformations on the affected and the neighbouring gill lamellae. The 18S rDNA sequence of M. tambroides sp.n. did not show a close relationship with any other Myxobolus spp., represented in the GenBank. This might be an emerging parasite likely to impact the propagation of this fish.
Bacillus strain NS 8, a lipase-producing bacterium isolated from a Malaysian hot spring, is able to tolerate a broad range of temperature and pH, which makes it beneficial for this study. It generated PCR products with molecular weight of 1,532 bp, and the 16S rRNA sequence analysis identified it as Bacillus subtilis with accession number AB110598. It showed a 71% similarity index with B. subtilis using Biolog Microstation System. Its lipase production was optimized using a shake flask system by changing the physical (agitation speed, pH and temperature) and nutritional (nitrogen, carbon and minerals) factors. The most suitable combination of the basal medium for lipase production was 2.5% olive oil (carbon), 1.5% peptone (nitrogen), 0.1% MgSO(4) (mineral) at an optimum temperature of 50°C, pH 7.5 and 150 rpm agitation, giving an enzyme yield of 4.23 U/ml. Statistical optimization using response surface methodology was carried out. An optimum lipase production of 5.67 U/ml was achieved when olive oil concentration of 3%, peptone 2%, MgSO(4)·7H(2)O 0.2% and an agitation rate of 200 rpm were combined. Lipase production was further carried out inside a 2-liter bioreactor, which yielded an enzyme activity of 14.5 U/ml after 15 h of incubation.
Prevalence, distribution and antibiotic resistance of Arcobacter spp. were investigated in cattle, goats, floor and treated water samples in this study. The prevalence of Arcobacter in adult and young was recorded as 8/110 (7.27%) and 4/83 (4.81%), respectively, which showed insignificant difference (P = 0.3503) in detection rates between adult and young cattle. A total of 33.33% of the floor samples and 11.11% of the treated water samples analysed were determined as positive for Arcobacter. Among the species isolated, over all, A. butzleri (45%) was the most frequently detected species, followed by A. skirrowii (5%). A. butzleri was isolated from adult cattle, floor and water samples at the rates of 75.0%, 33.4% and 50%, respectively. Co-colonization of species was not uncommon, and 50% of the samples were carrying more than one Arcobacter species. Only 12.5% sample from cattle (adult) was detected positive for only A. skirrowii. All samples from young animals, floor and water contained mixed isolates. None of the samples from goat farm was found to be carrying Arcobacter species. On profiling of antimicrobial resistance patterns, it was found that only one A. butzleri isolate (3.7%) was sensitive to all nine antibiotics tested. A. butzleri was found highly resistant to ampicillin (55.6%), followed by cefotaxime (33.4%) and ciprofloxacin (33.4%). Overall, 20% of the isolates showed multidrug resistance (resistant ≥4 antibiotics). Gentamicin and enrofloxacin can be used as drugs of choice for the treatment for Arcobacter infections.
Dengue virus (DENV) circulates in human and sylvatic cycles. Sylvatic strains are both ecologically and evolutionarily distinct from endemic viruses. Although sylvatic dengue cycles occur in West African countries and Malaysia, only a few cases of mild human disease caused by sylvatic strains and one single case of dengue hemorrhagic fever in Malaysia have been reported. Here we report a case of dengue hemorrhagic fever (DHF) with thrombocytopenia (13000/µl), a raised hematocrit (32% above baseline) and mucosal bleeding in a 27-year-old male returning to Spain in November 2009 after visiting his home country Guinea Bissau. Sylvatic DENV-2 West African lineage was isolated from blood and sera. This is the first case of DHF associated with sylvatic DENV-2 in Africa and the second case worldwide of DHF caused by a sylvatic strain.
Phaleria macrocarpa (Scheff.) Boerl (Thymelaceae) is commonly known as 'Crown of God', 'Mahkota Dewa', and 'Pau'. It originates from Papua Island, Indonesia and it grows in tropical areas. Empirically, it is potent in treating the hypertensive, diabetic, cancer and diuretic patients. It has a long history of ethnopharmacological usage, and the lack of information about its biological activities led us to investigate the possible biological activities by characterisation of flavonoids and antimicrobial activity of various part of P. macrocarpa against pathogenic bacteria and fungi. The results showed that kaempferol, myricetin, naringin, and rutin were the major flavonoids present in the pericarp while naringin and quercetin were found in the mesocarp and seed. Furthermore, the antibacterial activity of different parts of P. macrocarpa fruit showed a weak ability to moderate antibacterial activity against pathogenic tested bacteria (inhibition range: 0.93-2.17 cm) at concentration of 0.3 mg/disc. The anti fungi activity was only found in seed extract against Aspergillus niger (1.87 cm) at concentration of 0.3 mg/well. From the results obtained, P. macrocarpa fruit could be considered as a natural antimicrobial source due to the presence of flavonoid compounds.
Studies were conducted to assess the genetic relationships between the parental palms (dura and pisifera) and performance of their progenies based on nine microsatellite markers and 29 quantitative traits. Correlation analyses between genetic distances and hybrids performance were estimated. The coefficients of correlation values of genetic distances with hybrid performance were non-significant, except for mean nut weight and leaf number. However, the correlation coefficient of genetic distances with these characters was low to be used as predicted value. These results indicated that genetic distances based on the microsatellite markers may not be useful for predicting hybrid performance. The genetic distance analysis using UPGMA clustering system generated 5 genetic clusters with coefficient of 1.26 based on quantitative traits of progenies. The genotypes, DP16, DP14, DP4, DP13, DP12, DP15, DP8, DP1 and DP2 belonging to distant clusters and greater genetic distances could be selected for further breeding programs.
This study was conducted to evaluate the effects of topical application of aqueous extract of Hericium erinaceus fruiting bodies (HEFB) on the rate of wound healing enclosure and histology of the healed wound. Five groups of male Sprague-Dawley rats were experimentally wounded in the posterior neck area. A uniform wound area of 2.00 cm in diameter, using a circular stamp, was excised from the nape of the dorsal neck of all rats with the aid of a round seal. The animal groups were topically treated, respectively, with 0.2 mL each of sterilized distilled water (sdH2O); Intrasite gel; and 20, 30, and 40 mg/mL HEFB. Macroscopically, those rats whose wounds were dressed with HEFB and those in the Intrasite gel-treated group healed earlier than those treated with sdH2O. Histological analysis of healed wounds dressed with HEFB showed less scar width at wound enclosure and the healed wound contained fewer macrophages and more collagen with angiogenesis, compared to wounds dressed with sdH2O. In conclusion, wounds dressed with HEFB significantly enhanced the acceleration of wound healing enclosure in rats.
The HIV-positive population, due to their immuno-compromised nature, is considered more susceptible to parasitic infections than other populations. However despite the reports of other opportunistic pathogens such as Cryptosporidium and tuberculosis reported in vulnerable communities, microsporidia have not been highlighted in the local HIV-positive population in Malaysia. This study aimed to provide preliminary information on the prevalence of microsporidia in the local HIV-population. Microsporidia were detected in 21/247 (8.5%) stool samples from the HIV-infected individuals, a significantly higher (P-value <0.05) prevalence than in the control group, in which 5/173 (2.9%) were positive. HIV patients were 3x more at risk for acquiring microspordium (OR: 3.12; 95% CI 1.15-8.44). Spores were ellipsoid in shape with outlines that stained dark pink with the interior a lighter shade. Approximately 21% of the positive specimens were from individuals in the 40-49 years age group. Ten individuals who were positive for microsporidia were also positive for other enteric parasites such as Blastocystis hominis and Giardia lamblia. We detected Encephalitozoon intestinalis DNA following nested PCR from three of 10 samples analysed, as demonstrated by an amplicon of 370bp. From the findings reported, it appears that microsporidial infection in humans may actually be more common than reported. We strongly advocate greater emphasis on personal hygiene through public education on personal hygiene and the consumption of boiled or filtered water.
BACKGROUND AIMS. Dental pulp stromal cells (DPSC) are considered to be a promising source of stem cells in the field of regenerative therapy. However, the usage of DPSC in transplantation requires large-scale expansion to cater for the need for clinical quantity without compromising current good manufacturing practice (cGMP). Existing protocols for cell culturing make use of fetal bovine serum (FBS) as a nutritional supplement. Unfortunately, FBS is an undesirable additive to cells because it carries the risk of transmitting viral and prion diseases. Therefore, the present study was undertaken to examine the efficacy of human platelet lysate (HPL) as a substitute for FBS in a large-scale set-up. METHODS. We expanded the DPSC in Dulbecco's modified Eagle's medium-knock-out (DMEM-KO) with either 10% FBS or 10% HPL, and studied the characteristics of DPSC at pre- (T25 culture flask) and post- (5-STACK chamber) large-scale expansion in terms of their identity, quality, functionality, molecular signatures and cytogenetic stability. RESULTS. In both pre- and post-large-scale expansion, DPSC expanded in HPL showed extensive proliferation of cells (c. 2-fold) compared with FBS; the purity, immune phenotype, colony-forming unit potential and differentiation were comparable. Furthermore, to understand the gene expression profiling, the transcriptomes and cytogenetics of DPSC expanded under HPL and FBS were compared, revealing similar expression profiles. CONCLUSIONS. We present a highly economized expansion of DPSC in HPL, yielding double the amount of cells while retaining their basic characteristics during a shorter time period under cGMP conditions, making it suitable for therapeutic applications.