A new genus of onchidiid slugs, Wallaconchis Goulding & Dayrat, gen. n., is described, including ten species. Five species were previously described but known only from the type material: Wallaconchis ater (Lesson, 1830), W. graniferum (Semper, 1880), W. nangkauriense (Plate, 1893), W. buetschlii (Stantschinsky, 1907), and W. gracile (Stantschinsky, 1907), all of which were originally classified in Onchidium Buchannan, 1800. Many new records are provided for these five species, which greatly expand their known geographic distributions. Five species are new: Wallaconchis achleitneri Goulding, sp. n., W. comendadori Goulding & Dayrat, sp. n., W. melanesiensis Goulding & Dayrat, sp. n., W. sinanui Goulding & Dayrat, sp. n., and W. uncinus Goulding & Dayrat, sp. n. Nine of the ten Wallaconchis species are found in the Coral Triangle (eastern Indonesia and the Philippines). Sympatry is high, with up to six species found on the island of Bohol (Philippines) and eight species overlapping in northern Sulawesi (Indonesia). Wallaconchis is distinguished from other onchidiids by its bright dorsal colors (red, yellow, orange) but those are extremely variable and not useful for specific identification. Internally, the reproductive system can be used to identify all Wallaconchis species. The copulatory organs of Wallaconchis species are especially diverse compared to other onchidiid genera, and the possible role of reproductive incompatibility in species diversification is discussed. All specimens examined were freshly collected for the purpose of a worldwide revision of the Onchidiidae Rafinesque, 1815. The species are well delineated using DNA sequences and comparative anatomy. Mitochondrial DNA analysis yields thirteen molecular units separated by a large barcode gap, while nuclear DNA yields nine units. By integrating nuclear DNA and mitochondrial DNA with morphology, ten species are recognized. The natural history of each species (e.g., the microhabitat where they are found) is also documented. Nomenclature is addressed thoroughly (the types of all onchidiid species were examined, lectotypes were designated when needed, nomina dubia are discussed). Morphological characters, transitions to new microhabitats, and diversification processes are discussed in the context of a robust molecular phylogeny.
To investigate the epidemiological characteristics of influenza B viruses and explore the genetic evolution characteristics of the hemagglutinin(HA) and neuraminidase(NA) genes of local isolated strains in Ningbo, Southeast China, during 2010 to 2012.
To isolate and identify the pathogen of Dengue fever from Shenzhen city in 2005 - 2006, and to analyze the molecular characteristics of the isolated Dengue virus strain as well as to explore its possible origin.
Objective: To confirm the laboratory diagnosis of dengue bordline cases reported in Henan Province and trace its origin from molecular level in 2017. Methods: The study samples were blood samples (3-5 ml), which came from 8 suspected cases of dengue fever reported in the 2017 direct reporting system of Henan provincial infectious disease monitoring network. Meanwhile, case investigation was conducted according to National dengue fever surveillance programme. Serum were separated from blood samples and tested for Dengue NS1 antigen, IgM & IgG antibodies, and dengue RNA. According to dengue diagnosis criteria, confirmed cases were identified by testing results. Samples carried dengue RNA performed for real-time PCR genotyping and amplification of E gene. Then, the amplicons were sequenced and homological and phylogenetic analyses were constructed. Results: 8 serum samples of suspected dengue cases were collected in Henan Province, 2017. Six of them were diagnosed as dengue confirmed cases. All the dengue confirmed cases belonged to outside imported cases, 5 of them were positive by dengue RNA testing. Genotyping results showed there were 1 DENV1 case, 2 DENV2 cases and 2 DENV3 cases. A DENV2 case and a DENV3 case of this study were traced its origin successfully. The sequence of Pakistan imported DENV2 case belongs to cosmopolitan genotype, which was the most consistent with Pakistan's DENV2 KJ010186 in 2013 (identity 99.0%). The sequence of Malaysia imported DENV3 case belongs to genotype I, which was the most consistent with Singapore's DENV3 KX224276 in 2014(identity 99.0%). Conclusion: The laboratory diagnosis and molecular traceability of dengue cases in Henan Province in 2017 confirmed that all cases were imported and did not cause local epidemics.
To provide theoretical basis for the traceability and quality evaluation of edible bird's nests (EBNs), the Cytb sequence was applied to identify the origin of EBNs. A total of 39 experiment samples were collected from Malaysia, Indonesia, Vietnam and Thailand. Genomic DNA was extracted for the PCR reaction. The amplified products were sequenced. 36 sequences were downloaded from Gen Bank including edible nest swiftlet, black nest swiftlet, mascarene swiftlet, pacific swiftlet and germain's swiftlet. MEGA 7.0 was used to analyze the distinction of sequences by the method of calculating the distances in intraspecific and interspecific divergences and constructing NJ and UPMGA phylogenetic tree based on Kimera-2-parameter model. The results showed that 39 samples were from three kinds of EBNs. Interspecific divergences were significantly greater than the intraspecific one. Samples could be successfully distinguished by NJ and UPMGA phylogenetic tree. In conclusion, Cytb sequence could be used to distinguish the origin of EBNs and it is efficient for tracing the origin species of EBNs.
In a previous study, notable differences of several physicochemical properties, as well as the community structure of ammonia oxidizing bacteria as judged by 16S rRNA gene analysis, were observed among several disused tin-mining ponds located in the town of Kampar, Malaysia. These variations were associated with the presence of aquatic vegetation as well as past secondary activities that occurred at the ponds. Here, methane oxidizing bacteria (MOB), which are direct participants in the nutrient cycles of aquatic environments and biological indicators of environmental variations, have been characterised via analysis of pmoA functional genes in the same environments. The MOB communities associated with disused tin-mining ponds that were exposed to varying secondary activities were examined in comparison to those in ponds that were left to nature. Comparing the sequence and phylogenetic analysis of the pmoA clone libraries at the different ponds (idle, lotus-cultivated and post-aquaculture), we found pmoA genes indicating the presence of type I and type II MOB at all study sites, but type Ib sequences affiliated with the Methylococcus/Methylocaldum lineage were most ubiquitous (46.7 % of clones). Based on rarefaction analysis and diversity indices, the disused mining pond with lotus culture was observed to harbor the highest richness of MOB. However, varying secondary activity or sample type did not show a strong variation in community patterns as compared to the ammonia oxidizers in our previous study.
Disused tin-mining ponds make up a significant amount of water bodies in Malaysia particularly at the Kinta Valley in the state of Perak where tin-mining activities were the most extensive, and these abundantly available water sources are widely used in the field of aquaculture and agriculture. However, the natural ecology and physicochemical conditions of these ponds, many of which have been altered due to secondary post-mining activities, remains to be explored. As ammonia-oxidizing bacteria (AOB) are directly related to the nutrient cycles of aquatic environments and are useful bioindicators of environmental variations, the focus of this study was to identify AOBs associated with disused tin-mining ponds that have a history of different secondary activities in comparison to ponds which were left untouched and remained as part of the landscape. The 16S rDNA gene was used to detect AOBs in the sediment and water sampled from the three types of disused mining ponds, namely ponds without secondary activity, ponds that were used for lotus cultivation and post-aquaculture ponds. When the varying pond types were compared with the sequence and phylogenetic analysis of the AOB clone libraries, both Nitrosomonas and Nitrosospira-like AOB were detected though Nitrosospira spp. was seen to be the most ubiquitous AOB as it was present in all ponds types. However, AOBs were not detected in the sediments of idle ponds. Based on rarefaction analysis and diversity indices, the disused mining pond with lotus culture indicated the highest richness of AOBs. Canonical correspondence analysis indicated that among the physicochemical properties of the pond sites, TAN and nitrite were shown to be the main factors that influenced the community structure of AOBs in these disused tin-mining ponds.
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.
A chemically defined medium called KGm medium was used to isolate from a sample of sea water a bacterial strain, MW3A, capable of using N-3-oxohexanoyl-L: -homoserine lactone as the sole carbon source. MW3A was clustered closely to Pseudomonas aeruginosa by 16S ribosomal DNA sequence analysis. It degraded both N-acylhomoserine lactones (AHLs) with a 3-oxo group substitution and, less preferably, AHLs with unsubstituted groups at C3 position in the acyl side chain, as determined by Rapid Resolution Liquid Chromatography. Its quiP and pvdQ homologue gene sequences showed high similarities to those of known acylases. Spent supernatant of MW3A harvested at 8-h post inoculation was shown to contain long-chain AHLs when assayed with the biosensor Escherichia coli [pSB1075], and specifically N-dodecanoyl-L: -homoserine lactone and N-3-oxotetradecanoyl-L: -homoserine lactone by high resolution mass spectrometry. Hence, we report here a novel marine P. aeruginosa strain MW3A possessing both quorum-quenching and quorum-sensing properties.
Pleurotus citrinopileatus (yellow oyster mushroom) has an attractive shape and yellow colour but the fragile texture complicates packaging, and its strong aroma is unappealing to consumers. This study aimed to improve the characteristics and yield of P. citrinopileatus by interspecies mating between monokaryotic cultures of P. citrinopileatus and P. pulmonarius. Ten monokaryon cultures of the parental lines were crossed in all combinations to obtain hybrids. Eleven compatible mating pairs were obtained and cultivated to observe their sporophore morphology and yield. The selected hybrid, i.e. P1xC9, was beige in colour while hybrid P3xC8 was yellow in colour. Their sporophores had less offensive aroma, improved texture and higher yield. The DNA sequences of these hybrids were found to be in the same clade as the P. citrinopileatus parent with a bootstrap value of 99%. High bootstrap values indicate high genetic homology between hybrids and the P. citrinopileatus parent. The biological efficiencies of these hybrids P1xC9 (70.97%) and P3xC8 (52.14%) were also higher than the P. citrinopileatus parent (35.63%). Interspecies hybrids obtained by this mating technique can lead to better strains of mushrooms for genetic improvement of the Pleurotus species.
Bud rot disease is a damaging disease of oil palm in Colombia. The pathogen responsible for this disease is a species of oomyctes, Phytophthora palmivora which is also the causal pathogen of several tropical crop diseases such as fruit rot and stem canker of cocoa, rubber, durian and jackfruit. No outbreaks of bud rot have been reported in oil palm in Malaysia or other Southeast Asian countries, despite this particular species being present in the region. Analysis of the genomic sequences of several genetic markers; the internal transcribe spacer regions (ITS) of the ribosomal RNA gene cluster, beta-tubulin gene, translation elongation factor 1 alpha gene (EF-1α), cytochrome c oxidase subunit I & II (COXI and COXII) gene cluster along with amplified fragment length polymorphism (AFLP) analyses have been carried out to investigate the genetic diversity and variation of P. palmivora isolates from around the world and from different hosts in comparison to Colombian oil palm isolates, as one of the steps in understanding why this species of oomycetes causes devastating damage to oil palm in Latin America but not in other regions. Phylogenetic analyses of these regions showed that the Colombian oil palm isolates were not separated from Malaysian isolates. AFLP analysis and a new marker PPHPAV, targeting an unclassified hypothetical protein, was found to be able to differentiate Malaysian and Colombian isolates and showed a clear clade separations. Despite this, pathogenicity studies did not show any significant differences in the level of aggressiveness of different isolates against oil palm in glasshouse tests.
Banana is one of the most important fruits cultivated in Malaysia, and it provides many health benefits. However, bacterial wilt disease, which attacks bananas, inflicts major losses on the banana industry in Malaysia. To understand the complex interactions of the microbiota of bacterial wilt-diseased banana plants, we first determined the bacterial communities residing in the pseudostems of infected (symptomatic) and diseased-free (non-symptomatic) banana plants. We characterized the associated microorganisms using the targeted 16S rRNA metagenomics sequencing on the Illumina MiSeq platform. Taxonomic classifications revealed 17 and nine known bacterial phyla in the tissues of non-symptomatic and symptomatic plants, respectively. Cyanobacteria and Proteobacteria (accounted for more than 99% of the 16S rRNA gene fragments) were the two most abundant phyla in both plants. The five major genera found in both plant samples were Ralstonia, Sphingomonas, Methylobacterium, Flavobacterium, and Pseudomonas. Ralstonia was more abundant in symptomatic plant (59% out of the entire genera) as compared to those in the non-symptomatic plant (only 36%). Our data revealed that 102 bacterial genera were only assigned to the non-symptomatic plant. Overall, this study indicated that more diverse and abundant microbiota were associated with the non-symptomatic bacterial wilt-diseased banana plant as compared to the symptomatic plant. The higher diversity of endophytic microbiota in the non-symptomatic banana plant could be an indication of pathogen suppression which delayed or prevented the disease expression. This comparative study of the microbiota in the two plant conditions might provide caveats for potential biological control strategies.
Ganoderma boninense, the main causal agent of oil palm (Elaeis guineensis) basal stem rot (BSR), severely reduces oil palm yields around the world. To reduce reliance on fungicide applications to control BSR, we are investigating the efficacy of alternative control methods, such as the application of biological control agents. In this study, we used four Streptomyces-like actinomycetes (isolates AGA43, AGA48, AGA347 and AGA506) that had been isolated from the oil palm rhizosphere and screened for antagonism towards G. boninense in a previous study. The aim of this study was to characterize these four isolates and then to assess their ability to suppress BSR in oil palm seedlings when applied individually to the soil in a vermiculite powder formulation. Analysis of partial 16S rRNA gene sequences (512 bp) revealed that the isolates exhibited a very high level of sequence similarity (> 98%) with GenBank reference sequences. Isolates AGA347 and AGA506 showed 99% similarity with Streptomyces hygroscopicus subsp. hygroscopicus and Streptomyces ahygroscopicus, respectively. Isolates AGA43 and AGA48 also belonged to the Streptomyces genus. The most effective formulation, AGA347, reduced BSR in seedlings by 73.1%. Formulations using the known antifungal producer Streptomyces noursei, AGA043, AGA048 or AGA506 reduced BSR by 47.4, 30.1, 54.8 and 44.1%, respectively. This glasshouse trial indicates that these Streptomyces spp. show promise as potential biological control agents against Ganoderma in oil palm. Further investigations are needed to determine the mechanism of antagonism and to increase the shelf life of Streptomyces formulations.
Microbial flocs formed from raw textile wastewater in a prototype Aerobic Biofilm Reactor (ABR) system were characterised and studied for their potential use in the treatment of textile wastewater. After 90-100 days of operation, microbial flocs of loose irregular structures were obtained from the reactor with good settling velocity of 33 m/h and sludge volume index (SVI) of 48.2 mL/g. Molecular analysis of the flocs using PCR-amplified 16S rDNA sequence showed 98% homology to those of Bacillus sp, Paenibacillus sp and Acromobacter sp. Detection of Ca(2+)(131 mg/g) and Fe(2+)(131 mg/g) using atomic absorption spectrometer might be implicated with the flocs formation. In addition, presence of Co(2+) and Ni(2+) were indicative of the flocs ability to accumulate at least a fraction of the metals' present in the wastewater. When the flocs were used for the treatment of raw textile wastewater, they showed good removal of COD and colour about 55% and 70% respectively, indicating their potential application.
Textile wastewater, one of the most polluted industrial effluents, generally contains substantial amount of dyes and chemicals that will cause increase in the COD, colour and toxicity of receiving water bodies if not properly treated. Current treatment methods include chemical and biological processes; the efficiency of the biological treatment method however, remains uncertain since the discharged effluent is still highly coloured. In this study, granules consisting mixed culture of decolourising bacteria were developed and the physical and morphological characteristics were determined. After the sixth week of development, the granules were 3-10 mm in diameter, having good settling property with settling velocity of 70 m/h, sludge volume index (SVI) of 90 to 130 mL/g, integrity coefficient of 3.7, and density of 66 g/l. Their abilities to treat sterilised raw textile wastewater were evaluated based on the removal efficiencies of COD (initial ranging from 200 to 3,000 mg/L), colour (initial ranging from 450 to 2000 ADMI) of sterilised raw textile wastewater with pH from 6.8 to 9.4. Using a sequential anaerobic-aerobic treatment cycle with hydraulic retention time (HRT) of 24 h, maximum removal of colour and COD achieved was 90% and 80%, respectively.
Intensive aeration for nitrification is a major energy consumer in sewage treatment plants (STPs). Low-dissolved-oxygen (low-DO) nitrification has the potential to lower the aeration demand. However, the applicability of low-DO nitrification in the tropical climate is not well-understood. In this study, the potential of low-DO nitrification in tropical setting was first examined using batch kinetic experiments. Subsequently, the performance of low-DO nitrification was investigated in a laboratory-scale sequential batch reactor (SBR) for 42 days using real tropical sewage. The batch kinetic experiments showed that the seed sludge has a relatively high oxygen affinity. Thus, the rate of nitrification was not significantly reduced at low DO concentrations (0.5 mg/L). During the operation of the low-DO nitrification SBR, 90% of NH4-N was removed. The active low-DO nitrification was mainly attributed to the limited biodegradable organics in the sewage. Fluorescence in-situ hybridisation and 16S rRNA amplicon sequencing revealed the nitrifiers were related to Nitrospira genus and Nitrosomonadaceae family. Phylogenetic analysis suggests 47% of the operational taxonomic units in Nitrospira genus are closely related to a comammox bacteria. This study has demonstrated active low-DO nitrification in tropical setting, which is a more sustainable process that could significantly reduce the energy footprint of STPs.
Reserve lipids of microalgae are promising for biodiesel production. However, economically feasible and sustainable energy production from microalgae requires optimization of cultivation conditions for both biomass yield and lipid production of microalgae. Biomass yield and lipid production in microalgae are a contradictory problem because required conditions for both targets are different. Simultaneously, the mass cultivation of microalgae for biofuel production also depends extremely on the performance of the microalgae strains used. In this study a green unicellular microalgae Chlorella sorokiniana (DS6) isolated from the holding tanks of farm wastewater treatment plant using multi-step screening and acclimation procedures was found high-lipid producing facultative heterotrophic microalgae strain capable of growing on dairy farm effluent (DFE) for biodiesel feedstock and wastewater treatment. Morphological features and the phylogenetic analysis for the 18S rRNA identified the isolated strains. A novel three stage cultivation process of facultative strain of C. sorokiniana was examined for lipid production.
Mass mortalities of the larval stage of the giant freshwater prawn, Macrobrachium rosenbergii, have been occurring in Bangladesh since 2011. Mortalities can reach 100% and have resulted in an 80% decline in the number of hatcheries actively producing M. rosenbergii. To investigate a causative agent for the mortalities, a disease challenge was carried out using infected material from a hatchery experiencing mortalities. Moribund larvae from the challenge were prepared for metatranscriptomic sequencing. De novo virus assembly revealed a 29 kb single‑stranded positive-sense RNA virus with similarities in key protein motif sequences to yellow head virus (YHV), an RNA virus that causes mass mortalities in marine shrimp aquaculture, and other viruses in the Nidovirales order. Primers were designed against the novel virus and used to screen cDNA from larvae sampled from hatcheries in the South of Bangladesh from two consecutive years. Larvae from all hatcheries screened from both years were positive by PCR for the novel virus, including larvae from a hatchery that at the point of sampling appeared healthy, but later experienced mortalities. These screens suggest that the virus is widespread in M. rosenbergii hatchery culture in southern Bangladesh, and that early detection of the virus can be achieved by PCR. The hypothesised protein motifs of Macrobrachium rosenbergii golda virus (MrGV) suggest that it is likely to be a new species within the Nidovirales order. Biosecurity measures should be taken in order to mitigate global spread through the movement of post-larvae within and between countries, which has previously been linked to other virus outbreaks in crustacean aquaculture.
Coronavirus-like organisms have been previously identified in Arthropod ectoparasites (such as ticks and unfed cat flea). Yet, the question regarding the possible role of these arthropods as SARS-CoV-2 passive/biological transmission vectors is still poorly explored. In this study, we performed in silico structural and binding energy calculations to assess the risks associated with possible ectoparasite transmission. We found sufficient similarity between ectoparasite ACE and human ACE2 protein sequences to build good quality 3D-models of the SARS-CoV-2 Spike:ACE complex to assess the impacts of ectoparasite mutations on complex stability. For several species (e.g., water flea, deer tick, body louse), our analyses showed no significant destabilisation of the SARS-CoV-2 Spike:ACE complex, suggesting these species would bind the viral Spike protein. Our structural analyses also provide structural rationale for interactions between the viral Spike and the ectoparasite ACE proteins. Although we do not have experimental evidence of infection in these ectoparasites, the predicted stability of the complex suggests this is possible, raising concerns of a possible role in passive transmission of the virus to their human hosts.