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Presbytis neglectus or P. femoralis, Colobine Molecular Phylogenies, and GenBank Submission of Newly Generated DNA Sequences

Nijman V

Folia Primatol., 2020;91(3):228-239.
PMID: 31578029 DOI: 10.1159/000502093

Great progress has been made in unravelling the evolutionary history of Asian colobines, largely through the use of dated molecular phylogenies based on multiple markers. The Presbytis langurs are a case in point, with more allopatric species being identified, recognition of Presbytis thomasi from Sumatra rather than P. potenziani from the Mentawai Islands as being the most basal species of the group, and the discovery that P. rubicunda from Borneo is nested among the Sumatran species and only made it to Borneo in the last 1.3 million years. Based on variation in mitochondrial d-loop, it has recently been argued that Malaysia's P. femoralis femoralis is actually P. neglectus neglectus. Unfortunately, despite being available, sequences from the type locality, Singapore, were excluded from the analysis, and none of the newly generated sequences was deposited in GenBank. I manually reconstructed these sequences, which allowed me to present a molecular phylogeny that includes 8 additional sequences from West Malaysia and Singapore. P. neglectus from Malaysia and P. femoralis from Singapore form one monophyletic clade, with minimal divergence. I conclude that recognition of P. neglectus is erroneous and the name is a junior synonym of P. femoralis. Colobine taxonomy and systematics have advanced, and continue to advance, mostly by considering evidence from a wide range of individuals, species and data sets (molecular, behavioural and morphological) rather than focusing on single molecular markers from 1 or 2 species from one small geographic area. For an orderly taxonomic debate where evidence can be evaluated and reinterpreted it is essential that newly generated sequences are deposited in public repositories.

Matched MeSH terms: Sequence Analysis, DNA
Fulltext Construction of an escherichia coli expression vector for the non-structural (ns)-1 protein of avian influenza virus H5N1

Agusta, Istiqomah, Jacinta Santhanam, Yap, Wei Boon

Jurnal Sains Kesihatan Malaysia, 2020;18(2):73-81.
MyJurnal

In the search for universal vaccine candidates for the prevention of avian influenza, the non-structural (NS)-1 protein of avian influenza virus (AIV) H5N1 has shown promising potential for its ability to effectively stimulate the host immunity. This study was aimed to produce a bacterial expression plasmid using pRSET B vector to harbour the NS1 gene of AIV H5N1 (A/Chicken/Malaysia/5858/2004 (H5N1)) for protein expression in Escherichia coli (E. coli). The NS1 gene (687 bp) was initially amplified by polymerase chain reaction (PCR) and then cloned into a pGEM-T Easy TA vector. The NS1 gene was released from pGEM-T-NS1 using EcoRI and XhoI restriction enzymes (RE). The pRSET B vector was also linearized using the same RE. The digested NS1 gene and linearized pRSET B were ligated using T4 DNA ligase to form the expression plasmid, pRSET B-NS1. The NS1 gene sequence in pRSET B-NS1 was confirmed by DNA sequencing. To prepare recombinant bacterial cells for protein expression in the future, pRSET B-NS1 was transformed into E. coli strain BL21 (DE3) by heat-shock. Colonies bearing the recombinant plasmid were screened using PCR. The DNA sequencing analysis revealed that the NS1 gene sequence was 97% homologous to that of AIV H5N1 A/Chicken/Malaysia/5858/2004 (H5N1). These results indicated that the NS1 gene of influenza A/Chicken/Malaysia/5858/2004 (H5N1) was successfully amplified and cloned into a pRSET B vector. Bacterial colonies carrying pRSET B-NS1 can be used for the synthesis of NS1-based influenza vaccine in the future and thereby aid in the prevention of avian influenza.

Matched MeSH terms: Sequence Analysis, DNA
Sequencing and characterization of complete mitogenome DNA of Rasbora tornieri (Cypriniformes: Cyprinidae: Rasbora) and its evolutionary significance

Chung HH, Anak Kamar CK, Kit Lim LW, Roja JS, Liao Y, Tsan-Yuk Lam T, et al.

J Genet, 2020;99.
PMID: 32893838

The yellowtail rasbora (Rasbora tornieri) is a miniature ray-finned fish categorized under the genus Rasbora in the family of Cyprinidae. In this study, a complete mitogenome sequence of R. tornieri was sequenced using four primers targeting two halves of the mitogenome with overlapping flanking regions. The size of mitogenome was 16,573 bp, housing 22 transfer RNA genes, 13 protein-coding genes, two ribosomal RNA genes and a putative control region. Identical gene organization was detected between this species and other members of Rasbora genus. The heavy strand encompassed 28 genes while the light strand accommodated the other nine genes. Most protein-coding genes execute ATG as start codon, excluding COI and ND3 genes, which utilized GTG instead. The central conserved sequence blocks (CSB-E, CSB-F and CSB-D), variable sequence blocks (CSB-1, CSB-3 and CSB-2) as well as the terminal associated sequence (TAS) were conserved within the control region. The maximum likelihood phylogenetic family tree revealed the divergence of R. tornieri from the basal region of the Rasbora clade, where its evolutionary relationships with other Rasbora members are poorly resolved as indicated by the low bootstrap values. This work acts as window for further population genetics and molecular evolution studies of Rasbora genus in future.

Matched MeSH terms: Sequence Analysis, DNA/methods*
Fulltext Mitochondrial DNA mutations in Malaysian female breast cancer patients

Omasanggar R, Yu CY, Ang GY, Emran NA, Kitan N, Baghawi A, et al.

PLoS One, 2020;15(5):e0233461.
PMID: 32442190 DOI: 10.1371/journal.pone.0233461

Cancer development has been ascribed with diverse genetic variations which are identified in both mitochondrial and nuclear genomes. Mitochondrial DNA (mtDNA) alterations have been detected in several tumours which include lung, colorectal, renal, pancreatic and breast cancer. Several studies have explored the breast tumour-specific mtDNA alteration mainly in Western population. This study aims to identify mtDNA alterations of 20 breast cancer patients in Malaysia by next generation sequencing analysis. Twenty matched tumours with corresponding normal breast tissues were obtained from female breast cancer patients who underwent mastectomy. Total DNA was extracted from all samples and the entire mtDNA (16.6kb) was amplified using long range PCR amplification. The amplified PCR products were sequenced using mtDNA next-generation sequencing (NGS) on an Illumina Miseq platform. Sequencing involves the entire mtDNA (16.6kb) from all pairs of samples with high-coverage (~ 9,544 reads per base). MtDNA variants were called and annotated using mtDNA-Server, a web server. A total of 18 of 20 patients had at least one somatic mtDNA mutation in their tumour samples. Overall, 65 somatic mutations were identified, with 30 novel mutations. The majority (59%) of the somatic mutations were in the coding region, whereas only 11% of the mutations occurred in the D-loop. Notably, somatic mutations in protein-coding regions were non-synonymous (49%) in which 15.4% of them are potentially deleterious. A total of 753 germline mutations were identified and four of which were novel mutations. Compared to somatic alterations, less than 1% of germline missense mutations are harmful. The findings of this study may enhance the current knowledge of mtDNA alterations in breast cancer. To date, the catalogue of mutations identified in this study is the first evidence of mtDNA alterations in Malaysian female breast cancer patients.

Matched MeSH terms: Sequence Analysis, DNA
Fulltext Museomics for reconstructing historical floristic exchanges: Divergence of stone oaks across Wallacea

Strijk JS, Binh HT, Ngoc NV, Pereira JT, Slik JWF, Sukri RS, et al.

PLoS One, 2020;15(5):e0232936.
PMID: 32442164 DOI: 10.1371/journal.pone.0232936

Natural history collections and tropical tree diversity are both treasure troves of biological and evolutionary information, but their accessibility for scientific study is impeded by a number of properties. DNA in historical specimens is generally highly fragmented, complicating the recovery of high-grade genetic material. Furthermore, our understanding of hyperdiverse, wide-spread tree assemblages is obstructed by extensive species ranges, fragmented knowledge of tropical tree diversity and phenology, and a widespread lack of species-level diagnostic characters, prohibiting the collecting of readily identifiable specimens which can be used to build, revise or strengthen taxonomic frameworks. This, in turn, delays the application of downstream conservation action. A sizable component of botanical collections are sterile-thus eluding identification and are slowing down progress in systematic treatments of tropical biodiversity. With rapid advances in genomics and bioinformatic approaches to biodiversity research, museomics is emerging as a new field breathing life into natural collections that have been built up over centuries. Using MIGseq (multiplexed ISSR genotyping by sequencing), we generated 10,000s of short loci, for both freshly collected materials and museum specimens (aged >100 years) of Lithocarpus-a widespread tropical tree genus endemic to the Asian tropics. Loci recovery from historical and recently collected samples was not affected by sample age and preservation history of the study material, underscoring the reliability and flexibility of the MIGseq approach. Phylogenomic inference and biogeographic reconstruction across insular Asia, highlights repeated migration and diversification patterns between continental regions and islands. Results indicate that co-occurring insular species at the extremity of the distribution range are not monophyletic, raising the possibility of multiple independent dispersals along the outer edge of Wallacea. This suggests that dispersal of large seeded tree genera throughout Malesia and across Wallacea may have been less affected by large geographic distances and the presence of marine barriers than generally assumed. We demonstrate the utility of MIGseq in museomic studies using non-model taxa, presenting the first range-wide genomic assessment of Lithocarpus and tropical Fagaceae as a proof-of-concept. Our study shows the potential for developing innovative genomic approaches to improve the capture of novel evolutionary signals using valuable natural history collections of hyperdiverse taxa.

Matched MeSH terms: Sequence Analysis, DNA/methods*
Fulltext Whole-genome sequence of multi-drug resistant Pseudomonas aeruginosa strains UY1PSABAL and UY1PSABAL2 isolated from human broncho-alveolar lavage, Yaoundé, Cameroon

Madaha EL, Mienie C, Gonsu HK, Bughe RN, Fonkoua MC, Mbacham WF, et al.

PLoS One, 2020;15(9):e0238390.
PMID: 32886694 DOI: 10.1371/journal.pone.0238390

Pseudomonas aeruginosa has been implicated in a wide range of post-operation wound and lung infections. A wide range of acquired resistance and virulence markers indicate surviving strategy of P. aeruginosa. Complete-genome analysis has been identified as efficient approach towards understanding the pathogenicity of this organism. This study was designed to sequence the entire genome of P. aeruginosa UY1PSABAL and UY1PSABAL2; determine drug-resistance profiles and virulence factors of the isolates; assess factors that contribute toward stability of the genomes; and thereafter determine evolutionary relationships between the strains and other isolates from similar sources. The genomes of the MDR P. aeruginosa UY1PSABAL and UY1PSABAL2 were sequenced on the Illumina Miseq platform. The raw sequenced reads were assessed for quality using FastQC v.0.11.5 and filtered for low quality reads and adapter regions using Trimmomatic v.0.36. The de novo genome assembly was made with SPAdes v.3.13 and annotated using Prokka v.2.1.1 annotation pipeline; Rapid Annotation using Subsytems Technology (RAST) server v.2.0; and PATRIC annotation tool v.3.6.2. Antimicrobial resistance genes and virulence determinants were searched through the functional annotation data generated from Prokka, RAST and PATRIC annotation pipelines; In addition to ResFinder and Comprehensive Antibiotic Resistance Database (CARD) which were employed to determine resistance genes. The PHAge Search Tool Enhanced Release (PHASTER) web server was used for the rapid identification and annotation of prophage sequences within bacterial genome. Predictive secondary metabolites were identified with AntiSMASH v.5.0. Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) and cas genes regions were also investigated with the CRISPRone and CRISPRFinder server. The genome sizes of 7.0 and 6.4 Mb were determined for UY1PSABAL and UY1PSABAL2 strains with G+C contents of 66.1% and 66.48% respectively. β-lactamines resistance genes blaPAO, aminoglycoside phosphorylating enzymes genes aph(3')-IIb, fosfomycine resistance gene fosA, vancomycin vanW and tetracycline tetA were among identified resistance genes harboured in both isolates. UY1PSABAL bore additional aph(6)-Id, aph(3'')-Ib, ciprofloxacin-modifying enzyme crpP and ribosomal methylation enzyme rmtB. Both isolates were found harbouring virulence markers such as flagella and type IV pili; and also present various type III secretion systems such as exoA, exoS, exoU, exoT. Secondary metabolites such as pyochelin and pyoverdine with iron uptake activity were found within the genomes as well as quorum-sensing systems, and various fragments for prophages and insertion sequences. Only the UY1PSABAL2 contains CRISPR-Cas system. The phylogeny revealed a very close evolutionary relationship between UY1PSABAL and the similar strain isolated from Malaysia; the same trend was observed between UY1PSABAL2 and the strain from Chinese origin. Complete analyses of the entire genomes provide a wide range of information towards understanding pathogenicity of the pathogens in question.

Matched MeSH terms: Sequence Analysis, DNA
Fulltext Population Genomics of an Anadromous Hilsa Shad Tenualosa ilisha Species across Its Diverse Migratory Habitats: Discrimination by Fine-Scale Local Adaptation

Asaduzzaman M, Igarashi Y, Wahab MA, Nahiduzzaman M, Rahman MJ, Phillips MJ, et al.

Genes (Basel), 2019 12 30;11(1).
PMID: 31905942 DOI: 10.3390/genes11010046

The migration of anadromous fish in heterogenic environments unceasingly imposes a selective pressure that results in genetic variation for local adaptation. However, discrimination of anadromous fish populations by fine-scale local adaptation is challenging because of their high rate of gene flow, highly connected divergent population, and large population size. Recent advances in next-generation sequencing (NGS) have expanded the prospects of defining the weakly structured population of anadromous fish. Therefore, we used NGS-based restriction site-associated DNA (NextRAD) techniques on 300 individuals of an anadromous Hilsa shad (Tenualosa ilisha) species, collected from nine strategic habitats, across their diverse migratory habitats, which include sea, estuary, and different freshwater rivers. The NextRAD technique successfully identified 15,453 single nucleotide polymorphism (SNP) loci. Outlier tests using the FST OutFLANK and pcadapt approaches identified 74 and 449 SNPs (49 SNPs being common), respectively, as putative adaptive loci under a divergent selection process. Our results, based on the different cluster analyses of these putatively adaptive loci, suggested that local adaptation has divided the Hilsa shad population into two genetically structured clusters, in which marine and estuarine collection sites were dominated by individuals of one genetic cluster and different riverine collection sites were dominated by individuals of another genetic cluster. The phylogenetic analysis revealed that all the riverine populations of Hilsa shad were further subdivided into the north-western riverine (turbid freshwater) and the north-eastern riverine (clear freshwater) ecotypes. Among all of the putatively adaptive loci, only 36 loci were observed to be in the coding region, and the encoded genes might be associated with important biological functions related to the local adaptation of Hilsa shad. In summary, our study provides both neutral and adaptive contexts for the observed genetic divergence of Hilsa shad and, consequently, resolves the previous inconclusive findings on their population genetic structure across their diverse migratory habitats. Moreover, the study has clearly demonstrated that NextRAD sequencing is an innovative approach to explore how dispersal and local adaptation can shape genetic divergence of non-model anadromous fish that intersect diverse migratory habitats during their life-history stages.

Matched MeSH terms: Sequence Analysis, DNA
Variations in mitochondrial DNA control region among Malay and Chinese subpopulations (sequence 16000-16200)

Ishar SM, Parameswaran K, Masduki NS, Rus Din RD

Mitochondrial DNA A DNA Mapp Seq Anal, 2019 12;30(8):843-847.
PMID: 31709874 DOI: 10.1080/24701394.2019.1687693

DNA variations are alterations found in DNA sequence, occurring in both nuclear DNA and mitochondrial DNA. Variations might differ in individual following population, respectively. The aim of this study was to find variations in target sequence of mtDNA (16000-16200) to be used as marker in Malay and Chinese population. A total of 30 buccal swab samples from 20 Malay and 10 Chinese subjects were collected and preserved on FTA card. The FTA card that contained DNA sample was punched to be included into polymerase chain reaction mixture. Amplification was carried out and the products were sequenced. Sequence variations were found in both Malay and Chinese populations. A total of nine variations (16129, 16108, 16162, 16172, 16148, 16127, 16173, 16099 and 16100) were found in Malay population while a total of seven variations (16129, 16104, 16111, 16109, 16164, 16170 and 16136) were found in Chinese population. Nucleotide position 16129 was found as variation in both Malay and Chinese populations. This study implies that np 16129 can be used as a marker for Malaysian population. For further investigation, the length of the target sequence may be increased to obtain more variations that can be used as markers. This will increase the discrimination power of Malaysian population.

Matched MeSH terms: Sequence Analysis, DNA*
Fulltext Genetic diversity of Nile tilapia (Oreochromis niloticus) throughout West Africa

Lind CE, Agyakwah SK, Attipoe FY, Nugent C, Crooijmans RPMA, Toguyeni A

Sci Rep, 2019 11 14;9(1):16767.
PMID: 31727970 DOI: 10.1038/s41598-019-53295-y

Nile tilapia (Oreochromis niloticus) is a globally significant aquaculture species rapidly gaining status as a farmed commodity. In West Africa, wild Nile tilapia genetic resources are abundant yet knowledge of fine-scale population structure and patterns of natural genetic variation are limited. Coinciding with this is a burgeoning growth in tilapia aquaculture in Ghana and other countries within the region underpinned by locally available genetic resources. Using 192 single nucleotide polymorphism (SNP) markers this study conducted a genetic survey of Nile tilapia throughout West Africa, sampling 23 wild populations across eight countries (Benin, Burkina Faso, Côte d'Ivoire, Ghana, Togo, Mali, Gambia and Senegal), representing the major catchments of the Volta, Niger, Senegal and Gambia River basins. A pattern of isolation-by-distance and significant spatial genetic structure was identified throughout West Africa (Global FST = 0.144), which largely corresponds to major river basins and, to a lesser extent, sub-basins. Two populations from the Gambia River (Kudang and Walekounda), one from the western Niger River (Lake Sélingué) and one from the upper Red Volta River (Kongoussi) showed markedly lower levels of diversity and high genetic differentiation compared to all other populations, suggesting genetically isolated populations occurring across the region. Genetic structure within the Volta Basin did not always follow the pattern expected for sub-river basins. This study identifies clear genetic structuring and differentiation amongst West African Nile tilapia populations, which concur with broad patterns found in previous studies. In addition, we provide new evidence for fine-scale genetic structuring within the Volta Basin and previously unidentified genetic differences of populations in Gambia. The 192 SNP marker suite used in this study is a useful tool for differentiating tilapia populations and we recommend incorporating this marker suite into future population screening of O. niloticus. Our results form the basis of a solid platform for future research on wild tilapia genetic resources in West Africa, and the identification of potentially valuable germplasm for use in ongoing breeding programs for aquaculture.

Matched MeSH terms: Sequence Analysis, DNA/methods*
Indel-informed Bayesian analysis suggests cryptic population structure between Plasmodium knowlesi of humans and long-tailed macaques (Macaca fascicularis) in Malaysian Borneo

Wilcox JS, Kerschner A, Hollocher H

Infect Genet Evol, 2019 11;75:103994.
PMID: 31421245 DOI: 10.1016/j.meegid.2019.103994

Plasmodium knowlesi is an important causative agent of malaria in humans of Southeast Asia. Macaques are natural hosts for this parasite, but little is conclusively known about its patterns of transmission within and between these hosts. Here, we apply a comprehensive phylogenetic approach to test for patterns of cryptic population genetic structure between P. knowlesi isolated from humans and long-tailed macaques from the state of Sarawak in Malaysian Borneo. Our approach differs from previous investigations through our exhaustive use of archival 18S Small Subunit rRNA (18S) gene sequences from Plasmodium and Hepatocystis species, our inclusion of insertion and deletion information during phylogenetic inference, and our application of Bayesian phylogenetic inference to this problem. We report distinct clades of P. knowlesi that predominantly contained sequences from either human or macaque hosts for paralogous A-type and S-type 18S gene loci. We report significant partitioning of sequence distances between host species across both types of loci, and confirmed that sequences of the same locus type showed significantly biased assortment into different clades depending on their host species. Our results support the zoonotic potential of Plasmodium knowlesi, but also suggest that humans may be preferentially infected with certain strains of this parasite. Broadly, such patterns could arise through preferential zoonotic transmission of some parasite lineages or a disposition of parasites to transmit within, rather than between, human and macaque hosts. Available data are insufficient to address these hypotheses. Our results suggest that the epidemiology of P. knowlesi may be more complicated than previously assumed, and highlight the need for renewed and more vigorous explorations of transmission patterns in the fifth human malarial parasite.

Matched MeSH terms: Sequence Analysis, DNA/methods*
Fulltext Diversity, enumeration, and isolation of Arcobacter spp. in the giant abalone, Haliotis gigantea

Mizutani Y, Iehata S, Mori T, Oh R, Fukuzaki S, Tanaka R

Microbiologyopen, 2019 10;8(10):e890.
PMID: 31168933 DOI: 10.1002/mbo3.890

Arcobacter have been frequently detected in and isolated from bivalves, but there is very little information on the genus Arcobacter in the abalone, an important fishery resource. This study aimed to investigate the genetic diversity and abundance of bacteria from the genus Arcobacter in the Japanese giant abalone, Haliotis gigantea, using molecular methods such as Arcobacter-specific clone libraries and fluorescence in situ hybridization (FISH). Furthermore, we attempted to isolate the Arcobacter species detected. Twelve genotypes of clones were obtained from Arcobacter-specific clone libraries. These sequences are not classified with any other known Arcobacter species including pathogenic Arcobacter spp., A. butzleri, A. skirrowii, and A. cryaerophilus, commonly isolated or detected from bivalves. From the FISH analysis, we observed that ARC94F-positive cells, presumed to be Arcobacter, accounted for 6.96 ± 0.72% of all EUB338-positive cells. In the culture method, three genotypes of Arcobacter were isolated from abalones. One genotype had a similarity of 99.2%-100.0% to the 16S rRNA gene of Arcobacter marinus, while the others showed only 93.3%-94.3% similarity to other Arcobacter species. These data indicate that abalones carry Arcobacter as a common bacterial genus which includes uncultured species.

Matched MeSH terms: Sequence Analysis, DNA
Characterization and functional analysis of slc7a11 gene, involved in skin color differentiation in the red tilapia

Wang LM, Bu HY, Song FB, Zhu WB, Fu JJ, Dong ZJ

Comp Biochem Physiol A Mol Integr Physiol, 2019 10;236:110529.
PMID: 31310814 DOI: 10.1016/j.cbpa.2019.110529

Red tilapia has become more popular for aquaculture production in China in recent years. However, the pigmentation differentiation that has resulted from the process of genetic breeding and skin color variation during the overwintering period are the main problems limiting the development of commercial culture. The genetic basis of skin color differentiation is still not understood. Solute carrier family 7 member 11 (slc7a11) has been identified to be a critical genetic regulator of pheomelanin synthesis in the skin of mammals. However, little information is available about its molecular characteristics, expression, location and function in skin color differentiation of fish. In this study, three complete cDNA sequences (2159 bp, 2190 bp and 2249 bp) of slc7a11 were successfully isolated from Malaysian red tilapia, encoding polypeptides of 492, 525 and 492 amino acids respectively. Quantitative real-time PCR demonstrated that slc7a11 mRNA expression is high in the ventral skin of PR (pink with scattered red spots) fish. Immunofluorescence analysis revealed that xCT (the protein encoded by slc7a11) was concentrated mainly in the cytoplasm and nucleus of both the dorsal and ventral skin cells of fish. After RNA interference of slc7a11, slc7a11 and cbs mRNA expressions decreased, but the tyr mRNA expression increased in the skin of fish. Results suggest that slc7a11 plays an important role in skin color formation and differentiation of red tilapia through the melanogenesis pathway.

Matched MeSH terms: Sequence Analysis, DNA
Fulltext Blau Syndrome Associated with Nucleotide-binding Oligomerization Domain Containing 2 Mutation in a Baby from Malaysia

Leong KF, Sato R, Oh GGK, Surana U, Pramono ZAD

Indian J Dermatol, 2019 9 24;64(5):400-403.
PMID: 31543536 DOI: 10.4103/ijd.IJD_44_18

Blau syndrome (BS) is a very rare autosomal dominant juvenile inflammatory disorder caused by mutation in nucleotide-binding oligomerization domain containing 2 (NOD2). Usually, dermatitis is the first symptom that appears in the 1st year of life. About 220 BS cases with confirmed NOD2 mutation have been reported. However, the rarity and lack of awareness of the disease, especially in the regions where genetic tests are very limited, often result in late diagnosis and misdiagnosis. Here, we report a de novo BS case from Malaysia, which may be the first report from southeast Asia. PCR and DNA sequencing of peripheral blood mononuclear cells were performed to screen the entire coding region of NOD2 gene. A heterozygous c.1000C>T transition in exon 4, p. R334W, of the NOD2 gene was identified in the patient. This report further reaffirms the ubiquitousness of the disease and recurrency of p. R334W mutation.

Matched MeSH terms: Sequence Analysis, DNA
Evaluation of mitogenome sequence concordance, heteroplasmy detection, and haplogrouping in a worldwide lineage study using the Precision ID mtDNA Whole Genome Panel

Strobl C, Churchill Cihlar J, Lagacé R, Wootton S, Roth C, Huber N, et al.

Forensic Sci Int Genet, 2019 09;42:244-251.
PMID: 31382159 DOI: 10.1016/j.fsigen.2019.07.013

The emergence of Massively Parallel Sequencing technologies enabled the analysis of full mitochondrial (mt)DNA sequences from forensically relevant samples that have, so far, only been typed in the control region or its hypervariable segments. In this study, we evaluated the performance of a commercially available multiplex-PCR-based assay, the Precision ID mtDNA Whole Genome Panel (Thermo Fisher Scientific), for the amplification and sequencing of the entire mitochondrial genome (mitogenome) from even degraded forensic specimens. For this purpose, more than 500 samples from 24 different populations were selected to cover the vast majority of established superhaplogroups. These are known to harbor different signature sequence motifs corresponding to their phylogenetic background that could have an effect on primer binding and, thus, could limit a broad application of this molecular genetic tool. The selected samples derived from various forensically relevant tissue sources and were DNA extracted using different methods. We evaluated sequence concordance and heteroplasmy detection and compared the findings to conventional Sanger sequencing as well as an orthogonal MPS platform. We discuss advantages and limitations of this approach with respect to forensic genetic workflow and analytical requirements.

Matched MeSH terms: Sequence Analysis, DNA
Genetic characterisation of the erythrocyte-binding protein (PkβII) of Plasmodium knowlesi isolates from Malaysia

Fong MY, Lau YL, Jelip J, Ooi CH, Cheong FW

J Genet, 2019 Sep;98.
PMID: 31544794

Plasmodium knowlesi contributes to the majority of human malaria incidences in Malaysia. Its uncontrollable passage among the natural monkey hosts can potentially lead to zoonotic outbreaks. The merozoite of this parasite invades host erythrocytes through interaction between its erythrocyte-binding proteins (EBPs) and their respective receptor on the erythrocytes. The regionII of P. knowlesi EBP, P. knowlesi beta (PkβII) protein is found to be mediating merozoite invasion into monkey erythrocytes by interacting with sialic acid receptors. Hence, the objective of this study was to investigate the genetic diversity, natural selection and haplotype grouping of PkβII of P. knowlesi isolates in Malaysia. Polymerase chain reaction amplifications of PkβII were performed on archived blood samples from Malaysia and 64 PkβII sequences were obtained. Sequence analysis revealed length polymorphism, and its amino acids at critical residues indicate the ability of PkβII to mediate P. knowlesi invasion into monkey erythrocytes. Low genetic diversity (π = 0.007) was observed in the PkβII of Malaysia Borneo compared to Peninsular Malaysia (π = 0.015). The PkβII was found to be under strong purifying selection to retain infectivity in monkeys and it plays a limited role in the zoonotic potential of P. knowlesi. Its haplotypes could be clustered into Peninsular Malaysia and Malaysia Borneo groups, indicating the existence of two distinct P. knowlesi parasites in Malaysia as reported in an earlier study.

Matched MeSH terms: Sequence Analysis, DNA
Fulltext High-quality Schistosoma haematobium genome achieved by single-molecule and long-range sequencing

Stroehlein AJ, Korhonen PK, Chong TM, Lim YL, Chan KG, Webster B, et al.

Gigascience, 2019 Sep 01;8(9).
PMID: 31494670 DOI: 10.1093/gigascience/giz108

BACKGROUND: Schistosoma haematobium causes urogenital schistosomiasis, a neglected tropical disease affecting >100 million people worldwide. Chronic infection with this parasitic trematode can lead to urogenital conditions including female genital schistosomiasis and bladder cancer. At the molecular level, little is known about this blood fluke and the pathogenesis of the disease that it causes. To support molecular studies of this carcinogenic worm, we reported a draft genome for S. haematobium in 2012. Although a useful resource, its utility has been somewhat limited by its fragmentation.
FINDINGS: Here, we systematically enhanced the draft genome of S. haematobium using a single-molecule and long-range DNA-sequencing approach. We achieved a major improvement in the accuracy and contiguity of the genome assembly, making it superior or comparable to assemblies for other schistosome species. We transferred curated gene models to this assembly and, using enhanced gene annotation pipelines, inferred a gene set with as many or more complete gene models as those of other well-studied schistosomes. Using conserved, single-copy orthologs, we assessed the phylogenetic position of S. haematobium in relation to other parasitic flatworms for which draft genomes were available.
CONCLUSIONS: We report a substantially enhanced genomic resource that represents a solid foundation for molecular research on S. haematobium and is poised to better underpin population and functional genomic investigations and to accelerate the search for new disease interventions.

Matched MeSH terms: Sequence Analysis, DNA
Dickeya undicola sp. nov., a novel species for pectinolytic isolates from surface waters in Europe and Asia

Oulghazi S, Pédron J, Cigna J, Lau YY, Moumni M, Van Gijsegem F, et al.

Int J Syst Evol Microbiol, 2019 Aug;69(8):2440-2444.
PMID: 31166160 DOI: 10.1099/ijsem.0.003497

Strains 2B12T, FVG1-MFV-O17 and FVG10-MFV-A16 were isolated from fresh water samples collected in Asia and Europe. The nucleotide sequences of the gapA barcodes revealed that all three strains belonged to the same cluster within the genus Dickeya. Using 13 housekeeping genes (fusA, rpoD, rpoS, glyA, purA, groEL, gapA, rplB, leuS, recA, gyrB, infB and secY), multilocus sequence analysis confirmed the existence of a new clade. When the genome sequences of these three isolates and other Dickeya species were compared, the in silico DNA-DNA hybridization and average nucleotide identity values were found to be no more than 45.50 and 91.22 %, respectively. The closest relative species was Dickeya fangzhongdai. Genome comparisons also highlighted genetic traits differentiating the new strains from D. fangzhongdai strains DSM 101947T (=CFBP 8607T) and B16. Phenotypical tests were performed to distinguish the three strains from D. fangzhongdai and other Dickeya species. The name Dickeya undicola sp. nov. is proposed with strain 2B12T (=CFBP 8650T=LMG 30903T) as the type strain.

Matched MeSH terms: Sequence Analysis, DNA
Fulltext Genetic diversity of toxigenic Vibrio cholerae O1 from Sabah, Malaysia 2015

Zaw MT, Emran NA, Ibrahim MY, Suleiman M, Awang Mohd TA, Yusuff AS, et al.

J Microbiol Immunol Infect, 2019 Aug;52(4):563-570.
PMID: 29428381 DOI: 10.1016/j.jmii.2018.01.003

BACKGROUND: Cholera is an important health problem in Sabah, a Malaysian state in northern Borneo; however, Vibrio cholerae in Sabah have never been characterized. Since 2002, serogroup O1 strains having the traits of both classical and El Tor biotype, designated as atypical El Tor biotype, have been increasingly reported as the cause of cholera worldwide. These variants are believed to produce clinically more severe disease like classical strains.
PURPOSE: The purpose of this study is to investigate the genetic diversity of V.cholerae in Sabah and whether V.cholerae in Sabah belong to atypical El Tor biotype.
METHODS: ERIC-PCR, a DNA fingerprinting method for bacterial pathogens based on the enterobacterial repetitive intergenic consensus sequence, was used to study the genetic diversity of 65 clinical V.cholerae O1 isolates from 3 districts (Kudat, Beluran, Sandakan) in Sabah and one environmental isolate from coastal sea water in Kudat district. In addition, we studied the biotype-specific genetic traits in these isolates to establish their biotype.
RESULTS: Different fingerprint patterns were seen in isolates from these three districts but one of the patterns was seen in more than one district. Clinical isolates and environmental isolate have different patterns. In addition, Sabah isolates harbor genetic traits specific to both classical biotype (ctxB-1, rstRCla) and El Tor biotype (rstRET, rstC, tcpAET, rtxC, VC2346).
CONCLUSION: This study revealed that V.cholerae in Sabah were genetically diverse and were atypical El Tor strains. Fingerprint patterns of these isolates will be useful in tracing the origin of this pathogen in the future.

Matched MeSH terms: Sequence Analysis, DNA
Fulltext Microbial Diversity in Decaying Oil Palm Empty Fruit Bunches (OPEFB) and Isolation of Lignin-degrading Bacteria from a Tropical Environment

Tahir AA, Mohd Barnoh NF, Yusof N, Mohd Said NN, Utsumi M, Yen AM, et al.

Microbes Environ, 2019 Jun 27;34(2):161-168.
PMID: 31019143 DOI: 10.1264/jsme2.ME18117

Oil palm empty fruit bunches (OPEFB) are the most abundant, inexpensive, and environmentally friendly lignocellulosic biomass in Malaysia. Investigations on the microbial diversity of decaying OPEFB may reveal microbes with complex enzymes that have the potential to enhance the conversion of lignocellulose into second-generation biofuels as well as the production of other value-added products. In the present study, fungal and bacterial diversities in decaying OPEFB were identified using Illumina MiSeq sequencing of the V3 region of the 16S rRNA gene and V4 region of the 18S rRNA gene. Fungal diversity in decaying OPEFB was dominated by the phylum Ascomycota (14.43%), while most of the bacterial sequences retrieved belonged to Proteobacteria (76.71%). Three bacterial strains isolated from decaying OPEFB, designated as S18, S20, and S36, appeared to grow with extracted OPEFB-lignin and Kraft lignin (KL) as the sole carbon source. 16S rRNA gene sequencing identified the 3 isolates as Paenibacillus sp.. The molecular weight distribution of KL before and after degradation showed significant depolymerization when treated with bacterial strains S18, S20, and S36. The presence of low-molecular-weight lignin-related compounds, such as vanillin and 2-methoxyphenol derivatives, which were detected by a GC-MS analysis, confirmed the KL-degrading activities of isolated Paenibacillus strains.

Matched MeSH terms: Sequence Analysis, DNA
Fulltext Genetic diversity of Plasmodium falciparum histidine-rich protein 2 (PfHRP2) and its effect on the performance of PfHRP2-based rapid diagnostic tests

Mussa A, Talib M, Mohamed Z, Hajissa K

BMC Res Notes, 2019 Jun 11;12(1):334.
PMID: 31186056 DOI: 10.1186/s13104-019-4361-6

OBJECTIVE: Rapid diagnostic tests (RDTs) play a crucial role in the management and control of malaria infection. The histidine-rich protein 2 (PfHRP-2) based RDTs are the most commonly used RDTs for malaria diagnosis in Sudan. Deletion of pfhrp2 in Plasmodium falciparum genome affect the accuracy of PfHRP-2 based RDT kits. This study aimed to identify molecular variation of pfhrp2 among suspected malaria patients from different clinics in Omdurman, Sudan.
RESULTS: A noticeable variation between the RDT (Alltest Biotech, China) and nPCR results was observed, for RDT 78% (46/59) were P. falciparum positive, 6.8% (4/59) were co-infected with both P. falciparum and Plasmodium vivax, 15.3% (9/59) were negative by the RDT. However, when the nPCR was applied only 44.1% (26/59) and 55.9% (33/59) was P. falciparum positive and negative respectively. The pfhrp2 was further amplified form all nPCR positive samples. Only 17 DNA samples were positive from the 26 positive P. falciparum, interestingly, variation in band sizes was observed and further confirmed by DNA sequencing, and sequencing analysis revealed a high-level of genetic diversity of the pfhrp2 gene in the parasite population from the study area. However, despite extreme sequence variation, diversity of PfHRP2 does not appear to affect RDT performance.

Matched MeSH terms: Sequence Analysis, DNA

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