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DNA-RNA complementation on silicon wafer for thyroid cancer determination

Gopinath SCB, Xuan S

Biotechnol Appl Biochem, 2021 Jun;68(3):554-559.
PMID: 32460382 DOI: 10.1002/bab.1961

One of the current issues with thyroid tumor is early diagnosis as it makes the higher possibility of curing. This research was focused to detect and quantify the level of specific target sequence complementation of miR-222 with capture DNA sequence on interdigitated electrode (IDE) sensor. The aluminum electrode with the gap and finger sizes of 10 µm was fabricated on silicon wafer, further the surface was amine-functionalized for accommodating carboxylated-DNA probe. With DNA-target RNA complementation, the detection limit was attained to be 1 fM as estimated by a linear regression analysis [y = 1.5325x - 2.1171 R² = 0.9065] and the sensitivity was at the similar level. Current responses were higher by increasing the target RNA sequence concentrations. Control experiments with mismatched/noncomplementary sequences were failed to complement the capture DNA sequence immobilized on IDE, indicating the specific target validation. This research helps diagnosing and identifying the progression with thyroid tumor and miRNA being a potential "marker" in atypia diagnosis.

Matched MeSH terms: DNA/genetics*; Thyroid Neoplasms/genetics; Biomarkers, Tumor/genetics; MicroRNAs/genetics*
Fulltext Recovery of recombinant Mycobacterium tuberculosis antigens fused with cell wall-anchoring motif (LysM) from inclusion bodies using non-denaturing reagent (N-laurylsarcosine)

Mustafa AD, Kalyanasundram J, Sabidi S, Song AA, Abdullah M, Abdul Rahim R, et al.

BMC Biotechnol, 2019 05 14;19(1):27.
PMID: 31088425 DOI: 10.1186/s12896-019-0522-x

BACKGROUND: The current limitations of conventional BCG vaccines highlights the importance in developing novel and effective vaccines against tuberculosis (TB). The utilization of probiotics such as Lactobacillus plantarum for the delivery of TB antigens through in-trans surface display provides an effective and safe vaccine approach against TB. Such non-recombinant probiotic surface display strategy involves the fusion of candidate proteins with cell wall binding domain such as LysM, which enables the fusion protein to anchor the L. plantarum cell wall externally, without the need for vector genetic modification. This approach requires sufficient production of these recombinant fusion proteins in cell factory such as Escherichia coli which has been shown to be effective in heterologous protein production for decades. However, overexpression in E. coli expression system resulted in limited amount of soluble heterologous TB-LysM fusion protein, since most of it are accumulated as insoluble aggregates in inclusion bodies (IBs). Conventional methods of denaturation and renaturation for solubilizing IBs are costly, time-consuming and tedious. Thus, in this study, an alternative method for TB antigen-LysM protein solubilization from IBs based on the use of non-denaturating reagent N-lauroylsarcosine (NLS) was investigated.
RESULTS: Expression of TB antigen-LysM fusion genes was conducted in Escherichia coli, but this resulted in IBs deposition in contrast to the expression of TB antigens only. This suggested that LysM fusion significantly altered solubility of the TB antigens produced in E. coli. The non-denaturing NLS technique was used and optimized to successfully solubilize and purify ~ 55% of the recombinant cell wall-anchoring TB antigen from the IBs. Functionality of the recovered protein was analyzed via immunofluorescence microscopy and whole cell ELISA which showed successful and stable cell wall binding to L. plantarum (up to 5 days).
CONCLUSION: The presented NLS purification strategy enables an efficient and rapid method for obtaining higher yields of soluble cell wall-anchoring Mycobacterium tuberculosis antigens-LysM fusion proteins from IBs in E. coli.

Matched MeSH terms: Antigens, Bacterial/genetics; Mycobacterium tuberculosis/genetics; Recombinant Fusion Proteins/genetics; Tuberculosis Vaccines/genetics
Fulltext The Use of Genetic and Gene Technologies in Shaping Modern Rapeseed Cultivars (Brassica napus L.)

Ton LB, Neik TX, Batley J

Genes (Basel), 2020 09 30;11(10).
PMID: 33008008 DOI: 10.3390/genes11101161

Since their domestication, Brassica oilseed species have undergone progressive transformation allied with the development of breeding and molecular technologies. The canola (Brassica napus) crop has rapidly expanded globally in the last 30 years with intensive innovations in canola varieties, providing for a wider range of markets apart from the food industry. The breeding efforts of B. napus, the main source of canola oil and canola meal, have been mainly focused on improving seed yield, oil quality, and meal quality along with disease resistance, abiotic stress tolerance, and herbicide resistance. The revolution in genetics and gene technologies, including genetic mapping, molecular markers, genomic tools, and gene technology, especially gene editing tools, has allowed an understanding of the complex genetic makeup and gene functions in the major bioprocesses of the Brassicales, especially Brassica oil crops. Here, we provide an overview on the contributions of these technologies in improving the major traits of B. napus and discuss their potential use to accomplish new improvement targets.

Matched MeSH terms: Plant Diseases/genetics; Crops, Agricultural/genetics*; Brassica napus/genetics*; Disease Resistance/genetics
Population genetic variation of SLC6A4 gene, associated with neurophysiological development

Hande SH, Krishna SM, Sahote KK, Dev N, Erl TP, Ramakrishna K, et al.

J Genet, 2021;100.
PMID: 33764333

The serotonin transporter 5-HTT is encoded by a single gene SLC6A4. Polymorphisms in SLC6A4 has been associated with a wide variety of neurological and psychiatric disorders including increased risk of posttraumatic stress disorder, higher likelihood for depression, obsessive-compulsive disorder (OCD), increased hostility and criminal behaviour. Genes associated with complex diseases often exhibit strong signatures of purifying selection compared to others. Further, discernible population specific variation in the signature of natural selection have been observed for several complex disease-related genes. In this project we aimed to investigate the population genetic variation of the serotonin transporter gene (SLC6A4), focussing on the single nucleotide polymorphisms (SNPs). To this end, we employed 2504 individuals around the globe available in 1000 Genome project Phase III data and classified them into five ethnic groups: Americans (AMR), Europeans (EUR), Africans (AFR), East Asians (EAS) and South Asians (SAS). Principal component analysis (PCA) performed on all annotated SNPs of SLC6A4 depicted clear clustering between Africans and the rest of the world along PC1, and East Asians and other non-African populations along PC2. Further, these SNPs were found to be under strong selection pressure especially among East Asian populations with significantly high positive cross-population extended haplotype homozygosity scores compared to Africans, indicating that SLC6A4 has likely undergone a strong selective sweep among the East Asians in the recent past. Our study can potentially explain the association between polymorphisms in SLC6A4, and major depression and suicidal tendencies among people of East Asian ancestry and the absence of such associations among people of European ancestry.

Matched MeSH terms: Ethnic Groups/genetics*; Genetics, Population*; Serotonin Plasma Membrane Transport Proteins/genetics*
Fulltext Genetic relatedness and novel sequence types of clinical Aeromonas dhakensis from Malaysia

Lau TTV, Tan JMA, Puthucheary SD, Puah SM, Chua KH

Braz J Microbiol, 2020 Sep;51(3):909-918.
PMID: 32067209 DOI: 10.1007/s42770-020-00239-8

Aeromonas dhakensis is an emergent human pathogen with medical importance. This study was aimed to determine the sequence types (STs), genetic diversity, and phylogenetic relationships of different clinical sources of 47 A. dhakensis from Malaysia using multilocus sequence typing (MLST), goeBURST, and phylogenetic analyses. The analysis of a concatenated six-gene tree with a nucleotide length of 2994 bp based on six housekeeping genes (gyrB, groL, gltA, metG, ppsA, and recA) and independent analyses of single gene fragments was performed. MLST was able to group 47 A. dhakensis from our collection into 36 STs in which 34 STs are novel STs. The most abundant ST521 consisted of five strains from peritoneal fluid and two strains from stools. Comparison of 62 global A. dhakensis was carried out via goeBURST; 94.4% (34/36) of the identified STs are novel and unique in Malaysia. Two STs (111 and 541) were grouped into clonal complexes among our strains and 32 STs occurred as singletons. Single-gene phylogenetic trees showed varying topologies; groL and rpoD grouped all A. dhakensis into a tight-cluster with bootstrap values of 100% and 99%, respectively. A poor phylogenetic resolution encountered in single-gene analyses was buffered by the multilocus phylogenetic tree that offered high discriminatory power (bootstrap value = 100%) in resolving all A. dhakensis from A. hydrophila and delineating the relationship among other taxa. Genetic diversity analysis showed groL as the most conserved gene and ppsA as the most variable gene. This study revealed novel STs and high genetic diversity among clinical A. dhakensis from Malaysia.

Matched MeSH terms: Aeromonas/genetics*; DNA, Bacterial/genetics; Genes, Bacterial/genetics; Genes, Essential/genetics
Fulltext Over-expression of Nicotinamide phosphoribosyltransferase in mouse cells confers protective effect against oxidative and ER stress-induced premature senescence

Nuriliani A, Nakahata Y, Ahmed R, Khaidizar FD, Matsui T, Bessho Y

Genes Cells, 2020 Aug;25(8):593-602.
PMID: 32533606 DOI: 10.1111/gtc.12794

A main feature of aged organisms is the accumulation of senescent cells. Accumulated senescent cells, especially stress-induced premature senescent cells, in aged organisms lead to the decline of the regenerative potential and function of tissues. We recently reported that the over-expression of NAMPT, which is the rate-limiting enzyme in mammalian NAD+ salvage pathway, delays replicative senescence in vitro. However, whether Nampt-overexpressing cells are tolerant of stress-induced premature senescence remains unknown. Here, we show that primary mouse embryonic fibroblasts derived from Nampt-overexpressing transgenic mice (Nampt Tg-MEF cells) possess resistance against stress-induced premature senescence in vitro. We found that higher oxidative or endoplasmic reticulum (ER) stress is required to induce premature senescence in Nampt Tg-MEF cells compared to wild-type cells. Moreover, we found that Nampt Tg-MEF cells show acute expression of unfolded protein response (UPR)-related genes, which in turn would have helped to restore proteostasis and avoid cellular senescence. Our results demonstrate that NAMPT/NAD+ axis functions to protect cells not only from replicative senescence, but also from stress-induced premature senescence in vitro. We anticipate that in vivo activation of NAMPT activity or increment of NAD+ would protect tissues from the accumulation of premature senescent cells, thereby maintaining healthy aging.

Matched MeSH terms: Gene Expression Regulation/genetics; Gene Expression/genetics; Cell Aging/genetics; Nicotinamide Phosphoribosyltransferase/genetics*
Development of a species-specific PCR-RFLP targeting rpoD gene fragment for discrimination of Aeromonas species

Puah SM, Khor WC, Kee BP, Tan JAMA, Puthucheary SD, Chua KH

J Med Microbiol, 2018 Sep;67(9):1271-1278.
PMID: 30024365 DOI: 10.1099/jmm.0.000796

PURPOSE: The taxonomy of Aeromonas keeps expanding and their identification remains problematic due to their phenotypic and genotypic heterogeneity. In this study, we aimed to develop a rapid and reliable polymerase chain reaction-restriction fragment length polymorphism assay targeting the rpoD gene to enable the differentiation of aeromonads into 27 distinct species using microfluidic capillary electrophoresis.
METHODOLOGY: A pair of degenerate primers (Aero F: 5'-YGARATCGAYATCGCCAARCGB-3' and Aero R: 5'-GRCCDATGCTCATRCGRCGGTT-3') was designed that amplified the rpoD gene of 27 Aeromonas species. Subsequently, in silico analysis enabled the differentiation of 25 species using the single restriction endonuclease AluI, while 2 species, A. sanarelli and A. taiwanensis, required an additional restriction endonuclease, HpyCH4IV. Twelve type strains (A. hydrophila ATCC7966T, A. caviae ATCC15468T, A. veronii ATCC9071T, A. media DSM4881T, A. allosaccharophila DSM11576T, A. dhakensis DSM17689T, A. enteropelogens DSM7312T, A. jandaei DSM7311T, A. rivuli DSM22539T, A. salmonicida ATCC33658T, A. taiwanensis DSM24096T and A. sanarelli DSM24094T) were randomly selected from the 27 Aeromonas species for experimental validation.Results/key findings. The twelve type strains demonstrated distinctive RFLP patterns and supported the in silico digestion. Subsequently, 60 clinical and environmental strains from our collection, comprising nine Aeromonas species, were used for screening examinations, and the results were in agreement.
CONCLUSION: This method provides an alternative method for laboratory identification, surveillance and epidemiological investigations of clinical and environmental specimens.

Matched MeSH terms: Aeromonas/genetics*; Bacterial Proteins/genetics*; Sigma Factor/genetics*; DNA Primers/genetics
Simple and rapid detection of Salmonella strains by direct PCR amplification of the hilA gene

Pathmanathan SG, Cardona-Castro N, Sánchez-Jiménez MM, Correa-Ochoa MM, Puthucheary SD, Thong KL

J Med Microbiol, 2003 Sep;52(Pt 9):773-6.
PMID: 12909653

The suitability of a PCR procedure using a pair of primers targeting the hilA gene was evaluated as a means of detecting Salmonella species. A total of 33 Salmonella strains from 27 serovars and 15 non-Salmonella strains from eight different genera were included. PCR with all the Salmonella strains produced a 784 bp DNA fragment that was absent from all the non-Salmonella strains tested. The detection limit of the PCR was 100 pg with genomic DNA and 3 x 10(4) c.f.u. ml(-1) with serial dilutions of bacterial culture. An enrichment-PCR method was further developed to test the sensitivity of the hilA primers for the detection of Salmonella in faecal samples spiked with different concentrations of Salmonella choleraesuis subsp. choleraesuis serovar Typhimurium. The method described allowed the detection of Salmonella Typhimurium in faecal samples at a concentration of 3 x 10(2) c.f.u. ml(-1). In conclusion, the hilA primers are specific for Salmonella species and the PCR method presented may be suitable for the detection of Salmonella in faeces.

Matched MeSH terms: Bacterial Proteins/genetics; Salmonella/genetics; Salmonella typhimurium/genetics; Trans-Activators/genetics*
Fulltext DNA typing of Calliphorids collected from human corpses in Malaysia

Kavitha R, Tan TC, Lee HL, Nazni WA, Sofian-Azirun M

Trop Biomed, 2013 Mar;30(1):119-24.
PMID: 23665717 MyJurnal

Estimation of post-mortem interval (PMI) is crucial for time of death determination. The advent of DNA-based identification techniques forensic entomology saw the beginning of a proliferation of molecular studies into forensically important Calliphoridae (Diptera). The use of DNA to characterise morphologically indistinguishable immature calliphorids was recognised as a valuable molecular tool with enormous practical utility. The local entomofauna in most cases is important for the examination of entomological evidences. The survey of the local entomofauna has become a fundamental first step in forensic entomological studies, because different geographical distributions, seasonal and environmental factors may influence the decomposition process and the occurrence of different insect species on corpses. In this study, calliphorids were collected from 13 human corpses recovered from indoors, outdoors and aquatic conditions during the post-mortem examination by pathologists from the government hospitals in Malaysia. Only two species, Chrysomya megacephala and Chrysomya rufifacies were recovered from human corpses. DNA sequencing was performed to study the mitochondrial encoded COI gene and to evaluate the suitability of the 1300 base pairs of COI fragments for identification of blow fly species collected from real crime scene. The COI gene from blow fly specimens were sequenced and deposited in GenBank to expand local databases. The sequenced COI gene was useful in identifying calliphorids retrieved from human corpses.

Matched MeSH terms: Electron Transport Complex IV/genetics; Diptera/genetics; DNA, Mitochondrial/genetics; Protein Subunits/genetics
Fulltext Simulium (Nevermannia) chomthongense, a new species of black fly (Diptera: Simuliidae) from Chiang Mai, Thailand

Takaoka H, Srisuka W, Saeung A, Otsuka Y, Choochote W

Trop Biomed, 2012 Sep;29(3):381-90.
PMID: 23018501

Simulium (Nevermannia) chomthongense sp. nov. is described from female, male, pupal and larval specimens collected from Doi Inthanon National Park and Doi Phahompok National Park, Chiang Mai, Thailand. This new species, first reported as S. (Eusimulium) sp. A, and later regarded as S. (N.) caudisclerum Takaoka & Davies, described from peninsular Malaysia, is distinguished from S. (N.) caudisclerum in the male by the number of enlarged upper-eye facets and the relative size of the hind basitarsus against the hind tibia and femur, and in the pupa by the relative length of the stalks of paired filaments against the common basal stalk and the color of the dorsal surface of abdominal segments 1- 3 (or 4). Taxonomic and molecular notes are provided to separate this new species from four other known species of the vernum species-group, which share an accessory sclerite on the larval abdomen, a rare characteristic in this species-group.

Matched MeSH terms: DNA, Mitochondrial/genetics; DNA, Ribosomal/genetics; RNA, Ribosomal, 16S/genetics; Simuliidae/genetics
Fulltext Sequence analysis on the mitochondrial COXI gene of recent clinical isolates of Plasmodium knowlesi in Klang Valley, peninsular Malaysia

Fong MY, Lau YL, Chin LC, Al-Mekhlafi AM

Trop Biomed, 2011 Aug;28(2):457-63.
PMID: 22041769

The cytochrome oxidase subunit I (COXI) gene sequences of three recent (2007-2008) clinical Plasmodium knowlesi isolates from Klang Valley, peninsular Malaysia, were determined and compared with those of older (1960's) peninsular Malaysia, recent isolates from Sarawak (on Borneo Island), and an isolate from Thailand. Multiple alignment of the sequences showed that the three clinical isolates were more similar to the older peninsular Malaysia isolates than to those from Sarawak and Thailand. Phylogenetic tree based on the COXI sequences revealed three distinct clusters of P. knowlesi. The first cluster consisted of isolates from peninsular Malaysia, the second consisted of Sarawak isolates and the third composed of the Thailand isolate. The findings of this study highlight the usefulness of mitochondrial COXI gene as a suitable marker for phylogeographic studies of P. knowlesi.

Matched MeSH terms: Electron Transport Complex IV/genetics*; Mitochondria/genetics*; DNA, Protozoan/genetics; Plasmodium knowlesi/genetics*
Fulltext Characterization of vancomycin-resistant Enterococcus isolates from broilers in Selangor, Malaysia

Getachew YM, Hassan L, Zakaria Z, Saleha AA, Kamaruddin MI, Che Zalina MZ

Trop Biomed, 2009 Dec;26(3):280-8.
PMID: 20237442 MyJurnal

Vancomycin-resistant Enterococcus (VRE) is an emerging nosocomial pathogen in humans. The use of antibiotics in human therapy and in the production of food animals has been incriminated in the emergence of this organism. The present study describes the distribution of VRE species, the vancomycin-resistant genes detected, the vancomycin resistance pattern observed, and the genetic diversity of the isolates found in live broiler chickens in Malaysia. Overall 140 VRE were isolated with species comprising Enterococcus faecalis (48%), Enterococcus faecium (25.7%), Enterococcus gallinarum (12.1%), Enterococcus casseliflavus (1.4%) and other Enterococcus species (12.8%). Vancomycin resistance gene vanA and intrinsic genes vanC1 and vanC2/3 were detected in the study population. VanA was detected in 15 (63.9%) of E. faecium, 23 (22.4%) of E. faecalis and in 3 (17.6%) E. gallinarum isolates. E-test was conducted on randomly selected 41 of the isolates and the minimum inhibition concentration (MIC) of vancomycin for five (11.9%) of tested isolates is more than 256 μg/ml. Genotypic analysis using random amplified polymorphic DNA (RAPD) showed genetic diversity within the Enterococcus species.

Matched MeSH terms: Bacterial Proteins/genetics; Enterococcus/genetics*; Carbon-Oxygen Ligases/genetics; Vancomycin Resistance/genetics*
Fulltext Overexpression of WNT2 and TSG101 genes in colorectal carcinoma

Ma XR, Edmund Sim UH, Pauline B, Patricia L, Rahman J

Trop Biomed, 2008 Apr;25(1):46-57.
PMID: 18600204 MyJurnal

Colorectal carcinoma (CRC) arises as a result of mutational activation of oncogenes coupled with inactivation of tumour suppressor genes. Mutations in APC, K-ras and p53 have been commonly reported. In a previous study by our group, the tumour susceptibility gene 101 (TSG101) were found to be persistently upregulated in CRC cases. TSG101 was reported to be closely related to cancers of the breast, brain and colon, and its overexpression in human papillary thyroid carcinomas and ovarian carcinomas had previously been reported. The wingless-type MMTV integration site family member 2 (WNT2) is potentially important in the Wnt/beta-catenin pathway and upregulation of WNT2 is not uncommon in human cancers. In this study, we report the investigation for mutation(s) and expression pattern(s) of WNT2 and TSG101, in an effort to further understand their role(s) in CRC tumourigenesis. Our results revealed no mutation in these genes, despite their persistent upregulation in CRC cases studied.

Matched MeSH terms: DNA-Binding Proteins/genetics*; Transcription Factors/genetics*; Colorectal Neoplasms/genetics*; Wnt2 Protein/genetics*
Fulltext Partial characterization of genes encoding the ATP-binding cassette proteins of Cryptosporidium parvum

Li LC, Mun YF

Trop Biomed, 2005 Dec;22(2):115-22.
PMID: 16883276

The present study aims to explore the possible mechanisms underlying the multidrug resistance characteristic of Cryptosporidium parvum by detecting the presence of ATP-binding cassette (ABC) protein encoding genes, especially one that shows high similarity to members belonging to the multidrug resistance protein (MDR) and multidrug resistance associated protein (MRP) subfamilies. PCR using ABC-specific degenerate primers successfully amplified two unique fragments, designated Cpnbd1 and Cpnbd2, from C. parvum genomic DNA. Cpnbd1 exhibited high degree of homology (99-100%) with the nucleotide- binding domains (NBDs) at the NH2 -terminal halves of two previously reported ABC proteins (CpABC and CpABC1) of human and bovine origin C. parvum isolates. It is likely that CpABC, CpABC1 and Cpnbd1 were encoded by homologous genes of a type of ABC transporter protein found in different C. parvum isolates. However, Cpnbd2 showed moderate levels of similarities (28-49%) to the NBDs of four ABC proteins characterised in C. parvum to date. Therefore, Cpnbd2 could be a novel member of an ABC superfamily of proteins in C. parvum. Phylogenetic analyses on a list of ABC transporters known to associate with MDR phenotype has significantly linked Cpnbd1 and Cpnbd2 to these transporters, thus suggesting that Cpnbd1 and Cpnbd2 proteins may contribute to the intrinsic multidrug resistance phenotype of C. parvum.

Matched MeSH terms: Protozoan Proteins/genetics*; Drug Resistance, Multiple/genetics*; ATP-Binding Cassette Transporters/genetics*; Multidrug Resistance-Associated Proteins/genetics*
Fulltext Phylogenetic analyses uncover a novel clade of transferrin in nonmammalian vertebrates

Mohd-Padil H, Mohd-Adnan A, Gabaldón T

Mol Biol Evol, 2013 Apr;30(4):894-905.
PMID: 23258311 DOI: 10.1093/molbev/mss325

Transferrin is a protein super-family involved in iron transport, a central process in cellular homeostasis. Throughout the evolution of vertebrates, transferrin members have diversified into distinct subfamilies including serotransferrin, ovotransferrin, lactoferrin, melanotransferrin, the inhibitor of carbonic anhydrase, pacifastin, and the major yolk protein in sea urchin. Previous phylogenetic analyses have established the branching order of the diverse transferrin subfamilies but were mostly focused on the transferrin repertoire present in mammals. Here, we conduct a comprehensive phylogenetic analysis of transferrin protein sequences in sequenced vertebrates, placing a special focus on the less-studied nonmammalian vertebrates. Our analyses uncover a novel transferrin clade present across fish, sauropsid, and amphibian genomes but strikingly absent from mammals. Our reconstructed scenario implies that this novel class emerged through a duplication event at the vertebrate ancestor, and that it was subsequently lost in the lineage leading to mammals. We detect footprints of accelerated evolution following the duplication event, which suggest positive selection and early functional divergence of this novel clade. Interestingly, the loss of this novel class of transferrin in mammals coincided with the divergence by duplication of lactoferrin and serotransferrin in this lineage. Altogether, our results provide novel insights on the evolution of iron-binding proteins in the various vertebrate groups.

Matched MeSH terms: Bass/genetics*; Transferrin/genetics*; Vertebrates/genetics; Fish Proteins/genetics*
Antifungal Mechanism of Rhodotorula mucilaginosa and Aureobasidium sp. nov. Isolated from Cerbera manghas L. against the Growth of Destructive Molds in Post Harvested Apples

Sukmawati D, Shabrina A, Indrayanti R, Kurniati TH, Nurjayadi M, Hidayat I, et al.

Recent Pat Food Nutr Agric, 2020;11(3):219-228.
PMID: 32324527 DOI: 10.2174/2212798411666200423101159

BACKGROUND: Apples often experience postharvest damage due to being attacked by mold organisms. Several groups of molds such as Aspergillus sp., Penicilium expansum, Botrytis cinerea, and Venturia sp. can cause a serious postharvest disease exhibited as watery regions where areas of blue-green tufts of spores develop. Current methods using fungicides to control pathogenic fungi can cause resistance if applied in the long term. An alternative procedure using yeast as a biological agent has been found.
OBJECTIVE: The aim of this study is to screen potential yeast, which has the ability to inhibit the growth of Aspergillus brasielensis (isolate A1) and Aspergillus flavus section flavi (isolate A17) isolated from apple fruits.
METHODS: Antagonism test using YMA dual culture medium using in vitro assays and ITS rDNA identification were performed.
RESULTS: The result showed that 3 out of 19 yeast isolated from Cerbera manghas L, T1, T3 and T4, demonstrated the potential ability as a biocontrol agent. ITS rDNA identification demonstrated that T1 has a similarity to Rhodotorula mucilaginosa while T3 and T4 were identified as Aureobasidium sp. nov. The 3 isolates exhibited the ability to reduce the growth of A. brasiliensis sensu lato better than dithane 0.3% with a Disease Incidence (DI) of 100% and a Disease Severity (DS) value of 45%. Only isolate T1 and T3 were able to reduce decay symptoms in apples inoculated with A. flavus sensu lato (with DO and DS were 100% and 25%, respectively) compared to dithane pesticides 0.3%.
CONCLUSION: This study indicated that competition between nutrients occurs between pathogenic molds and under-yeast in vitro and in vivo conditions. However, further studies in the future might be able to elucidate the 'killer' activity and interaction with the pathogen cells and the bio-product production using Rhodotorula mucilaginosa and Aureoubasidium namibiae strains to control postharvest diseases.

Matched MeSH terms: Aspergillus/genetics; Aspergillus flavus/genetics; DNA, Ribosomal/genetics; Rhodotorula/genetics
Identification patterns of Trichoderma strains using morphological characteristics, phylogenetic analyses and lignocellulolytic activities

Asis A, Shahriar SA, Naher L, Saallah S, Fatihah HNN, Kumar V, et al.

Mol Biol Rep, 2021 Apr;48(4):3285-3301.
PMID: 33880673 DOI: 10.1007/s11033-021-06321-0

Trichoderma is a genus of soil-borne fungus with an abundance of reports of its economic importance in the agriculture industry. Thus, the correct identification of Trichoderma species is necessary for its commercial purposes. Globally, Trichoderma species are routinely identified from micro-morphological descriptions which can be tedious and prone to errors. Thus, we emphasize that the accurate identification of Trichoderma strains requires a three-pronged approach i.e. based on its morphological characteristics, multilocus gene sequences of the rDNA [internal transcribed spacer (ITS) 1 and 2 regions], translation elongation factor 1-α (TEF-1α), Calmodulin (CAL) and its lignocellulolytic activities. We used this approach to identify a total of 53 Trichoderma strains which were isolated from a wet paddy field located at Tuaran, Sabah, Malaysia. The 53 strains were positively identified as belonging to three Trichoderma species, namely T. asperellum (43 strains), T. harzianum (9 strains), and T. reesei (one strain) on the basis of its morphological characteristics and multilocus gene sequences. Phylogenetic trees constructed based on the UPGMA method of the ITS 1 and 2 regions of the rDNA, TEF-1α and CAL revealed three distinct groups with the T. asperellum, T. harzianum and T. reesei strains placed under the section of Trichoderma, Pachybasium and Longibrachiatum, respectively. In addition, the lignocellulolytic activities of the isolates were measured based on the diameters of the halo zones produced when degrading cellulose, lignin, and starch, respectively. This diagnostic assay can be used to identify Trichoderma as it produces polyphenol oxidase when Tannic Acid Media is used for the lignin test, endoglucanases when Jensen media is used for cellulose, and it hydrolyzes starch to glucose when the modified Melin-Nokrans media is used for the starch test. Accurate identification of Trichoderma species is needed as these strains can potentially be used as a biocontrol agent to prevent diseases and to increase yield in agriculture crops.

Matched MeSH terms: Cellulase/genetics; Catechol Oxidase/genetics; DNA, Ribosomal/genetics; Trichoderma/genetics
Fulltext Solar rhythm in the regulation of photoperiodic flowering of long-day and short-day plants

Yeang HY

J Exp Bot, 2013 Jul;64(10):2643-52.
PMID: 23645867 DOI: 10.1093/jxb/ert130

In photoperiodic flowering, long-day (LD) plants are induced to flower seasonally when the daylight hours are long, whereas flowering in short-day (SD) plants is promoted under short photoperiods. According to the widely accepted external coincidence model, flowering occurs in LD Arabidopsis when the circadian rhythm of the gene CONSTANS (CO) peaks in the afternoon, when it is light during long days but dark when the days are short. Nevertheless, extending this explanation to SD flowering in rice, Oriza sativa, requires LD and SD plants to have 'opposite light requirements' as the CO orthologue in rice, HEADING-DATE1 (Hd1), promotes flowering only under short photoperiods. This report proposes a role of the plant's solar rhythm in promoting seasonal flowering. The interaction between rhythmic genes entrained to the solar clock and those entrained to the circadian clock form the basis of an internal coincidence model that explains both LD and SD flowering equally well. The model invokes no presumption of opposite light requirements between LD and SD plants, and further argues against any specific requirement of either light or darkness for SD flowering. Internal coincidence predicts the inhibition of SD flowering of the rice plant by a night break (a brief interruption of light), while it also provides a plausible explanation for how a judiciously timed night break promotes Arabidopsis flowering even on short days. It is the timing of the light transitions (sunrise and sunset) rather than the duration of light or darkness per se that regulates photoperiod-controlled flowering.

Matched MeSH terms: Plant Proteins/genetics; Oryza/genetics; Arabidopsis/genetics; Flowers/genetics
Fulltext Progressive myoclonus epilepsies-Residual unsolved cases have marked genetic heterogeneity including dolichol-dependent protein glycosylation pathway genes

Courage C, Oliver KL, Park EJ, Cameron JM, Grabińska KA, Muona M, et al.

Am J Hum Genet, 2021 04 01;108(4):722-738.
PMID: 33798445 DOI: 10.1016/j.ajhg.2021.03.013

Progressive myoclonus epilepsies (PMEs) comprise a group of clinically and genetically heterogeneous rare diseases. Over 70% of PME cases can now be molecularly solved. Known PME genes encode a variety of proteins, many involved in lysosomal and endosomal function. We performed whole-exome sequencing (WES) in 84 (78 unrelated) unsolved PME-affected individuals, with or without additional family members, to discover novel causes. We identified likely disease-causing variants in 24 out of 78 (31%) unrelated individuals, despite previous genetic analyses. The diagnostic yield was significantly higher for individuals studied as trios or families (14/28) versus singletons (10/50) (OR = 3.9, p value = 0.01, Fisher's exact test). The 24 likely solved cases of PME involved 18 genes. First, we found and functionally validated five heterozygous variants in NUS1 and DHDDS and a homozygous variant in ALG10, with no previous disease associations. All three genes are involved in dolichol-dependent protein glycosylation, a pathway not previously implicated in PME. Second, we independently validate SEMA6B as a dominant PME gene in two unrelated individuals. Third, in five families, we identified variants in established PME genes; three with intronic or copy-number changes (CLN6, GBA, NEU1) and two very rare causes (ASAH1, CERS1). Fourth, we found a group of genes usually associated with developmental and epileptic encephalopathies, but here, remarkably, presenting as PME, with or without prior developmental delay. Our systematic analysis of these cases suggests that the small residuum of unsolved cases will most likely be a collection of very rare, genetically heterogeneous etiologies.

Matched MeSH terms: Introns/genetics; Mutation/genetics*; Myoclonic Epilepsies, Progressive/genetics*; DNA Copy Number Variations/genetics
Candida vulturna pro tempore sp. nov., a dimorphic yeast species related to the Candida haemulonis species complex isolated from flowers and clinical sample

Sipiczki M, Tap RM

Int J Syst Evol Microbiol, 2016 Oct;66(10):4009-4015.
PMID: 27411802 DOI: 10.1099/ijsem.0.001302

In a taxonomic study of yeasts isolated from flowers in Cagayan de Oro, Mindenao Island, The Philippines, strains were identified as representing Kabatiella microsticta, Metschnikowia koreensis and a hitherto undescribed dimorphic species. Sequences of the D1/D2 domains of the LSU 26S rRNA genes, the internal transcribed spacer (ITS) regions and the SSU 18S rRNA genes were identical in the strains of the last-named group and differed from the corresponding sequences of the type strain of the closest related species, Candida duobushaemulonii, by 4 % (D1/D2), 7 % (ITS) and 1 % (SSU). In an independent study, a strain with D1/D2 and ITS sequences very similar to those of the Philippine strains was isolated in Malaysia from the blood of a patient dying of aspiration pneumonia. Both groups of isolates were moderately sensitive to anidulafungin, caspofungin, fluconazole, itraconazole and voriconazole but resistant to amphotericin B. Molecular phylogenetic analysis of the sequences placed the Philippine and Malaysian isolates close to the Candida haemulonis complex of Candida species. To reflect the geographical location of the sites of sample collection, the novel species name Candida vulturna pro tempore sp. nov. is proposed to accommodate these strains. The type strain is 11-1170T (=CBS 14366T=CCY 094-001-001T=NCAIM-Y02177T) isolated in Cagayan de Oro, The Philippines. Mycobank: MB 817222.

Matched MeSH terms: Candida/genetics; DNA, Fungal/genetics; RNA, Ribosomal/genetics; DNA, Ribosomal Spacer/genetics

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