Roles of ß-Indole Acetic Acid (IAA) Producing Endophytic Bacteria on the Recovery of Plant Growth and Survival Ability of Sugarcane Infected White Leaf Disease (SWLD).

ß-Indole acetic acid is produced in the rhizosphere by endophytic bacteria and promotes plant growth. Effects of bacterial IAA producers (BIPs) on plant growth and recovery of sugarcane seedlings infected with phytoplasma causing white leaf disease (SWLD) were examined. Fifty-five endophytic bacteria isolated from rice roots were collected from the Mekong River Delta, Vietnam. Seven isolates showed ß-Indole acetic acid production in culture medium supplemented with tryptophan. Interestingly, two of them (BC17 and BTII2) produced the highest ß-Indole acetic acid after 4 days of culture. Based on 16S rRNA sequences and phylogenetic analysis, the BC17 and BTII2 isolates were identified as Delftia lacustris and Rahnella aquatilis, respectively. Plant growth induced by the BC17 and BTII2 isolates showed statistically significant differences in height, root length and fresh weight of rice seedlings compared with non-treatment as the control. Treatment of two bacterial isolates in SWLD infected sugarcane plants also showed differences in height of sugarcane seedlings, while gradual symptoms of exposure decreased plant mortality compared to non-treatment as the control. BIPs were shown to be efficient biofertilizer inoculants that promoted plant growth and also ameliorated damage caused by phytoplasma-associated diseases at the sugarcane seedling stage.

Identification of potential microRNA groups for the diagnosis of hepatocellular carcinoma (HCC) using microarray datasets and bioinformatics tools.

Hepatocellular carcinoma (HCC) is the most common primary liver malignancy and the third cause of cancer-related death worldwide. Potential microRNAs have been reported as biomarkers for early detection of HCC as well as novel molecular targets for HCC treatment. Various tissue expression profiles of miRNAs using three microarray datasets from groups in Asia (2), Europe, America (GSE147892, GSE21362, GSE74618, GSE40744) and multiple bioinformatics tools were integrated to determine the most significant miRNA groups to assist in the diagnosis of HCC. Statistical analyses identified at least 30 miRNAs with 17 up-regulated and 13 down-regulated in HCC-related tumor tissues. All the miRNAs also showed relevance to the hallmarks of cancer such as cell proliferation, invasion, metastasis, angiogenesis, metabolism, epithelial-mesenchymal transition and apoptosis. Expression levels of miRNAs observed in the European group showed up-regulation at 5-37% compared to both Asian and American groups. Interestingly, four miRNAs divided into two groups as miR-182-5p/miR-1269a and miR-199a/miR-422a were the most promising for diagnosis of HCC patients from healthy controls, with AUC values of 0.902 and 0.892, respectively. Results provided evidence of the correlation between potential miRNAs and HCC that could be useful for disease diagnosis based on in-depth analyses of large case numbers and cohort studies.

Unraveling the Host-Selective Toxic Interaction of Cassiicolin with Lipid Membranes and Its Cytotoxicity.

Cassiicolin (Cas), a toxin produced by Corynespora cassiicola, is responsible for Corynespora leaf fall disease in susceptible rubber trees. Currently, the molecular mechanisms of the cytotoxicity of Cas and its host selectivity have not been fully elucidated. Here, we analyzed the binding of Cas1 and Cas2 to membranes consisting of different plant lipids and their membrane disruption activities. Using high-speed atomic force microscopy and confocal microscopy, we reveal that the binding and disruption activities of Cas1 and Cas2 on lipid membranes are strongly dependent on the specific plant lipids. The negative phospholipids, glycerolipids, and sterols are more sensitive to membrane damage caused by Cas1 and Cas2 than neutral phospholipids and betaine lipids. Mature Cas1 and Cas2 play an essential role in causing membrane disruption. Cytotoxicity tests on rubber leaves of Rubber Research Institute of Vietnam (RRIV) 1, RRIV 4, and Prang Besar (PB) 255 clones suggest that the toxins cause necrosis of rubber leaves, except for the strong resistance of PB 255 against Cas2. Cryogenic scanning electron microscopy analyses of necrotic leaf tissues treated with Cas1 confirm that cytoplasmic membranes are vulnerable to the toxin. Thus, the host selectivity of Cas toxin is attained by the lipid-dependent binding activity of Cas to the membrane, and the cytotoxicity of Cas arises from its ability to form biofilm-like structures and to disrupt specific membranes.

Loop-mediated isothermal amplification (LAMP) assay for detection of sesame phyllody phytoplasmas in Vietnam.

Phloem-limiting phytoplasmas are known to be causal agents of phyllody, which is recognized by the abnormal development of floral structures resulting in serious yield losses in sesame plants. Currently, identification of the various groups of phytoplasmas that cause sesame phyllody (SP) is conducted by nested PCR, RFLP, and multiplex real-time qPCR assays. However, these methods require intensive labor and are costly and time-consuming so can only be undertaken in well-equipped labs. Here, diagnostic loop-mediated isothermal amplification (LAMP)-based assays allowing rapid detection of specific groups of phytoplasmas within 30 min were developed based on detection of the 16S rRNA sequence of phytoplasmas. Universal 16S rRNA phytoplasma primers and seven primer sets of different 16Sr group phytoplasmas (16SrI, 16SrII, 16SrIII, 16SrIV, 16SrV, 16SrX, 16SrXI) and universal plant cytochrome oxidase (cox) gene primers were used to detect 16S rRNA group phytoplasma sequences and the cox gene in sesame plants. The LAMP assays were carried out using a real-time fluorometer with amplification plots and annealing curves visualized directly. Results demonstrated that the 16SrI and 16SrII group phytoplasmas were causal agents of sesame phyllody in Vietnam. LAMP-based assays for in-field detection of sesame phyllody-causing phytoplasmas revealed advantages and potential applicability in comparison with conventional approaches. To the best of our knowledge, this is the first assessment of multiple phytoplasma infection associated with sesame phyllody disease in Vietnam using LAMP-based assays.

Development of a SCAR marker linked to fungal pathogenicity of rice blast fungus Magnaporthe Oryzae.

PCR-based molecular approaches including RAPD (random amplified polymorphic DNA), ISSR (inter-simple sequence repeat), and SRAP (sequence-related amplified polymorphism) are commonly used to analyze genetic diversity. The aims of this study are to analyze genetic diversity of M. oryzae isolates using PCR-based molecular approaches such as RAPD, ISSR, and SRAP and to develop SCAR marker linked to the pathogenicity of rice blast fungus. Twenty Magnaporthe oryzae isolates were collected mainly from the south of Vietnam and assessed for genetic variation by RAPD, ISSR, and SRAP methods. The comparison of those methods was conducted based on the number of polymorphic bands, percentage of polymorphism, PIC values, and phylogenetic analysis. Then, sequenced characterized amplified region (SCAR) markers were developed based on specific bands linked to fungal pathogenicity of rice blast fungus, M. oryzae. The results indicated that SRAP markers yielded the greatest number of polymorphic bands (174) and occupied 51.7% with polymorphism information content (PIC) value of 0.66. Additionally, the SRAP approach showed stability and high productivity compared with RAPD and ISSR. The SCAR marker developed from the SRAP method identified the presence of the avirulence AVR-pita1 gene involving fungal pathogenicity that can break down blast resistance in rice cultivars. The consistency of SCAR marker obtained in this study showed its efficiency in rapid in-field detection of fungal pathogenicity. SCAR marker developed from SRAP technique provides a useful tool for improving the efficiency of blast disease management in rice fields.

Effects of floral resources on the longevity and parasitism of Cotesia vestalis Haliday (Hymenoptera: Braconidae) on Plutella xylostella (L.) (Lepidoptera: Plutellidae) in Vietnam.

Parasitoids are important biological control of crop pests. In Vietnam, Cotesia vestalis is a native wasp species that has demonstrated its applicability as a natural insect pest enemy. Many adult parasitoids require food resources such as nectar and pollen to optimize their life cycles. Potential effects of yellow cosmos (Cosmos sulphureus), shrub verbena flower (Lantana camara), common zinnia (Zinnia elegans), and coriander flower (Coriandrum sativum L.) on the longevity and parasitism of C. vestalis were investigated. Results showed that nutrition resources from yellow cosmos (C. sulphureus) and shrub verbena flower (Lantana camara) significantly increased the longevity of C. vestalis in comparison with water (P < 0.001). Kaplan-Meier estimates of survival functions of female Cotesia vestalis exposed to different food sources indicated a significant difference in the treatments (P < 0.01). Male and female wasps feeding on Cosmos sulphureus, Lantana camara, and Coriandrum sativum survived significantly longer than wasps fed on water or Zinnia elegans. Parasitism efficiency of C. vestalis was tested on diamondback moth Plutella xylostella (L.) larvae for different food resources. Cosmos sulphureus, Lantana camara, Coriandrum sativum treatments gave the different mean number of emergence parasitoids per day compared to water treatment. Results implied that food resources from flowers benefitted the longevity of C. vestalis.

Expression of Plasma hsa-miR122 in HBV-Related Hepatocellular Carcinoma (HCC) in Vietnamese Patients.

BACKGROUND: Hepatocellular carcinoma (HCC) is the leading cause of cancer-related death in the world and considered as one of the most susceptible cancers in humans. The microRNA molecule, hsa-miR122, considered as a potential biological marker linked with the injury of hepatocellular tissue, is the most common microRNA in human liver cancer. Understanding the expression profile of hsa-miR122 plays an important role in the diagnosis of HCC. OBJECTIVE: Identification and comparison of cut-off values of plasma hsa-miR122 expression were conducted in blood samples of healthy control, HBV infected and HBV-related HCC Vietnamese patients. METHODS AND RESULT: Fifty-two blood samples of healthy control and HBV-related HCC cases, collected between 2015 and 2017 were obtained from Ho Chi Minh City Oncology Hospital, Vietnam. Written informed consent was attained from all patients and the Human Research Ethics Committee, Oncology Hospital (#08/BVUB-HDDD) approved the research protocol. Total RNA was isolated from blood samples with TrizolTM Reagent (Thermo Fisher Scientific, USA). To analyze the expression level of hsa-miR122, miRNA specific reverse transcription was performed using Sensi- FASTTM cDNA Synthesis Kit (Bioline, UK) as described by the manufacturer, followed by running RT-qPCR with SensiFASTTMSYBR No-ROX Kit (Bioline, UK). The housekeeping gene, GAPDH (glyceraldehyde-3-phosphate dehydrogenase) was used for normalization. The presence of hsamiR122 and HBV-DNA was identified in human blood using RT-PCR and LAMP techniques. Downregulation of plasma hsa-miR122 was observed in HBV-related HCC patients with a .Ct value of 7.9 ± 2.1 which was significantly lower than found in healthy control (p<0.01). The loss of hsa-miR122 expression was observed in HBV infected patients. We also identified the difference of diagnostic values of this microRNA in different populations and provided a high diagnostic accuracy of HCC (AUC = 0.984 with sensitivity and specificity of 96% and 94%, respectively). CONCLUSION: hsa-miR122 was downregulated in HBV-related HCC patients and found to be lower by approximately 10 fold than in healthy control, resulting in a potential biomarker for microRNA based diagnosis of HCC in human blood.

Closed tube loop-mediated isothermal amplification assay for rapid detection of hepatitis B virus in human blood.

Recently, many studies have demonstrated the significant advantages of loop-mediated isothermal amplification (LAMP) based methods over serological tests and PCR for rapid detection of microbial pathogens. Here, a rapid LAMP assay was developed to detect the hepatitis B virus (HBV) from DNA, and particularly, blood samples from infected patients using a commercially available master mix and portable real-time fluorometer. The final optimized fluorescence-based LAMP assay provided significant amplification time of less than 15 minutes compared with over 1 hour for PCR and an opened tube LAMP system described previously. Results indicated that fluorescence-based LAMP assay was more sensitive than PCR as a rapid, sensitive, efficient, and highly reliable approach for rapid detection of HBV.

The Role of Cell Wall Degrading Enzymes in Pathogenesis of Magnaporthe oryzae.

The plant cell wall is always the physical barrier in which phytopathogenic fungi must overcome by producing an array of cell wall degrading enzymes (CWDEs) that allow them to invade host tissues through the degradation of cell wall components of plants. Magnaporthe oryzae is a causal agent of blast disease, one of the most devastating disease in rice resulting significant crop losses worldwide. The penetration of plant cuticle and cell walls induced by infection structures of M. oryzae has been known to be acquired by the association of turgor pressure and CWDEs for successful infection of M. oryzae. In this review, we focus on recent discoveries of M. oryzae CWDEs, gene regulation and their biological roles as fungal virulence factors and elicitors of host defense response leading to plant resistance against fungal pathogens.