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Endophytic lactic acid bacteria (LAB) isolated from papaya seeds, including a consortium of two LAB isolates, Weissella cibaria PPKSD19 and Lactococcus lactis subsp. lactis PPSSD39 could previously inhibit papaya dieback disease causative agent, Erwinia mallotivora BT-MARDI in vitro, indicating their potential as biofertilizer. However, further characterizations on other plant growth-promoting (PGP) properties of the LABs are prerequisite to use in agricultural settings as bio-inoculum. Hence, this study aimed to evaluate PGP potentials further and in vitro antifungal activity of the LABs against various plant pathogens. The LAB isolates were tested positive in indole-3-acetic acid (IAA), siderophore, and ammonia production and could solubilize phosphate. Weissella cibaria PPKSD19 and L. lactis subsp. lactis PPSSD39 exhibited the strongest in vitro antifungal activity against Fusarium oxysporum TKA and Curvularia lunata. Inoculum concentration of 1x10 cfu/ml of W. cibaria PPKSD19 and L. lactis subsp. lactis PPSSD39 mixture showed the highest increment in shoot and root dry weight. In conclusion, W. cibaria PPKSD19 and L. lactis subsp. lactis PPSSD39 consortium displayed promising plant probiotic potential. These results highlighted the possibility of the bacterial consortium to be exploited as bioinoculant to promote plant growth and inhibit phytopathogens causing plant diseases.

Aims: Leaf blight disease caused by Pantoea spp. reduces rice yields in numerous nations. However, the exact strategy to combat Pantoea spp. has yet to be determined. Bactronophorus thoracites is a promising source of natural antimicrobial agents due to their potential as a substrate to generate peptides with high antimicrobial activity. This study determined the effects of lactic acid fermentation using Lactobacillus casei ATCC334 as a starter culture on antimicrobial activity against rice pathogens, proximate composition, and amino acid profiles from B. thoracites crude extract. Methodology and results: Bactronophorus thoracites was washed and deshelled to collect the flesh and homogenised at 4 °C before freeze-drying. The freeze-dried samples were fermented with L. casei for 4 to 8 days at 37 °C. The antimicrobial activity, MIC and MBC were determined using a spectrometer. The fermented protein was subjected to proximate and amino acid analyses. The antimicrobial activity of fermented B. thoracites protein (FBTP) was significantly (p<0.05) decreased with the increased fermentation days (from 4 to 8 days). The antimicrobial activity was also increased when the glucose concentration increased from 2% to 3%. However, raising the glucose concentration to 4% decreased the antimicrobial activity. The antimicrobial activity was significantly (p<0.05) increased when the substratewater (S/W) ratio increased from 0.84% to 0.96%. The FBTP (4 days, 3% glucose concentration and 0.96% S/W ratio) showed high antimicrobial activity against Pantoea ananatis and P. stewartii. The MIC and MBC values for FBTP were 500 µg/mL and 250 µg/mL against P. ananatis and P. stewartii. The zones of inhibition value for FBTP were 16.0 ± 0.5 mm (1000 µg/mL) and 9.33 ± 0.57 mm (500 µg/mL) for P. ananatis, and 11.7 ± 0.61 mm (1000 µg/mL), 9.33 ± 0.58 mm (500 µg/mL) and 7.17 ± 0.77 mm (250 µg/mL) for P. stewartii. The proximate composition and amino acid profiles of the freeze-dried protein hydroly

Fusarium oxysporum f. sp. cubense tropical race 4 (FocTR4) is a destructive necrotrophic fungal pathogen afflicting global banana production. Infection process involves the activation of programmed cell death (PCD). In this study, seven Musa acuminata vacuolar processing enzyme (MaVPE1–MaVPE7) genes associated with PCD were successfully identified. Phylogenetic analysis and tissue-specific expression categorized these MaVPEs into the seed and vegetative types. FocTR4 infection induced the majority of MaVPE expressions in the susceptible cultivar “Berangan” as compared to the resistant cultivar “Jari Buaya.” Consistently, upon FocTR4 infection, high caspase-1 activity was detected in the susceptible cultivar, while low level of caspase-1 activity was recorded in the resistant cultivar. Furthermore, inhibition of MaVPE activities via caspase-1 inhibitor in the susceptible cultivar reduced tonoplast rupture, decreased lesion formation, and enhanced stress tolerance against FocTR4 infection. Additionally, the Arabidopsis VPE-null mutant exhibited higher tolerance to FocTR4 infection, indicated by reduced sporulation rate, low levels of H2O2 content, and high levels of cell viability. Comparative proteomic profiling analysis revealed increase in the abundance of cysteine proteinase in the inoculated susceptible cultivar, as opposed to cysteine proteinase inhibitors in the resistant cultivar. In conclusion, the increase in vacuolar processing enzyme (VPE)-mediated PCD played a crucial role in modulating susceptibility response during compatible interaction, which facilitated FocTR4 colonization in the host.

Water stress is one of the major challenges to food security, causing a significant economic loss for the nation as well for growers. Accurate assessment of water stress will enhance agricultural productivity through optimization of plant water usage, maximizing plant breeding strategies, and preventing forest wildfire for better ecosystem management. Recent advancements in sensor technologies have enabled high-throughput, non-contact, and cost-efficient plant water stress assessment through intelligence system modeling. The advanced deep learning sensor fusion technique has been reported to improve the performance of the machine learning application for processing the collected sensory data. This paper extensively reviews the state-of-the-art methods for plant water stress assessment that utilized the deep learning sensor fusion approach in their application, together with future prospects and challenges of the application domain. Notably, 37 deep learning solutions fell under six main areas, namely soil moisture estimation, soil water modelling, evapotranspiration estimation, evapotranspiration forecasting, plant water status estimation and plant water stress identification. Basically, there are eight deep learning solutions compiled for the 3D-dimensional data and plant varieties challenge, including unbalanced data that occurred due to isohydric plants, and the effect of variations that occur within the same species but cultivated from different locations.

Curcuma aeruginosa from the Zingiberaceae family has been cultivated for its traditional and medicinal properties in Malaysia. This study aimed to investigate the volatile constituents of C. aeruginosa essential oil from Malaysia and evaluate its antimicrobial properties compared to C. aeruginosa essential oil from other countries. The essential oils were obtained from fresh and dried leaves and rhizome of C. aeruginosa using the hydrodistillation method and analyzed using gas chromatography-mass spectrometry (GC/MS). The highest essential oil yield was obtained from dried rhizome at 0.63 % (w/w). The major volatile constituent found in the essential oil isolated from fresh leaves were 1,8-cineole (7.91 %), curzerene (5.16 %), and germacrone (4.91 %); dried leaves contained 1,8-cineole (17.21 %), camphor (6.55 %) and curzene (3.85 %); fresh rhizomes contained 1,8-cineole (20.53 %) and camphor (6.08 %) whereas dried rhizomes contained 1,8- cineole (18.41 %), camphor (9.89 %), and DL-isoborneol (3.13 %). The essential oils showed good antibacterial and antifungal activity against Staphylococcus aureus, Bacillus subtilis Escherichia coli, Aspergillus niger Candida albicans but were not effective against Serratia marcescens. This study highlights the variation in the volatile constituents of C. aeruginosa essential oil from different countries and the antimicrobial potential of C. aeruginosa essential oil from Malaysia, which may be used against various bacterial and fungal species.

The usage of plant growth-promoting (PGP) Streptomyces to improve plant growth is an alternative to existing strategies of chemical fertilizers that commonly caused environmental pollution. The aim of this study was to isolate and characterize PGP Streptomyces from Malaysian rice fields with antagonistic activity against Bacterial Panicle Blight disease in rice caused by Burkholderia glumae. A total of 50 bacterial isolates were recovered from the soil, rhizosphere, and endosphere; 22% showed antagonistic activity against B. glumae. Molecular identification using 16S rRNA amplification and phylogenetic tree analyses revealed that the antagonistic isolates belonged to the genus Streptomyces. Among the antagonistic isolates, Streptomyces sp. TBS5 (26.35 ± 0.14 mm) demonstrated the most antagonistic activity (p < 0.05) against B. glumae. Through API® ZYM analysis, the antagonistic isolates were revealed to have phosphoric hydrolase, ester hydrolase, peptide hydrolase, and glycosidase activities that play a crucial role in plant growth promotion. The antagonistic isolates demonstrated the highest (80%) PGP traits including able to fix nitrogen and solubilize phosphate, as well as produce siderophore and indole-3-acetic acid. Plant growth promotion assay under laboratory and greenhouse conditions were analyzed with the treatment of rice, Oryza sativa, seeds with the antagonistic isolates. Seeds treated with Streptomyces sp. TBS5, JAS2, R2-7, and TKR8 showed improvement in vigor index by more than 100% compared to water-treated control plants under both conditions. Augmentation of root length showed an overall increment of more than 101.5% under laboratory condition and 151.9% under greenhouse condition using Streptomyces sp. TBS5, JAS2, R2-7, and TKR8 treatments. Results suggest that Streptomyces sp. TBS5, JAS2, R2-7, and TKR8 are a good candidate to be developed as both biocontrol agent against B. glumae and PGP agent in rice.

The failure of bio-control agents (BCAs) in suppressing Fusarium oxysporum f. sp. cubense tropical race 4 (Foc-TR4) was reported in vivo although antagonism was obtained from in vitro dual culture assay. This study aims to develop rapid assays to validate the antagonism of BCAs using detached banana leaves and roots. Both Trichoderma harzianum and Pseudomonas aeruginosa reduced root lesion length (2.7–3.1 cm) compared to 4.9 cm in TR4 treatment. BCA-treated leaves reduced wilting severity from 93.75 to 33.75%. Trypan blue staining showed lower cell death area in BCA-treated roots and leaves. Reduced leaf wilting was indicated by lower red ink content and higher chlorophyll content in BCA-treated leaves. Both defense-related genes, PR-1 and PR-3 were expressed in BCA-treated roots and leaves which could be associated with reduced root necrosis and leave wilting. In conclusion, both assays could be served as rapid methods to select potential BCAs before attempting large-scale trials.

Conventional refolding methods are associated with low yields due to misfolding and high aggregation rates or very dilute proteins. In this study, we describe the optimization of the conventional methods of reverse dilution and affinity chromatography for obtaining high yields of a cysteine rich recombinant glycoside hydrolase family 19 chitinase from Streptomyces griseus HUT6037 (SgChiC). SgChiC is a potential biocontrol agent and a reference enzyme in the study and development of chitinases for various applications. The overexpression of SgChiC was previously achieved by periplasmic localization from where it was extracted by osmotic shock and then purified by hydroxyapatite column chromatography. In the present study, the successful refolding and recovery of recombinant SgChiC (r-SgChiC) from inclusion bodies (IB) by reverse dilution and column chromatography methods is respectively described. Approximately 8 mg of r-SgChiC was obtained from each method with specific activities of 28 and 52 U/mg respectively. These yields are comparable to that obtained from a 1 L culture volume of the same protein isolated from the periplasmic space of E. coli BL21 (DE3) as described in previous studies. The higher yields obtained are attributed to the successful suppression of aggregation by a stepwise reduction of denaturant from high, to intermediate, and finally to low concentrations. These methods are straight forward, requiring the use of fewer refolding agents compared with previously described refolding methods. They can be applied to the refolding of other cysteine rich proteins expressed as inclusion bodies to obtain high yields of actively folded proteins. This is the first report on the recovery of actively folded SgChiC from inclusion bodies.

Respiratory burst oxidase homologs (RBOH) is the key enzyme responsible for the production of reactive oxygen species which act as important signal during plant responses to abiotic and biotic stresses. However, RBOH homologs have not been characterized in banana. In this study, we have identified twelve Rboh genes distributed on eight chromosomes of Musa acuminata subsp. malaccensis (DH Pahang) through a genome-wide analysis. MaRbohs exist as sibling paralogs with variable exon-intron structures and highly conserved functional domain. Phylogenetic analysis clustered MaRBOH into four distinct subgroups (I, II, III and IV). The expression of MaRbohs following 24 hours of inoculation with Fusarium oxysporium f. sp. cubense Tropical Race 4 revealed diverse patterns in root tissues. MaRbohH1 and MaRbohA2 were strongly upregulated and downregulated, respectively in response to the root-dip inoculation. This is the first report on genome-wide characterization of Rboh genes from banana and their expression in response to the fungal pathogen TR4. This research provides a basis for exploration of the role of NADPH oxidase in banana defence against the root pathogen.

Fusarium wilt of banana, caused by Fusarium oxysporum f. sp. cubense (Foc), is a major factor limiting banana production in Malaysia. To date, SSR, ISSR, AFLP and RAPD markers are widely used in PCR-based molecular characterization of Foc. Although these markers are effective in characterizing Foc, they all have their own limitations. An effector gene known as Secreted in Xylem (SIX) has been explored to broaden the molecular diagnostic toolbox for Foc. This study aims to investigate the diversity of Foc-TR4 in Peninsular Malaysia based on the phylogenetic analysis of SIX sequences. The virulence potential of twenty-seven Fusarium wilt of banana based on pathogenicity test and the presence of SIX1, SIX7 and SIX8 genes were determined. All of twenty-seven Foc ‘Tropical Race 4’ (TR4) (VCG 01213/160) isolates were obtained from Biological Control Laboratory, Department of Plant Protection, Universiti Putra Malaysia (UPM). The results showed that all Foc-TR4 isolates were pathogenic towards banana plantlets at different severity levels under greenhouse condition. Pathogenicity assays showed that the level of aggressiveness differs between isolates. The Foc-TR4 isolates were screened for the presence of three SIX genes (SIX1, SIX7 and SIX8) and the genetic differentiation of Foc-TR4 was evaluated by PCR analysis using three specific primers. Among these three SIX genes, SIX1 and SIX8 genes were detected in all twenty-seven Foc-TR4 isolates from Peninsular Malaysia. Phylogenetic analysis using SIX1 and SIX8 sequences showed that they were related to TR4 isolates (VCG 01213/16) from Australia and Indonesian with bootstrap values of 94% and 100% respectively. No variation was observed in the SIX1 and SIX8 sequences since all 27 isolates were clustered into the same clade. Virulence level also did not correlate with the existence or lack of these genes. Since Foc could hamper the local banana production in Malaysia, characterization of these isolates through SIX genes provid

Salinization of rice cultivation land has progressively enlarged, thus negatively impaired the world's rice bowl. Due to the polygenic nature and complexity of salinity tolerance mechanisms in plants, the development of new rice varieties with better adaptation to salinity has become a great challenge. Regarding this, transcriptomic profiling has been seen as a promising approach for a holistic understanding of salinity tolerance mechanisms in rice. Using Illumina RNA-Seq method, transcriptomes of two contrasting Malaysian rice varieties named as salt-tolerant MR211 and salt-sensitive MR220 were analysed within a short-term exposure (9 h) to salt-shock treatment (12 dS m-1) at early seedling stage. Transcriptomic analysis using Tuxedo package enabled the identification of 252 differentially expressed genes (DEGs). Interestingly, 93.3% of the DEGs (n=235) were identified as higher and specifically expressed in salt-tolerant MR211 compared to the sensitive variety (MR220's FPKM ≤ 0). This group of DEGs was assigned in 33 KEGG pathways, with the highest number of transcripts accounted in purine and thiamine metabolism pathways. Meanwhile, functional annotation analyses revealed the presence of regulatory genes, annotated functional and unknown genes involved in various salt adaptation mechanisms in the salt-tolerant variety MR211. The expression accuracy and reproducibility of the 252 DEGs identified from the RNA-Seq experiment were further verified through semi-quantitative RT-PCR followed by real time PCR analysis.

The global banana production is threatened by Fusarium wilt, a fungal soil-borne disease caused by Fusarium oxysporum f. sp. cubense (Foc). In Malaysia, Foc Tropical Race 4 (Foc-TR4) is the most devastating disease that has completely wiped out the Cavendish plantations in 1990s. The TR4 strain attacks a diverse range of bananas and plantains, making disease management challenging. In this review, we have summarized the current status of Fusarium wilt in Malaysia, current knowledge on the epidemiology of Foc and recommended control measures outlined by the Department of Agriculture (DOA) in preventing and/or avoiding spread of the disease. The research progress on the control of Foc-TR4 in Malaysia was also presented to highlight the current findings and knowledge gaps in each approach. Recommendations for future studies were included to improve the current disease management approaches while possible challenges were provided for researchers, farmers and policy makers to consider.

Dieback disease caused by Erwinia mallotivora is a major threat to papaya plantation in Malaysia. The current study was conducted to evaluate the potential of endophytic lactic acid bacteria (LAB) isolated from papaya seeds for disease suppression of papaya dieback. Two hundred and thirty isolates were screened against E. mallotivora BT-MARDI, and the inhibitory activity of the isolates against the pathogen was ranging from 11.7–23.7 mm inhibition zones. The synergistic experiments revealed that combination of W. cibaria PPKSD19 and Lactococcus lactis subsp. lactis PPSSD39 increased antibacterial activity against the pathogen. The antibacterial activity was partially due to the production of bacteriocin-like inhibitory substances (BLIS). The nursery experiment confirmed that the application of bacterial consortium W. cibaria PPKSD19 and L. lactis subsp. lactis PPSSD39 significantly reduced disease severity to 19% and increased biocontrol efficacy to 69% of infected papaya plants after 18 days of treatment. This study showed that W. cibaria PPKSD19 and L. lactis subsp. lactis PPSSD39 are potential candidate as biocontrol agents against papaya dieback disease.

Aims: This study sought to investigate the effect of bioformulation on the biocontrol efficacy, microbial viability and storage stability of a consortium of Pseudomonas aeruginosa DRB1 and Trichoderma harzianum CBF2 against Foc Tropical Race 4 (Foc-TR4). Materials and Results: Four bioformulations consisting of dry (pesta granules, talc powder and alginate beads) and liquid formulations were evaluated for their ability to control Foc-TR4, sustain microbial populations after application and maintain microbial stability during storage. All tested bioformulations reduced disease severity (DS) by more than 43·00% with pesta granules producing the highest reduction in DS by 66·67% and the lowest area under the disease progress curve value (468·75) in a glasshouse trial. Microbial populations of DRB1 and CBF2 were abundant in the rhizosphere, rhizoplane and within the roots of bananas after pesta granules application as compared to talc powder, alginate beads and liquid formulations 84 days after inoculation (DAI). The stability of both microbial populations after 180 days of storage at 4°C was the greatest in the pesta granule formulation. Conclusion: The pesta granule formulation was a suitable carrier of biological control agents (BCA) without compromising biocontrol efficacy, microbial population and storage stability as compared to other bioformulations used in this study. Significance and Impact of the Study: Pesta granules could be utilized to formulate BCA consortia into biofertilizers. This formulation could be further investigated for possible applications under agricultural field settings.