Browsing by Author "Nurul Hidayah Samsulrizal, Ph.D"

Edwardsiella ictaluri is the etiological agent of Bacillary Necrosis of Pangasius (BNP) in Pangasianodon hypophthalmus, causing substantial economic losses in freshwater aquaculture. Infected fish commonly exhibit clinical signs such as erratic swimming behaviour, patchy liver, and congested kidney. Despite its significance, the genomic characteristics, population structure, and epidemiological context of E. ictaluri in Malaysia, particularly in major production areas such as Temerloh and Pekan, remain limited. This knowledge gap hinders effective disease surveillance and control strategies in local aquaculture systems. Therefore, this study aimed to investigate the genomic characteristics, antimicrobial resistance determinants, and population-level genetic variation of E. ictaluri isolates from Temerloh and Pekan using whole-genome sequencing (WGS) and polymorphism-based approaches. Eleven E. ictaluri isolates were recovered from the tissues of clinically diseased P. hypophthalmus cultured in riverine cage systems in these areas. As all isolates originated from the same water body, one representative isolate, PLS31B, obtained from the brain of an infected fish, was selected for whole-genome sequencing (WGS) using Oxford Nanopore long-read technology. The assembled genome was 3.8 Mbp in size, with a high-contiguity N50 of 3,761,778 bp and a GC content of 57.39%. Antimicrobial resistance (AMR) analysis revealed resistance to fluoroquinolones, macrolides, and penams, highlighting potential challenges for disease management in aquaculture. Functional annotation identified 3,620 protein-coding genes and 126 RNA genes. Comparative genomic analysis indicated that PLS31B is closely related to E. ictaluri strains from China and Vietnam, suggesting potential regional connectivity or transboundary dissemination associated with the importation of fish seed. A large conserved core genome comprising of 3,246 genes was observed alongside of 51 strain-specific genes linked to those encoding outer membrane protein C precursor (OMP), ATP-binding protein, acid shock protein, and hemolysin secretion protein. Meanwhile, genetic polymorphism among all isolates was further assessed using Random Amplified Polymorphic DNA (RAPD) profiling and Sanger sequencing. Seven 10-mer primers were used for RAPD analysis, while the 23S rRNA gene region was sequenced for comparative analysis. Polymorphism analysis revealed minimal genetic differentiation among the isolates, irrespective of sampling locations or host age groups. This low genetic diversity is likely influenced by a shared aquatic environment and interconnected farming systems. In conclusion, these findings highlight the epidemiological relevance of regional dissemination pathways and guide the development of targeted control strategies, including early detection tools, preventive measures, and effective treatment approaches for BNP in P. hypophthalmus aquaculture.

Mitragyna speciosa, commonly known as kratom or “ketum”, is a tropical plant belonging to the Rubiaceae (coffee) family, predominantly found in Southeast Asia, particularly in Malaysia, Indonesia and Thailand. It is widely recognized for its medicinal properties, primarily due to mitragynine, a major psychoactive compound with stimulant and analgesic effects. Mitragynine biosynthesis involves a complex metabolic pathway in which the enzyme tryptophan decarboxylase (TDC) plays a critical role by converting tryptophan into tryptamine, a key alkaloid precursor. This study aimed to identify and characterize the TDC gene from the M. speciosa using molecular techniques and bioinformatics, focusing on its gene structure, three-dimensional modelling and mechanistic interaction, particularly the protein-ligand interaction between the TDC and tryptophan. Functional analysis revealed that the region spanning amino acid positions 57 to 430 represents a conserved TDC protein domain involved in metal-ion interaction and ligand binding within a full-length protein of 506 amino acids. The TDC gene was successfully isolated using the conventional Cetyltrimethylammonium bromide (CTAB), followed by Polymerase Chain Reaction (PCR) amplification. The resulting 1,599 bp nucleotide sequence was validated via Needleman-Wunsch pairwise alignment. Phylogenetic analysis was conducted using MEGA-X, and domain prediction with INTERPRO confirmed the presence of the Pyridoxal-Dependent Decarboxylase domain, which binds pyridoxal phosphate (PLP) and is essential for catalyzing the decarboxylation of amino acids, enabling the conversion of L-tryptophan to tryptamine. Physicochemical analysis using PROTPARAM classified TDC as a polar protein with an optimum isoelectric point (pI) of 6.05. The three-dimensional structure modelling identified the AlphaFold model as the most reliable, with ERRAT quality score of 95.7983. Tunnel visualization revealed a ligand-accessible channel measuring 4.7 Å in length with a bottleneck radius of 2.25 Å, suggesting ideal path for substrate transport. Molecular docking analysis using Webina and SeamDock platforms showed that Webina provided a more stable and stronger interaction, with a binding affinity of -9.901 kcal/mol, compared to -5.4 kcal/mol for SeamDock. The protein–ligand complex was further refined using YASARA energy minimization, resulting in a reduced total potential energy of −6.688 × 10⁴ kcal/mol, indicating a stable and optimized complex structure. In conclusion, this study elucidates the functional role of TDC in the biosynthesis of tryptamine-derived alkaloids in M. speciosa. The findings provide a molecular basis for future metabolic engineering and synthetic biology studies targeting TDC to enhance the production of specific pharmaceutically important alkaloids, particularly mitragynine.

Species identification based on hypanthium structure and herbarium samples were insufficient for Melastoma. Melastoma flowers are very delicate and tend to become incomplete specimens. Therefore, it is impossible to rely solely on flower characteristics and herbarium samples for the identification process. Hence, a comprehensive study was set up to identify varietal differences among Melastoma species found in Peninsular Malaysia. Our objectives can be restated as follows 1) to identify and describe the morphological characteristics of Melastoma species, 2) to identify and describe the anatomical and micromorphological characteristics of Melastoma species and 3) to identify and characterize the gene of Melastoma species. A total of 26 Melastoma accession were collected and given a final of 87 vegetative structures and 144 reproductive structures. This study has identified hypanthium, leaf, and twigs indumentum as important morphological structures for species identification. Meanwhile, 54 anatomical characters were identified and recorded among 20 selected taxa. Besides, 15 types of trichomes were determined to serve as supporting characters for Melastoma in the absence of reproductive structure. DNA analysis conducted on 11 selected taxa of Melastoma were subjected to DNA extraction, PCR with four new primers (triosephosphate isomerase (tpi) gene, photosystem II protein D1, partial cds; chloroplast (psbA) gene, granule-bound starch synthase (gbss) gene and vacuolar invertase (vr) gene), and DNA sequencing to obtain the final result. Nevertheless, only the tpi genes showed a promising result with convincing bootstrap support in the phylogenetic study. Of the eleven taxa, nine proved to be distinct species based on genetic characters. In conclusion, this study identified 14 taxa and seven M. malabathricum accessions. Out of 14 taxa, 6 were newly described, and three were reinstatement as species. These results reinforce the hypothesis that morphological structure will be the most important source of information for the identification of Melastoma.