White tail disease in Macrobrachium rosenbergii is caused by M. rosenbergii nodavirus (MrNV) infection, resulting in up to 100% mortality in larvae and post-larvae stages, severely impacting aquaculture production. Existing genome-based detection methods for MrNV are costly and time-consuming, highlighting the need for rapid and cost-effective diagnostic tests. This study evaluated the effects of truncating selected aptamer on its binding affinity to the MrNV capsid protein. The previously isolated and identified aptamer through magnetic-capture SELEX and Next Generation Sequencing demonstrated high binding affinity to the MrNV capsid protein. Truncation at the primer overhang was found to improve binding affinity, reducing the dissociation constant from 347 nM to 30.1 nM. The calculated limit of detection for the truncated aptamer decreased from 5.64 nM to 1.7 nM, while the limit of quantification decreased from 17.1 nM to 5.16 nM. These reductions indicate that the truncated aptamer has higher sensitivity compared to the full-length aptamer. In tests with MrNV-infected M. rosenbergii samples, both the enzyme-linked aptamer assay and the gold nanoparticle aptasensor assay showed consistent results when 0.5 μg of total protein lysate was used. This indicates that the prawn protein concentration interferes with the detection of the viral protein. These findings suggest the potential application of the truncated aptamer as a sensor in the development of a practical aptamer-based diagnostic kit. For instance, an aptamer-based lateral flow assay test kit could provide a user-friendly, cost-effective solution that eliminates the need for sophisticated instrumentation for diagnosis or data interpretation, making it ideal for detecting MrNV infection in M. rosenbergii aquaculture.
Three novel endophytic streptomycetes have been isolated and characterized from plants with ethnobotanical uses on the Malay Peninsula including: Thottea grandiflora (family -Aristolochiaceae), Polyalthia spp. (family -Annonaceae), and Mapania sp. (family -Cyperaceae). Each isolate, as studied by scanning electron microscopy, has small hyphae, and produces typical barrel-shaped spores arising by hyphal fragmentation. Interestingly, although none has any detectable antibacterial killing properties, each has demonstrable killing activity against one or more pathogenic fungi including organisms such as Phytophthora erythroseptica, Pythium ultimum, Sclerotinia sclerotiorum, Mycosphaerella fijiensis and Rhizoctonia solani. Molecular biological studies on the rRNA gene sequence of each isolate revealed that it is distinct from all other genetic accessions of streptomyectes in GenBank, and each bears some genetic similarity to other streptomycetes. The bioactivity of each microbe was extractable in various organic solvents.
The giant freshwater prawn holds a significant position as a valuable crustacean species cultivated in the aquaculture industry, particularly well-known and demanded among the Southeast Asian countries. Aquaculture production of this species has been impacted by Macrobrachium rosenbergii nodavirus (MrNV) infection, which particularly affects the larvae and post-larvae stages of the prawn. The infection has been recorded to cause mortality rates of up to 100% among the affected prawns. A simple, fast, and easy to deploy on-site detection or diagnostic method is crucial for early detection of MrNV to control the disease outbreak. In the present study, novel single-stranded DNA aptamers targeting the MrNV capsid protein were identified using the systematic evolution of ligands by exponential enrichment (SELEX) approach. The aptamer was then conjugated with the citrate-capped gold nanoparticles (AuNPs), and the sensitivity of this AuNP-based aptasensor for the detection of MrNV capsid protein was evaluated. Findings revealed that the aptamer candidate, APT-MrNV-CP-1 was enriched throughout the SELEX cycle 4, 9, and 12 with the sequence percentage of 1.76%, 9.09%, and 12.42%, respectively. The conjugation of APT-MrNV-CP-1 with citrate-capped AuNPs exhibited the highest sensitivity in detecting the MrNV capsid protein, where the presence of 62.5 nM of the viral capsid protein led to a significant agglomeration of the AuNPs. This study demonstrated the practicality of an AuNP-based aptasensor for disease diagnosis, particularly for detecting MrNV infection in giant freshwater prawns.