Agroathelia rolfsii (A. rolfsii) is a fungal infection and poses a significant threat to over 500 plant species worldwide. It can reduce crop yields drastically resulting in substantial economic losses. While conventional detection methods like PCR offer high sensitivity and specificity, they require specialized and expensive equipment, limiting their applicability in resource-limited settings and in the field. Herein, we present an integrated workflow with nucleic acid extraction and isothermal amplification in a lab-on-a-chip cartridge based on immiscible filtration assisted by surface tension (IFAST) to detect A. rolfsii fungi in soil for point-of-need application. Our approach enabled both DNA extraction of A. rolfsii from soil and subsequent colorimetric loop-mediated isothermal amplification (LAMP) to be completed on a single chip, termed IFAST-LAMP. LAMP primers targeting ITS region of A. rolfsii were newly designed and tested. Two DNA extraction methods based on silica paramagnetic particles (PMPs) and three LAMP assays were compared. The best-performing assay was selected for on-chip extraction and detection of A. rolfsii from soil samples inoculated with concentrations of 3.75, 0.375 and 0.0375 mg fresh weight per 100-g soil (%FW). The full on-chip workflow was achieved within a 1-h turnaround time. The platform was capable of detecting as low as 3.75 %FW at 2 days after inoculation and down to 0.0375 %FW at 3 days after inoculation. The IFAST-LAMP could be suitable for field-applicability for A. rolfsii detection in low-resource settings.

The red‐whiskered bulbul (Pycnonotus jocosus) is a popular avian species in Thailand and many other countries. The red‐whiskered bulbul has a high economic value, but breeding is challenging since sex identification is difficult. The PCR method is now used for sex identification. However, PCR amplification and post‐PCR analysis necessitate the use of a laboratory equipped with specialized scientific instruments, which is inconvenient for field operations. This research describes a method for amplification of DNA samples using the loop‐mediated isothermal amplification (LAMP) approach, which is a molecular biology methodology for isothermal amplification that is extremely sensitive, fast, and easy for post‐LAMP product visualization. Herein, total of 23 blood samples were collected and DNA was extracted. Two sets of LAMP primers were designed for CHD‐Z and CHD‐W genes. The colorimetric assay was used to investigate the best conditions for LAMP reactions and post‐LAMP product visualization. LAMP reactions for sex identification were compared to traditional PCR in terms of sensitivity and specificity. LAMP reactions were found to be 10‐fold more sensitive than PCR at 1 ng of DNA. When compared to electrophoresis analysis, the visualization with colorimetric assay using GelRed® and SYTO™ 9 was 100% accurate. The optimal LAMP condition tested simple DNA extracted from bird feathers using the HotSHOT technique. The result showed that the optimal condition could distinguish the sex of red‐whiskered bulbuls totally and accurately. A powerful method for red‐whiskered bulbul sex identification is demonstrated in this study, which can be used in field studies because it is quick and easy to perform, has high sensitivity, and does not require advanced scientific equipment.

Cadmium (Cd) is produced mainly as a by-product of zinc mining. In Thailand, the largest zinc mine is located in the Mae Sot district, Tak Province. Samples of Monopterus albus were collected from paddy fields in 4 sites, three downstream and one upstream from the zinc mine. The upstream site was considered to be uncontaminated while the three downstream sites were considered to be contaminated with Cd. Studies on the accumulation level of cadmium were conducted on the liver of the fish using the atomic absorption spectrophotometer technique. The metallothionein (MT) gene expression level in the liver, as a potential biomarker for long-term Cd exposure in their natural habitat, was also assessed. The level of hepatic MT gene expression was performed by quantitative real-time PCR. The result showed that Cd accumulation in the liver was much higher in swamp eels collected from the downstream sites when compared to those collected from the upstream site. The hepatic MT level in the upstream site was 0.75-fold, while the other three downstream sites were 0.36-, 4.44- and 0.94-fold. There is no parallel correlation between hepatic cadmium levels and hepatic MT gene expression. This study then suggests that MT gene expression biomarkers might be not suitable for swamp eels with prolonged exposure to Cd.

The giant freshwater prawn (Macrobrachium rosenbergii) has economic significance in the aquatic industry, but production is impacted by infectious diseases induced by various pathogens. Herein, we investigated the impact of adding feed additives to the diet of M. rosenbergii to promote health. Diets were formulated to contain different levels of Centella asiatica (L.) Urb. crude extracts (1, 5, and 10 g kg⁻¹), with growth performance and innate immune parameters assessed after 28 days of feeding. Real-time quantitative PCR (RT-qPCR) was employed to determine the mRNA levels of serine proteinase inhibitor (SPI) and alpha-2 macroglobulin (Mr-2α2M) from 12 h to 28 days of feeding. Prawns feeding at 5 and 10 g kg⁻¹ showed statistically significant differences in specific growth rate, lysozyme assay, and phenoloxidase activity. The expression levels of all the immune-related genes studied were significantly upregulated in prawns fed with supplemented diets compared to the control group. Findings revealed that the observed upregulations varied in response to alterations in feeding time and concentration. Furthermore, 16S rRNA analysis showed that the supplemented diets at 10 g kg⁻¹ supplementation increased beneficial bacteria (Lactococcus sp.) and reduced pathogenic taxa (e.g., Candidatus Hepatoplasma, Flavobacteriaceae, Weeksellaceae, Thiothrix sp.).

α-Hemoglobin (Hb) variants are common genetic mutations worldwide. The appropriate techniques must be followed to scan and identify these variants, particularly in routine investigations for thalassemia and hemoglobinopathies in countries with high prevalence of α and β-thalassemia. High resolution melting (HRM) analysis is a popular and effective technique for identifying genetic variations with rapid output results. This study designed four newly developed primer pairs that had full coverage of the HBA genes for detection of α-Hb variants using real-time PCR with HRM analysis. Forty-one blood samples were collected from individuals with known or suspected α-Hb variants. The results demonstrated clearly distinguished melting patterns of nine α-Hb variants including Hb Constant Spring, Hb Q-Thailand, Hb Pakse', Hb Hekinan, Hb Nakhon Ratchasima, Hb Siam, Hb Thailand, Hb Queens, and Hb Quong Sze compared with the wild-type sample pattern. All mutations were confirmed by DNA nucleotide sequencing. This study presents the first case report of the combination of Hb Shaare Zedek co-inherited with Hb Hekinan in a Thai patient. Interactions between these two Hb variants displayed a high level of Hb F (23.5%) on an HPLC Hb chromatogram and a mild symptom phenotype with low mean corpuscular volume (71.3 fL) and mean corpuscular hemoglobin (21.8 pg) in the proband. Overall, HRM analysis is a suitable, rapid, and powerful technique for the identification of gene mutations, and for the diagnosis of common and rare α-Hb variants to prevent and control thalassemia in Thailand.