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Background: Gastrointestinal cancer (GIC) ranks as the highest cause of cancer-related deaths globally. GIC patients are often diagnosed at advanced stages, limiting effective treatment options. Chemotherapy, the common GIC recommendation, has significant disadvantages such as toxicity and adverse effects. Natural products contain substances with diverse pharmacological characteristics that promise for use in cancer therapeutics. In this study, the flower of renowned Asian medicinal plant, Shorea roxburghii was collected and extracted to investigate its phytochemical contents, antioxidant, and anticancer properties on GIC cells., Methods: The phytochemical contents of Shorea roxburghii extract were assessed using suitable methods. Phenolic content was determined through the Folin-Ciocalteu method, while flavonoids were quantified using the aluminum chloride (AlCl 3 ) method. Antioxidant activity was evaluated using the FRAP and DPPH assays. Cytotoxicity was assessed in GIC cell lines via the MTT assay. Additionally, intracellular ROS levels and apoptosis were examined through flow cytometry techniques. The correlation between GIC cell viability and phytochemicals, 1 H-NMR analysis was conducted., Results: Among the four different solvent extracts, ethyl acetate extract had the highest phenolic and flavonoid contents. Water extract exhibited the strongest reducing power and DPPH scavenging activity following by ethyl acetate. Interestingly, ethyl acetate extract demonstrated the highest inhibitory activity against three GIC cell lines (KKU-213B, HepG2, AGS) with IC 50 values of 91.60 µg/ml, 39.38 µg/ml, and 35.59 µg/ml, while showing less toxicity to normal fibroblast cells. Ethyl acetate extract induced reactive oxygen species and apoptosis in GIC cell lines by downregulating anti-apoptotic protein Bcl-2. Metabolic profiling-based screening revealed a positive association between reduced GIC cell viability and phytochemicals like cinnamic acid and its derivatives, ferulic acid and coumaric acid., Conclusions: This study highlights the potential of natural compounds in Shorea roxburghii in the development of more effective and safer anticancer agents as options for GIC as well as shedding light on new avenues for cancer treatment., (© 2024. The Author(s).)

Detection of worm antigen in urine is a sensitive diagnostic method for opisthorchiasis, particularly for light-intensity infections; however, the presence of eggs in feces is essential for validating results from the antigen assay. To address the issue of low sensitivity of fecal examination, we modified the protocol for the formalin-ethyl acetate concentration technique (FECT) and compared it against urine antigen measurements for detection of the parasite Opisthorchis viverrini. First, we optimized the FECT protocol by increasing the number of drops for examinations from the standard two drops to a maximum of eight. We were able to detect additional cases after examination of ≥ 3 drops, and the prevalence of O. viverrini saturated after examination of ≥ 5 drops. We then compared the optimized FECT protocol (examining five drops of suspension) against urine antigen detection for the diagnosis of opisthorchiasis in field-collected samples. The optimized FECT protocol detected O. viverrini eggs in 25 of 82 individuals (30.5%) who had positive urine antigen tests but were fecal egg negative by the standard FECT protocol. The optimized protocol also retrieved O. viverrini eggs in 2 of 80 antigen-negative cases (2.5%). In comparison with the composite reference standard (combined FECT and urine antigen detection), the diagnostic sensitivity of examining two and five drops of FECT and the urine assay was 58.2, 67, and 98.8%, respectively. Our results show that multiple examinations of fecal sediment increase the diagnostic sensitivity of FECT and thus provide further support for the reliability and utility of the antigen assay for diagnosis and screening of opisthorchiasis.

Detection of IgG in urine is an efficient method comparable to that in serum for diagnosis of strongyloidiasis, but the effects of daily variation in urine dilution on diagnostic accuracy are not clearly known. This study evaluated the effects of urine concentration on the detection of parasite-specific IgG by urine enzyme-linked immunosorbent assay (ELISA), particularly in individuals with borderline results or false-negative diagnosis. Optimal concentration conditions were established by comparing Strongyloides -specific IgG antibody levels between unconcentrated and concentrated urine in participants with different infection intensities, namely, healthy control (HC), low-negative (LN), high-negative (HN), and low-positive (LP) groups. The optimal condition was selected and validated in a field trial study. The final urine concentration protocol required centrifugation at 4,000 × g at 4°C for 10 mins using the Amicon concentrator tube. This protocol was validated in groups of participants with various diagnoses according to urine ELISA and fecal examination ( n  = 148). The concentrated-urine ELISA increased the proportion of positive results in the LN group by 68.2% and by 100% in the HN group. Significantly elevated IgG antibody levels were seen in the LP group. In the group that was false negative by urine ELISA but positive by fecal examination ( n  = 28), concentrated-urine ELISA yielded 100% positive results. Overall, the frequency estimates of Strongyloides stercoralis were 23.6% by fecal culture, 27% by standard urine ELISA, and 90.5% by concentrated-urine ELISA. The concentration of urine samples prior to analysis by ELISA improved the sensitivity for diagnosis and is potentially useful in the diagnosis of strongyloidiasis in immunocompromised individuals or in low-prevalence areas.

To combat and eventually eliminate the transmission of the liver fluke Opisthorchis viverrini, an accurate and practical diagnostic test is required. A recently established urine antigen detection test using monoclonal antibody-based enzyme-linked-immunosorbent assay (mAb-ELISA) has shown promise due to its high diagnostic accuracy and the use of urine in place of fecal samples. To further test the utility of this urine assay, we performed a cross sectional study of 1,043 people in 3 opisthorchiasis endemic communities in northeast Thailand by applying urine antigen detection together with copro-antigen detection methods. The quantitative formalin-ethyl acetate concentration technique (FECT) was concurrently performed as a reference method. The prevalence of O. viverrini determined by urine antigen detection correlated well with that by copro-antigen detection and both methods showed 10-15% higher prevalence than FECT. Within the fecal negative cases by FECT, 29% and 43% were positive by urine and copro-antigen detection, respectively. The prevalence and intensity profiles determined by antigen detection and FECT showed similar patterns of increasing trends of infection with age. The concentration of antigen measured in urine showed a positive relationship with the concentration of copro-antigen, both of which were positively correlated with fecal egg counts. The data observed in this study indicate that urine antigen detection had high diagnostic accuracy and was in concordance with copro-antigen detection. Due to the ease and noninvasiveness of sample collection, the urine assay has high potential for clinical diagnosis as well as population screening in the program for the control and elimination of opisthorchiasis., Competing Interests: The authors have declared that no competing interests exist.

Contaminated liver fluke egg in the environment has led to the high prevalence of human opisthorchiasis associated with cholangiocarcinoma in Southeast Asia. To find the effective lessening methods of Opisthorchis viverrini eggs in the contaminated environment, we investigated the temperature conditions for killing of these trematode eggs in vitro. Numerous O. viverrini eggs were obtained in the proximal part of uteri of adult worms from experimental hamsters. Mature eggs with miracidium were allocated by experimental groups (2 control: positive and negative and 4 treatment: 50, 60, 70, and 80°C) with 0.85% saline, and treated by the experimental plan. Eggs in each experimental groups were observed under the confocal microscope after stain with Propidium Iodide (PI) to evaluate the effect of temperatures. Eggs in 70 and 80°C groups were all killed after over 10 min heated. Majority of eggs in 60°C (10, 15, and 30 min heated), 70 and 80°C (5 min heated) groups were inactivated. However in 50°C group, below half of eggs were to be killed in all time lapse (10, 15 and 30 min). In order to prevent O. viverrini infection and cholangiocarcinoma, direct treatment of sewage by heating at 70 or 80°C at least 10 min is essential. Therefore, treatment of O. viverrini eggs at a high temperature is a potential method for controlling egg contamination in sewage.

Background: Many strategies to control opisthorchiasis have been employed in Thailand, but not in the other neighbouring countries. Specific control methods include mass drug administration (MDA) and health education to reduce raw fish consumption. These control efforts have greatly shifted the epidemiology of Opisthorchis viverrini (OV) infection over the last decade from presenting as densely concentrated "heavy" infections in single villages to widespread "light" OV infections distributed over wide geographical areas. Currently, the "gold standard" detection method for OV infection is formalin ethyl-acetate concentration technique (FECT), which has limited diagnostic sensitivity and diagnostic specificity for light OV infections, with OV eggs often confused with eggs of minute intestinal flukes (MIFs) in feces. In this study, we developed and evaluated the diagnostic performance of a monoclonal antibody-based enzyme-linked immunosorbent assay for the measurement of OV excretory-secretory (ES) antigens in urine (urine OV-ES assay) for the diagnosis of opisthorchiasis compared to the gold standard detection FECT method., Methodology: We tested several methods for pre-treating urine samples prior to testing the diagnostic performance of the urine OV-ES assay. Using trichloroacetic acid (TCA) pre-treated urine, we compared detection and quantification of OV infection using the urine OV-ES assay versus FECT in OV-endemic areas in Northeastern Thailand. Receiver operating characteristic (ROC) curves were used to determine the diagnostic sensitivity and specificity of the urine OV-ES assay using TCA pre-treated urine, and to establish diagnostic positivity thresholds. The Positive Predictive Value as well as the likelihood of obtaining a positive test result (LR+) or a negative test result (LR-) were calculated for the established diagnostic positivity threshold. Diagnostic risks (Odds Ratios) were estimated using logistic regression., Results: When urine samples were pre-treated with TCA prior to use in the urine OV-ES assay, the analytical sensitivity was significantly improved. Using TCA pre-treatment of urine, the urine OV-ES assay had a limit of detection (LoD) of 39 ng/ml compared to the LoD of 52 ng/mL reported for coprological antigen detection methods. Similarly, the urine OV-ES assay correlated significantly with intensity of OV infection as measured by FECT. The urine OV-ES assay was also able to detect 28 individuals as positive from the 63 (44.4%) individuals previously determined to be negative using FECT. The likelihood of a positive diagnosis of OV infection by urine OV-ES assay increased significantly with the intensity of OV infection as determined by FECT. With reference to FECT, the sensitivity and specificity of the urine OV-ES assay was 81% and 70%, respectively., Conclusion: The detection of OV-infection by the urine OV-ES assay showed much greater diagnostic sensitivity and diagnostic specificity than the current "gold standard" FECT method for the detection and quantification of OV infection. Due to its ease-of-use, and noninvasive sample collection (urine), the urine OV-ES assay offers the potential to revolutionize the diagnosis of liver fluke infection and provide an effective tool for control and elimination of these tumorigenic parasites.

Free-grazing ducks play a major role in the rural economy of Eastern Asia in the form of egg and meat production. In Thailand, the geographical location, tropical climate conditions and wetland areas of the country are suitable for their husbandry. These environmental factors also favor growth, multiplication, development, survival, and spread of duck parasites. In this study, a total of 90 free-grazing ducks from northern, central, and northeastern regions of Thailand were examined for intestinal helminth parasites, with special emphasis on zoonotic echinostomes. Of these, 51 (56.7%) were infected by one or more species of zoonotic echinostomes, Echinostoma revolutum, Echinoparyphium recurvatum, and Hypoderaeum conoideum. Echinostomes found were identified using morphological criteria when possible. ITS2 sequences were used to identify juvenile and incomplete worms. The prevalence of infection was relatively high in each region, namely, north, central, and northeast region was 63.2%, 54.5%, and 55.3%, respectively. The intensity of infection ranged up to 49 worms/infected duck. Free-grazing ducks clearly play an important role in the life cycle maintenance, spread, and transmission of these medically important echinostomes in Thailand.

Opisthorchis viverrini is an important food-borne trematode in Southeast Asia. The infection causes significant morbidity in terms of hepatobiliary diseases and cholangiocarcinoma. The aim of this study was to improve the sensitivity of the PCR-based diagnosis of O. viverrini infection. A new fecal DNA extraction protocol for the detection of O. viverrini DNA using cetyltrimethyl-ammoniumbromide to remove PCR inhibitor was used and compared with the commercial stool kit method. The sensitivity of the new test was 79.3%, compared with the 44.8% of the previous method (P < 0.01). PCR-positive tests identified several cases judged parasite negative by the parasitological method (28.6%), indicating the new test's advantage in the diagnosis of individuals with light infections.