Multiplex RT Real-Time PCR Based on Target Failure to Detect and Identify Different Variants of SARS-CoV-2: A Feasible Method That Can Be Applied in Clinical Laboratories.

Shortly after its emergence, Omicron and its sub-variants have quickly replaced the Delta variant during the current COVID-19 outbreaks in Vietnam and around the world. To enable the rapid and timely detection of existing and future variants for epidemiological surveillance and diagnostic applications, a robust, economical real-time PCR method that can specifically and sensitively detect and identify multiple different circulating variants is needed. The principle of target- failure (TF) real-time PCR is simple. If a target contains a deletion mutation, then there is a mismatch with the primer or probe, and the real-time PCR will fail to amplify the target. In this study, we designed and evaluated a novel multiplex RT real-time PCR (MPL RT-rPCR) based on the principle of target failure to detect and identify different variants of SARS-CoV-2 directly from the nasopharyngeal swabs collected from COVID-19 suspected cases. The primers and probes were designed based on the specific deletion mutations of current circulating variants. To evaluate the results from the MPL RT-rPCR, this study also designed nine pairs of primers for amplifying and sequencing of nine fragments from the S gene containing mutations of known variants. We demonstrated that (i) our MPL RT-rPCR was able to accurately detect multiple variants that existed in a single sample; (ii) the limit of detection of the MPL RT-rPCR in the detection of the variants ranged from 1 to 10 copies for Omicron BA.2 and BA.5, and from 10 to 100 copies for Delta, Omicron BA.1, recombination of BA.1 and BA.2, and BA.4; (iii) between January and September 2022, Omicron BA.1 emerged and co-existed with the Delta variant during the early period, both of which were rapidly replaced by Omicron BA.2, and this was followed by Omicron BA.5 as the dominant variant toward the later period. Our results showed that SARS-CoV-2 variants rapidly evolved within a short period of time, proving the importance of a robust, economical, and easy-to-access method not just for epidemiological surveillance but also for diagnoses around the world where SARS-CoV-2 variants remain the WHO's highest health concern. Our highly sensitive and specific MPL RT-rPCR is considered suitable for further implementation in many laboratories, especially in developing countries.

Outcome prediction model and prognostic biomarkers for COVID-19 patients in Vietnam.

Background: Accurate prognosis is important either after acute infection or during long-term follow-up of patients infected by severe acute respiratory syndrome coronavirus 2. This study aims to predict coronavirus disease 2019 (COVID-19) severity based on clinical and biological indicators, and to identify biomarkers for prognostic assessment. Methods: We included 261 Vietnamese COVID-19 patients, who were classified into moderate and severe groups. Disease severity prediction based on biomarkers and clinical parameters was performed by applying machine learning and statistical methods using the combination of clinical and biological data. Results: The random forest model could predict with 97% accuracy the likelihood of COVID-19 patients who subsequently worsened to the severe condition. The most important indicators were interleukin (IL)-6, ferritin and D-dimer. The model could still predict with 92% accuracy after removing IL-6 from the analysis to generalise the applicability of the model to hospitals with limited capacity for IL-6 testing. The five most effective indicators were C-reactive protein (CRP), D-dimer, IL-6, ferritin and dyspnoea. Two different sets of biomarkers (D-dimer, IL-6 and ferritin, and CRP, D-dimer and IL-6) are applicable for the assessment of disease severity and prognosis. The two biomarker sets were further tested through machine learning algorithms and relatively validated on two Danish COVID-19 patient groups (n=32 and n=100). The results indicated that various biomarker sets combined with clinical data can be used for detection of the potential to develop the severe condition. Conclusion: This study provided a simple and reliable model using two different sets of biomarkers to assess disease severity and predict clinical outcomes in COVID-19 patients in Vietnam.

Anorectal manometry findings in relation with long-term functional outcomes of the patients operated on for Hirschsprung's disease compared to the reference-based population.

PURPOSE: This study investigated anorectal manometry (AM) findings and bowel function of patients operated on for Hirschsprung's disease (HD). METHODS: A cross-sectional study was conducted at Children's Hospital 2. Patients operated on for HD from January 2015 to January 2020 were reviewed. Their clinical characteristics, bowel function, and manometric findings were investigated and compared with the references. RESULTS: Ninety-five patients and 95 references were enrolled. Mean ages were 6.6 ± 2.2 years and 7.2 ± 2.9 years,; fecal incontinence rates were 25.3% and 2.1%, and constipation rates were 12.6% and 4.2 for the patients versus the references, respectively. Anal resting pressures were significantly decreased in the patients compared to the references (53.2 ± 16.1 mmHg versus 62.2 ± 14.0 mmHg; p < 0.05). Among the patients, the anal resting pressure was significantly decreased in the incontinents than in the continents (46.0 ± 10.6 mmHg versus 55.6 ± 16.9 mmHg, p < 0.05). During the sensation test, the value of maximum tolerated volume was significantly decreased in the incontinents than in the continents (135.9 ± 47.9 mL versus 166.6 ± 58.3 mL, p < 0.05). CONCLUSION: AM is an objective method providing beneficial information that could guide a more adapted management in HD patients with defecation disorders.

Nutrient dynamics and eutrophication assessment in the tropical river system of Saigon - Dongnai (southern Vietnam).

Saigon-Dongnai Rivers in Southern Vietnam is a complex lowland hydrological network of tributaries that is strongly influenced by the tidal cycles. The increasing economic, industrial and domestic developments in and around Ho Chi Minh City (HCMC) have led to serious impacts on water quality due to lack of appropriate wastewaters treatment. Drinking water production is impacted and the large aquaculture production areas may also be affected. We analyzed spatial and seasonal variability of nutrient concentrations (Phosphorus, Nitrogen and Silica) and eutrophication indicators (Organic Carbon, Chlorophyll-a and Dissolved Oxygen) based on bi-monthly monitoring during two hydrological cycles (July 2015-December 2017). Four monitoring sites were selected to assess the impact of HCMC: two upstream stations on the Saigon River and Dongnai River branches to provide the reference water quality status before reaching the urbanized area of HCMC; one monitoring station in the city center to highlight Saigon River water quality within the heart of the megacity; the fourth station downstream of the confluence to evaluate the impact of HCMC on the estuarine waters. This study points to excess nutrients in HCMC's water body with concentrations of NH4+ and PO43- averaging to 0.7 ±â€¯0.6 mgN L-1 and 0.07 ±â€¯0.06 mgP L-1, respectively in mean over the monitored period and rising up to 3 mgN L-1 and 0.2 mgP L-1, in extreme conditions. During the dry season, we evidenced that untreated domestic discharges leads to degradation of the Saigon River's water quality with extreme values of algal biomass (up 150 µChl-a L-1) and hypoxic conditions occurring episodically (DO < 2 mg L-1) in the heart of the megacity. Until now, eutrophication in the urban center has had no clear effect downstream because eutrophic water mass from the Saigon River is efficiently mixed with the Dongnai River and sea water masses during the successive semi-diurnal tidal cycles.

Long-Term Outcomes and Quality of Life in Patients after Soave Pull-Through Operation for Hirschsprung's Disease: An Observational Retrospective Study.

INTRODUCTION: Patients after pull-through operation for Hirschsprung's disease (HD) are at high risk of defecation disorders. This study aimed at investigating their long-term outcomes and quality of life (QoL) in comparison with controls. PATIENTS AND METHODS: Patients older than 5 years operated on for HD were interviewed to complete detailed questionnaires on bowel function. Patients without neurologic impairment were enrolled in a QoL survey to compare with controls matched for sex and age and selected randomly from the general population using sampling set in a ratio of four controls to one case of HD. RESULTS: In total, 53 operated patients were enrolled. Mean age of the patients was 16 ± 8 years, with 68% boys. Rectosigmoid aganglionosis was the most seen form of HD in 38 (72%) cases. Open Soave was performed in 40 (75.5%) cases, and minimally invasive surgery Soave (MIS Soave) in 13 (24.5%) cases. At investigation, prevalence of fecal incontinence and constipation were 22.6 and 13.2%, respectively. Regarding QoL survey, 45 patients and 180 controls were enrolled, excluding 8 patients with neurologic impairment. Thirty-seven (82.2%) patients were classified as having a good QoL (score ≥ 9 points); whereas six had a fair QoL (5-8 points) and two had a poor QoL (< 5 points). QoL score in the cases and the controls were 10.2 ± 2.5 and 11.9 ± 0.4 points, respectively. Long aganglionosis form of HD was significantly associated with a low QoL (score < 8 points), adjusted odds ratio = 9, 95% confidence interval [1.3; 64.1] (p < 0.05). In subscales analyses, the prevalence of each dimension including fecal continence, school absenteeism, unhappiness or anxiety, food restriction, and peer rejection was significantly higher in operated patients than in controls (p <0.001). CONCLUSION: Although the QoL of patients operated on for HD in general was with good outcomes, fecal incontinence and constipation still are problematic issues and challenges in a high percentage of patients. Therefore, a long-term and multidisciplinary follow-up is essentially required for these patients.

Rectal suction biopsy with calretinin immunohistochemistry in patients suspected with residual aganglionosis after operation for Hirschsprung disease.

INTRODUCTION: This study investigates the use of rectal suction biopsy (RSB) with calretinin immunohistochemical staining (CIS) in patients suspected of having abnormally innervated bowel after pull through operation for Hirschsprung disease (HD). METHOD: This study was conducted in Children's Hospital 2, Ho Chi Minh City from January 2015 through June 2016. Patients suspected with abnormally innervated bowel after pull through operation for HD were submitted for a RSB with CIS. Following histological results, the patients underwent an appropriate treatment (redo pull through operation or non-operative medical management) and followed up at least 6 months to evaluate their outcomes. RESULTS: Ten patients were enrolled in our study. The median age was 5.6 (range 1.4-20) years. The diagnosis of normally innervated bowel was made in five patients by showing positive reactivity of CIS on RSB, whereas five others were diagnosed with abnormally innervated bowel. Final diagnosis of the five latter patients was confirmed by analyses of the resected bowel after a redo pull through operation (including 4 cases with residual aganglionosis and one case with transition zone bowel). CONCLUSIONS: RSB with CIS is a reliable and simple method for diagnosis of abnormally innervated bowel after pull through operation for HD. LEVEL OF EVIDENCE: III TYPE OF STUDY: Study of diagnostic test (evaluate the effectiveness of a diagnostic test or outcome assessment).

Diagnostic value of rectal suction biopsies using calretinin immunohistochemical staining in Hirschsprung's disease.

BACKGROUND: The study investigates the diagnostic value of calretinin immunohistochemical staining (CIS) on rectal suction biopsies (RSB) in Hirschsprung's disease (HD). METHODS: A prospective study was conducted at Children's Hospital 2 in Ho Chi Minh City, Vietnam, from January through December 2015. Patients suspected of HD during this period underwent RSB and were followed in order to assess the accuracy of the diagnostic test with CIS compared with conventional histology (H&E). RESULTS: A total of 188 children with RSB were investigated. Median age was 7.1 (range 0.2-159) months with 65.4% boys. HD was confirmed in 80 (42.6%) children. There were 1 false positive and no false-negative cases. The sensitivity and specificity were 100% (80/80) and 99.1% (107/108) for CIS and 100% and 85.2% for H&E, respectively. Cohen's kappa coefficient was 0.9891 with a diagnostic accuracy of 99.5% for CIS, compared with 0.8303 and 91.5% for H&E, respectively. There were no serious complications related to the RSB. CONCLUSION: RSB with CIS is a useful diagnostic method for HD, with easy interpretation and no need for cryostat. CIS has a high diagnostic accuracy and should be considered as the primary method for the diagnosis of HD by RSB. LEVEL OF EVIDENCE: Diagnostic Studies - Level I.