A Simple Method to Detect SARS-CoV-2 in Wastewater at Low Virus Concentration.

Background: Since its initial appearance in December 2019, coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has spread globally. Wastewater surveillance has been demonstrated as capable of identifying infection clusters early. The purpose of this study was to investigate a quick and simple method to detect SARS-CoV-2 in wastewater in Thailand during the early stages of the second outbreak wave when the prevalence of the disease and the virus concentration in wastewater were low. Methods: Wastewater samples were collected from a hospital caring for patients with COVID-19 and from 35 markets, two of which were associated with recently reported COVID-19 cases. Then, samples were concentrated by membrane filtering prior to SARS-CoV-2 detection by RT-qPCR. Results: SARS-CoV-2 RNA was detected in the wastewater samples from the hospital; the Ct values for the N, ORF1ab, and S genes progressively increased as the number of patients admitted to the treatment floor decreased. Notably, the ORF1ab and S genes were still detectable in wastewater even when only one patient with COVID-19 remained at the hospital. SARS-CoV-2 RNA was detected in the wastewater samples from fresh market where COVID-19 cases were reported. Conclusions: Our findings suggest that wastewater surveillance for SARS-CoV-2 is sensitive and can detect the virus even in places with a high ambient temperature and relatively low prevalence of COVID-19.

Analysis of SCA8, SCA10, SCA12, SCA17 and SCA19 in patients with unknown spinocerebellar ataxia: a Thai multicentre study.

BACKGROUND: About 50 % of Thai patients with adult-onset spinocerebellar ataxia (SCA) was Machado-Joseph disease (MJD), SCA1, SCA2 and SCA6. The author investigated further on less common SCAs in the patients without any known mutations. METHODS: DNA samples of 82 index patients who were genetically excluded MJD, SCA1, SCA2, SCA6, SCA7 and dentatorubro-pallidoluysian atrophy (DRPLA) were examined. Analysis of SCA8, SCA10, SCA12, SCA17 and SCA19 genes were comprehensively performed. Normal range of trinucleotide repeat expansion sizes of TATA-box-binding protein gene (TBP) were also determined in 374 control subjects. RESULTS: Eight patients carried ≥42 CAG/CAA repeat allele in the TBP consistent with SCA17. The pathological repeat alleles ranged from 42 to 57 repeats. All patients had significant degree of cognitive dysfunction. Other non-ataxic phenotypes comprised of parkinsonism, chorea, dystonia and myoclonus. A sporadic patient carried a heterozygous 41-repeat allele developed chronic progressive cerebellar degeneration commenced at the age of 28 years. Whilst, 2 % of the control subjects (8/374) carried the 41-repeat allele. Five of the carriers were re-examined, and revealed that four of them had parkinsonism and/or cognitive impairment without cerebellar signs. Analysis of other types of SCAs was all negative. CONCLUSIONS: This is the first study of SCA8, SCA10, SCA12, SCA17 and SCA19 in Thais. SCA17 appears to be an important cause of ataxia in Thailand. Although, the pathological cut-off point of the TBP repeat allele remains unclear, the finding suggests that the 41-repeat may be a pathological allele resulting late-onset or mild phenotype. Apart from ataxia, cognitive impairment and parkinsonism may be clinical presentations in these carriers.

Confirmation of the association between LRRK2 R1628P variant and susceptibility to Parkinson's disease in the Thai population.

OBJECTIVE: LRRK2 p.R1628P (c.4883G > C) is associated with Parkinson's disease (PD) in Chinese and Thais. However, some studies in other East Asian ethnic groups did not observe this association. Carriers of p.R1628P are about 3-5% Chinese and Thais. In contrast, Japanese, Koreans and Malays are much less prevalent (0-<1%). The contradictory results may be caused by insufficient sample sizes especially studies in ethnic groups with low prevalence, which, theoretically need a much larger sample size. We conducted a case-control Thai PD study with appropriate size in order to support the role of p.R1628P related to susceptibility to PD. METHODS: Estimated total sample size of 958 Thai subjects was needed. 485 PD patients and 480 controls were recruited. The p.R1628P was screened by RFLP and confirmed by direct sequencing. Clinical characteristics were compared between PD patients with and without p.R1628P. RESULTS: 54 PD patients (11%) and 29 control subjects (6%) carried p.R1628P. Multiple logistic regression analysis showed that GC and CC genotypes were significantly higher in PD patients than in controls (OR = 1.81, 95%CI = 1.10-2.97). The PD patients carrying p.R1628P had earlier age at onset (56 ± 13 vs 60 ± 12; P = 0.021) and a more rapidly progressive course (P < 0.001) than the patients carrying wild-type nucleotide. CONCLUSIONS: We confirm the association between p.R1628P and risk of developing PD in the appropriated sample-sized cohort. Certain LRRK2 variants appear to be generally distributed among East Asians, however, in widely different frequencies. In order to study role of such variants in PD, it should be carefully estimated the appropriate sample size.

Glucocerebrosidase mutations in Thai patients with Parkinson's disease.

BACKGROUND: GBA mutations are an important risk factor in developing Parkinson's disease (PD) worldwide. The study aimed to determine the frequency and clinical characteristics of GBA mutations in a Thai PD cohort of 480 patients and 395 control subjects. METHODS: Direct sequencing of GBA was performed in all early-onset PD patients (EOPD: n = 108) and 100 PD patients with age at onset over 50 years (AAO > 50y-PD). The study subsequently screened all identified mutations in the remaining AAO > 50y-PD patients and all control subjects. Predictive factors associated with risk of developing PD were analyzed. Comparisons of clinical characteristics of PD patients with and without GBA mutations were also carried out. RESULTS: Heterozygous GBA mutations were identified in 24 patients (5%) and 2 controls (0.5%). Seven identified GBA point mutations comprised p.L444P, p.N386K, p.P428S, IVS2+1G > A, IVS9+3G > C, IVS10-9_10GT > AG and c.1309delG, of which five mutations were novel. Multiple logistic regression analysis revealed that GBA mutations were more frequent in EOPD than AAO > 50y-PD groups (OR = 4.64, P < 0.022). Patients with GBA mutations had mean age at onset (43.1 ± 10.2, mean ± standard deviation) earlier than patients without GBA mutations (54.4 ± 13.9, P = 0.002). The patients with GBA mutations also had a more rapid progressive course, in which they were more likely to have higher Hoehn and Yahr staging (OR = 4.20, P = 0.006) and slightly lower means of Schwab-England ADL score [74.1 ± 17.1 vs. 81.0 ± 18.08 (OR = 0.98, 95%CI = 0.96-1.01, P = 0.162)]. CONCLUSION: GBA mutations are an important risk of PD in the Thai population. Patients having the mutations are likely to have early onset and may exhibit more rapid motor progression.

Clinical analysis of adult-onset spinocerebellar ataxias in Thailand.

BACKGROUND: Non-ataxic symptoms of spinocerebellar ataxias (SCAs) vary widely and often overlap with various types of SCAs. Duration and severity of the disease and genetic background may play a role in such phenotypic diversity. We conducted the study in order to study clinical characteristics of common SCAs in Thailand and the factors that may influence their phenotypes. METHODS: 131 (49.43%) out of 265 Thai ataxia families with cerebellar degeneration had positive tests for SCA1, SCA2, Machado-Joseph disease (MJD) or SCA6. The study evaluated 83 available families including SCA1 (21 patients), SCA2 (15), MJD (39) and SCA6 (8). Comparisons of frequency of each non-ataxic sign among different SCA subtypes were analysed. Multivariate logistic regression analyses were undertaken to analyze parameters in association with disease severity and size of CAG repeat. RESULTS: Mean ages at onset were not different among patients with different SCAs (40.31 ± 11.33 years, mean ± SD). Surprisingly, SCA6 patients often had age at onset and phenotypes indistinguishable from SCA1, SCA2 and MJD. Frequencies of ophthalmoparesis, nystagmus, hyperreflexia and areflexia were significantly different among the common SCAs, whilst frequency of slow saccade was not. In contrast to Caucasian patients, parkinsonism, dystonia, dementia, and facial fasciculation were uncommon in Thai patients. Multivariate logistic regression analysis demonstrated that ophthalmoparesis (p < 0.001) and sensory impairment (p = 0.025) were associated with the severity of the disease. CONCLUSIONS: We described clinical characteristics of the 4 most common SCAs in Thailand accounting for almost 90% of familial spinocerebellar ataxias. There were some different observations compared to Caucasian patients including earlier age at onset of SCA6 and the paucity of extrapyramidal features, cognitive impairment and facial fasciculation. Severity of the disease, size of the pathological CAG repeat allele, genetic background and somatic heterogeneity of pathological alleles may influence clinical expressions of these common SCAs.

Prevalence, clinical features and treatment outcomes of patients with myasthenia gravis positive for antibodies to muscle-specific kinase in Thailand.

A small but variable subgroup of patients with myasthenia gravis (MG) who have antibodies to muscle-specific kinase (MuSKAb-MG) can present with distinct phenotypes and are often treatment-resistant. The prevalence, clinical phenotypes and outcomes of treatment of patients with MuSKAb-MG in Thailand were determined. Eight (16.3%) of the 49 patients with generalized MG who were negative for acetylcholine receptor antibodies (AChRAb) were positive for muscle-specific kinase antibodies. Most patients had predominant oculobulbar features and respiratory failure occurred in three. At follow up, three out of the seven patients who underwent thymectomy were in complete stable remission and four had improved and were on reduced immunosuppression medication, suggesting a possible benefit of thymectomy.

Prophylactic botulinum type A toxin complex (Dysport®) for migraine without aura.

OBJECTIVE: To evaluate the efficacy, safety, and optimum dose of a highly purified Clostridium botulinum type A toxin-hemagglutinin complex (Dysport) for migraine prophylaxis. BACKGROUND: Botulinum toxin type-A has demonstrated good efficacy in several open-label studies of patients with migraine, involving either individualized or standardized protocols, although data from placebo-controlled trials have been conflicting. METHODS: A 12-week, double-blind, randomized trial of Dysport (120 or 240 units) vs placebo was conducted in 6 centers in Thailand to evaluate the efficacy, safety, and optimum dose of botulinum toxin type-A (Dysport) for migraine prophylaxis. A total of 128 patients with migraine without aura were enrolled. The primary end point was the change in the mean number of migraine attacks per 4-week period from the pre-treatment period to 8-12 weeks post injection. Secondary efficacy measures included the change in the mean total intensity score from the pre-treatment period to 8-12 weeks, the investigator and patient global assessments of change at each visit compared with pre-treatment, and Migraine Disability Assessment and Short Form-36 scores. RESULTS: Change in number of migraine attacks from pre-treatment to weeks 8-12 was not significantly different. There was a greater improvement in total intensity score at weeks 8-12 with Dysport-240 (not significant), and interim visit data showed that this was significant at weeks 0-4 (P = .03 Dysport-240 vs placebo). The mean duration of headache during weeks 0-4 was lower with Dysport-240 (P = .04 vs placebo). Improvements in patient and investigator global assessments of change between weeks 0-4 and 8-12 were significant for the Dysport-240 group (both P < .05 vs placebo). CONCLUSIONS: Limitations in study design and assessment tools employed may have contributed to the inconclusive nature of the primary end point data. Dysport-240 showed significant benefit over placebo at some end points and further trials with more appropriate outcome measures are required to evaluate effectively this treatment.