Burden of Chikungunya Virus Infection during an Outbreak in Myanmar.

Chikungunya virus (CHIKV) infection is a re-emerging arboviral disease with no approved vaccine, although numerous options are in development. Before vaccine implementation, disease burden, affected age group, and hospitalization rate information should be documented. In 2019, a sizeable outbreak of the East Central South African genotype of CHIKV occurred in Myanmar, and during this period, a cross-sectional study was conducted in two regions, Mandalay and Yangon, to examine the molecular and seropositivity rate of the CHIKV infection. The participants (1124) included dengue-suspected pediatric patients, blood donors, and healthy volunteers, who were assessed using molecular assays (quantitative real-time RT-PCR), serological tests (anti-CHIKV IgM capture and IgG indirect enzyme-linked immunosorbent assays), and neutralization tests. The tests confirmed the following positivity rates: 11.3% (127/1124) for the molecular assay, 12.4% (139/1124) for the anti-CHIKV IgM Ab, 44.5% (500/1124) for the anti-CHIKV IgG Ab, and 46.3% (520/1124) for the CHIKV neutralizing Ab. The highest rate for the molecular test occurred with the dengue-suspected pediatric patients. The seroprevalence rate through natural infection was higher in the healthy volunteers and blood donors than that in the pediatric patients. The results of this study will help stakeholders determine the criteria for choosing appropriate recipients when a CHIKV vaccine is introduced in Myanmar.

Coinfection and circulation of chikungunya virus and dengue virus in pediatric patients in Myanmar, 2019.

Myanmar is an endemic country for arboviruses, and outbreaks occur frequently. A cross-sectional analytical study was conducted during the peak season of the chikungunya virus (CHIKV) outbreak in 2019. A total of 201 patients with acute febrile illness who were admitted to the 550-bedded Mandalay Children Hospital in Myanmar were enrolled in the study, and virus isolation, serological tests, and molecular tests for the dengue virus (DENV) and CHIKV were performed for all samples. Out of 201 patients, 71 (35.3%) were only DENV-infected, 30 (14.9%) were only CHIKV-infected and 59 (29.4%) were coinfected with DENV and CHIKV. The viremia levels of the DENV- and CHIKV- mono-infected groups were significantly higher than those of the group coinfected with DENV and CHIKV. Genotype I of DENV-1, genotypes I and III of DENV-3, genotype I of DENV-4 and the East/Central/South African genotype of CHIKV were co-circulating during the study period. Two novel epistatic mutations of CHIKV (E1:K211E and E2:V264A) were noted. This study highlighted that there were many coinfection cases during the outbreak and that the co-circulation of both viruses in DENV-endemic regions warrants effective monitoring of these emerging pathogens via comprehensive surveillance to facilitate the implementation of effective control measures.

Evidence of Chikungunya virus seroprevalence in Myanmar among dengue-suspected patients and healthy volunteers in 2013, 2015, and 2018.

INTRODUCTION: Chikungunya virus (CHIKV) is a mosquito-borne virus known to cause acute febrile illness associated with debilitating polyarthritis. In 2019, several institutions in Myanmar reported a CHIKV outbreak. There are no official reports of CHIKV cases between 2011 and 2018. Therefore, this study sought to determine the seroprevalence of CHIKV infection before the 2019 outbreak. METHODS: A total of 1,544 serum samples were collected from healthy volunteers and patients with febrile illnesses in Yangon, Mandalay, and the Myeik district in 2013, 2015, and 2018. Participants ranged from one month to 65 years of age. Antibody screening was performed with in-house anti-CHIKV IgG and IgM ELISA. A neutralization assay was used as a confirmatory test. RESULTS: The seroprevalence of anti-CHIKV IgM and anti-CHIKV IgG was 8.9% and 28.6%, respectively, with an overall seropositivity rate of 34.5%. A focus reduction neutralization assay confirmed 32.5% seroprevalence of CHIKV in the study population. Age, health status, and region were significantly associated with neutralizing antibodies (NAbs) and CHIKV seropositivity (p < 0.05), while gender was not (p = 0.9). Seroprevalence in 2013, 2015, and 2018 was 32.1%, 28.8%, and 37.3%, respectively. Of the clinical symptoms observed in participants with fevers, arthralgia was mainly noted in CHIKV-seropositive patients. CONCLUSION: The findings in this study reveal the circulation of CHIKV in Myanmar's Mandalay, Yangon, and Myeik regions before the 2019 CHIKV outbreak. As no treatment or vaccine for CHIKV exists, the virus must be monitored through systematic surveillance in Myanmar.

An Outbreak of Dengue Virus Serotype 2 Cosmopolitan Genotype in Nepal, 2017.

Dengue virus (DENV) is one of the most prevalent neglected tropical diseases, with half of the world's population at risk of infection. In Nepal, DENV was first reported in 2004, and its prevalence is increasing every year. The present study aimed to obtain and characterize the full-length genome sequence of DENV from the 2017 outbreak. Hospital-based surveillance was conducted in two provinces of Nepal during the outbreak. Acute-phase serum samples were collected from 141 clinically suspected dengue patients after the rainy season. By serological and molecular techniques, 37 (26.9%) and 49 (34.8%), respectively, were confirmed as dengue patients. The cosmopolitan genotype of DENV-2 was isolated from 27 laboratory-confirmed dengue patients. Genomic analysis showed many amino acid substitutions distributed mainly among the E, NS3, and NS5 genes. Phylogenetic analyses of the whole genome sequence revealed two clades (Asian and Indian) among DENV-2 isolates from Nepal. The DENV isolates from hilly and Terai areas were similar to Asian and Indian strains, respectively. Further genomic study on different DENV serotypes is warranted to understand DENV epidemics in Nepal, where there are limited scientific resources and infrastructure.

Detection of genotype-1 of dengue virus serotype 3 for the first time and complete genome analysis of dengue viruses during the 2018 epidemic in Mandalay, Upper Myanmar.

BACKGROUND: Dengue (DEN) is a neglected tropical disease, and surveillance of dengue virus (DENV) serotypes and genotypes is critical for the early detection of outbreaks. Risk factors for outbreaks include the emergence of new genotypes and serotype shifting. METHODOLOGY AND PRINCIPAL FINDINGS: To understand the genomic and viral characteristics of DENV-infected patients, we conducted a cross-sectional descriptive study among pediatric patients admitted at the 550-bedded Mandalay Children Hospital during the 2018 DEN endemic season. We conducted virus isolation, serological tests, viremia level measurement, and whole-genome sequencing. Among the 202 serum samples, we detected 85 samples with DENV (46 DENV-1, 10 DENV-3, 26 DENV-4 and three multiple serotype co-infections) via reverse transcription quantitative/real-time PCR (RT-qPCR), and we obtained 49 DENV isolates (31 DENV-1, 10 DENV-3 and 8 DEN-4). We did not detect DENV-2 in this study. The viral genome levels in serum did not differ significantly among virus serotypes, infection status (primary versus secondary) and disease severity. Based on the phylogenetic analysis, we identified DENV-1 genotype-1, DENV-4 genotype-1 and DENV-3 genotype-3 and genotype-1 which was detected for the first time. Next-generation sequencing analysis revealed greater frequencies of nonsynonymous and synonymous mutations per gene in the nonstructural genes. Moreover, mutation rates were also higher among DENV-1. CONCLUSION/SIGNIFICANCE: In conclusion, there was an increasing trend of DENV-3 cases during DENV endemic season in 2018 with the first detection of the genotype 1. However, DENV-1 has remained the predominant serotype in this study area since 2013, and we identified stop codon mutations in the DENV-1 genome. This report is the first to feature a complete genome analysis of the strains of DENV-3 and DENV-4 circulating among pediatric patients in Myanmar. This study highlighted the importance of annual surveillance for a better understanding of the molecular epidemiology of DENVs.

Effectiveness of the SA 14-14-2 Live-Attenuated Japanese Encephalitis Vaccine in Myanmar.

Myanmar is an endemic country for the Japanese encephalitis virus (JEV), and the SA-14-14-2 live-attenuated JEV vaccine was first introduced as a catch-up vaccination campaign in 2017. To determine the effectiveness of vaccination by means of neutralizing antibody titers against JEV, a cross-sectional descriptive study was conducted among five to 15-year-old monastic school children in Mandalay, Myanmar. A total of 198 students who had received vaccines were recruited, and single-time investigation of anti-JEV IgG and neutralizing antibodies against wild-type JEV were determined using anti-JEV IgG ELISA and plaque reduction neutralization tests (PRNT50). All students 100% (198/198) showed positive results on the anti-JEV IgG ELISA, and 87% (172/198) of the students had neutralizing antibodies against JEV six months after immunization. The geometric mean titers of both IgG antibodies and neutralizing antibodies increased with the participants' age groups, and statistically significant differences in anti-JEV IgG titers were noted across age groups. In this study, we could not investigate the persistence of neutralizing antibodies as only single-time blood collection was done. This study, which is the first report of JEV vaccination among children in Myanmar, showed similar neutralizing antibody production rates among vaccinated individuals as did studies in other countries.

Surge of severe acute respiratory syndrome coronavirus 2 infections linked to single introduction of a virus strain in Myanmar, 2020.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is a major health concern globally. Genomic epidemiology is an important tool to assess the pandemic of coronavirus disease 2019 (COVID-19). Several mutations have been reported by genome analysis of the SARS-CoV-2. In the present study, we investigated the mutational and phylogenetic analysis of 30 whole-genome sequences for the virus's genomic characteristics in the specimens collected in the early phase of the pandemic (March-June, 2020) and the sudden surge of local transmission (August-September, 2020). The four samples in the early phase of infection were B.6 lineage and located within a clade of the samples collected at the same time in Singapore and Malaysia, while five returnees by rescue flights showed the lineage B. 1.36.1 (three from India), B.1.1 (one from India) and B.1.80 (one from China). However, there was no evidence of local spread from these returnees. Further, all 19 whole-genome sequences collected in the sudden surge of local transmission showed lineage B.1.36. The surge of the second wave on SARS-CoV-2 infection was linked to the single-introduction of a variant (B.1.36) that may result from the strict restriction of international travel and containment efforts. These genomic data provides the useful information to disease control and prevention strategy.

Chikungunya Virus Infection in Blood Donors and Patients During Outbreak, Mandalay, Myanmar, 2019.

In 2019, an outbreak of chikungunya virus infection occurred in Mandalay, Myanmar, and 3.2% of blood donors and 20.5% of patients who were children were confirmed as being infected. The prevalence rate was up to 6.3% among blood donors. The East Central/South African genotype was predominantly circulating during this outbreak.

Inapparent dengue virus infection among students in Mandalay, Myanmar.

BACKGROUND: A school- and laboratory-based cross-sectional descriptive study was conducted to find out the burden of inapparent dengue virus (DENV) infection in Mandalay where DENV is endemic and there is circulation of all four DENV serotypes. METHODS: A total of 420 students who had no history of fever and visited the hospital within 6 months were recruited from three monastic schools. Serum samples were collected and the DENV genome was checked by conventional one-step RT-PCR and anti-DENV IgM and IgG antibodies were determined. Inapparent dengue (DEN) infection is defined as individuals who were either RT-PCR-positive or anti-DENV IgM-positive with no clinical manifestations or mild symptoms, and which are not linked to a visit to a healthcare provider. RESULTS: Among 420 students, 38 students (9.0%, 95% CI, 6.4 to 12.2) were confirmed as recent inapparent DEN infection. The DENV serotype-1 was detected in six students. Thirty-one out of 38 (81.6%) laboratory-confirmed inapparent DEN-infected students had primary infections and seven (18.4%) had secondary infections. CONCLUSION: This study explored the prevalence of inapparent DEN infection rate in urban monastic schools in Mandalay and showed that the rate of primary infection among inapparent DENV-infected children was high.

Evaluation of commercially available three dengue rapid diagnostic test kits for diagnosis of acute dengue virus infection at the point-of-care setting in Myanmar.

Early and accurate diagnosis of dengue virus (DENV) infection is very important and Rapid Diagnostic Test (RDT) Kits are been used as a point-of-care test to check DENV infection. A Hospital and Laboratory-based descriptive study was conducted at 550-bedded Mandalay Children Hospital in 2018. Acute-phase serum samples were collected from 202 dengue suspected patients to evaluate the efficacy of RDT Kits for the diagnosis of DENV infection. Commercially available three test kits which include: ((i) CareUs Dengue Combo, Korea, (ii) Humasis Dengue Combo, Korea and (iii) Wondfo Dengue Combo, China) were validated against WHO-based reference standard tests. 140/202 patients (69.3%) was confirmed to have DENV infection. All four serotypes of dengue viruses (57 DENV-1, 7 DENV-2, 6 DENV-3 and 10 DENV-4) were identified from 80 dengue confirmed patients and DENV-1 was the dominant serotype. Combining the NS-1 antigen and IgM antibody results from the CareUs Dengue Combo Kit gave the best sensitivity (92.1%, 95% CI 86.4%-96.0%) and specificity (75.8%, 95%CI 63.3%-85.8%). Among the three RDT Kits, the performance of CareUS Kit was better than the other two. This study explored the evidence of the usefulness of RDT Kits at the point-of-care setting for diagnosis of acute dengue infection.

Japanese Encephalitis- and Dengue-Associated Acute Encephalitis Syndrome Cases in Myanmar.

This study was conducted to find the burden of dengue virus (DENV) and Japanese encephalitis virus (JEV) among children under the age of 13, who presented with acute encephalitis syndrome at Mandalay Children Hospital in Myanmar in 2013. Molecular and serological investigations were performed on 123 cerebrospinal fluid (CSF) samples collected from these patients. By neutralization tests and/or virus isolation, four (3.3%) JEV- and one DENV-associated encephalitis cases (0.8%) were confirmed. Antibody titer against JEV Genotype 3 was the highest among the laboratory-confirmed JEV cases. One strain of DENV-1 with Genotype 1 was isolated from the CSF sample of the dengue encephalitis patient; this was similar to the virus circulating in the study area and neighboring countries. This study shows that flaviviruses are important pathogens causing encephalitis in Myanmar. Active disease surveillance, vector control, and vaccination programs should be enforced to reduce the morbidity and mortality caused by flavivirus encephalitis.

Comparison of Neutralizing Antibody Titers against Japanese Encephalitis Virus Genotype V Strain with Those against Genotype I and III Strains in the Sera of Japanese Encephalitis Patients in Japan in 2016.

Japanese encephalitis (JE) is an acute viral disease caused by the Japanese encephalitis virus (JEV). JEV strains are classified into 5 genotypes (I-V). JEV genotype V strains have never been detected in Japan to date, but they were recently detected in South Korea. In the present analysis, we tried to determine if a JEV genotype V strain caused any JE case in Japan in 2016. Serum and cerebrospinal fluid samples were collected from 10 JE patients reported in Japan in 2016. JEV RNA was not detected in any of the samples. Although JEV is a single-serotype virus, it can be expected that the neutralizing antibody titers against JEV genotype V strains are higher than those against genotype I and III strains in the serum of patients with JE in Japan whose causative JEV was the genotype V strain. The neutralizing antibody titers against the JEV genotype V strain were not higher than those against the genotype I or III strain in any serum samples. Therefore, the evidence that the JEV genotype V strain caused any JE case in Japan in 2016 was absent.

Detection of Zika Virus Infection in Myanmar.

Zika virus (ZIKV), an emerging mosquito-borne flavivirus, causes a dengue-like infection that has recently caught global attention. The infection, which also includes some birth defects, has been documented in the Americas, Pacific Islands, and some parts of Africa and Asia. There are no published reports on local ZIKV transmission in Myanmar. In this study, a total of 462 serum samples from patients and asymptomatic persons were collected in Myanmar from 2004 to 2017. They were analyzed for ZIKV infection by immunoglobulin M capture enzyme-linked immunosorbent assay (ELISA), immunoglobulin G indirect ELISA, neutralization test, real-time polymerase chain reaction (PCR), and conventional PCR. Our study confirmed ZIKV infection in 4.9% of patients with clinical dengue symptoms and in 8.6% of persons who were asymptomatic. This is the first report on ZIKV infection in Myanmar and it suggests the occurrence of ZIKV infection in two geographically distinct sites in this country since at least 2006.

Characterization of the 2013 dengue epidemic in Myanmar with dengue virus 1 as the dominant serotype.

In 2013 in Myanmar, dengue epidemic occurred with 20,255 cases including 84 deaths. This study aimed to determine the serological and molecular characteristics of dengue virus (DENV) infection among children with clinical diagnosis of dengue hemorrhagic fever (DHF) or dengue shock syndrome (DSS) during this period. Single acute serum samples were collected from 300 children in Mandalay Children Hospital, Mandalay, Myanmar. Out of the 300 children, 175 (58.3%) and 183 (61%) were positive for anti-dengue IgM and anti-dengue IgG, respectively. Among the IgM positives, 41 (23.4%) had primary DENV infection. Thirty-nine DENV strains (23 DENV-1, 10 DENV-2 and 6 DENV-4) were successfully isolated after inoculation of the patient serum samples onto C6/36 cells. DENV 1 was the dominant serotype in the 2013 epidemic. There was no correlation between the infecting serotypes and clinical severities. The DENV-1 strains belonged to three lineages of the genotype 1; the DENV-2 strains were of the Asian I genotype and were separated into two lineages; and DENV-4 strains belonged to the same lineage of genotype I. It is of interest to note the diversity of DENV-1 and -2 circulating in the same location during June-August 2013. These DENV isolates were genetically close (98%-100%) to the other previously reported isolates from Myanmar and its neighboring countries, namely China, Thailand, Sri Lanka, Cambodia and Vietnam. Primary DENV infection was still high among the severe dengue cases. Different serotypes of DENV were co-circulating in 2013, however, genotype shift was not observed. Additionally, amino acid mutations were detected in the study strains not seen in the previously reported strains from other countries and Myanmar. This paper provided information on the circulating serotypes for the last 15years and the recent dengue situation in Mandalay, Myanmar after 2006.

Detection of east/central/south African genotype of chikungunya virus in Myanmar, 2010.

In 2010, chikungunya virus of the East Central South African genotype was isolated from 4 children in Myanmyar who had dengue-like symptoms. Phylogenetic analysis of the E1 gene revealed that the isolates were closely related to isolates from China, Thailand, and Malaysia that harbor the A226V mutation in this gene.

Promoting antenatal care services for early detection of pre-eclampsia.

A prospective, quasi-experimental study was carried out in 2009 at urban health centres (UHCs) of five townships of Mandalay, Myanmar, to improve the skill of midwives (MWs) in diagnosis and referral of pre-eclampsia (PE) from UHC to the Central Women's Hospital (CWH) and to enhance the supervision of midwives by lady health visitors (LHVs). The intervention was training on quality antenatal care focusing on PE using an updated training manual. Altogether, 75 health care providers (MWs & LHVs) participated. In this study, data were extracted from patient registers and monthly reports of UHCs and CWH. Interviewers were trained regarding the conduct of semi-structured questionnaires to elicit knowledge and to use checklists in observation of skills in screening of PE, measuring blood pressure and urine protein (dipstick test). A guide for LHVs was also used to obtain data, and data was collected six months prior to and after the intervention. Significant improvements from baseline to endline survey occurred in the knowledge (p<0.001) and skill levels (p<0.001) including skills for screening, measuring blood pressure and urine protein. At CWH, there was an increase in referred cases of PE after the intervention, from 1.25% to 2.56% (p<0.001). In conclusion, this study highlights the early detection of pre-eclampsia by widespread use of quality antenatal care, education and training of health-care providers to improve their performance and increase human resources for health care, in order to enable women in our society to have healthy pregnancies and healthy babies.