Development and field validation of a reverse transcription loop-mediated isothermal amplification assay (RT-LAMP) for the rapid detection of chikungunya virus in patient and mosquito samples.

OBJECTIVES: We aimed to develop a reverse transcription loop-mediated isothermal amplification (RT-LAMP) platform for the rapid detection of chikungunya virus (CHIKV) in both patient and mosquito samples from Brazil. METHODS: We optimized an RT-LAMP assay and then evaluated the specificity and sensitivity using visual detection. In comparison with the RT-qPCR reference method, we validated the utility of this assay as a molecular diagnostic test in a reference laboratory for arbovirus diagnostics using 100 serum samples collected from suspected CHIKV cases. RESULTS: Our RT-LAMP assay specifically detected CHIKV without cross-reactivity against other arboviruses. The limit of detection of our RT-LAMP was estimated in -1.18 PFU (confidence interval [CI] ranging from -2.08 to 0.45), resulting in a similar analytical sensitivity when directly compared with the reference standard RT-qPCR assay. Then, we demonstrate the ability of our RT-LAMP assay to detect the virus in different human specimens (serum, urine, and saliva), and crude lysate of Aedes aegypti mosquitoes in as little as 20-30 minutes and without a separate RNA isolation step. Lastly, we showed that our RT-LAMP assay could be lyophilized and reactivated by adding water, indicating potential for room-temperature storage. Our RT-LAMP had a clinical sensitivity of 100% (95% CI, 90.97-100.00%), clinical specificity of 96.72% (95% CI, 88.65-99.60%), and overall accuracy of 98.00% (95% CI, 92.96-99.76%). DISCUSSION: Taken together, these findings indicate that the RT-LAMP assay reported here solves important practical drawbacks to the deployment of molecular diagnostics in the field and can be used to improve testing capacity, particularly in low- and middle-income countries.

Simultaneous Circulation of DENV, CHIKV, ZIKV and SARS-CoV-2 in Brazil: an Inconvenient Truth.

The emergence of SARS-CoV-2 has prompted the mobilization of a network of public health laboratories to diagnose COVID-19 patients, trace contacts and identify hot-spot areas for active community transmission at the expense of arbovirus diagnosis and control practices. In this article, we discuss the unprecedented challenges faced by the Brazilian public health system in dealing with the incursion of SARS-CoV-2 in the midst of ongoing triple arboviral epidemics caused by dengue, chikungunya, and Zika virus. Finally, we highlight the importance of the introduction of one health approach as an effective inter-disciplinary response and management to mitigate the catastrophic effect caused by these pathogens.

Survival analysis in non-congenital neurological disorders related to dengue, chikungunya and Zika virus infections in Northeast Brazil.

The Northeast of Brazil has experienced a triple epidemic, with the simultaneous circulation of dengue virus (DENV), chikungunya virus (CHIKV) and Zika virus (ZIKV), which may have contributed to the observed increase across this region of atypical forms of disease and deaths. In view of this fact, non-congenital neurological disorders related to arboviruses were compared with other etiologies, mortality and survival rates of patients admitted to referral neurology hospitals in Pernambuco State, Northeast Brazil, from 2015 to 2018. Blood and cerebrospinal fluid samples were collected and tested using molecular and serological assays. The arbovirus-exposed groups were compared with respect to epidemiological, clinical and neurologic characteristics by using the Pearson's chi-square test. For the survival analysis, the Kaplan-Meier and Hazard Ratio (HR) tests were used, with a 95% confidence interval (CI). Encephalitis and encephalomyelitis were more frequent in arboviruses, while myelitis predominated in the neurological disorders of other etiologies. Guillain-Barré Syndrome (GBS) was similarly distributed amongst the groups. Exposure to one of the arboviruses caused a six-fold increase in the risk of death (HR: 6.37; CI: 2.91 - 13.9). Amongst the arbovirus-exposed groups, infection (DENV/CHIKV) increased nine times the risk of death (HR: 9.07; CI: 3.67 - 22.4). The survival curve indicates that have been exposed to some arbovirus decreased the likelihood of survival compared to those with other etiologies (Log-Rank: p<0.001). Within this scenario, neurologic manifestations of DENV, CHIKV and ZIKV have the potential to increase mortality and decrease survival, and concomitant infection (DENV/CHIKV) is an aggravating factor in reducing the likelihood of survival when compared to monoinfections.

Epidemiological and clinical characteristics of the first 557 successive patients with COVID-19 in Pernambuco state, Northeast Brazil.

BACKGROUND: South America is the current epicenter of COVID-19 pandemic. Yet, the epidemiological and clinical features of the disease have not been described in Brazil, the third most affected country in the world. METHODS: In this retrospective study, we describe the demographics, epidemiology and clinical features of the first 557 consecutive patients positive for SARS-CoV-2 living in Pernambuco state, Northeast Brazil. RESULTS: The first COVID-19 cases occurred in the high income population. The age of infected patients ranged from 27 days to 97 years with a median of 47 years. The ratio of males to female in the SARS-CoV-2-infected group was 0.83:1. The most common symptom was cough (74.51%), followed by fever (66.79%), dyspnea (56.01%), sore throat (28.19%) and O2 saturation <95% (24.42%). 86.44% of the lethal cases were patients older than 51 years. The median time from illness onset to diagnosis was 4.0 days (range 0-39 days) Severe patients diagnosed after 14 days of symptoms onset had higher viral load than patients with mild disease. CONCLUSIONS: Our study provides important information about COVID-19 in the tropics and will assist physicians and health officials to face the current pandemics as SARS-CoV-2 continues to spread in the human population.