Monitoramento das Produções

by R. Tedjo Sasmono, Anne-Frieda Taurel, Ari Prayitno, Hermin Sitompul, Benediktus Yohan, Rahma F. Hayati, Alain Bouckenooghe, Sri Rezeki Hadinegoro, Joshua Nealon
Background Dengue is a febrile illness transmitted by mosquitoes, causing disease across the tropical and sub-tropical world. Antibody prevalence data and serotype distributions describe population-level risk and inform public health decision-making. Methodology/Principal findings In this cross-sectional study we used data from a pediatric dengue seroprevalence study to describe historical dengue serotype circulation, according to age and geographic location. A sub-sample of 780 dengue IgG-positive sera, collected from 30 sites across urban Indonesia in 2014, were tested by the plaque reduction neutralization test (PRNT) to measure the prevalence and concentration of serotype-specific neutralizing antibodies according to subject age and geography. PRNT results were obtained from 776 subjects with mean age of 9.6 years. 765 (98.6%) neutralized one or more dengue serotype at a threshold of >10 (1/dil). Multitypic profiles were observed in 50.9% of the samples; a proportion which increased to 63.1% in subjects aged 15–18 years. Amongst monotypic samples, the highest proportion was reactive against DENV-2, followed by DENV-1, and DENV-3, with some variation across the country. DENV-4 was the least common serotype. The highest anti-dengue antibody titers were recorded against DENV-2, and increased with age to a geometric mean of 516.5 [1/dil] in the oldest age group. Conclusions/Significance We found that all four dengue serotypes have been widely circulating in most of urban Indonesia, and more than half of children had already been exposed to >1 dengue serotype, demonstrating intense transmission often associated with more severe clinical episodes. These data will help inform policymakers and highlight the importance of dengue surveillance, prevention and control.

by Thomas J. Hladish, Carl A. B. Pearson, Diana Patricia Rojas, Hector Gomez-Dantes, M. Elizabeth Halloran, Gonzalo M. Vazquez-Prokopec, Ira M. Longini
Background Historically, mosquito control programs successfully helped contain malaria and yellow fever, but recent efforts have been unable to halt the spread of dengue, chikungunya, or Zika, all transmitted by Aedes mosquitoes. Using a dengue transmission model and results from indoor residual spraying (IRS) field experiments, we investigated how IRS-like campaign scenarios could effectively control dengue in an endemic setting. Methods and findings In our model, we found that high levels of household coverage (75% treated once per year), applied proactively before the typical dengue season could reduce symptomatic infections by 89.7% (median of 1000 simulations; interquartile range [IQR]:[83.0%, 94.8%]) in year one and 78.2% (IQR: [71.2%, 88.0%]) cumulatively over the first five years of an annual program. Lower coverage had correspondingly lower effectiveness, as did reactive campaigns. Though less effective than preventative campaigns, reactive and even post-epidemic interventions retain some effectiveness; these campaigns disrupt inter-seasonal transmission, highlighting an off-season control opportunity. Regardless, none of the campaign scenarios maintain their initial effectiveness beyond two seasons, instead stabilizing at much lower levels of benefit: in year 20, median effectiveness was only 27.3% (IQR: [-21.3%, 56.6%]). Furthermore, simply ceasing an initially successful program exposes a population with lowered herd immunity to the same historical threat, and we observed outbreaks more than four-fold larger than pre-intervention outbreaks. These results do not take into account evolving insecticide resistance, thus long-term effectiveness may be lower if new, efficacious insecticides are not developed. Conclusions Using a detailed agent-based dengue transmission model for Yucatán State, Mexico, we predict that high coverage indoor residual spraying (IRS) interventions can largely eliminate transmission for a few years, when applied a few months before the typical seasonal epidemic peak. However, vector control succeeds by preventing infections, which precludes natural immunization. Thus, as a population benefits from mosquito control, it gradually loses naturally acquired herd immunity, and the control effectiveness declines; this occurs across all of our modeled scenarios, and is consistent with other empirical work. Long term control that maintains early effectiveness would require some combination of increasing investment, complementary interventions such as vaccination, and control programs across a broad region to diminish risk of importation.

by Kishor Kumar Paul, Parnali Dhar-Chowdhury, C. Emdad Haque, Hasan Mohammad Al-Amin, Doli Rani Goswami, Mohammad Abdullah Heel Kafi, Michael A. Drebot, L. Robbin Lindsay, Gias Uddin Ahsan, W. Abdullah Brooks

Dengue viruses are responsible for over 100 million infections a year worldwide and are a public health concern in Bangladesh. Although risk of transmission is high, data on vector population characteristics are scanty in Bangladesh; therefore, a comprehensive prediction of the patterns of local virus transmission is not possible. Recognizing these gaps, multi-year entomological surveys were carried out in Dhaka, where the disease is most frequently reported. The specific objectives of the present study are threefold: i) to determine the risk factors for the presence of Aedes mosquitoes; ii) to identify the types of most productive and key containers; and iii) to estimate the effects of climatic factors on Aedes abundance in the city of Dhaka, Bangladesh. Entomological surveys were conducted in 12 out of 90 wards in Dhaka. These wards were selected using a probability proportional sampling procedure during the monsoon seasons in 2011, 2012 and 2013 and in the dry season in 2012. All containers inside and around sampled households were inspected for mosquito larvae and pupae, and containers were classified according to their relative size, use pattern, and materials of construction. During the study period (2011–2013), 12,680 larvae and pupae were collected. About 82% of the identified immature mosquitoes were Aedes aegypti, while the remainder were Ae. albopictus and other mosquito species. The largest number of immature mosquitoes was collected from tires and refrigerator trays during 2011 and 2012 monsoon seasons. Conversely, plastic drums were the most productive during the 2012 dry and 2013 monsoon season. Vehicle parts and discarded construction materials were the most efficient producers of Aedes mosquitoes in all surveys. The presence of Aedes mosquitoes was significantly (p 0.05) higher in low socio-economic zones of Dhaka. Container location, presence of vegetation, and availability of shade for containers were also significantly associated with finding immature Aedes mosquitoes, based on multivariable analysis after confounder adjustment. Rainfall, temperature, and relative humidity also significantly affected the mean abundance of mosquitoes. Proper use, disposal, and recycling of the containers that effectively produce large numbers of Aedes vector mosquitoes may decrease the risk of arboviral transmission.

by Bradley J. Main, Jay Nicholson, Olivia C. Winokur, Cody Steiner, Kasen K. Riemersma, Jackson Stuart, Ryan Takeshita, Michelle Krasnec, Christopher M. Barker, Lark L. Coffey

Zika virus (ZIKV) has emerged since 2013 as a significant global human health threat following outbreaks in the Pacific Islands and rapid spread throughout South and Central America. Severe congenital and neurological sequelae have been linked to ZIKV infections. Assessing the ability of common mosquito species to transmit ZIKV and characterizing variation in mosquito transmission of different ZIKV strains is important for estimating regional outbreak potential and for prioritizing local mosquito control strategies for Aedes and Culex species. In this study, we evaluated the laboratory vector competence of Aedes aegypti, Culex quinquefasciatus, and Culex tarsalis that originated in areas of California where ZIKV cases in travelers since 2015 were frequent. We compared infection, dissemination, and transmission rates by measuring ZIKV RNA levels in cohorts of mosquitoes that ingested blood meals from type I interferon-deficient mice infected with either a Puerto Rican ZIKV strain from 2015 (PR15), a Brazilian ZIKV strain from 2015 (BR15), or an ancestral Asian-lineage Malaysian ZIKV strain from 1966 (MA66). With PR15, Cx. quinquefasciatus was refractory to infection (0%, N = 42) and Cx. tarsalis was infected at 4% (N = 46). No ZIKV RNA was detected in saliva from either Culex species 14 or 21 days post feeding (dpf). In contrast, Ae. aegypti developed infection rates of 85% (PR15; N = 46), 90% (BR15; N = 20), and 81% (MA66; N = 85) 14 or 15 dpf. Although MA66-infected Ae. aegypti showed higher levels of ZIKV RNA in mosquito bodies and legs, transmission rates were not significantly different across virus strains (P = 0.13, Fisher’s exact test). To confirm infectivity and measure the transmitted ZIKV dose, we enumerated infectious ZIKV in Ae. aegypti saliva using Vero cell plaque assays. The expectorated plaque forming units PFU varied by viral strain: MA66-infected expectorated 13±4 PFU (mean±SE, N = 13) compared to 29±6 PFU for PR15-infected (N = 13) and 35±8 PFU for BR15-infected (N = 6; ANOVA, df = 2, F = 3.8, P = 0.035). These laboratory vector competence results support an emerging consensus that Cx. tarsalis and Cx. quinquefasciatus are not vectors of ZIKV. These results also indicate that Ae. aegypti from California are efficient laboratory vectors of ancestral and contemporary Asian lineage ZIKV.

by Rebekah Elliott, Tuhina Banerjee, Santimukul Santra

by Janet Ong, Xu Liu, Jayanthi Rajarethinam, Suet Yheng Kok, Shaohong Liang, Choon Siang Tang, Alex R. Cook, Lee Ching Ng, Grace Yap
Background Singapore experiences endemic dengue, with 2013 being the largest outbreak year known to date, culminating in 22,170 cases. Given the limited resources available, and that vector control is the key approach for prevention in Singapore, it is important that public health professionals know where resources should be invested in. This study aims to stratify the spatial risk of dengue transmission in Singapore for effective deployment of resources. Methodology/principal findings Random Forest was used to predict the risk rank of dengue transmission in 1km2 grids, with dengue, population, entomological and environmental data. The predicted risk ranks are categorized and mapped to four color-coded risk groups for easy operation application. The risk maps were evaluated with dengue case and cluster data. Risk maps produced by Random Forest have high accuracy. More than 80% of the observed risk ranks fell within the 80% prediction interval. The observed and predicted risk ranks were highly correlated (ρ ≥0.86, P

by Chaitawat Sa-ngamuang, Peter Haddawy, Viravarn Luvira, Watcharapong Piyaphanee, Sopon Iamsirithaworn, Saranath Lawpoolsri

Differentiating dengue patients from other acute febrile illness patients is a great challenge among physicians. Several dengue diagnosis methods are recommended by WHO. The application of specific laboratory tests is still limited due to high cost, lack of equipment, and uncertain validity. Therefore, clinical diagnosis remains a common practice especially in resource limited settings. Bayesian networks have been shown to be a useful tool for diagnostic decision support. This study aimed to construct Bayesian network models using basic demographic, clinical, and laboratory profiles of acute febrile illness patients to diagnose dengue. Data of 397 acute undifferentiated febrile illness patients who visited the fever clinic of the Bangkok Hospital for Tropical Diseases, Thailand, were used for model construction and validation. The two best final models were selected: one with and one without NS1 rapid test result. The diagnostic accuracy of the models was compared with that of physicians on the same set of patients. The Bayesian network models provided good diagnostic accuracy of dengue infection, with ROC AUC of 0.80 and 0.75 for models with and without NS1 rapid test result, respectively. The models had approximately 80% specificity and 70% sensitivity, similar to the diagnostic accuracy of the hospital’s fellows in infectious disease. Including information on NS1 rapid test improved the specificity, but reduced the sensitivity, both in model and physician diagnoses. The Bayesian network model developed in this study could be useful to assist physicians in diagnosing dengue, particularly in regions where experienced physicians and laboratory confirmation tests are limited.

Title: Ensaio real triplex para HBV e DENGUE, visando ampliação de alvos do kit NAT HIV/HCV Bio-Manguinhos
Authors: Ferreira, Antonio G. P.; Andrade, Elisabete; Rocha, Daniele; Fontana, Marcela; Alvarez, Patrícia

Title: Lateral flow immunochromatography as a potential test for yellow fever diagnostic
Authors: Freire, Diana Praia Borges; Jabor, Alfredo; Silva, Edimilson Domingos da; Silva Junior, Jose Godinho da; Ano Bom, Ana Paula Dinis

by Despina Contopoulos-Ioannidis, Shoshana Newman-Lindsay, Camille Chow, A. Desiree LaBeaud
Background Chikungunya virus (CHIKV) is an emerging arboviral infection with a global distribution and may cause fetal and neonatal infections after maternal CHIKV-infections during gestation. Methodology We performed a systematic review to evaluate the risk for: a) mother-to-child transmission (MTCT), b) antepartum fetal deaths (APFD), c) symptomatic neonatal disease, and d) neonatal deaths from maternal CHIKV-infections during gestation. We also recorded the neonatal clinical manifestations after such maternal infections (qualitative data synthesis). We searched PubMed (last search 3/2017) for articles, of any study design, with any of the above outcomes. We calculated the overall risk of MTCT, APFDs and risk of symptomatic neonatal disease by simple pooling. For endpoints with ≥5 events in more than one study, we also synthesized the data by random-effect-model (REM) meta-analysis. Principal findings Among 563 identified articles, 13 articles from 8 cohorts were included in the quantitative data synthesis and 33 articles in the qualitative data synthesis. Most cohorts reported data only on symptomatic rather than on all neonatal infections. By extrapolation also of these data, the overall pooled-MTCT-risk across cohorts was at least 15.5% (206/1331), (12.6% by REMs). The pooled APFD-risk was 1.7% (20/1203); while the risk of CHIKV-confirmed-APFDs was 0.3% (3/1203). Overall, the pooled-risk of symptomatic neonatal disease was 15.3% (203/1331), (11.9% by REMs). The pooled risk of symptomatic disease was 50.0% (23/46) among intrapartum vs 0% (0/712) among antepartum/peripartum maternal infections. Infected newborns, from maternal infections during gestation were either asymptomatic or presented within their first week of life, but not at birth, with fever, irritability, hyperalgesia, diffuse limb edema, rashes and occasionally sepsis-like illness and meningoencephalitis. The pooled-risk of neonatal death was 0.6% (5/832) among maternal infections and 2.8% (5/182) among neonatal infections; long-term neurodevelopmental delays occurred in 50% of symptomatic neonatal infections. Conclusions/Significance Published cohorts with data on the risk to the fetus and/or newborn from maternal CHIKV-infections during gestation were sparse compared to the number of recently reported CHIKV-infection outbreaks worldwide; however perinatal infections do occur, at high rates during intrapartum period, and can be related to neonatal death and long-term disabilities.

by David S. Kang, Martin S. Barron, Diane D. Lovin, Joanne M. Cunningham, Matthew W. Eng, Dave D. Chadee, Jun Li, David W. Severson

Populations of Aedes aegypti naturally exhibit variable susceptibility to dengue viruses. This natural variation can be impacted by nutritional stress resulting from larval-stage crowding, indicating the influence of environment components on the adult mosquito immune response. In particular, larval crowding was previously shown to reduce the susceptibility of adult females of a Trinidad field isolate of A. aegypti to the dengue serotype 2 (JAM1409) virus. Here, we present the first whole transcriptome study to address the impact of environmental stress on A. aegypti response to dengue virus. We examined expression profiles of adult females resulting from crowded and optimum reared larvae from the same Trinidad isolate at two critical early time points—3 and 18 hours post dengue virus infected blood meal. We exposed specimens to either a dengue or naïve blood meal, and then characterized the response in ten gene co-expression modules based on their transcriptional associations with environmental stress and time. We further analyzed the top 30 hub or master regulatory genes in each of the modules, and validated our results via qRT-PCR. These hub genes reveal which functions are critical to the mechanisms that confer dengue virus refractoriness or susceptibility to stress conditioned A. aegypti, as well as the time points at which they are most important.

by Nildimar Alves Honório, Keenan Wiggins, Daniel Cardoso Portela Câmara, Bradley Eastmond, Barry W. Alto

Chikungunya virus is a vector-borne alphavirus transmitted by the bites of infected female Ae. aegypti and Ae. albopictus. In Brazil between 2014 and 2016 almost 320 thousand autochthonous human cases were reported and in Florida numerous imported CHIKV viremic cases (> 3,800) demonstrate the potential high risk to establishment of local transmission. In the present study, we carried out a series of experiments to determine the viral dissemination and transmission rates of different Brazilian and Florida populations of Ae. aegypti and Ae. albopictus at 2, 5, and 13 days post-infection for the emergent Asian genotype of CHIKV. Our results show that all tested populations of Ae. aegypti and Ae. albopictus have a high proportion (> 0.80) of individuals with disseminated infection as early as 2 days-post exposure. We found no significant treatment effects of mosquito population origin effects on viral dissemination rates. Transmission rates had a heterogeneous pattern, with US Ae. aegypti and Brazilian Ae. albopictus having the highest proportion of individuals with successful infection (respectively 0.50 and 0.82 as early as 2 days-post infection). Model results found significant effects of population origin, population origin x species, population origin x days post-infection and population origin x species x days post infection.

by Mohammad Sorowar Hossain, Md. Mahbub Hasan, Muhammad Sougatul Islam, Salequl Islam, Miliva Mozaffor, Md. Abdullah Saeed Khan, Nova Ahmed, Waheed Akhtar, Shahanaz Chowdhury, S. M. Yasir Arafat, Md. Abdul Khaleque, Zohora Jameela Khan, Tashmim Farhana Dipta, Shah Md. Zahurul Haque Asna, Md. Akram Hossain, KM Sultanul Aziz, Abdullah Al Mosabbir, Enayetur Raheem
Background Chikungunya virus causes mosquito-transmitted infection that leads to extensive morbidity affecting substantial quality of life. Disease associated morbidity, quality of life, and financial loss are seldom reported in resources limited countries, such as Bangladesh. We reported the acute clinical profile, quality of life and consequent economic burden of the affected individuals in the recent chikungunya outbreak (May to September 2017) in Dhaka city, Bangladesh. Methods We conducted a cross-sectional study during the peak of chikungunya outbreak (July 24 to August 5, 2017) to document the clinical profiles of confirmed cases (laboratory test positive) and probable cases diagnosed by medical practitioners. Data related to clinical symptoms, treatment cost, loss of productivity due to missing work days, and quality of life during their first two-weeks of symptom onset were collected via face to face interview using a structured questionnaire. World Health Organization endorsed questionnaire was used to assess the quality of life. Results A total of 1,326 chikungunya cases were investigated. Multivariate analysis of major clinical variables showed no statistically significant differences between confirmed and probable cases. All the patients reported joint pain and fever. Other more frequently reported symptoms include headache, loss of appetite, rash, myalgia, and itching. Arthralgia was polyarticular in 56.3% of the patients. Notably, more than 70% patients reported joint pain as the first presenting symptom. About 83% of the patients reported low to very low overall quality of life. Nearly half of the patients lost more than 10 days of productivity due to severe arthropathy. Conclusions This study represents one of the largest samples studied so far around the world describing the clinical profile of chikungunya infection. Our findings would contribute to establish an effective syndromic surveillance system for early detection and timely public health intervention of future chikungunya outbreaks in resource-limited settings like Bangladesh.

Title: Early molecular markers predictive of dengue hemorrhagic fever
Authors: Calzavara-Silva, Carlos E.; Gomes, Ana L. V.; Maia, Rita C. C.; Acioli-Santos, Bartolomeu; Gil, Laura H. V. G.; Marques, Ernesto T. A.
Abstract: O manejo de pacientes infectados pelo dengue ainda é um problema desafiador. A maioria dos casos de dengue é benigna mas parte desses casos pode evoluir para o desenvolvimento de vasculopatia severa conhecida como dengue hemorrágica, que pode ser letal. Os sintomas iniciais da dengue e sua forma hemorrágica são bastante similares. O desenvolvimento de um teste diagnóstico que seja rápido e capaz de diferenciar as duas formas clínicas da dengue é crucial para o cuidado adequado de pacientes. O presente estudo avalia, através da PCR quantitativa em tempo real, o potencial preditivo dos níveis de expressão de RNAm candidatos a marcadores da dengue hemorrágica, previamente identificados por estudos genômicos funcionais. Um conjunto de seis marcadores moleculares para a dengue hemorrágica foi avaliado e apresentou correlação entre seus níveis de transcrição e o posterior desenvolvimento da vasculopatia severa. Os marcadores selecionados foram capazes de indicar, nos momentos iniciais dos sintomas, a evolução de um paciente infectado pelo dengue para a forma severa da doença. O painel de candidatos a marcadores de prognóstico obtido demonstrou um bom potencial para uso clínico na forma de um ensaios rápido baseado em amostras de sangue.

by Izabela Maurício de Rezende, Lívia Sacchetto, Érica Munhoz de Mello, Pedro Augusto Alves, Felipe Campos de Melo Iani, Talita Émile Ribeiro Adelino, Myrian Morato Duarte, Ana Luísa Furtado Cury, André Felipe Leal Bernardes, Tayrine Araujo Santos, Leonardo Soares Pereira, Maria Rita Teixeira Dutra, Dario Brock Ramalho, Benoit de Thoisy, Erna Geessien Kroon, Giliane de Souza Trindade, Betânia Paiva Drumond
Background Yellow fever (YF) is endemic in the Brazilian Amazon Basin, and sporadic outbreaks take place outside the endemic area in Brazil. Since 2016, YF epidemics have been occurring in Southeast Brazil, with more than 1,900 human cases and more than 1,600 epizooties of non-human primates (NHPs) reported until April 2018. Previous studies have demonstrated that Yellow fever virus (YFV) causing outbreaks in 2017 formed a monophyletic group. Methodology/Principal findings Aiming to decipher the origin of the YFV responsible for the recent epidemics, we obtained nucleotide sequences of YFV detected in humans (n = 6) and NHPs (n = 10) from Minas Gerais state during 2017–2018. Next, we performed evolutionary analyses and discussed the results in the light of epidemiological records (official numbers of YFV cases at each Brazilian Federative unit, reported by the Brazilian Ministry of Health). Nucleotide sequences of YFV from Southeast Brazil from 2016 to 2018 were highly conserved and formed a monophyletic lineage (BR-YFV_2016/18) within the genotype South America I. Different clusters were observed within lineage YFV-BR_2016/18, one containing the majority of isolates (from humans and NHPs), indicating the sylvatic transmission of YFV. We also detected a cluster characterized by two synapomorphies (amino acid substitutions) that contained YFV only associated with NHP what should be further investigated. The topology of lineage BR-YFV_2016/18 was congruent with epidemiological and temporal patterns of the ongoing epidemic. YFV isolates detected in 2016, in São Paulo state were located in the most basal position of the lineage, followed by the isolates from Minas Gerais and Espírito Santo obtained in 2017 and 2018. The most recent common ancestor of the lineage BR-YFV_2016/18 dated to 2015 (95% credible intervals = 2014–2016), in a period that was coincident with the reemergence of YFV in the Midwest region of Brazil. Conclusions The results demonstrated a single introduction of YFV in the Southeast region and the silent viral circulation before the onset of the outbreaks in 2016. Evolutionary analyses combined with epidemiological records supported the idea that BR-YFV_2016/18 was probably introduced from the Midwest into the Southeast region, possibly in São Paulo state. The persistence of YFV in the Southeast region, causing epidemics from 2016 to 2018, suggests that this region presents suitable ecological and climatic conditions for YFV maintenance during the epidemic and interepidemic seasons. This fact poses risks for the establishing of YF enzootic cycles and epidemics, outside the Amazon Basin in Brazil. YF surveillance and studies of viral dynamics deserve particular attention, especially in Midwest, Southeast and neighbor regions which are the main areas historically associated with YF outbreaks outside the Amazon Basin. YFV persistence in Southeast Brazil should be carefully considered in the context of public health, especially for public health decision-makers and researchers.

by Raquel Burger-Calderon, Karla Gonzalez, Sergio Ojeda, José Victor Zambrana, Nery Sanchez, Cristhiam Cerpas Cruz, Harold Suazo Laguna, Fausto Bustos, Miguel Plazaola, Brenda Lopez Mercado, Douglas Elizondo, Sonia Arguello, Jairo Carey Monterrey, Andrea Nuñez, Josefina Coloma, Jesse J. Waggoner, Aubree Gordon, Guillermina Kuan, Angel Balmaseda, Eva Harris

Zika virus (ZIKV) infection recently caused major epidemics in the Americas and is linked to congenital birth defects and Guillain-Barré Syndrome. A pilot study of ZIKV infection in Nicaraguan households was conducted from August 31 to October 21, 2016, in Managua, Nicaragua. We enrolled 33 laboratory-confirmed Zika index cases and their household members (109 contacts) and followed them on days 3–4, 6–7, 9–10, and 21, collecting serum/plasma, urine, and saliva specimens along with clinical, demographic, and socio-economic status information. Collected samples were processed by rRT-PCR to determine viral load (VL) and duration of detectable ZIKV RNA in human bodily fluids. At enrollment, 11 (10%) contacts were ZIKV rRT-PCR-positive and 23 (21%) were positive by IgM antibodies; 3 incident cases were detected during the study period. Twenty of 33 (61%) index households had contacts with ZIKV infection, with an average of 1.9 (range 1–6) positive contacts per household, and in 60% of these households, ≥50% of the members were positive for ZIKV infection. Analysis of clinical information allowed us to estimate the symptomatic to asymptomatic (S:A) ratio of 14:23 (1:1.6) among the contacts, finding 62% of the infections to be asymptomatic. The maximum number of days during which ZIKV RNA was detected was 7 days post-symptom onset in saliva and serum/plasma and 22 days in urine. Overall, VL levels in serum/plasma, saliva, and urine specimens were comparable, with means of 5.6, 5.3 and 4.5 log10 copies/ml respectively, with serum attaining the highest VL peak at 8.1 log10 copies/ml. Detecting ZIKV RNA in saliva over a similar time-period and level as in serum/plasma indicates that saliva could potentially serve as a more accessible diagnostic sample. Finding the majority of infections to be asymptomatic emphasizes the importance of silent ZIKV transmission and helps inform public health interventions in the region and globally.

by Sarah K. White, Carla Mavian, Maha A. Elbadry, Valery Madsen Beau De Rochars, Taylor Paisie, Taina Telisma, Marco Salemi, John A. Lednicky, J. Glenn Morris Jr.

In the context of recent arbovirus epidemics, questions about the frequency of simultaneous infection of patients with different arbovirus species have been raised. In 2014, a major Chikungunya virus (CHIKV) epidemic impacted the Caribbean and South America. As part of ongoing screening of schoolchildren presenting with acute undifferentiated febrile illness in rural Haiti, we used RT-PCR to identify CHIKV infections in 82 of 100 children with this diagnosis during May—August 2014. Among these, eight were infected with a second arbovirus: six with Zika virus (ZIKV), one with Dengue virus serotype 2, and one with Mayaro virus (MAYV). These dual infections were only detected following culture of the specimen, suggesting low viral loads of the co-infecting species. Phylogenetic analyses indicated that the ZIKV and MAYV strains differ from those detected later in 2014 and 2015, respectively. Moreover, CHIKV and ZIKV strains from co-infected patients clustered monophyletically in their respective phylogeny, and clock calibration traced back the common ancestor of each clade to an overlapping timeframe of introduction of these arboviruses onto the island.

by Getachew Ferede, Moges Tiruneh, Ebba Abate, Yitayih Wondimeneh, Demekech Damtie, Endalamaw Gadisa, Rawleigh Howe, Abraham Aseffa, Belay Tessema
Background Dengue is one of the most serious and rapidly spreading arboviral diseases in the world. Despite many acute febrile illnesses in Ethiopia, the burden of illness due to dengue in the country is largely unknown. Thus, the present study aimed to provide the first baseline data on seroprevalence and associated risk factors of dengue virus (DENV) infection in the country. Methods A cross-sectional study of febrile patients who were visiting Metema and Humera hospitals in Northwest Ethiopia from March 2016 to May 2017 was conducted. Blood samples were collected from each participant and serum samples were separated and tested for IgM and IgG antibodies against DENV infection by enzyme-linked immunosorbent assay (ELISA). Risk factors associated with the prevalence of anti-DENV antibodies were tested using logistic regression analysis. Results Of the 600 samples tested, the overall seroprevalence against DENV infection was 33.3%, while the seroprevalence by the study area was 40% in Metema and 27.5% in Humera. The overall prevalence of IgM and IgG antibodies against DENV infection was 19% and 21% respectively. Of these, 6.7% were positive for both IgM and IgG antibodies. Residence and occupational status were significantly associated with the prevalence of anti-DENV IgM seropositivity and anti-DENV IgM-/G+serostatus. The seasonal variation was significantly associated with the prevalence of anti-DENV IgM but not with anti-DENV IgM-/G+serostatus. The prevalence of anti-DENV IgM-/G+serostatus was significantly higher in Metema than Humera. High prevalence of anti-DENV IgM seropositivity was found in the summer and spring, with a peak in the month of August. The presence of uncovered water either indoor or outdoor and lack of mosquito net use was identified as risk factors for DENV infection. Conclusions These findings provide the preliminary data on seroprevalence and associated risk factors of DENV infection in the country. The presence of antibodies against DENV infection indicates dengue as one of the causes of undifferentiated febrile illnesses in the study areas. This suggests that prevention and control measures should be designed considering the risk factors identified by this study. Furthermore, we recommend a large-scale study to include DENV infection in the differential diagnosis of all febrile illnesses in Ethiopia.

by Benjamin Lopez-Jimena, Stefanie Wehner, Graham Harold, Mohammed Bakheit, Sieghard Frischmann, Michaël Bekaert, Oumar Faye, Amadou Alpha Sall, Manfred Weidmann
Background A single-tube one-step real-time reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay for rapid detection of chikungunya virus (CHIKV) targeting the conserved 6K-E1 target region was developed. The assay was validated with sera collected from a CHIKV outbreak in Senegal in 2015. Methodology/Principal findings A novel design approach by combining Principal Component Analysis and phylogenetic analysis of 110 available CHIKV sequences and the LAMP oligonucleotide design software LAVA was used. The assay was evaluated with an External Quality Assessment panel from the European Network for Diagnostics of "Imported" Viral Diseases and was shown to be sensitive and specific and did not cross-detect other arboviruses. The limit of detection as determined by probit analysis, was 163 molecules, and 100% reproducibility in the assays was obtained for 103 molecules (7/8 repetitions were positive for 102 molecules). The assay was validated using 35 RNA samples extracted from sera, and results were compared with those obtained by quantitative RT-PCR carried out at the Institut Pasteur Dakar, demonstrating that the RT-LAMP is 100% sensitive and 80% specific, with a positive predictive value of 97% and negative predictive value of 100%. Conclusions/Significance The RT-LAMP appeared to show superior performance with material stored for months compared to qRT-PCR and can be therefore recommended for use in infrastructures with poor settings.

by Benjamin Lopez-Jimena, Michaël Bekaert, Mohammed Bakheit, Sieghard Frischmann, Pranav Patel, Etienne Simon-Loriere, Louis Lambrechts, Veasna Duong, Philippe Dussart, Graham Harold, Cheikh Fall, Oumar Faye, Amadou Alpha Sall, Manfred Weidmann
Background 4 one-step, real-time, reverse transcription loop-mediated isothermal amplification (RT-LAMP) assays were developed for the detection of dengue virus (DENV) serotypes by considering 2,056 full genome DENV sequences. DENV1 and DENV2 RT-LAMP assays were validated with 31 blood and 11 serum samples from Tanzania, Senegal, Sudan and Mauritania. DENV3 and DENV4 RT-LAMP assays were validated with 25 serum samples from Cambodia Methodology/Principal findings 4 final reaction primer mixes were obtained by using a combination of Principal Component Analysis of the full DENV genome sequences, and LAMP primer design based on sequence alignments using the LAVA software. These mixes contained 14 (DENV1), 12 (DENV2), 8 (DENV3) and 3 (DENV4) LAMP primer sets. The assays were evaluated with an External Quality Assessment panel from Quality Control for Molecular Diagnostics. The assays were serotype-specific and did not cross-detect with other flaviviruses. The limits of detection, with 95% probability, were 22 (DENV1), 542 (DENV2), 197 (DENV3) and 641 (DENV4) RNA molecules, and 100% reproducibility in the assays was obtained with up to 102 (DENV1) and 103 RNA molecules (DENV2, DENV3 and DENV4). Validation of the DENV2 assay with blood samples from Tanzania resulted in 23 samples detected by RT-LAMP, demonstrating that the assay is 100% specific and 95.8% sensitive (positive predictive value of 100% and a negative predictive value of 85.7%). All serum samples from Senegal, Sudan and Mauritania were detected and 3 untyped as DENV1. The sensitivity of RT-LAMP for DENV4 samples from Cambodia did not quite match qRT-PCR. Conclusions/Significance We have shown a novel approach to design LAMP primers that makes use of fast growing sequence databases. The DENV1 and DENV2 assays were validated with viral RNA extracted clinical samples, showing very good performance parameters.

by Patricia L. Schirmer, Aaron Wendelboe, Cynthia A. Lucero-Obusan, Russell A. Ryono, Mark A. Winters, Gina Oda, Mirsonia Martinez, Sonia Saavedra, Mark Holodniy
Background Zika virus (ZIKV) is an important flavivirus infection. Although ZIKV infection is rarely fatal, risk for severe disease in adults is not well described. Our objective was to describe the spectrum of illness in U.S. Veterans with ZIKV infection. Methodology Case series study including patients with laboratory-confirmed or presumed positive ZIKV infection in all Veterans Health Administration (VHA) medical centers. Adjusted odds ratios of clinical variables associated with hospitalization and neurologic complications was performed. Principal findings Of 1,538 patients tested between 12/2015-10/2016 and observed through 3/2017, 736 (48%) were RT-PCR or confirmed IgM positive; 655 (89%) were male, and 683 (93%) from VA Caribbean Healthcare System (VACHCS). Ninety-four (13%) were hospitalized, 91 (12%) in the VACHCS. Nineteen (3%) died after ZIKV infection. Hospitalization was associated with increased Charlson co-morbidity index (adjusted odds ratio [OR] 1.2; 95% confidence interval [CI], 1.1–1.3), underlying connective tissue disease (OR, 29.5; CI, 3.6–244.7), congestive heart failure (OR, 6; CI, 2–18.5), dementia (OR, 3.6; CI, 1.1–11.2), neurologic symptom presentation (OR, 3.9; CI, 1.7–9.2), leukocytosis (OR, 11.8; CI, 4.5–31), thrombocytopenia (OR, 7.8; CI, 3.3–18.6), acute kidney injury (OR, 28.9; CI, 5.8–145.1), or using glucocorticoids within 30 days of testing (OR, 13.3; CI 1.3–133). Patients presenting with rash were less likely to be hospitalized (OR, 0.29; CI, 0.13–0.66). Risk for neurologic complications increased with hospitalization (OR, 5.9; CI 2.9–12.2), cerebrovascular disease (OR 4.9; CI 1.7–14.4), and dementia (OR 2.8; CI 1.2–6.6). Conclusion Older Veterans with multiple comorbidities or presenting with neurologic symptoms were at increased risk for hospitalization and neurological complications after ZIKV infection.

Title: Aprimoramento da produção do vírus da febre amarela em biorreatores de tanque agitado de 3 Litros
Authors: Silva, Marlon Vicente da; Mattos, Diogo Araujo de; Souza, Marta Cristina de Oliveira; Silva, Andréa Nazaré Monteiro Rangel da; Lima, Sheila Barbosa de; Castilho, Leda dos Reis; Gaspar, Luciane Pinto; Caride, Elena Siqueira Campos; Freire, Marcos da Silva

Title: Desenvolvimento do processo "downstream" na produção de uma vacina tetravalente recombinante contra dengue
Authors: Yorio, Anna Paula; Martins, Maria Carolina de Souza; Fernandes, André Tavares da Silva; Archer, Márcia

by Mabel L. Taracena, Vanessa Bottino-Rojas, Octavio A. C. Talyuli, Ana Beatriz Walter-Nuno, José Henrique M. Oliveira, Yesseinia I. Angleró-Rodriguez, Michael B. Wells, George Dimopoulos, Pedro L. Oliveira, Gabriela O. Paiva-Silva

Aedes aegypti is the vector of some of the most important vector-borne diseases like dengue, chikungunya, zika and yellow fever, affecting millions of people worldwide. The cellular processes that follow a blood meal in the mosquito midgut are directly associated with pathogen transmission. We studied the homeostatic response of the midgut against oxidative stress, as well as bacterial and dengue virus (DENV) infections, focusing on the proliferative ability of the intestinal stem cells (ISC). Inhibition of the peritrophic matrix (PM) formation led to an increase in reactive oxygen species (ROS) production by the epithelial cells in response to contact with the resident microbiota, suggesting that maintenance of low levels of ROS in the intestinal lumen is key to keep ISCs division in balance. We show that dengue virus infection induces midgut cell division in both DENV susceptible (Rockefeller) and refractory (Orlando) mosquito strains. However, the susceptible strain delays the activation of the regeneration process compared with the refractory strain. Impairment of the Delta/Notch signaling, by silencing the Notch ligand Delta using RNAi, significantly increased the susceptibility of the refractory strains to DENV infection of the midgut. We propose that this cell replenishment is essential to control viral infection in the mosquito. Our study demonstrates that the intestinal epithelium of the blood fed mosquito is able to respond and defend against different challenges, including virus infection. In addition, we provide unprecedented evidence that the activation of a cellular regenerative program in the midgut is important for the determination of the mosquito vectorial competence.

by John H. Huber, Marissa L. Childs, Jamie M. Caldwell, Erin A. Mordecai

Dengue, chikungunya, and Zika virus epidemics transmitted by Aedes aegypti mosquitoes have recently (re)emerged and spread throughout the Americas, Southeast Asia, the Pacific Islands, and elsewhere. Understanding how environmental conditions affect epidemic dynamics is critical for predicting and responding to the geographic and seasonal spread of disease. Specifically, we lack a mechanistic understanding of how seasonal variation in temperature affects epidemic magnitude and duration. Here, we develop a dynamic disease transmission model for dengue virus and Aedes aegypti mosquitoes that integrates mechanistic, empirically parameterized, and independently validated mosquito and virus trait thermal responses under seasonally varying temperatures. We examine the influence of seasonal temperature mean, variation, and temperature at the start of the epidemic on disease dynamics. We find that at both constant and seasonally varying temperatures, warmer temperatures at the start of epidemics promote more rapid epidemics due to faster burnout of the susceptible population. By contrast, intermediate temperatures (24–25°C) at epidemic onset produced the largest epidemics in both constant and seasonally varying temperature regimes. When seasonal temperature variation was low, 25–35°C annual average temperatures produced the largest epidemics, but this range shifted to cooler temperatures as seasonal temperature variation increased (analogous to previous results for diurnal temperature variation). Tropical and sub-tropical cities such as Rio de Janeiro, Fortaleza, and Salvador, Brazil; Cali, Cartagena, and Barranquilla, Colombia; Delhi, India; Guangzhou, China; and Manila, Philippines have mean annual temperatures and seasonal temperature ranges that produced the largest epidemics. However, more temperate cities like Shanghai, China had high epidemic suitability because large seasonal variation offset moderate annual average temperatures. By accounting for seasonal variation in temperature, the model provides a baseline for mechanistically understanding environmental suitability for virus transmission by Aedes aegypti. Overlaying the impact of human activities and socioeconomic factors onto this mechanistic temperature-dependent framework is critical for understanding likelihood and magnitude of outbreaks.

Title: Proposta de revisão da RDC 69 8/12/2014, para a produção de vacina de febre amarela de subunidade utilizando plataforma vegetal
Authors: Guimarães, Rosane Cuber; Freire, Marcos da Silva; Almeida, Antônio Eugenio Castro Cardoso de

Title: Avaliação do risco de biossegurança para a produção de vacina febre amarela de subunidade utilizando plataforma vegetal
Authors: Guimarães, Rosane Cuber; Freire, Marcos da Silva; Almeida, Antônio Eugenio Castro Cardoso de

Title: Teste de ativação de monócitos para monitoramento da atividade pirogênica na vacina de febre amarela
Authors: Silva, Vítor Fernandes; Almeida, Alessandra Santos; Navega, Elaine Cristina Azevedo; Silva, Cristiane Caldeira da; Presgrave, Octávio Augusto França; Delgado, Isabella Fernandes; Mattos, Katherine Antunes de

by Laith Hussain-Alkhateeb, Axel Kroeger, Piero Olliaro, Joacim Rocklöv, Maquins Odhiambo Sewe, Gustavo Tejeda, David Benitez, Balvinder Gill, S. Lokman Hakim, Roberta Gomes Carvalho, Leigh Bowman, Max Petzold
Background Dengue outbreaks are increasing in frequency over space and time, affecting people’s health and burdening resource-constrained health systems. The ability to detect early emerging outbreaks is key to mounting an effective response. The early warning and response system (EWARS) is a toolkit that provides countries with early-warning systems for efficient and cost-effective local responses. EWARS uses outbreak and alarm indicators to derive prediction models that can be used prospectively to predict a forthcoming dengue outbreak at district level. Methods We report on the development of the EWARS tool, based on users’ recommendations into a convenient, user-friendly and reliable software aided by a user’s workbook and its field testing in 30 health districts in Brazil, Malaysia and Mexico. Findings 34 Health officers from the 30 study districts who had used the original EWARS for 7 to 10 months responded to a questionnaire with mainly open-ended questions. Qualitative content analysis showed that participants were generally satisfied with the tool but preferred open-access vs. commercial software. EWARS users also stated that the geographical unit should be the district, while access to meteorological information should be improved. These recommendations were incorporated into the second-generation EWARS-R, using the free R software, combined with recent surveillance data and resulted in higher sensitivities and positive predictive values of alarm signals compared to the first-generation EWARS. Currently the use of satellite data for meteorological information is being tested and a dashboard is being developed to increase user-friendliness of the tool. The inclusion of other Aedes borne viral diseases is under discussion. Conclusion EWARS is a pragmatic and useful tool for detecting imminent dengue outbreaks to trigger early response activities.

Title: First detection of dengue virus in the saliva of immunocompetent murine model
Authors: Rasinhas, Arthur da Costa; Silva, Marcos Alexandre Nunes da; Caldas, Gabriela Cardoso; Jácome, Fernanda Cunha; Leonardo, Raphael; Santos, Flávia Barreto dos; Nunes, Priscila Conrado Guerra; Barth, Ortrud Monika; Vieira, Debora Ferreira Barreto