Mass trapping with MosquiTRAPs does not reduce Aedes aegypti abundance

The objective of this study was to evaluate the effectiveness of Aedes aegyptimass trapping using the sticky trap MosquiTRAP (MQT) by performing a cluster randomised controlled trial in Manaus, state of Amazonas, Brazil. After an initial questionnaire and baseline monitoring of adultAe. aegyptiabundance with BG-Sentinel (BGS) traps in six clusters, three clusters were randomly assigned to the intervention arm where each participating household received three MQTs for mass trapping during 17 months. The remaining three clusters (control arm) did not receive traps. The effect of mass trapping on adult Ae. aegyptiabundance was monitored fortnightly with BGS traps. During the last two months of the study, a serological survey was conducted. After the study, a second questionnaire was applied in the intervention arm. Entomological monitoring indicated that MQT mass trapping did not reduce adult Ae. aegyptiabundance. The serological survey indicated that recent dengue infections were equally frequent in the intervention and the control arm. Most participants responded positively to questions concerning user satisfaction. According to the results, there is no evidence that mass trapping with MQTs can be used as a part of dengue control programs. The use of this sticky trap is only recommendable for dengue vector monitoring.

Profils de réponse immunitaire innée et adaptative lors d'infections aiguës par le virus de l'hépatite B, de la dengue et influenza A

Les réponses immunitaires innées et adaptatives déclenchées par une infection virale chez l'humain sont classiquement décrites comme une succession d'événements communs à tous les virus- la réponse innée, caractérisés par la libération rapide de cytokines antivirales et des chémokines, recrutant monocytes, NK et lymphocytes Τ vers le site d'infection, suivis par l'activation de l'immunité adaptative. Notre compréhension de la dynamique de ces mécanismes dynamiques est limitée chez l'humain. En effet, il existe peu d'études portant sur la cinétique et l'analyse quantitative de la réponse Τ spécifique au virus, parallèlement aux aspects plus qualitatifs de cette réponse (cytokines sériques produites lors de différentes infections virales, notamment). Méthode: Nous avons étudiés trois groupes de patients tous recrutés au cours de la phase aiguë d'une infection par le virus de la dengue (28 patients), le virus influenza A (13 patients) et le virus de l'hépatite Β (HBV) (13 patients). Nous avons analysé le profil d'activation (CD38, HLA-DR) et de prolifération (Ki-67, Bcl-2) des lymphocytes Τ CD8+ (par cytométrie de flux), de façon longitudinale à différents timepoints (depuis le début des symptômes jusqu'à rémission totale) en quantifiant 15 cytokines et chémokines (par Luminex multiplex biométrie immunoassay) dans le sérum des patients infectés. Résultats: Nous avons comparé le profil des réponses innée et adaptative chez les 3 types d'infection virales; les patients infectés par l'HBV ont une fréquence élevée de CD8+ spécifiques activés et proliférant ainsi que des taux sériques élevés de TNF-α et d'IFN-γ. Les patients infectés par le virus de la dengue et par le virus Influenza présentent quant à eux une activation CD 8+ moins intense mais une forte expression de la réponse innée, marquée par une élévation des cytokines IFN-α, IFN-γ, et TNF-α. De plus, une particularité des patients infectés par le virus de la dengue est de présenter une élévation marquée des cytokines immunorégulatrices (IL-10, IL- 1RA). Conclusion: Ces résultats permettent de montrer que la réponse immunologique consécutive à une infections virale spécifique est caractérisée par sa propre signature, tant au niveau de la production de cytokines/chemokines que de la quantité des lymphocytes Τ CD+8+ spécifiques activés et proliférantes. Ce travail contribue ainsi à une meilleure compréhension de l'immunité antivirale chez les humains, grâce à la description de la cinétique et de la quantification des cellules Τ CD8+ activées et des taux de cytokines dans chaque infection étudiée. Abstract Knowledge of innate and adaptive immune parameters triggered by viral infections is limited but important for understanding disease pathogenesis. We performed a comparative longitudinal analysis of serum cytokines/chemokines and of virus-activated CD8 Τ cells population in patients with acute dengue, influenza A or HBV infections from onset to disease recovery. We observed that each viral infection is characterized by its own signature of cytokines/chemokines production and size of activated and proliferating CD8 Τ cell pool. This is, to our knowledge, the first comparative longitudinal study of the immune response in human subjects in three distinct viral infections.

Improved immunogenicity against HIV-1 clade C antigens by using NYVAC-C with restored replication competence.

In order to improve the immunogenicity of currently available non-replicating pox virus HIV vaccine vectors, NYVAC was genetically modified through re-insertion of two host range genes (K1L and C7L), resulting in restored replicative capacity in human cells. In the present study these vectors, expressing either a combination of the HIV-1 clade C antigens Env, Gag, Pol, Nef, or a combination of Gal, Pol, Nef were evaluated for safety and immunogenicity in rhesus macaques, which were immunized at weeks 0, 4 and 12 either by scarification (conventional poxvirus route of immunization), intradermal or by intramuscular injection (route used in previous vaccine studies).Replication competent NYVAC-C-KC vectors induced higher HIV-specific responses, as measured by IFN- ELISpot assay, than the replication defective NYVAC-C vectors. Application through scarification only required one immunization to induce maximum HIV-specific immune responses. This method simultaneously induced relatively lower anti-vector responses. In contrast, two to three immunizations were required when the NYVAC-C-KC vectors were given by intradermal or intramuscular injection and this method tended to generate slightly lower responses. Responses were predominantly directed against Env in the animals that received NYVAC-C-KC vectors expressing HIV-1 Env, Gag, Pol, Nef, while Gag responses were dominant in the NYVAC-C-KC HIV-1 Gag, Pol, Nef immunized animals.The current study demonstrates that NYVAC replication competent vectors were well tolerated and showed increased immunogenicity as compared to replication defective vectors. Further studies are needed to evaluate the most efficient route of immunization and to explore the use of these replication competent NYVAC vectors in prime/boost combination with gp120 protein-based vaccine candidates. This studies was performed within the Poxvirus T-cell Vaccine Discovery Consortium (PTVDC) which is part of the CAVD program.

Evaluation of oviposition traps as an entomological surveillance method for Aedes aegypti (Diptera, Culicidae)

This study aimed to describe the behavior of oviposition traps for Aedes aegypti over time, to compare it with the larval survey and to investigate the association with climatic variables. It was conducted in São José do Rio Preto city, São Paulo. Daily climatic data and fortnightly measurements for oviposition traps and larval infestation were collected from October 2003 to September 2004. Three different periods were identified in the behavior of oviposition traps' positivity and mean number of eggs: increase, plateau and decrease in values. These measurements followed the variation of climatic data from the first and third periods. High correlation was obtained between the positivity and the mean number of eggs. The oviposition traps showed higher capacity to detect the vector than did larval survey. It was observed that the first (October to December) and third (May to September) periods were considered to be the most suitable to use oviposition traps than larval surveys.

Impact of small variations in temperature and humidity on the reproductive activity and survival of Aedes aegypti (Diptera, Culicidae)

In short space of time increase in temperature and rainfall can affect vector populations and, consequently, the diseases for them transmitted. The present study analyzed the effect of small temperature and humidity variations on the fecundity, fertility and survival of Aedes aegypti. These parameters were analyzed using individual females at temperatures ranging from 23 to 27 °C (mean 25 °C); 28 to 32 °C (mean 30 °C) and 33 to 37 °C (mean 35 ºC) associated to 60±8% and 80±6% relative humidity. Females responded to an increase in temperature by reducing egg production, oviposition time and changing oviposition patterns. At 25 ºC and 80% relative humidity, females survived two-fold more and produced 40% more eggs when compared to those kept at 35 ºC and 80% relative humidity. However, in 45% of females kept at 35 ºC and 60% relative humidity oviposition was inhibited and only 15% females laid more than 100 eggs, suggesting that the intensity of the temperature effect was influenced by humidity. Gradual reductions in egg fertility at 60% relative humidity were observed with the increase in temperature, although such effect was not found in the 80% relative humidity at 25 º C and 30 º C. These results suggest that the reduction in population densities recorded in tropical areas during seasons when temperatures reach over 35 ºC is likely to be strongly influenced by temperature and humidity, with a negative effect on several aspects of mosquito biology.

Isolation of Bacillus thuringiensis from the state of Amazonas, in Brazil, and screening against Aedes aegypti (Diptera, Culicidae)

We investigated the use of Bacillus thuringiensis isolated in the state of Amazonas, in Brazil, for the biological control of the dengue vector Aedes aegypti. From 25 soil samples collected in nine municipalities, 484 bacterial colonies were obtained, 57 (11.78%) of which were identified as B. thuringiensis. Six isolates, IBt-03, IBt-06, IBt-07, IBt-28, IBt-30, and BtAM-27 showed insecticidal activity, and only BtAM-27 presents the five genes investigated cry4Aa, cry4Ba, cry10Aa, cry11Aa, and cry11Ba. The IBt-07 and IBt- 28, with lower LC50 values, showed equal toxicity compared to the standards. The isolates of B. thuringiensisfrom Amazonas constitute potential new means of biological control for A. aegypti, because of their larvicidal activity and the possibility that they may also contain new combinations of toxins.

Robust vaccine-elicited cellular immune responses in breast milk following systemic Simian Immunodeficiency Virus DNA prime and live virus vector boost vaccination of lactating rhesus monkeys.

Breast milk transmission of HIV remains an important mode of infant HIV acquisition. Enhancement of mucosal HIV-specific immune responses in milk of HIV-infected mothers through vaccination may reduce milk virus load or protect against virus transmission in the infant gastrointestinal tract. However, the ability of HIV/SIV strategies to induce virus-specific immune responses in milk has not been studied. In this study, five uninfected, hormone-induced lactating, Mamu A*01(+) female rhesus monkey were systemically primed and boosted with rDNA and the attenuated poxvirus vector, NYVAC, containing the SIVmac239 gag-pol and envelope genes. The monkeys were boosted a second time with a recombinant Adenovirus serotype 5 vector containing matching immunogens. The vaccine-elicited immunodominant epitope-specific CD8(+) T lymphocyte response in milk was of similar or greater magnitude than that in blood and the vaginal tract but higher than that in the colon. Furthermore, the vaccine-elicited SIV Gag-specific CD4(+) and CD8(+) T lymphocyte polyfunctional cytokine responses were more robust in milk than in blood after each virus vector boost. Finally, SIV envelope-specific IgG responses were detected in milk of all monkeys after vaccination, whereas an SIV envelope-specific IgA response was only detected in one vaccinated monkey. Importantly, only limited and transient increases in the proportion of activated or CCR5-expressing CD4(+) T lymphocytes in milk occurred after vaccination. Therefore, systemic DNA prime and virus vector boost of lactating rhesus monkeys elicits potent virus-specific cellular and humoral immune responses in milk and may warrant further investigation as a strategy to impede breast milk transmission of HIV.

The yellow fever 17D vaccine virus: molecular basis of viral attenuation and its use as an expression vector

The yellow fever (YF) virus is the prototype flavivirus. The use of molecular techniques has unraveled the basic mechanisms of viral genome structure and expression. Recent trends in flavivirus research include the use of infectious clone technology with which it is possible to recover virus from cloned cDNA. Using this technique, mutations can be introduced at any point of the viral genome and their resulting effect on virus phenotype can be assessed. This approach has opened new possibilities to study several biological viral features with special emphasis on the issue of virulence/attenuation of the YF virus. The feasibility of using YF virus 17D vaccine strain, for which infectious cDNA is available, as a vector for the expression of heterologous antigens is reviewed

Attenuation and immunogenicity of recombinant yellow fever 17D-dengue type 2 virus for rhesus monkeys

A chimeric yellow fever (YF)-dengue serotype 2 (dengue 2) virus was constructed by replacing the premembrane and envelope genes of the YF 17D virus with those from dengue 2 virus strains of Southeast Asian genotype. The virus grew to high titers in Vero cells and, after passage 2, was used for immunogenicity and attenuation studies in rhesus monkeys. Subcutaneous immunization of naive rhesus monkeys with the 17D-D2 chimeric virus induced a neutralizing antibody response associated with the protection of 6 of 7 monkeys against viremia by wild-type dengue 2 virus. Neutralizing antibody titers to dengue 2 were significantly lower in YF-immune animals than in YF-naive monkeys and protection against challenge with wild-type dengue 2 virus was observed in only 2 of 11 YF-immune monkeys. An anamnestic response to dengue 2, indicated by a sharp increase of neutralizing antibody titers, was observed in the majority of the monkeys after challenge with wild-type virus. Virus attenuation was demonstrated using the standard monkey neurovirulence test. The 17D-D2 chimera caused significantly fewer histological lesions than the YF 17DD virus. The attenuated phenotype could also be inferred from the limited viremias compared to the YF 17DD vaccine. Overall, these results provide further support for the use of chimeric viruses for the development of a new live tetravalent dengue vaccine.

Clonación y expresión de una proteína recombinante del Gen M del virus del Dengue Serotipo 2.

Tesis (Maestro en Ciencias con Acentuación en Inmunolobiología) UANL, 2011.

Identificación y análisis de las variantes genéticas del virus del dengue y su asociación en la dinámica de su transmisión.

Tesis (Maestría en Ciencias con Orientación en Biología Molecular e Ingeniería Genética) UANL, 2011.

Generación de un vector adenoviral para la producción de anticuerpos policlonales contra la proteína ENV del XMRV (Xenotropic murine leukemia virus-related virus).

Tesis (Maestría en Ciencias con Orientación en Biología Molecular e Ingeniería Genética) UANL, 2011.

Diseño de un sistema de expresión heteróloga de proteínas recombinantes de envoltura del virus del dengue.

Tesis (Maestro en Ciencias con orientación en Biología Molecular e Ingeniería Genética) UANL, 2013.

Identificación y análisis de las variantes genómicas del virus del dengue y su asociación con la presentación clínica en una zona de baja y otra de alta endemicidad en México.

Tesis (Maestro en Ciencias con orientación en Biología Molecular e Ingeniería Genética) UANL, 2013.