Effect of Larval Food Availability on Adult Aedes Aegypti (Diptera: Culicidae) Fitness and Susceptibility to Zika Infection.

Aedes (Stegomyia) aegypti (Linnaeus, 1762) is a mosquito species of significant medical importance. The use of this vector in research studies usually requires a large number of mosquitoes as well as rearing and maintenance in a laboratory-controlled environment. However, laboratory conditions may be different from field environments, presenting stressful challenges such as low food concentration, especially during larval stages, which may, in turn, impair vector biology. Therefore, we tested herein if larval food availability (0.004, 0.009, 0.020, and 0.070% diets) would affect overall adult insect fitness. We observed slower development in mosquitoes fed a 0.004% diet 15 d post-eclosion (DPE) and shorter mean time in mosquitoes fed a 0.020% diet (7 DPE). Larval diet and adult mosquito weight were positively correlated, and heavier females fed higher larval diets exhibited greater blood feeding capacity and oviposition. In addition, larval diet concentrations led to median adult lifespan variations (male/female in days-0.004%: 30 ± 1.41, 45 ± 1.3; 0.009%: 31.5 ± 1.33, 41 ± 1.43; 0.020%: 26 ± 1.18, 41 ± 1.45; 0.070%: 29 ± 1.07, 44 ± 1.34), reduced tolerance to deltamethrin (1 mg/m2) and changes in detoxification enzyme activities. Moreover, in the larval 0.070% diet, females presented higher Zika susceptibility (plaque-forming unit [PFU]: 1.218 × 106) compared with other diets (0.004%: 1.31 × 105; 0.009%: 2.0 × 105; 0.020%: 1.25 × 105 PFU). Altogether, our study demonstrates that larval diet restriction results not only in larval developmental arrest but also in adult fitness impairment, which must be considered in future assessments.

Accelerating the Morphogenetic Cycle of the Viral Vector Aedes aegypti Larvae for Faster Larvicidal Bioassays.

Any bioassay to test new chemically synthesized larvicides or phytolarvicides against Culicidae and more harmful mosquito species, such as Aedes aegypti and Aedes albopictus, which specifically transmit dengue, yellow fever, chikungunya viral fevers as well as Zika virus, or Anopheles gambiae, a vector for malaria and philariasis, requires thousands of well-developed larvae, preferably at the fourth instar stage. The natural morphogenetic cycle of Aedes spp., in the field or in the laboratory, may extend to 19 days at room temperature (e.g., 25°C) from the first permanent contact between viable eggs and water and the last stage of larval growth or metamorphosis into flying adults. Thus, accelerated sequential molting is desirable for swifter bioassays of larvicides. We achieved this goal in Aedes aegypti with very limited strategic and low-cost additions to food, such as coconut water, milk or its casein, yeast extract, and to a lesser extent, glycerol. The naturally rich coconut water was excellent for quickly attaining the population of instar IV larvae, the most advanced one before pupation, saving about a week, for subsequent larvicidal bioassays. Diluted milk, as another food source, allowed an even faster final ecdysis and adults are useful for mosquito taxonomical purpose.

Kdr genotyping in Aedes aegypti from Brazil on a nation-wide scale from 2017 to 2018.

Insecticide resistance is currently a threat to the control of Aedes agypti, the main vector of arboviruses in urban centers. Mutations in the voltage gated sodium channel (NaV), known as kdr (knockdown resistance), constitute an important selection mechanism for resistance against pyrethroids. In the present study, we investigated the kdr distribution for the Val1016Ile and Phe1534Cys alterations in Ae. aegypti from 123 Brazilian municipalities, based on SNP genotyping assays in over 5,500 mosquitoes. The alleles NaVS (1016Val+ + 1534Phe+), NaVR1 (1016Val+ + 1534Cyskdr) and NaVR2 (1016Ilekdr + 1534Cyskdr) were consistently observed, whereas kdr alleles have rapidly spread and increased in frequency. NaVS was the less frequent allele, mostly found in Northeastern populations. The highest allelic frequencies were observed for NaVR1, especially in the North, which was fixed in one Amazonian population. The double kdr NaVR2 was more prevalent in the Central-west and South-eastern populations. We introduce the 'kdr index', which revealed significant spatial patterns highlighting two to three distinct Brazilian regions. The 410L kdr mutation was additionally evaluated in 25 localities, evidencing that it generally occurs in the NaVR2 allele. This nationwide screening of a genetic mechanism for insecticide resistance is an important indication on how pyrethroid resistance in Ae. aegypti is evolving in Brazil.

Insecticide resistance, fitness and susceptibility to Zika infection of an interbred Aedes aegypti population from Rio de Janeiro, Brazil.

BACKGROUND: Aedes aegypti is a vector of high relevance, since it transmits several arboviruses, including dengue, chikungunya and Zika. Studies on vector biology are usually conducted with laboratory strains presenting a divergent genetic composition from field populations. This may impair vector control policies that were based on laboratory observations employing only long maintained laboratory strains. In the present study we characterized a laboratory strain interbreed with Ae. aegypti collected from five different localities in Rio de Janeiro (Aedes Rio), for insecticide resistance (IR), IR mechanisms, fitness and Zika virus infection. METHODS: We compared the recently established Aedes Rio with the laboratory reference strain Rockefeller. Insecticide resistance (deltamethrin, malathion and temephos), activity of metabolic resistance enzymes and kdr mutation frequency were determined. Some life table parameters (longevity, blood-feeding, number and egg viability) and Zika virus susceptibility was also determined. RESULTS: Aedes Rio showed resistance to deltamethrin (resistance ratio, RR50 = 32.6) and temephos (RR50 = 7.0) and elevated activity of glutathione S-transferase (GST) and esterases (α-EST and pNPA-EST), but not acetylcholinesterase (AChE). In total, 92.1% of males genotyped for kdr presented a "resistant" genotype. Weekly blood-fed females from both strains, presented reduced mortality compared to sucrose-fed mosquitoes; however, Aedes Rio blood-fed females did not live as long (mean lifespan: Rockefeller = 70 ± 3.07; Aedes Rio = 53.5 ± 2.16 days). There were no differences between strains in relation to blood-feeding and number of eggs, but Aedes Rio eggs presented reduced viability (mean hatch: Rockefeller = 77.79 ± 1.4%; Aedes Rio = 58.57 ± 1.77%). Zika virus infection (plaque-forming unit, PFU) was similar in both strains (mean PFU ± SE: Aedes Rio: 4.53 × 104 ± 1.14 × 104 PFU; Rockefeller: 2.02 × 104 ± 0.71 × 104 PFU). CONCLUSION: Selected conditions in the field, such as IR mechanisms, may result in pleiotropic effects that interfere in general physiology of the insect. Therefore, it is important to well characterize field populations to be tested in parallel with laboratory reference strains. This practice would improve the significance of laboratory tests for vector control methods.

Perfil de susceptibilidad a temefos en poblaciones de aedes aegypti (diptera: culicidae) de ciudad del este - alto Paraná, Paraguay / Aedes aegypti (díptera-culicidae) susceptibility to temephos in ciudad del este, alto Paraná, Paraguay

En Paraguay, el control del mosquito Aedes aegypti involucrado en la transmisión de varias arbovirosis implica la utilización del Temefos, un organofosforado que ha sido utilizado por los programas nacionales para el control vectorial por más de dos décadas en busca de la reducción de los estadios larvarios. En vista de la necesidad de evaluar periódicamente la actividad larvicida del compuesto químico mencionado, este estudio tuvo como objetivo monitorear el perfil de susceptibilidad de larvas de Ae. aegypti al Temefos. Para ello se aplicó un estudio analítico experimental con ensayos biológicos tipo dosis - respuesta, utilizando larvas del tercer estadio de la primera generación procedentes de una colonia de mosquitos colectada en Ciudad del Este. Las larvas fueron expuestas a la acción del Temefos a diferentes concentraciones definidas por un pre- test. Se registraron valores correspondientes al número de larvas expuestas y mortalidad al término de cada ensayo. Los resultados fueron concentración letal CL50 = 0,00966 mg/L y CL90 = 0,03015mg/L, a partir de estos valores se obtuvieron los indicadores cuantitativos de resistencia, Razón de resistencia RR50 = 2,3734 y RR90 = 4,1643 respectivamente. Este último es un indicativo de resistencia baja en las poblaciones de Ae. aegypti evaluadas, acorde con rangos estandarizados (RR>3<5). Los resultados observados en las poblaciones silvestres de larvas revelan una situación de alerta, considerando que el presente estudio evidenció un proceso de resistencia incipiente al Temefos. Finalmente, basados en los resultados se recomienda plantear y ejecutar estrategias basadas en acciones que permitan preservar la actividad larvicida de este compuesto, evitando el aumento progresivo de resistencia en las poblaciones silvestres.

In Paraguay, the control of Aedes aegypti mosquitoes, involved in the transmission of several arboviruses, implies the use of temephos, an organophosphate that has been used by national vector control programs for more than two decades in search of reducing larval stages. In view of the need to evaluate periodically the larvicidal activity of this chemical compound, this study aimed to monitor the susceptibility profile to temephos of larval stages of Ae. aegypti. For this, an experimental analytical study with bioassays dose rate –response was applied, using third instar larva e of the first generation from a mosquitoescolony collected in Ciudad del Este. The larvae were exposed to different concentrations ranges of themephos that were defined by a pre-test. The number of larva e exposed and mortality at the end of each trial were recorded. The results determined the LC50 =0.00966 mg/L and LC90 = 0.03015 mg/L, from which quantitative indicators of resistance were obtained, resistance ratio RR = 2.3734 and 4.1643 respectively. The latter valueindicates characteristics related to low resistance in populations of Ae. aegypti evaluateda ccording to standardized ranges (RR> 3 <5). The results observed in larval wild populations indicate an alert situation, considering that our study showed an incipient process of resistance to temephos. Finally, based on these results, proposing and implementing strategies based in actions that allow to preserve the larvicidal activity of this compound are recommended to avoiding the progressive increase of resistance in wild populations.

Bacterial microbiomes from vertically transmitted fungal inocula of the leaf-cutting ant Atta texana.

Microbiome surveys provide clues for the functional roles of symbiotic microbial communities and their hosts. In this study, we elucidated bacterial microbiomes associated with the vertically transmitted fungal inocula (pellets) used by foundress queens of the leaf-cutting ant Atta texana as starter-cultures for new gardens. As reference microbiomes, we also surveyed bacterial microbiomes of foundress queens, gardens and brood of incipient nests. Pseudomonas, Acinetobacter, Propionibacterium and Corynebacterium were consistently present in high abundance in microbiomes. Some pellet and ant samples contained abundant bacteria from an Entomoplasmatales-clade, and a separate PCR-based survey of Entomoplasmatales bacteria in eight attine ant-genera from Brazil placed these bacteria in a monophyletic clade within the bacterial genus Mesoplasma. The attine ant-Mesoplasma association parallels a similar association between a closely related, monophyletic Entomoplasmatales-clade and army ants. Of thirteen A. texana nests surveyed, three nests with exceptionally high Mesoplasma abundance died, whereas the other nests survived. It is unclear whether Mesoplasma was the primary cause of mortality, or Mesoplasma became abundant in moribund nests for non-pathogenic reasons. However, the consistent and geographically widespread presence of Mesoplasma suggests an important functional role in the association with attine ants.