14 research outputs found
The Effects of Infection on Mosquito Rhythmic Behavior
Most organisms live in a rhythmic world, where daily environmental variation has a profound effect on their behavior and physiology. In addition to abiotic influence, interactions with other organisms that have their own particular cycles are also part of circadian rhythm formation. In this chapter, we present aspects of the biology of mosquito vectors, more precisely Aedes aegypti, which is a vector of arboviruses of great epidemiological importance, like dengue, Zika, and chikungunya. The successful transmission of the virus depends on the coordination of several behavioral and physiological traits involved in the virus-vector-host interaction. Thus, understanding the mechanisms of endogenous control of rhythmic traits of the mosquito vector and the impact that both environmental variation and virus infection can have on this regulation is key for a reliable estimate of the vectorial capacity. We discuss the infection-driven changes in traits used to calculate parameters of the vectorial capacity, and finally, we review the current knowledge on the molecular mechanisms underlying vector rhythmic behavior and the potential cellular targets of arbovirus infection
Embryonic desiccation resistance in Aedes aegypti: presumptive role of the chitinized Serosal Cuticle
<p>Abstract</p> <p>Background</p> <p>One of the major problems concerning dengue transmission is that embryos of its main vector, the mosquito <it>Aedes aegypti</it>, resist desiccation, surviving several months under dry conditions. The serosal cuticle (SC) contributes to mosquito egg desiccation resistance, but the kinetics of SC secretion during embryogenesis is unknown. It has been argued that mosquito SC contains chitin as one of its components, however conclusive evidence is still missing.</p> <p>Results</p> <p>We observed an abrupt acquisition of desiccation resistance during <it>Ae. aegypti </it>embryogenesis associated with serosal cuticle secretion, occurring at complete germ band extension, between 11 and 13 hours after egglaying. After SC formation embryos are viable on dry for at least several days. The presence of chitin as one of the SC constituents was confirmed through Calcofluor and WGA labeling and chitin quantitation. The <it>Ae. aegypti </it>Chitin Synthase A gene (<it>AaCHS1</it>) possesses two alternatively spliced variants, <it>AaCHS1a </it>and <it>AaCHS1b</it>, differentially expressed during <it>Ae. aegypti </it>embryonic development. It was verified that at the moment of serosal cuticle formation, <it>AaCHS1a </it>is the sole variant specifically expressed.</p> <p>Conclusion</p> <p>In addition to the peritrophic matrix and exoskeleton, these findings confirm chitin is also present in the mosquito serosal cuticle. They also point to the role of the chitinized SC in the desiccation resistance of <it>Ae. aegypti </it>eggs. <it>AaCHS1a </it>expression would be responsible for SC chitin synthesis. With this embryological approach we expect to shed new light regarding this important physiological process related to the <it>Ae. aegypti </it>life cycle.</p
Avaliação do efeito de Novaluron, um inibidor da Síntese de Quitina sobre a formação de larvas de Aedes aegypti (Linnaeus, 1762)
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Previous issue date: 2014-11-18Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Rio de Janeiro, RJ, BrasilVárias populações de Aedes aegypti, vetor da dengue e da febre amarela urbana, apresentaram resistência aos inseticidas clássicos utilizados em seu controle. Sendo assim, torna-se imperativo o estabelecimento de estratégias alternativas. Nesse sentido, a biossíntese de quitina é um alvo potencial. Em artrópodes, a quitina é considerada o principal constituinte da cutícula (ou exoesqueleto). A quitina Sintase (do inglês Chitin Synthase, CHS) é uma enzima-chave na biossíntese desta molécula, e os compostos benzoil-fenil-uréias (BPUs) possuem atividade inibidora da síntese de quitina, interferindo na formação da cutícula em diversas espécies de insetos. Novaluron é um BPU recentemente recomendado pela WHO para uso em água potável, o que o qualifica como uma alternativa viável para o controle de larvas do vetor da dengue. Neste trabalho investigamos, durante o desenvolvimento larvar de Ae. aegypti, a influência de novaluron sobre: 1) o conteúdo de quitina; 2) a expressão do gene AaCHS1, evidenciada por seus transcritos alternativos AaCHS1a e AaCHS1b e, 3) a estrutura do exoesqueleto e da membrana peritrófica. Para tanto, inicialmente foi necessário observar os momentos das ecdises larvais. Os pontos definidos para a coleta das amostras levaram em conta o processo de síntese de cutícula a cada instar larvar. Foram escolhidos pontos tentativamente representativos do início (i), meio (int) e fim (f) do terceiro (L3) e do quarto (L4) instares. Os pontos de L3i, L3int, L3f, L4i, L4int e L4f foram definidos como 53, 59,5, 68, 75, 92,5 e 98 horas após a eclosão (HAE), respectivamente.
Novaluron foi então administrado continuamente a larvas L3, a partir de 51 HAE, em concentrações definidas em relação ao seu percentual de inibição de emergência do adulto (IE): IE20, IE50 e IE99. Foi observado um aumento significativo no conteúdo de quitina de larvas controle, desde L3i até L4f. Novaluron afetou significativamente a produção de quitina de forma dose-dependente, ao longo do desenvolvimento larvar. A abundância relativa de mRNA dos transcritos AaCHS1a e AaCHS1b foi avaliada por PRC quantitativo em tempo real, de L3i e L4f, em larvas controle e tratadas com dose de IE99. O perfil de expressão dos transcritos AaCHS1a e AaCHS1b é similar em larvas controle, ainda que a variação temporal de AaCHS1b seja maior. O BPU parece ter um efeito mais proeminente sobre o perfil de expressão de AaCHS1b, que se acentua nas primeiras horas após tratamento. Com base nos resultados obtidos, duas hipóteses são discutidas: mecanismo de feedback positivo ou atraso fisiológico do desenvolvimento. Ensaios histológicos preliminares validaram a metodologia utilizada para preparo e observação da morfologia de larvas tratadas e controle. Cortes longitudinais evidenciaram a preservação de estruturas internas, inclusive as quitinosas, o que abre a perspectiva de ensaios futuros de marcação específica para quitinaSeveral populations of
Aedes aegypti
, vector of dengue and urban yellow fever, exhibit
resistance to insecticides used on their control. T
herefore, the establishment of alternative strategi
es
becomes imperative. In this regard, chitin biosynth
esis is a potential target. In arthropods, chitin i
s
considered the main constituent of the cuticle (or
exoskeleton). Chitin Synthase (CHS) is a key
enzyme in the biosynthesis of this molecule and, Be
nzoyl-phenyl-urea (BPU) compounds inhibit chitin
synthesis, interfering with cuticle formation in se
veral insect species. Novaluron was recently
recommended by WHO for use in potable water, which
qualifies this BPU as a viable alternative to the
control of the dengue vector larvae. In the present
work we investigated the influence of novaluron,
during larval development of
Ae. aegypti
, on the: 1) chitin content, 2)
AaCHS1
gene expression,
through its alternative spliced forms
AaCHS1a
and
AaCHS1b
, and 3) exoskeleton and peritrophic
membrane structure. For this purpose, it was first
necessary to observe the timing of larval ecdysis.
The time points selected for sampling took into acc
ount the process of cuticle synthesis at each larva
l
instar. Tentative initial (i), intermediary (int) a
nd final (f) representative time points were chosen
for the
third (L3) and fourth (L4) larval instars. The poin
ts of L3i, L3int, L3f, L4i, L4int and L4f were defi
ned as
53, 59.5, 68, 75, 92.5 and 98 hours after hatching
(HAH), respectively. Novaluron was continuously
administered to L3 larvae, from 51 HAH on at concen
trations defined with respect to the percentage of
adult emergence inhibition (EI): EI
20
, EI
50
and EI
99
. In control larvae, a significant increase of the
chitin
content, from L3i until L4f, was observed. Novaluro
n significantly affected chitin production in a dos
e
dependent manner, during larval development. The re
lative abundance of
AaCHS1a
and
AaCHS1b
transcripts was evaluated by quantitative real time
PCR, from L3i to L4f, on both control and novaluro
n
IE
99
larvae. The expression of
AaCHS1a
the treated larvae were similar to control. The ex
pression
profile of both
AaCHS1a
and
AaCHS1b
transcripts is similar in control larvae, even if
variations in
AaCHS1b
are higher. BPU treatment effect seems more pronou
nced on
AaCHS1b
expression profile,
that is activated in the first hours immediately fo
llowing treatment. Two hypotheses are raised: a
positive feedback mechanism or a physiological deve
lopment delay. Preliminary histological
observations validated the methodology employed for
preparing and observing control and treated
larvae morphology. Longitudinal sections put in evi
dence the preservation of internal structures,
including chitinized ones, opening the perspective
of future assays for specific chitin
Culicídeos vetoresdiferenças e semelhanças fisiológicas e estruturais relacionadas ao processo de resistência dos ovos à dessecação
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Previous issue date: 2016-01-13Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Rio de Janeiro, RJ, Brasil.Diversos parasitas, agentes etiológicos de uma série de agravos, são transmitidos por mosquitos vetores em todo o mundo. A despeito de sua importância médica, ainda existem muitas lacunas no conhecimento da embriogênese desses insetos. Ovos de espécies do gênero Aedes resistem à dessecação, e são capazes de sobreviver vários meses no seco em estado de quiescência ao final da embriogênese. Entretanto, ovos de espécies de outros gêneros, como Anopheles e Culex, não possuem capacidade de resistir a longos períodos de exposição ao seco, apesar de adquirirem algum grau de impermeabilidade. Nos mosquitos Aedes aegypti e Anopheles gambiae tem sido mostrado que, durante a embriogênese, a síntese da cutícula serosa (CS), que contém quitina, coincide com a aquisição da resistência dos ovos à dessecação (ROD). O objetivo deste trabalho foi investigar, comparativamente, a embriogênese de diferentes mosquitos vetores, para elucidar semelhanças e diferenças na ROD. A maior parte do trabalho foi realizada com as espécies Ae. aegypti, An. aquasalis e Cx. quinquefasciatus. Verificamos que para An. aquasalis e Cx. quinquefasciatus a formação da CS também é simultânea à aquisição de ROD e contém quitina. O tempo requerido para a finalização da embriogênese de Ae. aegypti, An. aquasalis e Cx. quinquefasciatus é, respectivamente, 77,4, 51,3, e 34,2 horas após a postura
A formação da CS ocorre em estágio embrionário equivalente para Ae. aegypti e An. aquasalis e um pouco mais tardio para Cx. quinquefasciatus. Ao final da embriogênese diferentes níveis de ROD foram detectados: alto para Ae. aegypti, médio para An. aquasalis e baixo para Cx. quinquefasciatus. Com o objetivo de entender essas diferenças, avaliamos, comparativamente, alguns aspectos físicos dos ovos: comprimento, largura, área, volume, peso e a superfície da casca. Apesar de diferenças interessantes e significativas terem sido observadas, não foi possível estabelecer correlação direta entre ROD e tais aspectos, quando avaliados individualmente. Quantificação do polissacarídeo quitina revelou tendência de maior conteúdo em cascas de ovos de espécies mais resistentes ao seco. Além disso, diferenças significativas foram encontradas no grau de melanização dos ovos das diferentes espécies, sugerindo que ovos mais melanizados são mais resistentes ao seco. Essa hipótese foi testada utilizando o mutante GORO da espécie An. quadrimaculatus. Nesta linhagem a melanização não ocorre corretamente, e as cascas de seus ovos têm aparência dourada. Em contraste, seu equivalente selvagem (WT) possui ovos de casca escura
Ovos GORO possuem ROD deficiente: quando expostos ao seco por 10 horas após a formação da CS, apenas 12,3% são viáveis, em contraste com 66,5% de ovos WT. Nossos resultados confirmam de forma direta que, além da quitina da CS, a melanização da casca dos ovos é importante para a impermeabilização dos ovos de insetosSeveral parasites, etiologic agents of a number of diseases, are transmitted by mosquito vectors worldwide. Despite their medical importance, many gaps in our knowledge about the embryogenesis of these insects still remain. Eggs from the Aedes genus resist to desiccation, and are able to survive several months in dry conditions in a quiescent status at the end of embryogenesis. In contrast, eggs belonging to other genera, like Anopheles and Culex, do not withstand long periods of exposure in dry conditions, although they acquire some degree of impermeability. It has already been demonstrated that, in the course of Aedes aegypti and Anopheles gambiae embryogenesis, the synthesis of the serosal cuticle (SC), which contains chitin, coincides with the acquisition of egg resistance to desiccation (ERD). The aim of the present work was to investigate, under a comparative basis, the embryogenesis of different mosquito vectors, in order to elucidate ERD similarities and differences. Most of the work was performed with Ae. aegypti, Anopheles aquasalis and Culex quinquefasciatus. We found that, for An. aquasalis and Cx. quinquefasciatus, ERD acquisition is also simultaneous to the development of SC, a layer containing chitin. Time required for embryogenesis completion is 77.4, 51.3, and 34.2 hours after egglaying for respectively Ae. aegypti, An. aquasalis and Cx. quinquefasciatus. The formation of SC occurs in an equivalent embryonic stage for Ae. aegypti and An. aquasalis, but a little later for Cx. quinquefasciatus. At the end of embryogenesis different ERD levels were detected: high for Ae. aegypti, medium for An. aquasalis and low for Cx. quinquefasciatus. In order to understand these differences, we comparatively evaluated some physical aspects of the eggs: length, width, area, volume, weight and eggshell surface appearance. Although interesting and significant differences have been observed, it was not possible to establish any direct correlation between ERD and these physical aspects, when individually considered. Quantification of chitin revealed tendency of higher amount of this polysaccharide in the eggshells of species more resistant to desiccation. Furthermore, significant differences were found in the degree of melanization of eggs from the different species, suggesting that more melanized eggs are also more resistant to dry conditions. This hypothesis was tested using the An. quadrimaculatus GORO mutant. In this strain, melanization does not occur properly, and the eggs have a golden appearance. Eggshells from the wild type (WT) strain, in contrast, are dark. GORO eggs exhibit a defective ERD: when exposed to dry conditions during 10 hours after the SC formation only 12.3% are viable, in contrast with 66.5% of WT eggs. Our results confirm, directly, that beyond the chitin present in the SC, the eggshell melanization is an important factor towards insect eggs impermeabilit
Experiências em divulgação científica e sensibilização da população: importância do controle mecânico do vetor Aedes aegypti
Aedes aegypti é o principal vetor de dengue, Zika e chikungunya, doenças para as quais não existem vacinas totalmente eficazes. Alternativas de controle visando mitigar essas arboviroses são primordiais. Dentre essas, o controle mecânico aborda práticas de eliminação e/ou limpeza de criadouros do vetor. Neste relato, apresentamos e avaliamos criticamente ações realizadas pelo grupo, ocorridas entre 2016 e 2019, onde divulgamos informação científica clara através do diálogo com a população. Os métodos utilizados foram: 1) Palestras em escolas (público infantojuvenil) utilizando slides, fotos e vídeos; 2) Oficinas (público misto), stand com material in vivo do ciclo de vida do Aedes, jogos e desenhos. Analisamos dez palestras em escolas do ensino fundamental e médio e 20 oficinas realizadas em diferentes regiões do Brasil. Concluímos que tais ações e suas análises críticas devem ser realizadas continuamente para que sejam bem-sucedidas
Embryonic development of Aedes aegypti (Diptera: Culicidae): influence of different constant temperatures
Despite its vector importance little attention is given to Aedes
aegypti embryonic development. In this study, temperature influence
on time course of Ae. aegypti larvae hatching and egg viability were
evaluated. The dormancy state at the end of embryogenesis could be
interrupted with a proper stimulus. Temperatures tested ranged between
12-36°C; the maximum temperature limit is 35°C and the
minimum one is below 12°C. Egg viability between 16-31°C was
above 80%. The definition of physiological embryonic parameters at this
temperature range corroborates Ae. aegypti presence on tropical and
subtropical world regions
The influence of a light and dark cycle on the egg laying activity of Aedes aegypti (Linnaeus, 1762) (Diptera: Culicidae)
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Previous issue date: 2018Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Biologia Molecular de Insetos. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Biologia Molecular de Insetos. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Biologia Molecular de Insetos. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Biologia Molecular de Insetos. Rio de Janeiro, RJ, Brasil / Instituto Nacional de Ciência e Tecnologia em Entomologia Molecular. Rio de Janeiro, RJ, Brasil.The epidemiological importance of the mosquito Aedes aegypti as a vector of multiple human pathogens has generated a growing number of studies on the physiology and behaviour of its blood-feeding females. The activity of oviposition is one of the critical elements contributing to the expansion of Ae. aegypti's populations. Although there is a vast literature about oviposition behaviour, significant specific knowledge about egg viability and female fertility under light and dark conditions is still lacking
Serosal cuticle formation and distinct degrees of desiccation resistance in embryos of the mosquito vectors Aedes aegypti, Anopheles aquasalis and Culex quinquefasciatus
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Previous issue date: 2014Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Fisiologia e Controle de Artrópodes Vetores. Rio de Janeiro, RJ, Brasil / Instituto de Biologia do Exército. Laboratório de Entomologia. Rio de Janeiro, RJ, Brasil / Universidade Estadual do Norte Fluminense Darcy Ribeiro. Centro de Biociências e Biotecnologia. Laboratório de Química e Função de Proteínas e Peptídeos. Campos dos Goytacases, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Fisiologia e Controle de Artrópodes Vetores. Laboratório de Biologia Molecular de Flavivírus. Rio de Janeiro, RJ, Brasil / Instituto de Biologia do Exército. Laboratório de Entomologia. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Fisiologia e Controle de Artrópodes Vetores. Rio de Janeiro, RJ, Brasil / Instituto de Biologia do Exército. Laboratório de Entomologia. Rio de Janeiro, RJ, Brasil / Instituto Nacional de Ciência e Tecnologia em Entomologia Molecular. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Biologia Molecular de Flavivírus. Rio de Janeiro, RJ, Brasil / Instituto Nacional de Ciência e Tecnologia em Entomologia Molecular. Rio de Janeiro, RJ, Brasil.Universidade Estadual do Norte Fluminense Darcy Ribeiro. Centro de Biociências e Biotecnologia. Laboratório de Química e Função de Proteínas e Peptídeos. Campos dos Goytacases, RJ, Brasil / Instituto Nacional de Ciência e Tecnologia em Entomologia Molecular. Rio de Janeiro, RJ, Brasil.Given their medical importance, mosquitoes have been studied as vectors of parasites since the late
1800’s. However, there are still many gaps concerning some aspects of their biology, such as embryogenesis.
The embryonic desiccation resistance (EDR), already described in Aedes and Anopheles gambiae mosquitoes,
is a peculiar trait. Freshly laid eggs are susceptible to water loss, a condition that can impair their
viability. EDR is acquired during embryogenesis through the formation of the serosal cuticle (SC), protecting
eggs from desiccation. Nevertheless, conservation of both traits (SC presence and EDR acquisition)
throughout mosquito evolution is unknown. Comparative physiological studies with mosquito embryos
from different genera, exhibiting distinct evolutionary histories and habits is a feasible approach. In this
sense, the process of EDR acquisition of Aedes aegypti, Anopheles aquasalis and Culex quinquefasciatus at
25 C was evaluated. Completion of embryogenesis occurs in Ae. aegypti, An. aquasalis and Cx. quinquefasciatus
at, respectively 77.4, 51.3 and 34.3 hours after egg laying, Cx. quinquefasciatus embryonic development
taking less than half the time of Ae. aegypti. In all cases, EDR is acquired in correlation with SC
formation. For both Ae. aegypti and An. aquasalis, EDR and SC appear at 21% of total embryonic development,
corresponding to the morphological stage of complete germ band elongation/beginning of germ
band retraction. Although phylogenetically closer to Ae. aegypti than to An. aquasalis, Cx. quinquefasciatus
acquires both EDR and serosal cuticle later, with 35% of total development, when the embryo already progresses
to the middle of germ band retraction. EDR confers distinct egg viability in these species. While
Ae. aegypti eggs demonstrated high viability when left up to 72 hours in a dry environment, those of An.
aquasalis and Cx. quinquefasciatus supported these conditions for only 24 and 5 hours, respectively. Our
data suggest that serosa development is at least partially uncoupled from embryo development and that,
depending upon the mosquito species, EDR bestows distinct levels of egg viability
Physical features and chitin content of eggs from the mosquito vectors Aedes aegypti, Anopheles aquasalis and Culex quinquefasciatus: Connection with distinct levels of resistance to desiccation
Physical features and chitin content of eggs from the mosquito vectors Aedes aegypti, Anopheles aquasalis and Culex quinquefasciatus: Connection with distinct levels of resistance to desiccation
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Previous issue date: 2015Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Bilogia Molecular de Insetos. Rio de Janeiro, RJ, Brasil / Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Fisiologia e Controle de Artrópodes Vetores. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Biologia Celular. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Fisiologia e Controle de Artrópodes Vetores. Rio de Janeiro, RJ, Brasil / Instituto Nacional de Ciência e Tecnologia em Entomologia Molecular. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Biologia Molecular de Flavivírus. Rio de Janeiro, RJ, Brasil / Instituto Nacional de Ciência e Tecnologia em Entomologia Molecular. Rio de Janeiro, RJ, Brasil.Universidade Estadual do Norte Fluminense Darcy Ribeiro. Centro de Biociências e Biotecnologia. Laboratório de Química e Função de Proteínas e Peptídeos. Campos de Goytacazes, RJ, Brasil / Instituto Nacional de Ciência e Tecnologia em Entomologia Molecular. Rio de Janeiro, RJ, Brasil.Mosquito eggs are laid in water but freshly laid eggs are susceptible to dehydration, if their surroundings
dry out at the first hours of development. During embryogenesis of different mosquito vectors the serosal
cuticle, an extracellular matrix, is produced; it wraps the whole embryo and becomes part of the eggshell.
This cuticle is an essential component of the egg resistance to desiccation (ERD). However, ERD is variable
among species, sustaining egg viability for different periods of time. While Aedes aegypti eggs can survive
for months in a dry environment (high ERD), those of Anopheles aquasalis and Culex quinquefasciatus in the
same condition last, respectively, for one day (medium ERD) or a few hours (low ERD). Resistance to desiccation
is determined by the rate of water loss, dehydration tolerance and total amount of water of a
given organism. The ERD variability observed among mosquitoes probably derives from diverse traits.
We quantified several attributes of whole eggs, potentially correlated with the rate of water loss: length,
width, area, volume, area/volume ratio and weight. In addition, some eggshell aspects were also evaluated,
such as absolute and relative weight, weight/area relationship (herein called surface density) and
chitin content. Presence of chitin specifically in the serosal cuticle as well as aspects of endochorion
external surface were also investigated. Three features could be related to differences on ERD levels:
chitin content, directly related to ERD, the increase in the egg volume during embryogenesis and the eggshell
surface density, which were both inversely related to ERD. Although data suggest that the amount of
chitin in the eggshell is relevant for egg impermeability, the participation of other yet unidentified eggshell
attributes must be considered in order to account for the differences in the ERD levels observed
among Ae. aegypti, An. aquasalis and Cx. quinquefasciatus