Dengue viruses infect up to 50 million people each year, causing more than 20,000 deaths [1], [2]. These flaviviruses are mainly transmitted by Aedes aegypti, but also Ae. albopictus [3]. Native to the Oriental Region and some islands in the Indian Ocean [3], Ae. albopictus has become well-established in the Western hemisphere where it is the second main vector of dengue [4]. It is also an important vector of yellow fever and various types of encephalitis virus, as well as a competent vector of at least 23 other arboviruses under laboratory conditions [5], [6], [7]. It is well adapted to peridomestic environments with its larvae breeding in artificial containers and adults aggressively biting humans and different animals during the day [3].

Efforts to control dengue have mainly involved insecticide spraying programs, but this strategy has proven ineffectual [8]. While a vaccine is currently under development, without immediate prospects for success, vector control remains the only viable method to prevent dengue transmission [9], [10], [11], [12]. Improved knowledge regarding egg-laying behavior is relevant because it underpins the primary surveillance method, i.e., ovitraping [13], [14]. However, the most commonly used ovitrap, the CDC gravid trap, is not appropriate for capturing Ae. albopictus [15], [16].

Blood feeding in mosquitoes represents phenotypic expression of reproductive investment as it is the acquisition of resources specifically for reproduction [17]. Reproductive output represents the energy allocated to egg production and oviposition that could otherwise be allocated to maintenance of somatic function, and the act of oviposition is associated with a risk to survival [18]. There has been a great deal of research regarding the variations of reproductive investment and outcome. Overall, increases in both number and size of blood meals result in increased individual egg mass and number of eggs [18]. Clearly, in the field an increased frequency of blood uptake will tend to require host – mosquito contact and expose hosts to a greater risk of disease transmission.

Ae. albopictus has been occasionally incriminated in dengue epidemics in Asian countries [19], [20], [21]. The first report of a dengue epidemic in Malaysia was on Penang Island, and dengue fever with hemorrhagic manifestations was also reported from this Island [22]. These epidemics were accompanied by increased populations of Ae. albopictus. Increased invasiveness [23] and increased population densities of [24] have been recently been reported in the Malaysian peninsula. Ae. albopictus females was commonly observed developing in indoor/domestic sites throughout Penang Island during our 2009 entomological survey at Balik Pulau (Sungai Pinang and Sg. Burung), Gelugor, Jelutong, Air Itam (Kampung Relau), and other sites. This shift from outside to inside human dwellings is likely to increase the opportunities for females to obtain blood meals. This indoor immature breeding has raised the question of whether this mosquito may exhibit increased gonotrophic activity (GA) in response to potentially greater blood meal sources.

As the present study was begun with a wild population, it was first necessary to determine whether they could oviposit under laboratory conditions. Oviposition trials indicated that FWMs can lay eggs in the laboratory and that the patterns of egg–laying were associated with blood feeding time. The findings of the present study were consistent with the natural oviposition behavior of this mosquito [38], [39], [40], [41], therefore allowing the long-term experiments required for this study.

We found a major effect of level of adaptation to indoor/domestic environment on the number of GCs in the laboratory. d5FWMs showed a much higher number of GCs than their FWMs counterparts. In mosquitoes, the nutritional history of the parents is influential in determining the fecundity of daughters [42]. This study showed that daughters from parents reared in a food-limited environment produced more eggs than those from parents reared under high food conditions. They suggested that this increased fecundity arose to compensate for expected decreased longevity in stressing environments. Here, females derived from wild mosquitoes (FWMs) and females derived from these FWMs after five generations achieved 7 and 14 GCs, respectively. d5FWMs survival rate was higher than that of FWMs mosquitoes. Although the mechanisms underlying these observations are not yet clear, they could be the result of at least two processes. First, the highly nutritious food conditions in the laboratory could lengthen the mosquito lifespan, and thus increase the probability that it reproduces. Second, the shift from a complex wild environment to a simple environment, such as that in the laboratory, may result in physiological changes that increase the allocation of energy to functions other than egg production, thereby increasing the probability of survival. Longer living females may take more blood meals and reproduce more simply because of the increased availability of meals. Epidemiologically, an increased number of GCs will tend to increase the probability of disease occurrence. With an extended period of GA, females have a higher probability of picking up and transmitting a disease agent as well as an increased lifespan as blood provides an alternative energy source for survival [43]. The increased period of reproduction of the d5FWMs will also lead to a higher mosquito population density, which is also likely to be associated with increased occurrence of disease. Population levels expressed as larval [44], [45], [46], pupal [47], and adult indices [48] are often associated with levels of risk for dengue transmission. Dengue outbreaks occurred in Singapore [49] but not in Brazil [50] when the national overall percentage of houses positive for larvae (HI) was below 1%. In Puerto Rico, the incidence of dengue increased one month after larval density peaked [51], whereas in Brazil, dengue seroincidence increased when the HI was above 3% [52]. The present study was prompted by the permanent presence of biting adults and immature stages of Ae. albopictus within residences in Teluk Tempoyok and Balik Pulau located in northern peninsular Malaysia and a lack of information regarding the epidemiological implications. Although we did not determine whether these populations were infected with dengue viruses, the focal point of this study was that the presence of larvae strongly suggests that at least a GC has been achieved. Thus, infection would occur if the virus was present. Note that this species is competent for many viruses [6], [7], but has only occasionally been incriminated in minor dengue epidemics all the world, e.g., in Hawai in 2001–2002 [53].

In FWMs and d5FWMs, the number of eggs oviposited tended to decrease as GC rank progressed, but this effect was most marked in the second group. As in most anautogenous mosquitoes, the production of eggs in Ae. albopictus requires the ingestion of blood [54], [55]. The female converts about 20% of the ingested blood meal into egg constituents [56]. Several groups have reported that the degree to which eggs are produced depends largely on meal size [32], [57], [58]. Indeed, a female that ingests a large blood meal size will tend to invest more in egg production than a female with a small blood meal. In the present study, we have used females adapted to laboratory conditions and females derived from wild pupae. It is often assumed that wild insects are subject to much harsher environmental conditions that trigger small body size and that they have lower levels of energetic reserves than laboratory strains. Clearly, in the laboratory, the highly nutritious larval diet will tend to produce large bodied mosquitoes capable of blood feeding for long periods due to little or no host defense behavior from anesthetized hosts [32]. Here, FWMs and d5FWMs were similar in body size and had the same feeding time, so differential egg production due to differences in meal size is unlikely. These discrepancies may be explained by differential utilization of blood. Adult mosquitoes feed on blood for immediate energy needs [57], [59], but in some cases, they use blood as an alternative energy source for survival [43]. There is evidence that colonization alters reproductive traits [60], in particular offspring fecundity [42]. Therefore, it is tempting to suggest that the accommodation of the wild strain to the laboratory environment has occurred in addition to physiological changes relative to blood use. It is possible that the reduced level of egg production observed with increasing generations represented a compensation for better acclimation to the laboratory environment. Presumably, protein use by d5FWMs offsets the costs associated with egg production and facilitates population maintenance in this environment. In support of this suggestion, it has been reported that Ae. albopictus may use some blood proteins for maintenance [61].

The pattern of egg production was similar between females of both types, but lifetime fecundity was greater in d5FWMs. This difference was the result of their greater survival. Females with a long lifespan may take more blood meals and reproduce more simply because of the increased availability of meals. Easy access to blood sources in any host – vector interaction can be of crucial epidemiological significance because increased frequency of host-biting may favor the spread of infectious disease present. The presence of Ae. albopictus inside houses, that was observed in many residences throughout Penang Island, appears to facilitate human blood feeding. In this context, biting activity during both the day and night may increase mosquito – human contact. In addition, dengue viruses can be transmitted sexually from male to female Ae. albopictus [62]. Therefore, the increased fecundity of d5FWMs may contribute to virus propagation.

This study emphasizes the invasive properties of Ae. albopictus and importantly shows the acquisition of an indoor breeding behavior by this major vector of dengue viruses. This behavioral change may lead to increased vectorial capacity. Several parameters come into play when the vectorial capacity of a mosquito for an arbovirus is considered [63]. In particular, host availability and population density are very influential to the competence of a vector. Indeed the more individual vectors are present, the more likely they will be able to transmit a pathogen. Adaption to the indoor/domestic environment, which triggers increased human-vector contacts, will presumably stimulate feeding behavior. In the neighboring Thailand, 100% of field-collected populations of this mosquito fed on humans [64]. Theoretically, such an affinity for feeding on human blood in a wild context will tend to increase inside residences. Ae. albopictus exhibited a high biting activity, showing a shorter feeding time and a greater blood feeding success when compared to Ae. aegypt at night. This period is the time when residents exhibit low defensive responses to mosquito feeding. In our study, the epidemiological implications were also approached from gonotrophic performance and survival because biting activity is a pivotal factor in the continuation of both pathogen transmission and vector generation [65]. Furthermore, the mosquito individuals that are most easily infected and most likely to incubate a pathogen to an infectious level are those that live long enough [66]. Clearly, adaption to the indoor/domestic environment may produce more competent vectors, since it favors long life and increased lifetime reproductive output.