The results of this study demonstrate that two local Aedinae of the temperate zone,
Ae. notoscriptus and
Ae. camptorhynchus, can be infected by Zika virus and deliver infectious virus in their saliva. Similar to the results of Hall-Mendelin et al. [
18] for
Ae. notoscriptus and
Aedes vigilax (a saltmarsh species belonging to the same subgenus as
Ae. camptorhynchus), our results showed moderate infection rates and low dissemination rate for these
Aedes species. However, in contrast to Hall-Mendelin et al., our results showed presence of virus in the saliva of these mosquitoes’ Victorian populations: 42% and 13.5% of
Ae. notoscriptus and
Ae. camptorhynchus, respectively. The prevalence of virus in the saliva was lower than that of
Ae. aegypti (87%) (Table
1). Hall-Medellin et al. reported viral RNA prevalence in saliva at 27% for
Ae. aegypti at D14. Our results are closer to the 100% transmission results reported by Li et al. [
7] who, as we did for saliva, used virological methods (CPE) rather than molecular. Our qPCR molecular test was assessed for relative sensitivity by a dilution curve which showed positive results down to 10
−4-fold dilution of the initial cDNA from a TCID50 10
6/ml stock solution, being inconsistent at the 10
−5 and negative at the 10
−6 dilution (Fig.
3a). As a further insight of the viral dynamics within the mosquitoes, we tested three whole samples of freshly blood-fed
Ae. camptorhynchus which gave an average CT value of 30.03 (SD 1.12), as a relative measure of the initial virus load.
The specimens of the population of this mid-sized species were among the larger of tested samples of the different species. The quantity of ingested virus was probably among the higher. The average CT value for
Ae. camptorhynchus midgut samples is lower than this threshold, indicating a probable local replication after the blood meal. For
Ae. notoscriptus, the midgut average CT values are similar to the viral input average represented by the whole freshly blood-fed
Ae. camptorhynchus specimens (Fig.
3b). However, when positive, the carcass of
Ae. notoscriptus shows lower CT values, indicative of replication (Fig.
3c). Our molecular test could have missed some specimens with low level virus load, either in the midgut or in the carcass. The virological method of virus detection by CPE could be a more robust and reliable method and the most informative indicator of transmission.
Beyond the different methodological approaches, the different results between the vector competence assessments performed on two different populations of
Ae. notoscriptus, from either tropical or temperate zones, could be linked to the vector’s differences in genetic background. Indeed, Endersby et al. [
41] hypothesised that
Ae. notoscriptus could be a complex of cryptic species. Also, screening for
Wolbachia by conventional PCR on individual carcass (including ovaries) cDNA shows a very low prevalence of
Wolbachia for the temperate zone population of
Ae. notoscriptus and absence of positive molecular signal on pools of 5 cDNAs by qPCR. In contrast, a high prevalence of
Wolbachia natural infection has previously been described in tropical
Ae. notoscriptus populations either from a colony or field samples collected in Brisbane [
42], with this being the same origin as the tropical population tested for Zika virus vector competence [
18]. Although natural
Wolbachia infection did not influence the dengue virus competence of infected
Ae. notoscriptus populations [
42], our study cannot dismiss the possibility of a potential protective effect of natural
Wolbachia infection on
Ae. notoscriptus for Zika virus infection and could justify further research. It is to be noted that, despite the distance,
Ae. notoscriptus has been described as potentially invasive for USA [
43].
Aedes notoscriptus from Brisbane has been shown to transmit yellow fever virus better than
Ae. aegypti [
44], however
Ae. notoscriptus of unknown origin failed to develop infections with the same virus [
45].
Aedes notoscriptus from Brisbane area has been previously shown as a poor vector for the four serotypes of dengue virus [
46]. Indeed, populations of this mosquito in its southern distribution range, e.g. the state of Victoria, have not been documented for flavivirus vector competence. No dengue outbreak or documented secondary cases of dengue have been recorded in this region, in the absence of the known dengue vector
Ae. aegypti. The contact between human populations and
Ae. notoscriptus has been documented in Queensland but not in temperate regions.
Aedes notoscriptus has been shown to bite humans 19% of the time for blood meals in residential areas of Brisbane [
47]. Its dispersal and survival rates in the same area [
48] are similar to
Ae. aegypti and identify it as a potential vector in urban areas. Although it is known to be a backyard pest in the southern zones, much of its biology remains to be well defined in order to properly assess risk for transmission. The case of
Ae. camptorhynchus is quite different: its ecology is restricted to coastal areas, or around inland brackish zones [
49], with a lower abundance in urban areas [
50], with local exceptions in suburb areas around Melbourne, Victoria (SL, pers. Comm.). It has been shown to be an efficient experimental vector for the Alphavirus, Ross River virus [
51], and epidemiologically important in the natural transmission of this virus [
37]. This includes the possibility of vertical transmission for Ross River and Sindbis viruses [
52]. Except this current study, its status as vector for flavivirus is unknown. Similar to
Ae. notoscriptus, but to a lesser degree,
Ae. camptorhynchus showed intermediate levels of vector competence for Zika virus. Both are probably not primary vectors, meaning they may be unable to sustain large outbreaks on their own, unlike
Ae. aegypti in tropical areas, however they could be potential candidates to trigger few secondary cases and reinforce outbreaks in the presence of primary vectors.
Aedes notoscriptus is known as a backyard pest, due to its abundance and taste for human blood.
Aedes camptorhynchus has been shown to be responsible for Ross River virus outbreaks, here too displaying a close proximity with humans. However, a big gap in our knowledge remains: the experiments were performed at 27.5 °C. Whilst the state of Victoria can experience very high temperatures and heat waves, on average, summer temperatures are much lower than 25 °C. Performing vector competence experiments at lower temperatures would be useful in establishing a proper risk assessment for local transmission of Zika virus, both by local and potentially invasive species.