María V. Micieli

Global genetic diversity of Aedes aegypti

Mosquitoes, especially Aedes aegypti, are becoming important models for studying invasion biology. We characterized genetic variation at 12 microsatellite loci in 79 populations of Ae. aegypti from 30 countries in six continents, and used them to infer historical and modern patterns of invasion. Our results support the two subspecies Ae. aegypti formosus and Ae. aegypti aegypti as genetically distinct units. Ae. aegypti aegypti populations outside Africa are derived from ancestral African populations and are monophyletic. The two subspecies co‐occur in both East Africa (Kenya) and West Africa (Senegal). In rural/forest settings (Rabai District of Kenya), the two subspecies remain genetically distinct, whereas in urban settings, they introgress freely. Populations outside Africa are highly genetically structured likely due to a combination of recent founder effects, discrete discontinuous habitats and low migration rates. Ancestral populations in sub‐Saharan Africa are less genetically structured, as are the populations in Asia. Introduction of Ae. aegypti to the New World coinciding with trans‐Atlantic shipping in the 16th to 18th centuries was followed by its introduction to Asia in the late 19th century from the New World or from now extinct populations in the Mediterranean Basin. Aedes mascarensis is a genetically distinct sister species to Ae. aegypti s.l. This study provides a reference database of genetic diversity that can be used to determine the likely origin of new introductions that occur regularly for this invasive species. The genetic uniqueness of many populations and regions has important implications for attempts to control Ae. aegypti, especially for the methods using genetic modification of populations.

Somatic Wolbachia (Rickettsiales: Rickettsiaceae) Levels in Culex quinquefasciatus and Culex pipiens (Diptera: Culicidae) and Resistance to West Nile Virus Infection

ABSTRACT The endosymbiotic bacteria Wolbachia pipientis Hertig infects a wide variety of insect species and can increase viral resistance in its host. Wolbachia naturally infects Culex quinquefasciatus Say and Culex pipiens L. mosquitoes, both vectors of West Nile virus (WNV). We recently demonstrated that Wolbachia infection of Cx. quinquefasciatus laboratory strain Ben95 increases host resistance to WNV infection, reducing vector competence. This observation raised the possibility that Wolbachia could impact vector competence in other populations of Cx. quinquefasciatus or Cx. pipiens. To investigate this possibility, Wolbachia densities were measured in Ben95 Cx. quinquefasciatus and compared with densities in a newly established colony of Cx. quinquefasciatus, and in field-collected and colonized Cx. pipiens. Wolbachia densities in somatic tissues of Ben95 Cx. quinquefasciatus were significantly higher than densities in the other mosquito populations tested. There was also no significant spatiotemporal variation in Wolbachia density in the field-collected Cx. pipiens, although significant familial differences were observed. Correlating Wolbachia densities and vector competence in individual colonized Cx. pipiens indicated that the densities of somatic Wolbachia observed in the mosquitoes other than Ben95 Cx. quinquefasciatus were too low to inhibit WNV infection and reduce vector competence. These results suggest that the high Wolbachia densities capable of inducing resistance to WNV in Ben95 Cx. quinquefasciatus sue not a general characteristic of Cx. quinquefasciatus or Cx. pipiens mosquitoes and that the impact of Wolbachia on vector competence in field populations of Cx. quinquefasciatus and Cx. pipiens, if any, is likely to be limited to specific populations.

Life cycle and description of Amblyospora camposi n. sp. (Microsporidia: Amblyosporidae) in the mosquito Culex renatoi (Diptera, Culicidae) and the copepod Paracyclops fimbriatus fimbriatus (Copepoda, Cyclopidae).

Abstract The life cycle of Amblyospora camposi n. sp. is described from the mosquito Culex renatoi and the copepod Paracyclops fimbriatus fimbriatus collected in the leaf axils of the plant Eryngium cabrerae in Argentina. Meiospores of A. camposi (5.8 × 4.1 μm) were infectious per os to female adults of the copepod P. f. fimbriatus. All developmental stages in the copepod had unpaired nuclei, with sporulation involving the formation of a sub-persistent, sporontogenic, interfacial envelope and the production of a second type of uninucleate spore. These spores, formed in the ovaries of P. f. fimbriatus, were large, pyriform, and measured 10.70 × 3.85 μm. When ingested they infected C. renatoi larvae to initiate a sequence that involves schizogony and gametogony and ends with plasmogamy and nuclear association to form diplokaryotic meronts. Oblong ovate binucleate spores (7.86 × 2.96 μm) are formed in the adult mosquito and are responsible for vertical transmission to the filial generation. This is the first report of an Amblyospora species from a mosquito that inhabits the small-water bodies held in parts of terresterial plants (phytotelmata).

Laboratory evaluation of Mesocyclops annulatus (Wierzejski, 1892) (Copepoda: Cyclopidea) as a predator of container-breeding mosquitoes in Argentina

In laboratory bioassays we tested the predatory capacity of the copepod Mesocyclops annulatus on Aedes aegypti and Culex pipiens larvae. A single adult female of M. annulatus caused 51.6% and 52.3% mortality of 50 first instar larvae of Ae. aegypti and Cx. pipiens respectively, in a 72 h test period. When alternative food was added to the containers, mortality rates declined to 16% and 10.3% for Ae. aegypti and Cx. pipiens respectively. When 50 first instar larvae of each of the two mosquito species tested were placed together with a single adult female of M. annulatus, mortality rates were 75.5% for Ae. aegypti larvae and 23.5% for Cx. pipiens larvae in a three day test period. Different density of adult females of M. annulatus ranged from 5 to 25 females produced mortality rates of Ae. aegypti first instar larvae from 50% to 100% respectively. When a single adult female of M. annulatus was exposed to an increasing number of first-instar Ae. aegypti larvae ranging from 10 to 100, 100% mortality was recorded from 1 to 25 larvae, then mortality declined to 30% with 100 larvae. The average larvae killed per 24 h period by a single copepod were 29.

SEASONALITY AND ABUNDANCE OF THE MOSQUITO ISOSTOMYIA PARANENSIS FROM PHYTOTELMATA IN TEMPERATE ARGENTINA

ABSTRACT Sampling was conducted for 1 year in a marsh near Buenos Aires from axils of Scirpus giganteus, a larval habitat of the poorly known sabethine mosquito, Isostomyia paranensis. Immatures of this species were recovered on every sampling date, averaging 3–4/plant in April and decreasing to 0–1/plant in October–December. The spatial distribution of Is. paranensis immatures was clumped, and larval age skewed toward 1st instars. The percentage of mosquito-positive S. giganteus was negatively correlated with accumulated rainfall 1 wk before collection. Microcrustacea were the only other invertebrates common in this phytotelmata, and no parasites or pathogens were detected in Is. paranensis. Fourth instars of this species attacked and killed one another in the laboratory, but only algae were recovered from dissected digestive tracts of field-collected larvae. Adult females of this species emerged from independent collections of pupae refused blood, but females captured at human bait readily consumed human blood. Mean (±SD) number of eggs developed by females collected at human bait and fed with blood (77.4 ± 22.8) was not significantly different from the mean number of eggs developed by females collected as pupae and fed on sugar (72.0 ± 23.0).

Susceptibility of neotropical mosquito larvae (Diptera: Culicidae) and non-target aquatic organisms to the entomoparasitic nematode Strelkovimermis spiculatus Poinar & Camino, 1986 (Nematoda: Mermithidae)

Fourteen species of mosquito and several species of non-target aquatic organisms were tested for susceptibility to the neotropical mermithid nematode, Strelkovimermis spiculatus . All species of Aedes, Anopheles, Culex, Isostomyia and Ochlerotatus, plus one of the two Psorophora species exposed to S. spiculatus preparasites, were parasitised. None of the non-target aquatic organisms exposed to S. spiculatus were infected, with the exception of Chironomidae larvae in which nematode penetration was observed although host reaction in the form of nematode melanisation occurred 24-48 h post-infection, the nematodes dying within the host.

Evaluation of Mesocyclops annulatus (Copepoda: Cyclopoidea) as a control agent of Aedes aegypti (Diptera: Culicidae) in Argentina

We evaluated the potential of Mesocyclops annulatus as a control agent of Aedes aegypti in La Plata city (Argentina). Mosquito larval survivorship due to predation by these copepods was estimated at weekly intervals during the oviposition period of A. aegypti. Mean weekly A. aegypti larval survivorship in cylindrical plastic containers (12 cm height and 11 cm diameter) with copepods was significantly lower than in control containers. Furthermore, weekly larval survival was negatively correlated with M. annulatus adult density, and approximately 23 adult copepods/container would be a threshold density over which the weekly mosquito larval survivorship approached zero. The copepods were able to persist in all containers during approximately 100 days (in three of them until the end of the experiment: 155 days) without the resource represented by A. aegypti larvae. The predation and persistence observed suggest that M. annulatus is a potential control agent to be considered in biological control programs.

Pre-parasitic juveniles of Strelkovimermis spiculatus Poinar & Camino, 1986 (Nematoda: Mermithidae) predated upon by freshwater copepods (Crustacea: Copepoda)

Five copepod species, Acantocyclops robustus, Macrocyclops albidus, Mesocyclops annulatus, Thermocyclops sp. (all Cyclopoidea) and Argyrodiaptomus bergi (Calanoidea), were evaluated under laboratory conditions as potential predators of preparasites of the mermithid nematode Strelkovimermis spiculatus. Adults of all five copepod species consumed 70-100% of the preparasites within 24 h. Copepodids and adults of M. annulatus predated upon 93 and 100% of the nematode pre-parasites, respectively. An average of 197 pre-parasites was daily predated upon by an adult M. annulatus. Copepod density affected prevalence and intensity of S. spiculatus on mosquito larvae. Prevalence and intensity of S. spiculatus on Aedes aegypti larvae was reduced from 98% and 2.8 nematodes per larva in control containers free of copepods to 61.4% and 1.4 nematodes per larva in containers with 50 M. annulatus females per l.

First record of a mosquito iridescent virus in Culex pipiens L. (Diptera: Culicidae)

The mosquito iridescent viruses (MIVs) are large icosahedral DNA viruses that replicate and assemble in the cytoplasm of the host. Paracrystalline arrangements of virions that accumulate in the cytoplasm produce an iridescent color that is symptomatic of acute infections. In August 2010, we found larvae of Culex pipiens with these symptoms in suburban ditches around the city of La Plata, Argentina. Electron microscope studies, DNA sequencing, and phylogenetic analysis of the major capsid protein confirmed this as the first record of an MIV in C. Pipiens.

Effect of temperature, photoperiod, flooding, and drying on the hatching pattern of the eggs of Strelkovimermis spiculatus (Nematoda: Mermithidae).

We assessed the number of Strelkovimermis spiculatus preparasites obtained from a known initial number of nematode eggs and the effect of abiotic conditions (temperature, photoperiod, flooding-drying) on the number of emerged preparasites. Two egg groups were maintained: one continuously flooded, another with flooding-drying cycles (every 15, 30, 60 days). Each egg group was studied at 25°C and 14:10 (L:D) and 16°C and 12:12 (L:D). The flooded eggs contained a higher overall percentage of S. spiculatus preparasites compared to the wet-dry-cycle eggs. The conditions of continuous flooding at 16°C and 12:12 (L:D) produced the maximum percent of emerged J2s (30±15%). Preparasites were recorded by 7 (25°C) and 14 (16°C) days, suggesting this period as the minimum time for embryonic development. The preparasite-emergence time observed from the same flooded-egg batch (98 and 112 days at 25°C and 16°C, respectively) suggested a nonsynchronous hatching, possibly through nonuniform egg embryonation. The time of exposure to drought in the assays did not significantly affect the total average percentage of J2s obtained at 25°C and 14:10 (L:D), whereas at 16°C the number of emerged J2s diminished with a prolongation of the drying period. The oviposition period was also recorded only at 16°C and 12:12 (L:D): S. spiculatus eggs were detected at 12.6 days after postparasite emergence, and oviposition was complete at 51days under those conditions. We propose a flooding schedule to optimize the mass-rearing of S. spiculatus.