h Malaria transmission-blocking vaccines (TBVs) represent a promising approach for the elimination and eradication of this disease. AnAPN1 is a lead TBV candidate that targets a surface antigen on the midgut of the obligate vector of the Plasmodium parasite, the Anopheles mosquito. In this study, we demonstrated that antibodies targeting AnAPN1 block transmission of Plasmodium falciparum and Plasmodium vivax across distantly related anopheline species in countries to which malaria is endemic. Using a biochemical and immunological approach, we determined that the mechanism of action for this phenomenon stems from antibody recognition of a single protective epitope on AnAPN1, which we found to be immunogenic in murine and nonhuman primate models and highly conserved among anophelines. These data indicate that AnAPN1 meets the established target product profile for TBVs and suggest a potential key role for an AnAPN1-based panmalaria TBV in the effort to eradicate malaria.

Malaria sporozoites are deposited into the skin by mosquitoes and infect hepatocytes. The molecular basis of how Plasmodium falciparum sporozoites migrate through host cells is poorly understood, and direct evidence of its importance in vivo is lacking. Here, we generated traversal-deficient sporozoites by genetic disruption of sporozoite microneme protein essential for cell traversal (PfSPECT) or perforin-like protein 1 (PfPLP1). Loss of either gene did not affect P. falciparum growth in erythrocytes, in contrast with a previous report that PfPLP1 is essential for merozoite egress. However, although traversal-deficient sporozoites could invade hepatocytes in vitro, they could not establish normal liver infection in humanized mice. This is in contrast with NF54 sporozoites, which infected the humanized mice and developed into exoerythrocytic forms. This study demonstrates that SPECT and perforin-like protein 1 (PLP1) are critical for transcellular migration by P. falciparum sporozoites and demonstrates the importance of cell traversal for liver infection by this human pathogen.

Aedes albopictus is a potential West Nile virus bridge vector in Northern Virginia; however, information regarding its virus transmission dynamics is limited, as this species is not readily collected in existing traps. This study used 5 replicates of a 5 x 5 Latin square to evaluate the efficiency and effectiveness of 2 novel host-seeking mosquito traps (the BG-Sentinel and the Collapsible Mosquito Trap (CMT-20) in collecting Ae. albopictus, relative to a carbon dioxide (CO2)-baited Centers for Disease Control and Prevention (CDC) miniature light trap. When used with CO2, the BG-Sentinel (with BG-Lure) collected 33 times more female Ae. albopictus per 24-h trapping period than did the CO2-baited CDC light trap. Without CO2, the BG-Sentinel (with BG-Lure) still collected over 6 times as many female Ae. albopictus as the CO2-baited CDC trap. Both configurations of the BG-Sentinel were significantly more effective than the other traps. The BG-Sentinel was also significantly more efficient in collecting Ae. albopictus and collected a high proportion of this species, both with CO2 and without CO2. The CMT-20 (with SkinLure) collected significantly more Ae. albopictus when used with CO2 than without CO2, but did not collect significantly more Ae. albopictus than the CO2-baited CDC light trap. The proportion of Ae. albopictus collected in the CMT-20 with CO2 and without CO2 did not differ significantly from the proportion of Ae. albopictus collected in the CDC trap.

fVaccines have been at the forefront of global research efforts to combat malaria, yet despite several vaccine candidates, this goal has yet to be realized. A potentially effective approach to disrupting the spread of malaria is the use of transmissionblocking vaccines (TBV), which prevent the development of malarial parasites within their mosquito vector, thereby abrogating the cascade of secondary infections in humans. Since malaria is transmitted to human hosts by the bite of an obligate insect vector, mosquito species in the genus Anopheles, targeting mosquito midgut antigens that serve as ligands for Plasmodium parasites represents a promising approach to breaking the transmission cycle. The midgut-specific anopheline alanyl aminopeptidase N (AnAPN1) is highly conserved across Anopheles vectors and is a putative ligand for Plasmodium ookinete invasion. We have developed a scalable, high-yield Escherichia coli expression and purification platform for the recombinant AnAPN1 TBV antigen and report on its marked vaccine potency and immunogenicity, its capacity for eliciting transmission-blocking antibodies, and its apparent lack of immunization-associated histopathologies in a small-animal model.

Aedes albopictus (Skuse) and Aedes japonicus (Theobald) are two of the most recent and widespread invasive mosquito species to have become established in the United States. The two species co-occur in water-filled artificial containers, where crowding and limiting resources are likely to promote inter- or intraspecific larval competition. The performance of northern Virginia populations of Ae. japonicus and Ae. albopictus competing as larvae under field conditions was evaluated. Per capita rates of population increase for each species were estimated, and the effects of species composition and larval density were determined. In water-containing cups provided with oak leaves, Ae. albopictus larvae exhibited a competitive advantage over Ae. japonicus as a consequence of higher survivorship, shorter developmental time, and a significantly higher estimated population growth rate under conditions of interspecific competition. Intraspecific competition constrained population performance of Ae. albopictus significantly more than competition with Ae. japonicus. In the context of the Lotka-Volterra model of competition, these findings suggest competitive exclusion of Ae. japonicus in those habitats where this species co-occurs with Ae. albopictus.

Mosquito-based malaria transmission-blocking vaccines (mTBVs) target midgut-surface antigens of the Plasmodium parasite's obligate vector, the Anopheles mosquito. The alanyl aminopeptidase N (AnAPN1) is the leading mTBV immunogen; however AnAPN1's role in Plasmodium infection of the mosquito and how anti-AnAPN1 antibodies functionally block parasite transmission remains elusive. Here we present the 2.65 Å crystal structure of AnAPN1 and the immunoreactivity and transmission-blocking profile of three AnAPN1 monoclonal antibodies (mAb), including mAb 4H5B7, which effectively block transmission of natural strains of Plasmodium falciparum. Utilizing the AnAPN1 structure we map the conformation-dependent 4H5B7 neo-epitope to a previously uncharacterized region on domain 1, and further demonstrate that non-human primate neo-epitope-specific IgG also block parasite transmission. We discuss the Users may view, print, copy, and download text and data-mine the content in such documents, for the purposes of academic research, subject always to the full Conditions of use:http://www.nature.com/authors/editorial_policies/license.html#terms Correspondence should be addressed to: Natalie A. Borg, PhD., Department of Biochemistry and Molecular Biology, Monash University, Clayton, 3800, Victoria, Australia, natalie.borg@monash.edu, Tel: +613-9902-9369, Fax: +613-9902-9500; Rhoel R. HHS Public AccessAuthor manuscript Nat Struct Mol Biol. Author manuscript; available in PMC 2016 January 01. Published in final edited form as:Nat Struct Mol Biol. 2015 July ; 22(7): 532-539. doi:10.1038/nsmb.3048. Author Manuscript Author ManuscriptAuthor ManuscriptAuthor Manuscript prospect of a novel biological function of AnAPN1 as a receptor for Plasmodium in the mosquito midgut and the implications for redesigning the AnAPN1 mTBV.Malaria exacts a deadly toll on human populations worldwide. A new era in the fight against the disease has seen multiple malaria vaccines entering or completing advanced clinical study 1 , including malaria transmission-blocking vaccines (mTBVs) 2 . Malaria transmission requires the establishment of Plasmodium in the Anopheles mosquito, which is contingent on the parasite's ookinete stage successfully traversing the midgut epithelium to initiate sporogonic development 3,4 . TBVs disrupt this obligatory step in the parasite life cycle 5 , limiting the number of infectious mosquito vectors and introducing local herd immunity 6 . The concept of mTBVs is simple: antibodies against specific, mosquito midgut antigens circulating in the peripheral blood are ingested by the mosquito while feeding on immunized hosts. These antibodies, as well as complement, can survive in the mosquito midgut for up to 24 hours post-blood feeding and prevent parasite access to host ligands that mediate midgut cell adhesion and invasion. Unable to establish infection in the vector, progression of parasite development and transmission to new human hosts is arrested or reduced.We have shown that AnAPN1, an alanyl aminopeptidase N present on...

The success of an invasive species in a new region depends on its interactions with ecologically similar resident species. Invasions by disease vector mosquitoes are important as they may have ecological and epidemiological consequences. Potential interactions of a recent invasive mosquito, Aedes japonicus Theobald, with resident species in Virginia were evaluated by sampling larvae from containers and trapping adults. Distinct species compositions were observed for artificial containers and rock pools, with Ae. albopictus most abundant in the former and Ae. japonicus in the latter. However, these two species were found to co-occur in 21.2% of containers sampled. Among the six mosquito species most common in containers from May through September, 2006, only interspecific associations of Ae.japonicus with Aedesalbopictus(Skuse) and Aedestriseriatus(Say) were significant, and both were negative. In addition to differences in habitat preference, mean crowding estimates suggest that interspecific repulsion may contribute to the significant negative associations observed between these species. High relative abundances of late instars and pupae of Ae. japonicus seem to provide this species with a mechanism of evading competition with Ae. albopictus, facilitating their coexistence in artificial containers. Although annual fluctuations were observed, trends in adult populations over a 6-yr period provide no evidence of declines. In summary, this survey of diverse container types and all life stages provided only limited evidence for competitive displacements or reductions of resident container species by Ae. japonicus, as observed elsewhere in its invasive range.

The success of an invasive species becoming established in a new region often depends on its interactions with ecologically similar resident species. The propensity of the newly-established mosquito Aedes japonicus to inhabit rock pools throughout the eastern United States provides a natural setting for interspecific larval competition with the native Aedes atropalpus. A laboratory experiment conducted in simulated rock pools to evaluate larval interactions between and within these two species suggested that the performance of both species was more significantly impacted by intraspecific conditions than interspecific conditions of the same mosquito density. Aedes atropalpus was apparently more sensitive to larval densities than Ae. japonicus because it reproduces autogenously, requiring a lengthened period of larval development to obtain nutrient reserves for egg development, which may ultimately put Ae. atropalpus at a disadvantage under larval conditions of competition and limited resources. Excessively stressful experimental conditions, as evidenced by reduced body size, and thus fecundity and estimated finite rate of increase, may have obscured the effects of larval competition between these species. The impact of larval competition between these species in rock pool communities warrants further investigation under more ecologically realistic experimental conditions.