Entomopathogenic fungi and their potential for the management of Aedes aegypti (Diptera: Culicidae) in the Americas.

Classical biological control has been used extensively for the management of exotic weeds and agricultural pests, but never for alien insect vectors of medical importance. This simple but elegant control strategy involves the introduction of coevolved natural enemies from the centre of origin of the target alien species. Aedes aegypti - the primary vector of the dengue, yellow fever and Zika flaviviruses - is just such an invasive alien in the Americas where it arrived accidentally from its West African home during the slave trade. Here, we introduce the concept of exploiting entomopathogenic fungi from Africa for the classical biological control of Ae. aegypti in the Americas. Fungal pathogens attacking arthropods are ubiquitous in tropical forests and are important components in the natural balance of arthropod populations. They can produce a range of specialised spore forms, as well as inducing a variety of bizarre behaviours in their hosts, in order to maximise infection. The fungal groups recorded as specialised pathogens of mosquito hosts worldwide are described and discussed. We opine that similar fungal pathogens will be found attacking and manipulating Ae. aegypti in African forests and that these could be employed for an economic, environmentally-safe and long-term solution to the flavivirus pandemics in the Americas.

Temperature and parasite life-history are important modulators of the outcome of Trypanosoma rangeli-Rhodnius prolixus interactions.

Trypanosoma rangeli is a protozoan parasite, which does not cause disease in humans, although it can produce different levels of pathogenicity to triatomines, their invertebrate hosts. We tested whether infection imposed a temperature-dependent cost on triatomine fitness using T. rangeli with different life histories. Parasites cultured only in liver infusion tryptose medium (cultured) and parasites exposed to cyclical passages through mice and triatomines (passaged) were used. We held infected insects at four temperatures between 21 and 30 °C and measured T. rangeli growth in vitro at the same temperatures in parallel. Overall, T. rangeli infection induced negative effects on insect fitness. In the case of cultured infection, parasite effects were temperature-dependent. Intermoult period, mortality rates and ecdysis success were affected in those insects exposed to lower temperatures (21 and 24 °C). For passaged-infected insects, the effects were independent of temperature, intermoult period being prolonged in all infected groups. Trypanosoma rangeli seem to be less tolerant to higher temperatures since cultured-infected insects showed a reduction in the infection rates and passaged-infected insects decreased the salivary gland infection rates in those insects submitted to 30 °C. In vitro growth of T. rangeli was consistent with these results.

Attracted to the enemy: Aedes aegypti prefers oviposition sites with predator-killed conspecifics.

Oviposition habitat choices of species with aquatic larvae are expected to be influenced by both offspring risk of mortality due to predation, and offspring growth potential. Aquatic predators may indirectly influence growth potential for prey by reducing prey density and, for filter-feeding prey, by increasing bacterial food for prey via added organic matter (feces, partially eaten victims), creating the potential for interactive effects on oviposition choices. We tested the hypothesis that the mosquito Aedes aegypti preferentially oviposits in habitats with predatory Toxorhynchites larvae because of indirect effects of predation on chemical cues indicating bacterial abundance. We predicted that A. aegypti would avoid oviposition in sites with Toxorhynchites, but prefer to oviposit where bacterial food for larvae is abundant, and that predation by Toxorhynchites would increase bacterial abundances. Gravid A. aegypti were offered paired oviposition sites representing choices among: predator presence; the act of predation; conspecific density; dead conspecific larvae; and bacterial activity. A. aegypti preferentially oviposited in sites with Toxorhynchites theobaldi predation, and with killed conspecific larvae, but failed to detect preferences for other treatments. The antibiotic tetracycline eliminated the strongest oviposition preference. Both predation by Toxorhynchites and killed larvae increased bacterial abundances, suggesting that oviposition attraction is cued by bacteria. Our results show the potential for indirect effects, like trophic cascades, to influence oviposition choices and community composition in aquatic systems. Our results suggest that predators like Toxorhynchites may be doubly beneficial as biocontrol agents because of the attraction of ovipositing mosquitoes to bacterial by-products of Toxorhynchites feeding.

Juvenile responses to immune challenges are not carried through to subsequent life stages in an insect.

Environmental variability can significantly impact individual survival and reproduction. Meanwhile, high population densities can lead to resource scarcity and increased exposure to parasites and pathogens. Studies with insects can offer valuable insights into eco-immunology, allowing us to explore the connections between these variables. Here we use the moth Anticarsia gemmatalis to examine how increases in population density and immunological challenge during the larval stage shape its investment in immune defence and reproduction. Larvae reared at a high population density exhibited greater lytic activity against bacteria compared to those reared at low density, whilst bacterial challenge (i.e. bacteria-immersed needles) also increased lytic activity. There was no interaction between the variables population density and bacterial challenge, indicating that these are independent. Surprisingly, neither increase in lytic activity carried through to activity in prepupal haemolymph. Rearing of larvae at a high density delayed pupation and decreased pupal weight. The immunological stimulus did not significantly influence pupal development. Lower population density as a larva resulted in greater adult weight, but did not significantly influence lytic activity in the eggs or the number of eggs laid. Negative correlations were found between lytic activity in the eggs and the number of eggs, as well as between adult weight and the number of eggs. Overall, this study demonstrates that high population density and immune challenge trigger increased lytic activity in caterpillars, but this effect is transient, not persisting into later stages. The trade-offs observed, such as delayed pupation and reduced prepupal weights under high density, suggest a balancing act between immune investment and developmental aspects. The findings hint at a short-term adaptive response rather than a sustained strategy. The implications of delayed pupation and smaller adult moths could influence the moth's life history strategy, impacting its role in the ecosystem. Further research tracking larval immune investment and subsequent reproductive success will unveil the evolutionary dynamics of this relationship in changing environments.

Cordyceps cateniannulata: An endophyte of coffee, a parasite of coffee leaf rust and a pathogen of coffee pests.

Here, we report on a Cordyceps species entering into a multi-trophic, multi-kingdom association. Cordyceps cateniannulata, isolated from the stem of wild Coffea arabica in Ethiopia, is shown to function as an endophyte, a mycoparasite and an entomopathogen. A detailed polyphasic taxonomic study, including a multilocus phylogenetic analysis, confirmed its identity. An emended description of C. cateniannulata is provided herein. Previously, this species was known as a pathogen of various insect hosts in both the Old and New World. The endophytic status of C. cateniannulata was confirmed by re-isolating it from inoculated coffee plants. Inoculation studies have further shown that C. cateniannulata is a mycoparasite of Hemileia vastatrix, as well as an entomopathogen of major coffee pests; infecting and killing Hypothenemus hampei and Leucoptera coffeella. This is the first record of C. cateniannulata from Africa, as well as an endophyte and a mycoparasite. The implications for its use as a biocontrol agent are discussed.

Foraging ants trade off further for faster: use of natural bridges and trunk trail permanency in carpenter ants.

Trail-making ants lay pheromones on the substrate to define paths between foraging areas and the nest. Combined with the chemistry of these pheromone trails and the physics of evaporation, trail-laying and trail-following behaviours provide ant colonies with the quickest routes to food. In relatively uniform environments, such as that provided in many laboratory studies of trail-making ants, the quickest route is also often the shortest route. Here, we show that carpenter ants (Camponotus rufipes), in natural conditions, are able to make use of apparent obstacles in their environment to assist in finding the fastest routes to food. These ants make extensive use of fallen branches, twigs and lianas as bridges to build their trails. These bridges make trails significantly longer than their straight line equivalents across the forest floor, but we estimate that ants spend less than half the time to reach the same point, due to increased carriage speed across the bridges. We also found that these trails, mainly composed of bridges, are maintained for months, so they can be characterized as trunk trails. We suggest that pheromone-based foraging trail networks in field conditions are likely to be structured by a range of potentially complex factors but that even then, speed remains the most important consideration.

What prevents phenological adjustment to climate change in migrant bird species? Evidence against the "arrival constraint" hypothesis.

Phenological studies have demonstrated changes in the timing of seasonal events across multiple taxonomic groups as the climate warms. Some northern European migrant bird populations, however, show little or no significant change in breeding phenology, resulting in synchrony with key food sources becoming mismatched. This phenological inertia has often been ascribed to migration constraints (i.e. arrival date at breeding grounds preventing earlier laying). This has been based primarily on research in The Netherlands and Germany where time between arrival and breeding is short (often as few as 9 days). Here, we test the arrival constraint hypothesis over a 15-year period for a U.K. pied flycatcher (Ficedula hypoleuca) population where laying date is not constrained by arrival as the period between arrival and breeding is substantial and consistent (average 27 ± 4.57 days SD). Despite increasing spring temperatures and quantifiably stronger selection for early laying on the basis of number of offspring to fledge, we found no significant change in breeding phenology, in contrast with co-occurring resident blue tits (Cyanistes caeruleus). We discuss possible non-migratory constraints on phenological adjustment, including limitations on plasticity, genetic constraints and competition, as well as the possibility of counter-selection pressures relating to adult survival, longevity or future reproductive success. We propose that such factors need to be considered in conjunction with the arrival constraint hypothesis.

Realism in Immune Ecology Studies: Artificial Diet Enhances a Caterpillar's Immune Defense but Does Not Mask the Effects of a Plastic Immune Strategy.

The immune system is considered a functional trait in life-history theory and its modulation is predicted to be costly and highly dependent on the host's nutrition. Therefore, the nutritional status of an individual has a great impact on an animal's immune ecology. Herbivorous insects are commonly used as model organisms in eco-immunology studies and the use of an artificial diet is the predominant rearing procedure to test them. However, this diet differs from what herbivores experience in nature and it is unclear to what degree this distinction might impact on the relevance of these studies for the real world. Here, we compared plant-based vs. artificial diet in a set of three experiments to investigate the interaction of both diets with a plastic immune strategy known as Density-Dependent Prophylaxis (DDP). We used as a model organism the velvetbean caterpillar Anticarsia gemmatalis, which is known to adjust its immune defense in line with the DDP hypothesis. Our main results showed that larvae fed with artificial diet had 20.5% more hemocytes circulating in the hemolymph and died 20% more slowly when infected with an obligate (viral) pathogen. Crucially, however, we did not find any indication of fitness costs related to DDP. The use of artificial diet did not interact with that of DDP except in the case of host survival after infection, where the DDP effect was only observable in this diet. Our findings suggest the use of an artificial diet does not mask resource allocation conflicts between immune investment and fitness related traits, but to some extent it might lead to an overestimation of immune parameters and host survival time after infection. We believe that this is the first study to compare an artificial diet and a host plant covering all these aspects: immune parameters, life-history traits, and host survival after infection. Here we provide evidence that, besides the quantitative effects in immune parameters and host survival time, the use of artificial diet interacts only marginally with a density-dependent immune response. This provides support for the use of artificial diets in eco-immunology studies with insects.

Experimental evolution of cowpea mild mottle virus reveals recombination-driven reduction in virulence accompanied by increases in diversity and viral fitness.

Major themes in pathogen evolution are emergence, evolution of virulence, host adaptation and the processes that underlie them. RNA viruses are of particular interest due to their rapid evolution. The in vivo molecular evolution of an RNA plant virus was demonstrated here using a necrotic isolate of cowpea mild mottle virus (CPMMV) and a susceptible soybean genotype submitted to serial inoculations. We show that the virus lost the capacity to cause necrosis after six passages through the host plant. When a severe bottleneck was imposed, virulence reduction occurred in the second passage. The change to milder symptoms had fitness benefits for the virus (higher RNA accumulation) and for its vector, the whitefly Bemisia tabaci. Genetic polymorphisms were highest in ORF1 (viral replicase) and were independent of the symptom pattern. Recombination was a major contributor to this diversity - even with the strong genetic bottleneck, recombination events and hot spots were detected within ORF1. Virulence reduction was associated with different sites in ORF1 associated to recombination events in both experiments. Overall, the results demonstrate that the reduction in virulence was a consequence of the emergence of new variants, driven by recombination. Besides providing details of the evolutionary mechanisms behind a reduction in virulence and its effect under viral and vector fitness, we propose that this recombination-driven switch in virulence allows the pathogen to rapidly adapt to a new host and, potentially, switch back.

Density-dependent prophylactic immunity reconsidered in the light of host group living and social behavior.

According to the density-dependent hypothesis (DDP), hosts living at high densities suffer greater risk of disease and so invest more in immunity. Although there is much empirical support for this, especially from invertebrate systems, there are many exceptions, notably in social insects. We propose that (A) density is not always the most appropriate population parameter to use when considering the risks associated with disease and (B) behavioral defenses should be given a greater emphasis in considerations of a host's repertoire of immune defenses. We propose a complementary framework stressing the connectivity between and within populations as a starting point and emphasizing the costs represented by disease above the risk of disease per se. We consider the components of immune defense and propose that behaviors may represent lower-cost defenses than their physiological counterparts. As group-living and particularly social animals will have a greater behavioral repertoire, we conclude that with group living comes a greater capacity for behavioral immune defense, most particularly for social insects. This may escape our notice if we consider physiological parameters alone.

A standardizable protocol for infection of Rhodnius prolixus with Trypanosoma rangeli, which mimics natural infections and reveals physiological effects of infection upon the insect.

Trypanosoma rangeli is a protozoan parasite that shares hosts - mammals and triatomines - with Trypanosoma cruzi, the etiological agent of Chagas disease. Although T. rangeli is customarily considered to be non-pathogenic to human hosts, it is able to produce pathologies in its invertebrate hosts. However, advances are hindered by a lack of standardization of infection procedures and these pathologies need documentation. To establish a suitable, and standardizable, infection protocol, the duration of the fourth instar was evaluated in nymphs infected by injection into the thorax with different concentrations of parasites, and compared with nymphs infected naturally (i.e. orally). We demonstrate that delays in moult were attributable to the presence of the parasite in the haemolymph (vs. the gut) and propose that the protocol presented here simulates closely natural infections. This methodology was then used for the evaluation of physiological parameters and several hitherto unreported effects of T. rangeli infection on Rhodnius prolixus were revealed. Haemolymph volume was greater in infected than uninfected nymphs but this alteration could not be attributed to water retention, since infected insects lost the same amount of water as controls. However, we found that lipid content and fat body weight were both increased in insects infected by T. rangeli. We propose that this is due to the parasite's sequestration of host blood lipids and carrier proteins. With these findings, we have taken a few first steps to unravelling physiological details of the host-parasite interaction. We suggest future directions towards a fuller understanding of mechanistic and adaptive aspects of triatomine-trypanosomatid interactions.

Quantifying the reproduction of Bacillus thuringiensis HD1 in cadavers and live larvae of Plutella xylostella.

The Bacillus cereus group comprises a range of micro-organisms with diverse habits, including gut commensals, opportunistic pathogens and soil saprophytes. Using quantitative microbiological methods we tested whether Bacillus thuringiensis (Bt) could reproduce in cadavers of Plutella xylostella killed by Bt, or in the gut of live insects, or be transmitted vertically from females to their offspring. We also tested whether diverse Bt strains could grow in high nutrient broth at a pH similar to that in the larval midgut. Low levels of reproduction were found in insect cadavers but there was no evidence of vertical transmission, or of significant reproduction in live insects. Four strains of B. thuringiensis var. kurstaki and one of B. thuringiensis var. tenebrionis were found to be capable of growth at high pH. Greater spore recovery rates in frass were found in hosts that were resistant or tolerant of infection. We concluded that that spores recovered in frass represent, in general, an ungerminated fraction of ingested inoculum and that germination rates are reduced in unsuitable hosts.

Failure of the mite-pathogenic fungus Neozygites tanajoae and the predatory mite Neoseiulus idaeus to control a population of the cassava green mite, Mononychellus tanajoa.

Monitoring of a population of the phytophagous cassava green mite, Mononychellus tanajoa (Bondar), and its natural enemies was undertaken in central Bahia, Brazil, in mid-1996. In spite of the presence of extremely high densities of the predatory phytoseiid mite Neoseiulus idaeus Denmark & Muma, the phytophagous mite population reached such high densities itself that there was total overexploitation of the cassava plants, leading to total leaf loss. Meanwhile, the mite-pathogenic fungus Neozygites tanajoae Delalibera, Humber & Hajek did not affect the M. tanajoa population in its growth phase as there was no inoculum present, even though we predict from a simple regression model that there was the potential for epizootics at that time. Soon after the M. tanajoa population crashed due to defoliation, there could have been an epizootic but there were simply no mite hosts to infect. These data demonstrate the ineffectiveness of one natural enemy (the predator) in terms of prey population regulation and demonstrate the importance of timing in the possible effectiveness of the other (the pathogen). For the pathogen, this probably explains its sporadic effect on host populations as previously reported. We conclude that the fungus is likely to be most useful as an adjunct to biological control with predatory mites other than N. idaeus.

Witch's Broom Disease of Lime (Candidatus Phytoplasma aurantifolia): Identifying High-Risk Areas by Climatic Mapping.

Biological invasions of vectorborne diseases can be devastating. Bioclimatic modeling provides an opportunity to assess and predict areas at risk from complex multitrophic interactions of pathogens, highlighting areas in need of increased monitoring effort. Here, we model the distribution of an economically critical vectorborne plant pathogen 'Candidatus Phytoplasma aurantifolia', the etiological agent of Witches' Broom Disease of Lime. This disease is a significant limiting factor on acid lime production (Citrus aurantifolia, Swingle) in the Middle East and threatens its production globally. We found that temperature, humidity, and the vector populations significantly determine disease distribution. Following this, we used bioclimatic modeling to predict potential novel sites of infections. The model outputs identified potential novel sites of infection in the citrus producing regions of Brazil and China. We also used our model to explore sites in Oman where the pathogen may not be infectious, and suggest nurseries be established there. Recent major turbulence in the citrus agricultural economy has highlighted the importance of this work and the need for appropriate and targeted monitoring programs to safeguard lime production.

Putting the waste out: a proposed mechanism for transmission of the mycoparasite Escovopsis between leafcutter ant colonies.

The attine ant system is a remarkable example of symbiosis. An antagonistic partner within this system is the fungal parasite Escovopsis, a genus specific to the fungal gardens of the Attini. Escovopsis parasitizes the Leucoagaricus symbiont that leaf-cutting ants (Acromyrmex, Atta) have been farming over the past 8-12 Myr. However, it has been a puzzle how Escovopsis reaches its host. During a seasonal survey of nests of Acromyrmex subterraneus subterraneus in Atlantic rainforest in Brazil, Escovopsis was detected in all the sampled fungal garden waste tips or middens (n = 111). Middens were built strategically; always below the nest entrances. Here, we report the first evidence of a putative mechanism for horizontal transmission of Escovopsis between attine colonies. It is posited that leaf-cutting ants pick up the spores from soil and litter during foraging and vector the mycoparasite between attine colonies. Field and laboratory experiments, using At. laevigata and Ac. subterraneus subterraneus, confirm that Escovopsis spores are phoretic, and have an inbuilt dormancy, broken by the presence of their Leucoagaricus host. However, in the coevolutionary arms race, Atta ants may lose out-despite most species in the genus investing in a more advanced waste disposal system-due to the insanitary habits of their Acromyrmex neighbours.

The Spread of Helicoverpa armigera (Lepidoptera: Noctuidae) and Coexistence with Helicoverpa zea in Southeastern Brazil.

Helicoverpa armigera, one of the world's most destructive crop pests, was first documented in Brazil in 2013. Within a few months, this polyphagous insect had spread over the Northeast and Central-West of Brazil, causing great agricultural losses. With several reports of populations resistant to pesticides and Bt crops around the world, there is great concern about the spread of this pest in Brazil. There is confusion about the actual distribution of this species due to the high morphological similarity with the native corn earworm Helicoverpa zea, which may also coexist with H. armigera in the field. Our aims here were (i) to confirm its presence in the State of Minas Gerais, one of the most important agricultural regions in the country; and (ii) to assess the co-occurrence of this pest with the congeneric corn earworm H. zea. Using molecular screening, we confirmed the presence of H. armigera in Bt-crops of soybean and cotton, and non-Bt-crops of soybean, cotton and maize. Mixed infestations of H. armigera with H. zea were found in non-Bt maize (Viçosa, Southeastern Minas Gerais). These results highlight the need for adequate control strategies for H. armigera in Brazil, to deal with its polyphagous feeding habits, high dispersal capacity and possible risks of hybridization with congeneric species.

Recognition of endophytic Trichoderma species by leaf-cutting ants and their potential in a Trojan-horse management strategy.

Interactions between leaf-cutting ants, their fungal symbiont (Leucoagaricus) and the endophytic fungi within the vegetation they carry into their colonies are still poorly understood. If endophytes antagonistic to Leucoagaricus were found in plant material being carried by these ants, then this might indicate a potential mechanism for plants to defend themselves from leaf-cutter attack. In addition, it could offer possibilities for the management of these important Neotropical pests. Here, we show that, for Atta sexdens rubropilosa, there was a significantly greater incidence of Trichoderma species in the vegetation removed from the nests-and deposited around the entrances-than in that being transported into the nests. In a no-choice test, Trichoderma-infested rice was taken into the nest, with deleterious effects on both the fungal gardens and ant survival. The endophytic ability of selected strains of Trichoderma was also confirmed, following their inoculation and subsequent reisolation from seedlings of eucalyptus. These results indicate that endophytic fungi which pose a threat to ant fungal gardens through their antagonistic traits, such as Trichoderma, have the potential to act as bodyguards of their plant hosts and thus might be employed in a Trojan-horse strategy to mitigate the negative impact of leaf-cutting ants in both agriculture and silviculture in the Neotropics. We posit that the ants would detect and evict such 'malign' endophytes-artificially inoculated into vulnerable crops-during the quality-control process within the nest, and, moreover, that the foraging ants may then be deterred from further harvesting of 'Trichoderma-enriched' plants.

Temperature and population density: interactional effects of environmental factors on phenotypic plasticity, immune defenses, and disease resistance in an insect pest.

Temperature and crowding are key environmental factors mediating the transmission and epizooty of infectious disease in ectotherm animals. The host physiology may be altered in a temperature-dependent manner and thus affects the pathogen development and course of diseases within an individual and host population, or the transmission rates (or infectivity) of pathogens shift linearly with the host population density. To our understanding, the knowledge of interactive and synergistic effects of temperature and population density on the host-pathogen system is limited. Here, we tested the interactional effects of these environmental factors on phenotypic plasticity, immune defenses, and disease resistance in the velvetbean caterpillar Anticarsia gemmatalis. Upon egg hatching, caterpillars were reared in thermostat-controlled chambers in a 2 × 4 factorial design: density (1 or 8 caterpillars/pot) and temperature (20, 24, 28, or 32°C). Of the immune defenses assessed, encapsulation response was directly affected by none of the environmental factors; capsule melanization increased with temperature in both lone- and group-reared caterpillars, although the lone-reared ones presented the most evident response, and hemocyte numbers decreased with temperature regardless of the population density. Temperature, but not population density, affected considerably the time from inoculation to death of velvetbean caterpillar. Thus, velvetbean caterpillars succumbed to Anticarsia gemmatalis multiple nucleopolyhedrovirus (AgMNPV) more quickly at higher temperatures than at lower temperatures. As hypothesized, temperature likely affected caterpillars' movement rates, and thus the contact between conspecifics, which in turn affected the phenotypic expression of group-reared caterpillars. Our results suggest that environmental factors, mainly temperature, strongly affect both the course of disease in velvetbean caterpillar population and its defenses against pathogens. As a soybean pest, velvetbean caterpillar may increase its damage on soybean fields under a scenario of global warming as caterpillars may reach the developmental resistance faster, and thus decrease their susceptibility to biological control by AgMNPV.

Screening of Fungi for Biological Control of a Triatomine Vector of Chagas Disease: Temperature and Trypanosome Infection as Factors.

Entomopathogenic fungi have been investigated as an alternative tool for controlling various insects, including triatomine vectors of the protozoan Trypanosoma cruzi, the etiological agent of Chagas disease. Here we tested the pathogenicity and virulence of ten isolates of the fungi Metarhizium spp. and Beauveria bassiana against Rhodnius prolixus and found all of the isolates to be virulent. We used two isolates (URPE-11 Metarhizium anisopliae and ENT-1 Beauveria bassiana) for further screening based on their prolific sporulation in vitro (an important property of fungal biopesticides). We characterized their virulences in a dose-response experiment and then examined virulence across a range of temperatures (21, 23, 27 and 30°C). We found isolate ENT-1 to maintain higher levels of virulence over these temperatures than URPE-11. We therefore used B. bassiana ENT-1 in the final experiment in which we examined the survival of insects parasitized with T. cruzi and then infected with this fungus (once again over a range of temperatures). Contrary to our expectations, the survival of insects challenged with the pathogenic fungus was greater when they had previously been infected with the parasite T. cruzi than when they had not (independent of temperature). We discuss these results in terms of aspects of the biologies of the three organisms. In practical terms, we concluded that, while we have fungal isolates of potential interest for development as biopesticides against R. prolixus, we have identified what could be a critical problem for this biological tool: the parasite T. cruzi appears to confer a measure of resistance to the insect against the potential biopesticide agent so use of this fungus as a biopesticide could lead to selection for vector competence.

Invasive mutualisms between a plant pathogen and insect vectors in the Middle East and Brazil.

Complex multi-trophic interactions in vectorborne diseases limit our understanding and ability to predict outbreaks. Arthropod-vectored pathogens are especially problematic, with the potential for novel interspecific interactions during invasions. Variations and novelties in plant-arthropod-pathogen triumvirates present significant threats to global food security. We examined aspects of a phytoplasma pathogen of citrus across two continents. 'Candidatus Phytoplasma aurantifolia' causes Witches' Broom Disease of Lime (WBDL) and has devastated citrus production in the Middle East. A variant of this phytoplasma currently displays asymptomatic or 'silent' infections in Brazil. We first studied vector capacity and fitness impacts of the pathogen on its vectors. The potential for co-occurring weed species to act as pathogen reservoirs was analysed and key transmission periods in the year were also studied. We demonstrate that two invasive hemipteran insects-Diaphorina citri and Hishimonus phycitis-can vector the phytoplasma. Feeding on phytoplasma-infected hosts greatly increased reproduction of its invasive vector D. citri both in Oman and Brazil; suggesting that increased fitness of invasive insect vectors thereby further increases the pathogen's capacity to spread. Based on our findings, this is a robust system for studying the effects of invasions on vectorborne diseases and highlights concerns about its spread to warmer, drier regions of Brazil.