RNA interference as a next-generation control method for suppressing Varroa destructor reproduction in honey bee (Apis mellifera) hives.

BACKGROUND: The Varroa mite (Varroa destructor) is considered to be the greatest threat to apiculture worldwide. RNA interference (RNAi) using double-stranded RNA (dsRNA) as a gene silencing mechanism has emerged as a next-generation strategy for mite control. RESULTS: We explored the impact of a dsRNA biopesticide, named vadescana, designed to silence the calmodulin gene in Varroa, on mite fitness in mini-hives housed in a laboratory. Two dosages were tested: 2 g/L dsRNA and 8 g/L dsRNA. Vadescana appeared to have no effect on mite survival, however, mite fertility was substantially reduced. The majority of foundress mites exposed to vadescana failed to produce any offspring. No dose-dependent effect of vadescana was observed, as both the low and high doses inhibited mite reproduction equally well in the mini-hives and neither dose impacted pupal survival of the honey bee. Approximately 95% of bee pupae were alive at uncapping across all treatment groups. CONCLUSION: These findings suggest that vadescana has significant potential as an effective alternative to conventional methods for Varroa control, with broader implications for the utilization of RNAi as a next-generation tool in the management of pest species. © 2024 The Author(s). Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Effects of Deformed Wing Virus-Targeting dsRNA on Viral Loads in Bees Parasitised and Non-Parasitised by Varroa destructor.

The Varroa destructor mite is a devastating parasite of honey bees; however the negative effects of varroa parasitism are exacerbated by its role as an efficient vector of the honey bee pathogen, Deformed wing virus (DWV). While no direct treatment for DWV infection is available for beekeepers to use on their hives, RNA interference (RNAi) has been widely explored as a possible biopesticide approach for a range of pests and pathogens. This study tested the effectiveness of three DWV-specific dsRNA sequences to lower DWV loads and symptoms in honey bees reared from larvae in laboratory mini-hives containing bees and varroa. The effects of DWV-dsRNA treatment on bees parasitised and non-parasitised by varroa mites during development were investigated. Additionally, the impact of DWV-dsRNA on viral loads and gene expression in brood-parasitising mites was assessed using RNA-sequencing. Bees parasitised during development had significantly higher DWV levels compared to non-parasitised bees. However, DWV-dsRNA did not significantly reduce DWV loads or symptoms in mini-hive reared bees, possibly due to sequence divergence between the DWV variants present in bees and varroa and the specific DWV-dsRNA sequences used. Varroa mites from DWV-dsRNA treated mini-hives did not show evidence of an elevated RNAi response or significant difference in DWV levels. Overall, our findings show that RNAi is not always successful, and multiple factors including pathogen diversity and transmission route may impact its efficiency.

Gene drives for invasive wasp control: Extinction is unlikely, with suppression dependent on dispersal and growth rates.

Gene drives offer a potentially revolutionary method for pest control over large spatial extents. These genetic modifications spread deleterious variants through a population and have been proposed as methods for pest suppression or even eradication. We examined the influence of local dispersal, long-distance and/or human-mediated dispersal, and variation in population growth on the success of a gene drive for the control of invasive social wasps (Vespula vulgaris). Our simulations incorporated a spatially realistic environment containing variable habitat quality in New Zealand. Pest eradication was not observed, except in extreme and unrealistic scenarios of constant, widespread, and spatially intense releases of genetically modified individuals every year for decades. Instead, the regional persistence of genetically modified and wild-type wasps was predicted. Simulations using spatially homogeneous versus realistic landscapes (incorporating uninhabitable areas and dispersal barriers) showed little difference in overall population dynamics. Overall, little impact on wasp abundance was observed in the first 15 years after introduction. After 25 years, populations were suppressed to levels <95% of starting populations. Populations exhibited "chase dynamics" with population cycles in space, with local extinction occurring in some areas while wasps became abundant in others. Increasing the wasps' local dispersal distance increased the spatial and temporal variability of the occupied area and population suppression. Varying levels of human-associated long-distance dispersal had little effect on population dynamics. Increasing intrinsic population growth rates interacted with local dispersal to cause higher mean populations and substantially higher levels of variation in population suppression and the total amount of landscape occupied. Gene drives appear unlikely to cause a rapid and widespread extinction of this and probably other pests but could offer long-term and cost-effective methods of pest suppression. The predicted level of <95% pest suppression would substantially reduce the predation pressure and competitive interactions of this invasive wasp on native species. However, the predicted long-term persistence of genetically modified pests will influence the ethics and likelihood of using gene drives for pest control, especially given concerns that modified wasps would eventually be transported back to their home range.

An invasive ant increases deformed wing virus loads in honey bees.

The majority of invasive species are best known for their effects as predators. However, many introduced predators may also be substantial reservoirs for pathogens. Honey bee-associated viruses are found in various arthropod species including invasive ants. We examined how the globally invasive Argentine ant (Linepithema humile), which can reach high densities and infest beehives, is associated with pathogen dynamics in honey bees. Viral loads of deformed wing virus (DWV), which has been linked to millions of beehive deaths around the globe, and black queen cell virus significantly increased in bees when invasive ants were present. Microsporidian and trypanosomatid infections, which are more bee-specific, were not affected by ant invasion. The bee virome in autumn revealed that DWV was the predominant virus with the highest infection levels and that no ant-associated viruses were infecting bees. Viral spillback from ants could increase infections in bees. In addition, ant attacks could pose a significant stressor to bee colonies that may affect virus susceptibility. These viral dynamics are a hidden effect of ant pests, which could have a significant impact on disease emergence in this economically important pollinator. Our study highlights a perhaps overlooked effect of species invasions: changes in pathogen dynamics.

Gene silencing for invasive paper wasp management: Synthesized dsRNA can modify gene expression but did not affect mortality.

Invasive paper wasps such as Polistes dominula are a major pest and problem for biodiversity around the globe. Safe and highly targeted methods for the control of these and other social wasp populations are needed. We attempted to identify potentially-lethal gene targets that could be used on adult paper wasps in a gene silencing or RNA interference (RNAi) approach. Double-stranded RNA (dsRNA) was designed to target genes for which silencing has proven lethal in other insects. dsRNA was provided either orally to foragers or directly injected into the wasps. We also provided the dsRNA unprotected or protected from degradation by gut nucleases in two different forms (lipofectamine and carbon quantum dots). The effects of oral delivery of 22 different gene targets to forager wasps was evaluated. The expression of five different genes was successfully reduced following dsRNA ingestion or injection. These gene targets included the FACT complex subunit spt16 (DRE4) and RNA-binding protein fusilli (FUSILLI), both of which have been previously shown to have potential as lethal targets for pest control in other insects. However, we found no evidence of significant increases in adult wasp mortality following ingestion or injection of dsRNA for these genes when compared with control treatments in our experiments. The methods we used to protect the dsRNA from digestive degradation altered gene expression but similarly did not influence wasp mortality. Our results indicate that while many of the same gene targets can be silenced and induce mortality in other insects, dsRNA and RNAi approaches may not be useful for paper wasp control.

Can immune gene silencing via dsRNA feeding promote pathogenic viruses to control the globally invasive Argentine ant?

Pest control methods that can target pest species with limited environmental impacts are a conservation and economic priority. Species-specific pest control using RNA interference is a challenging but promising avenue in developing the next generation of pest management. We investigate the feasibility of manipulating a biological invader's immune system using double-stranded RNA (dsRNA) in order to increase susceptibility to naturally occurring pathogens. We used the invasive Argentine ant as a model, targeting the immunity-associated genes Spaetzle and Dicer-1 with dsRNA. We show that feeding with Spaetzle dsRNA can result in partial target gene silencing for up to 28 days in the laboratory and 5 days in the field. Dicer-1 dsRNA only resulted in partial gene knockdown after 2 days in the laboratory. Double-stranded RNA treatments were associated with significant gene expression disruptions across immune pathways in the laboratory and to a lower extent in the field. In total, 12 viruses and four bacteria were found in these ant populations. Some changes in viral loads in dsRNA-treated groups were observed. For example, Linepithema humile Polycipivirus 2 (LhuPCV2) loads increased after 2 days of treatment with Spaetzle and Dicer-1 dsRNA treatments in the laboratory. After treatment with the dsRNA in the field, after 5 days the virus Linepithema humile toti-like virus 1 (LhuTLV1) was significantly more abundant. However, immune pathway disruption did not result in a consistent increase in microbial infections, nor did it alter ant abundance in the field. Some viruses even declined in abundance after dsRNA treatment. Our study explored the feasibility of lowering a pest's immunity as a control tool. We demonstrate that it is possible to alter immune gene expression of pest species and pathogen loads, although in our specific system the affected pathogens did not appear to influence pest abundance. We provide suggestions on future directions for dsRNA-mediated immune disruption in pest species, including potential avenues to improve dsRNA delivery as well as the importance of pest and pathogen biology. Double-stranded RNA targeting immune function might be especially useful for pest control in systems in which viruses or other microorganisms are prevalent and have the potential to be pathogenic.

Identity, Prevalence, and Pathogenicity of Entomopathogenic Fungi Infecting Invasive Polistes (Vespidae: Polistinae) Paper Wasps in New Zealand.

Two species of entomogenous fungi were discovered infecting the invasive paper wasp Polistes chinensis during an ecological study on Farewell Spit, New Zealand. We sequenced two nuclear ribosomal RDNA genes, the internal transcribed spacer (ITS) and the small ribosomal subunit 18S, and one protein-coding gene, the translation elongation factor 1-alpha (ef1 α). Combining sequence information with morphological examination, we identified these species as Beauveria malawiensis and Ophiocordyceps humbertii. We estimated that these fungi produce infection in approximately 3.3% of colonies in our study population. In bioassays, we successfully infected P. chinensis individuals from healthy colonies with B. malawiensis, with significant effects on adult mortality. This is the first record of both B. malawiensis and O. humbertii from Polistine hosts in New Zealand, and the first investigation into disease causality by these pathogens in P. chinensis. Our findings may contribute to the future development of biological control agents for paper wasps in New Zealand and elsewhere around the world.

Viral communities in the parasite Varroa destructor and in colonies of their honey bee host (Apis mellifera) in New Zealand.

The parasitic mite Varroa destructor is a leading cause of mortality for Western honey bee (Apis mellifera) colonies around the globe. We sought to confirm the presence and likely introduction of only one V. destructor haplotype in New Zealand, and describe the viral community within both V. destructor mites and the bees that they parasitise. A 1232 bp fragment from mitochondrial gene regions suggests the likely introduction of only one V. destructor haplotype to New Zealand. Seventeen viruses were found in bees. The most prevalent and abundant was the Deformed wing virus A (DWV-A) strain, which explained 95.0% of the variation in the viral community of bees. Black queen cell virus, Sacbrood virus, and Varroa destructor virus 2 (VDV-2) played secondary roles. DWV-B and the Israeli acute paralysis virus appeared absent from New Zealand. Ten viruses were observed in V. destructor, with > 99.9% of viral reads from DWV-A and VDV-2. Substantially more variation in viral loads was observed in bees compared to mites. Where high levels of VDV-2 occurred in mites, reduced DWV-A occurred in both the mites and the bees co-occurring within the same hive. Where there were high loads of DWV-A in mites, there were typically high viral loads in bees.

A global review of socioeconomic and environmental impacts of ants reveals new insights for risk assessment.

Risk assessments are fundamental to invasive species management and are underpinned by comprehensive characterization of invasive species impacts. Our understanding of the impacts of invasive species is growing constantly, and several recently developed frameworks offer the opportunity to systematically categorize environmental and socioeconomic impacts of invasive species. Invasive ants are among the most widespread and damaging invaders. Although a handful of species receives most of the policy attention, nearly 200 species have established outside their native range. Here, we provide a global, comprehensive assessment of the impacts of ants and propose a priority list of risk species. We used the Socioeconomic Impact Classification for Alien Taxa (SEICAT), Environmental Impact Classification for Alien Taxa (EICAT) and Generic Impact Scoring System (GISS) to analyze 642 unique sources for 100 named species. Different methodologies provided generally consistent results. The most frequently identified socioeconomic impacts were to human health. Environmental impacts were primarily on animal and plant populations, with the most common mechanisms being predation and competition. Species recognized as harmful nearly 20 years ago featured prominently, including Wasmannia auropunctata (little fire ant, electric ant), Solenopsis invicta (red imported fire ant), Anoplolepis gracilipes (yellow crazy ant), and Pheidole megacephala (African big-headed ant). All these species except W. auropunctata have been implicated in local extinctions of native species. Although our assessments affirmed that the most serious impacts have been driven by a small number of species, our results also highlighted a substantial number of less well publicized species that have had major environmental impacts and may currently be overlooked when prioritizing prevention efforts. Several of these species were ranked as high or higher than some of the previously recognized "usual suspects," most notably Nylanderia fulva (tawny crazy ant). We compared and combined our assessments with trait-based profiles and other lists to propose a consensus set of 31 priority species. Ever-increasing global trade contributes to growing rates of species introductions. The integrated approaches we used can contribute to robust, holistic risk assessments for many taxa entrained in these pathways.

A Diverse Viral Community from Predatory Wasps in Their Native and Invaded Range, with a New Virus Infectious to Honey Bees.

Wasps of the genus Vespula are social insects that have become major pests and predators in their introduced range. Viruses present in these wasps have been studied in the context of spillover from honey bees, yet we lack an understanding of the endogenous virome of wasps as potential reservoirs of novel emerging infectious diseases. We describe the characterization of 68 novel and nine previously identified virus sequences found in transcriptomes of Vespula vulgaris in colonies sampled from their native range (Belgium) and an invasive range (New Zealand). Many viruses present in the samples were from the Picorna-like virus family (38%). We identified one Luteo-like virus, Vespula vulgaris Luteo-like virus 1, present in the three life stages examined in all colonies from both locations, suggesting this virus is a highly prevalent and persistent infection in wasp colonies. Additionally, we identified a novel Iflavirus with similarity to a recently identified Moku virus, a known wasp and honey bee pathogen. Experimental infection of honey bees with this novel Vespula vulgaris Moku-like virus resulted in an active infection. The high viral diversity present in these invasive wasps is a likely indication that their polyphagous diet is a rich source of viral infections.

Nesting Ecology and Colony Survival of Two Invasive Polistes Wasps (Hymenoptera: Vespidae) in New Zealand.

We examined the abundance, nesting ecology, and colony survival of two invasive species of paper wasp, Polistes dominula Christ (Hymenoptera: Vespidae) and Polistes chinensis Pérez (Hymenoptera: Vespidae), within their invaded range in New Zealand. The more recent invader, P. dominula, exhibited a strong habitat preference, reaching the highest abundances within suburban areas with an average of 87.4 wasps per 1,000 m2. Coastal habitats were also found to be suitable environments for P. dominula, although wasp abundance in these areas was comparatively lower than suburban sites at 26.5 wasps per 1,000 m2. Although P. chinensis were observed to build more nests in coastal habitats, this was not reflected in the abundance of adult wasps in these areas. Nests of P. dominula were larger and more productive, likely a result of the multiple founding and earlier emergence of workers compared to P. chinensis. Both species exhibited significant differences in nest survival, with P. dominula observed to have a higher colony survival rate, particularly in suburban habitats where this species utilized man-made substrates as nesting sites. Neither species nested within forest sites and translocated nests of P. dominula failed to thrive within forest habitats. Findings of this research suggest that P. dominula will not pose a threat to species inhabiting forested areas. Instead, biodiversity managers should focus their efforts on suburban and coastal environments as native species in these areas will require the greatest protection.

Bioclimatic Modelling Identifies Suitable Habitat for the Establishment of the Invasive European Paper Wasp (Hymenoptera: Vespidae) across the Southern Hemisphere.

Species distribution models (SDMs) are tools used by ecologists to help predict the spread of invasive species. Information provided by these models can help direct conservation and biosecurity efforts by highlighting areas likely to contain species of interest. In this study, two models were created to investigate the potential range expansion of Polistes dominula Christ (Hymenoptera: Vespidae) in the southern hemisphere. This palearctic species has spread to invade North and South America, South Africa, Australia, and more recently New Zealand. Using the BIOCLIM and MAXENT modelling methods, regions that were suitable for P. dominula were identified based on climate data across four regions in the southern hemisphere. In South America areas of central Chile, eastern Argentina, parts of Uruguay, and southern Brazil were identified as climatically suitable for the establishment of P. dominula. Similarly, southern parts of South Africa and Australia were identified by the model to be suitable as well as much of the North Island and east of the South Island of New Zealand. Based on outputs from both models, significant range expansion by P. dominula is possible across its more southern invaded ranges.

Polistes versicolor (Hymenoptera: Vespidae), an Introduced Wasp in the Galapagos Islands: Its Life Cycle and Ecological Impact.

The yellow paper wasp, Polistes versicolor (Olivier) was first recorded in the Galapagos archipelago in 1988. Its life cycle and ecological impacts were studied on two islands 11 yr after it was first discovered. This invasive wasp adapted quickly and was found in most environments. Colony counts and adult wasp monitoring showed a strong preference for drier habitats. Nest activities were seasonally synchronized, nest building followed the rains in the hot season (typically January-May), when insect prey increases, and peaked as temperature and rains started to decline. Next, the number of adult wasps peaked during the cool season when there is barely any rain in the drier zones. In Galapagos, almost half of the prey loads of P. versicolor were lepidopteran larvae, but wasps also carried spiders, beetles, and flies back to the colonies. An estimated average of 329 mg of fresh insect prey was consumed per day for an average colony of 120-150 wasp larvae. The wasps preyed upon native and introduced insects, but likely also affect insectivorous vertebrates as competitors for food. Wasps may also compete with native pollinators as they regularly visited flowers to collect nectar, and have been recorded visiting at least 93 plant species in Galapagos, including 66 endemic and native plants. Colonies were attacked by a predatory moth, Taygete sphecophila (Meyrick) (Lepidoptera: Autostichidae), but colony development was not arrested. High wasp numbers also affect the activities of residents and tourists. A management program for this invasive species in the archipelago is essential.

Sub-lethal effects of permethrin exposure on a passerine: implications for managing ectoparasites in wild bird nests.

Permethrin is increasingly used for parasite control in bird nests, including nests of threatened passerines. We present the first formal evaluation of the effects of continued permethrin exposure on the reproductive success and liver function of a passerine, the zebra finch (Taeniopygia guttata), for two generations. We experimentally treated all nest material with a 1% permethrin solution or a water control and provided the material to breeding finches for nest building. The success of two consecutive clutches produced by the parental generation and one clutch produced by first-generation birds were tracked. Finches in the first generation were able to reproduce and fledge offspring after permethrin exposure, ruling out infertility. Permethrin treatment had no statistically significant effect on the number of eggs laid, number of days from clutch initiation to hatching, egg hatch rate, fledgling mass or nestling sex ratio in either generation. However, treating nest material with permethrin significantly increased the number of hatchlings in the first generation and decreased fledgling success in the second generation. Body mass for hatchlings exposed to permethrin was lower than for control hatchlings in both generations, but only statistically significant for the second generation. For both generations, an interaction between permethrin treatment and age significantly affected nestling growth. Permethrin treatment had no effect on liver function for any generation. Permethrin was detected inside 6 of 21 exposed, non-embryonated eggs (28.5% incidence; range: 693-4781 ng of permethrin per gram of dry egg mass). Overall, results from exposing adults, eggs and nestlings across generations to permethrin-treated nest material suggest negative effects on finch breeding success, but not on liver function. For threatened bird conservation, the judicious application of this insecticide to control parasites in nests can result in lower nestling mortality compared to when no treatment is applied. Thus, permethrin treatment benefits may outweigh its sub-lethal effects.

High-Quality Assemblies for Three Invasive Social Wasps from the Vespula Genus.

Social wasps of the genus Vespula have spread to nearly all landmasses worldwide and have become significant pests in their introduced ranges, affecting economies and biodiversity. Comprehensive genome assemblies and annotations for these species are required to develop the next generation of control strategies and monitor existing chemical control. We sequenced and annotated the genomes of the common wasp (Vespula vulgaris), German wasp (Vespula germanica), and the western yellowjacket (Vespula pensylvanica). Our chromosome-level Vespula assemblies each contain 176-179 Mb of total sequence assembled into 25 scaffolds, with 10-200 unanchored scaffolds, and 16,566-18,948 genes. We annotated gene sets relevant to the applied management of invasive wasp populations, including genes associated with spermatogenesis and development, pesticide resistance, olfactory receptors, immunity and venom. These genomes provide evidence for active DNA methylation in Vespidae and tandem duplications of venom genes. Our genomic resources will contribute to the development of next-generation control strategies, and monitoring potential resistance to chemical control.

The potential for a CRISPR gene drive to eradicate or suppress globally invasive social wasps.

CRISPR gene drives have potential for widespread and cost-efficient pest control, but are highly controversial. We examined a potential gene drive targeting spermatogenesis to control the invasive common wasp (Vespula vulgaris) in New Zealand. Vespula wasps are haplodiploid. Their life cycle makes gene drive production challenging, as nests are initiated by single fertilized queens in spring followed by several cohorts of sterile female workers and the production of reproductives in autumn. We show that different spermatogenesis genes have different levels of variation between introduced and native ranges, enabling a potential 'precision drive' that could target the reduced genetic diversity and genotypes within the invaded range. In vitro testing showed guide-RNA target specificity and efficacy that was dependent on the gene target within Vespula, but no cross-reactivity in other Hymenoptera. Mathematical modelling incorporating the genetic and life history traits of Vespula wasps identified characteristics for a male sterility drive to achieve population control. There was a trade-off between drive infiltration and impact: a drive causing complete male sterility would not spread, while partial sterility could be effective in limiting population size if the homing rate is high. Our results indicate that gene drives may offer viable suppression for wasps and other haplodiploid pests.

Genetic Strain Diversity of Multi-Host RNA Viruses that Infect a Wide Range of Pollinators and Associates is Shaped by Geographic Origins.

Emerging viruses have caused concerns about pollinator population declines, as multi-host RNA viruses may pose a health threat to pollinators and associated arthropods. In order to understand the ecology and impact these viruses have, we studied their host range and determined to what extent host and spatial variation affect strain diversity. Firstly, we used RT-PCR to screen pollinators and associates, including honey bees (Apis mellifera) and invasive Argentine ants (Linepithema humile), for virus presence and replication. We tested for the black queen cell virus (BQCV), deformed wing virus (DWV), and Kashmir bee virus (KBV) that were initially detected in bees, and the two recently discovered Linepithema humile bunya-like virus 1 (LhuBLV1) and Moku virus (MKV). DWV, KBV, and MKV were detected and replicated in a wide range of hosts and commonly co-infected hymenopterans. Secondly, we placed KBV and DWV in a global phylogeny with sequences from various countries and hosts to determine the association of geographic origin and host with shared ancestry. Both phylogenies showed strong geographic rather than host-specific clustering, suggesting frequent inter-species virus transmission. Transmission routes between hosts are largely unknown. Nonetheless, avoiding the introduction of non-native species and diseased pollinators appears important to limit spill overs and disease emergence.

The association between mitochondrial genetic variation and reduced colony fitness in an invasive wasp.

Despite the mitochondrion's long-recognized role in energy production, mitochondrial DNA (mtDNA) variation commonly found in natural populations was assumed to be effectively neutral. However, variation in mtDNA has now been increasingly linked to phenotypic variation in life history traits and fitness. We examined whether the relative fitness in native and invasive common wasp (Vespula vulgaris) populations in Belgium and New Zealand (NZ), respectively, can be linked to mtDNA variation. Social wasp colonies in NZ were smaller with comparatively fewer queen cells, indicating a reduced relative fitness in the invaded range. Interestingly, queen cells in this population were significantly larger leading to larger queen offspring. By sequencing 1,872 bp of the mitochondrial genome, we determined mitochondrial haplotypes and detected reduced genetic diversity in NZ. Three common haplotypes in NZ frequently produced many queens, whereas the four rare haplotypes produced significantly fewer or no queens. The entire mitochondrial genome for each of these haplotypes was sequenced to identify polymorphisms associated with fitness reduction. We found 16 variable sites; however, no nonsynonymous mutation that was clearly causing impaired mitochondrial function was detected. We discuss how detected variants may alter secondary structures, gene expression or mito-nuclear interactions, or could be associated with nuclear-encoded variation. Whatever the ultimate mechanism, we show reduced fitness and mtDNA variation in an invasive wasp population as well as specific mtDNA variants associated with fitness variation within this population. Ours is one of only a few studies that confirm fitness impacts of mtDNA variation in wild nonmodel populations.

Different bacterial and viral pathogens trigger distinct immune responses in a globally invasive ant.

Invasive species populations periodically collapse from high to low abundance, sometimes even to extinction. Pathogens and the burden they place on invader immune systems have been hypothesised as a mechanism for these collapses. We examined the association of the bacterial pathogen (Pseudomonas spp.) and the viral community with immune gene expression in the globally invasive Argentine ant (Linepithema humile (Mayr)). RNA-seq analysis found evidence for 17 different viruses in Argentine ants from New Zealand, including three bacteriophages with one (Pseudomonas phage PS-1) likely to be attacking the bacterial host. Pathogen loads and prevalence varied immensely. Transcriptomic data showed that immune gene expression was consistent with respect to the viral classification of negative-sense, positive-sense and double-stranded RNA viruses. Genes that were the most strongly associated with the positive-sense RNA viruses such as the Linepithema humile virus 1 (LHUV-1) and the Deformed wing virus (DWV) were peptide recognition proteins assigned to the Toll and Imd pathways. We then used principal components analysis and regression modelling to determine how RT-qPCR derived immune gene expression levels were associated with viral and bacterial loads. Argentine ants mounted a substantial immune response to both Pseudomonas and LHUV-1 infections, involving almost all immune pathways. Other viruses including DWV and the Kashmir bee virus appeared to have much less immunological influence. Different pathogens were associated with varying immunological responses, which we hypothesize to interact with and influence the invasion dynamics of this species.