Immune system modulation & virus transmission during parasitism identified by multi-species transcriptomics of a declining insect biocontrol system.

BACKGROUND: The Argentine stem weevil (ASW, Listronotus bonariensis) is a significant pasture pest in Aotearoa New Zealand, primarily controlled by the parasitoid biocontrol agent Microctonus hyperodae. Despite providing effective control of ASW soon after release, M. hyperodae parasitism rates have since declined significantly, with ASW hypothesised to have evolved resistance to its biocontrol agent. While the parasitism arsenal of M. hyperodae has previously been investigated, revealing many venom components and an exogenous novel DNA virus Microctonus hyperodae filamentous virus (MhFV), the effects of said arsenal on gene expression in ASW during parasitism have not been examined. In this study, we performed a multi-species transcriptomic analysis to investigate the biology of ASW parasitism by M. hyperodae, as well as the decline in efficacy of this biocontrol system. RESULTS: The transcriptomic response of ASW to parasitism by M. hyperodae involves modulation of the weevil's innate immune system, flight muscle components, and lipid and glucose metabolism. The multispecies approach also revealed continued expression of venom components in parasitised ASW, as well as the transmission of MhFV to weevils during parasitism and some interrupted parasitism attempts. Transcriptomics did not detect a clear indication of parasitoid avoidance or other mechanisms to explain biocontrol decline. CONCLUSIONS: This study has expanded our understanding of interactions between M. hyperodae and ASW in a biocontrol system of critical importance to Aotearoa-New Zealand's agricultural economy. Transmission of MhFV to ASW during successful and interrupted parasitism attempts may link to a premature mortality phenomenon in ASW, hypothesised to be a result of a toxin-antitoxin system. Further research into MhFV and its potential role in ASW premature mortality is required to explore whether manipulation of this viral infection has the potential to increase biocontrol efficacy in future.

Chromosome-level genome assemblies of two parasitoid biocontrol wasps reveal the parthenogenesis mechanism and an associated novel virus.

BACKGROUND: Biocontrol is a key technology for the control of pest species. Microctonus parasitoid wasps (Hymenoptera: Braconidae) have been released in Aotearoa New Zealand as biocontrol agents, targeting three different pest weevil species. Despite their value as biocontrol agents, no genome assemblies are currently available for these Microctonus wasps, limiting investigations into key biological differences between the different species and strains. METHODS AND FINDINGS: Here we present high-quality genomes for Microctonus hyperodae and Microctonus aethiopoides, assembled with short read sequencing and Hi-C scaffolding. These assemblies have total lengths of 106.7 Mb for M. hyperodae and 129.2 Mb for M. aethiopoides, with scaffold N50 values of 9 Mb and 23 Mb respectively. With these assemblies we investigated differences in reproductive mechanisms, and association with viruses between Microctonus wasps. Meiosis-specific genes are conserved in asexual Microctonus, with in-situ hybridisation validating expression of one of these genes in the ovaries of asexual Microctonus aethiopoides. This implies asexual reproduction in these Microctonus wasps involves meiosis, with the potential for sexual reproduction maintained. Investigation of viral gene content revealed candidate genes that may be involved in virus-like particle production in M. aethiopoides, as well as a novel virus infecting M. hyperodae, for which a complete genome was assembled. CONCLUSION AND SIGNIFICANCE: These are the first published genomes for Microctonus wasps which have been deployed as biocontrol agents, in Aotearoa New Zealand. These assemblies will be valuable resources for continued investigation and monitoring of these biocontrol systems. Understanding the biology underpinning Microctonus biocontrol is crucial if we are to maintain its efficacy, or in the case of M. hyperodae to understand what may have influenced the significant decline of biocontrol efficacy. The potential for sexual reproduction in asexual Microctonus is significant given that empirical modelling suggests this asexual reproduction is likely to have contributed to biocontrol decline. Furthermore the identification of a novel virus in M. hyperodae highlights a previously unknown aspect of this biocontrol system, which may contribute to premature mortality of the host pest. These findings have potential to be exploited in future in attempt to increase the effectiveness of M. hyperodae biocontrol.

The venom composition and parthenogenesis mechanism of the parasitoid wasp Microctonus hyperodae, a declining biocontrol agent.

A biocontrol system in New Zealand using the endoparasitoid Microctonus hyperodae is failing, despite once being one of the most successful examples of classical biocontrol worldwide. Though it is of significant economic importance as a control agent, little is known about the genetics of M. hyperodae. In this study, RNA-seq was used to characterise two key traits of M. hyperodae in this system, the venom, critical for the initial success of biocontrol, and the asexual reproduction mode, which influenced biocontrol decline. Expanded characterisation of M. hyperodae venom revealed candidates involved in manipulating the host environment to source nutrition for the parasitoid egg, preventing a host immune response against the egg, as well as two components that may stimulate the host's innate immune system. Notably lacking from the venom-specific expression list was calreticulin, as it also had high expression in the ovaries. In-situ hybridisation revealed this ovarian expression was localised to the follicle cells, which may result in the deposition of calreticulin into the egg exochorion. Investigating the asexual reproduction of M. hyperodae revealed core meiosis-specific genes had conserved expression patterns with the highest expression in the ovaries, suggesting M. hyperodae parthenogenesis involves meiosis and that the potential for sexual reproduction may have been retained. Upregulation of genes involved in endoreduplication provides a potential mechanism for the restoration of diploidy in eggs after meiosis.

Iatrogenic Atrial Septal Defect Closure Through the Steerable Guide Catheter: Description of Technique and Single-Center Experience.

OBJECTIVES: To introduce a novel method of direct iatrogenic atrial septal defect (iASD) closure through the MitraClip steerable guide catheter (SGC). BACKGROUND: MitraClip implantation requires transseptal puncture and the creation of an iASD. There are relatively rare instances, such as hemodynamically significant shunting or concerns for embolus, where iASD must be closed during index procedure. In these instances, it may be beneficial to not give up access to left atrium. METHODS: We retrospectively reviewed all iASD closures during MitraClip implantation at our institution from 2015 to 2020. Cases where an ASD occluder was deployed directly through SGC were included. RESULTS: Eleven patients had immediate iASD closure through the SGC. Indications for using this method included concern for paradoxical embolus, large defect size and/or significant shunting. Closure device sizes ranged from 8 to 22 mm. Mean time from removal of clip delivery system to occlusion of iASD was 14.6 minutes. There were no procedural complications related to iASD closure using this method. CONCLUSION: Closure of iASD intra-procedurally directly through transseptal guide sheath via the method described was safe and allowed for continuous left atrium access.

A resilience sensing system for the biosphere.

We are in a climate and ecological emergency, where climate change and direct anthropogenic interference with the biosphere are risking abrupt and/or irreversible changes that threaten our life-support systems. Efforts are underway to increase the resilience of some ecosystems that are under threat, yet collective awareness and action are modest at best. Here, we highlight the potential for a biosphere resilience sensing system to make it easier to see where things are going wrong, and to see whether deliberate efforts to make things better are working. We focus on global resilience sensing of the terrestrial biosphere at high spatial and temporal resolution through satellite remote sensing, utilizing the generic mathematical behaviour of complex systems-loss of resilience corresponds to slower recovery from perturbations, gain of resilience equates to faster recovery. We consider what subset of biosphere resilience remote sensing can monitor, critically reviewing existing studies. Then we present illustrative, global results for vegetation resilience and trends in resilience over the last 20 years, from both satellite data and model simulations. We close by discussing how resilience sensing nested across global, biome-ecoregion, and local ecosystem scales could aid management and governance at these different scales, and identify priorities for further work. This article is part of the theme issue 'Ecological complexity and the biosphere: the next 30 years'.

Genomic signatures of parallel alpine adaptation in recently evolved flightless insects.

Natural selection along elevational gradients has potential to drive predictable adaptations across distinct lineages, but the extent of such repeated evolution remains poorly studied for many widespread alpine taxa. We present parallel genomic analyses of two recently evolved flightless alpine insect lineages to test for molecular signatures of repeated alpine adaptation. Specifically, we compare low-elevation vs. alpine stonefly ecotypes from parallel stream populations in which flightless upland ecotypes have been independently derived. We map 67,922 polymorphic genetic markers, generated across 176 Zelandoperla fenestrata specimens from two independent alpine stream populations in New Zealand's Rock and Pillar Range, to a newly developed plecopteran reference genome. Genome-wide scans revealed 31 regions with outlier single nucleotide polymorphisms (SNPs) differentiating lowland vs. alpine ecotypes in Lug Creek, and 37 regions with outliers differentiating ecotypes in Six Mile Creek. Of these regions, 13% (8/60) yielded outlier SNPs across both within-stream ecotype comparisons, implying comparable genomic shifts contribute to this repeated alpine adaptation. Candidate genes closely linked to repeated outlier regions include several with documented roles in insect wing-development (e.g., dishevelled), suggesting that they may contribute to repeated alpine wing reduction. Additional candidate genes have been shown to influence insect fecundity (e.g., ovo) and lifespan (e.g., Mrp4), implying that they might contribute to life history differentiation between upland and lowland ecotypes. Additional outlier genes have potential roles in the evolution of reproductive isolation among ecotypes (hedgehog and Desaturase 1). These results demonstrate how replicated outlier tests across independent lineages can potentially contribute to the discovery of genes underpinning repeated adaptation.

MitraClip Implantation Guided by Volumetric Intracardiac Echocardiography: Technique and Feasibility in Patients Intolerant to Transesophageal Echocardiography.

Transesophageal echo (TEE) guidance is essential for successful MitraClip implantation. In patients intolerant to TEE, options are limited. Three patients, with contraindications to TEE, underwent MitraClip implantation using volumetric intracardiac echo (vICE). All patients had severe symptomatic degenerative mitral regurgitation (MR) and had successful implantation of at least one clip with reduction of MR to ≤2+ maintained at 30d follow-up. All patients had improvement in functional status without any adverse clinical, clip, or vICE related events. We discuss in detail the technical considerations and limitations to performing the MitraClip procedure with vICE guidance.

Human liver-derived MAIT cells differ from blood MAIT cells in their metabolism and response to TCR-independent activation.

Mucosal associated invariant T (MAIT) cells are anti-microbial innate-like T cells that are abundant in blood and liver. MAIT cells express a semi-invariant T-cell receptor (TCR) that recognizes a pyrimidine ligand, derived from microbial riboflavin synthesis, bound to MR1. Both blood and liver derived (ld)-MAIT cells can be robustly stimulated via TCR or by cytokines produced during bacterial or viral infection. In this study, we compared the functional and transcriptomic response of human blood and ld-MAIT cells to TCR signals (Escherichia coli or the pyrimidine ligand) and cytokines (IL-12 + IL-18). While the response of blood and ld-MAIT cells to TCR signals were comparable, following cytokine stimulation ld-MAIT cells were more polyfunctional than blood MAIT cells. Transcriptomic analysis demonstrated different effector programmes of ld-MAIT cells with the two modes of activation, including the enrichment of a tissue repair signature in TCR-stimulated MAIT cells. Interestingly, we observed enhancement of IL-12 signaling and fatty acid metabolism in untreated ld-MAIT cells compared with blood MAIT cells. Additionally, MAIT cells from blood and liver were modulated similarly by TCR and cytokine signals. Therefore, we report that blood and ld-MAIT cells are fundamentally different but undergo conserved changes following activation via TCR or by cytokines.

Genomics Reveals Widespread Ecological Speciation in Flightless Insects.

Recent genomic analyses have highlighted parallel divergence in response to ecological gradients, but the extent to which altitude can underpin such repeated speciation remains unclear. Wing reduction and flight loss have apparently evolved repeatedly in montane insect assemblages and have been suggested as important drivers of hexapod diversification. We test this hypothesis using genomic analyses of a widespread wing-polymorphic stonefly species complex in New Zealand. We identified over 50,000 polymorphic genetic markers generated across almost 200 Zelandoperla fenestrata stonefly specimens using a newly generated plecopteran reference genome, to reveal widespread parallel speciation between sympatric full-winged and wing-reduced ecotypes. Rather than the existence of a single, widespread, flightless taxon (Zelandoperla pennulata), evolutionary genomic data reveal that wing-reduced upland lineages have speciated repeatedly and independently from full-winged Z. fenestrata. This repeated evolution of reproductive isolation between local ecotype pairs that lack mitochondrial DNA differentiation suggests that ecological speciation has evolved recently. A cluster of outlier single-nucleotide polymorphisms detected in independently wing-reduced lineages, tightly linked in an approximately 85 kb genomic region that includes the developmental "supergene" doublesex, suggests that this "island of divergence" may play a key role in rapid ecological speciation. [Ecological speciation; genome assembly; genomic island of differentiation; genotyping-by-sequencing; incipient species; plecoptera; wing reduction.].

Population genomics of the critically endangered kakapo.

The kakapo is a flightless parrot endemic to New Zealand. Once common in the archipelago, only 201 individuals remain today, most of them descending from an isolated island population. We report the first genome-wide analyses of the species, including a high-quality genome assembly for kakapo, one of the first chromosome-level reference genomes sequenced by the Vertebrate Genomes Project (VGP). We also sequenced and analyzed 35 modern genomes from the sole surviving island population and 14 genomes from the extinct mainland population. While theory suggests that such a small population is likely to have accumulated deleterious mutations through genetic drift, our analyses on the impact of the long-term small population size in kakapo indicate that present-day island kakapo have a reduced number of harmful mutations compared to mainland individuals. We hypothesize that this reduced mutational load is due to the island population having been subjected to a combination of genetic drift and purging of deleterious mutations, through increased inbreeding and purifying selection, since its isolation from the mainland ∼10,000 years ago. Our results provide evidence that small populations can survive even when isolated for hundreds of generations. This work provides key insights into kakapo breeding and recovery and more generally into the application of genetic tools in conservation efforts for endangered species.

Adjunctive use of fluoroscopy during MitraClip implantation reduces procedural complexity: The parallax technique.

BACKGROUND: During MitraClip implantation sub-valvular correction of trajectory and/or alignment may increase adverse clip or leaflet events. With systematic adjunctive use of fluoroscopy ("Parallax technique"), we aimed to assess parameters that minimize the need for corrective measures and help increase procedural efficiency. METHODS: We retrospectively analyzed 30 patients without (Fl-) and 39 patients utilizing adjunctive fluoroscopy (Fl+) during MitraClip implantation. After establishing trajectory and supra-valvular alignment, the Parallax technique was utilized. Trajectory and alignment are maintained during advancement. RESULTS: All patients had 3 or 4+ MR. There were no differences in baseline demographics. The average number of clips (Fl- vs Fl+) was 1.72 ± 0.8 vs 1.59 ± 0.5, p = .57. For the first clip, the need for sub-valvular alignment (80% vs. 36%, p = .0001), eversion with retraction back to left atrium (23% vs. 10%, p = .001) and the number of grasps (2.3 ± 1.2 vs 1.4 ± 0.9) was reduced. The time from transseptal puncture to first clip deployment (71 ± 21 vs 44 ± 16 min, p = .01) was reduced. Procedural success was achieved in all but one patient in the Fl- group (p = ns). There were no differences noted for in-hospital or 30-day outcomes. CONCLUSIONS: Systematic use of a simple and easy to implement "Parallax technique" was associated with reduced need for sub-valvular manipulation and was associated with improved procedural times. Further larger scale studies are needed to assess the applicability of the technique.

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.

Genetic Diversity in Invasive Populations of Argentine Stem Weevil Associated with Adaptation to Biocontrol.

Modified, agricultural landscapes are susceptible to damage by insect pests. Biological control of pests is typically successful once a control agent has established, but this depends on the agent's capacity to co-evolve with the host. Theoretical studies have shown that different levels of genetic variation between the host and the control agent will lead to rapid evolution of resistance in the host. Although this has been reported in one instance, the underlying genetics have not been studied. To address this, we measured the genetic variation in New Zealand populations of the pasture pest, Argentine stem weevil (Listronotus bonariensis), which is controlled with declining effectiveness by a parasitoid wasp, Microctonus hyperodae. We constructed a draft reference genome of the weevil, collected samples from a geographical survey of 10 sites around New Zealand, and genotyped them using a modified genotyping-by-sequencing approach. New Zealand populations of Argentine stem weevil have high levels of heterozygosity and low population structure, consistent with a large effective population size and frequent gene flow. This implies that Argentine stem weevils were able to evolve more rapidly than their biocontrol agent, which reproduces asexually. These findings show that monitoring genetic diversity in biocontrol agents and their targets is critical for long-term success of biological control.

Transjugular mitral valve repair with the MitraClip: A step-by-step guide.

The MitraClip procedure is carried out almost exclusively via the transfemoral approach. However, in some patients transfemoral delivery of MitraClip is not technically feasible (e.g., occluded inferior vena cava or tortuous/obstructive iliofemoral venous anatomy). The technical considerations and challenges of the MitraClip procedure are amplified when an alternate route is considered. We describe a successful case of MitraClip performed via the right internal jugular (IJ) approach in a patient with a flail A3 scallop and previous mitral valve repair. We reviewed prior cases in the literature and discuss step-by-step the pertinent clinical and technical considerations for performing this procedure via the transjugular route. In summary, the right IJ access site provides a reasonable alternative to perform the MitraClip procedure in cases where femoral access is contraindicated or prohibitive. A thorough understanding of the technical considerations is crucial in improving procedural success rates.

Transseptal access for left heart structural interventions in the setting of prior atrial septal defect closure.

A transseptal puncture is critical for "left-sided" structural heart interventions. Procedures such as transcatheter edge-to-edge repair (MitraClip) and left atrial appendage (LAA) closure (Watchman) require precise puncture of the interatrial septum (IAS), and the presence of a prior atrial septal defect (ASD) closure device poses a challenge. We aim to present a successfully completed case of MitraClip and Watchman in the presence of ASD closure device in two different patients. A review of the literature will be reported, and pertinent clinical and technical considerations will be discussed in depth to achieve procedural success. In summary, transseptal puncture for left heart structural interventions is feasible in the presence of a prior ASD/patent foramen ovale closure device. A detailed understanding of the anatomical considerations as well as the use of multimodality imaging to evaluate the IAS will aid in improving procedural success rates.

The Pacific Biosciences de novo assembled genome dataset from a parthenogenetic New Zealand wild population of the longhorned tick, Haemaphysalis longicornis Neumann, 1901.

The longhorned tick, Haemaphysalis longicornis, feeds upon a wide range of bird and mammalian hosts. Mammalian hosts include cattle, deer, sheep, goats, humans, and horses. This tick is known to transmit a number of pathogens causing tick-borne diseases, and was the vector of a recent serious outbreak of oriental theileriosis in New Zealand. A New Zealand-USA consortium was established to sequence, assemble, and annotate the genome of this tick, using ticks obtained from New Zealand's North Island. In New Zealand, the tick is considered exclusively parthenogenetic and this trait was deemed useful for genome assembly. Very high molecular weight genomic DNA was sequenced on the Illumina HiSeq4000 and the long-read Pac Bio Sequel platforms. Twenty-eight SMRT cells produced a total of 21.3 million reads which were assembled with Canu on a reserved supercomputer node with access to 12 TB of RAM, running continuously for over 24 days. The final assembly dataset consisted of 34,211 contigs with an average contig length of 215,205 bp. The quality of the annotated genome was assessed by BUSCO analysis, an approach that provides quantitative measures for the quality of an assembled genome. Over 95% of the BUSCO gene set was found in the assembled genome. Only 48 of the 1066 BUSCO genes were missing and only 9 were present in a fragmented condition. The raw sequencing reads and the assembled contigs/scaffolds are archived at the National Center for Biotechnology Information.

TCR- or Cytokine-Activated CD8+ Mucosal-Associated Invariant T Cells Are Rapid Polyfunctional Effectors That Can Coordinate Immune Responses.

Mucosal-associated invariant T (MAIT) cells can be activated via either their T cell receptor (TCR), which recognizes MR1-bound pyrimidines derived from microbial riboflavin biosynthesis, or via cytokines. These two modes of activation may act in concert or independently, depending upon the stimulus. It is unknown, however, how MAIT cell responses differ with the mode of activation. Here, we define transcriptional and effector responses of human CD8+ MAIT cells to TCR and cytokine stimulation. We report that MAIT cells rapidly respond to TCR stimulation, producing multiple cytokines and chemokines, altering their cytotoxic granule content and transcription factor expression, and upregulating co-stimulatory proteins. In contrast, cytokine-mediated activation is slower and results in a more limited response. Therefore, we propose that, in infections by riboflavin-synthesizing bacteria, MAIT cells play a key early role in effecting and coordinating immune responses, while in the absence of TCR stimulation, their role is likely to differ.