Spatial and temporal mapping of de novo mutations in schizophrenia to a fetal prefrontal cortical network.

Genes disrupted in schizophrenia may be revealed by de novo mutations in affected persons from otherwise healthy families. Furthermore, during normal brain development, genes are expressed in patterns specific to developmental stage and neuroanatomical structure. We identified de novo mutations in persons with schizophrenia and then mapped the responsible genes onto transcriptome profiles of normal human brain tissues from age 13 weeks gestation to adulthood. In the dorsolateral and ventrolateral prefrontal cortex during fetal development, genes harboring damaging de novo mutations in schizophrenia formed a network significantly enriched for transcriptional coexpression and protein interaction. The 50 genes in the network function in neuronal migration, synaptic transmission, signaling, transcriptional regulation, and transport. These results suggest that disruptions of fetal prefrontal cortical neurogenesis are critical to the pathophysiology of schizophrenia. These results also support the feasibility of integrating genomic and transcriptome analyses to map critical neurodevelopmental processes in time and space in the brain.

G-U base-paired hpRNA confers potent inhibition of small RNA function in plants.

MicroRNA (miRNA) target mimicry technologies, utilizing naturally occurring miRNA decoy molecules, represent a potent tool for analyzing miRNA function. In this study, we present a highly efficient small RNA (sRNA) target mimicry design based on G-U base-paired hairpin RNA (hpG:U), which allows for the simultaneous targeting of multiple sRNAs. The hpG:U constructs consistently generate high amounts of intact, polyadenylated stem-loop (SL) RNA outside the nuclei, in contrast to traditional hairpin RNA designs with canonical base pairing (hpWT), which were predominantly processed resulting in a loop. By incorporating a 460-bp G-U base-paired double-stranded stem and a 312-576 nt loop carrying multiple miRNA target mimicry sites (GUMIC), the hpG:U construct displayed effective repression of three Arabidopsis miRNAs, namely miR165/166, miR157, and miR160, both individually and in combination. Additionally, a GUMIC construct targeting a prominent cluster of siRNAs derived from cucumber mosaic virus (CMV) Y-satellite RNA (Y-Sat) effectively inhibited Y-Sat siRNA-directed silencing of the chlorophyll biosynthetic gene CHLI, thereby reducing the yellowing symptoms in infected Nicotiana plants. Therefore, the G-U base-paired hpRNA, characterized by differential processing compared to traditional hpRNA, acts as an efficient decoy for both miRNAs and siRNAs. This technology holds great potential for sRNA functional analysis and the management of sRNA-mediated diseases.

Rapid adaptation and interspecific introgression in the North American crop pest Helicoverpa zea.

Insect crop pests threaten global food security. This threat is amplified through the spread of non-native species and through adaptation of native pests to control measures. Adaptations such as pesticide resistance can result from selection on variation within a population, or through gene flow from another population. We investigate these processes in an economically important noctuid crop pest, Helicoverpa zea, which has evolved resistance to a wide range of pesticides. Its sister species Helicoverpa armigera, first detected as an invasive species in Brazil in 2013, introduced the pyrethroid resistance gene CYP337B3 to South American H. zea via adaptive introgression. To understand whether this could contribute to pesticide resistance in North America, we sequenced 237 H. zea genomes across 10 sample sites. We report H. armigera introgression into the North American H. zea population. Two individuals sampled in Texas in 2019 carry H. armigera haplotypes in a 4Mbp region containing CYP337B3. Next, we identify signatures of selection in the panmictic population of non-admixed H. zea, identifying a selective sweep at a second cytochrome P450 gene: CYP333B3. We estimate that its derived allele conferred a ∼5% fitness advantage and show that this estimate explains independently observed rare nonsynonymous CYP333B3 mutations approaching fixation over a ∼20-year period. We also detect putative signatures of selection at a kinesin gene associated with Bt resistance. Overall, we document two mechanisms of rapid adaptation: the introduction of fitness-enhancing alleles through interspecific introgression, and selection on intraspecific variation.

Targeted long-read sequencing identifies missing disease-causing variation.

Despite widespread clinical genetic testing, many individuals with suspected genetic conditions lack a precise diagnosis, limiting their opportunity to take advantage of state-of-the-art treatments. In some cases, testing reveals difficult-to-evaluate structural differences, candidate variants that do not fully explain the phenotype, single pathogenic variants in recessive disorders, or no variants in genes of interest. Thus, there is a need for better tools to identify a precise genetic diagnosis in individuals when conventional testing approaches have been exhausted. We performed targeted long-read sequencing (T-LRS) using adaptive sampling on the Oxford Nanopore platform on 40 individuals, 10 of whom lacked a complete molecular diagnosis. We computationally targeted up to 151 Mbp of sequence per individual and searched for pathogenic substitutions, structural variants, and methylation differences using a single data source. We detected all genomic aberrations-including single-nucleotide variants, copy number changes, repeat expansions, and methylation differences-identified by prior clinical testing. In 8/8 individuals with complex structural rearrangements, T-LRS enabled more precise resolution of the mutation, leading to changes in clinical management in one case. In ten individuals with suspected Mendelian conditions lacking a precise genetic diagnosis, T-LRS identified pathogenic or likely pathogenic variants in six and variants of uncertain significance in two others. T-LRS accurately identifies pathogenic structural variants, resolves complex rearrangements, and identifies Mendelian variants not detected by other technologies. T-LRS represents an efficient and cost-effective strategy to evaluate high-priority genes and regions or complex clinical testing results.

Multidisciplinary Surgical Approach to Increase Survival for Advanced Ovarian Cancer in a Tertiary Gynaecological Oncology Centre.

PURPOSE: The purpose of this paper is to report on changes in overall survival, progression-free survival, and complete cytoreduction rates in the 5-year period after the implementation of a multidisciplinary surgical team (MDT). METHODS: Two cohorts were used. Cohort A was a retrospectively collated cohort from 2006 to 2015. Cohort B was a prospectively collated cohort of patients from January 2017 to September 2021. RESULTS: This study included 146 patients in cohort A (2006-2015) and 174 patients in cohort B (2017-2021) with FIGO stage III/IV ovarian cancer. Median follow-up in cohort A was 60 months and 48 months in cohort B. The rate of primary cytoreductive surgery increased from 38% (55/146) in cohort A to 46.5% (81/174) in cohort B. Complete macroscopic resection increased from 58.9% (86/146) in cohort A to 78.7% (137/174) in cohort B (p < 0.001). At 3 years, 75% (109/144) patients had disease progression in cohort A compared with 48.8% (85/174) in cohort B (log-rank, p < 0.001). Also at 3 years, 64.5% (93/144) of patients had died in cohort A compared with 24% (42/174) of cohort B (log-rank, p < 0.001). Cox multivariate analysis demonstrated that MDT input, residual disease, and age were independent predictors of overall (hazard ratio [HR] 0.29, 95% confidence interval [CI] 0.203-0.437, p < 0.001) and progression-free survival (HR 0.31, 95% CI 0.21-0.43, p < 0.001). Major morbidity remained stable throughout both study periods (2006-2021). CONCLUSIONS: Our data demonstrate that the implementation of multidisciplinary-team, intraoperative approach allowed for a change in surgical philosophy and has resulted in a significant improvement in overall survival, progression-free survival, and complete resection rates.

Historical museum samples reveal signals of selection and drift in response to changing insecticide use in an agricultural pest moth.

In response to environmental and human-imposed selective pressures, agroecosystem pests frequently undergo rapid evolution, with some species having a remarkable capacity to rapidly develop pesticide resistance. Temporal sampling of genomic data can comprehensively capture such adaptive changes over time, for example, by elucidating allele frequency shifts in pesticide resistance loci in response to different pesticides. Here, we leveraged museum specimens spanning over a century of collections to generate temporal contrasts between pre- and post-insecticide populations of an agricultural pest moth, Helicoverpa armigera. We used targeted exon sequencing of 254 samples collected across Australia from the pre-1950s (prior to insecticide introduction) to the 1990s, encompassing decades of changing insecticide use. Our sequencing approach focused on genes that are known to be involved in insecticide resistance, environmental sensation, and stress tolerance. We found an overall lack of spatial and temporal population structure change across Australia. In some decades (e.g., 1960s and 1970s), we found a moderate reduction of genetic diversity, implying stochasticity in evolutionary trajectories due to genetic drift. Temporal genome scans showed extensive evidence of selection following insecticide use, although the majority of selected variants were low impact, and alternating trajectories of allele frequency change were suggestive of potential antagonistic pleiotropy. Our results provide new insights into recent evolutionary responses in an agricultural pest and show how temporal contrasts using museum specimens can improve mechanistic understanding of rapid evolution.

Clinically Complex LRBA Deficiency Due to a Founder Allele in the Georgian Jewish Population.

Pathogenic variants in LRBA, encoding the LPS Responsive Beige-Like Anchor (LRBA) protein, are responsible for recessive, early-onset hypogammaglobulinemia, severe multi-organ autoimmunity, and lymphoproliferation, with increased risk for malignancy. LRBA deficiency has a wide clinical spectrum with variable age of onset and disease severity. Three apparently unrelated patients with LRBA deficiency, of Georgian Jewish descent, were homozygous for LRBA c.6640C > T, p.R2214*, leading to a stop upstream of the LRBA BEACH domain. Despite carrying the same LRBA genotype, the three patients differed in clinical course: the first patient was asymptomatic until age 25 years; the second presented with failure to thrive at age 3 months; and the third presented at age 7 years with immune cytopenias and severe infections. Two of the patients developed malignancies: the first patient was diagnosed with recurrent Hodgkin's disease at age 36 years, and the second patient developed aggressive gastric cancer at age 15 years. Among Georgian Jews, the carrier frequency of the LRBA p.R2214* allele was 1.6% (4 of 236 Georgian Jewish controls). The allele was absent from other populations. Haplotype analysis showed a shared origin of the mutation. These three patients revealed a pathogenic LRBA founder allele in the Georgian Jewish population, support the diverse and complex clinical spectrum of LRBA deficiency, and support the possibility that LRBA deficiency predisposes to malignancy.

The redlegged earth mite draft genome provides new insights into pesticide resistance evolution and demography in its invasive Australian range.

Genomic data provide valuable insights into pest management issues such as resistance evolution, historical patterns of pest invasions and ongoing population dynamics. We assembled the first reference genome for the redlegged earth mite, Halotydeus destructor (Tucker, 1925), to investigate adaptation to pesticide pressures and demography in its invasive Australian range using whole-genome pool-seq data from regionally distributed populations. Our reference genome comprises 132 autosomal contigs, with a total length of 48.90 Mb. We observed a large complex of ace genes, which has presumably evolved from a long history of organophosphate selection in H. destructor and may contribute towards organophosphate resistance through copy number variation, target-site mutations and structural variants. In the putative ancestral H. destructor ace gene, we identified three target-site mutations (G119S, A201S and F331Y) segregating in organophosphate-resistant populations. Additionally, we identified two new para sodium channel gene mutations (L925I and F1020Y) that may contribute to pyrethroid resistance. Regional structuring observed in population genomic analyses indicates that gene flow in H. destructor does not homogenize populations across large geographic distances. However, our demographic analyses were equivocal on the magnitude of gene flow; the short invasion history of H. destructor makes it difficult to distinguish scenarios of complete isolation vs. ongoing migration. Nonetheless, we identified clear signatures of reduced genetic diversity and smaller inferred effective population sizes in eastern vs. western populations, which is consistent with the stepping-stone invasion pathway of this pest in Australia. These new insights will inform development of diagnostic genetic markers of resistance, further investigation into the multifaceted organophosphate resistance mechanism and predictive modelling of resistance evolution and spread.

Diagnostic yield of chromosomal microarray and trio whole exome sequencing in cryptogenic cerebral palsy.

OBJECTIVE: To determine the yield of genetic diagnoses using chromosomal microarray (CMA) and trio whole exome sequencing (WES), separately and combined, among patients with cryptogenic cerebral palsy (CP). METHODS: Trio WES of patients with prior CMA analysis for cryptogenic CP, defined as disabling, non-progressive motor symptoms beginning before the age of 3 years without known cause. RESULTS: Given both CMA analysis and trio WES, clinically significant genetic findings were identified for 58% of patients (26 of 45). Diagnoses were eight large CNVs detected by CMA and 18 point mutations detected by trio WES. None had more than one severe mutation. Approximately half of events (14 of 26) were de novo. Yield was significantly higher in patients with CP with comorbidities (69%, 22 of 32) than in those with pure motor CP (31%, 4 of 13; p=0.02). Among patients with genetic diagnoses, CNVs were more frequent than point mutations among patients with congenital anomalies (OR 7.8, 95% CI 1.2 to 52.4) or major dysmorphic features (OR 10.5, 95% CI 1.4 to 73.7). Clinically significant mutations were identified in 18 different genes: 14 with known involvement in CP-related disorders and 4 responsible for other neurodevelopmental conditions. Three possible new candidate genes for CP were ARGEF10, RTF1 and TAOK3. CONCLUSIONS: Cryptogenic CP is genetically highly heterogeneous. Genomic analysis has a high yield and is warranted in all these patients. Trio WES has higher yield than CMA, except in patients with congenital anomalies or major dysmorphic features, but these methods are complementary. Patients with negative results with one approach should also be tested by the other.

Genomic analysis of inherited hearing loss in the Palestinian population.

The genetic characterization of a common phenotype for an entire population reveals both the causes of that phenotype for that place and the power of family-based, population-wide genomic analysis for gene and mutation discovery. We characterized the genetics of hearing loss throughout the Palestinian population, enrolling 2,198 participants from 491 families from all parts of the West Bank and Gaza. In Palestinian families with no prior history of hearing loss, we estimate that 56% of hearing loss is genetic and 44% is not genetic. For the great majority (87%) of families with inherited hearing loss, panel-based genomic DNA sequencing, followed by segregation analysis of large kindreds and transcriptional analysis of participant RNA, enabled identification of the causal genes and mutations, including at distant noncoding sites. Genetic heterogeneity of hearing loss was striking with respect to both genes and alleles: The 337 solved families harbored 143 different mutations in 48 different genes. For one in four solved families, a transcription-altering mutation was the responsible allele. Many of these mutations were cryptic, either exonic alterations of splice enhancers or silencers or deeply intronic events. Experimentally calibrated in silico analysis of transcriptional effects yielded inferences of high confidence for effects on splicing even of mutations in genes not expressed in accessible tissue. Most (58%) of all hearing loss in the population was attributable to consanguinity. Given the ongoing decline in consanguineous marriage, inherited hearing loss will likely be much rarer in the next generation.

Molecular diagnosis of childhood immune dysregulation, polyendocrinopathy, and enteropathy, and implications for clinical management.

BACKGROUND: Most patients with childhood-onset immune dysregulation, polyendocrinopathy, and enteropathy have no genetic diagnosis for their illness. These patients may undergo empirical immunosuppressive treatment with highly variable outcomes. OBJECTIVE: We sought to determine the genetic basis of disease in patients referred with Immune dysregulation, polyendocrinopathy, enteropathy, X-linked-like (IPEX-like) disease, but with no mutation in FOXP3; then to assess consequences of genetic diagnoses for clinical management. METHODS: Genomic DNA was sequenced using a panel of 462 genes implicated in inborn errors of immunity. Candidate mutations were characterized by genomic, transcriptional, and (for some) protein analysis. RESULTS: Of 123 patients with FOXP3-negative IPEX-like disease, 48 (39%) carried damaging germline mutations in 1 of the following 27 genes: AIRE, BACH2, BCL11B, CARD11, CARD14, CTLA4, IRF2BP2, ITCH, JAK1, KMT2D, LRBA, MYO5B, NFKB1, NLRC4, POLA1, POMP, RAG1, SH2D1A, SKIV2L, STAT1, STAT3, TNFAIP3, TNFRSF6/FAS, TNRSF13B/TACI, TOM1, TTC37, and XIAP. Many of these genes had not been previously associated with an IPEX-like diagnosis. For 42 of the 48 patients with genetic diagnoses, knowing the critical gene could have altered therapeutic management, including recommendations for targeted treatments and for or against hematopoietic cell transplantation. CONCLUSIONS: Many childhood disorders now bundled as "IPEX-like" disease are caused by individually rare, severe mutations in immune regulation genes. Most genetic diagnoses of these conditions yield clinically actionable findings. Barriers are lack of testing or lack of repeat testing if older technologies failed to provide a diagnosis.

Gut Microbial Perturbation and Host Response Induce Redox Pathway Upregulation along the Gut-Liver Axis during Giardiasis in C57BL/6J Mouse Model.

Apicomplexan infections, such as giardiasis and cryptosporidiosis, negatively impact a considerable proportion of human and commercial livestock populations. Despite this, the molecular mechanisms of disease, particularly the effect on the body beyond the gastrointestinal tract, are still poorly understood. To highlight host-parasite-microbiome biochemical interactions, we utilised integrated metabolomics-16S rRNA genomics and metabolomics-proteomics approaches in a C57BL/6J mouse model of giardiasis and compared these to Cryptosporidium and uropathogenic Escherichia coli (UPEC) infections. Comprehensive samples (faeces, blood, liver, and luminal contents from duodenum, jejunum, ileum, caecum and colon) were collected 10 days post infection and subjected to proteome and metabolome analysis by liquid and gas chromatography-mass spectrometry, respectively. Microbial populations in faeces and luminal washes were examined using 16S rRNA metagenomics. Proteome-metabolome analyses indicated that 12 and 16 key pathways were significantly altered in the gut and liver, respectively, during giardiasis with respect to other infections. Energy pathways including glycolysis and supporting pathways of glyoxylate and dicarboxylate metabolism, and the redox pathway of glutathione metabolism, were upregulated in small intestinal luminal contents and the liver during giardiasis. Metabolomics-16S rRNA genetics integration indicated that populations of three bacterial families-Autopobiaceae (Up), Desulfovibrionaceae (Up), and Akkermanasiaceae (Down)-were most significantly affected across the gut during giardiasis, causing upregulated glycolysis and short-chained fatty acid (SCFA) metabolism. In particular, the perturbed Akkermanasiaceae population seemed to cause oxidative stress responses along the gut-liver axis. Overall, the systems biology approach applied in this study highlighted that the effects of host-parasite-microbiome biochemical interactions extended beyond the gut ecosystem to the gut-liver axis. These findings form the first steps in a comprehensive comparison to ascertain the major molecular and biochemical contributors of host-parasite interactions and contribute towards the development of biomarker discovery and precision health solutions for apicomplexan infections.

Mutational spectrum of breast cancer susceptibility genes among women ascertained in a cancer risk clinic in Northeast Brazil.

PURPOSE: There is a paucity of data on the spectrum and prevalence of pathogenic variants among women of African ancestry in the Northeast region of Brazil. METHODS: We performed BROCA panel sequencing to identify inherited loss-of-function variants in breast cancer susceptibility genes among 292 Brazilian women referred to a single institution cancer risk assessment program. RESULTS: The study included a convenient cohort of 173 women with invasive breast cancer (cases) and 119 women who were cancer-free at the time of ascertainment. The majority of the women self-reported as African-descended (67% for cases and 90.8% for unaffected volunteers). Thirty-seven pathogenic variants were found in 36 (20.8%) patients. While the spectrum of pathogenic variants was heterogeneous, the majority (70.3%) of the pathogenic variants were detected in high-risk genes BRCA1, BRCA2, PALB2, and TP53. Pathogenic variants were also found in the ATM, BARD1, BRIP1, FAM175A, FANCM, NBN, and SLX4 genes in 6.4% of the affected women. Four recurrent pathogenic variants were detected in 11 patients of African ancestry. Only one unaffected woman had a pathogenic variant in the RAD51C gene. Different risk assessment models examined performed well in predicting risk of carrying germline loss-of-function variants in BRCA1 and/or BRCA2 in breast cancer cases. CONCLUSION: The high prevalence and heterogenous spectrum of pathogenic variants identified among self-reported African descendants in Northeast Brazil is consistent with studies in other African ancestry populations with a high burden of aggressive young onset breast cancer. It underscores the need to integrate comprehensive cancer risk assessment and genomic testing in the management of newly diagnosed Black women with breast cancer across the African Diaspora, enabling improved cancer control in admixed underserved and understudied populations.

Text Messaging, Telephone, or In-Person Outpatient Visit to the Surgical Clinic: A Randomized Trial.

INTRODUCTION: Routine outpatient follow-up visits for surgical patients are a source of strain on health-care resources and patients. With the COVID-19 pandemic adding a new urgency to finding the safest follow-up arrangement, text message follow-up might prove an acceptable alternative to a phone call or an in-person clinic visit. METHODS: An open-label, three-arm, parallel randomized trial was conducted. The interventions were traditional in-person appointment, a telephone call, or a text message. The primary outcome was the number of postdischarge complications identified. The secondary outcomes were patient satisfaction with follow-up, future preference, default to follow-up, and preference to receiving medical information by text message. RESULTS: Two hundred eight patients underwent randomization: 50 in the in-person group, 80 in the telephone group, and 78 in the text message group. There was no difference in the number of reported complications: 5 (10%) patients in the in-person group, 7 (9%) patients in the text group, and 11 (14%) patients in the telephone group (P = 0.613). The preferred method of follow-up was by telephone (106, 61.6%). The least preferred was the in-person follow-up (15, 8.7%, P = 0.002), which also had the highest default rate (44%). CONCLUSIONS: There was no evidence that text messages and telephone calls are unsafe and ineffective methods of follow-up. Although most patients are happy to receive results by text message, the majority of patients would prefer a telephone follow-up and are less likely to default by this method. Health-care systems should develop telehealth initiatives when planning health-care services in the wake of the COVID-19 pandemic.

CRISPR-Cas9/long-read sequencing approach to identify cryptic mutations in BRCA1 and other tumour suppressor genes.

Current clinical approaches for mutation discovery are based on short sequence reads (100-300 bp) of exons and flanking splice sites targeted by multigene panels or whole exomes. Short-read sequencing is highly accurate for detection of single nucleotide variants, small indels and simple copy number differences but is of limited use for identifying complex insertions and deletions and other structural rearrangements. We used CRISPR-Cas9 to excise complete BRCA1 and BRCA2 genomic regions from lymphoblast cells of patients with breast cancer, then sequenced these regions with long reads (>10 000 bp) to fully characterise all non-coding regions for structural variation. In a family severely affected with early-onset bilateral breast cancer and with negative (normal) results by gene panel and exome sequencing, we identified an intronic SINE-VNTR-Alu retrotransposon insertion that led to the creation of a pseudoexon in the BRCA1 message and introduced a premature truncation. This combination of CRISPR-Cas9 excision and long-read sequencing reveals a class of complex, damaging and otherwise cryptic mutations that may be particularly frequent in tumour suppressor genes replete with intronic repeats.

Characterization of splice-altering mutations in inherited predisposition to cancer.

Mutations responsible for inherited disease may act by disrupting normal transcriptional splicing. Such mutations can be difficult to detect, and their effects difficult to characterize, because many lie deep within exons or introns where they may alter splice enhancers or silencers or introduce new splice acceptors or donors. Multiple mutation-specific and genome-wide approaches have been developed to evaluate these classes of mutations. We introduce a complementary experimental approach, cBROCA, which yields qualitative and quantitative assessments of the effects of genomic mutations on transcriptional splicing of tumor suppressor genes. cBROCA analysis is undertaken by deriving complementary DNA (cDNA) from puromycin-treated patient lymphoblasts, hybridizing the cDNA to the BROCA panel of tumor suppressor genes, and then multiplex sequencing to very high coverage. At each splice junction suggested by split sequencing reads, read depths of test and control samples are compared. Significant Z scores indicate altered transcripts, over and above naturally occurring minor transcripts, and comparisons of read depths indicate relative abundances of mutant and normal transcripts. BROCA analysis of genomic DNA suggested 120 rare mutations from 150 families with cancers of the breast, ovary, uterus, or colon, in >600 informative genotyped relatives. cBROCA analysis of their transcripts revealed a wide variety of consequences of abnormal splicing in tumor suppressor genes, including whole or partial exon skipping, exonification of intronic sequence, loss or gain of exonic and intronic splicing enhancers and silencers, complete intron retention, hypomorphic alleles, and combinations of these alterations. Combined with pedigree analysis, cBROCA sequencing contributes to understanding the clinical consequences of rare inherited mutations.

Inherited predisposition to malignant mesothelioma and overall survival following platinum chemotherapy.

Survival from malignant mesothelioma, particularly pleural mesothelioma, is very poor. For patients with breast, ovarian, or prostate cancers, overall survival is associated with increased sensitivity to platinum chemotherapy due to loss-of-function mutations in DNA repair genes. The goal of this project was to evaluate, in patients with malignant mesothelioma, the relationship between inherited loss-of-function mutations in DNA repair and other tumor suppressor genes and overall survival following platinum chemotherapy. Patients with histologically confirmed malignant mesothelioma were evaluated for inherited mutations in tumor suppressor genes. Survival was evaluated with respect to genotype and site of mesothelioma. Among 385 patients treated with platinum chemotherapy, median overall survival was significantly longer for patients with loss-of-function mutations in any of the targeted genes compared with patients with no such mutation (P = 0.0006). The effect of genotype was highly significant for patients with pleural mesothelioma (median survival 7.9 y versus 2.4 y, P = 0.0012), but not for patients with peritoneal mesothelioma (median survival 8.2 y versus 5.4 y, P = 0.47). Effect of patient genotype on overall survival, measured at 3 y, remained independently significant after adjusting for gender and age at diagnosis, two other known prognostic factors. Patients with pleural mesothelioma with inherited mutations in DNA repair and other tumor suppressor genes appear to particularly benefit from platinum chemotherapy compared with patients without inherited mutations. These patients may also benefit from other DNA repair targeted therapies such as poly-ADP ribose polymerase (PARP) inhibitors.

Genomics and transcriptomics yields a system-level view of the biology of the pathogen Naegleria fowleri.

BACKGROUND: The opportunistic pathogen Naegleria fowleri establishes infection in the human brain, killing almost invariably within 2 weeks. The amoeba performs piece-meal ingestion, or trogocytosis, of brain material causing direct tissue damage and massive inflammation. The cellular basis distinguishing N. fowleri from other Naegleria species, which are all non-pathogenic, is not known. Yet, with the geographic range of N. fowleri advancing, potentially due to climate change, understanding how this pathogen invades and kills is both important and timely. RESULTS: Here, we report an -omics approach to understanding N. fowleri biology and infection at the system level. We sequenced two new strains of N. fowleri and performed a transcriptomic analysis of low- versus high-pathogenicity N. fowleri cultured in a mouse infection model. Comparative analysis provides an in-depth assessment of encoded protein complement between strains, finding high conservation. Molecular evolutionary analyses of multiple diverse cellular systems demonstrate that the N. fowleri genome encodes a similarly complete cellular repertoire to that found in free-living N. gruberi. From transcriptomics, neither stress responses nor traits conferred from lateral gene transfer are suggested as critical for pathogenicity. By contrast, cellular systems such as proteases, lysosomal machinery, and motility, together with metabolic reprogramming and novel N. fowleri proteins, are all implicated in facilitating pathogenicity within the host. Upregulation in mouse-passaged N. fowleri of genes associated with glutamate metabolism and ammonia transport suggests adaptation to available carbon sources in the central nervous system. CONCLUSIONS: In-depth analysis of Naegleria genomes and transcriptomes provides a model of cellular systems involved in opportunistic pathogenicity, uncovering new angles to understanding the biology of a rare but highly fatal pathogen.

User Evaluation of a Novel Smart Insufflator for Laparoscopic Surgery-The EVA-15.

INTRODUCTION: There is resurging interest in the importance of effective, nuanced insufflation and personalised pneumoperitoneal pressure-management during laparoscopy. Here, we present user-evaluation data from a regulated, prospective, multispecialty study of a new insufflator (EVA-15, Palliare, Galway, Ireland) which provides high-frequency pressure-sensing, built-in smoke evacuation with pedal activation and highly responsive, high-flow gas provision. METHODS: With institutional ethics and regulatory body approval, a non-randomised, prospective clinical investigation was performed on 30 subjects undergoing laparoscopic surgery using an EVA-15 device. Cases were selected from a variety of specialties on a near-consecutive basis without specific exclusion criteria. Users (both surgeons and operating room nurses) completed a survey at case completion to capture ordinal categorical data on a 5-point Likert agreement scale (1 - Strongly disagree to 5 - Strongly agree) concerning (i) Settings and Setup Evaluations, (ii) Alarms and Displays Evaluations, (iii) Short Instruction Guide, and (iv) Insufflator Performance along with any additional feedback. RESULTS: Operations on 30 patients (mean age 54 y, 15 males) were studied with a questionnaire completed by operating room teams after individual consent. The procedures included general (n=13), upper (n=3) and lower (n=6) gastrointestinal surgery, bariatric (n=3), hepatobiliary (n=2) urology (n=2, both robotic prostatectomy) and gynaecology (n=1) operations. In all cases, the laparoscopic component was completed capably with the use of the EVA-15 device. The insufflator evaluation score across all categories was a median of 4, demonstrating satisfactory use and performance in all regards. CONCLUSION: The EVA-15 is a smart insufflator system that is capable of satisfactory performance across a spectrum of cases among different specialties.

Adaptive Introgression across Semipermeable Species Boundaries between Local Helicoverpa zea and Invasive Helicoverpa armigera Moths.

Hybridization between invasive and native species has raised global concern, given the dramatic increase in species range shifts and pest outbreaks due to anthropogenic dispersal. Nevertheless, secondary contact between sister lineages of local and invasive species provides a natural laboratory to understand the factors that determine introgression and the maintenance or loss of species barriers. Here, we characterize the early evolutionary outcomes following secondary contact between invasive Helicoverpa armigera and native H. zea in Brazil. We carried out whole-genome resequencing of Helicoverpa moths from Brazil in two temporal samples: during the outbreak of H. armigera in 2013 and 2017. There is evidence for a burst of hybridization and widespread introgression from local H. zea into invasive H. armigera coinciding with H. armigera expansion in 2013. However, in H. armigera, the admixture proportion and the length of introgressed blocks were significantly reduced between 2013 and 2017, suggesting selection against admixture. In contrast to the genome-wide pattern, there was striking evidence for adaptive introgression of a single region from the invasive H. armigera into local H. zea, including an insecticide resistance allele that increased in frequency over time. In summary, despite extensive gene flow after secondary contact, the species boundaries are largely maintained except for the single introgressed region containing the insecticide-resistant locus. We document the worst-case scenario for an invasive species, in which there are now two pest species instead of one, and the native species has acquired resistance to pyrethroid insecticides through introgression.