Mapping three-dimensional intratumor proteomic heterogeneity in uterine serous carcinoma by multiregion microsampling.

BACKGROUND: Although uterine serous carcinoma (USC) represents a small proportion of all uterine cancer cases, patients with this aggressive subtype typically have high rates of chemotherapy resistance and disease recurrence that collectively result in a disproportionately high death rate. The goal of this study was to provide a deeper view of the tumor microenvironment of this poorly characterized uterine cancer variant through multi-region microsampling and quantitative proteomics. METHODS: Tumor epithelium, tumor-involved stroma, and whole "bulk" tissue were harvested by laser microdissection (LMD) from spatially resolved levels from nine USC patient tumor specimens and underwent proteomic analysis by mass spectrometry and reverse phase protein arrays, as well as transcriptomic analysis by RNA-sequencing for one patient's tumor. RESULTS: LMD enriched cell subpopulations demonstrated varying degrees of relatedness, indicating substantial intratumor heterogeneity emphasizing the necessity for enrichment of cellular subpopulations prior to molecular analysis. Known prognostic biomarkers were quantified with stable levels in both LMD enriched tumor and stroma, which were shown to be highly variable in bulk tissue. These USC data were further used in a comparative analysis with a data generated from another serous gynecologic malignancy, high grade serous ovarian carcinoma, and have been added to our publicly available data analysis tool, the Heterogeneity Analysis Portal ( https://lmdomics.org/ ). CONCLUSIONS: Here we identified extensive three-dimensional heterogeneity within the USC tumor microenvironment, with disease-relevant biomarkers present in both the tumor and the stroma. These data underscore the critical need for upfront enrichment of cellular subpopulations from tissue specimens for spatial proteogenomic analysis.

Host-induced RNA interference targeting the neuromotor gene FMRFamide-like peptide-14 (Mi-flp14) perturbs Meloidogyne incognita parasitic success in eggplant.

KEY MESSAGE: The study demonstrates the successful management of Meloidogyne incognita in eggplant using Mi-flp14 RNA interference, showing reduced nematode penetration and reproduction without off-target effects across multiple generations. Root-knot nematode, Meloidogyne incognita, causes huge yield losses worldwide. Neuromotor function in M. incognita governed by 19 neuropeptides is vital for parasitism and parasite biology. The present study establishes the utility of Mi-flp14 for managing M. incognita in eggplant in continuation of our earlier proof of concept in tobacco (US patent US2015/0361445A1). Mi-flp14 hairpin RNA construct was used for generating 19 independent transgenic eggplant events. PCR and Southern hybridization analysis confirmed transgene integration and its orientation, while RT-qPCR and Northern hybridization established the generation of dsRNA and siRNA of Mi-flp14. In vitro and in vivo bio-efficacy analysis of single-copy events against M. incognita showed reduced nematode penetration and development at various intervals that negatively impacted reproduction. Interestingly, M. incognita preferred wild-type plants over the transgenics even when unbiased equal opportunity was provided for the infection. A significant reduction in disease parameters was observed in transgenic plants viz., galls (40-48%), females (40-50%), egg masses (35-40%), eggs/egg mass (50-55%), and derived multiplication factor (60-65%) compared to wild type. A unique demonstration of perturbed expression of Mi-flp14 in partially penetrated juveniles and female nematodes established successful host-mediated RNAi both at the time of penetration even before the nematodes started withdrawing plant nutrients and later stage, respectively. The absence of off-target effects in transgenic plants was supported by the normal growth phenotype of the plants and T-DNA integration loci. Stability in the bio-efficacy against M. incognita across T1- to T4-generation transgenic plants established the utility of silencing Mi-flp14 for nematode management. This study demonstrates the significance of targeting Mi-flp14 in eggplant for nematode management, particularly to address global agricultural challenges posed by M. incognita.

Dynamics of diurnal cortisol and alpha-amylase secretion and their associations with PTSD onset in recent interpersonal trauma survivors.

BACKGROUND: Dysfunction in major stress response systems during the acute aftermath of trauma may contribute to risk for developing posttraumatic stress disorder (PTSD). The current study investigated how PTSD diagnosis and symptom severity, depressive symptoms, and childhood trauma uniquely relate to diurnal neuroendocrine secretion (cortisol and alpha-amylase rhythms) in women who recently experienced interpersonal trauma compared to non-traumatized controls (NTCs). METHOD: Using a longitudinal design, we examined diurnal cortisol and alpha-amylase rhythms in 98 young women (n = 57 exposed to recent interpersonal trauma, n = 41 NTCs). Participants provided saliva samples and completed symptom measures at baseline and 1-, 3-, and 6-month follow-up. RESULTS: Multilevel models (MLMs) revealed lower waking cortisol predicted the development of PTSD in trauma survivors and distinguished at-risk women from NTCs. Women with greater childhood trauma exposure exhibited flatter diurnal cortisol slopes. Among trauma-exposed individuals, lower waking cortisol levels were associated with higher concurrent PTSD symptom severity. Regarding alpha-amylase, MLMs revealed women with greater childhood trauma exposure exhibited higher waking alpha-amylase and slower diurnal alpha-amylase increase. CONCLUSIONS: Results suggest lower waking cortisol in the acute aftermath of trauma may be implicated in PTSD onset and maintenance. Findings also suggest childhood trauma may predict a different pattern of dysfunction in stress response systems following subsequent trauma exposure than the stress system dynamics associated with PTSD risk; childhood trauma appears to be associated with flattened diurnal cortisol and alpha-amylase slopes, as well as higher waking alpha-amylase.

Patient-reported tolerability of adjuvant ipilimumab (3 or 10 mg/kg) versus high-dose interferon alfa-2b for resected high-risk stage III-IV melanoma in phase III trial E1609.

PURPOSE: Trial E1609 demonstrated superior overall survival with ipilimumab 3 mg/kg (ipi3) compared to high-dose interferon (HDI) for patients with resected high-risk melanoma. To inform treatment tolerability, we compared health-related quality of life (HRQoL), gastrointestinal (GI), and treatment-specific physical and cognitive/emotional symptoms. We also compared treatment-specific concerns between all arms. METHODS: We assessed HRQoL using the Functional Assessment of Cancer Therapy-General, physical and cognitive/emotional concerns using the FACT-Biologic Response Modifier subscale, and GI symptoms with the Functional Assessment of Chronic Illness Therapy-Diarrhea subscale pre-treatment and every 3 months. The primary outcome was the difference in HRQoL at 3 months between ipi3/ipi10 vs. HDI. RESULTS: 549 patients (n = 158 ipi3; n = 191 ipi10; n = 200 HDI) were analyzed. 3-month completion was 58.7%. Compared to HDI, ipilimumab patients reported better HRQoL (ipi3 = 87.5 ± 14.6 vs. HDI = 74.7 ± 15.4, p < .001; ipi10 = 84.9 ± 16.5 vs. HDI, p < .001) and fewer physical (ipi3 = 22.3 ± 4.6 vs. HDI = 17.1 ± 5.4, p < .001; ipi10 = 21.8 ± 5.0 vs. HDI p < .001) and cognitive/emotional (ipi3 = 18.6 ± 4.4 vs. HDI = 15.0 ± 5.3, p < .001; ipi10 = 17.7 ± 4.8 vs. HDI p < .001) concerns, but worse GI symptoms (ipi3 = 40.8 ± 5.0 vs. HDI = 42.2 ± 2.9, p = .011; ipi10 = 39.5 ± 7.0 vs. HDI, p < .001). Fewer ipilimumab patients reported worsening treatment-specific concerns (e.g., 52% of ipi3 and 58% of ipi10 reported worsening fatigue vs. 82% HDI, p's < .001). CONCLUSION: PROs demonstrated less toxicity of ipi3 compared to HDI and ipi10. Priorities for symptom management among patients receiving ipilimumab include GI toxicities, fatigue, weakness, appetite loss, arthralgia, and depression. TRIAL REGISTRATION: NCT01274338, January 11, 2011 (first posted date) https://clinicaltrials.gov/ct2/show/NCT01274338?term=NCT01274338&draw=2&rank=1 .

Host-delivered RNAi-mediated silencing using fusion cassettes of different functional groups of genes precludes Meloidogyne incognita multiplication in Nicotiana tabacum.

KEY MESSAGE: This study demonstrates multi-gene silencing approach for simultaneous silencing of several functional genes through a fusion gene strategy for protecting plants against root-knot nematode, Meloidogyne incognita. The ability of root-knot nematode (RKN), Meloidogyne incognita, to cause extensive yield decline in a wide range of cultivated crops is well-documented. Due to the inadequacies of current management approaches, the alternatively employed contemporary RNA interference (RNAi)-based host-delivered gene silencing (HD-RNAi) strategy targeting different functional effectors/genes has shown substantial potential to combat RKNs. In this direction, we have explored the possibility of simultaneous silencing of four esophageal gland genes, six plant cell-wall modifying enzymes (PCWMEs) and a serine protease gene of M. incognita using the fusion approach. In vitro RNAi showed that combinatorial gene silencing is the most effective in affecting nematode behavior in terms of reduced attraction, penetration, development, and reproduction in tomato and adzuki beans. In addition, qRT-PCR analysis of M. incognita J2s soaked in fusion-dsRNA showed perturbed expression of all the genes comprising the fusion construct confirming successful dsRNA processing which is also supported by increased mRNA abundance of five key-RNAi pathway genes. In addition, hairpin RNA expressing constructs of multi-gene fusion cassettes were developed and used for generation of Nicotiana tabacum transgenic plants. The integration of gene constructs and expression of siRNAs in transgenic events were confirmed by Southern and Northern blot analyses. Besides, bio-efficacy analyses of transgenic events, conferred up to 87% reduction in M. incognita multiplication. Correspondingly, reduced transcript accumulation of the target genes in the M. incognita females extracted from transgenic events confirmed successful gene silencing.

Adversity type and timing predict temporal summation of pain in African-American adults.

African Americans are disproportionately exposed to adversity across the lifespan, which includes both stressful and traumatic events. Adversity, in turn, is associated with alterations in pain responsiveness. Racial differences in pain responsiveness among healthy adults are well established. However, the extent to which adversity type and timing are associated with alterations in pain responsiveness among healthy African-American adults is not well understood. The present study included 160 healthy African-American adults (98 women), ages 18 to 45. Outcome measures included pain tolerance and temporal summation of pain to evoked thermal pain. Composite scores were created for early-life adversity (childhood trauma, family adversity) and recent adversity (perceived stress, chronic stress burden). A measure of lifetime racial discrimination was also included. Higher levels of recent adversity were associated with higher temporal summation of pain, controlling for gender, age, and education. Neither early-life adversity nor lifetime racial discrimination were associated with temporal summation of pain. The present findings suggest that heightened temporal summation of pain among healthy African-American adults is associated with exposure to recent adversity events. Improved understanding of how recent adversity contributes to heightened temporal summation of pain in African Americans could help to mitigate racial disparities in pain experiences by identifying at-risk individuals who could benefit from early interventions.

Differential effects of rhizobacteria from uninfected and infected tomato on Meloidogyne incognita under protected cultivation.

Intermingled uninfected and root-knot nematode-infected tomato plants are commonly observed under protected cultivation. To understand the role of rhizobacteria underlying the susceptibility to nematode infectivity in these tomato plants, 36 rhizobacteria (18 from each type) with morphologically distinct colony characteristics were isolated from the rhizosphere of uninfected and root-knot nematode-infected tomato plants. The in vitro nematicidal potential of rhizobacteria from the uninfected rhizosphere was significantly higher than that from the infested rhizosphere. The three most effective antagonists were identified as Microbacterium laevaniformans, Staphylococcus kloosii, Priestia aryabhattai from root-knot-nematode-infected tomato rhizosphere and Staphylococcus sciuri, Bacillus pumilus, and Priestia megaterium from the rhizosphere of uninfected tomato. Volatile organic compounds from these rhizobacteria were characterized. Except for S. kloosi, the soil drenching with other rhizobacteria significantly reduced juvenile penetration (>60%) in tomato roots. Furthermore, the application of a single or consortium of these rhizobacteria affected nematode reproduction in tomato. Four consortia of rhizobacteria (S. sciuri + B. pumilus + P. megaterium), (B. pumilus + P. megaterium), (S. sciuri + B. pumilus), and (S. sciuri + P. megaterium) from uninfested rhizosphere and two consortia (M. laevaniformans + P. aryabhattai), (M. laevaniformans + S. kloosii + P. aryabhattai) from infested rhizosphere (IRh) effectively reduced M. incognita reproduction and considerably enhanced plant growth and yield in tomato. The nematicidal efficacy, however, decreased when S. kloosii was applied in the consortium. These distinctive effects illustrate how the plant susceptibility to nematode infectivity is modulated under natural conditions.

Stringent in-silico identification of putative G-protein-coupled receptors (GPCRs) of the entomopathogenic nematode Heterorhabditis bacteriophora.

The infective juveniles (IJs) of entomopathogenic nematode (EPN) Heterorhabditis bacteriophora find and infect their host insects in heterogeneous soil ecosystems by sensing a universal host cue (CO2) or insect/plant-derived odorants, which bind to various sensory receptors, including G protein-coupled receptors (GPCRs). Nematode chemosensory GPCRs (NemChRs) bind to a diverse set of ligands, including odor molecules. However, there is a lack of information on the NemChRs in EPNs. Here we identified 21 GPCRs in the H. bacteriophora genome sequence in a triphasic manner, combining various transmembrane detectors and GPCR predictors based on different algorithms, and considering inherent properties of GPCRs. The pipeline was validated by reciprocal BLAST, InterProscan, GPCR-CA, and NCBI CDD search. Functional classification of predicted GPCRs using Pfam revealed the presence of four NemChRs. Additionally, GPCRs were classified into various families based on the reciprocal BLAST approach into a frizzled type, a secretin type, and 19 rhodopsin types of GPCRs. Gi/o is the most abundant kind of G-protein, having a coupling specificity to all the fetched GPCRs. As the 21 GPCRs identified are expected to play a crucial role in the host-seeking behavior, these might be targeted to develop novel insect-pest management strategies by tweaking EPN IJ behavior, or to design novel anthelminthic drugs. Our new and stringent GPCR detection pipeline may also be used to identify GPCRs from the genome sequence of other organisms.

RNA-Sequencing of Heterorhabditis nematodes to identify factors involved in symbiosis with Photorhabdus bacteria.

BACKGROUND: Nematodes are a major group of soil inhabiting organisms. Heterorhabditis nematodes are insect-pathogenic nematodes and live in a close symbiotic association with Photorhabdus bacteria. Heterorhabditis-Photorhabdus pair offers a powerful and genetically tractable model to study animal-microbe symbiosis. It is possible to generate symbiont bacteria free (axenic) stages in Heterorhabditis. Here, we compared the transcriptome of symbiotic early-adult stage Heterorhabditis nematodes with axenic early-adult nematodes to determine the nematode genes and pathways involved in symbiosis with Photorhabdus bacteria. RESULTS: A de-novo reference transcriptome assembly of 95.7 Mb was created for H. bacteriophora by using all the reads. The assembly contained 46,599 transcripts with N50 value of 2,681 bp and the average transcript length was 2,054 bp. The differentially expressed transcripts were identified by mapping reads from symbiotic and axenic nematodes to the reference assembly. A total of 754 differentially expressed transcripts were identified in symbiotic nematodes as compared to the axenic nematodes. The ribosomal pathway was identified as the most affected among the differentially expressed transcripts. Additionally, 12,151 transcripts were unique to symbiotic nematodes. Endocytosis, cAMP signalling and focal adhesion were the top three enriched pathways in symbiotic nematodes, while a large number of transcripts coding for various responses against bacteria, such as bacterial recognition, canonical immune signalling pathways, and antimicrobial effectors could also be identified. CONCLUSIONS: The symbiotic Heterorhabditis nematodes respond to the presence of symbiotic bacteria by expressing various transcripts involved in a multi-layered immune response which might represent non-systemic and evolved localized responses to maintain mutualistic bacteria at non-threatening levels. Subject to further functional validation of the identified transcripts, our findings suggest that Heterorhabditis nematode immune system plays a critical role in maintenance of symbiosis with Photorhabdus bacteria.

A novel tasi RNA-based micro RNA-induced gene silencing strategy to tackle multiple pests and pathogens in cotton (Gossypium hirsutum L.).

MAIN CONCLUSION: This study demonstrates the combinatorial management of multiple pests through a trans-acting siRNA (tasiRNA)-based micro RNA-induced gene silencing (MIGS) strategy. Transgenic cotton events demonstrated improved efficacy against cotton leaf curl disease, cotton leaf hopper and root-knot nematode. Cotton (Gossypium hirsutum L.), an important commercial crop grown worldwide is confronted by several pests and pathogens, thus reiterating interventions for their management. In this study, we report, the utility of a novel Arabidopsis miRNA173-directed trans-acting siRNA (tasiRNA)-based micro RNA-induced gene silencing (MIGS) strategy for the simultaneous management of cotton leaf curl disease (CLCuD), cotton leaf hopper (CLH; Amrasca biguttula biguttula) and root-knot nematode (RKN, Meloidogyne incognita). Cotton transgenics were developed with the MIGS construct targeting a total of 7 genes by an apical meristem-targeted in planta transformation strategy. Stable transgenics were selected using stringent selection pressure, molecular characterization and stress-specific bio-efficacy studies. We identified 8 superior events with 50-100% resistance against CLCuD, while reduction in the root-knot nematode multiplication factor in the range of 35-75% confirmed resistance to RKN. These transgenic cotton events were also detrimental to the growth and development of CLH, as only 43.3-62.5% of nymphs could survive. Based on the corroborating evidences obtained by all the bioefficacy analyses, 3 events viz., L-75-1, E-27-11, E-27-7 were found to be consistent in tackling the target pests. To the best of our knowledge, this report is the first of its kind demonstrating the possibility of combinatorial management of pests/diseases in cotton using MIGS approach. These identified events demonstrate immense utility of the strategy towards combinatorial stress management in cotton improvement programs.

Negative affect, childhood adversity, and adolescents' eating following stress.

Obesity commonly emerges by adolescence and is associated with serious health consequences. Emotional eating (consuming calories, fats, and sugars in response to negative affect) may promote obesity; however, evidence is mixed as to whether negative affect increases obesogenic eating. Early-life adversity may shape malleable neurobiological systems that govern inhibitory control, physiological regulation, coping strategies, and eating behavior, contributing to greater obesogenic eating in response to negative affect. Therefore, this study tested whether childhood adversity moderates the association between negative affect and food consumption in a diverse sample of female adolescents. After completing a childhood adversity assessment, 157 female adolescents (13-17 years; 28.7% African American, 39.5% Hispanic/Latina, 31.8% Non-Hispanic White) rated their negative affect in response to a standard social stress paradigm before consuming a buffet lunch, which was evaluated for calories, added sugars, and solid fats consumed. Results did not support that negative affect exerted a main effect on eating behavior. However, negative affect and childhood adversity interacted to predict calories and solid fats consumed, such that negative affect was associated with more obesogenic eating for those with high adversity exposure but not for those with low adversity exposure. Adversity and affect did not interact to predict added sugars consumed. Findings support that eating patterns in response to negative affect may differ by childhood adversity history. Reducing children's adversity exposure and bolstering emotion regulation techniques for adolescents who have been exposed to adversity may provide pathways to protect health and well-being by reducing maladaptive eating patterns.

Amenability of Maruca vitrata (Lepidoptera: Crambidae) to gene silencing through exogenous administration and host-delivered dsRNA in pigeonpea (Cajanus cajan L.).

Insect pests are one of the major biotic stresses limiting yield in commercially important crops. The lepidopteran polyphagous spotted pod borer, Maruca vitrata causes significant economic losses in legumes including pigeonpea. RNA interference (RNAi)-based gene silencing has emerged as one of the potential biotechnological tools for crop improvement. We report in this paper, RNAi in M. vitrata through exogenous administration of dsRNA with sequence specificity to three functionally important genes, Alpha-amylase (α-amylase), Chymotrypsin-like serine protease (CTLP) and Tropomyosin (TPM) into the larval haemolymph and their host-delivered RNAi in pigeonpea. Significant decline in the expression of selected genes supported by over-expression of DICER and generation of siRNA indicated the occurrence of RNAi in the dsRNA-injected larvae. Additionally, the onset of RNAi in the herbivore was demonstrated in pigeonpea, one of the prominent hosts, by host-delivered dsRNA. Transgenics in pigeonpea (cv. Pusa 992), a highly recalcitrant crop, were developed through a shoot apical meristem-targeted in planta transformation strategy and evaluated. Plant level bioassays in transgenic events characterized and selected at molecular level showed mortality of M. vitrata larvae as well as reduced feeding when compared to wild-type. Furthermore, molecular evidence for down regulation of target genes in the insects that fed on transgenic plants authenticated RNAi. Considering the variability of gene silencing in lepidopteran pests, this study provided corroborative proof for the possibility of gene silencing in M. vitrata through both the strategies. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s12298-022-01133-3.

A rice root-knot nematode Meloidogyne graminicola-resistant mutant rice line shows early expression of plant-defence genes.

MAIN CONCLUSION: Resistance to rice root-knot nematode Meloidogyne graminicola in a mutant rice line is suggested to be conferred by higher expression of several genes putatively involved in damage-associated molecular pattern recognition, secondary metabolite biosynthesis including phytoalexins, and defence-related genes. Meloidogyne graminicola has emerged as the most destructive plant-parasitic nematode disease of rice (Oryza sativa L.). Genetic resistance to M. graminicola is one of the most effective methods for its management. A M. graminicola-resistant O. sativa ssp. indica mutant line-9 was previously identified through a forward genetic screen (Hatzade et al. Biologia 74:1197-1217, 2019). In the present study, we used RNA-Sequencing to investigate the molecular mechanisms conferring nematode resistance to the mutant line-9 compared to the susceptible parent JBT 36/14 at 24 h post-infection. A total of 674 transcripts were differentially expressed in line-9. Early regulation of genes putatively related to nematode damage-associated molecular pattern recognition (e.g., wall-associated receptor kinases), signalling [Nucleotide-binding, Leucine-Rich Repeat (NLRs)], pathogenesis-related (PR) genes (PR1, PR10a), defence-related genes (NB-ARC domain-containing genes), as well as a large number of genes involved in secondary metabolites including diterpenoid biosynthesis (CPS2, OsKSL4, OsKSL10, Oscyp71Z2, oryzalexin synthase, and momilactone A synthase) was observed in M. graminicola-resistant mutant line-9. It may be suggested that after the nematode juveniles penetrate the roots of line-9, early recognition of invading nematodes triggers plant immune responses mediated by phytoalexins, and other defence proteins such as PR proteins inhibit nematode growth and reproduction. Our study provides the first transcriptomic comparison of nematode-resistant and susceptible rice plants in the same genetic background and adds to the understanding of mechanisms underlying plant-nematode resistance in rice.

Host-mediated RNAi for simultaneous silencing of different functional groups of genes in Meloidogyne incognita using fusion cassettes in Nicotiana tabacum.

KEY MESSAGE: This study establishes possibility of combinatorial silencing of more than one functional gene for their efficacy against root-knot nematode, M. incognita. Root-knot nematodes (RKN) of the genus Meloidogyne are the key important plant parasitic nematodes (PPNs) in agricultural and horticultural crops worldwide. Among RKNs, M. incognita is the most notorious that demand exploration of novel strategies for their management. Due to its sustainable and target-specific nature, RNA interference (RNAi) has gained unprecedented importance to combat RKNs. However, based on the available genomic information and interaction studies, it can be presumed that RKNs are dynamic and not dependent on single genes for accomplishing a particular function. Therefore, it becomes extremely important to consider silencing of more than one gene to establish any synergistic or additive effect on nematode parasitism. In this direction, we have combined three effectors specific to subventral gland cells of M. incognita, Mi-msp1, Mi-msp16, Mi-msp20 as fusion cassettes-1 and two FMRFamide-like peptides, Mi-flp14, Mi-flp18, and Mi-msp20 as fusion cassettes-2 to establish their possible utility for M. incognita management. In vitro RNAi assay in tomato and adzuki bean using these two fusion gene negatively altered nematode behavior in terms of reduced attraction, invasion, development, and reproduction. Subsequently, Nicotiana tabacum plants were transformed with these two fusion gene hairpin RNA-expressing vectors (hpRNA), and characterized via PCR, qRT-PCR, and Southern blot hybridization. Production of siRNAs specific to Mi-flp18 and Mi-msp1 was also confirmed by Northern hybridization. Further, transgenic events expressing single copy insertions of hpRNA constructs of fusion 1 and fusion-2 conferred up to 85% reduction in M. incognita multiplication. Besides, expression quantification revealed a significant reduction in mRNA abundance of target genes (up to 1.8-fold) in M. incognita females extracted from transgenic plants, and provided additional evidence for successful gene silencing.

Nematode genome announcement: The draft genome sequence of entomopathogenic nematode Heterorhabditis indica.

Heterorhabditis indica is one of the most widely used entomopathogenic nematodes for the biological control of agricultural insect pests worldwide. The draft genome of H. indica was sequenced using three genomic libraries of 300 bp, 600 bp and 5 kb sizes by Illumina HiSeq platform. The size of the draft genome assembly was 91.26 Mb, comprising 3,538 scaffolds. Genome completeness analysis by BUSCO (Benchmarking Universal Single-Copy Orthologs) showed 84% complete, and 6.5% fragmented BUSCOs. Further, 10,494 protein-coding genes were predicted. The H. indica draft genome will enable comparative and functional genomic studies in Heterorhabditis nematodes.

Effect of fluensulfone on different functional genes of root-knot nematode Meloidogyne incognita.

Meloidogyne incognita is an obligate plant-parasitic nematode causing serious damage to agricultural crops. Major constraints in nematode management arose due to the limited availability of non-fumigant nematicides in conjunction with the considerable ill effects of fumigants on human and non-target organisms. Recently, fluensulfone has been reported to be an effective non-fumigant nematicide against plant-parasitic nematodes and the model nematode Caenorhabditis elegans. The nematicidal efficacy varies according to its concentration at the time of application, exposure timing, nematode species variability, and even across subpopulations within the same species. It interferes with the key physiological processes of nematodes, like motility, behavior, chemosensation, stylet thrusting, infectivity, metabolism, lipid consumption, tissue integrity, oviposition, egg hatching, and survival. However, the molecular basis of these multivariate physiological anomalies is still largely unknown. Quantitative real-time PCR was carried out to understand the acute transcriptional perturbation of 30 functional genes associated with key physiological and life processes in a M. incognita population, following exposure of 10, 50, and 100 ppm of fluensulfone for 5 and 10 hr. The chemical treatment resulted in significant downregulation of all the neuropeptidergic genes, with concomitant repression of majority of genes related to chemosensation, esophageal gland secretion, parasitism, fatty acid metabolism, and G-protein coupled receptors. Collectively, the parasitism genes were found to be perturbed at highest magnitude, followed by the GPCRs and neuropeptidergic genes. These results establish the wide ranging effect of fluensulfone on various metabolic and physiological pathways of nematode.

Impact of adjuvant treatment and prognostic factors in stage I uterine leiomyosarcoma patients treated in Commission on Cancer®-accredited facilities.

OBJECTIVES: Determine the impact of adjuvant chemotherapy (ACT) and prognostic factors in surgically managed patients with stage I uterine leiomyosarcoma (ULMS). METHODS: Women who underwent hysterectomy and were diagnosed with stage I ULMS between 2010 and 2014 in the National Cancer Database were eligible for this observation study. Inverse probability of treatment weighting based on propensity score was used to balance clinical characteristics between ACT and no ACT patients. Hazard ratio (HR) and 95% confidence interval (CI) were estimated from Cox modeling. RESULTS: There were 1059 eligible patients with stage I ULMS including 514 treated with ACT and 545 with no ACT. Patient characteristics and tumor features varied in patients treated with ACT vs. no ACT (P < .0001). Multivariate survival analysis demonstrated that patient age, comorbidity score, tumor size, lymphovascular space invasion (LVSI) and grade were independent prognostic factors. After propensity score weighting to control for imbalance of prognostic clinical factors, adjusted five-year survival was 61.7% vs. 61.3% and restricted mean survival time was 39.7 vs. 40.6 months for ACT vs. no ACT, respectively. Risk of death in a weighted Cox analysis of overall survival was similar (HR = 1.08, 95% CI = 0.85-1.37, P = .054) for ACT vs. no ACT patients. Subset analysis demonstrated that survival was similar in ACT vs. no ACT patients categorized by age, tumor size and LVSI or with high grade or ungraded tumors. In contrast, patients with low grade tumors had worse 5-year survival (82.3% vs. 91.5%) and an increased risk of death (HR = 3.79, 95% CI = 1.15-12.40, P = .028) following ACT vs. no ACT. CONCLUSIONS: ACT did not improve survival over no ACT in patients with stage I ULMS and was inferior in patients with low grade tumors.

Novel biotechnological strategies to combat biotic stresses: polygalacturonase inhibitor (PGIP) proteins as a promising comprehensive option.

Global climate change and combinatorial environmental stresses pose grave challenges to food security and agricultural sustainability. This calls for diverse and futuristic approaches for the development of crops with increased resilience to natural vagaries. Though innumerable strategies involving diverse genes/pathways are being deciphered in plants to aid stress mitigation, the hunt is still on. Furthermore, strategies that work to alleviate a combination of stresses are always pertinent. In this review, we discuss polygalacturonase inhibitor (PGIP) proteins as a plausible option to mitigate multiple biotic stresses. These are ubiquitous cell wall proteins that inhibit the pectin-depolymerizing activity of cell wall loosening enzymes, polygalacturonases (PGs). While plant PGs are those responsible for developmental activities like fruit ripening, pollen tube elongation, etc., PGs from various biotic stress factors like insects, fungal and bacterial pathogens aid in invasion by reducing the plant cell wall rigidity. To counteract, plants secrete PGIPs, which inhibit the pectin hydrolyzing activity of PGs from the attacking pests and pathogens. Multiple approaches in diverse crop species have demonstrated PGIP-based protection against pathogens and insect pests. Additionally, effectual interaction between PGs-PGIP is an important aspect for successful utilization of this approach. Molecular strategies leading to improved PG-PGIPs interaction is a highlight to demonstrate the use of PGIPs as an amenable stress mitigation approach. The review focuses on a comprehensive update on phylogeny of PGIPs, natural variation of resistance as well as their emerging translational utility towards mitigation of various biotic stresses.

Unraveling the proteomic changes involved in the resistance response of Cajanus platycarpus to herbivory by Helicoverpa armigera.

The pigeonpea wild relative Cajanus platycarpus is resistant to Helicoverpa armigera, one of the major pests responsible for yield losses in Cajanus cajan. Deciphering the molecular mechanism underlying host plant resistance is pertinent to identify proteins that aid in the mitigation of the insect pest. The present study adopted comparative proteomics as a tool to interpret the resistance mechanism(s) in C. platycarpus vis-à-vis C. cajan during continued herbivory (up to 96 h). Over-representation analysis of the differentially expressed proteins implicated a multi-dimensional resistance response accomplished by both physical and chemical barriers in C. platycarpus. While the chemical basis for resistance was depicted by the upregulation of proteins playing a rate limiting role in the phenylpropanoid pathway, the physical basis was provided by the regulation of proteins involved in microtubule assembly and synthesis of lignins. Upregulation of proteins in the polyamine pathway indicated the role of metabolite conjugates to be negatively affecting herbivore growth. Reallocation of resources and diversion of metabolic flux to support the production of secondary metabolites could be the probable approach in the wild relative against herbivory. Our study provided deeper insights into the pod borer resistance mechanism in C. platycarpus for utility in crop improvement. KEY POINTS: • Pod borer resistance in Cajanus platycarpus is multi-dimensional. • Pod borer resistance has been arbitrated to cell wall rigidity and secondary metabolites. • Phenylpropanoid pathway derivatives apparently shaped the plant chemical defense against pod borer.

Agrobacterium tumefaciens-mediated in planta transformation strategy for development of transgenics in cotton (Gossypium hirsutum L.) with GFP as a visual marker.

Cotton (Gossypium hirsutum L.), a mercantile crop plant, is grown worldwide for fiber and seed oil. As with other economically important crops, cotton is bogged down with many biotic and abiotic stress factors. Towards this, genetic engineering offers numerous protocols to engineer plants for better resilience. However, recalcitrance of cotton to plant tissue culture has been the major constraint for successful in vitro regeneration. Hence, alternate methods that evade tissue culture regeneration have been envisaged. Non tissue culture-based in planta transformation strategies are in vogue due to amenability and ease in the generation of transgenic plants. In the present study, we demonstrate the utility of an in planta transformation protocol and establishment of a stringent selection agent-based screening for the identification of transgenics. The genotype independent nature of the protocol was validated in cotton cv. Pusa 8-6 using GFP. Preliminary transformation efficiency of 28% was achieved with a screening efficiency of 20% in the presence of hygromycin. The proof of T-DNA integration by various molecular and expression analysis in T1 and T2 generations proved that this technique can be employed to generate transgenic cotton.