Nucleotide-binding leucine-rich repeat network underlies nonhost resistance of pepper against the Irish potato famine pathogen Phytophthora infestans.

Nonhost resistance (NHR) is a robust plant immune response against non-adapted pathogens. A number of nucleotide-binding leucine-rich repeat (NLR) proteins that recognize non-adapted pathogens have been identified, although the underlying molecular mechanisms driving robustness of NHR are still unknown. Here, we screened 57 effectors of the potato late blight pathogen Phytophthora infestans in nonhost pepper (Capsicum annuum) to identify avirulence effector candidates. Selected effectors were tested against 436 genome-wide cloned pepper NLRs, and we identified multiple functional NLRs that recognize P. infestans effectors and confer disease resistance in the Nicotiana benthamiana as a surrogate system. The identified NLRs were homologous to known NLRs derived from wild potatoes that recognize P. infestans effectors such as Avr2, Avrblb1, Avrblb2, and Avrvnt1. The identified CaRpi-blb2 is a homologue of Rpi-blb2, recognizes Avrblb2 family effectors, exhibits feature of lineage-specifically evolved gene in microsynteny and phylogenetic analyses, and requires pepper-specific NRC (NLR required for cell death)-type helper NLR for proper function. Moreover, CaRpi-blb2-mediated hypersensitive response and blight resistance were more tolerant to suppression by the PITG_15 278 than those mediated by Rpi-blb2. Combined results indicate that pepper has stacked multiple NLRs recognizing effectors of non-adapted P. infestans, and these NLRs could be more tolerant to pathogen-mediated immune suppression than NLRs derived from the host plants. Our study suggests that NLRs derived from nonhost plants have potential as untapped resources to develop crops with durable resistance against fast-evolving pathogens by stacking the network of nonhost NLRs into susceptible host plants.

Vaginal vault drainage as an effective and feasible alternative in laparoscopic hysterectomy.

OBJECTIVE: Infected pelvic hematoma is a serious complication of hysterectomies. Pelvic drainage can help reduce complications. In this study, we evaluated the efficacy and safety of vaginal vault drainage in patients who underwent laparoscopic hysterectomy for benign gynecological diseases. METHODS: Patients who underwent laparoscopic hysterectomy and pelvic drain insertion for benign gynecological diseases between January 2008 and December 2015 were enrolled retrospectively in the study. They were grouped according to drain insertion sites, that is, through the abdomen (group 1) and vaginal vault (group 2). The postoperative outcomes were compared between the two groups. RESULTS: A total of 504 women were included. No significant differences were observed in the prevalence of postoperative fever, readmission, and reoperation between the two groups. CONCLUSION: Given the discomfort associated with holding and removing the abdominal drain, inserting a closed pelvic gravity drain through the vaginal vault appears to be a feasible alternative to an abdominal drain.

Receptor-mediated nonhost resistance in plants.

Nonhost resistance (NHR) is a plant immune response that prevents many microorganisms in the plant's environment from pathogenicity against the plant. Since successful pathogens have adapted to overcome the immune systems of their host, the durable nature of NHR has potential in the management of plant disease. At present, there is genetic and molecular evidence that the underlying molecular mechanisms of NHR are similar to the plant immune responses that occur in host plants following infection by adapted pathogens. We consider that the molecular basis of NHR is multilayered, conferred by physicochemical barriers and defense responses that are induced following molecular recognition events. Moreover, the relative contribution of each component may depend on evolutionary distances between host and nonhost plants of given pathogen species. This mini-review has focused on the current knowledge of plant NHR, especially the recognition of non-adapted pathogens by nonhost plants at the cellular level. Recent gains in understanding the roles of plasma membrane-localized pattern-recognition receptors (PRRs) and the cytoplasmic nucleotide-binding leucine-rich repeat receptors (NLRs) associated with these processes, as well as the genes involved, are summarized. Finally, we provide a theoretical perspective on the durability of receptor-mediated NHR and its practical potential as an innovative strategy for crop protection against pathogens.

Plasma membrane-localized plant immune receptor targets H+ -ATPase for membrane depolarization to regulate cell death.

The hypersensitive response (HR) is a robust immune response mediated by nucleotide-binding, leucine-rich repeat receptors (NLRs). However, the early molecular event that links activated NLRs to cell death is unclear. Here, we demonstrate that NLRs target plasma membrane H+ -ATPases (PMAs) that generate electrochemical potential, an essential component of living cells, across the plasma membrane. CCA 309, an autoactive N-terminal domain of a coiled-coil NLR (CNL) in pepper, is associated with PMAs. Silencing or overexpression of PMAs reversibly affects cell death induced by CCA 309 in Nicotiana benthamiana. CCA 309-induced extracellular alkalization causes plasma membrane depolarization, followed by cell death. Coimmunoprecipitation analyses suggest that CCA 309 inhibits PMA activation by preoccupying the dephosphorylated penultimate threonine residue of PMA. Moreover, pharmacological experiments using fusicoccin, an irreversible PMA activator, showed that inhibition of PMAs contributes to CNL-type (but not Toll interleukin-1 receptor NLR-type) resistance protein-induced cell death. We suggest PMAs as primary targets of plasma membrane-associated CNLs leading to HR-associated cell death by disturbing the electrochemical gradient across the membrane. These results provide new insight into NLR-mediated cell death in plants, as well as innate immunity in higher eukaryotes.

Comparative analysis of de novo genomes reveals dynamic intra-species divergence of NLRs in pepper.

BACKGROUND: Peppers (Capsicum annuum L.) containing distinct capsaicinoids are the most widely cultivated spices in the world. However, extreme genomic diversity among species represents an obstacle to breeding pepper. RESULTS: Here, we report de novo genome assemblies of Capsicum annuum 'Early Calwonder (non-pungent, ECW)' and 'Small Fruit (pungent, SF)' along with their annotations. In total, we assembled 2.9 Gb of ECW and SF genome sequences, representing over 91% of the estimated genome sizes. Structural and functional annotation of the two pepper genomes generated about 35,000 protein-coding genes each, of which 93% were assigned putative functions. Comparison between newly and publicly available pepper gene annotations revealed both shared and specific gene content. In addition, a comprehensive analysis of nucleotide-binding and leucine-rich repeat (NLR) genes through whole-genome alignment identified five significant regions of NLR copy number variation (CNV). Detailed comparisons of those regions revealed that these CNVs were generated by intra-specific genomic variations that accelerated diversification of NLRs among peppers. CONCLUSIONS: Our analyses unveil an evolutionary mechanism responsible for generating CNVs of NLRs among pepper accessions, and provide novel genomic resources for functional genomics and molecular breeding of disease resistance in Capsicum species.

Pressurized intraperitoneal aerosol chemotherapy for recurrent ovarian, fallopian or primary peritoneal cancer with peritoneal carcinomatosis: a narrative review.

For recurrent ovarian, fallopian or primary peritoneal cancer with peritoneal carcinomatosis (PC), it is challenging to resect tumors completely or to get complete remission by intravenous (IV) chemotherapy, and many patients show the resistance to various chemotherapeutic agents for IV chemotherapy ultimately. As an alternative, pressurized intraperitoneal aerosol chemotherapy (PIPAC) has been introduced for treating the disease, which delivers chemotherapeutic agents as an aerosol form while maintaining high intraperitoneal (IP) pressure. Based on preclinical studies, PIPAC showed better penetration depth and distribution of drugs into the peritoneum in comparison to conventional IP chemotherapy. Tumor regression on histology and peritoneal carcinomatosis index (PCI) has also been shown in relevant studies. In addition, most of the PIPAC procedures were completed successfully with acceptable toxicity due to the use of a low dose of chemotherapeutic agents. For considering these advantages of PIPAC, we review the current status of PIPAC for treating recurrent ovarian, fallopian or primary peritoneal cancer through literature review.

Genome-wide functional analysis of hot pepper immune receptors reveals an autonomous NLR clade in seed plants.

Plants possess hundreds of intracellular immune receptors encoding nucleotide-binding domain leucine-rich repeat (NLR) proteins. Full-length NLRs or a specific domain of NLRs often induce plant cell death in the absence of pathogen infection. In this study we used genome-wide transient expression analysis to identify a group of NLRs (ANLs; ancient and autonomous NLRs) carrying autoactive coiled-coil (CCA ) domains in pepper (Capsicum annuum). CCA -mediated cell death mimics hypersensitive cell death triggered by the interaction between NLRs and pathogen effectors. Sequence alignment and mutagenesis analyses revealed that the intact α1 helix of CCA s is critical for both CCA - and ANL-mediated cell death. Cell death induced by CCA s does not require NRG1/ADR1 or NRC type helper NLRs, suggesting ANLs may function as singleton NLRs. We also found that CCA s localize to the plasma membrane, as demonstrated for Arabidopsis singleton NLR ZAR1. Extended studies revealed that autoactive CCA s are well conserved in other Solanaceae plants as well as in rice, a monocot plant. Further phylogenetic analyses revealed that ANLs are present in all tested seed plants (spermatophytes). Our study not only uncovers the autonomous NLR clade in plants but also provides powerful resources for dissecting the underlying molecular mechanism of NLR-mediated cell death in plants.

Feasibility of local bone dust as a graft material in anterior cervical discectomy and fusion.

OBJECTIVE: Local bone dust has been used previously as a substitute cage filling material for iliac bone grafts during anterior cervical discectomy and fusion (ACDF). However, the impacts of local bone dust on fusion rate and clinical results remain unclear. Extragraft bone bridging (ExGBB) is a reliable CT finding indicating segmental fusion. This study was conducted to compare fusion rates for the use of local bone dust or an iliac auto bone graft during ACDF surgery and to evaluate the effect of implanting bone graft outside the cage. METHODS: Ninety-three patients who underwent ACDF at a single institution were included. An iliac bone graft was used as the polyetheretherketone (PEEK) cage filling graft material in 43 patients (iliac crest [IC] group). In the IC group, bone graft material was inserted only inside the cage. Local bone dust was used in 50 patients (local bone [LB] group). Bone graft material was inserted both inside and outside the cage in the LB group. Segmental fusion was assessed by 1) interspinous motion (ISM), 2) intragraft bone bridging (InGBB), and 3) ExGBB. Fusion rates, visual analog scale (VAS) scores for neck and arm pain, and Neck Disability Index (NDI) scores were compared between the 2 groups. RESULTS: The neck and arm pain VAS scores and NDI score improved significantly in both groups. Fusion rates assessed by ISM and InGBB did not differ significantly between the groups. However, the fusion rate in the LB group was significantly higher than that in the IC group when assessed by ExGBB (p = 0.02). CONCLUSIONS: Using local bone dust as cage filling material resulted in fusion rates similar to those for an iliac bone graft, while avoiding potential complications and pain caused by iliac bone harvesting. A higher rate of extragraft bone bridge formation was achieved by implanting local bone dust outside the cage.

Practice patterns of surgery for advanced ovarian cancer: analysis from international surveys.

BACKGROUND: We investigated the practice patterns of surgery for advanced ovarian cancer (AOC) through relevant international surveys. METHODS: After searching for 878 studies on surgery for AOC till 2017, we extracted 18 questions with similar query and answer formats from eight studies. Among them, 5 and 13 were classified as comprehensive and procedure information. RESULTS: In comprehensive information, there was a higher preference for optimal cytoreduction defined as no visible tumor (44%) compared with residual tumors <1 cm (38%) or <2 cm (2%) and omental disease involving the spleen or pancreas was more important as an intraoperative finding precluding optimal cytoreduction (35%) since 2010. The preference for neoadjuvant chemotherapy was the highest at its use for 1-10% (36%), which was preferred in Europe over USA. The positive expectation of preoperative determination of optimal cytoreduction was higher in Europe than in USA (44 vs. 27%; P < 0.05). In procedure information, conventional gynecological surgery was mainly performed by gynecological oncologists, whereas more than 50% of upper abdominal or urological surgeries were conducted by other surgeons. European clinicians showed a higher response rate of diaphragmatic stripping and resection than those from USA (88 vs. 60%; 69 vs. 24%; P < 0.05). CONCLUSION: No visible tumor as the criterion for optimal cytoreduction was preferred in AOC, and aggressive surgery beyond conventional gynecological surgery tended to be performed by other surgeons. Moreover, the preference of neoadjuvant chemotherapy and the positive expectation of preoperative determination of optimal cytoreduction were higher in Europe than in USA.

Development of a novel nomogram for predicting ongoing pregnancy after in vitro fertilization and embryo transfer.

OBJECTIVE: This study aimed to develop a nomogram that predicts ongoing pregnancy after in vitro fertilization and embryo transfer (IVF-ET) using patient age and serum hormonal markers. METHODS: A total of 284 IVF-ET cycles were retrospectively analyzed. At 14 days post-oocyte pick-up (OPU), the serum human chorionic gonadotropin (HCG) and progesterone levels were measured. The main predicted outcome was ongoing pregnancy. RESULTS: Patient age and serum of HCG and progesterone levels at 14 days post-OPU were good predictors of ongoing pregnancy. The cut-off value and area under the curve (AUC) (95% confidence interval) were 36.5 years and 0.666 (0.599-0.733), respectively, for patient age; 67.8 mIU/mL and 0.969 (0.951-0.987), respectively, for serum HCG level; and 29.8 ng/mL and 0.883 (0.840-0.925), respectively, for serum progesterone level. When the prediction model was constructed using these three parameters, the addition of serum progesterone level to the prediction model did not increase its overall predictability. Furthermore, a high linear co-relationship was found between serum HCG and progesterone levels. Therefore, we developed a new nomogram using patient age and HCG serum level only. The AUC of the newly developed nomogram for predicting ongoing pregnancy after IVF-ET cycles using patient age and serum HCG level was as high as 0.975. CONCLUSION: We showed that ongoing pregnancy may be predicted using only patient age and HCG serum level. Our nomogram could help clinicians and patients predict ongoing pregnancy after IVF-ET if the serum JCG level was ≥5 IU/L at 14 days post-OPU.

HSP70s Enhance a Phytophthora infestans Effector-Induced Cell Death via an MAPK Cascade in Nicotiana benthamiana.

A destructive pathogen, Phytophthora infestans, secretes hundreds of effectors for successful survival in its host plants. The effectors modulate the plant defense system at diverse cellular compartments to take an advantage of pathogen survivals. A few research studies have shown the mode of action of each effector and their interacting proteins in plant cells. Here, we investigated the mode of action of a P. infestans effector, Pi23226, which induces cell death in Nicotiana benthamiana. To identify its host factors, we performed coimmunoprecipitation and liquid chromatography-mass spectrometry, and selected members of heat shock protein 70 (HSP70s) as candidates. These HSP70s, known to function as chaperones, were associated with Pi23226 in planta and accelerated Pi23226-induced cell death. Additionally, they were found to be involved in plant basal defense by suppressing the growth of P. infestans. We also found that specific components of a mitogen-activated protein kinase cascade were involved in Pi23226-induced cell death. Our findings show that HSP70s functions in defense systems by regulating effector-triggered cell death and by suppressing the growth of the pathogen. This suggests that host plants manipulate the ubiquitous proteins to detect pathogen effectors for functioning in the defense system.

New reference genome sequences of hot pepper reveal the massive evolution of plant disease-resistance genes by retroduplication.

BACKGROUND: Transposable elements are major evolutionary forces which can cause new genome structure and species diversification. The role of transposable elements in the expansion of nucleotide-binding and leucine-rich-repeat proteins (NLRs), the major disease-resistance gene families, has been unexplored in plants. RESULTS: We report two high-quality de novo genomes (Capsicum baccatum and C. chinense) and an improved reference genome (C. annuum) for peppers. Dynamic genome rearrangements involving translocations among chromosomes 3, 5, and 9 were detected in comparison between C. baccatum and the two other peppers. The amplification of athila LTR-retrotransposons, members of the gypsy superfamily, led to genome expansion in C. baccatum. In-depth genome-wide comparison of genes and repeats unveiled that the copy numbers of NLRs were greatly increased by LTR-retrotransposon-mediated retroduplication. Moreover, retroduplicated NLRs are abundant across the angiosperms and, in most cases, are lineage-specific. CONCLUSIONS: Our study reveals that retroduplication has played key roles for the massive emergence of NLR genes including functional disease-resistance genes in pepper plants.

Current Understandings of Plant Nonhost Resistance.

Nonhost resistance, a resistance of plant species against all nonadapted pathogens, is considered the most durable and efficient immune system of plants but yet remains elusive. The underlying mechanism of nonhost resistance has been investigated at multiple levels of plant defense for several decades. In this review, we have comprehensively surveyed the latest literature on nonhost resistance in terms of preinvasion, metabolic defense, pattern-triggered immunity, effector-triggered immunity, defense signaling, and possible application in crop protection. Overall, we summarize the current understanding of nonhost resistance mechanisms. Pre- and postinvasion is not much deviated from the knowledge on host resistance, except for a few specific cases. Further insights on the roles of the pattern recognition receptor gene family, multiple interactions between effectors from nonadapted pathogen and plant factors, and plant secondary metabolites in host range determination could expand our knowledge on nonhost resistance and provide efficient tools for future crop protection using combinational biotechnology approaches. [Formula: see text] Copyright © 2017 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license .

Discovery of Q203, a potent clinical candidate for the treatment of tuberculosis.

New therapeutic strategies are needed to combat the tuberculosis pandemic and the spread of multidrug-resistant (MDR) and extensively drug-resistant (XDR) forms of the disease, which remain a serious public health challenge worldwide. The most urgent clinical need is to discover potent agents capable of reducing the duration of MDR and XDR tuberculosis therapy with a success rate comparable to that of current therapies for drug-susceptible tuberculosis. The last decade has seen the discovery of new agent classes for the management of tuberculosis, several of which are currently in clinical trials. However, given the high attrition rate of drug candidates during clinical development and the emergence of drug resistance, the discovery of additional clinical candidates is clearly needed. Here, we report on a promising class of imidazopyridine amide (IPA) compounds that block Mycobacterium tuberculosis growth by targeting the respiratory cytochrome bc1 complex. The optimized IPA compound Q203 inhibited the growth of MDR and XDR M. tuberculosis clinical isolates in culture broth medium in the low nanomolar range and was efficacious in a mouse model of tuberculosis at a dose less than 1 mg per kg body weight, which highlights the potency of this compound. In addition, Q203 displays pharmacokinetic and safety profiles compatible with once-daily dosing. Together, our data indicate that Q203 is a promising new clinical candidate for the treatment of tuberculosis.

ClpS1 is a conserved substrate selector for the chloroplast Clp protease system in Arabidopsis.

Whereas the plastid caseinolytic peptidase (Clp) P protease system is essential for plant development, substrates and substrate selection mechanisms are unknown. Bacterial ClpS is involved in N-degron substrate selection and delivery to the ClpAP protease. Through phylogenetic analysis, we show that all angiosperms contain ClpS1 and some species also contain ClpS1-like protein(s). In silico analysis suggests that ClpS1 is the functional homolog of bacterial ClpS. We show that Arabidopsis thaliana ClpS1 interacts with plastid ClpC1,2 chaperones. The Arabidopsis ClpS1 null mutant (clps1) lacks a visible phenotype, and no genetic interactions with ClpC/D chaperone or ClpPR core mutants were observed. However, clps1, but not clpc1-1, has increased sensitivity to the translational elongation inhibitor chloramphenicol suggesting a link between translational capacity and ClpS1. Moreover, ClpS1 was upregulated in clpc1-1, and quantitative proteomics of clps1, clpc1, and clps1 clpc1 showed specific molecular phenotypes attributed to loss of ClpC1 or ClpS1. In particular, clps1 showed alteration of the tetrapyrrole pathway. Affinity purification identified eight candidate ClpS1 substrates, including plastid DNA repair proteins and Glu tRNA reductase, which is a control point for tetrapyrrole synthesis. ClpS1 interaction with five substrates strictly depended on two conserved ClpS1 residues involved in N-degron recognition. ClpS1 function, substrates, and substrate recognition mechanisms are discussed.

Modified Clp protease complex in the ClpP3 null mutant and consequences for chloroplast development and function in Arabidopsis.

The plastid ClpPRT protease consists of two heptameric rings of ClpP1/ClpR1/ClpR2/ClpR3/ClpR4 (the R-ring) and ClpP3/ClpP4/ClpP5/ClpP6 (the P-ring) and peripherally associated ClpT1/ClpT2 subunits. Here, we address the contributions of ClpP3 and ClpP4 to ClpPRT core organization and function in Arabidopsis (Arabidopsis thaliana). ClpP4 is strictly required for embryogenesis, similar to ClpP5. In contrast, loss of ClpP3 (clpp3-1) leads to arrest at the hypocotyl stage; this developmental arrest can be removed by supplementation with sucrose or glucose. Heterotrophically grown clpp3-1 can be transferred to soil and generate viable seed, which is surprising, since we previously showed that CLPR2 and CLPR4 null alleles are always sterile and die on soil. Based on native gels and mass spectrometry-based quantification, we show that despite the loss of ClpP3, modified ClpPR core(s) could be formed, albeit at strongly reduced levels. A large portion of ClpPR subunits accumulated in heptameric rings, with overaccumulation of ClpP1/ClpP5/ClpP6 and ClpR3. Remarkably, the association of ClpT1 to the modified Clp core was unchanged. Large-scale quantitative proteomics assays of clpp3-1 showed a 50% loss of photosynthetic capacity and the up-regulation of plastoglobules and all chloroplast stromal chaperone systems. Specific chloroplast proteases were significantly up-regulated, whereas the major thylakoid protease (FtsH1/FtsH2/FtsH5/FtsH8) was clearly unchanged, indicating a controlled protease network response. clpp3-1 showed a systematic decrease of chloroplast-encoded proteins that are part of the photosynthetic apparatus but not of chloroplast-encoded proteins with other functions. Candidate substrates and an explanation for the differential phenotypes between the CLPP3, CLPP4, and CLPP5 null mutants are discussed.