T3SS protein EsrC binds to the lacI-like operator of type 1 fimbrial operon to suppress adhesion of Edwardsiella piscicida.

Type 1 fimbria, the short hair-like appendage assembled on the bacterial surface, plays a pivotal role in adhesion and invasion in Edwardsiella piscicida. The type III secretion system (T3SS), another bacterial surface appendage, facilitates E. piscicida's replication in vivo by delivering effectors into host cells. Our previous research demonstrated that E. piscicida T3SS protein EseJ inhibits adhesion and invasion of E. piscicida by suppressing type 1 fimbria. However, how EseJ suppresses type 1 fimbria remains elusive. In this study, a lacI-like operator (nt -245 to -1 of fimA) upstream of type 1 fimbrial operon in E. piscicida was identified, and EseJ inhibits type 1 fimbria through the lacI-like operator. Moreover, through DNA pull-down and electrophoretic mobility shift assay, an AraC-type T3SS regulator, EsrC, was screened and verified to bind to nt -145 to -126 and nt -50 to -1 of fimA, suppressing type 1 fimbria. EseJ is almost abolished upon the depletion of EsrC. EsrC and EseJ impede type 1 fimbria expression. Intriguingly, nutrition and microbiota-derived indole activate type 1 fimbria through downregulating T3SS, alleviating EsrC or EseJ's inhibitory effect on lacI-like operator of type 1 fimbrial operon. By this study, it is revealed that upon entering the gastrointestinal tract, rich nutrients and indole downregulate T3SS and thereof upregulate type 1 fimbria, stimulating efficient adhesion and invasion; upon being internalized into epithelium, the limit in indole and nutrition switches on T3SS and thereof switches off type 1 fimbria, facilitating effector delivery to guarantee E. piscicida's survival/replication in vivo.IMPORTANCEIn this work, we identified the lacI-like operator of type 1 fimbrial operon in E. piscicida, which was suppressed by the repressors-T3SS protein EseJ and EsrC. We unveiled that E. piscicida upregulates type 1 fimbria upon sensing rich nutrition and the microbiota-derived indole, thereof promoting the adhesion of E. piscicida. The increase of indole and nutrition promotes type 1 fimbria by downregulating T3SS. The decrease in EseJ and EsrC alleviates their suppression on type 1 fimbria, and vice versa.

Hybridization and divergent climatic preferences drive divergence of two allopatric Gentiana species on the Qinghai-Tibet Plateau.

BACKGROUND AND AIMS: Exploring how species diverge is vital for understanding the drivers of speciation. Factors such as geographical separation and ecological selection, hybridization, polyploidization and shifts in mating system are all major mechanisms of plant speciation, but their contributions to divergence are rarely well understood. Here we test these mechanisms in two plant species, Gentiana lhassica and G. hoae, with the goal of understanding recent allopatric species divergence on the Qinghai-Tibet Plateau (QTP). METHODS: We performed Bayesian clustering, phylogenetic analysis and estimates of hybridization using 561 302 nuclear genomic single nucleotide polymorphisms (SNPs). We performed redundancy analysis, and identified and annotated species-specific SNPs (ssSNPs) to explore the association between climatic preference and genetic divergence. We also estimated genome sizes using flow cytometry to test for overlooked polyploidy. KEY RESULTS: Genomic evidence confirms that G. lhassica and G. hoae are closely related but distinct species, while genome size estimates show divergence occurred without polyploidy. Gentiana hoae has significantly higher average FIS values than G. lhassica. Population clustering based on genomic SNPs shows no signature of recent hybridization, but each species is characterized by a distinct history of hybridization with congeners that has shaped genome-wide variation. Gentiana lhassica has captured the chloroplast and experienced introgression with a divergent gentian species, while G. hoae has experienced recurrent hybridization with related taxa. Species distribution modelling suggested range overlap in the Last Interglacial Period, while redundancy analysis showed that precipitation and temperature are the major climatic differences explaining the separation of the species. The species differ by 2993 ssSNPs, with genome annotation showing missense variants in genes involved in stress resistance. CONCLUSIONS: This study suggests that the distinctiveness of these species on the QTP is driven by a combination of hybridization, geographical isolation, mating system differences and evolution of divergent climatic preferences.

Brain structural changes in preschool children with MRI-negative epilepsy.

PURPOSE: To investigate abnormalities in cortical and subcortical structures of the brain in preschool children with MRI-negative epilepsy. METHODS: Cortical thickness, cortical mean curvature, cortical surface area, cortical volume, and volumes of subcortical structures were measured using Freesurfer software in preschool children with epilepsy and age-matched controls. RESULTS: Findings showed cortical thickening in the left fusiform gyrus, left middle temporal gyrus, right suborbital sulcus, and right gyrus rectus, and cortical thinning mainly in the parietal lobe of preschool children with epilepsy compared to controls. The difference in cortical thickness in the left superior parietal lobule remained after correction for multiple comparisons and was negatively correlated with duration of epilepsy. Cortical mean curvature, surface area, and volume were mainly altered in the frontal and temporal lobes. Changes in mean curvature in the right pericallosal sulcus were positively correlated with age at seizure onset, and changes in mean curvature in the left intraparietal sulcus and transverse parietal sulcus were positively correlated with frequency of seizures. There were no significant differences in the volumes of the subcortical structures. CONCLUSION: Changes in preschool children with epilepsy occur in the cortical rather than subcortical structures of the brain. These findings further our understanding of the effects of epilepsy in preschool children and will inform management of epilepsy in this patient population.

Development of a hyper-adhesive and attenuated Edwardsiella ictaluri strain as a novel immersion vaccine candidate in yellow catfish (Pelteobagrus fulvidraco).

Edwardsiella ictaluri, a Gram-negative intracellular pathogen, is the causative agent of enteric septicemia in channel catfish, and catfish aquaculture in China suffers heavy economic losses due to E. ictaluri infection. Vaccination is an effective control measure for this disease. In this study, an attenuated E. ictaluri strain was acquired through deletion mutation of the T3SS protein eseJei, and the ΔeseJei strain fails to replicate in the epithelioma papillosum of carp cells. The type 1 fimbria plays a pivotal role in the adhesion of E. ictaluri, and it was found in this study that deletion of -245 to -50 nt upstream of fimA increases its adhesion to around five times that of the WT strain. A hyper-adhesive and highly attenuated double mutant (ΔeseJeiΔfimA-245--50 strain) was constructed, and it was used as a vaccine candidate in yellow catfish via bath immersion at a dosage of 1 × 105 CFU/mL. It was found that this vaccine candidate can stimulate protection when challenged with E. ictaluri HSN-1 at 5 × 107 CFU/mL (∼20 × LD50). The survival rate was 83.61% for the vaccinated group and 33.33% for the sham-vaccinated group. The RPS (relative percent of survival) of the vaccination trial reached 75.41%. In conclusion, the ΔeseJeiΔfimA-245--50 strain developed in this study can be used as a vaccine candidate. It excels in terms of ease of delivery (via bath immersion) and is highly efficient in stimulating protection against E. ictaluri infection.

Orf1B controls secretion of T3SS proteins and contributes to Edwardsiella piscicida adhesion to epithelial cells.

Edwardsiella piscicida is a Gram-negative enteric pathogen that causes hemorrhagic septicemia in fish. The type III secretion system (T3SS) is one of its two most important virulence islands. T3SS protein EseJ inhibits E. piscicida adhesion to epithelioma papillosum cyprini (EPC) cells by negatively regulating type 1 fimbria. Type 1 fimbria helps E. piscicida to adhere to fish epithelial cells. In this study, we characterized a functional unknown protein (Orf1B) encoded within the T3SS gene cluster of E. piscicida. This protein consists of 122 amino acids, sharing structural similarity with YscO in Vibrio parahaemolyticus. Orf1B controls secretion of T3SS translocon and effectors in E. piscicida. By immunoprecipitation, Orf1B was shown to interact with T3SS ATPase EsaN. This interaction may contribute to the assembly of the ATPase complex, which energizes the secretion of T3SS proteins. Moreover, disruption of Orf1B dramatically decreased E. piscicida adhesion to EPC cells due to the increased steady-state protein level of EseJ within E. piscicida. Taken together, this study partially unraveled the mechanisms through which Orf1B promotes secretion of T3SS proteins and contributes to E. piscicida adhesion. This study helps to improve our understanding on molecular mechanism of E. piscicida pathogenesis.

Natural bioactive compounds from marine fungi (2017-2020).

Secondary metabolites generated by marine fungi have relatively small molecular weights and excellent activities and have become an important source for developing drug lead compounds. The review summarizes the structures of novel small-molecule compounds derived from marine fungi in recent years; introduces representative monomers in antimicrobial, antitumor, anti-viral, and anti-neuritis aspects; and discusses their biological activities and molecular mechanisms. This review will act as a guide for further discovering marine-derived drugs with novel chemical structures and specific targeting mechanisms.

Population subdivision and hybridization in a species complex of Gentiana in the Qinghai-Tibetan Plateau.

BACKGROUND AND AIMS: Hosting several global biodiversity hotspots, the region of the Qinghai-Tibetan Plateau (QTP) is exceptionally species-rich and harbours a remarkable level of endemism. Yet, despite a growing number of studies, factors fostering divergence, speciation and ultimately diversity remain poorly understood for QTP alpine plants. This is particularly the case for the role of hybridization. Here, we explored the evolutionary history of three closely related Gentiana endemic species, and tested whether our results supported the mountain geo-biodiversity hypothesis (MGH). METHODS: We genotyped 69 populations across the QTP with one chloroplast marker and 12 nuclear microsatellite loci. We performed phylogeographical analysis, Bayesian clustering, approximate Bayesian computation and principal components analysis to explore their genetic relationship and evolutionary history. In addition, we modelled their distribution under different climates. KEY RESULTS: Each species was composed of two geographically distinct clades, corresponding to the south-eastern and north-western parts of their distribution. Thus Gentiana veitchiorum and G. lawrencei var. farreri, which diverged recently, appear to have shared at least refugia in the past, from which their range expanded later on. Indeed, climatic niche modelling showed that both species went through continuous expansion from the Last Interglacial Maximum to the present day. Moreover, we have evidence of hybridization in the northwest clade of G. lawrencei var. farreri, which probably occurred in the refugium located on the plateau platform. Furthermore, phylogenetic and population genetic analyses suggested that G. dolichocalyx should be a geographically limited distinct species with low genetic differentiation from G. lawrencei var. farreri. CONCLUSIONS: Climatic fluctuations in the region of the QTP have played an important role in shaping the current genetic structure of G. lawrencei var. farreri and G. veitchiorum. We argue that a species pump effect did occur prior to the Last Interglacial Maximum, thus lending support to the MGH. However, our results do depart from expectations as suggested in the MGH for more recent distribution range and hybridization dynamics.

Talarolactone A, an Isocoumarin Derivative Fused with Dihydrothiophene with Selective Antimigratory Activity from the Endolichenic Fungus Talaromyces sp.

A 3,4-dihydroisocoumarin derivative fused with dihydrothiophene, talarolactone A (1), and two known compounds, terreusinone (2) and 4,6-dihydroxy-5-methylphthalide (3), were isolated from Talaromyces sp. associated with Xanthoparmelia angustiphylla. The structure of 1 was deduced from extensive spectroscopic data, electronic circular dichroism calculations, and X-ray diffraction analyses. A plausible biosynthetic pathway of 1 was further proposed. Compound 1 showed selective antimigratory activity in a wound-healing assay without appreciable cytotoxic activity.

Efficacy and safety of urate-lowering treatments in patients with hyperuricemia: A comprehensive network meta-analysis of randomized controlled trials.

WHAT IS KNOWN AND OBJECTIVE: Hyperuricemia (HUA) and gout are considerable public health problems because of their increasing incidence and interactions with other diseases. We aimed to evaluate the efficacy and safety of urate-lowering therapies (ULTs) for patients. METHODS: A systematic literature review was conducted, and a network meta-analysis was performed on the included studies using the Markov Chain Monte Carlo simulation method and a Bayesian statistical framework. We calculated surface under the cumulative ranking curve (SUCRA) values and performed clustered ranking to combine the efficacy and safety results. RESULTS: Twenty-two randomized controlled studies were identified for the efficacy analysis, and 20 studies were identified for the safety analysis. Compared with the placebo, the ULTs were efficient and safe. Febuxostat 120 mg/d and allopurinol 200 mg/d had the highest SUCRA scores for efficacy and safety, respectively. Clustered ranking results showed that febuxostat 120 mg/d was the best in terms of efficacy and safety, topiroxostat 120/160 mg/d was similar to febuxostat 80 mg/d in terms of efficacy but safer, and allopurinol was not inferior to topiroxostat. WHAT IS NEW AND CONCLUSION: Febuxostat had the best efficacy and safety results among the tested agents, and topiroxostat and allopurinol appeared to have fewer adverse events.

Unveiling the Photophysical and Photochemical Reaction Process of Naproxen via Ultrafast Femtosecond to Nanosecond Laser Flash Photolysis.

Naproxen is a nonsteroidal anti-inflammatory drug that exhibits phototoxic side effects in humans, but its mechanism of phototoxicity is ambiguous. To uncover photophysical and photochemical reaction processes of naproxen, femtosecond to nanosecond transient absorption spectroscopies were employed to directly detect excited and transient states of naproxen upon UV irradiation in pure acetonitrile, acetonitrile:water 1:1, and acetonitrile:PBS 1:1 solutions. The transient absorption data together with time-dependent density functional theory analysis-predicted absorption spectra of selected intermediates were integrated to elucidate photochemical mechanisms for reactions of naproxen in different solutions. Femtosecond transient absorption results demonstrated that naproxen has two different photochemical pathways at the early delay time before the formation of final products in various solutions. In a pure acetonitrile solvent, naproxen undergoes charge transfer to solvent to generate a radical cation intermediate, which decarboxylates to generate a radical 2B intermediate. While in an acetonitrile:PBS 1:1 solution, naproxen predominantly deprotonates first and is promoted to the singlet exited state (1NPX-), which undergoes intersystem crossing to give rise to the lowest-lying triplet states (T1). T1 then undergoes decarboxylation reaction and produces a radical 2B species. Kinetic characterization of these processes reveals that the decarboxylation reaction in an acetonitrile:PBS 1:1 solution is faster than that in a pure acetonitrile solvent. Deep studies on photophysical and photochemical processes of NPX will aid us to better understand the toxicology mechanisms associated with NPX in different conditions.

Active Metabolites from Endolichenic Fungus Talaromyces sp.

The active metabolites investigation of Talaromyces sp. (strain No. MH551540) associated with Xanthoparmelia angustiphylla afforded one new δ-lactone, talaromycin A (1), together with six known compounds, clearanol A (2), 6-methylbiphenyl-3,3',4,5'-tetraol (3), desmethylaltenusin (4), ergone (5), ergosterol (6), and palmitic acid (7). The structures of these compounds were elucidated by a combination of spectroscopic-data interpretation and single-crystal X-ray diffraction analysis. The cytotoxicities of 1-7 and the antioxidant activities of 3 and 4 were also evaluated.

Chemical identification and quality evaluation of Lycopus lucidus Turcz by UHPLC-Q-TOF-MS and HPLC-MS/MS and hierarchical clustering analysis.

Lycopus lucidus Turcz has been used as a traditional phytomedicine for menstrual disorder, amenorrhea, menstrual cramps, inflammation and cardiovascular diseases. However, there is not enough information about identification and quantification for the chemical constituents of L. lucidus Turcz. In this work, a simple, rapid and sensitive UHPLC-Q-TOF-MS method was developed for characterization and identification of the phytochemical compositions in L. lucidus Turcz in negative ion mode. A total of 37 compounds, including 15 phenolic acids, 12 flavonoids, three triterpenoids and seven organic acids were tentatively characterized and identified by means of the retention time, accurate mass and characteristic fragment ions. Thirteen compounds were reported for the first time in L. lucidus Turcz. Among of them, 11 compounds were further quantified by multiple reactions monitoring. The results showed good performance with respect to linearity (r > 0.9959), repeatability (RSD < 2.6%), intra- and inter-day precision (RSD < 3.2%), recovery (93.1-104.9%), and lower limit of quantification (5-50 ng/mL). Subsequently, the results were analyzed and classified by hierarchical cluster analysis. The research could be applied for identification and quality evaluation for L. lucidus Turcz.

Comparison of maternal and fetal complications in elective and emergency cesarean section: a systematic review and meta-analysis.

BACKGROUND: Though the same types of complication were found in both elective cesarean section (ElCS) and emergence cesarean section (EmCS), the aim of this study is to compare the rates of maternal and fetal morbidity and mortality between ElCS and EmCS. METHODS: Full-text articles involved in the maternal and fetal complications and outcomes of ElCS and EmCS were searched in multiple database. Review Manager 5.0 was adopted for meta-analysis, sensitivity analysis, and bias analysis. Funnel plots and Egger's tests were also applied with STATA 10.0 software to assess possible publication bias. RESULTS: Totally nine articles were included in this study. Among these articles, seven, three, and four studies were involved in the maternal complication, fetal complication, and fetal outcomes, respectively. The combined analyses showed that both rates of maternal complication and fetal complication in EmCS were higher than those in ElCS. The rates of infection, fever, UTI (urinary tract infection), wound dehiscence, DIC (disseminated intravascular coagulation), and reoperation of postpartum women with EmCS were much higher than those with ElCS. Larger infant mortality rate of EmCS was also observed. CONCLUSION: Emergency cesarean sections showed significantly more maternal and fetal complications and mortality than elective cesarean sections in this study. Certain plans should be worked out by obstetric practitioners to avoid the post-operative complications.

Targeting STAT3/miR-21 axis inhibits epithelial-mesenchymal transition via regulating CDK5 in head and neck squamous cell carcinoma.

BACKGROUND: Abnormal activation of STAT3 and miR-21 plays a vital role in progression and invasion of solid tumors. The cyclin-dependent kinase 5 (CDK5) is reported to contribute to cancer metastasis by regulating epithelial-mesenchymal transition (EMT). However, the role of STAT3/miR-21 axis and CDK5 in head and neck squamous cell carcinoma remains unclear. METHODS: We measured the expression of miR-21, CDK5 and EMT markers in 60 HNSCC tumor samples. We used Immunohistochemistry and in situ hybridization assay to examine the role of STAT3/miR-21 axis and CDK5 activation in the invasiveness of HNSCC. The clinical survival relevance was analyzed by Kaplan-Meier analysis and univariate/multivariate COX regression model. Multiple approaches including scratch, transwell chamber assay and other molecular biology techniques were used to validate the anti-invasion effect of targeting miR-21 in Tca8113 and Hep-2 cell lines in vitro. Furthermore, whether miR-21 depletion inhibits HNSCC invasion in vivo was confirmed in Tca8113 xenograft tumor model. RESULTS: The expression of miR-21 and CDK5 were significantly correlated with lymph node metastasis in HNSCC. Hep-2 and Tca8113 cell lines showed co-overexpression of miR-21 and CDK5. WP1066 or asON-miR-21 treatment depleted miR-21 and CDK5 expression and significantly inhibited migration or invasion in Hep-2 and Tca8113 cells. The expression levels of CDK5/p35, N-cadherin, vimentin, ß-catenin were inhibited while E-cadherin level was increased by miR-21 depletion in vitro and in vivo. Conversely, ectopic CDK5 overexpression significantly induced tumor cell motility and EMT. Moreover, ectopic CDK5 overexpression in Hep-2 and Tca8113 cells rescued the observed phenotype after miR-21 silencing or WP1066 treatment. CONCLUSIONS: miR-21 cooperates with CDK5 to promote EMT and invasion in HNSCC. This finding suggests that CDK5 may be an important cofactor for targeting when designing metastasis-blocking therapy by targeting STAT3/miR-21 axis with STAT3 inhibitor or miR-21 antisense oligonucleotide. This is the first demonstration of the novel role of STAT3/miR-21 axis and CDK5/CDK5R1 (p35) in metastasis of HNSCC.

Functional study on the mutations in the silkworm (Bombyx mori) acetylcholinesterase type 1 gene (ace1) and its recombinant proteins.

The acetylcholinesterase of Lepidoptera insects is encoded by two genes, ace1 and ace2. The expression of the ace1 gene is significantly higher than that of the ace2 gene, and mutations in ace1 are one of the major reasons for pesticide resistance in insects. In order to investigate the effects of the mutations in ace1's characteristic sites on pesticide resistance, we generated mutations for three amino acids using site-directed mutagenesis, which were Ala(GCG)303Ser(TCG), Gly(GGA)329Ala(GCA) and Leu (TCT)554Ser(TTC). The Baculovirus expression system was used for the eukaryotic expression of the wild type ace1 (wace1) and the mutant ace1 (mace1). SDS-PAGE and Western blotting were used to detect the targeting proteins with expected sizeof about 76 kDa. The expression products were purified for the determination of AChE activity and the inhibitory effects of physostigmine and phoxim. We observed no significant differences in the overall activity of the wild type and mutant AChEs. However, with 10 min of physostigmine (10 µM) inhibition, the remaining activity of the wild type AChE was significantly lower than that of the mutant AChE. Ten min inhibition with 33.4 µM phoxim also resulted in significantly lower remaining activity of the wild type AChE than that of the mutant AChE. These results indicated that mutations for the three amino acids reduced the sensitivity of AChE to physostigmine and phoxim, which laid the foundation for future in vivo studies on AChE's roles in pesticide resistance.

Cryptic diversity and rampant hybridization in annual gentians on the Qinghai-Tibet Plateau revealed by population genomic analysis.

Understanding the evolutionary and ecological processes involved in population differentiation and speciation provides critical insights into biodiversity formation. In this study, we employed 29,865 single nucleotide polymorphisms (SNPs) and complete plastomes to examine genomic divergence and hybridization in Gentiana aristata, which is endemic to the Qinghai-Tibet Plateau (QTP) region. Genetic clustering revealed that G. aristata is characterized by geographic genetic structures with five clusters (West, East, Central, South and North). The West cluster has a specific morphological character (i.e., blue corolla) and higher values of FST compared to the remaining clusters, likely the result of the geological barrier formed by the Yangtze River. The West cluster diverged from the other clusters in the Early Pliocene; these remaining clusters diverged from one another in the Early Quaternary. Phylogenetic reconstructions based on SNPs and plastid data revealed substantial cyto-nuclear conflicts. Genetic clustering and D-statistics demonstrated rampant hybridization between the Central and North clusters, along the Bayankala Mountains, which form the geological barrier between the Central and North clusters. Species distribution modeling demonstrated the range of G. aristata expanded since the Last Interglacial period. Our findings provide genetic and morphological evidence of cryptic diversity in G. aristata, and identified rampant hybridization between genetic clusters along a geological barrier. These findings suggest that geological barriers and climatic fluctuations have an important role in triggering diversification as well as hybridization, indicating that cryptic diversity and hybridization are essential factors in biodiversity formation within the QTP region.

Subjective learning gain from a simulation-based health management course: a mixed methods study.

Objectives: Simulations are increasingly being offered as part of the educational experience of healthcare students. We used a Health Management Scenario Simulation system to create a course. This study aimed to evaluate learning gains before and after the course. Methods: Based on the learning strategies of framing, simulation, and debriefing, the Health Management Scenario Simulation course lasted 4 weeks and was conducted online. Learning gain was assessed using a comparative self-assessment questionnaire administered electronically at the beginning and end of the course. We organized focus group interviews and collected quantitative data after students completed the simulations and the questionnaire. These data were subjected to descriptive statistical analysis and thematic grouping using frequency counting. Results: There were 195 health management students enrolled in the course. In total, 265 anonymously completed questionnaires were received, 141 (72.31%) on the pre-simulation and 124 (63.59%) on the post-simulation. All questionnaire item gain values were positive, except the item "I can identify common health risk factors," which showed no change. The skills domain showed the highest learning gain, ranging from 16 to 22%. Six students participated in the focus-group study. The main themes that emerged from students' reflections were learner-centeredness, competencies, and career development. Conclusion: Students acquired health management skills through the simulation, which contributed to the development of basic attitudes and skills in their professional careers. Students' comments highlighted the value of practicing health management skills in a simulated environment.

Rampant intraspecific variation of plastid genomes in Gentiana section Chondrophyllae.

Exploring the level of intraspecific diversity in taxa experienced radiation is helpful to understanding speciation and biodiversity assembly. Gentiana section Chondrophyllae sensu lato encompasses more than 180 species and occupies more a half of species in the genus. In this study, we collected samples across the range of three species (Gentiana aristata, G. crassuloides and G. haynaldii) in section Chondrophyllae s.l., and recovered the intra-species variation by comparing with closely related taxon. Using 25 newly sequenced plastid genomes together with previously published data, we compared structural differences, quantified the variations in plastome size, and measured nucleotide diversity in various regions. Our results showed that the plastome size variation in the three Chondrophyllae species ranged from 285 to 628 bp, and the size variation in LSC, IR and SSC ranged from 236 to 898 bp, 52 to 393 bp and 135 to 356 bp, respectively. Nucleotide diversity of plastome or any of the four regions was much higher than the control species. The average nucleotide diversity in plastomes of the three species ranged from 0.0010 to 0.0023 in protein coding genes, and from 0.0023 to 0.0061 in intergenic regions. More repeat sequence variations were detected within the three Chondrophyllae species than the control species. Various plastid sequence matrixes resulted in different backbone topology in two target species, showed uncertainty in phylogenetic relationship based inference. In conclusion, our results recovered that species of G. section Chondrophyllae s.l. has high intraspecific plastome variation, and provided insights into the radiation in this speciose lineage.

Phytochemical and underlying mechanism of Mikania micrantha Kunth on antibiotic resistance genes, and pathogenic microbes during chicken manure composting.

Chicken manure is a source of antibiotic resistance genes (ARGs) and pathogenic microbes. Mikania micrantha Kunth (MM) is an invasive plant containing phytochemicals as antimicrobial agents. To explore its impacts on ARGs and pathogen-host interactions (PHIs), MM was added to composting mixtures. The findings indicated that compared with control (CK), MM significantly improved the phytochemical abundances, particularly stilbenoids and diarylheptanoids (4.87%), and ubiquinones (2.66%) in the treatment (T) compost. Besides, significant ARGs reduction was noted, where rpoB2, RbpA, FosB1, vatC, and vatB were removed from T compost. PHIs significantly declined in T compost, where the growth of Xanthomonas citri, Streptococcus pneumoniae, Fusarium graminearum, Vibrio cholerae, and Xanthomonas campestris were inhibited. Multiple variable analyses demonstrated that temperature and pH revealed a significant role in ARGs and PHIs decline. Accordingly, this study considerably recommends MM as a promising compost additive in terms of its antimicrobial potential toward pathogenic microbes and ARGs.

Unraveling and characterization of novel T3SS effectors in Edwardsiella piscicida.

Type III secretion system (T3SS) facilitates survival and replication of Edwardsiella piscicida in vivo. Identifying novel T3SS effectors and elucidating their functions are critical in understanding the pathogenesis of E. piscicida. E. piscicida T3SS effector EseG and EseJ was highly secreted when T3SS gatekeeper-containing protein complex EsaB-EsaL-EsaM was disrupted by EsaB deficiency. Based on this observation, concentrated secretomes of ΔesaB strain and ΔesaBΔesaN strain were purified by loading them into SDS-PAGE gel for a short electrophoresis to remove impurities prior to the in-the gel digestion and mass spectrometry. Four reported T3SS effectors and two novel T3SS effector candidates EseQ (ETAE_2009) and Trx2 (ETAE_0559) were unraveled by quantitative comparison of the identified peptides. EseQ and Trx2 were revealed to be secreted and translocated in a T3SS-dependent manner through CyaA-based translocation assay and immunofluorescent staining, demonstrating that EseQ and Trx2 are the novel T3SS effectors of E. piscicida. Trx2 was found to suppress macrophage apoptosis as revealed by TUNEL staining and cleaved caspase-3 of infected J774A.1 monolayers. Moreover, Trx2 has been shown to inhibit the p65 phosphorylation and p65 translocation into the nucleus, thus blocking the NF-κB pathway. Furthermore, depletion of Trx2 slightly but significantly attenuates E. piscicida virulence in a fish infection model. Taken together, an efficient method was established in unraveling T3SS effectors in E. piscicida, and Trx2, one of the novel T3SS effectors identified in this study, was demonstrated to suppress apoptosis and block NF- κB pathway during E. piscicida infection. IMPORTANCE Edwardsiella piscicida is an intracellular bacterial pathogen that causes intestinal inflammation and hemorrhagic sepsis in fish and human. Virulence depends on the Edwardsiella type III secretion system (T3SS). Identifying the bacterial effector proteins secreted by T3SS and defining their role is key to understanding Edwardsiella pathogenesis. EsaB depletion disrupts the T3SS gatekeeper-containing protein complex, resulting in increased secretion of T3SS effectors EseG and EseJ. EseQ and Trx2 were shown to be the novel T3SS effectors of E. piscicida by a secretome comparison between ∆esaB strain and ∆esaB∆esaN strain (T3SS mutant), together with CyaA-based translocation assay. In addition, Trx2 has been shown to suppress macrophage apoptosis and block the NF-κB pathway. Together, this work expands the known repertoire of T3SS effectors and sheds light on the pathogenic mechanism of E. piscicida.