Staphylococcus aureus surgical site infection (SSI) is a prerequisite for implementing effective preventive interventions. The objective of this study was to develop a risk prediction model for S. aureus SSI or bacteremia after cardiothoracic surgery based on pre-operative variables.Identifying patients undergoing cardiothoracic surgery at high risk of S. aureus vaccine (V710-P003) clinical trial were analyzed. In this randomized placebo-controlled trial, the effect of preoperative vaccination against S. aureus was investigated in patients undergoing cardiothoracic surgery. The primary outcome was deep/superficial S. aureus SSI or S. aureus bacteremia through day 90 after surgery. Performance, calibration, and discrimination of the final model were assessed.Data from the Merck Phase IIb/III S. aureus infection . Independent risk factors for developing the primary outcome were pre-operative colonization with S. aureus , diabetes mellitus , BMI , and CABG . Although vaccination had a significant  protective effect, it was omitted from the model because its addition did not significantly change the coefficients of the final model and V710-vaccine development has been discontinued due to insufficient efficacy. The final prediction model had moderate discriminative accuracy .Overall 164 out of 7,647 included patients (2.1%) developed S. aureus colonization status, diabetes mellitus, BMI, and type of surgical procedure moderately predicted the risk of S. aureus SSI and/or bacteremia among patients undergoing cardiothoracic surgery.Pre-operative  Staphylococcus aureus [S. aureus SSI or post-operative bacteremia is inadequate [Surgical site infection (SSI) with or without bacteremia is a common post-operative complication responsible for increased morbidity, mortality, and health care costs–3. The ms aureus –6 which s aureus ,8. Yet tadequate . As preeadequate .S. aureus vaccine study (V710-P003) were analyzed [S. aureus on the incidence of postoperative S. aureus bacteremia and/or deep sternal wound infection in adult patients undergoing cardiothoracic surgery through postoperative day 90. V710 was not sufficiently efficacious in preventing the primary endpoint by prespecified criteria, and overall mortality rates for the placebo or vaccine group were not significantly different. The trial was stopped prematurely after interim analysis showed lack of efficacy as well as a numerically higher mortality rate in the subset of vaccine recipients developing S. aureus infections. Pre-operative S. aureus colonization status was documented as part of protocol-stipulated procedures.For this study, data from the Merck Phase IIb/III analyzed . This dopost hoc analysis of the prospectively collected data from this clinical trial, we aimed to develop a pathogen-specific risk prediction model for S. aureus SSI and/or bacteremia in patients after cardiothoracic surgery based on information ascertainable preoperatively.In the current S. aureus vaccine  were used for this post hoc analysis [Data from the randomized, double-blind, placebo-controlled trial of Merck Phase IIb/III analysis . BecauseAdult patients undergoing elective cardiothoracic surgery were eligible for inclusion. Exclusion criteria, described in more detail elsewhere, included active infection, pregnancy, and immunosuppression,12.S. aureus diagnoses: deep/superficial sternal wound infection (including mediastinitis), deep/superficial harvest site infection, and bacteremia (defined as at least one positive blood culture growing S. aureus). All cases were adjudicated by an independent committee using diagnostic criteria established by the Center for Disease Control and Prevention (CDC) [The primary outcome was a binary (yes/no) composite endpoint through day 90 after surgery, which included at least one of the following on (CDC) .S. aureus colonization status, pre-operative antibiotic use, diabetes mellitus, type of cardiothoracic procedure, body mass index (BMI), age, and sex.A list of candidate predictors was defined prior to initiating this analysis, based on clinical judgment and availability in the database, including pre-operative S. aureus carriage was documented by culture at any moment before surgery. This assumption was chosen because literature indicates that colonization status is largely dependent on the patient’s constitution and thus relatively constant over time[We defined a patient to be colonized if nasal over time.Pre-operative antibiotic use was defined as any systemic antibiotic use within 6 months before surgery, excluding pre-operative prophylaxis. A timeframe of 6 months pre-operatively was chosen, considering that previous studies had shown that the microbiome can be affected after antibiotic usage for this period of time. DiabeteS. aureus colonization status were imputed using multiple imputation techniques[Age and BMI were used as continuous variables; it was checked whether fractional polynomials improved model performance. Missingechniques.S. aureus colonization status and BMI or diabetes mellitus were performed .Univariate logistic regression analysis was performed on the mentioned variables. Variables with a univariate p≤0.157 were entered into the final multivariable model, roughly corresponding to the selection threshold based on the Akaike information criterion when considering p-values . Tests o2)[A logistic regression model was fitted with the variables described above. Overall model performance was assessed by measuring the explained variation (Nagelkerke R2). CalibraPatients might have died within 90 days post-surgery without reaching the primary outcome, which means that death is a competing event for the primary outcome. As a sensitivity analysis, a Fine & Gray model was fitted to account for the time-to-event, considering death as a competing event . SubdistConsidering that we used a slightly different primary outcome compared to the initial study , it was assessed whether a vaccine-effect was present  and whether adding vaccination to the model significantly altered the effect estimates.All statistical analyses were performed using R version 2.10.00. S. aureus SSI and/or bloodstream infection, including 122 (1.6%) patients with SSI without bacteremia, 28 (0.4%) patients with bacteremic SSI, and 15 patients (0.2%) with post-operative bacteremia without SSI.In the final analysis, 7,647 patients were included. Their baseline characteristics are described in S. aureus , diabetes mellitus , CABG , and BMI . No significant interaction was found between pre-operative S. aureus colonization and either BMI or diabetes mellitus .Several pre-operative variables were univariately associated with the primary outcome: pre-operative colonization status with Independent risk factors identified during multivariate analysis were pre-operative colonization status , diabetes mellitus , CABG  and BMI  .2 was 0.08. The distribution of predicted risks for the event of interest was highly skewed to the left, with more patients in the low risk categories than in the high-risk categories. Only 8.2% of the patients had a risk of ≥5%. Of the 209 S. aureus colonized, diabetic patients undergoing CABG , the risk of developing the event was 11% (n = 23). Of the 3012 patients without any preoperative risk factor, 28 (0.9%) developed the event.The mean explained variation of the model as indicated by the Nagelkerke R2 of 0.07 and AUC of 0.72 after correction for optimism. The Somers’ Dxy rank correlation between predicted probabilities and observed responses was 0.43 (0 indicating completely random predictions and 1 indicating perfect predictions).The stability of the final model was further assessed in 200 bootstrap samples. Using these samples, we derived an RA total number of 236 patients died within 90 days post-surgery. Of these, 229 had not yet developed the primary event of interest. Using the Fine & Gray competing risks analysis to assess whether the subdistribution hazard ratios differ from the odds ratios from the logistic regression model, the estimates did not change significantly (maximum observed change was 2%). Hence, the effect of death as a competing risk can largely be ignored.S. aureus infection , and remained so after correction for other predictors . However, other predictor estimates did not change significantly after incorporating vaccination status, indicating a lack of confounding effect. Furthermore, because the development of this specific vaccine has been discontinued, vaccination was not included as a predictor in the final model.Vaccination was univariately associated with the primary outcome. V710 was protective against S. aureus SSI and/or bacteremia after cardiothoracic surgery. We identified S. aureus colonization, diabetes, increasing BMI, and CABG surgery as independent risk factors. The final prediction model using these readily available predictors performed satisfactorily.In this analysis, we built a risk prediction model to determine which preoperative characteristics put patients at higher risk of developing S. aureus infections are scarce. Pathogen-specific prediction may be preferable, anticipating the arrival of targeted preventive measures in the near future [S. aureus infections are at substantial risk for bad outcomes and incur higher health care costs[S. aureus SSI of approximately 5000 patients across Europe undergoing different types of surgery and is currently ongoing.As the frequency and impact of post-surgical infections remain substantial, the relevance of an accurate prediction model remains. Many previous studies have developed and validated risk prediction tools for all-cause surgical site infection in cardiothoracic patients, some of which are frequently used in practice ,23. Hower future ,24–26. Fare costs–30. Thisare costs. Better are costs,32. ThisGlobal guidelines on the prevention of surgical site infection’ specifically stress the need for such a simple, inexpensive screening process, considering that in low- and middle-income countries the logistical and financial burdens that come with a screening and decolonization intervention may be too burdensome to implement on all preoperative patients [A possible option for new model developers could be to use an established, validated prediction score like Euroscore and assess whether adding pathogen-specific variables like colonization status can make the model pathogen-specific. This copatients .S. aureus colonization unlikely to occur outside the setting of a clinical trial. Last, but not least, the statistical analyses performed here, including the sensitivity analyses taking into account competing risks were sophisticated and comprehensive.A major strength of the current study is the size of the study and the number of participating countries/centers. Furthermore, data collection and patient follow-up was stipulated by protocol and closely monitored, minimizing the amount of missing data during follow-up, and ensuring a high proportion of patients screened for S. aureus were neither standardized nor documented. Decolonization methods were likely applied to colonized patients at a majority of the sites [There are several limitations to this analysis. First of all, decolonization strategies for he sites . If indehe sites ,37.S. aureus colonization, thus, carriage on skin or at other sites may have been missed. In other words, there is potential misclassification bias, since some of the “non-colonized” patients may have been colonized elsewhere. This misclassification likely would be independent of S. aureus bacteremia and SSI, giving rise to a non-differential misclassification of the S. aureus carrier status. The non-differential misclassification may have biased our estimates towards the null and reduced the discriminative effect of the new prediction model.Furthermore, in this study only nares were screened for Despite the limitations described above, the model performed moderately well. In its present form it may only be useful to indicate an especially high risk for patients having all three risk factors. For subtler prediction and external validation, further enhancement of the model is necessary.S. aureus colonization gives a 3x higher OR for S. aureus SSI / bacteremia in the unsubstantiated (but likely) presence of decolonization procedures. Without decolonization, the risk is likely to be higher. This model that included colonization status, diabetes, and CABG had overall average performance.From this analysis, we can conclude that pre-operative S1 File(DOC)Click here for additional data file.


Dugesia ryukyuensis (Tricladida). These sex-inducing substances may provide clues to the molecular mechanism of this reproductive switch. However, limited information about these sex-inducing substances is available.Turbellarian species can post-embryonically produce germ line cells from pluripotent stem cells called neoblasts, which enables some of them to switch between an asexual and a sexual state in response to environmental changes. Certain low-molecular-weight compounds contained in sexually mature animals act as sex-inducing substances that trigger post-embryonic germ cell development in asexual worms of the freshwater planarian D. ryukyuensis is useful for evaluating sex-inducing activity. We used the freshwater planarians D. ryukyuensis and Bdellocephala brunnea (Tricladida), land planarian Bipalium nobile (Tricladida), and marine flatworm Thysanozoon brocchii (Polycladida) as sources of the sex-inducing substances. Using an assay system, we showed that the three Tricladida species had sufficient sex-inducing activity to fully induce hermaphroditic reproductive organs in asexual worms of D. ryukyuensis. However, the sex-inducing activity of T. brocchii was sufficient only to induce a pair of ovaries. We found that yolk glands, which are found in Tricladida but not Polycladida, may contain the sex-inducing substance that can fully sexualize asexual worms of D. ryukyuensis.Our assay system based on feeding sex-inducing substances to asexual worms of D. ryukyuensis; namely, the crucial sex-inducing substance  produced in yolk glands.Our results suggest that within Tricladida, there are one or more common compounds or functional analogs capable of fully sexualizing asexual worms of The online version of this article (10.1186/s40851-018-0096-9) contains supplementary material, which is available to authorized users. Aurelia aurita, seasonally switches their life cycle between asexual polyps and sexual medusae , 22Dugesubstance . HoweverD. ryukyuensis, where possible, we used the land planarian Bipalium nobile  and marine flatworm Thysanozoon brocchii (Polycladida), with D. ryukyuensis and Bd. brunnea as sources of a sex-inducing substance , a natural food source for Bi. nobile, was also used  and nine microturbellarians , 25. Micnce Fig. . A slug,sed Fig. . To examD. ryukyuensis . The homogenate was centrifuged at 16000×g for 30 min at 4 °C. The supernatant, or cytosolic fraction, was filtrated using a 0.2 μm filter  and then centrifuged at 120000×g for 1 h at 4 °C. The cytosolic fraction was loaded onto a Sep-Pak® Light tC18 Cartridge  and eluted with 0, 10, and 100% aqueous methanol to create the following fractions: Fr. M0, Fr. M10, and Fr. M100. The dry weight of each fraction was measured  control. Thirty test worms were fed a piece of food daily for 4 weeks.Each of the test worms developed a pair of ovaries, testes, yolk glands, and a copulatory apparatus in sequence if the test food contained a sufficient quantity and quality of the sex-inducing substance. This morphological change allowed us to divide the sexual induction process into five distinct stages Fig.  22]. Af. Af22]. This experimental sexual induction has a point-of-no-return between stages 2 and 3. Worms at stages 1 and 2 return to the asexual state if the administration of the sex-inducing substance is stopped, whereas from stages 3 onward worms will continue to develop sexual organs, even if the administration of the sex-inducing substance is stopped, which suggests reversible and irreversible phases as evidenced by the point-of-no-return from external observations , which is a powerful enzyme for the elimination of peptides/proteins . The reaction solutions were incubated at 37 °C for 16 h, and then boiled for 15 min to deactivate Actinase E. As a control (− Actinase E), the solutions containing the Fr. M0 or M10 from approximately 4 g wet weight and 0.1% Actinase E solution were independently incubated and boiled, and finally mixed. To produce the test food for the bioassay, we mixed each dried four sample  with 200 μL of chicken liver homogenate, and then freeze-dried the mixture. Thirty test worms were fed a piece of food daily for 4 weeks.Foods were prepared for bioassay of the Fr. M0 and M10 fractions of v/v) HCl in 5/8 Holtfreter’s solution [Test worms were relaxed in cold 2% ) ) Fig. . Each prv)) Fig. .D. ryukyuensis and Bd. brunnea that had been eluted with water (Fr. M0) and 10% methanol (Fr. M10) exhibited sufficient sex-inducing activity in asexual worms of D. ryukyuensis to overcome the point-of-no-return [Bd. brunnea  did not develop reproductive organs. Concerning the crucial sex-inducing substance contained in the cytosolic fractions, the activity needed for overcoming the point-of-no-return was recognized in both Fr. M0 and M10 from ile Fig. . We alsoile Fig.  to confinea Fig. , whereasile Fig. . It shouile Fig.  and 5a–ele Table . In partle Figs. .Fig. 5HiT. brocchii (Polycladida) is a more distant species from D. ryukyuensis (Tricladida) than the other two turbellarian species. Although the administration of the Fr. M0, M10, and M100 fractions from T. brocchii induced a pair of ovaries externally in a statistically significant number of test worms, these fractions did not contain enough sex-inducing activity to overcome the point-of-no-return , did not have significant sex-inducing activity, although a pair of small ovaries became visible in a few asexual worms of D. ryukyuensis in this assay ies Fig. .D. ryukyuensis and Bd. brunnea is hydrophilic [Bi. nobile, strong sex-inducing activity was recovered in Fr. M0 and M10 , the existence of intact yolk gland cells in fresh cocoons collected within a day of deposition has been suggested by quantitative reverse transcription polymerase chain reaction analysis of a yolk gland marker gene [In some orders of turbellarians, worms have yolk glands, a reproductive organ filled with nurse cells; namely, yolk gland cells. Their eggs are ectolecithal (cocoons), which have several fertilized eggs and numerous yolk gland cells . In D. rker gene .D. ryukyuensis is present in worms, at least in Tricladida but not Polycladida. An anatomically crucial difference between Tricladida and Polycladida is the presence or absence of the yolk glands. This finding led us to examine the sex-inducing activity of yolk glands (cocoons) in freshwater planarians D. ryukyuensis and Bd. brunnea. As we expected, asexual test worms overcame the point-of-no-return when they were fed cocoons of D. ryukyuensis and Bd. brunnea daily for 4 weeks  [D. ryukyuensis, sexual induction has a point-of-no-return between stages 2 and 3  of the freshwater planarian esiidae)  is usefukyuensis . HoweverD. ryukyuensis was carried out using the freshwater planarians D. ryukyuensis and Bd. brunnea, land planarian Bi. nobile , and marine flatworm T. brocchii (Polycladida) as sources of the sex-inducing substance. A slug Ambigolimax valentianus (Mollusca) was also used.In this study, to further estimate a phylogenetic relationship of species containing the crucial sex-inducing substance, a comprehensive comparison of sex-inducing activity in asexual worms of Bd. brunnea and Bi. nobile  to produce strong sex-inducing activity in asexual worms of D. ryukyuensis (Dugesiidae) may be consistent with the aforementioned phylogenetic relationship.The present study clearly showed that in the cytosolic fractions, the probability of conspecific worms displaying sex-inducing activity was always lower than that of le Table . In partes Figs.  and 2. Mes Figs. . The abiT. brocchii  acts as a neuropeptide [A. valentianus must contain free d-Trp as a degradant of this neuropeptide. In the fractionation procedure, d-Trp is recovered primarily in Fr. M0 [D. ryukyuensis fed with the test food containing the Fr. M0 of A. valentianus developed a pair of ovaries   Fig. . This surn Figs.  and 6. TD. ryukyuensis lacking testes after treatment with the RNAi of Dr-nanos and Dr-piwi1 could not maintain their acquired sexuality [Schmidtea mediterranea [npy-8 in sexually mature worms results in the regression of the testes, which acts to maintain planarian sexuality. However, to date, the yolk gland has not been a candidate for the source of the crucial sex-inducing substance.There is much debate on the identity of the organs or tissues responsible for producing the crucial sex-inducing substance. One theory is that the putative hormone produced by the testes is responsible for the development of the copulatory apparatus . Indeed,exuality , 43. Theexuality –46. Inteterranea . The RNAD. ryukyuensis. Furthermore, the crucial sex-inducing substance needed to overcome the point-of-no return in asexual worms of D. ryukyuensis may be contained in worms of Tricladida, but not those of Polycladida. An anatomically crucial difference between Tricladida and Polycladida is the presence or absence of yolk glands. Immediately after the point-of-no-return (stage 3), primordial yolk glands emerged in D. ryukyuensis  with the sex-inducing activity in asexual worms of rn stage , primordion Fig. . Besidesion Fig. . We concA. valentianus, is a food for the land planarian Bi. nobile in nature. There were no fractions from A. valentianus that produced significant sex-inducing activity in asexual worms of D. ryukyuensis  in Turbellaria as sources of a sex-inducing substance. Using this assay system, we showed that the three species in Order Tricladida have strong sex-inducing activity and can fully sexualize asexual worms of D. ryukyuensis. Interestingly, the sex-inducing activity displayed by the conspecific sexual worms was not higher than that of the freshwater planarian Bd. brunnea or land planarian Bi. nobile, which belong to the same order. In contrast, the sex-inducing activity displayed by the marine flatworm T. brocchii, which belongs to Order Polycladida, was extremely low. On the basis of these results, we found that yolk glands, which exist in Tricladida but not Polycladida, possibly contain the crucial sex-inducing substance  that can fully sexualize asexual worms of D. ryukyuensis. The results obtained in this study will contribute to the identification of the crucial sex-inducing substance.Certain low-molecular-weight compounds found in sexually mature animals act as sex-inducing substances during post-embryonic germ cell development when the animals alternate from an asexual to a sexual state . The crucial sex-inducing substance responsible for the sexual induction of freshwater planarians has not yet been identified. An assay system that involves feeding asexual worms of the freshwater planarian Additional file 1:Table S1.. Relationship between test worm and food in the experimental sexual induction. (PDF 71 kb)Additional file 2:Table S2. Dry weight of fractions derived from the cytosolic fraction. (PDF 69 kb)Additional file 3:Table S3. Weight of precipitates and EtOAc layers. (PDF 69 kb)Additional file 4:Figure S1. Preparation of foods for the bioassay on Fr. M0 and M10 of Bd. brunnea treated with Actinase E. (PDF 303 kb)


XPG rs17655 G>C polymorphism and cancer risk has been investigated extensively, but the results remain contradictory. To get a more accurate conclusion, we performed a comprehensive meta-analysis of 60 case-control studies, involving 27,098 cancer cases and 30,535 healthy controls. Crude odds ratios (ORs) and 95% confidence interval (CIs) were calculated to determine the association of interest. Pooled analysis indicated that the XPG rs17655 G>C polymorphism increased the risk of overall cancer . Stratification analysis by cancer type further showed that this polymorphism was associated with increased risk of gastric cancer and colorectal cancer. This meta-analysis indicated that the XPG gene rs17655 G>C polymorphism was associated with increased overall cancer risk, especially the risk of gastric cancer and colorectal cancer. Further validation experiments are needed to strength our conclusion.Xeroderma pigmentosum group G (XPG), a key component in nucleotide excision repair pathway, functions to cut DNA lesions during DNA repair. Genetic variations that alter DNA repair gene expression or function may decrease DNA repair ability and impair genome integrity, thereby predisposing to cancer. The association between  Cancer-related deaths continue to rise in both developed and developing countries. In 2012, there were about 14.1 million new cancer cases and 8.2 million cancer-related deaths all over the world. Lung and breast cancer are the most common forms of cancer in human beings. Moreover, the incidences of liver, stomach and colorectal cancer are also very high in men and stomach, while cervix uteri and colorectal cancer prevail in women. Cancer is a complex disease. A variety of cancer risk factors have been recognized, such as smoking, drinking, lack of exercise, poor diet, reproductive changes, and genetic lesions  have been well studied, which participate in DNA repair, capable of preserving genetic integrity to prevent cells from malignant transformation [Genetic DNAs of living organisms are constantly subjected to various types of damages caused by environmental agents and byproducts  of cellular metabolic processes. To maintain genome integrity, human beings possess a number of systems for the prevention and restoration of DNA damage. Reduced DNA repair ability is a predisposing factor to cancer . Five coormation .ERCC5/XPG is located on chromosome 13q22-33, consisting of 15 exons and 14 introns . Its protein product is a 1,186 amino acid structure-specific endonuclease, and plays an essential role in the two incision steps of NER [XPG is highly polymorphic. Among known single nucleotide polymorphisms (SNPs) in this gene, a nonsynonymous Asp1104His  polymorphism is most frequently studied for its association with cancer risk [XPG gene rs17655 G>C polymorphism and cancer risk.cer risk –38. HoweXPG gene rs17655 polymorphism and cancer risk. And then, full texts of remaining articles were evaluated. Two publications [As shown in ications ,40 were ications –51 becauications –56 preseications –61 due tications ,31,37,38ications ,59,62–66ications ,20,25,67ications ,21,68,69ications ,70–74, fications ,75–77, eications ,35,78–80ications ,81,82, tications ,83, laryications , and proications ,28. Moreications , hepatocications , esophagications , oral sqications , sarcomaications  and braiications . Among tXPG gene rs17655 G>C polymorphism was associated with increased overall cancer susceptibility . Stratification analysis further indicated that the XPG gene rs17655 G>C polymorphism was associated with increased risk of gastric cancer  and colorectal cancer  . Moreover, the association remained significant in the subgroups with quality score ≤ 9  and hospital-based studies .As we can see in P=0.011) . We alsoP=0.808; CG vs. GG: P=0.050; CC vs. CG+GG: P=0.806; CG+CC vs. GG: P=0.047; C vs. G: P=0.240).Symmetry in the funnel plot  suggesteXPG gene rs17655 G>C polymorphism and cancer risk based on 60 eligible case-control studies with a total of 27,098 cancer cases and 30,535 healthy controls. Pooled risk estimates revealed that this polymorphism was significantly associated with an increased risk of overall cancer, especially with the risk of gastric cancer and colorectal cancer.In the current meta-analysis, we estimated the association between the XPG gene, one of the eight vital genes in the NER pathway, is responsible for recognizing and excising DNA lesions on the 3’ side [XPG gene over the past decades, among which the rs17655 polymorphism has revoked great attention for its association with cancer risk. The rs17655 polymorphism, leading to the replacement of aspartate with histidine at codon 1104 in ERCC5 protein, may cause an alteration in the protein function, thereby likely affecting DNA repair ability, genome integrity, and cancer predisposition.The etiology of cancer is multifactorial . Abnorma 3’ side . Loads oXPG gene, consisting of 177 patients and 237 controls. They found that the rs17655 polymorphism was associated with an increased risk of gastric cancer. This association was reconfirmed in different types of cancer, including breast cancer by Hsu et al. [XPG rs17655 polymorphism might be associated with reduced gastric cancer risk. Additionally, Ruiz-Cosano et al. [http://compbio.cs.queensu.ca/F-SNP/) [in vitro and in vivo data are needed to elucidate biological function of this variant.Numerous studies were performed to explore the association between the rs17655 polymorphism and the risk of various types of cancer. Feng et al.  carried u et al.  with 401u et al.  with 878u et al.  revealedo et al.  reportedo et al.  selectedo et al. –91. As co et al. . Moreove/F-SNP/) ; howeverThere are advantages that strengthened the robustness of our findings. First, we searched five databases to include most of the publications written in English or Chinese. The large sample size provided adequate statistical power. Second, stratified analyses were performed by cancer type, quality score, and source of control. Third, we used the Begg’s funnel plot and Egger’s linear regression test to assess the possible publication bias.However, several limitations still existed in this meta-analysis. Firstly, selection bias might occur because only publications written in English or Chinese were included. Researches in other languages were missed. Secondly, the number of individual studies for some cancer types, like HNC and prostate cancer (<5 studies), may be inadequate. Third, more than half of included studies had relative low quality scores (≤ 9). Our results should be interpreted cautiously. Further studies with high quality scores are needed to verify the real association.Additionally, age, sex, living habits, virus infections or some environmental factors may also influence cancer risk. Our findings based on unadjusted estimates for lack of access to original data might suffer from potential confounding bias. Therefore, the results should be interpreted with caution. Finally, lack of biological evidence of the implication of the rs17655 polymorphism in cancer is also a drawback of the study. Mechanistic studies of the rs17655 polymorphism with cancer should be performed in the future.XPG rs17655 G>C polymorphism is significantly associated with an increased overall cancer risk, especially with the risk of gastric cancer and colorectal cancer. Moreover, large-scale, well-designed studies in different cancers should be conducted to corroborate our findings.In conclusion, this meta-analysis suggests that the ERCC5 or XPG”, “polymorphism or variant”, and “cancer or carcinoma or neoplasm or malignance” in PubMed, EMBASE, CNKI, WANFANG, and Vip databases . We also manually searched the references of the retrieved publications for additional relevant eligible studies.We searched for relevant articles using the following terms: “XPG gene rs17655 polymorphism and the risk of one or more types of cancer; (3) case-control study. If studies had overlapping subjects, the publication including the largest number of individuals were selected.The publications contained in the meta-analysis had to meet the following criteria: (1) the study was only written in English or Chinese; (2) the study investigated the association between the Exclusion criteria were as follows (1) the study did not report sufficient genotype data to calculate odds ratio (OR) and 95% confidence interval (CI); (2) the study included survival data only. (3) the genotype frequencies of the rs17655 G>C and other polymorphisms were deviated from Hardy-Weinberg equilibrium (HWE) in the controls.Two investigators (Chen SS and Zhao J) extracted the following information from each publication independently: first author, publication year, cancer type, country of origin, race, genotyping method, source of controls , the genotype counts of cases and controls for the rs17655 G>C polymorphism. We also calculated the score of each publication based on the quality score assessment as described before . All conXPG rs17655 G>C polymorphism and overall cancer risk under the homozygous (CC vs. GG), heterozygous (CG vs. GG), recessive (CC vs. CG+GG), dominant (CG+CC vs. GG), and allele contrast (C vs. G) models. We carried out stratification analyses by cancer type , score (>9 and ≤9), and study design . We also calculated between-study heterogeneity using the Chi square-based Q-test. When P>0.1 indicating lack of heterogeneity, a fixed-effect model was adopted. Otherwise, a random-effect model would be applied [P values were two-tailed. P<0.05 was considered statistically significant. All data analyses were performed by the STATA software .We evaluated crude ORs and 95% CIs to assess the association between  applied . The pot applied . All of Supplementary File


H. pylori in the human stomach cause chronic gastritis, peptic ulcers, and gastric cancers. H. pylori infection also induces insulin resistance and has been defined as a predisposing factor to T2D development. Gastric and fecal microbiota may have been changed in H. pylori-infected persons and mice to promote gastric inflammation and specific diseases. However, the interaction of H. pylori and gut microbiota in regulating host metabolism also remains unknown. Further studies aim to identify the H. pylori-microbiota-host metabolism axis and to test if H. pylori eradication or modification of gut microbiota can improve the control of human metabolic disorders.As a barrier, gut commensal microbiota can protect against potential pathogenic microbes in the gastrointestinal tract. Crosstalk between gut microbes and immune cells promotes human intestinal homeostasis. Dysbiosis of gut microbiota has been implicated in the development of many human metabolic disorders like obesity, hepatic steatohepatitis, and insulin resistance in type 2 diabetes (T2D). Certain microbes, such as butyrate-producing bacteria, are lower in T2D patients. The transfer of intestinal microbiota from lean donors increases insulin sensitivity in individuals with metabolic syndrome, but the exact pathogenesis remains unclear.  The human gut, including the stomach and intestine, is inhabited by a vast number of microorganisms that host microbiota . Gut micH. pylori can persistently colonize the gastric epithelium by interacting with bacterial adhesion molecules and gastric receptors [H. pylori infection have been reported to change the microbial-origin fatty acid and lipid profiles in host blood [H. pylori infection was shown to disturb the carbohydrate and amino acid metabolism of the host [H. pylori. However, the exact mechanism remains unclear. This review article focused on the metabolic interaction between gut microbiota and H. pylori, as well as the related consequences of such interaction on the host’s health.eceptors ,10. The st blood . In an athe host . In addiet al. demonstrated that a single segmented filamentous bacterium can induce effector T cell differentiation in the lamina propria [et al. reported that intestinal macrophages could sense microbial signals to induce the RORγt+ innate lymphoid cells (ILCs) to produce colony-stimulating factor 2 (CSF2), thereby promoting intestinal homeostasis [Gut microbial communities are known to be inherited from the mother ,3. Sever propria . Other r propria ,16. Morteostasis . All of eostasis .et al. showed that significant concentrations of microbial long chain fatty acids (LCFAs) were present in human blood and corresponded to specific microbial compounds [Bacteroides and Bifidobacterium possessed abundant polysaccharides and starch breakdown genes [Crosstalk and signaling between the host and microbiota occur at both the cellular and metabolic levels. Ktsoyan ompounds . Furtherwn genes ,19,20.et al. demonstrated that the most common butyrate-producing bacteria were Clostridium spp., particular in clusters XIVa and IV in the human feces [et al. revealed that microbe-derived SCFAs, particularly butyrate, stimulated epithelial metabolism and decreased intracellular O2, resulting in the stabilization of the transcription factor hypoxia-inducible factor-1 (HIF-1) and epithelial barrier function [et al. reported that rats fed a high butyrate-producing fiber diet (wheat bran) had significantly fewer tumors and less tumor mass than those given low butyrate-production fiber diet [The other important energy source of bacterial colonizers of the epithelium, especially butyrate fermenting bacteria, is short chain fatty acids (SCFA) . Microbean feces . Decreasfunction . In a raber diet . These ret al. transferred intestinal microbiota from lean donors to recipient males with metabolic syndrome. Six weeks later, the insulin sensitivity of the recipients increased along with levels of butyrate-producing intestinal microbiota [An altered balance between gut microbiota and the host contributes to a spectrum of immune, inflammatory, and metabolic disorders. A metagenome-wide association study using deep shotgun sequencing of the gut microbial DNA demonstrated that type 2 diabetes (T2D) in Chinese patients had moderate degrees of gut microbiota dysbiosis, particularly decreased butyrate-producing bacteria . Based ocrobiota .et al. used a type 1 diabetes (T1D) non-obese diabetic (NOD) mice model to suggest that signaling through the MyD88 adaptor was critical for T1D development. This effect depended on commensal microbes because germ-free MyD88-negative NOD mice developed robust diabetes [Wen diabetes . MoreoveH. pylori can cause chronic gastritis and peptic ulcer disease [H. pylori as a group I carcinogen, emphasizing its association with gastric cancer [H. pylori have also been associated with several extra-gastric diseases like iron deficiency anemia, idiopathic thrombocytopenic purpura, and childhood growth [H. pylori and diseases is still being debated, bacterial eradication results in long-term benefits [H. pylori infection was also related to lipid and glucose metabolism [H. pylori-seropositive exhibited significantly higher low-density lipoprotein (LDL) cholesterol levels and significantly lower high-density lipoprotein (HDL) cholesterol levels than H. pylori-seronegative subjects [et al. suggested that lower HDL cholesterol was associated with H. pylori infection but did not correlate with the severity of coronary atherosclerosis [H. pylori can prevent or improve hyperlipidemia and atherosclerosis.Identified by Marshall and Warren in 1984,  disease ,28. The c cancer . H. pylod growth ,31,32. Abenefits ,34. Recetabolism ,36. A lasubjects . Jia et clerosis . A randoH. pylori on the host’s glucose metabolism remains controversial. In a hospital-based study, dyspeptic patients with chronic H. pylori infection had significantly higher HbA1c level and type 2 diabetes in participants older than 65 years of age, but had decreased insulin secretion and sensitivity in participants younger than 45 years old [H. pylori infection was associated with higher HbA1c, but not self-reported diabetes [The influence of ears old . A largediabetes . Howeverdiabetes .H. pylori-induced insulin resistance and diabetes development were studied in a mice model. Zhou et al. reported that H. pylori infection caused hepatic insulin resistance by mediating the c-Jun/miR-203/SOCS3 signaling pathway [H. pylori eradication on glycemic control. Vafaeimanesh et al. treated 93 consecutive patients with type 2 diabetes mellitus (T2DM) and found that H. pylori eradication had no role in glycemic control [et al. suggested that the eradication of H. pylori in T1DM patients could lead to a short-term decrease in HbA1c level [H. pylori in patients with insulin resistance and diabetes are still not routinely done in clinical practice. Further large-scale investigations are needed to clarify the association of H. pylori and diabetes.The underlying mechanisms of  pathway . However control . In cont1c level . HoweverH. pylori and gut microbiota both influence the host’s metabolism. Stearns et al. studied bacterial bio-geography from different GI sites in four healthy volunteers. Their results revealed larger Shannon and phylogenetic diversities in mouth samples than in samples from the stomach, duodenum, and colon [H. pylori infection on changes in stomach or colon microbiota has been studied in animals and humans [et al. investigated stomach microbes by deep sequencing of 16S-rRNA in rhesus macaque pre- and post-inoculation of H. pylori. They found no significant difference in the average relative abundance of non-Helicobacter taxa in the antrum or corpus [et al. demonstrated an increment in Enterococcus spp. and Staphylococcus aureus in the stomach and duodenum, and a downshift of Lactobacillus spp. in the stomach after H. pylori inoculation in Mongolian gerbils [et al. demonstrated that mice infected with immuno-pathologic H. pylori B8 could increase the luminal load of E. coli and enterococci in the cecum and of Bacteroides/Prevotella spp. in the colon [nd colon . They ald humans ,48,49,50r corpus . A cultu gerbils . In the H. pylori-infected and non-infected persons [et al. found that the gastric bacterial composition is similar between H. pylori-infected and non-infected subjects at the phylotype level [et al. reveal marked differences in the structure of gastric bacterial community in H. pylori-infected stomach, with an increase in Proteobacteria and a decrease in Actinobacteria, Bacteroidetes, and Firmicutes [H. pylori infection and changes in gastric and colonic microbiota is still being debated, more studies on the influence of interactions between commensal colonization and the microbiota–host–environment are critical for understanding how H. pylori influences the gut microbiota and human health [Using deeper sequencing methods, human studies have tried to determine the differences in gastric microbiota between  persons ,50. Bik pe level . In contn health ,52.Faecalibacterium and an increase in the genus Bacteroides in adults with CD and active disease [Crohn’s disease (CD) and ulcerative colitis (UC) are both immune-related chronic inflammatory bowel diseases (IBD). Clinical trials have shown that a dysbiosis of fecal microbiota, including changes in microbial diversity and an abundance of certain genus are closely related to the development and maintenance of IBD in adults and children ,54,55,56 disease . Unlike Clostridium difficile infections have good responses in colonic and frozen-capsulized fecal microbiota transplantation (FMT) [et al. reported that FMT enema once weekly for six weeks significantly improved disease remission by week 7 in patients with UC (24% vs. 5%) [Recurrent and serious on (FMT) ,58. Moay vs. 5%) . MoreoveH. pylori infection and IBD are both related to changes in gut microbiota, is there a link between H. pylori infection and IBD through alterations in gut microbiota? Luther et al. conducted a meta-analysis and concluded a significantly lower rate of H. pylori infection in IBD patients than in controls (27.1% vs. 40.9%), with a relative risk of 0.64  [H. pylori protection in IBD were fully discussed by Papamichael et al. [H. pylori can induce a Treg response and down-regulate the pro-inflammatory Th1/Th17 pathway on the gastric mucosa and in peripheral blood [If 54–0.75) . The potl et al. . Like geal blood ,62,63.H. pylori DNA or live bacteria to experimental mice can ameliorate colonic inflammation induced by dextran sulfate sodium (DSS) and Salmonella, respectively [H. pylori infection against IBD, evidence on the increased development of IBD in patients after the eradication of H. pylori is needed.Theoretically, such immune changes may be beneficial in protecting against the development of CD, a characteristic disease that up-regulates type 1 helper-T immune responses . In animectively ,66. Protectively . HoweverH. pyloriHelicobacter species (NHPH) and human gastrointestinal diseases, including Crohn’s disease, hepatobiliary disease, and gastritis [Helicobacteraceae were detected by PCR sequencing in 59% of fecal samples from children with Crohn’s disease, compared to 9% in healthy controls [Helicobacter spp. in intestinal specimens [H. pylori and IBD, should be investigated in the future. Nagalingam et al. reported that H. hepaticus triggered colonic inflammation only in conventional raised IL-10−/− C57BL/6 mice, but not in germ-free mice [H. hepaticus-induced colonic inflammation. Moreover, H. hepaticus infection altered the structure of intestinal microbiota even with prior antibiotic treatment.Recent studies focused on the association between non-astritis ,69,70. Eree mice , implyinet al. demonstrated that H. hepaticus-induced colonic inflammation was different in the IL-10−/− C57BL/6 mice obtained from two facilities. They further revealed different susceptibilities of H. hepaticus-induced colonic inflammation that originated from distinct microbiota among the groups. Further studies are therefore warranted in order to compare the discrepancy in microbiota structure between pre- and post-H. pylori eradication, and to investigate the structure susceptible to IBD development after H. pylori eradication.Interestingly, Yang H. pylori infection [H. pylori alters gut microbiota, with simultaneous changes in carbohydrate and amino acid metabolisms [et al. demonstrated that H. pylori-infected specific pathogen-free (SPF) mice had elevated levels of leptin, insulin, and peptide YY metabolites. Nonetheless, their growth curves remained inconclusive. Interestingly, germ-free mice with H. pylori infection also have increased levels of ghrelin and peptide YY, but with concomitant weight loss and malnutrition [H. pylori infection alters the composition of fecal Bifidobacterium/E. coli in children, which, in turn, is improved by ingesting probiotics-containing yogurt [H. pylori increases the growth in weight and height among children, together with increased serum acylated ghrelin level [H. pylori infection alters host metabolism and that commensal microbiota can attenuate the growth delay caused by H. pylori. The detailed mechanisms of metabolic interaction between H. pylori and gut microbiota in hosts warrant more studies.Although numerous studies suggest that gastric and colonic microbiota communities are changed after nfection ,47,50, oabolisms . Recentlutrition . Experieg yogurt . Lastly,in level . These fH. pylori and gut microbiota regulate the host metabolism. Some clinical observations and animal studies support the relationship of H. pylori infection and dysbiosis of gut microbiota to metabolic disorders like insulin resistance and diabetes [H. pylori and gut microbiota interact with each other to regulate the host metabolism remains unknown. Further studies are warranted to identify the H. pylori–microbiota–host metabolism axis and to test if the eradication of H. pylori or the modification of gut microbiota can control or treat human metabolic disorders.Both diabetes ,25,26. H


Poa pratensis L. is a turf grass and forage crop used worldwide. Being a facultative apomictic species, P. pratensis has a highly variable chromosome number. Chromosomal markers constitute a powerful tool for chromosome identification and for various aspects of genomic research. However, currently, no chromosomal markers are available for P. pratensis.Poa pratensis. Three tandemly repetitive sequences  were characterized as subtelomeric. Monomers of 318 bp, 189 bp and 189 bp were identified in PpTR-1, PpTR-2, and PpTR-3, respectively. One tandemly repetitive sequence (PpCR-1) was shown to be centromeric or pericentromeric, and it had a monomer of 27 bp. The distribution patterns of PpTR-1, PpTR-2, and PpTR-3 were highly conserved across different P. pratensis cultivars and in the distantly related Poa species, whereas PpCR-1 was conserved across different P. pratensis cultivars, but less conserved across Poa species.Four novel chromosome markers were isolated from a screen of Cot-1 DNA libraries, combined with fluorescence in situ hybridization (FISH) in P. pratensis. These chromosomal markers are powerful tools for accurate assessment of chromosome count, genomic and phylogenetic analyses, as well as studies of apomixis in P. pratensis.In this study, we report the identification and characterization of four novel chromosomal markers in The online version of this article (doi:10.1186/s13039-017-0307-7) contains supplementary material, which is available to authorized users. Poa pratensis L. (Kentucky bluegrass), which belongs to the Gramineae family, is a perennial herbaceous plant with strong regenerative ability, high fecundity, cold resistance, drought tolerance, and rapid colonization [nization . It is unization , 3. Thronization –6.Poa were determined by conducting chromosome counts of root tip cells. However, given their relatively small size, high preponderance and morphological similarity, the chromosome number of P. pratensis is difficult to accurately quantify [. Flow cytometry is the commonly used method for the measurement of DNA content in somatic cells of P. pratensis to determine the ploidy level [P. pratensis as an accurate method to evaluate ploidy level in a large number of individual plants [Traditionally, the ploidy levels in quantify . Flow cydy level . This apl plants , 10. Howl plants .Brachypodium pinnatum chromosomes could be accurately identified using rDNA-based and species-crossing BAC-based FISH probes [P. pratensis.The repetitive sequences are commonly used as probes in FISH (fluorescence in situ hybridization) assay for various applications, including identification of individual chromosomes, study of karyotype evolution, and screening for chromosome aberrations –14. AmonH probes . HoweverMedicago sativa), and this approach was shown to be the most accurate to identify alfalfa chromosomes [Cot-1 DNA is enriched with highly and moderately repetitive sequences. The labelled Cot-1 DNA could be used (as a probe) to localize heterochromatin in chromosomes . A numbeomosomes .P. pratensis Cot-1 library and subsequently conducted FISH screening with labelled clones on mitotic chromosomes. The clones showing strong hybridization signals were further investigated and several distinct chromosomal markers were identified. In this paper, we discuss the application of these chromosomal markers for the determination of ploidy level, assessment of genome composition, phylogenetic analysis, and identification of different types of apomixis.In this study, we first constructed a Poa cultivars and five introduced P. pratensis cultivars were used in this study  were identified by blue/white screening, as per the manufacturer’s instructions.The procedure for Cot-1 DNA library construction was adopted from Yu et al. , with miP. pratensis, as described by Fukui et al. [The 5S rDNA was amplified by polymerase chain reaction (PCR) using genomic DNA of i et al. . Insertsi et al. . The pWrRoot tips, with a length of approximately 1–2 cm, were excised and pre-treated in ice-cold water at 0 °C for 24 h, and fixed in ethanol:glacial acetic acid  for 24 h at room temperature. Each root tip was squashed in a drop of 45% acetic acid. The slides were kept at -80 °C in the Ultra-low temperature freezer for more than half an hour.FISH experiments were carried out as described by Dou et al. , with miCot-1 DNA cloned products were sequenced by Sangon Biotech Co., Ltd. . The DNAman software package  was used to analyse the sequence data. Sequence similarities were queried against the NCBI nucleotide database using BLASTN.P. pratensis interphase nuclei and mitotic metaphase chromosomes. The results showed that 14 clones, accounting for 31.8% of those tested, produced distinct “dot” or “block” hybridization signals on the chromosomes. To characterize the signals, we categorized the positive clones into two types, according to their chromosomal distribution . Type 2 showed hybridization signals on the subtelomeric or telomeric regions; most of the clones (92.3%) belonged to this group.All 14 clones that we screened were subjected to sequencing. To characterize the sequences of the inserts, we performed homology searches against the existing nucleotide sequences in the NCBI database. The search results revealed no significant sequence similarities. Sequence comparisons revealed that some screened clones had high homology to each other. Finally, data analysis revealed four unique sequences among the 14 positive clones, namely, clone 1, clone 6, clone 23 and clone 94.PREMIER Biosoft international, Canada)  across different cultivars, and each targeted chromosome carried only one PpCR-1 hybridization site  across different cultivars  were parameters used to more accurately evaluate the variability and conservation of repeat distribution across cultivars. The statistical analyses showed that the cytogenetic characters of ‘Qinghai’ were significantly different from those of the other cultivars  in all investigated cultivars except P. pratensis ‘Qinghai’, and PpCR-1 exhibited the lowest percentage of variation. This result suggests that the total chromosome number can be accurately determined by counting PpCR-1 hybridization signals. PpCR-1 is a useful chromosomal marker for chromosome number determination in P. pratensis.The chromosome number of 8 to 154 –7. The oP. pratensis, despite a highly variable number of chromosomes [P. pratensis in Missouri, USA suggested an allopolyploid origin of the species [P. pratensis is, at least, partly allopolyploid [P. pratensis from Qinghai, China, showed distinctly fewer PpCR-1 and PpTR-3 sites, compared with cultivars from the USA. This result implies a possible large genomic divergence between these cultivars. The genome composition of Qinghai P. pratensis presents as more likely to be allopolyploid than those of cultivars from the USA. This observation means that the ability to produce polyploidy with facultative apomictic reproduction may make the genome composition of P. pratensis more diverse and complex. Chromosomal markers, using comparative FISH analyses, would be helpful in elucidating plant genome composition in phylogenetic analysis. Comparative karyotyping of Brachypodium pinnatum was done using cross- species BAC-FISH [Poa species using the chromosomal markers identified in this study may provide valuable information about the origin of P. pratensis.A basic genome with a chromosome number of seven was determined in omosomes . Observa species . Analysiolyploid . In thisBAC-FISH . PhylogeBAC-FISH . Thus, cPoa pratensis has been described as an aposporous and pseudogamous facultative apomictic species [P. pratensis plants in P. pratensis cultivars and core accessions [P. pratensis has been confirmed in a five-locus model with differences in gene expressivity and penetrance [P. pratensis plants can be molecularly and cytogenetically studied by comparing the cytotypes of the parents and progeny, and apomictic inheritance analysis may be aided by finding apomixis-associated chromosomes or chromosome regions. species . Flow cycessions , 20. Thenetrance . HoweverPoa pratensis. Three  were characterized as subtelomeric, and one (PpCR-1) was centromeric or pericentromeric. The chromosomal markers, PpTR-1, PpTR-2, and PpTR-3, were stably detected across different P. pratensis cultivars and in the distantly related Poa species. However, PpCR-1 was conserved across various P. pratensis cultivars but less conserved across Poa species. These chromosomal markers will be powerful tools for chromosome counting, genomic and phylogenetic analyses, and studies of apomixis in P. pratensis.In this study, we report four novel chromosome markers that were developed by screening Cot-1 DNA libraries, combined with FISH assay in


Helicobacter pylori infection is associated with chronic gastritis, peptic ulcer, and gastric carcinoma. Due to the increased side effects of the treatment regimens and the development of antimicrobial resistance, a number of natural compounds have been tested as potential alternatives. In this review, we will examine the current knowledge on the effect of Citrus fruits and their derivatives against H. pylori, highlighting the remaining outstanding questions on the development of novel therapeutic strategies.It is known that Citrus plants  Swingle) or between Citrus aurantium L. (sour orange) and Citrus limon L. Burm. f. (lemon) or a mutation of the latter [ Citrus species flavanone rutinosides and neohesperidosides derived from naringenin, eriodictyol, and hesperetin. Commercial pectinolytic and cellulolytic enzymes were able to hydrolyze the flavonoid glycosides and the pectic oligosaccharides present in bergamot peel [ Escherichia coli, Pseudomonas putida, and Salmonella enterica [ Citrus bergamia derivatives are summarised in tivation . It is ce latter . Bergamoe latter . Recentle latter  and antie latter , 41 effee latter , 43, ante latter , 44–47, e latter –31, 48, e latter , 50 havemot peel . We havemot peel . A prebimot peel . The resHelicobacter pylori is a microaerophilic Gram-negative bacterium able to colonize the human gastric mucosa. It has been identified as group 1 carcinogen by the International Agency for Research on Cancer and it is known to be associated with chronic gastritis, peptic ulcers, atrophic gastritis, intestinal metaplasia, and lymphoma or cancer development [ H. pylori, with prevalence rates ranging from 20% to more than 80% in certain countries [ H. pylori remains unclear, the faecal-oral route represents an important pathway for its spread [elopment , 55. Altountries . The estountries . Althougs spread . H. pylori virulence, the cytotoxin-associated gene (cagA) and the vacuolating cytotoxin gene (vacA) play a crucial role. VacA, which is present in all H. pylori strains, contains at least two virulence parts [ cagA, not present in every H. pylori strain, is a marker for a pathogenicity island (PAI) [ cagA seems to be associated with more severe clinical outcomes and an increased risk of developing gastric cancer and peptic ulcer [ cagA and vacA signal sequence type s1 has been reported with strains carrying s1 m1 mosaic combination being able to secrete a vacuolating cytotoxin [ H. pylori antigens to the gastric mucosal barrier demonstrated a deleterious dose-influence effect of the lipopolysaccharide (LPS) [ H. pylori uses a combination of antimicrobial and antiacid agents [ H. pylori infection in humans and to the increased number of side effects to common treatments, more effort is needed to find novel natural or synthetic anti-H. pylori agents. The high infection rate of H. pylori in low-income countries, together with the poor management in the application of antibiotics, has increased the need for finding novel anti-H. pylori agents from medicinal plants. As a result, several naturally occurring substances have been investigated as potential alternatives for the treatment of H. pylori infection [ H. pylori [ H. pylori and its complications [ H. pylori infection.Amongst the genetic determinants causingce parts , whereasnd (PAI) . In partic ulcer . Type s1ytotoxin , 62. A rde (LPS) . The curd agents . A tripld agents . Neverthnfection –68. For . pylori . A recenications . Clinica Citrus intake has been associated with a lower risk of stomach cancer [ H. pylori infection [ Citrus fruits and their derivatives against H. pylori both in vitro and in vivo.It is well established that diet plays a crucial role in gastric carcinogenesis andh cancer , 71. Outnfection . Therefo H. pylori in vitro and the potential therapeutic combination between bergamot juice and the antibiotics AMX, CLA, and MTZ [ H. pylori (ATCC 43504 and ATCC 49503) and thirty-two clinical isolates recovered from gastric biopsy samples of dyspeptic adults undergoing digestive endoscopy. The 2.5% (v/v) concentration of BJ inhibited the growth of the clinical isolates tested by 50%, while 5% of BJ reduced H. pylori viability to 90%. Furthermore, concentration dependent killing was observed with BJ against all the tested strains. A >3log10 difference in colony-forming unit (CFU) between BJ 1.25% and BJ 20.0% with H. pylori ATCC 49503 was achieved after 2 h, whereas complete killing was achieved with BJ (10% and 20%) after 8 h and with BJ (5%) after 24 h. BJ gave less killing against H. pylori ATCC 43504, with a complete bacterial killing after 6 h exposure (20% BJ). This outcome could be explained by the increased antibiotic resistance of H. pylori ATCC 43504 compared to H. pylori ATCC 49503 [10 for H. pylori ATCC 49503 after 8 h and for H. pylori ATCC 43504 after 24 h exposure to BJ. The association of BJ with the three reference antibiotics was more effective against H. pylori ATCC 49503 than H. pylori ATCC 43504. The most effective combination was BJ and CLA against the clinical isolate HP6 and HP61, with more rapid bacterial killing achieved with the cagA-positive strain (HP6).We have recently investigated the effect of bergamot juice against and MTZ . We used H. pylori. BJ was effective against H. pylori strains, both alone or in combination with antibiotics, and could therefore be used as novel strategy for the treatment of antibiotic resistance. Although the exact mechanism of action is not known, we believe that the synergistic effect may be due to the initial damage of the microbial lipid membrane by the plant compound, which would increase the permeability of the bacteria to the antibiotic.This data confirmed the combination between BJ and the three reference antibiotics AMX, CLA, and MTZ had a synergistic effect against ATCC and clinical isolates of Citrus fruits and derivatives against H. pylori: two extracts (hexane and acetone) obtained from the leaves of Citrus unshiu, Citrus sinensis, Citrus paradisi, and Laurus nobilis demonstrated antimicrobial activity in vitro against a clinical isolate of H. pylori, with zone of inhibition diameters ranging within 0–30 mm and minimal inhibitory concentration (MIC) values of 1 : 512–1 : 4096 dilutions [ H. pylori: Citrus reticulata had MIC values close to 60 μg/mL [Other authors have also reported on the activity ofilutions . The ant60 μg/mL . Citrus lemon essential oil and its major compounds limonene and β-pinene and their activity in vitro against H. pylori ATCC 43504: results showed a gastroprotective effect of Citrus lemon and limonene with MIC values of 125 and 75 μg/mL, respectively. The role of β-myrcene, a minor constituent of essential oil from Citrus aurantium, in preventing peptic ulcer disease has been evaluated: results showed β-mircene was an effective inhibitor of gastric and duodenal ulcers through an increase in the levels of gastric mucosa defence factors [O-glucoside, produced by fungal enzymatic conversion of Citrus hesperidin, was active in vitro against H. pylori, also inhibiting human intestinal maltase and HMG-CoA reductase [ Citrus fruits, such as 4′-geranyloxyferulic acid and boropinic acid, as well as sudachitin and 3′-demethoxysudachitin, were found active against H. pylori in vitro [Rozza et al.  have rep factors . Hespereeductase . Other bin vitro , 79. H. pylori colonization of the gastric mucosa showed that a dietary supplementation with 50 ppm of auraptene suppressed H. pylori colonization, although it did not reduce gastric inflammation [ H. pylori colonization, expression of CD74, and production of proinflammatory mediators in mice [ H. pylori infection [In vivo studies on the effect of auraptene onammation . The att in mice , 82. Thenfection . H. pylori is a crucial enzyme able to protect the bacteria within the acidic environment of the human stomach. The Iranian Citrus aurantifolia and Citrus aurantium extracts revealed a strong urease inhibitory activity, with IC50 values of 432 and 465 μg/mL [A number of studies have investigated the effect of plant extracts against urease activity , 85. It 65 μg/mL , 85. Citrus fruits and their bioactive compounds could represent appropriate sources for the development of novel effective compounds against H. pylori, additional in vivo toxicological evaluations are warranted. Intervention studies should also take into account the fact that the pharmaceutical doses supporting a protective effect may derive from a synergistic action of the numerous bioactive compounds present in a plant extract. Tables H. pylori activity of Citrus fruits derivatives and their compounds.Even though the data presented indicate thatH. pylori agents has been forced by the increased side effects to common treatments and the spread of antibiotic resistance. The use of plant extracts could help control H. pylori infection in low-income countries, where the rate of infection is particularly high. However, more clinical investigations evaluating the toxicity, chemical or physical stability, and bioavailability of plant bioactives are warranted. Several studies have evaluated the effectiveness of Citrus fruits derivatives and their bioactive compounds against H. pylori, gastric carcinoma, and urease activity, suggesting that, either alone or in combination with antibiotics, they could represent useful sources to help eradicate H. pylori and avoid gastric ulcer relapse.The renewed interest in exploring natural sources of alternative anti-


Plasmodium have potential to inform on transmission dynamics, but few studies have evaluated this on a local spatial scale. We used microsatellite genotyping to characterise the micro-epidemiology of P. vivax and P. falciparum diversity to inform malaria control strategies in Timika, Papua Indonesia.Genetic analyses of P. falciparum and P. vivax isolates from a cross-sectional household survey and clinical studies conducted in Timika. Standard population genetic measures were applied, and the data was compared to published data from Kalimantan, Bangka, Sumba and West Timor.Genotyping was undertaken on 713 sympatric HE = 0.847 vs 0.625; p = 0.017) and polyclonality  were observed in P. vivax versus P. falciparum. Distinct P. falciparum substructure was observed, with two subpopulations, K1 and K2. K1 was comprised solely of asymptomatic infections and displayed greater relatedness to isolates from Sumba than to K2, possibly reflecting imported infections.Higher diversity (P. vivax versus P. falciparum to control measures, and risk of distinct parasite subpopulations persisting in the community undetected by passive surveillance. These findings highlight the need for complimentary new surveillance strategies to identify transmission patterns that cannot be detected with traditional malariometric methods.The results demonstrate the greater refractoriness of  Anopheles vector distributions, and ethnic and socio-economic features [Indonesia has one of the highest burdens of malaria in Southeast Asia. Despite recent progress in reducing case incidence, more than a quarter of a million clinical cases were reported in 2013 with 230 million people at risk of infection [Plasmodium that commonly infect humans are present in Indonesia [P. falciparum has historically been the predominant species, in recent years the proportion of P. vivax cases has risen [P. vivax species, particularly in Papua Province [Further complicating the picture, all five species of ndonesia . Differeas risen . This trProvince , 5.P. vivax and P. falciparum diversity and structure, and inform associated patterns of transmission intensity, foci and stability, thus complementing traditional malariometric surveillance [P. vivax and P. falciparum transmission between four sites in Indonesia with varying endemicity, highlighting regional heterogeneity within and between species [Population genetic approaches can define eillance –9. A rec species . However species , and litP. falciparum and P. vivax circulating in Timika, the capital of Mimika district, Papua Province. The region is an epicentre for antimalarial resistance [The aim of this study was to describe the population diversity and structure of co-endemic sistance , 5 and tP. vivax and 5.2% due to P. falciparum) [The study was conducted in Timika, Papua Province in eastern Indonesia, which has the greatest burden of malaria in the country. Timika is located within Mimika Baru, one of the largest sub-districts in Mimika district. The population of 202,350 people is increasing in size by an estimated 16% per year, mainly due to economic migration, and is composed of different ethnic groups including Highland Papuans, Lowland Papuans, and Non-Papuans. Unstable malaria transmission occurs mainly in the lowland areas, where approximately 90% of the population lives. In 2013, malaria prevalence by microscopy was estimated to be 12.2% (5.7% due to Plasmodium species are treated with dihydroartemisinin-piperaquine (DHP).The Rumah Sakit Mitra Masyarakat (RSMM) is the main hospital in the district, serving a population of approximately 150,000 people. Located in the lowland area, RSMM provides medical healthcare to approximately 300 patients per day. Since 2006, local antimalarial guidelines have recommended that all patients with uncomplicated malaria due to any Plasmodium cases was performed using ArcGIS software (version 10.2.1).Parasite DNA was collected within the framework of two studies. The first was an ongoing ex-vivo drug susceptibility surveillance study conducted between March 2011 and August 2014. The second was a cross-sectional study conducted between April and July 2013. Patient and parasitological details were recorded for each case. For the cross-sectional study, GPS coordinates were recorded of the residence of all individuals assessed. The majority of houses were located in one of 16 villages, within the 3 subdistricts Mimika Baru, Mimika Timur, and Kuala Kencana. Geospatial mapping of residences with Plasmodium species was undertaken using a nested PCR protocol with 2 μL gDNA template in duplicate [P. falciparum, P. vivax, P. malariae, and P. ovale small-subunit rRNA DNA clones  were used as positive controls.For parasitaemia detected by microscopy, species and parasite density were determined by examination of Giemsa-stained thick blood films. Genomic DNA (gDNA) was extracted from 50 μL packed red blood cell (RBC) pellets using the QIAamp 96 DNA Blood Kit (Qiagen) for the cross-sectional study and from 2 mL of packed RBCs using the QIAamp DNA Mini Kit (Qiagen) for the ex-vivo surveillance study cases. Confirmation of uplicate . P. falcP. falciparum and P. vivax genotyping was performed using short tandem repeat (STR) markers. Nine STR markers  were amplified in P. falciparum infections using previously described methods [P. vivax, nine STR markers that have been defined as a consensus panel within the Asia Pacific Malaria Elimination Network (APMEN)  were amplified using the APMEN protocol [ methods . For P. protocol –16. CapiHE), population differentiation was measured using the pairwise FST metric. Both HE and FST were calculated using Arlequin software (version 3.5) [P. vivax, measures of the HE, genetic differentiation and LD (described below) were undertaken on the full marker set and on a sub-set of five markers  defined as exhibiting balanced diversity in a recent study [Samples with more than one allele at any locus were defined as polyclonal infections. The multiplicity of infection (MOI) for each sample was defined as the maximum number of alleles at any locus. Population diversity was measured using the expected heterozygosity (ion 3.5) , 18. Fornt study .K (number of sub-populations), ranging from 1–10. The model parameters included admixture with correlated allele frequencies. The most probable K was derived by applying the delta K method [IAS) using the web-based LIAN 3.5 software [STRUCTURE software version 2.3.3 was used to assess population structure . The simK method . STRUCTUK method . Multi-lsoftware . The sigsoftware . Unrootesoftware . Mantel’software .HE was undertaken using the Mann-Whitney U test and Kruskal-Wallis test.Statistical analysis was conducted using SPSS v.20.0 for windows software (IBM SPSS Statistics), with a significance threshold of 0.05. Data on patient gender, ethnicity and proportion of polyclonal infections was compared between subgroups using Pearson’s Chi-square test. The significance of difference between patient age, parasite density, MOI and Ethical approval for this study was obtained from the Eijkman Institute Research Ethics Commission, Eijkman Institute for Molecular Biology, Jakarta, Indonesia (EIREC no. 47/2010 and EIREC no. 67/2013), Faculty of Medicine, University of Gadjah Mada (Ref: KE/FK/763/EC), and the Human Research Ethics Committee of the Northern Territory Department of Health & Families and Menzies School of Health Research, Darwin, Australia (Ref: HREC-2010-1434).P. vivax and 94 P. falciparum cases were collected from the surveillance study, with genotyping successfully completed in 96% (104) of P. vivax and all P. falciparum cases. The cross-sectional household survey, undertaken between April and July 2013, assessed 2,476 participants by blood film examination and PCR analysis, of whom 877 (35.4%) had peripheral parasitaemia detected by PCR. P. vivax and P. falciparum accounted for 749 (85.4%) malaria cases. Genotyping was successful in 60% (240/397) of P. vivax and 80% (280/352) of P. falciparum cases. A total of 73 (30.4%) P. vivax and 21 (7.5%) P. falciparum samples failed to amplify any of the markers; all of these had submicroscopic parasitaemia.Between March 2011 and August 2014, 108 P. falciparum or P. vivax parasitaemia are presented in P. vivax and P. falciparum cases did not reveal any local high prevalence clusters for either species in the cross-sectional study Demographic and parasitological details on the individuals with P. falciparum and P. vivax markers are summarised in P. falciparum markers (TA42 HE = 0.315 and TA109 HE = 0.176), all markers exhibited a minimum 5% minor allele frequency. With the exception of TA42 (20.6%) in P. falciparum populations, all other markers exhibited >80% genotyping success in both species. The MS8 assay presented artefacts that affected the reliability of the allele calling and therefore, this locus was excluded from further analysis.The diversity and genotyping success rate for each of the P. vivax exhibited higher genetic diversity than P. falciparum  and a higher proportion of polyclonal infections . The mean MOI was higher for P. vivax (1.6) than P. falciparum (1.2) (p<0.001) , even after excluding complex infections . There was no evidence of LD after restricting to the 5 balanced markers , although LD became significant after removing complex samples . LD analysis using the unique haplotypes found no evidence of epidemic transmission in either species (p<0.001) . In contP. vivax . Analysi species .P. falciparum MLGs (106 infections) were observed in more than one individual (range 2–13). Only 22 MLGs (68 cases) had GPS metadata available. Amongst these, 3 pairs of infections (8.8% (6/68)), were shared between members of the same household. Seven pairs of infections ) were less than 1 Km apart. The median distance between infections was 5.3 Km (range 2 meters to 19.5 Km) (Thirty three 19.5 Km) .P. falciparum, no substructure was found in the P. vivax population. The neighbour-joining analysis  to K1 and the remaining 89.4% of isolates demonstrating predominant ancestry to K2 .In contrast to analysis  illustrary to K2 . Neighbory to K2 . AssessmP. falciparum subpopulations  (p<0.001) (IAS of 0.041 (p<0.01) for K1 and 0.033 (p<0.01) for K2  . Howeverp<0.001) . LD anal) for K2 ; the marp = 0.084), sex (p = 0.734), ethnicity (p = 0.972), or occupation (p = 0.096) (p <0.001) (p = 0.762). The geographical distribution of both subpopulations is shown in p = 0.851).Investigation of the demographic profiles of the K1 and K2 clusters did not find any significant difference in terms of age (= 0.096) . Interes <0.001) . No signP. falciparum population, the Timika genotypes (Papua) were compared with published data on 168 P. falciparum isolates collected from 4 surveys in other Indonesian islands  underlying the sub-structure in the st Timor . Principst Timor . Furtherst Timor ; Table 5 islands .P. falciparum and P. vivax populations circulating in Timika, Papua Indonesia. At this high spatial resolution, we demonstrate contrasting transmission patterns of both co-existing species, and reveal admixture of two divergent P. falciparum subpopulations, one comprised of submicroscopic infections with high genetic relatedness to isolates from other Indonesian islands.This study utilised a comprehensive collection of clinical and community samples to describe the genetic microepidemiology of P. vivax compared to P. falciparum populations [P. falciparum, there was no evidence of substructure in P. vivax. LD was also markedly lower in P. vivax than P. falciparum, even after accounting for admixture in the latter. These patterns are consistent with more intense and stable transmission in the local P. vivax population relative to P. falciparum. The patterns of diversity in the P. falciparum population were similar to those found in low to meso-endemic regions in Southeast Asia, but markedly less diverse than in hyper-holoendemic regions in Africa [P. vivax, the patterns of P. vivax diversity in Timika were most comparable to those reported in other hypo-mesoendemic settings in South-East Asia and the Pacific [P. vivax compared to P. falciparum transmission in this location, including recurrences from relapsing infections, the earlier appearance of the transmissible sexual stages in P. vivax, and the larger reservoir of submicroscopic infections in this species [Similarly to previous reports, we found higher population and within-host diversity in co-endemic ulations , 27–30. n Africa , 31. Alt Pacific . Several species .P. falciparum population exhibited several clusters of identical infections, neither species demonstrated evidence of epidemic transmission dynamics, indicating moderately stable transmission at the time of the study. The identical infections enabled assessment of shared origins of infection, revealing a small proportion (8.8% (6/68)) of isolates that were shared amongst household members, likely reflecting transmission events taking place in the vicinity of the household. However, the large majority of shared infections (70.5%) were over 1 Km apart and more likely to reflect a reservoir outside of the household. Together, the patterns of parasite relatedness, diversity and LD suggest that broad ranging interventions are still needed to further interrupt transmission of P. falciparum and P. vivax before more targeted strategies can be implemented in the area. Furthermore, although the widespread deployment of ACT has been effective in reducing the P. falciparum population, radical treatment targeting the dormant liver stages will be required to make a similar impact on P. vivax transmission.Although the P. falciparum population was the distinct substructure at this small spatial scale. The marked differentiation, elevated IAS on pooling the K1 and K2 subpopulations, lack of evidence of epidemic transmission, together with the moderate genetic diversity within each subpopulation was indicative of admixture. However, it is unclear how sympatric P. falciparum subpopulations are genetically isolated with no sign of geographical, temporal or demographic boundaries. We contemplated three possibilities: 1) the importation of these cases, 2) the rise of a new, fitter population, and 3) an unidentified biological barrier.The most notable revelation in the Timika P. falciparum genotypes with data from Bangka, Kalimantan, Sumba and West Timor revealed that the K1 isolates were more closely related to non-Papuan than Papuan isolates, and K2 was strikingly different from the rest of Indonesia. In accordance with geographic distance, the K1 sub-population demonstrated the greatest genetic similarity to the two most proximal islands, Sumba and West Timor, where P. falciparum remains hypo-mesoendemic. Owing to the mining industry and the National Transmigration Programme, there has been a great deal of human movement in Timika. Thus, it is feasible that the K1 subpopulation is the product of infections acquired outside Timika. Imported infections undermine local intervention efforts and present the threat of drug resistance introduction. Our results emphasise the need to develop geographical markers to help to differentiate between imported and local malaria cases. A recent study presented a barcode of organellar genome markers to determine the geographic origin of P. falciparum isolates [Comparison of the Papuan isolates , althougP. falciparum structure in Timika is the rise and spread of a fitter subpopulation in response to recent selective pressure such as the change in policy from chloroquine to ACT treatment in 2006. Parasites carrying mutations conferring chloroquine resistance may lose their survival advantage after drug pressure is removed, as has been demonstrated by the decrease in K76T pfcrt frequency in several African countries [P. falciparum and P. vivax, this treatment regimen may have had a greater impact on P. falciparum, since ACT alone has no activity against the dormant liver stages of P. vivax. However, more studies are needed to confirm this.The second possibility for the unusual ountries . AssuminAnopheline populations and their vectorial capacity for K1 and K2 is needed to address the first hypothesis. Although we did not find any evidence of association between K1 or K2 with Lowland, Highland or non-Papuan ethnicity, it is possible that a common erythrocyte polymorphism(s) defining parasite binding and invasions preference and that this transcends the different ethnic groups, and differentiates K1 and K2. Genomic analysis of these isolates might provide further insights.A further possible explanation for the sympatric co-existence of the K1 and K2 subpopulations is an unidentified biological barrier such as mutually exclusive vector preferences or differential erythrocyte binding specificities. Further investigation of the local P. falciparum substructure in Timika. However, similar substructure has been observed in Cambodia, where it was postulated to support the emergence of drug resistance, highlighting a need for intense surveillance strategies [Although we conducted a thorough characterisation of the subpopulations, we cannot conclusively determine the factors underlying the marked rategies . It is iOur study demonstrates considerable genetic heterogeneity between co-endemic species and distinct substructure within species that can be observed at small spatial scales, and may be reliant on passive as well as active case detection. These findings highlight the need for complementary new surveillance strategies to identify local transmission patterns that cannot be detected with traditional malariometric methods.S1 FigThe plot was generated using ArcGIS software on GPS coordinate data from the individuals identified with malaria parasitaemia. Each dot presents an individual case, with colour-coding by species according to PCR data.(TIFF)Click here for additional data file.S2 FigP. falciparum population. Different MLGs are distinguished with different colours and shapes. Circles, pentagons and diamonds indicate MLGs found in two, three, and more than three individuals respectively.This plot was generated using ArcGIS software on the GPS coordinates for the 22 identical MLGs in the (TIFF)Click here for additional data file.S3 FigP. falciparum case, with colour-coding by subpopulation as defined by STRUCTURE.The plot was generated using ArcGIS software on GPS coordinate data. Each dot presents an individual (TIFF)Click here for additional data file.S4 Figet al., with the addition here of a site label for Timika, Papua Indonesia [P. falciparum parasite rate in the 2–10 year age group (PfPR2–10) within the stable spatial limits of transmission in 2010. The approximate locations of the study sites described here are indicated with black stars.This map is a modified version of ndonesia . The ori(TIF)Click here for additional data file.S5 FigP. vivax population and the other four Indonesian islands.This plot illustrates the limited differentiation between the Timika (TIFF)Click here for additional data file.S1 Table(DOCX)Click here for additional data file.