Pseudapanteles dignus (Hymenoptera: Braconidae) early immature stages (egg and larva) encapsulation by the host Tuta absoluta (Lepidoptera: Gelechiidae), and 2) to determine the occurrence of self-superparasitism and the rate of escaping to encapsulation of this parasitoid. Knowledge of host-parasitoid immunological interaction is crucial when evaluating the potential of an endoparasitoid as a biological control candidate. Parasitoid-exposed T. absoluta larvae were dissected in vivo under light stereoscope microscope at 24-h intervals, for five days after exposition to detect encapsulation. The preimaginal stages of P. dignus and numbers of healthy and encapsulated immature parasitoids per host were recorded. Samples of parasitoid eggs and larvae were processed for SEM visualization of encapsulation. Necropsies evidenced that only the early first larval instar of P. dignus (up to 96 h-old) was partially or completely encapsulated. A non-melanized capsule, formed by layers of granulocyte-type hemocytes enveloping around the parasitoid body, was recorded. Approximately 50% of the parasitized T. absoluta larvae had significantly only one P. dignus egg, meanwhile supernumerary parasitization yielded up to seven immature parasitoids per host. The proportion of single-early first larval instar of P. dignus reached ≈ 0.5 and decreased significantly as the number of parasitoid individuals per host increased. P. dignus encapsulation and its ability to overcome with the host immune defense through self-superparasitism indicate that T. absoluta is a semi-permissive host for this parasitoid.Endoparasitoids can be killed by host encapsulation, a cellular-mediated host immunological response against parasitism that involves hemocytes aggregation. As a counteracting strategy, many parasitoids can evade this host response through self-superparasitism. The objectives of this study were: 1) to describe the parasitoid  Parasitoid-host interactions involve biological and physiological processes. Regarding host defense against endoparasitoids, i.e. those which juvenile instars develop inside the host body, a cellular-mediated response is implicated in forming a multi-layered sheath of blood cells or hemocytes to its surface. Several differentiated types of hemocytes have been morphologically and functionally characterized, being the symmetrically-spread, adhesive, phagocytic granulocyte- type the most abundant blood cells in Lepidoptera. Granulocytes strongly adhere to the parasitoid’s surface and can form a capsule surrounding the parasitoid body totally, eliminating intruders by asphyxiation, starvation or physical prevention of development, or partially, what enables parasitoids to continue developing normally , 2. EncaTo avoid encapsulation, endoparasitoids display several mechanisms, such as superparasitism and injection of immune-suppressors (e.g. venoms and/or polydnaviruses) , 6. SupeEncapsulation may adversely affect biological control success, by preventing the effective establishment of exotic parasitoids in new regions, or by reducing the parasitoid´s efficacy against targeted pest(s) , 10. A hPseudapanteles dignus (Muesebeck) (Hymenoptera: Braconidae) and the tomato leafminer, Tuta absoluta (Meyrick) (Lepidoptera: Gelechiidae). P. dignus is a solitary endoparasitoid reported for few gelechiids, including T. absoluta [T. absoluta—P. dignus interaction pointed out the potential of this parasitoid as a biocontrol agent against the pest in tomato crops [We examined in this paper some aspects of the parasitoid host immune relationship between absoluta , 12, 13,absoluta , 15, 16.to crops , 18, 19.P. dignus early immature stages (egg and larva) encapsulation by T. absoluta, and 2) to determine the occurrence of self-superparasitism and the rate of escaping to encapsulation of this parasitoid. We addressed the hypothesis that self-superparasitism increases P. dignus successful development as a solitary endoparasitoid, by encapsulating totally or partially supernumerary immature individuals. We conclude that knowledge on encapsulation and superparasitism reported here may be useful to P. dignus rearing protocols for its mass production.The objectives of this study were: 1) to describe Tuta absoluta and P. dignus colonies were established in the laboratory following Luna et al. [a et al.  from damT. absoluta pupation. Pupae were placed in meshed cages (30× 30× 30 cm), and when emerged, adults were fed with a honey solution (50%) ad libitum. Periodically, three-expanded leaves potted tomato plants were provided to adults for oviposition, and to start a new generation.Leaves were kept in plastic boxes (500 ml) until P. dignus colony was originated from cocoons (pupae) emerged from T. absoluta larvae collected in the field, and kept until wasp emergence in polystyrene Petri dishes (50 ml). Couples of adult wasps were placed in polypropylene cups (7.5 cm diameter and 18 cm height), provided with a honey solution (50%) as food, and batches of T. absoluta larvae settled in tomato leaves to oviposit. Exposed host larvae were maintained as described for healthy T. absoluta larvae in plastic boxes until P. dignus wasps were obtained, which were used in the experiments.Both colonies were held in a walk-in environmental room at 25 ± 2°C, 70 ± 5% relative humidity and L14:D10 photoperiod, and checked every two days.P. dignus encapsulation process by T. absoluta. The experimental unit consisted of a polypropylene cup (7.5 cm diameter and 18 cm height), where 10 T. absoluta larvae (second and third instar stages), installed in tomato leaf mines, were exposed to one P. dignus female (48-h old and 24-h mated), provided with 50% honey solution. Oviposition was allowed for 24 h to ensure parasitism, as previously reported [T. absoluta larvae used. T. absoluta larvae were conditioned with fresh tomato foliage as food and maintained in an acclimatized walk-in rearing room as described above.We conducted an experiment to study the sequence of events involved in the reported . At the in vivo suspended in a drop of Ringer’s saline on a concave glass slide under a BX51 stereoscope microscope (Olympus UK Ltd.), from 0 to 5 d after P. dignus oviposition , and daily up to 168 h, to search for parasitoid immature stages and detect encapsulation. Samples of immature parasitoids were prepared for observation using light  and scanning electron microscopy (SEM). For the SEM study, we followed the protocol described by Hayat [P. dignus immature stages are minute of size (<1 mm of length), the preparation for critical-point dry was carried out by relocating each parasitoid specimen individually in a 316 L- stainless steel vial designed for this study (11 mm diameter and 20 mm high) covered inside with a 100% polyester serigraphic mesh to avoid the loss of material , and two hours in 100% acetone analytical grade. After dehydration and critical-point drying, the parasitoids were extracted from the vial with a fine brush and carefully mounted for gold coating using a microscope.T. absoluta to P dignus was corroborated and described. Three degrees of P. dignus encapsulation were distinguished: encapsulated , partially encapsulated (loose hemocytes aggregation attached to the parasitoid´s body that still remained its mobility), and non-encapsulated. The number of P. dignus individuals per host and the degree of encapsulation were recorded.The encapsulation response by P. dignus reproduction behavior were analyzed: 1) superparasitism, as the proportion of host larvae containing one to the maximum number of immature P. dignus registered upon dissections, and 2) encapsulation rate, as the proportion of encapsulated larvae over the number of P. dignus larvae found per host. To analyze superparasitism, the number of immature P. dignus found upon dissections from 16 to 48 hours intervals were considered, since supernumerary P. dignus could be eliminated by host encapsulation and resorption after that period.From dissections, some aspects of the T. absoluta larvae (dependent variable) among the number of immature P. dignus registered per host (predictor variable). Previously, a test of homogeneity of variances was used (Levene´s test). Means were separated using a Student-Neuman-Keul (SNK) test with α = 0.05.Means and standard errors were calculated for each variable. A one way-ANOVA was performed to compare the proportion of P. dignus by T. absoluta was analyzed by fitting a logistic regression between the proportion of parasitized hosts (p) and the number of parasitoid individuals per host (x). Because of the proportion of parasitism is a dependent categorical variable , which errors are assumed to be binomially distributed, the logistic model is more suitable [0 and ß1:1 is similar to a slope parameter .The rate of encapsulation of immature suitable . The modThe program Statistica  was emplIn vivo host dissections under the light microscope allowed observing the chronology of the immature P. dignus development when parasitizing T. absoluta larvae . By the fifth day after oviposition, ca. 40% of larvae reached the second and third larval instars. Four out of 40 females failed ovipositing and then the hosts exposed were eliminated for further analyses.P. dignus first larval instar (24–36 hour- old) had symptoms of encapsulation (n = 78 parasitized host larvae)  Figs .Hemocytes were adhered around the parasitoid body, with a flattened and symmetrically spread aspect, in accordance to as a granulocyte-type description. Light and SEM studies revealed the three degrees of encapsulation, from a dense capsule to partially encapsulated, leaving the parasitoid larva partly free and mobile, being still alive. The capsule did not show signs of melanization.P. dignus predominantly behaved as a solitary species, but superparasitism also occurred. Approximately 50% of the parasitized T. absoluta larvae significantly had only one parasitoid egg deposited  = 11.57; p< 0.01). Supernumerary parasitization yielded mainly two to four P. dignus immatures per host, although a host with up to seven parasitoid eggs was occasionally registered , ß1 = -0.22; Χ2: 124.34; p< 0.001).A single parasitized creasing  as the nP. dignus-T. absoluta interaction. The first larval instar of P. dignus was found floating freely in the body cavity of the host, and the anatomical description was coincident with that reported by Cardona and Oatman [Keiferia lycopersicella  (Lepidoptera: Gelechiidae). The mature P. dignus first larval instar is reached ≈ 1 day longer in T. absoluta than in K. lycopersicella.The present work provides evidences of the anatomical and immunological aspects in the d Oatman  for otheP. dignus first-larval instar is encapsulated by T. absoluta, meanwhile other instars  escape from this immunological response. The capsule is formed by aggregated, granulocyte- or hyperphagocytic-type hemocytes described for other Lepidoptera species [P. dignus egg stage is not encapsulated may result from its rapid development (< 24 h after oviposition). A fast egg parasitoid hatching is considered a strategy to anticipate the host immune system reaction, an event that usually takes longer time [Only the early  species , 23, andger time . The preger time , 24, 25.P. dignus superparasitizes T. absoluta, as it does in K. lycopersicella [P. dignus larval encapsulation for single vs. multiple-parasitized T. absoluta larvae resulted more frequent than that observed for K. lycopersicella [In solitary parasitoid species, only one individual larva is able to successfully complete its development in a host. If more than a single egg is laid in a host by itself or a conspecific female, the supernumerary larvae will be eliminated , 26. In rsicella . The decrsicella . This brrsicella , 17. Thersicella .P. dignus immature parasitoids found for T. absoluta was greater than that reported for Cardona and Oatman [K. lycopersicella. The known mechanisms involved in reducing the brood size in solitary parasitoids are: 1) fighting and killing by siblings , and 2) superparasitism, that helps overcoming the host´s hemocytic reaction for the surviving larva [in vivo dissection study done with the light microscope, evidences for fighting among parasitoid juveniles were not detected. Otherwise, many supernumerary larvae were observed as encapsulated. Thus, we conclude that multiple oviposition by P. dignus is an strategy to successfully parasitize T. absoluta.The maximum number of d Oatman , whom reng larva . Based oet al. [T. absoluta is a semi-permissive host for P. dignus. This braconid is considered an oligophagous species, since it has been reported parasitizing only members of gelechiid lepidopterans, including T. absoluta, K. lycopersicella, the potato tuberworm, Phthorimaea operculella (Zeller), and the pepper flower-bud moth, Symmetrischema capsica (Bradley and Polovny) [Parasitoid-host physiological interactions help in determining host suitability and especially for endoparasitoids, the host specificity , 28. Cuiet al.  classifiPolovny) , 13, 14.P. dignus early-first-instar larvae can be encapsulated by T. absoluta larvae, displaying this parasitoid a mechanism of superparasitism to limit the host capacity to encapsulate supernumerary parasitoid larvae per host. We also found that T. absoluta acts as a semi-permissive host for P. dignus. To increase knowledge of the use of P. dignus as a potential biocontrol candidate for T. absoluta, next research should address the outcome of the encapsulation and superparasitism under field conditions, since they can be affected by other abiotic and biotic factors, as temperature, host and parasitoid species densities, and by virus-like particles and other symbiontic infections in the parasitoid, among others.In conclusion, our laboratory studies revealed that S1 Table(DOCX)Click here for additional data file.S2 Table(DOCX)Click here for additional data file.


Escherichia coli (E. coli) has long been a popular model organism for basic biological research. In this paper, weighted gene co-expression network analysis (WGCNA) algorithm was applied to construct gene co-expression networks in E. coli. Thirty-one gene co-expression modules were detected from 1391 microarrays of E. coli data. Further characterization of these modules with the database for annotation, visualization, and integrated discovery (DAVID) tool showed that these modules are associated with several kinds of biological processes, such as carbohydrate catabolism, fatty acid metabolism, amino acid metabolism, transportation, translation, and ncRNA metabolism. Hub genes were also screened by intra-modular connectivity. Genes with unknown functions were annotated by guilt-by-association. Comparison with a previous prediction tool, EcoliNet, suggests that our dataset can expand gene predictions. In summary, 31 functional modules were identified in E. coli, 24 of which were functionally annotated. The analysis provides a resource for future gene discovery.Network-based systems biology has become an important method for analyzing high-throughput gene expression data and gene function mining.  Escherichia coli (E. coli) is an abundant bacteria in the intestine of humans and animals. It is a single-celled prokaryote that is widely used as a model organism in biology research. The E. coli genome size is about 4.64 M and encodes 5416 genes. Well-developed microarray technology has been used to investigate genome-wide gene expression thanks to its low cost. For convenience, we refer to gene as the corresponding probesets on the microarray throughout the manuscript when focusing on genes rather than probesets. There is extensive E. coli transcriptome data deposited in the public databases. These data include gene expression data under various conditions, such as different nutrients, growing stages, and gene mutations [E. coli has not yet been comprehensively reported. Although the metabolic network of E. coli has been constructed [E. coli gene network from 524 arrays, which focused on the statistical aspect by comparing four algorithms, including WGCNA [E. coli using hundreds of microarrays [E. coli genes, collects co-functional gene networks for E. coli from seven distinct types of data, including co-expression pattern data from 132 microarrays [utations . It is sstructed ,4, only roarrays ,8,9. Theroarrays .E. coli biological functions by modules, systematically; and (3) to provide organization or connection information within module genes, which may infer the potential function of unannotated genes by guilt-by-association rationale [The aims of our analysis were as follows: (1) to integrate the independent datasets, which will help to overcome the limitations caused by sample size and statistical method bias; (2) to summarize the thousands of genes on the array into tens of modules, which will help simplify data complexity and clarify the ationale . E. coli was constructed from 1391 microarrays. Thirty-one modules were identified, representing various aspects of E. coli biological functions. Hub genes bearing high connectivity may play roles in module function. Unknown genes were also annotated.Here, the gene co-expression network for The biggest gene co-expression network to date has been constructed . The topWGCNA identified 31 stable co-expressed modules within which the genes have a similar expression pattern . To testE. coli functions can be categorized into relatively independent functional modules. For instance, Blue is associated with secretion, whose coding proteins localize outside the cell. Magenta and Lightyellow both encode the peptidoglycan-based cell wall, but Lightyellow involves fatty acid oxidation and Magenta involves amine biosynthesis. Seven of these modules did not detect any function: Floralwhite, Lightgreen, Lightsteelblue1, Purple, Skyblue3, Steelblue, and Violet, corresponding to 34, 109, 37, 276, 45, 59, and 53 probesets, respectively. Among them, Floralwhite, Lightgreen, Lightsteelblue1, Skyblue3, and Steelblue are the eight least stable modules. The absence of DAVID terms being detected may be due to their relatively fewer probesets and lower module stability.The database for annotation, visualization, and integrated discovery (DAVID) online tool was used to characterize the function of these identified modules. Twenty-four of them were annotated with significant Gene Ontology (GO) or Kyoto Encyclopedia of Genes and Genomes (KEGG) terms . E. coliE. coli gene co-expression modules, each gene has a different contribution, that is, different connectivity. The higher the connectivity, the more important the gene in the module. Therefore, the contribution of a gene to a module can be simply described by its connectivity. The hub genes were screened by their connectivity in each module [In h module . The scrAnalysis of gene expression variation in each module found modules performing housekeeping functions. Tan, Green, and Darkgrey have relatively low gene expression variation (lowest three), suggesting their housekeeping functions, such as ncRNA metabolism, translation, and lipopolysaccharide biosynthesis. Brown4, Darkorange, and Plum1 have relatively high gene expression variation (top three), mainly corresponding to stress responses, such as iron transmembrane transporter activity, pathogenesis, and anaerobic respiration .Module gene connectivity includes intramodule connectivity and intermodule connectivity. Intramodule connectivity represents the relationship between genes within a specific module. Intermodule connectivity equals the total network connectivity minus intramodule connectivity, indicating the contribution of genes in coordinating functions across modules. Module-based gene connectivity analysis revealed that Black has the lowest intermodule connectivity while Darkorange has the highest intermodule connectivity . DarkoraModules can be independent units that perform biological functions. Linking the modular gene expression with experimental conditions may help to discover modules functioning in specific conditions. For example, inorganic phosphate limitation induces the highest Darkorange expression . DarkoraE. coli is cultured with bovine serum albumin (BSA) at the logarithmic growth phase [E. coli. Many Darkred genes are related to flagella, such as flgF (1759235_at), fliN (1765241_s_at), fliG (1766530_s_at), and flgG (1767435_s_at) (The highest Darkred expression is induced when th phase . Darkred35_s_at) . SeventeTo compare our modules with those identified by a community detection algorithm , we overE. coli gene function predictions, such as EcoliNet. An example here is the Darkred module, where all module genes should be associated with flagella. The 11 genes with unknown function were submitted to EcoliNet for function prediction, which found that 9 of these genes are predicted to be associated with flagella , carbohydrate metabolism (Black), lipopolysaccharide biosynthesis , sulfur metabolism , and glycerol metabolism . Ion transport modules are Ivory and Grey60. The behavior module is Darkred. The pathogenesis modules are Blue, Darkmagenta, and Darkorange. Each module has a different function, suggesting the robustness of WGCNA.Gene function prediction by co-expression approaches is based on the similarity of gene expression profiles across multiple samples or conditions. More data would help to increase the predictive power . TherefoE. coli pathogenesis [E. coli gene co-expression network and provides clues for candidate genes suitable for experimental validation.Further identification of hub genes may help to reveal their important role in modules. For instance, the Darkorange SepD hub gene may play roles in ogenesis ; Black ZE. coli were downloaded from the National Center for Biotechnology Information Gene Expression Omnibus (GEO) database [E. coli Genome 2.0 Array data were processed. Raw gene chip data were analyzed by Expression Console (Version 1.2) by MAS5.0 method. Probe-level gene expression data were retrieved.A total of 165 GEO Series (GSE) datasets from The WGCNA package was run on R (Version 2.3.2) to construct a gene co-expression network and identify modules with the following parameters: networkType = ‘signed’, softPower = 12, minModuleSize = 30, deepSplit = 4. Briefly, a weighted correlation network was created by calculating the correlation coefficients with the power β . The β =Module stability was tested as the average correlation between the original connectivity and the connectivity from 1391 samples that were randomly sampled 1000 times. The process was run for every module.p-value. To visualize the whole network or specific module, the WGCNA exportNetworkToCytoscape function and Cytoscape tool [Genes in each of the identified co-expressed modules were annotated by the database for annotation, visualization and integrated discovery (DAVID 6.7) . In DAVIape tool  were useFor gene expression variation analysis, the relative standard deviation for each gene in a module was calculated and the average values for each module were visualized. For connectivity analysis, the intermodule connectivity/total connectivity value was calculated for every gene and then the average value for each module was visualized.E. coli network constructed by Trevino [The  Trevino  and the  Trevino  were use Trevino  for pred


Mycobacterium abscessus complex, the recommended macrolide-containing combination therapy has poor kill rates. However, clinical outcomes are unknown. We searched the literature for studies published between 1990 and 2017 that reported microbial outcomes in patients treated for pulmonary M. abscessus disease. A good outcome was defined as sustained sputum culture conversion (SSCC) without relapse. Random effects models were used to pool studies and estimate proportions of patients with good outcomes. Odds ratios (OR) and 95% confidence intervals (CI) were computed. Sensitivity analyses and metaregression were used to assess the robustness of findings. In 19 studies of 1,533 patients, combination therapy was administered to 508 patients with M. abscessus subsp. abscessus, 204 with M. abscessus subsp. massiliense, and 301 with M. abscessus with no subspecies specified. Macrolide-containing regimens achieved SSCC in only 77/233 (34%) new M. abscessus subsp. abscessus patients versus 117/141 (54%) M. abscessus subsp. massiliense patients . In refractory disease, SSCC was achieved in 20%  of patients, which was not significantly different across subspecies. The estimated recurrent rates per month were 1.835%  for M. abscessus subsp. abscessus versus 0.683%  for M. abscessus subsp. massiliense . The proportion of patients with good outcomes was 52/223 (23%) with M. abscessus subsp. abscessus versus 118/141 (84%) with M. abscessus subsp. massiliense disease . M. abscessus subsp. abscessus pulmonary disease outcomes with the currently recommended regimens are atrocious, with outcomes similar to those for extensively drug-resistant tuberculosis. Therapeutically, the concept of nontuberculous mycobacteria is misguided. There is an urgent need to craft entirely new treatment regimens.In pharmacokinetic/pharmacodynamic models of pulmonary  Mycobacterium abscessus complex members are rapidly growing mycobacteria associated with a wide spectrum of disease in humans, of which pulmonary disease is the most recalcitrant , leading to the moniker “the antibiotic nightmare” , and an aminoglycoside given in the first 2 to 4 months  criteria. The remainder, 12/19 (63%), were at moderate or serious risk of bias due to missing data across all domains and due to confounding . Eleven (58%) of the 19 studies were from Northeast Asia , 7 (37%) were from North America, and one (5%) was from the Netherlands.Of the 1,166 unique citations identified through systematic search, 27 were eligible for full review, and 19 of these were included in our analyses  4, 6, 71416, 19–3619–M. abscessus complex patients, of whom 1,013 (66%) were started on antimicrobial combination chemotherapy (M. abscessus subsp. abscessus was isolated in 508 (50%) patients, M. abscessus subsp. massiliense in 204 (20%) patients, and M. abscessus subspecies bolletii in only 3 patients (<1%). The remainder, 301 (30%), did not have isolates identified by subspecies and were termed M. abscessus with no subspecies specified. Three patients had mixed infections, i.e., had more than one subspecies  studies reported deaths. The pooled incidence was 4%  with an I2 value of 0%. Given the critical risk of bias, further analyses of death as an outcome were not pursued.Death was inconsistently reported, and its effect size was considered a critical risk of bias . MoreoveOnly a single study reported use of macrolide-free regimens in 120 patients. Ten of 120 patients attained sustained sputum conversion. This translates to a sputum conversion rate of 8.33% .I2 = 91%; P < 0.001). However, there was lower heterogeneity among the seven M. abscessus subsp. abscessus disease studies  (P > 0.346 by Egger's test), suggesting that effect estimates from studies reflect true observations in study patients . The forest plot in M. abscessus subsp. abscessus attained sustained sputum conversion, which was significantly lower than the 117/141 patients with M. abscessus subsp. massiliense disease. The sputum conversion rates were 35%  in M. abscessus subsp. abscessus patients and 79%  in M. abscessus subsp. massiliense patients. The odds ratio (OR) of sustained sputum conversion in M. abscessus subsp. abscessus versus M. abscessus subsp. massiliense diseases was 0.108  (P < 0.001).Regarding to macrolide-containing regimens as the initial therapy, there was significant heterogeneity among the 13 studies that examined sustained sputum culture ( = 0.01) . There wI2 value among the 5 studies that examined sustained sputum culture of macrolide-containing regimens in refractory patients was 72% (P < 0.001), indicating significant heterogeneity. However, there was no significant publication bias or small-study effects (P > 0.399) . M. abscessus species. The pooled culture conversion rate was 20% , which on follow-up after stopping therapy for 12 months was not significantly different across the mycobacterial species. Comparison between M. abscessus subsp. abscessus- and M. abscessus subsp. massiliense-infected patients revealed no statistical difference in sputum culture conversion: the odds ratio was 3.316 , and the P value was 0.205. New investigational drugs and novel delivery systems were applied in these combination regimens. The pooled sputum conversion rates in patients with M. abscessus with no subspecies specified after liposomal aminoglycoside therapy versus parenteral macrolide-containing therapy were 13.33% versus 5.88% (P = 0.589). Nonetheless, these results revealed that sputum culture conversion in refractory patients was very poor across all species and statistically not different from each other, regardless of route of administration of aminoglycosides.The I2 value was 77% (P < 0.001), indicating significant heterogeneity between the studies. There was also significant bias (P = 0.030 by Egger's test) observed with the recurrence outcome studies. However, this bias was expected, since fewer studies (10/19) with even fewer patients reported on this outcome. Overall, disease recurrence in M. abscessus subsp. abscessus-infected patients was 40%  versus 7%  in M. abscessus subsp. massiliense-infected patients . Since the follow-up duration was variable between studies and the risk of recurrence was significantly different between the isolated organisms, we adjusted for both and then expressed disease recurrence per month and per year of follow-up, as shown in M. abscessus subsp. abscessus disease and 0.683%  per month in M. abscessus subsp. massiliense disease. Thus, recurrence was significantly higher with M. abscessus subsp. abscessus infection across all studies regardless of the risk of bias of each study.patients . The oddM. abscessus subsp. abscessus versus 118/141 (84%) with M. abscessus subsp. massiliense disease. The odds ratio of good outcomes was 0.059  (P < 0.001).Finally, in patients with appropriate follow-up, we defined good outcomes as sustained sputum conversion rate without relapse, a composite outcome, while the alternative was a poor outcome. The proportion of patients with good outcome was 52/223 (23%) with M. abscessus complex disease was 48%  in North American patients versus 68%  in the Northeast Asian patients. The odds ratio for sputum conversion rate among North America patients was 0.458  compared to Northeast Asia patients (P = 0.015). However, when analysis was restricted to patients with M. abscessus subsp. abscessus infection, pooled sputum culture conversions were 37%  in North America versus 33%  in Northeast Asia. The odds ratio was 1.222  and not significantly different (P = 0.526). This suggests that differences in sputum conversion rates between the two locales are partially driven by differences in prevalence of infection with different subspecies. Finally, we also examined the differential effect of macrolides (clarithromycin versus azithromycin) on sputum culture conversion and recurrence. There was no significant difference between the two drugs for either organism . Risk of bias did not change our findings even after metaregression analyses, shown in Fig. S6.Sensitivity analysis revealed that pooled sustained sputum culture conversion in M. abscessus subsp. abscessus are dramatically worse than those for patients with MDR-TB and similar to those for patients with XDR-TB. Outcomes in patients with M. abscessus subsp. massiliense pulmonary disease were also poor but similar to those for patients with MDR-TB. In fact, we found outcomes are significantly worse than cure rates of >95% and 10% relapse over 10 years for the epic and ancient disease of leprosy (Mycobacterium avium complex (MAC) were roughly 50%, which is very poor but still better than that for M. abscessus subsp. abscessus disease , more than 80% of patients respond to therapy. For the much-feared multidrug-resistant TB (MDR-TB), microbial response rates are 50 to 90%; for the dreaded extensively drug-resistant TB (XDR-TB), favorable outcomes are found for 16% of patients at 24 months of follow-up . Here, w leprosy . In a re disease . Thus, tM. abscessus complex, perhaps based on outcomes in MAC. We show that even with macrolide regimens, outcomes in pulmonary M. abscessus complex are very poor. Recently, in a hollow-fiber pharmacokinetics/pharmacodynamics (PK/PD) model of M. abscessus subsp. abscessus disease, we demonstrated that even when drug concentrations were at their most optimal, the standard macrolide-containing regimen had poor maximal microbial kill (Emax) . Thus, it was not surprising that even inhalational therapy did not improve outcomes, since regimens cannot kill any more than their Emax  , 13. At eir Emax , 13. IndM. abscessus complex, which obviously do not respond. Our results show that there is no therapeutic benefit to conflating different mycobacterial species, even within M. abscessus complex itself: M. abscessus subsp. massiliense had dramatically different response rates than M. abscessus subsp. abscessus. Therefore, why conflate them or combine them in the same category? It is hoped that in the era of WGS and matrix-assisted laser desorption ionization–time-of-flight mass spectrometry, the designation of non-hyena animals will be ditched. No one ever calls a Shigella species a non-E. coli organism, even though Shigella species are considered to belong to the genus Escherichia in the Enterobacteriaceae family, a closer relationship than M. avium to M. abscessus subsp. abscessus. Therapies for M. abscessus subsp. abscessus and M. abscessus subsp. massiliense should be sought without reference to M. tuberculosis or even M. avium, which in any case have much more favorable outcomes than their cousins.We propose that the hope that macrolides could improve outcomes was partially based on the misguided idea of classifying highly virulent mycobacterial species as nontuberculous mycobacteria (NTM), equivalent to classifying lions and elephants in the African savanna as non-hyena animals. This led to conflation of improvement of different mycobacterial species (NTM) responses to macrolides with M. abscessus complex is based on the balance of potential risks and benefits for each individual. Patients who opted not to be treated even with severe disease were not included in the analyses, suggesting that the atrocious outcomes that were reported and we identified are more optimistic than findings in the clinic. As an example, our estimates of pooled mortality estimates are subject to critical risk of bias, while those for disease recurrence have moderate risk of bias.There are several strengths and limitations to our study. First, as stated throughout our results, there was much heterogeneity of studies. However, we used a validated instrument to assess the risk of bias and then applied random effects models with subgroup analyses in anticipation of much heterogeneity. Metaregression methods and sensitivity analyses supported this approach by consistently showing that our designated subgroups were homogenous, and the same estimates were obtained with these different approaches. This suggests that our estimates are robust and that our conclusions will be reproduced in future studies. A second important limitation is that the same authors from the same institutions performed a considerable number of the retrospective studies, albeit with different enrollment time frames and inclusion criterion but nonetheless drawing from the same databases. While we were able to exclude obvious duplicate studies of the same patient cohorts, in some instances we could have failed to decipher whether the same patients were reported in different studies. The third limitation relates to differential sputum sampling between patients within and between studies for both diagnosis and monitoring disease during therapy. Sputum sampling did not follow an identical schedule between studies. Fourth, the decision to treat pulmonary M. abscessus complex disease or infection, (ii) clear specification of combination therapy received by patients, and (iii) clear specification of microbial outcomes attained by patients after 12 months of treatment or at the end of treatment. Studies judged to be at (i) critical risk for bias or (ii) without adequate information upon which risk of bias for any of the outcomes evaluated in that study could be made were excluded from the meta-analyses. There is no consensus on definitions for treatment outcomes in nontuberculous mycobacterial pulmonary diseases, so we used the outcomes (including sputum culture conversion) stated by each study to define composite outcomes. Most studies defined sputum conversion as 2 to 3 or more consecutive negative cultures, consistent with ATS and IDSA criteria .The complete search strategy used with each database is included in the supplemental material. Briefly, we searched PubMed and Embase for reports published before 30 March 2017 that included MeSH terms or the free-text terms “nontuberculous mycobacteria,” “rapid growing mycobacteria,” “www.riskofbias.info). The ROBIN-I tool addresses several weaknesses identified in previous instruments used to measure study quality in observational studies, is easy to use, is easy to interpret, and is highly reproducible  number of patients enrolled, receiving therapy, and had outcomes evaluated, (iv) combination therapy regimens examined and duration of therapy, and (v) number of patients with outcomes. Data were extracted from tables, text, or referred articles. Two reviewers (J.G.P. and D.O.) independently examined the studies for bias and extracted the data into a prespecified electronic database. The two databases were compared for consistency, and disagreements were settled after consultation with a third reviewer (T.G.). ATS/IDSA standard definitions and terms were used throughout (The following data were extracted from each study: (i) the author(s) and the year the study was conducted and published, (ii) criteria used to establish pulmonary roughout .M. abscessus complex therapy  and their 95% confidence intervals (CI) across therapy regimens, stratified by organism and geographic locale. Since macrolides are considered essential for  therapy , regimenI2 statistic to quantify heterogeneity of the effect size estimates between patient groups and between studies . Freeman and Tukey double arcsine transformation was used to stabilize the variance . This al studies . Sensiti studies . Egger's


Helicobacter pylori infection. There is increasing evidence from epidemiological studies of the association of H. pylori infection and specific virulence factors with gastric cancer. Studies in animal models indicate H. pylori is a primary factor in the development of gastric cancer. One major virulence factor in H. pylori is the cytotoxin-associated gene A (cagA), which encodes the CagA protein in the cag pathogenicity island (cag PAI). Meta-analysis of studies investigating CagA seropositivity irrespective of H. pylori status identified that CagA seropositivity increases the risk of gastric cancer  relative to the risk of H. pylori infection alone . Eradicating H. pylori is a strategy for reducing gastric cancer incidence. A meta-analysis of six randomised controlled trials (RCTs) suggests that searching for and eradicating H. pylori infection reduces the subsequent incidence of gastric cancer with a pooled relative risk of 0.66 (95% CI: 0.46–0.95). The introduction in regions of high gastric cancer incidence of population-based H. pylori screening and treatment programmes, with a scientifically valid assessment of programme processes, feasibility, effectiveness and possible adverse consequences, would impact the incidence of H. pylori-induced gastric cancer. Given the recent molecular understanding of the oncogenic role of CagA, targeting H. pylori screening and treatment programmes in populations with a high prevalence of H. pylori CagA-positive strains, particularly the more oncogenic East Asian H. pylori CagA strains, may be worth further investigation to optimise the benefits of such strategies.Gastric cancer is a major health burden and is the fifth most common malignancy and the third most common cause of death from cancer worldwide. Development of gastric cancer involves several aspects, including host genetics, environmental factors, and  Gastric cancer is the fifth most common malignancy in the world . Globally, nearly one million  new cases and 723,000 deaths were estimated to have occurred from this disease in 2012 [Using the high-quality cancer registry data from the Cancer Incidence in Five Continents Volume XI (CI5 XI) , Table 1In general, the distribution of gastric cancer by subsite of the stomach has not been well characterised in international comparative datasets. A recent estimate of the proportion of cardia and non-cardia gastric cancers using the same data sources as above indicates that approximately 27% of gastric cancers  arise in the cardia region of the stomach with the remaining 73%  defined as non-cardia [Helicobacter pylori was initially considered and classified as carcinogenic to humans (Group I carcinogen) by an International Agency for Research on Cancer (IARC) Working Group in 1994 based on the results from a small number of studies then available (four cohort and nine case–control) that considered gastric carcinoma [H. pylori is a Group 1 carcinogen with sufficient evidence of causing non-cardia gastric carcinoma and low-grade B-cell gastric MALT lymphoma. arcinoma . In 2009H. pylori is a Group 1 carcinogen with sufficient evidence of causing non-cardia gastric carcinoma and low-grade B-cell gastric MALT lymphoma [In 2009, a new Working Group reviewed considerably more data which had become available since the previous evaluation and reconfirmed that chronic infection with lymphoma .H. pylori infection and non-cardia gastric carcinoma in studies using Western blotting assays compared with enzyme-linked immunosorbent assay (ELISA) for the analysis of serum IgG responses to H. pylori. Using ELISAs, statistically significant ORs varied widely from 1.6 to 7.9, whilst studies using Western blotting analysis to determine infection status reported an increase in the ORs up to 10.6 [H. pylori infection in IgG serological studies was more than three-fold higher by Western blot assay than by ELISA  [H. pylori have also been investigated. Resected non-involved gastric mucosa of patients with gastric cancer was cultured in vitro to investigate mucosal IgA responses to H. pylori. Interestingly, several H. pylori IgG serological negative patients with gastric cancer secreted mucosal IgA against multiple H. pylori proteins indicative of a previous positive H. pylori status [The 2009 Working Group noted a substantial increase in the estimated odds ratios (ORs) for the association between  to 10.6  or 17.8 i status .H. pylori in experimental rodent models [H. pylori-infected Mongolian gerbils developed gastric adenocarcinoma in the majority of studies [H. pylori infection, gastritis was evident whilst gastric adenocarcinoma did not develop [H. pylori showed synergistic effects in gastric cancer development [H. pylori infection of transgenic mice overexpressing gastrin could induce gastric adenocarcinoma [H. pylori eradication therapy was introduced [H. pylori eradication in mice deficient in p27 similarly reversed premalignant lesions [The IARC Working Group evaluation also considered experimental evidence in Mongolian gerbils and mice and concluded that there is sufficient evidence for the carcinogenicity of infection with t models . H. pylo studies ,16,17,18 develop ,20,21. I develop , p27 def develop  and Tref develop , infectiarcinoma , while gtroduced . H. pyloH. pylori-induced gastric pathology progresses from antral gastritis to pan gastritis with corpus atrophy in a similar manner to human gastric pathology, in H. pylori infected individuals [H. pylori eradication on the development of gastric cancer [H. pylori transactivates the epidermal growth factor receptor (EGFR) in gastric epithelial cells [H. pylori-induced pathology have been examined in gerbil and murine models. Treatment of H. pylori-infected gerbils with the selective EGFR inhibitor EKB-569 for 32 weeks significantly reduced corpus atrophy, mucous metaplasia, submucosal herniations  of these could be attributed to H. pylori infection with a further 13,000 gastric lymphomas also likely to be H. pylori-associated [H. pylori infection in gastric non-cardia cancer cases of 94.6% and a relative risk of 17.0. These estimates indicate that H. pylori infection should be considered the primary risk factor for gastric cancer, and it has been proposed that infection might be considered “a (close to) necessary cause of non-cardia gastric cancer” [A combination of the high prevalence of sociated . These e cancer” .H. pylori strains expressed a protein of circa 120 K and mucosal IgA responses to this protein are significantly associated with peptic ulcer disease and with active gastritis characterised by increased gastric epithelial neutrophil infiltration [cag pathogenicity island (cag PAI), a type IV secretory system [cag PAI also translocates peptidoglycan breakdown products into gastric epithelial cells, which activate intracellular NOD1 and NFκB [cag PAI-positive infections [Early studies found that not all ltration . The 120y system ,39. CagAy system ,41. Othey system ,43. The and NFκB ,45, thusfections ,44,45,46cag type IV secretory system into epithelial cells [H. pylori delivered via the cag type IV secretory system triggers innate immune inflammatory responses. Recent studies have reported that in addition to NOD-1 activation, bacterial heptose-1,7-bisphosphate (HBP), a metabolic precursor of lipopolysaccharide biosynthesis, is also delivered via the al cells ,48. IndeH. pylori CagA-positive strains by both gastric mucosal IgA and serum IgG responses [The CagA protein is immunodominant and is recognised immunologically early following infection with esponses . With thesponses ,51, ELISesponses ,53,54,55esponses ,57.H. pylori-positive patients after a time span of 11 years reported 62% of CagA IgG seropositive subjects (n = 24) had developed atrophic gastritis versus 32% of H. pylori-positive CagA IgG seronegative subjects (n = 34)  [H. pylori positive patients (n = 267) undergoing endoscopy in 14 European countries, the combined presence of IgG antibodies to CagA and the H. pylori vacuolating cytotoxin A (VacA) was also significantly associated with atrophic corpus gastritis  [H. pylori-positive subjects. Supporting evidence comes from animal studies. Infection of Mongolian gerbils with a wild-type H. pylori strain with a functional cag type IV secretory system and isogenic mutant strains lacking cagA or cagY indicate that a functional cag PAI facilitates corpus colonisation and corpus atrophy [Followup endoscopic studies on corpus atrophy-negative 2–12.18) . In dysp26–7.56) . Infecti atrophy .H. pylori IgG seropositive subjects who were CagA seropositive had significantly lower pepsinogen A/C ratios than H. pylori seropositive subjects who were CagA seronegative. These studies reinforce the association of CagA-positive strains with corpus atrophy in multiple European countries [The Eurogast study included 17 centres in 13 counties with 2850 randomly selected subjects in whom serum pepsinogen A/C ratio was determined as a marker of corpus atrophy . Assays ountries .H. pylori infection and corpus atrophy have to date been undertaken in developed countries. In developing countries, factors that may beneficially modulate H. pylori-induced pathology are co-infection with helminths, which increase T regulatory cells and polarise inflammation to Th2 responses [H. pylori-infected Chinese subjects in the Dongting Lake area indicate that co-infection with the digenean helminth Schistosoma japonicum significantly reduces gastric atrophy in H. pylori-infected subjects as determined by increased pepsinogen A:pepsinogen C ratio in CagA seronegative subjects [H. pylori-associated corpus gastritis are required.The majority of studies on the association of CagA-positive esponses . Clinicasubjects . Furthercag PAI-positive H. pylori strains [H. pylori strains and distal gastric cancer. Functional differences in CagA tyrosine phosphorylation sites between Eastern and Western CagA-positive H. pylori strains are evident [H. pylori strains and also stimulates stronger cellular Ras–ERK activation [H. pylori-positive individuals and age-matched uninfected controls [The marked global variation in both the incidence of gastric cancer ,2,3, Tab and the  strains  are impo evident ,62. Recetivation . CagA Igcontrols ,67,68,69H. pylori-positive CagA-negative subjects had an OR of 2.7 (95% CI: 1.3–5.6) for intestinal-type gastric cancer compared to uninfected controls [H. pylori and CagA seropositive however had an increased risk of intestinal-type gastric cancer with an OR of 4.1 (95% CI: 2.2–7.7) [H. pylori who were CagA seropositive similarly had increased risk of gastric cancer  relative to uninfected controls [p < 0.05) in patients with gastric cancer relative to non-cancer controls. The risk of gastric cancer with CagA seropositivity  was lower than that in an earlier study [H. pylori infection will decrease the density of H. pylori infection and thus CagA serological positivity. Furthermore, the high frequency of CagA-positive strains in Asia [In two gastric cancer case–control studies ,65 serumcontrols . Men who2.2–7.7) . In the controls  in 81 iner study  in emigrer study , in conter study . Increas in Asia  may impaH. pylori-positive CagA seronegative and H. pylori-positive CagA seropositive had an increased OR of gastric cancer [H. pylori-positive CagA seronegative subjects [A study in 103 younger (<40 years) Japanese patients with gastric cancer and age-matched controls found both c cancer . The majsubjects .H. pylori seropositives with positive low CagA antibody titres was greater  than that of subjects with high CagA antibody titres  [Ethnic differences in CagA antibody responses in non-cardia gastric cancer in two Brazilian case–control groups have also been reported . One gro= 0.002) . H. pylori status was 62.8% in cases (n = 1707) and 37.5% in controls (n = 2124). CagA seropositivity increased the risk of gastric cancer  relative to the risk of H. pylori infection alone  [Meta-analysis of CagA serology determined in 10 non-cardia gastric cancer case–control studies with age- and gender-matched controls reported CagA seropositivity irrespective of 58–3.39) . A more 58–3.39) . However58–3.39) , which icag PAI is key in triggering, respectively, oncogenic signalling pathways and inflammatory innate responses via NFkB activation  after adjusting for H. pylori infection, gastric diseases, gastroesophageal reflux disease, gastric polyp, and gastritis. Those subjects with distal gastric cancer (n = 4539) who had received previous long-term treatment with simvastatin had a relative risk of 0.65 (95% CI: 0.57–0.75) of gastric cancer compared to controls who had not received previous long-term treatment with simvastatin [H. pylori and gastric cancer risk.The use of certain pharmacological agents may also affect the outcomes of gastric cancer case–control studies investigating CagA seropositivity. The translocation of CagA and peptidoglycan breakdown products/ HBP into gastric epithelial cells via the tion see . H. pyloepletion ,74, suggvastatin . These sH. pylori CagA seropositivity and risk of distal gastric cancer, other important epidemiological studies have examined the association of CagA seropositivity with oesophageal adenocarcinoma (OAC) and oesophageal squamous cell carcinoma (OSCC). In contrast to gastric cancer, which is decreasing in many developed countries with the reduction in H. pylori infection, OAC is increasing. Ye et al. [n = 127) and OAC (n = 80). Whilst finding a significant association of H. pylori and CagA seropositivity compared to H. pylori seronegativity with distal gastric cancer , no association was evident between CagA seropositivity and OAC [cagA+ strains may be protective by reducing OAC [Whilst most studies to date have focused on e et al.  in a pope et al.  in a pop and OAC . The abocing OAC , potentiH. pylori infection in subjects with duodenal or gastric ulcers likely reflect different patterns of H. pylori gastric colonisation, acid secretion and associated genetic variation in acid regulatory cytokines [An interesting study examining retrospective cohorts of 61,548 unoperated patients with duodenal ulcers and 81,379 unoperated patients with gastric ulcers in the Swedish Inpatient Register surprisingly showed patients with duodenal ulcers had a 70% higher risk of OAC (standardised incidence ratio (SIR) = 1.7, 95% CI: 1.1–2.5) . Gastricytokines , as wellH. pylori-infected individuals. In a gerbil model, infection with a cagA-positive strain, but not the isogenic cagA mutant, resulted in development of gastric cancer [cagA-positive strains develop gastric cancer in Western populations [H. pylori strains express cagA; therefore, CagA status alone cannot explain the clinical outcomes [H. pylori strains have a closer association with gastric mucosal atrophy and higher incidence of gastric cancer than Western-type CagA H. pylori strains [cag PAI, known as the Japanese cluster. Individuals infected with the Japanese cluster strains were reported to have higher atrophic gastritis scores than the Western type counterpart [CagA has been recognised as an important factor in the development of gastric cancer in c cancer . Individulations ,64,65. Houtcomes . The mosoutcomes . Epidemi strains ,82,83. Tnterpart . Epithelnterpart .cagA expression was undertaken using array studies. The cagA expression was significantly higher at pH 6.0 compared to pH 7.0 [cagA expression was also observed at pH 4.0 [H. pylori. This study indicated that the expression of cagA, determined by the amount of cDNA, was two-fold higher under acid–stress conditions (pH 5.2) compared to a neutral environment (pH 7.0) [H. pylori, especially strains with a high number of EPIYA repeat regions [Apart from the internal mosaicism of CagA, which may reveal different activity of this oncogenic protein, environmental factors are also important in the regulation of CagA expression. Reported environmental factors that may be important are pH, salt and host serum iron concentrations ,87,88. To pH 7.0 ; a similt pH 4.0 . In a re(pH 7.0) . However regions .H. pylori [H. pylori CagA/cagA at both a protein and transcriptome level. RNA array analysis indicated cagA transcription had a two-fold increase in high salt concentrations [cagA transcriptome [cagA in in vivo studies. In cagA-positive H. pylori-infected gerbils, a high salt diet induced higher cagA expression compared to H. pylori-infected gerbils on a regular diet. The gerbil group on a high salt diet infected with cagA-positive H. pylori strains had higher levels of inflammatory cytokines , anti-inflammatory cytokines (IL-10), chemokines , and inducible nitric oxide synthase (iNOS). The higher expression of the H. pylori CagA oncogenic protein and the enhanced host immune response resulted in higher inflammation scores as well as a higher frequency of gastric carcinoma [High salt diets have been shown in epidemiological studies to be associated with an increased risk of gastric cancer. Intriguingly, recent studies indicated that a high salt diet modifies the expression of CagA in . pylori . High satrations . Subsequcriptome . Increascriptome . High saarcinoma . cag type IV secretion system, almost 2-fold higher, in nine visualised pili on strains from iron-replete versus 17 visualised pili on strains from iron-depleted gerbils. The translocation of CagA into gastric epithelial cells was also increased. Gerbils on iron-depleted diet infected with wild-type H. pylori, but not the cagA-negative isogenic mutant, were significantly quicker to develop dysplasia and adenocarcinoma (within 6 weeks of infection) than iron-replete gerbils [There is epidemiological evidence of a decreased risk of gastric cancer in subjects with a high iron status . Iron ha gerbils . In addi gerbils . These dcagA-negative H. pylori transform into the coccoid form, a non-viable H. pylori morphology, suggesting that CagA provides an advantage to the viability of H. pylori [H. pylori-related gastro-duodenal diseases and suggest iron-deficient subjects may have increased risk of developing gastric cancer.A subsequent study in iron-depleted gerbils indicated that . pylori . Interes. pylori . These s. pylori ,94 demonH. pylori infection suggests a clear path to preventing gastric cancer cases through eradication of the infection [H. pylori programmes have, therefore, been proposed as a means to prevent gastric cancer [H. pylori eradication in reducing gastric cancer and uncertainty about the possible harm of mass antibiotic treatment and its impact on gut microbiota [The strong evidence for the carcinogenicity of chronic nfection . Populatc cancer , and intc cancer ,98. Howecrobiota .H. pylori eradication is based on a small number of randomised controlled trials (RCT) of limited to moderate quality in healthy asymptomatic infected (mainly Asian) individuals [H. pylori infection reduces the subsequent incidence of gastric cancer with a pooled relative risk of 0.66 (95% CI: 0.46–0.95) [The current scientific evidence that gastric cancer risk is reduced by ividuals . A syste46–0.95) . The aut46–0.95) .H. pylori eradication therapy had a lower incidence rate of gastric cancer than those without eradication therapy (a pooled rate ratio of 0.53 (95% CI: 0.44–0.64) after adjustment for baseline gastric cancer incidence) [H. pylori eradication was greater in studies with higher baseline incidence rates of gastric cancer.A more recent systematic review and meta-analysis of 24 prospective studies also concluded that individuals receiving cidence) . Subgroucidence) . This stH. pylori eradication in gastric cancer, potential adverse consequences of mass therapy with antibiotics—for example, short- and long-term changes in the normal human microbiota, weight gain, gastroesophageal reflux disease and antibiotic resistance—have not been addressed in previous studies. In addition, before implementing large-scale programmes, further investigations are required regarding programme costs, feasibility, and appropriate target groups for the intervention, as well as region-specific data.Despite the observed preventive effect of H. pylori screening and treatment programmes and should resolve many related uncertainties. The largest of these was initiated in 2011 in Linqu County in China with 184,786 residents aged 25–54 from a high-risk population [H. pylori-positive participants ascertained by 13C-urea breath test were assigned to one of two groups receiving either a 10-day quadruple anti-H. pylori treatment or lookalike placebo together with a single dose of omeprazole and bismuth.Several clinical trials that are currently underway should help clarify whether and how to implement population-based pulation . H. pyloH. pylori Screening Study was initiated in 1997 to assess whether screening for and eradicating H. pylori infection in 56,000 healthy men aged 35–69 and women aged 45–69 would reduce the incidence of gastric cancer with a planned followup of 15 years or more after recruitment [H. pylori screening followed by eradication in infected participants and endoscopy of those with serological evidence of atrophic gastritis could reduce gastric cancer mortality continues, recruiting in Latvia and potentially in neighbouring countries with the target number of 30,000 after completion of its pilot phase in 2016 [H. pylori treatment on gastric cancer among 40–65 year-old participants recruited from the nationwide cancer screening programme was launched in 2014. The trial with a target number of 11,000 participants is scheduled to run for 10 years with biennial endoscopic followup within the screening programmes and populations [In the UK, the ruitment . A multi in 2016 . In the ulations .H. pylori antibiotic resistance [H. pylori antibiotic eradication therapies have been evaluated in multiple clinical studies before their application in controlled trials to evaluate their impact on gastric cancer. To date, no other clinically validated approach for the eradication of H. pylori is currently available, despite extensive preclinical vaccine studies on preventative and therapeutic H. pylori vaccines [Whilst there are concerns regarding increasing sistance  and othevaccines ,107. Futvaccines . CagA bivaccines . Modellivaccines .H. pylori as a strategy for gastric cancer prevention [H. pylori screening and treatment would be both feasible and cost-effective in preventing gastric cancer [H. pylori screening and treatment programmes but within the context of a scientifically valid assessment of programme processes, feasibility, effectiveness and possible adverse consequences [H. pylori screening and treatment programmes, with a scientifically valid assessment of programme processes, feasibility, effectiveness and possible adverse consequences would affect the incidence of H. pylori-induced gastric cancer.In December 2013, a Working Group meeting convened by IARC reviewed the evidence regarding eradication of evention . The parevention . The Groc cancer . The Groequences ,110. TheH. pylori screening and treatment programmes in populations with a high incidence of H. pylori CagA-positive strains, particularly the more oncogenic East Asian H. pylori CagA strains, may be worth further exploration to optimise the benefits of such programmes.Given the recent molecular understanding of the oncogenic role of CagA, targeting


Cryptococcus neoformans and C. gattii, have been commonly designated as either an opportunistic pathogen for the first species or as a primary pathogen for the second species. Such a distinction has been based on epidemiological findings that the majority of patients presenting meningoencephalitis caused by C. neoformans are immunocompromised while C. gattii infection has been reported more often in immunocompetent patients. A recent report, however, showed that GM-CSF (granulocyte-macrophage colony-stimulating factor) neutralizing antibodies were prevalent in the plasma of “apparently immunocompetent” C. gattii patients with meningoencephalitis. Because GM-CSF is essential for differentiation of monocytes to macrophages and modulating the immune response, it is not surprising that the lack of GM-CSF function predisposes otherwise healthy individuals to infection via inhalation of environmental pathogens such as C. gattii. Since the test for anti-GM-CSF autoantibodies is not included in routine immunological profiling at most hospitals, healthy patients with GM-CSF neutralizing antibodies are usually categorized as immunocompetent. It is likely that a comprehensive immunological evaluation of patients with C. gattii meningoencephalitis, who had been diagnosed as immunocompetent, would reveal a majority of them had hidden immune dysfunction. This paper reviews the relationship between GM-CSF neutralizing antibodies and the risk for C. gattii infection with CNS involvement.The two etiologic agents of cryptococcal meningoencephalitis,  Cryptococcus neoformans and C. gattii are sibling species (85%–90% genomic identity) [C. neoformans is worldwide in distribution and though it can cause disease in otherwise healthy individuals, a majority of the reported cases have been from patients whose immune system have been compromised by various causes such as HIV infection, corticosteroid treatment and other immunosuppressive underlying conditions [C. neoformans has been widely known as an opportunistic pathogen. Epidemiological deviancy in C. neoformans infection, however, has been reported among patients in the Far East countries: China, Korea and Japan. In these countries, most of the cryptococcosis patients were caused by C. neoformans and were HIV sero-negative with or without any apparent predisposing underlying condition [C. gattii in HIV+ patients have been reported mostly in tropical and subtropical regions of the world such as Africa [The etiologic agents of cryptococcosis, dentity)  that caudentity) ,3 Epidemnditions . This isondition ,7. Crypts Africa .C. gattii infects apparently healthy hosts with no known predisposing risk factor more frequently than immunocompromised individuals regardless of geographic regions and is therefore widely regarded as a primary pathogen [C. neoformans [C. gattii infrequently infects such patients compared to C. neoformans even in the C. gattii endemic regions of the world [C. neoformans includes strains of serotype A, D and AD which are generally unable to assimilate D-proline as a nitrogen source and glycine as a carbon source. C. gattii on the other hand can utilize D-proline as a nitrogen source and glycine as a source for both carbon and nitrogen [Several studies have confirmed that pathogen ,14,15,16oformans ,18, C. ghe world ,13,16. Dnitrogen ,20. Thesnitrogen , which initrogen  and the nitrogen .C. gattii infection in apparently healthy individuals is the result of increased environmental exposure to the fungus. Such speculation has been based on the over-representation of C. gattii infection among the Australian Aboriginal people living in rural areas [C. gattii such as Eucalyptus and other trees, organic debris and soil is likely higher than the population living in urban areas. Analysis of the risk factors for C. gattii infection among patients in British Columbia, Canada showed that, in addition to environmental exposure, certain underlying medical conditions to be of significant risk [C. gattii infection include preexisting oral steroid use, emphysema, chronic bronchitis, chronic obstructive pulmonary disease, sarcoidosis and pneumonia [It has been speculated that al areas ,23 whereant risk . These uneumonia . Environneumonia .C. gattii infection in individuals with these risk factors have not been evaluated. Countless numbers of tourists and inhabitants are exposed to the environment of C. gattii endemic regions such as Vancouver, Canada, Southeast Asia and South America each year but the number of patients afflicted by C. gattii has only been a fraction of the exposed populations: less than one case per million per year in Australia [C. gattii disease may be triggered by exposure, inhaled C. gattii cells in healthy individuals are readily eliminated by the host defense systems unless certain predisposing risk factors are present in these patients. Such endogenous risk factors may also contribute to increased persistence of dormant C. gattii or be required to trigger reactivation of dormant C. gattii to cause infection. GM-CSF neutralizing antibody appears to be one such host-dependent risk factor. Defining C. gattii as a primary pathogen may become obsolete as more subtle immune dysfunctions are recognized in otherwise healthy patients with C. gattii meningoencephalitis.The host specific mechanisms explaining the increased incidence of ustralia  and Coloustralia  to 5.8 custralia . This suet al., detected anti-GM-CSF autoantibodies which inhibited GM-CSF signaling in the plasma of four previously healthy HIV-negative patients suffering from cryptococcal meningoencephalitis [n = 64), diseased controls  or cryptococcosis patients with recognized immunodeficiency (n = 36). These results suggested that GM-CSF neutralizing antibodies could be a previously hidden risk factor for cryptococcal meningoencephalitis in HIV negative and otherwise healthy individuals. Of the seven cryptococcal strains (four from current and three from the archived cases) isolated from GM-CSF autoantibody positive patients, three had been reported as C. neoformans, one as C. gattii and three as Cryptococcus without identification at species level. The three Cryptococcus unspeciated strains had been isolated from the immunocompetent patients whose plasma samples had been archived and the strains had been maintained as lyophilized cultures.In 2013, Rosen phalitis . This fiC. gattii and one healthy volunteer, but none of the 20 patients infected by C. neoformans . The reason for higher association of GM-CSF autoantibodies with C. gattii infection than with C. neoformans infection is not clear. However, the higher prevalence of C. gattii than C. neoformans infection among patients with anti-GM-CSF autoantibodies could offer some clues to address the major differences in host’s immune responses to these two etiologic agents of cryptococcosis.Does this imply that GM-CSF neutralizing antibodies are a risk factor exclusively for infection due to C. neoformnans has been recognized since the early 1990s when GM-CSF neutralizing antibody increased mortality of C. neoformans infected mice with rapid progression of meningoencephalitis [−/− mice) failed to clear the fungus, which led to higher cryptococcal burdens in pulmonary cryptococcosis because GM-CSF was required for early influx of macrophages, CD4 and CD8 cells into the lung [Granulocyte-macrophage colony stimulating factor (GM-CSF) is one of the family of glycoprotein cytokines that mediates the survival, proliferation, differentiation and function of hematopoietic cells ,32,33. Iphalitis . Corrobothe lung . Additiothe lung ,43.et al., as “idiopathic PAP” [Furthermore, cryptococcosis is reported more frequently than other fungal disease in patients with acquired pulmonary alveolar proteinosis (PAP) . When auhic PAP” . This dihic PAP” , surfacthic PAP” . While ihic PAP” ,52,53, thic PAP” ,55. To ahic PAP” ,57.Cryptococcus. The etiologic agent of cryptococcosis in this patient could have been C. gattii rather than C. neoformans. The patient was working in a lumberyard in Massachusetts when he was diagnosed with cryptococcosis [C. gattii but the lumberyard could have been handling trees imported from regions where C. gattii is endemic. This case is reminiscent of the first autochthonous case of C. gattii infection in Germany in an apparently immunocompetent healthy male who had been working in sawmills and woodworking factories and exposed to high levels of wood dust, including that of imported tropical trees [Eucalyptus trees as the environmental source of C. gattii in Australia [Since autoantibodies to GM-CSF cause defects in chemotaxis, adhesion, phagocytosis, microbicidal activity and phagolysosome fusion of alveolar macrophages , patientcoccosis . The patal trees . This caustralia .Aspergillus fumigatus [Blastomyces dermatitidis case [Coccidioides case [Histoplasma capsulatum [H. capsulatum [H. capsulatum. The mode of anti-Histoplasma action enhanced by GM-CSF in mice was reported as the sequestration of labile Zn in infected macrophages by inducing metallothioneins in a STAT3 and STAT5 dependent manner [Histoplasma fails to replicate within the macrophages leading to fungal clearance [Aspergillus conidia by maintaining proinflammatory response by alveolar macrophages [Although less frequently reported than cryptococcosis ,63,64,65umigatus ,66,67,68dis case , one Cocdes case , one Mucdes case  and fourpsulatum ,71. GM-Cpsulatum . Neutralt manner ,74. Sinclearance ,74. GM-Crophages .C. gattii than C. neoformans infection? Before attempting to address this question, it is necessary to investigate whether anti-GM-CSF autoantibodies are also prevalent in pulmonary C. gattii infection without CNS involvement since evaluation of GM-CSF autoantibodies and cryptococcosis thus far has only focused on patients with CNS infection. Pulmonary infection without CNS involvement is more prevalent with C. gattii than with C. neoformans infection [C. gattii patients with CNS involvement, it suggests that GM-CSF may play a more important role in dissemination of C. gattii than C. neoformans to the brain from the lung and/or brain immunity against C.gattii than C. neoformans. Little is known about the role of GM-CSF in dissemination of Cryptococci to the brain or brain immunity against the fungus. Our study with GM-CSF−/− mice suggested that a lack of the cytokine function enhances dissemination of both the C. neoformans strain H99 and the C. gattii VG1 strain isolated from the GM-CSF antibody positive Chinese patient [vs. the VGI strain is currently underway in our laboratory and should shed more light as to the role of GM-CSF in dissemination and brain immunity against C. gattii. Although GM-CSF does not appear to influence brain microglial killing of C. neoformans serotype A strains [C. gattii by immune cells in the brain.Why do GM-CSF autoantibodies pose a higher risk for nfection ,16,76. I patient  from the strains , there hPneumocyctis carinii [P. jirovecii) or C. neoformans [et al. [C. gattii infection. There is a possibility that pulmonary infection with C. gattii stimulates GM-CSF production more than C. neoformans and the high cytokine production by C. gattii for a long period of time can cause induction of functional antibodies against the cytokine. Could C. gattii also exacerbate the production of GM-CSF autoantibodies more than C. neoformans in the patients who already are antibody positive? This question cannot be answered since the anti-GM-CSF antibody titers in these patients prior to C. gattii infection are unknown and there are no longitudinal studies on the GM-CSF autoantibody titers during the infection in such patients. Monitoring the health status and the GM-CSF autoantibody titers over time in the healthy Chinese volunteer positive for the cytokine antibodies [Before GM-CSF autoantibody was known as the cause of PAP, a hypothesis that PAP is an abnormal pulmonary response to an unusual infectious agent such as  carinii ,78,79 (noformans  gained soformans . Howeveroformans . Among toformans . In the oformans  as well  [et al. , none of [et al. . This sutibodies  could ofCryptococcus in otherwise healthy individuals. For some unknown reason, the neutralizing antibodies against GM-CSF appear to be a greater risk for cryptococcosis due to C. gattii than C. neoformans. This etiologic agent’s difference in the prevalence of GM-CSF neutralizing antibodies among otherwise healthy cryptococcosis patients may offer an important clue in order to address the differences in the host’s immunological response to the two pathogens. It is important to also include the GM-CSF autoantibody test during immunological profiling of cryptococcosis patients who appear to be otherwise healthy. This is not only for the assessment of subtle underlying immune disorders but also for better clinical management for those positive for GM-CSF antibodies since they may eventually develop PAP after meningoencephalitis.Anti-GM-CSF autoantibodies are apparently an important underlying risk factor for meningoencephalitis caused by


Helicobacter pylori infection and osteoporosis. A few studies have discussed the influence of H. pylori eradication therapy on bone mineral density.Previous studies have reported conflicting results on the association between H. pylori-infected population in Taiwan and the influence of early and late H. pylori eradication therapy on bone mineral density.We assessed the prevalence of osteoporosis among the H. pylori eradication therapy from 2000 to 2010 and 21,788 controls, frequency-matched according to age, sex, and year of receiving H. pylori eradication therapy. Those who received H. pylori eradication therapy were divided into two groups based on the time interval between the diagnosis of a peptic ulcer and commencement of eradication therapy. The risk of developing osteoporosis was higher in the early H. pylori treatment cohort  and late H. pylori treatment cohort , compared with the risk in the control cohort. When followed for less than 5 years, both the early and late cohorts had a higher risk of developing osteoporosis . However, when the follow-up period was over 5 years, only the late eradication group exhibited a higher incidence of osteoporosis .Using data from Taiwan's National Health Insurance Research Database, we identified 5,447 patients who received H. pylori infection may be associated with an increased risk of developing osteoporosis in Taiwan. Early eradication could reduce the influence of H. pylori infection on osteoporosis when the follow-up period is greater than 5 years. Further prospective studies are necessary to discover the connection of H. pylori and osteoporosis.The development of osteoporosis is complex and multi-factorial. Via this population-based cohort study and adjustment of possible confounding variables, we found  Osteoporosis is a common, but difficult to detect, disease among elderly people and is characterized by decreased bone mineral density. The World Health Organization claimed that osteoporotic fractures account for 2.8 million disability-adjusted life years annually in Americans and Europeans. Previous studies have reported age, sex, cigarette smoking, low body mass index, steroid use, and chronic alcohol consumption –4 as risGastrointestinal diseases are also believed to contribute to the development of osteoporosis . Both ToHelicobacter pylori infection is associated with several gastrointestinal diseases, such as chronic gastritis, peptic ulcers, gastric cancer, and mucosa-associated lymphoid tissue lymphoma [H. pylori, including coronary artery disease, chronic kidney disease, dementia, and anemia [Research suggests lymphoma –16. Reced anemia –19.H. pylori infection is a risk factor for osteoporosis remains controversial [H. pylori eradication therapy on bone mineral density. Since H. pylori infection is the most common chronic bacterial infection of the human upper gastrointestinal tract [H. pylori eradication therapy may have different durations of chronic H. pylori infection. We suspected that early H. pylori eradication therapy could minimize the influence of chronic H. pylori infection on bone mineral density. We conducted a population-based retrospective cohort study by using records from the Taiwan National Health Insurance Research Database (NHIRD) to investigate the incidence of osteoporosis among people who received H. pylori eradication therapy and the influence of early and late H. pylori eradication therapy on bone.Whether oversial –23. Howeal tract , 25, peoA retrospective cohort study was assembled using data from the Longitudinal Health Insurance Database 2000 (LHID2000) provided by Taiwan’s National Health Research Institute (NHRI), which includes information on outpatient, ambulatory, and hospital inpatient care as well as dental services. The National Health Insurance (NHI) program was launched on March 1, 1995 and covered almost 99% of the 23 million residents of Taiwan . The detH. pylori eradication therapy. According to the insurance benefits of the NHI system, H. pylori-related treatments are confirmed with gastroscopy and biopsy. H. pylori eradication treatment with triple or quadruple therapy was defined as involving multiple differential medicinal treatments: a proton pump inhibitor or H2 receptor blocker, clarithromycin or metronidazole, and amoxicillin or tetracycline, with or without bismuth  [H. pylori eradication therapy within 1 year of diagnosis of peptic ulcer disease were included in the early eradication cohort. Patients who received H. pylori eradication therapy after 1 year of being initially diagnosed were included in the late eradication cohort. The index date for patients was set as the date that they first received H. pylori eradication therapy. The exclusion criteria were missing data regarding date of birth, sex, or history of osteoporosis (ICD-9-CM codes 733.0–733.1) before the index date. Controls were randomly selected from the pool of participants without peptic ulcer disease who did not receive H. pylori eradication therapy. Four controls were frequency-matched to each H. pylori eradication case according to age (every 5 y span), sex, and year of receiving H. pylori eradication therapy.We recruited patients who were aged 20 years or above who had been diagnosed with peptic ulcer disease  and subsequently received viously) . These dThe general diagnostic process of the NHI program was based on physical examination and quantitative ultrasound. Diagnosis in partial patients was confirmed using dual energy x-ray absorptiometry following the guidance issued by the Health Promotion Administration, Ministry of Health and Welfare in Taiwan on osteoporosis diagnosis and treatment. All patients were followed until a diagnosis of osteoporosis was made or they were censored for loss to follow-up, withdrawal from the NHI program, or December 31, 2011, whichever occurred first. Coronary artery disease (CAD) (ICD-9-CM codes 410–414), alcohol-related illness , stroke (ICD-9-CM codes 430–438), chronic obstructive pulmonary disease (COPD) , asthma (ICD-9-CM code 493), rheumatoid arthritis (ICD-9-CM code 714), and steroid use were considered as covariates.t-test were used to determine differences in categorical and continuous variables between H. pylori eradication (including early and late H. pylori eradication therapy) and control cohorts. The cumulative incidence of osteoporosis among the three cohorts was plotted using the Kaplan–Meier method, and the difference was tested using a log-rank test. The incidence density rate of osteoporosis was calculated for each instance of early H. pylori eradication therapy, late H. pylori eradication therapy, and for the control cohort. The incidence rate ratios of the H. pylori eradication cohorts to that of the control cohort and the 95% confidence interval (CI) were estimated using a Poisson regression model. Multivariable Cox proportional hazard regression analysis was performed to estimate the relative hazard ratios (HRs) and 95% CIs of osteoporosis development for the H. Pylori eradication cohorts adjusted for age, sex, and comorbidities of CAD, alcohol-related illness, stroke, COPD, asthma, rheumatoid arthritis, and steroid use. All data analyses were performed using the SAS statistical package . A 2-tailed P<0.05 indicated statistical significance.The chi-square test and Student’s H. pylori eradication therapy—comprising 2,721 who received early H. pylori eradication therapy and 2,726 who received late H. pylori eradication therapy—and 21,788 controls were investigated . The mean age was 51.1±14.3 years in the H. pylori eradication cohorts and 50.6±14.7 in the control cohort. Both the early and late H. pylori eradication cohorts exhibited a higher prevalence of CAD, alcohol-related illness, COPD, asthma, rheumatoid arthritis, and steroid use compared with the control cohort (P<0.05). The mean follow-up periods were 5.91±3.10 years in the early H. pylori eradication cohort, 5.07±2.77 years in the late H. pylori eradication cohort, and 5.52±2.96 years in the control cohort. After 12 years of follow-up, the cumulative incidence of osteoporosis was higher in the early and late H. pylori eradication cohorts than in the control cohort .In this study, 5,447 patients who received stigated . Among tH. pylori eradication cohort, late H. pylori eradication cohort, and control cohort, respectively  and late H. pylori eradication cohort  than in the control cohort. The overall incidence and risk of osteoporosis were compared in the late H. pylori eradication cohort and control cohort according to several variables including age, sex, and comorbidities. The risk of osteoporosis in the late H. pylori eradication cohort was also higher than that of the control cohort. The risk of osteoporosis in patients receiving early H. pylori eradication therapy was also higher than that of the control cohort in all cases, except for those aged ≤49 years.After adjustment for age, sex, and comorbidities, the risk of developing osteoporosis was higher in the early H. pylori eradication cohort and the late H. pylori eradication cohort according to several variables including age, sex, and comorbidities. The risk of osteoporosis in patients receiving early H. pylori eradication therapy was lower than that of the late eradication cohort; however, this difference was not statistically significant  .For patients receiving early 32–2.16) . There wH. pylori eradication therapy, those though to have chronic H. pylori infection, have a higher incidence of osteoporosis. The late eradication group, which had a longer period of chronic H. pylori infection, had a higher incidence of osteoporosis than the early eradication group. When the follow-up period was over 5 years, the incidence of osteoporosis among the early H. pylori eradication cohort returned to the levels observed in the control cohort, while the incidence in the late eradication group remained higher. We believe the higher incidence of osteoporosis within the first 5 years could be the result of chronic H. pylori infection before H. pylori eradication therapy, and people could benefit from early eradication therapy when the follow-up period is greater than 5 years.The main finding of this population-based retrospective cohort study in Taiwan is that people who received The NHIRD covers almost 99% of the population of Taiwan and is a representative data source that includes age, sex, and comorbidity information. It enabled this study—the first of its type nationwide—to be conducted with population-based data and a near-decade follow-up period to date. Although our results do not directly indicate the causal relationship and etiological treatise for this association, based on literature review, we believe that there are several possible explanations.H. pylori infection may be the reason for the increased incidence of osteoporosis observed after eradication therapy. First, chronic H. pylori infection may alter calcium absorption. H. pylori can either increase gastric acid production, which predisposes one to duodenal ulceration, or decrease gastric acid production, thus leading to pangastritis or gastric ulceration [H. pylori results in atrophic gastritis, which increases the likelihood of osteoporosis [H. pylori infection increased the subsequent risk of end-stage renal disease (ESRD) [H. pylori infection had lower levels of vitamin B12 [Chronic ceration , 30. Theceration , 32. Theoporosis , 33. Sece (ESRD) . Vitamine (ESRD) , 35. Furamin B12 . In addiamin B12 .H. pylori infection may cause chronic systemic inflammation that negatively affects bone mineral density. H. pylori is the most common chronic bacterial infection of the human upper gastrointestinal tract [H. pylori eradication groups in our study. Early eradication of H. pylori may prevent chronic inflammation and the subsequent prolonged elevation of serum levels of inflammatory cytokines that lead to osteoporosis, as described by Moss et al. in 1994 [al tract , 38, andal tract , 40. An al tract . This is in 1994 .H. pylori infection before eradication therapy, rather than the effect of eradication therapy. If it were the effect of eradication therapy, the early eradication group might exhibit a higher incidence of osteoporosis, especially in the early years after therapy. In contrary, although non-significant, we observed a lower incidence of osteoporosis among the early H. pylori eradication group compared with the late H. pylori eradication group , laboratory data, or biomarkers. Osteoporosis could be affected by these confounding factors, and may have influenced our results. Second, the NHIRD database recorded only patients who received therapy for H. pylori infection. Early eradication of H. pylori had a relatively lower incidence of osteoporosis when compared with the late eradication group. When the follow-up period was over 5 years, there was no difference between the control group and early H. pylori eradication group. This provides evidence that encourages physicians to manage H. pylori infections early in patients with a high risk of osteoporosis. Additional studies are necessary to clarify the relationship between H. pylori infection and eradication, and osteoporosis, after adjustment for confounding factors, and to identify the mechanism of the relationship between H. pylori and osteoporosis.In this study, we found a higher incidence of osteoporosis among people with