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Eosinophils may reside in the lower intestine to play several homeostatic functions. Regulation of IgA+plasma-cell (PC) homeostasis is one of these functions. Here, we assessed regulation of expression for a proliferation-inducing ligand (APRIL), a key factor from the TNF superfamily for PC homeostasis, in eosinophils from the lower intestine. We observed a strong heterogeneity, since duodenum eosinophils did not produce APRIL at all, whereas a large majority of eosinophils from the ileum and right colon produced it. This was evidenced both in the human and mouse adult systems. At these places, the human data showed that eosinophils were the only cellular sources of APRIL. The number of IgA+PCs did not vary along the lower intestine, but ileum and right colon IgA+PC steady-state numbers significantly diminished in APRIL-deficient mice. Use of blood cells from healthy donors demonstrated that APRIL expression in eosinophils is inducible by bacterial products. Use of germ-free and antibiotics-treated mice confirmed the dependency on bacteria for APRIL production by eosinophils from the lower intestine. Taken together, our study shows that APRIL expression by eosinophils is spatially regulated in the lower intestine with a consequence on the APRIL dependency for IgA+PC homeostasis.The study demonstrates that microbiotal component(s) from the ileum and colon induce(s) APRIL expression in bloodborne eosinophils to mediate plasma-cell survival in the lamina propria. This APRIL-dependent survival pathway is not active in the duodenum.

ObjectiveOur aim was to prospectively assess the antibiotic resistance rates in Helicobacter pyloristrains in Europe in 2018 and to study the link between antibiotic consumption in the community and H. pyloriresistance levels in the different countries.DesignThe proportion of primary antibiotic resistance cases of H. pyloriand their corresponding risk factors were investigated in 24 centres from 18 European countries according to a standardised protocol. Data on antibiotic consumption in the community were collected for the period 2008–2017. The link between antibiotic consumption and resistance data was assessed using generalised linear mixed models. The model with the best fit was selected by means of the Akaike Information Criterion.ResultsH. pyloriresistance rates for the 1211 adult patients included were 21.4% for clarithromycin, 15.8% for levofloxacin and 38.9% for metronidazole and were significantly higher in Central/Western and Southern than in the Northern European countries.The best model fit was obtained for the Poisson distribution using 2013 consumption data. A significant association was found between H. pyloriclarithromycin resistance and consumption in the community of macrolides (p=0.0003) and intermediate-acting macrolides (p=0.005), and between levofloxacin resistance and consumption of quinolones (p=0.0002) and second-generation quinolones (p=0.0003).ConclusionThis study confirms the positive correlation between macrolide and quinolone consumption in the community and corresponding H. pyloriresistance in European countries. Hence, H. pyloritreatment with clarithromycin and levofloxacin should not be started without susceptibility testing in most European countries.

Gastric Mucosa Associated Lymphoid Tissue lymphoma (GML) development is triggered by Helicobacter pylori(H. pylori) infection. Little is known about the impact of H. pyloriinfection on gastric microbiota.

Aim:Helicobacter pyloriinfection is a worldwide infection, its eradication rates with conventional therapies have fallen to unacceptable levels. In this context we were interested in metformin, to determine its effect on H. pylorigrowth. Materials & methods:Antimicrobial susceptibility tests and survival curves were performed in vitroand a H. pylori-infected mice model was used to determine metformin effect in vivo. Results:Helicobacter pylorisurvival and growth were decreased in presence of metformin. Furthermore, metformin-treated mice had significantly less bacteria in their stomach than the untreated mice. Conclusion:Our work is the first to demonstrate a direct antimicrobial effect of metformin on H. pylori, indicating that this molecule has not yet revealed its full potential.

APRIL is a member of the tumor necrosis factor cytokine family involved in the regulation of B-cell immunity. We present a study of the infection by Helicobacterspecies of transgenic (Tg) C57BL6 mice, ectopically expressing the human form of APRIL. Wild-type (WT) and APRIL Tg mice were infected with Helicobacter felisand Helicobacter pyloriand compared with noninfected animals. Mice were euthanized 18 months after infection, and inflammatory responses and histologic alterations were analyzed. Flow cytometry results revealed that WT-infected mice had less leukocyte infiltration than APRIL Tg-infected mice. In WT-infected mice, infiltrates in gastric tissues were predominantly composed of T cells, mainly CD4+for H. pyloriand CD8+for H. felis. In APRIL Tg-infected mice, leukocyte infiltrates were composed of B cells with few CD4+T cells for both species. B cells expressed B surface markers compatible with a marginal zone origin. These results were confirmed by immunohistochemistry. B cells in particular were involved in lymphoepithelial lesions, a hallmark of gastric MALT lymphoma. Monoclonality was observed in a few infiltrates in the presence of lymphoepithelial lesions. These results confirm the importance of APRIL in the development of gastric lymphoid infiltrates induced by Helicobacterspecies in vivo. We believe that APRIL Tg mice infected by Helicobacterspecies may represent a novel animal model of gastric lymphomagenesis.

Antimicrobial susceptibility testing is needed to adapt Helicobacter pyloritreatment to obtain the best results. Beside the standard phenotypic methods, molecular methods are increasingly used. The value of these molecular tests is that they are quick, independent of the transport conditions, easy to standardize, and commercial kits are available. In this article, these methods are reviewed, focusing on the determination of H pyloriresistance to macrolides and fluoroquinolones, and mentioning also the methods used for tetracycline and rifampin.

Neonatal thymectomy in BALB/c mice has been described as a model of gastric mucosa-associated lymphoid tissue (MALT) lymphoma (GML). By using this experimental system, we screened, for the first time to our knowledge, Helicobacter pyloriGML-associated strains for their capacity to promote disease. A cohort of BALB/c mice underwent thymectomy at day 3 after birth (d3Tx). Successful thymic ablation was evaluated by the degree of lymphopenia in blood samples collected at 4 weeks of age. d3Tx and non-thymectomized controls were infected with either GML strains (B38 or B47) or control strains (SS1 or TN2GF4). Gastric samples collected at 6, 12, and 18 months after infection were studied for bacteria content, and submitted to histological, immunochemical, molecular, and immunological analyses. Severe gastric inflammation was only observed in d3Tx mice. In these animals, the gastric lamina propria was infiltrated with lymphoid cells organized in follicles composed of B cells with few infiltrating T cells. PCR of D/J IgH gene segments proved the monoclonality of infiltrating B cells, which strongly correlated with the presence of lymphoepithelial lesions. B-cell infiltrates were particularly prominent in mice infected with the B47-GML strain. No pathological changes were detected in noninfected d3Tx mice. We identified new H. pyloriisolates adapted to the mouse stomach with high potential of GML development, which is only revealed in hosts rendered lymphopenic by neonatal thymic ablation.

Kingella kingae is an emerging osteoarticular pathogen in young children. Its isolation by traditional culture methods remains difficult, underscoring the need to implement other diagnostic methods for its detection and identification, such as nucleic acid amplification tests. Although the genome of this bacterium has not yet been sequenced, a toxin named RTX has been identified. The goal of this study was to develop sensitive, specific, and rapid molecular methods based on the rtxA toxin gene sequence to diagnose this infection. Two real-time PCR assays (SYBR green and TaqMan chemistries) targeting this gene are reported. Sensitivity and specificity were first evaluated successfully with 67 strains: 31 Kingella kingae isolates and 36 strains from other bacterial species. Then, 52 clinical specimens positive or negative by culture and/or PCR (16S rRNA and cpn60 genes) were tested with these assays. A nested PCR assay with subsequent sequencing was also developed to confirm the presence of Kingella kingae isolates in these clinical specimens. The results obtained demonstrate that these assays are accurate for the diagnosis of Kingella kingae infection.

ABSTRACTKingella kingaeis an emerging osteoarticular pathogen in young children. Its isolation by traditional culture methods remains difficult, underscoring the need to implement other diagnostic methods for its detection and identification, such as nucleic acid amplification tests. Although the genome of this bacterium has not yet been sequenced, a toxin named RTX has been identified. The goal of this study was to develop sensitive, specific, and rapid molecular methods based on the rtxAtoxin gene sequence to diagnose this infection. Two real-time PCR assays (SYBR green and TaqMan chemistries) targeting this gene are reported. Sensitivity and specificity were first evaluated successfully with 67 strains: 31 Kingella kingaeisolates and 36 strains from other bacterial species. Then, 52 clinical specimens positive or negative by culture and/or PCR (16S rRNA and cpn60genes) were tested with these assays. A nested PCR assay with subsequent sequencing was also developed to confirm the presence of Kingella kingaeisolates in these clinical specimens. The results obtained demonstrate that these assays are accurate for the diagnosis of Kingella kingaeinfection.

Helicobacter pylori infection plays a causal role in the development of gastric mucosa-associated lymphoid tissue (MALT) lymphoma (LG-MALT) and duodenal ulcer (DU). Although many virulence factors have been associated with DU, many questions remain unanswered regarding the evolution of the infection toward this exceptional event, LG-MALT. The present study describes and compares the complexome of two H. pylori strains, strain J99 associated with DU and strain B38 associated with LG-MALT, using the two-dimensional blue native/SDS-PAGE method. It was possible to identify 90 different complexes (49 and 41 in the B38 and J99 strains, respectively); 12 of these complexes were common to both strains (seven and five in the membrane and cytoplasm, respectively), reflecting the variability of H. pylori strains. The 44 membrane complexes included numerous outer membrane proteins, such as the major adhesins BabA and SabA retrieved from a complex in the B38 strain, and also proteins from the hor family rarely studied. BabA and BabB adhesins were found to interact independently with HopM/N in the B38 and J99 strains, respectively. The 46 cytosolic complexes essentially comprised proteins involved in H. pylori physiology. Some orphan proteins were retrieved from heterooligomeric complexes, and a function could be proposed for a number of them via the identification of their partners, such as JHP0119, which may be involved in the flagellar function. Overall, this study gave new insights into the membrane and cytoplasm structure, and those which could help in the design of molecules for vaccine and/or antimicrobial agent development are highlighted.

Helicobacter pyloriinfection plays a causal role in the development of gastric mucosa-associated lymphoid tissue (MALT) lymphoma (LG-MALT) and duodenal ulcer (DU). Although many virulence factors have been associated with DU, many questions remain unanswered regarding the evolution of the infection toward this exceptional event, LG-MALT. The present study describes and compares the complexome of two H. pyloristrains, strain J99 associated with DU and strain B38 associated with LG-MALT, using the two-dimensional blue native/SDS-PAGE method. It was possible to identify 90 different complexes (49 and 41 in the B38 and J99 strains, respectively); 12 of these complexes were common to both strains (seven and five in the membrane and cytoplasm, respectively), reflecting the variability of H. pyloristrains. The 44 membrane complexes included numerous outer membrane proteins, such as the major adhesins BabA and SabA retrieved from a complex in the B38 strain, and also proteins from the horfamily rarely studied. BabA and BabB adhesins were found to interact independently with HopM/N in the B38 and J99 strains, respectively. The 46 cytosolic complexes essentially comprised proteins involved in H. pyloriphysiology. Some orphan proteins were retrieved from heterooligomeric complexes, and a function could be proposed for a number of them via the identification of their partners, such as JHP0119, which may be involved in the flagellar function. Overall, this study gave new insights into the membrane and cytoplasm structure, and those which could help in the design of molecules for vaccine and/or antimicrobial agent development are highlighted.

The discovery of Helicobacter pylori in 1982 was the starting point of a revolution concerning the concepts and management of gastroduodenal diseases. It is now well accepted that the most common stomach disease, peptic ulcer disease, is an infectious disease, and all consensus conferences agree that the causative agent, H. pylori, must be treated with antibiotics. Furthermore, the concept emerged that this bacterium could be the trigger of various malignant diseases of the stomach, and it is now a model for chronic bacterial infections causing cancer. Most of the many different techniques involved in diagnosis of H. pylori infection are performed in clinical microbiology laboratories. The aim of this article is to review the current status of these methods and their application, highlighting the important progress which has been made in the past decade. Both invasive and noninvasive techniques will be reviewed.

SUMMARYThe discovery of Helicobacter pyloriin 1982 was the starting point of a revolution concerning the concepts and management of gastroduodenal diseases. It is now well accepted that the most common stomach disease, peptic ulcer disease, is an infectious disease, and all consensus conferences agree that the causative agent, H. pylori, must be treated with antibiotics. Furthermore, the concept emerged that this bacterium could be the trigger of various malignant diseases of the stomach, and it is now a model for chronic bacterial infections causing cancer. Most of the many different techniques involved in diagnosis of H. pyloriinfection are performed in clinical microbiology laboratories. The aim of this article is to review the current status of these methods and their application, highlighting the important progress which has been made in the past decade. Both invasive and noninvasive techniques will be reviewed.

ABSTRACTPeptic ulcer disease (PUD) occurs after a long-term Helicobacter pyloriinfection. However, the disease can develop earlier, and rare cases have been observed in children, suggesting that these H. pyloristrains may be more virulent. We used suppressive subtractive hybridization for comparative genomics between H. pyloristrains isolated from a 5-year-old child with duodenal ulcer and from a sex- and age-matched child with gastritis only. The prevalence of the 30 tester-specific subtracted sequences was determined on a collection of H. pyloristrains from children (15 ulcers and 30 gastritis) and from adults (46 ulcers and 44 gastritis). Two of these sequences, jhp0562 (80.0% versus 33.3%, P= 0.008) and jhp0870 (80.0% versus 36.7%, P= 0.015), were highly associated with PUD in children and a third sequence, jhp0828, was less associated (40.0% versus 10.0%, P= 0.048). Among adult strains, none of the 30 sequences was associated with PUD. However, both jhp0562 and jhp0870 were less prevalent in adenocarcinoma strains than in PUD strains from children and adults, the difference being statistically significant for jhp0870. In conclusion, two H. pylorigenes were identified as being strongly associated with PUD in children, and their putative roles as an outer membrane protein for jhp0870 and in lipopolysaccharide biosynthesis for jhp0562, suggest that they may be novel virulence factors of H. pylori.

Helicobacter pylori has been associated with the development of two malignant diseases: gastric adenocarcinoma and gastric mucosa-associated lymphoid tissue (MALT) lymphoma. Although the cag pathogenicity island, especially the cagA gene, has been linked with adenocarcinoma, few data concerning H. pylori pathogenic factors involved in low-grade gastric MALT lymphoma are available. The goal of this study was to analyze the prevalence of and correlation between genes coding for seven H. pylori virulence factors (cagA, cagE, vacA, iceA, babA, hopQ, and oipA) and two novel adhesins (sabA and hopZ) by comparing a collection of 43 H. pylori strains isolated from patients with low-grade gastric MALT lymphoma to 39 strains isolated from age-matched patients with gastritis only. Our results show that taken individually, none of the nine genes tested can be considered associated with MALT strains and allow us to conclude that MALT pathogenesis is not linked with more proinflammatory H. pylori strains. We demonstrated that in patients infected with strains harboring the iceA1 allele, sabA functional status, and hopZ "off" status, the odds of developing a MALT lymphoma were 10 times higher. However, the low prevalence of such strains (10 of 43 MALT strains) renders this triple association a low-sensitivity marker for MALT strains. Our data confirmed that H. pylori virulence factors are correlated with one another. If the involvement of H. pylori in MALT lymphoma is well established, the pathomechanism by which gastric lymphoma occurs remains to be identified.

ABSTRACTHelicobacter pylorihas been associated with the development of two malignant diseases: gastric adenocarcinoma and gastric mucosa-associated lymphoid tissue (MALT) lymphoma. Although the cagpathogenicity island, especially the cagAgene, has been linked with adenocarcinoma, few data concerning H. pyloripathogenic factors involved in low-grade gastric MALT lymphoma are available. The goal of this study was to analyze the prevalence of and correlation between genes coding for seven H. pylorivirulence factors (cagA, cagE, vacA, iceA, babA, hopQ, and oipA) and two novel adhesins (sabAand hopZ) by comparing a collection of 43 H. pyloristrains isolated from patients with low-grade gastric MALT lymphoma to 39 strains isolated from age-matched patients with gastritis only. Our results show that taken individually, none of the nine genes tested can be considered associated with MALT strains and allow us to conclude that MALT pathogenesis is not linked with more proinflammatory H. pyloristrains. We demonstrated that in patients infected with strains harboring the iceA1allele, sabAfunctional status, and hopZ“off” status, the odds of developing a MALT lymphoma were 10 times higher. However, the low prevalence of such strains (10 of 43 MALT strains) renders this triple association a low-sensitivity marker for MALT strains. Our data confirmed that H. pylorivirulence factors are correlated with one another. If the involvement of H. pyloriin MALT lymphoma is well established, the pathomechanism by which gastric lymphoma occurs remains to be identified.

We report the case of a 13-year-old boy with cystic fibrosis with a pulmonary exacerbation concomitant to the first isolation of Pandoraea sputorum. The imipenem and trimethoprim–sulfamethoxazole treatments failed, with persistence of the bacteria, bronchial congestion and a decline in lung function. Pandoraea sp.is rarely isolated, with only 10 cases reported in France in 2011.

We report the existence of eight different interleukin-15 receptor α-chain (IL-15Rα) transcripts resulting from exon-splicing mechanisms within the IL-15Rα gene. Two main classes of transcripts can be distinguished that do or do not (Δ2 isoforms) contain the exon 2-coding sequence. Both classes were expressed in numerous cell lines and tissues (including peripheral blood lymphocytes) at comparable levels and could be transcribed in COS-7 cells, and the proteins were expressed at the cell surface. Both receptor forms displayed numerous glycosylation states, reflecting differential usage of a single N-glycosylation site as well as extensive O-glycosylations. Whereas IL-15Rα bound IL-15 with high affinity, Δ2IL-15Rα was unable to bind IL-15, thus revealing the indispensable role of the exon 2-encoded domain in cytokine binding. A large proportion of IL-15Rα was expressed at the nuclear membrane with some intranuclear localization, supporting a potential direct action of the IL-15·IL-15Rα complex at the nuclear level. In sharp contrast, Δ2IL-15Rα was found only in the non-nuclear membrane compartments, indicating that the exon 2-encoded domain (which is shown to contain a potential nuclear localization signal) plays an important role in receptor post-translational routing. Together, our data indicate that exon 2 splicing of human IL-15Rα is a natural process that might play regulatory roles at different levels.

ABSTRACTA large database of Campylobacterisolates precisely identified at the species level was used to compare patients' characteristics. In a multivariate analysis, Campylobacter coliwas found more often in older patients and in patients having traveled abroad and less often in summertime than Campylobacter jejuni. Campylobacter fetusinfection occurred in much older patients and in hospitalized patients with a systemic disease.

The bacteriological diagnosis of intestinal bacterial infections has historically been based on culture on agar plates. However, culture may lack sensitivity, and some enteropathogens, such as pathovars of Escherichia coli, may escape routine diagnosis. Our goal was to evaluate the analytical performance of the Novodiag Bacterial GE+ kit for the detection of enteropathogenic bacteria in acute community diarrhea. We included 251 stools in this study (198 retrospective and 53 prospective).

A whole-genome sequencing (WGS) approach was conducted in order to identify the molecular determinants associated with antimicrobial resistance in 12 multidrug-resistant Campylobacter jejuniand Campylobacter coliisolates, with a focus on aminoglycoside resistance determinants. Two variants of a new aminoglycoside phosphotransferase gene [aph(2?)-Ii1and aph(2?)-Ii2] putatively associated with gentamicin resistance were found.

ABSTRACTThe detection of campylobacters in stools is performed essentially by culture, but this technique has a low sensitivity. New detection methods are now available. Among them, immunochromatography tests (ICTs) are very attractive in that they offer a result within 15 min. However, previous studies suggest that these tests have a relatively low specificity. The objective of this study was to evaluate the performance of these tests. During the study period, all patients who consulted the emergency units and had a stool culture were included. Their stool samples were tested with two ICTs, Ridaquick Campylobacter and ImmunoCardSTAT! Campy. Stools were also tested by a home-made PCR and two commercially available enzyme-linked immunosorbent assays (ELISAs) when one of the ICTs was positive. The composite reference standard (CRS) was defined as positive if the culture was positive or, in case of a negative culture, if the PCR and one of the ELISAs were positive simultaneously. Three hundred and five patients were included. Among the 50 positive specimens with Ridaquick Campylobacter, 47 were considered true positives by the CRS, corresponding to a positive predictive value (PPV) of 94.0%. Among the 52 positive specimens with ImmunoCardSTAT! Campy, 44 were considered true positives by the CRS, corresponding to a PPV of 84.6%. The negative predictive values were estimated at 94.9 and 92.4% for the Ridaquick Campylobacter and ImmunoCardSTAT! Campy tests, respectively. ICTs appear to be very efficient and allow a very rapid detection of campylobacters, which is important for treating early campylobacter infections with an adapted antibiotherapy.

ABSTRACTHelicobacter pylorichronically infects the gastric mucosa in more than half of the human population; in a subset of this population, its presence is associated with development of severe disease, such as gastric cancer. Genomic analysis of several strains has revealed an extensive H. pyloripan-genome, likely to grow as more genomes are sampled. Here we describe the draft genome sequence (63 contigs; 26× mean coverage) of H. pyloristrain B45, isolated from a patient with gastric mucosa-associated lymphoid tissue (MALT) lymphoma. The major finding was a 24.6-kb prophage integrated in the bacterial genome. The prophage shares most of its genes (22/27) with prophage region II of Helicobacter acinonychisstrain Sheeba. After UV treatment of liquid cultures, circular DNA carrying the prophage integrase gene could be detected, and intracellular tailed phage-like particles were observed in H. pyloricells by transmission electron microscopy, indicating that phage production can be induced from the prophage. PCR amplification and sequencing of the integrase gene from 341 H. pyloristrains from different geographic regions revealed a high prevalence of the prophage (21.4%). Phylogenetic reconstruction showed four distinct clusters in the integrase gene, three of which tended to be specific for geographic regions. Our study implies that phages may play important roles in the ecology and evolution of H. pylori.IMPORTANCEHelicobacter pylorichronically infects the gastric mucosa in more than half of the human population, and while most of the infected individuals do not develop disease, H. pyloriinfection doubles the risk of developing gastric cancer. An abundance and diversity of viruses (phages) infect microbial populations in most environments and are important mediators of microbial diversity. Our finding of a 24.6-kb prophage integrated inside an H. pylorigenome and the observation of circular integrase gene-containing DNA and phage-like particles inside cells upon UV treatment demonstrate that we have discovered a viable H. pyloriphage. The additional finding of integrase genes in a large proportion of screened isolates of diverse geographic origins indicates that the prevalence of prophages may have been underestimated in H. pylori. Since phages are important drivers of microbial evolution, the discovery should be important for understanding and predicting genetic diversity in H. pylori.