The CpxRA two-component system of adherent and invasive Escherichia coli contributes to epithelial cell invasion and early-stage intestinal fitness in a dysbiotic mouse model mediated by type 1 fimbriae expression.

Adherent and invasive Escherichia coli (AIEC) is a pathobiont that is involved in the onset and exacerbation of Crohn's disease. Although the inducible expression of virulence traits is a critical step for AIEC colonization in the host, the mechanism underlying AIEC colonization remains largely unclear. We here showed that the two-component signal transduction system CpxRA contributes to AIEC gut competitive colonization by activating type 1 fimbriae expression. CpxRA from AIEC strain LF82 functioned as a transcriptional regulator, as evidenced by our finding that an isogenic cpxRA mutant exhibits reduced expression of cpxP, a known regulon gene. Transcription levels of cpxP in LF82 increased in response to envelope stress, such as exposure to antimicrobials compromising the bacterial membrane, whereas the cpxRA mutant did not exhibit this response. Furthermore, we found that the cpxRA mutant exhibits less invasiveness into host cells than LF82, primarily due to reduced expression of the type 1 fimbriae. Finally, we found that the cpxRA mutant is impaired in gut competitive colonization in a mouse model. The colonization defects were reversed by the introduction of a plasmid encoding the cpxRA gene or expressing the type 1 fimbriae. Our findings indicate that modulating CpxRA activity could be a promising approach to regulating AIEC-involved Crohn's disease.

Selective human milk oligosaccharide utilization by members of the Bifidobacterium pseudocatenulatum taxon.

Human milk oligosaccharides (HMOs) are essentially unaffected by the digestive enzymes of the nursling and are known for their ability to enrich certain microbial species in the infant gut microbiota, in particular bifidobacteria. HMO metabolism has been studied in various bifidobacterial species such as B. breve, B. bifidum, and B. longum subsp. infantis. In the current study, we describe differential growth abilities elicited by twenty-three newly isolated Bifidobacterium pseudocatenulatum strains on particular HMOs, such as 2'-fucosyllactose (2'FL), 3-fucosyllactose (3FL), lacto-N-tetraose (LNT), and lacto-N-neotetraose (LNnT). Through gene-trait matching and comparative genome analysis, we identified genes involved in the degradation of fucosylated HMOs in this strain set, while we employed a transcriptomic approach to facilitate the identification and characterization of genes and associated enzymes involved in LNT metabolism by strain B. pseudocatenulatum MM0196. A total of 252 publicly available genomes of the B. pseudocatenulatum taxon were screened for homologs of the glycosyl hydrolases (GHs) identified here as being required for selected HMO metabolism. From this analysis, it is clear that all members of this species possess homologs of the genes involved in LNT degradation, while genes required for degradation of fucosylated HMOs are variably present.IMPORTANCEOur findings allow a better understanding of the complex interaction between Bifidobacterium and its host and provide a roadmap toward future applications of B. pseudocatenulatum as a probiotic with a focus on infant health. Furthermore, our investigations have generated information on the role of HMOs in shaping the infant gut microbiota, thus also facilitating applications of HMOs in infant nutrition, with potential extension into the mature or adult gut microbiota. Supplementation of HMOs is known to result in the modulation of bacterial communities toward a higher relative abundance of bifidobacteria, which in turn enforces their ability to modulate particular immune functions and strengthen the intestinal barrier. This work may therefore inspire future studies to improve the formulation of neonatal nutritional products, aimed at facilitating the development of a healthy digestive and immune system and reducing the differences in gut microbiota composition observed between breastfed and formula-fed babies or full-term and preterm infants.

The role of bacterial metabolism in human gut colonization.

Can we anticipate the emergence of the next pandemic antibiotic-resistant bacterial clone? Addressing such an ambitious question relies on our ability to comprehensively understand the ecological and epidemiological factors fostering the evolution of high-risk clones. Among these factors, the ability to persistently colonize and thrive in the human gut is crucial for most high-risk clones. Nonetheless, the causes and mechanisms facilitating successful gut colonization remain obscure. Here, we review recent evidence that suggests that bacterial metabolism plays a pivotal role in determining the ability of high-risk clones to colonize the human gut. Subsequently, we outline novel approaches that enable the exploration of microbial metabolism at an unprecedented scale and level of detail. A thorough understanding of the constraints and opportunities of bacterial metabolism in gut colonization will foster our ability to predict the emergence of high-risk clones and take appropriate containment strategies.

Possible link between colonization of the gastrointestinal tract by Citrobacter rodentium in C57BL/6 mice and microbiota composition.

Colonization resistance, conferred by the host's microbiota through both direct and indirect protective actions, serves to protect the host from enteric infections. Here, we identified the specific members of the gut microbiota that impact gastrointestinal colonization by Citrobacter rodentium, a murine pathogen causing colonic crypt hyperplasia. The gut colonization levels of C. rodentium in C57BL/6 mice varied among breeding facilities, probably due to differences in microbiota composition. A comprehensive analysis of the microbiota revealed that specific members of the microbiota may influence gut colonization by C. rodentium, thus providing a potential link between the two.

Fatty Acid Homeostasis Tunes Flagellar Motility by Activating Phase 2 Flagellin Expression, Contributing to Salmonella Gut Colonization.

Long-chain-fatty-acid (LCFA) metabolism is a fundamental cellular process in bacteria that is involved in lipid homeostasis, energy production, and infection. However, the role of LCFA metabolism in Salmonella enterica serovar Typhimurium (S. Tm) gut infection remains unclear. Here, using a murine gastroenteritis infection model, we demonstrate involvement of LCFA metabolism in S. Tm gut colonization. The LCFA metabolism-associated transcriptional regulator FadR contributes to S. Tm gut colonization. fadR deletion alters the gene expression profile and leads to aberrant flagellar motility of S. Tm. Colonization defects in the fadR mutant are attributable to altered swimming behavior characterized by less frequently smooth swimming, resulting from reduced expression of the phase 2 flagellin FljB. Notably, changes in lipid LCFA composition by fadR deletion lead to reduced expression of fljB, which is restored by exogenous LCFA. Therefore, LCFA homeostasis may maintain proper flagellar motility by activating fljB expression, contributing to S. Tm gut colonization. Our findings improve the understanding of the effect of luminal LCFA on the virulence of enteric pathogens.

uvrY Deletion and Acetate Reduce Gut Colonization of Crohn's Disease-Associated Adherent-Invasive Escherichia coli by Decreasing Expression of Type 1 Fimbriae.

Adherent-invasive Escherichia coli (AIEC) is involved in onset and/or exacerbation of Crohn's disease (CD). AIEC adapts to the gut environment by altering gene expression programs, leading to successful gut-lumen colonization. However, the underlying mechanism of gut colonization is still far from clarified. Here, we show the role of UvrY, a response regulator of bacterial two-component signal transduction systems, in AIEC gut colonization. An AIEC mutant lacking the uvrY gene exhibited impairment of competitive colonization in the murine intestinal tract. UvrY contributes to functional expression of type 1 fimbriae by activating expression of small RNA CsrB, which confers adherence and invasion into epithelial cells on AIEC. In contrast, acetate suppresses the UvrY-dependent expression of type 1 fimbriae, resulting in less efficient cell invasion and attenuated gut colonization. Our findings might lead to therapeutic interventions for CD, in which inhibitions of UvrY activation and acetate supplementation reduce the colonization levels of AIEC by decreasing type 1 fimbria expression.

Comparison between Perianal Swab and Stool Specimens for Detecting Colonization with Extended-Spectrum Beta-Lactamase-Producing and Fluoroquinolone-Resistant Enterobacterales.

Stool specimens are frequently used to detect gastrointestinal tract colonization with antimicrobial-resistant enteric bacteria, but they cannot be rapidly collected. Perianal swab specimens can be collected more quickly and efficiently, but data evaluating their suitability as a specimen type for this purpose are sparse. We performed selective culture for extended-spectrum ß-lactamase-producing Enterobacterales (ESBL-E) and fluoroquinolone-resistant Enterobacterales (FQRE) using paired perianal swab and stool specimens that were collected within 1 day of each other from hematopoietic cell transplant recipients and patients with acute leukemia. Nineteen (7.6%) of 251 stool specimens yielded ESBL-E and 64 (26%) of 246 stool specimens yielded FQRE. The positive percent agreement of perianal swab specimens compared to stool specimens was 95% (18/19; 95% confidence interval [CI], 74% to 100%) for detecting ESBL-E and 95% (61/64; 95% CI, 87% to 99%) for detecting FQRE. The concordance between specimen types was 98% (95% CI, 97% to 100%). Perianal swabs are a reliable specimen type for surveillance of the gastrointestinal tract for ESBL-E and FQRE.

Bacterial Carriage of Genes Encoding Fibronectin-Binding Proteins Is Associated with Long-Term Persistence of Staphylococcus aureus in the Nasal and Gut Microbiota of Infants.

Staphylococcus aureus can colonize both the anterior nares and the gastrointestinal tract. However, colonization at these sites in the same individuals has not been studied, and the traits that facilitate colonization and persistence at these sites have not been compared. Samples from the nostrils and feces collected on 9 occasions from 3 days to 3 years of age in 65 infants were cultured; 54 samples yielded S. aureus. The numbers of nasal and fecal S. aureus strains increased rapidly during the first weeks and were similar at 1 month of age (>40% of infants colonized). Thereafter, nasal carriage declined, while fecal carriage remained high during the first year of life. Individual strains were identified, and their colonization patterns were related to their carriage of genes encoding adhesins and superantigenic toxins. Strains retrieved from both the nose and gut (n = 44) of an infant were 4.5 times more likely to colonize long term (≥3 weeks at both sites) than strains found only in the rectum/feces (n = 56) or only in the nose (n = 32) (P ≤ 0.001). Gut colonization was significantly associated with carriage of the fnbA gene, and long-term colonization at either site was associated with carriage of fnbA and fnbB. In summary, gut colonization by S. aureus was more common than nasal carriage by S. aureus in the studied infants. Gut strains may provide a reservoir for invasive disease in vulnerable individuals. Fibronectin-binding adhesins and other virulence factors may facilitate commensal colonization and confer pathogenic potential. IMPORTANCE S. aureus may cause severe infections and frequently colonizes the nose. Nasal carriage of S. aureus increases 3-fold the risk of invasive S. aureus infection. S. aureus is also commonly found in the gut microbiota of infants and young children. However, the relationships between the adhesins and other virulence factors of S. aureus strains and its abilities to colonize the nostrils and gut of infants are not well understood. Our study explores the simultaneous colonization by S. aureus of the nasal and intestinal tracts of newborn infants through 3 years of follow-up. We identify bacterial virulence traits that appear to facilitate persistent colonization of the nose and gut by S. aureus. This expands our current knowledge of the interplay between bacterial commensalism and pathogenicity. Moreover, it may contribute to the development of targeted strategies for combating S. aureus infection.

Is there evidence for bacterial transfer via the placenta and any role in the colonization of the infant gut? - a systematic review.

With the important role of the gut microbiome in health and disease, it is crucial to understand key factors that establish the microbial community, including gut colonization during infancy. It has been suggested that the first bacterial exposure is via a placental microbiome. However, despite many publications, the robustness of the evidence for the placental microbiome and transfer of bacteria from the placenta to the infant gut is unclear and hence the concept disputed. Therefore, we conducted a systematic review of the evidence for the role of the placental, amniotic fluid and cord blood microbiome in healthy mothers in the colonization of the infant gut. Most of the papers which were fully assessed considered placental tissue, but some studied amniotic fluid or cord blood. Great variability in methodology was observed especially regarding sample storage conditions, DNA/RNA extraction, and microbiome characterization. No study clearly considered transfer of the normal placental microbiome to the infant gut. Moreover, some studies in the review and others published subsequently reported little evidence for a placental microbiome in comparison to negative controls. In conclusion, current data are limited and provide no conclusive evidence that there is a normal placental microbiome which has any role in colonization of infant gut.

High prevalence of MDR gram-negative bacteria in feces of healthy blood donors in Mexico.

During the initial stage of a study to recruit universal intestinal microbiota donors in Mexico City, we found multiple "healthy" subjects that colonized with MDRO (Multidrug-resistant organisms). We aimed to describe clinical and demographic characteristics of these individuals. This was a prospective observational study. Participants were consecutively recruited among blood donors. A fecal sample was collected from each subject and analyzed at the same day in search of MDRO through chromographic culture media and, if growth observed, later confirmed by MALDI-TOF and susceptibility testing in Vitek 2 system. From July 2018 to March 2019, 85 individuals were screened for fecal colonization. Median age was 35 years (IQR 27-46 years), and 48/85 (56.4%) were males. Seventy-two (84.7%) subjects harbored at least one MDRO. ESBL-producing microorganisms were found in 72/85 (84.3%) subjects, and E. coli was the most frequent (63/85, 74.1%). Four samples (2 E. coli, 2 P. aeruginosa, 2.4% each) harbored carbapenem-resistant Enterobacteriaceae (CRE), together with an ESBL-producing microorganism. Antibiotic use (p = 0.06) and PPIs or H2-blockers intake (p = 0.03) were more common in the colonized subjects during the previous 6-month period. We report a high incidence of enteric colonization of healthy subjects with MDRO, a condition that may be related to antibiotics or PPIs/H2-blockers consumption. This surprisingly high MDRO colonization rate in potential FMT donors emphasizes the need for careful screening of donors to avoid possible transmission to FMT recipients.

Can a probiotic supplement in pregnancy result in transfer to the neonatal gut: A systematic review.

INTRODUCTION: The establishment of the neonatal gut microbiome is a crucial step that may have lifelong health implications. We aimed to systematically review evidence on maternal probiotic supplementation during pregnancy and vertical transfer of the corresponding strain to the infant gut. MATERIAL AND METHODS: Medline, CINAHL, Embase, Web of Science, and OVID were searched from inception to September 2018. Studies of maternal probiotic supplementation for a minimum duration of 2 weeks and analyses of neonatal stool samples were included. The primary outcome was presence of the specific probiotic strain in the infant stool. Electronic databases were searched for relevant studies and references were cross-checked. Risk of bias among included studies was assessed and data were extracted independently by two authors. RESULTS: Three studies were included in the review. Only one study was identified involving prenatal maternal probiotic supplementation alone. Neonatal colonization with the maternally administered probiotic was not demonstrated but supplementation with the probiotic influenced levels of a bacterial strain other than that found in the probiotic product. The other two studies identified included both prenatal and postnatal supplementation of either mother or infant. All three studies reported employing strain-specific isolation methodology to isolate the supplemented bacterial strain in infant stool but none used whole metagenome shotgun sequencing. CONCLUSIONS: Few studies investigating transfer of a specific probiotic bacterial strain from mother to infant were identified, showing inconclusive evidence of vertical transfer.

Multidrug-Resistant Gram-Negative Bacteria Decolonization in Immunocompromised Patients: A Focus on Fecal Microbiota Transplantation.

Antimicrobial resistance is an important issue for global health; in immunocompromised patients, such as solid organ and hematological transplant recipients, it poses an even bigger threat. Colonization by multidrug-resistant (MDR) bacteria was acknowledged as a strong risk factor to subsequent infections, especially in individuals with a compromised immune system. A growing pile of studies has linked the imbalance caused by the dominance of certain taxa populating the gut, also known as intestinal microbiota dysbiosis, to an increased risk of MDR bacteria colonization. Several attempts were proposed to modulate the gut microbiota. Particularly, fecal microbiota transplantation (FMT) was successfully applied to treat conditions like Clostridioides difficile infection and other diseases linked to gut microbiota dysbiosis. In this review we aimed to provide a look at the data gathered so far on FMT, focusing on its possible role in treating MDR colonization in the setting of immunocompromised patients and analyzing its efficacy and safety.

Salmonella enterica Serovar Typhimurium CpxRA Two-Component System Contributes to Gut Colonization in Salmonella-Induced Colitis.

Salmonella enterica, a common cause of diarrhea, has to colonize the gut lumen to elicit disease. In the gut, the pathogen encounters a vast array of environmental stresses that cause perturbations in the bacterial envelope. The CpxRA two-component system monitors envelope perturbations and responds by altering the bacterial gene expression profile. This allows Salmonella to survive under such harmful conditions. Therefore, CpxRA activation is likely to contribute to Salmonella gut infection. However, the role of the CpxRA-mediated envelope stress response in Salmonella-induced diarrhea is unclear. Here, we show that CpxRA is dispensable for the induction of colitis by S. enterica serovar Typhimurium, whereas it is required for gut colonization. We prove that CpxRA is expressed during gut infection and that the presence of antimicrobial peptides in growth media activates the expression of CpxRA-regulated genes. In addition, we demonstrate that a S Typhimurium strain lacking the cpxRA gene is able to cause colitis but is unable to continuously colonize the gut. Finally, we show that CpxRA-dependent gut colonization requires the host gut inflammatory response, while DegP, a CpxRA-regulated protease, is dispensable. Our findings reveal that the CpxRA-mediated envelope stress response plays a crucial role in Salmonella gut infection, suggesting that CpxRA might be a promising therapeutic target for infectious diarrhea.

The Genital Tract Virulence Factor pGP3 Is Essential for Chlamydia muridarum Colonization in the Gastrointestinal Tract.

The cryptic plasmid is essential for Chlamydia muridarum dissemination from the genital tract to the gastrointestinal (GI) tract. Following intravaginal inoculation, a C. muridarum strain deficient in plasmid-encoded pGP3 or pGP4 but not pGP5, pGP7, or pGP8 failed to spread to the mouse gastrointestinal tract, although mice infected with these strains developed productive genital tract infections. pGP3- or pGP4-deficient strains also failed to colonize the gastrointestinal tract when delivered intragastrically. pGP4 regulates pGP3, while pGP3 does not affect pGP4 expression, indicating that pGP3 is critical for C. muridarum colonization of the gastrointestinal tract. Mutants deficient in GlgA, a chromosome-encoded protein regulated by pGP4, also consistently colonized the mouse gastrointestinal tract. Interestingly, C. muridarum colonization of the gastrointestinal tract positively correlated with pathogenicity in the upper genital tract. pGP3-deficient C. muridarum strains did not induce hydrosalpinx or spread to the GI tract even when delivered to the oviduct by intrabursal inoculation. Thus, the current study not only has revealed that pGP3 is a novel chlamydial colonization factor in the gastrointestinal tract but also has laid a foundation for investigating the significance of gastrointestinal Chlamydia.

Tadpoles fed supplemented diet with probiotic bacterium isolated from the intestinal tract of bullfrog Lithobates catesbeianus: Haematology, cell activity and electron microscopy.

The aim of this study is to select and isolate autochthonous bacteria with probiotic potential for use in a supplemented diet for bullfrog tadpoles, Lithobates catesbeianus. A total of 20 strains of lactic acid bacteria were isolated. Nine out of these were used in the following in vitro assays: antagonism against pathogenic bacteria (ANT), antimicrobial activity from extracellular compounds (MIC), tolerance to bile salts (TBS), pH reduction, protease production, sensitivity to antimicrobial tetracycline, cell viability, growth rate and doubling time. Using these data was defined an ideotype (ideal strain) based on the best results. Distances were estimated with the Mahalanobis (D2) test, and the best candidates, presenting the shortest ideotype distances, were considered to be used. The best strain was found to be Lactobacillus plantarum because it presented 10.00 ± 0.50 mm of ANT against Aeromonas hydrophila, 3.99 ± 0.01 of MIC independent of pathogenic bacteria, 85.07 ± 0.01 of TBS, 4.20 ± 0.02 of final pH, 17.67 ± 1.15 of protease production, 13.50 ± 2.00 sensitivity to antimicrobial tetracycline, 9.36 ± 0.04 of cell viability, 0.20 ± 0.00 of growth rate and 3.46 ± 0.00 doubling time. Therefore this probiotic candidate was then supplemented (2.045 ± 1.07 × 107 colony forming unities. g-1) into the diets of bullfrog tadpoles for a period of 42 days. At the end of the trial, samples of blood and intestines were collected to verify the haematological alterations and the intestinal morphology using transmission and scanning electron microscopy. Tadpoles fed the supplemented diet showed successful lactic acid bacterium colonisation, an increased number of circulating thrombocytes, monocytes, eosinophil and LG-PAS+ and also an increase in the length and density of intestinal microvilli. This study shows the feasibility of using probiotics isolated from farmed bullfrogs as a supplement in the diets of tadpoles, providing a promising alternative for modulating the health of these animals.

Serum Antibody Profile during Colonization of the Mouse Gut by Candida albicans: Relevance for Protection during Systemic Infection.

Candida albicans is a commensal microorganism in the oral cavity and gastrointestinal and urogenital tracts of most individuals that acts as an opportunistic pathogen when the host immune response is reduced. Here, we established different immunocompetent murine models to analyze the antibody responses to the C. albicans proteome during commensalism, commensalism followed by infection, and infection (C, C+I, and I models, respectively). Serum anti-C. albicans IgG antibody levels were higher in colonized mice than in infected mice. The antibody responses during gut commensalism (up to 55 days of colonization) mainly focused on C. albicans proteins involved in stress response and metabolism and differed in both models of commensalism. Different serum IgG antibody-reactivity profiles were also found over time among the three murine models. C. albicans gut colonization protected mice from an intravenous lethal fungal challenge, emphasizing the benefits of fungal gut colonization. This work highlights the importance of fungal gut colonization for future immune prophylactic therapies.

Chlamydia muridarum with Mutations in Chromosomal Genes tc0237 and/or tc0668 Is Deficient in Colonizing the Mouse Gastrointestinal Tract.

Chlamydiae colonize the gastrointestinal tracts of both animals and humans. However, their medical significance remains unknown. We have previously shown that wild-type Chlamydia muridarum spreads to and establishes stable colonization of the gastrointestinal tract following intravaginal inoculation. In the present study, we found that C. muridarum with mutations in chromosomal genes tc0237 and/or tc0668 was defective in spreading to the mouse gastrointestinal tract, which correlated with its attenuated pathogenicity in the upper genital tract. This correlation was more consistent than that of chlamydial pathogenicity with ascending infection in the genital tract, since attenuated C. muridarum spread significantly less to the gastrointestinal tract but maintained robust ascending infection of the upper genital tract. Transcervical inoculation further confirmed the correlation between C. muridarum spreading to the gastrointestinal tract and its pathogenicity in the upper genital tract. Finally, defective spreading of C. muridarum mutants was due to their inability to colonize the gastrointestinal tract since intragastric inoculation did not rescue the mutants' colonization. Thus, promoting C. muridarum colonization of the gastrointestinal tract may represent a primary function of the TC0237 and TC0668 proteins. Correlation of chlamydial colonization of the gastrointestinal tract with chlamydial pathogenicity in the upper genital tract suggests a potential role for gastrointestinal chlamydiae in genital tract pathogenicity.

Fecal Microbiota Transplantation in Patients With Blood Disorders Inhibits Gut Colonization With Antibiotic-Resistant Bacteria: Results of a Prospective, Single-Center Study.

Background: Patients with blood disorders colonized with antibiotic-resistant bacteria (ARB) are prone to systemic infections that are difficult to treat. Reintroduction of commensal bacteria in a murine model of enterococcal colonization of the gut can lead to eradication of enterococci. We hypothesized that fecal microbiota transplantation (FMT) could be used to eradicate ARB in humans. Methods: Participants colonized with ARB were treated with intraduodenal FMT according to a prospective protocol (NCT02461199). The primary endpoint was complete ARB decolonization at 1 month after FMT. Secondary endpoints included safety assessment and partial ARB decolonization. Microbiome sequencing was performed to investigate the influence of microbial composition of the transplanted material on the outcome of FMT. Results: Twenty-five FMTs were performed in 20 participants (including 40% who had neutropenia) who were colonized by a median of 2 (range, 1-4) strains of ARB. The primary endpoint was reached in 15/25 (60%) of the FMTs and more frequently in cases in which there was no periprocedural use of antibiotics (79% vs 36%, P < .05). Among participants, 15/20 (75%) experienced complete ARB decolonization. There were no severe adverse events, and partial ARB decolonization was observed in 20/25 (80%) of the FMTs. The microbiota composition analysis revealed higher abundance of Barnesiella spp., Bacteroides, and Butyricimonas and greater bacterial richness in the fecal material, resulting in eradication of Klebsiella pneumoniae compared with nonresponders. Conclusions: FMT in patients with blood disorders is safe and promotes eradication of ARB from the gastrointestinal tract. Clinical Trials Registration: NCT02461199.

Risk Factors and Outcomes of Infections by Multidrug-Resistant Gram-Negative Bacteria in Patients Undergoing Hematopoietic Stem Cell Transplantation.

The objective of this study was to determine risk factors and outcomes of infections by multidrug-resistant gram-negative (MDR GN) bacteria in 241 recipients of hematopoietic stem cell transplantation (HSCT). The cumulative incidence of infections was 10.5% (95% CI, 12.0% to 25.8%), with 57% of infections occurring during the period of severe neutropenia (neutrophil count < .1 × 106/L). In multivariate analysis, allogeneic transplant and colonization with MDR GN bacteria at admission to the transplant unit were significantly associated with an increased risk of infection. Although we observed neither transplant-related mortality (TRM) nor deaths due to infections by MDR GN bacteria after autologous transplant, in the allogeneic setting a significant difference was reported in terms of overall survival (OS) and TRM between patients who developed infections and those who did not (1-year OS, 39% versus 68%; 1-year TRM, 42% versus 19%). In multivariate analysis, refractory disease and development of grades III to IV graft-versus-host disease (GVHD) were factors that affected both TRM and OS, whereas occurrence of infections by MDR GN pathogens significantly reduced OS. We conclude that eligibility to allogeneic HSCT in MDR GN bacteria carriers should be carefully evaluated together with all other factors that independently influence outcome (disease status, donor, and GVHD risk).

Using long-term experimental evolution to uncover the patterns and determinants of molecular evolution of an Escherichia coli natural isolate in the streptomycin-treated mouse gut.

Although microbial ecology of the gut is now a major focus of interest, little is known about the molecular determinants of microbial adaptation in the gut. Experimental evolution coupled with whole-genome sequencing can provide insights of the adaptive process. In vitro experiments have revealed some conserved patterns: intermediate convergence, and epistatic interactions between beneficial mutations and mutations in global regulators. To test the relevance of these patterns and to identify the selective pressures acting in vivo, we have performed a long-term adaptation of an E. coli natural isolate, the streptomycin-resistant strain 536, in the digestive tract of streptomycin-treated mice. After a year of evolution, a clone from 15 replicates was sequenced. Consistently with in vitro observations, the identified mutations revealed a strong pattern of convergence at the mutation, gene, operon and functional levels. Yet, the rate of molecular evolution was lower than in in vitro, and no mutations in global regulators were recovered. More specific targets were observed: the dgo operon, involved in the galactonate pathway that improved growth on D-galactonate, and rluD and gidB, implicated in the maturation of the ribosomes, which mutations improved growth only in the presence of streptomycin. As in vitro, the nonrandom associations of mutations within the same pathways suggested a role of epistasis in shaping the adaptive landscape. Overall, we show that 'evolve and sequence' approach coupled with an analysis of convergence, when applied to a natural isolate, can be used to study adaptation in vivo and uncover the specific selective pressures of that environment.