Daily yogurt consumption does not affect bone turnover markers in men and postmenopausal women of Caribbean Latino descent: a randomized controlled trial.

BACKGROUND: Caribbean Latino adults are at high risk for osteoporosis yet remain underrepresented in bone research. This increased risk is attributed to genetics, diet, and lifestyle known to drive inflammation and microbial dysbiosis. OBJECTIVE: The primary objective of this study was to determine whether consuming 5 oz of yogurt daily for 8wks improves bone turnover markers (BTMs) among Caribbean Latino adults > 50 years; and secondarily to determine the impact on the gut microbiota and markers of intestinal integrity and inflammation. METHODS: Following a 4wk baseline period, participants were randomized to an 8wk whole fat yogurt intervention (n = 10) daily, containing only Streptococcus thermophilus and Lactobacillus bulgaricus, or to an untreated control group that did not consume yogurt (n = 10). Blood and stool samples collected at week-0 and week-8 were used to assess BTMs, inflammation, intestinal integrity biomarkers, and gut microbiota composition, short chain fatty acids (SCFAs), respectively. Data were evaluated for normality and statistical analyses were performed. RESULTS: Participants were 55% women, with a mean age of 70 ± 9 years, BMI 30 ± 6 kg/m2, and serum C-reactive protein 4.8 ± 3.6 mg/L, indicating chronic low-grade inflammation. Following 8wks of yogurt intake, absolute change in BTMs did not differ significantly between groups (P = 0.06-0.78). Secondarily, absolute change in markers of inflammation, intestinal integrity, and fecal SCFAs did not differ significantly between groups (P range 0.13-1.00). Yogurt intake for 8wks was significantly associated with microbial compositional changes of rare taxa (P = 0.048); however, no significant alpha diversity changes were observed. CONCLUSIONS: In this study, daily yogurt did not improve BTMs, inflammation, intestinal integrity, nor SCFAs. However, yogurt did influence beta diversity, or the abundance of rare taxa within the gut microbiota of the yogurt group, compared to controls. Additional research to identify dietary approaches to reduce osteoporosis risk among Caribbean Latino adults is needed. TRIAL REGISTRATION: This study is registered to ClinicalTrials.gov, NCT05350579 (28/04/2022).

The intestinal microbiota predicts COVID-19 severity and fatality regardless of hospital feeding method.

SARS-CoV-2-positive patients exhibit gut and oral microbiome dysbiosis, which is associated with various aspects of COVID-19 disease (1-4). Here, we aim to identify gut and oral microbiome markers that predict COVID-19 severity in hospitalized patients, specifically severely ill patients compared to moderately ill ones. Moreover, we investigate whether hospital feeding (solid versus enteral), an important cofounder, influences the microbial composition of hospitalized COVID-19 patients. We used random forest classification machine learning models with interpretable secondary analyses. The gut, but not the oral microbiota, was a robust predictor of both COVID-19-related fatality and severity of hospitalized patients, with a higher predictive value than most clinical variables. In addition, perturbations of the gut microbiota due to enteral feeding did not associate with species that were predictive of COVID-19 severity. IMPORTANCE SARS-CoV-2 infection leads to wide-ranging, systemic symptoms with sometimes unpredictable morbidity and mortality. It is increasingly clear that the human microbiome plays an important role in how individuals respond to viral infections. Our study adds to important literature about the associations of gut microbiota and severe COVID-19 illness during the early phase of the pandemic before the availability of vaccines. Increased understanding of the interplay between microbiota and SARS-CoV-2 may lead to innovations in diagnostics, therapies, and clinical predictions.

Dietary Intake of Pregnant Women with and without Inflammatory Bowel Disease in the United States.

BACKGROUND: Pregnancy is a vulnerable time where the lives of mother and baby are affected by diet, especially high-risk pregnancies in women with inflammatory bowel disease (IBD). Limited research has examined diet during pregnancy with IBD. AIMS: Describe and compare the diet quality of pregnant women with and without IBD, and examine associations between dietary intake and guidelines during pregnancy. METHODS: Three 24 h recalls were utilized to assess the diets of pregnant women with IBD (n = 88) and without IBD (n = 82) during 27-29 weeks of gestation. A customized frequency questionnaire was also administered to measure pre- and probiotic foods. RESULTS: Zinc intake (p = 0.02), animal protein (g) (p = 0.03), and ounce equivalents of whole grains (p = 0.03) were significantly higher in the healthy control (HC) group than the IBD group. Nutrients of concern with no significant differences between groups included iron (3% IBD and 2% HC met the goals), saturated fat (only 1% of both groups met the goals), choline (23% IBD and 21% HC met the goals), magnesium (38% IBD and 35% HC met the goals), calcium (48% IBD and 60% HC met the goals), and water intake (49% IBD and 48% HC met the goals). CONCLUSIONS: Most pregnant women in this cohort fell short of the dietary nutrients recommended in pregnancy, especially concerning for women with IBD.

DietaAnti-Inflamatoria or DAIN: A Crohn's disease management strategy tailored for Puerto Ricans.

Diet has been increasingly shown to be of therapeutic benefit for patients with inflammatory bowel diseases (IBD), especially Crohn's disease (CD). Yet dietary guidelines are nonexistent. Moreover, diets tailored to Puerto Ricans with IBD living on the island, have not been developed and tested. The rising prevalence of IBD in Puerto Rico warrants exploring the use of diet as part of the treatment strategies for these patients [1]. Here, we describe the study design of "Dieta Anti-Inflamatoria" or DAIN, a parallel two-arm randomized pilot trial aiming at testing the efficacy of IBD-Anti-inflammatory diet (IBD-AID) adapted for adults with CD living in Puerto Rico (clinical trial registration number: NCT05627128). We tailored the IBD-AID to the local cuisine preferences and food availability by creating and adapting recipes consistent with the IBD-AID principles [2,3]. In focus groups with a Community Research Advisory Panel and one-on-one consultations with implementation experts, we identified several aspects of the intervention to adapt before the implementation. The objectives of the stakeholder/expert-informed adaptation were to improve feasibility and compliance while developing the culturally tailored dietary intervention. DAIN was designed for adults living in Puerto Rico with CD and geared to be affordable, appropriate, and acceptable for patients with mild-to-moderate CD. The significance of this work is the validation of culturally appropriate nutritional guidelines to help manage CD symptoms. DAIN provides a blueprint for a comprehensive nutritional program that can be adapted to regional preferences and local food availability allowing wider implementation of diet as an adjunct treatment in diverse clinical settings.

The microbiota of pregnant women with SARS-CoV-2 and their infants.

BACKGROUND: Infants receive their first bacteria from their birthing parent. This newly acquired microbiome plays a pivotal role in developing a robust immune system, the cornerstone of long-term health. RESULTS: We demonstrated that the gut, vaginal, and oral microbial diversity of pregnant women with SARS-CoV-2 infection is reduced, and women with early infections exhibit a different vaginal microbiota composition at the time of delivery compared to their healthy control counterparts. Accordingly, a low relative abundance of two Streptococcus sequence variants (SV) was predictive of infants born to pregnant women with SARS-CoV-2 infection. CONCLUSIONS: Our data suggest that SARS-CoV-2 infections during pregnancy, particularly early infections, are associated with lasting changes in the microbiome of pregnant women, compromising the initial microbial seed of their infant. Our results highlight the importance of further exploring the impact of SARS-CoV-2 on the infant's microbiome-dependent immune programming. Video Abstract.

SARS-CoV-2 Serosurveys: How Antigen, Isotype and Threshold Choices Affect the Outcome.

BACKGROUND: Evaluating the performance of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) serological assays and clearly articulating the utility of selected antigens, isotypes, and thresholds is crucial to understanding the prevalence of infection within selected communities. METHODS: This cross-sectional study, implemented in 2020, screened PCRconfirmed coronavirus disease 2019 patients (n 86), banked prepandemic and negative samples (n 96), healthcare workers and family members (n 552), and university employees (n 327) for antiSARS-CoV-2 receptor-binding domain, trimeric spike protein, and nucleocapsid protein immunoglobulin (Ig)G and IgA antibodies with a laboratory-developed enzyme-linked immunosorbent assay and tested how antigen, isotype and threshold choices affected the seroprevalence outcomes. The following threshold methods were evaluated: (i) mean 3 standard deviations of the negative controls; (ii) 100 specificity for each antigen-isotype combination; and (iii) the maximal Youden index. RESULTS: We found vastly different seroprevalence estimates depending on selected antigens and isotypes and the applied threshold method, ranging from 0.0 to 85.4. Subsequently, we maximized specificity and reported a seroprevalence, based on more than one antigen, ranging from 9.3 to 25.9. CONCLUSIONS: This study revealed the importance of evaluating serosurvey tools for antigen-, isotype-, and threshold-specific sensitivity and specificity, to interpret qualitative serosurvey outcomes reliably and consistently across studies.

Dietary manipulation of the gut microbiome in inflammatory bowel disease patients: Pilot study.

Diet is a modifiable, noninvasive, inexpensive behavior that is crucial in shaping the intestinal microbiome. A microbiome "imbalance" or dysbiosis in inflammatory bowel disease (IBD) is linked to inflammation. Here, we aim to define the impact of specific foods on bacterial species commonly depleted in patients with IBD to better inform dietary treatment. We performed a single-arm, pre-post intervention trial. After a baseline period, a dietary intervention with the IBD-Anti-Inflammatory Diet (IBD-AID) was initiated. We collected stool and blood samples and assessed dietary intake throughout the study. We applied advanced computational approaches to define and model complex interactions between the foods reported and the microbiome. A dense dataset comprising 553 dietary records and 340 stool samples was obtained from 22 participants. Consumption of prebiotics, probiotics, and beneficial foods correlated with increased abundance of Clostridia and Bacteroides, commonly depleted in IBD cohorts. We further show that specific foods categorized as prebiotics or adverse foods are correlated to levels of cytokines in serum (i.e., GM-CSF, IL-6, IL-8, TNF-alpha) that play a central role in IBD pathogenesis. By using robust predictive analytics, this study represents the first steps to detangle diet-microbiome and diet-immune interactions to inform personalized nutrition for patients suffering from dysbiosis-related IBD.

Food as Treatment of Inflammatory Bowel Diseases.

Inflammatory bowel diseases (IBD), namely, Crohn's disease (CD) and ulcerative colitis (UC), are lifelong and incurable chronic inflammatory diseases affecting 6.8 million people worldwide. By 2030, the prevalence of IBD is estimated to reach 1% of the population in Western countries, and thus there is an urgent need to develop effective therapies to reduce the burden of this disease. Microbiome dysbiosis is at the heart of the IBD pathophysiology, and current research and development efforts for IBD treatments have been focused on gut microbiome regulation. Diet can shape the intestinal microbiome. Diet is also preferred over medication, is safe, and has been proven to be an effective strategy for the management of IBD. Therefore, although often overlooked, dietary interventions targeting the microbiome represent ideal treatments for IBD. Here, I summarize the latest research on diet as a treatment for IBD from infancy to adulthood, compile evidence of the mechanisms of action behind diet as treatment, and, lastly, provide insights into future research focusing on culturally tailored diets for ethnic minority groups with increased incidence of IBD yet underrepresented in nutrition research.

SepA Enhances Shigella Invasion of Epithelial Cells by Degrading Alpha-1 Antitrypsin and Producing a Neutrophil Chemoattractant.

Shigella spp. are highly adapted pathogens that cause bacillary dysentery in human and nonhuman primates. An unusual feature of Shigella pathogenesis is that this organism invades the colonic epithelia from the basolateral pole. Therefore, it has evolved the ability to disrupt the intestinal epithelial barrier to reach the basolateral surface. We have shown previously that the secreted serine protease A (SepA), which belongs to the family of serine protease autotransporters of Enterobacteriaceae, is responsible for the initial destabilization of the intestinal epithelial barrier that facilitates Shigella invasion. However, the mechanisms used by SepA to regulate this process remain unknown. To investigate the protein targets cleaved by SepA in the intestinal epithelium, we incubated a sample of homogenized human colon with purified SepA or with a catalytically inactive mutant of this protease. We discovered that SepA targets an array of 18 different proteins, including alpha-1 antitrypsin (AAT), a major circulating serine proteinase inhibitor in humans. In contrast to other serine proteases, SepA cleaved AAT without forming an inhibiting complex, which resulted in the generation of a neutrophil chemoattractant. We demonstrated that the products of the AAT-SepA reaction induce a mild but significant increase in neutrophil transepithelial migration in vitro. Moreover, the presence of AAT during Shigella infection stimulated neutrophil migration and dramatically enhanced the number of bacteria invading the intestinal epithelium in a SepA-dependent manner. We conclude that by cleaving AAT, SepA releases a chemoattractant that promotes neutrophil migration, which in turn disrupts the intestinal epithelial barrier to enable Shigella invasion. IMPORTANCEShigella is the second leading cause of diarrheal death globally. In this study, we identified the host protein targets of SepA, Shigella's major protein secreted in culture. We demonstrated that by cleaving AAT, a serine protease inhibitor important to protect surrounding tissue at inflammatory sites, SepA releases a neutrophil chemoattractant that enhances Shigella invasion. Moreover, SepA degraded AAT without becoming inhibited by the cleaved product, and SepA catalytic activity was enhanced at higher concentrations of AAT. Activation of SepA by an excess of AAT may be physiologically relevant at the early stages of Shigella infection, when the amount of synthesized SepA is very low compared to the concentration of AAT in the intestinal lumen. This observation may also help to explain the adeptness of Shigella infectivity at low dose, despite the requirement of reaching the basolateral side to invade and colonize the colonic epithelium.

Gut microbiota regulation of P-glycoprotein in the intestinal epithelium in maintenance of homeostasis.

BACKGROUND: P-glycoprotein (P-gp) plays a critical role in protection of the intestinal epithelia by mediating efflux of drugs/xenobiotics from the intestinal mucosa into the gut lumen. Recent studies bring to light that P-gp also confers a critical link in communication between intestinal mucosal barrier function and the innate immune system. Yet, despite knowledge for over 10 years that P-gp plays a central role in gastrointestinal homeostasis, the precise molecular mechanism that controls its functional expression and regulation remains unclear. Here, we assessed how the intestinal microbiome drives P-gp expression and function. RESULTS: We have identified a "functional core" microbiome of the intestinal gut community, specifically genera within the Clostridia and Bacilli classes, that is necessary and sufficient for P-gp induction in the intestinal epithelium in mouse models. Metagenomic analysis of this core microbial community revealed that short-chain fatty acid and secondary bile acid production positively associate with P-gp expression. We have further shown these two classes of microbiota-derived metabolites synergistically upregulate P-gp expression and function in vitro and in vivo. Moreover, in patients suffering from ulcerative colitis (UC), we find diminished P-gp expression coupled to the reduction of epithelial-derived anti-inflammatory endocannabinoids and luminal content (e.g., microbes or their metabolites) with a reduced capability to induce P-gp expression. CONCLUSION: Overall, by means of both in vitro and in vivo studies as well as human subject sample analysis, we identify a mechanistic link between cooperative functional outputs of the complex microbial community and modulation of P-gp, an epithelial component, that functions to suppress overactive inflammation to maintain intestinal homeostasis. Hence, our data support a new cross-talk paradigm in microbiome regulation of mucosal inflammation. Video abstract.

Associations between Diet, the Gut Microbiome, and Short-Chain Fatty Acid Production among Older Caribbean Latino Adults.

BACKGROUND: Caribbean Latino adults have disproportionately high prevalence of chronic disease; however, underlying mechanisms are unknown. Unique gut microbiome profiles and relation to dietary quality may underlie health disparities. OBJECTIVES: To examine the dietary quality of an underrepresented group of Caribbean Latino older adults with high prevalence of chronic disease; characterize gut microbiome profiles in this cohort; determine associations between dietary quality, gut microbiome composition, and short-chain fatty acid (SCFA) production; examine associations of clinical factors (body mass index, type 2 diabetes [T2D] status, and laxative use) with gut microbiome composition. DESIGN: The study design was cross-sectional. PARTICIPANTS/SETTING: Recruitment and interviews occurred at the Senior Center in Lawrence, MA, from September 2016-September 2017. A total of 20 adults aged ≥50 years, self-identified of Caribbean Latino origin, without use of antibiotics in 6 months or intestinal surgery were included in the study. EXPOSURE AND OUTCOME MEASURES: Diet was assessed by two, 24-hour recalls and dietary quality was calculated using the Healthy Eating Index 2015 and the Mediterranean Diet Score. The gut microbiome was assessed by 16S rRNA sequencing and fecal SCFA content. Anthropometrics (ie, weight and height) were measured by a trained interviewer, and self-reported laxative use, and other self-report health outcomes (ie, T2D status) were assessed by questionnaire. STATISTICAL ANALYSES: Faith Phylogenetic Diversity (alpha diversity) and unique fraction metric, or UniFrac (beta diversity) and nonphylogenetic metrics, including Shannon diversity index (alpha diversity) were calculated. Spearman correlations and group comparisons using Kruskal-Wallis test between alpha diversity indexes and nutrient intakes were calculated. Patterns in the microbiome were estimated using a partitioning around medoids with estimation of number of clusters, with optimum average silhouette width. Log odds were calculated to compare predefined nutrients and diet score components between microbiome clusters using multivariable logistic regression, controlling for age and sex. Pearson correlation was used to relate SCFA fecal content to individual nutrients and diet indexes. Final models were additionally adjusted for laxative use. Differences in lifestyle factors by gut microbiome cluster were tested by Fisher's exact test. RESULTS: Generally, there was poor alignment of participant's diets to either the Mediterranean Diet score or Healthy Eating Index 2015. Range in the Healthy Eating Index 2015 was 36 to 90, where only 5% (n=1) of the sample showed high adherence to the Dietary Guidelines for Americans. Mediterranean Diet scores suggested low conformance with a Mediterranean eating pattern (score range=2 to 8, where 45% scored ≤3 [poor adherence]). The gut microbiome separated into two clusters by difference in a single bacterial taxon: Prevotella copri (P copri) (permutational multivariate analysis of variance [PERMANOVA] R2=0.576, ADONIS function P=0.001). Significantly lower P copri abundance was observed in cluster 1 compared with cluster 2 (Mann-Whitney P<0.0001). Samples in the P copri dominated cluster 2 showed significantly lower alpha diversity compared with P copri depleted cluster 1 (Shannon diversity index P=0.01). Individuals in the P copri dominated cluster showed a trend toward higher 18:3 α-linolenic fatty acid intakes (P=0.09). Percentage of energy from total fat intake was significantly, positively correlated with fecal acetate (r=0.46; P=0.04), butyrate (r=0.50; P=0.03) and propionate (r=0.52; P=0.02). Associations between dietary intake and composition of the gut microbiome were attenuated by self-report recent laxative use. Individuals with T2D exhibited a significantly greater abundance of the Enterobacteriales (P=0.01) and a trend toward lower fecal content of butyric acid compared to subjects without T2D (P=0.08). Significant beta diversity differences were observed by weight (Mantel P<0.003) and body mass index (Mantel P<0.07). CONCLUSIONS: Two unique microbiome profiles, identified by abundance of P copri, were identified among Caribbean Latino adults. Microbiome profiles and SCFA content were associated with diet, T2D, and lifestyle. Further research is needed to determine the role of P copri and SCFA production in the risk for chronic disease and associated lifestyle predictors.

A dietary intervention to improve the microbiome composition of pregnant women with Crohn's disease and their offspring: The MELODY (Modulating Early Life Microbiome through Dietary Intervention in Pregnancy) trial design.

Crohn's disease (CD), a type of inflammatory bowel disease (IBD), is a chronic condition of the gastrointestinal tract that is caused by the loss of mucosal tolerance towards the commensal bacteria resulting in inflammatory responses. It has long been postulated that the gut microbiota, a complex and dynamic population of microorganisms, plays a key role in the pathogenesis of IBD. Maternal diagnosis of IBD has been identified as the greatest risk factor for IBD in offspring increasing the odds of developing the disease >4.5-fold. Moreover, babies born to mothers with IBD have demonstrated reduced gut bacterial diversity. There is accumulating evidence that the early life microbiota colonization is informed by maternal diet within the 3rd trimester of pregnancy. While babies born to mothers with IBD would pose an ideal cohort for intervention, no primary prevention measures are currently available. Therefore, we designed the MELODY (Modulating Early Life Microbiome through Dietary Intervention in Pregnancy) trial to test whether the IBD-AID™ dietary intervention during the last trimester of pregnancy can beneficially shift the microbiome of CD patients and their babies, thereby promoting a strong, effective immune system during a critical time of the immune system development. We will also test if favorable changes in the microbiome can lead to a reduced risk of postpartum CD relapse and lower mucosal inflammation in the offspring. This study will help create new opportunities to foster a healthy microbiome in the offspring at high risk of other immune-mediated diseases, potentially reducing their risk later in life.

Dysbiosis in a canine model of human fistulizing Crohn's disease.

Crohn's disease (CD) is a chronic immune-mediated inflammatory condition caused by the loss of mucosal tolerance toward the commensal microbiota. On average, 29.5% and 42.7% CD patients experience perianal complications at 10 and 20 y after diagnosis, respectively. Perianal CD (pCD) result in high disease burden, diminished quality of life, and elevated health-care costs. Overall pCD are predictors of poor long-term outcomes. Animal models of gut inflammation have failed to fully recapitulate the human manifestations of fistulizing CD. Here, we evaluated dogs with spontaneous canine anal furunculosis (CAF), a disease with clinical similarities to pCD, as a surrogate model for understanding the microbial contribution of human pCD pathophysiology. By comparing the gut microbiomes between dogs suffering from CAF (CAF dogs) and healthy dogs, we show CAF-dog microbiomes are either very dissimilar (dysbiotic) or similar (healthy-like), yet unique, to healthy dog's microbiomes. Compared to healthy or healthy-like CAF microbiomes, dysbiotic CAF microbiomes showed an increased abundance of Bacteroides vulgatus and Escherichia coli and a decreased abundance of Megamonas species and Prevotella copri. Our results mirror what have been reported in previous microbiome studies of patients with CD; particularly, CAF dogs exhibited two distinct microbiome composition: dysbiotic and healthy-like, with determinant bacterial taxa such as E. coli and P. copri that overlap what it has been found on their human counterpart. Thus, our results support the use of CAF dogs as a surrogate model to advance our understanding of microbial dynamics in pCD.

Substantial Increase in Compliance with Saturated Fatty Acid Intake Recommendations after One Year Following the American Heart Association Diet.

The American Heart Association (AHA) dietary guidelines recommend 30⁻35% of energy intake (%E) be from total fat, <7%E from saturated fatty acids (SFA), and <1%E from trans fatty acid (TFA). This study evaluates the effect of AHA dietary counselling on fat intake. Between 2009 and 2014, 119 obese adults with metabolic syndrome (MetS), (71% women, average 52.5 years of age, and 34.9 kg/m² of body mass index), received individual and group counselling on the AHA diet, over a one-year study period. Each participant attended 2 individual sessions (months 1 and 12) and 12 group sessions, at one-month intervals. At baseline and one-year, we collected three random 24-h diet recalls (two weekdays and one weekend day). Fat intake patterns over time were analyzed using paired-t test and linear mixed-effect models. There was significant variation on SFA and TFA intake per meal, being highest at dinner, in restaurants, and on weekends. Over the one-year study period, daily intake of total fat, SFA, and TFA decreased by 27%, 37% and 41%, respectively (p-value < 0.01, each). Correspondingly, the percentage of participants complying with AHA's recommendations, increased from 25.2% to 40.2% for total fat (p-value = 0.02); from 2.5% to 20.7% for SFA (p-value < 0.01); and from 45.4% to 62% for TFA (p-value = 0.02). Additionally, SFA intake for all meal types at home decreased significantly (p-value < 0.05, each). AHA dietary counselling significantly increased the compliance with AHA dietary guidelines, with an eightfold increase in compliance in SFA intake. Nonetheless, ~80% of our participants still exceeded the recommended SFA intake. Substantial efforts are needed to encourage low-SFA and low-TFA food preparation at home, with strong public health policies to decrease SFA and TFA in restaurants and prepared foods.

Shigella depends on SepA to destabilize the intestinal epithelial integrity via cofilin activation.

Shigella is unique among enteric pathogens, as it invades colonic epithelia through the basolateral pole. Therefore, it has evolved the ability to breach the intestinal epithelial barrier to deploy an arsenal of effector proteins, which permits bacterial invasion and leads to a severe inflammatory response. However, the mechanisms used by Shigella to regulate epithelial barrier permeability remain unknown. To address this question, we used both an intestinal polarized model and a human ex-vivo model to further characterize the early events of host-bacteria interactions. Our results showed that secreted Serine Protease A (SepA), which belongs to the serine protease autotransporter of Enterobacteriaceae family, is responsible for critically disrupting the intestinal epithelial barrier. Such disruption facilitates bacterial transit to the basolateral pole of the epithelium, ultimately fostering the hallmarks of the disease pathology. SepA was found to cause a decrease in active LIM Kinase 1 (LIMK1) levels, a negative inhibitor of actin-remodeling proteins, namely cofilin. Correspondingly, we observed increased activation of cofilin, a major actin-polymerization factor known to control opening of tight junctions at the epithelial barrier. Furthermore, we resolved the crystal structure of SepA to elucidate its role on actin-dynamics and barrier disruption. The serine protease activity of SepA was found to be required for the regulatory effects on LIMK1 and cofilin, resulting in the disruption of the epithelial barrier during infection. Altogether, we demonstrate that SepA is indispensable for barrier disruption, ultimately facilitating Shigella transit to the basolateral pole where it effectively invades the epithelium.

Phylogeographic evidence of cognate recognition site patterns and transformation efficiency differences in H. pylori: theory of strain dominance.

BACKGROUND: Helicobacter pylori has diverged in parallel to its human host, leading to distinct phylogeographic populations. Recent evidence suggests that in the current human mixing in Latin America, European H. pylori (hpEurope) are increasingly dominant at the expense of Amerindian haplotypes (hspAmerind). This phenomenon might occur via DNA recombination, modulated by restriction-modification systems (RMS), in which differences in cognate recognition sites (CRS) and in active methylases will determine direction and frequency of gene flow. We hypothesized that genomes from hspAmerind strains that evolved from a small founder population have lost CRS for RMS and active methylases, promoting hpEurope's DNA invasion. We determined the observed and expected frequencies of CRS for RMS in DNA from 7 H. pylori whole genomes and 110 multilocus sequences. We also measured the number of active methylases by resistance to in vitro digestion by 16 restriction enzymes of genomic DNA from 9 hpEurope and 9 hspAmerind strains, and determined the direction of DNA uptake in co-culture experiments of hspAmerind and hpEurope strains. RESULTS: Most of the CRS were underrepresented with consistency between whole genomes and multilocus sequences. Although neither the frequency of CRS nor the number of active methylases differ among the bacterial populations (average 8.6 ± 2.6), hspAmerind strains had a restriction profile distinct from that in hpEurope strains, with 15 recognition sites accounting for the differences. Amerindians strains also exhibited higher transformation rates than European strains, and were more susceptible to be subverted by larger DNA hpEurope-fragments than vice versa. CONCLUSIONS: The geographical variation in the pattern of CRS provides evidence for ancestral differences in RMS representation and function, and the transformation findings support the hypothesis of Europeanization of the Amerindian strains in Latin America via DNA recombination.

Structure of the human gastric bacterial community in relation to Helicobacter pylori status.

The human stomach is naturally colonized by Helicobacter pylori, which, when present, dominates the gastric bacterial community. In this study, we aimed to characterize the structure of the bacterial community in the stomach of patients of differing H. pylori status. We used a high-density 16S rRNA gene microarray (PhyloChip, Affymetrix, Inc.) to hybridize 16S rRNA gene amplicons from gastric biopsy DNA of 10 rural Amerindian patients from Amazonas, Venezuela, and of two immigrants to the United States (from South Asia and Africa, respectively). H. pylori status was determined by PCR amplification of H. pylori glmM from gastric biopsy samples. Of the 12 patients, 8 (6 of the 10 Amerindians and the 2 non-Amerindians) were H. pylori glmM positive. Regardless of H. pylori status, the PhyloChip detected Helicobacteriaceae DNA in all patients, although with lower relative abundance in patients who were glmM negative. The G2-chip taxonomy analysis of PhyloChip data indicated the presence of 44 bacterial phyla (of which 16 are unclassified by the Taxonomic Outline of the Bacteria and Archaea taxonomy) in a highly uneven community dominated by only four phyla: Proteobacteria, Firmicutes, Actinobacteria and Bacteroidetes. Positive H. pylori status was associated with increased relative abundance of non-Helicobacter bacteria from the Proteobacteria, Spirochetes and Acidobacteria, and with decreased abundance of Actinobacteria, Bacteroidetes and Firmicutes. The PhyloChip detected richness of low abundance phyla, and showed marked differences in the structure of the gastric bacterial community according to H. pylori status.