Faculty Bibliography

Background. Diabetic foot infections are a leading cause of lower extremity amputations. Our study examines the microbiota of diabetic skin prior to ulcer development or infection. Methods. In a case-control study, outpatient males were recruited at a veterans hospital. Subjects were swabbed at 4 cutaneous sites, 1 on the forearm and 3 on the foot. Quantitative polymerase chain reaction (qPCR) with primers and probes specific for bacteria, Staphylococcus species, Staphylococcus aureus, and fungi were performed on all samples. High-throughput 16S ribosomal RNA (rRNA) sequencing was performed on samples from the forearm and the plantar aspect of the foot. Results. qPCR analysis of swab specimens from 30 diabetic subjects and 30 control subjects showed no differences in total numbers of bacteria or fungi at any sampled site. Increased log concentrations of Staphylococcus aureus, quantified by the number of nuc gene copies, were present in diabetic men on the plantar aspect of the foot. High-throughput 16S rRNA sequencing found that, on the foot, the microbiota in controls (n = 24) was dominated by Staphylococcus species, whereas the microbiota in diabetics (n = 23) was more diverse at the genus level. The forearm microbiota had similar diversity in diabetic and control groups. Conclusions. The feet of diabetic men had decreased populations of Staphylococcus species, increased populations of S. aureus, and increased bacterial diversity, compared with the feet of controls. These ecologic changes may affect the risk for wound infections.

Objective The purpose of this study was to analyze the association of maternal antenatal therapy on initial preterm infant oral microbial acquisition of gut metabolically important bacteria: Firmicutes, Bacteroidetes, Lactobacillus, Bifidobacterium, and Bacteroides species.Study Design Infant oral samples were collected prefeeding at 24 hours and analyzed using group-specific primers by real-time 16S rRNA quantitative polymerase chain reaction with analysis of variance and logistic regression to evaluate effect of antenatal exposure.Results Sixty-five infants <34 weeks' gestational age (GA) were evaluated; mean GA was 28.6 +/- 2.6 (standard deviation) weeks. Infants unexposed to antenatal treatment (n = 5) acquired <1% Firmicutes, which was composed of 100% Lactobacillus species with no detectable Bifidobacterium, Bacteroidetes, or Bacteroides species. Infants exposed to antibiotics (n = 7), acquired fivefold less total bacterial density (TBD) with 45% Firmicutes 1.3% Lactobacillus species, 23.5% Bacteroidetes and rare Bacteroides. Compared with unexposed infants, steroids (n = 26) or steroid and antibiotics (n = 27) exposure led to an eightfold increase in TBD with <1% Lactobacillus species and Bacteroides species 100% and 30%, respectively (p < 0.04). Bifidobacterium was undetectable in all groups.Conclusion Preterm infant exposure to routine maternal antenatal treatments influence early oral microbial acquisition during the primary hours related to establishment of gut commensal bacteria.

Psoriasis is a common chronic inflammatory disease of the skin. We sought to use bacterial community abundance data to assess the feasibility of developing multivariate molecular signatures for differentiation of cutaneous psoriatic lesions, clinically unaffected contralateral skin from psoriatic patients, and similar cutaneous loci in matched healthy control subjects. Using 16S rRNA high-throughput DNA sequencing, we assayed the cutaneous microbiome for 51 such matched specimen triplets including subjects of both genders, different age groups, ethnicities and multiple body sites. None of the subjects had recently received relevant treatments or antibiotics. We found that molecular signatures for the diagnosis of psoriasis result in significant accuracy ranging from 0.75 to 0.89 AUC, depending on the classification task. We also found a significant effect of DNA sequencing and downstream analysis protocols on the accuracy of molecular signatures. Our results demonstrate that it is feasible to develop accurate molecular signatures for the diagnosis of psoriasis from microbiomic data.

We examined whether colonization of selected oral pathogens is associated with gastric precancerous lesions in a cross-sectional study. A total of 119 participants were included, of which 37 were cases of chronic atrophic gastritis, intestinal metaplasia, or dysplasia. An oral examination was performed to measure periodontal indices. Plaque and saliva samples were tested with real-time quantitative PCR for DNA levels of pathogens related to periodontal disease (Porphyromonas gingivalis, Tannerella forsythensis, Treponema denticola, Actinobacillus actinomycetemcomitans) and dental caries (Streptococcus mutans and S. sobrinus). There were no consistent associations between DNA levels of selected bacterial species and gastric precancerous lesions, although an elevated but non-significant odds ratio (OR) for gastric precancerous lesions was observed in relation to increasing colonization of A. actinomycetemcomitans (OR = 1.36 for one standard deviation increase, 95% Confidence Interval = 0.87-2.12), P. gingivalis (OR = 1.12, 0.67-1.88) and T. denticola (OR = 1.34, 0.83-2.12) measured in plaque. To assess the influence of specific long-term infection, stratified analyses by levels of periodontal indices were conducted. A. actinomycetemcomitans was significantly associated with gastric precancerous lesions (OR = 2.51, 1.13-5.56) among those with >/= median of percent tooth sites with PD>/=3 mm, compared with no association among those below the median (OR = 0.86, 0.43-1.72). A significantly stronger relationship was observed between the cumulative bacterial burden score of periodontal disease-related pathogens and gastric precancerous lesions among those with higher versus lower levels of periodontal disease indices (p-values for interactions: 0.03-0.06). Among individuals with periodontal disease, high levels of colonization of periodontal pathogens are associated with an increased risk of gastric precancerous lesions.

Our aim was to examine the humoral immune response against Streptococcus gallolyticus subspecies gallolyticus antigens in individuals subjected to a routine colonoscopy in which colon adenomatous polyps were present or not. Serum samples from 133 individuals with adenomatous polyps and serum samples from 53 individuals with a normal colonoscopy were included. Western blot was performed in all subjects using a whole cell antigen from S. gallolyticus ATCC 9809, and rabbit antisera against the whole cell bacteria was prepared as a control. By analyzing the immune profile of the rabbit-immunized sera by Western-blot, at least 22 proteins were identified as immunogenic in S. gallolyticus. When we evaluated sera from human subjects, two proteins of approximately 30 and 22 kDa were most prominent. Based on this 2-protein band pattern, Western-blot profiles from human subjects were compared. The detection of a protein band of 22 kDa was associated with the presence of adenomatous polyps in colon [odds ratios (OR) 7.98, 95% confidence intervals (CI): 3.54-17.93], p < 0.001. When the presence of the 30 kDa protein alone or both the 22 and 30 kDa proteins were analyzed, the OR increased to 22.37 (95% CI: 3.77-131.64), p < 0.001. The specificity was 84.9 for the presence of the 22 kDa protein, and 98.1 for the presence of the 30 kDa protein alone or both 22 and 30 kDa bands. Serum from individuals with adenomatous polyps recognized two proteins from S. gallolyticus. This result confirmed the possible association of S. gallolyticus with adenomatous polyps in the colon.

Purpose: Bariatric surgery can achieve sustained weight loss compared to medical management. Among bariatric surgeries, laparoscopic adjustable gastric banding (LAGB) is less-invasive and potentially reversible. LAGB may decrease BMI through restriction of food intake, behavior changes, satiety and digestive hormone levels. The dramatic reduction of appetite observed with LAGB can be ameliorated if the band is underfilled. This effect has not been well evaluated in terms of patient behavior and hormonal changes. Our aim was to assess outcomes related to eating behavior, insulinotropic hormones, and weight change before and after temporary gastric band release. Methods: Adults >= 18 yeaars of age who previously underwent LAGB and achieved successful weight loss were enrolled. All patients underwent standardized evaluation including anthropometric measurements and completion of the Three-Factor Eating Questionnaire (TEFQ-R18) before and after a period of 14 days during which the band was completely loosened. At baseline and follow-up, blood was collected after an overnight fast and 1h after a standard high protein meal, and levels of insulinotropic hormones determined. Results: The mean age of the study cohort (9 women and 6 men) was 42 +/-14 years with mean pre-band adjustment BMI of 32.9 +/- 5.6 and mean waist circumference of 40 +/- 7 inches. All patients had >30% percent reduction in weight within 12-months of the LAGB and demonstrated a lower degree but continued weight loss in the 6-months before study enrollment. Compared to baseline values for the TEFQ-R18, within 2-weeks of loosening the band, cognitive restraint was reduced (11.2 +/- 3 vs. 10.4 +/- 4), while there was a significant increase in both disinhibition (6.4 +/- 3 vs. 9.4 +/- 3, p=0.004) and hunger scores (4.1 +/- 3 vs. 8.0 +/- 3, p=0.004). Compared to baseline, at follow-up insulin output in response to a meal showed a downward trend [Median (IQR) 1,110 (728-1,332) vs. 621 (375-1,325) pg/ml; p=0.21] while leptin was significantly elevated [10,400 (6,030-11,350) vs. 13,700 (10,500-43,900) pg/ml; p=0.001]. Consistent with these findings BMI significantly increased (32.9 +/- 5.6 vs. 34.5 +/- 5.6, p=0.001) along with waist size (40 +/- 7 vs 42 +/- 6, p=0.003). The amount of weight regained within two weeks, returned the cohort to the weight loss level noted at the 12-month post LAGB time point. Conclusion: LAGB adjustment continues to impact eating behavior, satiety hormones, and body weight beyond the initial 12-months following placement. Complete loosening of the LAGB can result in rapid changes in eating behavior, insulinotropic hormones, and significant changes in BMI. Careful adjustment of the band is necessary for continued maintenance of weight loss

ABSTRACT: BACKGROUND: Helicobacter pylori are successful colonizers of the human gastric mucosa. Colonization increases the risk of peptic ulcer disease and adenocarcinoma. However, potential benefits of H. pylori colonization include protection against early-onset asthma and against gastrointestinal infections. Campylobacter jejuni are a leading cause of bacterial diarrhea and complications include Guillain-Barre syndrome. Here, we describe the development of reliable serological assays to detect antibodies against those two bacteria in Rhesus macaques and investigated their distribution within a social group of monkeys. METHODS: Two cohorts of monkeys were analyzed. The first cohort consisted of 30 monkeys and was used to establish an enzyme-linked immunosorbent assay (ELISA) for H. pylori antibodies detection. To evaluate colonization of those macaques, stomach biopsies were collected and analyzed for the presence of H. pylori by histology and culture. C. jejuni ELISAs were established using human serum with known C. jejuni antibody status. Next, plasma samples of the 89 macaques (cohort 2) were assayed for antibodies and then statistically analyzed. RESULTS: An H. pylori IgG ELISA, which was 100% specific and 93% sensitive, was established. In contrast, the IgA ELISA was only 82% specific and 61% sensitive. The CagA IgG assay was 100% sensitive and 61% of the macaques were positive. In cohort 2, 62% macaques were H. pylori sero-positive and 52% were CagA positive. The prevalence of H. pylori IgG and CagA IgG increased with monkey age as described for humans. Of the 89 macaques 52% showed IgG against C. jejuni but in contrast to H. pylori, the sero-prevalence was not associated with increasing age. However, there was a drop in the IgG (but not in IgA) mean values between infant and juvenile macaques, similar to trends described in humans. CONCLUSIONS: Rhesus macaques have widespread exposure to H. pylori and C. jejuni, reflecting their social conditions and implying that Rhesus macaques might provide a model to study effects of these two important human mucosal bacteria on a population.

Background: The prevalence of Helicobacter pylori in Western populations has steadily decreased. This has been suggested as one of the factors involved in the recent increase of asthma and allergy. Some studies have reported a negative association between H. pylori and asthma and allergy, but data are inconsistent and there are a few studies in children. Aim: We investigated whether the prevalence of H. pylori was associated with asthma symptoms, allergic rhinitis, and atopic dermatitis in childhood. Methods: We determined IgG anti-H. pylori and CagA antibodies in serum of Dutch children, who took part in the PIAMA birth cohort study. Serum was collected from 545 children, aged 7-9 years (Dutch ethnicity 91.5%). Symptoms of asthma and atopy were assessed by yearly questionnaires. Chi-square tests and logistic regression were used. Results: We found 9%H. pylori and 0.9% CagA seropositivity. Twelve (5.9%) children with reported wheezing ever were H. pylori positive, compared to 37 (10.9%) of the non-wheezers (p = .05). No significant differences in H. pylori prevalence were found between children with or without allergic rhinitis (8.5% vs 9.5%), atopic dermatitis (8.7% vs 9.2%), and physician-diagnosed asthma (7.1% vs 9.4%). Multivariate analysis showed no significant associations between H. pylori seropositivity and wheezing (OR 0.52; 95% CI 0.25-1.06), allergic rhinitis (OR 0.96; 95% CI 0.51-1.81), atopic dermatitis (OR 1.05; 95% CI 0.56-1.98) or physician-diagnosed asthma (OR 0.87; 95% CI 0.37-2.08). Conclusion: We found a borderline significantly lower H. pylori seropositivity in children with wheezing compared to non-wheezers, but no association between H. pylori serum-antibody status and allergic rhinitis, atopic dermatitis, or asthma.