Genomic Determination of Relative Risks for Clostridioides difficile Infection From Asymptomatic Carriage in Intensive Care Unit Patients.

BACKGROUND: Clostridioides difficile infections (CDIs) are among the most prevalent hospital-associated infections (HAIs), particularly for intensive care unit (ICU) patients. The risks for developing active CDI from asymptomatic carriage of C. difficile are not well understood. METHODS: We identified asymptomatic C. difficile carriage among 1897 ICU patients using rectal swabs from an existing ICU vancomycin-resistant enterococci (VRE) surveillance program. C. difficile isolates from VRE swabs, and from C. difficile-positive stool samples, were genome sequenced. Spatial-temporal data from hospital records assessed genomically identified clusters for potential transmission events. RESULTS: Genomic analyses identified a diverse set of strains in infected patients and asymptomatic carriers. A total of 7.4% of ICU patients asymptomatically carried C. difficile; 69% of isolates carried an intact toxin locus. In contrast, 96% of C. difficile stool isolates were toxin encoding. CDI rates in asymptomatic carriers of toxin-encoding strains were 5.3% versus 0.57% in noncarriers. The relative risk for CDI with asymptomatic carriage of a toxin-encoding strain was 9.32 (95% confidence interval, 3.25-26.7). Genomic identification of clonal clusters supported analyses for asymptomatic transmission events, with spatial-temporal overlaps identified in 13 of 28 cases. CONCLUSIONS: Our studies provide the first genomically confirmed assessments of CDI relative risk from asymptomatic carriage of toxin-encoding strains and highlight the complex dynamics of asymptomatic transmission in ICUs. Asymptomatic carriers are an active reservoir of C. difficile in the nosocomial environment. C. difficile screening can be implemented within existing HAI surveillance programs and has the potential to support infection-control efforts against this pathogen.

Interval analysis of blood filtration efficacy in novel device for autotransfusion in postpartum hemorrhage.

Postpartum haemorrhage (PPH), the leading cause of maternal mortality, is particularly problematic in low resource settings where access to safe blood supplies and definitive medical treatment is limited. We describe the continued development of an autotransfusion device designed to treat PPH by collection, filtration and infusion of maternal blood. Previous study has demonstrated that the device effectively moves blood through a filtration apparatus and removes up to 97% of aerobic bacteria but had poor anaerobic bacteria reduction. In this study, we investigate the filtration efficacy of the device using configurations comprised of three different leukocyte depletion filter designs: the Pall Leukoguard RS leukocyte reduction filter (PLRF), the Haemonetics BPF4™ (BPF4) leukocyte reduction filter, and the Haemonetics SCRC Leukotrap® (SCRC) filter. All configurations performed well with reductions ranging from 49 to 98%. Configurations containing 2 Haemonetics SCRC Leukotrap®filters (configuration 5 and 6) consistently reduced anaerobic bacteria by at least 73%. These results indicate that utilising a combination of SCRC and PLRF filters confers a high level of microbial filtration with improved removal of anaerobic organisms.

Silver-Coated Endotracheal Tubes Cleaned With a Mechanism for Secretion Removal.

BACKGROUND: Biofilm on the surface of endotracheal tubes (ETTs) is associated with ventilator-associated pneumonia. The use of silver-coated ETTs has been suggested to reduce the occurrence of ventilator-associated pneumonia by preventing biofilm formation. However, mucus accumulation can reduce the antibacterial activity of silver-coated ETTs by isolating bacterial colonies from the silver surface. We hypothesized that, in mechanically ventilated subjects, periodic removal of secretions through the use of a cleaning device would enhance the antimicrobial properties of silver-coated ETTs and thus reduce bacterial colonization. METHODS: Subjects were randomized to either standard suctioning (blind tracheal suctioning, control group) or blind tracheal suctioning plus cleaning maneuver every 8 h (treatment group). Tracheal aspirates were collected immediately before extubation for microbiological culture. After extubation, ETTs were collected for both cultural and non-cultural microbiological analysis and biofilm isolation. RESULTS: 39 subjects expected to be ventilated for > 48 h were enrolled; 36 ETTs (18 control, 18 treatment) and 29 tracheal samples (15 control, 14 treatment) were collected. Among the ETTs positive for bacterial colonization (15 vs 9, P = .18), cleaning maneuvers did not reduce microbial load, shown as the decimal logarithm of colony-forming units (CFU) per mL (1.6 ± 1.2 vs 0.9 ± 1.2 logCFU/mL, P = .15). There was a trend toward decreased biofilm deposition (439.5 ± 29.0 vs 288.9 ± 157.7 mg, P = .09) in the treated ETTs. No significant differences were observed in the number of positive tracheal aspirates (13 vs 10, P = .39) or in the microbial load (4.8 ± 4.0 vs 4.2 ± 3.8 logCFU/mL, P = .70) of tracheal secretions. Finally, no differences in the microbial load of Gram-positive organisms, Gram-negative organisms, or yeasts were found between the ETTs and tracheal aspirates of the 2 groups. CONCLUSIONS: In 39 critically-ill subjects intubated with silver-coated ETTs, periodic cleaning maneuvers did not decrease bacterial colonization of the ETTs and did not lower respiratory tract colonization compared to the standard suctioning. (Clinicaltrials.gov registration NCT02120001.).

Genomically Informed Surveillance for Carbapenem-Resistant Enterobacteriaceae in a Health Care System.

UNLABELLED: Carbapenem-resistant Enterobacteriaceae (CRE) are an urgent public health concern. Rapid identification of the resistance genes, their mobilization capacity, and strains carrying them is essential to direct hospital resources to prevent spread and improve patient outcomes. Whole-genome sequencing allows refined tracking of both chromosomal traits and associated mobile genetic elements that harbor resistance genes. To enhance surveillance of CREs, clinical isolates with phenotypic resistance to carbapenem antibiotics underwent whole-genome sequencing. Analysis of 41 isolates of Klebsiella pneumoniae and Enterobacter cloacae, collected over a 3-year period, identified K. pneumoniae carbapenemase (KPC) genes encoding KPC-2, -3, and -4 and OXA-48 carbapenemases. All occurred within transposons, including multiple Tn4401 transposon isoforms, embedded within more than 10 distinct plasmids representing incompatibility (Inc) groups IncR, -N, -A/C, -H, and -X. Using short-read sequencing, draft maps were generated of new KPC-carrying vectors, several of which were derivatives of the IncN plasmid pBK31551. Two strains also had Tn4401 chromosomal insertions. Integrated analyses of plasmid profiles and chromosomal single-nucleotide polymorphism (SNP) profiles refined the strain patterns and provided a baseline hospital mobilome to facilitate analysis of new isolates. When incorporated with patient epidemiological data, the findings identified limited outbreaks against a broader 3-year period of sporadic external entry of many different strains and resistance vectors into the hospital. These findings highlight the utility of genomic analyses in internal and external surveillance efforts to stem the transmission of drug-resistant strains within and across health care institutions. IMPORTANCE: We demonstrate how detection of resistance genes within mobile elements and resistance-carrying strains furthers active surveillance efforts for drug resistance. Whole-genome sequencing is increasingly available in hospital laboratories and provides a powerful and nuanced means to define the local landscape of drug resistance. In this study, isolates of Klebsiella pneumoniae and Enterobacter cloacae with resistance to carbapenem antibiotics were sequenced. Multiple carbapenemase genes were identified that resided in distinct transposons and plasmids. This mobilome, or population of mobile elements capable of mobilizing drug resistance, further highlighted the degree of strain heterogeneity while providing a detailed timeline of carbapenemase entry into the hospital over a 3-year period. These surveillance efforts support effective targeting of infection control resources and the development of institution-specific repositories of resistance genes and the mobile elements that carry them.

Dynamics of the microbiota in response to host infection.

Longitudinal studies of the microbiota are important for discovering changes in microbial communities that affect the host. The complexity of these ecosystems requires rigorous integrated experimental and computational methods to identify temporal signatures that promote physiologic or pathophysiologic responses in vivo. Employing a murine model of infectious colitis with the pathogen Citrobacter rodentium, we generated a 2-month time-series of 16S rDNA gene profiles, and quantitatively cultured commensals, from multiple intestinal sites in infected and uninfected mice. We developed a computational framework to discover time-varying signatures for individual taxa, and to automatically group signatures to identify microbial sub-communities within the larger gut ecosystem that demonstrate common behaviors. Application of this model to the 16S rDNA dataset revealed dynamic alterations in the microbiota at multiple levels of resolution, from effects on systems-level metrics to changes across anatomic sites for individual taxa and species. These analyses revealed unique, time-dependent microbial signatures associated with host responses at different stages of colitis. Signatures included a Mucispirillum OTU associated with early disruption of the colonic surface mucus layer, prior to the onset of symptomatic colitis, and members of the Clostridiales and Lactobacillales that increased with successful resolution of inflammation, after clearance of the pathogen. Quantitative culture data validated findings for predominant species, further refining and strengthening model predictions. These findings provide new insights into the complex behaviors found within host ecosystems, and define several time-dependent microbial signatures that may be leveraged in studies of other infectious or inflammatory conditions.

Staphylococcal enterotoxin P predicts bacteremia in hospitalized patients colonized with methicillin-resistant Staphylococcus aureus.

BACKGROUND: Methicillin-resistant Staphylococcus aureus (MRSA) colonization predicts later infection, with both host and pathogen determinants of invasive disease. METHODS: This nested case-control study evaluates predictors of MRSA bacteremia in an 8-intensive care unit (ICU) prospective adult cohort from 1 September 2003 through 30 April 2005 with active MRSA surveillance and collection of ICU, post-ICU, and readmission MRSA isolates. We selected MRSA carriers who did (cases) and those who did not (controls) develop MRSA bacteremia. Generating assembled genome sequences, we evaluated 30 MRSA genes potentially associated with virulence and invasion. Using multivariable Cox proportional hazards regression, we assessed the association of these genes with MRSA bacteremia, controlling for host risk factors. RESULTS: We collected 1578 MRSA isolates from 520 patients. We analyzed host and pathogen factors for 33 cases and 121 controls. Predictors of MRSA bacteremia included a diagnosis of cancer, presence of a central venous catheter, hyperglycemia (glucose level, >200 mg/dL), and infection with a MRSA strain carrying the gene for staphylococcal enterotoxin P (sep). Receipt of an anti-MRSA medication had a significant protective effect. CONCLUSIONS: In an analysis controlling for host factors, colonization with MRSA carrying sep increased the risk of MRSA bacteremia. Identification of risk-adjusted genetic determinants of virulence may help to improve prediction of invasive disease and suggest new targets for therapeutic intervention.

Maternal microbe-specific modulation of inflammatory response in extremely low-gestational-age newborns.

The fetal response to intrauterine inflammatory stimuli appears to contribute to the onset of preterm labor as well as fetal injury, especially affecting newborns of extremely low gestational age. To investigate the role of placental colonization by specific groups of microorganisms in the development of inflammatory responses present at birth, we analyzed 25 protein biomarkers in dry blood spots obtained from 527 newborns delivered by Caesarean section in the 23rd to 27th gestation weeks. Bacteria were detected in placentas and characterized by culture techniques. Odds ratios for having protein concentrations in the top quartile for gestation age for individual and groups of microorganisms were calculated. Mixed bacterial vaginosis (BV) organisms were associated with a proinflammatory pattern similar to those of infectious facultative anaerobes. Prevotella and Gardnerella species, anaerobic streptococci, peptostreptococci, and genital mycoplasmas each appeared to be associated with a different pattern of elevated blood levels of inflammation-related proteins. Lactobacillus was associated with low odds of an inflammatory response. This study provides evidence that microorganisms colonizing the placenta provoke distinctive newborn inflammatory responses and that Lactobacillus may suppress these responses.

Bifidobacterium animalis subsp. lactis fermented milk product reduces inflammation by altering a niche for colitogenic microbes.

Intestinal health requires the coexistence of eukaryotic self with the gut microbiota and dysregulated host-microbial interactions can result in intestinal inflammation. Here, we show that colitis improved in T-bet(-/-)Rag2(-/-) mice that consumed a fermented milk product containing Bifidobacterium animalis subsp. lactis DN-173 010 strain. A decrease in cecal pH and alterations in short chain fatty acid profiles occurred with consumption, and there were concomitant increases in the abundance of select lactate-consuming and butyrate-producing bacteria. These metabolic shifts created a nonpermissive environment for the Enterobacteriaceae recently identified as colitogenic in a T-bet(-/-)Rag2(-/-) ulcerative colitis mouse model. In addition, 16S rRNA-based analysis of the T-bet(-/-)Rag2(-/-) fecal microbiota suggest that the structure of the endogenous gut microbiota played a key role in shaping the host response to the bacterial strains studied herein. We have identified features of the gut microbiota, at the membership and functional level, associated with response to this B. lactis-containing fermented milk product, and therefore this model provides a framework for evaluating and optimizing probiotic-based functional foods.

Enterobacteriaceae act in concert with the gut microbiota to induce spontaneous and maternally transmitted colitis.

Disruption of homeostasis between the host immune system and the intestinal microbiota leads to inflammatory bowel disease (IBD). Whether IBD is instigated by individual species or disruptions of entire microbial communities remains controversial. We characterized the fecal microbial communities in the recently described T-bet(-/-) ×Rag2(-/-) ulcerative colitis (TRUC) model driven by T-bet deficiency in the innate immune system. 16S rRNA-based analysis of TRUC and Rag2(-/-) mice revealed distinctive communities that correlate with host genotype. The presence of Klebsiella pneumoniae and Proteus mirabilis correlates with colitis in TRUC animals, and these TRUC-derived strains can elicit colitis in Rag2(-/-) and WT adults but require a maternally transmitted endogenous microbial community for maximal intestinal inflammation. Cross-fostering experiments indicated a role for these organisms in maternal transmission of disease. Our findings illustrate how gut microbial communities work in concert with specific culturable colitogenic agents to cause IBD.

Persistence survey of toxic shock syndrome toxin-1 producing Staphylococcus aureus and serum antibodies to this superantigen in five groups of menstruating women.

BACKGROUND: Menstrual Toxic Shock Syndrome (mTSS) is thought to be associated with the vaginal colonization with specific strains of Staphylococcus aureus TSST-1 in women who lack sufficient antibody titers to this toxin. There are no published studies that examine the seroconversion in women with various colonization patterns of this organism. Thus, the aim of this study was to evaluate the persistence of Staphylococcus aureus colonization at three body sites (vagina, nares, and anus) and serum antibody to toxic shock syndrome toxin-producing Staphylococcus aureus among a small group of healthy, menstruating women evaluated previously in a larger study. METHODS: One year after the completion of that study, 311 subjects were recalled into 5 groups. Four samples were obtained from each participant at several visits over an additional 6-11 month period: 1) an anterior nares swab; 2) an anal swab; 3) a vagina swab; and 4) a blood sample. Gram stain, a catalase test, and a rapid S. aureus-specific latex agglutination test were performed to phenotypically identify S. aureus from sample swabs. A competitive ELISA was used to quantify TSST-1 production. Human TSST-1 IgG antibodies were determined from the blood samples using a sandwich ELISA method. RESULTS: We found only 41% of toxigenic S. aureus and 35.5% of non-toxigenic nasal carriage could be classified as persistent. None of the toxigenic S. aureus vaginal or anal carriage could be classified as persistent. Despite the low persistence of S. aureus colonization, subjects colonized with a toxigenic strain were found to display distributions of antibody titers skewed toward higher titers than other subjects. Seven percent (5/75) of subjects became seropositive during recall, but none experienced toxic shock syndrome-like symptoms. CONCLUSIONS: Nasal carriage of S. aureus appears to be persistent and the best predicator of subsequent colonization, whereas vaginal and anal carriage appear to be more transient. From these findings, it appears that antibody titers in women found to be colonized with toxigenic S. aureus remained skewed toward higher titers whether or not the colonies were found to be persistent or transient in nature. This suggests that colonization at some point in time is sufficient to elevate antibody titer levels and those levels appear to be persistent. Results also indicate that women can become seropositive without experiencing signs or symptoms of toxic shock syndrome.

Active Video/Arcade Games (Exergaming) and Energy Expenditure in College Students.

Video games have become increasingly popular among young adults. The purpose of this pilot study was to determine if interactive video/arcade games, requiring physical activity to play, increase the energy expenditure (EE) and heart rate (HR) of young adults enough to elicit a training response. Thirteen male and female participants 26.6 ± 5.7 years of age were in the study. Participants were familiarized with equipment and allowed to practice with three games: (1) moving and striking lighted pads, (2) riding a bike to increase the pace of a race car, and (3) boxing against a video simulated opponent. A portable metabolic cart and HR monitor were attached to participants to measure baseline and exercise values. Participants could play any of the three games for 30 minutes while metabolic and HR data were collected. Exercise data were compared to baseline measures, and the 3 games were compared for EE. Paired sample t-tests showed baseline and exercise values differed for HR (t(12) = -18.91, p < 0.01), and EE (t(12) = -15.62, p < 0.01). The boxing game provided the highest VO2 (17.47 ± 4.79 ml·kg(·-1)min(-1)). Participants achieved 60% or better of their HR reserve (162.82 ± 10.78 beats·min(-1),) well within the ACSM guidelines for a training HR. Caloric expenditure during the 30-minute exercise session (226. 07 ± 48.68) is also within the ACSM recommendations for daily physical activity. Thus, interactive video/arcade games that require physical activity to play can be utilized as part of an overall aerobic exercise program.

Diversity of the gut microbiota and eczema in early life.

BACKGROUND: A modest number of prospective studies of the composition of the intestinal microbiota and eczema in early life have yielded conflicting results. OBJECTIVE: To examine the relationship between the bacterial diversity of the gut and the development of eczema in early life by methods other than stool culture. METHODS: Fecal samples were collected from 21 infants at 1 and 4 months of life. Nine infants were diagnosed with eczema by the age of 6 months (cases) and 12 infants were not (controls). After conducting denaturating gradient gel electrophoresis (DGGE) of stool samples, we compared the microbial diversity of cases and controls using the number of electrophoretic bands and the Shannon index of diversity (H') as indicators. RESULTS: Control subjects had significantly greater fecal microbial diversity than children with eczema at ages 1 (mean H' for controls = 0.75 vs. 0.53 for cases, P = 0.01) and 4 months (mean H' for controls = 0.92 vs. 0.59 for cases, P = 0.02). The increase in diversity from 1 to 4 months of age was significant in controls (P = 0.04) but not in children who developed eczema by 6 months of age (P = 0.32). CONCLUSION: Our findings suggest that reduced microbial diversity is associated with the development of eczema in early life.

Detection of bacteria in placental tissues obtained from extremely low gestational age neonates.

OBJECTIVE: The objective of the study was to quantify and identify aerobic and anaerobic bacteria as well as Mycoplasma and Ureaplasma in the chorionic parenchyma. STUDY DESIGN: A sample of the chorionic parenchyma from neonates delivered between 23-27 completed weeks was cultured and tested by polymerase chain reaction (PCR) methods using universal bacterial primers for the presence of bacteria and mycoplasmas. RESULTS: The culture positive rate was higher for vaginal deliveries (333/489; 68%) than for cesarean sections (363/876; 41%). Thirty percent of all culture-positive samples had only aerobic bacteria, 21% of the samples had only anaerobic bacteria, and 9% of the samples had only Mycoplasma/Ureaplasma. The mean concentration of Mycoplasma/Ureaplasma (4.00 +/- 1.11 log10 CFU/g) was significantly higher (P < .001) than the total count of either aerobes (3.24 +/- 1.12 log10 CFU/g) or anaerobes (2.89 +/- 0.99 log10 CFU/g). Staphylococcus sp. and Corynebacterium sp. as well as organisms associated with bacterial vaginosis were the most frequently recovered. A PCR product was not detected from either randomly selected or known culture-positive samples. CONCLUSION: Approximately half of second-trimester placentas harbor organisms within the chorionic plate. The chorion parenchyma appears to harbor constituents that prevent the identification of bacterial deoxyribonucleic acid by PCR methods.

Mode of delivery and cord blood cytokines: a birth cohort study.

BACKGROUND: The mechanisms for the association between birth by cesarean section and atopy and asthma are largely unknown. OBJECTIVE: To examine whether cesarean section results in neonatal secretion of cytokines that are associated with increased risk of atopy and/or asthma in childhood. To examine whether the association between mode of delivery and neonatal immune responses is explained by exposure to the maternal gut flora (a marker of the vaginal flora). METHODS: CBMCs were isolated from 37 neonates at delivery, and secretion of IL-13, IFN-gamma, and IL-10 (at baseline and after stimulation with antigens [dust mite and cat dander allergens, phytohemagglutinin, and lipopolysaccharide]) was quantified by ELISA. Total and specific microbes were quantified in maternal stool. The relation between mode of delivery and cord blood cytokines was examined by linear regression. The relation between maternal stool microbes and cord blood cytokines was examined by Spearman's correlation coefficients. RESULTS: Cesarean section was associated with increased levels of IL-13 and IFN-gamma. In multivariate analyses, cesarean section was associated with an increment of 79.4 pg/ml in secretion of IL-13 by CBMCs after stimulation with dust mite allergen (P < 0.001). Among children born by vaginal delivery, gram-positive anaerobes and total anaerobes in maternal stool were positively correlated with levels of IL-10, and gram-negative aerobic bacteria in maternal stool were negatively correlated with levels of IL-13 and IFN-gamma. CONCLUSION: Cesarean section is associated with increased levels of IL-13 and IFN-gamma, perhaps because of lack of labor and/or reduced exposure to specific microbes (e.g., gram-positive anaerobes) at birth.

Prevalence of toxic shock syndrome toxin 1-producing Staphylococcus aureus and the presence of antibodies to this superantigen in menstruating women.

Menstrual toxic shock syndrome (mTSS) is thought to be associated with colonization with toxic shock syndrome toxin 1 (TSST-1)-producing Staphylococcus aureus in women with insufficient antibody titers. mTSS has been associated with menstruation and tampon use, and although it is rare, the effects can be life threatening. It remains of interest because of the widespread use of tampons, reported to be about 70% of women in the United States, Canada, and much of Western Europe. This comprehensive study was designed to determine S. aureus colonization and TSST-1 serum antibody titers in 3,012 menstruating women in North America between the ages of 13 and 40, particularly among age and racial groups that could not be assessed reliably in previous small studies. One out of every four subjects was found to be colonized with S. aureus in at least one of three body sites (nose, vagina, or anus), with approximately 9% colonized vaginally. Eighty-five percent of subjects had antibody titers (> or =1:32) to TSST-1, and the vast majority (81%) of teenaged subjects (13 to 18 years) had already developed antibody titers. Among carriers of toxigenic S. aureus, a significantly lower percentage of black women than of white or Hispanic women were found to have antibody titers (> or =1:32) to TSST-1 (89% versus 98% and 100%). These findings demonstrate that the majority of teenagers have antibody titers (> or =1:32) to TSST-1 and are presumed to be protected from mTSS. These findings also suggest that black women may be more susceptible to mTSS than previously thought.