Forecasting Staphylococcus aureus Infections Using Genome-Wide Association Studies, Machine Learning, and Transcriptomic Approaches.

Staphylococcus aureus is a major human and animal pathogen, colonizing diverse ecological niches within its hosts. Predicting whether an isolate will infect a specific host and its subsequent clinical fate remains unknown. In this study, we investigated the S. aureus pangenome using a curated set of 356 strains, spanning a wide range of hosts, origins, and clinical display and antibiotic resistance profiles. We used genome-wide association study (GWAS) and random forest (RF) algorithms to discriminate strains based on their origins and clinical sources. Here, we show that the presence of sak and scn can discriminate strains based on their host specificity, while other genes such as mecA are often associated with virulent outcomes. Both GWAS and RF indicated the importance of intergenic regions (IGRs) and coding DNA sequence (CDS) but not sRNAs in forecasting an outcome. Additional transcriptomic analyses performed on the most prevalent clonal complex 8 (CC8) clonal types, in media mimicking nasal colonization or bacteremia, indicated three RNAs as potential RNA markers to forecast infection, followed by 30 others that could serve as infection severity predictors. Our report shows that genetic association and transcriptomics are complementary approaches that will be combined in a single analytical framework to improve our understanding of bacterial pathogenesis and ultimately identify potential predictive molecular markers. IMPORTANCE Predicting the outcome of bacterial colonization and infections, based on extensive genomic and transcriptomic data from a given pathogen, would be of substantial help for clinicians in treating and curing patients. In this report, genome-wide association studies and random forest algorithms have defined gene combinations that differentiate human from animal strains, colonization from diseases, and nonsevere from severe diseases, while it revealed the importance of IGRs and CDS, but not small RNAs (sRNAs), in anticipating an outcome. In addition, transcriptomic analyses performed on the most prevalent clonal types, in media mimicking either nasal colonization or bacteremia, revealed significant differences and therefore potent RNA markers. Overall, the use of both genomic and transcriptomic data in a single analytical framework can enhance our understanding of bacterial pathogenesis.

Galleria mellonella Larvae as an Infection Model to Investigate sRNA-Mediated Pathogenesis in Staphylococcus aureus.

Small regulatory RNAs (sRNAs) are key players in bacterial regulatory networks. Monitoring their expression inside living colonized or infected organisms is essential for identifying sRNA functions, but few studies have looked at sRNA expression during host infection with bacterial pathogens. Insufficient in vivo studies monitoring sRNA expression attest to the difficulties in collecting such data, we therefore developed a non-mammalian infection model using larval Galleria mellonella to analyze the roles of Staphylococcus aureus sRNAs during larval infection and to quickly determine possible sRNA involvement in staphylococcal virulence before proceeding to more complicated animal testing. We began by using the model to test infected larvae for immunohistochemical evidence of infection as well as host inflammatory responses over time. To monitor sRNA expression during infection, total RNAs were extracted from the larvae and invading bacteria at different time points. The expression profiles of the tested sRNAs were distinct and they fluctuated over time, with expression of both sprD and sprC increased during infection and associated with mortality, while rnaIII expression remained barely detectable over time. A strong correlation was observed between sprD expression and the mortality. To confirm these results, we used sRNA-knockout mutants to investigate sRNA involvement in Staphylococcus aureus pathogenesis, finding that the decrease in death rates is delayed when either sprD or sprC was lacking. These results demonstrate the relevance of this G. mellonella model for investigating the role of sRNAs as transcriptional regulators involved in staphylococcal virulence. This insect model provides a fast and easy method for monitoring sRNA (and mRNA) participation in S. aureus pathogenesis, and can also be used for other human bacterial pathogens.

Galleria mellonella as a Suitable Model of Bacterial Infection: Past, Present and Future.

The increasing interest for Galleria mellonella larvae as an infection model is evidenced by the number of papers reporting its use, which increases exponentially since the early 2010s. This popularity was initially linked to limitation of conventional animal models due to financial, technical and ethical aspects. In comparison, alternative models (e.g. models using Caenorhabditis elegans, Drosophila melanogaster or G. mellonella) were cheap, simple to use and not limited by ethical regulation. Since then, similar results have been established with G. mellonella model comparatively to vertebrates, and it is more and more often used as a robust model per se, not only as an alternative to the murine model. This review attempts to summarize the current knowledge supporting the development of this model, both on immunological and microbiological aspects. For that, we focus on investigation of virulence and new therapies for the most important pathogenic bacteria. We also discuss points out directions for standardization, as well as recent advances and new perspectives for monitoring host-pathogen interactions.

Adhesion of Staphylococcus aureus to epithelial cells: an in vitro approach to study interactions within the nasal microbiota.

Introduction. Staphylococcus aureus is a skin and mucous commensal bacterium of warm-blooded animals. In humans, the nose is the main ecological niche of S. aureus, and nasal carriage is a risk factor for developing an endogenous infection. S. aureus nasal colonization is a multifactorial process, involving inter-species interactions among the nasal microbiota.Aims. The objectives of this study were to characterize the microbiota of carriers and non-carriers of S. aureus and to demonstrate the importance of inter-species relationships in the adhesion of S. aureus, a key step in nasal colonization.Methodology. First, we characterized the nasal microbiota from 30 S. aureus carriers and non-carriers by a culturomic approach. We then evaluated the adhesion of S. aureus, first alone and then along with other bacteria of the nasal microbiota. To do that, we used an in vitro model to measure the interactions among bacteria in the presence of epithelial cells.Results. Analysis of the nasal microbiota of the carriers and non-carriers of S. aureus made it possible to observe that each microbiota has specific features in terms of composition. However, this composition differs significantly between carriers and non-carriers mainly through two bacterial groups: coagulase-negative staphylococci and corynebacteria. In a second part, adhesion of S. aureus to epithelial cells showed competition between S. aureus and these bacteria, suggesting a limitation of nasal colonization by S. aureus.Conclusion. These findings demonstrate the existence of a negative correlation between S. aureus and other species which inhibits adhesion and could limit nasal colonization.

Simplification of first-line antibacterial regimen for complicated appendicitis in children is associated with better adherence to guidelines and reduced use of antibiotics.

Acute appendicitis in children requires early surgery and short-course antibiotics active against Enterobacteriaceae and anaerobes. Although an aminoglycoside-containing three-drug regimen has been used successfully for decades, simpler regimens with similar efficacy are increasingly used. This study evaluated the impact of a switch from the combination of cefotaxime, metronidazole and gentamicin (regimen 1) to piperacillin/tazobactam (regimen 2) as first-line regimen for complicated acute appendicitis in children. In total, 171 children were enrolled [median (IQR) age, 10 (6-13) years], treated with regimen 1 (n = 80) or regimen 2 (n = 91) following surgery for complicated acute appendicitis. The two groups were comparable except for surgical approach (through laparoscopy in 46% vs. 88% for regimens 1 and 2, respectively; P < 0.001). Post-operative complications and duration of hospital stay were similar. Deviations from antibacterial treatment protocol decreased from 36% (29/80) to 14% (13/91) (P < 0.001), with a dramatic reduction in antibacterial treatment duration from median (IQR) of 15 (12-16) days to 5 (5-8) days (P < 0.001). Post-operative intra-abdominal abscess developed in 32 children (18.7%). Female sex (OR = 2.76, 95% CI 1.18-6.48; P = 0.02) and sepsis/septic shock on admission (OR = 4.72, 95% CI 1.12-19.97; P = 0.035) were independently associated with post-operative intra-abdominal abscess, but not antibacterial regimen. This study shows that simplification of first-line antibacterial regimen for complicated appendicitis in children was associated with reduced protocol deviation, reduced duration of antibiotics, and similar outcomes (post-operative complications and duration of hospital stay).

Staphylococcus aureus Regulatory RNAs as Potential Biomarkers for Bloodstream Infections.

Staphylococcus aureus is a commensal bacterium and pathogen. Identifying biomarkers for the transition from colonization to disease caused by this organism would be useful. Several S. aureus small RNAs (sRNAs) regulate virulence. We investigated presence and expression of 8 sRNAs in 83 S. aureus strains from 42 patients with sepsis or septic shock and 41 asymptomatic colonized carriers. Small pathogenicity island sRNAs sprB and sprC were clade specific. Six sRNAs had variable expression not correlated with clinical status. Expression of RNAIII was lower in strains from septic shock patients than in strains from colonized patients. When RNAIII was associated with expression of sprD, colonizing strains could be discriminated from strains in patients with bloodstream infections, including patients with sepsis and septic shock. Isolates associated with colonization might have sRNAs with target expression different from those of disease isolates. Monitoring expression of RNAIII and sprD could help determine severity of bloodstream infections.

Short-term decline in all-cause acquired infections with the routine use of a decontamination regimen combining topical polymyxin, tobramycin, and amphotericin B with mupirocin and chlorhexidine in the ICU: a single-center experience.

OBJECTIVES: In a multicenter, placebo-controlled, randomized, double-blind trial, we showed that acquired infections in intubated patients were reduced by the combination of topical polymyxin plus tobramycin and nasal mupirocin plus chlorhexidine body wash. Because intubated patients are particularly at risk for acquired infections, we reassessed the impact of this protocol as a routine procedure to control acquired infections in the ICU. DESIGN: Nonrandomized study comparing acquired infections in ICU patients during two 1-year periods: the last year before (group A, n = 925) and the first year after the implementation of the protocol (group B, n = 1,022). Acquired infections were prospectively recorded. SETTING: Polyvalent medical ICU at a university-affiliated hospital. PATIENTS: All patients admitted to the ICU. INTERVENTIONS: Administration of polymyxin/tobramycin/amphotericin B in the oropharynx and the gastric tube plus a mupirocin/chlorhexidine regimen in intubated patients and standard care in the other patients. MEASUREMENTS AND MAIN RESULTS: The comparison of acquired infection rates between groups was adjusted for differences at baseline. Infection rates were lower in group B compared with group A (5.3% vs 11.0%; p < 0.001), as were the incidence rates of total acquired infections (9.4 vs 23.6 per 1,000 patient-days; p < 0.001), intubation-related pneumonia (5.1 vs 17.1 per 1,000 ventilator-days; p < 0.001), and catheter-related bloodstream infections (1.0 vs 3.5 per 1,000 catheter-days; p = 0.03). There were fewer acquired infections caused by ceftazidime-resistant Enterobacteriaceae (0.8‰ vs 3.6‰; p < 0.001), ciprofloxacin-resistant Enterobacteriaceae (0.8‰ vs 2.5‰; p = 0.02), ciprofloxacin-resistant Pseudomonas aeruginosa (0.5‰ vs 1.6‰; p = 0.05), and colistin-resistant Gram-negative bacilli (0.7‰ vs 1.9‰; p = 0.04). Fewer patients got acquired infections due to multidrug-resistant aerobic Gram-negative bacilli (p = 0.008). CONCLUSIONS: In intubated patients, the use of topical polymyxin/tobramycin/amphotericin B plus mupirocin/chlorhexidine was associated with the reduction of all-cause ICU-acquired infections. Long-term emergence of multidrug-resistant organisms deserves further investigation.

Effect of oropharyngeal povidone-iodine preventive oral care on ventilator-associated pneumonia in severely brain-injured or cerebral hemorrhage patients: a multicenter, randomized controlled trial.

OBJECTIVE: To evaluate the efficacy and safety of oral care with povidone-iodine on the occurrence of ventilator-associated pneumonia in a high-risk population. DESIGN: A multicenter, placebo-controlled, randomized, double-blind, two-parallel-group trial performed between May 2008 and May 2011. SETTING: Six ICUs in France. PATIENTS: One hundred seventy-nine severely brain-injured patients (Glasgow Coma Scale ≤ 8) or cerebral hemorrhage expected to be mechanically ventilated for more than 24 hours. INTERVENTIONS: Participants were randomly assigned to receive oropharyngeal care with povidone-iodine (n = 91) or placebo (n = 88) six times daily until mechanical ventilation withdrawal. MEASUREMENTS AND MAIN RESULTS: Primary endpoint was the rate of ventilator-associated pneumonia. Secondary endpoint included the rates of ventilator-associated tracheobronchitis and acute respiratory distress syndrome and patient's outcome. The number of patients evaluable for the primary endpoint (preplanned modified intention-to-treat population) was 150 (78 in the povidone-iodine group, 72 in the placebo group). Ventilator-associated pneumonia occurred in 24 patients (31%) in the povidone-iodine group and 20 (28%) in the placebo group (relative risk, 1.11 [95% CI, 0.67-1.82]; p = 0.69). There was no significant difference between the two groups for ventilator-associated tracheobronchitis: eight patients (10%) in the povidone-iodine group and five patients (7%) in the placebo group (relative risk, 1.48 [95% CI, 0.51-4.31]; p = 0.47). Acute respiratory distress syndrome occurred in five patients in the povidone-iodine group but not in the placebo group (p = 0.06). There was no difference between groups for ICU and hospital lengths of stay, as well as ICU and 90-day mortality. CONCLUSIONS: There is no evidence to recommend oral care with povidone-iodine to prevent ventilator-associated pneumonia in high-risk patients. Furthermore, this strategy seems to increase the rate of acute respiratory distress syndrome.

Whole-Genome Sequences of Staphylococcus aureus ST398 Strains of Animal Origin.

Staphylococcus aureus sequence type 398 (ST398) was originally associated with animal infections. We announce the complete genome sequences of two ST398 methicillin-susceptible S. aureus strains from the livestock environment. These genome sequences assist in the characterization of interesting ST398 features relying on host tropism and epidemiological settings.

Analysis of prophages harbored by the human-adapted subpopulation of Staphylococcus aureus CC398.

Staphylococcus aureus clonal complex 398 is a livestock-associated pathogen that poses a worldwide threat because of its ability to colonize and infect both humans and animals. We used high-resolution whole-genome microarrays, prophage profiling, immune evasion cluster characterization and whole-genome sequencing to investigate the roles of prophages in the emerging human-adapted subpopulation of CC398 that has been associated with invasive infections in humans living in animal-free environments. We characterized one phage and two prophages specifically harbored by CC398 isolates belonging to the emerging subpopulation. We introduced the phage into permissive prophage-free isolates. We investigated the effects of lysogeny on the host ability to resist further phage infection and transformation, to acquire the capacity to invade human cells, and to express virulence factors encoded by prophages. We report evidence of a defective ϕMR11-like helper prophage, named StauST398-5pro, specifically associated with the emerging non-LA CC398 subpopulation. StauST398-5pro confers substantial protection against horizontal genetic transfer to its host. It interacts with a human-associated ß-converting prophage encoding immune-modulating proteins such that virulence genes are expressed during stress situations. Our findings provide insight into the role of phages in the expression of virulence and in the spread of genetic information among new host-adapted S. aureus isolates. We demonstrate that functional prophage elements can condition host specificity and confer new virulence traits on emerging intra-species clones of bacteria.

Clostridium difficile infections: do we know the real dimensions of the problem?

Clostridium difficile infection (CDI) is the primary cause of nosocomial diarrhoea in industrialised countries, usually occurring as a complication of antibiotic therapy in elderly patients. Landmark events contributed to boosting interest in CDI over the last 10 years, including the emergence of unusually severe and recurrent CDI due to the NAP1/BI/027 strain, as well as reports suggesting that CDI is also significantly encountered in patients previously considered at no risk, such as community-acquired CDI in patients with no recent antibiotic use, or CDI during pregnancy. Despite this growing interest from the medical community, we do not know the real dimensions of the disease for the following reasons: (i) despite comprehensive guidelines published in Europe and in the USA, most laboratories still use diagnostic tests with suboptimal sensitivity as a 'rule-out' test, hence a significant proportion of CDIs remain undiagnosed; (ii) use of PCR as a stand-alone test by others will probably overestimate the real incidence of CDI and jeopardise any comparison between institutions with different diagnostic procedures; and (iii) transversal studies, with optimum design and diagnostic tests, are rapidly outdated due to the dramatic changes in CDI epidemiology that may occur from one year to another. To get an accurate picture of the real dimensions of the CDI issue, we need more systematic use of an adequate and homogeneous diagnostic strategy in the field as well as the implementation of continuous monitoring of CDI incidence through surveillance programmes.

CC9 livestock-associated Staphylococcus aureus emerges in bloodstream infections in French patients unconnected with animal farming.

We report 4 bloodstream infections associated with CC9 agr type II Staphylococcus aureus in individuals without animal exposure. We demonstrate, by microarray analysis, the presence of egc cluster, fnbA, cap operon, lukS, set2, set12, splE, splD, sak, epiD, and can, genomic features associated with a high virulence potential in humans.

Methicillin-susceptible ST398 Staphylococcus aureus responsible for bloodstream infections: an emerging human-adapted subclone?

In the course of an annual 3-month bloodstream infections (BSI) survey conducted during a four-year period in 31 healthcare institutions located in three noncontiguous French regions, we report 18 ST398 Staphylococcus aureus BSI. ST398 BSI incidence showed a seven-fold increase during the study period (0.002 per 1,000 patient days in 2007 vs. 0.014 in 2010). ST398 BSI isolates differed from the pig-borne multiresistant clone: 17/18 BSI isolates were methicillin susceptible and none was of t011, t034 or t108 pig-borne spa-types. ST398 BSI isolates had homogenous resistance patterns (15/18 with only Ery(r)) and prophagic content (all harboured the hlb-converting Sau3int phage). The clustering of BSI and pig-borne isolates by spa-typing and MLVA, the occurrence of Sau3int phage in BSI isolates and the lack of this phage in pig-borne isolates suggest that the emergence of BSI isolates could have arisen from horizontal transfer, at least of the Sau3int phage, in genetically diverse MSSA ST398 isolates. The acquisition of the phage likely plays a role in the increasing ability of the lysogenic ST398 isolates to colonize human. The mode of acquisition of the non pig-borne ST398 isolates by our 18 patients remains unclear. ST398 BSI were diagnosed in patients lacking livestock exposure and were significantly associated with digestive portals of entry (3/18 [16.7%] for ST398 vs. 19/767 [2.5%] for non ST398 BSI; p = .012). This raises the question of possible foodborne human infections. We suggest the need for active surveillance to study and control the spread of this human-adapted subclone increasingly isolated in the hospital setting.

Randomized comparison of 2 protocols to prevent acquisition of methicillin-resistant Staphylococcus aureus: results of a 2-center study involving 500 patients.

OBJECTIVE: To compare an interventional protocol with a standard protocol for preventing the acquisition of methicillin-resistant Staphylococcus aureus (MRSA) in the intensive care unit (ICU). DESIGN: Prospective, randomized, controlled, parallel-group, nonblinded clinical trial. SETTING: Medical ICUs of 2 French university hospitals. PARTICIPANTS: Five hundred adults with an expected length of stay in the ICU greater than 48 hours. INTERVENTIONS: For the intervention group, the protocol required repeated MRSA screening, contact and droplet isolation precautions for patients at risk for MRSA at ICU admission and for MRSA-positive patients, and decontamination with nasal mupirocin and chlorhexidine body wash for MRSA-positive patients. For the standard group, the standard precautions protocol was used, and the results of repeated MRSA screening in the standard group were not communicated to investigators. MAIN OUTCOME MEASURE: MRSA acquisition rate in the ICU. An audit was conducted to assess compliance with hygiene and isolation precautions. RESULTS: In the intent-to-treat analysis ([Formula: see text]), the MRSA acquisition rate in the ICU was similar in the standard (13 [5.3%] of 243) and intervention (16 [6.5%] of 245) groups ([Formula: see text]). The audit showed that the overall compliance rate was 85.5% in the standard group and 84.1% in the intervention group ([Formula: see text]), although compliance was higher when isolation precautions were absent than when they were in place (88.2% vs 79.1%; [Formula: see text]). MRSA incidence rates were higher without isolation precautions (7.57‰) than with isolation precautions (2.36‰; [Formula: see text]). CONCLUSIONS: Individual allocation to MRSA screening, isolation precautions, and decontamination do not provide individual benefit in reducing MRSA acquisition, compared with standard precautions, although the collective risk was lower during the periods of isolation. TRIAL REGISTRATION: Clinicaltrials.gov identifier: NCT00151606.

Epidemiology and antimicrobial resistance of Streptococcus pneumoniae in France in 2007: data from the pneumococcus surveillance network.

Antimicrobial resistance of Streptococcus pneumoniae in France is closely monitored by the pneumococcus surveillance network, founded in 1995, which collects data from regional observatories (Observatoire Régionaux du Pneumocoque [ORP]). In 2007, 23 ORPs analyzed the antibiotic susceptibility of 5,302 isolates of S. pneumoniae recovered in France from cerebrospinal fluid, blood, middle ear fluid, and pleural fluid, as well as from adult respiratory samples. The study showed that 38.2% of the strains were nonsusceptible to penicillin, 19.3% nonsusceptible to amoxicillin, and 10.5% nonsusceptible to cefotaxime. The percentage of pneumococcus nonsusceptible to penicillin varied according to both the sample and the age of the patient (child/adult): blood (27.8%/32.5%), cerebrospinal fluid (33.7%/34.6%), middle ear fluid (60.2%/27.5%), and pleural fluid (50.0%/31.0%). Between 2003 and 2007, the frequency of penicillin resistance in invasive pneumococcal disease gradually decreased from 46.4% to 29.0% in children and from 43.8% to 32.7% in adults. This decrease coincided with the introduction of a seven-valent pneumococcal conjugate vaccine into immunization programs and with a general reduction in levels of antibiotic consumption in France.

Should we systematically perform central nervous system imaging in patients with Whipple's endocarditis?

BACKGROUND: Whipple's endocarditis is an uncommon disease, with approximately 100 cases reported to date. Case series suggest that Whipple's endocarditis usually presents without extracardiac manifestations of Whipple's disease. METHODS: We report 4 consecutive cases of Whipple's endocarditis associated with brain lesions. All patients fulfilled Duke Criteria for definite endocarditis. Whipple's disease was diagnosed through 16S rRNA polymerase chain reaction assays on valves excised from patients with culture-negative endocarditis (n=3) or through polymerase chain reaction and periodic acid staining-positive foamy macrophages on duodenal biopsy (n=1). RESULTS: All patients were male, aged 56 to 72 years. They presented with mitral (n=1), aortic (n=1), mitral and aortic (n=1), and tricuspid (n=1) endocarditis. Brain magnetic resonance imaging was performed because of mild-to-moderate cognitive disorders (n=3) or ataxia (n=1) and revealed multiple (n=3) or solitary (n=1) contrast-enhancing lesions. Cerebrospinal fluid studies revealed meningitis in 1 case. Polymerase chain reaction assays on cerebrospinal fluid were negative for all patients. All patients received intravenous ceftriaxone (2-4 weeks) associated with gentamicin (2 weeks), followed by 1 year of oral trimethoprim-sulfamethoxazole, with favorable outcomes. CONCLUSION: Whipple's associated central nervous system disease may be common but frequently undiagnosed, in patients with Whipple's endocarditis. Because treatment is different when neurologic disease is present (ie, trimethoprim-sulfamethoxazole vs doxycycline/hydroxychloroquine), clinicians should consider brain imaging in patients diagnosed with Whipple's endocarditis.

Constipation in long-term ventilated patients: associated factors and impact on intensive care unit outcomes.

OBJECTIVES: To characterize the factors associated with delayed defecation in long-term ventilated patients and to examine the relationship between delayed defecation and logistic organ dysfunction scores, acquired bacterial infections, and mortality in the intensive care unit. DESIGN: Prospective observational cohort study. SETTING: A 21-bed polyvalent intensive care unit in a university hospital. PATIENTS: A total of 609 adult patients admitted over a 41-month period who underwent mechanical ventilation for ≥ 6 days. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: Three hundred fifty-three patients (58%) passed stools ≥ 6 days after they were admitted to the intensive care unit ("late" defecation). Patients with early and late defecation had similar general characteristics when admitted to the intensive care unit and had similar logistic organ dysfunction scores on the first day of mechanical ventilation. Several variables were independently associated with a delay in defecation: a Pao2/Fio2 ratio of less than 150 mm Hg (adjusted hazard ratio 1.40; 95% confidence interval: 1.06-1.60; p = .0073), a systolic blood pressure between 70 and 89 mm Hg (adjusted hazard ratio 1.48; 95% confidence interval: 1.17-1.79; p = .002), and systolic blood pressure < 68 mm Hg (adjusted hazard ratio 1.29; 95% confidence interval: 1.01-1.60; p = .03). Logistic organ dysfunction scores were significantly higher on the fourth and ninth days of mechanical ventilation in patients with late defecation than in those with early defecation. The crude intensive care unit mortality rate was 18% in patients with early defecation and 30% in patients with late defecation (p < .001). Acquired bacterial infections at any site occurred in 34% of patients with early defecation and 66% of patients with late defecation (p < .001). CONCLUSION: A Pao2/Fio2 ratio of < 150 mm Hg and systolic blood pressure of < 90 mm Hg during the first 5 days of mechanical ventilation were independently associated with a delay in defecation. Our results suggest that constipation is associated with adverse outcomes in long-term ventilated patients.

Low occurrence of safety hazards in coagulase negative staphylococci isolated from fermented foodstuffs.

Some coagulase negative staphylococci (CNS) species play an important role in the fermentation of meat and milk products and are considered as food-grade. However, the increasing clinical significance of CNS and the presence of undesirable and unsafe properties in CNS question their presence or use in food. Our goal was to assess the safety of CNS by developing a diagnostic microarray targeting 268 genes corresponding to safety hazards in a food context i.e. toxins (especially enterotoxins) and determinants of antibiotic resistance and biogenic amine production. Target genes were selected among staphylococci and Gram-positive species that may be in contact with CNS in foodstuffs. The diagnostic microarray was used to screen 129 strains belonging to the 2 dominant species isolated from foodstuffs (S. equorum and S. xylosus) and the 2 main species isolated both in foodstuffs and clinical samples (S. epidermidis and S. saprophyticus). Microarray data were further completed by antibiograms and measurement of biogenic amine production. Safety hazards associated with CNS were mostly limited to the presence of antibiotic resistance. Seventy-one percent of the strains possessed at least one gene encoding antibiotic resistance, while only one strain carried an enterotoxin gene. Most strains did not carry any genes encoding staphylococcal toxins (68%), non-staphylococcal toxins (95%) or decarboxylases involved in biogenic amine production (78%). Food safety hazards were more pronounced in S. epidermidis than in the three other species regardless the food or clinical origin of the strains. Seventy-six percent of the strains carrying genes encoding staphylococcal toxin and 69% of strains carrying 5 or more antibiotic determinants belonged to S. epidermidis species. The dominant antibiotic resistance targeted erythromycin, tetracycline and penicillin and were generally traced back to the presence of tetK and blaZ in the two latest cases. Six percent of the food-related strains produced significant amounts of biogenic amines in vitro without any of the corresponding genes detected, reflecting a lack of knowledge on genetic determinants of such production in staphylococci. This work gives a first picture of safety hazards within four species of CNS frequently isolated from food or clinical environment.