Genomic epidemiology of the Escherichia coli O104:H4 outbreaks in Europe, 2011.

The degree to which molecular epidemiology reveals information about the sources and transmission patterns of an outbreak depends on the resolution of the technology used and the samples studied. Isolates of Escherichia coli O104:H4 from the outbreak centered in Germany in May-July 2011, and the much smaller outbreak in southwest France in June 2011, were indistinguishable by standard tests. We report a molecular epidemiological analysis using multiplatform whole-genome sequencing and analysis of multiple isolates from the German and French outbreaks. Isolates from the German outbreak showed remarkably little diversity, with only two single nucleotide polymorphisms (SNPs) found in isolates from four individuals. Surprisingly, we found much greater diversity (19 SNPs) in isolates from seven individuals infected in the French outbreak. The German isolates form a clade within the more diverse French outbreak strains. Moreover, five isolates derived from a single infected individual from the French outbreak had extremely limited diversity. The striking difference in diversity between the German and French outbreak samples is consistent with several hypotheses, including a bottleneck that purged diversity in the German isolates, variation in mutation rates in the two E. coli outbreak populations, or uneven distribution of diversity in the seed populations that led to each outbreak.

Detection of HPV DNA in cervical specimens collected in cytologic solution by ligation-dependent PCR.

OBJECTIVE: To determine the feasibility and sensitivity of detecting human papillomavirus (HPV) in specimens collected in Cytyc PreservCyt fluid (Boxborough, Massachusetts, U.S.A.) using ligation-dependent polymerase chain reaction (LD-PCR) and to demonstrate the diagnostic value of HPV DNA testing as an adjunct to cytology in the detection of cervical squamous intraepithelial lesions (SIL), especially in cases of atypical squamous cells of undetermined significance (ASCUS). STUDY DESIGN: LD-PCR is a recently invented DNA amplification technology that utilizes a capture probe for target isolation and 2 hemiprobes for target detection. The hemiprobes are designed in such a way that when they hybridize to their target, the 5' end of one probe and the 3' end of the other probe are brought together. Two hemiprobes can then be ligated into a full probe that can serve as a template for PCR amplification. A total of 94 cervical specimens were collected in cytologic fluid and tested with LD-PCR. The results were compared with those of the Digene Hybrid Capture II assay (HC II) (Beltville, Maryland, U.S.A.) and consensus PCR. RESULTS: The overall sensitivity for detecting HPV was 41.5% (39/94) by LD-PCR, 50% (47/94) by consensus PCR and 37.2% (35/94) by HC II. The prevalence of HPV by HC II, consensus PCR and LD-PCR were 87.5%, 100% and 87.5% in the high grade SIL group; 100%, 90.9% and 90.9% in the low grade SIL group; 30%, 52.5% and 40% in the ASCUS group; and 14.2%, 22.8% and 17.1% in women with normal cytology. These results indicate that all 3 methods have similar sensitivity in patients with SIL. However, there is greater variation in detection rates in the ASCUS and normal cytology groups. CONCLUSION: LD-PCR is a useful method of detecting HPV in liquid-based gynecologic cytologic preservatives, and HPV testing as a method adjunct to the liquid-based Pap test could be useful in detecting SILs, especially for the management of patients with ASCUS.

Variable effects of short chain fatty acids and lactic acid in inducing intestinal mucosal injury in newborn rats.

BACKGROUND: Short chain fatty acids and lactic acid are colonic bacterial fermentation products. METHODS: To evaluate the effects of these organic acids on the intestinal mucosa, a total of 72 newborn Sprague-Dawley rats (10 days old) were studied. A 3.5F catheter was inserted per rectum 4.0 cm deep into the proximal colon for organic acid administration at a volume of 0.1 ml/10 g body weight. The pH of organic acid solutions and normal saline was adjusted to 4.0. Group 1 (n = 10) received normal saline as a control. Group 2 (n = 11) received 150 mM acetic acid. Group 3 (n = 11) received 300 mM acetic acid. Group 4 (n = 10) received 150 mM butyric acid. Group 5 (n = 11) received 300 mM butyric acid. Group 6 (n = 7) received 150 mM lactic acid, and group 7 (n = 12) received 300 mM lactic acid. Animals were killed 24 hours after colonic installation of test solutions. RESULTS: Both 300 mM acetic acid and 300 mM butyric acid were associated with impaired weight gain, increased colon wet weight, and increased histologic injury scores in the colon and distal ileum (P < 0.05, analysis of variance). Both 150 mM acetic acid and butyric acid at 150 mmol/L induced minimal injury in the colon and distal ileum. Neither 150 mM nor 300 mM lactic acid induced any identifiable gross or microscopic intestinal mucosal injury. CONCLUSION: Luminal short chain fatty acids can induce dose-dependent intestinal mucosal injury in newborn rats, resembling the pathology seen in neonatal necrotizing enterocolitis. Overproduction/accumulation of short chain fatty acids, but not lactic acid, in the proximal colon and/or distal ileum may play a role in the pathogenesis of necrotizing enterocolitis in premature infants.

Vitamin A supplementation ameliorates butyric acid-induced intestinal mucosal injury in newborn rats.

Vitamin A (vit A) plays an important role in wound healing and therefore may help in repairing of intestinal mucosal injury. The purpose of this study was to determine if vit A supplementation could promote healing in intestinal mucosal injury as commonly seen in neonatal necrotizing enterocolitis (NEC). Mild intestinal mucosal injury was induced in 10-day-old Sprague-Dawley rats by luminal administration of 1.5% butyric acid (BA) at pH 4.0. Normal saline at the same pH was administered as control. Immediately after administrations of BA or normal saline, animals were randomly assigned to receive high dose vit A (20,000 IU/kg for one dose, i.p.), low dose vit A (5,000 IU/kg for two doses) or vehicle. Animals were followed for 48 hours and then sacrificed for histological examination. Rats with BA-induced intestinal mucosal injury had a reduction in daily weight gain (p < 0.05). Vit A supplementation significantly improved the daily weight gain in the rats with BA-induced intestinal mucosal injury and the effect is dose dependent. At sacrifice, the colon wet weight was significantly heavier and the histological injury scores from both ileum and proximal colon higher in the rats with BA-induced intestinal mucosal injury. All of those parameters were improved with vit A supplementation. We conclude that vit A supplementation ameliorates BA induced-intestinal mucosal injury in newborn rats.