Stephen C. Edberg

Recognition of Commensal Microflora by Toll-Like Receptors Is Required for Intestinal Homeostasis

Toll-like receptors (TLRs) play a crucial role in host defense against microbial infection. The microbial ligands recognized by TLRs are not unique to pathogens, however, and are produced by both pathogenic and commensal microorganisms. It is thought that an inflammatory response to commensal bacteria is avoided due to sequestration of microflora by surface epithelia. Here, we show that commensal bacteria are recognized by TLRs under normal steady-state conditions, and this interaction plays a crucial role in the maintenance of intestinal epithelial homeostasis. Furthermore, we find that activation of TLRs by commensal microflora is critical for the protection against gut injury and associated mortality. These findings reveal a novel function of TLRs-control of intestinal epithelial homeostasis and protection from injury-and provide a new perspective on the evolution of host-microbial interactions.

Escherichia coli : the best biological drinking water indicator for public health protection

Public health protection requires an indicator of fecal pollution. It is not necessary to analyse drinking water for all pathogens. Escherichia coli is found in all mammal faeces at concentrations of 10 log 9−1, but it does not multiply appreciably in the environment. In the 1890s, it was chosen as the biological indicator of water treatment safety. Because of method deficiencies, E. coli surrogates such as the ‘fecal coliform’ and total coliforms tests were developed and became part of drinking water regulations. With the advent of the Defined Substrate Technology in the late 1980s, it became possible to analyse drinking water directly for E. coli (and, simultaneously, total coliforms) inexpensively and simply. Accordingly, E. coli was re‐inserted in the drinking water regulations. E. coli survives in drinking water for between 4 and 12 weeks, depending on environmental conditions (temperature, microflora, etc.). Bacteria and viruses are approximately equally oxidant‐sensitive, but parasites are less so. Under the conditions in distribution systems, E. coli will be much more long‐lived. Therefore, under most circumstances it is possible to design a monitoring program that permits public health protection at a modest cost. Drinking water regulations currently require infrequent monitoring which may not adequately detect intermittent contamination events; however, it is cost‐effective to markedly increase testing with E. coli to better protect the publics health. Comparison with other practical candidate fecal indicators shows that E. coli is far superior overall.

Association of Streptococcus bovis with carcinoma of the colon.

Two patients with colonic adenocarcinoma and Streptococcus bovis endocarditis suggested a possible association between the two. Non-enterococcal Group D streptococci were isolated from fecal cultures of 11 of 105 controls, 35 of 63 patients with carcinoma of the colon, seven of 25 with inflammatory bowel disease, four of 21 with non-colonic neoplasms and five of 37 with other gastrointestinal disorders. All such streptococci examined for lactose fermentation were S. bovis. The prevalence of S. bovis in fecal cultures from patients with carcinoma of the colon was significantly increased (P less than 0.001) as compared to that in controls, and also to all other groups (P less than 0.001). No other group had results significantly different from those of controls (P less than 0.05) although patients with inflammatory bowel disease were more frequently carriers. The carrier state was unrelated to age, hospitalization status, colonic stasis, gastrointestinal bleeding or recent barium-enema examination. The implications of this association are unknown.

A ten-year review of neonatal sepsis and comparison with the previous fifty-year experience.

Records have been kept prospectively in our institution since 1928 of all positive blood cultures taken from neonates. Using a modification of objective Centers for Disease Control criteria to define sepsis, we reviewed the records of all neonates with positive blood cultures for the years 1979 to 1988 inclusive and found 270 cases of sepsis. The sepsis rate for infants less than or equal to 30 days of age was 2.7 cases/1000 live births, with a mortality rate from sepsis of 15.9%. There was an increase in sepsis due to commensal species (CS) over the period (P less than 0.007). The number of infants in the nursery who developed sepsis when more than 30 days of age also increased (P less than 0.002), as did the rate of sepsis from CS in this group (P less than 0.001). Isolation of CS from the blood with fulfillment of the modified Centers for Disease Control criteria was associated with a 13.7% mortality rate, whereas isolation of CS without fulfillment was associated with a 4% rate (P less than 0.01).

Streptococcus bovis Septicemia and Carcinoma of the Colon

We prospectively studied patients with Streptococcus bovis septicemia for the presence of gastrointestinal lesions. This study was prompted by our reported findings of the association of fecal carriage of S. bovis with carcinoma of the colon. We studied 29 patients with 30 episodes of S. bovis septicemia. Fifteen completed gastrointestinal evaluations that included colonscopy, surgery, or autopsy. Eight of these had carcinoma of the colon, three had adenomatous polyps of the colon without carcinoma, and two had carcinoma of the esophagus. The 14 patients who did not have complete evaluations included one each with carcinoma of the stomach, gastric lymphoma, and adenomatous polyp of the colon and three with colonic masses not further delineated. Nineteen patients had no gastrointestinal signs or symptoms or stools positive for occult blood at admission. The results of our study suggest that all patients with S. bovis septicemia need aggressive evaluation of the gastrointestinal tract, especially the colon.

Bacteriophages as indicators of enteric viruses and public health risk in groundwaters.

1. Introduction, 5

Diarrheagenic Escherichia coli Infection in Baltimore, Maryland, and New Haven, Connecticut

BACKGROUND Diarrhea remains a common complaint among US patients who seek medical attention. METHODS We performed a prospective study to determine the etiology of diarrheal illness among patients and control subjects of all ages presenting to the emergency departments and outpatient clinics of 2 large academic hospitals in Baltimore, Maryland, and New Haven, Connecticut. We used molecular methods to detect the presence of diarrheagenic Escherichia coli pathotypes, including enteroaggregative E. coli (EAEC), as well as Shiga toxin-producing, cytodetaching, enterotoxigenic and enteropathogenic E. coli. RESULTS Of the pathotypes sought, only EAEC was found in an appreciable proportion (4.5%) of case patients, and it was found more frequently among case patients than control subjects (P<.02). Surprisingly, EAEC was the most common bacterial cause of diarrhea in our population. EAEC was common in all age strata and was not associated with foreign travel or immunodeficiency. EAEC infection is frequently accompanied by fever and abdominal pain, though this did not happen more frequently in patients with EAEC infection than in patients with diarrhea due to other causes. CONCLUSIONS Our data suggest that EAEC infection should be considered among persons with diarrhea that does not yield another known etiologic agent.

Pseudomonas aeruginosa : Assessment of risk from drinking water

Pseudomonas aeruginosa is an ubiquitous environmental bacterium. It can be recovered, often in high numbers, in common food, especially vegetables. Moreover, it can be recovered in low numbers in drinking water. A small percentage of clones of P. aeruginosa possesses the required number of virulence factors to cause infection. However, P. aeruginosa will not proliferate on normal tissue but requires previously organs. Further narrowing the risk to human health is that only certain specific hosts are at risk, including patients with profound neutropenia, cystic fibrosis, severe burns, and those subject to foreign device installation. Other than these very well-defined groups, the general population is refractory to infection with P. aeruginosa. Because of its ubiquitous nature, it is not only not practical to eliminate P. aeruginosa from our food and drinking water, but attempts to do so would produce disinfection byproducts more hazardous than the species itself. Moreover, because there is no readily available sensitive and specific means to detect and identify P. aeruginosa available in the field, any potential regulation governing its control would not have a defined laboratory test measure of outcome. Accordingly, attempts to regulate P. aeruginosa in drinking water would not yield public health protection benefits and could, in fact, be counterproductive in this regard.

The value and limitations of the Gram stain examination in the diagnosis of intraamniotic infection

Gram stain examination of amniotic fluid is a method used for the rapid diagnosis of intraamniotic infection in patients with preterm premature rupture of membranes and preterm labor. The management of these patients relies heavily on the Gram stain results. Therefore, it is critical that the diagnostic value and limitations, optimal technique, and outcome correlates be precisely established. Most studies have focused on culture results rather than on Gram stain results to describe neonatal and maternal outcome. However, management is based on the Gram stain because culture results are not immediately available. One hundred eighty-seven amniocenteses were performed in 131 patients with preterm premature rupture of the membranes (n = 90) and preterm labor (n = 41). Spun and unspun Gram stains were performed. Centrifugation of the sample did not improve the sensitivity of the technique significantly. The agreement between the two methods was substantial (kappa index 0.89, p less than 0.001). The sensitivity of the Gram stain was 44.8% and the specificity was 97.6%. The sensitivity of the Gram stain was directly proportional to the number of bacteria present in amniotic fluid. In the presence of greater than 10(5) colony forming units per milliliter, 80% of the Gram stains were positive. The absence of both bacteria and white blood cells in a smear was associated with a negative culture of amniotic fluid in 95% of the cases. Clinical chorioamnionitis was associated with a positive Gram stain of amniotic fluid (p less than 0.001). There was a trend toward a higher incidence of endometritis in patients with a positive Gram stain compared with those with a negative Gram stain (p = 0.07). There was no neonatal infectious morbidity in patients with a true negative Gram stain. Patients with a false negative Gram stain had a 25% incidence of neonatal infectious complications (proved and suspected sepsis).

Gentamicin Uptake in Wild-Type and Aminoglycoside-Resistant Small-Colony Mutants of Staphylococcus aureus

Gentamicin uptake and killing were studied in aminoglycoside-susceptible wild-type Staphylococcus aureus strains and aminoglycoside-resistant small-colony mutants selected by gentamicin from these strains. In wild-type S. aureus three phases of gentamicin accumulation were noted, and killing occurred during the last and most rapid phase of uptake. Uptake and killing were abolished by anaerobic growth and sodium azide, suggesting that energy-dependent active drug transport required respiration. Treatment of wild-type strains with the uncouplers N,N′-dicyclohexyl carbodiimide (DCCD) and carbonyl cyanide-m-chlorophenyl hydrazone showed disparate effects on gentamicin uptake, producing enhanced and diminished accumulations, respectively. Small-colony mutants demonstrated markedly deficient uptake compared with the wild-type strains and were not killed by gentamicin in concentrations up to 10 μg/ml. Several classes of aminoglycoside-resistant mutant strains are described. One mutant strain was a menadione auxotroph which, when grown in the presence of menadione, exhibited normal gentamicin uptake and killing. Gentamicin uptake and killing in this strain were abolished by KCN when the strain was grown in a medium supplemented with menadione. The membrane adenosine triphosphatase inhibitor DCCD was lethal for this mutant but not for other mutants or wild-type strains. Preincubation with menadione prevented the lethal effect of DCCD, and this strain demonstrated normal gentamicin accumulation when exposed to both DCCD and menadione. A second mutant strain demonstrated both gentamicin uptake and killing in the presence but not the absence of DCCD. Studies with small-colony mutants of S. aureus indicated that the defect in aminoglycoside uptake is very likely related to an inability to generate or maintain energized membranes from respiration. These studies suggest that the membrane energization associated with active aminoglycoside accumulation requires electron transport for the generation of a protonmotive force.