An epidemiological study of enteric viruses in sewage with molecular characterization by RT-PCR and sequence analysis.

The aim of this study was to assess the presence and seasonal frequency of various enteric viruses in wastewater treatment. The detection of astrovirus, norovirus, enterovirus, hepatitis A virus (HAV) and rotavirus was carried out by molecular analyses in concentrated water samples collected over 18 months at the entrance and exit of an activated sludge sewage treatment plant. The reverse transcriptase-polymerase chain reaction (RT-PCR) results were confirmed by sequencing, and comparative phylogenetic analysis was performed on the isolated strains. Genomes of human astrovirus and human rotavirus were identified in 26/29 and 11/29 samples of raw sewage, respectively, and in 12/29 and 13/29 treated effluent samples, respectively. Some rotavirus sequences detected in environmental samples were very close to those of clinical strains. Noroviruses, enteroviruses and HAV were not detected during the study period. This could be related to the small sample volume, to the sensitivity of the detection methods or to local epidemiological situations. Frequent detection of viral RNA, whether infectious or not, in the exit effluent of sewage treatment indicates wide dispersion of enteric viruses in the environment. Consequently, viral contamination resulting from the use of these treated waters is a risk that needs to be addressed.

Capsid protein composition of reference strains and wild isolates of human astroviruses.

Astroviruses are small RNA viruses that are frequently associated with gastroenteritis in humans and animals. Despite much work on the genetic analysis of astrovirus strains, little progress has been made in the characterization of the proteins composing mature virions. We have analyzed the capsid protein composition of the reference strains and several wild isolates of human astroviruses using high-resolution polyacrylamide gradient gels. For reference strains of the seven serotypes analyzed, a consistent pattern of three infection-specific proteins--designated P1, P2, and P3 -was generally observed. The strains could be divided into two groups, based upon the reactivity of these proteins in immune precipitation assays that used homologous rabbit serum. One group included reference types 1 4 for which all three proteins were precipitated by homologous rabbit sera; for the other group, types 5 7, only proteins P2 and P3 were precipitated. When wild isolates from around the world were compared to the reference strains, a correlation between genetic type and the pattern of protein sizes and immune reactivity was observed for strains of the common types (1-4). Strains of types 2 and 4 consistently exhibited P3 proteins larger than those of types 1 and 3. Unusual patterns of proteins or immune reactivity were detected in strains of types 5-7, indicating that there may be incomplete processing of the capsid precursor during growth in cell culture.

RT-PCR identification and typing of astroviruses and Norwalk-like viruses in hospitalized patients with gastroenteritis: evidence of nosocomial infections.

BACKGROUND: Astroviruses (HAstVs) and 'Norwalk-like viruses' (NLV) are frequent causes of gastroenteritis worldwide, though no data on the strains in circulation or their prevalence is available for France. OBJECTIVES: We applied molecular methods to detect HAstVs and NLVs by reverse transcription-polymerase chain reaction (RT-PCR) in fecal samples collected during a 2-year period from children and adults hospitalized with gastroenteritis. STUDY DESIGN: All samples negative for rotavirus and adenovirus by latex agglutination which contained small (25-40 nm) viral particles observed by electron microscopy (EM) were examined by RT-PCR. RT-PCR products were sequenced to characterize the HAstV and NLV strains present. RESULTS: A total of 75 samples were analyzed by RT-PCR, of which 15 were positive for HAstV and 24 for NLV. Several distinct strains of serotype 1 HAstV, the predominant serotype, circulated during the period. Nineteen of the 24 NLVs were of the G2 genogroup including Mexico-like (n=10), Bristol-like (n=8), and Hawaii-like viruses (n=1); two were genogroup 1. Overall, seven (47%) of the 15 HAstV infections and nine (37.5%) of the 24 NLV infections appeared to be nosocomially acquired based on the date of admission in hospital and the date of illness. CONCLUSION: This study provides additional evidence of the importance of nosocomial infections caused by NLV and HAstV.

Effect of glutaraldehyde on the antigenicity and infectivity of hepatitis A virus.

The effect of glutaraldehyde on the antigenicity and infectivity of hepatitis A virus (HAV) was examined. The CF 53 strain, adapted to human hepatoma PLC/PRF/5 cells, was treated with glutaraldehyde using three different concentrations, 0.02, 0.10, and 0.50%, for various periods of time, 3, 10, and 30 min, respectively. After the virucidal assays, glutaraldehyde and HAV were separated by gel filtration, then the antigen (radioimmunoassay) titer and the infectivity titer were determined. The greatest antigen titer reduction was about 80% after 30 min using 0.10% glutaraldehyde and within only 3 min using 0.50% glutaraldehyde. Glutaraldehyde is an effective disinfectant against HAV: the infectious virus titer decreased by more than 3 log10 after 30 min using 0.10% glutaraldehyde and within only 3 min using 0.50% glutaraldehyde. Statistical studies showed that the decrease of antigen or infectious virus titer was affected by both glutaraldehyde concentration and exposure time.

An in-vitro evaluation of the activity of povidone-iodine against nosocomial bacterial strains.

Two povidone-iodine (PVP-I) preparations, one, an antiseptic handwash and one, a skin disinfectant, were tested against 504 bacterial strains isolated from nosocomial infections in 12 French hospitals. In vitro bactericidal activity was determined by a micromethod, using specific interfering substances over a range of dilutions, after 1, 3 and 5 min exposure times. A 5 log10 reduction of the challenge inoculum was considered as the criterion of efficacy. Any resistant strains were tested with the French Standard (T72300). When the micromethod was carried out at 20 degrees C, 10.7% (54/504) of the strains were resistant to the PVP-I skin disinfectant (dilution 1:10) and 1.6% (8/504) were resistant to the handwashing formulations (dilution 1:3) after 1 min exposure. By increasing the temperature to 32 degrees C, the resistance rate to the skin disinfectant fell to 1.9% (10/504). All of the 18 strains found resistant with the micromethod were sensitive using the French standard.

Role of infection control measures in limiting morbidity associated with multi-resistant organisms in critically ill patients.

A retrospective comparative study was performed to determine the impact of infection control measures (ICMs) on colonization and infections due to methicillin-resistant Staphylococcus aureus (MRSA), Klebsiella pneumoniae (producing transferable extended-spectrum beta-lactamase, KPESBL), and multi-resistant Enterobacter aerogenes (MREA) in intensive care unit patients. Infection Control Measures included surveillance cultures, isolation procedures and mupirocin for MRSA nasal carriage. The numbers of patients infected and/or colonized by MRSA, KPESBL or MREA were compared during two consecutive one-year periods (Period 1 before ICMs, and Period 2 after ICMs). The antibiotic consumption during the two periods was analysed. In Period 1 and Period 2, respectively, the rate of patients infected or colonized by at least one of the three organisms was 15% and 6.8% (P=0.001); by MRSA 7.7% and 2.6% (P=0. 004); by KPESBL 1.7% and 0% (P=0.25); and by MREA 5.6% and 4.3% (P=0. 47). During Period 2, there was a clear-cut decrease in the percentage of patients infected by MRSA (P=0.018), a non-significant decrease in those infected by KPESBL (P=0.06), and no decrease in patients infected by MREA (P=0.22). When calculated per 1000 patient-days, for Period 1 and Period 2, respectively, the rate of patients infected or colonized by at least one of the three organisms was 11.9 and 8.8; for MRSA it was 4 and 2.2; for KPESBL it was 1 and 0; and for MREA it was 4 and 4. Antibiotic cost was pound98.7 in Period 1 and pound62.7 in Period 2. ICMs contributed to the control of infections and colonizations due to MRSA and KPESBL but not those due to MREA.

Comparison of in-vivo antibacterial activity of two skin disinfection procedures for insertion of peripheral catheters: povidone iodine versus chlorhexidine.

Skin disinfection is a key step in the prevention of nosocomial infections especially prior to invasive procedures such as the insertion of peripheral catheters. Alcohol-based antiseptics improve bactericidal activity and decrease the time needed for skin disinfection in emergencies. A randomized study was performed in two groups of 22 volunteers to compare the in vivo bactericidal effect of two rapid disinfection procedures using povidone iodine (PVP-I) in scrub formulation followed by alcoholic PVP-I, or chlorhexidine in scrub formulation followed by alcoholic chlorhexidine. Bacteria were recovered using the cylinder scrub method. Comparison of reductions in the aerobic and anaerobic flora from baseline levels to each of the three sampling times (30 sec, 3 min, 2 h) showed no significant difference between the two procedures Log(10)reduction after 30 seconds was around 1.5 for the aerobic flora and 1.1 for the anaerobic flora. After 3 minutes the corresponding values were 2.1 and 1.8, and after 2 hours 2.0 and 1.3. The products were well tolerated in both groups. The two procedures had comparable rapid bactericidal activity in vivo.

Characteristics and diffusion in the rabbit of a phage for Escherichia coli 0103. Attempts to use this phage for therapy.

A bacteriophage for Escherichia coli 0103 was isolated during a study on E. coli diarrhoea in intensive breeding units of rabbits. The phage had an isometric head and a short tail and resembled coliphage N4 (Podoviridae). It had a very narrow host range and seemed to be specific for serogroup 0103, suggesting that it might be used for preliminary identification of E. coli strains of this serogroup instead of the usual slide agglutination. In view of its possible use as a therapeutic phage, we investigated its dissemination in rabbit organs after oral administration. The phage persisted in the spleen for at least 12 days. However, in vivo studies showed that this phage and a mixture of more virulent phages for E. coli 0103 were ineffective in preventing disease in rabbits inoculated with an enteropathogenic strain of E. coli 0103.

[Evaluation of 4 years of surveillance of enteroviruses in sewage. Correlation with human pathology]. / Bilan de quatre années de dépistage des entérovirus dans les eaux usées. Corrélations avec la pathologie humaine.

This survey is concerned with isolations of enteroviruses from sewage and stools of children admitted to pediatric wards of the Clermont-Ferrand hospital during 4 years (from january 1, 1980, to december 31, 1983). Some epidemics of different serotypes (Coxsackies B1, B4, B5, Echovirus 33) occurred, mainly between june and october. In some instances, virus isolation in sewage occurred a few weeks before epidemic of the same virus, giving some predictive value to pediatric pathology. This survey allowed us to adapt immunologic diagnosis of enteroviruses in children and limit the use of expensive reagents.

[A practical method of numerical evaluation applicable to miniaturized technics of plaque assay]. / Méthode pratique d'évaluation numérique applicable aux techniques miniaturisées de titrage en plages

When one wishes to titrate viruses by counting the number of infective foci on small surfaces of observation, one is rapidly limited by confluences which render counting impossible. Between the use of an all or nothing system which is not accurate and a total field of count, often impossible, there is an intermediate method already described and studied by Tippett, which is very little used in practice. We have performed the calculations again, drawn up reference values and given rules for choice of the field. Two examples show their practical use.

[Bacterial contamination of eyedrops in clinical use]. / Etude de la contamination bactérienne de collyres en usage clinique.

PURPOSE: The aim of this study is to assess the importance of bacterial contamination of multidose eyedrops in a routine clinical setting. MATERIAL AND METHODS: 406 eyedrop vials were cultured about one week after clinical use: 204 collected from an Ophthalmic Department, and 202 from a Nursing Home. The microbiological analysis was performed on the tip and the residual eyedrop, counting the number of bacterial colonies. RESULTS: 66 (16.3%) from the 406 analyzed vials were contaminated, and 5.4% out of these were severely affected. There was no significant difference between the "Ophthalmic Department" and the "Nursing Home". Commensal germs were the most frequently encountered in both groups. 4 gram negative organisms were isolated from the "Nursing Home" group. CONCLUSION: These results are in agreement with the literature. Comparison between our two groups is difficult because the eyedrops and uses were different. However, we notice the presence of gram negative organisms in the "Nursing Home". These severe contaminations due to opportunistic pathogen organisms are rare (0.75%), probably underestimated, and represent a real infectious risk during instillation. The study of the contamination site shows that the eyedrop is more often contaminated than the tip. This can be in relation to germ desicsation and to an aspiration phenomenon of contaminated fluid at the tip level. At last, the role of preservatives is not sufficient to ensure the sterility of multidose eyedrops during their use, and this justifies safer (single dose or filtration system) eyedrop vials.

[Laryngoscope. Evaluation of a device for preventing blade contamination]. / Laryngoscope. Evaluation d'un dispositif de prévention de la contamination des lames.

OBJECTIVE: To evaluate the efficacy and the difficulty of use of a disposable sheath which prevents the contamination of blades. STUDY DESIGN: Prospective bacteriological, virological and clinical evaluation. MATERIAL: A translucid cover sheath, made of polyethylene enclosing the blade of the laryngoscope, and delivered in clean, non-sterile packaging (Prolam, Péters). METHOD: 1) A control of sterility performed by setting-up a culture derived from the solution used for rinsing the device before its use. 2) An in vitro study of the effectiveness of preventing contamination of the blades by a polio virus/RT-PCR technique. 3) Clinical evaluation: after 200 orotracheal intubations by 12 anaesthesiologists and 15 nurse anaesthetists, a questionnaire on the ease of use was completed. RESULTS: The bacteriological study of the sheats before use showed an acceptable level of contamination. The sheath was an effective barrier against poliovirus, even after 12 h of immersion. Clinically, the sheath was easily adapted over the blade of the laryngoscope in 98% of the cases. Insertion in the mouth was considered as easy in 94% of the patients. The visualization was good or excellent in 83% of the cases and in 16% of the patients, the users experienced difficulties to intubate. CONCLUSION: The laryngoscope blade sheath is simple and easy to use, efficient and not expensive.

[A simple and rapid method to test the permeability of condoms to viruses]. / Une méthode simple et rapide de contrôle de l'imperméabilité aux virus des préservatifs.

phi x 174 and T7 coliphages can be used to find out if condoms are virus-proof. Protection against hepatitis B virus or papillomavirus contamination is not assured when T7, a 65 nm broad virus, leaks through the condom. The simple test can be used to establish a quality control norm for condoms.

Comparison of bacteriophage and enteric virus removal in pilot scale activated sludge plants.

AIMS: The aim of this experimental study was to determine comparatively the removal of two types of bacteriophages, a somatic coliphage and an F-specific RNA phage and of three types of enteric viruses, hepatitis A virus (HAV), poliovirus and rotavirus during sewage treatment by activated sludge using laboratory pilot plants. METHODS AND RESULTS: The cultivable simian rotavirus SA11, the HAV HM 175/18f cytopathic strain and poliovirus were quantified by cell culture. The bacteriophages were quantified by plaque formation on the host bacterium in agar medium. In each experiment, two pilots simulating full-scale activated sludge plants were inoculated with viruses at known concentrations, and mixed liquor and effluent samples were analysed regularly. In the mixed liquor, liquid and solid fractions were analysed separately. The viral behaviour in both the liquid and solid phases was similar between pilots of each experiment. Viral concentrations decreased rapidly following viral injection in the pilots. Ten minutes after the injections, viral concentrations in the liquid phase had decreased from 1.0 +/- 0.4 log to 2.2 +/- 0.3 log. Poliovirus and HAV were predominantly adsorbed on the solid matters of the mixed liquor while rotavirus was not detectable in the solid phase. In our model, the estimated mean log viral reductions after 3-day experiment were 9.2 +/- 0.4 for rotavirus, 6.6 +/- 2.4 for poliovirus, 5.9 +/- 3.5 for HAV, 3.2 +/- 1.2 for MS2 and 2.3 +/- 0.5 for PhiX174. SIGNIFICANCE AND IMPACT OF THE STUDY: This study demonstrates that the pilots are useful models to assess the removal of infectious enteric viruses and bacteriophages by activated sludge treatment. Our results show the efficacy of the activated sludge treatment on the five viruses and suggest that coliphages could be an acceptable indicator of viral removal in this treatment system.

[Influence of some substances on the titration of a poliovirus aqueous suspension (author's transl)]. / Influence de quelques substances sur le titre apparent d'un poliovirus en suspension aqueuse

Important changes of virus titer are observed when low concentration of mineral salts (NaCl, MgCl2, AlCl3) or detergents (Tween 80) are added to a poliovirus suspension in distilled water. Calf serum addition to these suspensions induces new variations of virus titer. The highest titre is obtained in isotonic medium with calf serum.