New patterns of drinking-water consumption: results of a pilot study.

A pilot study on water consumption was carried out in the Québec City region in April and May 1996 with 125 people using a 24-h recall plus a 2-day diary. Consumption of drinking water via liquid and food was assessed as well as the type of water consumed (tap, bottle or filtered water) and place of consumption (home or away from home). Most of the people (56%) were drinking some bottled water or filtered tap water and 25% of water intake was away from home. Food consumption was found to be a non-significant source of drinking-water intake. The average water consumption was nearly similar in exclusively tap-water consumers and bottled- or filtered-water consumers (1.5 vs. 1.7 l/day, P = 0.29) but two-thirds of the consumption in this last group is natural water, while it is mixed water in the bottled/filtered-water group. No significant difference in amounts consumed were found according to age, but older people drank hot beverages and soup more often. The present pilot-study was weakened by a low participation rate (14%). Incentive might be necessary to improve participation rate and data collection methods must also be simplified. A 24-h recall plus a 1-day diary seem sufficient and data on consumption could be limited to liquids, soups and cereals.

Aeromonas hydrophila isolated from food and drinking water: hemagglutination, hemolysis, and cytotoxicity for a human intestinal cell line (HT-29).

Aeromonas hydrophila isolated from food and drinking water was tested for pathogenicity by studying its hemolysis, hemagglutination, and cytotoxicity. Hemolysis, tested on erythrocytes from six different species, was more frequently seen with water isolates (64%) than with food isolates (48%). Hemagglutination was more frequently encountered with food isolates (92%) than with water isolates (73%). Cytotoxicity, evaluated on seven cell lines, was frequently observed with food isolates (92%) and with water isolates (73%). Heat treatment (56 degrees C for 10 min) of culture supernatant fluids inhibited the toxicity of some but not all toxin-producing isolates. Our results suggest that the human intestinal cell line HT-29 could be a useful complement for testing A. hydrophila exotoxins and for studying the enteropathogenicity of this species for humans.

Differential media for quantitative recovery of waterborne Aeromonas hydrophila.

Because of the ubiquity of Aeromonas spp., their prevalence in drinking water, and the increasing number of reports on Aeromonas sp.-related infections, a standard method for routine and quantitative recovery had to be defined. On the basis of a survey of 10 media for recovery analysis and subsequent differentiation assays in mixed cultures, we conclude that ampicillin-dextrin agar performed the best for the recovery of Aeromonas spp. in drinking water and the differentiation by simple criteria of that genus from other common waterborne bacteria.

Comparison of the microbiological quality of water coolers and that of municipal water systems.

The microbiological quality of tap water and that of water from 50 water coolers located in residences and workplaces were comparatively studied. In addition, difference factors that might influence the bacteriological contamination of water dispensers were examined. Aeorbic and facultative anaerobic heterotrophic bacteria, total coliforms, and two indicators for fecal contamination (fecal coliforms and fecal streptococci) as well as three types of pathogenic bacteria (Staphylococcus aureus, Pseudomonas aeruginosa, and Aeromonas spp.) were enumerated. It was found that 36 and 28% of the water dispenser samples from the residences and the workplaces, respectively, were contaminated by a least one coliform or indicator bacterium and/or at least one pathogenic bacterium. The respective proportions of tap water samples contaminated in a similar fashion were 18 and 22%, much less than those observed for water coolers (Chi2(1) = 3.71, P = 0.05). We were unable to discern the dominant factors responsible for the contamination of water coolers, but cleaning the water dispenser every 2 months seemed to limit the extent of contamination.

Isolation and sequence of a 2-kbp miniplasmid from Bacillus thuringiensis var. kurstaki HD-3a3b: relationship with miniplasmids of other B. thuringiensis strains.

The miniplasmid profiles of 18 Bacillus thuringiensis strains belonging to 8 different serotypes were determined using an alkaline hydrolysis method for isolation of low molecular mass plasmids. Nearly all the strains contained covalently closed circular (CCC) DNA species ranging from 2 to 5 species per strain and from 1.5 to 10.5 kbp in size (values corresponding to CCC forms). A 2-kbp plasmid from B. thuringiensis var. kurstaki HD-3a3b futura strain was used in Southern hybridization experiments to analyse relationships among the low molecular mass plasmids of different B. thuringiensis strains. This 2-kbp miniplasmid was present in most strains which show toxicity against lepidoptera. It was not present in those strains toxic against diptera (B. thuringiensis var. israelensis) or coleoptera (B. thuringiensis var. tenebrionis). The 2-kbp miniplasmid from B. thuringiensis var. kurstaki HD-3a3b futura was cloned and fully sequenced. Sequence analysis of the 2058 bp of the miniplasmid revealed the presence of an ORF (630 bp, 210 amino acids in size) that is preceded by a consensus sequence of B. thuringiensis crystal protein gene transcription promoters. No significant homology was observed with known B. thuringiensis toxin nucleic acid sequences or with other known sequences.

Effect of Feeding Lactic Acid Bacteria and Fermented Milk on Specific and Nonspecific Immune Reponses of Mice Infected With Klebsiella pneumoniae AD-1.

The diets of six groups of weaned mice were supplemented with ultra high temperature (UHT) milk containing a washed suspension of lactic acid bacteria (mixture of 8 strains) or with UHT milk fermented by the same strains and heat-treated or not. Control groups received physiological saline or UHT milk only. The mice were infected intranasally by Klebsiella pneumoniae AD-1 on the 13th d of feeding. The effect on the immune system (specific and nonspecific) before and after infection was evaluated by measuring the phagocytosis of alveolar macrophages (using zymosan particles) and by measuring of total immunoglobulin G and A levels in serum and in pulmonary fluid (using the enzyme-linked immunosorbent assay method). Postinfection survival was 0.7 d longer for mice receiving fermented milk than for the saline control group. The percent phagocytosis did not vary significantly, while serum immunoglobulin G levels differed between mice fed fermented milk and those fed bacterial suspensions in unfermented milk. Fermentation appears to be essential for the beneficial effects on the immune system and survival time; this effect no longer occurs after pasteurization of fermented milk.

[Biochemical diagnosis of the presence of poisoning by Bacillus thuringiensis serotype "H3a,3b" in two lepidopterans]. / Diagnostic biochimique de la présence d'une intoxication par Bacillus thuringiensis sérotype "H3a,3b" chez deux lépidoptères.

A detailed biochemical analysis has shown that during larval development on artificial medium, the amounts of K+, Na+, and Ca2+ in the hemolymph of healthy Choristoneura fumiferana varied from 85 to 110 mg/100 mL, 29 to 33 mg/100 mL, and 4.8 to 7.3 mg/100 mL, respectively. Similar results were obtained with Malacosoma disstria. Intoxication by Bacillus thuringiensis "H3a,3b" (B. t.) considerably modified the amounts of the cations. Thus, after 4 h, the quantity of K+ in M. disstria increased from 99 to 229 mg/100 mL and Na+ from 26.5 to 50.3 mg/100 mL while that of Ca2+ decreased from 5.8 to 1.2 mg/100 mL. Similar results were observed with C. fumiferana, but these variations occurred after 2 to 4 days of B. t. intoxication. The variations detected during the bacillosis, with respect to the cationic composition of the insect hemolymph, are rapidly detectable, well before light microscope observation can confirm the presence of this intoxication. Aspartate aminotransferase, alanine aminotransferase, alpha-hydroxybutyrate dehydrogenase, and isocitrate dehydrogenase activity fluctuated very slightly in the hemolymph of either healthy or bacillosed larvae of the two insects under study. These results suggest that it is possible to diagnose biochemically the presence of a B. t. intoxication in lepidopteran forest pests following treatments by this biological insecticide for their control.

Monoclonal antibodies to TEM-1 plasmid-mediated beta-lactamase.

At least 28 plasmid-mediated beta-lactamases have been described in gram-negative bacteria. To assess the relationship among these enzymes, we produced and characterized 28 murine monoclonal antibodies to the TEM-1 plasmid-mediated beta-lactamase. Radial immunodiffusion identified 3 monoclonal antibodies as immunoglobulin M (IgM), 18 as subclass IgG1, 2 as IgG2a, and 5 as IgG2b. Using a newly described enzyme immunoassay, cross-reactivity of 16 of these monoclonal antibodies was tested against 24 plasmid-determined beta-lactamases. The 16 monoclonal antibodies cross-reacted with TEM-2 and TLE-1 and, to a certain extent, SHV-1. Different levels of cross-reactivity were also observed with OXA-3 (11 of 16), OXA-7 (8 of 16), OXA-1 (2 of 16), OXA-6 (2 of 16), and AER-1 (2 of 16). Six monoclonal antibodies demonstrated partial neutralization of beta-lactamase activity. This study suggests that common epitopes are shared by nine biochemically distinct plasmid-mediated beta-lactamases. On the basis of cross-reactivities with these monoclonal antibodies, we identified four epitopes on TEM-1, TEM-2, TLE-1, and SHV-1 beta-lactamases.

Efficacy of different dosing schedules of tobramycin for treating a murine Klebsiella pneumoniae bronchopneumonia.

The efficacy of different dosing schedules of tobramycin for treating a murine Klebsiella bronchopneumonia was compared. Therapeutic efficacy depended upon dosing intervals. In mice treated for four days with a daily dose of 48 mg/kg, dosing intervals of 4 and 8 h allowed cure of 10/10 animals, whereas dosing intervals of 12 and 24 h yielded survival rates of only 6/10 (P less than 0.05) and 2/10 (P less than 0.01) respectively. Although observed in vitro, a post-antibiotic effect of tobramycin against K. pneumoniae was unlikely in vivo since residual concentrations of tobramycin were found in the lung tissue 12 h after dosing, when efficacy starts to decrease. Efficacy also depended to some extent upon the duration of therapy.

Ecological distribution of Legionellaceae in the Quebec city area.

One hundred environmental water samples, which were collected in the Quebec city area and cultured on buffered charcoal yeast extract medium and three selective media, were inoculated to guinea pigs and were screened by direct immunofluorescent staining (DFA) for the presence of Legionellaceae. Six isolates were made (four Legionella pneumophila and two Tatlockia (Legionella) micdadei: three by animal inoculation and three by culture). No samples were simultaneously positive by both methods. After screening by DFA, 43 of the 100 samples were positive for Legionellaceae and 27 of those contained more than one serogroup and (or) species of Legionellaceae. Legionella pneumophila (serogroups 1 to 6) was the most frequent species seen by DFA. These results clearly show that Legionellaceae are frequent members of the freshwater microbial flora of the Quebec city area.

Comparative study of the beta-lactamase activity found in Achromobacter.

A survey of 21 clinical isolates of Achromobacter species demonstrated a high level of beta-lactamase activity in all strains tested. The beta-lactamases were characterized by isoelectric focusing, purification by affinity chromatography, determination of molecular weight, immunological identity, and genetic analysis. At least three distinct patterns of beta-lactamases were found in 19 strains. The kinetic values Km and Vmax measured by a microacidimetric method showed that all three types of enzymes are cephalosporinases and did not hydrolyse oxacillin, cloxacillin, and methicillin. Two of the three types of cephalosporinases studied, namely MULB 901 (isoelectric point (pI)7.4) and MULB 905(pI 9.3) are enzymes mediated by genes of chromosomal origin. The MULB 906 (pI 8.1) enzyme, however, which has been previously shown to be mediated by an 8.2 MDal nonconjugative plasmid, showed hydrolysis of cefoxitime, cefotaxin, and moxalactam by the bioassay. In all cases, beta-lactamase synthesis appeared constitutive. This study confirms that beta-lactamase activity is commonly found in Achromobacter and that these enzymes are different and of clinical interest when compared with those observed in other Gram-negative bacteria.

A plasmid-mediated cephalosporinase from Achromobacter species.

An unusual cephalosporinase in Achromobacter species was characterized biochemically; the enzyme had a pI of 8.1 and a molecular mass of 36,200 daltons, and it was not inhibited by p-chloromercuribenzoate or cloxacillin. Specific antiserum neutralized enzymatic activity. Agarose gel electrophoresis of the DNA of two strains (MULB 906 and MULB 912) revealed at least three plasmid bands; cured strains demonstrated a simultaneous loss of beta-lactamase and plasmid DNA. Resistance to beta-lactam antibiotics was transferred by transformation of Escherichia coli strain HB101 with plasmid DNA. This plasmid-mediated beta-lactamase differed from the two types of chromosomal cephalosporinases (pI 7.4 and 9.3, respectively) found in a survey of clinical isolates of Achromobacter species. This enzyme also differed in its biochemical properties from all of the other known plasmid-mediated beta-lactamases.

Hydrolysis of Cefotaxime by a beta-lactamase from Bacteroides fragilis.

A beta-lactamase isolated from a strain of Bacteroides fragilis subsp. fragilis possessed hydrolytic activity toward cefotaxime. This antibiotic was degraded to a lower extent than was cephalothin, cephaloridine, and cefamandole, whereas cefoxitin remained unaffected by the enzyme. Kinetic parameters Vmax and Km for cefotaxime were calculated at 0.172 mumol/min and 1.1 X 10(-2) mM, respectively.

Investigation of antibodies against Fasciola hepatica in moose Alces americana.

A serological investigation was carried out to check the presence of antibodies against Fasciola hepatica in a collection of 125 sera moose Alces americana killed in the Province of Quebec. As revealed by hemagglutination tests, two moose had titers suggesting contact with this parasite.

Simultaneous production of two types of beta-lactamase in Escherichia coli and Providencia stuartii.

The production of beta-lactamase has been studied in two strains isolated from clinical samples: Providencia stuartii MULB 501 and Escherichia coli MULB 130. These strains were selected for their high resistance level to penicillins and cephalosporins. Determination and identification of beta-lactamase activity were achieved by combining several up-to-date methods including (i) neutralization by anti-beta-lactamase sera, (ii) purification by affinity chromatography on cephalosporin C linked Sepharose 4B, (iii) determination of substrate specificity and kinetic values (Km and Vmax) by a computerized microacidimetric method, and (iv) isoelectric focusing. The results clearly demonstrate that in these two strains there is a simultaneous production of different beta-lactamases: the first one is similar to the TEM penicillinase and the second one shares a typical cephalosporinase profile. This double beta-lactamase production is a relatively rare phenomenon.

Enzymatic and immunological characterization of a new cephalosporinase from Enterobacter aerogenes.

A hospital strain of Enterobacter aerogenes (MULB 250) isolated from a urinary tract infection was found to be cephalosporin and ampicillin resistant and carbenicillin susceptible. The beta-lactamase produced by this strain was extracted and purified by means of affinity chromatography, using a cephalosporin C-bound Sepharose 4B column. The purified enzyme was tested for hydrolysis of penicillin and various cephalosporins. The K(m) value is 11.8 muM for benzyl penicillin and 130 muM for cephalosporin C. The isoelectric point of the enzyme is 9.3, and its molecular weight is 29,500 +/- 1,000. Rabbit antiserum obtained against this MULB 250 beta-lactamase showed no cross-reaction with other penicillinases or cephalosporinases in neutralization tests. Comparisons of results obtained with other beta-lactamases, particularly from Enterobacter cloacae P99, indicate that the Enterobacter MULB 250 enzyme presents a typical cephalosporinase profile. As far as we know, this type of enzyme is relatively rare.

[Immobilized antibodies: a possible technic for the epidemiological study of antibiotic resistant bacteria]. / Les anticorps immobilisés: un outil possible pour l'étude épidémiologique des bactéries résistantes aux antibiotiques

An antibody specific to a cephalosporinase responsible for the resistance of bacteria to cephalosporines has been immobilized on agarose beads by the iminocarbonate method. The resin thus obtained has been studied at a biospecific point of view. Its high thermostability is also demonstrated. This biospecific resin allows the performance of epidemiologic experiments of bacteria resistant to beta lactamines.

[Purification of beta-lactamases by affinity chromatography]. / Purification des beta-lactamases par chromatographie d'affinité

Affinity columns able to purifie beta-lactamases have been prepared either by linking covalently reversible inhibitors or substrates to agarose beds. The enzyme is eluted with a gradient of sodium chloride or released with the substrate. This method is a pertinent one for the purification of these enzymes and for the study of bacteria harbouring more than one beta-lactamase.