Microarray analysis of Escherichia coli strains from interstitial beach waters of Lake Huron (Canada).

DNA microarray analyses revealed that clusters of repetitive extragenic palindromic PCR-related Escherichia coli isolates were isogenic only within interstitial Lake Huron beach water samples and not in surrounding waters. This suggested that adaptation and growth occurred within the interstitial water sites tested. All isolates were nonpathogenic, and three lake isolates possessed tetracycline resistance genes.

Decrease in Cryptosporidium parvum oocyst infectivity in vitro by using the membrane filter dissolution method for recovering oocysts from water samples.

Exposure of Cryptosporidium parvum oocysts to solutions used for cellulose acetate membrane (CAM) dissolution filtration reduced their infectivity in HCT-8 cells. Ethanol (95% [vol/vol] and 70% [vol/vol]) alone and short exposure times to acetone decreased infectivity. These findings contrast with similar experiments using excystation assays and infectivity in mice.

Improving the rate of infectivity of Cryptosporidium parvum oocysts in cell culture using centrifugation.

Centrifugation was evaluated as a method to improve infectivity assays of Cryptosporidium parvum in cell culture using the focus detection method, an immunofluorescence-based method for detecting infectious C. parvum oocysts in vitro. Human ileocecal adenocarcinoma (HCT-8) cells were grown for 48 hr on 13-mm cover slips in 24-well microtiter plates and infected with bleach-treated C. parvum oocysts. Plates were centrifuged at 228 g for 10 min and incubated at 37 C for 5, 12, 18, 24, and 48 hr. Foci of infection were stained by immunofluorescence and enumerated using epifluorescent microscopy. Results were compared to noncentrifuged controls. Foci in centrifuged samples could be enumerated after 18 hr. According to most probable number (MPN) analysis, the number of infectious oocysts estimated at 48 hr (13,326 infectious oocysts) was reached by 18 hr in centrifuged samples. After 48 hr, there was no significant difference (P < 0.05) between centrifuged and noncentrifuged samples enumerated by number of foci or the MPN of infectious oocysts. Centrifugation may expedite detection during C. parvum infectivity assays. Furthermore, multiwell plate formats are more cost effective than traditional chamber slides.

An uncommon Helicobacter isolate from blood: evidence of a group of Helicobacter spp. pathogenic in AIDS patients.

An unusual Helicobacter sp. was isolated from the blood of a human immunodeficiency virus (HIV)-infected patient. This organism had spiral morphology, with single amphitrichous flagella, and was negative for hippurate hydrolysis, production of urease, and reduction of nitrate. 16S rRNA gene sequence analysis verified that the isolate was a species of Helicobacter, most closely related to an undescribed Helicobacter-like isolate from Vancouver, British Columbia, Canada, and to Helicobacter westmeadii, a recently described species from Australia. Both organisms had also been isolated from the blood of HIV-infected patients. These blood isolates, along with Helicobacter cinaedi, form a cluster of closely related Helicobacter spp. that may represent an emerging group of pathogens in immunocompromised patients.

Detection of Legionella by PCR in respiratory specimens using a commercially available kit.

Using polymerase chain reaction (PCR) for the detection of pathogens that are difficult to grow, such as Legionella species, may reduce difficulties encountered with culture and immunofluorescent staining. We evaluated a commercial PCR and hybridization kit, designed for environmental samples, for the detection of Legionella in respiratory specimens. Sixteen Legionella species cultures tested positive with the Perkin Elmer Legionella EnviroAmp Amplification and Detection kits (Perkin Elmer, Foster City, Calif). The assay detected as few as 100 colony-forming units per milliliter of spiked bronchoalveolar lavage (BAL) fluid, and no false-negative results were obtained. PCR inhibition by blood in the specimens was removed by washing pelleted specimens in sterile distilled water. Of 126 specimens screened with the kit, 1 induced sputum and 3 BAL specimens were positive by PCR. All 4 were validated as true-positive results by culture or serologic testing. The entire PCR and hybridization assay can be completed in less than 6 hours, whereas isolation and identification by culture requires up to 12 days, and serologic conversion may not be demonstrated for weeks. Molecular techniques based on direct extraction and amplification of DNA from respiratory specimens nay be useful for the timely diagnosis of legionellosis.

Nutrient-enhanced survival of and phenanthrene mineralization by alginate-encapsulated and free Pseudomonas sp. UG14Lr cells in creosote-contaminated soil slurries.

The effects of nutrient amendment and alginate encapsulation on survival of and phenanthrene mineralization by the bioluminescent Pseudomonas sp. UG14Lr in creosote-contaminated soil slurries were examined. UG14Lr was inoculated into creosote-contaminated soil slurries either as a free cell suspension or encapsulated in alginate beads prepared with montmorillonite clay and skim milk. Additional treatments were free-cell-inoculated slurries amended with sterile alginate beads, free-cell-inoculated and uninoculated slurries amended with skim milk only, and uninoculated, unamended slurries. Mineralization was determined by measuring 14CO2 released from radiolabelled phenanthrene. Survival was measured by selective plating and bioluminescence. Inclusion of skim milk was found to enhance both survival of and phenanthrene mineralization by free and encapsulated UG14Lr cells.

Survival and respiratory activity of genetically engineered Pseudomonas spp. exposed to antimicrobial agents in broth and soil.

The effectiveness of seven chemical disinfectants were tested against genetically engineered Pseudomonas spp. under optimal growth conditions. Each chemical was tested to determine how quickly 10(8) cells/ml Pseudomonas fluorescens C5t (containing Bacillus thuringiensis endotoxin gene) were killed in King's B broth at 30 degrees C. The minimal bactericidal concentrations (MBC) for calcium hypochlorite, benzalkonium chloride, Germiphene and Spectrum Clear Bath were 0.06% (w/v), 0.01% (w/v), 0.08% (v/v) and 0.044 (v/v), respectively. Virocidin X and CanLab Neutral did not kill P. fluorescens C5t at concentrations up to 20% (v/v). Baxter Bacdown was ineffective as a killing agent at concentrations up to 10% (v/v). These agents were also tested on P. aureofaciens RNL11 (lacZY), P. putida strains PaW8 and PaW340 (containing the plasmid pLV1013 encoding the xylE gene) and P. aeruginosa UG2L (lacZY and luxAB). All strains were killed by similar disinfectant concentrations with the exception of P. aeruginosa UG2L. The MBCs for this strain were double the MBCs for P. fluorescens C5t with all disinfectants used except for Spectrum Clear Bath, which was similar. Oxygen consumption and carbon dioxide evolution were used to assess the activity of P. fluorescens C5t inoculated in non-sterile soil in the presence of antimicrobial agents. Concentrations chosen were 0.1 and 1.0% (w/v) calcium hypochlorite, 0.01 and 0.1% (w/v) benzalkonium chloride, 0.1 and 1.0% (v/v) Germiphene and 0.1 and 1.0% (v/v) Spectrum Clear Bath. Over a 15-day period, the antimicrobial agents did not reduce respiratory activity of P. fluorescens C5t and indigenous soil micro-organisms.(ABSTRACT TRUNCATED AT 250 WORDS)