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The viable but non-culturable (VBNC) stage of Helicobacter pylori may represent a problem of public health concern, since these cells cannot be detected by traditional culture methods. In this study, the direct viable count method (DVC) was modified and adapted to H. pylori analysis by testing different times of incubation and concentrations of DNA-gyrase inhibitors. The DVC procedure was combined with fluorescent in situ hybridization (FISH) for the specific detection of viable cells of H. pylori (DVC-FISH). Incubation with 0.5 microg/ml of novobiocin for 24 h provided the optimal conditions for obtaining 3-5 times the original size of Helicobacter viable cells. Field work performed with various types of water (freshwater and seawater) using the DVC-FISH approach enabled us to confirm the presence of VBNC H. pylori cells in 16 of the 45 analyzed samples. The combination of the modified DVC procedure with FISH can provide a rapid and specific method to detect and identify viable cells of H. pylori in environmental samples.

Background : Patients with the obesity-hypoventilation syndrome (OHS) benefit from non-invasive ventilatory support. We assessed the long-term physiopathological response to 12-months of nocturnal ventilatory assistance at home with bi-level positive airway pressure (BiPAP) via nasal mask in patients with this disease. Methods : A series of 13 non-consecutive patients diagnosed of OHS (5 men and 8 women) with a mean (sd) age of 61.9 (8) years, underwent the following studies before (baseline) and after 12 months of non-invasive domiciliary mechanical ventilation: arterial blood gases, nocturnal digital pulse oximetry, spirometry, body plethysmography, maximum muscular respiratory pressures and ventilatory pattern with measurement of occlusion pressure ( P 0.1) before and after hypercapnia. An overnight cardiorespiratory polygraphy was done at baseline. Results : After 12 months of non-invasive mechanical ventilation, there were significant ( P P aCO 2 ) and increases in arterial oxygen tension, forced vital capacity and ventilatory response to hypercapnia, as measured by the relationship between changes of P 0.1 and P aCO 2 (Δ P 0.1/Δ P aCO 2 ) as well as respiratory minute volume ( V E ) and P aCO 2 (Δ V E /Δ P aCO 2 ). Changes of the P aCO 2 correlated significantly with those in the (Δ P 0.1/Δ P aCO 2 ) slope ( r =0.576, P Conclusions : The present results confirm that non-invasive home mechanical ventilation is an effective approach for long-term treatment of OHS.

The objective of the present trial was to compare the efficacy, safety, and tolerability of moxifloxacin (400 mg) given intravenously (i.v.) once daily followed by oral moxifloxacin (400 mg) for 7 to 14 days with the efficacy, safety, and tolerability of co-amoxiclav (1.2 g) administered by i.v. infusion three times a day followed by oral co-amoxiclav (625 mg) three times a day, with or without clarithromycin (500 mg) twice daily (i.v. or orally), for 7 to 14 days in adult patients with community-acquired pneumonia requiring initial parenteral therapy. A total of 628 patients were enrolled and assessed by evaluation of their clinical and bacteriological responses 5 to 7 days and 21 to 28 days after administration of the last dose of study medication. Although the trial was designed, on the basis of predefined outcomes, to demonstrate the equivalence of the two regimens, the results showed statistically significant higher clinical success rates (for moxifloxacin, 93.4%, and for comparator regimen, 85.4%; difference [Δ], 8.05%; 95% confidence interval [CI], 2.91 to 13.19%; P = 0.004) and bacteriological success rates (for moxifloxacin, 93.7%, and for comparator regimen, 81.7%; Δ, 12.06%; 95% CI, 1.21 to 22.91%) for patients treated with moxifloxacin. This superiority was seen irrespective of the severity of the pneumonia and whether or not the combination therapy included a macrolide. The time to resolution of fever was also statistically significantly faster for patients who received moxifloxacin (median time, 2 versus 3 days), and the duration of hospital admission was approximately 1 day less for patients who received moxifloxacin. The treatment was converted to oral therapy immediately after the initial mandatory 3-day period of i.v. administration for a larger proportion of patients in the moxifloxacin group than patients in the comparator group (151 [50.2%] versus 57 [17.8%] patients). There were fewer deaths (9 [3.0%] versus 17 [5.3%]) and fewer serious adverse events (38 [12.6%] versus 53 [16.5%]) in the moxifloxacin group than in the comparator group. The rates of drug-related adverse events were comparable in both groups (38.9% in each treatment group). The overall incidence of laboratory abnormalities was similar in both groups. Thus, it is concluded that monotherapy with moxifloxacin is superior to that with a standard combination regimen of a β-lactam and a β-lactamase inhibitor, co-amoxiclav, with or without a macrolide, clarithromycin, in the treatment of patients with community-acquired pneumonia admitted to a hospital.

We developed a double-staining procedure involving NanoOrange dye (Molecular Probes, Eugene, Oreg.) and membrane integrity stains (LIVE/DEAD Bac Light kit; Molecular Probes) to show the morphological and membrane integrity changes of Campylobacter coli cells during growth. The conversion from a spiral to a coccoid morphology via intermediary forms and the membrane integrity changes of the C. coli cells can be detected with the double-staining procedure. Our data indicate that young or actively growing cells are mainly spiral shaped (green-stained cells), but older cells undergo a degenerative change to coccoid forms (red-stained cells). Club-shaped transition cell forms were observed with NanoOrange stain. Chlorinated drinking water affected the viability but not the morphology of C. coli cells.

The onset of fatal and near-fatal asthma attacks can be rapid. The objective of this study was to determine the frequency and clinical characteristics of rapid-onset asthma (ROA) in patients suffering fatal and near-fatal crises. Two-hundred and twenty patients with fatal or near-fatal attacks were enrolled in a multicentre, prospective study. ROA was defined as a crisis developing in ≤2 h. Data on patient and clinical characteristics were collected, and spirometric and allergy studies were performed when the patients were in a stable condition. Forty-five attacks (20%) were ROA and 175 (80%) were slow-onset asthma (SOA). The triggers for SOA and ROA attacks were different, with the ROA group having a significantly lower rate of suspected respiratory infection (7% versus 38%), higher rates of fume/irritant inhalation (9% versus 1%) and a higher intake of nonsteroidal anti-inflammatory drugs (14% versus 3%). The ROA group exhibited significantly higher rates of impaired consciousness (63% versus 44%), absence of lung sounds upon admission (68% versus 42%), fewer hours of mechanical ventilation (13 h versus 28 h) and fewer days of hospitalization (8 days versus 9.5 days) than the SOA group. The 20% frequency of rapid-onset fatal and near-fatal attacks in this study suggests that rapidly developing attacks may not be rare. These findings also support a distinct clinical profile for rapid-onset asthma marked by differences in triggers, severity of exacerbation and clinical course.

We have evaluated the use of PCR and fluorescent in situ hybridization (FISH) techniques for the detection of thermotolerant campylobacters in naturally contaminated chicken products. 16S rRNA sequence data was used to design two specific primers and an oligonucleotide probe for PCR and FISH analyses, respectively. The PCR protocol amplified a 439-bp fragment corresponding to a portion of specific 16S RNA gene from thermotolerant campylobacters. The detection range of the PCR assay varied between 10 cells (after enrichment) to 10(2) cells per mL (without enrichment). FISH probes were able to identify thermotolerant Campylobacter species in 'spiked' and 'unspiked' naturally contaminated samples. PCR and FISH were performed on naturally contaminated samples and compared with the isolation of cells on selective media. The in situ hybridization technique was less sensitive than PCR, although its sensitivity of detection was increased considerably after 22 h of enrichment. These results confirm the usefulness of 16S rRNA-based techniques for the direct detection of campylobacters in food samples.

This study compared the performance of LMX(R) broth (LMX), Chromocult Coliform(R) agar (CC) and Chromocult Coliform agar plus cefsulodin (10 microg ml-1) (CC-CFS), with standard methods multiple tube fermentation (MTF), for the enumeration of total coliforms and Escherichia coli from marine recreational waters. LMX and CC are two media designed to concurrently detect total coliform (TC) bacteria and E. coli by the specific action of beta-galactosidase (total coliforms) and beta-glucuronidase (E. coli). Overall results for the TC test showed that LMX, CC and MTF recovered 2.63, 1.95 and 1.90 times as many TCs as CC-CFS, respectively. Data from the multiple range test showed significant differences (P < 0.05) between TC counts on CC-CFS and LMX. The traditional MTF was less sensitive for E. coli enumeration. However, there was no statistically significant differences between LMX, CC, CC-CFS and the MTF method for E. coli enumeration. Background interference was reduced on CC-CFS and the counts obtained reflected more accurately the number of TCs. Therefore, the contribution of beta-galactosidase positive, non coliform bacteria (Aeromonas spp. and Vibrio spp.) to TC counts should not be neglected.

Cefsulodin was evaluated as a potential selective agent for aeromonads. Resistance of Aeromonas and coliform isolates was determined by using a standard disk diffusion technique. A total of 119 Aeromonas and 78 coliform strains were isolated. For 102 of 130 [corrected] Aeromonas isolates (environmental and reference strains), the MIC of cefsulodin was < 8 micrograms/ml. Results of MIC tests by the agar dilution method showed that a concentration of cefsulodin of 10 micrograms/ml or less inhibited the growth of 96% of isolates. In comparison, for 81 of 94 coliform isolates (environmental and reference strains), the MIC of cefsulodin was > 32 micrograms/ml. Because cefsulodin suppresses growth of Aeromonas and other oxidase-positive organisms, total coliform (TC) and Escherichia coli counts on Chromocult Coliform agar (CC agar) without cefsulodin and on CC agar with 10 mg of cefsulodin per liter (CC-CFS) were compared. Variance analysis of data from 14 sewage-polluted irrigation water specimens did not demonstrate any statistically significant difference in the enumeration of E. coli with CC and CC-CFS media. On average, the CC agar recovered 2.46 times as many TCs as CC-CFS. However, Aeromonas colonies made up an average of 58.6% of the TC counts on CC agar. Because no Aeromonas spp. were recovered on CC-CFS, background interference was eliminated and the counts that were obtained reflected more accurately the number of TCs. Results of this study suggest that cefsulodin may be a useful selective agent against Aeromonas spp. which should be included in coliform chromogenic media when high levels of accompanying flora are expected.

Thirty-two strains of thermophilic campylobacters isolated from marine recreational waters and seven reference strains were biotyped and analysed by chromosomal DNA Hae III ribopatterns and AP-PCR profiles based on a random 10-mer primer (5′-CAA TCG CCG T-3′). The majority of seawater isolates (90%) were Campylobacter coli, and three strains were Camp. jejuni. Southern blot hybridization analysis showed differences between the strains, and in a numerical analysis three main clusters were formed at the 45% similarity level, that corresponded to Camp. jejuni subsp. jejuni, Camp. coli, and a combination of Camp. coli and Camp. jejuni subsp. doylei. AP-PCR profiles also differentiated between the species but were less discriminatory than ribotyping because six strains (17%) could not be typed by this method. Numerical analysis gave four main clusters at the 45% similarity level, corresponding to Camp. jejuni subsp. jejuni, Camp. coli (two clusters) and Camp. lari. The study shows that strains within each species are diverse genomically. Both molecular methods were highly discriminatory, although some strains with identical ribotypes could be distinguished by AP-PCR, and they are valuable new alternatives to traditional typing in epidemiological studies of environmental campylobacters.

Two extraction procedures were examined, and it was found that DNA recovered from Campylobacter jejuni lysed by the cetyltrimethylammonium bromide (CTAB) method was more suitable for use as a PCR template than DNA released by the boiling method. The region targeted for PCR amplification was a 1.73-kb portion of the flagellin A gene of C. jejuni. The detection limit was lower than 30 cells per 100 ml in artificially contaminated waters. PCR assay and conventional culturing method had the same sensitivity, but results of the PCR technique were available within 48 h and so shortened the time necessary for detection by 48 h.

The performance of aeration, photodecomposition and biological degradation processes as methods to reduce molinate contamination levels in effluent water from rice fields was studied. Aeration produced a molinate dissipation of 84%, as against 22% without aeration. Application of UV-light to clean water solutions achieved a molinate photodecomposition of 96% in 24 h. Maximal degradation obtained in algal cultures was 55% in 20 days and 78% in 40 days. In micro-organism cultures, kept in darkness and with a continuous flow of aqueous solution of molinate and inorganic salts, a degradation of 97% was achieved.

This study compared the performance of a commercial chromogenic medium, CHROMagarECC (CECC), and CECC supplemented with sodium pyruvate (CECCP) with the membrane filtration lauryl sulfate-based medium (mLSA) for enumeration of Escherichia coli and non- E. coli thermotolerant coliforms (KEC). To establish that we could recover the maximum KEC and E. coli population, we compared two incubation temperature regimens, 41 and 44.5°C. Statistical analysis by the Fisher test of data did not demonstrate any statistically significant differences ( P = 0.05) in the enumeration of E. coli for the different media (CECC and CECCP) and incubation temperatures. Variance analysis of data performed on KEC counts showed significant differences ( P = 0.01) between KEC counts at 41 and 44.5°C on both CECC and CECCP. Analysis of variance demonstrated statistically significant differences ( P = 0.05) in the enumeration of total thermotolerant coliforms (TTCs) on CECC and CECCP compared with mLSA. Target colonies were confirmed to be E. coli at a rate of 91.5% and KEC of likely fecal origin at a rate of 77.4% when using CECCP incubated at 41°C. The results of this study showed that CECCP agar incubated at 41°C is efficient for the simultaneous enumeration of E. coli and KEC from river and marine waters.

Introduction Asthma Control Questionnaire (ACQ) is a validated tool to measure asthma control. Cut-off points that best discriminate “well-controlled” or “not well-controlled” asthma have been suggested from the analysis of a large randomized clinical trial but they may not be adequate for daily clinical practice. Aims To establish cut-off points of the ACQ that best discriminate the level of control according to Global Initiative for Asthma (GINA) 2006 guidelines in patients with asthma managed at Allergology and Pulmonology Departments as well as Primary Care Centers in Spain. Patients and methods An epidemiological descriptive study, with prospective data collection. Asthma control following GINA-2006 classification and 7-item ACQ was assessed. The study population was split in two parts: 2/3 for finding the cut-off points (development population) and 1/3 for validating the results (validation population). Results A total of 1,363 stable asthmatic patients were included (mean age 38 ± 14 years, 60.3% women; 69.1% non-smokers). Patient classification according to GINA-defined asthma control was: controlled 13.6%, partially controlled 34.2%, and uncontrolled 52.3%. The ACQ cut-off points that better agreed with GINA-defined asthma control categories were calculated using receiver operating curves (ROC). The analysis showed that ACQ Conclusion The ACQ cut-off points associated with GINA-defined asthma control in a real-life setting were