Improvement of an encapsulation process for the preparation of pro- and prebiotics-loaded bioadhesive microparticles by using experimental design.

The purpose of this study was to design a new vaginal bioadhesive delivery system based on pectinate-hyaluronic acid microparticles for probiotics and prebiotics encapsulation. Probiotic strains and prebiotic were selected for their abilities to restore vaginal ecosystem. Microparticles were produced by emulsification/gelation method using calcium as cross-linking agent. In the first step, preliminary experiments were conducted to study the influence of the main formulation and process parameters on the size distribution of unloaded microparticles. Rheological measurements were also performed to investigate the bioadhesive properties of the gels used to obtain the final microparticles. Afterwards an experimental design was performed to determine the operating conditions suitable to obtain bioadhesive microparticles containing probiotics and prebiotics. Experimental design allowed us to define two important parameters during the microencapsulation process: the stirring rate during the emulsification step and the pectin concentration. The final microparticles had a mean diameter of 137µm and allowed a complete release of probiotic strains after 16h in a simulated vaginal fluid at +37°C.

Zidovudine (AZT) has a bactericidal effect on enterobacteria and induces genetic modifications in resistant strains.

The spread of multiresistant bacteria increases the need for new antibiotics. The observation that some nucleoside analogues have antibacterial activity led us to further investigate the antimicrobial activity and resistance of zidovudine (AZT). We determined the minimum inhibition concentration (MIC), studied time-kill curves, induced resistant bacteria and sequenced the gene for thymidine kinase. We demonstrate that AZT has a bactericidal effect on some enterobacteria. However, AZT could induce resistance in Escherichia coli. These resistances were associated with various modifications in the thymidine kinase gene. In particular, we observed the presence in this gene of an insertion sequence (IS) similar to IS911 of Shigella dysenteriae in two resistant clones. No cross-resistance with classical antibiotics in strains with modified thymidine kinase gene was observed. Finally, an additive or synergistic activity between AZT and the two aminoglycoside antibiotics amikacin and gentamicin was observed. We demonstrate the bactericidal activity of AZT and show synergy in association with gentamicin. Genetic modifications in resistant bacteria were identified. Our results indicate that AZT could potentially be added in the treatment of infections with enterobacteria or represent the basis for the development of derivatives with better activity and inducing less resistance.

Reliability of Pseudomonas aeruginosa semi-automated rep-PCR genotyping in various epidemiological situations.

The purpose of this study was to evaluate the possibility of using a semi-automated repetitive DNA sequences-based polymerase chain reaction (rep-PCR) for typing Pseudomonas aeruginosa isolates. rep-PCR profiles obtained by the DiversiLab system of 84 P. aeruginosa isolates from distinct epidemiological situations were obtained. rep-PCR groupings were in good agreement with the origin of these isolates. Linked rep-PCR profiles were observed for isolates recovered from a same family of cystic fibrosis (CF) patients, for the etiological agents of clustered cases of nosocomial infections, and for some isolates recovered from a same hospital room. rep-PCR and pulsed-field gel electrophoresis SpeI restricted genomic DNA (PFGE-SpeI) profiles were compared. In a few instances, rep-PCR revealed genetic divergences among isolates of a same group of PFGE-SpeI profiles. These divergences could reflect genetic drifts among closely related isolates, as illustrated by those observed between clinical and environmental isolates of a same group of PFGE-SpeI profiles. The interpretation of such differences will require further studies, but the rep-PCR analysis of P. aeruginosa diversity appeared to be an appropriate method to investigate infra-specific genetic relatedness.

Spatio-temporal analysis of infra-specific genetic variations among a Pseudomonas aeruginosa water network hospital population: invasion and selection of clonal complexes.

AIMS: To investigate infra-specific spatio-temporal dynamics of a hospital water network Pseudomonas aeruginosa population. To infer the origin of water network isolates and assess their potential health hazard. METHODS AND RESULTS: 168 P. aeruginosa strains were isolated from tap waters and swabs of tap nozzle aerators of a hospital unit, over 2 years, and from rectal swabs and nosocomial infections. Genetic diversity among this collection was assessed by pulsed field gel electrophoresis of SpeI restricted genomic DNA. Virulence gene sets, biofilm properties, and hypochlorite resistance were analysed. Exactly 68% of the water samples and 74% of the tap nozzle aerators harboured P. aeruginosa. The strains were divided into 22 clonal lineages, with one dominant clone shown to have been involved in a nosocomial infection. CONCLUSIONS: An important turnover among the P. aeruginosa hospital population was observed. Some clonal lineages were found to persist, spread in the unit, and diversify into clonal complexes. Rectal carriage appeared an important source of contamination of the water network. SIGNIFICANCE AND IMPACT OF THE STUDY: High P. aeruginosa infra-specific population diversity suggested a broad ability in colonizing water networks but persistence analysis indicated a strong selection leading to the emergence of dominant clones.

High prevalence of nosocomial infections in rehabilitation units accounted for by urinary tract infections in patients with spinal cord injury.

This study (part of the nationwide French prevalence survey of 2001) was organized to investigate the prevalence and risk factors of nosocomial infections (NIs) and the resistant flora in patients hospitalized in rehabilitation units. Two hundred and eighty-six patients were included from two hospitals in the 'Hospices Civils de Lyon' group. Patients were classified into those with and without a spinal cord injury (SCI). Seventy-eight (27.3%) patients had an SCI. They were younger and more often characterized by a low Activity of Daily Life score, bladder incontinence and chronic respiratory disease. Urinary catheterization and mechanical ventilation were more common in these patients. The NI prevalence rate was higher in the SCI group (21.8% vs 4.3%, P<0.00001), particularly for urinary tract infections (UTIs, 19.2% vs 3.4%, P<0.00001). There was a positive relationship between the number of risk factors and NI acquisition. Multi-variate analysis showed that the only independent risk factor for NI acquisition was indwelling urinary catheterization [odds ratio (OR): 11.64, 95% confidence intervals (CI): 2.53-53.65, P=0.002]. Marginally significant factors were chronic kidney or liver disease (OR: 5.84, 95%CI: 0.80-42.68, P=0.082) and SCI (OR: 2.97, 95%CI: 0.61-14.60, P=0.179). The prevalence of antibiotic-resistant micro-organisms was high (nine cases of resistant organisms for 31 infection sites), but there were no differences between the groups. The high rate of NIs, especially UTIs, in SCI patients was not due to an independent effect of SCI but was probably due to the high number of risk factors. These high-risk patients need targeted NI surveillance.

Comparison of the new API Candida system to the ID 32C system for identification of clinically important yeast species.

API Candida was evaluated in comparison with the ID 32C system for the identification of 619 yeast isolates. The sensitivity of API Candida for the identification of the 15 species it claims to identify with and without additional tests was 97.4% (593 of 609) and 75.2% (458 of 609), respectively. The API Candida system is easy to use and rapid (result in 18 to 24 h).

Evaluation of CPS ID2 medium for detection of urinary tract bacterial isolates in specimens from a rehabilitation center.

CPS ID2 medium (bioMérieux) enables the presumptive identification of Escherichia coli and enterococci as well as the detection of indologenous or nonindologenous Proteeae and bacteria belonging to the Klebsiella, Enterobacter, and Serratia group with a specificity ranging from 98 to 100%. When the cultures were polymicrobial, the sensitivity varied from 70 to 97%, and the sensitivity varied from 97 to 100% when they were monomicrobial.

[In vitro comparative study of the sensitivity of Aspergillus to antifungal agents]. / Etude comparative in vitro de la sensibilité des Aspergillus aux antifongiques.

Minimum inhibitory concentrations (MIC) of amphotericin B, 5-fluorocytosine, miconazole, tioconazole, econazole and ketoconazole were determined for 310 Aspergillus strains belonging to four different species isolated from clinical specimens. Econazole exhibited the best in vitro activity with MIC less than or equal to 3.12 micrograms/ml for 96% of strains and less than or equal to 1.56 micrograms/ml for 68%. For amphotericin B, 69% of strains were less than or equal to 1.56 micrograms/ml and, for 5-fluorocytosine, 50% were less than or equal to 25 micrograms/ml. Miconazole and ticonazole exhibited comparable in vitro activities with MIC less than or equal to 6.25 micrograms/ml for 66% and 58% of strains respectively. Ketoconazole was the less active agent in vitro, with MIC less than or equal to 6.25 micrograms/ml for only 27% of strains. Moreover, this study demonstrated significant differences in susceptibility from species to species. A. fumigatus was susceptible to amphotericin B (75%) and 47% of strains were susceptible to 5-fluorocytosine. A. niger showed high susceptibility to amphotericin B (82%) and 5-fluorocytosine (82%). A. flavus exhibited the highest resistance to 5-fluorocytosine (14%), the highest susceptibility to ketoconazole (72%), and average susceptibility to amphotericin B (41%). A. nidulans was rarely susceptible to amphotericin B (21%) or 5-fluorocytosine (29%) and 46% of strains were susceptible to miconazole and ketoconazole. These results provide useful guidelines for the choice of antifungal agents in the difficult treatment of aspergillosis.

Laser nephelometric semi-automated system for rapid bacterial susceptibility testing.

Experiments with a semi-automated susceptibility test system are described. The system was based on a laser nephelometer which was used to estimate extinction in broth cultures, and ready-to-use cuvettes containing antibiotics. Each antibiotic was tested at two concentrations, the cuvettes were manually inoculated and the results were obtained after 3-5 h incubation in an agitator-incubator. The system was used to test 70 strains (mainly Gram-negative) for susceptibility to 14 antibiotics. The results were compared with the minimal inhibitory concentration (MIC) determined in a solid medium and partially with the ABAC semi-automatic antibiotic sensitivity test. No major discrepancy was observed between the semi-automated laser nephelometric system and MIC and the results were in complete agreement in 91.2%. The laser nephelometric system was found slightly superior to the ABAC system which showed a full accord rate of 88% with MIC. Since the method used a common apparatus in protein immunochemistry, it was found applicable in routine diagnostic microbiology laboratories.

Serological grouping of streptococci: a collective evaluation in six laboratories of three rapid methods.

Three serological methods for streptococci grouping: latex agglutination after pronase extraction (LAP), latex agglutination after nitrous acid extraction (LAN) and counterimmunoelectrophoresis after nitrous acid extraction (CIEN) were evaluated in six laboratories with 310 strains of streptococci. To reduce the possible influence of a particular laboratory, each serological procedure was performed in two different laboratories with identical protocols and strains. Between the three procedures, a total agreement of 81.3% with six identical results was obtained. Since only one discordant result was obtained in 12.2% (CIEN 4.8%-LAN 5.8%-LAP 1.6%) and two discordant results with two different serological methods in 2.6%, it could be concluded that agreement between the three procedures was 96%. For B (99 strains tested) and D(100 strains tested) groups, total agreements of respectively 95% and 92% were achieved. In conclusion, LAP, LAN, and CIEN gave equivalent results for streptococci grouping, but LAN was the most simple needing no heating, centrifugation, or electrophoresis equipment and lacking cross-reactivity of group C antibody.