Pulmonary tuberculosis due to Mycobacterium microti: a study of six recent cases in France.

Human tuberculosis caused by Mycobacterium microti is rare, but its prevalence and clinical significance may have been underestimated. To the best of our knowledge, 21 cases have been reported in the literature in the last decade. We report six recent pulmonary cases caused by M. microti over a period of 5 years detected in French clinical mycobacteriology laboratories of the hospital network. Our data confirm the potential of M. microti to cause clinical illness in immunocompetent patients. M. microti grew slowly from specimens, delaying the final microbiological diagnosis. Therefore, patients with tuberculosis caused by M. microti could benefit from the use of rapid diagnostic molecular techniques directly on clinical samples. From a review of the literature and this study, a classical antituberculous therapy seems effective in treating patients with M. microti disease.

[Infection due to Mycobacterium malmoense in an immunocompetent patient]. / Infection à Mycobacterium malmoense chez un patient immunocompétent.

INTRODUCTION: Mycobacterium malmoense (MM) is an atypical mycobacterium responsible for opportunistic infection. The clinical and radiological picture is non-specific. The infection develops most frequently in a dystrophic lung. CASE REPORT: A patient of 52 years was admitted with an extensive multifocal pneumonia which later proved to be due to infection with MM. Empirical treatment was started with the combination of rifampicin, isoniazid, pyrazinamide (rifater) and ethambutol (myambutol). Subsequently, cultures showed sensitivity to rifampicin, ethambutol, oxfloxacin, clarithromycin (MIC < 2 mg/l) and rifabutin (MIC < 0.5 mg/l). More than two weeks after the start of treatment, material aspirated at fibroscopy showed the persistence of numerous acid-alcohol fast bacilli, an increase, compared with the original examination, from 5 to 25 per field on day 2, to 20 to 100 per field on day 19. Despite the late addition of clarithromycin there was a progressive deterioration in the pulmonary condition. CONCLUSION: There is little correlation between the in vivo and in vitro sensitivities of MM to antibiotics. In our patient the progress was unfavourable, even though the mycobacterium was sensitive to the combination of antibiotics used, with the exception of isoniazid that was not tested. In vitro isoniazid does not seem to be active against MM. There is no consensus of opinion on the antibiotic treatment of MM infections.

Determination of new types of DNA lesions in human sperm.

Careful attention has been focused recently on DNA quality in human IVF. Therefore a variety of methods has been developed to evaluate DNA integrity, especially concerning fragmentation. Using liquid chromatography and mass spectrometry (LC/MS/MS) for our best sperm samples, we have established reference values for several oxidative lesions, in order to gain insights into the cause of DNA lesions. Besides 8-oxodeoxyguanosine, we found rather high levels of two ethenonucleosides: 1,N6-ethenoadenosine and 1,N2-ethenoguanosine. These compounds probably arise from a reaction with 4-hydroxy-2-nonenal, the main aldehyde compound released during lipid peroxidation, or after occupational exposure to vinyl chloride. The quantity of chlorinated bases detected is low. All of this decay has to be repaired by the oocytes at the time of fertilization or immediately after. This aspect should not be overlooked in assisted reproductive technology, in order to understand risks and limitations.

Surgery combined with peritonectomy procedures and intraperitoneal chemohyperthermia in abdominal cancers with peritoneal carcinomatosis: a phase II study.

PURPOSE: To evaluate the tolerance of peritonectomy procedures (PP) combined with intraperitoneal chemohyperthermia (IPCH) in patients with peritoneal carcinomatosis (PC), a phase II study was carried out from January 1998 to September 2001. PATIENTS AND METHODS: Fifty-six patients (35 females, mean age 49.3) were included for PC from colorectal cancer (26 patients), ovarian cancer (seven patients), gastric cancer (six patients), peritoneal mesothelioma (five patients), pseudomyxoma peritonei (seven patients), and miscellaneous reasons (five patients). Surgeries were performed mainly on advanced patients (40 patients stages 3 and 4 and 16 patients stages 2 and 1) and were synchronous in 36 patients. All patients underwent surgical resection of their primary tumor with PP and IPCH (with mitomycin C, cisplatinum, or both) with a closed sterile circuit and inflow temperatures ranging from 46 degrees to 48 degrees C. Three patients were included twice. RESULTS: A macroscopic complete resection was performed in 27 cases. The mortality and morbidity rates were one of 56 and 16 of 56, respectively. The 2-year survival rate was 79.0% for patients with macroscopic complete resection and 44.7% for patients without macroscopic complete resection (P =.001). For the patients included twice, two are alive without evidence of disease, 54 and 47 months after the first procedure. CONCLUSION: IPCH and PP are able to achieve unexpected long-term survival in patients with bulky PC. However, one must be careful when selecting the patients for such an aggressive treatment, as morbidity rate remains high even for an experienced team.

[Development of a method for determining lead blood levels with chronopotentiometry]. / Développement d'une méthode de dosage de la plombémie par chronopotentiométrie.

A potentiometric stripping analysis method for blood lead determination, using Radiometer Tracelab PSU 22, has been investigated. Hemoglobin precipitation by hydrochloric and perchloric acids mixture allows to obtain the same signal for the same lead concentrations in aqueous standards and various blood samples. Thus it is possible to use a time saving calibration procedure. Equilibration time increase improved method sensitivity and precision. The analysis time was 3 minutes per sample. The detection limit was 0.006 microM/L. The relative standard deviation was 3.6% for 1.351 microM/L and 6% for 0.203 microM/L. The results of 66 determinations obtained by this method and by graphite furnace atomic absorption spectrometry were compared (R=0.991). Potentiometric stripping analysis is less expensive and very convenient for clinical laboratories.

Population pharmacokinetics of cisplatin in patients with advanced ovarian cancer during intraperitoneal hyperthermia chemotherapy.

The objective of this study was to determine the pharmacokinetic profile of total platinum administered by hyperthermic peritoneal perfusion (HPP) in 16 patients with ovarian cancer. The patients had a performance status of lIIalpha/b/c on the FIGO scale. They received 60, 80 or 100 mg of cisplatin. The percent of cisplatin remaining in the body after the peritoneum was emptied averaged 65% (41.7-85.4%). The average ratio between peritoneal drug concentrations and plasma concentrations was 73. A Bayesian estimation of individual phamacokinetic parameters was carried out using the non-linear mixed-effect modeling approach as implemented in the NONMEM computer program. A two-compartment model with an additional peritoneal cavity compartment was used to fit the data. Large interindividual variability of the pharmacokinetic parameters occurred The maximum platinum concentration in plasma was reached between 1 and 1.5 hours after the beginning of administration; it ranged from 0.37 to 1.7 microg/ml (1.9 to 8.72 microM). The elimination half-life was 80 hours (48-152 hours and the area under the plasma concentration time curve normalized to a 100 mg cisplatin dose was 79 mg/liter x hours. The simultaneous fit of perfusate and plasma concentrations allowed us to estimate the percent of cisplatin that reached the systemic circulation at about 20%. At time infinity, the urinary cisplatin recovery accounted for only 20% of the administered dose. The results in this study showed that a high proportion of the cisplatin dose was absorbed by target tumor cells. In spite of the advanced disease of patients at the time of HPP, 37.5% of them were still alive three years after HPP (ie., 3-6 years after cancer diagnosis) and 12.5%, 7 years after HPP (i.e., 8 years after cancer diagnosis).

Mutations in the rpoB gene of rifampicin-resistant Mycobacterium tuberculosis strains isolated in Brazil and France.

We evaluated the mutations in a 193bp of the rpoB gene by automated sequencing of rifampicin (RMP)-resistant and susceptible Mycobacterium tuberculosis strains isolated from Brazil (25 strains) and France (37 strains). In RMP-resistant strains, mutations were identified in 100% (16/16) from France and 89% (16/18) from Brazil. No mutation was detected in the 28 RMP-susceptible strains. Among RMP-resistant or RMP-susceptible strains deletion was observed. A double point mutation which had not been reported before was detected in one strain from France. Among French resistant strains mutations were found in codons 531 (31.2%), 526, 513 and 533 (18.7% each). In Brazilian strains the most common mutations were in codons 531 (72.2%), 526 (11.1%) and 513 (5.5%). The heterogeneity found in French strains may be related to the fact that most of those strains were from African or Asian patients.

Mutations in the rpoB gene of rifampicin-resistant Mycobacterium tuberculosis strains isolated in Brazil and France

We evaluated the mutations in a 193bp of the rpoB gene by automated sequencing of rifampicin (RMP)-resistant and susceptible Mycobacterium tuberculosis strains isolated from Brazil (25 strains) and France (37 strains). In RMP-resistant strains, mutations were identified in 100 percent (16/16) from France and 89 percent (16/18) from Brazil. No mutation was detected in the 28 RMP-susceptible strains. Among RMP-resistant or RMP-susceptible strains deletion was observed. A double point mutation which had not been reported before was detected in one strain from France. Among French resistant strains mutations were found in codons 531 (31.2 percent), 526, 513 and 533 (18.7 percent each). In Brazilian strains the most common mutations were in codons 531 (72.2 percent), 526 (11.1 percent) and 513 (5.5 percent). The heterogeneity found in French strains may be related to the fact that most of those strains were from African or Asian patients

Intraperitoneal chemohyperthermia with mitomycin C for digestive tract cancer patients with peritoneal carcinomatosis.

BACKGROUND: Most patients with peritoneal carcinomatosis of digestive tract origin die within 6 months. Intraperitoneal chemohyperthermia (IPCH) associated with surgery has been reported as a possible new therapeutic approach. METHODS: A prospective Phase II trial was carried out with 83 patients who had digestive tract cancer and peritoneal carcinomatosis to evaluate the tolerance and efficacy of IPCH with mitomycin C (MMC) associated with surgery. Eighty-six IPCH treatments with MMC were given as complementary therapy after surgery (peritoneal perfusate with a 10 mg/L dose of MMC; inflow temperature, 46-49 degrees C; use of a closed circuit; duration, 90 minutes). Primary tumors were mainly gastric (in 42 cases) or colorectal (in 27 cases). RESULTS: Mortality and morbidity occurred in 3 of 83 cases and 8 of 83 cases, respectively. For patients with resectable tumors, the median survival time was 16 months when carcinomatosis was Stage I and II (malignant granulations less than 5 mm in greatest dimension), whereas it was 6 months when carcinomatosis was Stage III and IV (malignant granulations more than 5 mm in greatest dimension). For patients with resectable gastric cancer and Stage I and II carcinomatosis, 1-, 2-, and 3-year actuarial survival rates were 80%, 61%, and 41%, respectively, whereas the rate was 10% at 1 year for patients with bulky disease (Stage III and IV). CONCLUSIONS: IPCH appears to be a promising new approach to treating patients with digestive tract cancers and peritoneal carcinomatosis with small, malignant granulations (Stage I and II).

Intraperitoneal chemo-hyperthermia with mitomycin C for gastric cancer patients with peritoneal carcinomatosis.

OBJECTIVE: This study evaluates the tolerance and efficacy of Intraperitoneal Chemo-hyperthermia (IPCH) with Mitomycin C (MMC) associated with surgery, in peritoneal carcinomatosis of gastric origin. BACKGROUND: Most patients with peritoneal carcinomatosis of gastric origin die within 6 months, and IPCH associated with surgery has been reported as a possible new therapeutic approach. METHODS: A prospective non randomized trial was carried out on 42 patients with gastric cancers and peritoneal carcinomatosis. Fourty-three IPCH with MMC were used as complementary treatment after surgery (peritoneal perfusate with a 10 mg/l dose of MMC, inflow temperature 46 to 49 degrees C, use of a closed circuit, duration 90 minutes). Fourteen primary tumors were unresectable ones and 12 patients had large malignant preoperative ascites. RESULTS: Mortality and morbidity rates were 2/42 and 4/42 respectively. For resectable gastric cancers with stage 1 and 2 carcinomatosis (malignant granulations less than 5 mm in diameter), one, two and three year survival rates were 80, 61 and 41% respectively. For unresectable primary tumors and for stage 3 and 4 carcinomatosis (granulations larger than 5 mm in diameter), six and twelve month survival rates were 50% and 10% respectively. CONCLUSIONS: IPCH appears as a safe new therapeutic approach in gastric cancers with peritoneal carcinomatosis with small malignant granulations (stage 1 and 2) and randomized trials are now needed to clearly evaluate its efficacy.

Carrier-mediated system for pefloxacin uptake in human monocytes.

Protein transporters present in cellular membranes can influence the intracellular concentration of a variety of drugs. Pefloxacin is a fluoroquinolone which is commonly used in systemic infections. Its transport mechanisms have never been described. In the present study, we demonstrated that pefloxacin uptake is carrier-mediated. Pefloxacin crosses the membrane in zwitterionic form. The transport seems to differ from that of other fluoroquinolones which use amino acid or organic anion transporters. Pefloxacin uptake is not influenced by the presence of hexose in the incubation medium. It does not use a nucleoside transport system to penetrate the cells but adenosine increases the uptake. The carrier is inhibited by verapamil at 60 min and is activated by a Ca2+-dependent system.

Tonsillar diffusion kinetics of amoxycillin after oral administration of 1 g to adults.

Although amoxycillin is widely used to treat pharyngo-tonsillitis, the kinetics of tonsillar diffusion have rarely been studied. The aim of this work was to study the kinetics of amoxycillin diffusion in the tonsils of adults, by measuring tonsillar concentrations 1.5, 3, 6 and 12 h after oral administration of 1 g of amoxycillin (Clamoxyl dispersible tablet, 1 g) relative to concomitant serum levels by using an HPLC method and a microbiological technique. At enrollment, the 20 patients were randomly divided into four groups of five, corresponding to the time intervals (1.5, 3, 6 or 12 h) between the third amoxycillin intake and the start of surgery. The results given by the two assay methods correlated well (r = 0.92-0.99). Amoxycillin showed excellent penetration into the tonsils, with both tonsillar and serum concentrations exceeding the MIC for most pathogens encountered in this setting (including Streptococcus pyogenes), even 12 h after repeated dosing with 1 g of amoxycillin.

Mycobacterium avium complex strains from AIDS patients belong to a homogeneous group ascribed by rRNA typing methods.

16S rRNA RFLP analysis of Mycobacterium avium complex (MAC) strains isolated from 25 AIDS patients led to identification of seven ribotypes. The same ribotype was determined for strains from 19 patients with and without disseminated disease. When isolates representing the seven ribotypes were examined for their internal transcribed spacer (ITS) between the 16S and 23S rRNA gene nucleotide sequence, four different sequences, including a new ITS type, were recovered. All isolates with the most common ribotype belonged to the sequevar Mav-B. When MAC strains from AIDS patients were compared by ITS sequencing and ribotyping, a significant degree of homogeneity was observed. The discriminatory level reached with ribotyping might be useful for grouping isolates from different clinical sources.

Is the uptake of pefloxacin in human blood monocytes a simple diffusion process?

Effective therapy of infections caused by facultative intracellular micro-organisms, which persist after ingestion by mononuclear phagocytic cells, requires antibiotics which inactivate these intraphagocytic bacteria. A previous study demonstrated that intracellular concentrations of normethylpefloxacin, Ci, were the result of simple diffusion and an active efflux involving organic anion transporters. In our work, we studied pefloxacin transport in human monocytes. A kinetic approach was followed to establish the processes involved. Extracellular concentrations, Ce, corresponding to in-vivo pharmacological levels of the drug were used. Uptake of the antibiotic was assessed principally by HPLC. Pefloxacin was accumulated by the cells (Ci/Ce = 3). The uptake was rapid, non saturable, reversible and temperature dependent. At 4 degrees C, the Ci/Ce was equal to 1. At high temperatures, an active and initial process appeared that was not visible at lower temperatures and disappeared in presence of energetic metabolism inhibitor: At 42 degrees C, NaCN and 2.4 DNP (1 mM) reduced the initial transport but they had no significant effect on the subsequent curve.

Aerosolized interferon gamma for Mycobacterium avium-complex lung disease.

It has recently been shown that human alveolar macrophages can be selectively activated without systemic effect by the use of aerosolized interferon-gamma (IFN gamma), a cytokine that enhances macrophage oxidative and antimicrobial activity. We report the case of a 38-yr-old man negative for human immunodeficiency virus (HIV), with silicosis and advanced cavitary lung disease due to Mycobacterium avium intracellulare (MAI), who failed to improve despite 3 yr of continuous medical therapy with three or more drugs. He received three courses of aerosolized IFN gamma (500 micrograms 3 d per week for 5 wk in two courses and 200 micrograms 3 d a week for 5 wk after a short single trial of subcutaneous IFN gamma). The numbers of MAI decreased in the sputum during therapy, but cultures of the organism remained positive at the same level for the first two treatment periods. The patients sputum became AFB smear negative and the number of colonies decreased significantly after the third course of IFN gamma therapy. Cessation of IFN gamma was associated with a rapid increase in the numbers of MAI in the sputum. Aerosolized IFN gamma can be considered as an adjuvant to conventional drug therapy, with a good tolerance, in cases of lung disease caused by resistant MAI.

Discrimination of Mycobacterium avium-Mycobacterium intracellulare strains by genomic DNA fingerprinting with a 16S rRNA gene probe.

Ribotyping was investigated as a means of distinguishing ten different serotyped reference strains and seven epidemiologically unrelated isolates of Mycobacterium avium-Mycobacterium intracellulare using a labelled 16S rDNA probe. Thirteen restriction enzymes were screened towards an accurate discrimination of strains. Two selected restriction enzymes (SacI and ClaI) enabled us to classify the 17 strains into ten ribotypes with an index of discrimination of 0.897. Typeability and reproducibility of the method reached 100%. The patterns obtained exhibited polymorphism of RE fragments within and outside the 16S rRNA gene and may be useful for epidemiological studies.

Routine identification of Mycobacterium tuberculosis complex isolates by automated hybridization.

Methodologies for biochemical identification of mycobacteria isolated from clinical samples are still cumbersome, taking skilled technicians 3 to 6 weeks. We describe here a 2-h identification system for mycobacterial isolates belonging to the Mycobacterium tuberculosis complex using a DNA probe. After 30 min of hands-off sample preparation, the 1.5-h hybridization test is totally automated in the newly developed VIDAS system (bioMérieux, Marcyl'Etoile, France), which performs solid-phase specific hybridization of 16S rRNA at 37 degrees C. The strain collection of actinomycetes tested was composed of 662 isolates from 27 species: 461 members of the M. tuberculosis complex (443 M. tuberculosis, 10 M. bovis, and 8 M. bovis BCG isolates) and 201 isolates of other species, including 55 M. avium-intracellulare isolates). They were identified by traditional methods: growth rate, colonial morphology, pigmentation, and biochemical profiles. The automated probe assay displayed an excellent correlation with the reference results. The four members of the Nocardia and Rhodococcus genera tested did not cross-hybridize. This flexible random-access and automated technology was shown to suit the routine context of the laboratory by rapidly delivering the results.

Accumulation of pefloxacin in the lower respiratory tract demonstrated by bronchoalveolar lavage.

The in-vivo pulmonary disposition of pefloxacin in alveolar macrophages alveolar macrophages and in the alveolar epithelial lining fluid recovered by bronchoalveolar lavage was studied in 10 healthy volunteers. Bronchoalveolar lavage was performed either 2 or 4 h after oral intake of 800 mg of the drug. The recovered fluid was immediately centrifuged and processed for the assays. Pefloxacin was assayed by High Pressure Liquid Chromatography (HPLC) and by a microbiological method. The mean concentrations of pefloxacin assayed by HPLC were 106 +/- 11.1 mg/L in alveolar macrophages and 88.2 +/- 10 mg/L in the epithelial lining fluid, whereas the mean serum concentration was 6.67 +/- 0.47 mg/L. Therefore, pefloxacin accumulated rapidly in human alveolar macrophages. The high epithelial lining fluid concentrations may be attributed to lipophilicity of the drug and to rapid diffusion from blood, pulmonary cells and interstitium during the bronchoalveolar lavage procedure. The substantial accumulation of pefloxacin in alveolar components (alveolar macrophages and epithelial lining fluid) endorses its use in the treatment of intracellular bacterial infections such as legionellosis; for these diseases, pefloxacin represents an alternative to the macrolide antibiotics.

Regional chemotherapy (with mitomycin C) and intra-operative hyperthermia for digestive cancers with peritoneal carcinomatosis.

Intraperitoneal chemo-hyperthermia with mitomycin C was used to treat 28 patients with far advanced digestive adenocarcinoma and histologically confirmed peritoneal carcinomatosis. Surgical resection of the primary tumor was possible in 17 cases. After closure of the abdominal wall, intraperitoneal chemo-hyperthermia was performed for 90 to 120 minutes under general anesthesia and 32 degrees C hypothermia, through 3 intraperitoneal drains forming a closed circuit, using 10 mg/l of mitomycin C in 6 liters of peritoneal dialysate heated to an inflow temperature of 46-49 degrees C. No mortality occurred, and there were 2 post-operative complications, with transitory biological side effects. In 9 out of 10 patients with preoperative malignant ascites, the ascites cleared after treatment. One-year survival rate was 54.2%. These encouraging preliminary results show that intraperitoneal chemohyperthermia with mitomycin C is a safe and reliable treatment for peritoneal carcinomatosis in far advanced digestive cancers.