Clostridium difficile infection after allogeneic hematopoietic stem cell transplantation: incidence, risk factors, and outcome.

Clostridium difficile (C. difficile) infection was observed in 13% of recipients after hematopoietic stem cell transplantation (HSCT), mainly in the first month posttransplantation. Risk factors were cord blood as the source of stem cells, acute graft-versus-host disease (GVHD), and total body irradiation (TBI). No association was found with an increased risk of mortality. The purpose of this study was to evaluate the incidence, risk factors, and outcome of C. difficile infection (CDI) after HSCT. We conducted a single-center, retrospective, cohort study on all patients who received an allogeneic HSCT from January 2004 to December 2007. All patients with diarrhea in the first year after HSCT were tested for the presence of C. difficile in stools. Among the 407 assessable patients, 53 presented at least 1 CDI in the first year post-HSCT. The total incidence rate was 5.6 cases of CDI per 10,000 patient-days. Fifty percent of cases were diagnosed in the first month after HSCT, and 95% occurred during the first 6 months. Fewer than 5% of patients with CDI had severe diarrhea and severe complications were never observed. TBI in the conditioning regimen, cord blood as the source of stem cells, and acute graft-versus-host disease (aGVHD) were independently associated with CDI. Six patients (11%) had a recurrence of CDI. Four patients required second-line treatment with vancomycin. With a median follow-up of 22 months, the 2-year overall survival rates were similar between patients who presented a CDI and those who did not. CDI was observed in approximately 13% of recipients after HSCT, mainly in the first month posttransplantation and was associated with CB, aGVHD, and TBI. CDI was not associated either with severe complications or with an increased risk of mortality in this large cohort of patients.

Absence of community-acquired methicillin-resistant Staphylococcus aureus carriage or infection in a cohort of 300 men who have sex with men attending an inner-city sexually transmitted disease clinic in Paris, France.

BACKGROUND: Community-acquired methicillin-resistant Staphylococcus aureus (caMRSA) skin and soft-tissue infections are a major cause for concern. Communities of men who have sex with men (MSM) in the USA being prone to caMRSA epidemics, we investigated whether MSM attending our sexually transmitted disease clinic were carriers of or infected by caMRSA. METHODS: 300 MSM were prospectively recruited, interrogated and examined. Swabs of their nares and of their pubic and perineal skin were taken and cultured. RESULTS: No evidence of carriage or infection by caMRSA was found. Nasal carriage of methicillin-sensitive S. aureus was within French estimates. Perineal carriage significantly correlated with the number of sexual partners. CONCLUSION: Although caMRSA carriage or infection does not seem to affect Paris MSM, prospective sampling may not be the best method to monitor them. The correlation between number of partners and perineal carriage suggests that thorough hygiene after intercourse could limit the spread of S. aureus in MSM with multiple partners.

Molecular and biochemical characterization of the natural chromosome-encoded class A beta-lactamase from Pseudomonas luteola.

Pseudomonas luteola (formerly classified as CDC group Ve-1 and named Chryseomonas luteola) is an unusual pathogen implicated in rare but serious infections in humans. A novel beta-lactamase gene, bla(LUT-1), was cloned from the whole-cell DNA of the P. luteola clinical isolate LAM, which had a weak narrow-spectrum beta-lactam-resistant phenotype, and expressed in Escherichia coli. This gene encoded LUT-1, a 296-amino-acid Ambler class A beta-lactamase with a pI of 6 and a theoretical molecular mass of 28.9 kDa. The catalytic efficiency of this enzyme was higher for cephalothin, cefuroxime, and cefotaxime than for penicillins. It was found to be 49% to 59% identical to other Ambler class A beta-lactamases from Burkholderia sp. (PenA to PenL), Ralstonia eutropha (REUT), Citrobacter sedlakii (SED-1), Serratia fonticola (FONA and SFC-1), Klebsiella sp. (KPC and OXY), and CTX-M extended-spectrum beta-lactamases. No gene homologous to the regulatory ampR genes of class A beta-lactamases was found in the vicinity of the bla(LUT-1) gene. The entire bla(LUT-1) coding region was amplified by PCR and sequenced in five other genetically unrelated P. luteola strains (including the P. luteola type strain). A new variant of bla(LUT-1) was found for each strain. These genes (named bla(LUT-2) to bla(LUT-6)) had nucleotide sequences 98.1 to 99.5% identical to that of bla(LUT-1) and differing from this gene by two to four nonsynonymous single nucleotide polymorphisms. The bla(LUT) gene was located on a 700- to 800-kb chromosomal I-CeuI fragment, the precise size of this fragment depending on the P. luteola strain.

Etiology of genital ulcer disease. A prospective study of 278 cases seen in an STD clinic in Paris.

OBJECTIVES: The goal of this study was to identify the causes and factors associated with genital ulcer disease (GUD) among patients attending a sexually transmitted disease (STD) clinic in Paris. METHODS: This study was a prospective investigation of GUD cases. Data were collected from 1995 to 2005. In each case, a Dark Field Examination (DFE), Gram stain, inoculation onto Thayer Martin agar, Columbia agar and chocolate agar with 1% isovitalex and 20% fetal calf serum, PCR Chlamydia trachomatis (Amplicor Roche), culture for herpes simplex virus (HSV) on MRC 5 cells and PCR HSV (Argene Biosoft) were obtained from the ulceration. First Catch Urine (FCU) PCR for Chlamydia trachomatis and syphilis, HIV, HSV, and HBV serologies were also performed. RESULTS: A total 278 cases of GUD were investigated, 244 (88%) in men and 34 (12%) in women. Primary syphilis accounted for 98 cases (35%), genital herpes for 74 (27%), chancroid for 8 (3%), other infections for 12 (5%). In 91 (32%) patients, no identifiable microorganism was documented. Primary syphilis was more prevalent in MSMs (P < 0.0001), while genital herpes and chancroid were significantly associated with heterosexuality (both P < 0.0001). A high level of HIV infection (27%) was found, particularly in patients with primary syphilis (33%). In the univariate analysis, no statistical difference was found between syphilis and herpes according to clinical presentation, pain being the only item slightly more frequent in herpes (P = 0.06). In the multivariable model syphilis was associated with being MSM (OR: 51.3 [95% CI: 14.7-178.7], P < 0.001) and with an ulceration diameter >10 mm (OR: 9.2 [95% CI: 2.9-30.7], P < 0.001). Genital herpes was associated with HIV infection in the subgroup of MSWs (OR: 24.4 [2.4-247.7], P = 0.007). We did not find significant differences in the clinical presentation of the ulcers according to HIV status. CONCLUSION: The profound changes of the epidemiology of GUD during the decade, due to disappearance of chancroid and reemergence of infectious syphilis have led to a new distribution of pathogens, genital herpes, primary syphilis and GUD from unknown origin, accounting each for one third of cases. No clinical characteristic is predictive of the etiology, underlining the importance of performing a thorough microbiologic evaluation. Close association with HIV is still a major public health problem.

Multiple-antibiotic resistance in Salmonella enterica serotype Paratyphi B isolates collected in France between 2000 and 2003 is due mainly to strains harboring Salmonella genomic islands 1, 1-B, and 1-C.

This study was conducted to investigate the occurrence of multiple-antibiotic resistance among 261 clinical isolates of Salmonella enterica serotype Paratyphi B strains collected between 2000 and 2003 through the network of the French National Reference Center for Salmonella. The 47 multidrug-resistant (MDR) isolates identified (18%), were characterized on the basis of the presence of several resistance genes (bla(TEM), bla(PSE-1), bla(CTX-M), floR, aadA2, qacEdelta1, and sul1), the presence of Salmonella genomic island 1 (SGI1) by PCR mapping and hybridization, and the clonality of these isolates by several molecular (ribotyping, IS200 profiling, and pulsed-field gel electrophoresis [PFGE]) and phage typing methods. The results of PCR and Southern blot experiments indicated that 39 (83%) of the 47 S. enterica serotype Paratyphi B biotype Java MDR isolates possessed the SGI1 cluster (MDR/SGI1). Among these 39 MDR/SGI1 isolates, only 3 contained variations in SGI1, SGI1-B (n = 1) and SGI1-C (n = 2). The 39 MDR/SGI1 isolates showed the same specific PstI-IS200 profile 1, which contained seven copies from 2.6 to 18 kb. Two PstI ribotypes were found in MDR/SGI1 isolates, RP1 (n = 38) and RP6 (n = 1). Ribotype RP1 was also found in two susceptible strains. Analysis by PFGE using XbaI revealed that all the MDR/SGI1 isolates were grouped in two related clusters, with a similarity percentage of 82%. Isolation of MDR/SGI1 isolates in France was observed mainly between the second quarter of 2001 and the end of 2002. The source of the contamination has not been identified to date. A single isolate possessing the extended-spectrum beta-lactamase bla(CTX-M-15) gene was also identified during the study.

Evaluation of three techniques for detection of low-level methicillin-resistant Staphylococcus aureus (MRSA): a disk diffusion method with cefoxitin and moxalactam, the Vitek 2 system, and the MRSA-screen latex agglutination test.

Very-low-level methicillin-resistant Staphylococcus aureus (MRSA), or class 1 MRSA, is often misdiagnosed as methicillin-susceptible S. aureus (MSSA). We evaluated the performances of three methods for detection of low-level methicillin resistance: the disk diffusion method using the cephamycin antibiotics cefoxitin and moxalactam, the Vitek 2 system (bioMérieux), and the MRSA-screen test (Denka). Detection of the mecA gene by PCR was considered to be the "gold standard." We also determined the sensitivity of the oxacillin disk diffusion method with 5- and 1-microg disks and that of the Oxascreen agar assay with 6 mg of oxacillin liter(-1) for detection of MRSA. We compared the distributions of MICs of oxacillin and cefoxitin by the E-test (AB Biodisk), and those of moxalactam by dilutions in agar, for MRSA and MSSA isolates. The 152 clinical isolates of S. aureus studied were divided into 69 MSSA (mecA-negative) and 83 MRSA (mecA-positive) isolates, including 63 heterogeneous isolates and 26 class 1 isolates (low-level resistance). The cefoxitin and moxalactam disk diffusion tests detected 100% of all the MRSA classes: cefoxitin inhibition zone diameters were <27 mm, and moxalactam inhibition zone diameters were <24 mm. The Vitek 2 system and the MRSA-screen test detected 94 and 97.6% of all MRSA isolates, respectively. The sensitivities of the 5- and 1-microg oxacillin disks were 95.2 and 96.4%, respectively, whereas that of the Oxascreen agar screen assay was 94%. All of the tests except the 1-microg oxacillin disk test were 100% specific. For the class 1 MRSA isolates, the sensitivity of the Vitek 2 test was 92.3%, whereas those of the MRSA-screen test and the disk diffusion method with cefoxitin and moxalactam were 100%. Therefore, the cefoxitin and moxalactam disk diffusion methods were the best-performing tests for routine detection of all classes of MRSA.

An intrinsic control element for translational initiation in class 1 integrons.

Integrons are genetic elements able to capture anti-biotic resistance and other genes and to promote their transcription. Here, we have investigated integron-dependent translation of an aminoglycoside 6'-N-acetyltransferase gene (aac(6')-Ib7) inserted at the attI1 site. N-terminal sequencing revealed that translation of this gene was initiated at a GTG codon, which is not part of a plausible translation initiation region (TIR). A short open reading frame (called ORF-11) overlapping the attI1 site was probed by site-directed mutagenesis for its contribution to aac(6')-Ib7 translation. When ORF-11 and its TIR were deleted en bloc, translational efficiency dropped by over 80%, as determined with an acetyltransferase- luciferase fusion product. Invalidation of the ATG start codon of ORF-11 or its putative Shine-Dalgarno sequence resulted in a decrease of over 60%, whereas the decrease was much less pronounced when the amino acid sequence of the putative ORF-11-encoded peptide was altered or when the distance between ORF-11 and aac(6')-Ib7 was doubled. This demonstrates that aac(6')-Ib7 translation is dependent upon the translation of ORF-11, but almost certainly not upon the corresponding peptide. These results lead us to conclude that an intrinsic short ORF present in the 5'-conserved segment of many class 1 integrons may substantially enhance expression at the translational level of captured TIR-deficient anti-biotic resistance genes.

Fluoroquinolone resistance linked to GyrA, GyrB, and ParC mutations in Salmonella enterica typhimurium isolates in humans.

We report two cases of infection with clonally unrelated, high-level ciprofloxacin-resistant, b-lactamase-producing strains of Salmonella enterica Typhimurium. Resistance was caused by four topoisomerase mutations, in GyrA, GyrB, and ParC and increased drug efflux. Ciprofloxacin treatment failed in one case. In the second case, reduced susceptibility to third-generation cephalosporins occurred after initial treatment with these drugs and may explain the treatment failure with ceftriaxone.

Salmonella enterica serovar Typhimurium bla(PER-1)-carrying plasmid pSTI1 encodes an extended-spectrum aminoglycoside 6'-N-acetyltransferase of type Ib.

We have studied the aminoglycoside resistance gene, which confers high levels of resistance to both amikacin and gentamicin, that is carried by plasmid pSTI1 in the PER-1 beta-lactamase-producing strain of Salmonella enterica serovar Typhimurium previously isolated in Turkey. This gene, called aac(6')-Ib(11), was found in a class 1 integron and codes for a protein of 188 amino acids, a fusion product between the N-terminal moiety (8 amino acids) of the signal peptide of the beta-lactamase OXA-1 and the acetyltransferase. The gene lacked a plausible Shine-Dalgarno (SD) sequence and was located 45 nucleotides downstream from a small open reading frame, ORF-18, with a coding capacity of 18 amino acids and a properly spaced SD sequence likely to direct the initiation of aac(6')-Ib(11) translation. AAC(6')-Ib(11) had Leu118 and Ser119 as opposed to Gln and Leu or Gln and Ser, respectively, which were observed in all previously described enzymes of this type. We have evaluated the effect of Leu or Gln at position 118 by site-directed mutagenesis of aac(6')-Ib(11) and two other acetyltransferase gene variants, aac(6')-Ib(7) and -Ib(8), which naturally encode Gln118. Our results show that the combination of Leu118 and Ser119 confers an extended-spectrum aminoglycoside resistance, with the MICs of all aminoglycosides in clinical use, including gentamicin, being two to eight times higher for strains with Leu118 and Ser119 than for those with Gln118 and Ser119.

High prevalence of Mycoplasma genitalium in the lower genitourinary tract of women attending a sexually transmitted disease clinic in Paris, France.

BACKGROUND: Previous studies have suggested that Mycoplasma genitalium is associated with cervicitis and may be a cause of pelvic inflammatory disease. This study attempted to investigate further the possible role of M genitalium in genital symptoms of women attending a sexually transmitted disease (STD) clinic. GOAL: To determine the prevalence of and the association of clinical and microbiologic features with M genitalium in women presenting with genital symptoms. STUDY DESIGN: Between April 1994 and June 1996 a prospective study of 170 consecutive women with abnormal vaginal discharge, with or without urethral itching, dysuria, or pelvic pain, was conducted at the STD clinic at Saint-Louis Hospital in Paris. Information was collected on each subject's characteristics, and a clinical vulvar, vaginal, and cervical examination was performed. Cervical, vaginal, and urethral samples were obtained to identify infecting organisms. RESULTS: DNA of M genitalium was identified by polymerase chain reaction (PCR) at one or more genital sites in 65 women (38%; 95% CI, 31-46%) and was statistically more frequently detected in the vagina (39%) than in the cervix (21%) or urethra (28%) (P = 0.001 and 0.048, respectively). PCR inhibitors were detected in 17 specimens (4%). M genitalium was found in 6/14 (43%) positive for Chlamydia trachomatis, but no significant association between M genitalium and any demographic, clinical, or microbiologic data was noted in univariable or multivariable analysis. CONCLUSION: This study indicates that M genitalium is frequently encountered in the female genital tract and is not associated with cervicitis or any particular clinical or microbiologic data.