Prospective study comparing the conventional and same-day strategies to diagnose pulmonary tuberculosis.

OBJECTIVE: The WHO recommends same-day sputum smear microscopy for the diagnosis of smear-positive tuberculosis (TB) in countries with high TB burden for earlier diagnosis and treatment, a cornerstone to prevent air-borne transmission. We aimed to compare the conventional strategy (sputum collection on three consecutive days) and the same-day strategy (hour h, h+1, h+2) in France, a country with low TB burden. PATIENTS AND METHODS: Over a six-month period, all adult individuals presenting with presumptive smear-positive TB were eligible for the study, registered in https://clinicaltrials.gov/ ID (NCT02961569). Sputum specimens were collected three times the first day, then once on the second day and once on the third day. The concordance between the two strategies regarding smears and cultures were assessed. RESULTS: Of the 131 eligible individuals, 34 were given a TB treatment. Smears from hour h, h+1, h+2, day two and three were negative in 19 of these 34 patients. Positive smears were obtained in 15, 14, 15, 14, and 14 patients at hour h, h+1, h+2, on day two and three, respectively. Concordance regarding smear or culture was good, with Kappa 0.69 and 0.64, respectively. CONCLUSION: The same-day strategy seems to be a good alternative to the conventional strategy.

Multidisciplinary advisory teams to manage multidrug-resistant tuberculosis: the example of the French Consilium.

SETTING: The World Health Organization (WHO) recommends that multidrug-resistant tuberculosis (MDR-TB) treatment should be managed in collaboration with multidisciplinary advisory committees (consilia). A formal national Consilium has been established in France since 2005 to provide a centralised advisory service for clinicians managing MDR-TB and extensively drug-resistant (XDR-TB) cases.OBJECTIVE: Review the activity of the French TB Consilium since its establishment.DESIGN: Retrospective description and analysis of the activity of the French TB Consilium.RESULTS: Between 2005 and 2016, 786 TB cases or contacts of TB cases were presented at the French TB Consilium, including respectively 42% and 79% of all the MDR-TB and XDR-TB cases notified in France during this period. Treatment regimens including bedaquiline and/or delamanid were recommended for 42% of the cases presented at the French TB Consilium since 2009. Patients were more likely to be presented at the French TB Consilium if they were born in the WHO Europe Region, had XDR-TB, were diagnosed in the Paris region, or had resistance to additional drugs than those defining XDR-TB.CONCLUSION: The French TB Consilium helped supervise appropriate management of MDR/XDR-TB cases and facilitated implementation of new drugs for MDR/XDR-TB treatment.

Installing biosafety level 3 containment laboratories in low- and middle-income countries: challenges and prospects from Mali's experience.

In Mali, the incidence of tuberculosis (TB) is estimated at 56 cases per 100 000 people, with a prevalence of multidrug-resistant TB in new cases of 1.7% (range, 0.3-3.1%) and in retreatment cases of 17% (range, 4.4-30%). Appropriate biosafety conditions for performing routine TB culture and antimicrobial susceptibility testing have been lacking. In 2015, a biosafety level 3 (BSL3) laboratory set up in a shipping container was donated to the Malian Ministry of Health and Public Hygiene to provide capacity for TB testing. This laboratory is now managed by Malian laboratory staff and is processing samples at the national level. We explain the necessary steps for establishing and running a BSL3 laboratory. Despite the acute need for functioning and sustainable BSL3 laboratories, low- and middle-income countries are faced with a complex process and must overcome many challenges.

Prospective study on antimicrobial resistance in leprosy cases diagnosed in France from 2001 to 2015.

Antimicrobial resistance (AMR) in leprosy is mostly unknown because Mycobacterium leprae does not grow in vitro and bacteriologic investigations have been abandoned. However, molecular detection of resistance can be applied to multibacillary cases. Patients living in France mainland or in the French territories and diagnosed with leprosy from 2001 to 2015 were prospectively studied for AMR by detecting mutations in rpoB for rifampicin resistance, in folP1 for dapsone and in gyrA for ofloxacin. Single nucleotide polymorphism (SNP) genotypes 1-4 were determined for resistant strains. Of 334 skin biopsy samples received for suspicion of leprosy, 184 (55.1%) were positive for M. leprae (acid-fast bacilli and M. leprae-specific repetitive element PCR) corresponding to 160 multibacillary cases. AMR was detected in 18 cases (11.3%): 13 cases (8.1%) of dapsone resistance, three (1.9%) rifampicin and two (1.3%) ofloxacin. There were no strains with multidrug resistance. The mutations (numbering system of M. leprae TN strain genome) found were P55L (n = 7), T53I (n = 5), T53A (n = 1) in folP1; S456L (n = 2) and S456F (n = 1) in rpoB; and A91V (n = 2) in gyrA. Resistance proportion differ significantly between new and relapse cases (9/127, 7.0%, vs. 9/33, 25.7%, p 0.003). The frequency distribution of SNP1-4 types of resistant strains was two, one, 12 and three with five SNP3 cases from New Caledonia harbouring the same T53I FolP1 substitution. This is the first report of AMR surveillance for new and relapse cases of leprosy in Europe. Detection of resistance helped in individual treatment as well as in epidemiologic investigations.

Comparison of methods available for identification of Mycobacterium chimaera.

OBJECTIVES: Mycobacterium chimaera is a recently described nontuberculous mycobacterium belonging to the Mycobacterium avium complex (MAC). Because this species is implicated in a worldwide outbreak due to contaminated heater-cooler unit water tanks during open-heart surgery, it has become mandatory for clinical microbiology laboratories to be able to differentiate M. chimaera from the other MAC species, especially M. intracellulare. Such identification has so far been restricted to specialized laboratories because it required the analysis of several gene sequences. The aim of this study was to evaluate commercial methods for identifying M. chimaera with regard to the reference gene sequencing ITS, the internal transcribed spacer 16-23S. METHODS: Forty-seven clinical and environmental isolates including 41 MAC were identified by (a) PCR sequencing of the ITS and hsp65 genes, (b) three molecular biology kits (INNO-LiPA Mycobacteria, GenoType Mycobacterium CM and GenoType NTM-DR) and (c) matrix-assisted desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) using Microflex LT. RESULTS: There was a high concordance for species determination between the reference ITS sequencing and the GenoType NTM-DR test (39/41, 95%), the INNO-LiPA Mycobacteria test (38/41, 93%) and the hsp65 sequencing (38/41, 93%). The GenoType Mycobacterium CM test did not distinguish M. chimaera from M. intracellulare. MALDI-TOF MS distinguished two M. chimaera-M. intracellulare groups separated from M. avium and from the other mycobacterial species on a score-oriented dendrogram, but it also failed to differentiate the two species. CONCLUSIONS: INNO-LiPA Mycobacteria and GenoType NTM-DR are efficient assays for M. chimaera identification in clinical microbiology laboratories.

Monitoring antimicrobial resistance (AMR) using CUSUM control charts.

We evaluated the use of the Cumulative Summation (CUSUM) control chart methodology for detection of an excessive increase in antimicrobial-resistant (AMR) bacteria acquisition. We used administrative, clinical and bacteriological data from all 157,570 patients hospitalized for at least 48 h from January 1, 2010 to December 31, 2015 in a 654-bed university teaching hospital in Paris, France. Monthly computed CUSUM were evaluated for the detection of out-of-control situations, defined as incidence rates of acquired AMR bacterial colonization exceeding acceptable thresholds at the hospital and ward levels (based on six selected wards) for AMR bacteria overall and Extended-spectrum beta-lactamases Enterobacteriaceae (ESBL-E) and Methicillin-resistant Staphylococcus aureus (MRSA), specifically. During the study period, 1,403 samples of acquired AMR bacteria were identified including 1,129 ESBL-E and 151 MRSA. The incidence rate of acquired AMR bacteria was stable at the hospital and the wards level. When based on AMR bacteria overall, CUSUM alarms were triggered at the hospital level and at the ward level in four units. For ESBL-E, CUSUM tests generated alarms at the hospital level and for the same four wards, and for MRSA, CUSUM tests detected out-of-control situations in all the wards. The CUSUM approach appears complementary with hospital infection control strategies currently in practice and appears of interest in common practice as a simple tool for AMR surveillance.

[Cutaneous tuberculosis of the ear due to Mycobacterium bovis]. / Tuberculose cutanée de l'oreille à Mycobacterium bovis.

INTRODUCTION: Isolated cutaneous tuberculosis is uncommon, accounting for only 0.14 to 5% of Mycobacterium tuberculosis infections. We report a rare case of ear cutaneous tuberculosis due to Mycobacterium bovis in an immunocompetent woman. CASE REPORT: A 59-year-old woman presented an erythematous and scaly lesion of the ear present for two years. The histological findings were compatible with a diagnosis of sarcoidosis, with non-necrotic granuloma. After failure of dermal corticosteroid therapy, a further biopsy identified M. bovis; the patient was cured following anti-tubercular treatment. DISCUSSION: Ear lesions are predominantly associated with tumors, fungal infections, chondritis, lupus and sarcoidosis. The ear, like the face in general, is a classic localization of lupus vulgaris, a chronic form of confined tuberculosis infection with progressive evolution. The paucibacillary nature of these lesions is the reason why their diagnosis is based in some cases on clinical, histological and immunological findings without bacteriological evidence. However, given the potential therapeutic implications, it is important to push the microbiological analysis as far as possible. In our case, culture and identification provided evidence of M. bovis infection, enabling suitable and effective therapy to be given.

High medical impact of implementing the new polymeric bead-based BacT/ALERT® FAPlus and FNPlus blood culture bottles in standard care.

Blood culture (BC) efficiency is critical for the diagnosis of bloodstream infection (BSI). We evaluated the impact on standard care of implementing the new BacT/ALERT® FAPlus and FNPlus BC bottles containing antibiotic-binding polymeric beads. We measured positivity rates and time to detection (TTD) during the first 10 months of implementation (PF) and during the previous 10-month period (PS) during which we were using standard aerobic (SA) or standard anaerobic (SN) BC bottles. For each period, the same number of consecutive patients (n = 3,918) was included. Per patient, a median of 1 BC set (1 aerobic and 1 anaerobic bottles) has been sampled. A higher positivity rate was measured during PF than PS when counting per BC bottle (7.0 % vs 5.8 % with 1,456 and 1,237 positive bottles respectively, P < 0.0001) and per BC set (9.6 % vs 7.8 % with 995 and 832 positive BC sets respectively, P < 0.0001). In PF, an increased number of cases due to staphylococci (P < 0.0001) and to Gram-negative bacilli (P < 0.005) was observed, whereas the contamination rate was similar during the two periods (2.4 % of BC sets in PF and 2.3 % in PS). Although antibiotic consumption and medical activity were similar during the two periods, BSI case detection increased from 2.2 to 2.6 per 1,000 hospital-days, especially in intensive care units (ICU; 35.1 to 55.7). Mean TTD for pathogenic microorganisms was significantly shorter in PF than in PS (15.5 h vs 18.0 h, P < 0.01). In conclusion, the use of the new FAPlus/FNPlus BC bottles improved the diagnosis of bacteremia in our hospital, especially in ICU patients.

Revisiting susceptibility testing in MDR-TB by a standardized quantitative phenotypic assessment in a European multicentre study.

OBJECTIVES: Treatment outcome of MDR-TB is critically dependent on the proper use of second-line drugs as per the result of in vitro drug susceptibility testing (DST). We aimed to establish a standardized DST procedure based on quantitative determination of drug resistance and compared the results with those of genotypes associated with drug resistance. METHODS: The protocol, based on MGIT 960 and the TB eXiST software, was evaluated in nine European reference laboratories. Resistance detection at a screening drug concentration was followed by determination of resistance levels and estimation of the resistance proportion. Mutations in 14 gene regions were investigated using established techniques. RESULTS: A total of 139 Mycobacterium tuberculosis isolates from patients with MDR-TB and resistance beyond MDR-TB were tested for 13 antituberculous drugs: isoniazid, rifampicin, rifabutin, ethambutol, pyrazinamide, streptomycin, para-aminosalicylic acid, ethionamide, amikacin, capreomycin, ofloxacin, moxifloxacin and linezolid. Concordance between phenotypic and genotypic resistance was >80%, except for ethambutol. Time to results was short (median 10 days). High-level resistance, which precludes the therapeutic use of an antituberculous drug, was observed in 49% of the isolates. The finding of a low or intermediate resistance level in 16% and 35% of the isolates, respectively, may help in designing an efficient personalized regimen for the treatment of MDR-TB patients. CONCLUSIONS: The automated DST procedure permits accurate and rapid quantitative resistance profiling of first- and second-line antituberculous drugs. Prospective validation is warranted to determine the impact on patient care.

Predictive factors for extended-spectrum beta-lactamase producing Enterobacteriaceae causing infection among intensive care unit patients with prior colonization.

We investigated the predictive factors for extended-spectrum beta-lactamase producing Enterobacteriaceae (ESBL-PE) causing infections among intensive care unit patients with prior documented ESBL-PE colonization. Using multivariate analysis, referral from medical ward, nursing home or rehabilitation center [Odds ratio (OR), 2.5; 95 % confidence interval (CI), [1.3-5.0]; p = 0.007], previous fluoroquinolone treatment (OR, 3.4; CI, [1.1-10.5]; p = 0.003), extracorporeal membrane oxygenation (OR, 4.6; CI, [1.3-15.9]; p = 0.02), and absence of prior positive ESBL-PE rectal swab culture (OR, 5.0; CI, [1.6-10.0]; p = 0.0009) were risk factors for ESBL-PE infection. Easily identifiable factors may help with targeting carbapenem prescriptions.

Outbreak in a haematology unit involving an unusual strain of glycopeptide-resistant Enterococcus faecium carrying both vanA and vanB genes.

OBJECTIVES: To report an outbreak due to an unusual strain of Enterococcus faecium containing both the vanA and vanB genes, in France, where the rate of glycopeptide-resistant enterococci (GRE) is below 1%. METHODS: Cases were patients infected or colonized with GRE on the haematology ward. Contact patients were screened by real-time PCR performed on rectal swabs. Clinical features were compared for GRE-positive and GRE-negative patients. GRE isolates were characterized by phenotypic and molecular methods including PFGE. Conjugation experiments were performed to identify van genetic support. RESULTS: After the index patient presented a bacteraemia with vanA/vanB E. faecium, 56 contact patients were screened, 7 of whom were found to be GRE positive: 6 additional cases with vanA/vanB E. faecium and 1 with GRE carrying vanA only. PFGE confirmed the clonal relationship of the seven vanA/vanB E. faecium strains, whereas the vanA isolate was distinct. Only the vanA gene could be transferred to enterococcal recipients by conjugation, and it was probably localized on a mobile genetic element. All isolates were resistant to vancomycin (MIC > 256 mg/L) and teicoplanin (MIC = 24-32 mg/L), but were susceptible to tigecycline (MIC = 0.09 mg/L), linezolid (MIC = 0.75 mg/L) and daptomycin (MIC = 1-2 mg/L). Significant differences (P < 0.001) between carriers and non-carriers of GRE were observed for the median duration of hospitalization (57 days versus 16.5 days) and of neutropenia (40 days versus 6 days), the median number of antibiotics used (5 versus 1.5) and the duration of glycopeptide treatment (14.5 days versus 0 days). CONCLUSIONS: vanA/vanB E. faecium strains, although rare, can emerge in the absence of previous outbreaks of vanA-GRE or vanB-GRE.

[Enterobacteriaceae producing extended spectrum beta-lactamase: epidemiology, risk factors, and prevention]. / Entérobactéries productrices de bêta-lactamases à spectre élargi : épidémiologie, facteurs de risque et mesures de prévention.

Multidrug-resistant bacteria are a major worldwide health public concern. It results from the growing increase in antibiotic prescriptions, which are responsible for selection pressure on bacteria. In France like in other countries, enterobacteriaceae producing extended spectrum beta-lactamase (EESBL) are the predominant multidrug-resistant bacteria. EESBL may be responsible for severe infections and require prescription of broad-spectrum antibacterial agents. The current EESBL outbreak is different from methicillin-resistant Staphylococcus aureus outbreak that occurred in the early 1980. Consistently, EESBL are isolated both in hospital and community. Moreover, standard hygiene measures appear ineffective since EESBL prevalence is still increasing. The current inability to contain EESBL outbreak is due to several factors, including the existence of a wide community- and hospital-acquired tank of EESBL, failure to follow strict rules for hygiene, and the current irrational prescription of antibiotics.

Appearance of aac(6')-Ib-cr gene among extended-spectrum beta-lactamase-producing Enterobacteriaceae in a French hospital.

OBJECTIVES: The aac(6')-Ib gene encodes many variants of an aminoglycoside-acetyltransferase enzyme that is responsible for amikacin resistance. Recently, a new variant aac(6')-Ib-cr capable of modifying aminoglycosides and fluoroquinolones has been described. The aim of our study was to observe the appearance and the location of the aac(6')-Ib gene in extended-spectrum beta-lactamase (ESBL)-producing Enterobacteriaceae strains. METHODS: Sixty-six and nine non-clonal ESBL-producing Enterobacteriaceae strains were isolated, respectively, for one 3-year period from 1999 to 2001 and one 2-month period in 2005 in a French Hospital (Paris, France). RESULTS: Among these isolates, 35 of them carried the aac(6')-Ib gene. Fourteen out of the aac(6')-Ib genes of the period 1 and two of the period 2 were genes cassette located within class 1 integrons, whereas 16 and 3, respectively, were outside integrons. One of these encoded an aminoglycoside-acetyltransferase enzyme leading to an acetyltransferase that confers resistance to all aminoglycosides. The new -cr variant of aac(6')-Ib was detected in three Escherichia coli isolates in 2005 always associated with CTX-M-15 enzyme. CONCLUSIONS: The aac(6')-Ib-cr gene, responsible for antibiotic resistance to two very different drugs, is emerging in ESBL-producing Enterobacteriaceae strains isolated in France especially in strains carrying the bla(CTx-M-15) gene.

Cervical necrotizing fasciitis: 8-years' experience of microbiology.

Cervical necrotizing fasciitis (CNF) is a life-threatening complication of pharyngeal or dental infections. The aim of this paper was to investigate whether dental or pharyngeal source result from different pathogen(s) in CNF and whether antibiotics, given before admission, influence the antimicrobial resistance of pathogens. In 152 CNF patients, Streptococcus milleri group and Prevotella species were the predominant isolates, frequently copathogens, mostly in dental CNF samples. Penicillin and clindamycin resistance were observed in 39% and 37% of cases, respectively, independently of any previous antibiotic therapy. Thus, a combined aerobe-anaerobe infection may have a synergistic effect, which allows the infection to spread in cervical tissues.

Improved diagnosis specificity in bone and joint infections using molecular techniques.

OBJECTIVES: The microbiological diagnosis of osteoarticular infections currently relies on microbiological cultures, to specifically target treatment. However, these conventional methods sometimes lack of sensitivity and of specificity to establish definitive diagnosis. This study was conducted to determine whether molecular method could improve bacterial bone and joint infection diagnosis. METHODS: We evaluated the performance of nucleic acid extraction with the semi-automated NucliSens miniMAG instrument coupled to 16S rDNA sequencing on 76 samples collected from 51 patients with suspected infections: prosthetic-joint infection (15), spondylodiscitis (7) acute septic arthritis (11) and 18 controls. No pre-treatment of the sample was done before nucleic acid extraction. Classification in infected group required an accumulation of arguments. RESULTS: Our molecular method identified a broad spectrum of pathogenic bacteria including Staphylococcus aureus, Streptococcus pyogenes, Streptococcus agalactiae, Enterococcus faecalis, Salmonella enterica, Escherichia coli, Pseudomonas aeruginosa, and fastidious bacteria like Neisseria gonorrhoeae and Fusobacterium nucleatum. The overall PCR sensitivity was 73.3%: 53.8% for prosthetic-joint infections and 88.2% for infections without prostheses. The overall PCR specificity was 95.2%, whereas culture specificity was only 85.7%. CONCLUSIONS: The instrument was simple to use and provided nucleic acids free of PCR inhibitors and free of contamination by foreign bacterial DNA. Our study highlights the need for still improved molecular diagnostic sensitivity.

Prospective, randomized, parallel-group trial to evaluate the effects of lactulose and polyethylene glycol-4000 on colonic flora in chronic idiopathic constipation.

BACKGROUND: Although lactulose and polyethylene glycol are osmotic laxatives widely used in the treatment of chronic constipation, no study has been conducted to compare their actions on the colonic bacterial ecosystem, which has an important influence on host health. AIM: To assess the effects of lactulose and polyethylene glycol on the composition and metabolic indices of the faecal flora in patients with chronic idiopathic constipation. METHODS: Sixty-five patients with chronic idiopathic constipation were included in this controlled, multi-centre, randomized, parallel-group study. Participants received lactulose (Duphalac) or polyethylene glycol-4000 (Forlax) powders for the first week at a fixed dosage at night (20 g/day); in the second week, patients were given the option to vary the dose according to efficacy and tolerance (10-30 g/day); for the last 2 weeks, treatment was administered at a fixed dosage based on the results of the second week (10-30 g/day). Stools were recovered for bacteriological analysis at days -1, 21 and 28. RESULTS: Clinical efficacy and tolerance were similar with both treatments. In the lactulose group, an increase in faecal bifidobacteria counts (P = 0.04) and beta-galactosidase activity (P < 0.001) was observed from day -1 to day 28, whereas, in the polyethylene glycol group, there was a decrease in total short-chain fatty acids (P = 0.02), butyrate (P = 0.04), acetate (P = 0.02) and faecal bacterial mass (P = 0.001). No differences were observed in stools with regard to the following parameters: counts of Lactobacillus, clostridial spores, Bacteroides and enterobacteria, pH, biliary acids and neutral sterol concentrations. CONCLUSIONS: Both lactulose and polyethylene glycol are efficacious and well tolerated. However, although lactulose can be considered as a pre-biotic in constipated patients, polyethylene glycol produces signs of decreased colonic fermentation in the stool.