Genomic analysis of the international high-risk clonal lineage Klebsiella pneumoniae sequence type 395.

BACKGROUND: Klebsiella pneumoniae, which is frequently associated with hospital- and community-acquired infections, contains multidrug-resistant (MDR), hypervirulent (hv), non-MDR/non-hv as well as convergent representatives. It is known that mostly international high-risk clonal lineages including sequence types (ST) 11, 147, 258, and 307 drive their global spread. ST395, which was first reported in the context of a carbapenemase-associated outbreak in France in 2010, is a less well-characterized, yet emerging clonal lineage. METHODS: We computationally analyzed a large collection of K. pneumoniae ST395 genomes (n = 297) both sequenced in this study and reported previously. By applying multiple bioinformatics tools, we investigated the core-genome phylogeny and evolution of ST395 as well as distribution of accessory genome elements associated with antibiotic resistance and virulence features. RESULTS: Clustering of the core-SNP alignment revealed four major clades with eight smaller subclades. The subclades likely evolved through large chromosomal recombination, which involved different K. pneumoniae donors and affected, inter alia, capsule and lipopolysaccharide antigen biosynthesis regions. Most genomes contained acquired resistance genes to extended-spectrum cephalosporins, carbapenems, and other antibiotic classes carried by multiple plasmid types, and many were positive for hypervirulence markers, including the siderophore aerobactin. The detection of "hybrid" resistance and virulence plasmids suggests the occurrence of the convergent ST395 pathotype. CONCLUSIONS: To the best of our knowledge, this is the first study that investigated a large international collection of K. pneumoniae ST395 genomes and elucidated phylogenetics and detailed genomic characteristics of this emerging high-risk clonal lineage.

Detection of carbapenemase-producing Enterobacterales by means of matrix-assisted laser desorption ionization time-of-flight mass spectrometry with ertapenem susceptibility-testing disks as source of carbapenem substrate.

MALDI-TOF mass spectrometry has become widely used in clinical microbiology and has proved highly accurate for detection of carbapenemases in Gram-negative bacteria. However, the use of carbapenem-hydrolysis assays in routine diagnostics is hampered by the need for antibiotic substances and for making their fresh solutions each time an assay is conducted. Here, we evaluated the use of commercial antibiotic susceptibility-testing disks as source of ertapenem substrate in MALDI-TOF MS-based assay for detection of carbapenemase-producing Enterobacterales (CPE). The assay was validated on 48 CPE isolates of 8 different species expressing NDM-, VIM-, KPC- and OXA-48-type carbapenemases and exhibiting various levels of resistance to carbapenems (MIC range: 0.25- > 32 mg/l), as well as on 48 carbapenemase-non-producing isolates. The assay conditions were optimized as follows: 10-µl loopful of bacterial colonies was suspended in 150 µl 0.01 M Na-PBS buffer, pH 7.4, a 10 µg ertapenem susceptibility-testing disk was immersed in the suspension and incubated 3 h at 35°C, after which supernatant was obtained by centrifugation and applied on a target plate with alpha-cyano-4-hydroxycinnamic acid matrix. Mass spectra were analyzed between 440 and 560 m/z. Carbapenemase activity was detected in all tested CPE isolates by the appearance of m/z peaks corresponding to ertapenem hydrolysis products: [Mh + H]+:494.2, [Mh + Na]+:516.2, [Mh + 2Na]+:538.2, [Mh/d + H]+:450.2, [Mh/d + Na]+:472.2, and simultaneous decrease or loss of peaks of intact antibiotic: [M + H]+:476.2, [M + Na]+:498.1, [M + 2Na]+:520.1. No hydrolysis peaks or loss of intact ertapenem peaks were observed for carbapenemase-negative strains. We therefore report the development of a sensitive, specific and cost-effective MALDI-TOF MS-based assay for detection of CPE, which makes use of antibiotic disks readily available in most laboratories.

AMRmap: An Interactive Web Platform for Analysis of Antimicrobial Resistance Surveillance Data in Russia.

Surveillance of antimicrobial resistance (AMR) is crucial for identifying trends in resistance and developing strategies for prevention and treatment of infections. Globally, AMR surveillance systems differ in terms of organizational principles, comprehensiveness, accessibility, and usability of data presentation. Until recently, the data on AMR in Russia were scarcely available, especially to international community, despite the fact that the large prospective multicenter surveillance in Russia was conducted and data were accumulated for over 20 years. We describe the source of data, structure, and functionality of a new-generation web platform, called AMRmap (https://amrmap.net/), for analysis of AMR surveillance data in Russia. The developed platform currently comprises susceptibility data of >40,000 clinical isolates, and the data on abundance of key resistance determinants, including acquired carbapenemases in gram-negatives, are updated annually with information on >5,000 new isolates. The AMRmap allows smart data filtration by multiple parameters and provides interactive data analysis and visualization tools: MIC and S/I/R distribution plots, time-trends and regression plots, associated resistance plots, prevalence maps, statistical significance graphs, and tables.

Complete Genome Sequence of Acinetobacter baumannii Phage BS46.

Acinetobacter myovirus BS46 was isolated from sewage by J. S. Soothill in 1991. We have sequenced the genome of BS46 and found it to be almost unique. BS46 contains double-stranded DNA with a genome size of 94,068 bp and 176 predicted open reading frames. The gene encoding the tailspike that presumably possesses depolymerase activity toward the capsular polysaccharides of the bacterial host was identified.

Vaccine Prophylaxis of Pneumococcal Infections in Children under Conditions of Severe Flood in the Amur River Basin.

BACKGROUND: Pneumococcal infection being one of the dominant causes of acute respiratory diseases and exacerbation of chronic ones is a serious problem for human health and society. The flood in the Amur river basin in the summer of 2013 created a special zone and risk conditions for the formation of respiratory pathology in the Far-Eastern region of Russia. We aimed to give clinical and epidemiological assessment of the effectiveness of vaccination programs of respiratory viral and pneumococcal infections and generalization of regional experience in the organization of a set of measures aimed at their prevention in the postflood period in the Far-Eastern region. METHODS: The monitoring program includes children aged 2 to 5 years in the amount of 4988 with risk factors for pneumococcal infection. The pneumococcal conjugate vaccine Prevenar-13 was used for immunization. Data on the incidence of ARVI and pneumonia in children in pre- and postvaccination periods were to be recorded. The indicators and special criteria were used to assess the effectiveness of vaccination. To study the circulation of serovariants of pneumococcus in inflammatory diseases of the respiratory tract and nasopharyngeal carrier, bacteriological and molecular genetic methods (RT-PCR in the mode of multiprime detection) were used. RESULTS: Differences in the frequency and range of serovariants of circulating isolates of pneumococcus in the postvaccinal period and in unvaccinated children, elimination of a number of serotypes, and appearance of circulation of nonvaccinated strains were revealed. The incidence of acute respiratory diseases and pneumonia among the vaccinated population for 2 years in the region decreased by 2.5 times. The coefficient of effectiveness of vaccination according to the indicator of morbidity of children with pneumonia reaches 75-100% with direct dependence on the age of children (r=0.98). CONCLUSION: Comparative statistical analysis revealed a high degree of effectiveness of regional programs with the methods of immunoprophylaxis of pneumococcal infections.

Eremomycin pyrrolidide: a novel semisynthetic glycopeptide with improved chemotherapeutic properties.

PURPOSE: Development of new semisynthetic glycopeptides with improved antibacterial efficacy and reduced pseudoallergic reactions. METHODS: Semisynthetic glycopeptides 3-6 were synthesized from vancomycin (1) or eremomycin (2) by the condensation with pyrrolidine or piperidine. The minimum inhibitory concentration (MIC) for the new derivatives was measured by the broth micro-dilution method on a panel of clinical isolates of Staphylococcus and Enterococcus. Acute toxicity (50% lethal dose, maximum tolerated doses), antibacterial efficacy on model of systemic bacterial infection with S. aureus and pseudoallergic inflammatory reaction (on concanavalin A) of eremomycin pyrrolidide (5) were evaluated in mice according to standard procedures. RESULTS: The eremomycin pyrrolidide (5) was the most active compound and showed a high activity against Gram-positive bacteria: vancomycin-susceptible staphylococci and enterococci (minimum inhibitory concentrations [MICs] 0.13-0.25 mg/L), as well as vancomycin-intermediate resistant Staphylococcus aureus (MICs 1 mg/L). Antimicrobial susceptibility tested on a panel of 676 isolates showed that 5 had similar activity for the genera Staphylococcus and Enterococcus with MIC90=0.5 mg/L, while vancomycin had MIC90=1-2 mg/L. The number of resistant strains of Enterococcus faecium (vancomycin-resistant enterococci) (MIC =64 mg/L) with this value was 7 (8%) for vancomycin (1) and 0 for the compound 5. In vivo comparative studies in a mouse model of systemic bacterial infection with S. aureus demonstrated that the efficacy of 5 was notably higher than that of the original antibiotics 1 and 2. In contrast to 1, compound 5 did not induce pseudoallergic inflammatory reaction (on concanavalin A). CONCLUSION: The new semisynthetic derivative eremomycin pyrrolidide (5) has high activity against staphylococci and enterococci including vancomycin-resistant strains. Compound 5 has a higher efficacy in a model of staphylococcal sepsis than vancomycin (1) or eremomycin (2). In striking contrast to natural antibiotics, the novel derivative 5 does not induce a pseudoallergic inflammatory reaction to concanavalin A and therefore has no histamine release activity. These results indicate the advantages of a new semisynthetic glycopeptide antibiotic eremomycin pyrrolidide (5) which may be a prospective antimicrobial agent for further pre-clinical and clinical evaluations.

Epidemiology and Genetic Diversity of Colistin Nonsusceptible Nosocomial Acinetobacter baumannii Strains from Russia for 2013-2014.

A high level of resistance to carbapenems in Acinetobacter baumannii strains severely limits therapeutic possibilities. Colistin is the last resort drug against such strains, although the cases of resistance to this drug have become more frequent. This article presents the epidemiological features and genetic diversity of colistin nonsusceptible A. baumannii strains collected as part of a national multicenter epidemiological study of the antibiotic resistance of pathogens of nosocomial infections (MARATHON), which was conducted in 2013-2014 in Russia. A total of 527 A. baumannii isolates were collected, 10 (1.9%) of which were nonsusceptible to colistin. The majority of nonsusceptible A. baumannii isolates to colistin showed resistance to carbapenems and had the genes of the acquired OXA-40-like carbapenemases (n = 6). In one case, a combination of OXA-23-like + OXA-40-like (n = 1) genes was identified. One strain had the multidrug-resistant (MDR) phenotype, 6 isolates had extensively drug-resistant (XDR) phenotype, and 3 isolates had pandrug-resistant (PDR) phenotype. Among the colistin nonsusceptible A. baumannii isolates, 6 individual genotypes were identified, most of which belonged to successful international clones (CC92OXF/CC2PAS, n = 4; CC944OXF/ST78PAS, n = 4; CC109OXF/CC1PAS, n = 1).

In Vitro Activity of Oral Antimicrobial Agents against Pathogens Associated with Community-Acquired Upper Respiratory Tract and Urinary Tract Infections: A Five Country Surveillance Study.

INTRODUCTION: Bacterial infections that cause community-acquired urinary tract infections (CA-UTI) and upper respiratory tract infections (CA-URTI) are most frequently treated empirically. However, an increase in antimicrobial resistance has become a problem when treating outpatients. METHODS: This study determined the in vitro activities of oral antibiotics among 1501 pathogens from outpatients with CA-UTI and CA-URTI in medical centers during 2012 and 2013 from Argentina, Mexico, Venezuela, Russia, and the Philippines. Minimal inhibitory concentrations (MICs) were determined using broth microdilution and susceptibility defined by Clinical Laboratory Standards Institute (CLSI) and European Committee for Antimicrobial Susceptibility Testing (EUCAST) criteria. RESULTS: Ceftibuten (MIC50, ≤0.25 mg/L) was more potent in vitro compared to other ß-lactams against Enterobacteriaceae from CA-UTI. Susceptibility to fluoroquinolones using CLSI criteria varied: Argentina and Mexico (50%), the Philippines (60%), Venezuela (70%), and Russia (80%). Fosfomycin susceptibility was >90% against Enterobacteriaceae in each country. Susceptibility among Enterobacteriaceae to trimethoprim-sulfamethoxazole was 30.6-75.6% and nitrofurantoin susceptibility also varied among the countries and was higher when EUCAST breakpoints were applied (65->90%) compared to CLSI (52-84%). All Haemophilus influenzae isolates from CA-URTI were susceptible to ceftibuten, cefixime, cefpodoxime, and cefuroxime using CLSI breakpoint criteria. EUCAST criteria produced intermediate and resistant MIC values for these oral cephalosporins. Country-specific susceptibility variation for fluoroquinolones, macrolides, and trimethoprim-sulfamethoxazole was observed among Streptococcus pneumoniae and Streptococcus pyogenes from CA-URTI. CONCLUSION: This study demonstrated that antimicrobial susceptibility patterns varied in the five countries investigated among pathogens from CA-UTI and CA-URTI. FUNDING: Merck & Co. Inc., Kenilworth, New Jersey, USA.

Long-term dissemination of CTX-M-5-producing hypermutable Salmonella enterica serovar typhimurium sequence type 328 strains in Russia, Belarus, and Kazakhstan.

In this paper, we present evidence of long-term circulation of cefotaxime-resistant clonally related Salmonella enterica serovar Typhimurium strains over a broad geographic area. The genetic relatedness of 88 isolates collected from multiple outbreaks and sporadic cases of nosocomial salmonellosis in various parts of Russia, Belarus, and Kazakhstan from 1996 to 2009 was established by multilocus tandem-repeat analysis (MLVA) and multilocus sequence typing (MLST). The isolates belong to sequence type 328 (ST328) and produce CTX-M-5 ß-lactamase, whose gene is carried by highly related non-self-conjugative but mobilizable plasmids. Resistance to nalidixic acid and low-level resistance to ciprofloxacin is present in 37 (42%) of the isolates and in all cases is determined by various single point mutations in the gyrA gene quinolone resistance-determining region (QRDR). Isolates of the described clonal group exhibit a hypermutable phenotype that probably facilitates independent acquisition of quinolone resistance mutations.

Spread of extensively resistant VIM-2-positive ST235 Pseudomonas aeruginosa in Belarus, Kazakhstan, and Russia: a longitudinal epidemiological and clinical study.

BACKGROUND: Multidrug-resistant and extensively-drug-resistant Pseudomonas aeruginosa are increasing therapeutic challenges worldwide. We did a longitudinal epidemiological and clinical study of extensively-drug-resistant P aeruginosa in Belarus, Kazakhstan, and Russia. METHODS: The study was done in three prospectively defined phases: Jan 1, 2002-Dec 31, 2004; Jan 1, 2006-Dec 31, 2007; and Jan 1, 2008-Dec 31, 2010. The first two phases were in Russia only. All consecutive, non-duplicate, nosocomial isolates and case report forms were sent to the coordinating centre (Institute of Antimicrobial Chemotherapy, Smolensk, Russia), where species reidentification, susceptibility testing, and molecular typing of isolates were done. We did susceptibility testing by agar dilution. The presence of metallo-ß-lactamase (MBL) genes was established by PCR and sequencing, and class 1 integrons containing MBL gene cassettes were analysed by the PCR restriction fragment length polymorphism approach. Strain relatedness was analysed by multiple loci variable-number tandem-repeat (VNTR) analysis (at six VNTR loci) and multilocus sequence typing. RESULTS: In 2002-04, 628 of 1053 P aeruginosa isolates were insusceptible to carbapenems and 47 (4.5%) possessed MBLs. In 2006-07, 584 of 787 isolates were insusceptible to carbapenems and 160 (20.3%) possessed MBLs. In 2008-10, 1238 of 1643 Russian P aeruginosa isolates were insusceptible to carbapenems and 471 (28.7%) possessed MBLs. Additionally, the 32 P aeruginosa isolates from Belarus and Kazakhstan were all carbapenem insusceptible and all possessed MBLs. More than 96% of MBL-positive P aeruginosa isolates were resistant to all antibiotics except colistin (ie, extensively drug resistant), and, in 2010, 5·9% were resistant to colistin. 685 (96.5%) of 710 MBL-positive P aeruginosa belonged to ST235. bla(VIM-2) genes were detected in 707 (99.6%) of 710 MBL-positive isolates. INTERPRETATION: Extensively-drug-resistant ST235 P aeruginosa has rapidly spread throughout Russia and into Belarus and Kazakhstan via clonal dissemination. Increases in the use of colistin will probably result in further spread of ST235 P aeruginosa resistant to all drugs. FUNDING: HEFC, Ministry of Health of the Russian Federation, Government of the Republic of Belarus, Government of the Republic of Kazakhstan, European Union, Medical Research Council UK-Canada partnership.

Estimating the burden of pneumococcal pneumonia among adults: a systematic review and meta-analysis of diagnostic techniques.

BACKGROUND: Pneumococcal pneumonia causes significant morbidity and mortality among adults. Given limitations of diagnostic tests for non-bacteremic pneumococcal pneumonia, most studies report the incidence of bacteremic or invasive pneumococcal disease (IPD), and thus, grossly underestimate the pneumococcal pneumonia burden. We aimed to develop a conceptual and quantitative strategy to estimate the non-bacteremic disease burden among adults with community-acquired pneumonia (CAP) using systematic study methods and the availability of a urine antigen assay. METHODS AND FINDINGS: We performed a systematic literature review of studies providing information on the relative yield of various diagnostic assays (BinaxNOW® S. pneumoniae urine antigen test (UAT) with blood and/or sputum culture) in diagnosing pneumococcal pneumonia. We estimated the proportion of pneumococcal pneumonia that is bacteremic, the proportion of CAP attributable to pneumococcus, and the additional contribution of the Binax UAT beyond conventional diagnostic techniques, using random effects meta-analytic methods and bootstrapping. We included 35 studies in the analysis, predominantly from developed countries. The estimated proportion of pneumococcal pneumonia that is bacteremic was 24.8% (95% CI: 21.3%, 28.9%). The estimated proportion of CAP attributable to pneumococcus was 27.3% (95% CI: 23.9%, 31.1%). The Binax UAT diagnosed an additional 11.4% (95% CI: 9.6, 13.6%) of CAP beyond conventional techniques. We were limited by the fact that not all patients underwent all diagnostic tests and by the sensitivity and specificity of the diagnostic tests themselves. We address these resulting biases and provide a range of plausible values in order to estimate the burden of pneumococcal pneumonia among adults. CONCLUSIONS: Estimating the adult burden of pneumococcal disease from bacteremic pneumococcal pneumonia data alone significantly underestimates the true burden of disease in adults. For every case of bacteremic pneumococcal pneumonia, we estimate that there are at least 3 additional cases of non-bacteremic pneumococcal pneumonia.

Comparative activity of ceftobiprole against Gram-positive and Gram-negative isolates from Europe and the Middle East: the CLASS study.

OBJECTIVES: to assess the in vitro activity of ceftobiprole and comparators against a recent collection of Gram-positive and Gram-negative pathogens, in order to detect potential changes in susceptibility patterns, and to evaluate the Etest assay for ceftobiprole susceptibility testing. METHODS: contemporary Gram-positive and Gram-negative isolates (excluding extended-spectrum ß-lactamase-producing isolates) from across Europe and the Middle East were collected, and their susceptibility to ceftobiprole, vancomycin, teicoplanin, linezolid, ceftazidime and cefepime was assessed using the Etest method. Quality testing [using Etest and broth microdilution (BMD)] was conducted at a central reference laboratory. RESULTS: some 5041 Gram-positive and 4026 Gram-negative isolates were included. Against Gram-positive isolates overall, ceftobiprole had the lowest MIC50 (0.5 mg/L), compared with 1 mg/L for its comparators (vancomycin, teicoplanin and linezolid). Against methicillin-resistant Staphylococcus aureus, all four agents had a similar MIC90 (2 mg/L), but ceftobiprole had a 4-fold better MIC90 (0.5 mg/L) against methicillin-susceptible strains. Only 38 Gram-positive isolates were confirmed as ceftobiprole resistant. Among Gram-negative strains, 86.9%, 91.7% and 95.2% were susceptible to ceftobiprole, ceftazidime and cefepime, respectively. Pseudomonas aeruginosa was less susceptible to all three antimicrobials than any other Gram-negative pathogen. There was generally good agreement between local Etest results and those obtained at the reference laboratory (for ceftobiprole: 86.8% with Gram-negatives; and 94.7% with Gram-positives), as well as between results obtained by BMD and Etest methods (for ceftobiprole: 98.2% with Gram-negatives; and 98.4% with Gram-positives). CONCLUSIONS: ceftobiprole exhibits in vitro activity against a wide range of Gram-positive and Gram-negative pathogens, including multidrug-resistant strains. No changes in its known susceptibility profile were identified.

Clinical consensus conference: survey on Gram-positive bloodstream infections with a focus on Staphylococcus aureus.

The increased incidence over the past decade of bloodstream infections (BSIs) caused by gram-positive bacteria, particularly methicillin-resistant Staphylococcus aureus, highlights the critical need for a consistent approach to therapy. However, there is currently no international consensus on the diagnosis and management of gram-positive BSIs. The Clinical Consensus Conference on Gram-Positive Bloodstream Infections was convened as a session at the 9th International Symposium on Modern Concepts in Endocarditis and Cardiovascular Infections held in 2007. Participants discussed various aspects of the practical treatment of patients who present with gram-positive BSI, including therapeutic options for patients with BSIs of undefined origin, the selection of appropriate empirical therapy, and treatment of complicated and uncomplicated BSIs. The opinions of participants about these key issues are reflected in this article.

Management of nonsevere pneumonia in military trainees with the urinary antigen test for Streptococcus pneumoniae: an innovative approach to targeted therapy.

BACKGROUND: The drug of choice for treatment of Streptococcus pneumoniae infection is generally a penicillin (including amoxicillin). Targeted therapy is, however, rarely used, because results of definitive diagnostic tests for pneumonia are not available for several days. Thus, broad-spectrum antibiotics are used for empirical treatment of pneumonia to cover both typical and atypical pathogens. Our purpose was to assess the usefulness of a strategy of targeted antimicrobial therapy based on the results of a rapid urinary antigen test for S. pneumoniae. METHODS: Military trainees with pneumonia were prospectively assigned to 2 groups: patients with positive urinary antigen test results who were treated with amoxicillin (1000 mg 3 times per day), and patients with negative urinary antigen test results who were treated with clarithromycin (500 mg 2 times per day). The duration of therapy was 5-10 days for both groups. RESULTS: A total of 219 evaluable patients were enrolled in the study. The most common causes of pneumonia were S. pneumoniae, Chlamydia pneumoniae, and Mycoplasma pneumoniae. Patients with positive urinary antigen test results had illness of greater severity at the time of study entry. Twenty-two percent of patients had positive urinary antigen test results (i.e., the amoxicillin group), and 78% had negative urinary antigen test results (i.e., the clarithromycin group). The clinical success rates were 94% for the clarithromycin group and 90% for the amoxicillin group (P = not significant). None of the patients who were classified as having treatment failure died. Resolution of clinical manifestations was slower for patients with pneumococcal pneumonia defined by a positive urinary antigen test result. CONCLUSIONS: The urine antigen test allowed targeted use of a penicillin (amoxicillin) for young immunocompetent individuals with nonsevere, community-acquired pneumonia. Clarithromycin was highly effective against both S. pneumoniae pneumonia and pneumonia due to atypical pathogens.

Antistreptococcal activity of telithromycin compared with seven other drugs in relation to macrolide resistance mechanisms in Russia.

The susceptibilities of 468 recent Russian clinical Streptococcus pneumoniae isolates and 600 Streptococcus pyogenes isolates, from 14 centers in Russia, to telithromycin, erythromycin, azithromycin, clarithromycin, clindamycin, levofloxacin, quinupristin-dalfopristin, and penicillin G were tested. Penicillin-nonsusceptible S. pneumoniae strains were rare except in Siberia, where their prevalence rate was 13.5%: most were penicillin intermediate, but for three strains (two from Smolensk and one from Novosibirsk) the MICs of penicillin G were 4 or 8 micro g/ml. Overall, 2.5% of S. pneumoniae isolates were resistant to erythromycin. Efflux was the prevalent resistance mechanism (five strains; 41.7%), followed by ribosomal methylation encoded by constitutive erm(B), which was found in four isolates. Ribosomal mutation was the mechanism of macrolide resistance in three isolates; one erythromycin-resistant S. pneumoniae isolate had an A2059G mutation in 23S rRNA, and two isolates had substitution of GTG by TPS at positions 69 to 71 in ribosomal protein L4. All S. pyogenes isolates were susceptible to penicillin, and 11% were erythromycin resistant. Ribosomal methylation was the most common resistance mechanism for S. pyogenes (89.4%). These methylases were encoded by erm(A) [subclass erm(TR)] genes, and their expression was inducible in 96.6% of isolates. The rest of the erythromycin-resistant Russian S. pyogenes isolates (7.6%) had an efflux resistance mechanism. Telithromycin was active against 100% of pneumococci and 99.2% of S. pyogenes, and levofloxacin and quinupristin-dalfopristin were active against all isolates of both species.

International external quality assessment scheme for the laboratory diagnosis of diphtheria.

An international external quality assessment (EQA) scheme has been established so as to evaluate the proficiency of specialist, national diphtheria reference laboratories in the laboratory diagnosis of diphtheria. Six simulated clinical specimens were freeze-dried and distributed to 23 participants in 20 countries. Participants were asked to isolate, identify and perform toxigenicity testing on any corynebacteria present and to complete a simple questionnaire describing the procedures and reagents used. Only three laboratories obtained correct biochemical and toxigenicity results for all six specimens. The majority of laboratories performed better with toxigenicity testing than with the biochemical identification. Of concern were the results from three laboratories that failed to isolate any corynebacteria from four or more of the specimens. In one centre this was shown to be due to lack of availability of 'in-date' media and, in the other two, was presumed to be due to lack of experience in primary laboratory diagnostics for this organism. It is essential that countries, globally, maintain awareness and laboratory capabilities in this specialised area of microbiology. EQA is an invaluable process, which enables laboratories to monitor, evaluate and improve their own performance in such areas.