Csaba Ködmön

First Worldwide Proficiency Study on Variable-Number Tandem-Repeat Typing of Mycobacterium tuberculosis Complex Strains

ABSTRACT Although variable-number tandem-repeat (VNTR) typing has gained recognition as the new standard for the DNA fingerprinting of Mycobacterium tuberculosis complex (MTBC) isolates, external quality control programs have not yet been developed. Therefore, we organized the first multicenter proficiency study on 24-locus VNTR typing. Sets of 30 DNAs of MTBC strains, including 10 duplicate DNA samples, were distributed among 37 participating laboratories in 30 different countries worldwide. Twenty-four laboratories used an in-house-adapted method with fragment sizing by gel electrophoresis or an automated DNA analyzer, nine laboratories used a commercially available kit, and four laboratories used other methods. The intra- and interlaboratory reproducibilities of VNTR typing varied from 0% to 100%, with averages of 72% and 60%, respectively. Twenty of the 37 laboratories failed to amplify particular VNTR loci; if these missing results were ignored, the number of laboratories with 100% interlaboratory reproducibility increased from 1 to 5. The average interlaboratory reproducibility of VNTR typing using a commercial kit was better (88%) than that of in-house-adapted methods using a DNA analyzer (70%) or gel electrophoresis (50%). Eleven laboratories using in-house-adapted manual typing or automated typing scored inter- and intralaboratory reproducibilities of 80% or higher, which suggests that these approaches can be used in a reliable way. In conclusion, this first multicenter study has documented the worldwide quality of VNTR typing of MTBC strains and highlights the importance of international quality control to improve genotyping in the future.

Inventory study of non-tuberculous mycobacteria in the European Union

BackgroundSince non-tuberculous mycobacteria (NTM) disease is not notifiable in most European Union (EU) and European Economic Area (EEA) countries, the epidemiological situation of the >150 NTM species is largely unknown. We aimed to collect data on the frequency of NTM detection and NTM species types in EU/EEA countries.MethodsOfficially nominated national tuberculosis reference laboratories of all EU/EEA countries were asked to provide information on: laboratory routines for detection and identification of NTM, including drug sensitivity testing (DST) methods; data on the number and type of NTM species identified; coverage and completeness of the provided data on NTM; type and number of human specimens tested for NTM; and number of specimens tested for Mycobacterium tuberculosis complex and NTM. This information was summarized and the main results are described.ResultsIn total, 99 different NTM species were identified with M. avium, M. gordonae, M. xenopi , M. intracellulare, and M. fortuitum identified most frequently. Seven percent of the NTM species could not be identified. NTM was cultured from between 0.4-2.0% of the specimens (data from four countries). The laboratories use culturing methods optimised for M. tuberculosis complex. Identification is mainly carried out by a commercial line probe assay supplemented with sequencing. Most laboratories carried out DST for rapid growers and only at the explicit clinical request for slow growers.ConclusionIt is likely that the prevalence of NTM is underestimated because diagnostic procedures are not optimized specifically for NTM and isolates may not be referred to the national reference laboratory for identification. Due to the diagnostic challenges and the need to establish the clinical relevance of NTM, we recommend that countries should concentrate detection and identification in only few laboratories.

The epidemiology of invasive meningococcal disease in EU/EEA countries, 2004–2014

BACKGROUND Invasive meningococcal disease (IMD) is a major cause of bacterial meningitis and septicaemia although infection by some serogroups may be prevented through vaccination. We aimed to describe the epidemiology of IMD in EU/EEA countries during 2004-2014 to monitor serogroup- and age-specific trends, and compare country trends by the period of meningococcal C conjugate (MCC) vaccine introduction. METHODS We analysed IMD surveillance data by age, gender, serogroup, country and outcome. We estimated the percentage change in annual notification rate (NR), using linear regression analysis of the log of the annual NR. We grouped countries by the year they introduced MCC vaccination into their routine immunisation programmes. RESULTS The overall NR was 0.9/100 000 population, and decreased 6.6% (95%CI: -8.0%;-5.1%) annually. Infants had the highest NR (16.0/100 000), and there were decreasing trends in all age groups <50years. Serogroup B (SgB) caused 74% of all cases, and the majority of cases in all age groups. There were decreasing trends in SgB and serogroup C (SgC) and an increasing trend in serogroup Y. Countries that introduced MCC vaccination before, and between 2004 and 2014, had decreasing trends in NR of SgC, but not countries without routine MCC vaccination. CONCLUSIONS Our findings support evidence that routine MCC vaccination was the driving force behind the decreasing SgC trend. Vaccinating against SgB in the first year of life could help reduce the burden of IMD due to this serogroup. Changing serogroup-specific NR trends highlight the need for high-quality surveillance data to accurately assess the changing epidemiology of IMD, the effectiveness and impact of implemented vaccines, and the need for future vaccines.

Molecular surveillance of multi- and extensively drug-resistant tuberculosis transmission in the European Union from 2003 to 2011.

The European Centre for Disease Prevention and Control (ECDC) initiated a project on the molecular surveillance of multi- and extensively drug-resistant tuberculosis (MDR-/XDR-TB) transmission in the European Union (EU) in the period from 2009 to 2011. In total, 2,092 variable number of tandem repeat (VNTR) patterns of MDR-/XDR-TB Mycobacterium tuberculosis isolates were collected, originating from 24 different countries in the period 2003 to 2011. Of the collected VNTR patterns, 45% (n=941) could be assigned to one of the 79 European multiple-country molecular fingerprint clusters and 50% of those (n=470) belonged to one extremely large cluster caused by Beijing strains of one genotype. We conclude that international transmission of MDR-/XDR-TB plays an important role in the EU, especially in the eastern part, and is significantly related to the spread of one strain or clone of the Beijing genotype. Implementation of international cluster investigation in EU countries should reveal underlying factors of transmission, and show how TB control can be improved regarding case finding, contact tracing, infection control and treatment in order to prevent further spread of MDR-/XDR-TB in the EU.

Migration-related tuberculosis: epidemiology and characteristics of tuberculosis cases originating outside the European Union and European Economic Area, 2007 to 2013.

Migrants arriving from high tuberculosis (TB)-incidence countries may pose a significant challenge to TB control programmes in the host country. TB surveillance data for 2007-2013 submitted to the European Surveillance System were analysed. Notified TB cases were stratified by origin and reporting country. The contribution of migrant TB cases to the TB epidemiology in EU/EEA countries was analysed. Migrant TB cases accounted for 17.4% (n = 92,039) of all TB cases reported in the EU/EEA in 2007-2013, continuously increasing from 13.6% in 2007 to 21.8% in 2013. Of 91,925 migrant cases with known country of origin, 29.3% were from the Eastern Mediterranean, 23.0% from south-east Asia, 21.4% from Africa, 13.4% from the World Health Organization European Region (excluding EU/EEA), and 12.9% from other regions. Of 46,499 migrant cases with known drug-susceptibility test results, 2.9% had multidrug-resistant TB, mainly (51.7%) originating from the European Region. The increasing contribution of TB in migrants from outside the EU/EEA to the TB burden in the EU/EEA is mainly due to a decrease in native TB cases. Especially in countries with a high proportion of TB cases in non-EU/EEA migrants, targeted prevention and control initiatives may be needed to progress towards TB elimination.

Tuberculosis treatment outcome monitoring in European Union countries: systematic review

Treatment success measured by treatment outcome monitoring (TOM) is a key programmatic output of tuberculosis (TB) control programmes. We performed a systematic literature review on national-level TOM in the 30 European Union (EU)/European Economic Areas (EEA) countries to summarise methods used to collect and report data on TOM. Online reference bibliographic databases PubMed/MEDLINE and EMBASE were searched to identify relevant indexed and non-indexed literature published between January 2000 and August 2010. The search strategy resulted in 615 potentially relevant indexed citations, of which 27 full-text national studies (79 data sets) were included for final analysis. The selected studies were performed in 10 EU/EEA countries and gave a fragmented impression of TOM in the EU/EEA. Publication year, study period, sample size, databases, definitions, variables, patient and outcome categories, and population subgroups varied widely, portraying a very heterogeneous picture. This review confirmed previous reports of considerable heterogeneity in publications of TOM results across EU/EEA countries. PubMed/MEDLINE and EMBASE indexed studies are not a suitable instrument to measure representative TOM results for the 30 EU/EEA countries. Uniform and complete reporting to the centralised European Surveillance System will produce the most timely and reliable results of TB treatment outcomes in the EU/EEA.

Eligibility for shorter treatment of multidrug-resistant tuberculosis in the European Union

In May 2016, the World Health Organization (WHO) updated its treatment guidelines for drug-resistant tuberculosis (TB) and included a recommendation for the use of the shorter multidrug-resistant (MDR) TB regimen [1]. The WHO update is based on information provided by observational studies coordinated by the International Union Against Tuberculosis and Lung Disease, Médecins sans Frontières and the Damien Foundation which showed that the shorter treatment regimen resulted in a higher likelihood of treatment success compared to the longer conventional treatment in the study settings [2–5]. The shorter MDR-TB regimen consists of a standardised treatment course lasting 9–12 months and includes kanamycin, moxifloxacin, prothionamide, clofazimine, pyrazinamide, isoniazid and ethambutol. It is recommended for patients with rifampicin-resistant (RR) or MDR-TB (cases with TB bacilli resistant to at least isoniazid and rifampicin) who have not been treated previously with second-line drugs and in whom resistance to fluoroquinolones and second-line injectable agents has been excluded or is considered highly unlikely. It is not recommended for pregnant women and patients with extrapulmonary TB. In the European Union, 11% of all MDR-TB cases are eligible for the shorter MDR-TB regimen http://ow.ly/kukg306AcFl

Second worldwide proficiency study on variable number of tandem repeats typing of Mycobacterium tuberculosis complex

BACKGROUND The quality of variable number of tandem repeats (VNTR) typing of Mycobacterium tuberculosis was first investigated in 2009 in 37 laboratories worldwide. The results revealed an inter- and intra-laboratory reproducibility of respectively 60% and 72%. These data spurred an improvement in laboratory-specific assays and global standardisation of VNTR typing. OBJECTIVE To measure the effects of the technical improvements and increased standardisation, a test panel consisting of 30 M. tuberculosis complex DNA samples was distributed for VNTR typing in 41 participating laboratories from 36 countries. RESULTS The inter- and intra-laboratory reproducibility increased overall to respectively 78% and 88%. The 33 laboratories that participated in both the first and second proficiency studies improved their inter- and intra-laboratory reproducibility from 62% and 72% to respectively 79% and 88%. The largest improvement in reproducibility was detected in 10 laboratories that use an in-house polymerase chain reaction technique and perform amplicon sizing using gel electrophoresis. Detailed error analysis revealed a reduction in the number of systematic errors, sample exchange events and non-amplifiable loci. CONCLUSION This second worldwide proficiency study indicates a substantial increase in the reproducibility of VNTR typing of M. tuberculosis. This will contribute to a more meaningful interpretation of molecular epidemiological and phylogenetic studies on the M. tuberculosis complex.

The effect of migration within the European Union/European Economic Area on the distribution of tuberculosis, 2007 to 2013.

Immigration from tuberculosis (TB) high-incidence countries is known to contribute notably to the TB burden in low-incidence countries. However, the effect of migration enabled by the free movement of persons within the European Union (EU)/European Economic Area (EEA) on TB notification has not been analysed. We analysed TB surveillance data from 29 EU/EEA countries submitted for the years 2007-2013 to The European Surveillance System. We used place of birth and nationality as proxy indicators for native, other EU/EEA and non-EU/EEA origin of the TB cases and analysed the characteristics of the subgroups by origin. From 2007-2013, a total of 527,467 TB cases were reported, of which 129,781 (24.6%) were of foreign origin including 12,566 (2.4%) originating from EU/EEA countries other than the reporting country. The countries reporting most TB cases originating from other EU/EEA countries were Germany and Italy, and the largest proportion of TB cases in individuals came from Poland (n=1,562) and Romania (n=6,285). At EU/EEA level only a small proportion of foreign TB cases originated from other EU/EEA countries, however, the uneven distribution of this presumed importation may pose a challenge to TB programmes in some countries.

External Quality Assessment for Tuberculosis Diagnosis and Drug Resistance in the European Union: A Five Year Multicentre Implementation Study.

Background External quality assurance (EQA) systems are essential to ensure accurate diagnosis of TB and drug-resistant TB. The implementation of EQA through organising regular EQA rounds and identification of training needs is one of the key activities of the European TB reference laboratory network (ERLTB-Net). The aim of this study was to analyse the results of the EQA rounds in a systematic manner and to identify potential benefits as well as common problems encountered by the participants. Methods The ERLTB-Net developed seven EQA modules to test laboratories’ proficiency for TB detection and drug susceptibility testing using both conventional and rapid molecular tools. All National TB Reference laboratories in the European Union and European Economic Area (EU/EEA) Member States were invited to participate in the EQA scheme. Results A total of 32 National TB Reference laboratories participated in six EQA rounds conducted in 2010–2014. The participation rate ranged from 52.9% - 94.1% over different modules and rounds. Overall, laboratories demonstrated very good proficiency proving their ability to diagnose TB and drug-resistant TB with high accuracy in a timely manner. A small number of laboratories encountered problems with identification of specific Non-tuberculous Mycobacteria (NTMs) (N = 5) and drug susceptibility testing to Pyrazinamide, Amikacin, Capreomycin, and Ethambutol (N = 4). Conclusions The European TB Reference laboratories showed a steady and high level of performance in the six EQA rounds. A network such as ERLTB-Net can be instrumental in developing and implementing EQA and in establishing collaboration between laboratories to improve the diagnosis of TB in the EU/EEA.