Jeroen W. Den Boer

A large outbreak of Legionnaires' disease at a flower show, the Netherlands, 1999.

In 1999, an outbreak of Legionnaires’ disease affected many visitors to a flower show in the Netherlands. To identify the source of the outbreak, we performed an environmental investigation, as well as a case-control study among visitors and a serologic cohort study among exhibitors to measure exposure to possible sources. Of 77,061 visitors, 188 became ill (133 confirmed and 55 probable cases), for an attack rate of 0.23% for visitors and 0.61% for exhibitors. Two whirlpool spas in halls 3 and 4 of the exhibition and a sprinkler in hall 8 were culture positive for Legionella pneumophila. One of three genotypes found in both whirlpool spas was identical to the isolates from 28 of 29 culture-positive patients. Persons who paused at the whirlpool spa in hall 3 were at increased risk for becoming ill. This study illustrates that whirlpool spas may be an important health hazard if disinfection fails.

Legionnaires' Disease at a Dutch Flower Show: Prognostic Factors and Impact of Therapy

After a large outbreak of Legionnaires’ disease in the Netherlands, we determined risk factors for intensive care unit (ICU) admission and death and the impact of adequate therapy on ICU-free survival among 141 hospitalized patients. Overall mortality rate was 13%, and ICU mortality rate was 36%. Smoking, temperature >38.5°C, and bilateral infiltrates shown on chest x-ray were independent risk factors for ICU admission or death (all p<0.05). Starting adequate therapy within 24 hours after admission resulted in a higher ICU-free survival rate compared to therapy initiation after 24 hours: 78% versus 54%, respectively (p=0.005). However, delay in providing therapy to patients with urinary antigen tests with negative results did not influence outcome. These data suggest that by using the urinary antigen test on admission a more tailored approach to patients with community-acquired pneumonia may be applied.

Wild-type MIC distribution and epidemiological cut-off values in clinical Legionella pneumophila serogroup 1 isolates.

OBJECTIVES The purpose of this study was to establish wild-type (WT) distributions and determine the epidemiological cut-off values (ECOFF) in clinical L. pneumophila serogroup 1 isolates for 10 antimicrobials commonly used for the treatment of Legionella infections using a method feasible in a routine clinical laboratory. METHODS MICs of 183 clinical L. pneumophila serogroup 1 isolates, collected as part of an outbreak detection program, were tested using E-test methodology on buffered charcoal yeast extract agar supplemented with α-ketoglutarate (BCYE-α). The MICs were read after 2 days of incubation at 35 °C with increased humidity and without CO(2). ECOFFs were determined according to EUCAST methodology and expressed as WT ≤ X mg/L. RESULTS All antimicrobials showed a WT distribution, although the width varied from 2 two-fold dilutions to 8 dilutions, depending on antibiotic class. The ECOFFs determined were 1.0 mg/L for ciprofloxacin, 0.50 mg/L for levofloxacin, 1.0 mg/L for moxifloxacin, 1.0 mg/L for erythromycin, 1.0 mg/L for azithromycin, 0.50 mg/L for clarithromycin, 1.0 mg/L for cefotaxime, 0.032 mg/L for rifampicin, 16 mg/L for tigecycline, and 8 mg/L for doxycycline. CONCLUSION All isolates were inhibited by low concentrations of the fluoroquinolones and macrolides tested, with somewhat higher MICs for the fluoroquinolones. Rifampicin was found to be the most active against L. pneumophila isolates in vitro. These data can be used as a reference for the detection of resistance in clinical L. pneumophila isolates and as a setting of clinical breakpoints.

Legionnaires' disease and Pontiac fever after using a private outdoor whirlpool spa

Abstract Pontiac fever and Legionnaires’ disease are regarded as clinically and epidemiologically distinct diseases, caused by bacteria of the genus Legionella. Although several outbreaks of either Pontiac fever or Legionnaires’ disease have been reported, they are rarely seen simultaneously. In this report we describe such a simultaneous outbreak of Pontiac fever and Legionnaires’ disease that occurred in the Netherlands. In August 2009, 1 patient with Legionnaires’ disease and 3 patients with Pontiac fever, all from a single family, were reported to the Municipal Health Service. All family members had been exposed to the private whirlpool spa in the garden of their home. A sampling investigation by the Legionella Source Identification Unit (LSIU) showed that a sample from the whirlpool spa, as well as a sample from the garden shower and 2 samples from a garden hose were positive for Legionella pneumophila serogroup 1, and genotyping results indicated the AFLP-type 004 Lyon (ST47) to be present in these samples.

Isolation of ciprofloxacin-resistant Legionella pneumophila in a patient with severe pneumonia

Sir, Legionella species are responsible for 1%– 5% of cases of community-acquired pneumonia. Legionella pneumophila serogroup 1 (SG1) accounts for .90% of Legionnaires’ disease (LD) in North America and Europe and is the cause of significant mortality. The mortality rate among patients with L. pneumophila infections continues to be high, up to 26%. The antimicrobial agents most commonly used for treatment of LD are fluoroquinolones (e.g. ciprofloxacin or levofloxacin) and macrolides. In recent studies, we established wild-type distributions and determined the epidemiological cut-off values (ECOFFs) in clinical L. pneumophila SG1 isolates for 10 antimicrobials commonly used for the treatment of Legionella infections. A patient sought care at his general practitioner after several days of falling and body pains. On examination, the patient appeared ill and was sent to the emergency department of a nearby hospital. The initial chest radiograph demonstrated an infiltrate of the left lower lung field. The patient was admitted to the intensive care unit. Blood cultures were taken and antibiotic treatment was started with cefazolin and gentamicin. Urine was examined for the presence of Legionella antigens and when this test was reported positive, treatment was switched to 400 mg of ciprofloxacin intravenously twice daily. After initial improvement the clinical condition of the patient deteriorated, leading to intubation and mechanical ventilation. A new chest radiograph revealed a diffuse interstitial pneumonia. Bronchoalveolar lavage (BAL) was performed 4 days after treatment with ciprofloxacin was started and the patient slowly recovered. Eventually, culture of the BAL grew L. pneumophila SG1 after 4 days of ciprofloxacin treatment. After 10 days, the patient could be transferred to the ward. Therapy was then switched to 500 mg of clarithromycin orally twice daily. The patient’s further recovery was uncomplicated. The L. pneumophila SG1 strain was sent, as part of a national Legionella outbreak detection programme, to the reference laboratory for Legionella in Haarlem, The Netherlands. Susceptibility testing for ciprofloxacin was performed with Etest and an MIC value of ciprofloxacin of 2 mg/L was found. This value is outside the wild-type distribution range ECOFF1⁄41 mg/L as previously described and therefore potentially resistant. For sequencing of gyrA and gyrB (DNA gyrase) and parC and parE (topoisomerase IV) genes, extraction of L. pneumophila DNA was performed. The DNA extraction was performed by use of Qiagen’s BioRobot EZ1 (Hilden, Germany) according to the manufacturer’s instructions. The sequencing reaction was performed twice by using primer systems previously described for the L. pneumophila SG1 strain Paris. A comparative analysis of the obtained sequences was done using the published L. pneumophila genomes and data from the literature describing mutations in the quinolone resistance-determining region (QRDR) of type II topoisomerase of L. pneumophila by using DNAStar (WI, USA) software and the NCBI database. For control experiments, the wild-type strain MTZ OLDA and a spontaneous quinolone-resistant mutant of this strain were used. MTZ OLDA is an environmental isolate (L. pneumophila SG1), isolated from the water supply of a large building. A point mutation in the QRDR of the gyrA gene was identified and this mutation led to an amino acid exchange at position 83 (Escherichia coli numbering system). The result of this amino acid exchange is a change in ciprofloxacin susceptibility. Mutation at the same position (amino acid 83) has also been reported for other spontaneous quinolone-resistant mutants (Table 1). It is known that, in general, pathogens can become resistant during the course of a patient’s therapy and also induction of resistance upon exposure to antibiotics has been described. The origin of resistance in the clinical isolate is as yet unclear. There are two possibilities. The first is that the patient contracted an L. pneumophila SG1 strain with this point mutation from the environment. Alternatively, the mutation occurred during the Research letters

Pragati: an empowerment programme for female sex workers in Bangalore, India.

Objectives : To describe the effects of a broad empowerment programme among female sex workers (FSWs) in Bangalore,1 1Meaning ‘progress’ (in Kannada – an Indian language) India, which seeks to develop the capacities of these women to address the issues that threaten their lives and livelihoods. Design : This study is based on a comprehensive, on-going HIV-prevention and empowering programme, known as Pragati, which reaches out to approximately 10,000–12,000 FSWs in Bangalore each year. The programme has been designed in collaboration with the sex worker community and provides a personalised set of services, which include STI prevention and treatment services, crisis-response facilities, de-addiction services, and microfinance support all of which have been tailored to adequately fulfil each womans needs. During the period examined by this study, the programme reached out to 20,330 individual FSWs [median (IQR) age 28 (24–35) years]. The programmes personal records of the participating FSWs were used for this descriptive study. Results : Between 2005 and 2010, the number of participating FSWs increased from 2,307 to 13,392. These women intensified their contact with the programme over time: the number of programme contacts increased from 10,351 in 2005 to 167,709 in 2010. Furthermore, data on the effects of crisis-response facilities, de-addiction and microfinance services, condom distribution schemes, and STI diagnosis and treatment showed an accumulating involvement of the participating FSWs in these programme services. Conclusion : This programme, which focuses on social and economic empowerment among FSWs, is successful in reaching and involving the target population.

Reduction in STIs in an empowerment intervention programme for female sex workers in Bangalore, India: the Pragati programme

Background/Objective The Pragati programme is an on-going empowerment programme for female sex workers (FSWs) working and living in Bangalore, India. Pragati aims to reduce transmission of HIV and sexually transmitted infections (STIs) among FSWs. This study describes the STI incidence rate, contact rate, and condom use during follow-up years. Design Between April 2005 and November 2010, 20,330 FSWs participated in the programme. Outcome measures were programme exposure (number of contacts per person-year), STI incidence rate, and condom use. All analyses were stratified by year of follow-up. STIs were diagnosed by syndromic case management in either programme or referral clinics. We restricted our analyses to the period between April 2005 and July 2008 (when the majority of STIs were diagnosed in programme clinics), in order to minimise the possible influence of differences in STI diagnosis between clinic types. Results Results showed a significant increase of programme exposure (p-value for trend < 0.001) and a significant decrease in the STI incidence rate (p-value for trend < 0.001) over the follow-up time (between April 2005 and July 2008). Reported condom use at last paid sex increased from 77.6% in year 1 to 100% in year 4 of follow-up (p-value for trend < 0.001). Conclusion Our data seem to suggest that the Pragati programme had a positive effect on the STI incidence rate and condom use, possibly as a result of increased programme exposure. We recommend for future studies to invest more in the study design, type of data collection, and recording mechanisms before starting with an intervention. Incorporation of empowerment strategies as an approach in HIV prevention programmes can have a beneficial effect on the lives and livelihoods of FSWs.Background/Objective The Pragati programme is an on-going empowerment programme for female sex workers (FSWs) working and living in Bangalore, India. Pragati aims to reduce transmission of HIV and sexually transmitted infections (STIs) among FSWs. This study describes the STI incidence rate, contact rate, and condom use during follow-up years. Design Between April 2005 and November 2010, 20,330 FSWs participated in the programme. Outcome measures were programme exposure (number of contacts per person-year), STI incidence rate, and condom use. All analyses were stratified by year of follow-up. STIs were diagnosed by syndromic case management in either programme or referral clinics. We restricted our analyses to the period between April 2005 and July 2008 (when the majority of STIs were diagnosed in programme clinics), in order to minimise the possible influence of differences in STI diagnosis between clinic types. Results Results showed a significant increase of programme exposure (p-value for trend < 0.001) and a significant decrease in the STI incidence rate (p-value for trend < 0.001) over the follow-up time (between April 2005 and July 2008). Reported condom use at last paid sex increased from 77.6% in year 1 to 100% in year 4 of follow-up (p-value for trend < 0.001). Conclusion Our data seem to suggest that the Pragati programme had a positive effect on the STI incidence rate and condom use, possibly as a result of increased programme exposure. We recommend for future studies to invest more in the study design, type of data collection, and recording mechanisms before starting with an intervention. Incorporation of empowerment strategies as an approach in HIV prevention programmes can have a beneficial effect on the lives and livelihoods of FSWs.

Results from the National Legionella Outbreak Detection Program, the Netherlands, 2002–2012

This program has provided insights into the transmission, diagnosis, source investigation, and genotypic strain characteristics of the disease.

Comparative genome analysis of a large Dutch Legionella pneumophila strain collection identifies five markers highly correlated with clinical strains

BackgroundDiscrimination between clinical and environmental strains within many bacterial species is currently underexplored. Genomic analyses have clearly shown the enormous variability in genome composition between different strains of a bacterial species. In this study we have used Legionella pneumophila, the causative agent of Legionnaires disease, to search for genomic markers related to pathogenicity. During a large surveillance study in The Netherlands well-characterized patient-derived strains and environmental strains were collected. We have used a mixed-genome microarray to perform comparative-genome analysis of 257 strains from this collection.ResultsMicroarray analysis indicated that 480 DNA markers (out of in total 3360 markers) showed clear variation in presence between individual strains and these were therefore selected for further analysis. Unsupervised statistical analysis of these markers showed the enormous genomic variation within the species but did not show any correlation with a pathogenic phenotype. We therefore used supervised statistical analysis to identify discriminating markers. Genetic programming was used both to identify predictive markers and to define their interrelationships. A model consisting of five markers was developed that together correctly predicted 100% of the clinical strains and 69% of the environmental strains.ConclusionsA novel approach for identifying predictive markers enabling discrimination between clinical and environmental isolates of L. pneumophila is presented. Out of over 3000 possible markers, five were selected that together enabled correct prediction of all the clinical strains included in this study. This novel approach for identifying predictive markers can be applied to all bacterial species, allowing for better discrimination between strains well equipped to cause human disease and relatively harmless strains.

Legionnaires' disease after using an industrial pressure test pump: a case report

IntroductionLegionnaires’ disease is an acute pneumonia caused by inhalation or aspiration of aerosols contaminated with Legionella bacteria. The majority (>90%) of Legionnaires’ disease cases are caused by the species Legionella pneumophila, and about 85% more specifically by L. pneumophila serogroup 1 that can be detected by a fast and easy to perform urinary antigen test. Previously reported sources of infection include cooling towers, plumbing systems of hospitals, and whirlpool spas, but for the majority of cases of Legionnaires’ disease the source of infection remains unknown.Case presentationA 52-year-old Caucasian man was admitted to a Dutch hospital with pneumonia, where a culture of the available bronchial lavage was found positive for L. pneumophila serogroup 3, confirming the diagnosis of Legionnaires’ disease. An environmental investigation identified a manually operated pressure test pump at the metal processing company where he worked as the source of infection: the water sample from the pump contained 9·8×103 colony forming units/L L. pneumophila, and sequence-based typing showed the same sequence type (ST93) for both the clinical and environmental strains.ConclusionThis case shows that Legionnaires’ disease can be acquired by exposure to relatively rare sources that are not considered in regular control and prevention measures.