Legionella

Legionnaires’ disease (LD) treatment recommendations are supported by data obtained from in vitro and cellular studies, experimental studies with animal models, and observational studies, some of which come from prospective clinical studies of community-acquired pneumonia (CAP). There is some controversy about the use of new fluoroquinolones versus macrolides as the treatment of choice for LD. The least inflammation is found in the animal model with azithromycin, while the most is observed with erythromycin, with the quinolones being intermediate. Extrapulmonary manifestations of legionellosis, which are rare but sometimes observed in the immunocompromized host, may indicate significant therapeutic connotations. Mixed infections in legionellosis should be kept in mind concerning the inmunocompromized population since there are many reports of death when clinicians fail to identify and treat the dual component of infection. Some international guidelines recommend combined therapy for severe episodes but no consistent evidence supports this suggestion. For most patients monotherapy with a macrolide or a selected fluoroquinolone usually leads to a more cost-effective outcome. Possible toxicities of adding more than one antibiotic may be a concern, particularly in the intensive care unit setting. In vitro and in vivo evidence shows a highest potency of rifampin against Legionella, although the theoretical possibility of rapid induction of rifampin-resistant strains when it is administered alone precludes its use as sole therapy. The only variable that remained statistically significant on multivariate logistic regression analysis was the APACHE score (OR 1.86) at intensive care unit admission.

This chapter deals about the direct detection of Legionella from hospital water samples by commercial real-time (RT)-PCR system, that offers rapid results and reduced risk of cross-contamination. The preliminary results of 61 water samples from 6 hospitals and 15 water samples from air conditioning systems are presented and compared with those of the isolation method. Water samples were concentrated by filtration through cellulose acetate membrane filter and resuspended in 10 ml of the same water. The principle steps were cell lysis with proteinase K, purification on spin column and elution. Culture was positive in 28 (45.9%) hospital water samples, and the strains were identified as Legionella pneumophila serogroup 1, 3, 6, and 2-14 and Legionella spp. The results of qualitative RT-PCR were as follows: Legionella spp. was positive (detection limit: 133 GU/liter) in 60 (98.4%) samples and L. pneumophila was positive in 44 (72.1%). The results of culture and qualitative analysis for L. pneumophila by RT-PCR are shown. All hospitals examined showed some positive samples for culture and PCR, except for one that was culture negative and PCR positive. All water samples from air conditioning systems were culture negative, but RT-PCR was positive in 15 water samples for Legionella spp. and in 2 for L. pneumophila. In conclusion, Legionella detection by RT-PCR from environmental methods seems to be promising. However, at the moment, correlation between the method-result interpretations and public health significance need further evaluations.

The usefulness of sampling waters for the detection of Legionella remains a point of discussion among researchers and industry for a variety of reasons. This chapter describes a novel detection system utilizing two liquid media (medium 1 and medium 2, formulations are currently under provisional patent applications) and real-time PCR that is capable of detecting viable Legionella in a variety of waters within 2 to 3 days. Over 30 strains of Legionella have been tested by this method, and the real-time PCR detection system has been validated by authors' laboratory. Real-time PCR utilizing Legionella 16S rRNA primers was performed on a Corbett Rotor-Gene 3000 utilizing SYTO9 as a DNA intercalating dye for PCR. The GMS method was challenged with 82 field samples (cooling tower, evaporative tower, and potable waters) and run in parallel to AS/NZS 3896:1998. Comparison of methods using field samples showed that the GMS method is 100% specific (no false positives) and 92% sensitive, with a single discrepant sample. The GMS method described is rapid, simple, cost-effective, less laborious than conventional culture, and easy to implement in any modern day microbiology laboratory.

Metalworking fluids (MWFs) are highly susceptible to microbiological contamination from the environment due to high water content, available nutrients, and optimum growth temperature for most environmental microorganisms. Outbreaks of Pontiac fever and Legionnaires' disease were associated with MWFs at two unrelated automotive facilities. Legionella feeleii and L. pneumophila serogroup 1 (sg 1) were identified as the probable causative organisms. The objective of this chapter was to develop a selective and sensitive culture method for detection of Legionella in MWFs in order to prevent and control possible occupational health-related problems. Recovery efficiency was evaluated by spiking MWF field samples, highly contaminated (107 to 108/ml) with nonlegionella organisms with high levels (106 to 107/ml) of L. pneumophila sg 1 and L. feeleii. Several sample treatment conditions to reduce the interference of the high background population, including acidification, heat treatment, and use of antibiotics in the recovery medium, were evaluated.

Acute-care and long term-care facilities continue to experience cases of hospital-acquired Legionnaires’ disease. One of the major unresolved issues is whether the recommendations found in the guidelines will, if followed, result in the control and prevention of hospital-acquired Legionnaires’ disease. An evidence-based approach has been suggested as a way to resolve many of these issues. If applied to a guideline, the criteria should be that (i) the recommendations should be prospectively validated under controlled studies using a step-wise approach, (ii) the evaluation should be a prolonged observational period (>1 year) to evaluate the efficacy of the recommendations, and (iii) the recommended approach/actions should achieve the expected result-prevention of the disease through environmental control. Legionnaires’ disease is an environmentally acquired illness. One recommendation often found in guidance documents is “remove showerheads and aerators monthly for cleaning with chlorine bleach’’. The role of environmental monitoring in Legionella prevention has been a source of debate for many years. A number of disinfection methods have been used for control of Legionella in hospital water systems. These include thermal eradication (heat and flush), hyperchlorination, copper-silver ionization, point-of-use filters, and chlorine dioxide. Each of these methods has completed some of the evaluation criteria. All four steps of the evaluation criteria have been fulfilled for copper-silver ionization. The path to effective control of hospital acquired Legionnaires’ disease must be evidence based. Patients and healthcare facilities suffer when unconfirmed and untested recommendations become part of prevention guidelines.

This chapter investigates the overall Legionella colonization of thermal water and cold and hot water of particular hotels, including the detailed identification and typing of species and serogroups. The aim was also to evaluate the risk of particular hydrotherapy procedures, to identify the source of the Philadelphia strain in the spa complex, and to propose corrective actions to minimize the risk of future Legionella infections. Permanent chlorine dioxide disinfection was recommended for the hot water systems of the two hotels. A dilemma caused by the legislation has arisen as to how to treat the thermal water and its distribution system. Oxidizing biocides and ionizers are not permitted by law. Regular thermal disinfection is recommended, but its application is often unfeasible for the rapid settling of mineral deposits and scale, which usually clog pipes and valves. The bath equipment (mostly very sophisticated) contained many different kinds of tubing and hoses and air and water jets that were almost all colonized with biofilms harboring Legionella. Thermal and hot water distribution systems and hydrotherapy procedures present the sources, while aerosol inhalation and drinking water appear to be the transmission.

Different methods have been applied to control Legionella in water systems such as copper-silver ionization, chlorine dioxide, thermal treatments, hyperchlorination, UV radiation. In this chapter, authors evaluated the efficacy of a continuous system based on a silver-hydrogen peroxide mixture in two nursing home water systems. Recent studies showed a decline of Legionnaire's disease (LD) in health care facility-acquired cases where environmental monitoring of the water system and installation of a disinfection method were performed. Therefore, when the plumbing system is heavily contaminated by Legionella, it is important to apply control measures to reduce Legionella colonization and infection risk. Studies on the efficacy of the silver-hydrogen peroxide disinfection system are currently lacking, and to our knowledge, the present study is the first reported for Legionella control in hot water systems. In vitro studies demonstrated that hydrogen peroxide is a weak disinfectant in the absence of a catalyst. The combined action of hydrogen peroxide with silver as the catalyst characterize the disinfection power of the mixture used in this study. Results of this study, although preliminary, show that such disinfection systems can effectively control Legionella water system colonization even if some increase of Legionella levels at distal sites is observed.

Legionnaires’ disease is a potentially fatal form of pneumonia caused by the bacterium Legionella. Legionella control in cooling water systems depends on many parameters (conception, hydraulic management, maintenance) favorable to biological deposits. Any variation in the parameters measured (microbiological, physicochemical and environmental parameters) made it possible to identify and destroy Legionella growth sources. The cooling tower system is an ecosystem in which Legionella is always present and can proliferate any time of year. The percentage of samples in which Legionella is detected increased after April. A meticulous analysis of the risks made it possible to identify and manage all the factors of risk of proliferation of Legionella. The Legionella risk management cannot be carried out without taking account of the protozoa which are resistant to biocides. To fight continuously against the formation of biofilm seems to be the method most adapted to fight effectively against the proliferation of Legionella and limit the use of biocides.

Despite extensive efforts, the etiology of about 40 to 60% of community-acquired pneumonia remains unclear; a number of these cases are attributed to Legionella. The incidence in this group may be as high as 38%. Cultivation is still supposed to be the gold standard, but there is doubt about the sensitivity in clinical practice. Researchers established a new method for the detection of Legionella by using PCR to amplify genomic ribosomal DNA and a reverse dot blot for the detection of L. pneumophila and non-pneumophila species. The authors used a multiplex-PCR and the coamplification of human DNA (pyruvate dehydrogenase gene) as an internal control to avoid false-positive results. To avoid false-positive results, incubation with uracil-N-glycosylase was used, and for avoidance of false-negative results the coamplification of the pyruvate dehydrogenase gene was used, creating an amplificate of 185 bp in length in each PCR. The authors used the established method to investigate clinical specimens of 93 patients taken out of a prospective randomized study about the epidemiology of community-acquired pneumonia in Berlin, Germany, from 1991 to 1992. They were able to demonstrate the ability to detect Legionella using PCR in conjunction with reverse dot blotting. This method allows the enhanced detection of non-pneumophila species in addition to established methods. Interpretation of the results for the incidence of non-pneumophila infections remains controversial because of a lack of facts about pathogenity of this group and comparable results in other studies.

Legionella is an important cause of both community-acquired and nosocomial pneumonias. In this chapter, the authors produced 17 monoclonal antibodies (mAbs) against the recombinant PAL (rPAL) cloned from Legionella pneumophila serogroup (sg) 1 and investigated antigenic diversity of the peptidoglycan-associated lipoprotein (PAL) proteins from soluble fractions of 16 Legionella species including 22 serogroups by mAb reactivity patterns in ELISA. Soluble antigens from Legionella strains (L. pneumophila sg 1, 3, 4, 5, and 6; L. annisa; L. dumoffii; L. gormanii; L. jordanis; L. micdadei; L. oakridgensis; L. sainthelensi; L. bozemanii sg 1 and 2; L. longbeachae sg 1 and 2; L. Hackeliae sg 1) were prepared by the method of Berdal et al. The authors concluded that antigenic diversity of a Legionella species–common PAL protein is present among Legionella species and that developing diagnostic agents using a Legionella antigen, even if it is conserved, might be better for separately detecting L. pneumophila species and non-pneumophila Legionella species.