Legionella Dr. Harriet Whiley Most undervalued profession It is responsible for almost everything you take for granted living in a developed country.

Legionellosis Legionnaires disease Pontiac Fever A sever atypical pneumonia like infection Infection Pontiac Fever A mild flu like illness Allergic reaction to live/dead Legionella + endotoxins from non Legionella bacteria? Asymptomatic Legionella Increased Legionella antibodies with no symptoms

Incidence of Legionellosis Across Europe (2011) = 4897 Legionellosis cases incidence rate of 0.97 cases per 100,000 (European Centre for Disease Prevention and Control, 2013) Untied Stated (2011) = 4,202 Legionellosis cases incidence rate of 1.36 cases per 100,000 (Centers for Disease Control and Prevention 2013) Australia (2014) = 427 Legionellosis cases (40 from SA) incidence rate of 1.8 per 100,000 (0.4 per 100,000 for SA) 149 Meningococcal cases 1840 dengue fever cases 12785 salmonellosis cases 82625 Chlamydia infection cases (Department of Health, National Notifiable Disease Surveillance System 2015).

True incidence of Legionellosis could be 20 times greater than the currently reported incidence. In the US Legionella is the leading cause of all drinking water outbreaks (Centers for Disease Control and Prevention 2013). The annual cost of hospitalisations due to Legionellosis in the United States is estimated to exceed US$716 million (Giambrone 2013) Melbourne aquarium outbreak (2000) 125 cases, 95 hospitalisations, 4 deaths loss of business/tourism etc. $35 million legal claims

Electron microscope image of L. pneumophila Legionellosis 19 species of Legionella cause human infection L. pneumophila (aquatic) 80% of Legionella cases worldwide L. longbeachae (potting mix) more common in Australia, New Zealand and China Electron microscope image of L. pneumophila

Sources of L. pneumophila Humidifiers Cooling towers Hydrotherapy pools Birthing pool Fountains Respirator Ice machines Present in potable water – main reason its able to survive within these environments is its association with biofilms showers Dental units Feeding tubes Spas

For an aerosol ‘size is important!’ >10 µm nose and throat 5-10 µm upper and lower respiratory tract 2-5 µm lungs and conducting airways >10 µm >5-10 µm 2-5 µm Eg. 90% shower aerosols 1-5 µm

L. pneumophila survival: Biofilm Biofilm growth on hot water pipe Biofilm growth on cooling tower pipe Biofilm growth on plastic tubing

L. pneumophila survival - Amoeba Electron micrographs of U937 macrophages (human white blood cell) (A) and Acanthamoeba polyphaga (B) infected by L. pneumophila (strain AA100) at 24 h. The infection is very similar in both host cells (Molmeret, et al. 2005).

L. pneumophila on BYCE GVPC agar Legionella detection methods Culture – the ‘gold standard’ PCR / qPCR Fluorescent in situ hybridization (FISH) solid phase cytometry optical wavelight spectroscopy Enzyme-Amplified Electrochemical Detection with DNA probe Surface plasmon resonance immunosensor Detection methods L. pneumophila on BYCE GVPC agar

L. pneumophila on BYCE GVPC agar Legionella detection methods Culture – the ‘gold standard’ Limit of detection Viable but non culturable (VBNC) Presence of disinfectants High temperature Low nutrients If the legionella is at the flagellated stage of the life cycle Present within a protozoa FASLE NEGATIVES!!!! Detection methods L. pneumophila on BYCE GVPC agar

Cooling tower mandatory reporting to Department of Health NSW / QLD / WA – No mandatory reporting VIC – 3 consecutive >10 Legionella CFU/mL ACT / SA / TAS - >1000 Legionella CFU/mL

Legionella targets for potable water pipelines using culture (gold standard) detection Country CFU/mL Comments France <1000   <100 <50 Target for general public facilities Target for prevention of nosocomial Infections Target where at‑risk patients are hospitalized Germany The Netherlands Guideline target United Kingdom

Investigation into South Australian potable water Maximum concentrations 105 copies/mL Legionella spp, 103 copies/mL L. pneumophila qPCR Doesn’t differentiate between live and dead Significant increases in concentration occurred during summer, low disinfection residual and dead end. (Whiley et al. 2014)

Take home message Assume its always there!!! Prevent/remove biofilm Maintain disinfection residual For hot water systems make sure the temperature are being reached – don’t ever have warm water sitting in the pipes Protect the high risk groups

Questions?  Harriet.Whiley@flinders.edu.au

References Centers for Disease Control and Prevention (CDC). (2013). Surveillance for waterborne disease outbreaks associated with drinking water and other nonrecreational water-United States, 2009-2010. MMWR. Morbidity and mortality weekly report, 62(35), 714. European Centre for Disease Prevention and Control. (2014) Legionnaires’ disease in Europe, 2012. Stockholm: ECDC. Department of Health. (2015). National Notifiable Disease Surveillance System, Australian Government Giambrone, G. P. (2013). National Cost of Hospitalization for Legionnaires' Disease, 2001-2009. 2013 CSTE Annual Conference, Cste. Molmeret, M., M. Horn, M. Wagner, M. Santic, and Y. Abu Kwaik. (2005). Amoebae as training grounds for intracellular bacterial pathogens. Appl Environ Microbiol 71:20–8 Whiley, H., Keegan, A., Fallowfield, H., & Bentham, R. (2014). Detection of Legionella, L. pneumophila and Mycobacterium avium complex (MAC) along potable water distribution pipelines. International journal of environmental research and public health, 11(7), 7393-7405.