Current and Emerging Legionella Diagnostics

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National Institute for Communicable Diseases nicolew@nicd.

1 Current and Emerging Legionella Diagnostics Nicole Wolter Centre for Respiratory Diseases and Meningitis (CRDM) National Institute for Communicable Diseases 7 th FIDSSA Conference, Cape Town, 9-11 November 2017

2 Overview What is Legionellosis? Who gets Legionnaires disease? Why is Legionnaires disease important? How is Legionnaires disease diagnosed? Current methods Emerging methods When should Legionella testing be requested? Where can Legionella testing be done? 2

in environment, mostly natural water sources Amplification occurs in warm

decorative fountains, spa pools) Transmitted by inhalation/aspiration of

3 What is Legionellosis? Caused by Legionella bacteria, commonly Legionella pneumophila Ubiquitous in environment, mostly natural water sources Amplification occurs in warm water environments (cooling towers, hot water tanks, plumbing systems, decorative fountains, spa pools) Transmitted by inhalation/aspiration of contaminated aerosols No person to person transmission 3 Phin et al., Lancet Infect Dis, 2014; Mercante and Winchell, CMR, 2015

4 Presence in water supply Amplification Aerosol production Inhalation by susceptible host 4 Mercante and Winchell, Clin. Microbiol. Rev., 2015

5 What is Legionellosis? Legionnaire s Disease: Pneumonia, dry cough, fever, headaches, chest pain 2-14 days incubation period Severe, commonly requires hospitalisation, CFR 5-30% Requires antibiotic treatment (macrolide or fluoroquinolone) Pontiac fever: Mild flu-like illness (no pneumonia) Occurs 2-3 days after exposure Does not require hospitalisation or antibiotic treatment 5 Phin et al., Lancet Infect Dis, 2014; Mercante and Winchell, CMR, 2015

6 Who gets Legionnaires disease? CDC (NICD) Older people ( 50 years of age) (any adult >18 years) More common in males Current or former smoker Chronic lung/heart disease (previous or current TB) Immunocompromised (diabetes, cancer) (HIV) Immunosuppressed Wolter et al., Emerg Infect Dis,

Why is Legionnaires disease important? 1. Notifiable disease 2. Environmental source and therefore preventable 3. Severe pneumonia, commonly requiring ICU and high CFR 4.

7 Why is Legionnaires disease important? 1. Notifiable disease 2. Environmental source and therefore preventable 3. Severe pneumonia, commonly requiring ICU and high CFR 4. Radiologically and clinically indistinguishable from other causes of pneumonia 5. Empiric antibiotic therapy ineffective 5. Disease incidence is underestimated Lack of awareness of disease Low index of suspicion Inadequate and unavailable diagnostic tests Limited reporting of cases Limited / sub-optimal surveillance systems 7

8 Why is Legionnaires disease important? 6. Can cause major outbreaks 1976 outbreak at American Legion Convention in Philadelphia 221 suspected cases and 34 deaths 2015 outbreak in New York City approximately 120 cases and 12 deaths 8

9 Why is Legionnaires disease important? 9

10 How is Legionnaires disease diagnosed? Legionnaires disease case definition: Clinical/radiological evidence of pneumonia + Detection of Legionella spp. from clinical specimen 10

11 How is Legionnaires disease diagnosed? Test Specimen Advantages Disadvantages Culture* Sputum Pleural fluid BAL Bronchial aspirates Lung tissue Detects all species and serogroups 100% specific Molecular comparison Identify exposure Culture/PCR should be used in conjunction with UAT Urinary antigen* Urine Rapid (same day) Ease of sample collection Ease of use Slow (>5 days) Low sensitivity (20-80%) Affected by antibiotic treatment Specialised media (BCYE agar) Only detects Lp1 Does not allow molecular comparison PCR Sputum Pleural fluid BAL Bronchial aspirates Lung tissue Rapid Sensitive and specific Serology Serum Less affected by antibiotic treatment 80-90% sensitive >99% specific Assays vary by laboratory Expensive equipment Requires paired sera Retrospective diagnosis DFA Lung tissue Lower respiratory tract secretion Rapid >95% specific 25-75% sensitive Invasive specimen 11 * UAT AND culture are preferred diagnostic tests Phin et al., Lancet Infect Dis, 2014; Mercante and Winchell, CMR, 2015

12 How is Legionnaires disease diagnosed? 10 Europe* 10 USA* UAT Culture PCR UAT Culture/PCR South Africa, UAT PCR 12 *International Legionella Conference, Rome, 2017

13 How is Legionnaires disease diagnosed? Emerging methods: Isothermal nucleic acid amplification Amplification of DNA/RNA at constant temperature Rapid target detection, does not require expensive equipment Microfluidic Taqman Array Cards (TAC) Simultaneous detection of >20 respiratory pathogens Rapid and simple set-up Mass spectrometry (MALDI-TOF) Legionella species identification and typing Fast Amoebic co-culture Improve recovery of Legionella from culture-negative clinical specimens

14 How is Legionnaires disease diagnosed? Emerging methods: Microbial 16S rrna sequencing Identify bacteria in clinical specimen Detection directly from specimens Whole-genome sequencing (WGS) Microbial identification and typing Phylogenetic relationships Outbreak investigations Antibiotic resistance and virulence determinants Core genome multilocus sequence typing scheme (~50 genes)

15 How is Legionnaires disease diagnosed? Neonate case identified as positive for L. pneumophila SG1 Environmental sampling of the hospital and patients home Table: Sequence based typing results of clinical and environmental Legionella pneumophila isolates obtained during a Legionella case investigation, Western Cape, South Africa, Number Isolate Number Source of Isolate Location/Type of specimen Culture Result Serogroup Sequence type Clinical Isolates Liver Tissue L. pneumophila Serogroup 1 ST Clinical Isolates Liver abscess L. pneumophila Serogroup 1 ST Environmental isolate Household Bath tap L. pneumophila Serogroup 2-14 ST HM64758 Environmental isolate Household Kitchen tap L. pneumophila Serogroup 2-14 ST Environmental isolate Household Shower tap L. pneumophila Serogroup 1 ST1 6 HM64775 Environmental isolate Hospital Chilled water tank L. pneumophila Serogroup 2-14 new ST* 7 HM64776 Environmental isolate Hospital Water Tower Negative HM64777 Environmental isolate Hospital Labour ward Negative HM64778 Environmental isolate Hospital Surgical ward 2 Negative Environmental isolate Hospital High care L. pneumophila Serogroup 1 ST Environmental isolate Hospital Nursery L. pneumophila Serogroup 1 ST Environmental isolate Hospital Hot water L. pneumophila Serogroup 1 ST Environmental isolate Hospital ICU L. pneumophila Serogroup 1 ST1 14 HM64787 Environmental isolate Hospital Paediatric High Care Unit Negative HM64788 Environmental isolate Hospital Neonatal ICU cold water Unit Negative - -

16 Clinical Clinical Hospital Household Maximum-likelihood neighbour-joining tree of core genome SNPs (with the scale bar representing the nucleotide substitutions per site) of all Legionella pneumophila clinical and environmental isolates, Western Cape, South Africa, 2016.

17 When should Legionella testing be requested? Legionella testing is indicated in the following patients: 1. Severe pneumonia, in particular requiring intensive care 2. Failed antibiotic treatment for CAP 3. Immunocompromised patients with pneumonia 4. Healthcare-associated pneumonia 5. Travel history 17

18 Where can Legionella testing be done? Culture CRDM, NICD, Johannesburg UAT Submit urine and lower respiratory tract specimens for diagnosis CRDM, NICD, Johannesburg NHLS Infection Control Laboratory, Johannesburg Groote Schuur laboratory, NHLS, Cape Town Private laboratories PCR CRDM, NICD, Johannesburg Private laboratories 18

19 Case notification

20 Legionella resources 20

21 Conclusion Legionellosis in South Africa is severely underestimated Need to improve awareness for testing, availability and use of current diagnostic methods and notification/investigation of cases Source Prevent cases and outbreaks CIF Case notification Test Legionella positive Legionella testing available Legionella test requested Legionellosis clinically suspected 21

22 Thank you 22

Legionnella. You can t t see it but you can catch it. Dr Charmaine Gauci MD, MSc, PhD Head of Disease Surveillance Unit

Legionnella. You can t t see it but you can catch it. Dr Charmaine Gauci MD, MSc, PhD Head of Disease Surveillance Unit Legionnella You can t t see it but you can catch it Dr Charmaine Gauci MD, MSc, PhD Head of Disease Surveillance Unit Legionella infections First documented outbreak name from 221 cases of pneumonia occurred