Enhancing Legionella Testing Capacity for Clinical and Environmental Samples at

the Washington State PHLJeff Lahti

Lead MicrobiologistWashington State Department of Health

Jeff.lahti@doh.wa.gov

• Pre 2017 testing capabilities in Washington

• Our thought process on which tests to implement

• A brief look at the tests that were implemented

• A look at our future plans

• Helpful validation ideas

Outline

Legionnaires’ Disease Cases Reported to WA DOH by Year of Onset

0

10

20

30

40

50

60

70

1985

1986

1987

1988

1989

1990

1991

1992

1993

1994

1995

1996

1997

1998

1999

2000

2001

2002

2003

2004

2005

2006

2007

2008

2009

2010

2011

2012

2013

2014

2015

Case

Cou

nt

Year of Onset

Case counts are on an increasing trend nationally as well

Legionella Testing in Washington• Most hospitals do not collect a respiratory specimen, instead

rely on the urine antigen test

• There were only few facilities that maintained the capacity to culture a clinical specimen

• There were only 2 facilities that participated in the ELITE program

• If a large outbreak occurred, the state lab had no procedures for culturing clinical or environmental samples

– We are working to revise the WAC to state “isolates or the respiratory specimen associated with a positive result be submitted to the state”

What capabilities were needed

– Culture

– Increase sensitivity and turn-around-time of samples using a molecular assay

• After examining what the state lab can offer, we identified the areas that needed improvement

– Rapid identification of isolates

– Training for microbiologist to identify and isolate Legionella from specimens

Where to Start

– We had a microbiologist with experience Legionella isolates

• We decided to concentrate on enhancing clinical testing first

– In clinical micro a procedure existed for identification of isolates

– We still had unresolved issues regarding testing in our environmental lab, i.e. incubators, BSC, filtering equipment

– Clinical testing was considered “easier” to implement since there were more resources available

Isolate Identification

• First step was to expand identification capabilities

• Microgen Latex Kit

• Easy to use

• Rapid identification

• Added the ability to identify L. pneumophila SG1

• This kit can be used for clinical and environmental isolates

Photo curtesy of Microgen Bioproducts https://microgenbioproducts.com

Add Selective Media

• Next we expanded our media to include BCYE-PAV and BCYE-GPVC– We chose to add these two antibiotic plates since they were

cited the most for use in clinical and environmental isolation

• BCYE-PAC was also evaluated…– BCYE-PAC worked great for pneumophila,

but very poor with other species. – 6/9 species didn’t grow – 4/9 species had poor recovery

Validate Culture and PCR• Last step was to validate the culture and PCR• These validations were done at the same time

– Material was limited– The matrix was spiked– Had one dedicated microbiologist working on the

validation– Able to dedicate a BSC and ABI 7500 FAST

Future Testing

• WGS

• MALDI-TOF

• Legiolert

Helpful Validation Hints• Secure the testing material early on

– Isolates– Matrix

• Write a very detailed validation plan

• Risk Assessment

• Have one person in charge of the validation. They write the plan, oversee the testing and present the data for approval

Helpful Validation Hints• Create a flow diagram of the validation plan

Helpful Validation Hints

• Dedicate lab space for the validation

• Seek out training opportunities– CDC– Old ELITE PT

References

CDC’s ELITE programwwwn.cdc.gov/ELITE/public/elitehome.aspx

Garcia, Lynne Shore; Isenberg, Henry D. “Legionella Cultures 3.11.4” Clinical Microbiology Procedures Handbook 3rd ed., Lynne S. Garcia. 2007. Washington D.C.: ASM, c2010.

ISO 11731:2017 Water quality – Enumeration of Legionella

Jorgensen, Hames H, Pfaller, Michael A, Carroll, Karen C. Manual of Clinical Microbiology 11th ed. Washington D.C : ASM, 2015 887-904

References“Legionella (Legionnaires' Disease and Pontiac Fever).” Centers for Disease Control and Prevention, Centers for Disease Control and Prevention, 30 Apr. 2018, www.cdc.gov/legionella/labs/procedures-manual.html

Rice, E.W., Baird, R.B., Eaton, L.S., Clesceri. Standard Methods for the Examination of Water and Wastewater. 20th ed. 1998. Washington, DC: American Public Health Association, 2012. Print.