Updating Technology of Water Microbiology

20th September 2023

 Speaker Introduction
Wan Teng is a seasoned professional
in the water industry. With a career
spanning back to 2013, Wan Teng
embarked on her journey in this vital
sector with a water treatment
company. There, she excelled in a
multitude of supporting functions,
seamlessly blending technical
expertise with commercial acumen.

Beginning as a Technical Support

Specialist, she lend her expertise to
support the dynamic needs of
countries in Southeast Asia. Her
passion and proficiency soon
Wong Wan Teng propelled her to her current role,
Technical Sales, SEA Water where she spearheads technical
sales in the bustling hub of
Singapore.

© 2023 IDEXX Laboratories, Inc. All rights reserved.

Legionella pneumophila

Legiolert®
 Indonesia Swimming Pool and Spa Water
Requirements
o Permenkes no 2 Tahun 2023

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 Legionella pneumophila L. pneumophila is the
primary cause of
Legionnaires’ disease
Culture-confirmed cases of
Legionnaires’ disease
EU/EAA 2009-2014

L. pneumophila is the
most clinically relevant
Legionella species in
building water

Source: Europe Centre for Disease Control & Prevention When L. pneumophila is
controlled, all other
Legionella spp. are too

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 Regulation specifying L. pneumophila
New Brunswick (Canada)

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 Legiolert

Nutrient Indicator = Proprietary

Enzyme = Proprietary

SUGAR Coloured Indicator

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 ASTM D8429-21
Legiolert

o American Society for

Testing Materials (ASTM)
International: D8429-21

• Standard Test Method for

Legionella pneumophila in
Water Samples Using
Legiolert

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 Water Types
Potable Water Non-Potable Water
Definition Water which is fit for consumption by Water that is not of drinking water
humans, also called drinking water in quality, but may be used for other
reference to it’s intended use. purposes depending upon the quality.
Common Drinking Water, Drinking Fountains, Cooling Towers, Evaporative
Examples Kitchen taps/sinks, Bathroom Condensers, Fluid Coolers, Chilled-
taps/sinks, Showers, Eyewash water Return tanks, Header tanks
Stations, Bath Taps, Ice Machines Air Conditioners, Machine/utility
waters, Whirlpools, Pools, Spas,
Baths, Decorative fountains,
Untreated waters (springs, ponds,
surface waters), Car Washes,
Rainwater harvest storage tanks

© 2023 IDEXX Laboratories, Inc. All rights reserved.

Sample Preparation
Non-potable Water
 Equipment for non-potable water sample

Legiolert
Reagent Sterile 100mL vessel Pretreatment Kit

Quanti-Tray/Legiolert Incubator Quanti-Tray MPN Table

(QTL) and rubber insert Sealer PLUS
© 2023 IDEXX Laboratories, Inc. All rights reserved.
 Legiolert pretreatment kit
o The Legiolert pretreatment kit consists of 4 pretreatment
vessels
• containing a dry powder reagent to be reconstituted in sterile
water

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 Non-potable water – Legiolert pretreatment

Add sterile, DI
to the 100mL
fill line

Pre-treatment Shake to Store at RT

vessel dissolve

• Can be used to run up to 500 Legiolert tests

• Reconstituted Legiolert Pretreatment is stable at ambient
temperature (15–30°C) until expiration date
© 2023 IDEXX Laboratories, Inc. All rights reserved.
 Non-potable water - 1mL Test procedure
1 2 Water 2ml
sample 3 4

Legiolert 2ml
pre-
treatment
60s
Add reagent to the Treat sample with Add 2mL Pour into Quanti-
100ml diluent Legiolert pretreated sample Tray/Legiolert
pretreatment to reagent mixture

5 6 7

Seal using Quanti-Tray Incubate at 37°C± 0.5°C Count positive wells and
Sealer PLUS for 7 days refer to MPN Table
© 2023 IDEXX Laboratories, Inc. All rights reserved.
Quanti-Tray Legiolert (QTL) and
Results Interpretation
 Reading positive wells on Quanti-Tray/Legiolert

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 Interpreting Results with Quanti-Tray/Legiolert
MPN Table

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 MPN & CFU are interchangeable per ISO 6107:2021
definition
Acceptable units to estimate the
number of cultural microorganisms:

- CFU
- MPN

CFU and MPN are interchangeable!

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 Significantly easier to interpret results

becomes

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 Legionella Pneumophila
Proven AFNOR, ASTM, UK Blue Book, and more*

Easy Eliminates media preparation, no glassware cleaning or colony counting

Ease of use simplifies training
Easy quality control (15 minutes)
Single test for 100mL sample (vs. multiple plates and steps for traditional methods)
Any brown color and/or turbidity indicates a positive for Legionella pneumophila
Rapid Less than two minutes of hands-on time
Confirmed result in 7 days (vs. 10-14 days for traditional methods)
No confirmations needed
Accurate Detects 1 CFU/100mL
Eliminates subjective interpretation required by traditional methods
Specifically detects Legionella pneumophila, the primary cause of Legionnaires’
disease
Flexible Presence / absence or quantification (Quanti-Tray)
Validated on a wide range of water types

© 2023 IDEXX Laboratories, Inc. All rights reserved. *Note: Review IDEXX Water Community for full approval list
IDEXX-QC Kits
 Standard Method 9020
Quality Assurance/Quality Control

© 2023 IDEXX Laboratories, Inc. All rights reserved.

 IDEXX-QC Kits
o IDEXX-QC is a set of bacterial cultures for use in quality control
testing or as performance evaluation standards.

o The viable organisms are each contained in a colored disc, which

resides on top of a cotton plug and silica gel desiccant in the vial.

© 2023 IDEXX Laboratories, Inc. All rights reserved.

 IDEXX- QC Kits
Organisms and Quantity Included in
IDEXX-QC Kit For which Reagent Organisms and Expected Reactions
Each Kit
3 – Escherichia coli E. coli: Total Coliform +; E. coli +
Colilert, Colilert-18,
Coliform & E. coli 3 – Klebsiella variicola K. variicola: Total Coliform +; E. coli -
Colisure
3 – Pseudomonas aeruginosa P. aeruginosa: Total Coliform -; E. coli -
3 – Escherichia coli E. coli: Fecal coliform +
Fecal coliform Colilert-18
3 – Pseudomonas aeruginosa P. aeruginosa: Fecal coliform -
3 – Enterococcus faecalis E. faecalis: Enterococci +
Enterococci Enterolert 3 – Escherichia coli E. coli: Enterococci - (gram-negative)
3 – Streptococcus bovis S. bovis: Enterococci - (gram-positive)
3 – Pseudomonas aeruginosa P. aeruginosa: P. aeruginosa +
Pseudomonas
Pseudalert 3 - Escherichia coli E. coli: P. aeruginosa -
aeruginosa
3 – Pseudomonas fluorescens P. fluorescens: P. aeruginosa -
E. faecalis: HPC +
(use a blank made with sterile water as
HPC Simplate 3 – Enterococcus faecalis
the
negative control)
Colilert, Colilert-18,
E. coli 9 – Escherichia coli E. coli: Total Coliform +; E. coli +
Colisure

Legionella 3 – Legionella pneumophila L. pneumophila: L. pneumophila +

Legiolert
pneumophila 3 – Enterococcus faecalis E. faecalis: L. pneumophila -

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 Procedures
1 2

Remove required vial(s) from freezer. Aseptically transfer the pellet to vessel
Equilibrate at room temperature for 15 mins. containing 100 mL sterile water.

3 4

Swirl the sample and allow to stand for up to Refer to the acceptance range for each
5 mins. Mix by inverting the vessel 10 times. organism on the COA.
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 IDEXX-QC Legionella pneumophila
Certificate of Analysis

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© 2023 IDEXX Laboratories, Inc. All rights reserved.