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ACTIVE BIOMASS QUANTIFICATIONUSING QUENCH-GONE™ AQUEOUS (QGA™) KIT AND TOTAL CONTROL MICROBIOLOGY (TCM™) KIT
Quench-Gone Aqueous (QGA™) kit allows fast measurement of total viable fl ora present in water samples through the quantifi cation of intracellular Adenosine Triphosphate (ATP). ATP molecule is the energy transporter inside all living organisms and is degraded immediately when released in the environment. The principle of ATP measurement is based on a bioluminescence technique, in which one photon of light is produced in the enzymatic reaction between Luciferase enzyme and an ATP molecule. Intensity of light is measured with a luminometer. The quantity of produced light is directly proportional to the active biomass present in the sample.
ATP + luciferin + O2 AMPP + PPi + oxyluciferin + LIGHT LIGHT Mg++
luciferase
QGA™ kits quantify only ATP contained in living microorganisms (Intracellular ATP or cATP). Intracellular ATP is obtained after fi ltration of the sample (50ml) and lyses of bacteria retained on the fi lter for release of ATP. Total time to result is 6min. The quantity of produced light is directly proportional to active biomass: primary results expressed in RLU (Relative Luminescence Unit) are converted in pg ATP/ml or Equivalent microorganisms/ml* using ATP calibrated standard.
1. FILTRATION 2. EXTRACTION 3. RESULT
Intracellular ATP is the true indicator of living total fl ora. Thus measured, active biomass includes cultivable, non cultivable or diffi cult to culture microorganisms.
Total Control Microbiology (TCM™) kit is a tool for specifi c evaluation of biomass stress (BSI™ = Biomass Stress Index) through the measurement of total ATP (tATP) and extracellular ATP (dATP, issued from microorganisms mortality): BSI = dATP/tATP.
Recommended cATP and BSI thresholds:
Process Param. Good Qualtiy
Preventive Action
Corrective Action
Potable Water cATP(pg/ml)
< 0,5 0,5 à 10 > 10
Cooling Water(Non-Oxidizing Biocides
cATP(pg/ml)
<100 100 à 1.000 > 1.000
Cooling Water(Oxidizing Biocides)
cATP(pg/ml) <10 10
à 100 >100
Surface orRecycle water cATP
(pg/ml) < 5 5 à 100 > 100
STUDY OF DIFFERENT CONTINUOUS OXIDIZING BIOCIDE TREATMENTS USING ACTIVE BIOMASS QUANTIFICATION
Impact of oxygenated water stabilized with silver ions (H2O2/Ag) and hypobromous acid (HOBr) treatments on active biomass: During H2O2/Ag treatment active biomass concentration varied from 104 and 106 microbial equivalents/ml (10 to 1000 pg ATP/ml). Residual H
2O
2
concentration varied between 1 and 200 mg/L.With HOBr treatment, active biomass decreased to 102 and 104 microbial equivalents/ml (under 10 pg ATP/ml) for concentrations varying between 0.5 and 10mg/L.
Active biomass through ATP : GF CTF and FS
Biomass Stress Index (BSI), representing effi ciency of biocides, during H202/Ag and HOBr treatment:
Biomass Stress Index follow-up at change of treatment
Stress of active biomass during H20
2/Ag treatment varied from 20%
to 40%. HOBr treatment showed higher effi ciency as BSI reached 100%.
IMPLEMENTATION OF LOCAL BIOCIDE INJECTION POINTS AT EACH COOLING CIRCUIT WITH DIRECT INJECTION OF THE PRODUCT
A solution containing 90% of hypochlorous acid and 10% of chlorine dioxide produced by electrolysis (OVIPUR system - BIO-DES technology ; Marin Development, France) was tested. The injection of HOCl/ClO
2 mix obtained by electrolysis is depending on the
redox potential in each circuit. Tests on 3 circuits showed that a redox potential of 600mV is required for maintaining cATP and Legionella spp PCR values conform to guidelines.
Active biomass GF11
Intracellular ATP follow-up on a circuit treated successively by H2O2/Ag fi rst, then by HOBr and a mixture of HOCl/ClO2
PARALLEL MEASUREMENTS USING LEGIONELLA SPP. PCR METHOD AND QGA ATP-METRY TEST
The measures performed in parallel by Legionella spp PCR method and cATP method showed that results followed the same trend, thus for 10 cooling circuits and with different treatments applied.
GF12 : legionella spp (UG/L) – active biomass (microbial eq/L)
Follow-up of intracellular ATP and of L.spp PCR on cooling circuit GF12
esta
mpe
s.co
m
ATP-METRY FOR SELF-INSPECTION ON COOLING CIRCUITS:use of new analytical tools for Legionella risk monitoring and control of biocide effi cienciesCarine Magdo, Altis Semiconductors – Veliana Todorova, Aqua-tools – Marc Raymond, Aqua-tools
INTRODUCTIONScientifi c data show that Legionella proliferation is linked to the presence of biofi lms and important microbiological fl ora in water circuits. Management of Legionella risk in cooling towers requires the implementation of control plans based on physico-chemical and microbiological indicators. Methods for fast detection of total microbiological proliferation in water and biofi lms are useful for Legionella risk monitoring in cooling circuits. Altis Semiconductors factory runs an important cooling circuits installation. Concerned by optimization of monitoring, a new ATP-metry technique has been tested at Altis premises during 1 month. Quench-Gone™ Aqueous kits (Luminultra™ (Canada); commercialized by Aqua-tools in Europe and EMEA) are dedicated for microbiological control of water networks and allow measurement of only intracellular ATP, representative of viable microbial fl ora. The study performed at Altis included the follow-up of intracellular ATP on 10 cooling circuits for evaluating the effi ciency of different biocide treatment strategies. Parallel measurements between ATP-metry and Legionella species PCR were performed.
Our way to manage sanitary risks and improve environmental conformity.
*(1pg/ml of ATP is equivalent to 1000 microorganisms/ml - van Crombrigge J ; Was G (1991) : ATPmethod - Bulletin Fil/IDF 256/1991).
INTERPRETATIONThe different measures performed on the cooling towers showed that active biomass measurement by QGA™ kit is a reliable and fast tool for monitoring cooling circuits installations. Guideline values and action thresholds were defi ned for this indicator: their day-to-day interpretation in addition to the other physico-chemical analysis allows to determine actions to undertake. At Altis, the target value for the cooling towers was fi xed at 1pg of ATP/ml, representative of a microbial population under control and an effi cient treatment. An alarm threshold at 10pg of ATP/ml was implemented, indicating a drift in the process linked to a risk of results for Legionella culture excesses.
CONCLUSIONATP measurement with QGA™ kit has several benefi ts for an industrial site. First, this is an instantaneous indicator of the level of microorganisms that can be used as a tool for regular follow-up of the process. The quantifi cation of biomass using ATP-metry is the only true indicator of the total fl ora, as viable cultivable and viable non cultivable bacteria present in the installations are both quantifi ed with this method. Additionally, analyses are performed in laboratory on-site and take 6min per sample. At last, the cost of this measure is low compared to other available analytical measures.
The purposes for the follow-up of an installation, using ATP-metry are numerous: Verifi cation of effectiveness of biocide treatment on viable cultivable and viable non•cultivable micro-organisms, complementary to Legionella spp PCR.- Improvement of effi ciency of treatments- Optimization of concentration of biocides- Validation of disinfectants effectiveness
Autocontrol:•- Reaction time: results in 6min (compared to 10 days per culture and 48h per PCR)- Anticipation of water quality derivationIdentifi cation of pollutions and zones of bacterial proliferation•Cost effectiveness: optimize budget for analysis, using the most discerning indicators•
to reduce the budget for chemical treatments.
The use of ATP-metry method on Altis Semiconductor site allowed the qualifi cation of a new treatment (production of HOCl and ClO2
by electrolyze) on a group of cooling towers. This treatment will afterwards be generalized on the entire site leading to important reduction of chemical treatments (6T/year) and improvement of environmental impact by eliminating bromates. Measurement of intracellular ATP allowed furthermore validation of biocide concentration and contact time used for shock disinfection treatments; this comforted Altis in their decision to stop some biocide treatments (isothialozone, H2O
2/Ag).
Parameter
Goodactivity
Preventiveactivity
Correctiveactivity
BSI* (%)
> 75
50 to 75
< 50
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