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Sensorer til måling af mikrobiel vandkvalitet SEMIVA projektet – hvad findes og hvad er undervejs

Hans-Jørgen Albrechtsen

Professor, Head of Section - Urban Water Systems

DANVA

Drikkevandssensorer 2016 – udstilling

og vidensdeling

27. januar 2016,Vandhuset,

Skanderborg

SEMIVA SEnsorer til måling af MIkrobiel VAndkvalitet

HOFOR A/S (Ann-Katrin Pedersen)

Aarhus Vand A/S (Jørn-Ole Andreasen)

VandCenter Syd A/S (Anne Esbjørn)

Kalundborg Forsyning A/S (Pernille Ingildsen)

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’Den ideelle sensor’ Forsyningernes definition

Installeres online eller at-line

Detektere indikatororganismer (E. coli og total coliforme)

Have høj følsomhed til detektion af lave koncentrationer af mikroorganismer i drikkevand

Give hurtigt svar

Kræve lav vedligeholdelse

Give lave forekomst af falske-positive

Arbejdsmetode

Opsummering af erfaringer med sensorer til monitering af mikrobiel vandkvalitet fra danske vandforsyninger - Spørgeskema

Fokuseret internet søgning

Opgørelse af relevante forskningsprojekter

Dialog med sensorproducenter

Telefoninterviews med fagfolk

Gennemgang af den videnskabelige litteratur

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Teknologier

på markedet

under udvikling eller i dokumentationsfase

på forskningsniveau

Teknologier på markedet Påvisning af coliforme og E. coli

Enzymatisk

Samme princip som Colilert

E.coli β-galactosidase activity (yellow)

Coliforme β-glucuronidase activity (fluorescence)

P. aeruginosa substrate hydrolysation

Total bacteria alkaline phosphatase activity

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Påvisning af coliforme og E. coli

Technology name

ALARM CALM Coliguard

BACTcontrol) aquaBio TECTA ColiMinder

Company Colifast A/S Colifast A/S MicroLAN

(mbOnline) ADASA Veolia

VMW (Vienna Monitoring Solutions)

Country Norway Norway The Netherlands Spain Canada/

Switzerland Austria

Analysed parameter

TC or EC TC, EC, P.

Aeruginosa (two simultaneously)

TC or EC TC and EC

simultaneously TC and EC

simultaneously TC, EC, or total

bacteria

Application DW Raw water,

process water, wastewater

DW, bathing water, process

water

DW, wastewater, bathing water

DW, wastewater, reclaim water

Sewage, surface water, DW soon

tested

Validation ETV EU project

Demowatercoli Research paper Currently not ETV N/A

Measuring unit

+/- +/-, MPN, CFU pmol

MUF/min/100 mL MPN/100 mL

+/- estimation of bacteria level

MUF/100 mL

Detection limit

1 cfu/100 mL 1 CFU/mL dep. on measuring mode

0.1 pmol MUF/100 mL

1 cfu/100 mL 1 cfu/100 mL 0.8 mMFU/100 mL

Installation at-line at-line at-line at-line Manual sampling

loading/ autosampling

at-line

Response time*

6-15 h (max 1 sample/day)

4-12 h 2-4 h 3-18 h 2-18 h 10-30 min (only

tested for surface water)

Sampling volume

100 mL 25-100 mL 20-3000 mL N/A N/A 6.5 mL

Maintenance Reagent refill

every 20 tests, yearly service

Reagent refill weekly to

monthly. Service twice a year

Reagents refill every 1-3 months for 4 tests/day13

Every 15 d reagent

replacement & monthly cleaning

N/A N/A for drinking water – currently

being tested

Market 190,000 DKK More expensive

than ALARM 300,000 DKK or

rental

174,000 DKK + 15,000 DKK yr maintenance

135,000 DKK For rent

Comments

HOFOR A/S, Trondheim and

Stavanger utilities

Stavanger utility

Enterococci i 2016? Experience by HOFOR A/S and

Nordvand A/S

Did not reply to inquiry. Svendborg

Vand A/S, Nordvand A/S,

HOFOR A/S with manual version

Currently working on new version

that incorporates pc

Coliforme / E. coli – sensorer

Teknologier:

‘ALARM’, ‘CALM’, ‘Coliguard’, ‘aquaBio’, ‘TECTA’, ‘Coliminder’

Automatiserede (excl. TECTA)

Enzymbaseret

Inkuberingstid til vækst af bakterier (2-18 timer)

Konvertering af enzym aktivitet til bakterietal - udfording

Forbrug af reagenser (refill uge/måned)

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Coliminder, aquaBio

Ingen dansk erfaring

ALARM

HOFOR A/S: data every 24 hours

Disadvantage: not for both TC EC at the same time)

Presence/absence.

Operation is quite stable, without excessive maintenance needs

Trondheim utility, 200,000 people: 3-4 years. infiltrated surface water, raw water intake to monitor for contamination

Several manure or sewage contamination detected, correlated roughly with grab sampling, good contamination indicator

User friendly system, only requires a short training course

Planning one more ‘ALARM’ at the lake water intake point

Rogaland (Stavanger) 300,000 people: 2 ‘ALARM’ sensors

Surface water intake points and monitor for E. coli every 4 h

Contamination cases detected, correlated roughly with weekly grab samples experience

positive

Concern: high initial and analysis cost, planning to continue using them

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CALM

Rogaland (Stavanger) 300,000 people

2 ‘ALARM’ sensors and 2 ‘CALM’ sensors

Overall experience is positive

Only expressed concern: high initial and analysis cost, especially of the ‘CALM’ sensor

Planning to continue using them

Coliguard

Nordvand A/S and HOFOR A/S

measure E. coli and total coliforms

4 times a day, low detection limit.

high maintenance needs

consider it an expensive technology

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TECTA

Nordvand A/S, HOFOR A/S, Svendborg Vand A/S

manual version

rather positive, user-friendly

response time of 2-18 hours is shorter than the traditional culture based methods, yet giving accurate results

only partly quantitative

mainly used during holidays, when laboratory closed

Påvisning af coliforme og E. coli

Technology name

ALARM CALM Coliguard

BACTcontrol) aquaBio TECTA ColiMinder

Company Colifast A/S Colifast A/S MicroLAN

(mbOnline) ADASA Veolia

VMW (Vienna Monitoring Solutions)

Country Norway Norway The Netherlands Spain Canada/

Switzerland Austria

Analysed parameter

TC or EC TC, EC, P.

Aeruginosa (two simultaneously)

TC or EC TC and EC

simultaneously TC and EC

simultaneously TC, EC, or total

bacteria

Application DW Raw water,

process water, wastewater

DW, bathing water, process

water

DW, wastewater, bathing water

DW, wastewater, reclaim water

Sewage, surface water, DW soon

tested

Validation ETV EU project

Demowatercoli Research paper Currently not ETV N/A

Measuring unit

+/- +/-, MPN, CFU pmol

MUF/min/100 mL MPN/100 mL

+/- estimation of bacteria level

MUF/100 mL

Detection limit

1 cfu/100 mL 1 CFU/mL dep. on measuring mode

0.1 pmol MUF/100 mL

1 cfu/100 mL 1 cfu/100 mL 0.8 mMFU/100 mL

Installation at-line at-line at-line at-line Manual sampling

loading/ autosampling

at-line

Response time*

6-15 h (max 1 sample/day)

4-12 h 2-4 h 3-18 h 2-18 h 10-30 min (only

tested for surface water)

Sampling volume

100 mL 25-100 mL 20-3000 mL N/A N/A 6.5 mL

Maintenance Reagent refill

every 20 tests, yearly service

Reagent refill weekly to

monthly. Service twice a year

Reagents refill every 1-3 months for 4 tests/day13

Every 15 d reagent

replacement & monthly cleaning

N/A N/A for drinking water – currently

being tested

Market 190,000 DKK More expensive

than ALARM 300,000 DKK or

rental

174,000 DKK + 15,000 DKK yr maintenance

135,000 DKK For rent

Comments

HOFOR A/S, Trondheim and

Stavanger utilities

Stavanger utility

Enterococci i 2016? Experience by HOFOR A/S and

Nordvand A/S

Did not reply to inquiry. Svendborg

Vand A/S, Nordvand A/S,

HOFOR A/S with manual version

Currently working on new version

that incorporates pc

8

Andre teknologier på markedet Technology name Aquascope

Desktop microscope

7000 RMS BACMON EZ-ATP Biocounter

Company Biotrack 4-deep Mettler Toledo A/S Grundfos A/S Applitek Biosensores

Country The Netherlands Canada International Denmark Israel,

Belgium Spain

Parameter Total bacteria, E. coli, Enterococci

or Legionella

Particles, bacteria or other

organisms

Bacteria and particles

(simultaneously)

Particles and total bacteria

ATP N/A

Principle Filter cytometry

& FISH Optical Optical Optical

Chemiluminescence

N/A

Application N/A Medical (cancer research, urine and blood tests)

Pharmaceutical waters

Drinking water Drinking

water and wastewater

Wastewater, beverage

Validation N/A N/A N/A MS in prep N/A N/A

Measuring unit N/A Particles or cells/mL

Bacteria and particles/mL

Cells and particles/mL

N/A N/A

Detection limit 1 cell/mL N/A for drinking

water 1 Biocount (no

conversion) 160 cells/mL N/A 103 cells/ mL

Installation N/A + flow cell at-line at-line at-line at-line

Response time 20-45 min Immediate N/A 10 min N/A 15 min

Sampling volume 1-500 mL N/A N/A 6 µL fixed in the flow cell

N/A N/A

Maintenance N/A Every day wiping

glass and flushing flow cell

N/A Regular change

of flow N/A N/A

Market N/A

Rental, 200,000 DKK + min.

40,000 DKK for training

N/A For rent N/A N/A

Comments No reply Microscope

needs operator at least remotely

US technology 'RMS-ON90' just

acquired Information not

available

Experience TREFOR VAND A/S, Aarhus Vand A/S,

HOFOR A/S

No reply No reply

Aquascope

Company: Biotrack

Country The Netherlands

Parameter Total bacteria, E. coli, Enterococci or Legionella

Principle Filter cytometry & FISH (Fluorescence in situ hybridization)

Detection limit 1 cell/mL

Response time 20-45 min

Sampling volume 1-500 mL

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Desktop microscope

Company 4-deep

Country Canada

Parameter Particles, bacteria or other organisms

Principle Optical, morfologi

Application Medical (cancer research, urine and blood)

Validation N/A

Measuring unit Particles or cells/mL

Detection limit N/A for drinking water

Installation + flow cell

Response time Immediate

Sampling vol. N/A

Maintenance Every day wiping glass, flushing flow cell

Market Rental, 200,000 DKK + >40,000 DKK for training

Comments Needs operator at least remotely

7000 RMS (RMS-ON90)

Company Mettler Toledo A/S

Country International

Parameter Bacteria and particles (simultaneously)

Principle Optical

Application Pharmaceutical waters

Validation N/A

Measuring unit Bacteria and particles/mL

Detection limit 1 Biocount (no conversion)

Installation at-line

Response time N/A

Sampling volume N/A

Maintenance N/A

Market N/A

Comments US technology just acquired Information not available

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BACMON (Zebra)

Company Grundfos A/S

Country Denmark

Parameter Particles and total bacteria

Principle Optical

Application Drinking water

Validation MS in prep

Measuring unit Cells and particles/mL

Detection limit 160 cells/mL

Installation at-line

Response time 10 min

Sampling volume 6 µL fixed in the flow cell

Maintenance Regular change of flow cell

Market For rent

BACMON

Erfaring: TREFOR VAND A/S, Aarhus Vand A/S, HOFOR A/S

Positivt: stabil drift, god brugerinterface, lille vedligeholdelse

Ulemper: Vanskeligt at tolke resultat,

men fortsat anvendelse

HIOFOR: installeret i ledningsnet, lang opholdstid

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EZ-ATP

Company Applitek

Country Israel, Belgium

Parameter ATP

Principle Chemiluminescence

Application Drinking water and wastewater

Validation N/A

Measuring unit N/A

Detection limit 103 cells/ mL

Installation at-line

Response time 15 min

Sampling volume N/A

Maintenance N/A

Market N/A

Comments No reply

Biocounter

Company Biosensores

Country Spain

Parameter N/A

Principle N/A

Application Wastewater, beverage

Validation N/A

Measuring unit N/A

Detection limit 103 cells/ mL

Installation at-line

Response time 15 min

Sampling volume N/A

Maintenance N/A

Market N/A

Comments No reply

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At-line teknologier under udvikling Name Bactiline N/A MiniLab MicroLab N/A

Company Mycometer A/S SBT Aqua ApS Optiqua (Optisense) Nwater Blusense

Diagnostics ApS

Background Sells Bactiquant test kit for detection of bacteria

in water

Spin-off from DTU Nanotech

Sells the EventLab technology for

chemical contaminant monitoring

N/A Spin-off from DTU

Nanotech

Country Denmark Denmark NL / Singapore Finland Denmark

Parameter Activity of several gram positive and negative

bacteria Total bacteria Total bacteria N/A

Specific bacteria strains

Principle Enzymatic activity and

fluorescence Impedance flow

cytometry

Optical chip inferometry with specific antibody

binding

Optical

Molecular extraction and

detection of specific genes

Validation/ development

stage

Developed prototype(s) are currently validated in the laboratory. To follow

validation at Vitens water-works (NL)

2 months continuous operation at Vilstrup

Vandværk. No measurements, only

robustness of microfluidic system

Manual handling. No test in drinking water. Further development

and automation during 2016/2017

Prototype developed

Prototype for detection of

proteins in blood and bacteria in

urine. Not tested in drinking water

Exp. release 2016 End 2016 2017 2017 End 2016

Comments An automated version of

the Bactiquant test -

Automated combined version of the

EventLab and MiniLab (on market)

- -

Funding EU (FP 7) via

SmartWater4Europe project

MUDP and VTUF EU (H2020) via

aquaSHIELD project N/A VTUF

Information og validering

Mangel på transparens i information fra forhandler

interest is to promote their products.

technical details are not disclosed due to commercialization interests (patent)

arbitrary validation tests

difficult to realise how far these technologies are in the development process

as manufacturing companies tend to be too optimistic about their progress and the expected time of market release.

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Validation

Metoder på forskningsplan

Method Immunoassays Polymerase

chain reaction (PCR)

Fluorescence in situ hybridization

(FISH) Electrical Flow cytometry

Raman spectroscopy

Principle

Antigen-antibody specific binding Used to

capture the target microorganisms and are typically combined with a

detection method

DNA amplification and detection

by dyes or probes

RNA hybridization and microscopic

detection of fluorescent probes

bound in cells

Different methods where

signal is detected by an electrode.

Are typically combined with

immunomagnetic separation

Labelled cells are suspended in a flow stream that passes

through a laser beam89-90

Measurement of inelastically

scattered light after excitation4.

Provides quantitative and

qualitative information

Sub- categories

Fluorescent immunoassays

Immunomagnetic assays Immunosorbent assays

- -

Electrochemical Electrical

impedance Piezoelectric

-

Surface enhanced Raman

spectroscopy (SERS) Lazer

tweezer Raman spectroscopy

(LTRS)

Current application

Various types of water and food to detect E. coli Salmonella Legionella

Enterococci

Various types of water

Groundwater to detect E. coli and

bacteria involved in chromium dioxide

reduction in a contaminated site

E. coli Staphylococcus enterotoxin B Salmonella

Cryptosporidium

Bacteria Yeast Algae Protozoa Viruses

E. coli Streptococcus Enterococcus

Bacteria spores

Advantages

High automation potential Can be

combined with several detection methods and can target a wide range

of microorganisms Potential for lab-on-a-chip implementation

Highly sensitive Specific Rapid Potential to

detect several micro-

organisms in a single reaction Potential for lab-on-a-chip

implementation

Allows the detection of viable but non-culturable

cells Can be combined with flow

cytometry in a microfluidic system

Potential for online and lab-

on-a-chip implementation

Rapid and accurate Various staining and labelling methods An

automated and on-line system and in lab-on-a-

chip systems Implemented in an automated and on-line system and in lab-on-

a-chip systems

Very sensitive Identification of

single bacteria cells Molecular

information with no reagents use Potential for microfluidic

implementation

Dis- advantages

Capturing of non-targeted microorganisms

Rapid but not quantitative Sample

concentration needed

Complex method

Enrichment steps may be required

Piezoelectric sensors require incubation time

Subjective data interpretation Bacteria detection requires high sensitivity of detection instruments High cost

Cells must be immobilised by

physical or chemical contact

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Større igangværende danske og EU projekter

Future water WP 8- Biosensors

Detection of E. coli DNA in distribution

systems

Real-time water quality monitoring by

electrical detection

AQUAWARN SMARTWATER4-EUROPE

Aquavalens Aquashield AQUAVIR

Partners

Ringkøbing-Skjern Forsyning A/S, VIA University College, Amphi-Bac ApS,

Alectia A/S, Minus 10dB ApS

Blusense Diagnostics

SBT AquaVerdo Vand A/S Svendborg Vand A/S

Sønderborg Vandforsyning

A/S Guldborgsund

Forsyning/ Nykøbing

Waterworks

6 EU partners Mycometer A/S and 19 EU partners

DTU FOOD DTU

Environment Højmarklaboratoriet A/S IPU A/S Nordvand A/S and 34 EU

partners

Optiqua (Optisens)

DTU Nanotech Unisensor A/S Delta A/S DHI A/S and 10 EU

partners

Country Denmark Denmark Denmark UK and other EU countries

Several EU countries

Several EU countries

The Netherlands

Several EU countries

Duration Sept 2013 - dec-15 Jul 2015-mar

2016 Jan 2015-sep

2016 Dec 2013-nov

2015 Jan 2014-dec 2017

Feb 2013-jan 2018

Jan 2015-jan 2017

Nov 2013-okt 2016

Relevant aim

Ddevelopment of single

components for alarm, automated

sampling and characterization of a

contamination

Development of a cheap sensor to monitor E.

coli in the distribution

system within 30 min

Further development of the SBT Aqua

sensor

Integrated detection of pollution in water using state-of-the-

art microfluidic technology

Further development

and integration of sensor

technologies

Development of automated

detection methods for

microbial water quality monitoring

Integrated sensor to detection

chemical and microbial

contamination

Development of a

microfluidic automated

virus analyser

Funding VTUF, MUDP VTUF VTUF, MUDP EU (FP7-SME-

2013) EU (FP7-CP) EU (FP7) EU (H2020) EU (FP7-CP)

Funding – EU-projekter

SMARTWATER4EUROPE 10,043,233 EUR

3 år, December 2017.

1 WP: Yderligere udvikling og validering. ‘Bactiline’ fra Mycometer A/S

Aquavalens 11,909,166 EUR

5 år, January 2018.

Udvikling af metoder og teknologier til at påvise pathogener i drikkevand og vand i fødevareproduktion. DTU FOOD og DTU Environment deltager i WP med udvikling af ATP-sensor

aquaSHIELD 1,123,136 EUR

Jan 2017

Udvikling af integreret sensor Optiqua (NL). Kombinere 2 sensorkomponenter: chlorine residual og risikostoffer

AQUAVIR 5,246,429 EUR

3 år, October 2016

Udvikling af bærbar on-site microfluidic system til at påvise virus i ferskvand. Koordineret af DTU Nanotech

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Status i dag? Området i hastig udvikling

Ofte optimistiske forventninger fra udvikler

Ofte begrænset dokumentation

Konkurrencehensyn

Er der tilstrækkelig dokumentation (også objektivt?)

Lovende teknologier, men udfordringer (undervurderet) på markedet:

Høj følsomhed

Kort svartid

Lav vedligeholdelse

Tekniske udfordringer i drikkevand

Få target-organismer

Lav mikrobiel aktivitet

Interferens

Udfældninger

Den ‘ideelle sensor’ er (endnu) ikke helt på plads

Status i dag?

Den ‘ideelle sensor’ er (endnu) ikke helt på plads

Coliforme og E. coli detektion tilgængeligt

Enzymatisk (colilert-baseret)

Responstid: 2-18 timer

Forbrug af reagenser, refill

Alternativer til coliforme og E. coli?

Totalt antal bakterier

~real-tid

Flere detektionsprincipper: optisk, enzymatisk, ATP

Langt i udvikling

Udvikling tager

Tid (5-10 år)

Betydelige investeringer

16

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Information og validation

lack of transparency in the information provided by the manufacturing companies, as their main interest is to promote their products. Often, detailed validation information or technical details are not disclosed due to commercialization interests. Therefore the information provided by the manufacturing companies should be interpreted with care, especially when conclusions are drawn from arbitrary validation tests and when satisfactory performance in other fields is assumed to apply also for drinking water.

Compiling technical details about technologies under development is even more challenging than the methods currently on the market, because companies are particularly reluctant to disclose any information before they launch their product. Often, it is also difficult to realise how far these technologies are in the development process, as manufacturing companies tend to be too optimistic about their progress and the expected time of market release.

Information og validering

Mangel på transparens i information fra forhandler

interest is to promote their products.

technical details are not disclosed due to commercialization interests (patent)

arbitrary validation tests

difficult to realise how far these technologies are in the development process

as manufacturing companies tend to be too optimistic about their progress and the expected time of market release.

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Funding – EU-projekter

‘SMARTWATER4EUROPE’ 10,043,233 EUR

three-year period ending December 2017107. one specific WP aims to further develop sensor technologies and to validate their use at selected demonstration sites108. The ‘Bactiline’ technology developed by Mycometer A/S

‘Aquavalens’ 11,909,166 EUR

five years and ends January 31st 2018.

develop methods and practices to detect pathogens in drinking water and in water used for food preparation. DTU Environment is involved in a work package that aims to develop an automated platform for detection, based on ATP

‘aquaSHIELD1,123,136 EUR

ends January 1st 2017

further develop an integrated sensor solution developed by the Dutch company Optiqua67. Specifically, the aim of the project is to combine two already developed sensor components for online monitoring, which monitor chlorine residual, and performs rapid screening of a set of high priority threat substances67. The project involves only Optiqua

‘AQUAVIR’ 5,246,429 EUR

three years, and ends October 31st 2016

The project aims to develop a portable, on-site microfluidic system to detect viruses in different freshwater water bodies. coordinated by DTU Nanotech and involves 13 more partners

Større igangværende danske og EU projekter

19

Metoder på forskningsplan