Conferencia a cargo de Mireia Lázaro, Responsable Comercial de Biótica Bioquímica Analítica, explicando el método SIM (Separación Inmunomagnética) para detectar de forma rápida legionella en aguas, en la Jornada organizada por Frilabo en Lisboa el pasado 13 noviembre.
3. Reunião de esclarecimiento SIM
• Innovative company of technological base (EIBT).
• Located at Scientific Park–Univ. Jaime I of Castellón
(Espaitec).
• Develop methods to detect fastly microorganisms.
• ISO 9001:2008.
www.biotica.es
The company
5. Reunião de esclarecimiento SIMReunião de esclarecimiento SIM
Bacteria
separation
Fast
methods
Conventional
Methods
Detection Identification
RISK DAMAGE
Complex matrix
Clean matrix
The analytical problem
6. Reunião de esclarecimiento SIMReunião de esclarecimiento SIM
Alive
CV
Yes
Transitions
Damaged
Envelop¿?
Grows
¿?
No
Yes
VBNC
No Dead
NV
DebrisRecoverables
Legionella shows different states
8. Reunião de esclarecimiento SIMReunião de esclarecimiento SIM
What is Legipid®?
• Fast method to determine Legionella spp. in water.
• It uses magnetic particles and antibodies against
Legionella spp.
• Validated and Certified (AOAC-RI)
• Considered in UNE 100030:2017
• Acreditable (ex. ENAC, ACCREDIA, NATA, UKAS…)
10. Reunião de esclarecimiento SIMReunião de esclarecimiento SIM
Polystyrene
Magnetite
Magnetic particles
anchorage
Antibody
recognition
0.9 μm
11. Reunião de esclarecimiento SIMReunião de esclarecimiento SIM
•Capture
•Marking
•Detection
•(Prefilter)
•Membrane filter
•Elution flask
• Visual
•Colorimeter
• CFU eq
Stages of Legipid® Test
Filtration Analysis Result
12. Reunião de esclarecimiento SIMReunião de esclarecimiento SIM
•Inmuno magnetic Separation and enzyme immunoassay
Principle of the Legipid® test
13. Reunião de esclarecimiento SIMReunião de esclarecimiento SIM
Capture: molecular velcro
MAGNETIC
PARTICLE
LEGIONELLA
CELL
14. Reunião de esclarecimiento SIMReunião de esclarecimiento SIMD
A B C
DAMAGEA B C
INTERACION
STRENGH DAMAGE
ALIVE
DAMAGED DEAD
Capture of an intact cell
17. Reunião de esclarecimiento SIMReunião de esclarecimiento SIM
Result: visual interpretation
UFCeq/vol examined
18. Reunião de esclarecimiento SIMReunião de esclarecimiento SIM
Result: optical reading
• Negative results— (Ar < 0.04 u.a): ND
• Positive results— (Ar ≥ 0.04 u.a): D
Log10 C = 2.3061 Log10 (Ar) +
4.9815
CFUeq/vol examined
Semi-micro cuvettes
Sample
(T)
Negative
Control
(C)
19. Reunião de esclarecimiento SIMReunião de esclarecimiento SIM
It is the quantity of CFU that should appear in a
culture of the same concentrated sample, in
maximum conditions of cultivability.
100 % cells able to
grow and accessible
0 % interfere
Result: CFU equivalent
21. Reunião de esclarecimiento SIMReunião de esclarecimiento SIM
2
Legipid® Formats: devices
Manual
shaking
Orbital
shaker
22. Reunião de esclarecimiento SIMReunião de esclarecimiento SIM
Orbital shaker Plataform
Elution flask
MP4
Reagents
Tapete
Filtration
system
Reagents
Colorimeter
23. Reunião de esclarecimiento SIMReunião de esclarecimiento SIM
Analytical application
IMS
Fast methods Conventional
methods
Detection Identification
RISK DAMAGE
Complex matrix
Clean matrix
24. Reunião de esclarecimiento SIMReunião de esclarecimiento SIM
• Results the same day
• Detection of viable and VBNC
• Useful in complex matrices
• Good sensibility
• Low cost
• CFUeq vs CFU
• Easy implementation
• Publication peer-reviewed available
• Validated and Certified (AOAC)
• Accomplish regulation UNE100030:2017
• Useful in routine and outbreaks
• Public Health Benefit
Legipid®’ s
profits
25. Reunião de esclarecimiento SIMReunião de esclarecimiento SIM
Determination by Immuno Magnetic Separation
(IMS)
• False positives in samples strongly colored.
• No others are known.
Disadvantages
26. Reunião de esclarecimiento SIM
FACILITY/S
OWNER
PUBLIC HEALTH
AUTHORITY
MAINTENANCE
COMPANY LABORATORY
ISO 17025
HAVING A FAST RESULT, I
WILL REDUCE
COSTS(sanctions, media
damage, production stop,
etc.)
I CAN MAKE MORE
SCREENINGS, HIGHER
AND CHEAPER
I CAN ANALYSE ON THE FIELD
I CAN ANALYZE MORE
POINTS AT THE SAME
PRICE.
I USE LESS SPACE
NO SPECIAL
INSTRUMENTAL
REQUIRED.
I CAN BE
ACCREDITED
I DON’T NEED
QUALIFIED STAFF
I CAN SEND RESULTS
THE SAME DAY TO MY
CLIENT
IN AN OUTBREAK, I NEED
MAXIMUM SPEED AND
FIABILITY.
OFFER THIS
SERVICE TO MY
CLIENTS IS A
BENEFIT BETWEEN
ME AND MY
COMPETITIVE
IS THE TECHNIQUE WITH
LESS WORKFOCE
HAS DE DISINFECTION BEEN
EFFECTIVE? NOW I CAN NOW
IT
CAN DO IT MYSELF!
28. Reunião de esclarecimiento SIMReunião de esclarecimiento SIM
Compendium International Certificate Published validation
Accreditation Guide
(Guide 20, ENAC
2017)
Internal quality control Collaborative
29. Reunião de esclarecimiento SIM
Validation and certification
• Validation is conducted and certified by a competent
and independent body
o Validation is provided by the manufacturer
o User doesn´t validate methods
User verifies their method´s performance
User needs an Accreditation Guide
30. Reunião de esclarecimiento SIMReunião de esclarecimiento SIM
Technical Sheet and International Certificate
32. Reunião de esclarecimiento SIMReunião de esclarecimiento SIM
Routine
Cooling
Towers
Shower
Spas,
jacuzzi
Decorative
fountains
Floors
Municipal
water
• Earlier
detection
• Immediate
action
• Reduced
exposition
33. Reunião de esclarecimiento SIMReunião de esclarecimiento SIM
Routine: Legionella sp en swimming-pools
2 days - PCR
48 hour-results
(1 inconclusive result)
1 hour - Legipid®
(Conclusive results in 1 hour)
Law compliance (RD742/2013)
Sample taking
Analysis on the same day
Census
Identified risk facilities
10-12 days - Culture
(unconclusive results,
MICROBIOTA)
Optimal cost
(reduce long closing and
reopening periods )
Protected users
(without activities
interruption)
Advanced prevention
(Prevention Plan with fast
technique)
Law RD742/2013, Norm UNE.EN
100030
34. Reunião de esclarecimiento SIMReunião de esclarecimiento SIM
FOTOcontrol DBNPA
Study in collaboration with:
FOTOcontrol Hypochlorite
Another uses: the efect of biocides
35. Reunião de esclarecimiento SIM
277 AFFECTED
4 DECEASED
FERNANDO CEBRIÁN, Head of Environmental
Laboratory Service. of the Castilla-La Mancha
Community Council.
Quick analysis help to early identification of
potential risk sources in this Legionella
outbreak, by reducing decision making process
and the exposure time of the population.
FIRST CASES,
12/11/2015
MUESTRAS LAST
CASE
SOCIAL
ALARM
ACTIONS
END
02/03/2016
LEGIPID® RESULTS
2 DAYS
52 DAYS
Outbreak: Manzanares
2015-2016
How could the inspection control an outbreak
as aggressive as the Manzanares outbreak.
41. Reunião de esclarecimiento SIMReunião de esclarecimiento SIM
Ultra Legionella Inmunoanalysis
System for Early Sensing (ULISENS)
42. Reunião de esclarecimiento SIMReunião de esclarecimiento SIM
Ultra Legionella Inmunoanalysis System for
Early Sensing (ULISENS)
“This project has received funding from the European Union’s
Horizon 2020 research and innovation programme under
grant agreement No 726499”.
43. Reunião de esclarecimiento SIMReunião de esclarecimiento SIM
SAMPLE TAKING
Outside water loop
FILTRATION
Filtration of a defined water
Sample volume and its elution
on the reagents cartridge
ANALYSIS
Robotized arm manages
the reagents
RESULT
Report of CFUeq which could
be remotely managed together
with other parameters
44. Reunião de esclarecimiento SIMReunião de esclarecimiento SIM
Legiolab is an online system
for Cooling Tower Legionella
detection and quantification.
Legiolab automatically
performs sampling and
analysis of Legionella providing
a reliable result within an hour.
45. Reunião de esclarecimiento SIMReunião de esclarecimiento SIM
•Early alert system/ continuous monitorization
• Integration with movile phones
• Results in situ
• Aplication in other pathogen
• Use in field and/or laboratory
• Combines sample preparation and analysis
• Integration with other paramethers
• Action of Cleaning and Disinfection
Profit