Plant biostimulants - Determination of Escherichia coli

This document gives general guidelines for the detection and identification of the specified microorganism Escherichia coli in technical and formulated biostimulant products, both in liquid and solid state, and also the horizontal method for the enumeration of ß-glucuronidase-positive Escherichia coli in plant biostimulants products (both in liquid and solid state).
The qualitative method described in this document is based on the detection of Escherichia coli in a non-selective liquid medium (enrichment broth), followed by isolation on a selective agar medium. Other methods can be appropriate, depending on the level of detection required.
NOTE   For the detection of Escherichia coli, subcultures can be performed on non-selective culture media followed by suitable identification steps (e.g. using identification kits).
The quantitative method described in this document uses a colony-count technique at 44 °C on a solid medium containing a chromogenic ingredient for detection of the enzyme ß-glucuronidase.
WARNING - Strains of Escherichia coli which do not grow at 44 °C and, in particular, those that are ß-glucuronidase negative, such as Escherichia coli O157, will not be detected.

Pflanzen-Biostimulanzien - Bestimmung von Escherichia coli

Dieses Dokument enthält allgemeine Leitlinien für den Nachweis und die Identifizierung des festgelegten Mikroorganismus Escherichia coli in technischen und formulierten Produkten von Biostimulanzien (sowohl in flüssigem als auch in festem Zustand) sowie das horizontale Verfahren für die Zählung von β Glucuronidase-positiven Escherichia coli in Produkten von Pflanzen-Biostimulanzien (sowohl in flüssigem als auch in festem Zustand).
Das in diesem Dokument beschriebene qualitative Verfahren basiert auf dem Nachweis von Escherichia coli in einem nicht-selektiven flüssigen Medium (Anreicherungsbouillon), gefolgt von der Isolierung auf einem selektiven Agar. Je nach gewünschtem Nachweisgrad können auch andere Methoden geeignet sein.
ANMERKUNG   Für den Nachweis von Escherichia coli können Subkulturen auf nicht-selektiven Nährmedien durchgeführt werden, gefolgt von geeigneten Identifizierungsschritten (z. B. unter Verwendung von Identifizierungskits).
Das in diesem Dokument beschriebene quantitative Verfahren verwendet ein Koloniezählverfahren bei 44 °C auf einem festen Medium, das einen chromogenen Bestandteil zum Nachweis des Enzyms β Glucuronidase enthält.
WARNUNG — Stämme von Escherichia coli, die bei 44 °C nicht wachsen, und insbesondere solche, die β Glucuronidase-negativ sind, wie z. B. Escherichia coli O157, werden nicht erkannt.

Biostimulants des végétaux - Détermination des Escherichia coli

Le présent document donne des lignes directrices générales pour la détection et l’identification du microorganisme spécifié Escherichia coli dans les produits biostimulants techniques et formulés, aussi bien à l’état liquide qu’à l’état solide, ainsi que la méthode horizontale pour le dénombrement des Escherichia coli ß-glucuronidase-positive dans les produits biostimulants des végétaux (aussi bien à l’état liquide qu’à l’état solide).
La méthode qualitative décrite dans le présent document repose sur la détection des Escherichia coli dans un milieu liquide non sélectif (bouillon d’enrichissement), suivie de l’isolement sur un milieu gélosé sélectif. D’autres méthodes peuvent être appropriées, en fonction du niveau de détection requis.
NOTE   Pour la détection des Escherichia coli, des subcultures peuvent être effectuées sur des milieux de culture non sélectifs, suivies d’étapes d’identification appropriées (par exemple, à l’aide de kits d’identification).
La méthode quantitative décrite dans le présent document utilise une technique de dénombrement des colonies à 44 °C sur un milieu solide contenant un ingrédient chromogène pour la détection de l’enzyme ß-glucuronidase.
AVERTISSEMENT - Certaines souches d’Escherichia coli qui ne poussent pas à 44 °C et, en particulier, celles qui sont ß-glucuronidase-négative, telles que les Escherichia coli O157, ne seront pas mises en évidence.

Rastlinski biostimulanti - Določanje Escherichia coli

General Information

Not Published
Publication Date
Current Stage
5020 - Submission to Vote - Formal Approval
Start Date
Due Date
Completion Date


Buy Standard

prEN 17716:2023
English language
25 pages
sale 10% off
sale 10% off
e-Library read for
1 day

Standards Content (Sample)

oSIST prEN 17716:2023
Rastlinski biostimulanti - Ugotavljanje prisotnosti in števila Escherichia coli
Plant biostimulants - Determination of Escherichia coli
Pflanzen-Biostimulanzien - Bestimmung von Escherichia coli
Biostimulants des végétaux - Détermination de Escherichia coli
Ta slovenski standard je istoveten z: prEN 17716
65.080 Gnojila Fertilizers
oSIST prEN 17716:2023 en,fr,de
2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.

oSIST prEN 17716:2023
oSIST prEN 17716:2023
prEN 17716
April 2023
ICS 65.080 Will supersede CEN/TS 17716:2022
English Version
Plant biostimulants - Determination of Escherichia coli
Biostimulants des végétaux - Détermination de Pflanzen-Biostimulanzien - Bestimmung von
Escherichia coli Escherichia coli
This draft European Standard is submitted to CEN members for enquiry. It has been drawn up by the Technical Committee
CEN/TC 455.
If this draft becomes a European Standard, CEN members are bound to comply with the CEN/CENELEC Internal Regulations
which stipulate the conditions for giving this European Standard the status of a national standard without any alteration.

This draft European Standard was established by CEN in three official versions (English, French, German). A version in any other
language made by translation under the responsibility of a CEN member into its own language and notified to the CEN-CENELEC
Management Centre has the same status as the official versions.

CEN members are the national standards bodies of Austria, Belgium, Bulgaria, Croatia, Cyprus, Czech Republic, Denmark, Estonia,
Finland, France, Germany, Greece, Hungary, Iceland, Ireland, Italy, Latvia, Lithuania, Luxembourg, Malta, Netherlands, Norway,
Poland, Portugal, Republic of North Macedonia, Romania, Serbia, Slovakia, Slovenia, Spain, Sweden, Switzerland, Türkiye and
United Kingdom.
Recipients of this draft are invited to submit, with their comments, notification of any relevant patent rights of which they are
aware and to provide supporting documentation.

Warning : This document is not a European Standard. It is distributed for review and comments. It is subject to change without
notice and shall not be referred to as a European Standard.



CEN-CENELEC Management Centre: Rue de la Science 23, B-1040 Brussels
© 2023 CEN All rights of exploitation in any form and by any means reserved Ref. No. prEN 17716:2023 E
worldwide for CEN national Members.

oSIST prEN 17716:2023
prEN 17716:2023 (E)
Contents Page
European foreword . 4
Introduction . 5
1 Scope . 6
2 Normative references . 6
3 Terms and definitions . 6
4 Principle . 7
4.1 Qualitative method . 7
4.2 Quantitative method . 7
5 Diluent and culture media . 7
5.1 General . 7
5.2 Broth and culture media in the qualitative method . 8
5.2.1 Enrichment broth . 8
5.2.2 Selective culture media: Tryptone-bile-glucuronic medium (TBX) for isolation of
Escherichia coli . 8
5.3 Diluent and culture media in the quantitative method . 8
5.3.1 Diluent . 8
5.3.2 Culture media: Tryptone-bile-glucuronic medium (TBX) . 8
6 Apparatus and glassware . 8
7 Handling of plant biostimulants products and sampling . 8
8 Procedure. 8
8.1 General . 8
8.2 Qualitative method . 9
8.2.1 General . 9
8.2.2 Solid formulations: wettable powder (WP), water dispersible granules (WDG),
pellets, granules, microgranules (slow release) formulation . 9
8.2.3 Liquid formulations: water-based formulations and oil-based (emulsifiable
concentrate - EC) formulations . 9
8.2.4 Incubation of the inoculated enrichment broth. 9
8.2.5 Detection and identification of Escherichia coli . 9
8.3 Quantitative method . 10
8.3.1 Test portion and initial suspension . 10
8.3.2 Serial dilutions . 11
8.3.3 Inoculation (surface plate method) and incubation . 11
8.3.4 Counting the colony-forming units . 11
9 Expression of results . 11
9.1 Expression of results in the qualitative test . 11
9.2 Expression of results in the quantitative test . 12
10 Test report . 12
Annex A (informative) Enrichment broth in the qualitative method . 14
A.1 General . 14
oSIST prEN 17716:2023
prEN 17716:2023 (E)
A.2 Enrichment broth . 14
A.3 Other enrichment broth . 15
Annex B (informative) Selective agar medium in the qualitative and quantitative method . 16
B.1 General . 16
B.2 Selective agar medium for isolation . 16
B.3 Other selective agar medium for isolation . 17
B.4 Selective agar medium for confirmation. 17
Annex C (informative) Diluent in the quantitative method . 19
C.1 General . 19
C.2 Phosphate buffered saline (PBS) . 19
Annex D (informative) Neutralization of the antimicrobial properties of the product . 20
D.1 General . 20
D.2 Suitability of the detection method . 20
D.3 Interpretation of suitability test results . 21
Annex E (informative) Interlaboratory study . 22
E.1 Materials used in the interlaboratory study . 22
E.2 Results of the interlaboratory study . 22
Annex ZA (informative) Relationship of this European Standard and the essential
requirements of Regulation (EU) 2019/1009 making available on the market of EU
fertilising products aimed to be covered . 24
Bibliography . 25
oSIST prEN 17716:2023
prEN 17716:2023 (E)
European foreword
This document (prEN 17716:2023) has been prepared by Technical Committee CEN/TC 455 “Plant
Biostimulants”, the secretariat of which is held by AFNOR.
This document is currently submitted to the CEN enquiry.
This document will supersede CEN/TS 17716:2022.
This document has been prepared under a Standardization Request given to CEN by the European
Commission and the European Free Trade Association, and supports essential requirements of EU
Directive(s) / Regulation(s).
For relationship with EU Directive(s) / Regulation(s), see informative Annex ZA, which is an integral part
of this document.
oSIST prEN 17716:2023
prEN 17716:2023 (E)
This document was prepared by the experts of CEN/TC 455 “Plant Biostimulants”. The European
Committee for Standardization (CEN) was requested by the European Commission (EC) to draft
European standards or European standardization deliverables to support the implementation of
Regulation (EU) 2019/1009 of 5 June laying down rules on the making available on the market of EU
fertilizing products (“FPR” or “Fertilising Products Regulation”). This Standardization Request, presented
as SR M/564 and M/564 Amd1, also contributes to the Communication on “Innovating for Sustainable
Growth: A Bio economy for Europe”. Working Group 5 “Labelling and denominations”, was created to
develop a work program as part of this request.
Technical committee CEN/TC 455 “Plant Biostimulants” was established to carry out the work program
that will prepare a series of standards. The interest in biostimulants has increased significantly in Europe
as a valuable tool to use in agriculture. Standardization was identified as having an important role in
order to promote the use of biostimulants. The work of CEN/TC 455 seeks to improve the reliability of
the supply chain, thereby improving the confidence of farmers, industry, and consumers in biostimulants,
and will promote and support commercialisation of the European biostimulant industry.
Biostimulants used in agriculture can be applied in multiple ways: on soil, on plants, as seed treatment,
etc. A microbial plant biostimulant consists of a microorganism or a consortium of microorganisms, as
referred to in Component Material Category 7 of Annex II of the EU Fertilising Products Regulation.
This document is applicable to all microbial biostimulants in agriculture.
Table 1 summarizes many of the agro-ecological principles and the role played by biostimulants.
Table 1 — Agro-ecological principles and the role played by biostimulants [1]
Increase biodiversity
By improving soil microorganism quality/quantity
Reinforce biological regulation and interactions
By reinforcing plant-microorganism interactions
— symbiotic exchanges i.e. mycorrhize
— symbiotic exchanges i.e. rhizobiaciae/fava
— secretions mimicking plant hormones (i.e. trichoderma)
By regulating plant physiological processes
— e.g. growth, metabolism, plant development
Improve biogeochemical cycles
— improve absorption of nutritional elements
— improve bioavailability of nutritional elements in the soil
— stimulate degradation of organic matter
WARNING — Persons using this document should be familiar with normal laboratory practice. This
document does not purport to address all of the sa

Questions, Comments and Discussion

Ask us and Technical Secretary will try to provide an answer. You can facilitate discussion about the standard in here.