CEN/TS 17715:2022
(Main)Plant biostimulants - Detection of Shigella spp.
Plant biostimulants - Detection of Shigella spp.
This document provides a method for verifying that the pathogen Shigella spp. is not present in microbial plant biostimulants in a concentration that exceeds the respective limits outlined in the EU Regulation on Fertilising Products.
The detection method for Shigella pathogens is not sensitive and quantification is rarely performed. Detection is usually performed using an enrichment medium followed by subculturing onto a variety of selective media.
Pflanzen-Biostimulanzien - Nachweis von Shigella spp.
Dieses Dokument liefert ein Verfahren zum Nachweis, dass das Pathogen Shigella spp. in mikrobiellen Pflanzen-Biostimulanzien nicht in einer Konzentration vorhanden ist, die die jeweiligen Grenzwerte überschreitet, die in der EU-Düngeprodukteverordnung festgelegt sind.
Das Nachweisverfahren für Shigella-Pathogene ist nicht trennscharf, und eine Quantifizierung wird selten durchgeführt. Der Nachweis erfolgt in der Regel mit einem Anreicherungsmedium und anschließender Subkultivierung auf verschiedenen Selektivmedien.
Biostimulants des végétaux - Recherche de Shigella spp.
Le présent document fournit une méthode permettant de vérifier que le pathogène Shigella spp n’est pas présent dans les biostimulants microbiens des végétaux à une concentration dépassant les limites correspondantes indiquées dans le Règlement UE sur les fertilisants.
La méthode de détection des pathogènes Shigella n’est pas sensible et la quantification est rarement réalisée. La détection est généralement réalisée en utilisant un milieu d’enrichissement, puis en procédant à un repiquage sur une variété de milieux sélectifs.
Rastlinski biostimulanti - Ugotavljanje prisotnosti Shigella spp.
Ta dokument zagotavlja metodo za preverjanje, da patogen Shigella spp. ni prisoten v mikrobnih rastlinskih biostimulantih v koncentraciji, ki presega ustrezne omejitve, določene v Uredbi EU o sredstvih za gnojenje.
Metoda ugotavljanja prisotnosti patogenov Shigella ni občutljiva in kvantifikacija se redko izvaja. Prisotnost patogenov se običajno ugotavlja z uporabo obogatitvenega gojišča, ki mu sledi subkultura na različnih selektivnih gojiščih.
General Information
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Standards Content (Sample)
SLOVENSKI STANDARD
01-februar-2023
Rastlinski biostimulanti - Ugotavljanje prisotnosti Shigella spp.
Plant biostimulants - Detection of Shigella spp
Pflanzen-Biostimulanzien - Nachweis von Shigella spp.
Biostimulants des végétaux - Recherche de Shigella spp.
Ta slovenski standard je istoveten z: CEN/TS 17715:2022
ICS:
65.080 Gnojila Fertilizers
2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.
CEN/TS 17715
TECHNICAL SPECIFICATION
SPÉCIFICATION TECHNIQUE
March 2022
TECHNISCHE SPEZIFIKATION
ICS 65.080
English Version
Plant biostimulants - Detection of Shigella spp.
Biostimulants des végétaux - Recherche de Shigella Pflanzen-Biostimulanzien - Nachweis von Shigella spp.
spp.
This Technical Specification (CEN/TS) was approved by CEN on 3 January 2022 for provisional application.
The period of validity of this CEN/TS is limited initially to three years. After two years the members of CEN will be requested to
submit their comments, particularly on the question whether the CEN/TS can be converted into a European Standard.
CEN members are required to announce the existence of this CEN/TS in the same way as for an EN and to make the CEN/TS
available promptly at national level in an appropriate form. It is permissible to keep conflicting national standards in force (in
parallel to the CEN/TS) until the final decision about the possible conversion of the CEN/TS into an EN is reached.
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, Turkey and
United Kingdom.
EUROPEAN COMMITTEE FOR STANDARDIZATION
COMITÉ EUROPÉEN DE NORMALISATIO N
EUROPÄISCHES KOMITEE FÜR NORMUN G
CEN-CENELEC Management Centre: Rue de la Science 23, B-1040 Brussels
© 2022 CEN All rights of exploitation in any form and by any means reserved Ref. No. CEN/TS 17715:2022 E
worldwide for CEN national Members.
Contents Page
European foreword . 3
Introduction . 4
1 Scope . 5
2 Normative references . 5
3 Terms and definitions . 5
4 Principle . 5
4.1 General. 5
4.2 Enrichment in selective liquid medium . 5
4.3 Plating out and identification of colonies . 5
4.4 Biochemical and serological confirmation . 6
5 Culture media, reagents and antisera . 6
6 Apparatus and glassware . 6
7 Sampling . 6
8 Procedure . 6
8.1 General. 6
8.2 Test portion . 6
8.3 Enrichment . 6
8.4 Plating out and colony selection . 7
8.5 Confirmation of colonies . 7
8.6 Serological confirmation . 12
9 Expression of results . 13
10 Test report . 13
Annex A (normative) Diagram of test procedure . 14
Annex B (normative) Composition and preparation of culture media and reagents . 15
Annex C (normative) Description of Shigella colony morphology and colour on selective agars, for
both identification and quality control purposes . 26
Bibliography . 27
European foreword
This document (CEN/TS 17715:2022) has been prepared by Technical Committee CEN/TC 455 “Plant
Biostimulants”, the secretariat of which is held by AFNOR.
Attention is drawn to the possibility that some of the elements of this document may be the subject of
patent rights. CEN shall not be held responsible for identifying any or all such patent rights.
This document has been prepared under a Standardization Request given to CEN by the European
Commission and the European Free Trade Association.
Any feedback and questions on this document should be directed to the users’ national standards body.
A complete listing of these bodies can be found on the CEN website.
According to the CEN/CENELEC Internal Regulations, the national standards organisations of the
following countries are bound to announce this Technical Specification: 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, Turkey and the United
Kingdom.
Introduction
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 2019 laying down rules on the making available on the market of
EU fertilizing products (“FPR” or “Fertilising Products Regulation”).
This standardization request, presented as M/564, also contributes to the Communication on “Innovating
for Sustainable Growth: A Bio economy for Europe”. The Working Group 5 “Labelling and
denominations”, was created to develop a work program as part of this request. The 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 commercialization 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.
The table below 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. Mycorrhizae
— symbiotic exchanges i.e. Rhizobiaceae/Faba
— 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 safety problems, if any, associated with its use. It is the
responsibility of the user to establish appropriate safety and health practices and to ensure compliance
with any national regulatory conditions.
IMPORTANT — It is absolutely essential that tests conducted in accordance with this document be
carried out by suitably trained staff.
1 Scope
This document provides a method for verifying that the pathogen Shigella spp. is not present in microbial
plant biostimulants in a concentration that exceeds the respective limits outlined in the EU Regulation on
Fertilising Products.
The detection method for Shigella pathogens is not sensitive and quantification is rarely performed.
Detection is usually performed using an enrichment medium followed by subculturing onto a variety of
selective media.
2 Normative references
The following documents are referred to in the text in such a way that some or all of their content
constitutes requirements of this document. For dated references, only the edition cited applies. For
undated references, the latest edition of the referenced document (including any amendments) applies.
CEN/TS 17708, Plant biostimulants — Preparation of sample for microbial analysis
CEN/TS 17724, Plant Biostimulants — Terminology
3 Terms and definitions
For the purposes of this document, the terms and definitions given in CEN/TS 17724 and the following
apply.
3.1
Shigella spp.
microorganisms which form typical colonies on solid selective media described and which display the
morphological, physiological and biochemical characteristics described when the analysis is carried out
in accordance with this document
3.2
detection of Shigella spp.
determination of the presence or absence of these microorganism in a particular mass of product, when
tests are carried out in accordance with this document
4 Principle
4.1 General
Detection of Shigella spp. will be conducted according to the sections specified in EN ISO 21567:2004 and
with the following four successive stages (see Annex A, Figure A.1).
4.2 Enrichment in selective liquid medium
A test portion is inoculated into Shigella broth containing 0,5 µg/ml of novobiocin, then incubated
anaerobically at (41,5 ± 1) °C for 16 h to 20 h.
4.3 Plating out and identification of colonies
From the enrichment culture obtained, three selective differential media are inoculated: MacConkey agar
with low selectivity; XLD agar with moderate selectivity; Hektoen enteric agar with the greatest
selectivity. All are incubated at 37 °C for 20 h to 24 h.
4.4 Biochemical and serological confirmation
Typical and suspect colonies are selected from each of the three selective agars. The colonies are purified
on nutrient agar, then biochemical and serological characterizations are carried out using the tests
described.
5 Culture media, reagents and antisera
Use only reagents of recognized analytical grade, unless otherwise specified, and distilled or
demineralized water or water of equivalent purity.
See Annex B for descriptions of all media, reagents and antisera.
Commercially available dehydrated media should give more consistent results than media prepared from
their component parts in the laboratory. Follow the manufacturer’s instructions exactly, as small changes
in preparation can significantly change the performance of selective media. Excessive heating of the
selective agars used in this document by autoclaving, storage and then re-heating for use may result in
loss of selectivity.
6 Apparatus and glassware
Disposable equipment is an acceptable alternative to re-usable glassware if it has suitable specifications.
Usual microbiological laboratory equipment according to CEN/TS 17708 shall be used.
7 Sampling
Sampling is not part of the method specified in this document (see the specific European
...
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