Water quality - Guidance on field and laboratory procedures for quantitative analysis and identification of macroinvertebrates from inland surface waters

This standard gives guidance on the estimation of abundance and identification of macro-invertebrates in samples taken from inland waters. The procedure deals with pre-treatment (cleaning), sub-sampling, sorting and final identification of organisms from preserved and live samples originating from natural habitats or artificial substrates.

Wasserbeschaffenheit - Anleitung zu Feld- und Laborverfahren zur quantitativen Analyse und Identifizierung von Makroinvertebraten aus Inland-Oberflächengewässern

Dieses Dokument ist eine Anleitung zur quantitativen Beurteilung der Abundanz und Bestimmung von Makroinvertebraten in Proben aus Binnengewässern. Das Verfahren befasst sich mit der Vorbehandlung (Reinigung), Herstellen von Teilproben, Sortierung und endgültigen Bestimmung von Organismen aus konservierten und lebenden Proben, die aus natürlichen Lebensräumen oder künstlichen Substraten stammen, und ihrem Transport zum Labor. Für die DNA-Analyse wird eine spezifische Anleitung zur Konservierung gegeben.

Qualité de l'eau - Guide sur les modes opératoires de terrain et de laboratoire pour l'analyse quantitative et d'identification des macro-invertébrés des eaux de surface intérieures

Le présent document donne des conseils sur l’estimation quantitative de l’abondance et l’identification des macro-invertébrés dans des échantillons provenant de milieux aquatiques continentaux. La procédure s’applique au prétraitement (nettoyage), au sous-échantillonnage, au tri et à l’identification finale des organismes provenant d’échantillons avec ou sans conservateurs, issus d’habitats naturels ou d’échantillonneurs de colonisation, ainsi qu’à leur transport au laboratoire. Des conseils spécifiques sont donnés au sujet de la conservation en vue de l’analyse ADN.

Kakovost vode - Navodilo za terenske in laboratorijske postopke kvantitativne analize in identifikacije velikih nevretenčarjev v celinskih površinskih vodah

Ta standard podaja navodilo za oceno številčnosti in identifikacijo velikih nevretenčarjev v vzorcih iz celinskih voda. Postopek obravnava predobdelavo (čiščenje), podvzorčenje, razvrščanje in dokončno identifikacijo organizmov iz ohranjenih in živih vzorcev, ki izhajajo iz naravnih habitatov ali umetnih substratov.

General Information

Status
Published
Public Enquiry End Date
04-Aug-2017
Publication Date
10-Apr-2019
Technical Committee
Current Stage
6060 - National Implementation/Publication (Adopted Project)
Start Date
13-Mar-2019
Due Date
18-May-2019
Completion Date
11-Apr-2019

Buy Standard

Standard
SIST EN 17136:2019 - BARVE na PDF-str 18,19
English language
18 pages
sale 10% off
Preview
sale 10% off
Preview

e-Library read for
1 day

Standards Content (sample)

2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.YDQWLWDWLYQHWasserbeschaffenheit - Anleitung zu Feld- und Laborverfahren zur quantitativen Analyse und Identifizierung von Makroinvertebraten aus Inland-OberflächengewässernQualité de l'eau - Guide sur les modes opératoires de terrain et de laboratoire pour l'analyse quantitative et d'identification des macro-invertébrés des eaux de surface intérieuresWater quality - Guidance on field and laboratory procedures for quantitative analysis and identification of macroinvertebrates from inland surface waters13.060.70Preiskava bioloških lastnosti vodeExamination of biological properties of water13.060.10Voda iz naravnih virovWater of natural resourcesICS:Ta slovenski standard je istoveten z:EN 17136:2019SIST EN 17136:2019en,fr,de01-maj-2019SIST EN 17136:2019SLOVENSKI

STANDARD
SIST EN 17136:2019
EUROPEAN STANDARD NORME EUROPÉENNE EUROPÄISCHE NORM
EN 17136
February
t r s { ICS
s uä r x rä s r English Version

Water quality æ Guidance on field and laboratory procedures for quantitative analysis and identification of macroinvertebrates from inland surface waters Qualité de l 5eau æ Guide sur les procédures de terrain et de laboratoire pour l 5analyse quantitative et l 5identification des macroæinvertébrés des eaux de surface continentales

Wasserbeschaffenheit æ Anleitung zu Feldæ und Laborverfahren zur quantitativen Analyse und Bestimmung von Makroinvertebraten aus Binnenoberflächengewässern This European Standard was approved by CEN on

s v December
t r s zä

egulations which stipulate the conditions for giving this European Standard the status of a national standard without any alterationä Upætoædate lists and bibliographical references concerning such national standards may be obtained on application to the CENæCENELEC Management Centre or to any CEN memberä

translation under the responsibility of a CEN member into its own language and notified to the CENæCENELEC Management Centre has the same status as the official versionsä

CEN members are the national standards bodies of Austriaá Belgiumá Bulgariaá Croatiaá Cyprusá Czech Republicá Denmarká Estoniaá Finlandá Former Yugoslav Republic of Macedoniaá Franceá Germanyá Greeceá Hungaryá Icelandá Irelandá Italyá Latviaá Lithuaniaá Luxembourgá Maltaá Netherlandsá Norwayá Polandá Portugalá Romaniaá Serbiaá Slovakiaá Sloveniaá Spainá Swedená Switzerlandá Turkey and United Kingdomä

EUROPEAN COMMITTEE FOR STANDARDIZATION COMITÉ EUROPÉEN DE NORMALISATION EUROPÄISCHES KOMITEE FÜR NORMUNG

CEN-CENELEC Management Centre:
Rue de la Science 23,
B-1040 Brussels

t r s { CEN All rights of exploitation in any form and by any means reserved worldwide for CEN national Membersä Refä Noä EN

s y s u xã t r s { ESIST EN 17136:2019

EN 17136:2019 (E) 2 Contents Page European foreword ....................................................................................................................................................... 3 Introduction .................................................................................................................................................................... 4 1 Scope .................................................................................................................................................................... 5 2 Normative references .................................................................................................................................... 5 3 Terms and definitions ................................................................................................................................... 5 4 Principle ............................................................................................................................................................. 5 5 Equipment, preservatives and reagents ................................................................................................. 6 5.1 Sieves ................................................................................................................................................................... 6 5.2 Microscope......................................................................................................................................................... 6 5.3 Fixatives and preservatives ......................................................................................................................... 6 5.3.1 General ................................................................................................................................................................ 6 5.3.2 Formaldehyde, 37 % (volume fraction) .................................................................................................. 6 5.3.3 Ethanol, C2H5OH ............................................................................................................................................. 7 5.4 Reagents for examination using compound microscopes ................................................................ 7 5.4.1 General ................................................................................................................................................................ 7 5.4.2 Koenike ............................................................................................................................................................... 7 5.4.3 Laevulose syrup ............................................................................................................................................... 7 6 Pre-laboratory procedure ............................................................................................................................ 7 6.1 General ................................................................................................................................................................ 7 6.2 Sample processing in the field .................................................................................................................... 8 6.3 Transport and storing of samples ............................................................................................................. 8 7 Laboratory procedure ................................................................................................................................... 9 7.1 General ................................................................................................................................................................ 9 7.2 Pre-treatment prior to sorting ................................................................................................................ 10 7.2.1 General ............................................................................................................................................................. 10 7.2.2 Pre-sorting ...................................................................................................................................................... 11 7.2.3 Rinsing ............................................................................................................................................................. 11 7.2.4 Decanting ........................................................................................................................................................ 11 7.2.5 Sub-sampling ................................................................................................................................................. 11 7.3 Quantitative sorting .................................................................................................................................... 12 7.3.1 General ............................................................................................................................................................. 12 7.3.2 Quantitative sorting on a tray .................................................................................................................. 13 7.3.3 Quantitative sorting using a stereo-zoom microscope ................................................................... 14 7.3.4 Screening ......................................................................................................................................................... 14 8 Identification ................................................................................................................................................. 14 8.1 General ............................................................................................................................................................. 14 8.2 Arachnida ........................................................................................................................................................ 15 8.3 Chironomids ................................................................................................................................................... 15 8.4 Oligochaetes ................................................................................................................................................... 15 8.5 Platyhelminthes ............................................................................................................................................ 15 9 Quality assurance ......................................................................................................................................... 15 Annex A (informative)

Examples of sample splitters and sorting tray .................................................... 16 Bibliography ................................................................................................................................................................. 18

SIST EN 17136:2019

EN 17136:2019 (E) 3 European foreword This document (EN 17136:2019) has been prepared by Technical Committee CEN/TC 230 “Water analysis”, the secretariat of which is held by DIN. This European Standard shall be given the status of a national standard, either by publication of an identical text or by endorsement, at the latest by August 2019, and conflicting national standards shall be withdrawn at the latest by August 2019. 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. According to the CEN-CENELEC Internal Regulations, the national standards organisations of the following countries are bound to implement this European Standard: Austria, Belgium, Bulgaria, Croatia, Cyprus, Czech Republic, Denmark, Estonia, Finland, Former Yugoslav Republic of Macedonia, France, Germany, Greece, Hungary, Iceland, Ireland, Italy, Latvia, Lithuania, Luxembourg, Malta, Netherlands, Norway, Poland, Portugal, Romania, Serbia, Slovakia, Slovenia, Spain, Sweden, Switzerland, Turkey and the United Kingdom. SIST EN 17136:2019

EN 17136:2019 (E) 4 Introduction WARNING — Working in or around water is inherently dangerous. Care should be taken when working with different and generally toxic preservatives. This standard does not purport to address the safety problems associated with its use. It is the responsibility of the user to adopt appropriate health and safety practices in compliance with national regulatory conditions. Macroinvertebrates are important in water quality assessment and form one of the biological quality elements in the EC Water Framework Directive (WFD 2000/60/EC). Macroinvertebrates are a very diverse group of organisms for which an efficient and effective procedure for analysis is not always obvious. Their large size range, very different appearances and the diverse range of habitats they live in produce specific problems for collecting and analysis. This standard aims to provide a general procedure for the quantitative analysis of macroinvertebrate samples for the WFD in particular and scientific studies in general. It is not intended to give a full and exact overview of all possible methods. Although specific situations and investigation objectives may require adjustments to the outlined procedure the general principles given in this guideline should still be valid. SIST EN 17136:2019

EN 17136:2019 (E) 5 1 Scope This document gives guidance on the quantitative estimation of abundance and identification of macroinvertebrates in samples taken from inland waters. The procedure deals with pre-treatment (cleaning), sub-sampling, sorting, and final identification of organisms from preserved and unpreserved samples originating from natural habitats or artificial substrates and their transport to the laboratory. Specific guidance is given for preservation for DNA-analysis. 2 Normative references There are no normative references in this document. 3 Terms and definitions For the purposes of this document, the following terms and definitions apply. ISO and IEC maintain terminological databases for use in standardization at the following addresses: — IEC Electropedia: available at http://www.electropedia.org/ — ISO Online browsing platform: available at http://www.iso.org/obp

3.1 fixation protection from disintegration of the morphological structure of organisms 3.2 macroinvertebrate invertebrate that is easily visible without magnification (> 0,5 mm) and generally belongs to the group of organisms that live for at least one life stage on or in the bottom substrate or vegetation of inland surface waters 3.3 matrix components of a sample other than the macroinvertebrates 3.4 preservation process that protects organic substances from decay 4 Principle The identification and quantification of macroinvertebrates comprises both field and laboratory procedures. After an optional pre-treatment of rinsing and sieving in the field, macroinvertebrate samples are transported to the laboratory where the samples are further processed. Macroinvertebrates are taken out of the matrix material by a sample dependent technique, quantified and sorted in functional groups for identification using a microscope or DNA-analysis. Several techniques exist to clean (remove unwanted matrix material) the sample and sort the organisms. The most suitable technique should be selected by visual inspection of the individual sample. Dependent on the objective of the analysis samples can be preserved in the field or cooled and processed unpreserved. SIST EN 17136:2019

EN 17136:2019 (E) 6 5 Equipment, preservatives and reagents 5.1 Sieves Each new sieve should be checked for the right mesh size before first use. The smallest (typically 500 µm) sieve (having a significant effect on the validity of the analysis) is critical, its mesh should be guaranteed by the manufacturer attestation or control, providing metrological traceability. It should regularly visually be inspected for visible damage before every use For DNA-based studies the removal of larger, visible contaminants by washing the equipment in water only is not sufficient. Instead sieves should be cleaned preferably with an appropriate bleach solution for each consecutive sample location. 5.2 Microscope A low-power stereo-zoom microscope (typical magnification ~10x to 60x) can be used for accurate quantitative sorting and most identifications. For the species identification of specific groups like Chironomidae and Oligochaeta a compound microscope (typical magnification 40x to 400x) should be used. 5.3 Fixatives and preservatives 5.3.1 General There is a distinct difference between fixation and preservation. In general, fixation is preferred for morphological analysis. Within the context of freshwater macroinvertebrate analysis ethanol as a preservative is the most often used chemical for preserving samples for quantification and identification. When ethanol is used as a preservative formaldehyde fixation can be left out for macroinvertebrate analysis. WARNING — Formaldehyde is hazardous to humans (CMR substance, see ECHA) even at very low concentrations in air. Formaldehyde fixation should be avoided as much as possible. Preservation by ethanol and formaldehyde fixation affects the estimation of biomass. Although weight loss by ethanol is considered to be more significant than by formaldehyde no unambiguous scientific data are available on the exact effect of either preservative. Therefore, prior to estimating biomass on the basis of preserved organisms the bias by using preserved material should be assessed for the specific taxonomic groups under investigation and the relevant conditions of sampling and preservation. After the fixative or preservative has been added the sample should be gently turned to disperse the preservative. 5.3.2 Formaldehyde, 37 % (volume fraction) Formaldehyde 37 % should be added to an undrained sample with a final concentration of 4 % to 6 %. This concentration provides a good fixation of organisms (avoiding damaging and shrinking). Formaldehyde is not a suitable fixative for DNA analysis. Formaldehyde samples should always be thoroughly rinsed with water on a sieve prior to processing. Long-term storage in formaldehyde can seriously affect the colour of specimens and dissolves the shells of Mollusca (if not buffered). To prevent this, formaldehyde fixed samples can be transferred to 70 % ethanol. Transferring the sample to ethanol can already be done after 24 h of fixation. The sample should be thoroughly rinsed with water to remove the free formaldehyde before adding the ethanol. SIST EN 17136:2019

EN 17136:2019 (E) 7 5.3.3 Ethanol, C2H5OH Before addition of ethanol the sample should be drained to minimize the dilution with excess sample water. In most cases ethanol 96 % should be added to compensate for dilution by the residual sample water. In samples with absorbing material and/or relatively large organisms (e.g. numerous molluscs) the initial 96 % ethanol should be replaced after a few days to ensure a permanent concentration of approximately 70 % in the sample. The sample container should not be filled for more than one third with material. Ethanol should be added until the sample is completely immersed and the container is filled to achieve a concentration of 75 %. Ethanol concentrations well above 70 % will harden many organisms rendering them difficult to manipulate and identify. Ethanol concentrations below 70 % will lead to tissue degradation. Initial ethanol concentrations should be checked sample-wise with an alcohol meter. Unfixed macroinvertebrate samples preserved with ethanol can be stored for several years. Macroinvertebrate samples used for DNA studies or DNA-based identification that need to be stored for extended times before analysis (e.g. DNA barcoding) should be placed within two days of sampling in

· 90 % ethanol and preferably stored in a cool and dark place. At concentrations below 80 % DNA will degenerate. 5.4 Reagents for examination using compound microscopes 5.4.1 General Many chemicals are known to enhance the microscope image quality in order to be able to accurately describe diagnostic features. The most relevant chemicals including mounting media can be found in group specific identification literature. Koenike and Laevulose syrup, presented below, are the most often used reagents. Koenike can also be used as a preservative for Arachnida (mites). 5.4.2 Koenike Mix 50 ml of glycerol with 30 ml of water and add 20 ml of acetic acid. This clear solution can be stored at room temperature for an unlimited period of time. 5.4.3 Laevulose syrup Dilute 25 g D(-)Fructose in 25 ml water by proper stirring (magnetic stirring bar). The solution can be slightly heated. Thereafter add 25 ml of lactic acid and stir again. This clear solution can be stored at room temperature for an unlimited period of time. 6 Pre-laboratory procedure 6.1 General The pre-laboratory procedure starts with a visual inspection of the sample to assess the amount of matrix material (silt, clay, sand, macrophytes) and the number of organisms. With this information the right procedure for pre-treatment and sorting can be selected. It can be done in the field directly after sampling or in the laboratory. The general objective is to select the organisms in the most effective way (least amount of time). If sub-sampling is necessary it should be done in a random way. When not all organisms have to be identified selectivity in sorting of taxa should be avoided. Before collecting the organisms the sample should be rinsed and cleaned and/or sub-divided in smaller portions. In case of unpreserved samples macroinvertebrates are collected alive in larger trays with the naked eye or with the use of a magnifying glass. SIST EN 17136:2019

EN 17136:2019 (E) 8 In case of preserved sampl
...

Questions, Comments and Discussion

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