SIST EN 16698:2016
(Main)Water quality - Guidance on quantitative and qualitative sampling of phytoplankton from inland waters
Water quality - Guidance on quantitative and qualitative sampling of phytoplankton from inland waters
Development of a method for quantitative and qualitative sampling of phytoplankton from inland waters. The method includes all common existing European sampling strategies.
The main aspects for phytoplankton sampling in lakes covered by this EN are the effects of sampling on phytoplankton biomass and composition
- if sampled in different seasons;
- if euphotic or epilimnetic zone;
- if number of sampling sites for large water bodies is one or three;
- if mixing samples from depth-step-wise sampling or from integrated samplers or by flexible tubes for integrated sampling;
- if replicate sampling is 1 or 5 at one site (aspects of reproducibility);
- if sampling is done in unusual sampling designs as sampling from the shore side or at the outflow compared to the deepest point of the lake.
Wasserbeschaffenheit - Anleitung für die quantitative und qualitative Probenahme von Phytoplankton aus Binnengewässern
Diese Europäische Norm legt Verfahren für die Probenahme von Phytoplankton in Binnengewässern fest und beschreibt Verfahren von Probenahmetechniken für Phytoplankton in Binnengewässern (z. B. Flüssen und Kanälen bzw. Seen, Teichen, Stauseen und anderen künstlichen Wasserkörpern).
Diese Europäische Norm gibt eine Anleitung zur Probenahme von Phytoplankton für qualitative und quantitative limnologische Untersuchungen und die Überwachung der Wasserbeschaffenheit, z. B. des ökologischen Zustandes.
Qualité de l'eau - Lignes directrices sur l'échantillonnage quantitatif et qualitatif du phytoplancton dans les eaux intérieures
La présente Norme européenne spécifie les procédures d'échantillonnage du phytoplancton dans les eaux intérieures et décrit, en outre, des méthodes et des techniques d'échantillonnage du phytoplancton dans les eaux intérieures (par exemple les rivières et les canaux, ou les lacs, les étangs, les retenues d'eau et autres masses d'eau artificielles, respectivement).
La présente Norme européenne fournit des lignes directrices pour l'échantillonnage du phytoplancton en vue de mener des études limnologiques qualitatives et quantitatives et de surveiller la qualité de l'eau (par exemple, l'état écologique).
Kakovost vode - Navodilo za kvantitativno in kvalitativno vzorčenje fitoplanktona v celinskih vodah
Razvoj metode za kvantitativno in kvalitativno vzorčenje fitoplanktona v celinskih vodah. Metoda vključuje vse običajne obstoječe evropske strategije vzorčenja.
Glavni vidiki vzorčenja fitoplanktona v jezerih, ki jih pokriva ta standard EN, so učinki vzorčenja na biomaso in sestavo fitoplanktona
– če vzorčenje poteka v različnih letnih časih;
– če gre za evfotično ali epilimnetično cono;
– če je število mest vzorčenja za velika vodna telesa ena ali tri;
– če se mešajo vzorci globinskega vzorčenja po korakih ali iz integriranih vzorcev ali prek gibljivih cevi za integrirano vzorčenje;
– če je ponovljeno vzorčenje 1 ali 5 na enem mestu (vidiki možnosti reprodukcije);
– če je vzorčenje opravljeno v nenavadni obliki vzorčenja kot vzorčenje ob obali ali ob iztoku v primerjavi z najglobljo točko jezera.
General Information
Standards Content (Sample)
2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.Wasserbeschaffenheit - Anleitung für die quantitative und qualitative Probenahme von Phytoplankton aus BinnengewässernQualité de l'eau - Lignes directrices sur l'échantillonnage quantitatif et qualitatif du phytoplancton dans les eaux intérieuresWater quality - Guidance on quantitative and qualitative sampling of phytoplankton from inland 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 16698:2015SIST EN 16698:2016en,fr,de01-februar-2016SIST EN 16698:2016SLOVENSKI
STANDARD
SIST EN 16698:2016
EUROPEAN STANDARD NORME EUROPÉENNE EUROPÄISCHE NORM
EN 16698
October
t r s w ICS
s uä r x rä y r English Version
Water quality æ Guidance on quantitative and qualitative sampling of phytoplankton from inland waters Qualité de l 5eau æ Lignes directrices sur l 5échantillonnage quantitatif et qualitatif du phytoplancton dans les eaux intérieures
Wasserbeschaffenheit æ Anleitung für die quantitative und qualitative æ Probenahme von Phytoplankton aus Binnengewässern This European Standard was approved by CEN on
z August
t r s wä
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 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:
Avenue Marnix 17,
B-1000 Brussels
9
t r s w CEN All rights of exploitation in any form and by any means reserved worldwide for CEN national Membersä Refä Noä EN
s x x { zã t r s w ESIST EN 16698:2016
EN 16698:2015 (E) 2 Contents Page European foreword . 4 Introduction . 5 1 Scope . 6 2 Normative references . 6 3 Terms and definitions . 6 4 Principles of phytoplankton sampling . 8 4.1 General . 8 4.2 Selection of sampling sites . 9 4.2.1 General . 9 4.2.2 Sampling sites in rivers and streams . 9 4.2.3 Sampling sites in lakes . 9 4.3 Sampling frequency and replicates . 10 5 Equipment and preservatives . 10 6 Procedure. 11 6.1 General requirements for phytoplankton sampling . 11 6.2 Sampling in rivers . 12 6.3 Sampling in lakes . 12 6.3.1 General . 12 6.3.2 Sampling in polymictic lakes . 13 6.3.3 Sampling in stratified lakes during circulation. 13 6.3.4 Sampling in stratified lakes during phase of summer stagnation . 13 6.4 Preparation of mixed samples of a water column . 14 6.4.1 Preparation of mixed samples using an integral water sampler . 14 6.4.2 Preparation of mixed samples using other water samplers . 14 6.5 Bottling and fixation of samples . 15 6.6 Storage and transport of the samples . 15 6.7 Additional samples for analysis of diatoms . 16 6.8 Qualitative sampling . 16 7 Measurements of accompanying parameters . 16 7.1 General . 16 7.2 Secchi depth . 16 7.3 Water temperature . 17 7.4 Dissolved oxygen . 17 7.5 pH . 17 7.6 Chlorophyll-a . 17 8 Quality Assurance . 17 Annex A (informative)
Description of methodology . 18 A.1 Water colour, Secchi-depth and euphotic depth . 18 A.2 Secchi depth – practical hints . 19 A.3 Sampling frequency – examples. 19 Annex B (informative)
Examples for suitable water samplers . 22 B.1 Examples for sampling devices in rivers . 22 SIST EN 16698:2016
EN 16698:2015 (E) 3 B.1.1 General requirements . 22 B.1.2 Horizontal sampler . 22 B.2 Examples for sampling devices in lakes . 22 B.2.1 General requirements . 22 B.2.2 Hose sampler . 23 B.2.3 Tube integrating sampler . 25 B.2.4 Mechanical integrating water sampler . 27 B.2.5 Hydrostatic integrating water sampler . 27 B.2.6 Electronic integrating water sampler . 28 B.3 Sampling equipment cleaning . 29 Annex C (informative)
Determination of the depth gradient . 30 Annex D (informative)
Example for a sampling protocol . 32 Bibliography . 34
SIST EN 16698:2016
EN 16698:2015 (E) 4 European foreword This document (EN 16698:2015) 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 April 2016, and conflicting national standards shall be withdrawn at the latest by April 2016. Attention is drawn to the possibility that some of the elements of this document may be the subject of patent rights. CEN [and/or CENELEC] shall not be held responsible for identifying any or all such patent rights. This document has been prepared under a mandate given to CEN by the European Commission and the European Free Trade Association. According to the CEN-CENELEC Internal Regulations, the national standards organizations 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, Slovakia, Slovenia, Spain, Sweden, Switzerland, Turkey and the United Kingdom. SIST EN 16698:2016
EN 16698:2015 (E) 5 Introduction WARNING —Working in or around water is inherently dangerous, Persons using this European standard should be familiar with usual field and laboratory practice. This standard 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 health and safety practices and to ensure compliance with any national regulatory guidelines. IMPORTANT — It is absolutely essential that tests conducted according to this European Standard be carried out by suitably trained staff. Series of phytoplankton samples provide information on the taxonomic composition as well as the spatial occurrence of the individual taxa and their relative abundances. They allow the calculation of the biomass of the individual taxa as well as for the phytoplankton assemblage as a whole. For the purpose of limnological investigations like monitoring and status assessment of surface waters representative phytoplankton samples are necessary. Therefore suitable sampling methods and monitoring strategies are needed depending on the objectives of the investigation and the given natural conditions. SIST EN 16698:2016
EN 16698:2015 (E) 6 1 Scope This European Standard specifies procedures for phytoplankton sampling in inland waters and describes methods of sampling techniques for phytoplankton in inland waters (e.g. rivers and channels, or lakes, ponds, reservoirs and other artificial water bodies, respectively). This European Standard gives guidance for sampling of phytoplankton for qualitative and quantitative limnological investigations and monitoring of water quality, e.g. ecological status. 2 Normative references The following documents, in whole or in part, are normatively referenced in this document and are indispensable for its application. For dated references, only the edition cited applies. For undated references, the latest edition of the referenced document (including any amendments) applies. EN 15204, Water quality — Guidance standard on the enumeration of phytoplankton using inverted microscopy (Utermöhl technique) EN ISO 5814, Water quality — Determination of dissolved oxygen — Electrochemical probe method (ISO 5814) EN ISO 7027, Water quality — Determination of turbidity (ISO 7027) EN ISO 10523, Water quality — Determination of pH (ISO 10523) ISO 17289, Water quality — Determination of dissolved oxygen — Optical sensor method 3 Terms and definitions For the purposes of this document, the following terms and definitions apply. 3.1 deep chlorophyll maximum DCM local chlorophyll maximum below the epilimnion where the maximum chlorophyll concentration is at least 1,5 times higher than the average chlorophyll concentration measured in the epilimnion 3.2 depth gradient parameter to distinguish between polymictic and di- or monomictic lakes as a measure for the mixing intensity of a water body calculated as the quotient of maximum depth and theoretical epilimnion depth
Note 1 to entry: See Annex C. Note 2 to entry: Values > 1,5 indicate a thermally stable stratified lake. For further details see Annex C and [18]. 3.3 dimictic lake lake with uniform water temperature and holomictic conditions twice a year: during autumn (before ice cover) and during spring (after ice cover) 3.4 epilimnion zone of the water body between surface and thermocline in which the water temperature and density is approximately uniform, showing a temperature gradient of < 1 °C/m SIST EN 16698:2016
EN 16698:2015 (E) 7 3.5 euphotic zone zone of the water body where light intensity is sufficient for photosynthetic production Note 1 to entry: The euphotic zone is the layer with more than 1 % of incident light intensity of subsurface light (see [13]). The euphotic depth is approximately 2,5 times Secchi depth [1], with exception of humic lakes (0,8 to 2,1 times Secchi depth) [13]. 3.6 holomictic lake lake that shows complete mixing of the water body every year 3.7 integral sampler water sampler which provides a representative sample of a predefined continuous water column 3.8 mean depth depth value obtained from dividing a lake's volume by its surface area Note 1 to entry: To obtain the mean depth in metres (m), the lake's volume should be specified in cubic metres (m3) and the surface area in square meters (m2). 3.9 metalimnion thermocline water zone between epi- and hypolimnion with the greatest vertical density gradient caused by temperature gradients > 1 °C/m 3.10 complete mixing full mixing of the whole water body, indicated by uniform temperature along the vertical axis Note 1 to entry: Complete mixing of water is only possible when the density in the whole water column is equal which usually means the temperature is the same in every depth. 3.11 monomictic lake lake with uniform water temperature and holomictic conditions from autumn to spring (no ice cover in winter) during which the water body is completely mixed 3.12 polymictic lake lake with more than two circulation phases per year comprising the whole water body (depth gradient
¶ 1,5) Note 1 to entry: See also [17]. 3.13 shallow lake in this European Standard shallow lakes are defined as polymictic lakes (3.12) – based on the depth gradient - independent of their mean or maximum depth SIST EN 16698:2016
EN 16698:2015 (E) 8 3.14 secchi depth visibility through the water column (transparency measured using a white plate lowered into the water) 3.15 stratification state of a water body during which a vertical density gradient in the water column (caused by a gradient of temperature or dissolved matter concentration) prevents its complete mixing 4 Principles of phytoplankton sampling 4.1 General Phytoplankton samples are collected and preserved for later microscopic investigation. Phytoplankton samples are most often collected in lakes, reservoirs or ponds and in larger rivers where residence time and light conditions enable phytoplankton growth. The following sections provide detailed recommendations on when and where samples should be taken to enable a more or less detailed study of phytoplankton in any location. Species composition, number of individuals and biomass of phytoplankton vary throughout the seasons as well as spatially across water bodies and within the water column. When setting up a sampling program it is important to be aware of this variability and adjust the sampling programmes accordingly. There is a range of recognized equipment and procedures used for the collection of phytoplankton samples and these can be selected according to the precise requirements of any particular study or sampling programme. Most sampling procedures are based on the collection of water samples which contain phytoplankton in its natural composition and abundances. Preservation and subsequent sedimentation of the phytoplankton enable the investigation according to Utermöhl (EN 15204). This allows taxa identification and quantitative assessment of abundance and biomass. Samples taken using a plankton net concentrate the phytoplankton, increasing the likelihood of picking up rare taxa, but cannot be used for quantitative assessment of abundance. Subsidiary information could include physical data such as water depth, temperature and oxygen profiles, assessment of euphotic zone and chemical determinants such as chlorophyll and nutrient concentrations. The frequency, location and type of samples collected to analyse phytoplankton in lakes and rivers should be determined by the requirements of the monitoring programme or study for which they are required. Generally, it is distinguished between sampling in lakes and sampling in rivers. In rivers which are assumed to be vertically completely mixed, it is sufficient to take a sample from the main flow. In lakes the mixed water layer or the zone in which phytoplankton is produced (euphotic zone) should be sampled. Generally, a vertical mixed sample should be taken. This is possible either with an integral sampler or by mixing of sub-samples from all depths of the mixed or euphotic water column. In clear lakes (euphotic depth > epilimnetic depth) a mixed sample should be taken from the euphotic zone. In turbid and humic lakes (euphotic depth < epilimnetic depth) it is adequate to take the sample from the mixed zone (epilimnion in stable stratified lakes, whole water column in polymictic lakes). The spatial extent of these zones is determined by recording Secchi depth and depth profiles of water temperature. For ambiguous temperature-depth profiles also other parameters such as dissolved oxygen and pH should be measured. It is recommended to determine the vertical distribution of the phytoplankton by measuring chlorophyll-a using a fluorescence probe in order to define the sampling zone. SIST EN 16698:2016
EN 16698:2015 (E) 9 Samples for phytoplankton and other parameters (e.g. chlorophyll-a and nutrients) should be taken at the same time. The number of samples, sampling depth range and location of sample sites should be determined by the purpose of the study or sampling programme. 4.2 Selection of sampling sites 4.2.1 General The location(s) where samples are taken should take into account the spatial variability of phytoplankton. 4.2.2 Sampling sites in rivers and streams Sampling should ideally take place in the main flow of the river. Samples may be taken from a bridge or boat if it is not safe to enter the river or obtain access to the appropriate location from the river bank. If samples are collected from engineered stretches, for example those with reinforced banking, they should be representative of the river section as a whole, including the more natural areas. For this reason it is recommended that the width of the river at the sampling point is not larger than twice the mean width in more natural sections, and that the average cross-sectional depth does not exceed the depth of the natural section by more than one third. If lateral heterogeneity is known or expected, several samples along a cross section should be taken and either be mixed or analysed separately. Samples should ideally be collected from a completely mixed water column. This can be confirmed by vertical measurements of temperature and, whenever possible, chlorophyll fluorescence. River sections which are not completely mixed, such as harbors and impoundments, should be sampled using methods suitable for lakes (see 4.2.3 and 6.3). 4.2.3 Sampling sites in lakes The spatial distribution of phytoplankton in lakes can be very variable, with vertical variation typically greater than horizontal variation. Wherever possible, phytoplankton samples should be taken from the open water as this enables consideration to be given to the variation in abundance and species composition within the water column; integrated samples can be taken through a known water depth or discrete samples can be taken at intervals through the water column. Only open water sampling locations allow to gather temperature and oxygen profile data to determine the degree of stratification and position of the thermocline. Additionally, to indicate variability in nutrient or chlorophyll concentrations in the whole water column, samples for chemical analysis can be collected. The location of open water sampling sites are depending on the bathymetry of the lake. Generally, more sampling stations are recommended for elongated fjord lakes and reservoirs (riverine/transitional/lacustrine zone [4]), for lakes with several basins separated by shallow sills, and for lakes with isolated large bays [2], [13], [14]. For lakes with a homogenous morphology, at least one sample should normally be taken at the bathymetric deepest point (zmax) as this allows the most complete profile of probe data to be gathered; the centre of the lake may also be used, although this may not be the deepest point. The position of sampling points should be recorded (e.g. by GPS) and should preferably be used again for successive surveys. NOTE It is well known that in lakes patchiness (small-scale spatial variability of the phytoplankton abundance and/or species composition) can occur. For some monitoring purposes it is advisable to integrate this variability by pooling samples taken from different points. Pooling reduces error in the characterization of the lake phytoplankton community without increasing the number of samples that need to be processed; however information about the horizontal patchiness is necessarily lost [4].
SIST EN 16698:2016
EN 16698:2015 (E) 10 In lakes that are segmented by a distinctive ground topography, samples generally should be taken in each basin and treated separately, i.e. they should not be pooled. If the objective is to investigate the patchiness, all samples should be taken using the same sampling method. Open water sampling usually requires the use of a boat. Provided that the potential limitations of shore sampling are considered, samples can also be collected from the lake shore, ideally at or near the outflow, or from a jetty or promontory. These can be collected using a bottle fixed to a long pole or a weighted bottle with float, on the end of a long rope thrown out into the lake, away from the immediate influence of the shore. Samples should not be collected near the inflow, or in areas likely to be influenced by hotspots of pollution, such as effluent discharges. Shore/outflow samples collect only the subsurface phytoplankton, and results can differ from open water sampling. Furthermore this method of sampling does not allow collection of information on stratification, or spatial distribution of phytoplankton. Samples collected from the shore/outflow provide an indication of the lake phytoplankton community and biomass and can be used in the context of large-scale monitoring programmes where some data from many lakes covering a wide geographic area is required rather than more detailed information from relatively few lakes. 4.3 Sampling frequency and replicates The sampling frequency is very important. The required number of samples per year depends on the objectives of the investigation and the intended type of data evaluation. The period without ice cover (which may be all year depending upon local climatic conditions) is also important. At least monthly sampling during the growing season is recommended; the growing season is often March/April to October/November, but can be shorter or longer depending upon local weather patterns. For a reliable assessment of a water body’s ecological quality, an investigation period of at least three years is recommended. In particular cases the sampling frequency may be reduced in accordance with the details given in Table A.2. Phytoplankton species composition and biomass varies strongly through the seasons (associated with, for example, spring peak, clear water phase, stratification period, circulation period). Thus, the more samples are collected, the more representative the results will be for the actual status of the water body. Infrequent sampling increases the likelihood of misrepresenting the actual conditions (see A.3, [13]). Replicates (multiple samples taken at the same location and same time) are only required when it is an objective of the investigation to get information on the spatial variability of phytoplankton biomass (or chlorophyll concentration) results received from the individual samples taken from one sampling point. 5 Equipment and preservatives 5.1 Equipment for sampling: 5.1.1 Suitable water sampler.
See Annex B for examples of suitable samplers; B.1 for samplers for rivers and B.2 for samplers for lakes. 5.1.2 Electrochemical/photo-optical probe with depth sensor to measure water temperature and dissolved oxygen according to EN ISO 5814 or ISO 17289. 5.1.3 pH probe (optional), e.g. according to EN ISO 10523. 5.1.4 Secchi disk according to EN ISO 7027 (see A.2 for further information). 5.1.5 Chlorophyll fluorescent probe with depth sensor (optional). A probe for total chlorophyll measurement is sufficient. SIST EN 16698:2016
EN 16698:2015 (E) 11 5.1.6 Boat (optional), suitable for local conditions with appropriate safety equipment. 5.1.7 Detailed bathymetric maps of the area of interest for preparation of the sampling design and localization of the sampling points. 5.1.8 Echo sounder and GPS, to localize the deepest point and the sampling stations on a water body. 5.1.9 Winch, with counter or rope with metre mark. 5.1.10 Mixing container, for example a plastic cask or bucket with lid (to protect the samples from sun light), for homogenizing integrated samples.
The size of the container depends on the sampled volumes. 5.1.11 Plankton net, generally for qualitative sampling only; mesh width from 5 µm to 25 µm, depending on the taxonomic target. 5.1.12 Sample bottles, according to EN 15204. 5.1.13 Lightproof closed containers, suitable for the transport of the sample bottles (e.g. cooling boxes). 5.2 Fixatives and preservatives: Lugol’s reagent is suitable for Utermöhl phytoplankton analysis, depending on pH (see EN 15204). See 5.2.1 and 5.2.2. for further specifications. 5.2.1 Alkaline Lugol's iodine, with sodium acetate for preservation of plankton from neutral or alkaline waters. 5.2.2 Acidic Lugol's iodine, with acetic acid for preservation of plankton from acidic waters. 5.2.3 Ethanol, C2H5OH, volume fraction 90 % to 96 %, for preservation of diatom or picoplankton samples. 5.2.4 Formaldehyde (optional). Formaldehyde is appropriate for preservation of diatom or picoplankton samples. 6 Procedure 6.1 General requirements for phytoplankton sampling A representative phytoplankton sample should be taken with
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SLOVENSKI STANDARD
oSIST prEN 16698:2014
01-januar-2014
.DNRYRVWYRGH1DYRGLOR]DNYDQWLWDWLYQRLQNYDOLWDWLYQRY]RUþHQMHILWRSODQNWRQDY
FHOLQVNLKYRGDK
Water quality - Guidance on quantitative and qualitative sampling of phytoplankton from
inland waters
Wasserbeschaffenheit - Anleitung für die quantitative und qualitative Probenahme von
Phytoplankton aus Binnengewässern
Ta slovenski standard je istoveten z: prEN 16698
ICS:
13.060.10 Voda iz naravnih virov Water of natural resources
13.060.70 Preiskava bioloških lastnosti Examination of biological
vode properties of water
oSIST prEN 16698:2014 en,fr,de
2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.
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oSIST prEN 16698:2014
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oSIST prEN 16698:2014
EUROPEAN STANDARD
DRAFT
prEN 16698
NORME EUROPÉENNE
EUROPÄISCHE NORM
December 2013
ICS 13.060.70
English Version
Water quality - Guidance on quantitative and qualitative
sampling of phytoplankton from inland waters
Qualité de l'eau - Lignes directrices sur l'échantillonnage Wasserbeschaffenheit - Anleitung für die quantitative und
quantitatif et qualitatif du phytoplancton dans les eaux qualitative Probenahme von Phytoplankton aus
intérieures Binnengewässern
This draft European Standard is submitted to CEN members for enquiry. It has been drawn up by the Technical Committee CEN/TC 230.
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, Former Yugoslav Republic of Macedonia, France, Germany, Greece, Hungary, Iceland, Ireland, Italy, Latvia, Lithuania,
Luxembourg, Malta, Netherlands, Norway, Poland, Portugal, Romania, Slovakia, Slovenia, Spain, Sweden, Switzerland, Turkey 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.
EUROPEAN COMMITTEE FOR STANDARDIZATION
COMITÉ EUROPÉEN DE NORMALISATION
EUROPÄISCHES KOMITEE FÜR NORMUNG
CEN-CENELEC Management Centre: Avenue Marnix 17, B-1000 Brussels
© 2013 CEN All rights of exploitation in any form and by any means reserved Ref. No. prEN 16698:2013 E
worldwide for CEN national Members.
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oSIST prEN 16698:2014
prEN 16698:2013 (E)
Contents
Page
Foreword .3
Introduction .4
1 Scope .5
2 Normative references .5
3 Terms and definitions .5
4 Principles of phytoplankton sampling .6
4.1 General .6
4.2 Selection of sampling sites .7
4.2.1 Sampling sites in rivers and streams .7
4.2.2 Sampling sites in lakes .7
4.2.3 Horizontal patchiness in lakes .7
4.3 Sampling frequency and replicates .8
5 Equipment and preservatives .8
5.1 Equipment for sampling .8
5.2 Fixatives and preservatives .9
6 Procedure .9
6.1 General requirements for phytoplankton sampling .9
6.2 Sampling in rivers . 10
6.3 Sampling in lakes. 10
6.3.1 Phases of circulation . 10
6.3.2 Phase of summer stagnation . 10
6.4 Preparation of mixed samples . 12
6.4.1 Preparation of mixed samples using an integral water sampler . 12
6.4.2 Preparation of mixed samples using other water samplers . 12
6.5 Bottling and fixation of samples . 12
6.6 Storage of the samples . 12
6.7 Additional samples for analysis of diatoms . 13
7 Measurements of accompanying parameters . 13
7.1 General . 13
7.2 Secchi depth . 13
7.3 Water temperature . 13
7.4 Dissolved oxygen . 14
7.5 Chlorophyll-a . 14
8 Quality Assurance . 14
Annex A (informative) Description of methodology and gear for sampling phytoplankton . 15
A.1 Water colour, Secchi-depth and euphotic depth . 15
A.2 Sampling position . 16
A.3 Sampling frequency – examples . 16
A.4 Storage and transport of water samples . 17
A.5 Example for a hose-integrated sample . 17
A.6 Tube integrated sample . 19
A.7 Hydrostatic integrated water sample . 20
A.8 Mechanic integrated water sample . 21
A.9 Electronic integrated water sample . 22
Bibliography . 23
2
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oSIST prEN 16698:2014
prEN 16698:2013 (E)
Foreword
This document (prEN 16698:2013) has been prepared by Technical Committee CEN/TC 230 “Water analysis”,
the secretariat of which is held by DIN.
This document is currently submitted to the CEN Enquiry.
This document has been prepared under a mandate given to CEN by the European Commission and the
European Free Trade Association, and supports essential requirements of the Water Framework Directive
(2000/60/EC) (WFD), and the Directive on Environmental Quality Standards (Directive 2008/105/EC).
WARNING —Working in or around water is inherently dangerous, Persons using this European
standard should be familiar with usual field and laboratory practice. This standard 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 health and safety practices and to ensure compliance with any national
regulatory guidelines.
IMPORTANT — It is absolutely essential that tests conducted according to this European Standard be
carried out by suitably trained staff.
3
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oSIST prEN 16698:2014
prEN 16698:2013 (E)
Introduction
Phytoplankton samples provide information on the taxonomic composition as well as the spatial density of the
individual taxa and their relative abundances. They allow the calculation of the productivity and biomass of the
phytoplankton assemblage as a whole as well as for the individual taxa.
For the purpose of limnological investigations and monitoring of surface waters representative phytoplankton
samples are necessary. Therefore suitable sampling methods are needed depending on the aims of the
investigation and the given natural conditions.
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1 Scope
This European standard gives guidance for sampling of phytoplankton from inland waters for quantitative and
qualitative limnological investigations and monitoring of water quality, e. g. ecological status.
This European standard is intended to ensure uniform conditions for phytoplankton sampling in inland waters
and describes methods of sampling techniques for phytoplankton in inland waters (rivers, channels, lakes,
ponds, reservoirs and other artificial water bodies).
2 Normative references
The following documents, in whole or in part, are normatively referenced in this document and are
indispensable for its application. For dated references, only the edition cited applies. For undated references,
the latest edition of the referenced document (including any amendments) applies.
EN 15204, Water quality – Guidance standard on the enumeration of phytoplankton using inverted microscopy
(Utermöhl technique).
EN 14996, Guidance on assuring the quality of biological and ecological assessments in the aquatic
environment.
EN ISO 7027, Water quality – Determination of turbidity (ISO 7027).
EN ISO 5814, Water quality – Determination of dissolved oxygen; electrochemical probe method (ISO 5814).
3 Terms and definitions
For the purposes of this document, the following terms and definitions apply.
3.1
holomictic lake
lake that shows complete mixing of the water body every year
3.2
monomictic lake
lake with uniform water temperature and holomictic conditions during the autumn to spring phase (no ice
cover in winter) during which the water body is completely circulated
3.3
dimictic lake
lake with uniform water temperature and holomictic conditions twice a year: during autumn (before ice cover)
and during spring (after ice cover)
3.4
polymictic lake
shallow lake which has more than two circulation phases per year comprising the whole water body
3.5
shallow lake
polymictic lake with a maximum depth usually ≤ 10 m
3.6
mixing of water
complete mixing of the whole water body
Note 1 to entry: Complete mixing of water is only possible when the density in the whole water column is equal, that
means usually the temperature is the same in every depth.
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Note 2 to entry: In the context of this European Standard shallow lakes are defined as polymictic lakes.
3.7
metalimnion
thermocline
water zone with the greatest vertical density gradient caused by temperature gradients in a lake > 1 °C/m
3.8
stratification
state of a water body during which a vertical density gradient in the water column prevents its complete mixing
3.9
epilimnion
z
epi
zone of the water body between surface and thermocline in which the water temperature and density is
approximately uniform, showing a temperature gradient of < 1 °C/m
3.10
euphotic zone
z
eu
zone of the water body below the surface, in which the photosynthetic production takes place
Note 1 to entry: Layer with more than 1% of incident light intensity (or subsurface light see [11]). The euphotic depth is
approximately 2,5fold Secchi depth [1], with exception of humic lakes or lakes showing algal scums (0,8 to 3,0 fold Secchi
depth) [11]
3.11
mean depth
3 2
depth value (in metres) obtained from dividing a lake's volume (m ) by its surface area (m )
3.12
integral sampler
water sampler which provides a representative sample of a defined continuous water column
3.13
Secchi depth
maximum depth at which a white disk according to Secchi that is lowered into the water is still just visible; it is
the average of the depth at which the disk being lowered disappears and the depth at which it reappears when
it is subsequently raised
3.14
stream centre line
connecting line between the positions of the main flow in transsects of a running water
4 Principles of phytoplankton sampling
4.1 General
A representative phytoplankton sample is taken and preserved for later microscopic investigation (e.g.
according to EN 15204). Generally, it is distinguished between sampling in lakes and sampling in rivers. In
rivers that are completely mixed, it is sufficient to take a sample from the water column in the stream centre
line.
In temporarily stratified waters the mixed water layer (epilimnion) or the zone in which phytoplankton is
produced (euphotic zone) shall be sampled. In these waters, a vertical mixed sample shall be taken. This is
possible either with an integral sampler or by mixing of sub-samples from all depths of the mixed or euphotic
water column.
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In clear lakes a mixed sample shall be taken from the euphotic zone. In turbid and humic lakes it is adequate
to take the sample from the mixed zone (epilimnion in stratified lakes, whole water column in polymictic lakes).
The depth of these zones shall be determined by recording Secchi depth, and depth profiles of water
temperature and dissolved oxygen. The distribution of the phytoplankton should be determined by measuring
chlorophyll-a using a fluorescence sensor.
For special objectives samples may be collected individually from fixed depths (e.g. drinking water intake).
4.2 Selection of sampling sites
4.2.1 Sampling sites in rivers and streams
The width of the river at the sampling point may not be larger than twice the mean width in adjacent
unobstructed sections (i. e., sections without bank fixation). The average cross-sectional depth may not
exceed the depth of the unobstructed upstream section by more than one third. Sampling shall take place in
the stream centre line. Samples from large and deep rivers may be taken from a bridge or a boat.
Complete mixing of the water column shall be confirmed by vertical measurements of temperature and
chlorophyll fluorescence. Artificially widened and consolidated stream sections (harbours, locks, sites directly
before or behind barrages etc.) are no representative sampling points for phytoplankton in rivers since the flow
velocity at these places is highly modified, which may result in stratification or sedimentation of the
phytoplankton. River sections that are not completely mixed, like harbours and impoundments, shall be
sampled according to the instructions for lakes (see 4.2.2 and 6.3).
4.2.2 Sampling sites in lakes
) because the
In lakes the sampling of phytoplankton shall take place at the bathymetric deepest point (z
max
depth profiles of temperature and oxygen shall as well be taken at the point of maximum depth. Sampling at a
single station at the centre of the lake may be sufficient [2], [13], but may differ from the deepest point. In
lakes that are segmented by a distinctive ground relief forming several basins, sampling generally shall be
performed in each basin. The samples from each basin shall be treated separately, i. e. they shall not be
pooled with samples from another basin. Lakes with longitudinal gradients (reservoirs, channel lakes) shall be
sampled at different points reflecting the biological differences to be expected between the sampling points
(e. g., in reservoirs: riverine zone, transitional zone and lacustrine zone [4]).
Only in exceptional cases (e. g. when no boat can be hoisted because of very steep shores) sampling can be
conducted from the shore line or outflow respectively. These samples are not vertically integrated and are
restricted to qualitative or semiquantitative results with regard to the phytoplankton community of the surface
water body. No additional information on stratification, light climate and metalimnic or hypolimnic nutrients and
deep chlorophyll maxima (DCM) are possible.
4.2.3 Horizontal patchiness in lakes
It is well known that in lakes patchiness (small-scale spatial variability of the phytoplankton abundance and/or
species composition) can occur. For some monitoring purposes it is advisable to integrate samples over some
area, thus removing the variability within the area integrated. The loss of information caused by this pooling of
samples should be weigthed against the gain in accuracy achieved by the reduction of an error component [3].
If the objective is to investigate the patchiness, all samples shall be taken using the same sampling method.
In valley lakes, especially in reservoirs, the phytoplankton biomass is lower at a point near the outflow or dam
than in the longitudinal centre of the lake. In this case sampling at the end of the lake would lead to an
underestimation of the phytoplankton biomass and trophic level of the lake [4]. Therefore, in valley lakes at
2
least one additional point at the longitudinal centre shall be sampled. In large lakes (> 100 ha or 1 km ) more
than one sampling point is recommended to take into account differences in the phytoplankton composition
that may occur along the longitudinal axis of the water body. Strongly isolated bays shall also be sampled
separately [2].
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4.3 Sampling frequency and replicates
The sampling frequency is very important. The required number of samples per year depends on the
objectives of the investigation, the intended type of data evaluation and the duration of the period without ice
cover. During the growing season (March/April to October) at least monthly sampling is recommended. For a
reliable assessment of a water body's ecological quality, an investigation period of at least three years is
advisable. If necessary, the sampling frequency may be reduced in accordance with the details given in
Table A.2.
Depending on the objectives of the investigation and taking into account the seasonal processes in the water
body (spring peak, clear water phase, stratification period, full mixing etc.) the maxima of phytoplankton
development shall be covered. The probability of trophic mis-classification depends on the frequency of
sampling (see Annex A and [12]).
Replicates are required when it is an objective of the investigation to get information on the variability of
phytoplankton biomass (or chlorophyll content) results received from the individual samples taken from one
sampling point. In regular monitoring programs replicates are not necessary, but it is recommended to take
several samples from one sampling point and pool them.
5 Equipment and preservatives
5.1 Equipment for sampling
To prevent spreading of flora and fauna between water bodies, the sampling equipment shall be cleaned
between samplings in different water bodies. After each day of sampling, plankton nets shall be washed in
warm freshwater with detergent or in an ultrasonic water-bath in order to reduce clogging and ensure optimum
filtration capacity. Equipment such as water samplers, hoses, plankton nets and mixing containers shall be
rinsed in freshwater and dried before they are stored for future use. Hoses shall be hung up to dry. Rapid
drying is essential to prevent unwanted growth within hoses.
5.1.1 Suitable water sampler. See Annex A for examples of suitable samplers.
5.1.2 Electrochemical probe with depth sensor to measure water temperature and dissolved oxygen
according to EN ISO 5814.
5.1.3 Secchi disk
5.1.4 Chlorophyll fluorescent probe with depth sensor (optional).
5.1.5 Boat, suitable for local conditions with appropriate safety equipment.
Equipment for communication from the boat to designated shore-based staff, with access to rescue services is
recommended. An experienced boat driver familiar with local conditions should be deployed.
5.1.6 Detailed bathymetric maps of the surveyed area
5.1.7 Echo sounder, to localize the deepest point of a lake
5.1.8 Winch, with counter or rope with metre marks for deep lakes.
5.1.9 Mixing container, for example a plastic can or bucket with lid, for homogenising integrated samples.
5.1.10 Plankton net, mesh width 10 µm to 25 µm.
5.1.11 Sample bottles, with a volume of 100 ml, 250 ml or 500 ml.
Use clear transparent narrow-neck glass bottles with a volume of 100 ml, 250 ml or 500 ml (see 6.6).
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5.1.12 Dark closed containers, suitable for the transport of the sample bottles (e.g. cooling boxes).
5.2 Fixatives and preservatives
Two different reagents shall be used for phytoplankton sample preservation (for details see EN 15204).
5.2.1 Alkaline Lugol's iodine, with sodium acetate for preservation of plankton from neutral or alkaline
waters (pH ≥ 7).
5.2.1 Acidic Lugol's iodine, with acetic acid for preservation of plankton of acidic waters (pH < 7).
5.2.3 Ethanol, C H OH, volume fraction 90% to 96 %, for preservation of diatom samples.
2 5
6 Procedure
6.1 General requirements for phytoplankton sampling
A representative phytoplankton sample shall be taken using a suitable water sampler (see Annex A for
examples). During sampling, the sampling gear shall not be allowed to touch the bottom, because this might
contaminate the water samples.
The phytoplankton sample shall be filled immediately or after mixing in sampling bottles and preserved for
later microscopical investigation according to EN 15204.
Sample bottles shall be marked before sampling to avoid mixing up the samples. Waterproof marker pen or
pencil shall be used (ballpoint pens or regular marker pens are not appropriate). Writing directly on the glass
is not recommended. Instead, the use of water-resistant adhesive tape that can be labelled is advisable.
As a minimum, the following information shall be given for each sample (included on the label or in the
sampling protocol):
name of locality;
sampling person,
order number or order designation;
sampling date and time;
sampling position if appropriate;
sampling depth (mixed, epilimnetic or euphotic depth);
type of sample (e. g. phytoplankton or pelagic diatoms; in case of net samples mesh width and volume of
water filtered through the net).
The sample shall be labelled to ensure that it can be clearly identified using the information given in the
sampling protocol.
The sample bottles shall be transported to the laboratory in dark, and if necessary cooled, non-transparent
containers.
The sub-samples for phytoplankton and other parameters (e. g. chlorophyll-a and nutrients) shall be taken
from the same mixed sample. This means that the sample volume should be sufficiently large. Nutrient
analyses shall only be done from mixed euphotic samples, if the euphotic zone does not reach into the
hypolimnion. If the euphotic zone stretches into the hypolimnion a separate mixed sample from the epilimnion
shall be taken for nutrient analysis.
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When using the water sampler or hose, it shall be checked that the closing mechanism is working properly to
ensure that unhindered water flow-through is achieved and the valve is closed. The water sampler is lowered
vertically to the required depth at an even, moderate speed (not exceeding 1 m/s). When integrating samplers
or hose samplers are used, it is important that the device is lowered slowly to the required depth.
6.2 Sampling in rivers
The samples shall be taken with a suitable sampler from the mixed water column in the stream centre line of
the river. Sampling of the water surface shall be avoided.
If the Secchi depth is less than 1 m a second sample should be taken directly from the water surface and
mixed with the sample from the water column with a ratio of 1:1. Since phytoplankton organisms (especially
diatoms) of rivers usually settle very rapidly, samples shall be transferred quickly into the sampling bottles and
only after prior homogenisation [5].
6.3 Sampling in lakes
6.3.1 Phases of circulation
During phases of circulation in shallow lakes the whole water column shall be sampled to a depth of 0,5 m to
1 m above the sediment surface.
During phases of circulation sampling in deep lakes shall take place to a maximum depth of 20 m or 1 m
above the sediment surface, respectively.
The position of the sampling stations shall be recorded by GPS to be found and used for each survey.
6.3.2 Phase of summer stagnation
During summer stagnation in polymictic (shallow) lakes (usually with z ≤ 10 m) the sampling zone
max
comprises the water column down to a depth of 6 m, or 0,5 m to 1 m above sediment surface, respectively.
For sampling in stratified (deep) lakes (usually with z > 10 m) during summer stagnation a distinction is
max
drawn between two different states (see Figure 1):
1) Turbid and humic lakes with euphotic depth < epilimnion depth (z < z ): Take a mixed sample
eu epi
from the epilimnic zone.
2) Clear lakes with euphotic depth > epilimnion depth (z > z ): Take a mixed sample from the
eu epi
euphotic zone.
If a chlorophyll probe is available, the vertical extension of the euphotic zone can be determined exactly, so
that a deep chlorophyll maximum (DCM) will not be missed. See also 6.1 concerning an euphotic zone
reaching into the hypolimnion.
If no chlorophyll probe is available, the euphotic depth can be determined using Equation (1):
z = t⋅ z
eu secc
where
z is the euphotic depth in meters (m);
eu
z is the secchi depth in meters (m);
secc
t is an empiric constant depending on the turbidity of the lake ranging from 0,8 to 3,0 [11]:
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under usual conditions in clear and turbid lakes t = 2,5;
when algal scum is observed at the lake surface t = 3,0;
in humic lakes: t = 0,8 to 2,1, depending on the degree of colouration, in accordance with the
correlation given in [11] (see also Annex A, Figure A1).
Figure 1 provides a decision scheme for sampling in deep stratified lakes.
The determination of sampling depth can also be based on informa
...
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