The quantitative separation of meiofauna

Helgoland Marine Research, May 1973

1. This paper presents the results of a meeting on the evaluation of quantitative procedures for the separation of meiofauna, held at the Marine Station of the Biologische Anstalt Helgoland, in May 1972. Close co-operation on the part of the participants (p. 194) has allowed assessment of advantages and disadvantages of the separation methods applied. The time needed for preserved methods can be reduced by changing to scanning rather than picking out for sorting and counting. 2. Sorting without concentrations: This method is suitable for very fine-grained sediments, oozes; it is generally applied for preserved samples; adequate for hard fauna; insufficient for soft fauna; very time consuming. 3. Decantation: Suitable for sandy sediments; generally applied for preserved samples; adequate for hard fauna; insufficient for soft fauna; time consuming. 4. Elutriation: Suitable for sandy sediments; for unpreserved samples only with anaesthetization, more effective with preserved samples; limited for total live fauna. With preserved samples, adequate for hard fauna; insufficient for soft fauna; quick method. 5. Warm-water elutriation: Suitable for sandy sediments; designed for live hard fauna (especially nematodes, ostracods); quick method. 6. Sea-water ice treatment: Suitable for sandy sediments with microporal structure; only for live extraction; limited for hard fauna; well suited for soft fauna (including ciliates and flagellates); time consuming.

A PDF file should load here. If you do not see its contents the file may be temporarily unavailable at the journal website or you do not have a PDF plug-in installed and enabled in your browser.

Alternatively, you can download the file locally and open with any standalone PDF reader:

https://link.springer.com/content/pdf/10.1007%2FBF01609968.pdf

The quantitative separation of meiofauna

Helgol~inder wiss. Meeresunters. The quantitative separation of meiofauna G. UHLIG 0 H. THIEL ~x: J. S. GRAY 0 0 1 Biologische Anstalt HelgoIand (Meeresstation); Helgoland, Federal Republic of Germany and 2 Institut fiir Hydrobiologie und Fischereiwissenschafl, Universitlit Hamburg; Hamburg, Federal Republic of Germany and 3 University of Leeds, Wellcome Marine Laboratory; Robin Hood's Bay , Yorkshire, England A comparison of methods KURZFASSUNG: Die quantitative Isolierung der Meiofauna. Ein Methodenvergleich. Im Mai 1972 land an der Meeresstation der Biologischen Anstalt Helgoland ein Arbeitstreffen mit dem Ziel statt, die vers&iedenen Methoden zur quantitativen Isolierung der Meiofauna aus dem Sediment in ihrer Effektivit~it miteinander zu vergleichen. Die Verfahren zur Anreicherung der Meiofauna in der Probe, des Sortierens und Ausz~ihlens werden beschriebenund die dutch die Teilnehmer am Arbeitstreffen gemeinsarnerzielten Ergebnisse diskutiert. Die Wettung der einzelnen Metboden zur Erfassung der Gesamtfauna, der ,,harten" und der ,weichen" Meiofauna, yon konservierten und nichtkonservierten Organismen sowie die Anwendung der Methoden auf verschiedene Sedimenttypen werden durch statistische Analysen abgesichert. Eine Zusammenfassung der Arbeitsverfahren sowie deren Charakterisierung und Leistungsf~higkeit beschlief~t die Darstellung. - meiofauna have become increasingly important. Consequently, separation procedures have become more laborious, with the need for extra precision. In 1969, the Mediterranean Marine Sorting Center in Salammbo, Tunisia, invited several meiobenthologists to discuss the central problems in meiofauna investigations. The results of this meeting were published in the "Manual for the study of meiofauna" (HuLINaS & GRAY 1971). This manual represents the first attempt to summarize recent experience in this fidd. Many recommendations are given for the measurement of environmental parameters, sampling and separation of the meiofauna from the sediment, as well as preparation methods for the main representative groups of mdofauna. In the foreword of the "Proceedings of the First International Conference on Meiofauna" HULINGS (1971) pronounces: "Statistical evaluation of various techniques of sampling . . . are imperative." Yet, concerning separation methods, no comparison could be made during this Tunisia meeting. For the separation of meiofauna several methods have been published which essentially are modifications of a few basic procedures. Generally, the choice of a suitable method depends on the aim of the scientific problem studied. This includes the decision, whether the meiofauna (certain species, important representatives, or the total meiofauna) has to be separated alive, or whether it can be handled preserved. The efficiency of the commonly used methods has been evaluated with varying results by different authors. Howe~er, no statistical analyses are available which can offer reliable comparisons. In order to solve this problem a number of specialists were invited to a meeting. All the participants were asked to present their quantitative separation procedures for comparison, using replicate samples. This meeting was held on Helgoland at the Marine Station of the Biologische Anstalt Helgoland from May 4~hto May 12% 1972, in co-operatlon with the working group on meiofauna of the Baltic Marine Biologists (Convenor: H. TmEL). The participants at the Helgoland meeting, who are listed on page 194, have authorised us to present here the data and results obtained in close co-operation and discussion at that meeting. D E S C R I P T I O N OF T H E S E P A R A T I O N M E T H O D S USED Separation of meiofauna from the sediment involves two different procedures: (1) concentration and (2) sorting and counting. We define these terms as follows: C o n c e n t r a t i o n : The enrichment of meiofauna in respect of sediment particles in the sample. S o r t i n g a n d c o u n t i n g : The classification of fauna into taxonomical groups and the counting of specimens. The essential methodological differences are restricted to the concentration procedure. These will be described therefore in separate sections, while sorting and counting as done by the participants is combined in one paragraph. Concentration methods are based largely on the hydrodynamic behaviour of the sediment particles. This depends on different characteristics of the particles: Size, specific weight, form and surface structure. Sinking rate and transport by water flow are determined by such characteristics. Since there is no suitable short expression for particles with different hydrodynamic behaviour we have used the terms, "lighter" particles and "heavier" particles in the subsequent text. These terms may be defined as" "lighter" particles, those with a low sinking rate or easily transported by a definite water flow and "'heavier" particles those with a high sinking rate or less easily transported by the same water flOW. C o n c e n t r a t i o n o f m e i o f a u n a Decantation and sieving The method of decantation and sieving was practised by R. ELMGaeN (EL) and H. T~tleL (TH). The most simple treatment for the concentration of meiofauna specimens in. a sediment sample is decantation. The sample is stirred up in a dish, glass beaker or measuring glass. ARer allowing a short time for settlement of "heavier" sand grains or other constituents of the sample, the supernatant water is poured out through a sieve or a set of sieves. Larger particles, including the organisms wanted, are retained, while mud particles pass through. This procedure may be repeated several times until most or all of the fauna is washed out of the coarse fraction of the sample. Repeated decantation and subsequent sorting and counting allows an estimation of the efficiency of the procedure, since the residue can be checked periodically. In total fauna separation, decantation is applied generally for preserved material. Concentration by sieving is restricted to the separation of a coarser fraction containing the organisms and the finer fraction, which is discarded. The mesh size of the sieve used depends on the size of animals to be considered in the investigation. The sieving of the coarser fraction into size groups makes sorting and counting easier and more efficient (p. 180) but no concentration effect is achieved. Normally, both these treatments are combined into one procedure. Etutriation methods The elutriation method was introduced by BOlSSEAU(1957). In the years following, this method has been used for sandy sediments by several authors and the equipment has &anged slightly. The general idea is the partition of particles by their different characteristics in response to a continous water stream. The "lighter" particles, including the meiofauna organisms, are washed out of the sample and are collected on a sieve. (a) M o d i f i c a t i o n s o f t h e BOISSEAU m e t h o d . Elutriation methods of the t~OlSSEAUtype were practised by I. Di~zYclMs~i (DR), J. S. GI~AY(GR), and A. D. MCINTYI~ (McI) in 3 modifications. Two of them used preserved (McI) or unpreserved (DR) material in an open water stream while in the third apparatus (GR) a closed water circuit was used for narcotized living organisms. The essential part of the apparatus (Fig. 1) is a separation funnel which contains the sample. The open circuit system is connected to the main cold-water supply or to some filtered sea-water reservoir. The water stream passes through the separation funnel, containing the sample, and the overflow tube. The meiofauna washed out of the sample is concentrated on the sieve. In the closed-circuit system a water pump circulates water from a reservoir. The flow rates are adjusted by the two valves. Fig. 1: Elutriation apparatus, combined for open and closed system. BP; by-pass, CC: closed circuit, OC: open circuit, OT: overflow tube, RV: reflex valve, S: sieve, SF: separation funnel, V: valve, WP: water pump, WR: water reservoir great quantity. The animals retained on the 45 micron sieve were then washed into a 9 cm diameter bacteriological Petri dish (or a 14 cm diameter dish when faunal density was high), dispersed with water from a wash bottle, allowed to settle and counted. Fig. 2: Apparatus for warm-water elutriation. BP: by-pass, CB: collecting bottle, CR: collecting receptacle, LC: Liebig condensor, OT: overflow tube, SR: sample receptacle, ST: settling tube, VT: ventilation tube, WR: water reservoir, 1-5: valves. (After OHD~, unpublished) Because of the much higher proportion of silt in the muddy sand sample, elutriation rates were reduced so that small fractions of the total sample could be examined separately. Thus, the sample was examined in 4 or 5 fractions in which the numbers of animals decreased very substantially with each successive "washing". The amount of material "washed off" in any one fraction was controlled, so that the sediment deposit in the petri dish would not obscure fauna for the scanning operation. Total dutriation time was approximately the same as for other sediments. (b) T h e w a r m - w a t e r e l u t r i a t i o n . The warm-waterelutriationwas designed and practised by J. OHDE(OH) (unpublished). The apparatus (Fig. 2) consists of the receptacle for the sample, connected by a spiral settling tube with a Liebig condensor. At its lower end a collecting receptacle connects to a water in- and outlet, as well as to a by-pass which runs ba& into the settling tube. The overflow tube leads into the collecting bottle or a sieve. In the concentration procedure, the whole system is first filled with filtered water from the reservoir. The sample is rinsed into the sample receptacle and, by opening of the valves 1 and 2, is allowed to flow down the settling tube and to sink through the inner pipe of the condensor into the coIlection receptacle. The jacket tube of the condensor is charged with water of 55 ° C constant temperature. By this means the fluid and the sample in the inner tube of the condensor are warmed. Under these conditions nematodes are stretched and ostracods open their shells, resulting in a higher water resistance, i.e. a better separation efficiency. After closing valve 2 and opening valves 3, 4, and 5, water flow from the reservoir carries the particles up the by-pass into the spiral settling tube. The water stream transports the "lighter" particles and fauna via the overflow tube into the collecting bottle. "Heavier" particles run slowly down the wall of the spiral settling tube and again sink down the condensor. The flow rate is regulated by changing the vertical position of the sample receptacle. The procedure can be repeated at will. Air bubbles originating from gases dissolved in the water may be let out of the system through the ventilation tube. Finally, the sediment and the water are drained out of the apparatus and the tubes are washed thoroughly. For maximum efficiency, the water may be run through a sieve to collect specimens stuck to the tube walls. Sea-water ice treatment This method was practised by E. HARTWm & S. HOXHOLD (HH) and by G. UHLm (UH). The sea-water ice treatment, described by UHLm (1964, 1968), is based on changing the intensity of environmental factors in the sediment samples treated. The infauna of a sample is influenced mainly by two ecological parameters: (1) The change from high to low salinity of meIting sea water and (2) a low perfusion and flow of water through the sediment. Temperature, also, may have a slight effect. Under these conditions the interstitial fauna migrates actively out of the sediment and is collected alive in a culture dish, almost free of sediment. One end of a plastic tube (Fig. 3) is covered with a tightly fitting nylon gauze, the mesh size of which depends on the main grain size and then the tube is filled with crushed sea-water ice, prepared in a deep freezer. Insulation is placed between sediment and ice. A Petri- or culture dish, filled with filtered sea water, is placed under the tube so that the nylon gauze is just in contact with the sea-water surface. Both the culture dish and the tube are in a fixed position. The Petri dish is placed in a second larger one, into which the water can overflow. Plastic tubes of different size and diameter were used by (HH) and (UH). In that way, (HH) exposed a larger sediment surface PT / / / / / / / / / Fig. 3: Sea-water ice treatment. I: Insulation material, NG: nylon gauze, P1, P2: Petri- or culture dishes, PT: Plastic tube, S: Sediment, SI: Sea-water ice, SW: Sea water, TH: Tube holder S o r t i n g c o u n t i n g o f m e i o f a u n a Sorting and counting under a stereoscopic microscope is common to all the separation methods tested. O n l y small differences were encountered between the procedures of the participants but personal experience is important. This part is, therefore, described together in one paragraph. For preservation 4-5 °/0 formalin was used for at least 20 minutes. In samples to be stored f o r a long time the formalin should be neutralized with 100 g hexamethylene tetramine per 1000 cm3 of 40 0/0 formaldehyde. A ~ e r washing out the formalin, the sample was stained with rose bengal + phenol (THI~L 1966) for at least 15 minutes. S t a i n i n g s o 1 u t i o n : 100 ml of water + 1 ml of concentrated rose bengal solution + 5 ml of concentrated aqueous solution of phenol (for p H adjustment). S t a i n i n g p r o c e d u r e : (1) The sample is washed from the sampling bottle into a sieve of the smallest mesh size required in the respective research. (2). The sample is rinsed in a beaker of tap water until no preservation fluid remains. (3) The sample in the sieve is placed in the staining solution for about 15 minutes. (4) The sample in the sieve is again rinsed either in a beaker of tap water until the water is not coloured immediately or by elutriation. Staining was followed either by direct sorting and counting or by elutriation or decantation, described in the previous section under methods for concentration. Mechanical hand counters were used to record numerically dominant groups. In the case of (McI) animals were counted in situ in the dish and organisms were ex~tracted (by pipette) only when identification under high-power microscope was required. Counting was carried out with a Petri dish resting on a squared paper grid (1 cm square units) and using a Vickers Extenda-arm scanning microscope, 20 × magnification. The field of view is thus approximately 1 cm~. In this way, with the Petri dish fixed in position, the bottom of the dish and the surface of the water can be systematically scanned. In situ counting is less time consuming than the simultaneous extraction of the organisms with a pipette (which all other participants did and Petfi dishes were used for sorting under stereoscopic microscopes). With decantation, (EL) produced 2 fractions of t00 l~m and 37 l~m. These were transferred to a Bogorov-tray (RusSELL& COLM~N 1931, ARLT 1973). The tray was carefully searched and all animals found pi&ed out. The contents of the tray were then thoroughly agitated with the pipette, and allowed to settle before a new search was made. This was repeated once more, so that three searches were made for each portion examined, and the animals found in each search noted (further discussion in ELMCR~N1973). The decantation technique omitted the sorting of coarse fractions. Instead, small fractions were used by (TH) but all the sediment was searched through, as indeed has to be done with very fine-grained sediments such as deep-sea oozes. Before sorting and counting, the samples were sieved into four fractions of > 42, > 65, > 100 and > 150/~m. Working with a narrow grain-size range was found to be more effective. Small dishes were used with the stereoscopic microscope and limited portions were sorted successively in order to produce a thin sediment layer in the dish. The samples were sorted twice. SAMPLING, SEDIMENT ANALYSES AND FAUNA It was the intention of the working group to analyse the efficiency of the methods on a wide spectrum of sediments. Within the available time we were able to study four different types of substrata (Fig. 4). The sediments from each of the three sublittoral stations were homogenized by manual mixing to omit natural variability between samples, while the samples from the beach were taken in situ. S e d i m e n t d a t a a n d h a n d l i n g (1) Medium sand with fine sand, well sorted - termed medium sand (Fig. 4a); Helgoland Reede - near Youth Hostel; Sublittoral, 6-8 m. The material was collected by divers scraping surface sediment into bu&ets. In a constant-temperature 0.63 o,68 o,~2 o,'25 6.s '~ o,63 o,6~ Q;2 o.2~ o]5 Fig. 4: Cumulative grain-size curve for the four types of sediments used in the experiments. A Medium sand w i t h fine sand, well sorted. (a) Q1 = 259, Q3 = 366, M d = 291, So = 1,19; (b) Q1 = 259, Q3 = 361, M d = 287, So = 1,18; (c) Q1 = 260, Q8 = 379, M d = 297, So = 1,12; (abbreviation: Medium sand, corresponding to Tables 3, 7, 10). B Coarse sand w i t h shell gravel, fairly well sorted. (a) Q1 = 493, Q3 = 702, M d = 563, So = 1,19; (b) Q1 = 507, Q~ = 707, Md = 563, So = 1,18; (c) Q1 = 543, Q8 = 889, M d = 707, So = 1,28; (abbreviation: coarse sand, corresponding Tables 4, 8, 11). C Fine sand, well sorted. (a) Q1 = 203, Q3 = 277, Md = 233, So = 1,16; (b) Q1 = 203, Q3 = 279, Md = 237, So = 1,17; (abbreviation: fine sand, corresponding to Table 6). D Medium sand w i t h fine sand, less sorted. (a) Q1 = 254, Q8 = 841, M d = 354, So = 1 , 8 2 ; ( b ) Q1 = 259, Q3 = 637, M d = 337, So = 1,57; (c) Q1 = 277, Q8 = 229, Md = 384, So = 2,88; (abbreviation: m u d d y sand, corresponding to Tables 5, 9, 12) room (15 ° C), the sediment was transferred into a t a n k of 50 × 90 cm under intensive manual mixing. For sampling, a grid was laid over the t a n k with 4 × 8 fields of 10 × 10 cm, leaving 5 cm margin around the tank. The 4 longitudinal rows, each with 8 fields, are called levels A, B, C, D (Fig. 5). Sediment was cored from each field centre. For statistical analyses the distribution of samples between participants was arranged according to a table of r a n d o m numbers. Each method received 4 samples, one from each level. The samples covered a surface area of 4 cm ~ and reached 4 cm into the sediment. OH HH UH EL TH GR EL 1013 Fig. 5: Pattern of random sample distribution of the coarse-sand sediment (Table 4). Longitudinal: levels A-D; vertical: rows 1-8. The figures within the fields represent the total number of animals found by the different methods as marked by symbols I n o r d e r to d e m o n s t r a t e the f r e q u e n c y of o c c u r r e n c e o f the different taxa, the n u m b e r of organisms c o u n t e d by the eight m e t h o d s a r e listed in T a b l e 1. T a b l e 1 also illustrates t h e high v a r i a t i o n b e t w e e n the m e t h o d s f o r certain taxa. T a b l e 2 gives the Number of organisms counted per taxon, and different methods used on coarse sand with shell gravel. Abbreviations see text Elutriation GR McI Decantation total sorting EL T H Total number of organisms counted per sediment type. Individual numbers for the fine-sand sample from Helgoland Dtine apply to a sediment surface area of 104 cm2 and 4 cm sediment depth Medium sand Total % Coarse sand Total °/0 Muddy sand Total °/0 t o t a l n u m b e r , t o g e t h e r w i t h t h e p e r c e n t a g e of t h e d o m i n a n t t a x a f r o m t h e f o u r sediments f o r all methods, e x c l u d i n g the ciliates. The group " o t h e r s " includes the f o l l o w ing t a x a : Halammohydra, rotifers, tardigrades, nemerteans, otigochaetes a n d bivalves. T h e y w e r e represented in o n l y small numbers. S T A T I S T I C A L A N A L Y S E S I n i t i a l analyses of the r a w d a t a showed t h a t the variance was p r o p o r t i o n a l to the mean, for the different methods. F u r t h e r analyses, therefore, were done on transformed data, using logl0(n), where n was the number of the r a w data. T w o - w a y analyses of variance were conducted on the total fauna d a t a (with the exception of the beach data), for each t y p e of sediment, in order to test for differences between methods, a n d between levels in the t a n k of sediment. The interaction between levels and methods was used as the error term since there were no replicates in this analysis. O n l y in the m u d d y sand sample (Table 6) were significant differences between levels in the t a n k found. Elimination of samples f r o m level A in this t a n k (which had significantly smaller numbers of animals than the other levels), rendered the differences between levels non-significant. Thus, the tanks of sediment could be considered homogenous (since differences between levels were not significant), and the four samRaw data and statistical analyses: total fauna (corresponding to Tables 7, 10 and Fig. 4a). Medium sand with fine sand, well sorted (medium sand). X: mean of transformed data; S~: variance of transformed data; dr: degrees of freedom; S.S.: sum of squares; M.S.: mean square; F: ratio; p: level o~f probabdlty. ". Significant at less than p - 0.05, Ies than p ~ 0.01; *** less than p = 0.001. - - Non significantly different means (within bars). + Preserved material used in elutriation. (Further explanations in the text) Decantation total sorting EL TH Elutriation OH 176 242 372 110 126 205 294 180 96 255 673 173 101 316 306 194 2 . 0 8 3 1 2.4004 2.5881 2.2O56 0.0143 0.0060 0.0275 0.0124 Sea-water ice technique H H UH One-way analysis of variance (after transformation to logl0) Source of variation Between methods Within methods 0.001 7,24 = 5.23 LSD test of ranked means (LSD = 0.1789) LSD = t(v)]//-~---Ms within and t(~) is the value of t from standard tables for a probability p --- 0.05 and with (v) degrees of freedom, where v is the within group degrees of freedom from the analysis of variance table, n is the number of samples per group and MS within is the within group mean square from the analysis of variance table. In the following examples the means are ranked from left to right and non-significantly different means are included within bars. The LSD is subtracted successively from each mean until all means are included. Raw data and statistical analyses: total fauna (corresponding to Tables 8, 11 and Fig. 4b). Coarse sand with shell gravel, fairly welI sorted (coarse sand). (Explanations see Table 3 and text) Decantation total sorting EL TH Elutriation OH 404 480 378 171 346 242 260 258 342 215 796 197 278 391 932 397 2.5309 2.4957 2.7157 2.3845 0.0044 0.0276 0.0694 0.0259 Sea-water ice technique H H UH LSD test of ranked means (LSD) == 0.2446) H H UH TH DR + GR 2.6990 2.6857 2.5619 2.5309 2.4957 C O M P A R I S O N A N D E V A L U A T I O N O F S E P A R A T I O N M E T H O D S F o r the general evaluation o f the separation methods tested b y our experiments, " K e n d a l l ' s Tau r a n k correlation" was applied (So~:AL & ROHL~ 1969). The various methods were ranked against numbers of organisms from total fauna data, N e m a t o d a data, h a r d fauna and soft fauna d a t a to give a total r a n k for the methods. U n d e r this solely numerical comparison, decantation and elutriation ranked highest and sea-water ice t r e a t m e n t next, while d u t r i a t i o n with anaesthetization and the w a r m - w a t e r method were less effective. H o w e v e r , such comparison does n o t t a k e i n t o consideration that (1) the various animal groups are not separated w i t h the same accuracy, (2) the organisms are collected alive in some methods a n d d e a d in others, (3) the separation efficiency differs from one sediment t y p e to another and (4) the time required for concentration, sorting and counting is unequal. These topics will be discussed in the following paragraphs. Raw data and statistical analyses: total fauna (corresponding to Tables 9, 12 and Fig. 4c). Fine sand, well sorted (fine sand). (Explanations see Table 3 and text) Decantation total sorting EL TH Elutriation OH One-way analysis of variance (after transformation to logl0) Source of variation Between methods Within methods LSD test of ranked means (LSD = 0.2343) 98 9i i68 33 79 87 77 21 31 127 76 21 36 112 74 54 25 69 190 88 121 78 1.6669 1.9781 2.0165 1 . 5 6 8 0 0.0685 0.0105 0.0315 0.0730 Sea-water ice technique H H UH 1 . 9 5 6 8 1.7773 0.0058 0.0285 Investigations of benthic biocoenoses are generally concerned with the total fauna. Thus, we tried to separate the total meiofauna from our samples. The results are given in Tables 3-6. The statistical evaluation of the methods is demonstrated b y the Least Significance Difference (LSD) test of r a n k e d means. The non-significantly different means are included w i t h i n the bars. I n Table 3 the highest mean (McI) was n o t signific a n t l y different f r o m (EL), ( U H ) or ( T H ) b u t was significantly better t h a n (GR), ( H H ) , ( O H ) and (DR). S i m i l a r l y (EL) was not different from (UH), ( T H ) and (GR) but better t h a n ( H H ) , ( O H ) a n d (DR). The largest break occurred between ( H H ) a n d ( O H ) w i t h three groups terminating here. The difference between the t w o sea-water ice procedures ( U H ) a n d ( H H ) resulted from the splitting of the total sample into three grain-size fractions b y ( U H ) and consequently in a m o r e thorough treatment. Raw data and statistical analyses, total fauna (corresponding to Fig. 4d). Medium sand, poorly sorted (muddy sand). (Explanations see Table 3 and text) Decantation total sorting EL TH Elutriation Sea-water ice technique H H UH Two-way analysis of variance (aider transformation to logl0) 0.01 6,16 = 4.20 p 0.05 3,16 = 3.24 One-way anova without level A (after transformation to log10) Source of variation Between methods Within methods 0.01 6,13 = 4.62 a n d d e a d s e p a r a t i o n It was one of our intentions at the Helgoland meeting, to compare the efficiencies of methods for the separation of living and preserved meiofauna. The question posed Raw data and statisticaI analyses: hard fauna (corresponding to Tables 3, t0 and Fig. 4a). Medium sand with fine sand, well sorted (medium sand). (Explanations see Table 3 and text) Decantation total sorting EL TH OH 154 192 228 101 101 177 t75 t73 83 203 521 155 84 245 201 189 2.0088 2.3070 2.4052 2.1773 0.0157 0.0036 0.0454 0.0145 Sea-water ice te&nique H H UH Source of variation Between methods Within methods Elutriation 356 430 178 170 289 205 192 257 307 167 578 187 179 328 803 394 2.4381 2.4209 2.5501 2,3769 0.0167 0.0351 0.1105 0.0271 one-way analysis of variance (after transformation to tog10) Decantation total sorting EL T H Elutriation 72 65 126 27 66 70 69 19 19 112 62 20 34 94 50 50 19 60 156 87 99 6O 1.5531 1.8917 1.9139 1.5299 0.0786 0.0131 0.0341 0.0804 0.05 7,34 .... 2.55 Raw data and statistical analyses: h a r d fauna (corresponding to Tables 4, 11 and Fig. 4b). Coarse sand with shell gravel, fairly well sorted (coarse sand). (Explanations see Table 3 and text) Raw data and statistical analyses: hard fauna (corresponding to Tables 5, 12 and Fig. 4c). Fine sand, well sorted (fine sand). Explanations see Table 3 and text) One-way analysis of variance (after transformation to logl0) Source of variation Between methods Within methods LSD test of ranked means (LSD = 0.2505) was, to w h a t extent can living extractions be considered as quantitative methods? The live methods were the sea-water ice treatments ( H H ) , ( U H ) , elutriation (DR), and elutriation w i t h anaesthetization (GR). Live elutriation w i t h o u t anaesthetization is less effective, as obviously m a n y organisms remain a t t a & e d to the sediment grains during the procedure. Better results were obtained with an anaesthetic in a closedcircuit system ( G R : Tables 3-5). The sea-water ice methods involved both sea-water ice treatment followed by preservation and staining of the sand residue, a t w o - p a r t te&nique (Tables 3-6). They r a n k at about the: same, level as elutriation w'ith anaesthetic (GR). Furthermore, the effectiveness of sea-water ice w i t h o u t a d d i t i o n a l preservation, was examined. By this test, the ( H H ) method, which involves exposing a larger sediment surface to ice than in the ( U H ) method, is clearly more effective. H o w e v e r , as sole treatments, both are tess efficient than preservation and elutriation o r decantation (McI) and (EL). The i m p o r t a n t advantages of live extraction b y the sea-water ice t r e a t m e n t are the isolation of meiofauna (especially the sot% fauna) in unharmed condition, and the almost q u a n t i t a t i v e isolation of ciliates and flagellates, not recovered with any other method (Tables 1 and 2). Raw data and statistical analyses: soft fauna (corresponding to Tables 3, 7 and Fig. 4a). Medium sand with fine sand, well sorted (medium sand). (Explanations see Table 3 and text) Decantation total sorting EL T H Elutrlation 22 50 143 9 26 28 118 7 13 52 153 15 17 7i 120 5 1 . 2 7 5 4 1.6783 2.1228 0.9186 0.0t73 0.0284 0.0031 0.0404 LSD test of ranked means (LSD = 0.1972) EL H H TH GR 2 . 0 9 6 8 1 . 9 5 6 3 1 . 7 3 0 0 1 . 6 7 8 3 It was thought that some methods may be more effective in extracting hard fauna (Nematoda, Ostracoda, Copepoda and Halacarida), whilst others may be better for sot~ fauna (Turbellaria, Gastrotricha, Archiannelida, Polychaeta, Nemertea and solitary Cnidaria). Tables 7, 8 and 9 show the data for hard fauna and Tables 10, 11 and 12 show .the data for soft fauna. The split into hard and soft: fauna was not made for m u d d y sand since the fauna was composed almost exclusively of nematodes, i.e. the results correspond more or less with the total fauna (Table 6). Tables 7 and 9 indicate that there was little difference between methods, only (DR) in Table 7 and (DR) and (OH) in Table 9 being significantly less effective. In Table 8, however, the t w o - w a y analysis of variance showed a high between-level variance and a non-significant between-method variance. Consideration of the raw data demonstrated that the large variation in numbers of Ostracoda probably accounted for the fact that no difference could be detected between methods (Table 8). Tables 10, I1 and 12 show that for the soft fauna the preservation methods of (EL), (McI) and the sea-water ice methods (UH) and ( H H ) were consistently the most Raw data and statistical analyses: soft fauna (corresponding to Tables 4, 8 and Fig. 4b). Coarse sand with shell gravel, fairly well sorted (coarse sand). (Explanations see Table 3 and text) Decantation total sorting EL TH OH 48 50 199 1 57 37 68 1 35 48 217 10 99 63 I28 3 1.7442 1.6869 2.1438 0 . 3 6 9 3 0.0358 0.0089 0.0531 0.2274 Sea-water ice technique H H UH successful. T h e preservation and direct sorting method ( T H ) was m o r e variable whilst anaesthetization (GR), the (DR +) preservation and w a r m - w a t e r ( O H ) methods were significantly less effective. H o w e v e r , one must consider t h a t preservation does not a l l o w accurate identification of sot~ fauna representatives. Ciliates, for instance, may easily be confused with turbellarians. S e d i m e n t l o s s i n s e p a r a t i o n t~aw data and statistical analyses: soft fauna (corresponding to Tables 5, 9 and Fig. 4c) Fine sand, well sorted (fine sand). (Explanations see Table 3 and text) Decantation total sorting EL TH 1.5116 1 . 3 2 7 9 0.0126 0.0488 Elutriation OH 26 26 42 6 13 27 8 2 12 15 14 1 2 18 24 4 6 9 34 1 22 18 0.9374 1.2464 1.3117 0.3362 0.1774 0.0382 0.0581 0.1233 Sea-water ice technique H H UH 1 . 3 2 0 0 1.1358 0.0323 0.0483 LSD test of ranked means (LSD = 0.3506) H H McI GR UH 1 . 3 2 0 0 1.3117 1.2464 1 . 1 3 5 8 In general, elutriation and decantation are the quicker concentration procedures, while in sea-water ice treatment the time required depends on the melting process. Concerning sorting and counting, a surprisingly quick procedure was demonstrated by direct in situ counting (p. 180). Contrary to this, other participants counted the fauna by picking out with pipettes, which is much more time consuming. Differences in time loss relate to the sediment quota in the counting dish. Under this aspect seawater ice treatment ranges over elutriation, decantation and total sorting. SUMMARY 1. This paper presents the results of a meeting on the evaluation of quantitative procedures for the separation of meiofauna, held at the Marine Station of the Biologis&e Anstalt Helgoland, in May 1972. Close co-operation on the part of the participants (p. 194) has allowed assessment of advantages and disadvantages of the separation methods applied. The time needed for preserved methods can be reduced by changing to scanning rather than pi&ing out for sorting and counting. 2. Sorting without concentrations- This method is suitable for very fine-grained sediments, oozes; it is generally applied for preserved samples; adequate for hard fauna; insufficient for soft fauna; very time consuming. 3. Decantation: Suitable for sandy sediments; generally applied for preserved samples; adequate for hard fauna; insufficient for sot~fauna; time consuming. 4. Elutriation: Suitable for sandy sediments; for unpreserved samples only with anaesthetization, more effective with preserved samples; limited for total live fauna. With preserved samples, adequate for hard fauna; insufficient for sot~ fauna; quick method. 5. Warm-water elutriation" Suitable for sandy sediments; designed for live hard fauna (especially nematodes, ostracods); quick method. 6. Sea-water ice treatment: Suitable for sandy sediments with microporal structure; only for live extraction; limited for hard fauna; well suited for sof~ fauna (including ciliates and flagellates); time consuming. Acknowledgements. Travelling funds for all participants were granted by their National Agencies. Substantial support by the BiologischeAnstalt Helgoland and financial aid by the "Sonderforschungsbereich 94 - Meeresforschungan der Universidit Hamburg" is gratefully a&nowledged. Thanks are due to Dr. K. FmGE,Deutsches Hydrographisches Institut (Hamburg), for the grain-size analyses. LITERATURE CITED L I S T O F P A R T I C I P A N T S 2 Hamburg 50, Palmaille 55, Federal Republic of Germany. 2 Hamburg 50, Palmaille 55, Federal Republic of Germany. 2 Hamburg 50, Palmaille 55, Federal Republic of Germany. 2 Hamburg 50, Palmaille 55, Federal Republic of Germany. Urn.m, G., Biologis&e Anstalt Helgoland (Meeresstation), 2192 HelgoIand, Federal Republic of Germany. Unable to attend: Visitors to the meeting: G. UHLm , H . THIEL & J. S. GRAY ALT, G. , 1973 . On the vertical and horizontal microdistribution of the meiofauna in the Greifswalder Bodden . Oikos (Suppl.) 14 (in press). Boiss~Au , J. P. , 1957 . Techniquepour l' ~tude quantitative de la faune interstitielle des sables . C. r. Congr. Socs say. Paris Sect. Sci., 117 - 119 . G. UHLIG , H. THIEL & J. S. GRAY ELMCREN,R., 1973 . Methods of sampIing sublittoral sol[ bottom meiofauna . Oikos (Suppl.) 14 (in press). HULmGS , N. C. (Ed.), 1971 . Proceedings of the First International Conference on Meiofauna. Smithson. Contr. Zool . 76 , 1 - 205 . - - & GRAY,J . S. (Eds), 1971 . A manual for the study of meiofauna . Smithson. Contr. Zool . 78 , 1 - 83 . RUSSELL , F. S. & COLMAN, T. E. , 1931 . The zooplankton I. Gear, methods and station lists . Scient. Rep. Gt. Barrier Reef Exped . 2 ( 2 ), 1 - 35 . SO ~AL, R. P. & ROHLF , F. T. , 1969 . Biometry. Freeman, San Francisco, 776 pp. Tm ~L, H. , 1966 . Quantitative Untersuchungen fiber die Meiofauna des Tiefseebodens . VerSff. Inst. Meeresforsch. Bremerh. (Sonderbd) 2 , 131 - 148 . UttLm , G. , 1964 . Eine einfache Methode zur Extraktion der vagilen, mesopsammalen Mikrofauna . Helgol~inder wiss . Meeresunters . 11 , 178 - 185 . - - 1968 . Quantitative methods in the study of interstitial fauna . Trans. microsc. Soc . 87 , 226 - 232 . At ~L~;, G. , Fachbereich Meeres- und Fischereibiologie, Sektion Biologie der Universit~t Rostock , 25 Rostock, Freiligrathstrafle 7 /8, German Democratic Republic. SWEr)MAR~, B. , Kristineberg Zoologiska Station , S-45034 Fiskeb~ickskil, Sweden. G. UHLIG , H . THIEL & J. S. GRAY GEgLACH, S. , Institut fiir Meeresforschung , 285 Bremerhaven, Am Handelshafen 12, Federal Republic of Germany. JtsARio , J. V. , Institut fiir Meeresforschung , 285 Bremerhaven, Am Handelshafen 12, Federal Republic of Germany. Rhc~toll, E. , Institut ftir Meeresforschung , 285 Bremerhaven, Am Handelshafen 12, Federal Republic of Germany. WALTER , M. , Institut fiir Hydrobiologie und Fischereiwissenschafi der Universit~it Hamburg, 2 Hamburg 50 , Palmaille 55 , Federal Republic of Germany. First authors's address: Dr. G. UHLIG Biologische Anstalt Helgoland (Meeresstation) 2192 Helgoland Federal Republic of Germany


This is a preview of a remote PDF: https://link.springer.com/content/pdf/10.1007%2FBF01609968.pdf

G. Uhlig, H. Thiel, J. S. Gray. The quantitative separation of meiofauna, Helgoland Marine Research, 1973, 173-195, DOI: 10.1007/BF01609968