A sampling device for soil microarthropods

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The Berlese funnel method for the removal of arthropods from samples of soils and other materials has been used by numerous investigators for qualitative col-.
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A SAMPLING DEVICE FOR SOIL MICROARTHROPODS BY

S. I. AUERBACH and D. A. CROSSLEY, Jr. (Health Physics Division, Oak Ridge National Laboratory, Oak Ridge, Tennessee)

1.

INTRODUCTION. The Berlese funnel method for the removal of arthropods from samples of soils and other materials has been used by numerous investigators for qualitative collections. Many modifications of the apparatus, designed to operate on varions materials and for the collection of varions arthropods under differing conditions, have been publi~hed. Ali use heat, light; dessication, fumigants, or a combination of these applied to a sainple of soil or other material .contained in the top of a funnel. Arthropods are driven out of the sample and fall into a collecting jar fastened to the neck of the funnel. The most extensive use of the Berlese funnel has been in the procurement of specimens for taxonomie purposes. Less frequently the method has been employed for quantitative estimates of the arthropod fauna. Historically the developrhent of a quantitative application has been characterized by a controversy over the choice of a method which will yield the closest approximation to the total number of arthropods !na given medium. One school of thought has favored the "funnel" technique while a second school has advocated a " flotation " technique. MACFADYEN (1955) has ably reviewed and discussed the arguments for each of these general approaches. Insofar as the so-called microarthropods, particularly the mites and collembolans, are concerned, the general criteria for a practical extraction apparatus favor sorne type of funnel . deviee. Essentially what is needed is a method which will permit sampling and extraction of soil arthropods under conditions which may be duplicated easily for numbers of samples and which will be efficient for the majority of the microarthropods. This paper describes a sampling tool and an extraction apparatus which were designed to meet these qualifications. MACFADYEN (1953) has summerized certain milestones in the development of quantitative Berlese funnel method~, as follows : TuLLGREN (rgr8) first used an I.

Operated by Union Carbide Company for the U. S. Atomic Energy Commission. Acarologia, t. II, fasc. 3, rg6o.

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electric light bulb as a heat source and thus also added light as a stimulus. FoRD (1937) cornbined a battery·of small TullgJ;"en funnels for statistical treatment of large numbers of samples. HAARL0V (1947) vented the funJ:].els to prevent the condensation of moisture in them. HAMMER (1944) instituted the practice of placing undisturbed samples in the funnels in an inverted position, rather than breaking the samples apart. We would add that MACFADYEN (1953) combined these developments into a compact set of small funnels designed to produce a high gradient of temperature and humidity in the samples. The funnel system described in the present paper embodies the major features of Macfadyen's highgradien t design . THE SAMPLING

TOOL.

The sampling tool (Fig. r) is a stainless steel cylinder approximately 7 in. long, in. outside diameter, and I 3/4 in. inside diameter. One end of the _cylinder (the cutting end of the tool) is increased to I 5/8 in. inside diameter over the first half inch of its length,- so that a shelf is formed inside the cylinder. This end is also beveled to form a cutting edge. The slope of the bevel is such that it provides ease of entry into the soil with a minimum of lateral compression of the sample. In addition, the slightly greater diameter of the top of the bevel as compared to the rest of the cylinder reduées adhesion of the tube to the surrounding soil. The sample holders are hollow plastic cylinders 2 in. long and r 3/4 in. outside diameter (Fig. 2 and 3). The cylinders are made of nie thyl methacrylate (" Lu cite " , " Perspex ", " Plexiglas ") which is easily machined. These are inserted in the top of the sampling tool and come to rest on the shelf near the bottom. A solid "lucite " cylinder of I 3/4 in. diameter is then inserted to fill the remainder of the tube and the sampling tool is ready for use. Aluminum cylinders may be used in place of plastic ones for sample containers. The plastic cylinders; hm·vever, are rouch lighter (r5 g vs 40 g) and thus permit more accurate weighing of the contained sample. Also, the transparency of the plastic sample containers is an advantage. In practice, the sampling tool with the sample container is pressed into the soil to a depth of 2 rj2 inches. An ~ngraved line on the outer surface of the tool 1 . serves as a constant depth marker. When the tool is withdrawn, a solid " lucite " cylinder of I 5/8 in. diameter is inserted into the cutting end and used to push the sample container and sample out the top of the tool. The sample container with the contained sample is then wrapped with aluminum foil for transport to the laboratory. The original design of this instrument allowed three of the "lucite " containers ,to be placed in the tube, the intention being to take samples of six-inch depth. The plan was to press the tube to that depth in the soil and thereby fill the three containers. After removal of the containers from the tube they were to be separab:!d by cutting the soil between the tubes and each container processed separately. 2

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In practice this method was not feasible. Unless the soil was of an optimum composition and plasticity, ·effective penetration and filling of ali three tubes was difficult to obtain. Subsequent removal of ali three containers with intact soil samples also proved difficult to achieve. Because of these limitations only one sample container has been used in each sampÜng.

THE EXTRACTION DEVICE. The extraction deviee was modeled closely after that of MACFADYEN (1953). Major differences between his- apparatus and ours include r) the use of methyl methacrylate instead of wood and metal for most of the structural components, 2) a compressed air inlet coupling located in the lower left front portion of the container (Fig. 2), 3) an air exhaust located on the upper right wall to the rear (Fig. 3), 4) an ammeter connected in series with the heaters, and 5) special trays which hold the sample containers a fixed distance above the glass funnels . A "lucite" box, r6 X ro r j2 X ro rj2 in., comprises the extraction apparatus (Fig. 2 and 3) 1 . Contained in it are removable trays to hold I5 sample containers and I5 small funnels . Walis of the box are made of "lucite" 3/8 in. thick. The sample holders are arranged in 3 banks of 5 each, with " lucite " partitions separating the banks. Nichrome wire in a refl.ector is used as a heat source, with separate heaters being provided for each bank. Removable trays hold the sample containers in position about r inch below the reflectors (Fig. 2) . A metal insert with a flange which fits into an opening in the tray, holds each sample container. The length of the metal insert is such that approximately half the length of the sample container is above the tray. A removable, circular piece of screen fits in the bottom of the metal insert. Since the screen is a separate item, a variety of mesh sizes can be used. For our purposes ro-mesh screen has proved to be satisfactory, A second set of trays holds the small glass funnels immediately below the sample container. \iVhen in position in the apparatus the bottom of the sample containers extends below the lip of the funnel. Because of this configuration it is necessary to load the trays and position the funnels outside the box. For this purpose the trays are mounted in an open rack, loaded, and then both trays containing samples and funnels are siqmltaneously inserted into the box, where they slide into grooves eut in the walls and partitions. Collecting tubes are attached to the stems of the funnels with rubber tubing. Several funnel and tube sizes have been used. Commercially made glass funnels (Pyrex 6o 0 , 65 mm ID, Corning No. 6r8o) with the stems removed and replacéd by larger (25 mm) diameter glass tubing are satisfactory. This size of funnel stem permits the use of Kimbal " Opticlear" 7-dram vials for collecting the arthropods. The partitions between the banks allow complete circulation of air below the I.

Detailed construction bluepriri.ts are a vailable from the authors upon request.

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-282funnels. The trays are partially vented, but also . serve as heat baffles to retain the heat in the sample compartments, thus making for a high gradient of temperature. The upper partitions between the banks are vented baffl.~s permitting partial circulation of heat above the samples. The banks are connected in parallel to the switch box making it possible to operate any combination of the three. However, the voltage supply cannot be varied independently for the three banks. During the operation compressed air is fed iri.to the apparatus in the lower left front of the box and emerges from 'the upper right rear (Fig. 2 and 3). Forced air circulation prevents moisture condensation :ln the funnels. This circulation is especially necessary during the first 48 hours of an extraction. The insulating action of the trays and partitions results in a steep gradient of temperature from the tops to the bottoms of the samples and, presumably, similar gradients of humidity are produced also. RESULTS AND DISCUSSION. The sample taken with the sampling tool is smaller than that used in older studies but is of similar size to those used in more recent investigations (MAcFADYEN, 1953 ; MuRPHY, rg56). The surface area of the samples is 2.r in.2 (r3.5 cm 2) and ·the depth of the samples varies between r and 2 in., so that the volume of the samples is about 2-4 in. 3 (35-70 cm 3). The average number of arthropods recovered is about so-rso per sample, depending upon such factors as litter type and time of year. These numbers are satisfactory for statistical validation of many ecological studies, and are not so large as to render accurate counts · too time-consuming: Routinely, weights of samples are taken before and after treatment with the extraction deviee. ·For a given soil, differences in dry weight of samples refiect differences in depth of sample only, since the area covered is constant. Usually, the number of arthropods per sample varies independently of the weight of the sample. This suggests that most of the arthropods are in the top portion of the soil, so that weight differences due to varying amounts of deeper soil have little effect. Rarely, the heavier samples contain fewer arthropods thau do the lighter ones. Figure 4 shows the relation between weight of core an~ number of animais recovered for a series of cores in which the heavier samples contained fewer arthropods than did the lighter ones. In most cases the soils contained in series of samples may be considered to be of similar volume or surface area and differences between dry weights .of samples can be ignored. MACFADYEN (r953) has demonstrated that a high gradient of temperature and humidity is desirable in samples for most efficient recovery of micro-arthropods. The apparatus used by Macfadyen produced a sharp gradient in temperature, in contrast to the conventional Tullgren type funnel. Also, samples in the Tullgren funnel were subjected to considerable diurnal fluctuation in temperature due to varying room temperatures, while the high-gradient apparatus was relatively

independent of diurnal variation. . A steep gradient of humidity also was found in the high-gradient apparatus. We did not attempt to measure carefully the gradients of temperature or humidity in our apparatus. A difference of 8° C between the tops and bottoms of the soil cores was obtained after 2 days of operation; this is similar to the result obtained in Macfadyen's more detailed studies. In order to assess the relative efficiency of the extraction deviee, a comparison was made with a series of Tullgren funnels. These latter funnels were large ones (ro in. dia.) with steep sides. · Duplicate soil core sainples were extracted simul- · taneously in the high-gradient apparatus and in the Tullgren funnels. Both 40 and 6o watt bulbs were used in the Tullgren funn~ls as heat sources. The results are summarized in Table I. It is seen that the high-gradient apparatus has no advantage over the Tullgren funnel except in compactness. The recovery of 346 arthropods from a single sample (40 watt bulb in Tullgren apparatus) is unusual and can probably be disregarded; if this is done there is virtually no difference in efficiency between the three treatments. TABLE

I. Numbers of microarthropods recovered in two types of Berlese Appm·atuses

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(Several replicates).

Collembola

Funnel type

Other Insects

Trombi- Mesostig- Oribatei diformes mata

Other

Total

---6o-watt bulb ( in Tullgren 1 1 1

40-watt bulb in Tullgren

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High-gradient apparatus -

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39 .42 r6

2 r 3

72 44

TJ 45 56 '

29

6 2 5

37 44

4 3 3

46 65 148

15 4 r2

47 26 108

5

49 44 55 35

6 6 5 5

56

57 '

I4 6 9

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190 132 106

7

136 153 346

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29 36

13

162 94 128 106

10 19

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36 23 31 16

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7

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These results, in comparison with those of Macfadyen, suggest that a sharp gradient of temperature in the sample may not be so important as the stability of the temperature. The Tullgren funnels were maintained in a constant laboratory climate (24 ± 1° C and 50 ± 5 % R. H.) without diurnal fluctuations which may account for their relative success. The temperature gradient between the tops and bottoms of the samples was 2° C or less, in comparison to the 8° C gradient in the high-gradient apparatus. Temperatures on the tops of the samples· under the 40 watt bulbs reached about 36° C, as did samples in the high-gradient apparatus. These temperatures under the 6o watt bulbs were about 45° C. Tullgren funnels,

when used in an environment subject to ·diurnal fluctuations, may show similar fluctuations in temperatures of the soil samples. The high-gradient apparatus is sufficiently insulated by the air spaces so that environmental fluctuations in temperature have little effect. ; A further comparison of the high-gradient and the Tullgren deviees was made by examining day-to-day recoveries of arthropods. Figure 5 shows the numbers of three arthropod groups collected in the first five days of extraction. The Tullgren funnel with a 6o watt bulb was used. The histograms indicate ·t hat most of the animais in the high-gradient samples were recovered on the second day, while greatest recovery in the Tullgren samples occurred on the first day. Also, the recovery of the thin-skinned prostigmatic mites ceased after the second day in the Tullgren samples. These comparisons suggest that the establishment of temperature and humidity gradients occurs m~re slowly in the high-gradient apparatus than in the Tullgren funnel. The result would be a gentler action by the highgradient apparatus. Recovery for those animais which are the more sensitive to low humidities is probably greater in the high-gradient apparatus than in the Tullgren funnels. It is concluded that the sampling tool and extraction deviee described produce adequate samples of arthropods. The advantages of the high-gradient apparatus ovet the Tullgren funnel are _those of compactness and greater control of the variables (temperature and humidity) which govern extraction efficiency. Our comparisons havebeen made by use of the total number of arthropods extracted, which is a poor measure of the utility of the deviee, By discovering the optimum temperature and air flow, the high-gradient deviee could deliver the optimum recovery for an arthropod species or group of species. , Such manipulations would prove difficult with the Tullgren funnel. The high-gradient apparatus is not well suited, however, to studies of the larger arthropods such as carabid beetles, nor is it applicable for qualitative investigations requiring processing of large volumes of material. For , these purposes the Tullgren funnels are superior. When large volumes of material are used, the temperature and humidity conditions in the samples are probably more satisfactory than those produced in the smaller samples for quantitative estimates of microarthropods.

SUMMARY I. A sampling tool and a high-gradient Berlese-type apparatus are described which will permit sampling and extraction of soil arthropods under conditions which easily may be duplicated for numbers of samples and which will be efficient for the majority of the inicroarthropods. 2. The efficiency of the high-gradient apparatus was compared to that of a standard Tullgren-type funnel with both kept in a constant environment. No difference in efficiency between the two types was found.

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Sicle view of high-gradient apparatus.

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