Bijdragen tot de Dierkunde, 55 (1): 88-94 — 1985

The assimilation of heavy metals by variegatus and

Glomeris marginata (Chilopoda; Diplopoda)

by

S.P. Hopkin K. Watson M.H. Martin & M.L. Mould

Departments of Botany and Zoology, Woodland Road, The University, Bristol BS8 1UG, United Kingdom

which 10 times than in Abstract litter are to 20 greater

similar uncontaminated woodlands. Concur- Specimens of the and the

rent with this contamination is a thick of Glomeris marginata collected from uncontaminated layer leaflitter which sites, and sites contaminated with heavy metals, have been undecomposed represents more fed tissues from the on terrestrial isopod asellus and than 20 of accumulation years (Coughtrey et leaf litter, respectively, containing different concentrations al., 1979). of lead and The which zinc, cadmium, copper. extent to A study of the soil/litter fauna has shown these elements are accumulated by Lithobius variegatus or that there differences in the extent to Glomeris the of metals in the clearly are marginata depends on amounts

food the which different taxonomic of in- and degree of contamination of the site from groups

which the and collected. are The the Some vertebrates are affected by pollution. tissues within the where metals stored body are are com- groups such as millipedes and earthworms are pared in the two and suggestions are made of the reduced greatly in numbers whereas others which in of areas are most need further research. such and woodlice as centipedes large are ap-

parently unaffected (table I). The reduction in

the numbers of and earthworms INTRODUCTION millipedes almost certainly accounts for the reduction in

One of the world's lead largest primary zinc, decomposition oflitter at the contaminated sites

and cadmium works is smelting situated at since previous studies have shown that there is

Avonmouth, 10 km to the north-west of Bristol no reduction in abundance or diversity of in south-west the of England. Despite presence microorganisms and fungi as a result of the which sophisticated filtering equipment pollution (Martin et al., 1980). The springtails,

98 the metals removes in excess of % of from the larvae and beetles mites, Diptera predatory are

waste amounts of zinc, cadmium, in unit gases, large present greater numbers per area, prob-

lead and emitted each fine copper are year as ably because the larger standing crop of leaflit-

which than particles, some of are carried more of material ter supports a larger amount fungal

20 km to the north-east the south- mites by prevailing on which and springtails feed, which in

westerly winds. for insects. turn provide more prey predatory

Our studies have been concerned In de- mainly this presentation, experiments are

with the of this on the impact pollution ecology scribed on the assimilation of heavy metals by of two oak-hazel woodlands, Hallen Wood and the centipede Lithobius variegatus Leach which is

Haw Wood, which are situated on either side of common in the contaminated woodlands, and the M5 3 km downwind of the motorway, the pill millipede Glomeris marginata (Villers) works. These smelting woodlands are heavily which is absent (table I). The detailed results of contaminated with lead and zinc, cadmium, this work will be presented elsewhere (Hopkin &

which reach concentrations in the leaf copper Martin, 1984a; Hopkin et al., in prep.).

of this is However, the purpose paper to sum-

marise the main of this * findings work, to com- Present address: Department of Pure and Applied for the first time the assimilation of Zoology, University of Reading, Whiteknights, Reading pare zinc,

RG6 United 2AJ, Kingdom. cadmium, lead and copper in Lithobius variegatus BIJDRAGEN TOT DE DIERKUNDE, 55 (1) - 1985 89

TABLE I

of invertebrate ofindividuals in leaf litter and Density major groups (number m-²)

soil from contaminated km works a woodland 3 downwind of a smelting (Hallen)

and a similar but uncontaminated site (Wetmoor).

Hallen Wetmoor

2 Litter 14.28 1.35 standing crop (kg m dry wt.)

Isopoda Linnaeus 56 20

Trichoniscus pusillus Brandt 0 151

Diplopoda Polydesmidae 8 79

Julidae 0 11

Glomeridae 0 21

Chilopoda 112 116

Geophilomorpha 328 263

Arachnida Acari 129000 19400

Araneae 248 81

Pseudoscorpionidae 200 67

Insecta Collembola 20800 8688

Coleoptera 902 48

Coleoptera (larvae) 120 4

Diptera (larvae) 4590 291

Annelida Lumbricus rubellus Hoffmeister 17 3

Lumbricus terrestris Linnaeus 0 29

Allolobophora longa Ude 0 4

Allolobophora caliginosa Savigny 0 30

Octolasium 0 9 cyaneum Savigny

and Glomeris and The of Oniscus asellus consists of four marginata to suggest areas hepatopancreas tubules. Within individual blind-ending animals, the con- which are in most need of further research. centrations of lead and the zinc, cadmium, copper are

in same all four lobes (Hopkin & Martin, 1982b).

if the left removed and fed MATERIALS AND METHODS Therefore, pair are to a cen-

tipede, the amounts of metals in the food can be deter- Lithobius variegatus mined from the concentrations in the right pair of tubules.

of adult Lithobius and the Within each of the of fourteen Specimens variegatus two groups centipedes,

fed tubules from the Oniscus asellus (Linnaeus) were collected from Wetmoor seven were on hepatopancreas of

Wood (British Ordnance Survey grid reference ST 743 woodlice from Wetmoor and seven on tubules from the

876) and Haw Wood (ST 560 798) duringMay 1982. Wet- hepatopancreas of woodlice from Haw. The tissue was

each small of moor is an uncontaminated site, whereas Haw is 3 km presented to on a piece woodlouse

cuticle downwind of a primary smelting works and is heavily con- which does not contain detectable amounts ofzinc, taminated with lead and cadmium, lead or The other lobes stored zinc, cadmium, copper (Martin et copper. two were

small of filter Each al., 1982). on a piece Millipore paper. centipede

The woodlice were maintained in plastic tanks on leaf grasped the piece of cuticle with the poison claws and ate litter from the collection site. The starved the soft within about minutes. centipedes were tissue, usually ten The piece for four allow the of the be voided. of which removed the days to contents gut to cuticle, was never eaten, was on At the of fourteen the three the faeces end this period, centipedes of 2-3 cm in following day. During next days, void- length were selected from each site population and ed by the centipedes were collected for analysis. The

of metals transferred to individual Petri dishes. The dishes were kept amounts taken in by the centipedes could be

moist base within determined the on a a covered plastic tank and were thus accurately by subtracting amounts of all maintained at the same relative humidity and metals remainingon the piece of cuticle from the amounts

for the duration of the in the tubules. temperature(14-18 °C) experiment. opposite pair of hepatopancreas The cen- 90 PROCEEDINGS 6th INTERNATIONAL CONGRESS OF MYRIAPODOLOGY

Fig. 1. Diagrammatic representation of the experimental procedure used to

measure the assimilation of metals by Lithobius variegatus (for further details

see text).

few of the tipedes were fed on six occasions at intervals of four days field sites within a days of the end experiment,

The experimental design is summarised in fig. 1. starved for four days and dissected in the same manner.

Centipedes which died during the experiment were The tissues and samples of faeces were placed on small

filter dried 70°C dissected immediately. However, after the sixth feeding pieces of Millipore paper, overnight at

occasion, all the centipedes from Wetmoor fed onwoodlice and weighed on a microbalance. The samples were

from Haw had died (table II). Centipedes which were still digested in 2 ml of boiling Aristar grade concentrated

alive this for four dissected diluted ml with deionised distilled at stage were starved days and nitric acid and to 5

also. The of all when removed The were for cadmium, lead guts centipedes were empty water. digests analysed zinc,

from the and flame flameless body cavity. copper by (Varian AA775) or (Varian

GTA95) atomic absorption spectrophotometry (AAS).

Figures for the net fluxes of metals through Lithobius

TABLE II variegatus between the beginningand end of the experiment

calculated the (figs. 2-4) were by balancing amounts Number ofspecimens ofLithobius variegatus surviving after measured in the faeces with changes in the levels in the eating two lobes of the hepatopancreas of six specimens of tissues on the assumption that the concentrations prior to Oniscus asellus (n = 7 at start of experiment). the first feeding occasion were the same as in centipedes

collected from the field. The estimates of the amounts of

Lithobius this shown variegatus metals in the food calculated on basis and on

left hand side of the in the figures were good agreement Haw Wetmoor

with the amounts of metals in the opposite pair oftubules (con- (uncon- of the hepatopancreas of Oniscus asellus fed to the cen- taminated) taminated) tipedes.

Oniscus asellus Haw 5 0

Wetmoor 7 7 Glomeris marginata

Specimens of Glomeris marginata were collected from

Midger Wood (ST 796 893) and from Charterhouse (ST

Each centipede was dissected into ten tissue fractions 503 555) on the Mendip Hills to the south ofBristol during

(Hopkin & Martin, 1983). However, for the sake of October 1983. Midger Wood is an uncontaminated oak-

simplicity, the results of the analyses have been hazel woodland, whereas Charterhouse is rough grassland

into three fractions in this 'exo' the of a disused lead mine contaminated with amalgamated paper, 'gut', on spoil tips

the connective tissues lead and (the exoskeleton including adhering zinc, cadmium, copper (Hopkin & Martin,

inner and the 'rest' fraction of to the surface) consisting a 1982a).

ofall the in the In of twelve from each pooled sample other organs body cavity. Groups millipedes population were addition, seven centipedes were collected from each of the placed in factorial combination in individual Petri dishes BIJDRAGEN TOT DE DIERKUNDE, 55 (1) - 1985 91

leaf litter from Wood Acer on Midger (field maple, campestre that Lithobius from the suggests variegatus con- L.), Charterhouse (grass) and Haw Wood (field maple) taminated site have acquired tolerance to from which Glomeris marginata is absent. The millipedes elevated levels of metals in their diet. were maintained for two months in plastic tanks in the

The extent to which some metals are same manner as Lithobius variegatus. Faecal pellets were

removed few and stored for assimilated both the levels in the every days analysis. depends on

At the end of the small number of the experiment, only a food and the degree of contaminationof the site

millipedes had died The survivors were (table III). from which the collected. For centipedes were dissected into three tissue fractions, 'gut' (from which the if of Lithobius eat 'fat 'rest' example, specimens variegatus contents were removed), body' and a pooled food which contains levels of zinc than sample which consisted of the exoskeleton and other higher

within the In twelve would in their organs body cavity. addition, they normally expect diet, they

collected from each of the field sites within millipedes were assimilate the metal (fig. 2a). However, if they

a few days of the end ofthe experiment and dissected in the lose consume food containing lower levels, they The concentrations of lead same manner. zinc, cadmium, three times more zinc in the faeces than they and in the tissue and the fluxes copper fractions, between take in with the hepatopancreas tubules (fig. tissues during the experiment were calculated in the same

for Lithobius it difficult Most of this metal is lost from the and way as variegatus. However, proved 2b). gut

determine the of leaf litter loss of to consumption by weight it is interesting to note that our preliminary between leaves in the Petri dishes the beginning and end of the this observations on ultrastructure of organ the experiment due to fluctuations in the wet weight of this have revealed the of intracellular presence material. Consequently, the relative rates of consumption in which and of food determined from the of faeces granules calcium, phosphorus were weight pro- be duced. zinc can detected by X-ray microanalysis.

found These granules can also be in the faeces

and which zinc may provide a route by excess TABLE III can be removed from the body.

There differences in the Survival and daily production of faeces as a percentage of are important extent

Glomeris from uncontaminated body weightby marginata an which different take metals which to organs up woodland (Midger) and a disused lead mine would be overlooked if only overall assimilation (Charterhouse) fed on contaminated and uncontaminated rates were considered. For example, Lithobius leaf litter for two months (n = 12 at start of experiment). variegatus from Wetmoor assimilate only 7.2%

of the cadmium when fed on the faeces hepatopan-

creas of woodlice from Haw Millipedes Leaf litter n surviving production (fig. 3). However,

the concentration of this metal in the in- (%) gut

than 50 times creases by more during the ex- Midger Wood Midger Wood 10 6.4 The cells of this epithelial organ are Charterhouse 11 6.4 periment. barrier the of this Haw Wood 12 3.0 clearly acting as a to passage

potentially toxic element into the rest of the

Charterhouse Midger Wood 12 10.4 body tissues. Charterhouse 11 10.0 Lead is not assimilated by Lithobius variegatus, Haw Wood 12 2.0 when even the centipedes are fed on the tissues

of woodlice containing much higher concentra-

tions of this element than would in they expect

RESULTS AND DISCUSSION the wild (fig. 4). However, it is not known

whether this is due lead in in- to the being an Lithobius variegatus soluble form in the digestive fluids of the cen-

from the contaminated site survive whether Centipedes tipedes, or they possess specific longer than those from the uncontaminated site mechanisms which prevent its uptake. when both fed the The concentrations cadmium and populations are on of copper

increase in the of hepatopancreas of Oniscus asellus containing gut tissues centipedes fed on high concentrations of metals (table II). This the hepatopancreas of woodlice from their 92 PROCEEDINGS 6th INTERNATIONAL CONGRESS OF MYRIAPODOLOGY

'own' site. This suggests that adult Oniscus woodlice in which the concentrations of metals asellus do much not form a major proportion of the are lower (Hopkin & Martin, 1984b).

diet of Lithobius variegatus in deciduous

woodlands although they probably consume Glomeris marginata

large numbers of newly-released juvenile Glomeris marginata from both sites ate significant-

ly less leaf litter from Haw Wood than from

Wood Charterhouse or Midger (table III).

Presumably, they find some aspect of the com-

position of the leaves, which might include the

high concentrations of metals, 'distasteful'. In-

terestingly, Glomeris marginata from Charter-

house of both of leaf than ate more types litter

millipedes from Midger although the reason for

this difference is not clear. All the millipedes

initially lost weight in the first two weeks of the

those litter experiment although fed on from

Midger and Charterhouse showed signs of

of this loss the of recovery weight towards end

the fed leaflit- experiment. Glomeris marginata on

from Haw did this initial ter not recover weight

due loss, probably to their low relative con-

sumption of food.

fraction far the The 'rest' was by most impor-

tant tissue fraction for the storage of zinc (fig. 5)

and whereas for cadmium copper, (fig. 6),

significant amounts of this metal were also con-

is tained in the gut. It clear that the gut

is far less epithelium of Glomeris marginata suc-

cessful the of cadmium at preventing passage

from the food in the lumen across the

into the tissues the epithelium body than gut of

Lithobius variegatus. The fat body of Glomeris

marginata is not of major importance in the

of of the four metals. storage any

Figs. 2-4. Estimated assimilation rates (%) and net fluxes

in amounts of metals through Lithobius variegatus and be-

tween the ‘gut’, ‘exo’ and ‘rest’ tissue fractions between

the beginning and end of the experiment. The value given

in the left hand box of each tissue fraction represents the

of metal estimated be the of amount to present at start the

The value in the hand box the experiment. right represents

the amount of metal present at end. (ND, not detected.)

2a: Zinc in (nmol) Wetmoor centipedes fed on Haw

woodlice (WC/HW). 2b: Zinc (nmol) in Haw centipedes

fed onWetmoor woodlice (HC/WW). 3: Cadmium (nmol)

in Wetmoor fed Haw woodlice centipedes on (WC/HW).

4: Lead (nmol) in Wetmoor centipedes fed on Haw

woodlice (WC/HW). - 93 BIJDRAGEN TOT DE DIERKUNDE, 55 (1) 1985

Glomeris like Lithobius than from the marginata, variegatus, copper millipedes uncon- does assimilate of taminatedsite when not significant amounts lead, both populations are fed on

when leaf even feeding on litter which contains contaminatedlitter (e.g. cadmium, fig. 6). This concentrations of this metal times is of the many greater difference between the populations one than would in the wild which needs be fur- they experience (fig. 7). many topics to investigated

Glomeris from Charterhouse ther. marginata con-

assimilate cadmium sistently more zinc, and

CONCLUSIONS AND SUGGESTED TOPICS FOR

FURTHER RESEARCH

The the assimilation of ability to regulate essen-

tial non-essential metals be the and may major

selective in sites that deter- pressure polluted

mines which of centipedes and

able survive and It millipedes are to reproduce.

is therefore essential that more extensive studies

carried into the of metal are out dynamics up-

take and excretion in myriapods. Some of the

lines of research which might be profitable to

follow are outlined below.

1. A detailed of the concentrations of survey

metals in of of a range species centipedes and

millipedes from polluted and unpolluted sites.

Some contaminated have unusual areas may an

fauna as a result of differential resistance of dif-

ferent species to pollutants.

2. Ultrastructural studies the in which on ways

the cells of different organs, particularly the

gut, deal with metals.

3. Breeding experiments to determine whether

the differences in the extent to which different

populations of Lithobius variegatus and Glomeris

Figs. 5-7. Estimated assimilation rates (%) and net fluxes

in amounts of metals through Glomeris marginata and be-

tween the ‘gut’, ‘fat body’ and ‘rest’ tissue fractions be-

tween the beginning and end ofthe experiment. The value

in the left hand box of each tissue given fraction represents

the of metal estimated the amount to be present at start of

the The value in hand box experiment. the right represents

the amount present at the end. 5: Zinc (nmol) in Charterhouse millipedes fed on Haw litter (CM/HL). 6a:

Cadmium in (pmol) Midger millipedes fed on Haw litter

(MM/HL). 6b: Cadmium (pmol) in Charterhouse

fed millipedes on Haw litter (CM/HL). 7: Lead (pmol) in

fed Charterhouse Midger millipedes on litter (MM/CL). 94 PROCEEDINGS 6th INTERNATIONAL CONGRESS OF MYRIAPODOLOGY

metals due to lead and within the of wood- marginata assimilate heavy are ac- copper hepatopancreas a

climation of the louse. Tiss. Cell, 14: 703-715. physiological processes during

& 1983. metals in the centipede Litho- , Heavy lifetime of the animal, or to the evolution of bius variegatus (Chilopoda). Environ. Pollut., (B) 6: mechanisms which can be genetically inherited. 309-318.

& , 1984a. The assimilation of zinc, cad-

lead and the Lithobius mium, copper by centipede ACKNOWLEDGEMENT variegatus (Chilopoda). J. Appl. Ecol., 21:535-546.

1984b. metals in woodlice. In: research & , Heavy S. This work was funded by a grant from the

L. SUTTON & D. M. HOLDICH eds., The of Natural Environment Research Council. biology

terrestrial isopods. Symp. zool. Soc. Lond., 53: 143-

166.

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