G Loranger

J F Ponge

& P Lavelle

Humus forms in two secondary semi-evergreen tropical forests

The dynamics and function of humus forms in tropical forests are still poorly understood. Humus profiles in two secondary semi-evergreen woodlands in Guadeloupe (French West Indies) were analysed micromorphologically. The humus forms are described under the canopy of five dominant tree species at two sites: under Pisonia subcordata and Bursera simaruba in a secondary forest on a Leptosol (Rendzina), and under Swietenia macrophylla, Tabebuia heterophylla and B. simaruba in a plantation on a calcareous Vertisol.

In the secondary forest, two distinct humus forms were observed. A calcareous Amphimull, characterized by an OH horizon comprising the faecal pellets of millipedes, is formed under the canopy of P. subcordata, which produces a litter that is rich in nitrogen. A Dysmull with a thick root mat (OF Rh horizon) develops under the canopy of B. simaruba, which produces a litter rich in lignin and phenol that is consumed slowly by soil fauna. In the plantation on the Vertisol, the activity of the endoanecic earthworm Polypheretima elongata has led to the rapid disappearance of litter and the mixing of organic and mineral material. The humus form is an Eumull and is similar under all three tree species present.

Introduction

Secondary forests in the tropics provide timber and fire wood, maintain biodiversity and organic matter in the soil, and they are a source of medicinal plants. Currently, 40% of the tropical woodlands are secondary forests and each year 9 million hectares are cut [START_REF] Brown | Tropical secondary forests[END_REF]). In the Caribbean the primeval forests have decreased during the four last centuries due to agriculture and urban development. The secondary forests resulting from human activities, are still much exploited and might disappear in the near future. On the island of Grande-Terre (Guadeloupe), secondary semi-evergreen forests cover only 2730 ha, i.e. 5% of the total surface. These forests contain few tree species, and several precious plant species, for example Calophyllum calaba, have disappeared. [START_REF] Huc | Premiers résultats expérimentaux sur le comportement de Tabebuia heterophylla (D.C.) Britton et de Hymenaea courbaril L., vis-à-vis de la sécheresse[END_REF], [START_REF] Lugo | Tropical forests in the Caribbean[END_REF] and [START_REF] Lugo | Comparison of tropical tree plantation with secondary forest of similar age[END_REF] have studied the vegetation in these forests, but no one has studied the dynamics of the soil organic matter.

The organic layers that form forest humus on the soil's surface are biologically very active, and decomposition is at its maximum there. The humus under temperate forests has been studied for over a century, and its three main forms, mull, moder and mor, are well established. In these forests, earthworms and white-rots (lignin-degrading fungi) produce mull in which litter decomposition is rapid and the organo-mineral horizon has a macrostructure. Mull humus profiles are distinct from profiles in which the activity of arthropods and enchytraeids dominate. In the latter, the moder and mor, litter decomposes slowly and organic and mineral materials are not mixed. A few investigations, such as [START_REF] Bernhard-Reversat | Litter incorporation to soil organic matter in natural and planted tree stands in Senegal[END_REF] and [START_REF] Leroy | Variations des caractéristiques de l'humus forestier d'un sol ferrallitique (Guyane) selon l'essence arborée considérée. Résultats préliminaires[END_REF], have studied the dynamics and function of humus forms in tropical forest ecosystems, and they are still poorly understood.

The value of humus classification based on that in the temperate zone is still uncertain.

Here we describe the structure of the humus forms under five tree species that produce litter with contrasting chemical qualities in two semi-evergreen woodlands in Guadeloupe, and we attempt to identify the biological processes influencing their development. Our hypothesis is that in these tropical forests, the quality of litter and soil fauna are the main factors determining the development of humus profiles, whereas microclimatic and edaphic properties are less important [START_REF] Lavelle | A hierarchical model for decomposition in terrestrial ecosystems: application to soils of the humid tropics[END_REF][START_REF] Aerts | Climate, leaf litter chemistry and leaf litter decomposition in terrestrial ecosystems: a triangular relationship[END_REF]. We examined the different components of humus horizons with a light microscope and described them in detail to understand their functioning. This method is one that we used and improved for the temperate zone [START_REF] Ponge | Ecological study of a forest humus by observing a small volume. I. Penetration of pine litter by mycorrhizal fungi[END_REF][START_REF] Bernier | Humus form dynamics during the sylvogenetic cycle in a mountain spruce forest[END_REF][START_REF] Peltier | Humus forms in Mediterranean scrublands with aleppo pine[END_REF], where it helped us to discern the role of soil organisms in the transformation of organic matter and the formation of horizons. Here it is tested for the first time for the tropics.

Materials and methods

Site description

The study was carried out in two semi-evergreen woodlands, at Port-Louis, in North Grande-Terre (Guadeloupe). This region has a 6-month dry season (December to May). The annual rainfall is 1250 mm on average, and the mean annual temperature is 26°C. During the dry season, the deciduous plant species loose their leaves and some of their twigs. The two plots have the same bedrock, a pure hard coral limestone from tertiary era (Pleistocene). A stratification exists within this thick (200 m) marine deposit. At the base (in contact with the volcanic shelf) are found softer sediments made of a mixing of calcareous shells, marine sands and volcanic debris. Above them was built the reef, now a wide plateau which has been inclinated and fractured by tectonic activity of the Caribbean plate.

The first site is a natural secondary forest relict of the primeval dry forest. It is one of the last such forests in Guadeloupe and is on a steep slope that may reach 45%. This slope corresponds to the NW border of a fracture between two plateaux. The maximum altitude is 79 m. The soil is a shallow calcareous Leptosol (FAO classification), which corresponds to a Rendzina in the CPCS classification [START_REF] Duchaufour | Abrégé de Pédologie. Sol, Végétation[END_REF]. It has a silt loam texture (71% silt in the upper 10 cm and 28% silt at 30 to 40 cm depth). In the top 10 cm, the soil is rich in organic matter (21% C) and has a C:N ratio of 12.5. The pH (in water) is 7.5 in the top 10 cm and 7.8 from 30 to 40 cm depth. The main canopy species are Pisonia subcordata L. and Bursera simaruba (L.) Sarg., two native deciduous tree species, which cover 32% and 24% of the total area, respectively [START_REF] Loranger | Déterminants de la Décomposition de la Litière dans une Forêt semi-décidue de la Guadeloupe[END_REF]. We described the humus profiles and soil macrofauna under the canopy of these two species.

In this secondary forest we observed that the calcareous rock attacked by HCl disintegrated most easily on the lower slopes. For an unknown reason, probably due to differences in the nature of the original coral population [START_REF] Gaiffe | Origine paléoécologique de l'aptitude des calcaires jurassiques à la fracturation. Conséquences tectoniques, pédogénétiques et écologiques[END_REF], the underlying calcareous rock becomes increasingly more resistant to weathering, and the root systems of trees become progressively less developed when going upwards. For both tree species (B. simaruba and P. subcordata), one humus profile was described on the lower, middle and upper slope (six humus profiles in total). Preliminary observations established that two distinct humus forms related to the two tree species occur in all three positions of the slope.

The second site is a 50-year-old plantation on a calcareous Vertisol on a plateau. This soil has a clay texture (78% clay in the upper 10 cm and 76% clay from 30 to 40 cm depth). The top 10 cm contains 5.3% C and has a C:N ratio of 12. The pH (in water) is 7.7 in the top 10 cm and 8 from 30 to 40 cm depth.

Originally, several species were planted for timber production by the National Office for Forests. Currently, the principal canopy species are Swietenia macrophylla King, Tabebuia heterophylla DC. Britton and B. simaruba, which cover 32%, 30% and 7% of the total area, respectively [START_REF] Loranger | Déterminants de la Décomposition de la Litière dans une Forêt semi-décidue de la Guadeloupe[END_REF]. S. macrophylla is an exotic species, while T. heterophylla and B. simaruba are native.

We have studied one humus profiles and the soil's macrofaunal communities under the canopy of these three deciduous species. All three species give rise to distinctive humus forms.

Soil macrofauna

We sampled the macrofauna in November 1997 (during the wet season) in the soil under the main tree species in both the secondary forest and the plantation, using the modified Tropical Soil Biology and Fertility method [START_REF] Anderson | Tropical Soil Biology and Fertility: a Handbook of Methods[END_REF].

Ten individual trees of each species were randomly chosen in the two woodlands.

Under the canopy of each tree, soil animals were collected and sorted by hand from a soil block (30 cm × 30 cm × 30 cm), which was dug out with a spade then sprinkled over a plastic sheet. Invertebrates were identified (a) at the species level for earthworms and millipedes and (b) as morphospecies for other groups.

Chemical leaf analyses

Freshly fallen leaves of the four canopy species, P. subcordata, S. macrophylla, T. heterophylla and B. simaruba, were collected from the forest floor. The leaves were air-dried and milled and the chemical composition determined.

The total nitrogen content was measured by the Kjeldahl method. Lignin and cellulose were analysed by sequential digestion of fibres [START_REF] Van Soest | Use of detergents in the analysis of fibrous feeds. II. A rapid method for determination of fibre and lignin[END_REF].

Samples were first extracted with neutral detergent. Lignocellulose (acid detergent fibre) was determined after extraction with acid detergent. Lignin (acid detergent lignin) was measured after hydrolysis with 72% H 2 SO 4 . The cellulose content corresponds to the difference between acid detergent fibre and acid detergent lignin. Total soluble phenols were extracted with 70% methanol then measured colorimetrically using the Folin-Ciocalteu method [START_REF] Marigo | Sur une méthode de fractionnement et d'estimation des composés phénoliques chez les végétaux[END_REF]. Tannins were measured with a colorimeter after precipitation with bovine serum albumin [START_REF] Hagerman | Protein precipitation method for the quantitative determination of tannins[END_REF].

Humus profiles

Humus profiles were sampled using the method described by [START_REF] Ponge | Etude écologique d'un humus forestier par l'observation d'un petit volume, premiers résultats. I.-La couche L 1 d'un moder sous pin sylvestre[END_REF]. A block of surface soil 5 cm x 5 cm x 15 cm (L x w x h) was cut with a sharp knife, with as little disturbance as possible, and the litter and soil surrounding it were removed and the block excavated. Each humus block was separated in the field by eye into its obvious layers, without reference to any pre-conceived classification of horizons [START_REF] Ponge | Horizons and humus forms in beech forests of the Belgian Ardennes[END_REF][START_REF] Peltier | Humus forms in Mediterranean scrublands with aleppo pine[END_REF]. The thickness of each layer was measured. They were then classified into OL (entire leaves), OF (fragmented leaves), OH (holorganic faecal pellets) and A (organo-mineral material) horizons, according to the classification of [START_REF] Brêthes | Classification of forest humus forms: a French proposal[END_REF], and their subdivision numbered from the top downwards, OL 1,2,… , OF 1,2,… , OH 1,2,… and A 1,2,… etc. Fifty six horizons were sampled in the secondary forest and 21 in the plantation. All 77 layers were immediately fixed in 95% ethanol and transported to the laboratory.

The composition of each humus layer was analysed under a dissecting microscope at ×40 magnification. In all, 45 types of material were recognized, 42 in the secondary forest and 28 in the plantation (Table 1). Most of the materials were plant litter at different stages of decomposition or comminution by soil fauna.

The volume ratio of each identified type was measured using the count point method [START_REF] Jongerius | Optic-volumetric measurements on some humus forms[END_REF][START_REF] Bal | Morphological investigation in two moder-humus profiles and the role of the soil fauna in their genesis[END_REF][START_REF] Bernier | Humus form dynamics during the sylvogenetic cycle in a mountain spruce forest[END_REF]. The 45 types were grouped into seven main categories which were used to build simplified humus diagrams (Figures 1, 2 and5). This micromorphological method enables the exploration of a greater volume of soil than the thin section method, but the pore patterns cannot be evaluated.

Millipede faecal pellets and organo-mineral material

After components of humus horizons had been identified and counted under the dissecting microscope, aggregates were inspected at the magnification ×400, using the method described by [START_REF] Bernier | Humus form dynamics during the sylvogenetic cycle in a mountain spruce forest[END_REF], to determine their internal composition. They were broken down gently in a drop of methyl-blue-lacto-phenol then observed under a phase contrast microscope. Three samples each of three categories (faecal pellets of millipedes, fresh earthworm casts, and organo-mineral material) were analysed. Eleven types of material were recognized under the microscope: plant fragments, mineral particles, free amorphous organic matter, amorphous organic matter bound to minerals, coal, pollen, plant epidermal cells, fungal hyphae, bacteria, earthworm cells and miscellaneous. The volume ratio of each identified component was again estimated using the count-point method.

Results

Soil macrofauna

Fifty-one species or morphospecies of soil macrofauna were found in the two forests (Table 2), 39 in the secondary forest and 28 in the plantation. The two sites had 16 morphospecies in common.

Millipedes, insect larvae, ants and termites were the most abundant arthropod groups. Millipedes (29 to 206 individuals m -2 ) dominated the soil macrofauna. The main millipede species was Anadenobolus monilicornis, which is known to inhabit dry soils [START_REF] Mauriès | Diplopodes Chilognates de la Guadeloupe et ses dépendances[END_REF].

Two earthworm species were found: Dichogaster sp., a small epigeic species, and Polypheretima elongata, an endoanecic species. Dichogaster sp.

was present in both sites (Table 2). P. elongata was found only in the Vertisol under the plantation.

Litter chemical analyses

Chemical analyses (Table 3) showed that freshly fallen leaves of B. simaruba and S. macrophylla had the biggest content of phenols and tannins. Due to their large lignin content, these leaves had the smallest cellulose content. The leaves of P. subcordata had more nitrogen (2.5%) than those of all other species.

Description and development of humus profiles: the secondary forest

In the secondary forest two different humus forms were observed. The first (Fig. 1) is characterized by a 1.5 cm thick OH horizon, which has a granular structure and consists of faecal pellets of millipedes. The organo-mineral A horizon below has a crumb structure. This humus form resembles a calcareous Amphimull as defined by [START_REF] Brêthes | Classification of forest humus forms: a French proposal[END_REF], but there is evidence of much millipede activity. This humus form is typical of the middle slope plot under P. subcordata.

The second humus form, which has a 7 cm thick root mat, or OF Rh horizon [START_REF] Loranger | Formes d'humus originales dans une forêt tropicale semidécidue de la Guadeloupe[END_REF], can be classified as a Dysmull [START_REF] Brêthes | Classification of forest humus forms: a French proposal[END_REF]. A similar root horizon has also been observed in tropical rain forests elsewhere [START_REF] Leroy | Variations des caractéristiques de l'humus forestier d'un sol ferrallitique (Guyane) selon l'essence arborée considérée. Résultats préliminaires[END_REF]. The Dysmull with its thick root mat is typical of the upper slope plot under B. simaruba (Fig. 2). In these profiles, the litter is usually transformed mainly by epigeic arthropods, but we found only a few animal faeces in the organic layers.

The organo-mineral A horizon has a crumb structure.

The influence of site and tree species on the composition of horizons was analysed by grouping the types of materials into broader categories, the percentage volume of which was calculated for the layer in which they were most abundant. The proportions of fragmented leaves (at 4-6 cm depth), organo-mineral aggregates (at 4-6 cm depth) and roots (at 8-10 cm depth) varied greatly between plots and trees (Table 4). Between 4 and 6 cm depth there were more fragmented leaves and fewer organo-mineral aggregates in the profiles on the upper slope than those on the two lower plots, indicating that litter was less rapidly incorporated on the upper slope than elsewhere. Between 8 and 10 cm depth, there were more roots under B. simaruba than under P. subcordata, showing that B. simaruba tends to develop of a root mat (OF Rh horizon).

Description and development of humus profiles: the plantation

In the plantation, humus profiles are similar for all three tree species in spite of differences in litter quality (Table 3). The OL horizon is discontinuous and comprises intact leaves, twigs, and wood. This horizon is 0.2 to 2 cm thick. OF and OH horizons are absent, and there are few millipede faeces in spite of the abundance of these animals (Table 2). The transition to the underlying organomineral A horizon, which has a crumb structure, is sharp. The humus form (Figure 3) is an Eumull [START_REF] Brêthes | Classification of forest humus forms: a French proposal[END_REF].

Bursera simaruba belongs to a family of trees known for their disinfecting properties [START_REF] Stehlé | Classification des arbres à latex et à sécrétion de gomme, résines et matières colorantes des Antilles françaises[END_REF]. Its leaves contain much lignin (23% of dry matter) and phenols (10% of dry matter) and decompose slowly [START_REF] Loranger | Déterminants de la Décomposition de la Litière dans une Forêt semi-décidue de la Guadeloupe[END_REF]. Both of these factors may slow the biodegradation of its litter. The OF Rh horizon is best developed in the plot on the upper slope where the parent material was a hard and pure calcareous rock. This weakly weathered bedrock might favour the development of a thick superficial root mat because roots cannot penetrate.

In the plantation, we found two macrofaunal communities, a) an epigeic (litter-dwelling) one dominated by millipedes and an endogeic (soil-dwelling) one dominated by the endoanecic earthworm P. elongata which feeds on organomineral material and probably also on faeces of the former group. The mixing role of this earthworm favours the formation of clay/humus complexes [START_REF] Bouché | Étude de l'alimentation et du brassage pédo-intestinal du lombricien Nicodrilus velox (Annelida, Lumbricidae) par l'analyse élémentaire[END_REF][START_REF] Bernier | Earthworm feeding activity and development of the humus profile[END_REF], which is shown by the abundance of organic matter bound to minerals (Table 5). Their activity maintains a macroporous crumb structure [START_REF] Alegre | Dynamics of soil physical properties in Amazonian agroecosystems inoculated with earthworms[END_REF][START_REF] Blanchart | Regulation of soil structure by geophagous earthworm activities in humid savannas of Cote d'Ivoire[END_REF] and forms an Eumull humus profile, whatever the tree species. They also bury litter under their casts thereby accelerating the incorporation of litter into the soil. The Vertisol has a large water capacity, providing suitable conditions for the establishment of P. elongata. When worm activity is lesser, as observed in the plantation under B. simaruba, the thickness of the OL layer increases. Despite the abundance of millipedes, only earthworm activity is expressed in the composition of the humus profile.

The organo-mineral aggregates in the plantation soil probably result from a combination of earthworm activity (rounded aggregates) and shrinking and swelling of smectites (polyhedral aggregates). In contrast, in the humus profiles of the secondary forest, the organo-mineral horizon has a macrostructure, the origin of which is still poorly understood. The organo-mineral aggregates of the secondary forest could be derived by the precipitation of colloidal organic matter by clay minerals [START_REF] Chenu | Influence of a fungal polysaccharide, scleroglucan, on clay microstructures[END_REF], followed by seasonal shrinking and swelling of smectites. It is possible, however, that in the sites we studied some millipedes might have ingested humified organic matter and deposited organo-mineral faeces in their galleries [START_REF] Loranger | Déterminants de la Décomposition de la Litière dans une Forêt semi-décidue de la Guadeloupe[END_REF], so that we can find old millipede organo-mineral faeces bound together in the A horizon. However, these millipedes cannot create large crumb structures as earthworms do.

In tropical semi-evergreen forests, as in temperate forests, the activity of endoanecic earthworms gives mull humus profiles, whatever the quality of the litter. These worms mix mineral and organic material, homogenize the upper the soil profile and help to maintain porosity. Where endoanecic earthworms are absent, as in the secondary forest, humus forms are more varied and depend on parent rock, litter quality, and millipede activity.

We conclude that soil fauna and litter quality are key factors in the development of humus profiles in tropical semi-evergreen forests and that parent rock and edaphic factors can be equally important in some calcareous soils. We believe, however, that our conclusions may not apply to non calcareous tropical soils.

Figure captions Figure 1 Figure 2 Figure 3

 captions123 Figure captions

Table 1

 1 Types of humus material in two semi-evergreen forests in Guadeloupe

	Categories

Table 2

 2 Densities of soil macrofauna (mean numbers of individuals per square metre with standard errors in brackets) under four tree species in two secondary semi-evergreen forests in Guadeloupe.

	Only Diplopoda (millipede) and Oligochaeta (earthworm) species are included
		Swietenia	Tabebuia	Bursera	Pisonia	Bursera
		macrophylla	heterophylla	simaruba	subcordata	simaruba
		(plantation)	(plantation)	(plantation)	(secondary	(secondary
					forest)	forest)
	Diplopoda (7 species)
	Orthomorpha coarctata	0	0	0	1.6 (1.7)	1.6 (1.7)
	Anadenobolus monilicornis	96.0 (26.7)	123.2 (30.8) 46.4 (19.1)	105.6 (50.6) 25.6 (11.3)
	Trigoniulus corallinus	0	0	1.6 (1.7)	1.6 (1.7)	1.6 (1.7)
	Spilostrophus naresi	33.6 (28.2)	24 (13.1)	51.2 (34.1)	19.2 (14.4)	0
	Epinannolene pittieri	4.8 (3.6)	59.2 (30.0)	4.8 (2.6)	0	0
	Pseudospirobolellus avernus	3.2 (2.3)	0	9.6 (5.3)	6.4 (4.5)	0
	Siphonophora filiformis	0	0	0	1.6 (1.7)	0
	Oligoaeta (2 species)
	Dichogaster sp.	3.2 (3.4)	4.8 (2.6)	9.6 (4.5)	0	4.8 (3.6)
	Polypheretima elongata	8.0 (3.8)	6.4 (3.7)	1.6 (1.7)	0	0
	Chilopoda (2 morphospecies)	9.6 (5.2)	4.8 (2.6)	3.2 (2.3)	4.8 (3.9)	9.6 (7.1)
	Coleoptera (5 morphospecies)	8.0 (3.8)	1.6 (1.7)	0	3.2 (3.4)	6.4 (4.3)
	Formicida (11 morphospecies)	6.4 (6.8)	4.8 (5.1)	12.8 (11.7)	41.6 (35.4) 35.2 (20)
	Isoptera (1 morphospecies)	57.6 (60.7)	17.6 (11.9)	9.6 (8.4)	1.6 (1.7)	0
	Insect larvae (10 morphospecies)	9.6 (5.2)	94.4 (27.2) 80.0 (57.0)	9.6 (9)	19.2 (9.4)
	Isopoda (3 morphospecies)	1.6 (1.7)	28.8 (8.3)	19.2 (13.7)	40.0 (20.2)	9.6 (6)
	Dermaptera (1 morphospecies)	0	0	0	3.2 (2.3)	9.6 (5.2)
	Blattodea (3 morphospecies)	0	0	3.2 (2.3)	4.8 (5.1)	9.6 (6.8)
	Araneidae (3 morphospecies)	0	0	0	3.2 (3.4)	3.2 (3.4)
	Heteroptera (1 morphospecies)	0	0	0	0	1.6 (1.7)
	Homoptera (1 morphospecies)	0	0	0	4.8 (5.1)	0
	Orthoptera (1 morphospecies)	0	3.2 (2.3)	1.6 (1.7)	0	0
	TOTAL	242 (151)	373 (139)	255 (167)	253 (164)	138 (82)

Table 3

 3 Chemical composition (mean percent of dry matter with standard error) of leaves of the four selected tree species. Values are means of three replicates

		Pisonia	Swietenia	Tabebuia	Bursera
		subcordata	macrophylla	heterophylla	simaruba
	Total nitrogen	2.5 (0.1)	1.1 (0.1)	0.9 (0.1)	1.1 (0.1)
	Cellulose	19.2 (0.6)	19.8 (4.1)	32.1 (2.3)	20.9 (0.4)
	Lignin	29.5 (15.1)	29.1 (10)	12.0 (2.1)	22.8 (9.6)
	Soluble phenols	6.7 (0.6)	12.7 (0.6)	4.5 (2.3)	10.0 (1.0)
	Tannins	1.8 (0.8)	2.9 (0.6)	0.3 (0.1)	3.3 (0.7)

Table 4

 4 Secondary forest. Influence of position on the slope and tree species on bulk types of material identified under the dissecting microscope. Mean volume of material type is indicated in percent of the total matrix with standard error between

	brackets
		Fragmented	Organo-mineral	Roots
		leaves	aggregates	8-10 cm
		4-6 cm	4-6 cm
	Lower slope (n = 2)	2.0 (0.3)	53.9 (3.8)	29.2 (9.0)
	Middle slope (n = 2)	1.8 (0.1)	33.3 (9.1)	22.9 (4.0)
	Upper slope (n = 2)	63.1 (0.1)	0.0 (0.04)	26.1 (3.6)
	Bursera simaruba (n = 3)	15.1 (0.4)	17.9 (7.1)	34.6 (5.5)
	Pisonia subcordata (n = 3)	5.9 (0.2)	23.3 (8.2)	11.0 (4.3)

Table 5

 5 Aggregate components identified under the phase contrast microscope.Comparisons between five types of aggregates with three replicates each. Mean volume of each component is indicated in percent with standard error between

	brackets
		Plant	Free amorphous	Amorphous organic	Mineral
		fragments	organic matter	matter bound to	material
				minerals
	Millipede organic
	faeces (secondary	93.4 (1.8)	5.3 (0.9)	0	0.1 (0.1)
	forest)
	Millipede organo-
	mineral faeces	16.4 (1.6) 69.2 (5.5)	10.7 (3.9)	3.5 (0.6)
	(secondary forest)
	Earthworm casts, (plantation)	62.3 (14.6) 14.9 (7.9)	16.0 (0.3)	11.0 (6.1)
	Organo-mineral
	aggregates	27.1 (11.9) 45.4 (20.3)	7.4 (3.0)	17.1 (3.4)
	(plantation)
	Organo-mineral
	aggregates	8.7 (1.3) 53.8 (4.0)	8.8 (1.9)	12.1 (3.4)
	(secondary forest)

Acknowledgements

We thank Professor Paul Leadley (Université d'Orsay, France) and the deputy editor J.M. Hodgson (Rothamsted, UK) for revising our English language, and Professor Jean-Paul Mauriès (Muséum National d'Histoire Naturelle, Paris) for identifying the millipedes at the species level.

Microscopic assessment of organic and organo-mineral aggregates

The millipede organic faeces contain more plant fragments than all other aggregates (Table 5). For example they contain six times more (93.4±1.8%) than the millipede organo-mineral faeces (16.4±1.6%). Organo-mineral earthworm casts in the plantation were richer in plant fragments than other (non-animal) organo-mineral aggregates. In particular they contain twice as many plant fragments (62.3±14.6%) as organo-mineral aggregates of non-animal origin (27.1±11.9%). Amorphous organic matter bound to minerals was more abundant in earthworm casts (16.0±0.3%) than in other organo-mineral aggregates (7.4±3.0% in the plantation, 8.8±1.9% in the secondary forest).

Discussion

In the secondary forest we found an epigeic macrofaunal community dominated by millipedes. These animals ingest fresh organic matter and deposit their faecal pellets at the soil surface. The endoanecic earthworm P. elongata was absent, probably because this species cannot tolerate dehydration and needs to aestivate when the soil dries out, which is impossible in the shallow Leptosol.

Millipedes are especially abundant under the canopy of P. subcordata (136 individuals.m -2 ), probably because the foliage of this tree species is rich in nitrogen (2.5%). As a result, millipedes produce abundant faecal pellets which accumulate to form a thick superficial OH horizon. This is possible only in the absence of endoanecic earthworms that would ingest and incorporate them into an A horizon.


Corinne Sarthou
email: sarthou@mnhn.fr

Jean-François Villiers

& Ponge

Shrub thicket vegetation on tropical granitic inselbergs (French Guiana)

Keywords: Correspondence Analysis, Granitic outcrops, Phytosociology, Shrub communities, Thickets, Vegetation dynamics

In French Guiana, inselbergs are granite outcrops rising abruptly from the surrounding rain forest. They constitute isolated islands of a special type of vegetation restricted to this peculiar substrate. Shrub granitic vegetation, organised in thickets on open exposed rocks of inselbergs, are described using the Braun-Blanquet method combined with numerical analysis (Correspondence Analysis). This phytosociological study revealed only one particular shrub community on each inselberg, including predominantly evergreen and sclerophyllous shrubs, especially microphanerophytes, belonging to Clusiaceae, Myrtaceae and Bombacaceae. These insular outcrop communities exhibit species endemic to the Guianas region and also species rare in French Guiana. Affinities with flora of other inselbergs and vegetation types in South America are examined and discussed. Reasons for observed floristic and structural changes in each community are also discussed.

Introduction

Inselbergs are isolated rocky outcrops consisting generally of Precambrian granite or gneiss. The geomorphology and the geology of inselbergs have been studied worldwide, a survey being provided by [START_REF] Bremer | Inselbergs: geomorphology and geoecology[END_REF]. These particular geomorphological formations constitute natural laboratories to address some prevailing questions concerning the past and future of biotic diversity. Inselbergs offer models for the quantitative analysis of species diversity and biotic communities in the frame of vegetation dynamics and island ecology. [START_REF] Porembski | Inselbergs. Biotic diversity of isolated rock outcrops in tropical and temperate regions[END_REF] gave a synthesis of research work done on inselberg vegetation throughout all continents. The study of plant communities on temperate inselbergs has a long tradition in North America and Australia. Such studies are lacking for South America and specially for Guianas where inselbergs need helicopter transport to be accessible. It is obvious that more extensive research needs to be done on Guianan inselbergs where only qualitative floristic data are available.

In South America, classical whale back and sugar loaf shaped inselbergs are scattered throughout Guyana and Brazilian Shields (up to East Bolivia). These outcrops rise abruptly from the surrounding plain landscape and represent singular habitats in tropical rain forests.

Thus, they reflect a clear habitat fragmentation and constitute functional terrestrial islands [START_REF] Prance | Islands in Amazonia[END_REF]. As a consequence, insular outcrop communities are of interest because they exhibit specific, and sometimes rare, species and community types. Some floristic studies have been carried out in French Guiana [START_REF] De Granville | Aperçu sur la végétation des inselbergs du sud-ouest de la Guyane française[END_REF][START_REF] De Granville | Recherches sur la flore et la végétation guyanaise[END_REF][START_REF] Sarthou | Dynamique de la végétation pionnière sur un inselberg en Guyane française[END_REF], in Venezuela [START_REF] Steyermark | Flora of the Venezuelan Guayana. VII Contributions of the Flora of the Cerro Aracamuni, Venezuela[END_REF][START_REF] Steyermark | Flora of the Venezuelan Guyana[END_REF], in Brazil [START_REF] Barthlott | Some remarks on the vegetation of tropical inselbergs: diversity and ecological differenciation[END_REF][START_REF] Porembski | Diversity and ecology of saxicolous vegetation mats on inselbergs in the Brazilian Atlantic rainforest[END_REF]) and in Bolivia [START_REF] Ibisch | Floristic, biogeographical, and vegetational aspects of Pre-Cambrian rock outcrops (inselbergs) in eastern Bolivia[END_REF]. Due to poorly developed soils and extreme microclimate variations, outcrop vegetation is characterised by low diversity and oligotrophic or, in some instances, xerophytic characters which contrast abruptly with surrounding rain forest vegetation. A few semi-quantitative studies described typical herbaceous communities such as monocotyledonous mats made up of Bromeliaceae and Velloziaceae, swamp vegetation and plant communities living seasonally in rock pools [START_REF] Sarthou | Epilithic plant communities on inselbergs in French Guiana[END_REF][START_REF] Meirelles | The vegetation of granite rock outcrops in Rio de Janeiro, Brazil, and the need for its protection[END_REF]. Woody patches seem to be characteristic of Guyana shield inselbergs but they received less attention. In French Guiana, they constitute numerous shrub islands mainly composed of Clusia spp. on granitic outcrops [START_REF] De Granville | Aperçu sur la végétation des inselbergs du sud-ouest de la Guyane française[END_REF][START_REF] De Granville | Recherches sur la flore et la végétation guyanaise[END_REF]. But despite their distinctiveness and relative simplicity of species composition, no quantitative data exist on shrub communities from Guianas inselbergs. This paper is the first phytosociological study of shrubby thickets on French Guianan inselbergs. Phytosociological data could also provide avenues for studies on vegetation dynamics occurring within each of the three investigated plant communities.

Methods

Study area

In French Guiana, inselbergs, 100-800 m high, consist of Precambrian granites.

Detailed accounts about the geomorphology and geology of rock outcrops in the study area were provided by [START_REF] Hurault | Recherches sur les inselbergs granitiques nus en Guyane française[END_REF], [START_REF] Gruau | Rb-Sr and Sm-Nd geochronology of lower proterozoic granite-green stone terrains in French Guiana, South America[END_REF] and [START_REF] Teixeira | A review of the geochronology of the Amazonian craton: tectonic implications[END_REF]. These granitic outcrops are unique because they are located in the undisturbed rain forest while most inselbergs known in Africa, Brazil and Venezuela are situated within landscapes dominated by human activities. The open areas of exposed rocks support numerous scattered herbaceous and shrubby patches separated by bare rock, forming a mosaic called locally "savane-roche".

Data regarding inselberg flora are available in the Aublet data base [START_REF] Hoff | La banque de données "AUBLET" de l'Herbier du Centre ORSTOM de Cayenne (CAY)[END_REF] but are still fragmentary. The first vegetation studies were based on qualitative field observations made in SW French Guiana [START_REF] De Granville | Aperçu sur la végétation des inselbergs du sud-ouest de la Guyane française[END_REF][START_REF] De Granville | Recherches sur la flore et la végétation guyanaise[END_REF].

Recently, six herbaceous and suffrutescent associations and three sub-associations have been described on the base of semi-quantitative data [START_REF] Sarthou | Epilithic plant communities on inselbergs in French Guiana[END_REF].

The field work was undertaken during the rainy season in the course of three botanical expeditions on Mont-Chauve (3° 49' N, 52° 44' W, alt. 265 m) in April 1997, Trinité (4° 35' N, 53° 21' W, alt. 400 m) in May 1998 and Nouragues (4° 3' N, 52° 42' W, alt. 410 m) in April 1999, respectively (Fig. 1). The annual average rainfall amounts to 3000-3250 mm at Mont Chauve and Nouragues and 2000-2250 mm at Trinité [START_REF] Boyé | Climatologie: Atlas des départements français d'Outre-Mer -IV-La Guyane[END_REF]. The studied inselbergs are dome-shaped (whale-back type). They belong to the "cyanobacteria type" i.e. the exposed rock is covered by films of cyanobacteria [START_REF] Sarthou | Cyanophycées de l'inselberg des Nouragues (Guyane française)[END_REF], in contrast to the "lichen type" which is characteristic for drier conditions [START_REF] Büdel | Cyanobacteria and cyanobacterial lichens from inselbergs of the Ivory Coast, Africa[END_REF]. Cyanobacteria films give a red-brownish to blackish-green colour to the inselberg except on areas with intense water flow.

A detailed investigation on the Nouragues inselberg showed that most extreme environmental conditions prevailed on these rocky outcrops, due to very high daily fluctuations in temperature and relative humidity [START_REF] Sarthou | Dynamique de la végétation pionnière sur un inselberg en Guyane française[END_REF]. The mean temperature ranges between 18-55°C and the mean humidity between 20-100%. The open exposed rock is submitted to intense sunlight with high evaporation rate and rapid runoff following precipitation. The temperature of the bare rock surface reaches 50°C (up to 75°C in dry season). Cyanobacteria constitute the primary factor in the establishment of nutrient cycles on the rock surface [START_REF] Sarthou | Mécanismes de colonisation par la végétation d'un inselberg granitique en Guyane française[END_REF]. Soils are characterised by shallowness (1-20 cm for herbaceous patches, 10-40 cm for shrubby patches), marked acidity (pH from 3.5 to 5.5), coarse material (quartz sand) and low nutrient content [START_REF] Sarthou | Mécanismes de colonisation par la végétation d'un inselberg granitique en Guyane française[END_REF].

Plots

A total of 151 phytosociological relevés were made on the three inselbergs, considering the whole set of each outcrop, in floristically and structurally homogeneous thicket stands. Forty four relevés (1-44) were sampled on Mont Chauve inselberg, 50 on Trinité and 57 (95-151) on Nouragues. Relevés were done on plots which were distributed on the whole surface of each inselberg, thus they could be considered as representing the whole population. Plot sizes were small, in accordance with the size of vegetation patches, and distances from a plot to another were also variable, folllowing vegetation distribution patterns. The area was fixed to 2 x 4 m (8m²) on Mont Chauve, to 2 x 7 m (14 m²) on Trinité and to 2 x 8 m (16 m²) on Nouragues. For each relevé, cover abundance data for all terrestrial pteridophytes and spermaphytes were recorded according to Braun-Blanquet approach [START_REF] Braun-Blanquet | Pflanzensoziologie[END_REF]Westhoff & van der Maarel 1973). Specimens from all plant species were collected then identified by the authors using collections from two herbaria (Cayenne and Paris). Voucher specimens were deposited at the Herbarium of French Guiana (CAY) in Cayenne and at the National Museum of Natural History in Paris (P).

A synoptic table of constancy classes (Braun-Blanquet 1979) including the three shrub communities was elaborated (Table 1). Original data, including abundance-dominance of plant species in the 151 relevés, together with slope and aspect, can be provided upon request to the corresponding author.

Analysis

For numerical analysis, cover-abundance values (according to Braun-Blanquet scale) were transformed into the 1-9 ordinal scale of [START_REF] Van Der Maarel | Transformation of cover-abundance values in Phytosociology and its effects on community similarity[END_REF]. Plots were ordinated according to their floristic composition using Correspondence Analysis, a multivariate method using the chi-square distance [START_REF] Greenacre | Theory and applications of correspondence analysis[END_REF]. Simple correspondence analysis was used in place of detrended correspondence analysis [START_REF] Hill | Detrended correspondence analysis: an improved ordination technique[END_REF] because not any horseshoe effect was expected to occur given our data set embracing a variety of environmental and phytogeographical conditions. This analysis was performed using the STATBOX program on a matrix crossing plant species present in at least three plots (66 taxa) with the 151 relevés. Species characterised by cover-abundance values were used as active variables (contributing to factorial axes). Passive variables, describing environmental conditions such as orientation and slope, and structural characteristics such as the total number of species, the total number of epiphytes and the number of different life forms per relevé were added to measure their degree of relationship with the factorial axes. Active and passive data were reweighted and focused (mean fixed to 20 and standard deviation fixed to 1) and each structural variable was associated with a conjugate, varying in an opposite sense (complement to 40), according to a method used by [START_REF] Peltier | Establishment of Fagus sylvatica and Fraxinus excelsior in an old-growth beech forest[END_REF] for the study of plant communities. This allows factorial coordinates of variables to be proportional to their contribution to the axes, as in principal components analysis [START_REF] Hotelling | Analysis of complex of statistical variables into principal components[END_REF]) and gradients of global vegetation abundance to be depicted [START_REF] Greenacre | Theory and applications of correspondence analysis[END_REF]. Thus, each species will be represented by two points, indicating higher and lower values, respectively.

Results

Flora

A total of 94 plant species were recorded in the three shrub communities (Appendix 1). The species belong to 78 genera distributed into 40 families. Dominant families are Myrtaceae (9.6% of total number of species), Bromeliaceae (8.5%) and Orchidaceae (8.5%).

Other widely dispersed tropical families were also encountered, such as Rubiaceae (6.4%), Melastomataceae (4.3%), Clusiaceae (4.3%), Euphorbiaceae (4.3%) and Mimosaceae (4.3%).

Well represented genera are Myrcia, Clusia and Croton followed by Calyptranthes, Psychotria, Erythroxylum, Encyclia, Aechmea, Vriesea and Sauvagesia. Woody forms (63%) are mainly constituted by shrubs (31%), small trees (17%), climbers (10%) and lianescent sub-shrub parasites (5%) (Appendix 1). Herbaceous forms (37%) are mainly represented by erect herbs, rosettes or tussocks and by one palm species restricted to granitic outcrops in Guianas. Among life-forms, phanerophytes (49%), especially microphanerophytes, clearly dominated the communities, followed by hemicryptophytes (28%), lianas (10%), chamaephytes (7%) and parasites (5%) (Appendix 1). Geophytes were represented by only one species.

Among a total of 94 plant species, 35% were endemic to Guianas (Appendix 1). These Our list of 94 species includes plants also found in other habitats such as the rain forest (terrestrial or epiphyte vegetation), rocks on river sides or in the rain forest, savannas, secondary forest, white sand forest. Among inventoried species, 20% are confined to granitic inselbergs in French Guiana (Appendix 1).

Ordination

In the plane of axes 1 and 2, CA differentiates three plant communities corresponding to the three studied sites (Fig. 2). Despite the common occurrence of some taxa with a high constancy, which were generally placed not far from the origin (Table 1, Fig. 2: species with border), each community exhibits a particular floristic group described below. Given the clear delimitation of characteristic groups, no numerical classification was judged necessary to separate them. The floristic composition of the three groups is outlined in Table 1.

All shrub communities present on exposed rock surfaces are evergreen and form individualised dense thickets, 3-5 m tall, generally separated by areas of bare rock or by herbaceous communities. Occurring both on slopes (0-63%) and in depressions, they represent the main functional woody unit of the "savane-roche" ecosystem. These communities are physiognomically dominated by Clusiaceae and Myrtaceae species. Species richness averages 12, 13, and 14 species per plot on Trinité, Mont Chauve and Nouragues respectively, which was considered similar given small differences in the size of sampling areas (varying from 8 to 16 m 2 ). Three layers can be recognised in each of these vegetation units.

A dominant canopy stratum 3-5 m in height with rare emergents reaching 6-8 m in height is characterised by microphanerophytes (70-100% cover). Shrubs with a high constancy in all communities (Table 1), such as Clusia nemorosa, Myrcia saxatilis, Psychotria hoffmannseggiana, Miconia ciliata and Erythroxylum citrifolium, are also important structural components of the thickets. Other life forms associated with the crown space of thickets, such as lianas, parasites or epiphytes occur but with a small number of individuals. The dominant layer is complemented by a smaller stratum 0.5-1.5 m in height, typified by nanophanerophytes (5-40% cover). A herbaceous layer (50-100% cover) is often composed by dense colonies of spiny bromeliads, Cyperaceae or graminoids, small populations of orchids and species occasionally found growing in shady sites. Two species, Pitcairnia geyskesii and Scleria cyperina, occur with a high constancy and sometimes a high cover rate on the three inselbergs (Table 1).

Although above mentioned physiognomic features can be found in all investigated inselbergs, a particular community has been recognised on each inselberg, made of distinct species as well as species differing in their constancy among the three inselbergs.

The Quapoya scandens-Rhodognaphalopsis flaviflora community (Table 1, Fig. 2) occurs on Trinité inselberg. This shrub unit is characterised by the Guianan endemics Quapoya scandens and Rhodognaphalopsis flaviflora, two shrubs rare in French Guiana and restricted to rocky outcrops. The upper layer includes Eugenia macrocalyx, a tree also found in the adjacent forest. The tall shrub layer is composed of the nanophanerophyte Turnera rupestris var. frutescens, endemic to the Guianas region. The Cyperaceae Mapania effusa and the epilithic orchid Encyclia granitica predominate in the herb layer.

The Clusia minor community (Table 1, Fig. 2), occurring on the Nouragues inselberg, is mainly characterised by this Clusiaceae shrub species. Two Myrtaceae, Myrcia guianensis and M. fallax, endemics to the Guianas, were also found in the shrub layer. The lower layer is physiognomically dominated by populations of the Cyclanthaceae, Stelestylis surinamensis, 1-2m height, rare in French Guiana. The perennial grass, Axonopus ramosus, another Guianan endemic species, is well represented in some plots. Anthurium jenmanii and Epidendrum nocturnum, also found on the Trinité inselberg, are more constant in this community (Table 1).

The Calyptranthes lepida-Guapira sp. community (Table 1, Fig. 2) is found on Mont Chauve inselberg. This unit is characterised by Calyptranthes lepida, Guapira sp. and the palm Syagrus stratincola, only found on granite outcrops in the Guianas. Lianas such as Dioscorea pubescens and Dioclea virgata are also specific to the dominant layer. The orchid Sobralia stenophylla grows in small populations in the herb layer. The woody species Clusia nemorosa, Eriotheca surinamensis and Tabebuia capitata, also found in other communities, are important structural components of this community, here more constant and with higher cover-abundance values than in other communities (Table 1). The orchid Encyclia ionosma and the terrestrial bromeliad Guzmannia lingulata are also better represented on this outcrop (Table 1).

Other marked differences in the floristic composition are reflected along axis 3 of CA, especially within the Trinité shrub unit (Fig. 3), and to a lesser extent in the Nouragues inselberg. Axis 3 could indicate a successional gradient, as suggested by the ordination of life-forms along this axis, and the fact that environmental variables such as slope and exposure contributed poorly to this axis (near to the origin, not represented in Fig. 3).

Hemicryptophytes and nanophytes were projected mainly on the positive side of Axis 3, while microphanerophytes, lianas and epiphytes were projected on its negative side, suggesting a gradient in vegetation height and biodiversity, increasing from positive to negative values of Axis 3.

Discussion

The shrub vegetation of the inselbergs clearly shows a typical flora with many endemics to the Guianas region in each community. These insular outcrop communities are of interest because they exhibit specific and also rare species, due to isolation (McArthur & Wilson 1967) and particular environmental conditions. The low capacity of long-distance dispersal of most outcrop species (most species being dispersed by territorial birds and ants, personal observations) explain probably the differences in vegtation composition from an inselberg to another.

Ecological constraints, especially geology and temperature, are evident for species living in the studied rocky habitats where most extreme environmental conditions prevail [START_REF] Sarthou | Dynamique de la végétation pionnière sur un inselberg en Guyane française[END_REF]. In contrast, in the Amazonian Basin, mountains reaching 100-200 m are scarce and humid conditions are predominant. We presume that many of the inventoried species are specifically adapted to the particular combination of high temperatures, intense runoff and chemically poor substrates which prevails on tropical granitic outcrops. Many of these species have been also found in open areas such as savannas, caatingas and also forests on white sand soils (Appendix 1).

No classification of French Guianan outcrop shrub vegetation exists owing to difficult access to inselbergs, and previous data were exclusively based on qualitative field observations. The present study constitutes the first semi-quantitative approach to the inselberg shrub flora of French Guiana, but further phytosociological analyses are required before a syntaxonomical outline can be drawn. Some authors have qualitatively described Clusiaceae thickets on Tumuc Humac inselbergs in SW French Guiana, a dryer region (< 2000 mm/year) at 450-800 m altitude (de [START_REF] De Granville | Aperçu sur la végétation des inselbergs du sud-ouest de la Guyane française[END_REF][START_REF] De Granville | Recherches sur la flore et la végétation guyanaise[END_REF]. As in our study, their results seem to confirm that, despite some common taxa, a specific floristic group can be recognised on each inselberg.

Similarly organised shrub islands have been qualitatively described on inselbergs in Venezuela [START_REF] Steyermark | Flora of the Venezuelan Guyana[END_REF], Bolivia [START_REF] Ibisch | Floristic, biogeographical, and vegetational aspects of Pre-Cambrian rock outcrops (inselbergs) in eastern Bolivia[END_REF] and Africa [START_REF] Bonardi | Contribution à l'étude botanique des inselbergs de Côte d'Ivoire forestière[END_REF][START_REF] Jenik | Plant communities of the Accra Plains, Ghana[END_REF]. Thus, affinities with flora of saxicolous shrub communities of Venezuelan inselbergs (or lajas) was especially observed at the family and also at the genus level [START_REF] Steyermark | Flora of the Venezuelan Guayana. VII Contributions of the Flora of the Cerro Aracamuni, Venezuela[END_REF][START_REF] Steyermark | Flora of the Venezuelan Guyana[END_REF]. Indeed, the evergreen shrub islands on lajas harbour woody species mostly belonging to Melastomataceae, Clusiaceae, Bignoniaceae, Bombacaceae, Erythroxylaceae, Euphorbiaceae, Ochnaceae, Rubiaceae and Theaceae. Among the most important colonising herbs are also several species of Bromeliaceae and terrestrial Orchidaceae. Some genera such as Tabebuia, Erythroxylum, Clusia, Ternstroemia are also found on lajas. The occurrence of the gregarious palm Syagrus orinocensis in the understory of lajas low forest can also be compared to that of Syagrus stratincola in Trinité thickets. However, the flora of Venezuelan lowland granitic outcrops generally exhibits a higher level of endemism than that of French Guianan outcrops. This could be explained by higher elevation (>1000 m), a higher habitat diversity, or by hypothetical historical features, a more ancient isolation allowing more speciation.

Similarities are also noticed with the vegetation of Guianas savannas both floristically (at a genus and family level) and physiognomically. We can compare the studied communities with the sclerophyllous thickets described by [START_REF] Hoock | Les savanes guyanaises: Kourou. Essai de phytoécologie numérique[END_REF] in French Guianan coastal savannas. However, the life-form spectrum of outcrop shrub communities, with the predominance of microphanerophytes and hemicryptophytes, is more similar to the peripheral forest group of coastal savannas. Outcrop communities also have affinities with some savanna-bushes described by van Donselaar (1965) and [START_REF] Teunissen | Vegetation and flora of the savannas in the Brickheuvel Nature Reserve, Northern Suriname[END_REF] in Surinam such as Clusia-Scleria bushes on wet white sand and Marlierea bushes on wet loamy sand.

Affinities may be also pointed out at the family and genus levels with other vegetation types such as low forests in French Guiana [START_REF] Larpin | The low forest (Nouragues Inselberg)[END_REF], shrublands of the Venezuelan Guyana [START_REF] Huber | Shrublands of the Venezuelan Guyana[END_REF], scrub caatingas and campos rupestres in Brazil (Pires-O'Brien 1992; [START_REF] Alves | Plant species endemism in savanna vegetation on table mountains (Campo Rupestre) in Brazil[END_REF][START_REF] Da Silva | Aspectos ecologicos da vegetaçao rupestre da serra dos carajas, estado do Para, Brasil[END_REF] and white sand and rocky substrates at the top of the Araracuara sandstone plateau in Colombian Amazonia [START_REF] Duivenvoorden | Amazonian savanna vegetation on the sand stone plateau near Araracuara, Colombia[END_REF].

Parallels in the floristic composition and structure of these vegetation types seem to be determined by particular edaphic factors. A combination of extremely poor and acid (pH<4) soil conditions and the periodic desiccation of the shallow soil profile characterises thicket vegetation on French Guianan inselbergs [START_REF] Sarthou | Mécanismes de colonisation par la végétation d'un inselberg granitique en Guyane française[END_REF]. Thus, periodic water stress may explain the xerophytic character of some components of this vegetation. A similar periodic water deficit has been also reported in other shrub communities described by [START_REF] Duivenvoorden | Amazonian savanna vegetation on the sand stone plateau near Araracuara, Colombia[END_REF], [START_REF] Steyermark | Flora of the Venezuelan Guyana[END_REF] and [START_REF] Larpin | The low forest (Nouragues Inselberg)[END_REF].

Floristic differences observed between the three studied inselbergs (Axes 1 and 2 of CA) could be related to local historical features, depending on initial outcrop colonisation. We can suppose that the neighbouring Nouragues and Mont Chauve inselbergs (projected on the positive side of Axis 1, separated by Axis 2) have a more common history of land colonisation than Trinité (projected on the negative side of Axis 1), located further north.

The presence of vegetation gradients was apparent within each shrub community, more especially at the Trinité inselberg (Axis 3 of CA). These gradients are probably associated with thicket ageing (plant succession), since orientation and exposure did not explain the variation displayed by Axis 3. But we cannot discard other factors not acounted for in our study, such as differences in edaphic constraints and runoff, for instance.

The present study pointed on the isolation of several plant communities the floristic composition of which could vary according to geographical distance. A thorough investigation of a wide range of French Guianan inselbergs will help to better discern compositional trends (gradients) at the regional scale, in order to test the hypothesis of a past development of the savane-roche in place of the present rain forest. respectively. Associated with correspondence analysis of passive variables: life-forms [START_REF] Raunkier | The life-forms of plants and statistical plant geography[END_REF], number of epiphytes and total number of species per relevé. Bold or italic means higher or lower values, respectively. Abbreviations: Ch: Chamaephytes; Hc: Hemicryptophytes; Na: Nanophanerophytes; Mi: Microphanerophytes; L: Lianas; TotS: total number of species per relevé; NbEp: number of epiphytes per relevé.

Table 1. Floristic composition of the shrub communities. Constancy-classes: I: 1-20%; II: 21-40%; III: 41-60%; IV: 61-80%; V: 81-100%. Species reaching only constancy class I in all the communities were omitted. Arabic numbers following indicate the range of coverabundance values. *: without epiphytes. Species with border are species in common to all three groups (see also Figure 2). Species in bold are endemic species to the Guianas region.

  include many species characteristic and abundant in shrub inselberg communities such as Calyptranthes lepida, Syagrus stratincola, Quapoya scandens, Rhodognaphalopsis flaviflora, Stelestylis surinamensis, Myrcia guianensis, Myrcia fallax, Myrcia saxatilis and Pitcairnia geyskesii. These insular outcrop communities are also of interest because they exhibit species rare in French Guiana such as Quapoya scandens, Rhogdophanalopsis flaviflora, Eriotheca surinamensis, Syagrus stratincola, Stelestylis surinamensis, Erythroxylum ligustrinum, Croton argyrophylloides, C. hostmannii, Zygia tetragona, Eugenia macrocalyx and Ternstroemia dentata.

  van Donselaar, J. 1965. An ecological and phytogeographic study of northern Surinam savannas. Wentia 14: 1-163.Westhoff, V. & van der Maarel, E. 1973. The Braun-Blanquet approach. In: Whittaker, R.H. (ed.) Ordination and classification of plant communities, pp. 617-737. Junk, The Hague.

Figure 1 .

 1 Figure 1. Location of the inselbergs studied in French Guiana: Mont-Chauve (MC), Nouragues (N), and Trinité (T).

Figure 2 .

 2 Figure 2. Correspondence analysis of the 151 relevés (sharing 66 species) combined with vegetation data on the three inselbergs: projection in the plane of the two first axes with 16.1 % and 12.3 % inertia, respectively. Trinité: Quapoya scandens-Rhodogphanalopsis flaviflora community (). Mont Chauve: Calyptranthes lepida-Guapira sp. Community (). Nouragues: Clusia minor community (). Species with border are common to all three groups (see also Table1). Species in bold are endemic species to the Guianas.

Figure 3 .

 3 Figure 3. Correspondence analysis of the 151 relevés (sharing 66 species) and associated sites: projection in the plane of the axis 2 and 3 with 12.3 % and 4.6 % inertia,

  Fig. 1

  Table 1). Species in bold are endemic species to the Guianas. Abbreviations of taxa names: AXO: Axonopus ramosus; CAL: Calyptranthes lepida; CLM: Clusia minor; CLN: Clusia nemorosa; DIO: Dioscorea pubescens; DIV: Dioclea aff. virgata; ENG: Encyclia granitica; ERI: Eriotheca surinamensis; ERN: Ernestia

granvillei; ERY: Erythroxylum citrifolium; EUG: Eugenia macrocalyx; GUA: Guapira sp.; MAP: Mapania effusa; MAT: Matelea sastrei; MIC: Miconia ciliata; MYF: Myrcia fallax; MYG: Myrcia guianensis; MYS: Myrcia saxatilis; PIT: Pitcairnia geyskesii; PPT: Piptocoma schomburgkii; PSY: Psychotria hoffmannseggiana; QUA: Quapoya scandens; RHO: Rhodognaphalopsis flaviflora; SAT: Satyria cerander; SCL: Scleria cyperina; SOB: Sobralia stenophylla; STE: Stelestylis surinamensis; SYA: Syagrus stratincola; TAB: Tabebuia capitata; TUR: Turnera rupestris var. frutescens.

Acknowledgements. The studies on Mont Chauve and Trinité inselbergs were supported by a grant from the French Ministry of Environment (SOFT Research Program) and from the National Office of Forests, respectively. Investigations on the Nouragues inselberg were conducted at the Nouragues Field Research Station (UPS-CNRS 656). The authors thank Philippe Gaucher for field assistance on Trinité and Georges Cremers for helpful suggestions to improve the manuscript.


Nassima Sadaka

Jean-François F Ponge
email: jean-francois.ponge@wanadoo.fr

FUNGAL COLONIZATION OF PHYLLOSPHERE AND LITTER OF QUERCUS ROTUNDIFOLIA LAM. IN A HOLM OAK FOREST (HIGH ATLAS, MOROCCO)

Keywords: Decomposition, Fungal succession, Isolation methods, Biodiversity

The microfungal flora of holm oak living, senesced and litter leaves was studied at five different stages of decomposition using three different isolation methods.

Holm oak leaves are first colonized on the tree by a variety of primary saprophytes such as Trichothecium, Aureobasidium, Cladosporium, Epicoccum and Alternaria. After leaf fall there is an intensive development of the fungal flora, including both species already present in the phyllosphere and new colonizers from the litter layer. With increasing decomposition initial colonizers gradually disappear, being replaced by other forms. When all isolation methods were pooled, maximum biodiversity (species richness) of the fungus flora was observed during the first three stages of leaf litter decomposition, but strong variation occurred according to the isolation method. Sterilization of the leaf material revealed that a number of fungal strains were present at the inside of holm oak leaves before abscission, increasing from living to senescent stage, and that a strong decrease in the internal colonization of leaf litter was observed at late decomposition stages.

Introduction:

Fungi play fundamental roles in leaf litter decomposition processes within forest ecosystems [START_REF] Swift | Decomposition in terrestrial ecosystems[END_REF][START_REF] Cooke | Ecology of saprotrophic fungi[END_REF]. Fungal species distribution and successional changes occurring during the decomposition process have been extensively investigated on several litter types and using different isolation methods [START_REF] Kendrick | Biological aspects of the decay of Pinus sylvestris leaf litter[END_REF][START_REF] Struwe | Changes in population structure during decomposition[END_REF][START_REF] Rosenbrock | Fungal succession and changes in the fungal degradation potential during the initial stage of litter decomposition in a black alder forest [Alnus glutinosa (L.) Gaertn.][END_REF]. Mechanisms underlying fungal successions as well as advantages and biases of the different methods have been reviewed and discussed by Frankland (1992Frankland ( , 1998)).

The decomposition of tree leaves is not entirely confined to the litter layer on the forest floor [START_REF] Ruinen | The phyllosphere. I. An ecologically neglected milieu[END_REF]Davenport 1976). In fact, decay processes are initiated as soon as the leaf is formed, and the large surface of this plant organ is exposed to microbial and faunal attack during its entire life, senescence and death. Relatively few studies have been designed to follow the colonization of leaves from their initial expansion to their incorporation into humus [START_REF] Ruscoe | Mycoflora of living and dead leaves of Nothofagus truncata[END_REF]Wildman and Parkinson 1979;[START_REF] Mishra | Phylloplane and litter fungi of Ilex aquifolium[END_REF].

Less is known about long-living evergreens leaves, though [START_REF] Ruinen | The phyllosphere. I. An ecologically neglected milieu[END_REF] demonstrated that persistent leaves of tropical plants supported complex and extensive microbial populations. The paucity of information concerning qualitative aspects of fungi inhabiting holm oak leaves in Mediterranean areas [START_REF] Vardavakis | The mycoflora, production and decomposition of leaf litter in an Orno-Quercetum ilicis association[END_REF], and particularly in Morocco, prompted the investigation reported here. The present study examined the fungal colonization of holm oak leaves from phyllosphere (living and senescent leaves) to leaf litter at five different stages of decomposition.

MATERIAL AND METHODS

Study Material

Holm oak (Quercus rotundifolia Lam.) forms typical climax forests that are widely distributed throughout the western Mediterranean area (Maghreb, Central Spain), mostly in mountains where it tolerates a dry and cold climate [START_REF] Achhal | A propos de la valeur bioclimatique et dynamique de quelques essences forestières au Maroc[END_REF][START_REF] Arx | The genera of fungi sporulating in pure culture[END_REF]. It is characterized by persistent, spiny, sclerophyllous foliage. Litter falls throughout the year, mainly from April to June [START_REF] Lossaint | La forêt méditerranéenne de chênes verts[END_REF]. The thickness of litter is determined by seasonal litter fall, decomposition rate, and biennial cycles of large and small amounts of litter input [START_REF] Rapp | Cycle de la matière organique et des éléments minéraux dans quelques écosystèmes méditerranéens[END_REF][START_REF] Poli | Produzione di littiera nella lecceta del M. Minardo (Etna) nel periodo Settembre 1970-Giugnio 1974[END_REF][START_REF] Sadaka-Laulan | Comparative leaf decomposition within the holm oak complex[END_REF].

Study area

The investigation was performed in a holm oak forest dominated by Quercus rotundifolia Lam. (3-7 m height, 85-95 % cover), located at Toufliht on the northern slope of the High Atlas, Morocco. The climate is of the sub-humid to semi-arid Mediterranean type, with most precipitation from October to February and a long warm dry period from May to September (mean annual rainfall 840 mm; maximum summer temperature 30.5°C and minimum winter temperature 1°C).

Sampling method

Samples were collected in May 1999, at the time of optimum litter-fall. Leaves were collected directly on the phyllosphere and in several litter horizons from ten trees growing not far from each other within a one are area. Still attached leaves were separated into living (green) and senescent (yellow) leaves, irrespective of their age. The mean duration of life of holm oak leaves has been estimated to two years, but leaf fall may occur during the first as well as during the third year of life [START_REF] Rapp | Production de litière et apport au sol d'éléments minéraux dans deux écosystèmes méditerranéens: la forêt de Quercus ilex L. et la guarrigue de Quercus coccifera L[END_REF]. Leaves from the litter layer were divided into five stages of decomposition (Sadaka-Laulan and [START_REF] Sadaka-Laulan | Comparative leaf decomposition within the holm oak complex[END_REF] according to morphological criteria such as colour, thickness and hardness (Table 1).

After pooling and throroughly mixing the samples per category, forty-eight leaves were randomly selected within each of the seven categories of phyllosphere and litter leaves.

Fungal isolation

A combination of isolation methods was applied in order to obtain more information about the composition of leaf fungal flora and to differentiate epiphytic from endophytic flora.

One leaf disk was punched from each collected leaf with a sterile cork borer (6mm in diameter), avoiding primary veins. Leaf disks obtained were subdivided into three parts, corresponding to the three techniques used: washing, surface-sterilization, moist chambers.

*Washed leaves:

This method removes all easily detachable fungal propagules on the surface and isolates only mycelia growing actively on and in the leaf [START_REF] Kendrick | Biological aspects of the decay of Pinus sylvestris leaf litter[END_REF]Macauley and Thrower 1966;[START_REF] Osono | Decomposing ability of interior and surface fungal colonizers of beech leaves with reference to lignin decomposition[END_REF]. Sixteen disks of each leaf type were serially washed in a sterile tube by mechanical shaking. First, the disks were washed during 5 min in two changes of sterile distilled water containing one drop of Tween 80, a wetting agent, then rinsed eight times with sterile distilled water (2 min per washing). After blotting them on sterile filter paper, leaf disks were plated out on 2% Malt Extract Agar (MEA) added with Chloramphenicol, an antibacterial agent. This medium isolates a large spectrum of fungal strains and also allows rapid fungal growth (Wildman and Parkinson 1979).

For each leaf type, four Petri dishes were plated with four disks each. Since each disk was taken from a single leaf, we can consider that we have 16 replicates in total, taking into account that variation between individual leaves was more important than variation between Petri dishes.

After 48 h of incubation at 23°C in darkness, the plates were observed daily during 10 days, until no new colonies appeared. As soon as they were observed, fungal colonies were transferred to fresh MEA plates for isolation. The identification was done at the genus level using [START_REF] Barron | The genera of hyphomycetes from soil[END_REF], [START_REF] Arx | The genera of fungi sporulating in pure culture[END_REF]Carmichael et al. (1980), then at the species level using many specialized monographs. *Surface sterilized leaves: designed to isolate internal fungi (Wildman and Parkinson 1979;Espinosa-Garcia and Langenheim 1990). The leaf disks were surface sterilized for 60 sec by immersion in 1% sterile AgNO 3 , followed by two 2-min washings in 1% NaCl and five 2-min washings in sterile distilled water. Concerning stages IV and V, owing to their advanced decomposition, the duration of immersion in AgNO 3 has been reduced to 30 sec. The disks were then treated as for washed leaves. Cultures were incubated for 2 weeks and examined regularly after the second or third day. *Moist chambers incubation: used in an attempt to induce sporulation of fungi which might have been unable to compete with faster growers on nutrient media. Leaf disks were washed as for the first method and subsequently kept in sterile moist chambers [START_REF] Keyworth | A Petri dish moist chamber[END_REF] for 30 days. This long period enables fungi originally present as propagules to sporulate [START_REF] Lindsey | A survey of methods in the study of microfungal succession on leaf surfaces[END_REF][START_REF] Gourbière | Structure spatio-temporelle de la mycoflore des premiers stades de décomposition des aiguilles d'Abies alba[END_REF]).

The results were expressed in terms of numbers of isolates for each species from the 16 leaf disks treated by each method. Species were then ranked according to, first, their preferential stage and, second, their coefficient of variation (%). The preferential stage was determined using the following formula: (1.X 1 +2.X 2 +3.X 3 +4.X 4 +5.X 5 +6.X 6 +7.X 7 ) / (X 1 +X 2 +X 3 +X 4 +X 5 +X 6 +X 7 ) where numbers 1 to 7 correspond to the rank attributed to the leaf stage, 1 for living, 2 for senescent, until 7 for stage V. X 1 to X 7 correspond to the total number of isolates for each species at each stage using the three isolation methods. It should be noted that this calculation method allowed to determine the mean stage at which a given fungal strain could be found, not necessarily its preferential stage, which should be the mode of the distribution.

Data analysis

Frequency data (numbers of isolates of each fungal species at each leaf stage using each of the three isolation methods) were subjected to correspondence analysis, a multivariate method using the chi-square distance between individuals and between variables in a symmetrical manner [START_REF] Greenacre | Theory and applications of correspondence analysis[END_REF]. The different fungal species were the active variables. Leaf stages (living leaves, senescent leaves, stages I, II, III, IV and V), and isolation methods (washing, surface sterilization and moist chambers) were treated as passive variables, i.e. they were projected on the factorial axes as if they had been involved in the analysis, without contributing to the factorial axes. They were coded as 1 or 0. In order to give the same weight to all variables (active and passive) they were transformed to mean 20 and unit variance by X = (x-m)/s + 20, where x is the original value, m is the mean of a given variable and s is its standard deviation [START_REF] Ponge | The heterogeneity of humus profiles and earthworm communities in a virgin beech forest[END_REF]. The addition to each standardized variable of a constant 20 made all values positive, because correspondence analysis deals only with positive numbers, commonly counts [START_REF] Greenacre | Theory and applications of correspondence analysis[END_REF]. Following this transformation, factorial coordinates of variables can be interpreted directly in terms of their contribution to the factorial axes. The further a variable is projected from the origin of the axes (barycentre) along a factorial axis, the more it contributes to this axis.

RESULTS

Table 2 lists the 36 fungal species and their frequencies recorded by stage and isolation method. Results revealed successional changes in the fungal flora of phyllosphere and litter. Figure 1 shows an increase in species richness from living to freshly fallen leaves (stage I), followed by a decrease until stage V, the most decomposed litter, just before incorporation into humus. The mean number of fungal colonies (total numbers for 16 disks, averaged among the three isolation methods) shows the same bell-shaped curve, but with a slight decrease from green to moribund leaves while the maximum extended to the first three stages of decomposition.

The phyllosphere microflora consisted of relatively few species (13 among 36).

Trichothecium roseum and Ulocladium chartarum exhibited a high levels of detection on green leaves. Aureobasidium pullulans, Cladosporium cladosporioides, Alternaria alternata and Epicoccum purpurascens were found on living as well as on senecent leaves, with maximal frequencies of isolation on senescent leaves. Species from the latter group were also recorded in most litter stages, except the final one (stage V). The appearance of Acremonium sp, Fusarium oxysporum and white sterile mycelium coincided with senescence.

Twenty-three species were recorded only on litter. There was a progressive change in the fungus flora of holm oak leaves during decomposition. Some species appeared as soon as leaf fall (Marasmius quercophilus, Trichoderma koningii, sterile mycelia) while others were late colonizers (Aspergillus glaucus, Penicillium chrysogenum, Phialophora sp.). Phyllosphere inhabitants such as C. cladosporioides and A. pullulans were also isolated frequently from freshly fallen litter and tended to decrease progressively until stage IV, whereas species of Mucor and Trichoderma were rarely found at an early stage of decomposition. From older litter stages high colonization rates were registered for the genera Trichoderma, Mucor, Penicillium and Aspergillus. Some species were isolated only from one (Trichoderma pseudokoningii) or two stages (Basidiomycetes S41 and S42, green sterile mycelium, Stachybotrys atra).

Serial washing, surface-sterilization and moist chambers yielded 42%, 15% and 43% of total colonies, respectively (Table 2). The low number of records from surfacesterilized leaves may indicate that relatively few species were able to penetrate internal tissues. The three methods combined revealed that 27% of the total fungal flora came from the phyllosphere and 73% came from the litter. The omnipresent C. cladosporioides All isolation methods established that fungal diversity increased with senescence and after leaf-fall (Fig. 2). As the decomposition of oak litter proceeded, species richness decreased progressively while the mean isolation frequency varied only little before stage V. During later stages (IV and V) few species have been isolated by surface sterilization, which could however indicate that the leaf inside was affected by sterilization in spite of our effort to reduce the time of immersion.

When considering coefficients of variation of isolation frequencies, some species appeared to be restricted to some particular stages, while others exhibited a large spectrum of occurrence (Table 2). For instance, T. roseum was nearly restricted to living leaves (C.V.=232%) while U. chartarum, B. cinerea and A. alternata seemed to prefer senescent leaves but were also found in other stages (149, 135% and 113%, respectively). Trichoderma pseudokoningii was isolated only at stage III and exhibited the highest coefficient of variation (265%). It should be noted that all species preferring stage V (A. glaucus, P. chrysogenum and Phialophora sp.) occurred only at this stage (C.V.=265%).

Axes 1 and 2 of correspondence analysis extracted 19.6% and 16.2% of the total variance, respectively. Fungal species (active variables), leaf stages and isolation methods (passive variables) were projected in the plane of the first two factorial axes (Fig. 3). Correspondence analysis showed that phyllosphere and stage I species were mostly detected by washing and to a lesser extent by surface-sterilization while litter species, and more particularly stages III to V, were mostly detected by moist chambers. Other results concerned the classification of fungal species along a gradient of increasing ageing of leaves, from positive values of Axis 1 (right side of the graph) to negative values (left side of the graph). Stages were spread along Axis 1 in the same order as they were stratified in the ecosystem, except senescent leaves, which were projected somewhat farther from the origin than living leaves. Axis 2 opposed washing to sterilization methods, highlighting advantages of one or the other method for the different fungal strains. For instance, sterilization revealed sterile basidiomycetes (S41 and S42), green and brown sterile mycelia and Chaetomium globosum, while washing revealed all Trichoderma species and many other highly-sporulating micromycetes.

DISCUSSION

The phyllosphere harboured relatively few fungal species such as Trichothecium roseum, Aureobasidium pullulans, Cladosporium cladosporioides, Alternaria alternata, Ulocladium chartarum, Botrytis cinerea and Epicoccum purpurascens. These fungi have been detected by all methods and are grouped into the common primary saprophytes (Hudson 1968), except Botrytis cinerea, a plant pathogen [START_REF] Smith | An introduction to industrial mycology[END_REF]Dickinson 1976), which has been isolated only by moist chambers in green and senescent leaves, and by washing only in senescent and stage I leaves. Isolation methods showed that these fungi were both superficial growers and internal colonists of green and senescent leaves. Hudson (1968) considered them to be present on living leaves as spores which become vegetatively active only at leaf senescence. However, the present investigation and others [START_REF] Ruscoe | Mycoflora of living and dead leaves of Nothofagus truncata[END_REF][START_REF] Lindsey | Succession of microfungi on attached leaves of Hippophaë rhamnoides[END_REF]Dickinson 1976;Wildman and Parkinson 1979) have shown that these fungi are active prior to leaf senescence.

As a whole, senescence is accompanied by a slight increase in species richness (Table 2, Fig. 2), but also by an increase in the internal colonisation of leaves, as depicted by higher colonial records after surface sterilization. However, among the fungi reported as endophytes by several authors, a number of taxa known to be frequent epiphytes are able to live endophytically within plant tissues [START_REF] Petrini | Fungal endophytes of tree leaves[END_REF]. For instance, A. alternata and C. cladosporioides are phyllosphere colonizers able to penetrate into leaving leaf tissues at the onset of the senescence process. Epicoccum purpurascens may behave in a similar way. The senescence process apparently modifies the ecological niche provided by leaf tissues to "true" endophytes and allows the development of organisms that are usually better adapted to saprotrophic life [START_REF] Petrini | Fungal endophytes of tree leaves[END_REF].

The small range of fungi growing on phyllosphere indicates a rather extreme environment, probably due to the exposure to changing weather conditions, limited availability of nutrients, and the presence of antimicrobial compounds [START_REF] O'donnell | Pathogenicity of Alternaria and Cladosporium isolates on Phaseolus[END_REF][START_REF] Petrini | Fungal endophytes of tree leaves[END_REF]. Some primary saprophytes such as Cladosporium, Alternaria and Epicoccum are able to survive on leaf surfaces exposed to ultra-violet radiation and dryness by forming dark, thick-walled clamydospores and microsclerotia (Pugh andBuckley 1971, Dickinson 1981).

All species of the phyllosphere, except Trichothecium roseum, persisted in fallen litter and tended to disappear in oldest stages of decomposition (stages IV and V). Freshly fallen leaves (stage I) were invaded by several new colonists, which increased dramatically the species richness (Table 2, Fig. 1). Among new colonizers appeared Marasmius quercophilus and species of Trichoderma, Mucor, Penicillium and Aspergillus, which are considered as secondary saprophytes (Hudson 1968).

This study showed many similarities with results obtained by workers having studied very different leaf materials, highlighting that the successional process we observed seems to be universal, occurring in deciduous as well as evergreen leaves.

However, the time needed for leaves to fall on the ground allows some fungus genera to develop in internal tissues a long time before leaf abscission. Everything happens as if the first stage of decomposition occurred on the tree, although with a restricted array of fungal strains.

Members of the genus Trichoderma are known to be secondary colonizers of a wide variety of forest litters and have been mostly recorded in relatively well decomposed materials (Domsch et al. 1980), but species differed in their affinity for temperature and moisture [START_REF] Widden | Competition between Trichoderma species: effects of temperature and litter type[END_REF], which may explain why our six Trichoderma species were found at so different stages. Trichoderma species are also known for their ability to produce cellulases [START_REF] Danielson | The abundance of Trichoderma propagules and the distribution of species in forest soils[END_REF], thus they could play a role in the loss of weight of holm oak litter (Sadaka-Laulan and Ponge 2000), together with actively decomposing basidiomycetes such as Marasmius spp., as this has been shown to occur on inoculated Scots pine needles by [START_REF] Cox | Effects of fungal inocula on the decomposition of lignin and structural polysaccharides in Pinus sylvestris litter[END_REF].

Physical and chemical characteristics of the leaf surface play an important role in governing the success or failure of fungal growth on, and subsequently in, the leaf.

Hardness and thickness may be significant in the resistance of leaves to fungal penetration [START_REF] Allen | Influence of leaf surface features on spore deposition and the epiphytic growth of phytopathogenic fungi[END_REF], which could explain the lower number of isolates obtained after surface sterilization (Fig. 2).

In a previous study (Sadaka-Laulan and Ponge 2000) we used the weight per unit area in order to follow the senescence and decomposition of holm oak leaves. We showed that leaves exhibited an increase in weight during senescence then stage I, followed by a decrease until stage V. A similar trend has been found here for the mycoflora and a significant correlation was obtained between leaf areal weight and total fungal isolates (r=0.793, P = 0.017). This allows us to hypothesize that the habitat size (here expressed by the areal weight) is an important determinant of the fungal population size, which can be explained by synergistic effects of space and nutrient availability [START_REF] Kinkel | Fungal community dynamics[END_REF].

  and A. pullulans and other species such as Trichothecium roseum, Ulocladium chartarum and Alternaria alternata were isolated by all methods. Most species, and particularly fastspreading micromycetes such as Trichoderma, Mucor and Penicillium have been detected simultaneously by washing and moist chambers. Marasmius quercophilus was observed only in moist chambers while Acremonium sp., Alternaria tenuissima, Ulocladium atrum and Epicoccum purpurascens were never recorded by this method. Few internal colonists have been isolated by surface-sterilization only, such as Chaetomium globosum, Basidiomycetes S41 and S42, hyaline and dark sterile mycelia.

Figure legends Figure 1 :

 legends1 Figure legends

Figure 2 :

 2 Figure 2: Species richness and total frequency of isolation from phyllosphere and leaf

Figure 3 :

 3 Figure 3: Correspondence Analysis. Projection of active variables (fungal species) and

  Fig.1

Fig. 3

 3 Fig. 3

Table 2 .

 2 Stage I Yellow, externally intact, thick, hard, lower side densely covered with green hairs Stage II Yellow, slightly decayed but still thick and hard, hair cover on the lower side invaded by fungi Stage III Brown, thin, soft, decayed, a few hairs still present Stage IV Brown, thin, very soft, strongly decayed but still recognizable, no hair present Stage V Bleached, very thin, brittle and more or less aggregated by fungal mycelium

Table 1 :

 1 Morphological features of the five decomposition stages of the holm oak leaf litter