Earthworms (Annelida: Oligochaeta) of Sao Tomé

During a soil zoology expedition to Sao Tomé Island, among other members of the soil fauna, earthworms were collected. During this collecting trip some 170 earthworm specimens belonging to 18 species were gathered of which two, Dichogaster (Diplothecodrilus) coeruleoviridis and Dichogaster (Diplothecodrilus) zicsii, proved to be new to science. In addition, a small sample from the same locality was also examined and Dichogaster (Dichogaster) thomeana Cognetti, 1910 was removed from synonymy of Dichogaster (Dichogaster) greeffi Michaelsen, 1902.


Introduction
Sao Tomé is a volcanic island in the gulf of Guinea, some 200 km off the West African coast. Together with Annobon, Principé, and Bioko islands it belongs to the Cameroon volcanic line. The origin of the island chain is somewhat debated but the newest data favour a linear mantle upwelling zone origin contrary to a single hotspot hypothesis (Meyers et al. 1998), so the age of the islands does not necessarily correspond with the position in the volcanic line. It has been shown that most of the islands are of Miocene age, and Sao Tomé is dated about 15 million years (Akaa et al. 2001). The island was unpopulated until the 16th century, when Portuguese colonists settled and introduced Angolan slaves. Since then, owing to agriculture, the lowland regions of the island have been heavily disturbed and many species have been introduced, but the region of the central volcano remained relatively untouched and harbours a number of autochthonous species.
In 2000, as a member of the joint expedition of the Hungarian Public Television and the Hungarian Academy of Sciences organized by the late Prof. Dr János Balogh, I had the opportunity to collect earthworms in different parts of the island including the most natural central volcano region. This collection, together with a smaller sample from Dr John Measey (IRD France), resulted in some 170 earthworm specimens, distributed among 18 species and six families.

Materials and methods
Earthworms were collected by digging and searching under the bark of fallen logs. The animals were killed in 75% ethanol, preserved in 4% formol and after several days transferred into 75% ethanol. The penial setae were removed by dorsal dissection of the specimens and mounted in Euparal for light microscopic study. Setae of the species of taxonomic importance were also investigated using a Hitachi SN-2600N scanning electron microscope. For SEM studies they were glued on aluminium stubs using double-sided carbon tape and sputter-coated with gold.
All the material is deposited in the Oligochaeta collection of the Hungarian Natural History Museum. Abbreviations used in the text: L., length; D., diameter.  Figure 1). Spermathecae skittle-shaped, duct short muscular, diverticula acinous ( Figure 2). Calciferous glands three pairs in 15-17, but the first pair is very small. Meronephridia six or seven on each side of the intestine. Penial setae uniform, the fully adult seta 1.3-1.4 mm long and in the middle 0.02 mm wide.

Systematics
The ectal third of the seta slightly undulated and the tip is sharply hooked. Each seta bears a characteristic ornamentation of large slightly abutting scales ( Figure 3).

Diagnosis
L. 55-60 mm, D. 2.5-3 mm, number of segments 117-126. Colour alive purplish brown, conserved pale. First dorsal pore in 12/13. Clitellum 13-20, prostatic pores in segment 17, 19, spermathecal pores in 7/8, 8/9 (Figure 4). Spermatheca ampulla barbell-shaped, duct long muscular, diverticula empty, unilocular with long duct ( Figure 5). Calciferous glands three pairs in 15-17, the first pair is slightly smaller than the others. Meronephridia four to five on each side of the intestine. Penial setae uniform, the fully adult seta 1.7-1.8 mm long and in the middle 0.04 mm wide. The seta is slightly curved and the spoon-shaped tip is reclinate. Each seta bears a characteristic ornamentation which are large serrated sculptures on the ental part becoming large thorns towards the ectal part ( Figure 6).

Remarks
This species was previously known only from the original description. Based on it Csuzdi (1996) placed D. feai into the subgenus Dichogaster. The well-preserved new material revealed that the first dorsal pore is located in 5/6 and the spermatheca is subdivided, possessing a simple or sometimes bifurcated diverticulum, proving that this species belongs to the subgenus Diplothecodrilus. The moderate number of meronephridia (six to seven per side) also corroborates this fact.

Remarks
This species was transferred to Benhamia Michaelsen, 1889 based on the investigation of the softened and somewhat macerated type specimen (Csuzdi and Zicsi 1994). In the present collection we found two preadult exemplars of this species. A thorough examination of the excretory system revealed that this species belongs to the genus Dichogaster, and shows high affinity with D. (Dt.) feai but there are important differences; the greater body size, the presence of extremely thickened dissepiments in 11/12, 12/13, and 13/14 and the more numerous meronephridia (9-10 on each side). There are differences also in the penial setae. In case of D.

Internal characters
No septa notably thickened. Oesophageal gizzards two, in segments 5 and 6. The three pairs of calciferous glands are of about equal size, located in segments 15-17. Excretory system meronephridial, with two to three meronephridia on each side. Paired hearts are present in segments 10, 11, and 12. Testes are paired in 10 and 11 enclosed into free sperm mass filling the cavity of the segments. Seminal vesicles lacking, ovary small in segment 13. Seminal duct convoluted, discharging through a small muscular atrium in segment 18. Two pairs of small prostatic glands are present in 17 and 19, confined into their own segment and each provided with a penial setal sack containing two different types of penial setae. The larger one is about 0.4 mm long and 0.008 mm wide with spatula-like tip and minute serrated ornamentation. The smaller one is smooth, about 0.3 mm long and 0.006 mm wide with sharply pointed tip ( Figure 10). There are two pairs of spermathecae in segments 8 and 9. Each has an elongated skittle-shaped ampulla and an equally long duct. The lower part of the ampulla bears a small, sometimes bifid diverticulum ( Figure 11).

Remarks
Dichogaster (Diplothecodrilus) zicsii, bearing two types of penial setae, belongs to the bolaui species group (Csuzdi 1996) and shows affinities with D. (Dt.) bolaui (Michaelsen, 1891) and D. (Dt.) amphibiotica Dahl, 1957 but differs from both species in the paired female pores, in the shape and ornamentation of the penial setae and furthermore in the presence of a muscular thickening at the end of the male duct.

Derivatio nominis
The name of this species refers to its characteristic bluish green colour.

Internal characters
There are no septa notably strengthened. Oesophageal gizzards two, in segments 5 and 6. Three pairs of calciferous glands of about equal size, situated in segments 15-17. Excretory system meronephridial, with five meronephridia on each side of the intestine. Paired lateral hearts are present in segments 10, 11, and 12. Testes are paired in 10 and 11 enclosed into perioesophageal sperm sacs. Two pairs of small seminal vesicles present in 11 and 12, and a pair of large racemose ovaries pendent from the posterior face of the septum 12/13. Seminal duct convoluted forming a big bow in segment 15, discharging through a small thickened ectal part, as wide as the prostatic duct, in segment 18. The two pairs of prostatic glands are moderately large occupying two to three segments. Each prostate accompanied by a penial setal sac containing three to four setae of the same type. The penial setae are about 1.4 mm long and 0.02 mm wide with a somewhat hooked tip. The bent part is smooth without any sculpture, below the elbow it has a characteristic ornamentation of scattered thorns and five protruding denticles (Figure 13). Two pairs of spermathecae present in segments 8 and 9. Each consists of a long duct and subdivided ampulla. The lower part of the ampulla is narrow, slightly curved and at the junction with the duct bears a small, sometimes bifurcating diverticulum (Figure 14).

Discussion
Earthworms are low dispersers and due to their intolerance of salt water they were thought to be lacking from true volcanic islands (Sims 1980). The rich earthworm fauna, including six endemic species, found in Sao Tomé seems to contradict this widely accepted opinion (Omodeo 1963;Sims 1980;Csuzdi 1994) and together with a previous report of James (1996) indicates that a small-scale over-water dispersal could not be excluded.
Except for D. pinguis all the endemic species of Sao Tomé live in fallen trunks, indicating that rafting tree trunks, at least over small distances, might be an important factor for over-water dispersals, even in the case of earthworms. Sea-level lowering that occurred several times during the Tertiary and Quaternary might have facilitated dispersal events.
In the first account of the earthworm fauna of Sao Tomé (Cognetti 1910), apart from the four endemic species only one peregrine worm, the widely introduced tropical Eudrilus eugeniae, was reported. In the present survey, some hundred years later, 18 species are reported including six endemic ones. All the six species thought to be autochtonous to the island were collected in the more natural central volcano region. In the costal plains that are converted mostly to plantations (cocoa, oil palm etc.) and arable fields, only widely introduced peregrine species have been collected, demonstrating the fragility of the endemic earthworm fauna.