Scuttle Flies (Diptera: Phoridae) Reared from Fungi in Benin

ABSTRACT Ten species of Phoridae reared from fungi in Benin are reported, including the descriptions of the following: Megaselia cakpoae Disney, sp. n., Megaselia fuscilobulorum Disney, sp. n., Megaselia kurinai Disney, sp. n. and Megaselia tedersooi Disney, sp. n. Megaselia termitomyca Disney, 1989 is synonymised with Megaselia labiata Borgmeier, 1967.


INTRODUCTION
The larvae of many scuttle flies are associated with fungi, some feeding on the fun gus, others preying upon or parasitizing the larvae of other fungus feeding Diptera (e.g. Disney 1994). The species associated with fungi in the Afrotropical Region have been little studied, mainly because they mostly belong to the huge genus Megaselia Rondani.
Of species reared from fungi the best known are the pest species of cultivated mushrooms. The latter include M. sandhui Disney, which is abundant in the Afrotropical part of Arabia, and the introduced Megaselia halterata (Wood). Both species are serious pests of cultivated agaric mushrooms, such as Agaricus bisporus (Disney 1981(Disney , 2008b, but M. halterata larvae feed on the mycelium whereas those of M. sandhui feed on the sporophore. The larvae of some species, such as Dohrniphora diminuens (Schmitz), have been reported feeding on the fungus combs of Termitomyces species (Lyophyllaceae) in termite nests (Disney & Kistner 1989). The sporophores of these fungi are exploited by a different suite of species, such as Megaselia labiata Borgmeier (= M. termitomyca Disney, see below) and M. zariaensis Disney (Disney 1989). Decaying fun gus sporophores are exploited by other species such as M. scalaris (Loew) (Disney 2008a) and Chonocephalus fletcheri Schmitz (Disney 2005).
In this paper we report on species reared by LT and YC from fungus sporophores in Benin (Table 1). The Phoridae were sorted from the other reared Diptera by OK and have been identified by RHLD, who describes four new spe cies below.
(Fabaceae). Fresh sporophores were wrapped in aluminium foil to pre vent desiccation and placed in 750 ml plastic containers covered with nylon gauze to ensure air circulation. Mixed hardwood sawdust was used as a pupation substrate for emerging insects. The adults that emerged were preserved in 70 % alcohol and RHLD subsequently mounted them in Berlese Fluid on slides (Disney 2001).
Approximately 25 % of the fresh sporophores of each fungus species was removed from the incubated sample, air-dried at 35 °C, labelled and deposited in the Herbarium of the Tartu University (TU). The DNA extracted from each sample was subjected to PCR amplification and sequencing of the rDNA Internal Transcribed Spacer region as de scribed in Tedersoo et al. (2011). Metadata of all sporophores are deposited in the pub lic UNITE database (Abarenkov et al. 2010a, b). As many of the fungi in our study area are undescribed they are identified to different taxonomic levels.
The type material and other voucher specimens of Phoridae are deposited in the University of Cambridge Museum of Zoology (UCMZ). These are all mounted on slides. For the new species, other specimens remaining in alcohol are given in brackets but are not treated as being part of the type series. Each fungal fruit body is supplemented by a corresponding voucher number of the sample from the same fruit body in the TU. The reference numbers on the slide labels, e.g. 42-6, refer to RHLD's notebook (42) and page (6). TAXONOMY Genus Megaselia Rondani, 1856 All but one specimen belong to the genus Megaselia. The recognition of the species in this huge genus is based on the male hypopygium in the first instance. Beyer has  (Beyer 1965). However, he either fails to illustrate the hypopygia or his figures are sketchy and often mis leading. Indeed it has been generally judged to be difficult to use Beyer's keys with confidence. This is partly due to runs of couplets being based on male features only interspersed with couplets based on female characters only. Furthermore, Beyer described some species from females only, including cases where he described the male as a separate species. In addition his keys are notable for errors and omissions, such as the species from the Seychelles described by Collin, apart from there being many sub sequently described species. Both sexes of the four new species run to M. ahmedseifi Disney, 2009 in the keys to Arabian species (Disney 2009b). With it these five species form a distinct species group. A key to all five species is given below. Otherwise the way the species run down in Beyer's (1965) keys are indicated for each of the new species. Disney, 2009 This species was previously known from Arabia. Both sexes are keyed by Disney (2009b

Male.
Frons yellow to orange brown, clearly broader than long, with 40-60 hairs and dense but very fine microtrichia. Only 2 SAs. Antials lower on frons than ALs, and about as far from SAs as either is from AL. Pre-ocellars at most as far apart as SAs and closer together than either is from ML bristle, which is at about the same height on frons.
Cheek with 0-2 bristles and jowl with 2 longer bristles. Subglobose postpedicels yellow, without subcutaneous pit sensilla (SPS vesicles). Palps pale yellow, only about 0.2× as broad as postpedicel and 1.1-1.2× as long as breadth of latter, with 4 short bristles (longest, apical, bristle being 0.08-0.09 mm) and up to twice as many hairs. Labrum pale yellow and about 0.7× as wide as postpedicel. Labella coloured as palps, with 2 or 3 hairs on each above and very few short spinules below. Thorax yellow apart from brown patch on pteropleuron. Three notopleural bristles and no cleft in front of these. Mesopleuron bare. Scutellum with anterior pair of fine hairs (subequal to those in middle of scutum) and posterior pair of bristles. Abdominal tergites yellow apart from brown areas on outer thirds of T3 and T4. T1 to T6 with small hairs, mainly at hind margins and towards the sides, and being longest posterolaterally from T2 onwards and at rear of T6. Venter yellow, with minute hairs on the sides behind spiracles and with small hairs below on segments 3-6. Hypopygium (Fig. 6) mainly brown, with yellow anal tube.
Legs yellow apart from brown patches on mid coxae. Fore tarsus with pd hair palisade on segments 1-5, segment 5 very slightly longer than 4. Dorsal hair palisade of mid tibia extends just over 0.75× of its length. Last segment of mid tarsus longer than 4 th (similar to Fig. 19). Hairs below basal half of hind femur longer than those of av row of outer half. Base of hind tibia as Figs 1 & 2, with 6 or 7 differentiated pd hairs beyond the modified basal region and spinules of apical combs simple. Wings 1.60-1.76 mm long. Costal index 0.45-0.47. Costal ratios 3.4-4.9:1.4-2.1:1. Costal cilia (section 3) 0.06-0.07 mm long. No hair at base of vein 3. With 2 axillary bristles, the outer being longer than costal cilia. Sc not reaching R1. Thick veins yellowish grey, thin veins 4-6 grey and 7 pale. Membrane only lightly tinged grey (not obviously evident to naked eye when viewed against a white background). Haltere knob yellow.

Female.
Head similar to male but with 2 or 3 bristles on cheek. Palps up to 1.4× as long as width of postpedicel and with 5 bristles. Labrum 0.9× as wide as diameter of postpedicel. Thorax as male. Abdominal tergites yellow, progressively a little narrower from T2 to T6. Venter yellow, with hairing of segments 3-6 similar to male. Segments 7 and 8 brown, elongated and retractile. Light brown tergite 7 about 4.5× as long as broad, with a pair of longer hairs at rear end and half a dozen shorter ones further forward. Sternite 7 light brown, about 4× as long as broad and with hairing similar to T7. Single lobe at rear of sternum 8 as Figs 5, 6. Cerci and epiproct as Fig. 3. Four rectal papillae. Furca not evident. Dufour's crop mechanism about 4× as long as broad and almost parallel sided with a rounded rear end. Legs similar to male except 5 th segment of mid tarsus being shorter than 4 th ; hind tibia not modified in its basal section, so there are about a dozen differentiated posterodorsal hairs. Wing as male except length is 1.61-1.92 mm. Costal index 0.44-0.47. Costal ratios 3.6-4.5:1.5-2.2:1. Costal cilia 0.07 mm. Haltere as male.  Remarks: In Beyer's (1965) keys the male runs to page 54, couplet 28 to M. tergatula Beyer, 1965, only known from the female, but the latter has brown postpedicels. Proceeding the male runs to couplet 30, lead 2, to M. nubila Colyer, 1952, but the latter has brown legs. The female on page 54 runs on to couplet 22, to M. incrassaticosta Bridarolli, 1951, only known from the male, but its thorax is brown rather than yellow. Proceeding to couplets 24 to 26 based on males only, one can continue on the basis of the costal index or costal cilia being too short for the species of these couplets. It then runs to couplet 30 to M. nubila again. Megaselia fuscilobulorum Disney, sp. n. Figs 7-10 Etymology: From Latin fuscus (dark) and lobus (lobe), with reference to the female's dark, sclerotised lobes at the rear of abdominal sternum 8. Description:

Male.
Frons brown, clearly broader than long, with 60-90 hairs and dense but very fine microtrichia. SAs with lower pair shorter than longest bristles on palps. The antials lower on frons than ALs, and further from upper SAs as either is from AL. Pre-ocellars about as far apart as upper SAs, and as either is from ML, which is very slightly higher on frons.
Cheek with 1 or 2 (usually 2) bristles and with two longer ones on jowl. Subglobose postpedicels pale dusky yellow without subcutaneous pit sensilla (SPS vesicles), but unusually with irregular basal rims surrounding the pedicels. Palps pale yellow, 1.6× as broad as labrum but slightly longer than breadth of postpedicel, with 5 or 6 bristles (longest being longer than lower SAs) and half as many hairs. Labrum pale yellow and about ⅔ as wide as postpedicel. Labella coloured as palps, each with 2 hairs above and with very few short spinules below. Thorax with orange to yellowish brown scutum, a darker scutellum and paler pleural regions apart from brown on pteropleura. Three notopleural bristles and no cleft in front of these. Mesopleuron bare. Scutellum with Figs 7-10. Megaselia fuscilobulorum sp. n., ♂ (7, 10) and ♀ (8,9): (7) anterior face of base of hind tibia, (8,9) lobes at rear of sternum 8 in two different specimens, (10) left face of hypopygium. Scale bar = 0.1 mm.
anterior pair of hairs (about equal to those in middle of scutum) and posterior pair of bristles. Abdominal tergites partly brown, and partly yellow. Typically T1 entirely brown, T2 and T6 yellow in their anterior halves at least, and T3 to T5 with variably yellow median patches in their anterior halves at least. Hairs tend to be most numerous towards hind margins and posterolaterally, and longest at T6 hind margin. Venter yellow, and with conspicuous hairs on segments 3-6. Hypopygium (Fig. 10) brown, with yellow anal tube. Right lobe of hypandrium vestigial. Legs yellow apart from brown patches on mid coxae and tips of hind femora. Fore tarsus with pd hair palisade on segments 1-5, but that on 5 is abbreviated, and segment 5 about as long as 4. Dorsal hair palisade of mid tibia extends almost 0.75× of its length. Segment 4 of mid tarsus longer than segment 5. Hairs below basal half of hind femur longer than those of av row of outer half. Hind tibia with 3 or 4 differentiated hairs in basal region (Fig. 7)

Female.
Head similar to male but labrum about 1.1× as wide as a postpedicel and inner edges of labella with 5 teeth. Thorax as male. Abdominal T1-5 with anterior halves to two thirds yellow and the rest brown, but T6 largely yellow with a little brown at rear. Small hairs on all tergites but 1 or 2 longer ones towards sides of T2 and at rear of T6. T7 subrectangular, but very slightly tapered, and largely light brown; and about 1.5× as long as broad with a dozen hairs in its rear half and 4 longer ones at its hind margin. Venter yellowish grey, with conspicuous hairs below on segments 3-6. Sternite 7 brown, 1. Remarks: In Beyer's (1965) keys the male runs to page 54, couplet 28 to M. tergatula, only known from the female, but the latter has brown postpedicels. Proceeding the male runs to couplet 30, lead 2, to M. nubila, but the latter has brown legs. Some males with a costal index of less than 0.44 will run down on page 56 to couplet 4, but its haltere knob being yellow not brown one proceeds to couplet 5. Its thorax being paler not dark brown means one then proceeds to couplet 10, where the yellow legs and thorax exclude both species. The female on page 54 runs to couplet 22, to M. incrassaticosta, only known from the male, but its thorax is dark brown rather than pale brown at most. Proceeding to couplets 24 to 26 based on males only, one can continue on the basis of the costal index or costal cilia being too short for the species of these couplets. It then runs to couplet 30 to M. nubila again.

Male.
Frons brown, clearly broader than long, with 28-52 hairs and dense but very fine micro trichia. Supra-antennal bristles (SAs) very unequal, lower pair being at most 0.75× as long as apical bristles of palps. The antials lower on frons than ALs and further from upper SAs as either is from AL. Pre-ocellars about as far apart than either is from ML, which is about the same level on frons. Cheek with 4-6 bristles and jowl with two longer ones. The subglobose postpedicels yellow, without subcutaneous pit sensilla (SPS) vesicles. Palps yellow, only about 0.2× as broad as postpedicel but at least as long as breadth of latter, with 4-6 bristles and as many hairs. Labrum pale yellow and about 3× as wide as a palp. Labella coloured as palps and with only a few short spinules below. Thorax yellow. Three notopleural bristles and no cleft in front of these. Mesopleuron bare. Scu tellum with anterior pair of hairs (about as long as those in middle of scutum) and pos terior pair of bristles. Abdominal T1 and T2 largely yellow but with brown patches in rear of outer thirds at most. T3 and T4 largely brown with yellow median bands. T5 yellow with small brown smudges at front in outer thirds. T6 entirely yellow. All tergites with fine hairs, which are longest at rear of T6. Venter yellow, with a few small hairs on seg ment 6 only. Hypopygium largely brown, with yellowish brown anal tube; right lobe of hypandrium small (Fig. 16). With 4 rectal papillae. Legs yellow apart from brown patches on mid coxae and slightly brown tips to hind femora. Fore tarsus with pd hair palisade on segments 1-5, and segment 5 about as long as 4. Dorsal hair palisade of mid tibia extends about ⅔ of its length. Segments 3-5 of mid tarsus as Fig. 12. Hairs below basal half of hind femur longer than those of av row of outer half. Base of hind tibia as Fig. 11, with about a dozen differentiated pd hairs and spinules of apical combs simple. Segments 3-5 of hind tarsus as Fig. 13. Wings 1.1-1.4  Sc not reaching R1. Thick veins yellowish grey, thin veins 4-6 grey and 7 very pale. Membrane lightly tinged grey (only just evident to naked eye when viewed against a white background). Haltere knob brown.

Female.
Head similar to male but labrum about 0.9× as wide as diameter of postpedicel but 3× as wide as a palp. Thorax as male. Abdominal tergites yellow with hairs mainly at hind margins and longest at rear of T6. Venter yellow and with a few hairs at rear of segment 6. Retractile terminal segments dark. Tergite and sternite 7 represented by a few hairs only. Posterolateral lobes at rear of sternum 8 with narrow, dark and bare tips (Figs 14,15). Cerci vestigial (Fig. 15). With 4 rectal papillae. Furca not evident. Dufour's crop mechanism at least twice as long as broad and rounded behind. Legs similar to male but segments 5 of the mid and hind tarsi shorter than 4 th segments. Wing as male except 1.4-1.5 mm long. Costal index 0.45-0.46. Costal ratios 4.3-5.3:2.9-3.0:1. Hal tere as male.
Holotype: ♂ BENIN: Koussanzamou, ex Russula sp. 2 (TU116098), 5.vii,. Paratypes and other material examined: 1♀ same data as holotype (42-9); 1♂ same locality, ex Tylopilus sp. Remarks: In Beyer's (1965) keys both sexes run to the same couplets on page 54 as the previous species, to M. nubila, and is likewise distinguished from this species. Speci mens with the costal index less than 0.44 will run to couplet 10 on page 57, but the combination of yellow legs and postpedicels and 3 bristles on the notopleuron rules out the two species of this couplet.
This species was described from the Congo, where it was associated with termites (Borgmeier 1967). Having been described after Beyer's (1965) keys, it was included in Borgmeier's (1968) catalogue. However, many of Beyer's species were omitted from this catalogue as Beyer declined to send Borgmeier a copy of his 1965 monograph. This was because Beyer had been upset by Borgmeier's criticisms of his work. Beyer's monograph only became known to Borgmeier in March 1969 when it was abstracted by the Zoological Record (Borgmeier 1971). Consequently Borgmeier was unable to indicate how M. labiata runs down in Beyer's keys. It was only when RHLD belatedly annotated these 1965 keys to include subsequently described species that it revealed M. termitomyca to be a synonym of M. labiata. This species is not covered by any key. In Beyer's (1965) keys both sexes run to couplet 19 on page 54. The male runs to lead 2, to M. nigricauda Beyer, 1965, but is immediately distinguished by its enlarged labella of the proboscis (Borgmeier 1967 : fig. 2). The female of M. labiata fits neither option in couplet 19, as the thorax is too pale for lead 2 and the frons too wide for lead 1.
It was previously reared from the stipes of Termitomyces sp. in Zaria, Nigeria (Disney 1989 Disney, 1991 This species has proved to be widespread and both sexes are included in recent keys (Disney 2009a, b). It has previously been reported from the Cape Verde Islands, Arabia and the Seychelles.
Megaselia scalaris (Loew, 1866) The larvae of this cosmopolitan species feed on a wide range of decaying organic materials and are occasionally facultative secondary feeders on fungi already spoiled by other fungus feeding insects (Disney 2008a). Both sexes are included in recent keys (Disney 2009b).

Male.
Frons brown, clearly broader than long, with 38-40 hairs and dense but very fine micro trichia. Supra-antennal bristles (SAs) very unequal, lower pair being a little shorter than apical bristles of palp. Antials lower on frons than ALs and further from upper SAs than either is from AL. Pre-ocellars slightly closer together than either is from ML, all 4 bristles at about the same level on frons. Cheek with 4 bristles and jowl with 2 longer ones. The subglobose postpedicels yellow, without subcutaneous pit sensilla (SPS) vesicles. Palps pale yellow, about 0.2× as broad as postpedicel but about as long as breadth of latter, with 6 bristles and as many hairs. Labrum pale yellow and about 0.7× as wide as a postpedicel. Labella coloured as palps and with only a few short spinules below. Thorax yellow. Three notopleural bristles and no cleft in front of these. Me sopleuron bare. Scutellum with anterior pair of hairs (about as strong as those in middle of scutum) and posterior pair of bristles. Abdominal T1, T5 and T6 yellow, T2 yellow with brown at rear margin in outer thirds, T3 and T4 brown with a median yellow stripe. All tergites with small hairs, mainly near hind margins, but some longer ones towards sides of T2 and clearly longer ones at rear of T6. Venter dusky yellow and with conspicuous hairs on segments 3-6. Hypopygium (Fig. 22) mainly brown, with yellowish brown anal tube, hypandrial lobes being vestigial. With 4 rectal papillae. Legs yellow except for brown patches on mid coxae. Fore tarsus with pd hair palisade on segments 1-5, segment 5 about as long as 4. Dorsal hair palisade of mid tibia extends about ⅔ of its length. Segments 3-5 of mid tarsus as Fig. 19. Hairs below basal half of hind femur longer than those of av row of outer half. Base of hind tibia as Figs 17 & 18, with 7 or 8 differentiated pd hairs beyond this modified region (the first and base of the second of these are evident in Fig. 17), and spinules of apical combs simple. Segment 5 of hind tarsus almost as long as segment 4. Wings 1.48-1.49 mm long. Costal index 0.44-0.45. Costal ratios 3.7:2.4-2.5:1. Costal cilia (section 3) 0.06-0.07 mm long. No hair at base of vein 3. With 2 axillary bristles, outer being longer than costal cilia. Sc not reaching R1. Thick veins yellowish grey, thin veins 4-6 grey and 7 pale. Mem brane lightly tinged grey (just evident to naked eye when viewed against a white back ground). Haltere knob yellow.

Female.
Head similar to male but labrum about 0.9× as wide as diameter of postpedicel. Thorax as male. Abdominal tergites yellow, with small hairs apart from a few longer ones towards sides of T2. Venter dusky yellow and with about a dozen hairs below seg ments 3-6. Segment 7 onwards darkened and retractile. Posterolateral lobes at rear of sternum 8 as Fig. 21. Cerci as Fig. 20. With 4 rectal papillae. Furca not evident. Dufour's crop mechanism at least 3× as long as broad and rounded behind. Legs similar to male but fifth segment of mid tarsus is slightly shorter than fourth and base of hind tibia not modified, so that there are about a dozen posterodorsal differentiated hairs. Wing as male except 1. Remark: In Beyer's (1965) keys both sexes run to the same couplets on page 54 as the previous species, to M. nubila, and is likewise distinguished from this species. The fact that four out of the nine species of Megaselia obtained were new species underlines our current ignorance of Afrotropical species of this huge genus. Furthermore, these four species evidently belong to a distinctive complex of sibling species. The do minance of female flies, at 62.6 %, is striking. These females could only be named be cause they were associated with their males in reared series. Indeed such associations, apart from pairs caught mating, are required for the identification of most females of Afrotropical species of Megaselia. In this case unassociated females could only have been assigned to the sibling species complex and not to the individual species represented by their males. When collections from a locality include specimens of both sexes belonging to different species complexes it is often possible to link females to their males on the basis of small details (e.g. Disney 2009a, b), but the use of molecular barcodes is likely to prove useful in the future (e.g. Boehme et al. 2010). Our records highlight a similar current ignorance of Afrotropical fungi.