Original article Evidence of reproductive isolation confirms that Apis andreniformis (Smith, 1858) is a separate species from sympatric Apis florea (Fabricius, 1787) S Wongsiri K Limbipichai P Tangkanasing M Mardan T Rinderer HA Sylvester G Koeniger G Otis 1 Bee Biology Research Unit, Faculty of Science, Chulalongkorn University, Bangkok 10330, Thailand; 2 Department of Plant Protection, Universiti Pertanian Malaysia, 43400 Serdang, Selangor, Malaysia; 3 Honey-Bee Breeding, Genetics & Physiology Research 1157 Ben Hur Road, Baton Rouge, Louisiana 70820, USA; 4 Institut für Bienenkunde D 6370 Oberursel 1, FRG (Received 7 September 1989; accepted 29 September 1989) Summary — The species Apis andreniformis (Smith, 1858), the small dwarf honey bee of South- east Asia, is recognized as a valid biological species. This recognition is based on distinctive endo- phallus characteristics in comparison with sympatric Apis florea (Fabricius, 1787). Additionally, scan- ning electron microscope images of drone basitarsi are presented, as are preliminary comparisons of wing venation. Apis florea / Apis andreniformis / taxonomy / reproductive isolation INTRODUCTION characteristics of Apis florea (Fabricius, 1787) that are reported for worker bees (Maa, 1953). In 1984, our group collected dwarf honey bees in Thailand in the province of Chan- Wu and Kuang (1986, 1987) reported taburi near the border with Kampuchea. that secondary sex characteristics differed Laboratory examinations of worker bees between drones of A florea and A andre- from these collections revealed that some formis. Specifically, both have a furcated bees had the species specific characteris- basitarsus, presumably modified to grasp tics of Apis andreniformis (Smith, 1858) queens during mating (see Ruttner, 1988). and that others had the species specific The furcated basitarsus is quite different in * Correspondence and reprints. the 2 groups. Following the report of Wu In contrast, a full stage 9 eversion of an and Kuang, we re-examined the sympatric A florea endophallus has a pair of bursal populations in 4 Thai provinces, as well as cornua that coil or generally fold twice, a bees from East and West Malaysia. pair of upper cornua at the sides of the Drones and workers were collected in vestibulum, each of which is composed of 1988 from nests of both A florea and A an- 3 joined protrusions; a continuous, non- dreniformis in the Thai provinces of Chan- indented hairy field on the ventral surface taburi (Southeastern), Uthaithanee (West- extending from the vestibulum; a small pro- ern), Chiang Rai (Northern) and trusion proximal to a small dorsal hairy Chumporn (Southern). A andreniformis patch; a fimbriate lobe having 3 protru- was collected in 1989 in Malaysia. sions and a comparatively thin and strong- curved terminal distal Comparative studies of endophalli re- ly portion extending vealed evidence of reproductive isolation to the fimbriate lobe. between the 2 groups, and hence con- Figure 1 illustrates these characters. firmed the biological validity of naming 2 The drawings are based on direct observa- species of dwarf bees. In addition, drone tions of several specimens as well as the hind legs were examined using electron accompanying photographs. microscopy, and worker bee wing venation Figure 2 shows photographs of the basi- was studied. tarsal extensions described by Wu and Kuang (1986, 1987). Quite clearly, the thumb-like extension is comparatively characteristics Species-specific short in A andreniformis, and was absent from the only drone of A andreniformis ex- amined from This exten- Comparisons of the endophalli of A an- Sabah, Malaysia. dreniformis and A florea reveal numerous sion presumably provides a "clasper or- structural characteristics which assure gan" that fits on the queen’s hind tibia complete reproductive isolation between during mating (Ruttner, 1975). However, whether or not this of function these species (fig 1). A full or stage 9 ever- description sion (Woyke and Ruttner, 1958; Ruttner et is accurate awaits confirmation from obser- vations of The venation of al, 1973) of an A andreniformis endophal- matings. wing andreniformis workers is more similar lus has a pair of bursal cornua that fold for- Apis to that of dorsata and koschevni- ward once near their end; a pair of upper Apis Apis cornua at the sides of the vestibulum, kovi than it is to Apis cerana or Apis florea. The cubital index of andreniformis is each of which has small protrusions; a Apis hairy field on the ventral surface that has 4 large (x for colonies = 6.07, N = 11 colo- separate hairy patches distal to the pri- nies, 10 worker bees each; ranges = 5.03 mary hairy area near the vestibulum; an in- to 9.21 for colony averages and 3.5 to 13.8 dented triangular hairy patch on the dorsal for individual bees) while the cubital index surface proximal to a fimbriate lobe; a fim- of A florea is comparatively small (x for col- briate lobe having 6 small protrusions; and onies = 2.78, N = 6 colonies, 10 worker a comparatively thick and straight terminal bees each; ranges = 2.53 to 3.14 for colo- portion extending distal to the fimbriate ny averages and 1.93 to 5.27 for individual lobe. bees). CONCLUSION trales (BC) et dorsales (UC), dans la sur- face pileuse ventrale (HF) et dorsale (HDP), dans le lobe feuilleté (FL) et dans andreniformis is a valid Apis (Smith, 1858) la et la forme de la termi- which is found in south- longueur partie biological species nale qui s’étend après le lobe feuilleté (T). east Asia and is with florea sympatric Apis Ces différences sont la preuve de l’isole- in much of Thailand. It is also found in the ment reproductif des deux espèces sympa- Southern China peninsula, and Malaysia triques et donc, la preuve qu’il s’agit bien Borneo but its exact distribution and sym- dans le cas présent de véritables espèces. with florea are unknown. patry Apis Equal- Les endophallus d’A andreniformis de unknown are most ly comparative aspects Thaïlande et de Malaysie sont semblables. of A andreniformis biology. Much that was Sont en outre des au published concerning A florea may be A jointes photos de l’extension ba- andreniformis biology. Extensive compara- microscope électronique sitarsale des anté- tive studies are required to describe and caractéristique pattes properly attribute known dwarf honey bee rieures des mâles des deux espèces anté- biology to the correct species and discover (fig 2). Les photographies de l’aile clairement what is unique to each species. rieure des 2 espèces montrent l’index cubital élevé, caractéristique d’A an- dreniformis (x = 6,07) et celui petit, carac- ACKNOWLEDGMENTS téristique d’A florea (x = 2,78) (fig 3). La répartition précise d’A andreniformis aussi bien sa sont encore in- In cooperation with the Louisiana Agricultural que biologie connues. Des études sont Experiment Station. Sincere appreciation is ex- comparatives certains pressed to Gerard Perrone for his work in pre- nécessaires, car dans le passé paring the figures. We thank H V Daly, N Koeni- caractères d’A andreniformis ont certaine- ger, B Oldroyd and F Ruttner for suggestions ment été attribués à tort à A florea. concerning the manuscript. G W Otis was sup- ported by a research leave grant from the Uni- versity of Guelph, Guelph, Ontario. Financial Apis florea / Apis andreniformis / systé- support was provided by the Research Affairs matique / isolement sexuel Division of Chulalongkorn University, Bangkok, Thailand. Zusammenfassung — Der Beweis für die reproduktive Isolierung bestätigt, Résumé — La preuve de l’isolement dass Apis andreniformis (Smith, 1858) sexuel confirme qu’Apis andreniformis eine von der sympatrischen Apis florea (Smith, 1858) et Apis florea (Fabricius, (Fabricius, 1787) getrennte Art ist. Apis 1787) sont deux espèces sympatriques andreniformis (Smith, 1858) und Apis séparées. Apis andreniformis (Smith, florea (Fabricius, 1787) wurden in 1858) et Apis florea (Fabricius, 1787) ont folgenden Provinzen von Thailand été trouvées en position de sympatrie dans sympatrisch nachgewiesen : Chantaburi, les régions thaïlandaises suivantes : Chan- Uthaithanee, Chiang Rai und Chumporn. taburi, Uthaithanee, Chiang Rai et Chum- Untersuchungen des Endophallus der porn. L’examen de l’endophallus des Drohnen beider Zwergbienen ergaben bei mâles de ces abeilles naines a révélé des folgenden strukturellen Einzelheiten différences dans presque tous les détails Unterschiede (Abb 1) : bei den ventralen de structure (fig 1 ) : dans les cornules ven- (BC) und dorsalen (UC) Hörnchen (Cornua), beim ventralen Haarfeld (HF), REFERENCES beim dorsalen Haarfeld (HDP), beim und in der und Federanhang (FL) Länge Fabricius JC (1787) Mantissa Insectorum Vol I Form des Endteils nach dem Federanhang Proft. Hafniae Diese Unterschiede sind Beweise für (T). Maa T (1953) An into the of eine Isolation zweier inquiry systematics reproduktive sympa- the tribus Apidini or honey bees Hym. Treu- trischer Formen, also Beweise, dass es bia 21, 525-640 sich hierbei um echte Arten handelt. Die Ruttner F (1975) Ein metatarsaler Haftapparat Endophalli von A andreniformis von bei den Drohnen der Gattung Apis (Hyme- Thailand und Malaysia sind ähnlich. noptera: Apidae). Entomol Germanica 2, 22- Zusätzlich wurden elektronenmikros- 29 kopische Fotos über den basitarsalen Ruttner F (1988) Biogeography and taxonomy of Fortsatz (Klammerorgan) des Hinterbeins honey bees. Springer Verlag, Berlin NY N der Drohnen von beiden Arten beigefügt Ruttner F, Woyke J, Koeniger (1973) Repro- der duction in Apis cerana 2. Reproductive or- (Abb 2). Fotografien Vorderflügel and natural insemination. J Res beider Arten deutlich den charak- gans Apic zeigen 12, 21-34 teristisch Kubital-Index von A grossen Smith F (1858) Catalogue of the Hymenopter- andreniformis = und den charak- (x 6,07) ous insects collected at Sarawak, Borneo; teristisch kleinen Kubital-Index von A Mount Ophir, Malakka; and at Singapore, florea (x = 2,78) (Abb 3). Wallace AR. J Proc Linn Soc London Zool 2, 42-130 Sowohl die genaue Verbreitung von A andreniformis als auch ihre Biologie sind Woyke J, Ruttner H (1958) An anatomical study noch unbekannt.