Zoomorphology (2012) 131:303–324 DOI 10.1007/s00435-012-0163-2

ORIGINAL PAPER

A comparative morphology of the male genitalia of Aphididae (Insecta, Hemiptera): part 2

Karina Wieczorek • Bartosz J. Płachno • Piotr S´wia˛tek

Received: 30 November 2011 / Revised: 24 May 2012 / Accepted: 29 May 2012 / Published online: 26 June 2012 Ó The Author(s) 2012. This article is published with open access at Springerlink.com

Abstract The present study provides new data related to aphids and differing little from the hypothetical condition. the morphology of the male genitalia of Aphididae. The In dwarfish males of Anoeciinae, Thelaxinae, Hormaph- structure of the male genitalia of 39 species from 23 genera idinae, and Eriosomatinae, the miniaturization of the body of Aphididae was studied using light and scanning electron size affects on the modification of genitalia, mostly para- microscopy. In the species studied, the genitalia of males meres. However, the homology of non-modified and consist of a phallus composed of the sclerotized basal part modified structures of parameres is not clear. with its articulation and a membranous apical part—an aedeagus as well as parameres. This state probably repre- Keywords External reproductive system Á Parameres Á sents the hypothetical plesiomorphic condition of the Phallus Á Insects Á SEM external male genitalia of aphids. According to the results of the present study, the male genitalia vary among sub- families (the most varied in Lachninae). Both the phallus Introduction and parameres show great variability in their form and the number of setae and may provide characters of taxonomic The male genitalia have proven to be of considerable tax- and diagnostic importance. The shape, size, and modifi- onomic value for various groups of insects. This paper cation of parameres are considered in conjunction with the continues our studies into comparative morphology and phylogenetic relationships among the studied taxa. Com- ultrastructure of the internal and external male genitalia of pared with Lachninae, Greenideinae, Aiceoninae, the Aphididae (Insecta, Hemiptera) (Wieczorek 2006, 2008; external genitalia of Aphidinae are less specialized, having Wieczorek and S´wia˛tek 2008, 2009; Wieczorek et al. 2011). many features in common with those of drepanosiphine In comparison with other hemipterans, the external male genitalia of aphid species are characterized by simplicity; however, they are well developed and typically consist of Communicated by T. Bartolomaeus. the phallus composed of the sclerotized basal part with its articulation and a membranous apical part—the aedeagus. & K. Wieczorek ( ) Laterally and above the phallus, there is a pair of setose Department of Zoology, University of Silesia, Bankowa 9, 40-007 Katowice, Poland parameres. In the first part of our studies devoted to external e-mail: [email protected] male genitalia (Wieczorek et al. 2011), the general mor- phologies of these structures were described and compared B. J. Płachno in those representatives of the Aphididae in which the Department of Plant Cytology and Embryology, Jagiellonian University, Grodzka 52, 31-044 Cracow, Poland genitalia are not modified—Drepanosiphinae, Chaitophor- e-mail: [email protected] inae, Calaphidinae, Phyllaphidinae, Saltusaphidinae, Lize- riinae, Spicaphidinae, Tamalinae, Parachaitophorinae, ´ P. Swia˛tek Phloeomyzinae, and Aphidinae. Department of Animal Histology and Embryology, University of Silesia, Bankowa 9, 40-007 Katowice, Poland In the present study, strongly modified structures of the e-mail: [email protected] male genitalia in dwarfish males of the Mindarinae, 123 304 Zoomorphology (2012) 131:303–324

Hormaphidinae, Anoeciinae, Thelaxinae, and Eriosomati- DMR light microscopes. Drawings were made with a nae as well as in normal-sized males of the Greenideinae, camera lucida. A magnified view is provided for each of Aiceoninae, and Lachninae are described. To date, the the photographs and drawings. external male genitalia of species belonging to these groups The procedure for preparing samples for SEM was as have only been studied in a limited number of species [e.g., described earlier (Płachno and S´wia˛tek 2009, 2010). Eriosoma lanigerum (Hausmann, 1802) (Baker 1915;de Briefly, whole specimens were fixed with 2.5 % glutaral- Fluiter 1931); Thelaxes dryophila (Schrank, 1801) (Pola- dehyde in a 0.1 M phosphate buffer (pH 7.4) for several szek 1987a); Greenidea (Trichosiphum) anonae (Pergande, days or fixed in 70 % ethanol. Later, the samples, which 1906) (Chaudhuri 1956); Aiceona japonica Takahashi, had been dehydrated in ethanol as well as an acetone series,

1960 (Takahashi 1960); Stomaphis species (Sorin 1965); were critical-point dried in liquid CO2 and coated with gold Cinara species (Eastop 1972); Eutrichosiphum assamense using a JEOL-JFC 1100E sputter coater. The specimens A. K. Ghosh, R. C Basu & D. N. Raychaudhuri, 1969 were viewed using a HITACHI S-4700 microscope (Chakrabarti and Maity 1980)]. They have usually been (Scanning Microscopy Laboratory of Biological and Geo- poorly described or simply illustrated. Only Polaszek logical Sciences, Jagiellonian University) at 20 kV. (1987b in lit.) provided a comprehensive study of the male genitalia of the entire family Aphididae, using numerous examples from most of these subfamilies. Results In this study, we present the general morphologies of the modified male genitalia of selected species of Aphididae, Morphology mainly the variability of parameres in their form and location, and consider these structures in conjunction with The external male genitalia of Aphididae consist of the the phylogenetic relationships among the entire family. phallus, which is composed of the partially sclerotized basal part with its articulation (sclerotized arms) and a membranous apical part, the aedeagus, as well as para- Materials and methods meres. These structures are defined in the following way. The phallus is the median intromittent organ as a whole. Taxon sampling The basal part of the phallus is a pair of partially sclerotized, various-shaped lobes which enfold the aedeagus and take part The structure of the male genitalia of 39 species belonging to in everting and maintaining it in position during copulation. the Mindarinae, Hormaphidinae, Anoeciinae, Thelaxinae, The sclerotized arms of the basal part of the phallus are a pair of Eriosomatinae, Greenideinae, Aiceoninae, and Lachninae sclerotized projections divided into distal and proximal part, was studied using light (LM) and scanning electron micros- giving support for the basal part of the phallus. copy (SEM). All of the species studied were borrowed from The aedeagus is a membranous, copulatory part of the the Muse´um national d’Histoire naturelle, Paris, France, with phallus, which is withdrawn within the body and everted the exception of Pachypappa tremulae (Linne, 1761), Proc- during copulation. iphilus fraxini (Fabricius, 1777), Anomalosiphum mendeli The parameres is usually a pair of various-shaped, setose Quednau & Martin 2006, and Schoutedenia ralumensis ventral processes positioned at the anterior end of the genital Ru¨bsaamen, 1905, which were borrowed from the Natural area. In some species of Aphididae, the parameres are History Museum, London, United Kingdom. The adult males strongly modified: completely fused into a single, sharply of Glyphina betulae (Linne, 1758), Lachnus roboris (Linne, pointed structure (Anoeciinae, Thelaxinae), reduced to 1758) and nymphs of Stomaphis quercus (Linne, 1758) were elongated projections supported by additional sclerotization collected in Poland in the years 2008–2011. Because of the (Eriosomatinae), divided into lobate parts which arise into rarity of males, sample sizes consisted of 1–10 individuals. various-shaped projections toward base of the phallus The genitalia of 39 species were studied from aphid material (Greenideinae, Aiceoninae and partially Lachninae). preserved in alcohol and slide-mounted specimens (LM); 13 The general morphologies of these structures of selected species were also observed using SEM techniques. The col- species of Aphididae were described and individual species lection data and the microscopic techniques used are sum- were then studied systematically (the classification of marized in Table 1. Aphidoidea is after Nieto Nafria et al. 1998)

Light and electron microscopy Mindarinae

Alcohol-preserved specimens and slides were examined Mindarus abietinus Koch, 1857 (male wingless, dwarf- and photographed using Nikon Eclipse 600 and Leica ish)—parameres are present, located above the basal part of 123 Zoomorphology (2012) 131:303–324 305

Table 1 Collection data for species included in the study Subfamily/tribe/species Collection locality Host plants Number of LM SEM studied specimens

Mindarinae Mindarus abietinus Koch, 1857 Brouvelieures, France Abies pectinata 10 ?? M. japonicus Takahashi, 1931 Osaka, Japan Cephalotaxus drupacea 2 ? Hormaphidinae: Nipponaphidini Neothoracaphis yanonis (Matsumura, 1917) Matsuzaka, Japan Distylium sp. 3 ? Nipponaphis distyliicola Monzen, 1934 Ise, Japan Distylium racemosum 1 ? Monzenia globuli (Monzen, 1934) Osaka, Japan Distylium sp. 4 ? Anoeciinae Anoecia vagans (Koch, 1856) Ailli, France Cornus sanguinea 1 ?? A. corni (Fabricius, 1775) Le Fayet, France Cornus sp. 10 ?? Thelaxinae Glyphina betulae (Linne, 1758) Katowice, Poland Betula sp. 10 ?? G. jacutensis Mordvilko, 1931 St. Etienne-de-Tinee, France Alnus incana 1 ? Eriosomatinae: Eriosomatini Tetraneura (Tetraneurella) Pont de l’Isere Ulmus sp. 2 ?? nigriabdominalis (Sasaki, 1899) Drome, France Eriosomatinae: Pemphigini Pemphigus versicarius Passerini, 1861 Orange, France Colutea arborescens 4 ?? Pachypappa tremulae (Linne, 1761) Aspley Heath, UK Picea abies 2 ? Prociphilus fraxini (Fabricius, 1777) Istanbul, Turkey Fraxinus sp. 9 ? P. ligustrifoliae (Tseng&Tao, 1938) Hakusan, Japan Ligustrum lucidum 4 ? Greenideinae: Greenideini Greenidea (Trichosiphum) carpini Tokyo, Japan Carpinus sp. 3 ? Takahashi, 1963 G. (T.) okajimai Suenaga, 1943 Osaka, Japan Shiia sp. 2 ? Eutrichosiphum sinense Kobe, Osaka, Japan Shiia sp. 6 ? D. N. Raychaudhuri, 1956 Mollitrochosiphum (Metatrichosiphon) Ise, Japan Meliosma rigida 1 ? yamabiwae Suenaga, 1943 Greenideinae: Cervaphidini Anomalosiphum mendeli Sabah, Borneo, Malaysia – 1 ? Quednau&Martin 2006 Greenideinae: Schoutedeniini Schoutedenia ralumensis Ru¨bsaamen, 1905 Mosman, Australia Breynia oblongifolia 1 ? Aiceoninae Aiceona japonica Takahashi, 1960 Kasugayama, Japan Cinnamomum camphora 1 ? Lachninae: Lachnini Lachnus pallipes (Hartig, 1841) Bouvieres, France Quercus sp. 10 ?? L. roboris (Linne, 1758) Katowice, Poland Quercus sp. 10 ?? Pterochloroides persicae Esfahan, Iran Prunus sp. 1 ? (Cholodkovsky, 1899) Stomaphis aceris Takahashi, 1960 Shikoku, Japan Acer sp. 4 ? S. graffi Cholodkovsky, 1894 France Quercus sp. 2 ? S. pini Takahashi, 1920 Mt. Shimaji, Japan Pinus densiflora 2 ? S. quercus (Linne, 1758) La Guyarde, France Juglans regia 10 ?? Katowice, Poland Quercus sp. S. yanonis Takahashi, 1918 Osaka, Japan Celtis sinensis 1 ? Maculolachnus submacula (Walker, 1848) La Varenne, France Rosa sp. 8 ?? Schizolachnus piniradiatae (Davidson, 1909) Orono, USA Pinus sp. 5 ?

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Table 1 continued Subfamily/tribe/species Collection locality Host plants Number of LM SEM studied specimens

Lachninae: Eulachnini Eulachnus agilis (Kaltenbach, 1843) Haute Alpes, France Pinus uncinata 10 ?? E. rileyi (Williams, 1911) Haute Alpes, France Pinus nigra 10 ?? Cinara cedri Mimeur, 1936 La Varenne, France Cedrus atlantica 6 ?? C. laricis (Hartig, 1839) Haute Alpes, France Larix europaea 2 ? C. maculipes Hille Ris Lambers 1966 Mouree, Pakistan Pinus excelsa 7 ? C. piceicola (Cholodkovsky, 1896) Haute Savoie, France Picea excelsa 7 ? C. todocola (Inouye, 1936) Tokyo, Japan Pinus sp. 3 ? C. (Cedrobium) laportei (Remaudiere, 1954) Ifrane, Marocco Cedrus atlantica 4 ? the phallus, pale, small, basally fused, elongated, with a phallus, strongly modified, completely fused into a single few setae on the outer margin (Fig. 1a). The basal part of triangular, sharply pointed structure, without setae on its tip the phallus in slide-mounted specimens (Fig. 1a) and (Fig. 2a). The basal part of the phallus is clearly visible, scanning electron microscope (Fig. 1b) is clearly visible, pale, oar-like, without setae. It is supported by rather long pale, triangular. Sclerotized arms are dark pigmented, with and thin sclerotized arms (Fig. 2b). The aedeagus is short a long distal part and a very short proximal part. The and oval shaped. aedeagus is rather short and oval in shape (Fig. 1b). Anoecia corni (Fabricius, 1775) (male wingless, M. japonicus Takahashi, 1931 (male wingless, dwarf- dwarfish)—external genitalia are very similar to those of ish)—external genitalia are very similar to those of M. abi- A. vagans, with less sharply pointed modified parameres etinus with a club-shaped the basal part of phallus (Fig. 2c). (Fig. 1c). Thelaxinae Hormaphidinae Glyphina betulae (Linne, 1758) (male wingless, dwarf- Neothoracaphis yanonis (Matsumura, 1917) (male wing- ish)—parameres are present, located above the basal part less, dwarfish)—parameres are located above the basal part of the phallus, strongly modified and fused into a single, of the phallus, very small and hardly visible in slide- dark, bulb-shaped structure with a few, short setae on its mounted specimens, with a few, short setae on the outer tip (Fig. 2d). The basal part of the phallus is present, but margin. The basal part of the phallus is clearly visible, pale, strongly reduced, dusky, triangular, without setae, visible and forceps-like, without setae. The sclerotized arms are both in the scanning electron microscope (Fig. 2e) and in rather short, of a similar length to the proximal and distal the slide-mounted specimens (Fig. 2f). Sclerotized arms parts. They are very thin and pale (Fig. 1d). The aedeagus are barely visible. The aedeagus is rather short with its is not visible. distal part wider than its basal part (Fig. 2f). Nipponaphis distyliicola Monzen, 1934 (male wingless, Glyphina jacutensis Mordvilko, 1931 (male wingless, dwarfish)—parameres are hardly visible. The basal part of dwarfish)—external genitalia are very similar to those of phallus is clearly visible, pale and lobe-shaped, without G. betulae with a barely visible the basal part of the setae (Fig. 1e). The sclerotized arms and the aedeagus are phallus. not visible. Monzenia globuli (Monzen, 1934) (male wingless, Eriosomatinae dwarfish)—parameres are hardly visible. The basal part of the phallus is clearly visible, pale and claw-like, without Tetraneura (Tetraneurella) nigriabdominalis (Sasaki, setae (Fig. 1f). The sclerotized arms and the aedeagus are 1899) (male wingless, dwarfish)—parameres are reduced not visible. to dusky, lobate projections (Fig. 3a). The basal part of the phallus is dusky, flattened and short, without setae Anoeciinae (Fig. 3b). The sclerotized arms are dusky, rather short, of a similar length to the proximal and distal parts. Anoecia vagans (Koch, 1856) (male wingless, dwarfish)— The most significant character of external genitalia, vis- parameres are present, located above the basal part of the ible in the slide-mounted specimens, is the additional

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Fig. 1 External genitalia of males. a, b Mindarus abietinus. a ventral distyliicola lateral view (light microscopy). f Monzenia globuli dorsal view (light microscopy), b lateral view (scanning electron micros- view (light microscopy). a aedeagus, bp basal part of phallus with copy). c M. japonicus ventral view (light microscopy). d Neothora- sclerotized arms consists of short proximal (solid arrow) and long caphis yanonis ventral view (light microscopy). e Nipponaphis distal (dotted arrow) part, p parameres

sclerotization of parameres projections reaching up to the similar length to the proximal and distal parts (Fig. 3c). VI abdominal sternite. This structure is formed by The aedeagus is not visible. elongated, parallel arms that are fused together along the Pachypappa tremulae (Linne, 1761) (male wingless, entire length (Fig. 3a). The aedeagus is short and oval dwarfish)—external genitalia are similar to those of Pem- shaped. phigus versicarius with much shorter projections of para- Pemphigus versicarius Passerini, 1861 (male wingless, meres and few short setae on the basal part of the phallus dwarfish)—parameres are reduced to pale, very long fin- (Fig. 3e). ger-like projections. Additional sclerotization of parameres Prociphilus fraxini (Fabricius, 1777) (male wingless, projections is formed by elongated, parallel arms that are dwarfish)—parameres are reduced to dusky, triangular fused together along the entire length. They reach up to the projections. Additional sclerotization of parameres projec- VIII abdominal sternite (Fig. 3c). The basal part of the tions, reaching up to the VIII abdominal sternite, is formed phallus is clearly visible, pale and lobate, without setae by elongated, separate, parallel arms that are fused together (Fig. 3d). The sclerotized arms are dusky, rather short, of a on the tip. The basal part of the phallus is clearly visible,

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Fig. 2 External genitalia of males. a, b Anoecia vagans. a ventral microscopy), e lateral view (scanning electron microscopy), f lateral view (scanning electron microscopy), b dorsal view (light micros- view (light microscopy). a aedeagus, bp basal part of phallus with copy). c Anoecia corni ventral view (scanning electron microscopy). sclerotized arms consists of short proximal (solid arrow) and long d, e, f Glyphina betulae. d ventral view (scanning electron distal (dotted arrow) part, c cauda, fp fused parameres dusky and flattened, without setae. The sclerotized arms are of the phallus. Lobate parts of parameres are basally fused. dusky, rather short, of a similar length to the proximal and The outer margin of lobate parts of parameres is covered by distal parts (Fig. 3f). The aedeagus is not visible. numerous, long setae, whereas on projections, setae are P. ligustrifoliae (Tseng & Tao, 1938) (male wingless, short and distributed on their apices. The basal part of the dwarfish)—external genitalia are similar to those of Proc- phallus is rather long, pale, sclerotized on the outer mar- iphilus fraxini with finger-like projections of parameres. gins, without setae. The sclerotized arms are dusky, rather The aedeagus is short and bulbous. short, of a similar length to the proximal and distal parts (Fig. 4a). The aedeagus is not visible. Greenideinae G. (T.) okajimai Suenaga, 1943 (male winged)—exter- nal genitalia are similar to those of Greenidea (Trichosip- Greenidea (Trichosiphum) carpini Takahashi, 1963 (male hum) carpinii with wider and much more setose lobate winged)—parameres are present, located above the basal parts of parameres and wider the basal part of the phallus. part of the phallus, pale, clearly divided into lobate parts Eutrichosiphum sinense D. N. Raychaudhuri, 1956 which arising into short, finger-like projections toward base (male winged)—parameres are present, clearly visible,

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Fig. 3 External genitalia of males. a, b Tetraneura (Tetraneurella) f Prociphilus fraxini ventral view (light microscopy). bp basal part of nigriabdominalis. a ventral view (light microscopy), b lateral view phallus with sclerotized arms consists of short proximal (solid arrow) (scanning electron microscopy). c, d Pemphigus versicarius. c ventral and long distal (dotted arrow) part, ap parallel arms of parameres view (light microscopy), d lateral view (scanning electron micros- projections, pp parameres projections copy). e Pachypappa tremulae ventral view (light microscopy). pale, basally fused, free at their finger-like projections. setae. The sclerotized arms are dusky, rather short, of a Lobate parts are setose in comparison with their projec- similar length to the proximal and distal parts (Fig. 4c). tions. The basal part of the phallus is rather short, pale, The aedeagus is not visible. without setae, fused, together forming an almost complete Anomalosiphum mendeli Quednau & Martin 2006 (male cylinder. The sclerotized arms are dusky, rather short, of a winged)—parameres are present, clearly visible, pale, similar length to the proximal and distal parts (Fig. 4b). basally fused, free only at their very short, finger-like The aedeagus is not visible. projections. They are almost hairless, having only a few Mollitrochosiphum (Metatrichosiphon) yamabiwae short setae on the outer margin of the lobate parts of Suenaga, 1943 (male winged)—parameres are present, parameres. The basal part of the phallus is rather short and clearly visible, dusky, basally fused, free at their finger-like flattened, pale, without setae. The sclerotized arms are projections. In comparison with the above-mentioned rather short, of a similar length to the proximal and distal species, both the lobate parts of parameres and their pro- parts (Fig. 4d). The aedeagus is not visible. jections are setose and much more robust. The basal part of Schoutedenia ralumensis Ru¨bsaamen, 1905 (male win- the phallus is long, flattened, hook-shaped, pale, without ged)—parameres are present, clearly visible, basally fused,

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Fig. 4 External genitalia of males. a Greenidea (Trichosiphum) view. f Aiceona japonica ventral view (light microscopy). bp basal carpini ventral view. b Eutrichosiphum sinense dorsal view. c Mol- part of phallus with sclerotized arms consists of short proximal (solid litrochosiphum (Metatrichosiphon) yamabiwae ventral view. d Anom- arrow) and long distal (dotted arrow) part, lp lobate parts of alosiphum mendeli ventral view. e Schoutedenia ralumensis ventral parameres, pp parameres projections pale, with the lobate parts arising into sharply pointed shaped, without setae. The sclerotized arms are clearly projections toward the base of the phallus. The projection visible and dusky with a long distal part and a short is short and hairless in comparison with the setose lobate proximal part (Fig. 4f). The aedeagus is not visible. part of parameres. The basal part of the phallus is short, hook-shaped, pale, without setae. The sclerotized arms are Lachninae barely visible (Fig. 4e). The aedeagus is not visible. Lachnus pallipes (Hartig, 1841) (male wingless)—para- Aiceoninae meres are present, clearly visible, separate, semicircular, dusky on the outer margin, with numerous, long setae Aiceona japonica Takahashi, 1960 (male winged)—para- mostly distributed on the outer margin (Fig. 5a). In com- meres are present, located above the basal part of the parison with the above-mentioned species, parameres are phallus, clearly visible, separate, pale, setose, with the positioned laterally to the basal part of the phallus lobate parts arising into sharply pointed projections toward (Fig. 5b). The basal part of the phallus is short, finger-like, the base of the phallus. The projection is short and hairless. dark pigmented, with only a few rather long setae (Fig. 5a). The basal part of the phallus is rather long, pale, hook- The sclerotized arms are of a similar length to the proximal

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Fig. 5 External genitalia of males. a, b, c Lachnus pallipes. a lateral view (light microscopy). f Pterochloroides persicae ventral view view (scanning electron microscopy), b dorsal view (scanning (light microscopy). a aedeagus, bp basal part of phallus with electron microscopy), c ventral view (light microscopy). d, e Lachnus sclerotized arms consists of short proximal (solid arrow) and long roboris. d ventral view (scanning electron microscopy), e ventral distal (dotted arrow) part, p parameres and distal parts, robust and clearly visible. Distal part ends dusky on the outer margin, with numerous, rather short in forked apices (Fig. 5c). The aedeagus is short and bul- setae on the entire surface. The basal part of the phallus is bous (Fig. 5a). short, flattened, oval paddle-shaped, dusky, with setae. The Lachnus roboris (Linne, 1758) (male winged)—para- sclerotized arms are clearly visible, very short, of a similar meres are present, located laterally to the basal part of the length to the proximal and distal parts (Fig. 5f). The ade- phallus, clearly visible, separate, semicircular, pale, with agus is not visible. numerous, long setae on the entire surface (Fig. 5d). The Stomaphis aceris Takahashi, 1960 (male wingless, basal part of the phallus is wide, pale, without setae dwarfish)—parameres are present, located above the basal (Fig. 5e). Sclerotized arms are similar to those of the part of the phallus, clearly visible, separate, pale, divided above-mentioned species but much wider. The aedeagus is into lobate parts arising into a finger-like projection toward short and bulbous (Fig. 5d). the base of the phallus. The projection is very thin and long Pterochloroides persicae (Cholodkovsky, 1899) (male (the longest among the species of the genus Stomaphis winged)—parameres are present, located above the basal studied and twice as long as the basal part of the phallus) part of the phallus, clearly visible, separate, lobe-shaped, and almost hairless in comparison with the lobate part of

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Fig. 6 External genitalia of males. a Stomaphis aceris ventral view microscopy), f nymph ventral view (light microscopy). a aedeagus, bp (light microscopy). b S. pini dorsal view (light microscopy). c, d, e, basal part of phallus with sclerotized arms consists of short proximal f Stomaphis quercus. c ventral view (scanning electron microscopy), (solid arrow) and long distal (dotted arrow) part, p parameres, lp d ventral view (light microscopy), e lateral view (scanning electron lobate parts of parameres, pp parameres projections paramere. The basal part of the phallus is short, finger-like, Stomaphis pini Takahashi, 1920 (male wingless, dark pigmented, sclerotized on inner margin, slightly dwarfish)—projections of parameres are very short, wide, setose. The sclerotized arms are clearly visible, of a similar setose, as long as the basal part of the phallus. The basal length to the proximal and distal parts, both very short, part of the phallus is thin and flattened, dark pigmented, dark pigmented (Fig. 6a). The adeagus is not visible. sclerotized on outer margin and slightly setose. The Stomaphis graffi Cholodkovsky, 1894 (male wingless, sclerotized arms are rather short, of a similar length to the dwarfish)—external genitalia are similar to those of the proximal and distal parts (Fig. 6b). The aedeagus is not above-mentioned species with a short and thin projection visible. (2 times shorter than the basal part of the phallus). The Stomaphis quercus (Linne, 1758) (male wingless, basal part of the phallus is long, finger-like, setose only on dwarfish)—projections of parameres are finger-like, rather apices. The aedeagus is not visible. Nymphs with clearly short and wide, covered by much shorter setae than the visible, broadly rounded buds of parameres situated in the lobate part of paramere (Fig. 6c). The basal part of the genital area. phallus is rather short, flattened, club-shaped, dusky,

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Fig. 7 External genitalia of males. a, b Maculolachnus submacula microscopy). a aedeagus, bp basal part of phallus with sclerotized a ventral view (light microscopy), b lateral view (scanning electron arms consists of short proximal (solid arrow), long distal (dotted microscopy). c Schizolachnus piniradiatae ventral view (light arrow) part and upper half-circle-shaped structure that surrounds the microscopy), d, e Eulachnus agilis d lateral view, e ventral view genital area (arrow-head), lp lobate parts of parameres, pp parameres (light microscopy). f Eulachnus rileyi ventral view (scanning electron projections sclerotized on inner margin, with numerous, long setae robust, paddle-shaped, dusky, with a few short setae. The (Fig. 6c). The sclerotized arms are dark pigmented and distal part of sclerotized arms is rather short and thin, and much more robust than in the above-mentioned species proximal part is robust, strongly sclerotized, and dark (Fig. 6d). The aedeagus is short (Fig. 6e). Nymphs with pigmented (Fig. 7a). The aedeagus is short and bulbous clearly visible, broadly rounded buds of parameres situated (Fig. 7b). in the genital area (Fig. 6f). Schizolachnus piniradiatae (Davidson, 1909) (male Stomaphis yanonis Takahashi, 1918 (male wingless, winged)—parameres are present, located above the basal dwarfish)—external genitalia are similar to those of part of the phallus, clearly visible, basally separate, divided S. aceris with shorter, paler and much more setose pro- into lobate parts arising into long, finger-like projection jections of parameres. toward the base of the phallus. The parameres are dusky, Maculolachnus submacula (Walker, 1848) (male wing- with numerous, long setae on the entire surface. The basal less)—parameres are present, located above the basal part part of the phallus is short, flattened, dusky, and setose. The of the phallus, clearly visible, dusky, with separate, setose sclerotized arms are clearly visible, dark pigmented with lobate parts and their projections fused into a single, tri- proximal part robust and ends in rounded apices. The distal angular structure. The basal part of the phallus is long, part is thinner and forms the upper half-circle-shaped

123 314 Zoomorphology (2012) 131:303–324 structure that surrounds the genital area (Fig. 7c). The proximal part is robust and ends in oval apex. The distal aedeagus is not visible. part is thinner and forms the upper half-circle-shaped Eulachnus agilis (Kaltenbach, 1843) (male winged)— structure that surrounds the genital area (Fig. 7e). The parameres are present, located above the basal part of the aedeagus is not visible. phallus, clearly visible, basally fused. Their lobate parts Eulachnus rileyi (Williams, 1911) (male winged)— (triangular in lateral view Fig. 7d) arise into a distinct, external genitalia are similar to those of E. agilis with a finger-like projection toward the base of the phallus. The thinner basal part of the phallus. The aedeagus is short and parameres are dark pigmented on the outer margin, with bulbous (Fig. 7f). numerous, long setae on the entire surface with the Cinara cedri Mimeur, 1936 (male winged)—parameres exception of a few, short setae distributed on the projec- are present, located above the basal part of the phallus, tions. The basal part of the phallus is short, flattened, pale, clearly visible, dark pigmented, with setose lobate parts with a few short setae. The sclerotized arms are clearly fused and arising into a finger-like projections toward base visible, strongly sclerotized, and dark pigmented. The of the phallus (Fig. 8a). The projection is long (twice as

Fig. 8 External genitalia of males. a, b Cinara cedri a ventral view (light microscopy). bp basal part of phallus with sclerotized arms (scanning electron microscopy), b dorsal view (light microscopy). consists of short proximal (solid arrow), long distal (dotted arrow) c Cinara maculipes ventral view (light microscopy). d Cinara part and upper half-circle-shaped structure that surrounds the genital piceicola ventral view (light microscopy). e Cinara todocola ventral area (arrow-head), lp lobate parts of parameres, pp parameres view (light microscopy). f Cinara (Cedrobium) laportei ventral view projections 123 Zoomorphology (2012) 131:303–324 315 long as the basal part of the phallus) and covered with characteristic mainly for taxa of a normal size (i.e., not numerous, long setae. The basal part of the phallus is rather dwarfish) and mostly for winged males. The present study short, flattened, dark pigmented, and slightly setose. The shows that in almost all of the species examined (repre- sclerotized arms are clearly visible, strongly sclerotized, sentatives of eight taxa traditionally classified as subfami- and dark pigmented. The proximal part is robust and ends lies within 24 subfamilies of the Aphididae), the external in oval apex. The distal part is thinner and forms the upper male genitalia are modified in both dwarfish and normal- half-circle-shaped structure that surrounds the genital area sized males (winged and wingless). (Fig. 8b). The aedeagus is not visible. Cinara laricis (Hartig, 1839) (male winged)—external The external genitalia of males—the phallus genitalia are similar to those of C. cedri with shorter and paler projections of parameres. In Aphididae, the phallus is composed of a membranous Cinara maculipes Hille Ris Lambers 1966 (male win- part—the aedeagus and the sclerotized basal part supported ged)—projections of parameres are sharply pointed, trian- by its articulations. The aedeagus is entirely unsclerotized gular, setose, dark pigmented (Fig. 8c). The basal part of and its shape is estimated mostly based on material pre- the phallus is slightly visible, in lateral view almost roun- served in alcohol or with scanning electron microscope. In ded with more sclerotized outer margin. The aedeagus is most of the species studied, it is clearly visible, although it not visible. is rather short and oval shaped (Lachnus pallipes Fig. 5a Cinara piceicola (Cholodkovsky, 1896) (male wing- and Stomaphis quercus Fig. 6e). Only in Glyphina betulae less)—lobate parts of parameres are broadly rounded, is the distal part of the aedeagus bulbous and wider than its setose, and the projections of parameres are very long, proximal part (Fig. 2f). Polaszek (1987b in lit.), according finger-like, setose and dark pigmented. The basal part of to the terminology of Bonhag and Wick (1953), distin- the phallus is clearly visible, very thin and long, almost guished the basal part of the aedeagus—a conjuctiva and its parallel, dark pigmented, setose. The sclerotized arms are distal part—a vesica in some species. The present study clearly visible. The proximal part is robust, dark pigmented does not support this division. Generally, in the species and ends in spiniform apices. The distal part is much studied, the aedeagus (when visible) is characterized by a thinner and forms the upper half-circle-shaped structure small amount of variation in its shape in comparison with that surrounds the genital area (Fig. 8d). The aedeagus is previously studied species of Drepanosiphinae, Calaphidi- not visible. nae, Chaitophorinae or Aphidinae. Cinara todocola (Inouye, 1936) (male winged)—lobate The basal part of the phallus (the so-called penis valves parts of parameres are crescent-shaped, and the projections by some authors), which is composed of the partially of parameres are short and sharply pointed, dark pig- sclerotized lobes, is present and clearly visible in all of the mented, setose on the entire surface. The basal part of the species studied. Only in the dwarfish males of Glyphina phallus is hook-shaped, short, pale, without setae. The betulae is it reduced to small, triangular plates (Fig. 2e). proximal part of sclerotized arms is robust, dark pigmented Polaszek (1987b in lit.) stated that in representatives of the and ends in a spiniform apices. The distal part is much genera Anoecia, Glyphina, Thelaxes, and Pemphigus (all thinner and forms the upper half-circle-shaped structure with dwarfish males), the basal part of the phallus is either that surrounds the genital area. (Fig. 8e). The aedeagus is absent, modified and fused or is replaced by an extension of not visible. the anal plate. The present study does not support this Cinara (Cedrobium) laportei (Remaudiere, 1954) (male observation. In all of the species with dwarfish males winged)—external genitalia are similar to those of C. cedri studied, representatives of the Mindarinae, Hormaphidinae, with slight projections of parameres and less robust scler- Anoeciinae, Thelaxinae and Eriosomatinae as well as otized arms (Fig. 8f). dwarf males of the genus Stomaphis (Lachninae), the basal part of the phallus is present and visible in the scanning electron microscope and the slide-mounted specimens. In Discussion most cases, the lobes are rather short and flattened (Nipp- onaphis distyliicola Fig. 1e, Pemphigus versicarius The previous study of the comparative morphology of the Fig. 3d), forceps-like (Neothoracaphis yanonis Fig. 1d, external male genitalia of Aphididae (Wieczorek et al. Monzenia globuli Fig. 1f), triangular (Mindarus abietinus 2011) showed that these structures are well developed and Fig. 1a, Anoecia vagans Fig. 2b) or finger-like (Stomaphis typically consist of the phallus, which is composed of the aceris Fig. 6a). Representatives of the Greenideinae, sclerotized basal part with its articulation and a membra- Aiceoninae, and Lachninae (in exception of the genus nous apical part, the aedeagus. Above the phallus, there is a Stomaphis) are characterized by normal-sized males. In pair of setose parameres. However, this scheme was most of species of the Greenideinae studied, the basal part 123 316 Zoomorphology (2012) 131:303–324 of the phallus is rather short and flattened (Anomalosiphum studied species of Drepanosiphinae, Calaphidinae, Chai- mendeli Fig. 4d) or hook-shaped (Mollitrochosiphum tophorinae, or Aphidinae. The dwarfish males of Hor- (Metatrichosiphon) yamabiwae Fig. 4c), usually without maphidinae (Neothoracaphis yanonis Fig. 1d) have a setae. However, the shape of basal part of the phallus in similar type of broadly rounded and barely visible para- Eutrichosiphum sinense differs from these structures in meres. On the other hand, parameres appear to be signifi- other aphid genera. The lobes are fused, free only at their cantly modified in the dwarfish males of the Anoeciinae, tips and together form an almost complete cylinder Thelaxinae, and Eriosomatinae. In representatives of the (Fig. 4b). Unfortunately, only slide-mounted males were genera Anoecia and Glyphina, parameres are fused into a available for Greenideinae so it was not possible to perform single, sharply pointed structure that is triangular without a detailed electron microscopy study of these structures. setae on the tip in Anoecia (Fig. 2a–c) or bulb-shaped with The basal part of the phallus of Aiceona japonica is similar few, short setae on the tip in Glyphina (Fig. 2d). In the to the hook-shape of the above-mentioned species of species of Eriosomatinae studied, parameres are reduced to Greenideinae; however, it is much longer (Fig. 4f). Lach- variable-shaped projections—mostly finger-like (Pemphi- ninae are characterized by the most various shapes of the gus versicarius Fig. 3c) but also lobate (Tetraneura basal part of the phallus. In general, the most common is (Tetraneurella) nigriabdominalis Fig. 3a) or triangular the finger-like shape (Lachnus roboris Fig. 5e, Cinara (Prociphilus fraxini Fig. 3f)—and are supported by addi- piceicola Fig. 8d) and rarely club-shaped (Stomaphis tional sclerotization that reaches up to the VI-VIII quercus Fig. 6d) or paddle-shaped (Maculolachnus sub- abdominal sternite. This structure has never been described macula Fig. 7a). It is joined laterally with the aedeagus and before and is formed by elongated, parallel arms fused attached to it (Figs. 5b, 6e). Moreover, the basal part of the together along the entire length (Fig. 3a) or fused only on phallus is more setose in Lachninae than in representatives the tip (Fig. 3f). Only de Fluiter (1931) called it a ‘‘ventrale of the other subfamilies. chitinestaaf’’ (a chitinized rod) and figured it in Eriosoma In all of the species studied, the basal part of the phallus lanigerum. As in a case of the sclerotized arms of basal part is additionally fortified by sclerotized arms. These struc- of the phallus, they are probably apophyses that provide tures are not uniform and vary in form and length. In rep- support for muscle attachment. In male insects, parameres resentatives of the Hormaphidinae, Anoeciinae (Anoecia are the grasping apparatus, which play an important role vagans Fig. 2b), Thelaxinae, Eriosomatinae (Pemphigus during copulation. It is possible that the modified para- versicarius Fig. 3c), and Greenideinae (Mollitrochosiphum meres of Eriosomatinae need additional support; however, (Metatrichosiphon) yamabiwae Fig. 4c), the sclerotized the mating behavior of these aphids has never been arms are barely visible, pale, rather short and of a similar observed. Greenideinae are also characterized by modified length to the proximal and distal parts. In Mindarinae parameres. They are clearly divided into lobate parts arise (Mindarus japonicus Fig. 1c) and Aiceoninae, the sclero- into short, usually finger-like projections toward the base of tized arms are clearly visible and dusky with a long distal the phallus. The lobate parts of parameres are fused basally part and a short proximal part. In Lachninae, the sclerotized and are free only at their projections. Polaszek (1987b in arms are the most variable: of a similar length to the lit.) stated that parameres of Greenideinae are reduced and proximal and distal parts in the genera Lachnus (Fig. 5c), only the structure he called the anterior genital process, Pterochloroides and Stomaphis (Fig. 6a) with a rather which forms a pair of finger-like processes, is developed. short and thin distal part and a robust proximal part in The present study does not support this observation. All of Maculolachnus submacula (Fig. 7a), or with a robust the species studied, especially representatives of the tribe proximal part ending in an oval apices and a much thinner Greneeidini (Fig. 4a–c), have large, well-developed para- distal part, which forms the upper half-circle-shaped meres that are clearly visible in the slide-mounted speci- structure that surrounds the genital area in the genera mens. Chaudhuri (1956) and Chakrabarti and Maity (1980) Schizolachnus (Fig. 7c), Eulachnus (Fig. 7e) and Cinara also described and illustrated parameres of Greneeidini as (Fig. 8b–f). Additionally, in all of the species of this composed of the base with lateral projections. Moreover, subfamily studied, the arms are heavily sclerotized, robust scanning electron micrographs of the external genitalia of and dark pigmented. males of Schoutedenia ralumensis presented by Polaszek (1987b in lit.) show that these structures are well devel- The external genitalia of males—parameres oped. Aiceona japonica (Aiceoninae) has a similar type of parameres as those found in Greenideinae (Fig. 4f). All of Parameres are the most variable part of the male external the mentioned species have parameres positioned above the genitalia of species studied. In Mindarus (Fig. 1a), which basal part of the phallus. The most diverse forms of para- has dwarfish males, parameres are rather small, fused meres are characterized for Lachninae. They are separate, basally and elongated, similar to those of previously semicircular or lobate without a finger-like projection in 123 Zoomorphology (2012) 131:303–324 317 the genera Lachnus (Fig. 5c, e) and Pterochloroides the aedeagus and the basal part of the phallus as well as the (Fig. 5f) (Lachnini) or divided into lobate parts (separate or length of the sclerotized arms is variable between different fused) arising into variable-shaped projections toward the genera and species, but varies very little within each species. base of the phallus in the remaining species of this sub- Thus, this characteristic is valuable for diagnosing species family that were studied. The projections of the lobate parts but is less important in phylogenetic considerations. of parameres (called the anterior genital process by Pola- szek 1987b in lit.) are variable between different genera Phylogenetic implications—what is important parameres and species, but they vary very little within each species. not modified or modified? Their shape and length is a very good diagnostic character for distinguishing males belonging to the different species On the other hand, parameres show the most striking of this subfamily. Those observed in the genera Stomaphis differences and appear to be highly variable among the (Lachnini—Fig. 6a, b, d) and Cinara (Eulachnini— different subfamilies studied. In Drepanosiphinae, Chaito- Fig. 8b–f) are the most variable. In Maculolachnus sub- phorinae, Calaphidinae, Phyllaphidinae, Saltusaphidinae, macula (Lachnini—Fig. 7a), the projections of the lobate Lizeriinae, Spicaphidinae, Tamalinae, Parachaitophorinae, parts of parameres are fused into a single, large, triangular Phloeomyzinae, and Aphidinae (Fig. 9a), parameres are structure. Moreover, nymphs of the genus Stomaphis have large and positioned at the anterior end of the genital area, clearly visible, broadly rounded buds of parameres situated above the basal part of the phallus (Wieczorek et al. 2011). in the genital area (Fig. 6f). Parameres of Lachninae are Similar types of parameres have been found in the dwarfish more setose than in the other subfamilies. In the genus males of the Mindarinae (Fig. 9b) and Hormaphidinae. The Lachnus they are positioned laterally to the basal part of the Mindarinae, represented by only one extant genus Minda- phallus (Lachnus pallipes Fig. 5b), whereas in the rus Koch, are considered to be the most primeval drepa- remaining species of this subfamily, parameres are located nosiphine aphids by some authors (Takahashi 1931; above the basal part of the phallus. Mordvilko 1934; Heie and We˛gierek 2009b; Quednau 2010) or are recognized as an independent subfamily (Heie The external genitalia of males—phylogenetic 1980; Remaudiere and Stroyan 1984). In comparison with implications other taxa with dwarfish males, parameres of M. abietinus and M. japonicus are not modified and are similar to those Comparative studies of male genitalia have formed the observed in drepanosiphine aphids. Thus, our study cor- basis of the taxonomic and phylogenetic studies of many roborates Quednau’s (2010) point of view, who claims that groups of insects (e.g., Zumpt and Heinz 1950; Eades this genus represents the most primordial group of the 2000). Males of most Aphididae, unlike males of other drepanosiphine aphids. The dwarfish males of Hormaph- Hemiptera, do not appear until autumn and have to mature idinae that were studied represent the same type of para- quickly. This is why the taxonomy and construction of the meres, although they are very small. According to Quednau higher classifications of this group of insects have gener- (2010), the Hormaphidinae together with Eriosomatinae ally been based on the morphology and the characters of and Anoeciinae form the common lineage. However, the females. However, a comparative, systematic study parameres of studied species belonging to these subfami- of the male genitalia of Aphididae has revealed a number lies differ significantly. In Eriosomatinae, parameres are of characters that are of potential use in discussions of the reduced to rather long projections, which are supported by phylogenetic relationships of this group of Hemiptera. additional sclerotization reaching up to the VI–VIII abdominal sternite (Fig. 9c). This unusual shape of para- Phylogenetic implications—why the phallus presents some meres may be regarded as the autapomorphy of this sub- stable characters. family. In Anoeciinae, on the other hand, parameres are fused into a single, sharply pointed structure, similar to that The phallus, which is composed of the sclerotized basal in Thelaxinae (Fig. 9d). It is not clear whether this for- part with its articulation and a membranous apical part— ward-pointing structure at the anterior end of the genital the aedeagus, presents some stable characters among all of area actually represents the parameres that have become the subfamilies studied, that is, with dwarfish, normal- fused, or whether it is derived from the fusion of para- sized, winged or wingless males. The stable character of meres’ projections or whether it has arisen in some other the components of the phallus is a result of their functions: way. According to the molecular studies of Ortiz-Rivas and the lateral sclerotization of the otherwise membranous Martinez-Torres (2010), Anoeciinae, Eriosomatinae, Hor- aedeagus enfolds it and supported by the sclerotized arms maphidinae, Mindarinae, and Thelaxinae are grouped takes part in everting the aedeagus and maintaining it in the together. Moreover, these subfamilies share a common right position during copulation. The shape and size of feature—the presence of a wingless, dwarfish sexual 123 318 Zoomorphology (2012) 131:303–324

Fig. 9 External genitalia of males. a Not modified in the Drepanos- fused into a single, sharply pointed structure in the Anoeciinae and iphinae, Chaitophorinae, Calaphidinae, Phyllaphidinae, Saltusaphid- Thelaxinae. a aedeagus, bp basal part of phallus with sclerotized arms inae, Lizeriinae, Spicaphidinae, Tamalinae, Parachaitophorinae, consists of short proximal (solid arrow) and long distal (dotted arrow) Phloeomyzinae and Aphidinae. b Not modified in the Mindarinae part, ap parallel arms of parameres projections, fp fused parameres, and Hormaphidinae. c Modified with parameres reduced to variable- p parameres, pp parameres projections. Drawings according to light shaped projections in the Eriosmatinae. d Modified with parameres microscopical preparations generation (feeding—Anoeciinae, Mindarinae, Thelaxinae, small and barely visible parameres in Hormaphidinae and Hormaphidinae or non-feeding—Eriosomatinae). How- very small the basal part of the phallus in Thelaxinae. ever, they do not share a common origin of host alternation Greenideinae (Fig. 10a) and Aiceoninae (Fig. 10b) are (dioecious—pemphigid host alternation—return of sexu- characterized by normal-sized males and large parameres parae to the primary host in Anoeciinae, Eriosomatinae, divided into lobate parts arising into short, usually finger- Hormaphidinae, and monoecious in Mindarinae and Thel- like projections toward the base of the phallus. The close axinae). Von Dohlen and Moran (2000) stated that host relations between these subfamilies were pointed out by alternation originated independently in the ancestors of Heie and We˛gierek (2009b); however, they remain un- these aphids. It is probable that the modifications in para- sampled for molecular data (von Dohlen 2009). According meres also arose independently. Moreover, reduction in the to Quednau and Martin (2006), the genus Aiceona Takah- size of sexual morphs may then have a direct effect on ashi may have evolved from Neophyllaphis-like ancestors, characters associated with external genitalia, that is, very which may also be the ancestors of primitive Lachninae. In

123 Zoomorphology (2012) 131:303–324 319

Fig. 10 External genitalia of males. a Modified with parameres Cinara, Eulachnus, Schizolachnus and f Stomaphis (Lachninae) with divided into lobate parts arising into finger-like projections in the parameres divided into lobate parts arising into finger-like projec- Greenideinae. b Modified with parameres divided into lobate parts tions. bp basal part of phallus with sclerotized arms consists of short arising into finger-like projections in the Aiceoninae. c Not modified proximal (solid arrow) and long distal (dotted arrow) part, lp lobate in genera Lachnus and Pterochloroides (Lachninae). d Modified in parts of parameres, p parameres, pp parameres projections. Drawings the genus Maculolachnus submacula (Lachninae) with parameres according to light microscopical preparations projections fused into single structure. e Modified in the genera fact, parameres of A. japonica are also very similar to those genus Lachnus, they are positioned laterally to the basal of some Cinara species. part of the phallus. In Maculolachnus submacula The most diverse forms of parameres, similar to almost (Fig. 10d), projections fused into a single structure, all of the types described above, are characterized for resemble those found in Anoeciinae and Thelaxinae, but Lachninae. Parameres of the genera Lachnus (Fig. 10c) and the lobate parts of parameres are large and well developed. Pterochloroides resemble those found in Aphidinae and In the genera Cinara (Fig. 10e), Eulachnus, Schizolachnus, drepanosiphine aphids in shape and size; however, in the and Stomaphis (Fig. 10f) (and in representatives of the

123 320 Zoomorphology (2012) 131:303–324 tribe Tramini Trama troglodytes von Heyden not studied Thelaxinae, Hormaphidinae and Eriosomatinae the minia- here (Blackman et al. 2001)), parameres divided into lobate turization of the body size affects on the modification of parts arising into projections, resemble those found in genitalia—mostly parameres. Various modified parameres Greenideinae and Aiceoninae; however, they are highly found in males of these four subfamilies do not show any diverse in the shape and size of their projections. Among strong similarites with each other (single structure in Aphididae, Lachninae are one of the most diverse groups in Anoeciinae and Thelaxinae, very small and barely visible host–plant association, ecological and life-history charac- in Hormaphidinae or reduced to elongated projections ters (Normark 2000) with males that are relatively small, supported by additional sclerotization in Eriosomatinae) but not dwarfish (in exception of the genus Stomaphis). and have developed in different way. It suggests that Whether the parameres of Lachninae are homologous with reduction in the size of sexual generation has occurred those found in the remaining subfamilies is not clear. independently in a number of distinct lineages within Because external male genitalia are composed of func- Aphididae. Whether the parameres of mentioned dwarf tionally different components (the phallus—a copulatory males are homologus with those found in the drepanosi- organ and parameres—the grasping apparatus), any unique phine aphids and Aphidinae is not clear. The question, combination of these components should be species-spec- whether they function as the grasping apparatus, is also ificity (Song 2009). Lachninae differ from other aphids, as open. It may therefore be that the parameres are non- many species may infest the same host plant. Thus, the functional. However, the mating behavior of these aphids highly variable genitalia of Lachninae could play an has never been observed. Compared with the drepanosi- important role in the context of sexual selection. According phine aphids and Aphidinae, the external genitalia of to the molecular studies of Ortiz-Rivas and Martinez- Lachninae (except for the genera Lachnus and Pterochlo- Torres (2010), Lachninae constitute an independent and roides), Aiceoninae, and Greenideinae are more special- basal lineage in Aphididae. However, both Heie and ized, that is, parameres are divided into lobate parts ended We˛gierek (2009a) and Quednau (2010) stated that Aice- by various-shaped projections. It is possible that the oninae, Greenideinae, Aphidinae, and Lachninae constitute extended region of parameres perform the same function, a complex of modern aphids. Our previous study (Wieczorek presumably that of gripping the female, as the parameres et al. 2011) showed that Aphidinae are characterized by do in other aphids. Although the character of parameres parameres that are not modified and are similar to those in the appears similar in Lachninae, Aiceoninae, and Greenidei- drepanosiphine aphids, whereas parameres of Lachninae nae, these subfamilies differ greatly otherwise. It seems (except for the genera Lachnus and Pterochloroides) likely that the modification of parameres have evolved resemble those in Aiceoninae and Greenideinae. independently in Lachninae as well as Aiceoninae and Greenideinae on the other hand. External genitalia—hypothetical plesiomorphic condition The most important characters of the external male and its modifications genitalia of Aphididae, which could be discussed in con- junction with the phylogenetic relationships among the The contribution of comparative morphology of male studied taxa, are compared in the Table 2. external genitalia, if studied alone, is limited and needs further morphological characters, when reconstructing the phylogeny. In addition, in the absence of data in the fossil Summary material, it is difficult to speculate which external genitalia characters should be interpreted as primitive (plesiomor- The males of most aphids, unlike the males of other phic) and which are derived. Aphids are characterized by Hemiptera, do not appear until autumn (or even early in the the ‘mosaic’ type of evolution—some features have summer) and have to mature quickly. Their mating evolved independently, several times, in various evolu- behavior characterized by many short copulations requires tionary lines, which makes the interpretation of data a certain degree of simplicity in the structure of the complicated (Heie and We˛gierek 2009a; Quednau 2010). reproductive system—both external and internal. The Probably, the hypothetical plesiomorphic condition of the external male genitalia of Aphididae are well developed external male genitalia of aphids is characterized by the and typically consist of the phallus, which is composed of rather large, lobate and setose parameres positioned at the the sclerotized basal part with its articulation and a mem- anterior end of the genital area as well as well developed branous apical part—the aedeagus as well as parameres. the basal part of the phallus which protects the aedeagus This state probably represents the hypothetical plesiomor- and is supported by the sclerotization. This type of external phic condition of the external male genitalia of aphids. The genitalia is found in most modern taxa of drepanosiphine phallus structure presents a stable character among all of aphids and Aphidinae. In dwarfish males of Anoeciinae, the subfamilies studied, whereas parameres appeared to be 123 omrhlg 21)131:303–324 (2012) Zoomorphology Table 2 The most important characters of the external male genitalia of Aphididae Subfamily/tribe/species 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31

Mindarinae Mindarus abietinus ?--??----??--?---?--??--?-?-??- Drepanosiphinae

Drepanosiphum platanoidis ?-?-?----??-?----?--??--?-?-?-? Chaitophorinae Chaitophorini Chaitophorus populeti ?-?-?----??-?----?--??-?--??--? Periphyllus coracinus ?-?-?----??-?----?-?-?--?-??--? Chaitophorinae Siphini Chaetosiphella stipae ?--??----??-?----?-?-?-?--?-??- Sipha (Rungsia) maydis ?--??----??-?----?-?--??--?-??- Laingia psammae ?--??----??-?----?-?-?-?--?-??- Calaphidinae Calaphidini Clethrobius comes ?-?-?----??-?----?--??--?-?-?-? Calaphidinae Panaphidini Appendiseta robiniae ?-?-?----??-?----?--??-?--?-?-? Mexicallis spinifer ?-?-?----??-?----?--??--?-?-??- Myzocallis (Lineomyzocallis) walshii ?-?-?----??--?---?--??--?-?-?-? Panaphis juglandis ?-?-?----??-?----?-?-?-?--?-?-? Phyllaphidinae Phyllaphis fagi ?-?-?----? ???----?-?-?-?--?-?-? Diphyllaphis mordvilkoi ?-?-?----?-??----?-?-?--?-?-??? Saltusaphidinae Thripsaphidini Subsaltusaphis flava ?-?-?----??--?---?-?-???--?-??? Lizeriinae Lizerius ocoteae ?-?-?----??-?----?--?-?-?-?-??? Spicaphidinae Neuquenaphis edwardsi ?-?-?----?-??----?--??--?-?-??? Tamalinae Tamalia sp. ?-?-?----?-??----?--?-?-?-?-?-? Parachaitophorinae Parachaitophorus yamashitai ?-?-?----??--?---?--??--?-??-?? Phloeomyzinae Phloeomyzus passerini ?-?-?----??-?----?--??-?--?-??? 123 Aphidinae Aphidini Aphis pomi ?-?-?----?-?--?--?-?-?--?-?-?-? 321 322 123 Table 2 continued Subfamily/tribe/species 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31

Aphidinae Macrosiphini Brachycaudus divaricatae ?-?-?----?-??----?--?-?-?-?-??- Cavariella saxifragae ?-?-?----?-??----?--??--?-?-?-? Hyperomyzus pallidus ?-?-?----?-?-?---?--?-?-?-?-??? Myzus cerasi ?-?-?----?-??----?--??--?-?-?-? Pterocomma populeum ?-?-?----?-??----?--??--?-?-?-? Hormaphidinae Nipponaphidini Neothoracaphis yanonis ?--??----? ??--?--?-?--??--?-??? Nipponaphis distyliicola ?--??----? ??--? --?-?--???-?-??? Monzenia globuli ?--??----? ??--? --?-?--???-?-??? Anoeciinae Anoecia vagans ?--?-?---?--?----?--?-??--?-??- Thelaxinae Glyphina betulae ?--?-?---?------???---???-?-??- Eriosomatinae Eriosomatini Tetraneura (Tetraneurella) ?-?---?-?--?-----?-?--??--?-??- nigriabdominalis Eriosomatinae Pemphigini Pemphigus versicarius ?-?---?-?--?-----?-?--??--?-??? Pachypappa tremulae ?-?---?-?--?-----?-?-?-?--?-??? Prociphilus fraxini ?-?---?-?--?-----?-?--??--?-??? Greenideinae Greenideini Greenidea (Trichosiphum) carpini ?-?----?-??---?--?--?-??--?-??? Eutrichosiphum sinense ?-?----?-??----?-?-?--??--?-??? Mollitrochosiphum (Metatrichosiphon) ?-?----?-??-?----?--?-??--?-??? yamabiwae

Cervaphidini 131:303–324 (2012) Zoomorphology Anomalosiphum mendeli ?-?----?-??---?--?-?--??--?-??? Schoutedeniini Schoutedenia ralumensis ?-?----?-??----?-?-?--???-?-??? Aiceoninae Aiceona japonica ?-?----?-?-?---?-?--?-?-?-?-??? Lachninae Lachnini Lachnus pallipes -??-?----?-?--?--?-?-?-?--?-??- Pterochloroides persicae ?-?-?----?-??----?-?-?-?--?-??? Stomaphis aceris ?-?----?-?-?---?-?-?-?-?--?-??? Zoomorphology (2012) 131:303–324 323 6

14 highly variable between the different subfamilies studied (the most varied in Lachninae). They are observed in taxa ?? ?? ?? ?? ?? with normal-sized (Aiceoninae, Greenideinae and Lachni- nae) and dwarfish males (Anoeciinae, Eriosomatinae and Thelaxinae) and could be useful in considerations of the relationships and phylogeny of aphids. However, the homology of non-modified and modified structures of distal parts of sclerotized parameres not modified, 28 5 parameres is not clear. In addition, both the phallus and basal part of the phallus with

22 parameres show the greatest variability in their form and

parameres covered by setae on the number of setae and may provide characters of taxonomic additional sclerotization of parameres 17 and diagnostic importance. 9

Acknowledgments We are sincerely grateful to Andrev Polaszek parameres small,

4 for access to his unpublished Ph. D. thesis concerning the compara- tive anatomy of the male aphid reproductive system. We would like to

parameres covered by setae on the whole surface, express our thanks to Georges Remaudie‘re, Daniele Matile-Ferrero

13 and Thierry Bourgoin, Museum national d’Histoire naturelle, Paris and to Jon Martin, the Natural History Museum London, for the loan basal part of the phallus long, of the material studied and to Łukasz Depa, University of Silesia, 21 parameres large, Katowice for his help in collecting the material of nymphs of S. 3 sclerotized arms: proximal part short and wide, distal part long and quercus. Special thanks go to Łukasz Junkiert, University of Silesia,

25 Katowice for preparing the drawings. B. J. Płachno gratefully

aedeagus long acknowledges the scholarship for Outstanding Young Scientists from parameres separate, 31 the Minister of Science and Higher Education. The authors are grateful the Reviewers and the Editor for all suggestions and com- 12 ments that improved the manuscript. parameres covered by setae on the lobate parts,

16 Open Access This article is distributed under the terms of the Creative Commons Attribution License which permits any use, dis-

aedeagus short, tribution, and reproduction in any medium, provided the original basal part of the phallus short,

30 author(s) and the source are credited. 20 parameres modified into lobate parts ended by projections, 8 distal parts of sclerotized arms without the upper half-circle-shaped structure, parameres fused basally, 27 11 References

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18 Heie OE (1980) The Aphidoidea (Hemiptera) of Fennoscandia and basal part of the phallus without setae, continued Denmark. I. General part. The families Mindaridae, Hormaph- distal parts of sclerotized arms with the upper half-circle-shaped structure,

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Character present, Fauna Ent. Scand., Scandinavian Science Press LTD, Klampen- Parameres positioned above the basal part of the phallus, C. (Cedrobium) laportei C. piceicola Cinara cedri Eulachnus agilis Maculolachnus submacula Schizolachnus piniradiatae S. quercus projections present, parameres covered by setae on the inner margin, parameres modified into single structure, projections, 1 ? setae, thin, arms with rounded processes, Lachninae Eulachnini Table 2 Subfamily/tribe/species 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19borg 20 21 9:1–235 22 23 24 25 26 27 28 29 30 31

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