Title: Morphological and histological study of the forewing of Aleyrodes proletella (Linnaeus 1758) (Sternorrhyncha, Hemiptera) with a comparative analysis of forewings among Sternorrhyncha infraorders Author: Barbara Franielczyk-Pietyra, Łukasz Depa, Piotr Wegierek Citation style: Franielczyk-Pietyra Barbara, Depa Łukasz, Wegierek Piotr. (2019). Morphological and histological study of the forewing of Aleyrodes proletella (Linnaeus 1758) (Sternorrhyncha, Hemiptera) with a comparative analysis of forewings among Sternorrhyncha infraorders. “Zoomorphology” (29 May 2019), doi 10.1007/s00435-019- 00449-1 Zoomorphology https://doi.org/10.1007/s00435-019-00449-1 ORIGINAL PAPER Morphological and histological study of the forewing of Aleyrodes proletella (Linnaeus 1758) (Sternorrhyncha, Hemiptera) with a comparative analysis of forewings among Sternorrhyncha infraorders Barbara Franielczyk‑Pietyra1 · Łukasz Depa1 · Piotr Wegierek1 Received: 16 January 2019 / Revised: 10 May 2019 / Accepted: 14 May 2019 © The Author(s) 2019 Abstract Identifcation of whitefies is based mainly on larval stages and generally very little is known about wings of these insects. Therefore, both sides of the forewings of Aleyrodes proletella were studied using histological methods, light and scanning electron microscopes. Studies confrm the occurrence of only three veins on forewings: the costal, radial and anal ones; only the anal vein lies under the anal fold. A campaniform and trichoid sensilla are present. The shape of wax secretions and wing margins is described. The comparative analysis of forewing structures contains new data for all Sternorrhyncha infraorders. The current results confrm the monophyly of the group, but place aphids closer to psyllids. The analysis of forewing base indicates that its general model is similar among Sternorrhyncha, but there occur some intergroup diferences. Ways of wing folding depend on the structure of thorax. Keywords Forewing morphology · Wing veins · Cross section · Aleyrodids · Sternorrhyncha Introduction representatives of Aleyrodidae are also named whitefies. They resemble tiny moths, which was probably the reason Sternorrhyncha is a Hemiptera suborder divided into four why Linnaeus in 1758 included A. proletella into that group infraorders: Psyllomorpha (jumping plant-lice), Aleyro- (Martin et al. 2000). Like other Sternorrhyncha representa- domorpha (whitefies), Aphidomorpha (aphids) and Coc- tives, whitefies are plant sap-sucking pests. Fossil inclu- comorpha (scale insects) (Heie and Wegierek 2009). Mor- sions can only be found in amber (Drohojowska and Szwedo phological studies have indicated that these hemimetabolous 2015; Szwedo and Drohojowska 2016). insects are a monophyletic group consisting of sister groups Whitefy identifcation keys have been based on the char- of aphids + coccoids and psyllids + aleyrodids (Hennig 1981; acteristics of larval stages (Hodges and Evans 2005). As a Carver et al. 1991). result, rarely have the imagines of whitefies been investi- The studied whitefy species, Aleyrodes proletella (Lin- gated. Sexual dimorphism among adult specimens is dem- naeus 1758) (Fig. 1a, b) is a small insect belonging to the onstrated in diferences in genitalia, the number of ventral Aleyrodidae family. This family comprises mostly extant abdominal wax plates and in a smaller size of males. These subfamilies (Evans 2007). Due to the strong wax covering features tend to be mentioned only in supplements to the keys describing the larval stages of aleyrodids (Hodges and Electronic supplementary material The online version of this Evans 2005). However, Drohojowska and Szwedo (2015) article (https ://doi.org/10.1007/s0043 5-019-00449 -1) contains studied fossil whitefy genera from Lebanese amber and supplementary material, which is available to authorized users. showed some diagnostic features for the adult stage. These * authors, as well as Martin (2007), emphasized that there is Barbara Franielczyk-Pietyra a lack of descriptive characters of adult whitefies, which [email protected] limits descriptive and comparative work between fossils and 1 Department of Zoology, Faculty of Biology extant taxa. Wing venation is often used in the systematics and Environmental Protection, University of Silesia, of whitefies (Shcherbakov 2000; Martin 2007). Bankowa 9, 40-007 Katowice, Poland Vol.:(0123456789)1 3 Zoomorphology Materials and methods Scanning electron microscopy (SEM) Forewings of A. proletella species were examined using scanning electron microscopy. Samples were fxed and stored in 70% ethanol and then prepared using ethanol dehydration and hexamethyldisilazane (HMDS) drying. After 70% ethanol fxation, material was dehydrated in a graded ethanol/water series of 75%, 80%, 90%, 96% and 100% for 10 min in each concentration and then there were three 100% ethanol changes. After dehydration, samples were treated with HMDS 3 × 10 min and remained in HMDS after third change until the solution evaporated (Kanturski et al. 2015). Because of their high sensitiv- ity, wings were not cleaned before SEM examination. Any cleaning attempt resulted in wings damage or curl- ing (Franielczyk-Pietyra, not published). It was difcult to avoid damages of wings even while using alcohol series. Samples were mounted on holders, sputter-coated with gold and examined using a scanning electron microscope Hitachi UHR FE-SEM SU 8010 (Tokyo, Japan) in the Scan- ning Electron Microscopy Laboratory at the Faculty of Biol- ogy and Environmental Protection, University of Silesia. Histology Specimens were collected to 70% ethanol and then trans- Fig. 1 a Light microscopy (LM): the forewing of Aleyrodes proletella ferred to 2.5% glutaraldehyde in a 0.05 M cacodylate (Linnaeus 1758), veins organization, dorsal view. Scale bar 250 µm, bufer (pH 7.4). After washing in a 0.1 M phosphate bufer b Scanning electron microscopy (SEM) showing a row of 7 hairs on the costal margin of the hindwing (white arrow). Scale bar 150 µm, c (pH 7.4), the material was postfxed for 2 h in 1% OsO4 in Scanning electron microscopy showing campaniform sensilla at the a phosphate bufer, dehydrated in a graded series of etha- beginning of radial vein. Scale bar 20 µm nol replaced by acetone and then embedded in an Epoxy Embedding Medium Kit (Sigma, St. Louis, MO). Semithin sections were cut from the root to the tip of the forewing Information on forewing venation in aleyrodids can on a Leica Ultracut UCT ultramicrotome (each section be found in Patch (1909), Schlee (1970), Szelegiewicz having a thickness of 700 nm) with diamond knife and (1971), Klimaszewski and Wojciechowski (1992) as well stained with methylene blue. as Shcherbakov (2000, 2007). Patch studied both wing Sectional cuts (Fig. 2) were analyzed using Nikon venation and tracheation. Other authors introduced some eclipse E600 light microscope and photographed with a mostly theoretical modifcations of venation. Nikon DS-Fi2 camera. The whole wing was cut into about It is the last part of the research on forewings in Ster- 700 semithin sections, from which 12 slices were selected norrhyncha representatives. The whole series was under- after very thorough examination. We divided the wing for taken to verify the following hypotheses: 12 sections on the course of the veins. The most damaged sections were rejected. Sections are aligned in Figs. 3 and 1. Morphologically fight apparatus does not difer much 4 in accordance with the white lines on SEM images (cos- among Sternorrhyncha infraorders. tal margin at the top, anal margin at the bottom, upper 2. Forewing external veins refect their internal structure. surface to the right). Some slices were positioned at the 3. The way of wing folding (fat or roof-like) in Aphido- angle to use the available space efciently. morpha infraorder is correlated with the structure of As the nomenclature of veins is still non-consistent, wing base. we decided to adopt that of Shcherbakov (2007), Szwedo 1 3 Zoomorphology 2016) using the “Traditional Search” algorithm to fnd the most parsimonious trees. The following parameters were used: “memory set—10,000 trees”; “tree bisection—recon- nection (TBR) branch-swapping algorithm” with “10 repli- cations”, “saving 100 trees per replicate” and “zero-length branches collapse after the search”. The consensus tree was calculated using the “majority rule consensus” method with a cutof threshold of 50%. All multistate characters were treated as unordered and equally weighted. The trees and features were studied using WinClada ver. 1.00.08 (Nixon 2002) and graphical processing was performed using Paint. NET 4.0.9. Fig. 2 Light microscopy: the forewing of Aleyrodes proletella (Lin- naeus 1758), places of sectional cuts. Scale bar 500 µm Results and Nel (2011) as well as Nel et al. (2012). The following SEM abbreviations are used in fgures and in the text: af—anal fold; alf—fold of anal lobe; anwp—anterior notal wing There are two types of exteroreceptors on the dorsal side of process; axc2—axillary cord; br—basiradiale; brb—basir- forewing—campaniform and trichoid sensilla. The former adial bridge; bsc—basisubcostale; dmp—distal median are cuticular processes (Fig. 5a) which are usually grouped plate; hp—humeral plate; m—mesonotum; mnwp— and found near the joints at the wing base. There are sev- median notal wing process nt1—pronotum; pmp—proxi- eral campaniform sensilla at the beginning of radial vein mal median plate; pnwp—posterior notal wing process; (Fig. 1c) and some along the course of this vein (Fig. 5b). ppt—parapterum;