Neotibicen similaris similaris ( Smith and Grossbeck, 1907 )

Marshall, David C. & Hill, Kathy B. R., 2017, A new Neotibicen cicada subspecies (Hemiptera: Cicadidae) from the southeastern USA forms hybrid zones with a widespread relative despite a divergent male calling song, Zootaxa 4272 (4), pp. 529-550 : 537-544

Neotibicen similaris similaris ( Smith and Grossbeck, 1907)

Cicada similaris Smith and Grossbeck, 1907: 125 . Rihana similaris Davis, 1912: 262 .

Tibicen similaris Van Duzee, 1916: 2 , 8, 31–33.

Neotibicen similaris, Hill et al., 2015: 220 , 226–227, 249–251. Paratibicen similaris Lee, 2016: 449 , 451, 453.

Type locality. Florida, Nassau County, Fernandina (now called Fernandina Beach) . Type specimen deposited in the USNM (see Sanborn 1999).

Specimens examined. Florida: Alachua Co. — 1♂, High Springs Campgrd., near High Springs at I75 nr jct with Rt. 236, 29°52.458'N 82°32.839'W 38ft. 24 July 2008. K. Hill & D. Marshall US.FL.HSC. Genitalia preparation TIB 26 ( MSM) . 1♂, same location and collectors, 1 Sep 2008, DNA voucher 08.US.FL.HSC.#20, genitalia dissected . 1♂, same location and collectors, 1 Sep 2008 . 2♀, same location and collectors, 0 2 Sep 2008, one with voucher number 02.US.FL.HSP.01 . Marion Co. – 1♀, Rainbow Garden Apartments, Dunnellon , 29°3.070N, 82°27.428W, 6m. 14 Sep 2009. K. Hill, D. Marshall, R. Veal . 1♀, same location, R. Veal, 12 Aug 2010. Collected emerging. Okaloosa Co. — 2♀, 1mi E of Rt 189 on Karick Lake Lower Rd., Blackwater River State Forest , 30°53.453'N 86°39.474W, 84m. 15 Sep 2009. K. Hill & D. Marshall. DNA vouchers 09.US.FL.KLR.01 and 09.US.FL.KLR.02 (legs in EtOH). St. Johns Co. — 1♂, along Hwy A1A, Matanzas River outlet/Bridge. 5 Aug 1984, F. Huber coll., specimen at UMMZ labelled UMMZ.3 . Taylor Co. — 1♂, 2.6mi N. of US98 on CR14 (Aucilla River Rd), SSW of Lamont , 30°10.422'N 83°53.300'W 37ft. 21 July 2008, K. Hill & D. Marshall. DNA voucher 08.US.FL.AUC.01 (legs in EtOH) . Georgia: Lowndes Co. — 1♂, W. side of Hahira , JCT I75 and Rt122, 30.992°N 83.387°W, 210ft. 24 Jul 2008. K. Hill & D. Marshall. DNA voucher 08.US.GA.HAH.01 (legs in EtOH). Seminole Co. — 1♂, Cummings Landing Park , near entrance. 2.3 mi S. of Rt. 263, off Rt. 39, 30°47.153'N 84°52.395'W 61ft. 22 July 2008. K. Hill & D. Marshall. US.GA.CUM. Thorax and abdomen only, no genitalia. Toombs Co. — 1♂, 0.45mi S of Jarhan Collins Rd on Rt86. 1.2mi S of Rt152. ~ 10mi ENE of Vidalia , 32.258°N 82.246°W, 276 ft. 25 Jul 2006. K. Hill & D. Marshall. DNA voucher 10.US.GA.SIM.01 (legs in EtOH) . For recording-only locality information see Supplementary Table 1. The type specimen of Neotibicen similaris was also examined at the USNM, and a dorsal photo is published in Sanborn and Heath (2012). All pinned material is stored in the KHDM collection unless otherwise noted.

Morphological description (see also Fig. 1 E–H, 2B). Smith and Grossbeck (1907) described the morphology of the nominate subspecies of Neotibicen similaris . Focusing on the unusual uncus, they also noted a male body size of 35 mm and the fact that the "lateral border" of the pronotal collar is largely green, a point made to contrast with N. lyricen . These features are shared with N. similaris apalachicola . We extend the morphological description to include the characters stated above for N. similaris apalachicola with the following modifications noted in the Distinguishing Characters section. Size measurements (in mm) for a sample of seven male specimens of Neotibicen similaris similaris are given in Table 1. Note that, as in the preceding section, subspecies identification of these specimens was inferred from geography, with all specimens collected from the region where only N. similaris similaris songs were heard.

Song. The following describes a single phrase of a free-flying male of Neotibicen similaris similaris recorded in the morning on 29 August 2008 in Nassau County, Florida, at the junction of Highway A1A and Lofton Creek, about 7 air miles west-southwest of Fernandina Beach, the approximate location of the original town of Fernandina (recording 08. US.FL.LCC.T04.WAV) and type locality. This recording will be deposited at the online repository BioAcoustica ( Baker et al., 2015) and at www.insectsingers.com ( Marshall and Hill, 2010). Much like N. similaris apalachicola , the song frequency ranges from around 3 to 19 kHz (approximately the limit of the microphone used) and contains the following three sections: (1) A leading section consisting of a uniform buzz which was audible in the field but is not clearly visible against the background sound in the recording. (2) A main phrase that alternates sharply between high-pitched echemes and low-pitched echemes or sets of echemes; as in apalachicola, the high-pitched echemes are also higher in amplitude. The main phrase contains two parts that differ in the rate of alternation and in the temporal pattern. For each cycle in part I (5.7 s in duration), a high-pitched echeme (main energy 8–13 kHz, duration ca. 0.013 s) containing 4–5 pulses is immediately followed by two low-pitched echemes (main energy with peaks at about 4.5 and 6.8 kHz), about 0.016 s and 0.013 s in duration, with the high-pitched echeme seamlessly "slurring" into the first low echeme. The two low-pitched echemes contain shorter pulses that are more numerous and more difficult to resolve than those in the high-pitched echemes. For each cycle in part II of the main phrase, each high-pitched echeme is followed by just one low-pitched echeme. The overall rate of alternation is 14.7 cycles/second in part I and 21.6/ sec in part II; note that these values are somewhat slower than in many of our other recordings measured (see below) probably because of a cool morning temperature. Figure 4 shows an example phrase from a higherquality recording made at a different location.

Song variation. Measurements of song characters from 15 phrases sampled from throughout the range of Neotibicen similaris similaris are given in Table 2 (see Supplementary Table 1 for source locations). Note that most song phrases from throughout the range, including at the holotype locality, exhibited isolated irregularly patterned oscillations in the transition from the alternating section of the main phrase to the trailing buzz.

Distinguishing characters. Neotibicen similaris apalachicola is easily distinguished from N. similaris similaris by features of the male calling song, especially a song phrase containing a single, slow rate of alternation (2–3 cycles/sec) between low-pitched and high-pitched sound, sounding like the word "easy" being slowly repeated. N. similaris similaris contains almost the same sound frequencies but alternates very rapidly between low- and high-pitched echemes at more than ten times the rate and increases the rate part-way into the phrase, forming a clacking rattle. The song of N. similaris apalachicola is superficially similar to that shared by N. winnemanna (Davis) and N. pruinosus (Say) , which also oscillates between high- and low-pitched sound, but the latter two species produce sound that is mostly below 8 kHz in pitch and the fine-scale structure of the sound is entirely unlike that of apalachicola (unpublished data). The song of N. similaris similaris superficially resembles the pulsed call phrase of N. tibicen (L.), but the rate of oscillation in the latter is only about half that of similaris and the fine-scale structure of the song is again entirely different.

Morphologically, Neotibicen similaris apalachicola cannot be consistently distinguished from the nominate subspecies, but on average it has a larger body size, a wider pronotal collar, darker forewing infuscation, and longer male opercula (more often extending beyond sternite IV). Although some character means are significantly different, the ranges show broad overlap in all cases ( Table 1). Subtle differences exist on average in the wing venation as well, with the forewing vein r-m between ulnar cell 2 and apical cell 3 more likely to be one-fourth or less of the length of forewing vein m between apical cell 4 and ulnar cell 2. We found no significant differences in the male genitalia of the two subspecies of N. similaris .

Both subspecies of Neotibicen similaris are easily distinguished from other USA Neotibicen species (see Table 3) by the recurved spines protruding from the uncus, which can be viewed without genitalic dissection if the pygofer is gently extruded with a pin while the specimen is soft. In addition, N. similaris possesses a partly to entirely black pronotal collar (generally present only in N. tibicen , N. lyricen and N. similaris ), a dark pronotum (green in N. tibicen ), and clear wing membranes (suffused with brown in N. lyricen ). Aberrant specimens of other eastern Neotibicen that usually have green pronotal collars, including linnei (Smith and Grossbeck) , winnemanna (Davis) , robinsonianus (Davis) and davisi , either have bright green markings on the mesothorax (the first three species) or are small, with rounded opercula and a wide head ( N. davisi ). Both subspecies of N. similaris are also clearly distinguished from all other USA cicadas by song.

Pronotal Male Mesothorax Wing suffused

collar color genitalia pattern color with brown distally?

N. similaris similaris

and N. s. apalachicola black recurved spine dull green/brown no N. tibicen black no spine green/black slightly N. lyricen lyricen black no spine green/brown slightly

(more in l. virescens) N. linnei , N. winnemanna green no spine green/black slightly in some N. robinsonianus dark green no spine green/brown no N. davisi green/brown no spine green/brown no

Ecology and behavior of Neotibicen similaris subspecies. Calendar dates for our records of adults and emerging cicadas of Neotibicen similaris ranged from 2 July to 28 September. Other sources show N. similaris active in Florida from mid-June until late October ( Sanborn et al. 2008; Walker 2000). Males sing mainly in bright sunshine. The average time of observation of singing cicadas in our study was 12:30 PM, with singing beginning around 8:30 AM and only rarely extending after 6:30 PM. We observed almost no dusk singing, compared to some species like N. linnei which are often active around sunset. Males of N. similaris apalachicola commonly flew to a new singing station after each song, especially in the morning hours, while males of N. similaris similaris appeared more likely to sing several song phrases from one location. When more than one song phrase was sung from a single perch, males of both subspecies produced a continuous low buzz between phrases, as in other Neotibicen cicadas. Interestingly, although some cicada species use vertical movements of the abdomen to alter song pitch (e.g., Magicicada septendecim , see Allard 1937), no such movement was observed when the holotype male of N. similaris apalachicola sang in a cage. Most males heard singing did so from very high stations (almost all over 6m, the maximum reach of our net poles, and most were much higher). Like all Neotibicen spp., mature males utter a loud alarm call when disturbed or handled.

Neotibicen similaris cicadas of both subspecies often sang from coniferous trees, which were present at nearly all of the field sites. During collections at the N. similaris apalachicola holotype locality from ~9:30 PM to ~12:30 AM on multiple evenings, emerging nymphs were found almost exclusively on large conifers despite the proximity of mature interspersed deciduous trees (Supplementary Fig. 3 ). Generally, emerging cicadas will climb up the nearest vertical surface, so possibly most of these cicadas had hatched from the branches of the pine trees. However, one female apalachicola was collected after she was observed ovipositing in a dead sycamore branch. Furthermore, males of both subspecies also sang from junipers and from deciduous trees like pecans, live oaks, laurel oaks and introduced gingko as long as they were large enough. Davis (1918) mentioned male Neotibicen similaris singing in small turkey oaks in Florida, and Sanborn and Phillips (2013) have observed N. similaris in large deciduous trees (subspecies unknown in both cases).

Distributions of Neotibicen similaris subspecies. Song-based records for the Neotibicen similaris complex extend across the southeastern USA from Mississippi to North Carolina, including northern and central Florida ( Fig. 5 ). Details of these locations are given in Supplementary Table 1. One record has been published from Louisiana ( Sanborn & Phillips 2013), one specimen is known from Pennsylvania (see below), and Davis (1918) included Virginia in the distribution of N. similaris without specific information. Sanborn et al. (2008, their Fig. 10) showed localities extending the distribution somewhat farther south in Florida, including a disjunct record in Lee Co., FL. The two subspecies inhabit parapatric (interlocking) ranges with hybrid songs evident in areas of contact, as discussed below. The new subspecies Neotibicen similaris apalachicola exclusively inhabits a compact section of the Florida panhandle approximately 125 km x 55 km in area, centered on the city of Tallahassee ( Fig. 5 c). In addition, two corridors of nearly pure N. similaris apalachicola populations extend north from Jackson and Leon Counties in Florida and surround a small region of pure N. similaris similaris centered on Decatur and Seminole Counties in Georgia. The two corridors of apalachicola join again, to the north of these similaris populations, and extend farther north to Stewart, Webster, and Sumter Counties in Georgia, after which they expand to the east and west into a region of south-central Alabama and Georgia. The northernmost records of N. similaris apalachicola songs are found, to date, in Peach Co. and Johnson Co., GA, and Russell Co., Bullock Co., and Pike Co., AL.

One male in the FAC collection with the label data "PA: BERKS Co.//Douglasville// 31-VIII-83 // F.W.Skillman" was examined for this study. The uncus exhibits the unmistakable inward curving double spines. The only Berks County in the United States is in Pennsylvania. We listened for cicadas around the specified location on the afternoon of 18 September 2009, under cool but sunny conditions, but we heard no Neotibicen similaris songs. Additional searching would be worthwhile because this record considerably expands the published range. It is possible that the specimen was mislabeled or collected in Berks County after having been moved there as an egg or a nymph on a transplanted tree (e.g., Chilcote & Stehr 1984).

Hybridization. Putative hybrid songs combining characteristics of Neotibicen similaris similaris and N. similaris apalachicola were observed in many locations where the subspecies come into contact and apparently interbreed. Examples are shown in Figure 6 and vary from songs more resembling subspecies similaris ( Fig. 6 a), to those more resembling apalachicola ( Fig. 6 b, c), to some with the characters dramatically shifting within the song ( Fig. 6 d). Fig. 6 e shows how the detailed structure of the song in Fig. 6 d includes elements of both apalachicola (the long high-pitched echeme — compare to Fig. 3 c) and similaris (alternating short echemes of high and low pitch, compare to Fig. 4 d). Note that, while it was our impression in the field that consecutive songs made by the same hybrid male resembled each other more than those of other males, we were unable to confidently track individual singing cicadas, and the degree of song variability in hybrids remains undocumented.

In the southern sector of the range of subspecies apalachicola ( Fig. 5 c), hybrid songs were heard only in a zone approximately 20 km wide separating the core apalachicola area from the surrounding populations of true similaris . Farther north, in Alabama and Georgia, pure and hybrid populations of both subspecies are less coherently distributed ( Fig. 5 b); we consider the significance of this pattern in the Discussion. Because our sampling was done rapidly by car, with many sites only briefly checked, our data do not resolve detailed spatial patterns within the hybrid zone, but the region with hybridization was obviously limited relative to the distributions of the parental subspecies. At 388 out of 438 sites where we found Neotibicen similaris cicadas, only one subspecies was heard. At 21 sites, normal songs of one or both parental subspecies were heard together with songs exhibiting hybrid influence. At 24 sites, all songs heard showed signs of hybrid influence, although again most of these were only very briefly sampled. The US.AL.GRN (N. side of Greenfield) and US.AL.WFC (Jct. 33/131 E. of Clio) sites were especially notable for the large number of varied hybrid songs heard (see Supplementary Table 1 for location details). There was no significant evidence of coexistence without hybridization: At five sites we noted both parental subspecies present without hybrid songs, but these were samples of less than one minute's duration with hybrid songs recorded at nearby sites.

Two specimens were collected at sites where hybrid songs were common, one from Stewart Co., GA ( DNA voucher 08. US.GA.LMK.01) and one from Mitchell Co., GA ( DNA voucher 08. US.GA.FLI.01, see Supplementary Table 1). These specimens are housed at the University of Connecticut. The alarm sound of the first male was recorded.

Figures

FIGURE 1. A – D, Neotibicen similaris apalachicola, n. subsp., dorsal view, ventral view, pygofer ventral view, pygofer lateral view (holotype specimen 08. US. FL. TRA. 03). E – H, N. similaris similaris, dorsal view, ventral view, pygofer ventral view, pygofer lateral view (specimen 08. US. GA. HAH. 01). Scale bars = 12.5 mm.

FIGURE 3. Neotibicen similaris apalachicola, n. subsp., field recording of male calling song from Wakulla Co., Florida (recording 08. US. FL. BXU. T 08. WAV). Each panel shows a waveform above a spectrogram; A, complete song phrase; B, partial zoom of the central portion of the phrase; C, further zoom of one of the four oscillations in B; D, segment of song that precedes the main phrase. Sound energy below 1.8 kHz has been removed.

FIGURE 4. Neotibicen similaris similaris, field recording of male calling song from Wakulla Co., Florida (recording 08. US. FL. FOR. T 01. WAV). Each panel shows a waveform above a spectrogram; A, complete song phrase, with annotation identifying two parts in the main phrase that differ in rate of alternation between high- and low-pitched echemes; B, zoom centered about 0.48 s into the clip in A, at the transition from part I to part II; C, further zoom of part I; D, further zoom of part II; E, segment of song that precedes and follows the main phrase. Sound energy below 1.8 kHz has been removed.

FIGURE 5. Distribution records of Neotibicen similaris similaris (blue), N. similaris apalachicola n. subsp. (orange), and hybrid cicadas (X and + symbols). Symbols with central dots or a + symbol indicate that a specimen was collected or a voucher recording was made; other records were aurally noted; A, regional scale map of southeastern USA with labeled states outlined in black and with state counties outlined in grey; zoomed section in B is indicated by the dashed line; B, moderate scale map, zoomed section in C is indicated by the dashed line; C, core N. similaris apalachicola distribution surrounding Tallahassee, FL (grey star).

FIGURE 6. Field recordings of putative hybrids between subspecies of Neotibicen similaris; A – D; spectrograms showing a range of phenotypes exhibiting different combinations of the parental song characters from four different locations (see track ID at bottom right); E, zoomed waveform and spectrogram of the song in D, showing the long high-pitched echeme of N. similaris apalachicola n. subsp. followed by a set of short echemes matching those of N. similaris similaris. Sound energy below 1.8 kHz has been removed.