Allonemobius Socius – Allonemobius Fasciatus)

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Biological Journal of the Linnean Society, 2008, 94, 777–796. With 4 ﬁgures

Scale-independent criteria and scale-dependent agents determining the structure of a ground cricket mosaic hybrid zone (Allonemobius socius – Allonemobius fasciatus)

CHARLES L. ROSS1*, JAMES H. BENEDIX JR2, CHRISTOPHER GARCIA3, KALLI LAMBETH3, RACHEL PERRY3†, VANESSA SELWYN3 and DANIEL J. HOWARD3

1School of Natural Sciences, Hampshire College, Amherst, MA 01002, USA 2Department of Biology, DePauw University, Greencastle, IN 46135, USA 3Department of Biology, New Mexico State University, Las Cruces, NM 88003, USA

Received 5 December 2007; accepted for publication 1 November 2007

The structure of the hybrid zone between the ground crickets, Allonemobius socius and Allonemobius fasciatus, approximates an environmental mosaic based on temperature/moisture at regional spatial scales. In the present study, we show that the micro-geographic spatial structure (i.e. single fields) of this hybrid zone is governed by the same criteria. Thus, the criteria that structure this hybrid zone are scale independent, even though the agents that implement these criteria may differ at various scales (climate, latitude, and elevation at regional scales; grass height, slope aspect, and field use at micro-geographic scales). Additionally, the study demonstrates a previously unknown barrier to genetic exchange in this system that acts before conspecific sperm precedence. © 2008 The Linnean Society of London, Biological Journal of the Linnean Society, 2008, 94, 777–796.

ADDITIONAL KEYWORDS: habitat segregation – microgeographic scale – reproductive isolation – spatial structure – speciation.

INTRODUCTION hybrid zone structure (i.e. the opportunity for individuals to interact) and maintenance (i.e. criteria for Hybrid zones may serve as arenas for the evolution favourable traits) (Cain, Andreasen & Howard, 1999); of barriers to gene exchange while still allowing (Sadedin & Littlejohn, 2003). Thus, trait differences the transfer of favourable alleles between species that inﬂuence local distribution within these hybrid (Harrison, 1990; Howard, 1993; Rieseberg, Whitton zones contribute to species coexistence and persis- & Gardner, 1999; Martinsen et al., 2001; Fitzpatrick tence in the face of hybridization. & Shaffer, 2004; Panithanarak et al., 2004; Payseur, The geographic structures of hybrid zones can Krenz & Nachman, 2004; Bouck et al., 2005). Differ- range from clinal (Endler, 1977; Barton & Hewitt, ential introgression of genetic linkage groups indi- 1985; Barton & Gale, 1993) to mosaic (Harrison & cates favoured, neutral, and disfavoured genomic Rand, 1989; Howard et al., 1993; Ross & Harrison, regions with respect to intrinsic and extrinsic envi- 2002; Vines, Köhler & Thiel, 2003; Yanchukov, ronments (Barton & Gale, 1993; Rieseberg, Baird & Hofman & Szymura, 2006). Mosaic zones are struc- Gardner, 2000; Panithanarak et al., 2004). Addition- tured by a patchwork of habitats, where each species ally, the ability of alleles to introgress will depend on is found in alternate habitats, and hybridization between parental types occurs primarily at patch boundaries or in intermediate habitats (Harrison, *Corresponding author. E-mail: [email protected] †Current address: 19111 Rustic Stone Court, Richmond, 1986, 1990; Howard, 1986; Harrison & Rand, 1989; TX 77469, USA. Ross & Harrison, 2002). Thus, the maintenance of

Figure 1. Distribution of Allonemobius socius and Allonemobius fasciatus in North America, with hybrid zone labeled. Dots indicate the general locations of the three study sites. these hybrid zones (and likely many clinal hybrid Hewitt, 1993; Patton, 1993; Szymura, 1993; Sites, zones) includes an extrinsic, environmental compo- Barton & Reed, 1995; Bridle, Vass-de-Zomba & nent (Harrison, 1986, 1990; Arnold, 1997; but see also Butlin, 2002; Ross & Harrison, 2002). Searle, 1993). However, the mechanism that drives On regional spatial scales (i.e. on a scale of habitat associations is not clear for many mosaic 10–100 km), the mosaic hybrid zone between the hybrid zones, and many hybrid zones may have mul- North American ground crickets, Allonemobius socius tiple mechanisms of maintenance. Regardless of the (Scudder) and Allonemobius fasciatus (De Geer), is mechanisms, these hybrid zones provide a superb structured by an environmental patchwork implicat- opportunity to understand how habitat segregation ing both climate and topography in its position and can play a prominent role in structuring and main- maintenance (Howard, 1986; Howard & Waring, taining hybrid zones, and thus create ecological and 1991; Britch, Cain & Howard, 2001). Allonemobius reproductive isolation. socius, the ‘southern ground cricket’, is found Understanding how spatial scale influences struc- throughout the southeastern USA from the eastern ture and maintenance in hybrid zones is critical for seaboard to Texas, extending North to Illinois and understanding those forces that are influential in the New Jersey (Howard & Furth, 1986) (Fig. 1). evolution that occurs within hybrid zones (Arntzen, Allonemobius fasciatus, the ‘striped ground cricket’, 1996; Ross & Harrison, 2002; Babik, Szymura & has a more northern distribution, occurring in south- Rafinski, 2003). Some criteria, such as length of ern Canada and the northeastern USA from the east growing season or population density, may affect coast to Illinois, extending as far south as New Jersey hybrid zone pattern and process only at specific scales and central Illinois (Howard & Furth, 1986) (Fig. 1). and thus may not be relevant for influencing the These morphologically indistinct, univoltine species evolution of certain kinds of genetic barriers that are (A. socius is bivoltine in the far south; Howard, 1982, effective at other spatial scales. On the other hand, 1983; Howard & Furth, 1986) hybridize in a narrow some criteria (e.g. temperature) may apply at all mosaic zone that extends from New Jersey to Illinois, spatial scales but may require scale-specific ‘agents’ to except in the Appalachian Mountains (Howard, 1986; implement them at distinct scales. Although the issue Howard et al., 1993), where the zone widens mark- of scale has rarely been studied explicitly, many edly due to topographic complexity (Howard & hybrid zones may vary in structure at different Waring, 1991) (Fig. 1). In the Appalachians, A. fascia- spatial scales (Searle, 1986; Cruzan & Arnold, 1993; tus populations are found at higher elevations than

© 2008 The Linnean Society of London, Biological Journal of the Linnean Society, 2008, 94, 777–796 CRICKET HYBRID ZONE STRUCTURE 779 nearby A. socius populations (Howard, 1986; Howard this habitat segregation, and the precursory role that & Waring, 1991), with distributions that likely reflect habitat segregation plays in the prezygotic isolation the climatic structuring in these mountains. Both that exists between these two species. species live in disturbed grassy environments such as pastures and fields (Howard & Furth, 1986; Howard et al., 1993); however, A. fasciatus is found in cooler, MATERIAL AND METHODS wetter habitats than A. socius (Howard & Furth, 1986). These local differences in distribution (i.e. CRICKET COLLECTION 1–10 km) may be influenced by factors such as veg- We collected crickets from two previously sampled etation length and density. For example, where the locations, JLW and Kenna, and one new location, JBY two species overlap in west-central Indiana, sites that (Fig. 1, Table 1). JLW is a cow pasture in Indiana are mowed or heavily grazed tend to have mostly with two small hills (northwest- and southeast-facing; A. socius, whereas those that are more lightly grazed NWF and SEF, respectively) separated by a flat area have an abundance of A. fasciatus (J. H. Benedix, that is intermittently water saturated (WF) (Fig. 2A). unpubl. data). A large silver maple (Acer saccharinum) tree is Even though environmental habitat associations located near the top of NWF and shades a significant influence the distributions of each cricket species portion of the hill until late afternoon. The pasture is outside and within the hybrid zone at regional and grazed frequently leading to relatively short grass local scales, it is not clear how habitat structures the height, but it contains a number of shrub species hybrid zone at micro-geographic levels (i.e. < 100 m). (family: Rosaceae) and nonpreferred forb species (e.g. However, the fine-scale habitat structure of this Vernonia sp.) that dot the landscape and protect taller hybrid zone, along with factors such as dispersal, will clumps of grass from grazing. Kenna is a cow pasture determine the opportunity for individuals of both in West Virginia with a flat area next to a creek and species to interact and potentially interbreed. If a large north-northwest-facing slope (Fig. 2B). The species have strict environmental/habitat require- flat area is intensively grazed, leading to very short ments, heterospecific interactions may be rare, and grass height, but the adjoining slope has taller grass other barriers to gene exchange need only be weak to due to less frequent grazing. JBY is the mowed maintain genetic isolation for most of the genomes grass yard of a residential house in Greencastle, IN, of the two species. In the case of A. fasciatus and where the front yard slopes gently toward the south- A. socius, however, extensive previous studies have southwest and the back yard is relatively level demonstrated that conspecific sperm precedence (Fig. 2C). We subdivided two of the locations, JLW (CSP) is a major barrier to gene flow between these and Kenna, into five subsites based on topography. At species and that this barrier is quite strong (Gregory JLW, these subsites comprise: (1) NWF; (2) NWF-WF & Howard, 1993, 1994; Howard et al., 1998a, b), transition; (3) WF; (4) WF-SEF transition; and (5) which suggests that micro-geographic environmental SEF. At Kenna, these subsites comprise: (1) Creek, (2) partitioning would not have to be as strong despite Field, (3) Hill, (4) Lower Slope, and (5) Upper Slope. evidence of local and regional habitat segregation. As JBY was subdivided into two subsites, the front noted by Coyne & Orr (2004: 70, 236), A. fasciatus (JBFY) and back (JBBY) yard. and A. socius represent a most unusual situation in Crickets from each location were sampled in July nature, namely a case in which extensive study has 2004 and 2005 (Fig. 2, Table 1). In 2004, samples identified only a single, strong barrier to gene flow were collected approximately every 5 m in approxi- between two closely-related species. mately straight-line transects across JLW (northwest- In the present study, we extend our investigations facing hill to the southeast-facing hill) and Kenna of barriers to gene exchange in the hybrid zone by (creek to the top of the north-facing hill). At JBY, one studying the micro-geographic structure of the zone. sample each was collected from the front (south side Specifically, we ask two questions. First, how effec- of house) and back (north side of house) yard, which tively can temperature, a strong factor that struc- were separated by approximately 20 m. In 2005, tures this mosaic hybrid zone at regional and local samples were collected in a two-dimensional array scales, explain hybrid zone structure at a micro- approximately 5 m apart (or as the terrain allowed) at geographic scale, the scale of a single field and the both JLW and Kenna after identifying areas of inter- same spatial resolution in which individual crickets est (Fig. 2A, B, Table 1). Thus, each sample repre- meet, mate, and potentially produce hybrid offspring? sents approximately a 5-m2 area of the field. At JBY, Second, how effectively can habitat segregation act as several samples were collected in the front and back a partial barrier to gene exchange? The study reveals yards (Fig. 2C, Table 1). Overall, we collected 108 patterns of habitat segregation at the level of single samples at JLW, 131 samples at Kenna, and nine pastures, temperature profiles that can account for samples at JBY. Each sample was cataloged as being

in one of the ﬁve subsites at JLW or Kenna, or one of 4 and

- two subsites at JBY. Locations for each sample at all sites are listed in Table 1. For each sample, we collected at least 20–30 crickets or until the sample area

CI score of was exhausted. Crickets were collected with sweep = nets, immediately snap-frozen in liquid nitrogen, and Ancestry 4soc’

- then transferred for long-term storage at –80 °C.

GENETIC ANALYSIS We assayed individuals at four variable allozyme loci that are informative for distinguishing A. socius from A. fasciatus: hexokinase (Hk), isocitrate dehydrogenase Idh-1), aspartate aminotransferase (Aat), and peptidase (Pep-3) (Howard, 1982, 1983). At Hk, A. fasciatus and A. socius are ﬁxed for alternate alleles. The other three loci (Idh-1, Aat, Pep-3) contain alleles that

s with a particular score and ancestry (e.g. ‘ are unique to one or the other species at high frequencies. We also scored individuals at an additional locus, Malate dehydrogenase (Mdh), which is used to discriminate A. socius and A. fasciatus from other Allonemobius species. Frozen crickets were homog- enized in water buffer and then subjected to electrophoresis on 11.75% horizontal potato starch gels

1soc 0mix 1mix 2mix 2fas 3mix 3fas 4mix 4fas 5mix 5fas 6fas socius mixed fasciatus (Starchart) to assess variation at the ﬁve loci. - Detailed methods can be found in Howard (1982,

1mix 1983). All gels were scored by one of the investigators - (C.L.R.). 2soc

- From the four variable loci, we determined a character-index (CI) score for each individual based 2mix

- on its multilocus genotype. As described previously (Howard, 1986; Howard & Waring, 1991), CI scores 3soc - were computed by assigning a score of +1 to alleles speciﬁc to A. fasciatus and a score of –1 to alleles 3mix - speciﬁc to A. socius. Because three loci (Hk, Pep-3,

4soc and Aat) have unique alleles in A. fasciatus, but only - two loci (Hk and Idh-1) have unique alleles in A. socius, the score of an individual was in the range

Elevation (m) –4 to +6. Individuals with exclusively A. fasciatus or A. socius alleles were considered to be pure parental types, whereas individuals harboring alleles from 87 228 51 1 33 4 7 3 0 2 0 1 0 0 0 0 1 0 1 1 0.87 0.10 0.03 878181 21581 76781 76981 156 77887 106 802 17 137 816 7 104 233 10 58 14 0 22 26 30 3 24 12 0 23 16 10 0 13 0 5 9 0 2 2 5 13 15 0 13 0 0 12 1 0 0 0 6 0 0 4 8 0 6 0 11 2 1 8 0 3 0 16 0 0 4 0 11 45 2 1 0 3 8 2 0 8 38 3 11 6 0 1 16 11 27 1 2 0 2 1 8 2 3 2 0 21 103 0 49 13 1 0 7 5 0 5 0 2 15 130 21 0 1 41 16 51 0.23 120 8 0 0 1 4 1 0.80 0.27 6 0.62 0.19 0.50 0.62 1 140 68 0.25 0.00 0 0.28 45 22 0.13 0.01 0.00 0.10 0.27 4 0.30 0.72 0.70 4 0.72 0.12 0.16 8787 21087 20987 358 203 115 208 52 13 200 134 110 26 37 56 15 13 23 82 44 38 23 6 13 14 2 5 10 13 22 0 8 0 11 8 0 19 9 8 1 4 4 26 7 0 4 1 8 0 3 59 9 1 5 48 2 14 1 7 10 181 234 0 2 0.41 9 2 0.21 1 7 0.38 5 0 28 0 50 27 0.52 25 0.22 3 0.63 0.26 0.23 1 0.77 0.15 0.20 0.02 ------Central meridian both species were classiﬁed as hybrids (‘mixed species’). For each sample and subsite, we compiled a CI proﬁle to summarize the genetic composition for particular groups. Easting (m) Zone

TEMPERATURE DATA Northing (m) At each site (JLW, JBY, Kenna), we collected tem- 341 4393615 504287 16 212 4393617 504249 16

Geographic coordinatesN Character index numbers perature data using up to seven HOBO temperature data loggers (Onset Computer Corp.). At JLW, three loggers were successfully deployed in NWF and four Geographic coordinates and character index values for subsites at each location in SEF (Fig. 2A, Table 2). At JBY, four loggers were Transition Transition deployed in JBFY (south side of house) and three were deployed in JBBY (an eighth data logger was JBY JBFY 107 4389599 513853 16 JBY JBBY 58 4389627 513860 16 Kenna Upper Slope 441 4280944 442668 17 Kenna Lower Slope 214 4280967 442666 17 Kenna Hill 581 4280999 442676 17 Kenna Field 273 4281012 442677 17 Kenna Creek 223 4281020 442678 17 JLW SEF 279 4393625 504211 16 JLW WF-SEF JLW WF 471 4393616 504267 16 JLW NWF-WF JLW NWF 1252 4393612 504301 16 pure socius ancestry). AncestryNWF, values north-west are facing; proportions SEF, south-east of facing; individuals WF, with intermittently a water particular saturated. ancestry in each subsite. Table 1. Location Subsite Coordinates and elevations are means for all GPS readings at each sampling location within a subsite. Character index valuesplaced are the number of cricket in the back yard, but it failed after 3.5 days).

Figure 2. Landscape proﬁles and distribution of sampling points for the study sites: JLW (A); Kenna (B); JBY (C). Numbers and ‘X’ indicate locations of data loggers. UTM, Universal Transverse Mercator coordinate system.

At Kenna, two loggers each were deployed at Creek (Table 3). The last day at Kenna (July 22) was and Lower Slope. The data loggers measured surface excluded from our analysis. Temperature profiles on temperature at 10-min intervals between 29 July to that day were outliers compared to previous days 10 August 2005 at JLW (12 days), 7–15 July 2005 at because a large storm system moved through the JBY (8 days), and 14–22 July 2006 at Kenna (8 days). area. Temperature readings at each time interval for each subsite (i.e. NWF versus SEF; JBFY versus JBBY, Creek versus Lower Slope) were averaged across data RESULTS loggers (2, 3, or 4), and then averaged across days. To test for differences in maximum daily temperature CHARACTER INDEX PROFILES between the subsites at each location, we performed Overall, we collected 2555 crickets from JLW, 1732 analysis of variance (ANOVA) with date, data logger crickets from Kenna, and 165 crickets from JBY. (nested within subsite), and subsite as fixed effects Among subsites at each location, CI profiles revealed

Table 2. Temperature values from each HOBO data logger TAL. ET

08TeLnenSceyo London, of Society Linnean The 2008 © Geographic Coordinates Temperature

Tukey multiple

Location Subsite HOBO Northing (m) Easting (m) zone central meridian mean Tmax Std Err comparison

JLW NWF 2 4393602 504291 16 -87 30.02 0.5245 A JLW NWF 3 4393606 504296 16 -87 JLW NWF 4 4393600 504291 16 -87 35.21 0.5245 CD JLW NWF 5 4393611 504299 16 -87 33.96 0.5245 BC JLW SEF 6 4393637 504253 16 -87 36.74 0.5245 D JLW SEF 7 4393626 504246 16 -87 35.24 0.5245 CD JLW SEF 8 4393617 504243 16 -87 32.08 0.5245 AB JLW SEF 9 4393619 504247 16 -87 33.99 0.5245 BC JBY JBBY 3 4389637 513852 16 -87 44.06 0.4660 FG

ilgclJunlo h ina Society Linnean the of Journal Biological JBY JBBY 5 4389626 513873 16 -87 39.79 0.3190 BC JBY JBBY 7 4389640 513844 16 -87 40.90 0.3190 CD JBY JBBY 9 4389627 513860 16 -87 36.21 0.3190 A JBY JBFY 2 4389601 513850 16 -87 41.44 0.3190 DE JBY JBFY 4 4389590 513853 16 -87 38.97 0.3190 B JBY JBFY 6 4389609 513860 16 -87 42.37 0.3190 EF JBY JBFY 8 4389582 513846 16 -87 44.85 0.3190 G Kenna Creek 10 4281015 442690 17 -81 47.08 0.6404 B Kenna Creek 11 4281011 442688 17 -81 49.27 0.6404 A Kenna Lower Slope 12 4280982 442669 17 -81 47.21 0.6404 B Kenna Lower Slope 13 4280976 442672 17 -81 46.89 0.6404 B

Mean Tmax is the maximum daily temperature averaged over all days measured (see text). Turkey’s multiple comparisons show values that are signiﬁcantly different. For each location, any two means that do not share a letter are signiﬁcantly different from one another. HOBO #3 at JLW failed to collect any data. At JBY, HOBO #3 collected only 3.5 days of data. 2008, , NWF, north-west facing; SEF, south-east facing. 94 777–796 , CRICKET HYBRID ZONE STRUCTURE 783

Table 3. Analysis of variance for maximum daily temperature at JLW and JBY

Source d.f. Sequential SS Adjusted MS FP

JLW Subsite 1 46.669 46.669 13.05 0.001 Day 12 342.976 28.581 7.99 0.000 HOBO (subsite) 5 343.036 68.607 19.18 0.000 Error 72 257.533 3.577 Total 90 990.216 JBY Subsite 1 39.049 39.049 48.13 0.000 Day 7 95.962 11.988 14.77 0.000 HOBO (subsite) 6 358.372 53.619 66.09 0.000 Error 45 36.510 0.811 Total 59 529.893 Kenna Subsite 1 8.657 8.657 7.539 0.013 Day 6 110.084 18.347 15.978 0.000 HOBO (subsite) 2 17.584 8.792 7.656 0.004 Error 45 20.670 1.148 Total 59 156.994

All factors were treated as fixed effects. d.f., degrees of freedom. a marked spatial segregation between crickets of dif- These transitions in character profiles from A. fas- ferent ancestry over the spatial scale of an individual ciatus to A. socius occur on a spatial scale of less than field. At JLW, individuals with pure A. fasciatus 20 m at all locations. At JLW, obvious physical fea- ancestry were mainly located in NWF and decreased tures correspond to changes in species composition. in abundance across subsites from NWF to SEF The decline in A. fasciatus abundance as one moves (Fig. 3A, Table 1). Allonemobius socius individuals from NWF to SEF occurs in WF, a level area of showed the opposite trend. Proportions of crickets pasture that is bordered by small water-saturated with pure A. socius genotypes gradually decreased strips, including a small stream between WF and from SEF to NWF, although they still constituted 41% SEF. At Kenna, the transition between areas domi- of individuals in NWF (Fig. 3A, Table 1). Crickets of nated by different species occurs in the Field subsite. mixed ancestry remained constant at approximately Finer transect sampling of this subsite (not shown) 21% in all subsites. A similar pattern emerged at revealed that the CI profile transition occurs approxi- Kenna. Pure A. fasciatus crickets dominated the Hill, mately 15–20 m from the creek at the border between Lower Slope, and Upper Slope subsites but were the Creek and Field subsites. As Hill, where A. fas- sparse in the Creek and Field subsites (Fig. 3B, ciatus predominates, is approximately 35 m from the Table 1). Pure A. socius individuals were very abun- creek, the transition from A. socius to A. fasciatus dant near the creek, and they decreased in frequency occurs half way between the creek and the beginning dramatically toward the slope subsites. Again, mixed of the north-facing slope. At this point in the field, a ancestry individuals remained relatively constant small ridge (approximately 30 cm higher than the across all subsites at approximately 27%. At both surrounding field) bisects the field, although it seems JLW and Kenna, the CI scores for individuals of unlikely that this ridge alone presents a barrier for mixed ancestry reflected the composition of the pure crickets. The transition between the two subsites at individuals in a subsite (i.e. higher CI scores where JBY was not sampled (i.e. the side yard of the house), A. fasciatus dominated and a broader range of CI but JBBY and JBFY are separated by approximately scores where proportions of pure individuals were 20 m. more equal). Finally, JBY produced a pattern similar to that of the other two locations. Here, A. socius dominated both subsites, but A. fasciatus occurred INFLUENCE OF SEX AND DEVELOPMENTAL STAGE almost exclusively in JBBY (Fig. 3C, Table 1). Like Males outnumbered females in eight of 12 subsites JLW and Kenna, mixed ancestry crickets remained across all locations (two-way ANOVA of subsite and constant (11%) across both subsites. sex assuming unequal variances among subsites:

© 2008 The Linnean Society of London, Biological Journal of the Linnean Society, 2008, 94, 777–796 iue3. Figure 784 hrce ne C)soe o lee tfu oi(e et.Hg crsaeidctv of indicative are scores High text). (see loci four at alleles for scores (CI) index character ihseicC cr.Pecat eeloealpootosfrasbie niiul fpure of Individuals subsite. a for proportions overall reveal charts Pie score. CI of speciﬁc indicative are with scores low and individuals ersne ybakbr/egs ie netyidvdasaege aswde;idvdaso pure of individuals bars/wedges; grey are individuals ancestry mixed bars/wedges; black by represented netyaewiebars/wedges. white are ancestry .L ROSS L. C. A

hrce ne rﬁe o l ustsfrtesuysts L A;Kna() B C.The (C). JBY (B); Kenna (A); JLW sites: study the for subsites all for proﬁles index Character Proportion Proportion Proportion Proportion Proportion 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.0 0.1 0.2 0.3 0.4 0.5 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.0 0.1 0.2 0.3 0.4 0.5 0.6 4- 2- 6 5 4 3 2 1 0 -1 6 -2 5 -3 4 -4 3 6 2 5 1 4 0 3 -1 6 2 -2 5 1 -3 4 -4 0 3 -1 2 -2 1 -3 -4 0 -1 -2 -3 -4 4- 2- 6 5 4 3 2 1 0 -1 -2 -3 -4 TAL. ET 08TeLnenSceyo London, of Society Linnean The 2008 © N apesize. sample , pure socius JLW bysubsite character indexscore NWF-WF transition WF-SEF transition loeoissocius Allonemobius NWF WF SEF mixed ilgclJunlo h ina Society Linnean the of Journal Biological The . pure fasciatus y ai hw rprino niiul tasubsite a at individuals of proportion shows -axis N=1252 N=212 N=341 N=279 N=471 loeoisfasciatus Allonemobius .socius A. 2008, , x netyare ancestry ai depicts -axis 94 .fasciatus A. 777–796 , CRICKET HYBRID ZONE STRUCTURE 785

B Kenna by subsite 0.5 N=223 0.4 1-Creek

0.3

0.2

Proportion 0.1

0.0 -4 -3 -2 -1 0 1 2 3 4 5 6

0.5 N=273 0.4 2-Field

0.3

0.2

Proportion 0.1

0.0 -4 -3 -2 -1 0 1 2 3 4 5 6

0.5 N=581 0.4 3-Hill

0.3

0.2

Proportion 0.1

0.0 -4 -3 -2 -1 0 1 2 3 4 5 6

0.5 N=214 0.4 4-Lower slope

0.3

0.2

Proportion 0.1

0.0 -4 -3 -2 -1 0 1 2 3 4 5 6

0.5 N=441 0.4 5-Upper slope

0.3

0.2

Proportion 0.1

0.0 -4 -3 -2 -1 0 1 2 3 4 5 6 character index score

pure socius mixed pure fasciatus

Figure 3. Continued

Fsex = 4.28, P = 0.0628, although P = 0.026 when JBY ability’ differences. Comparing the ratio of males to subsites with low sample sizes are excluded; Table 4). females for each ancestral group separately (i.e. pure The highest asymmetries occurred at NWF (JLW), fasciatus, mixed, and pure socius), sex ratios at where the male : female ratio = 1.36, and JBFY, two JLW subsites differed signiﬁcantly among the where the male : female ratio = 0.56. Greater male three genetic groups (likelihood ratio test: NWF, abundances may indicate asynchronous development c2 = 14.168, d.f. = 2, P = 0.0008; NWF-WF, c2 = 9.350, times between sexes, sex-speciﬁc dispersal, or ‘catch- d.f. = 2, P = 0.0093). In all other subsites, sex ratios

C JBY by subsite

0.5 N=58 0.4 JBBY

0.3

0.2

Proportion 0.1

0.0 -4 -3 -2 -1 0 1 2 3 4 5 6

0.5 N=107 0.4 JBFY

0.3

0.2

Proportion 0.1

0.0 -4 -3 -2 -1 0 1 2 3 4 5 6

pure socius mixed pure fasciatus

Figure 3. Continued

were not significantly different among ancestral encounter probability would be 0.28 (Table 5) if these groups. crickets were uniformly distributed throughout the The number of male and female crickets of different five subsites at JLW. ancestries (i.e. their relative proportions) will influ- For all subsites at our three study locations, the ence the potential encounter rates (and thus hybrid- probabilities of heterospecific encounters were signifi- ization rates) of the two species. For example, the cantly less than expected based on a homogenous interspecific encounter rate potentially will be rela- distribution of crickets at each location (Z61 = –1.7793, tively high in areas with a large proportion of A. fas- P = 0.0376 for one-tailed test of Ln transformed ciatus females (compared to all females) and a large values; juveniles are excluded because sex is undeter- proportion of A. socius males (compared to all males), mined). Conspecific encounter probabilities, however, but will not be high in areas with both A. fasciatus did not deviate from expectations (Z31 = –2.753, females and males at high proportions (relative to P = 0.6085). Moreover, the probabilities of heterospe- other ancestral groups of each sex). Assuming overall cific encounters between males and females (includ- cricket densities are equal across all subsites and ing mixed ancestry individuals) for all subsites were the numbers of crickets caught of different ancestries significantly less than the probabilities of conspecific are representative of their relative densities within encounters (all subsites t38.8 = 2.530, P = 0.0156, subsites, the probability of hetero- and conspecific assuming unequal variances; Table 5). These results encounters between individuals of opposite sex within suggest that habitat segregation provides a genuine a subsite can be estimated by multiplying the relative pre-zygotic barrier to mating and gene exchange proportions of males and females for each ancestral between these species even within a single pasture. class (Table 5). We then can compare these encounter We collected individuals from all developmental probabilities within each subsite to a ‘null’ expecta- stages (early-instar to adults; Table 6). Variation in tion; in this case, the ‘expected’ encounter probabili- the proportions of individuals at specific developmen- ties if males and females of all ancestral classes were tal stages can reflect selection, habitat preferences, evenly distributed across all subsites at a location and movement among habitats, or varying rates of (i.e. no habitat segregation). These expected encoun- development among sites or genotypes (Endler, 1986). ter probabilities are weighted by the relative overall Like the genotype-specific sex ratios of adults, the proportions within an ancestral class. For example, at proportions of crickets in various developmental JLW, the overall proportions of A. socius males and stages will influence rates of hybridization in this females compared to all males and females are 0.56 system. For our sampling sites, variation among and 0.50, respectively (Table 4). Thus, their expected developmental stages in the proportion of crickets

Table 4. Number of individuals (and proportion within sex) by sex and ancestry in each subsite

JLW Subsite

Sex Ancestry NWF NWF-WF WF WF-SEF SEF

Juvenile Mixed 65 (0.20) 12 (0.24) 6 (0.20) 2 (0.12) 6 (0.25) Juvenile Fasciatus 71 (0.21) 8 (0.16) 4 (0.13) 0 (0.00) 0 (0.00) Juvenile Socius 196 (0.59) 30 (0.60) 20 (0.67) 15 (0.88) 18 (0.75) Female Mixed 82 (0.21) 37 (0.29) 53 (0.27) 23 (0.27) 24 (0.18) Female Fasciatus 195 (0.50) 38 (0.30) 29 (0.15) 2 (0.02) 0 (0.00) Female Socius 112 (0.29) 53 (0.41) 118 (0.59) 61 (0.71) 111 (0.82) Male Mixed 116 (0.22) 25 (0.16) 47 (0.20) 18 (0.17) 24 (0.20) Male Fasciatus 206 (0.39) 39 (0.25) 36 (0.15) 3 (0.03) 1 (0.01) Male Socius 208 (0.39) 89 (0.58) 158 (0.66) 88 (0.81) 94 (0.79)

Kenna Subsite

Sex Ancestry Creek Field Hill Lower Slope Upper Slope

Juvenile Mixed 17 (0.22) 17 (0.18) 52 (0.23) 1 (0.50) 8 (0.40) Juvenile Fasciatus 4 (0.05) 8 (0.08) 103 (0.46) 1 (0.50) 11 (0.55) Juvenile Socius 56 (0.73) 70 (0.74) 68 (0.30) 0 (0.00) 1 (0.05) Female Mixed 25 (0.35) 23 (0.31) 48 (0.29) 26 (0.30) 43 (0.26) Female Fasciatus 9 (0.13) 10 (0.13) 93 (0.56) 62 (0.70) 119 (0.72) Female Socius 37 (0.52) 42 (0.56) 25 (0.15) 0 (0.00) 4 (0.02) Male Mixed 19 (0.29) 26 (0.27) 57 (0.31) 33 (0.35) 59 (0.27) Male Fasciatus 8 (0.15) 16 (0.24) 91 (0.44) 62 (0.50) 156 (0.56) Male Socius 39 (0.59) 55 (0.57) 38 (0.20) 0 (0.00) 0 (0.00)

JBY Subsite

Sex Ancestry JBBY JBFY

Juvenile Mixed 1 (0.25) 0 (0.00) Juvenile Socius 3 (0.75) 5 (1.00) Female Mixed 3 (0.10) 6 (0.09) Female Fasciatus 4 (0.13) 0 (0.00) Female Socius 24 (0.77) 58 (0.91) Male Mixed 3 (0.14) 5 (0.14) Male Fasciatus 5 (0.23) 3 (0.08) Male Socius 14 (0.64) 28 (0.78)

NWF, north-west facing; SEF, south-east facing; WF, intermittently water saturated.

with different ancestry occurred in three subsites proportion of adult crickets (Table 6). This trend at JLW (likelihood ratio test: NWF, c2 = 109.888, occurs at all subsites of JLW and Kenna where appre- d.f. = 10, P < 0.0001; NWF-WF, c2 = 16.430, d.f. = 8, ciable numbers of A. socius occur, even though it P = 0.0366; WF, c2 = 16.918, d.f. = 6, P = 0.0096) and is signiﬁcant in only ﬁve of the ten subsites. On two subsites at Kenna (likelihood ratio test: Field, the other hand, crickets with A. fasciatus ancestry c2 = 19.981, d.f. = 6, P = 0.0028; Hill, c2 = 17.545, increase in proportion in each progressive immature d.f. = 6, P = 0.0147). At JLW and Kenna, crickets with stage, but decrease as a proportion of the adult stage. A. socius ancestry decrease in proportion in each Again, this occurs at all subsites at JLW and Kenna progressive immature developmental stage (middle where A. fasciatus occurs at appreciable frequencies instar, penultimate instar, last instar; early instar is (except between middle instar and penultimate instar excluded due to low numbers) compared to crickets of at WF, and in Upper Slope and Lower Slope, where mixed or A. fasciatus ancestry, but comprise a larger A. fasciatus occurs at high proportions in all stages).

Table 5. Encounter probabilities for individuals of different sex and ancestry in each subsite

JLW Subsite

Male Female Type Exp. prob. NWF NWF-WF WF WF-SEF SEF

Socius Socius Con 0.282 0.113 0.241 0.387 0.573 0.649 Fasciatus Socius Het 0.126 0.111 0.106 0.088 0.020 0.007 Mixed Socius Het 0.102 0.063 0.068 0.115 0.117 0.166 Socius Fasciatus Het 0.164 0.196 0.173 0.095 0.019 0.000 Fasciatus Fasciatus Con 0.073 0.194 0.076 0.022 0.001 0.000 Mixed Fasciatus Het 0.059 0.109 0.049 0.028 0.004 0.000 Socius Mixed Het 0.107 0.082 0.168 0.174 0.216 0.140 Fasciatus Mixed Het 0.048 0.082 0.074 0.040 0.007 0.001 Mixed Mixed Con 0.039 0.046 0.047 0.052 0.044 0.036

Kenna Subsite

Male Female Type Exp prob. Creek Field Hill Lower Slope Upper Slope

Socius Socius Con 0.038 0.308 0.318 0.031 0.000 0.000 Fasciatus Socius Het 0.096 0.063 0.092 0.074 0.000 0.017 Mixed Socius Het 0.056 0.150 0.150 0.046 0.000 0.007 Socius Fasciatus Het 0.104 0.075 0.076 0.114 0.000 0.000 Fasciatus Fasciatus Con 0.262 0.015 0.022 0.274 0.460 0.520 Mixed Fasciatus Het 0.152 0.036 0.036 0.172 0.245 0.197 Socius Mixed Het 0.058 0.208 0.174 0.059 0.000 0.000 Fasciatus Mixed Het 0.147 0.043 0.051 0.141 0.193 0.188 Mixed Mixed Con 0.086 0.101 0.082 0.089 0.103 0.071

JBY Subsite

Male Female Type Exp prob. JBBY JBFY

Socius Socius Con 0.625 0.705 0.493 Fasciatus Socius Het 0.119 0.076 0.176 Mixed Socius Het 0.119 0.017 0.106 Socius Fasciatus Het 0.030 0.000 0.082 Fasciatus Fasciatus Con 0.006 0.000 0.029 Mixed Fasciatus Het 0.006 0.000 0.018 Socius Mixed Het 0.069 0.073 0.062 Fasciatus Mixed Het 0.013 0.008 0.022 Mixed Mixed Con 0.013 0.013 0.013

Con, conspeciﬁc encounter; Het, heterospeciﬁc encounter; Exp. prob., expected probability over all subsites; NWF, north-west facing; SEF, south-east facing; WF, intermittently water saturated. Juveniles are excluded from probability calculations.

Hence, these trends at JLW and Kenna occur at every TEMPERATURE DIFFERENCES subsite where A. fasciatus and A. socius both occur in at least moderate numbers. Unlike the other sites, Daily temperature proﬁles for JLW, Kenna, and JBY JBY does not show this pattern, although few pure indicate signiﬁcant differences between subsites A. fasciatus individuals were found at this site. (Fig. 4, Tables 2, 3). At JLW, the southeast-facing Notably, the only A. fasciatus crickets found at JBY hill (i.e. SEF, WF-SEF) experienced higher daily were adults, mostly females. Crickets of mixed ances- maximum temperatures than the northwest-facing try remained relatively constant in the proportion slope (NWF and NWF-WF) (P = 0.001; Table 3). Mean of individuals at each developmental stage for all daily maximum temperature (Tmax) at SEF averaged subsites. over all days and data loggers was 34.51 °C; at NWF,

Table 6. Number of individuals (and proportion within stage) by developmental stage and ancestry in each subsite

JLW Subsite

Stage Ancestry NWF NWF-WF WF WF-SEF SEF

EI Mixed 3 (0.60) 1 (0.50) 0 (0.00) 0 (0.00) 3 (0.60) EI Fasciatus 2 (0.40) 0 (0.00) 0 (0.00) 0 (0.00) 0 (0.00) EI Socius 0 (0.00) 1 (0.50) 0 (0.00) 0 (0.00) 2 (0.40) MI Mixed 66 (0.19) 10 (0.19) 8 (0.18) 5 (0.17) 10 (0.20) MI Fasciatus 80 (0.23) 8 (0.15) 6 (0.13) 0 (0.00) 0 (0.00) MI Socius 202 (0.58) 35 (0.66) 31 (0.69) 25 (0.83) 41 (0.80) PI Mixed 129 (0.24) 14 (0.15) 35 (0.28) 13 (0.21) 11 (0.17) PI Fasciatus 206 (0.39) 25 (0.27) 11 (0.09) 1 (0.02) 0 (0.00) PI Socius 199 (0.37) 54 (0.58) 77 (0.63) 47 (0.77) 52 (0.83) LI Mixed 47 (0.17) 27 (0.24) 27 (0.19) 17 (0.23) 18 (0.26) LI Fasciatus 159 (0.57) 37 (0.33) 34 (0.24) 1 (0.01) 1 (0.01) LI Socius 71 (0.26) 47 (0.42) 80 (0.57) 57 (0.76) 50 (0.72) Adult Mixed 18 (0.22) 21 (0.30) 36 (0.23) 8 (0.18) 12 (0.14) Adult Fasciatus 21 (0.26) 15 (0.21) 17 (0.11) 3 (0.07) 0 (0.00) Adult Socius 43 (0.52) 35 (0.49) 107 (0.67) 34 (0.76) 76 (0.86)

Kenna Subsite

Stage Ancestry Creek Field Hill Lower Slope Upper Slope

EI Mixed 0 (0.00) 0 (0.00) 0 (0.00) 0 (0.00) 1 (1.00) EI Fasciatus 0 (0.00) 0 (0.00) 0 (0.00) 1 (1.00) 0 (0.00) MI Mixed 17 (0.22) 17 (0.18) 52 (0.24) 2 (0.20) 7 (0.32) MI Fasciatus 4 (0.05) 8 (0.08) 102 (0.46) 8 (0.80) 14 (0.64) MI Socius 56 (0.73) 70 (0.74) 67 (0.30) 0 (0.00) 1 (0.05) PI Mixed 16 (0.31) 17 (0.25) 59 (0.28) 14 (0.30) 16 (0.22) PI Fasciatus 6 (0.12) 12 (0.18) 109 (0.52) 33 (0.70) 56 (0.78) PI Socius 29 (0.57) 39 (0.57) 40 (0.19) 0 (0.00) 0 (0.00) LI Mixed 15 (0.33) 15 (0.33) 31 (0.29) 34 (0.39) 61 (0.32) LI Fasciatus 6 (0.13) 12 (0.26) 61 (0.58) 54 (0.61) 127 (0.66) LI Socius 24 (0.53) 19 (0.41) 14 (0.13) 0 (0.00) 4 (0.02) Adult Mixed 13 (0.33) 17 (0.29) 15 (0.38) 10 (0.26) 22 (0.20) Adult Fasciatus 5 (0.13) 3 (0.05) 15 (0.38) 28 (0.74) 87 (0.80) Adult Socius 22 (0.55) 39 (0.66) 10 (0.25) 0 (0.00) 0 (0.00)

JBY Subsite

Stage Ancestry JBBY JBFY

MI Mixed 1 (0.25) 0 (0.00) MI Socius 3 (0.75) 5 (1.00) PI Mixed 1 (0.25) 3 (0.09) PI Socius 3 (0.75) 32 (0.91) LI Mixed 1 (0.08) 3 (0.13) LI Socius 12 (0.92) 20 (0.87) Adult Mixed 4 (0.11) 5 (0.12) Adult Fasciatus 9 (0.25) 3 (0.07) Adult Socius 23 (0.64) 34 (0.81)

EI, early instar; MI, middle instar; PI, penultimate instar; LI, last instar; NWF, north-west facing; SEF, south-east facing; WF, intermittently water saturated.

35 50 NWF Lower Slope SEF Creek 33 JLW Kenna 45 31

29 40

27 35

30 23 Mean Temperature (˚C) Mean Temperature (˚C) 21 25

19 20 17

15 15 0 0300 0600 0900 1200 1500 1800 2100 0 0300 0600 0900 1200 1500 1800 2100 Time of Day Time of Day

C 45 JBBY JBY JBFY

Mean Temperature (˚C) 25

15 0 0300 0600 0900 1200 1500 1800 2100 Time of Day

Figure 4. Diurnal temperature proﬁles: JLW (A); Kenna (B); JBY (C). For each graph, the curves depict for two subsites the average temperature (with standard error) during 10-min intervals over the period temperature was measured. Black curves show subsites where Allonemobius socius is dominant; grey curves show subsites where Allonemobius fasciatus is more abundant.

mean Tmax was 33.06 °C. At Kenna, mean Tmax at the shown), most of this separation was due to tempera- Creek subsite was signiﬁcantly higher than at the ture differences that occurred during the hottest part

Lower Slope subsite (48.16 °C versus 47.04 °C; of the day. As a result, Tmax is a good indicator of

P = 0.013). Finally, mean Tmax at JBFY was signifi- temperature-dependent effects between subsites. At cantly higher than JBBY (41.91 °C versus 40.24 °C; this time of the day, crickets likely experience the P < 0.001). Significant variation also was observed most heat and water stress, which may affect their among days and among data loggers at all locations ability to survive (especially softer-bodied immatures) (Table 3). and may influence behavioral taxis and development As shown in Fig. 4, the maximum temperatures at rate. all subsites occurred between 11.07 h and 16.24 h At JLW and Kenna, the ‘fasciatus’ sites (NWF and each day, with 50% of them occurring between 13.14 h Lower Slope, respectively) are significantly hotter and 14.39 h. Even though the average temperatures than their corresponding ‘socius’ sites (SEF and across an entire diurnal cycle were significantly dif- Creek) late in the day (approximately 14.00–19.00 h ferent between subsites at each location (data not in the afternoon), although NWF and Lower Slope

never experienced a higher Tmax than SEF or Creek. which can all affect micro-geographic variation in The sustained afternoon warming at NWF and Lower temperature and moisture (Waterhouse, 1955; More- Slope likely is due to fact that the sun late in the day croft, Taylor & Oliver, 1998; Collins et al., 2003). at this time in summer can directly impact these A previous study of this hybrid zone failed to detect slopes, but SEF and Creek are not receiving much significant habitat segregation on micro-geographic direct incident solar radiation. At JLW, this is scales. Specifically, Howard et al. (1993) found no enhanced because the northwest-facing hill is no evidence of structuring at a fine spatial scale (5-m longer shaded by the large tree in NWF at this time grids), although at least one of their subsites sug- of day. At Kenna, the Creek subsite is protected by a gested some nonrandom distributions of genotypes, number of trees and the bank of the creek itself. notably a larger than expected number of hybrid Furthermore, we recorded large amounts of variation individuals. Additionally, no heterogeneity was found in Tmax among data loggers. Tukey’s multiple compari- in the distribution of developmental stages across son tests within subsites indicate that, even though grids or by genotype (Howard et al., 1993). The most areas within subsites are similar with respect to obvious explanation for the difference in results is temperature profiles, there is still considerable varia- that, by contrast to the present study, the previous tion (Table 2). This may reflect exposure to sun (i.e. study emphasized sampling across homogeneous amount of incident radiation mentioned above), varia- areas that did not vary in aspect, height of grass, or tion due to vegetation, and position in the subsite pattern of utilization. The difference in findings (especially with respect to the tree in NWF). For underscores the fact that despite clear habitat parti- example, this may explain relatively high readings for tioning on very local scales, the two species occur one data logger in NWF (#4) and relatively low read- together in some habitat patches. ings for one in SEF (#8). There is some indication that specific samples nearest to these data loggers show more A. socius and A. fasciatus alleles, respectively FORCES INFLUENCING HABITAT SEGREGATION (data not shown), but this result is inconclusive and Habitat segregation in patchy environments is often will require more specific spatial mapping of geno- explained as the outcome of habitat choice or com- types and temperature, especially during diurnal petitive exclusion (Hanski, 1983; Shorrocks, 1990; cycles. Doebeli, 1996; Arlettaz, 1999; Schluter, 2000; Spina, 2000; Vernes, 2003; Friggens & Brown, 2005). In the case of A. fasciatus and A. socius, distinct behavioral DISCUSSION preferences are more likely to account for habitat The distribution of numerous pairs of related North- segregation than competitive exclusion. In laboratory ern hemisphere terrestrial species, such as A. socius experiments, A. socius tends to move to warmer and and A. fasciatus, are largely determined by regional drier microhabitats more than A. fasciatus, and these climatic effects (Howard & Waring, 1991; Britch et al., preferences are present in both nymphs and adults 2001). Northern range limits often are strongly influ- (J. H. Benedix, unpubl. data). Additionally, large scale enced by minimum winter temperatures and south- field experiments carried out more than 20 years ago ern range limits by maximum summer temperatures demonstrated that A. fasciatus has a clear preference (although other biotic and abiotic factors may also for wetter microhabitats in fields where it is sym- contribute) (Tauber, Tauber & Masaki, 1986; Parme- patric with a closely-related species, Allonemobius san & Yohe, 2003; Parmesan, Gaines & Gonzalez, allardi, resulting in habitat segregation on the scale 2005). In the present study, we have shown that of a few meters (Howard & Harrison, 1984a, b). habitat segregation between cricket species occurs on Although competition is thought to be a pervasive the ‘micro-geographic’ scale of a single field, over force in structuring animal and plant communities distances of approximately 20 m. Allonemobius socius (Wiens, 1977; Diamond, 1978; Schoener, 1983; Gold- is most abundant in warmer areas. By contrast, berg & Barton, 1992; Gurevitch et al., 1992), it may A. fasciatus occurs mostly in relatively cooler areas. be a rare occurrence in omnivores and detritivores The influence of regional climatic effects and large- (e.g. ground crickets) that experience little resource scale topography (general location and elevation) are limitation (Howard & Furth, 1986; Schluter, 2000). likely trivial on this small spatial scale, but the same Allonemobius fasciatus shows no competitive interac- criteria (i.e. temperature and moisture) can still tions with A. allardi when they are sympatric in a operate to determine species distributions. The scale- single field, even though both species show specific specific agents that influence these criteria may habitat associations (Howard & Harrison, 1984a, b). include slope aspect, grass height, variation in veg- Thus, habitat partitioning between A. socius and etation, and field utilization (cow grazing, sheep A. fasciatus likely represents current-day divergent grazing, cultivation for hay, frequency of mowing, etc). habitat choice. However, there is no evidence that this

© 2008 The Linnean Society of London, Biological Journal of the Linnean Society, 2008, 94, 777–796 792 C. L. ROSS ET AL. habitat choice evolved in the hybrid zone. The pat- Allonemobius socius, contrastingly, showed the oppo- terns of association of each species are consistent with site trend (mixed individuals remained constant). those seen outside the zone of overlap, implicating This observation is consistent throughout the study natural selection operating in allopatric populations. sites, suggesting that species- or sex-specific move- Habitat choice and thus segregation between A. fas- ment (at least within our sites) cannot explain these ciatus and A. socius appears to be relative to adjoin- trends. One potential explanation for this finding ing habitats crickets may encounter. The respective is that A. fasciatus individuals are more fit than mean Tmax for JLW subsites (NWF = 33.06 °C versus A. socius individuals early in development, but SEF = 34.51 °C) are both less than the mean Tmax for A. socius individuals are more fit during the last- JBY and Kenna subsites (JBBY = 40.24 °C versus instar molt and as adults. Earlier longitudinal studies JBFY = 41.91 °C; Lower Slope = 44.31 °C versus (following specific cohorts) by Howard et al. (1993) Creek = 45.42 °C). In each case, A. fasciatus is found demonstrated strong selection in favour of A. socius nearly exclusively in the subsites with lower Tmax over the course of the summer in mixed populations, even though JBBY and Lower Slope were warmer and long-term studies have documented the loss of on average than either JLW subsite. Allonemobius A. fasciatus in some mixed populations over the past socius, however, occurred at all subsites where tem- 20 years (Britch et al., 2001). Therefore, we favour the perature was recorded. This pattern suggests that hypothesis that A. fasciatus fares poorly at these sites A. fasciatus is more sensitive to microhabitat differ- during the height of the summer heat in July due to ences than A. socius. viability selection against adults. Up to this point, the maintenance of bimodality in character scores has been attributed to a single repro- SELECTION AGAINST HYBRIDS ductive barrier: CSP. It is now apparent that at least Although adaptive behavioral differences appear to be one other factor, habitat segregation, plays a role in important in the maintenance of this zone, the same limiting gene flow between A. fasciatus and A. socius. cannot be said for selection against hybrids. Offspring Because habitat segregation will decrease inter- of heterospecific and F2 crosses do not experience specific encounter rates in zones of overlap, it will significant post-zygotic selection affecting viability or augment any reproductive barrier, such as CSP, that development under laboratory conditions (Gregory & operates over short distances or after mating. Most Howard, 1993). In the field, hybrid individuals main- interspecific encounter probabilities we estimated tain constant proportions throughout development in were below a ‘random’ expectation, assuming no specific cohorts, and hybrid individuals at most loca- habitat segregation, and many were near zero. tions were at least as fit as A. fasciatus (Howard However, many of the predicted conspecific encoun- et al., 1993). Moreover, the cross-sectional sampling of ters rates were quite high. We did not measure move- this study found that the proportions of hybrid indi- ment of individuals directly among subsites; however, viduals were consistent in all developmental stages given likely habitat preferences, the movement of and in all subsites at every location. Therefore, individuals to preferred habitats would increase con- hybrids probably do not face serious viability selection specific and decrease heterospecific encounters. It is during hatchling-to-adult development compared to also possible that differences in development rate other genotypic classes, nor do they suffer differential contribute to isolation. Both species are univoltine fitness consequences in different habitats. Indeed, and short-lived in the contact zone. Thus, if the peaks hybrid crickets likely have intermediate fitness com- of adult emergence of the two species are offset, pared to individuals from the two parental classes in conspecific matings will be more common than het- all habitats (Howard et al., 1993), a situation under- erospecific matings, even in mixed populations with considered (Graham, Freeman & McArthur, 1995; equal abundances of the two species. Bell, 1997; Dessauer, Cole & Townsend, 2000; Barton, When multiple barriers are present, any one factor 2001; Burke & Arnold, 2001; Campbell, 2004) and need not be as strong to effectively prevent introgres- one that could result in asymmetrical selective forces sion. As most hybrid zones are associated with large against (or for) hybridization for the parental types. amounts of statistical (rather than genetic) linkage disequilibrium at their centers due to the mixing of distinct parental genomes (Barton, 1979a, b; Barton DEVELOPMENT, MICROGEOGRAPHIC STRUCTURE, & Hewitt, 1985), introgression of alleles at specific AND THE MAINTENANCE OF BIMODALITY loci are tied to their abilities to dissociate from At JLW and Kenna, the proportion of A. fasciatus the ‘superlocus’ of highly linked parental genomes. increased in more advanced juvenile developmental Because of high linkage disequilibrium, most loci will stages, but decreased as a proportion of adults, col- experience an ‘effective’ weak disruptive selection lected as a cross-sectional sample in July (Table 6). against hybrids because of specific loci that experi-

© 2008 The Linnean Society of London, Biological Journal of the Linnean Society, 2008, 94, 777–796 CRICKET HYBRID ZONE STRUCTURE 793 ence strong selection due to their involvement as predict that maximum summer high temperatures reproduction isolation factors (such as sperm utiliza- will increase in the Appalachian Mountains (Boer, tion factors in Allonemobius). If traits such as habitat Flato & Ramsden, 1999; Polsky et al., 2000), the preference and development rate involve many loci hybrid zone overall in this region should narrow due (Rieseberg et al., 1999; Harushima et al., 2001; Ortiz- to A. fasciatus getting ‘pushed’ off the top of moun- Barrientos et al., 2002; but, for a notable exception, tains (i.e. due to elevation-temperature covariances) see Martin, Bouck & Arnold, 2005, 2006), they will but, locally, this process should occur more quickly on link a higher proportion of the genome to a barrier to slopes with a southern aspect (warmer) than a north- gene exchange, limiting introgression to a greater ern aspect, and more slowly in fields with taller grass extent (perhaps non-additively) than when just one species that are resistant to climate change; for other barrier exists. examples, see Jeffree & Jeffree (1996) and Bull & Burzacott (2001). From ongoing surveys of specific populations within the hybrid zone, it is apparent MICRO-GEOGRAPHIC STRUCTURE IN HYBRID ZONES that A. socius alleles are increasing in frequency The habitat segregation between A. fasciatus and (Britch et al., 2001). Closer inspection of micro- A. socius demonstrates that closely-related species climate, landscape, and habitat associations with are capable of fine-scale partitioning of habitats such cricket genotypes should continue to prove useful in that only very detailed studies are capable of docu- understanding evolution in this hybrid zone. menting the partitioning. Thus, it seems likely that the phenomenon of habitat segregation is under- ACKNOWLEDGEMENTS reported in hybrid zone studies, particularly those that emphasize genetic analyses. At the same time, it We thank Steve Parsons and Ann Greene, and John is important to acknowledge that ‘micro-geographic’ and Linda Welker for allowing us to collect crickets in structure has been documented in several other their fields. Ron Ross provided valuable assistance hybrid zones. For example, in the Gryllus firmus/ with collecting crickets. This project was supported Gryllus pennsylvanicus (field cricket) hybrid zone, by National Science Foundation grant 0316194 to parental genotypes are strongly correlated with soil D.J.H., a Research Initiative for Scientific Enhance- type (Harrison, 1986; Rand & Harrison, 1989) and ment (RISE) award to C.G., a Minority Access to this association occurs at fine spatial scales (Ross & Research Career (MARC) award to V.S., and grants Harrison, 2002), even though the mosaic character of from the Student/Faculty Summer Research Fund at this hybrid zone is apparent only at intermediate DePauw University to J.H.B. 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