MOLECULAR ECOLOGY AND EVOLUTION Host Race Evolution in graminum (: ): Nuclear DNA Sequences

KEVIN A. SHUFRAN1

USDAÐARS, , Peanut, and Other Field Crops Research Unit, 1301 N. Western Road, Stillwater, OK 74075 Downloaded from https://academic.oup.com/ee/article-abstract/40/5/1317/420303 by guest on 13 October 2019

Environ. Entomol. 40(5): 1317Ð1322 (2011); DOI: http://dx.doi.org/10.1603/EN11103 ABSTRACT The greenbug , (Rondani) was introduced into the United States in the late 1880s, and quickly was established as a pest of wheat, oat, and barley. was also a host, but it was not until 1968 that greenbug became a serious pest of it as well. The most effective control method is the planting of resistant varieties; however, the occurrence of greenbug biotypes has hampered the development and use of plant resistance as a management technique. Until the 1990s, the evolutionary status of greenbug biotypes was obscure. Four mtDNA cytochrome oxidase subunit I (COI) haplotypes were previously identiÞed, suggesting that S. graminum sensu lato was comprised of host-adapted races. To elucidate the current evolutionary and taxonomic status of the greenbug and its biotypes, two nuclear genes and introns were sequenced; cytochrome c (CytC) and elongation factor 1-␣ (EF1-␣). Phylogenetic analysis of CytC sequences were in complete agreement with COI sequences and demonstrated three distinct evolutionary lineages in S. graminum. EF1-␣ DNA se- quences were in partial agreement with COI and CytC sequences, and demonstrated two distinct evolutionary lineages. Host-adapted races in greenbug are sympatric and appear reproductively isolated. Agricultural biotypes in S. graminum likely arose by genetic recombination via meiosis during sexual reproduction within host-races. The 1968 greenbug outbreak on sorghum was the result of the introduction of a host race adapted to sorghum, and not selection by host resistance genes in crops.

KEY WORDS biotypes, cytochrome c, elongation factor 1-␣, greenbug, host-plant resistance

The greenbug, Schizaphis graminum (Rondani), was The sudden appearance and wide range of the “sor- Þrst discovered in Virginia in 1882 and by 1890 large ghum biotype”, or biotype C as described by Harvey scale outbreaks on wheat, Triticum aestivum L., and and Hackerott (1969), was thought to be a result of an oat, sativa L., began to occur in Texas and introduction of a genotype specializing on that host Oklahoma, as well as other states including Missouri, (Eastop 1973, Blackman 1981). Biotype C was consid- Illinois, and Indiana (Webster and Phillips 1912). It ered morphologically distinct from previous popula- remained mostly relegated to these hosts until 1968 tions (biotype B) damaging to wheat (Blackman 1981, when grain sorghum, (L.) Moench, Fargo et al. 1986, Inayatullah et al. 1987). This was an was infested and injured over a large portion of the early indication that S. graminum sensu lato might be United States (Harvey and Hackerott 1969). Previ- comprised of evolutionary divergent host races or ously, sorghum had been reported as a host, but green- subspecies; however, this theory mostly was ignored bug had not been a signiÞcant pest. until biotypes were examined for variability by using At the time of the 1968 outbreak on sorghum, the molecular genetics approaches. A simplistic model of greenbug biotype phenomenon was well established biotype evolution, whereby biotype E evolved from C, (Porter et al. 1997). The occurrence of greenbug bio- and C evolved from B was generally accepted (In- types capable of injuring and killing resistant wheat ayatullah 1985). and sorghum was thought to be the result of selection Subsequent molecular ecology studies cast doubt pressure placed on single or major host plant resis- on this theory and instead proposed that S. graminum tance genes in a manner analogous to insecticide re- biotypes represented host-adapted races and were the sistance (Teetes 1980) and this theory persisted well products of hundreds if not thousands of years of into the late 20th century (Eisenbach and Mittler reproductive isolation on different hosts (Powers et al. 1987a, de Ponti and Mollema 1992, Lazar et al. 1995). 1989, Black et al. 1992, Black 1993). These studies led to a new theory of the evolution and occurrence of S. graminum biotypes, in which biotypes existed inde- pendently of agriculture and were maintained on The United States Department of Agriculture is an equal oppor- tunity provider and employer. noncultivated hosts (Porter et al. 1997). This new 1 Corresponding author, e-mail: [email protected]. paradigm was supported by sequencing of mtDNA

0046-225X/11/1317Ð1322$04.00/0 ᭧ 2011 Entomological Society of America 1318 ENVIRONMENTAL ENTOMOLOGY Vol. 40, no. 5

Table 1. Schizaphis graminum biotypes and mtDNA haplotype trons, cytochrome c (CytC) and elongation factor 1-␣ associations from which nDNA was sequenced (EF1-␣), were polymerase chain reaction (PCR) am- pliÞed and sequenced. mtDNA Biotype Reference Haplotype A 640 bp PCR amplicon of the CytC was produced and direct sequenced using the primers cytC-C-5Ј I C (Shufran et al. 2000) Ј Ј E (Shufran et al. 2000) (5 -AAGTGTGCYCARTGCCACAC-3 ) (Palumbi E-OK (Shufran and Puterka 2011) 1996) and cytC-B-3Ј (5Ј-CATCTTGGTGCCGGGG- I (Shufran et al. 2000) ATGTATTTCTT-3Ј) (Palumbi 1996). Amplicons J (Shufran et al. 2000) were puriÞed using the Wizard SV Gel and PCR K (Shufran et al. 2000) II F (Shufran et al. 2000) Clean-Up System (Promega, Madison, WI). A 1.2-kbp G (Shufran et al. 2000) amplicon of the EF1-␣ was produced using the primers

NY (Shufran et al. 2000) EF2Ð3Ј (5Ј-ATGTGAGCAGTGTGGCAATCCAA-3Ј) Downloaded from https://academic.oup.com/ee/article-abstract/40/5/1317/420303 by guest on 13 October 2019 ?-OK (Shufran and Puterka 2011) (Palumbi 1996) and EFS175 (5Ј-GGAAATGGG- III B (Shufran et al. 2000) Ј B-OK (Shufran and Puterka 2011) AAAAGGCTCCTTCAAGTAYGCYTGGG-3 ) (Mo- vaginatum (Shufran and Puterka 2011) ran et al. 1999). The EF1- ␣ amplicon was then cloned H H (Shufran et al. 2000) using the pGEM ÐT Easy Vector kit (Promega, Mad- ison, WI) and sequenced with the T7 and SP6 primers. DNA sequencing was conducted with 5ϫ to 6ϫ cov- cytochrome oxidase subunit I (COI), which deter- erage with BigDye-terminated reactions analyzed on mined S. graminum populations had three mtDNA an ABI model 3730 DNA Analyzer (Recombinant COI haplotypes (Shufran et al. 2000, Anstead et al. DNA/Protein Core Facility, Department of Biochem- 2002). These mtDNA sequencing results were consis- istry and Molecular Biology, Oklahoma State Univer- tent with the results of restriction fragment length sity, Stillwater, OK). Internal primers EF4 and EF5 polymorphism (RFLP) analysis of mtDNA (Powers et were used to complete sequencing (von Dohlen et al. al. 1989) and rDNA (Black 1993) and supported the 2002) DNA sequences were assembled and aligned by theory that S. graminum was comprised of host- the ClustalW method (Thompson et al. 1994) using adapted races or subspecies. However, studies of in- the LaserGene Ver. 8.0.2 software suite (DNASTAR, trabiotype variation demonstrated that biotypes ex- Inc., Madison, WI). Default alignment parameters isted independent of COI haplotypes (Anstead et al. were used; gap penalty 15.0, gap length penalty 6.66, 2002). Single biotypes were found in more than one delay divergent sequences 30%, and DNA transition COI haplotype. weight 0.5. Phylogenetic analyses were conducted us- The source of biotypic diversity was shown to be a ing the MEGA5 statistical software package (Tamura product of recombination of virulence genes during et al. 2011). Maximum likelihood (ML) method with meiosis, which occurs during the holocyclic life cycle 1,000 bootstraps was used based on the Tamura and phase of S. graminum (Puterka and Peters 1989, 1990, Nei (1993) model, with uniform substitution rates 1995). Crosses between and within biotypes resulted among sites, all sites (gaps and/or missing data) used, in new biotypic variation in the offspring. Multiple and the ML heuristic method Nearest-Neighbor-In- biotypes found among with different COI hap- terchange. Completed sequences were submitted to lotypes (Anstead et al. 2002) suggest there was gene GenBank with accession numbers: CytC, JF719744 - ßow between host races or homologous virulence JF719757; and EF1-␣, JF968544 - JF968556. Voucher genes existed between host races that were reproduc- specimens of S. graminum biotypes (sisters of speci- tively isolated. To answer this question, DNA se- mens used for DNA analysis) were deposited with the quence variation in the intron and coding regions of Museum of Entomology, FL State Collection of Ar- two nuclear genes was analyzed using the same S. thropods, Gainesville, FL. graminum populations of Shufran et al. (2000), plus four additional clones. Results In total, 518 nucleotides were sequenced from S. Materials and Methods graminum CytC. No nucleotide variation was found in Ten S. graminum biotypes (B, C, E, F, G, H, I, J, K, the coding region, i.e., the Þrst 80 bases, and all sub- and NY), preserved at Ϫ80ЊC from Shufran et al. stitutions, deletions, and insertions occurred in the (2000), three biotypes (E, B, and an unknown, des- 438-bp intron that followed. Phylogenetic analysis ignated as “?”) collected from wheat at Okeene, OK with maximum likelihood produced a dendrogram (Shufran and Puterka 2011), and the Paspalum vagi- with three major clades (Fig. 1a), although the clade natum Sw. (AKA Florida or P) biotype (Nuessly et al. containing biotypes C, E, E-OK, I, K and J was not well 2008) were used in the current study. The mtDNA supported by bootstrap analysis. Neighbor-joining COI haplotype associated with each is shown in Table and maximum parsimony analysis produced essen- 1. The prepGEM DNA extraction kit (ZyGEM tially identical results. Each clade corresponded to Corp. Ltd., Hamilton, New Zealand) was used accord- one of the three mtDNA clades of Shufran et al. ing to the manufacturerÕs instructions to extract (2000), except biotype H fell into the clade con- genomic DNA from a single apterous individual of taining biotypes F, G, NY, and the unknown OK each biotype. Two nuclear DNA genes and their in- biotype (Fig. 1a). October 2011 SHUFRAN:HOST RACE EVOLUTION IN SCHIZAPHIS GRAMINUM 1319

A sequence alignment of CytC from bases 240Ð310 shows differentiation of three clades corresponding to mtDNA haplotypes (Fig. 2a). Biotypes with mtDNA haplotype I contained an eight b insertion starting at position 310 and also differed by a SNP at position 278. Biotypes with mtDNA haplotypes II, III and H lacked this insertion, and mtDNA haplotypes II and III dif- fered by two SNPs at positions 261 and 298. In total, 1,165 nucleotides were sequenced from S. graminum EF1-␣. Three introns of 63, 72, and 70 bp were found and all nucleotide substitutions, inserts, and deletions occurred in these. The exons of all bio- Downloaded from https://academic.oup.com/ee/article-abstract/40/5/1317/420303 by guest on 13 October 2019 types had identical DNA sequences. Biotype J was not included because it is the rarest biotype and too few specimens remain in the collection. Biotype J has never been found a second time after its initial dis- covery. Phylogenetic analysis with maximum likeli- hood produced a dendrogram with three major clades (Fig. 1b). Neighbor-joining and maximum parsimony analysis produced essentially identical results (results not shown). One clade contained all biotypes having a mtDNA haplotype I (C, E, E-OK, I, and K); however, there were mtDNA haplotype II (F and NY) and haplotype III (Paspalum and B-OK) biotypes within. A second clade contained B-OK (Haplotype II and Fig. 1. Maximum likelihood tree of Schizaphis graminum biotype H), and a third clade contained biotypes G biotypes produced from nucleotide sequences of a: a) 518 bp and ?-OK (Haplotype II) and B (Haplotype III). fragment of the cytochrome c nDNA gene; b) 1162 bp frag- A sequence alignment of EF1- ␣ from bases 81Ð160 ment of the elongation factor 1-␣ nDNA gene. shows differentiation of biotypes corresponding to

Fig. 2. Nucleotide sequence of nDNA introns from Schizaphis graminum biotypes showing association with three mtDNA haplotypes, including biotype H: a) cytochrome c; b) elongation factor 1-␣. 1320 ENVIRONMENTAL ENTOMOLOGY Vol. 40, no. 5 mtDNA haplotypes (Fig. 2a.). All mtDNA Haplotype bach and Mittler (1987b) demonstrated sex phero- I biotypes (C, E, E-OK, I, and K had a three b deletion mone discrimination by males in two greenbug bio- beginning at position 95. Biotype F and the Paspalum types, C and E. Because there is no relation between biotype also had this deletion. Biotypes B, G, and ?-OK biotype and mtDNA haplotype (Anstead et al. 2002), were differentiated from Haplotype I biotypes by hav- the authors could have had two different host races in ing a three b insertion beginning at position 95 and their experiment. Unfortunately, no voucher speci- SNPs at positions 94, 98, 100, 101, and 111. The Pas- mens were kept by Eisenbach and Mittler (1987b) so palum biotype was identical to biotypes C, E, I, and F. it is impossible to determine if this was the case. Fur- Biotypes B-OK and H had unique sequences. A second ther investigation of sex pheromone discrimination in variable region of the EF1-␣ (data not shown) showed S. graminum is warranted. a two b deletion at positions 752Ð753 differentiation Puterka and Peters (1989, 1990, 1995) made interbio- between biotypes C, E, E-OK, I, K, F, NY, and Pas- type crosses (C, E, and F) in the laboratory. The results Downloaded from https://academic.oup.com/ee/article-abstract/40/5/1317/420303 by guest on 13 October 2019 palum from G, ?-OK, B, B-OK, and H. demonstrated that recombination of virulence genes re- sulted in biotypic diversity in S. graminum. Based on the molecular genetic data presented in the past and herein, Discussion it appears that biotypic diversity may result from sexual S. graminum in the United States appears to be com- reproduction within host-races, not between them. Vir- prised of two to three evolutionary distinct entities, ulence genes responsible for producing the same biotype which are most likely host-adapted races. This diver- in more than one host race (Anstead et al. 2002) would gence is made evident by the complete agreement of be a result of homologous genes occurring in different phylogenetic analysis of mtDNA COI (Shufran et al. host races. It is not known if there was forced mating 2000) and nDNA CytC sequences. The EF1-␣ sequence between mtDNA haplotypes, but it is likely because data were not in complete agreement with the COI and Puterka and Peters (1989, 1990, 1995) used the same CytC sequences; however it showed that all the mtDNA biotype F (Haplotype II) as this study, and because their haplotype I biotypes were in a single major clade, indi- C and E were collected from wheat. Crosses between cating a distinct evolutionary divergence from most of biotype F and biotypes C and E made by Puterka and the other biotypes and their representative mtDNA hap- Peters (1989, 1990, 1995) would not likely occur in na- lotypes. The EF1-␣ gene and its introns probably did not ture, because biotype F is associated with turf grasses and evolve at the same rates as COI or CytC, which could has not been found on wheat or sorghum. account for the discrepancies. Biotypic diversity in S. graminum now appears to be Mitochondrial and nuclear DNA sequence data sup- because of sexual reproduction within two or three port the existence of a sorghum host-race that Þrst host races (or perhaps subspecies), even though pop- occurred in the United States during 1968 and has ulations are sympatric. The evolutionary status of thereafter been predominant on wheat and sorghum. other aphid biotypes also has been determined The biotypes E, I, and K can be considered as members (e.g., Acyrthosiphon pisum (Via 1999, Hawthorne and of the sorghum biotype, which initially was named Via 2001, Via and Hawthorne 2002); Aphis gossypii biotype C. This grouping can be extended to green- (Vanlerberghe-Masutti and Chavigny 1998, Najar- bugs containing the mtDNA haplotype I, which is Rodrõ´guez et al. 2009); and Therioaphis trifolii (Sun- predominant on crops. Another designation other nucks et al. 1997). In these cases, the term biotype no than biotype can now be assigned for greenbugs that longer applies because speciÞc causes of intraspeciÞc occur on and are pests of sorghum. I propose that these variation were determined by molecular genetics populations and biotypes now be recognized as a true techniques, host utilization studies, and the develop- sorghum host race that was introduced into the United ment of quantitative trait loci (QTL) genomic maps. States during 1968. This hypothesis was originally sug- The production of a QTL genomic map in S. graminum gested by Blackman (1981) and further discussed by s. lat. would further elucidate the evolutionary and Blackman and Eastop (2007). Besides the data pre- biochemical basis of biotype, and perhaps lead to host- sented herein, most other molecular DNA data sup- race distinction. The subsequent identiÞcation of spe- port this hypothesis (Powers et al. 1989, Black 1993, ciÞc aphid genes virulent to wheat and sorghum re- Zhu-Salzman et al. 2003). sistance genes would further aid in the development Anstead et al. (2002) and J. D. Burd and K. A. and use of greenbug resistant crops. Shufran (J.D.B. and K.A.S., unpublished data), indi- cated that S. graminum host races, as identiÞed by mtDNA haplotypes, are sympatric in nature, occurring Acknowledgments on both crop and wild grasses. In addition to the data presented herein, most other molecular DNA data I thank John Burd for supplying specimens of the Paspalum support the hypothesis that S. graminum host races vaginatum biotype, and for determining the biotype of the were reproductively isolated (Powers et al. 1989, greenbugs collected near Okeene, OK. Barbara Driskel com- pleted the DNA extraction, PCR, cloning, and preparation of Black 1993, Zhu-Salzman et al. 2003). Little gene ßow samples for DNA sequencing. Mention of trade names or appears to have occurred between two or three ten- commercial products in this article is solely for the purpose tative host races. Host race speciÞc sex pheromones of providing speciÞc information and does not imply recom- produced by oviparae during the holocycle may be mendation or endorsement by the United States Department one isolating mechanism by which this occurs. Eisen- of Agriculture. October 2011 SHUFRAN:HOST RACE EVOLUTION IN SCHIZAPHIS GRAMINUM 1321

References Cited determination of greenbug, Schizaphis graminum (Hemiptera: Aphididae), on seashore paspalum turfgrass Anstead, J. A., J. D. Burd, and K. A. Shufran. 2002. Mito- (Paspalum vaginatum). Environ. Entomol. 37: 586Ð591. chondrial DNA sequence divergence among Schizaphis Palumbi, S. R. 1996. Nucleic acids II: the polymerase chain graminum (Hemiptera: Aphididae) clones from culti- reaction, pp. 205Ð247. In D. M. Hillis, C. Moritz, and B. K. vated and non-cultivated hosts: haplotype and host asso- Mable (eds.), Molecular systematics, 2nd ed. Sinauer, Inc. ciations. Bull. Entomol. Res. 92: 17Ð24. Publishers, Sunderland, MA. Black, W. C., IV. 1993. Variation in the ribosomal RNA cis- Porter, D. R., J. D. Burd, K. A. Shufran, J. A. Webster, and tron among host-adapted races of an aphid (Schizaphis G. L. Teetes. 1997. Greenbug (Homoptera: Aphididae) graminum). Insect Mol. Biol. 2: 59Ð69. Black, W. C., IV, N. M. DuTeau, G. J. Puterka, J. R. Nechols, biotypes: selected by resistant cultivars or preadapted and J. M. Pettorini. 1992. Use of the random ampliÞed opportunists? J. Econ. Entomol. 90: 1055Ð1065. polymorphic DNA polymerase chain reaction (RAPD- Powers, T. O., S. G. Jensen, S. D. Kindler, C. J. Stryker, and PCR) to detect DNA polymorphisms in aphids (Ho- L. J. Sandall. 1989. Mitochondrial DNA divergence Downloaded from https://academic.oup.com/ee/article-abstract/40/5/1317/420303 by guest on 13 October 2019 moptera: Aphididae). Bull. Entomol. Res. 82: 151Ð159. among greenbug (Homoptera: Aphididae) biotypes. Blackman, R. L. 1981. Aphid genetics and host plant resis- Ann. Entomol. Soc. Am. 82: 298Ð302. tance. Bull. Union Int. Sci. Biol., Sect. Reg. Ouest Pale- Puterka, G. J., and D. C. Peters. 1989. Inheritance of green- artique. 4: 13Ð19. bug, Schizaphis graminum (Rondani), virulence to Gb2 Blackman, R. L., and V. F. Eastop. 2007. Taxonomic issues, and Gb3 resistance genes in wheat. Genome. 32: 109Ð114. pp. 1Ð29. In H. F. van Emden and R. Harrington (eds.), Puterka, G. J., and D. C. Peters. 1990. Sexual reproduction Aphids as crop pests. CAB International, Oxfordshire, and inheritance of virulence in the greenbug, Schizaphis United Kingdom. graminum (Rondani), pp. 289Ð318. In R. K. Campbell de Ponti, O.M.B., and C. Mollema. 1992. Breeding strategies (ed.), Aphid-plant genotype interactions. Elsevier, Am- for insect resistance, pp. 323Ð346. In H. T. Stalker and J. P. sterdam, Netherlands. Murphy (eds.), Proc. Plant Breeding in the 1990s. CAB Puterka, G. J., and D. C. Peters. 1995. Genetics of greenbug International, Oxfordshire, United Kingdom. (Homoptera: Aphididae) virulence to resistance in sor- Eastop, V. F. 1973. Biotypes of aphids, pp. 40Ð51. In A. D. ghum. J. Econ. Entomol. 88: 421Ð429. Lowe (ed.), Perspectives in aphid biology. Bull. No. 2. Shufran, K. A., J. D. Burd, J. A. Anstead, and G. Lushai. 2000. Entomol. Soc. N. Z., Auckland. Mitochondrial DNA sequence divergence among green- Eisenbach, J., and T. E. Mittler. 1987a. Polymorphism of bug (Homoptera: Aphididae) biotypes: evidence for biotypes E and C of the aphid Schizaphis graminum (Ho- host-adapted races. Insect Mol. Biol. 9: 179Ð184. moptera: Aphididae) in response to different scoto- Shufran, K. A., and G. J. Puterka. 2011. DNA barcoding to phases. Environ. Entomol. 16: 519Ð523. identify all life stages of holocyclic cereal aphids Eisenbach, J., and T. E. Mittler. 1987b. Sex pheromone dis- (Hemiptera: Aphididae) on wheat and other . crimination by male aphids of a biotype of Schizaphis Ann. Entomol. Soc. Am. 104: 39Ð42. graminum. Entomol. Exp. Appl. 43: 181Ð182. Sunnucks, P., F. Driver, W. V. Brown, M. Carver, D. F. Hales, Fargo, W. S., C. Inayatullah, J. A. Webster, and D. Holbert. and W. M. Milne. 1997. Biological and genetic charac- 1986. Morphometric variation within apterous females of terization of morphologically similar Therioaphis trifolii Schizaphis graminum biotypes. Res. Popul. Ecol. 28: 163Ð (Monell) (Hemiptera: Aphididae) with different host 172. utilization. Bull. Entomol. Res. 87: 425Ð436. Harvey, T. L., and H. L. Hackerott. 1969. Recognition of a Tamura, K., and M. Nei. 1993. Estimation of the number of greenbug biotype injurious to sorghum. J. Econ. Entomol. nucleotide substitutions in the control region of the mi- 62: 776Ð779. tochondrial DNA in humans and chimpanzees. Mol. Biol. Hawthorne, D. J., and S. Via. 2001. Genetic linkage of eco- Evol. 10: 512Ð526. logical specialization and reproductive isolation in pea Tamura, K., D. Peterson, N. Peterson, G. Stecher, M. Nei, and aphids. Nature 412: 904Ð907. S. Kumar. 2011. MEGA5: Molecular evolutionary genet- Inayatullah, C. 1985. Greenbug biotypes in relation to host ics analysis using maximum likelihood, evolutionary dis- plant resistance. Pest Management Technical Report No. tance, and maximum parsimony methods. Mol. Biol. Evol. 1, Oklahoma State University and U.S. Department of (in press). Agriculture-ARS, Stillwater, OK. Teetes, G. L. 1980. Breeding resistant to , Inayatullah, C., J. A. Webster, and W. S. Fargo. 1987. Mor- phometric variation in the alates of greenbug (Ho- pp. 457Ð485. In F. G. Maxwell and P. R. Jennings (eds.), moptera: Aphididae) biotypes. Ann. Entomol. Soc. Am. Breeding plants resistance to insects. Wiley, New York. 80: 306Ð311. Thompson, J. D., D. G. Higgins, and T. J. Gibson. 1994. Lazar, M. D., G. J. Michels, Jr., and J. D. Booker. 1995. CLUSTAL W: improving the sensitivity of progressive Reproductive and developmental rates of two greenbug multiple sequence alignment through sequence weight- biotypes in relation to two wheat host resistance genes. ing, position speciÞc gap penalties and weight matrix Southwest. Entomol. 20: 467Ð482. choice. Nucleic Acids Res. 22: 4673Ð4680. Moran, N. A., M. E. Kaplan, M. J. Gelsey, T. G. Murphy, and Vanlerberghe-Masutti, F., and P. Chavigny. 1998. Host E. A. Scholes. 1999. Phylogenetics and evolution of the based genetic differentiation in the aphid Aphis gossypii aphid genus Uroleucon based on mitochondrial and nu- Glover, evidenced from RAPD Þngerprints. Mol. Ecol. 7: clear DNA sequences. Syst. Entomol. 24: 85Ð93. 905Ð914. Najar-Rodrı´guez, A. J., E. A. McGraw, C. D. Hull, R. K. Via, S. 1999. Reproductive isolation between sympatric Mensah, and G. H. Walter. 2009. The ecological differ- races of pea aphids. I. Gene ßow restriction and habitat entiation of asexual lineages of cotton aphids: alate be- choice. Evolution 53: 1446Ð1457. haviour, sensory physiology, and differential host associ- Via, S., and D. J. Hawthorne. 2002. The genetic architecture ations. Biol. J. Linn. Soc. 97: 503Ð519. of ecological specialization: correlated gene effects on Nuessly, G. S., R. T. Nagata, J. D. Burd, M. G. Hentz, A. S. host use and habitat choice in pea aphids. Am. Nat. 159: Carroll, and S. E. Halbert. 2008. Biology and biotype 76Ð87. 1322 ENVIRONMENTAL ENTOMOLOGY Vol. 40, no. 5 von Dohlen, C. D., U. Kurosu, and S. Aoki. 2002. Phylogenetics Zhu-Salzman, K., H. Li, P. E. Klein, R. L. Gorena, and R. A. and evolution of the eastern AsianÐeastern North American Salzman. 2003. Using high-throughput ampliÞed frag- disjunct aphid tribe, Hormaphidini (Hemiptera: Aphidi- ment length polymorphism to distinguish sorghum green- dae). Mol. Phylogen. Evol. 23: 257Ð267. bug (Homoptera: Aphididae) biotypes. Agric. For. En- Webster, F. M., and W. J. Phillips. 1912. The spring grain- tomol. 5: 311Ð315. aphis or “green bug”. Bulletin, Bureau of Entomology, United States Department of Agriculture 110: 1Ð153. Received 19 April 2011; accepted 27 June 2011. Downloaded from https://academic.oup.com/ee/article-abstract/40/5/1317/420303 by guest on 13 October 2019