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J. gen. Virol. (1989), 70, 2717-2727. Printed in Great Britain 2717 Key words: evolution/geminivirus/coat protein/taxonorny Phylogeny of Geminiviruses By A L A N J. H O W A R T H * AND G E O R G E J. V A N D E M A R K Department of Plant Pathology, University of Arizona, Tucson, Arizona 85721, U.S.A. (Accepted 14 June 1989) SUMMARY Amino acid sequences of 16 geminivirus replication-associated proteins and 15 coat proteins were aligned and a new computer program was used to calculate the minimum mutation distances for all possible pairwise comparisons. These data were used to construct phylogenetic trees. Trees based on coat proteins had two main branches which were positively correlated with vector specificities of the viruses. Trees based on replication-associated proteins also had two main branches which were positively correlated with viral host specificities for either monocotyledonous or dicotyledonous plants. Therefore, evolutionary pressures on coat proteins and replication-associated proteins are probably highly influenced by vectors and hosts, respectively. Geminiviruses that infect dicotyledonous plants may be divided further by geographical origins into Old World and New World viruses. These results suggest the possible geographical origins of some geminiviruses, that new taxa should be erected, and have implications for distinguishing viruses and strains. INTRODUCTION The taxonomy of viruses has improved greatly over the years as more information has become available about the physical and chemical nature of viruses. Early attempts at classifying viruses were based on characters such as symptomatology and host range which depended on the genotype of both viruses and host organisms (Matthews, 1983). Current classification schemes for distinguishing viruses into families and genera, or groups, rely on inherent traits of the viruses, including type of nucleic acid, particle morphology and mode of replication (Matthews, 1982). Within viral taxa distinctions between viruses are often based on characteristics such as host range and serological relationships (Francki, 1983). Geminiviruses are of world-wide importance as pathogens of crops including cereals such as maize and wheat (Goodman, 1981a). They are among the smallest known autonomously replicating viruses (Howarth & Goodman, 1982). As a result, geminiviruses are among the most intensively studied viruses for their potential usefulness in genetic engineering of plants (Buck & Coutts, 1983). The genomes of at least 16 geminivirus isolates have been sequenced (Table 1). Nucleic acid type (circular ssDNA) and a unique geminate particle morphology distinguish geminiviruses from all other known viruses (Matthews, 1982). However, a dichotomy among geminiviruses has been evident from the time that they were recognized as worthy of taxonomic distinction (Harrison et al., 1977). This dichotomy has been viewed in terms of vector specificity (leafhoppers or whiteflies), host range [monocotyledonous (monocots) or dicotyledonous (dicots) plants], and, more recently, genomic complexity (segmented or unsegmented; Fig. 1 ; Goodman, 1981 b; Stanley, 1983, 1985; Grimsley et al., 1987). Based on these criteria three subgroups have been suggested: (i) whitefly-transmitted, segmented genome, infects dicots; (ii) leafhoppertransmitted, unsegmented genome, infects dicots; (iii) leafhopper-transmitted, unsegmented genome, infects monocots. Although it has been useful to categorize geminiviruses in these ways, new sequence information provided an opportunity for further examination of relationships among geminiviruses. We have chosen to use molecular distance data to evaluate the relationships among geminiviruses. Only two proteins, coat protein (gene I) and the replication-associated protein 0000-8991 © 1989 SGM Downloaded from www.microbiologyresearch.org by IP: 88.99.165.207 On: Fri, 16 Jun 2017 22:41:01 Downloaded from www.microbiologyresearch.org by IP: 88.99.165.207 On: Fri, 16 Jun 2017 22:41:01 Mung bean yellow mosaic Tomato golden mosaic Tomato yellow leaf curl Wheat dwarf Cassava latentt Chloris striate mosaic Digitaria streak Maize streak Abutilon mosaic Beet curly top Bean golden mosaic Virus Brazil California, U.S.A. Puerto Rico Brazil Kenya Australia Vanuatu Kenya Nigeria South Africa Thailand Brazil Israel Czechoslovakia Czechoslovakia Sweden AbMV BCTV BGMV-PR BGMV-BZ CLV CSMV DSV MSV-K MSV-N MSV-S MYMV TGMV TYLCV WDV-C WDV-CJI WDV-S B. tabaci B. tabaci Nesoclutha pallida Nesoclutha declivata Cicadulina spp. Cicadulina spp. Cicadulina spp. B. tabaci B. tabaci B. tabaci Psammotettix alienus P. alienus P. alienus Bemisia tabaci Neoaliturus tenellus B. tabaci Vector Malvaceae 44 families* Leguminosae, Malvaceae Leguminosae Solanaceae Gramineae Gramineae Gramineae Gramineae Gramineae Leguminosae Solanaceae Solanaceae Gramineae Gramineae Gramineae Host Reference Gilbertson et al. (1988) Stanley & Gay (1983) Andersen et al. (1988) Donson et al. (1987) Howell (1984) Mullineaux et al. (1984) Lazarowitz (1988) Morinaga et al. (1987) Hamilton et al. (1984) N. Navot & H. Czosnek (unpublished) Commandeur et al. (1987) Woolston et al. (1988) MacDowell et al. (1985) Frischmuth et al. (1987) Stanley et al. (1986) Howarth et al. (1985) * All 44 families which contain hosts of BCTV are dicotyledonous (Bennett, 1971). t Cassava latent virus is the same as African cassava mosaic virus (Bock & Woods, 1983). Origin Abbreviation T a b l e 1. O r i g i n s , v e c t o r s a n d h o s t s o f 16 g e m i n i v i r u s isolates Z t7 rn Z t7 > o ~o t,~ Geminivirus phylogeny 2719 (a) DNA 1 (X (b) (c) MSV-N [!iiiiiiiiiili!ii!iiiii!!iii!!iiiiii!iiiiiii!ii!ii!i BGMV Fig. 1. Organization of geminivirus genomes and genes. The genomes of BGMV (a) and MSV (b) are drawn schematically as represenative of dicot- and monocot-infecting geminiviruses, respectively. Arrows depict the polarity and relative lengths of the conserved genes. Boxes are the common regions. The small arc shows the binding site of the 80 nucleotide (approx.) 'primer' which is found with DNA in virions of the monocot viruses. (c) Amino acid sequence similarity of MSV ORFs IIa and lib to BGMV ORF II is indicated. (gene II), are encoded in the genomes of all g e m i n i v i r u s e s characterized to date (Fig. 1 ; see references in T a b l e 1). Phylogenetic trees were constructed based o n sequence a l i g n m e n t s of each of these two proteins. I n a previous study i n v o l v i n g only six viruses, we used replicationassociated protein genes as the basis for a n a l y s i n g relationships b e t w e e n viruses ( H o w a r t h & G o o d m a n , 1986). Downloaded from www.microbiologyresearch.org by IP: 88.99.165.207 On: Fri, 16 Jun 2017 22:41:01 2720 A. J. HOWARTH AND G. J. VANDEMARK METHODS The viruses analysed in this study and their abbreviations are listed in Table 1. Amino acid sequence alignments of replication-associated proteins and coat proteins, respectively, were developed from the alignments of MacDowell et al. (1985) and Stanley et al. (1986). Gaps were added as necessary to maximize similarities of additional viral sequences. Then, each sequence was compared pairwise to every other sequence to determine minimum mutation distances (Fitch & Margoliash, 1967), the minimum number of mutations required to convert the sequence of a protein into the sequence of the protein with which it is compared. These numbers were normalized for sequence length (Fitch & Margolish, 1967). Comparisons involving gaps were assigned a value of zero and, thus, were omitted from the calculations. These computations were aided by a new computer program which requires as input the aligned sequences and which gives as output a matrix of normalized mutation distances. The program was designed to read an input file with a default length of l0 characters for the virus name and 600 characters of single-letter code for the amino acid sequence of the peptide. The program can compare up to 50 different viruses in one program execution. The name length, amino acid sequence length and maximum number of viruses compared in one execution can be increased beyond the default limits by simple modifications of the source code. The source code is a Pascal program which can be executed on any IBM-compatible personal computer with a Turbo Pascal compiler or on any mainframe computer by invoking the Pascal compiler. The program is available to interested users. Rooted and unrooted trees were constructed from the minimum mutation distances using the Fitch and Kitsch programs contained in the Phylip package of programs of Felsenstein 0985) as supplied for the IBM PC by G. D. F. Wilson (Scripps Institution of Oceanography, La Jolla, Ca. 92093, U.S.A.). Phylogenetic trees with the smallest sums of squares were obtained by at least three executions of each program using the jumble and global rearrangement options. RESULTS Comparison of replication-associated proteins Alignments of 16 amino acid sequences which correspond to the replication-associated proteins of sequenced geminiviruses were compared in every pairwise combination. Minimum mutation distances were calculated for each comparison (Table 2). After three executions of the tree-construction program, we concluded that the best tree had been established because each execution resulted in the same best tree and sum of squares (S.S. = 0.25). We used an F test (Felsenstein, 1984) to compare the rooted tree produced by the Kitsch program, which assumes a molecular clock, to the unrooted tree produced by the Fitch program, which assumes no clock. The resulting highly significant F statistic of 35-0 led us to reject the hypothesis that there was no difference between the rooted and unrooted trees. Therefore, we rejected the validity of the molecular clock for these data. However, the same clusters of viruses were obtained with both programs. Fig. 2 depicts the best tree of an average of 777 trees that were examined for each execution of the program. The average percent standard deviation of this tree is only 3.3. The principal feature of the tree is that there are two main branches which correspond to the host specificities of the viruses for either monocots or dicots. The horizontal lines in the monocot branch are longer than in the dicot branch, i.e. there is less divergence among dicot-infecting viruses than among monocot-infecting viruses. All of the viruses in the monocot branch of the tree have a gene organization as shown in Fig. 1 (b). The branch containing the viruses that infect dicots divides into two branches. The viruses in one branch (AbMV, BCTV, BGMV-PR, BGMV-BZ and TGMV) were collected in the New World whereas those in the other branch [CLV (ACMV), MYMV and TYCV] were collected in the Old World. BCTV is the only virus in this cluster which is not transmitted by the whitefly, Bemisia tabaci (Bennett, 1971). Further, BCTV has a genome composed of a 2993 nucleotide DNA, which is very similar in its gene organization D N A 1 of BGMV and other whitefly-transmitted viruses (Fig. 1a; Stanley et al., 1986). The other dicot-infecting viruses in Fig. 2 have two genomic DNAs and the gene organization depicted in Fig. 1 (a) (Lazarowitz, 1987). The normalized mutation distances (Table 2) showed that the viruses which are considered to be strains of WDV were very similar and had mutation distances of 10 or less, as did strains of MSV. Even the most similar of other viruses had scores at least 10-fold larger. BGMV-PR and BGMV-BZ were not each other's closest relative; BGMV-PR was nearest to AbMV and BGMV-BZ was nearest to TGMV. Downloaded from www.microbiologyresearch.org by IP: 88.99.165.207 On: Fri, 16 Jun 2017 22:41:01 Downloaded from www.microbiologyresearch.org by IP: 88.99.165.207 On: Fri, 16 Jun 2017 22:41:01 0 237 199 153 240 506 496 511 511 513 259 190 241 469 469 470 232 0 220 216 231 497 487 502 502 504 250 210 232 459 460 460 203 226 0 179 223 490 480 494 494 496 242 131 224 452 452 453 153 208 173 0 220 486 476 491 491 493 239 t69 221 448 449 449 242 239 239 225 0 476 466 481 481 483 194 214 176 438 439 439 498 474 482 483 477 0 358 372 372 374 495 480 477 357 358 358 508 480 471 473 460 355 0 205 205 207 485 470 467 347 348 348 527 481 490 502 456 374 208 0 5 7 500 485 482 362 362 363 527 481 490 502 459 374 205 5 0 2 500 485 482 362 362 363 529 483 491 501 461 374 205 8 2 0 502 487 484 364 364 365 245 262 245 244 205 498 483 492 495 494 0 233 158 457 458 458 196 218 131 167 225 486 480 497 497 497 216 0 215 442 442 443 226 248 221 227 169 497 504 501 504 503 158 195 0 439 440 440 485 448 445 453 415 360 345 360 364 367 467 456 460 0 8 3 481 446 442 450 413 360 344 362 366 368 466 453 459 8 0 9 481 446 442 449 413 360 343 360 363 365 466 453 461 3 9 0 AbMV BCTV BGMV-BZ BGMV-PR CLV CSMV DSV MSV-K MSV-N MSV-S MYMV T G M V TYLCV WDV-C WDV-CJI WDV-S Matrix of mutation distances* of geminivirus replication-associated proteins * Numbers abovethe diagonalare normalized m i n i m u m m u t a t i o n distances. Below the diagonalare distancesreconstructed from thetree with the smallestsum of squares. AbMV BCTV BGMV-BZ BGMV-PR CLV CSMV DSV MSV-K MSV-N MSV-S MYMV TGMV TYLCV WDV-C WDV-CJI WDV-S T a b l e 2. t~ to Downloaded from www.microbiologyresearch.org by IP: 88.99.165.207 On: Fri, 16 Jun 2017 22:41:01 ~ 0 4-2 ,,60 ....... 78.6 I7 173.6 17131 l ~ 1~0~. 1 ~2~ ~ 8 70"4 880 70.0 60-6 113.6 8%6 [ 66.2 Ab~V ~v~v BCTV - TGMV BGMV-BZ BGMV-PR CLV 86.8 146.7 lO8.2, 145.9 l 29.5 34-2 179.2 BGMV-PR BGMV-BZ I I 28.5 TYLCV 20.4 35.5 CLV 37 ] ' ~ 7 TGMV ~ 2 31.~'81 ~ 20-7 AbMV Fig. 3, Relationship of 15 geminiviruses based on coat protein alignments. The statistically best tree is shown. Lengths of horizontal arms are proportional to the mutation distances indicated by the numbers, Lengths of vertical lines are arbitrary. WDV-CJI 1-0 BCTV WDV-C 0-0 WDV-S 0)1 csMv C--] osv~2 67~ I 38.0 0.5 MSV-K ]~ .3 MSV-S 2 g[-- 69.8 MSV_N J ~ Fig, 2, Relationship of 16 geminiviruses based on replication-associated protein alignments. The tree with the smallest sum of squares (Felsenstein, 1984) as calculated by the Fitch program of the Phylip package is shown. Lengths of horizontal arms are proportional to the mutation distances indicated by the numbers. Lengths of vertical lines are arbitrary. WDV-CJ1 0I~~ wov_~ ~..... WDV-C 1.0 95.2 183,7 MSV-K 2.6 107.4 DSV CSMV 6 t MVS-S 2. MSV-N Z t7 Z ,.q o tO tO Downloaded from www.microbiologyresearch.org by IP: 88.99.165.207 On: Fri, 16 Jun 2017 22:41:01 0 387 32 47 118 413 410 414 415 416 38 111 423 423 423 377 0 378 374 381 318 315 319 320 321 372 374 328 328 328 32 377 0 38 108 404 401 405 405 407 29 101 414 414 414 49 375 38 0 105 400 397 401 402 403 33 98 410 410 410 AbMV BCTV BGMV-BZ BGMV-PR 110 381 116 105 0 407 404 408 409 410 103 64 417 417 417 392 318 394 398 402 0 214 218 218 220 398 400 285 285 285 CLV CSMV 406 308 401 400 400 215 0 139 139 141 395 397 282 282 282 413 318 413 412 410 218 139 0 4 3 400 401 287 287 287 409 317 409 409 408 218 138 4 0 6 400 402 287 287 287 DSV MSV-K MSV-N 412 318 412 411 411 218 142 3 6 0 402 403 289 289 289 40 373 28 32 105 393 401 410 407 409 0 96 408 408 408 MSV-S T G M V 104 383 108 96 64 391 397 410 408 410 97 0 410 410 410 TYLCV Matrix ofmutation distances*forgeminiv~us coatproteins 413 333 411 411 411 295 283 283 287 283 418 411 0 2 0 412 333 410 410 412 295 284 283 287 283 417 412 2 0 2 WDV-C WDV-CJI 413 333 411 411 411 295 283 283 287 283 418 411 0 2 0 WDV-S * Numbers above the diagonal are normalized minimum mutation distances. Below the diagonal are distances reconstructed from the tree with the smallest sum of squares. AbMV BCTV BGMV-BZ BGMV-PR CLV CSMV DSV MSV-K MSV-N MSV-S TGMV TYLCV WDV-C WDV-CJI WDV-S T a b l e 3. l,O 2724 A. J. HOWARTH AND G. J. VANDEMARK Comparison o f coat proteins Alignments of 15 coat protein amino acid sequences (the coat protein sequence of MYMV was not available) were compared in every pairwise combination, as described above for the replication-associated proteins. Minimum mutation distances were computed (Table 3). Three executions of the Fitch program of the Phylip package each yielded the same best tree and sum of squares (S.S. = 0.088). An average of 593 trees were examined per program execution. The best tree had an average percent standard deviation of only 2-0. An F test (F = 65.6) led us to reject the validity of the molecular clock, therefore Fig. 3 depicts an unrooted tree. The main feature of this tree is dichotomous branching into clusters of geminiviruses that correlate with their vectors, either whiteflies or leafhoppers. The horizontal lines of the leafhopper-transmitted viruses were longer than those of the whitefly-transmitted viruses, showing that divergence was greater among the leafhopper- than among the whitefly-transmitted viruses, a result also concluded from the tree based on replication-associated proteins (Fig. 2). According to this scheme, BCTV is nearest to viruses which are also transmitted by leafhopper vectors, but differs from them in having a host range restricted to dicots and a gene organization like DNA 1 of Fig. 1 (a). In other regards, the tree in Fig. 3 is similar to the tree in Fig. 2. Again, BGMV-PR and BGMV-BZ were each more closely related to another virus, TGMV, than to each other. DISCUSSION Serology or amino acid composition of capsid proteins are commonly used to suggest phylogenies of plant viruses (Shukla & Ward, 1988; Fauquet et al., 1986). Serological studies of geminiviruses have shown that whitefly-transmitted geminiviruses are related sufficiently to cross-react with polyclonal and monoclonal antisera; however, there is a general lack of serological cross-reactivity among leafhopper-transmitted geminiviruses (Roberts et al., 1984), with the exceptions of MSV and DSV (Dollet et al., 1986) and BCTV and tobacco yellow dwarf virus (Thomas & Bowyer, 1980). No antigenic cross-reactivity has been substantiated between whitefly- and leafhopper-transmitted geminiviruses (Thomas et al., 1986). These observations, together with the fact that coat protein is the only protein that has been purified from most geminiviruses, limits the usefulness of a serological approach in a comparative study. We have overcome these problems in determining phylogenetic relationships of geminiviruses by analysis of alignments of amino acid sequences of both gene products which all geminiviruses contain, the coat protein and the replication-associated protein (MacDowell et al., 1985 ; Stanley et al., 1986). Thus, even the most disparate geminiviruses were compared. The geminiviruses that infect either monocots or dicots havc different genome organizations. First, the monocot-type DNAs have two non-coding (intcrgenic) regions, whereas dicot-type DNAs have only one intergenic region in DNA 1. Second, the monocot-type viruses have small (approximately 80 nucleotides) complementary-strand DNAs which contain a few covalently bound, 5'-terminal ribonucleotides and are associated with and probably base-paired to viral DNA inside nucleocapsids (Howell, 1984; Donson et al., 1984, 1987; Andersen et al., 1988; Hayes et al., 1988; Lazarowitz, 1988). These small DNAs are thought to be primers of complementary strand DNA synthesis. No such 'primer' molecules have been reported for viruses with dicot-type DNAs. Third, monocot viruses encoded an 11K to 12K protein in a gene (alpha in Fig. 1 b) with the same polarity as, and just upstream from, the coat protein gene (Mullineaux et al., 1988). No comparable gene is conserved in the genomes of the dicot viruses. Similarly, monocot viruses lack genes III and IV that are conserved in dicot viruses. Fourth, monocot viruses have two open reading frames (ORFs) in the complementary strand (labelled IIa a n d l I b in Fig. 1b), the products of which have significant amino acid sequence similarities to the product of gene II of the dicot viruses (Mullineaux et al., 1985). Schalk et al. (1989) have shown that splicing of WDV RNA results in removal of an intron and, presumably, expression of only one protein from these two ORFs. These points suggest that although monocot and dicot geminiviruses are grossly similar in genetic organization, close examination reveals that there may be significant differences in their mechanisms of replication and gene expression. Downloaded from www.microbiologyresearch.org by IP: 88.99.165.207 On: Fri, 16 Jun 2017 22:41:01 Geminivirus phylogeny 2725 The replication-associated protein phylogenetic tree (Fig. 2) has two main branches one of which contains the viruses that infect dicots and the other those which infect monocots. These results suggest that this tree is consistent with classification of geminiviruses based on two constantly associated characteristics, host range limited to either monocots or dicots and type of genome organization. Further, the results also suggest that evolution of replication-associated proteins is influenced by viral hosts. Thus, the original divergence of monocot and dicot geminiviruses may have been, in part, due to evolution of the replication-associated protein genes to function eMciently in the different plant hosts. One could speculate, then, that gene organization has become an effective determinant, but not necessarily the only determinant, of host range. For example, the two ORFs that encode the replication-associated protein of the monocot viruses probably require RNA splicing as a necessary step in expression of that gene (Schalk et al., 1989). Perhaps dicots do not process these genes properly and are, thus, eliminated as potential hosts of these viruses. The coat protein tree (Fig. 3) is similar to the replication-associated protein tree (Fig. 2) in that it also divides into two main branches. Unlike the replication-associated protein tree, however, the phylogeny derived from analysis of coat proteins correlates with vector specificities of the viruses for either leafhoppers or whiteflies. The relationships among the monocot viruses are unchanged from the tree in Fig. 2. The whitefly-transmitted viruses have some small differences in that BGMV-PR and BGMV-BZ switch places in their relative associations with T G M V and AbMV. CLV and TYLCV, the only whitefly-transmitted viruses of the Old World for which coat protein sequences were available, form their own branch of the tree apart from the whiteflytransmitted New World viruses to suggest that, as with the replication-associated protein tree, geographical isolation has played a role in their phylogenetic history. The main difference between the coat protein and replication-associated protein trees is that the position of BCTV has changed from near the other viruses of dicots, all of which are whitefly-transmitted, to near the monocot viruses, all of which are leafhopper-transmitted. This result supports the hypothesis that coat protein is intimately associated with vector transmission (Roberts et al., 1984; Gardiner et al., 1988), the corollary of which is that evolution of the coat protein gene is constrained by the need to accommodate vector transmissibility. Thus, it seems logical that phylogeny based on coat proteins would correspond to vector specificity. We favour the replication-associated protein tree as the best representation of the phylogeny of geminiviruses because the genome organization, structure of its replication-associated protein gene and host range of BCTV are like those of other geminiviruses that infect dicots. Our results may have a bearing on the issue of origins of geminiviruses. For example, Oman (1969) suggested that BCTV originated in the New World and perhaps was transmitted by a less aggressive insect which has since been displaced by the introduced vector, Neoaliturus tenellus (Circulifer tenellus). Bennett (1971) argued that BCTV was introduced to North America along with the vector. On the face of it, our results with a California isolate of BCTV support Oman's conclusion because this isolate of BCTV lies within the cluster of New World viruses (Fig. 2). However, it is also possible that BCTV originated in the Old World and has mutated since its introduction to the New World, in response to new host plants, to become similar to other New World geminiviruses. Similar to the issue of the origin of BCTV is that of CLV. Our results suggest that CLV originated in Africa and was present there before the introduction of cassava from South America. The lack of a South American geminivirus of cassava also supports this idea. However, we cannot eliminate the possibility that CLV originated in South America and was introduced to Africa with cassava. Our results may also have a bearing on the definition of viruses and strains. We included examples of viruses that have been regarded as three strains each of MSV and WDV. Our results corroborate the use of the term 'strain' for these viruses. For example, MSV strains not only have the same vector and hosts, but they have only very small mutation distances (Tables 1 and 2). Their mutation distances were 8 or less in the replication-associated protein tree and 6 or less in the coat protein tree, whereas the smallest mutation distance between any other pair of viruses was more than 13-fold higher (131 for T G M V and BGMV-BZ) in the replication-associated protein tree and four-fold higher (28 for T G M V and BGMV-BZ) in the coat protein tree. It is Downloaded from www.microbiologyresearch.org by IP: 88.99.165.207 On: Fri, 16 Jun 2017 22:41:01 2726 A.J. H O W A R T H AND G . J . VANDEMARK interesting, but not too surprising, that the two viruses named BGMV-PR and BGMV-BZ are not each other's closest relative. Reasons for this may include that they were collected at distantly separated sites, Puerto Rico and Brazil; BGMV-PR was selected as a mechanically transmissible isolate and BGMV-BZ was not; BGMV-PR was originally collected from Phaseolus lunatus and BGMV-BZ was collected from Phaseolus vulgaris; ultimately, they were named BGMV because they cause golden mosaic symptoms on bean and are transmitted by whiteflies. We conclude that BGMV-PR and BGMV-BZ may not be strains of the same virus because of their closer relationships to other viruses than to each other. It seems clear that additional taxa should be erected to classify the two different types of geminiviruses that are included in the group. If current nomenclature is retained then subgroups for monocot- and dicot-infecting geminiviruses, respectively, should be defined. It would be simple to convert this hierarchical structure to a family with two genera (Matthews, 1985), if such a system is approved in the future. This classification would remove the problem of having MSV as the type geminivirus (Matthews, 1982; Stanley, 1985), a status which is not consistent with what we know about the structure and other features of these viruses. As more geminiviruses are characterized, standardization in naming the two D N A segments is desirable; the precedent of Stanley & Gay (1983) should be observed. We thank D. P. Maxwell, H. Jeske and T. Frischmuth, and H. Czosnek and N. Navot for sharing sequences prior to publication, G. D. F. Wilson for providing the MS-DOS version of the Phylip package, and J. G. Utermohlen, K. D. Hirschi, and M. C. Hawes for comments on the manuscript. This work was supported by USDA Hatch and C R G O (85-CRCR-l-1793) grants and this paper is no. 7043 in the journal series of the Arizona Agricultural Experiment Station. REFERENCES ANDERSEN,M. T., RICHARDSON,K. A., HARBISON,S. & MORRIS,B. A. M. (1988). Nucleotide sequence of the geminivirus Chloris striate mosaic virus. 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