Mosaic is the most destructive virus disease of sugarcane (Koike and Gillaspie, 1989; Xu et al., 2008). Sugarcane mosaic virus (SCMV; genus Potyvirus, family Potyviridae) is one of the causal pathogens of mosaic diseases on sugarcane, maize, sorghum and some other graminaceous species with worldwide distribution and significant yield losses (Achon et al., 2007; Alegria et al., 2003). Sugarcane originated from the island of New Guinea in the South Pacific and then spread to Northern India. It is now cultivated in several countries in the world, prominently in Brazil, India, China, Pakistan, Mexico, Thailand, Colombia, Australia, Indonesia, Philippines, and USA (He et al., 2016). In Iran, sugarcane is one of the most important commercial crops cultivated in north and southwest of the country. Viral diseases such as those caused by SCMV are a potential threat to sugarcane industry (Moradi et al., 2016). SCMV was first reported in sugarcane from Khuzestan Province of Iran in 1993 (Amiri and Izadpanah, 1993). Like other potyviruses, the virus has a positive single-stranded RNA genome encoding a single polyprotein and is transmitted by aphids in a non-persistent manner. The polyprotein is cleaved by three self-encoded proteinases to produce at least ten final protein products, which are multifunctional (Adams et al., 2012). The coat protein (CP) is a protein involved in several steps of the potyvirus life cycle: aphid transmission, cell-to-cell and systemic movement, encapsidation of the viral RNA, regulation of viral RNA amplification (Urcuqui-Inchima et al., 2001) and probably host specificity (Li et al., 2013; Shukla et al., 1991). The variable N-terminus of the potyvirus CP is necessary for aphid transmission, systemic infection, and adaptation of the virus to its host (Ullah et al., 2003). Differences in this region are important for the potyvirus discrimination (Handley et al., 1998; Shukla and Ward, 1989; Zhong et al., 2005). Evolutionary studies of viruses concentrated on understanding effects of variation caused by mutation, recombination, selection pressure, and host adaptation in viral populations. These studies help us to understand important aspects of viral biology such as changes in virulence, geographical spread and adaptation to new hosts or emergence of a new epidemic (Gibbs and Ohshima, 2010; Li et al., 2013; Moury et al., 2002). Hence, information on evolutionary history of viruses is crucial to design management and control strategies to contain their spread. Non-persistently transmission of SCMV by aphids makes the control of the virus vectors rather inefficient. Therefore, cultivation of resistant varieties is the most promising approach for control of SCMV (Dussle et al., 2002) which in turn requires a complete understanding of the genetic diversity of the virus. Several studies have been performed in recent decades on SCMV biology, genome characterization and sequence diversity (Achon et al., 2007; Alegria et al., 2003; Gao et al., 2011; Wang et al., 2010). Recently, Li et al. (2013) analyzed the genetic diversity and population structure of SCMV based on CP coding region. They suggested that gene flow, recombination and selection pressures were important driving forces for the evolution of SCMV. Based on complete genome sequence analysis, Xie et al. (2016) showed that host, negative selection and recombination were important evolutionary factors shaping the genetic structure of SCMV populations in Shanxi province of China. To our knowledge, little is available on the genetic variability and recombination analysis of the CP gene of SCMV population in Iran. The present study was carried out to investigate the prevalence and genetic diversity of the virus in Khuzestan and Mazandaran provinces of Iran, the major growing regions of sugarcane and maize in the country. To this end, the CP gene sequences of Iranian and worldwide isolates of SCMV were analyzed to know more about the molecular evolution of SCMV populations in Iran.

At least three viruses, including SCMV, Sorghum mosaic virus, and Sugarcane streak mosaic virus in the family Potyviridae infect sugarcane and cause mosaic disease (Perera et al., 2009; Xu et al., 2008, 2010). As such, the findings of this study suggest that SCMV is a common causative agent of mosaic in sugarcane and maize in Iran. In this study, SCMV was found to naturally infect sorghum and Johnson grass plants grown nearby or some distance away from sugarcane and maize fields. According to RT-PCR results, SCMV was present in 89.2% of the collected samples from all visited areas. The sugarcane varieties under cultivation in Iran are widely infected with SCMV as evidenced by more than 92% infection in our samples (Table 1). This high infection rate could be attributed to vector transmission manner (aphids), vegetative sett planting, and the presence of different host plants (crop and weed). For designing better epidemic control strategies, understanding the important features of RNA viruses, such as the molecular basis of virus geographical range, epidemiological routes, population structure, adaptation to new hosts or emergence of a new virus epidemic is crucial (Elena et al., 2011; Gibbs and Ohshima, 2010; Jones, 2009). For this purpose, the CP sequences of 27 SCMV isolates obtained from different hosts and distant geographic regions of Iran were determined, and their genetic relationships with CP sequences of 76 worldwide isolates of the virus were studied. Pairwise nucleotide sequence identity scores showed that global SCMV CP sequence similarity was between 75.2% and 99.9%. This genetic diversity divided the 103 isolates into five main divergent phylogenetic groups. The rate of genetic diversity and nonsynonymous to synonymous mutations (dN/dS) were significantly different between these five groups. In phylogenetic analysis of CP sequences, Iranian isolates were clustered together with isolates from Egypt, Australia, Argentina, USA, Brazil, India, Pakistan, South Africa, and China. Many studies have been previously done on the phylogenetic relationships of SCMV isolates (Achon et al., 2007; Alegria et al., 2003; Gao et al., 2011; Gemechu et al., 2006; Ha et al., 2008; Li et al., 2013; Wang et al., 2010; Xu et al., 2008). According to previous researches, SCMV isolates clustered in relation to their original hosts, and geographically distinct isolates from sugarcane or maize clustered differently (Li et al., 2013). Interestingly, based on a recent study, the complete genome sequences of two Iranian isolates could not be grouped according to geographical origin or host plant (Moradi et al., 2016) as well as GD (Guangdong, China, AJ310105) (Achon et al., 2007) and Ohio (USA) isolates. In the present study, genetic differentiation and phylogenetic analysis of CP gene of 103 SCMV isolates, consisting of 27 isolates from Iran, revealed that the population structure of the five SCMV phylogroups, were not correlated with their geographical origin or host. With the exception of Iranian isolates, from different hosts, group I consisted of members that originated from sugarcane, while isolates in groups II, III, IV, and V derived from sugarcane, maize, maize/sugarcane, and sugarcane, respectively. Among the five groups in the CP-based phylogenetic tree, groups II, IV, and V were consistent with the reported groups E: SO, C: SCE/MZ, and B: NSCE (Gao et al., 2011), respectively. However, all Iranian isolates and the previously reported isolates in D: SCE group were integrated into group I. Furthermore, isolates in A: MZ group were also integrated into group IV. Such structuring of virus isolates might be related to bottlenecks imposed by the host plant, potential arthropod vectors, and presence of physical and quarantine barriers between countries. Since SCMV can be transmitted through vegetative sett, the germplasm exchange could be another possible explanation for such high sequence similarities observed among the intercontinental isolates of SCMV (Padhi and Ramu, 2011). The higher genetic diversity between groups than within groups of SCMV populations suggests that there is more frequent gene flow within groups than between groups. The study conducted by Li et al. (2013) reported that SCMV populations between maize and sugarcane had relatively frequent gene flow (F_ST < 0.33); but within maize or sugarcane geography distinct isolates had infrequent gene flow (F_ST > 0.33). Similarly, no significant difference was detected between maize and sugarcane (also other plants) isolates from Iran (F_ST < 0.33), indicating that the gene flow between SCMV populations of maize and sugarcane was frequent. However, the close genetic affinities observed between some geographically distant SCMV isolates suggest long-distance migration, probably due to international traffic of propagative sugarcane materials. Historical accounts support the hypothesis that the SCMV isolates were most likely introduced into Iran via importation of sugarcane germplasm materials. For instance, the NRA and ZRA SCMV isolates were introduced into Iran probably from sugarcane-producing countries (Australia, USA, Argentina, India, and Egypt) via contaminated vegetative cuttings (Moradi et al., 2016). Subsequently, further spread of SCMV within Iran could have occurred via movement of contaminated vegetative cuttings across provinces to other host plants such as maize, sorghum, and so on and within provinces via aphid vectors. Mutation and recombination are two main sources of genetic variation in RNA viruses (Gell et al., 2015; Mangrauthia et al., 2008; Moreno et al., 2004; Padhi and Ramu, 2011). Recombination events are common in Potyvirus evolution and can be detected throughout the genome (Chare and Holmes, 2006; Revers et al., 1996; Sztuba-Solińska et al., 2011; Valli et al., 2007). Recombination may play an important role in SCMV evolution, and be an important reason for SCMV genetic diversity (Padhi and Ramu, 2011; Xie et al., 2016). Previously, several recombination events have been reported in SCMV and the probability of recombination was diverse for different genes (Achon et al., 2007; Moradi et al., 2016; Padhi and Ramu, 2011; Xie et al., 2016; Zhong et al., 2005). Based on the CP gene of SCMV, six recombinants were detected (Li et al., 2013). In addition, two isolates from Iran, NRA and ZRA were identified as recombinants in the P1 (with highly significant P-values), HC-Pro, CI, NIa-Vpg, and NIa-pro coding region (Moradi et al., 2016). These results point to the importance of genetic recombination in the evolution of global SCMV populations. However, no signature of recombination was detected in CP gene of Iranian SCMV isolates identified in this study. Natural selection is an important evolutionary mechanism limiting variation of virus populations (Pérez-Losada et al., 2008). In the present study, no significant (P > 0.10) deviation from neutrality was detected for almost any of the groups using MKT. On the other hand, a purifying selection was detected in CP genes of SCMV isolates, enhancing the speed of elimination of deleterious mutations in genes and shaping the stable genetic structure of the population. It is also noteworthy that most of the codons in the CP gene were under negative selection or neutral evolution except for codons 27 and 48, which were under positive selection (Li et al., 2013). In contrast to previous studies, Iranian SCMV isolates were not grouped according to host plant or geographical origin which could be explained by virus transmission characteristics, maize or sugarcane germplasm exchange, continental barriers, host plant, and so on. According to the results, SCMV isolated from maize or sorghum is greatly similar to SCMV isolated from sugarcane. One possibility is introduction of sugarcane from different sources to the country. On the other hand, in Iran, maize and sorghum are frequently planted nearby sugarcane crops and weed grasses such as Sorghum halepense, are commonly found in sugarcane fields. Based on the negative selection, SCMV could constantly accumulate the available variations to adapt to the different hosts. Meanwhile, those plants naturally infected by SCMV could serve as potential reservoirs of the virus in the field. Therefore, avoidance of planting maize near sugarcane crops and control of those weeds are important to mitigate the virus infection. Taken together with this study and previous researches (Alegria et al., 2003; Gao et al., 2011; Li et al., 2013; Xie et al., 2016), differences existed between isolates from different hosts or within the same host from different locations, indicating the complexity of genetic structure of the SCMV populations. Nevertheless, it seems that Iranian SCMV isolates based on complete genomic sequences (Moradi et al., 2016) and CP gene sequences are not grouped according to geographical origin or host plant. Frequent gene flow, negative selection and homologous recombination appear to be important driving forces acting synergistically to limit genetic variation within these SCMV isolates. The present study provides a comprehensive analysis of SCMV population structure based on CP gene and the driving forces involved in its evolution. This information will provide a base for evaluating the epidemiological features of SCMV in Iran and will be suitable in designing long-term, sustainable management strategies for SCMV. Further information of SCMV isolates from other geographic zones and hosts from Iran are needed to fully assess the molecular diversity and evolutionary history of the virus.