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Article

High-Throughput Sequencing Reveals Differential Begomovirus Species Diversity in Non-Cultivated Plants in Northern-Pacific Mexico

by
Edgar Antonio Rodríguez-Negrete
1,†,
Juan José Morales-Aguilar
2,†,
Gustavo Domínguez-Duran
2,
Gadiela Torres-Devora
2,
Erika Camacho-Beltrán
2,
Norma Elena Leyva-López
2,
Andreas E. Voloudakis
3,
Eduardo R. Bejarano
4 and
Jesús Méndez-Lozano
2,*
1
Consejo Nacional de Ciencia y Tecnología (CONACYT), Instituto Politécnico Nacional, CIIDIR-Unidad Sinaloa, Departamento de Biotecnología Agrícola, Guasave, Sinaloa 81101, Mexico
2
Instituto Politécnico Nacional, CIIDIR-Unidad Sinaloa, Departamento de Biotecnología Agrícola, Guasave, Sinaloa 81101, Mexico
3
Laboratory of Plant Breeding and Biometry, Agricultural University of Athens, 75 Iera Odos, Athens 11855, Greece
4
Instituto de Hortofruticultura Subtropical y Mediterránea “La Mayora”, Universidad de Málaga-Consejo Superior de Investigaciones Científicas (IHSM-UMA-CSIC), Universidad de Málaga, Campus Teatinos, 29071 Málaga, Spain
*
Author to whom correspondence should be addressed.
†
These authors contributed equally to this work.
Viruses 2019, 11(7), 594; https://0-doi-org.brum.beds.ac.uk/10.3390/v11070594
Submission received: 26 March 2019 / Revised: 28 May 2019 / Accepted: 17 June 2019 / Published: 29 June 2019
(This article belongs to the Special Issue Plant Virus Ecology and Biodiversity)
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Abstract

:
Plant DNA viruses of the genus Begomovirus have been documented as the most genetically diverse in the family Geminiviridae and present a serious threat for global horticultural production, especially considering climate change. It is important to characterize naturally existing begomoviruses, since viral genetic diversity in non-cultivated plants could lead to future disease epidemics in crops. In this study, high-throughput sequencing (HTS) was employed to determine viral diversity of samples collected in a survey performed during 2012–2016 in seven states of Northern-Pacific Mexico, areas of diverse climatic conditions where different vegetable crops are subject to intensive farming. In total, 132 plant species, belonging to 34 families, were identified and sampled in the natural ecosystems surrounding cultivated areas (agro-ecological interface). HTS analysis and subsequent de novo assembly revealed a number of geminivirus-related DNA signatures with 80 to 100% DNA similarity with begomoviral sequences present in the genome databank. The analysis revealed DNA signatures corresponding to 52 crop-infecting and 35 non-cultivated-infecting geminiviruses that, interestingly, were present in different plant species. Such an analysis deepens our knowledge of geminiviral diversity and could help detecting emerging viruses affecting crops in different agro-climatic regions.

Graphical Abstract

1. Introduction

Agroecosystems are used for the production of food, feed, fuel, fiber, and other harvestable goods providing human support and health [1]. Mexico has 196,437,500 ha, of which approximately 13% correspond to agricultural land. In 2016, 21.9 million ha were cultivated, with an agricultural production of 26,032 million tons with a value of 26,760 million dollars, which allowed the country to be ranked eleventh in the world´s crop production.
Plant diseases caused by begomoviruses and RNA viruses have been the main concern in Mexican horticulture throughout the years, having a significant negative impact on the crop production of tomato, pepper, bean, pumpkin, melon, soybean, tomatillo, tobacco, watermelon, and cotton [2,3,4,5,6,7]. Begomoviruses (family Geminiviridae) are characterized by their geminate particles that encapsidate a circular single-stranded (ss) DNA genome (monopartite or bipartite) of about 2.8 kb in size. They are transmitted via whitefly (Bemisia tabaci), infecting a large number of plant species worldwide and causing serious crop losses, and constitute a major global threat [8]. Important diseases caused by geminiviruses include maize streak disease [9], cassava mosaic disease [10], cotton leaf curl disease [11], and tomato leaf curl disease [12]. In Mexico, the first report of a disease caused in tomato by geminiviruses came from the Sinaloa state in 1970, and was later designated as Chino del tomato virus (CdTV) [13]. However, the first serious disease was reported in pepper crops and was named “rizado amarillo”, identified as the coinfection with Pepper huasteco yellow vein virus (PHYVV) and Pepper golden mosaic virus (PepGMV) [14]. Recently, the introduction of Tomato yellow leaf curl virus (TYLCV) in Sinaloa has dealt a devastating impact on tomato production. In different agro-climatic regions of Mexico, begomovirus-associated diseases are commonly caused by mixed infections with diverse begomoviruses, and affect tomato and pepper crops [5,15,16]. Coinfections with begomoviruses adapted to non-cultivated plants together with crop-adapted begomoviruses have been reported in soybean, tobacco, and pepper plants in the Sinaloa, Chiapas, and Jalisco states of Mexico [17,18,19]. Geminiviruses exhibit high mutation rates and recombination frequencies, both within and between species, resulting in rapid adaptive evolution. Several reports have indicated the emergence of recombinant species of geminiviruses [20,21,22,23]. For example, Tomato yellow leaf curl Malaga virus (TYLMaV) is a recombinant of Tomato yellow leaf curl Sardinia virus (TYLCSV) and Tomato yellow leaf curl virus mild strain (TYLCV) which, unlike its “parental” genomes, has the ability to infect common bean and wild Solanum nigrum [24]. More importantly, TYLMaV accumulated to the same levels in susceptible and resistant tomato, indicating that the recombination event and subsequent selection led to the generation of a resistance-breaking isolate [25].
It has been proposed that global warming will influence the epidemiology of virus-related plant diseases, mainly due to alterations in the distribution of insect vectors and in their host range [26,27]. Emerging diseases caused by geminiviruses have arisen more frequently in recent years, which emergence could be associated with climate change [28]. The viral quasispecies (non-identical but related genomes) present in a host plant could be generated by recombinant genomes providing an improved fitness potential to the viruses that may subsequently initiate an infection in new host species, or cause more severe disease symptoms in an established host by overcoming plant resistance. The generation of quasispecies depends on the host-virus interaction, the environmental conditions, and the cultivation practices [9,29,30,31].
New mutant or recombinant viral genomes that have arisen in non-cultivated plants could achieve successful infection of plant hosts to which they have newly adapted and cause significant crop damage, in a process subject to effective transmission between plant species. Vector-transmission is an evolutionary barrier for plant viruses to expand their host range. Alterations in vector-dependent transmission, most likely through mutations in the viral coat protein sequences, could increase the risk of emergence of a plant pathogen in a given crop. Begomoviruses are transmitted by the whitefly Bemisia tabaci, while aphid vectoring was recently confirmed for the Capulovirus Alfalfa leaf curl virus (ALCV) [32,33]. Vector metagenomics could contribute to surveying the diversity of geminiviruses present in their insect vectors and to defining the corresponding coat protein sequences. Although coat proteins have a structural role for the viral capsid, CP gene sequences are known to diverge [34] due to a high mutation rate. In addition, it is possible that such CP mutations could facilitate new means of transmission, such as seed transmission. Therefore, it is important to study the complexity and heterogeneity of geminiviral quasipecies found in wild reservoir hosts, and understand the factors behind the observed genetic variation since such knowledge will be extremely useful to develop resistance strategies (e.g., RNAi-based approaches) and, as a consequence, to prevent crop losses.
High-throughput sequencing (HTS) has provided the means to detect known viruses as well as to identify novel viruses in plants [35,36,37,38,39]. As a result, sensitive and accurate diagnosis of viral infection has been achieved, rendering this method extremely useful for quarantine purposes. The development of bioinformatic tools and the design of various pipelines have contributed significantly towards a deep analysis of the vast amount of HTS data produced [40].
The current next generation sequence (NGS) technologies have a couple of drawbacks: firstly, the contigs/singletons need to be annotated de novo via short read assembly, a process that may create chimeras deriving from the different genomes present in a sample, and secondly, the accurate differentiation of sequences; thus, confirmation is required by cloning followed by Sanger sequencing. The hope is that the latest single-molecule NGS technologies (3rd generation), where long reads are obtained, could address both of the above-mentioned issues, especially when their sequence reading error rates drop significantly compared to the levels of the previous NGS technologies. The resolution of the metagenome of a given sample could identify genetic variations in a viral population, providing the necessary input to study viral genome evolution, and determine which environmental factors might influence the generation of novel plant pathogens from previously benign viruses.
It is a well accepted notion that non-cultivated (wild) host plants play a key role in the generation of viral genetic variation, maintaining a certain degree of sequence heterogeneity convenient for viral adaptation in nature, without altering essential consensus sequences. Only recently, metagenomics studies on non-cultivated plant species have attracted the attention of plant researchers [38,41]. Mexico is considered to be one of the most megadiverse countries worldwide [42] and, despite the fact that some non-cultivated plant species have been reported as geminivirus reservoirs [16,43], to date the knowledge of geminivirus distribution in Mexican natural ecosystems is still limited. To this purpose, we aimed to determine the genetic diversity of begomoviruses in non-cultivated species that are present in the vicinity of cultivated crops (designated as the agro-ecological interface) in Northern-Pacific Mexico. In the present study, rolling circle amplification (RCA) was applied to DNA samples from wild plant species obtained during a five-year survey, and several begomoviral DNA signatures and begomoviral sequences were identified. Our results support the presence of a niche for begomoviral evolution in the neighborhood of important cultivation areas in Northern-Pacific Mexico.

2. Materials and Methods

2.1. Plant Sample Collection

A total of 422 non-cultivated plants (both symptomatic and asymptomatic) located in the area between crops and wild vegetation zones (agro-ecological interface), in seven states in the northern-pacific region of Mexico during the period 2012–2016, were collected, GPS-documented, photographed, and identified to the species level. Thus, 132 species of plants belonging to 34 families were identified. The sampling regions were grouped as follows: (1) Baja California (BC), (2) Sonora (SO), (3) Sinaloa (SI), (4) Colima-Nayarit (CN), and (5) Coahuila-Durango (CD) states of Mexico. Samples were placed on ice and brought to the laboratory and stored at −80 °C until processed. Plants were collected in non-protected areas; additionally, an herbarium was established with most of the plant specimens.

2.2. DNA Isolation, RCA, and Library Construction

Total DNA was extracted from individual plants using the CTAB method [44], and the isolated DNA, upon spectrophotometric estimation of its concentration, was used as template for PCR-based Begomovirus detection using degenerate universal primers (Supplementary Table S3). For each sampling region, total DNA from Begomovirus PCR-positive plants, belonging to the same plant species, was mixed in equimolar concentration. Following this procedure, 100 ng of each DNA mixture was used for circular DNA-molecule enrichment by rolling circle amplification (RCA) using the illustra TempliPhi DNA Amplification Kit (GE Healthcare, Milwaukee, MA, USA), following the manufacturer´s instructions. Then, all RCA products obtained per sampling region were pooled in equimolar concentrations, and cleaned using phenol:chloroform:isoamyl alcohol (25:24:1)/potassium acetate (5 M) and 100 ethanol % precipitation (1/10 v/v, 1/2 v/v, respectively). DNA integrity was analyzed by agarose gel electrophoresis, and the cleaned RCA mixtures were used for NGS library construction that was sequenced by a commercial facility (LANGEBIO, Irapuato, GTO, MX) using Illumina Nextera XT paired end 2 × 150 bp protocol on a MiSeq 500. The same procedure was followed for each sampling region to obtain one library per region (for a total of five libraries).

2.3. Metagenomic Analysis of Geminivirus-Related Signatures

Reads obtained from each library were trimmed employing the trimmomatic tool [45] (parameters (TRAILING: 30, HEADCROP:5) followed by quality check analysis by FASTQC (https://www.bioinformatics.babraham.ac.uk). Each library was filtered for human, bacteria, plant, and eukaryotic viruses reads using the ViromeScan pipeline [40] in order to obtain the geminivirus-related reads. All filtered libraries were subjected to de novo assembly using SPAdes [46]; both contigs (≥78 bp) and unassembled reads were compared against the GeneBank non-redundant database using BLASTn hosted in the Galaxy server [47]. Geminivirus-related signatures were sorted by contig length and analyzed manually. Contigs obtained in the present study are available at: https://www.dropbox.com/sh/ha6pkzls9217dhf/AAADNUa0TfYj3EZ8bb315cSga?dl=0.

2.4. Begomovirus Full-Length Genome Amplification, Cloning, and Sequence Analysis

Full-length geminiviral genomes were obtained following a previously described protocol [48]. In brief, the total DNA (100 ng) from selected non-cultivated plants was used as a template for viral circular DNA genome enrichment by RCA, as mentioned above. To obtain the viral monomeric full-length genomes, the RCA products were digested with a selected a single-site restriction enzyme (BamHI, EcoRI, XbaI, or XhoI, depending on the virus under analysis). The expected linearized geminivirus full-length genomes (~2.7 kb) were recovered from 1% ultrapure agarose gels using PureLink Quick Gel Extraction Kit (Thermo Fisher Scientific, USA) according to the manufacturer´s instructions. The fragments were ligated into linearized pGreen 0029 plasmid [49] that was digested with the corresponding restriction enzymes. The resulting recombinant plasmids were transformed in E. coli DH5α, and positive clones were subjected to Sanger genome walking sequencing. Genome assemblies were obtained using SeqMan (DNASTARInc, Madison, WI, USA) and SnapGene (GLS Biotech LLC, Chicago, IL, USA) software. All pairwise comparisons were performed using the MUSCLE algorithm implemented in Mega 7 [50] and maximum likelihood phylogenetic tree(s) were constructed on both begomoviral DNA components, with a 1000 bootstrap on both components to assess branch support. To analyze the nucleotide and amino acid identity, open reading frames (ORFs) were separated and individually compared with highest match homologous genome of each virus obtained from NCBI databank, using the ClustalW algorithm implemented in Mega 7.

3. Results and Discussion

It is an accepted notion that global warming will have an impact on global food security. In particular, crop yields are predicted to significantly decrease considering the “worst” CO2-emission scenario (A1FI) put forward by the Intergovernmental Panel on Climate Change [51]. Plant pathogens will have varying responses to climate change and plant-pathogen warfare is expected to be altered [52], imposing negative, neutral or positive effects on yields depending on the host-pathogen-environment tripartite interaction (known as the ‘disease triangle’). Disease pattern changes are anticipated due to alterations in the host range of plant pathogens, especially in rapidly evolving pathogens, while disease severity will be influenced by increased CO2, heavy rains, increased humidity, drought, and warmer winter temperatures [53]. Studies towards the understanding of the existing genetic diversity of plant viruses occurring in the agro-ecological interface, the generation of new genomes with features advantageous to the pathogen, and the relationship with their corresponding vectors will contribute significantly to humankind’s preparation to adapt to climate change and to sustain future food production. In this study, high-throughput sequencing (HTS) was employed to determine begomoviral diversity in plant samples collected in Northern-Pacific Mexico.

3.1. Non-Cultivated Plants from Northern-Pacific Mexico Region as a Reservoir of Begomoviruses

Plant viruses have generally been studied as disease-causing infectious agents that have a negative impact on their hosts [54]. Considering the wide diversity of geminiviruses, non-cultivated plants may serve as reservoirs for known, agriculturally relevant viruses: such weed-hosted viral species hold great potential to initiate the future emergence of new diseases in cultivated plants [55]. To determine begomoviral diversity in Northern-Pacific Mexico, a survey was performed in the agro-ecological interface in the vicinity of crop sites during 2012–2016. Sampling areas were divided in five regions: Baja California, Sonora, Sinaloa, Colima-Nayarit, and Coahulia-Durango, with three, four, seven, two, and four sampling points, respectively (Figure 1a). A total of 422 non-cultivated plants (both symptomatic and asymptomatic), belonging to 34 families and 132 species were identified (Supplementary Table S1 and Supplementary Figure S1). Among the different families identified in each region, the most commonly distributed were the Astaraceae, Solanaceae, Malvaceae, and Fabaceae. It is worth mentioning that those families were found to be the most predominant in natural ecosystems considering 200 plant families described in Mexico [42,56]. Using degenerate universal primers based on the DNA A genome of the genus Begomovirus, we were able to amplify the expected DNA fragment of 950–1100 bp in 252 out of the 422 (60%) tested individual plant specimens, indicating that begomoviruses were present in 29 plant families collected from all five regions sampled (Table 1). These results suggest that a number of non-cultivated plants widely distributed in Northern-Pacific Mexico represent a reservoir for begomoviruses.

3.2. Metagenomics Study Reveals a Number of Geminviruses from Non-Cultivated Plants

Following the pipeline described in the Materials and Methods section, NGS resulted in 16 to 215 million reads for the five libraries. After human, bacteria, plant, and eukaryotic virus sequence depletion, an NCBI-GenBank database search was performed to identify the most closely related geminivirus sequences, obtaining between 6000 and 4.6 million reads (Table 2). Subsequent annotation showed that more than 99% of the reads correspond to the genus Begomovirus, and the remaining 1% matches the Geminiviridae genera Curtovirus, Becurtovirus, Turncurtovirus, Topocuvirus and Mastrevirus. No sequences with homology to genera Capulavirus, Eragrovirus, and Grablovirus were identified in our study. The genus Begomovirus has been reported previously as the most widely distributed in Mexico [3,4,16,18,57,58,59]; in addition, sporadic reports on other genera like Curtovirus and Grablovirus were described [60,61]. Our data pointed out the abundance and importance of the genus Begomovirus in Mexico; however, follow up studies of the other genera are imperative.
The de novo assembly of geminivirus-related reads resulted in 24,289 geminivirus-related contigs ranging from 78 to 2858 bp in length (Table 2). The generated contigs were used to search the NCBI-GenBank database in order to identify the most closely related geminivirus exemplars at the species level (Supplementary Figure S2). Table 3 shows all geminivirus-related signatures ≥300 bp and with at least 80% similarity at the nucleotide level with the best match, regardless whether DNA A or B viral components were detected. Similar findings were described in a metagenomics analysis in whiteflies, in which only one DNA component of a bipartite begomovirus was retrieved [62]. Additionally, the geminivirus-related signatures with 100–300 bp and/or below <80% similarity at the nucleotide level with the best match in NCBI gene sequences are listed in Supplementary Table S2. It is important to note that short geminivirus-related signatures could hinder the correct classification of a begomovirus species or strain; nonetheless, profiling the phylogenetic composition of the viral communities is pivotal as a significant part of the different plant-virus-environment interaction.
The analysis of the highest geminivirus-related signature sequences revealed a list of both bipartite and monopartite begomoviruses, including crop-adapted viruses in different plant families; 14 with bipartite genomes such as Pepper huasteco yellow vein virus (PHYVV-signature) present in four regions, Pepper golden mosaic virus (PepGMV-signature) present in four regions, Pepper leafroll virus (PepLRV-signature) present in one region, Tomato chino la Paz virus (ToChLPV-signature) present in two regions, Tomato severe leaf curl virus (ToSLCV-signature) present in two regions, Tomato yellow spot virus (ToYSV) present in three regions, Potato yellow mosaic virus (PYMV) present in one region, Okra yellow mosaic mottle virus (OYMMV-signature) present in four regions, Cabbage leaf curl virus (CabLCV-signature) present in two regions, Bean calico mosaic virus (BCaMV-signature) present in four regions, Bean yellow mosaic Mexico virus (BYMMV-signature) present in one region, Vigna yellow mosaic virus (ViYMV-signature) present in two regions, Water melon chlorotic stunt virus (WmCSV-signature) present in one region and Squash leaf curl virus (SLCV-signature) present in three regions; and five with monopartite genomes, three belonging to the genus Begomovirus, namely Chilli leaf curl virus (ChiLCV-signature) present in one region, and Tomato yellow leaf curl virus (TYLCV-signature) present in four regions, Sweet potato leaf curl virus (SPLCV-signature) present in one region; additionally, one belonging to the genus Curtovirus, namely Beet curly top virus (BCTV-signature) and another belonging to genus Topocovirus, namely Tomato pseudo-curly top virus (TPCTV), both present in one region (Table 3). Furthermore, nine non-cultivated plant-adapted viruses included only begomoviruses with bipartite genomes such as Solanum mosaic Bolivia virus (SoMBoV-signature), present in two regions, Sida mosaic Sinaloa virus (SiMSiV-signature) present in five regions, Sida golden yellow spot virus (SiGYSV-singnature) present in one region, Malvastrum bright yellow mosaic virus (MaBYMV-signature) present in three region, Rhyncosia golden mosaic virus (RhGMV-signature) present in five regions, and Rhyncosia golden mosaic Sinaloa virus (RhGMSV-signature) present in two regions, Euphorbia mosaic virus (EuMV-signature) present in three regions, Euphorbia yellow mosaic virus (EuYMV) present in one region and Blechum leaf curl virus (BlelCV-signature) present in one region, (Table 3). Interestingly, the observation of Tomato pseudo-curly top virus (TPCTV) in samples from Colima-Nayarit is the first report of the genus Topocovirus in Mexico. The list of geminiviruses are grouped as a potential of new viruses or strain of the best match virus, with molecular and biological validation being necessary.
The genus Begomovirus comprises the most common DNA viruses responsible for several virus-associated plant diseases in Mexico. Among them PHYVV, an endemic virus, and PepGMV have been documented as the most widespread and predominant in pepper crops [5,14,57,63]. It is noteworthy that the introduction of the promiscuous TYLCV in Yucatán and later in Sinaloa states, with a dramatic impact on crop yield, became the major concern for tomato crops in Northern Mexico [64]. Interestingly, TYLCV did not exclude the “native” viruses, and its co-infection with PHYVV or PepGMV caused severe disease in pepper crops [65]. However, the presence of such viruses in non-cultivated plants, as alternative hosts in nature, increases the chances of viral evolution through recombination events or other mechanisms, representing a latent threat. In fact, a new isolate of PHYVV described in pepper had significant sequence changes on its DNA B genome, which confers a modified host range since this isolate was able to infect tomato plants causing severe symptoms [5,66]. The other invasive virus which was introduced in Sonora state, apparently from the Middle East, was WmCSV [6], and it was detected in the present study along with SLCV. Our results suggest that WmCSV and SLCV are present in non-cultivated plants collected in Coahuila-Durango region (Table 3), which implies that WmCSV virus has the potential to spread in Mexico, and by adapting to new environments it has the potential to initiate an emerging disease in a new region. It is important to mention that both viruses could interact in mixed infection inducing more severe symptoms on crops as reported in Jordan [67]. The detection of ToChLPV, ToSLCV, OYMMV, BCaMV, and BYMMV, that were reported previously in Mexico, indicate that they still occupy an ecological niche and could be a potential source of viral disease. The identification of SiMSiV and RhGMV in all regions sampled is intriguing. Perhaps both of them represent viruses that are well adapted to different hosts and environments with a potential risk to evolve into an emerging disease. SiMSinV was initially reported in Sinaloa state associated to Sida rombifolia [17] without a negative impact on crops described to date. On the other hand, RhGMV was previously reported as the cause of disease in tobacco and soybean [4,18]. It is well known that viruses adapted to non-cultivated plants do not normally induce disease symptoms in their hosts. Nonetheless, SiMSiV and RhGMV induce symptoms in their corresponding first reported hosts (Sida rombifolia and Rhyncosia minima, respectively), with different wild species (acting as reservoirs) being suspected as the origin of the inoculum. Moreover, EuMV and EuYMV were reported previously in Euphorbia heterophylla in Mexico and Brazil [68,69]; whereas BlelCV, is a novel virus recently described in Chiapas state [70]. To the best of our knowledge, the ChiLCV, PepLRV, ToYSV, PYMV, CabLCV, SPLCV, MaBYMV, and ViYMV geminivirus-related signatures have not been previously described and/or associated to disease in Mexico and represent potential strains and/or novel viruses whose biological role needs to be determined in the immediate future. It is worth mentioning that viruses like ChilCV, PepLRV, and ToYSV are already associated with pepper, bean, and tomato diseases in Pakistan, Peru, Ecuador and Brazil [71,72,73].

3.3. Molecular Validation of the Predominant Begomoviruses Identified by HTS

The ecology of begomoviruses identified by HTS studies in non-cultivated plants requires more effort in order to understand the contribution of these plants for disease development [74,75,76]. Non-cultivated plants could be a source of viral inoculum to cultivated plants [77,78,79,80,81,82,83] and could contribute to viral evolution. Here, we described some non-cultivated plants at the agro-ecological interface carrying geminivirus-signatures (Table 3 and Supplementary Table S2). The information acquired represents important progress towards elucidating the above-mentioned issues, but more work is needed for the validation of the described identities.
According to our survey, plants belonging to the Fabaceae, Malvaceae, and Solanaceae families are the most widely distributed in all sampled areas. The HTS analysis revealed that signatures corresponding to TYLCV, SiMSiV, and RhGMV were present in all five NGS libraries, whereas the RhGMSV-signature was detected only in two out of five NGS libraries, suggesting that these species of begomovirus are predominant. Initially, the presence of these four viruses was confirmed by using sequence-specific primers for each viral species (Supplementary Table S3), testing an individual plant of the corresponding family for viral presence. To obtain the full-length viral genome of detected viruses, total DNA of Nicotiana glauca from Sinaloa (TYLCV PCR-positive); of Sida acuta from Colima (SiMSiV PCR-positive), and two Rhynchosia minima both from Sinaloa (PCR-positive for RhGMV/RhGMSV), were used for RCA amplification and cloning. Sequence analysis of obtained clones is summarized in Table 4 and Table 5.
Clone LV15-Ng-04 (Accession number: MK643155) from N. glauca was 2781 nt in length and showed high nucleotide homology (99.9%) to TYLCV (Accession number: EF523478.1). Clones LV15-Sa-03 and LV15-Sa-02 (Accession numbers: MK636866, and MK643154), from S. acuta, were 2611 nt and 2583 nt in length and showed nucleotide homology of 95.1 and 91.3% with DNA-A and DNA-B of SiMSiV (Accession numbers: DQ520944.1, and DQ356428.1, respectively). Clones LV17-Rm-02 LV17-Rm-06 (Accession numbers: MK634355, and MK634539), from R. minima, were 2605 nt and 2568 nt in length and showed nucleotide homology of 98.6 and 91% with DNA-A and DNA-B of RhGMV (Accession numbers: EU339939.1, and DQ356429.1, respectively). Finally, clones LV15-Rm-02 LV15-Rm-08 (Accession numbers: MK618662, and MK618663), from R. minima, were 2578 nt and 2525 nt in length and showed nucleotide homology of 91.9 and 85.9% with DNA-A and DNA-B of RhGMSV (Accession numbers: DQ406672.1, and DQ406673.1, respectively). For all viral genomes obtained, the predicted stem-loop region containing the sequence TAATATTAC, found in the common region of family Geminiviridae, was identified. For bipartite begomoviruses, high nucleotide homology of the common region (CR), of 98, 91, and 87% (DNA-A versus DNA-B) was observed for SiMSiV, RhGMV, and RhGMSV, respectively. Furthermore, the array of regulatory elements (iterons and TATA boxes) was conserved in all cases, suggesting that corresponding DNA-A and DNB-B are cognates. According to the present taxonomic classification of ICTV [84], for family Geminiviridae, the clones of TYLCV, SiMSiV, and RhGMV obtained in this study are classified as strains (≥94% DNA-A nucleotide homology), whereas the RhGMSV clones are classified as different isolates (≥91% DNA-A nucleotide homology).
Phylogenetic trees based on the nucleotide alignment with selected begomoviruses from the GenBank database, are shown in Figure 2. The results show that TYLCV isolate LV15-Ng-04 from N. glauca cluster together with different TYLCV isolates from different regions of the world, and segregate more closely to Mexican and Asian isolates (Israel and China) (Figure 2A). These data are in agreement with the TYLCV classification by geographic area, where Asian and American isolates are placed in Group I [85]. SiMSiV is a Malvaceae-infecting virus, whereas RhGMV and RhGMSV are Fabaceae-infecting viruses. Phylogenetic analysis showed that SiMSiV (DNA A and B) isolated from S. acute is closely related to an isolate of SiMSiV from Sinaloa state (Figure 2B,C). Similarly, isolates of RhGMV (DNA A and B) isolated from R. minima is clustered with isolates previously reported from soybean and weeds from Sinaloa state (Figure 2B,C). Altogether, the HTS analysis strongly suggests the existence of biologically active viruses in the agro-ecological interface with the potential of developing novel or emerging crop diseases. Finally, infectious clones of TYLCV, SiMSiV, RhGMV and RhGMSV were also obtained (to be described elsewhere).

3.4. Ecogenomic Analyis of Predominant Begomoviruses

The metagenomic approach carried out in the present work, allowed us to identify the geminiviral diversity present in different plant families and geographical regions; however, it is crucial from an ecological point of view to determine the occurrence and dynamics of the viral community in non-cultivated plants.
Ecogenomic analyses of begomovirus were accomplished by sequence-specific PCR detection including PHYVV, TYLCV, SiMSiV, and RhGMV/RhGMSV as the most widely distributed species in Northern-Pacific Mexico (Table 3, Supplementary Table S3). Thus, a total of 126 individual species sorted into the predominant plant families (Fabaceae, Malvaceae, and Solanaceae) were examined for the dynamics of the individual plant-virus infections in the five sampling regions (Table 6, and Supplementary Figure S3). The analysis revealed that TYLCV was detected in 89 plants, emerging as the predominant virus, followed by SiMSiV, RhGMV/RhGMSV, and PHYVV with 63, 62, and 46 detections, respectively. Moreover, the number of single infections were lower, namely the observed corresponded to TYLCV (13.54%) followed by SiMSiV and RhGMV/RhGMSV (3.12 and 1.04%, respectively), suggesting that single infection is not common (Table 6, Figure 3). Interestingly, the dynamics of multiple (double, triple or quadruple) infections seem to be the rule in most of the plant species analyzed (Figure 3). The most common double viral infection detected was TYLCV-SiMSiV (14.58%), while TYLCV-SiMSiV-RhGMV/RhGMSV (11.45%) and TYLCV-PHYVV-RhGMV/RhGMSV (10.4%) were the most common triple infections. Furthermore, quadruple infections with TYLCV-PHYVV-SiMSiV-RhGMV/RhGMSV (31.25%) represented the highest viral complex found in different plant species and in the five agroecological regions included in these study (Table 6, and Figure 3).
TYLCV is the major viral concern for tomato production worldwide [78]. It was recently identified as a seed-transmitted virus [86] and is associated with crop diseases with other begomoviruses in both the Old and New World [87,88,89,90,91,92,93,94]. It has also been reported in many non-cultivated plant species worldwide including the families Amaranthaceae, Asteraceae, Chenopodiaceae, Convolvulaceae, Euphorbiaceae, Fabaceae, Malvaceae, and Solanaceae [95,96,97,98,99]. In the present work, TYLCV was detected in new host species of the Fabaceae family (Crotalaria juncea, Lonchocarpus lanceolatus, Macroptilium atropurpureum, Melilotus indicus, Rhychosia precatoria, Rhynchosia minima and Senna uniflora), the Malvaceae family (Abutilon trisulcatum, Anoda pentaschista, Herissantia crispa, kosteletzkua depressa, Malvastrum coromandelianum, Malvella leprosa, Melochia piramydata, Sida acuta, Sida rombifolia, Sidastrum lodiegensis and Sphaeralcea angustifolia), and the Solanaceae family (Datura discolor, Nicotiana plumbanginifolia, Nicotiana glauca, Solanum trydynamum, Solanum verbacifolium). PHYVV is an endemic virus of Mexico, described as a major concern for pepper production [5,14,63,65], additionally, it has been reported in several plant families [15,100]. Here, PHYVV was detected in new host species of the Fabaceae family (C. juncea, R. precatoria, R. minima and S. uniflora), the Malvaceae family (A. trisulcatum, H. crispa, K. depressa, M. coromandelianum, M. piramydata, S. acuta, S. rombifolia, S. lodiegensis), and the Solanaceae family (Datura stramonium, N. glauca, N. plumbanginifolia, Solanum elaegnifolium, Solanum rostrarum and Solanum verbascifolium). On the other hand, SiMSiV was reported infecting Sida rombifolia in Mexico; interestingly, this virus was detected not only in several other malvaceous plants (A. palmeri, A. pentaschista, H. crispa, K. depressa, Malva parviflora, M. coronomandelianum) but also in the Fabaceae family (C. juncea, L. lanceolatus, M. atropurpureum, R. precatoria, R. minima, S. uniflora) and the Solanaceae family (N. plumbanginifolia, Datrua stramonium, N. glauca, Solanum elaeagnifolium, Solanum nigrescens and S. rostrarum). Furthermore, RhGMV was first reported infecting Rhynchosia minima in Honduras [101] and after infecting tobacco and soybean crops in Chiapas and Sinaloa states from Mexico, respectively [4,18]. The data showed that RhGMV species (RhGMV and/or RhGMSV) were able to infect not only Rhynchosia minima but also other fabaceous plants (C. juncea, Macroptilium atropurpureum, R. precatoria, S. uniflora, Meliotus indica, and L. lanceolatus), and other hosts belonging to the Malvaceae (A. palmeri, A. trisulcatun, H. crispa, K. depressa, M. parviflora, M. coromandelianum, M. piramydata, S. acuta, S. rombifolia, and S. lodiegensis), and the Solanaceae (D. stramonium, N. glauca, N. plumbanginifolia, S. nigrescens, and S. rostrarum) families. This part of the study was oriented to individual plant samples, providing evidence about the ecology of the virus. The existence of several plant species harboring different viruses in multiple infections represents an important melting pot for the geminivirus to evolve in different directions triggered by vector and environmental factors [102]. Previous works have described extensively strains or new viruses in plants or whiteflies from different regions and some imply the host plant and biological properties [32,68,69,102]. Nonetheless, data from other studies were limited because the viral host was not determined [62,103]. The rate of discovery of new viral sequences is insufficient in terms of plant pathology [54,76], therefore an in-depth survey and biological determination is required to enhance our comprehension of the agroecological, environmental impact of viral evolution.
The evolution of plant viruses is a complex process involving multiple ecological and genetic factors resulting in host-pathogen co-evolution. Studies on viral diversity have been documented in a wide number of non-cultivated plants, with new entries described either as different strains of existing viruses or as new viruses [32,62,68,102,103,104,105,106]. However, it is noteworthy that viral species are often co-infecting the same plants with the possibility of genetic interaction, giving raise to inter- or intra-specific recombinant viruses resulting in more severe strains, as in the case of cassava mosaic diseases (CMD) [104,107] and tomato yellow curl diseases (TYLCD) [23,24,25,108]. Plant disease emergence requires that a virus from a reservoir host invades a new host resulting in new infection dynamics and viral adaptation [109]. In this sense, some geminiviruses acquired the ability to form a complex disease by assorting DNA A and B genomes, like ACMV and EACMV in Uganda [107] or those cases where the DNA A genome is similar to bipartite begomoviruses but the DNA B has not yet been described, such as TChLPV and ToSLCV in Mexico [7,58] or Datura leaf curl virus (DaLCV) in Sudan [110]. In the present work, the geminiviral diversity was described in different Mexican regions, showing that TYLCV, SiMSinV, PHYVV, RhGMV and RhGMSV are the predominant viruses. In addition, different multiviral complexes were detected in several plant species, highlighting the high frequency of mixed infections detected (Figure 3). However, a relevant issue is the wide host range observed for these viruses and that the genetic structure of the virome could be modified in an unpredictable manner. Moving forward, work is in progress aiming to determine whether new viruses or strains constitute a potential risk for Mexican agriculture.

Supplementary Materials

The following are available online at https://0-www-mdpi-com.brum.beds.ac.uk/1999-4915/11/7/594/s1, Figure S1: Representation of non-cultivated plants belonging to predominant plant families collected in natural ecosystem in five regions of northern-pacific Mexico, Figure S2: PCR detection of five geminivirus species in three predominant plant families of non-cultivated plants from northern-pacific Mexico, Table S1: Non-cultivated plants collected from northern-pacific regions of Mexico and determination of begomovirus host by PCR-test, Table S2: Geminivirus signatures obtained by de novo assembly from the metagenomic study, Table S3: List of PCR primers used in the present work title.

Author Contributions

Conceptualization, E.A.R.-N., A.E.V., and J.M.-L.; formal analysis, E.A.R.-N., J.J.M.-A., A.E.V., and J.M.-L.; investigation, E.A.R.-N., J.J.M.-A., G.D.-D., G.T.-D. and E.C.-B.; resources, J.M.-L.; data curation, E.A.R.-N., A.E.V. and J.M.-L. writing—original draft preparation, E.A.R.-N., A.E.V., and J.M.-L.; writing—review and editing, E.A.R.-N., A.E.V., N.E.L.-L., E.R.B., and J.M.-L.; supervision, A.E.V., N.E.L.-L. and J.M.-L.; project administration, J.M.-L.; funding acquisition, J.M.-L.

Funding

This research was supported by grants from CONACYT-Mexico (PDCPN No. 214950) and Instituto Politécnico Nacional (SIP 20131613, SIP20141282, SIP20151969, SIP20164812) and J.J.M.-A., G.D.-D., and G.T.-D were supported by a fellowship from CONACYT and BEIFI program of IPN.

Acknowledgments

We would like to thank Germán Bojórquez and José Saturnino Díaz for the valuable support in plant species identification. We thank M.A. Magallanes-Tapia for sample collection and support in the establishment of the herbarium. We also thank José Álvaro López, Patricia Castro, Claudia López, Edmycel Mandujano and especially for Dorin Ortiz Félix for their support in the administration of the grants. Finally, authors appreciate the English edition of the manuscript by Javier Ruiz Albert from University of Málaga.

Conflicts of Interest

The authors declare no conflict of interest.

References

  1. Garbach, K.; Milder, J.C.; Montenegro, M.; Karp, D.S.; DeClerck, F.A.J. Biodiversity and ecosystem services in agroecosystems. Encycl. Agric. Food Syst. 2014, 2, 21–40. [Google Scholar]
  2. Torres-Pacheco, I.; Garzón-Tiznado, J.A.; Herrera-Estrella, L.; Rivera-Bustamente, R.F. Complete nucleotide sequence of pepper huasteco virus: Analysis and comparison with bipartite geminiviruses. J. Gen. Virol. 1993, 74, 2225–2231. [Google Scholar] [CrossRef] [PubMed]
  3. Garrido-Ramirez, E.R.; Gilbertson, R.L. First report of tomato mottle geminivirus infecting tomatoes in Yucatan, Mexico. Plant Dis. 2007, 5, 592. [Google Scholar] [CrossRef] [PubMed]
  4. Méndez-Lozano, J.; Leyva-López, N.E.; Mauricio-Castillo, J.A.; Perea-Araujo, L.L.; Ruelas-Ayala, R.D.; Argüello-Astorga, G.R. A Begomovirus isolated from chlorotic and stunted soybean plants in Mexico is a new strain of rhynchosia golden mosaic virus. Plant Dis. 2006, 90, 972. [Google Scholar] [CrossRef] [PubMed]
  5. Melendrez-Bojorquez, N.; Rodríguez-Negrete, E.A.; Magallanes-Tapia, M.A.; Camacho-Beltran, E.; Armenta-Anaya, C.; Leyva-López, N.E.; Mendez-Lozano, J. Pepper huasteco yellow vein virus associated to sweet pepper disease in Sinaloa, Mexico. Plant Dis. 2016, 9, 2338. [Google Scholar] [CrossRef]
  6. Domínguez-Durán, G.; Rodríguez-Negrete, E.A.; Morales-Aguilar, J.J.; Camacho-Beltrán, E.; Romero-Romero, J.L.; Rivera-Acosta, M.A.; Leyva-López, N.E.; Arroyo-Becerra, A.; Méndez-Lozano, J. Molecular and biological characterization of Watermelon chlorotic stunt virus (WmCSV): An Eastern Hemisphere begomovirus introduced in the Western Hemisphere. Crop Prot. 2018, 103, 51–55. [Google Scholar] [CrossRef]
  7. Holguín-Peña, R.J.; Arguello-Astorga, G.R.; Brown, J.K.; Rivera-Bustamante, R.F. A new strain of tomato chino la paz virus associated with a leaf curl disease of tomato in Baja California Sur, Mexico. Plant Dis. 2007, 90, 973. [Google Scholar] [CrossRef] [PubMed]
  8. Moffat, A.S. Geminiviruses emerge as serious crop threat. Science 1999, 286, 1835. [Google Scholar] [CrossRef]
  9. Harkins, G.W.; Martin, D.P.; Duffy, S.; Monjane, A.L.; Shepherd, D.N.; Windram, O.P.; Owor, B.E.; Donaldson, L.; van Antwerpen, T.; Sayed, R.A.; et al. Dating the origins of the maize-adapted strain of maize streak virus, MSV-A. J. Gen. Virol. 2009, 90, 3066–3074. [Google Scholar] [CrossRef]
  10. Patil, B.L.; Fauquet, C.M. Cassava mosaic geminiviruses: Actual knowledge and perspectives. Mol. Plant Pathol. 2009, 10, 685–701. [Google Scholar] [CrossRef]
  11. Briddon, R.W.; Markham, P.G. Cotton leaf curl virus disease. Virus Res. 2000, 71, 151–159. [Google Scholar] [CrossRef]
  12. Accotto, G.P.; Navas-Castillo, J.; Noris, E.; Moriones, E.; Louro, D. Typing of tomato yellow leaf curl viruses in Europe. Eur. J. Plant Pathol. 2000, 106, 179–186. [Google Scholar] [CrossRef]
  13. Brown, J.K. Transmission, host range, and virus-vector relationships of chino del tomate virus, a whitefly-transmitted geminivirus from Sinaloa, Mexico. Plant Dis. 1988, 72, 866. [Google Scholar] [CrossRef]
  14. Garzon-Tiznado, J.A. Inoculation of peppers with infectious clones of a new geminivirus by a biolistic procedure. Phytopathology 1993, 83, 514. [Google Scholar] [CrossRef]
  15. Garzon-Tiznado, J.A.; Acosta-Garcia, G.; Torres-Pacheco, I.; Gonzalez-Chavira, M.; Rivera-Bustamante, R.F.; Maya-Hernandez, V.; Guevara-Gonzalez, R.G. Presence of geminivirus, pepper huasteco virus (PHV), texas pepper virus-variant Tamaulipas (TPV-T), and Chino del tomate virus (CdTV) in the states of Guanajuato, Jalisco and San Luis Potosi, Mexico. Rev. Mex. Fitopatol. 2002, 20, 45–52. [Google Scholar]
  16. Hernández-Zepeda, C.; Idris, A.M.; Carnevali, G.; Brown, J.K.; Moreno-Valenzuela, O.A. Preliminary identification and coat protein gene phylogenetic relationships of begomoviruses associated with native flora and cultivated plants from the Yucatan peninsula of Mexico. Virus Genes 2007, 35, 825–833. [Google Scholar] [CrossRef] [PubMed]
  17. Mauricio-Castillo, J.A.; Argüello-Astorga, G.R.; Bañuelos-Hernández, B.; Ambríz-Granados, S.; Velásquez-Valle, R.; Méndez-Lozano, J. A new strain of chino del tomate virus isolated from soybean plants (Glycine max L.) in Mexico. Rev. Mex. Cienc. Agríc. 2014, 5, 1441–1449. [Google Scholar]
  18. Ascencio-Ibáñez, J.T.; Argüello-Astorga, G.R.; Méndez-Lozano, J.; Rivera-Bustamante, R.F. First report of rhynchosia golden mosaic virus (RhGMV) infecting tobacco in Chiapas, Mexico. Plant Dis. 2002, 86, 692. [Google Scholar] [CrossRef]
  19. Gregorio-Jorge, J.; Argüello-Astorga, G.R.; Frías-Treviño, G.; Bernal-Alcocer, A.; Moreno-Valenzuela, O.; Alpuche-Solís, Á.G.; Bañuelos-Hernández, B.; Hernández-Zepeda, C. Analysis of a new strain of Euphorbia mosaic virus with distinct replication specificity unveils a lineage of begomoviruses with short Rep sequences in the DNA-B intergenic region. Virol. J. 2010, 7, 275. [Google Scholar] [CrossRef]
  20. Padidam, M.; Sawyer, S.; Fauquet, C.M. Possible emergence of new geminiviruses by frequent recombination. Virology 1999, 265, 218–225. [Google Scholar] [CrossRef]
  21. Hernández-Zepeda, C.; Varsani, A.; Brown, J.K. Intergeneric recombination between a new, spinach-infecting curtovirus and a new geminivirus belonging to the genus Becurtovirus: First New World exemplar. Arch. Virol. 2013, 158, 2245–2254. [Google Scholar] [CrossRef] [PubMed]
  22. Lefeuvre, P.; Moriones, E. Recombination as a motor of host switches and virus emergence: Geminiviruses as case studies. Curr. Opin. Virol. 2015, 10, 14–19. [Google Scholar] [CrossRef] [PubMed]
  23. Fiallo-Olivé, E.; Trenado, H.P.; Louro, D.; Navas-Castillo, J. Recurrent speciation of a tomato yellow leaf curl geminivirus in Portugal by recombination. Sci. Rep. 2019, 9, 1–8. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  24. Monci, F.; Sánchez-Campos, S.; Navas-Castillo, J.; Moriones, E. A natural recombinant between the geminiviruses Tomato yellow leaf curl Sardinia virus and Tomato yellow leaf curl virus exhibits a novel pathogenic phenotype and is becoming prevalent in Spanish populations. Virology 2002, 303, 317–326. [Google Scholar] [CrossRef] [PubMed]
  25. Díaz-Pendón, J.A.; Sánchez-Campos, S.; Fortes, I.M.; Moriones, E. Tomato yellow leaf curl sardinia virus, a begomovirus species evolving by mutation and recombination: A challenge for virus control. Viruses 2019, 11, 45. [Google Scholar] [CrossRef] [PubMed]
  26. Aregbesola, O.Z.; Legg, J.P.; Sigsgaard, L.; Lund, O.S.; Rapisarda, C. Potential impact of climate change on whiteflies and implications for the spread of vectored viruses. J. Pest Sci. (2004) 2018, 92, 381–392. [Google Scholar] [CrossRef] [Green Version]
  27. Anonymous. Scientific Opinion on the risks to plant health posed by Bemisia tabaci species complex and viruses it transmits for the EU territory. EFSA J. 2016, 11, 3162. [Google Scholar]
  28. Canto, T.; Aranda, M.A.; Fereres, A. Climate change effects on physiology and population processes of hosts and vectors that influence the spread of hemipteran-borne plant viruses. Glob. Chang. Biol. 2009, 15, 1884–1894. [Google Scholar] [CrossRef] [Green Version]
  29. Duffy, S.; Holmes, E.C. Phylogenetic evidence for rapid rates of molecular evolution in the single-stranded DNA begomovirus tomato yellow leaf curl virus. J. Virol. 2007, 82, 957–965. [Google Scholar] [CrossRef]
  30. Lefeuvre, P.; Martin, D.P.; Harkins, G.; Lemey, P.; Gray, A.J.; Meredith, S.; Lakay, F.; Monjane, A.; Lett, J.M.; Varsani, A.; et al. The spread of tomato yellow leaf curl virus from the Middle East to the world. PLoS Pathog. 2010, 6, e1001164. [Google Scholar] [CrossRef]
  31. Monjane, A.L.; Harkins, G.W.; Martin, D.P.; Lemey, P.; Lefeuvre, P.; Shepherd, D.N.; Oluwafemi, S.; Simuyandi, M.; Zinga, I.; Komba, E.K.; et al. Reconstructing the history of maize streak virus strain a dispersal to reveal diversification hot spots and its origin in Southern Africa. J. Virol. 2011, 85, 9623–9636. [Google Scholar] [CrossRef] [PubMed]
  32. Bernardo, P.; Golden, M.; Akram, M.; Naimuddin; Nadarajan, N.; Fernandez, E.; Granier, M.; Rebelo, A.G.; Peterschmitt, M.; Martin, D.P.; et al. Identification and characterisation of a highly divergent geminivirus: Evolutionary and taxonomic implications. Virus Res. 2013, 177, 35–45. [Google Scholar] [CrossRef] [PubMed]
  33. Varsani, A.; Roumagnac, P.; Fuchs, M.; Navas-Castillo, J.; Moriones, E.; Idris, A.; Briddon, R.W.; Rivera-Bustamante, R.; Murilo Zerbini, F.; Martin, D.P. Capulavirus and Grablovirus: Two new genera in the family Geminiviridae. Arch. Virol. 2017, 162, 1819–1831. [Google Scholar] [CrossRef] [PubMed]
  34. Rybicki, E.P. A phylogenetic and evolutionary justification for three genera of Geminiviridae. Arch. Virol. 1994, 139, 49–77. [Google Scholar] [CrossRef] [PubMed]
  35. Barba, M.; Czosnek, H.; Hadidi, A. Historical perspective, development and applications of next-generation sequencing in plant virology. Viruses 2014, 6, 106–136. [Google Scholar] [CrossRef] [PubMed]
  36. Massart, S.; Olmos, A.; Jijakli, H.; Candresse, T. Current impact and future directions of high-throughput sequencing in plant virus diagnostics. Virus Res. 2014, 188, 90–96. [Google Scholar] [CrossRef] [PubMed]
  37. Wu, Q.; Ding, S.-W.; Zhang, Y.; Zhu, S. Identification of viruses and viroids by next-generation sequencing and homology-dependent and homology-independent algorithms. Annu. Rev. Phytopathol. 2015, 53, 425–444. [Google Scholar] [CrossRef] [PubMed]
  38. Pooggin, M.M. Small RNA-omics for plant virus identification, virome reconstruction, and antiviral defense characterization. Front. Microbiol. 2018, 9, 1–20. [Google Scholar] [CrossRef] [PubMed]
  39. Pereira, A.J.; Alfenas-Zerbini, P.; Cascardo, R.S.; Andrade, E.C.; Murilo Zerbini, F. Analysis of the full-length genome sequence of papaya lethal yellowing virus (PLYV), determined by deep sequencing, confirms its classification in the genus Sobemovirus. Arch. Virol. 2012, 10, 2009–2011. [Google Scholar] [CrossRef] [PubMed]
  40. Rampelli, S.; Soverini, M.; Turroni, S.; Quercia, S.; Biagi, E.; Brigidi, P.; Candela, M. ViromeScan: A new tool for metagenomic viral community profiling. BMC Genomics 2016, 17, 165. [Google Scholar] [CrossRef] [PubMed]
  41. Bernardo, P.; Charles-Dominique, T.; Barakat, M.; Ortet, P.; Fernandez, E.; Filloux, D.; Hartnady, P.; Rebelo, T.A.; Cousins, S.R.; Mesleard, F.; et al. Geometagenomics illuminates the impact of agriculture on the distribution and prevalence of plant viruses at the ecosystem scale. ISME J. 2017, 12, 173–184. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  42. Llorente-Bousquets, J.; Ocegueda, S. Estado del conocimiento de la biota. in Capital natural de México: Conocimiento actual de la biodiversidad. Conabio Méx. 2008, 1, 283–322. [Google Scholar]
  43. Mauricio-Castillo, J.A.; Arguello-Astorga, G.R.; Ambriz-Granados, S.; Alpuche-Solís, A.G. First Report of Tomato golden mottle virus on Lycopersicon esculentum and Solanum rostratum in Mexico. Plant Dis. 2007, 11, 11–12. [Google Scholar] [CrossRef] [PubMed]
  44. Doyle, J.J.; Doyle, J.L. A rapid DNA isolation procedure for small quantities of fresh leaf tissue. Phytochem. Bull. 1987, 19, 11–15. [Google Scholar]
  45. Bolger, A.M.; Lohse, M.; Usadel, B. Trimmomatic: A flexible trimmer for Illumina sequence data. Bioinformatics 2014, 30, 2114–2120. [Google Scholar] [CrossRef] [PubMed]
  46. Kulikov, A.S.; Prjibelski, A.D.; Tesler, G.; Vyahhi, N.; Sirotkin, A.V.; Pham, S.; Dvorkin, M.; Pevzner, P.A.; Bankevich, A.; Nikolenko, S.I.; et al. SPAdes: A new genome assembly algorithm and its applications to single-cell sequencing. J. Comput. Biol. 2012, 5, 455–477. [Google Scholar]
  47. Altschul, S.F.; Gish, W.; Miller, W.; Myers, E.W.; Lipman, D.J. Basic Local Aligment Search Tool. J. Mol. Biol. 1990, 215, 403–410. [Google Scholar] [CrossRef]
  48. Inoue-Nagata, A.K.; Albuquerque, L.C.; Rocha, W.B.; Nagata, T. A simple method for cloning the complete begomovirus genome using the bacteriophage 29 DNA polymerase. J. Virol. Methods 2004, 116, 209–211. [Google Scholar] [CrossRef]
  49. Hellens, R.P.; Anne Edwards, E.; Leyland, N.R.; Bean, S.; Mullineaux, P.M. pGreen: A versatile and flexible binary Ti vector for agrobacterium-mediated plant transformation. Plant Mol. Biol. 2000, 42, 819–832. [Google Scholar] [CrossRef]
  50. Kumar, S.; Stecher, G.; Tamura, K. MEGA7: Molecular evolutionary genetics analysis version 7.0 for bigger datasets. Mol. Biol. Evol. 2016, 7, 1870–1874. [Google Scholar] [CrossRef]
  51. Livermore, M.; Fischer, G.; Rosenzweig, C.; Parry, M.; Iglesias, A. Effects of climate change on global food production under SRES emissions and socio-economic scenarios. Glob. Environ. Chang. 2004, 14, 53–67. [Google Scholar]
  52. Velásquez, A.C.; Castroverde, C.D.M.; He, S.Y. Plant–pathogen warfare under changing climate conditions. Curr. Biol. 2018, 28, 619–634. [Google Scholar] [CrossRef] [PubMed]
  53. Luck, J.; Spackman, M.; Freeman, A.; TreBicki, P.; Griffiths, W.; Finlay, K.; Chakraborty, S. Climate change and diseases of food crops. Plant Pathol. 2011, 60, 113–121. [Google Scholar] [CrossRef]
  54. Roossinck, M.J. Plant Virus Metagenomics: Biodiversity and Ecology. Annu. Rev. Genet. 2012, 46, 359–369. [Google Scholar] [CrossRef] [PubMed]
  55. Prajapat, R.; Marwal, A.; Gaur, R.K. Begomovirus associated with alternative host weeds: A critical appraisal. Arch. Phytopathol. Plant Prot. 2014, 2, 157–179. [Google Scholar] [CrossRef]
  56. Gutiérrez-García, J.A.; Hernández-Vizcarra, J.A.; Villaseñor, J.L.; Vega-Aviña, R.; Aguiar-Hernández, H.; Vega-López, I.F. Endemismo regional presente en la flora del municipio de Culiacán, Sinaloa, México. Acta Bot. Mex. 2017, 53, 1–15. [Google Scholar]
  57. Torres-Pacheco, I.; Garzón Tiznado, J.A.; Brown, J.K.; Becerra-Flora, A.; Rivera-Bustamante, R.F. Detection and distribution of geminiviruses in Mexico and the Southern United States. Phytopathology 1996, 11, 1186–1192. [Google Scholar] [CrossRef]
  58. Mauricio-Castillo, J.A.; Argüello-Astorga, G.R.; Alpuche-Solís, A.G.; Monreal-Vargas, C.T.; Díaz-Gómez, O.; De La Torre-Almaraz, R. First report of tomato severe leaf curl virus in México. Plant Dis. 2006, 90, 1116. [Google Scholar] [CrossRef] [PubMed]
  59. Idris, A.M.; Carnevali, G.; Brown, J.K.; Hernández-Zepeda, C.; Moreno-Valenzuela, O.A.; Argüello-Astorga, G.; Rivera-Bustamante, R.F. Characterization of Rhynchosia yellow mosaic Yucatan virus, a new recombinant begomovirus associated with two fabaceous weeds in Yucatan, Mexico. Arch. Virol. 2010, 155, 1571–1579. [Google Scholar]
  60. Reveles Torres, L.R.; Velásquez Valle, R.; Mauricio Castillo, J.A.; Salas Muñoz, S. Detection of mixed infections caused by begomovirus and curtovirus in chili pepper for drying plants in San Luis Potosi, Mexico. Rev. Mex. Fitopatol. 2012, 30, 155–160. [Google Scholar]
  61. Gasperin-Bulbarela, J.; Hernández-Martínez, R.; Licea-Navarro, A.F.; Pino-Villar, C.; Carrillo-Tripp, J. First report of grapevine red blotch virus in Mexico. Plant Dis. 2018, 2, 381. [Google Scholar] [CrossRef]
  62. Rosario, K.; Seah, Y.M.; Marr, C.; Varsani, A.; Kraberger, S.; Stainton, D.; Moriones, E.; Polston, J.E.; Duffy, S.; Breitbart, M. Vector-enabled metagenomic (VEM) surveys using whiteflies (Aleyrodidae) reveal novel begomovirus species in the new and old worlds. Viruses 2015, 7, 5553–5570. [Google Scholar] [CrossRef]
  63. Rodelo-Urrego, M.; García-Arenal, F.; Pagá, I. The effect of ecosystem biodiversity on virus genetic diversity depends on virus species: A study of chiltepin-infecting begomoviruses in Mexico. Virus Evol. 2015, 1, vev004. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  64. Ascencio-Ibáñez, J.T.; Diaz-Plaza, R.; Méndez-Lozano, J.; Monsalve-Fonnegra, Z.I.; Argüello-Astorga, G.R.; Rivera-Bustamante, R.F. First report of tomato yellow leaf curl geminivirus in Yucatán, México. Plant Dis. 1999, 83, 1178. [Google Scholar] [CrossRef] [PubMed]
  65. Morales-Aguilar, J.J.; Rodríguez-Negrete, E.A.; Camacho-Beltrán, E.; López-Luque, C.A.; Leyva-López, N.E.; Jiménez-Díaz, F.; Voloudakis, A.; Santos-Cervantes, M.E.; Méndez-Lozano, J. Identification of Tomato yellow leaf curl virus, Pepper huasteco yellow vein virus and Pepper golden mosaic virus associated with pepper diseases in northern Mexico. Can. J. Plant Pathol. 2019. [Google Scholar] [CrossRef]
  66. Moreno-Félix, M.L.; Rodríguez-Negrete, E.A.; Camacho-Beltrán, E.; Leyva-López, N.E.; Meléndrez-Bojórquez, N.; Méndez-Lozano, J. A new isolate of Pepper huasteco yellow vein virus (PHYVV) breaks geminivirus tolerance in tomato (Solanum lycopersicum) commercial lines. Acta Hortic. 2018, 35–44. [Google Scholar] [CrossRef]
  67. Abudy, A.; Sufrin-Ringwald, T.; Dayan-Glick, C.; Guenoune-Gelbart, D.; Livneh, O.; Zaccai, M.; Lapidot, M. Watermelon chlorotic stunt and Squash leaf curl begomoviruses—New threats to cucurbit crops in the Middle East. Isr. J. Plant Sci. 2010, 58, 33–42. [Google Scholar] [CrossRef]
  68. Hernández-Zepeda, C.; Idris, A.M.; Carnevali, G.; Brown, J.K.; Moreno-Valenzuela, O.A. Molecular characterization and phylogenetic relationships of two new bipartite begomovirus infecting malvaceous plants in Yucatan, Mexico. Virus Genes 2007, 35, 369–377. [Google Scholar] [CrossRef] [PubMed]
  69. Fernandes, F.R.; Albuquerque, L.C.; de Oliveira, C.L.; Cruz, A.R.R.; da Rocha, W.B.; Pereira, T.G.; Naito, F.Y.B.; De Dias, N.M.; Nagata, T.; Faria, J.C.; et al. Molecular and biological characterization of a new Brazilian begomovirus, euphorbia yellow mosaic virus (EuYMV), infecting Euphorbia heterophylla plants. Arch. Virol. 2011, 156, 2063–2069. [Google Scholar] [CrossRef] [Green Version]
  70. Cantú-Iris, M.; Pastor-Palacios, G.; Mauricio-Castillo, J.A.; Bañuelos-Hernández, B.; Avalos-Calleros, J.A.; Juárez-Reyes, A.; Rivera-Bustamante, R.; Argüello-Astorga, G.R. Analysis of a new begomovirus unveils a composite element conserved in the CP gene promoters of several Geminiviridae genera: Clues to comprehend the complex regulation of late genes. PLoS ONE 2019, 1, e0210485. [Google Scholar] [CrossRef]
  71. Martínez-Ayala, A.; Sánchez-Campos, S.; Cáceres, F.; Aragõn-Caballero, L.; Navas-Castillo, J.; Moriones, E. Characterisation and genetic diversity of pepper leafroll virus, a new bipartite begomovirus infecting pepper, bean and tomato in Peru. Ann. Appl. Biol. 2014, 164, 62–72. [Google Scholar] [CrossRef]
  72. Andrade, E.C.; Manhani, G.G.; Alfenas, P.F.; Calegario, R.F.; Fontes, E.P.B.; Zerbini, F.M. Tomato yellow spot virus, a tomato-infecting begomovirus from Brazil with a closer relationship to viruses from Sida sp., forms pseudorecombinants with begomoviruses from tomato but not from Sida. J. Gen. Virol. 2006, 87, 3687–3696. [Google Scholar] [CrossRef] [PubMed]
  73. Shih, S.L.; Tsai, W.S.; Green, S.K.; Khalid, S.; Ahmad, I.; Rezaian, M.A.; Smith, J. Molecular characterization of tomato and chili leaf curl begomoviruses from Pakistan. Plant Dis. 2007, 2, 200. [Google Scholar] [CrossRef] [PubMed]
  74. Malmstrom, C.M.; Melcher, U.; Bosque-Pérez, N.A. The expanding field of plant virus ecology: Historical foundations, knowledge gaps, and research directions. Virus Res. 2011, 159, 84–94. [Google Scholar] [CrossRef] [PubMed]
  75. Roossinck, M.J. Environmental viruses from biodiversity to ecology. Curr. Opin. Virol. 2011, 1, 50–51. [Google Scholar] [CrossRef] [PubMed]
  76. Stobbe, A.H.; Roossinck, M.J. Plant virus metagenomics: What we know and why we need to know more. Front. Plant Sci. 2014, 5, 1–4. [Google Scholar] [CrossRef]
  77. Aguiar, R.W.S.; Alves, G.B.; Queiroz, A.P.; Nascimento, I.R.; Lima, M.F. Evaluation of Weeds as Virus Reservoirs in Watermelon Crops. Planta Daninha 2017, 2016, 1–13. [Google Scholar] [CrossRef]
  78. Basak, J. Tomato yellow leaf curl virus: A serious threat to tomato plants world wide. J. Plant Pathol. Microbiol. 2016, 7, 346. [Google Scholar] [CrossRef]
  79. Paz-Carrasco, L.C.; Castillo-Urquiza, G.P.; Lima, A.T.M.; Xavier, C.A.D.; Vivas-Vivas, L.M.; Mizubuti, E.S.G.; Zerbini, F.M. Begomovirus diversity in tomato crops and weeds in Ecuador and the detection of a recombinant isolate of rhynchosia golden mosaic Yucatan virus infecting tomato. Arch. Virol. 2014, 159, 2127–2132. [Google Scholar] [CrossRef]
  80. Perry, K.L.; McLane, H.; Thompson, J.R.; Fuchs, M. A novel grablovirus from non-cultivated grapevine (Vitis sp.) in North America. Arch. Virol. 2018, 163, 259–262. [Google Scholar] [CrossRef]
  81. Bekele, B.; Kumari, S.; Ahmed, S.; Fininsa, C.; Yusuf, A.; Abraham, A. Non-cultivated grass hosts of yellow dwarf viruses in Ethiopia and their epidemiological consequences on cultivated cereals. J. Phytopathol. 2018, 166, 103–115. [Google Scholar] [CrossRef]
  82. Strydom, E.; Pietersen, G. Alternative hosts and seed transmissibility of soybean blotchy mosaic virus. Eur. J. Plant Pathol. 2018, 151, 263–268. [Google Scholar] [CrossRef]
  83. Mansoor, S.; Haider, M.; Tahir, M.; Briddon, R.; Amin, I. Ageratum enation virus—A begomovirus of weeds with the potential to infect crops. Viruses 2015, 7, 647–665. [Google Scholar]
  84. Brown, J.K.; Zerbini, F.M.; Navas-Castillo, J.; Moriones, E.; Ramos-Sobrinho, R.; Silva, J.C.F.; Fiallo-Olivé, E.; Briddon, R.W.; Hernández-Zepeda, C.; Idris, A.; et al. Revision of begomovirus taxonomy based on pairwise sequence comparisons. Arch. Virol. 2015, 160, 1593–1619. [Google Scholar] [CrossRef] [PubMed]
  85. Yuan, W.; Ruan, M.Y.; Yang, Y.J.; Zhou, G.Z.; Yao, Z.P.; Wang, R.Q.; Wan, H.J.; Ye, Q.J.; Li, Z.M. Assessment of the genetic diversity of tomato yellow leaf curl virus. Genet. Mol. Res. 2015, 1, 529–537. [Google Scholar]
  86. Kil, E.-J.; Kim, S.; Lee, Y.-J.; Byun, H.-S.; Park, J.; Seo, H.; Kim, C.-S.; Shim, J.-K.; Lee, J.-H.; Kim, J.-K.; et al. Tomato yellow leaf curl virus (TYLCV-IL): A seed-transmissible geminivirus in tomatoes. Sci. Rep. 2016, 6, 19013. [Google Scholar] [CrossRef] [PubMed]
  87. Moriones, E.; Navas-Castillo, J. Review. Rapid evolution of the population of begomoviruses associated with the tomato yellow leaf curl disease after, invasion of a new ecological niche. Span. J. Agric. Res. 2008, 6, 147–159. [Google Scholar] [CrossRef]
  88. Salati, R.; Nahkla, M.K.; Rojas, M.R.; Guzman, P.; Jaquez, J.; Maxwell, D.P.; Gilbertson, R.L. Tomato yellow leaf curl virus in the Dominican Republic: Characterization of an infectious clone, virus monitoring in whiteflies, and identification of reservoir hosts. Phytopathology 2002, 92, 487–496. [Google Scholar] [CrossRef]
  89. Qinones, M.; Fonseca, D.; Martinez, Y.; Accotto, G.P. First report of tomato yellow leaf curl virus infecting pepper plants in Cuba. Plant Dis. 2002, 86, 73. [Google Scholar] [CrossRef]
  90. Cardenas-Conejo, Y.; Argüello-Astorga, G.R.; Poghosyan, A.; Hernandez-Gonzalez, J.; Lebsky, V.; Holguin-Peña, J.; Medina-Hernandez, D.; Vega-Peña, S. First report of tomato yellow leaf curl virus co-infecting pepper with tomato chino La Paz virus in Baja California Sur, Mexico. Plant Dis. 2010, 94, 1266. [Google Scholar] [CrossRef]
  91. Brown, J.K.; Idris, A.M. Introduction of the exotic monopartite tomato yellow leaf curl virus into west coast Mexico. Plant Dis. 2006, 90, 1360. [Google Scholar] [CrossRef] [PubMed]
  92. Gamez-Jimenez, C.; Romero-Romero, J.L.; Santos-Cervantes, M.E.; Leyva-Lopez, N.E.; Mendez-Lozano, J. Tomatillo (Physalis ixocarpa) as a natural new host for tomato yellow leaf curl virus in Sinaloa, Mexico. Plant Dis. 2009, 93, 545. [Google Scholar] [CrossRef] [PubMed]
  93. Barboza, N.; Blanco-Meneses, M.; Hallwass, M.; Moriones, E.; Inoue-Nagata, A.K. First report of tomato yellow leaf curl virus in tomato in Costa Rica. Plant Dis. 2013, 98, 699. [Google Scholar] [CrossRef] [PubMed]
  94. Dong, J.H.; Luo, Y.Q.; Ding, M.; Zhang, Z.K.; Yang, C.K. First report of tomato yellow leaf curl virus infecting vommon bean in China. Plant Pathol. 2007, 56, 342. [Google Scholar] [CrossRef]
  95. Bedford, I.D.; Kelly, A.; Banks, G.K.; Briddon, R.W.; Cenis, J.L.; Markham, P.G. Solanum nigrum: An indigenous weed reservoir for a tomato yellow leaf curl geminivirus in southern Spain. Eur. J. Plant Pathol. 1998, 104, 221–222. [Google Scholar] [CrossRef]
  96. García-Andrés, S.; Monci, F.; Navas-Castillo, J.; Moriones, E. Begomovirus genetic diversity in the native plant reservoir Solanum nigrum: Evidence for the presence of a new virus species of recombinant nature. Virology 2006, 350, 433–442. [Google Scholar] [CrossRef] [PubMed]
  97. Kil, E.J.; Park, J.; Lee, H.; Kim, J.; Choi, H.S.; Lee, K.Y.; Kim, C.S.; Lee, S. Lamium amplexicaule (Lamiaceae): A weed reservoir for tomato yellow leaf curl virus (TYLCV) in Korea. Arch. Virol. 2014, 159, 1305–1311. [Google Scholar] [CrossRef] [PubMed]
  98. Kashina, B.D.; Mabagala, R.B.; Mpunami, A.A. Reservoir weed hosts of tomato yellow leaf curl begomovirus from Tanzania. Arch. Phytopathol. Plant Prot. 2003, 35, 269–278. [Google Scholar] [CrossRef]
  99. Jordá, C.; Font, I.; Martínez, P.; Juarez, M.; Ortega, A.; Lacasa, A. Current status and new natural hosts of tomato yellow leaf curl virus (TYLCV) in Spain. Plant Dis. 2007, 85, 445. [Google Scholar] [CrossRef]
  100. Cervantes-Díaz, L.; Zavaleta-Mejía, E.; Rojas-Martínez, R.I.; Alanís-Martínez, I.D.; Ochoa-Martínez, L.; Valadez-Moctezuma, E.; Grimaldo-Juárez, O. Detección de geminivirus asociados a la alstroemeria (Alstroemeria L.) en villa guerrero, estado de México. Interciencia 2009, 12, 903–908. [Google Scholar]
  101. Potter, J.L.; Roca de Doyle, M.M.; Nakhla, M.K.; Maxwell, D.P. First report and characterization of rhynchosia golden mosaic virus in Honduras. Plant Dis. 2007, 84, 1045. [Google Scholar] [CrossRef] [PubMed]
  102. Silva, S.J.C.; Castillo-Urquiza, G.P.; Hora-Júnior, B.T.; Assunção, I.P.; Lima, G.S.A.; Pio-Ribeiro, G.; Mizubuti, E.S.G.; Zerbini, F.M. Species diversity, phylogeny and genetic variability of begomovirus populations infecting leguminous weeds in northeastern Brazil. Plant Pathol. 2012, 61, 457–467. [Google Scholar] [CrossRef]
  103. Rosario, K.; Marr, C.; Varsani, A.; Kraberger, S.; Stainton, D.; Moriones, E.; Polston, J.E.; Breitbart, M. Begomovirus-associated satellite DNA diversity captured through vector-enabled metagenomic (VEM) surveys using whiteflies (Aleyrodidae). Viruses 2016, 8, 36. [Google Scholar] [CrossRef] [PubMed]
  104. Fondong, V.N.; Pita, J.S.; Rey, M.E.C.; De Kochko, A.; Beachy, R.N.; Fauquet, C.M. Evidence of synergism between African cassava mosaic virus and a new double-recombinant geminivirus infecting cassava in Cameroon. J. Gen. Virol. 2000, 81, 287–297. [Google Scholar] [CrossRef] [PubMed]
  105. Pita, J.S.; Fondong, V.N.; Sangaré, A.; Kokora, R.N.N.; Fauquet, C.M. Genomic and biological diversity of the African cassava geminiviruses. Euphytica 2001, 120, 115–125. [Google Scholar] [CrossRef]
  106. Ribeiro, S.G.; Ambrozevícius, L.P.; Ávila, A.C.; Bezerra, I.C.; Calegario, R.F.; Fernandes, J.J.; Lima, M.F.; De Mello, R.N.; Rocha, H.; Zerbini, F.M. Distribution and genetic diversity of tomato-infecting begomoviruses in Brazil. Arch. Virol. 2003, 148, 281–295. [Google Scholar] [CrossRef] [PubMed]
  107. Pita, J.S.; Fondong, V.N.; Sangaré, A.; Otim-Nape, G.W.; Ogwal, S.; Fauquet, C.M. Recombination, pseudorecombination and synergism of geminiviruses are determinant keys to the epidemic of severe cassava mosaic disease in Uganda. J. Gen. Virol. 2001, 82, 655–665. [Google Scholar] [CrossRef] [PubMed]
  108. Davino, S.; Napoli, C.; Dellacroce, C.; Miozzi, L.; Noris, E.; Davino, M.; Accotto, G.P. Two new natural begomovirus recombinants associated with the tomato yellow leaf curl disease co-exist with parental viruses in tomato epidemics in Italy. Virus Res. 2009, 143, 15–23. [Google Scholar] [CrossRef]
  109. Roossinck, M.J.; García-Arenal, F. Ecosystem simplification, biodiversity loss and plant virus emergence. Curr. Opin. Virol. 2015, 10, 56–62. [Google Scholar] [CrossRef]
  110. Mohammed, H.S.; El Siddig, M.A.; El Hussein, A.A.; Navas-Castillo, J.; Fiallo-Olivé, E. Complete genome sequence of datura leaf curl virus, a novel begomovirus infecting Datura innoxia in Sudan, related to begomoviruses causing tomato yellow leaf curl disease. Arch. Virol. 2018, 163, 273–275. [Google Scholar] [CrossRef]
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Figure 1. Northern-Pacific Mexico sampling areas and high-throughput sequencing (HTS) library construction. (a) Sampling areas were divided in five regions located in different biogeographic zones (according to the CONABIO classification): Baja California (BC), Sonora (SO), Sinaloa (SI), Colima-Nayarit (CN), and Coahulia-Durango (CD). BC-SQ: Baja California, San Quintin; BC-EN: Baja California, Ensenada; BC-ME: Baja California, Mexicali; SO-OB: Sonora, Obregon; SO-NA: Sonora, Navojoa; SO-HU: Sonora, Huatabampo; SO-RC: Sonora, Rio Colorado; SI-GV: Sinaloa, Guasave; SI-SL: Sinaloa, Sinaloa de Leyva; SI-MO: Sinaloa, Mocorito; SI-PC: Sinaloa, Playa Ceuta; SI-CO: Sinaloa, Concordia; SI-AC: Sinaloa, Agua Caliente; SI-RO: Sinaloa, El Rosario; CN-SO: Colima-Nayarit, Santa Maria del Oro; CN-TE: Colima-Nayarit, Tecoman; CD-TL: Coahuila-Durango, Tlahualilo; CD-LG: Coahuila-Durango, La Goma; CD-TO: Coahuila-Durango, Torreon; CD-PO: Coahuila-Durango, Poanas. Sampling areas are indicated in the map by colored squares. (b) Diagram of sample processing to obtain HTS libraries. For each sampling area, total DNA from Begomovirus PCR-positive plants belonging to the same species was pooled in equimolar concentrations. The resulting DNA mix was used as a template for rolling circle amplification (RCA)-mediated viral circular molecule enrichment. Finally, all resulting RCA products were pooled in equimolar concentrations and used for HTS library construction.
Figure 1. Northern-Pacific Mexico sampling areas and high-throughput sequencing (HTS) library construction. (a) Sampling areas were divided in five regions located in different biogeographic zones (according to the CONABIO classification): Baja California (BC), Sonora (SO), Sinaloa (SI), Colima-Nayarit (CN), and Coahulia-Durango (CD). BC-SQ: Baja California, San Quintin; BC-EN: Baja California, Ensenada; BC-ME: Baja California, Mexicali; SO-OB: Sonora, Obregon; SO-NA: Sonora, Navojoa; SO-HU: Sonora, Huatabampo; SO-RC: Sonora, Rio Colorado; SI-GV: Sinaloa, Guasave; SI-SL: Sinaloa, Sinaloa de Leyva; SI-MO: Sinaloa, Mocorito; SI-PC: Sinaloa, Playa Ceuta; SI-CO: Sinaloa, Concordia; SI-AC: Sinaloa, Agua Caliente; SI-RO: Sinaloa, El Rosario; CN-SO: Colima-Nayarit, Santa Maria del Oro; CN-TE: Colima-Nayarit, Tecoman; CD-TL: Coahuila-Durango, Tlahualilo; CD-LG: Coahuila-Durango, La Goma; CD-TO: Coahuila-Durango, Torreon; CD-PO: Coahuila-Durango, Poanas. Sampling areas are indicated in the map by colored squares. (b) Diagram of sample processing to obtain HTS libraries. For each sampling area, total DNA from Begomovirus PCR-positive plants belonging to the same species was pooled in equimolar concentrations. The resulting DNA mix was used as a template for rolling circle amplification (RCA)-mediated viral circular molecule enrichment. Finally, all resulting RCA products were pooled in equimolar concentrations and used for HTS library construction.
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Figure 2. Phylogenetic trees based on multiple sequence alignment of complete monopartite (A) and bipartite begomovirus DNA-A (B), and DNA-B (C) with selected isolates obtained from NCBI. Trees were constructed by the maximum likelihood method with 1000 bootstrap replicates using MEGA7. Virus acronyms: Bean golden yellow mosaic virus (BGYMV), Malvastrum bright yellow mosaic virus (MaBYMV), Okra yellow mosaic virus (OYMV), Rhynchosia golden mosaic Sinaloa virus (RhGMSV), Rhynchosia golden mosaic virus (RhGMV), Rhynchosia golden mosaic Yucatan virus (RhGMYuV), Sida golden mosaic Honduras virus (SiGMHoV), Sida mosaic Sinaloa virus (SiMSiV), Sida yellow mottle virus (SiYMV), Sida yellow mosaic Yucatan virus (SiYMYuV), Sida yellow vein virus (SiYVV), Soybean chlorotic spot virus (SoCSV). Viral genomes Accession numbers are shown. Countries codes are as follow: Brazil (BR), Cuba (CU), Guatemala (GU), Ecuador (EC), Honduras (HN), Israel (IR), Mexico (MX), Puerto Rico (PR) and United States of America (US). As an out-group, Beet curly top virus sequence (BCTV) was used. Malvaceae and Fabaceae infecting virus are highlighted in blue and green, respectively.
Figure 2. Phylogenetic trees based on multiple sequence alignment of complete monopartite (A) and bipartite begomovirus DNA-A (B), and DNA-B (C) with selected isolates obtained from NCBI. Trees were constructed by the maximum likelihood method with 1000 bootstrap replicates using MEGA7. Virus acronyms: Bean golden yellow mosaic virus (BGYMV), Malvastrum bright yellow mosaic virus (MaBYMV), Okra yellow mosaic virus (OYMV), Rhynchosia golden mosaic Sinaloa virus (RhGMSV), Rhynchosia golden mosaic virus (RhGMV), Rhynchosia golden mosaic Yucatan virus (RhGMYuV), Sida golden mosaic Honduras virus (SiGMHoV), Sida mosaic Sinaloa virus (SiMSiV), Sida yellow mottle virus (SiYMV), Sida yellow mosaic Yucatan virus (SiYMYuV), Sida yellow vein virus (SiYVV), Soybean chlorotic spot virus (SoCSV). Viral genomes Accession numbers are shown. Countries codes are as follow: Brazil (BR), Cuba (CU), Guatemala (GU), Ecuador (EC), Honduras (HN), Israel (IR), Mexico (MX), Puerto Rico (PR) and United States of America (US). As an out-group, Beet curly top virus sequence (BCTV) was used. Malvaceae and Fabaceae infecting virus are highlighted in blue and green, respectively.
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Figure 3. Venn diagram of predominant begomovirus detected. A total of 126 individual species were examined for the presence of predominant begomovirus in the five sampling regions. The number of begomovirus-positive plants (97) showing single, double, triple or quadruple infections is reported. The values are also reported in percentage in parenthesis. Virus acronyms: Pepper huasteco yellow vein virus (PHYVV), Rhynchosia golden mosaic virus, (RhGMV), Rhynchosia golden mosaic Sinaloa virus (RhGMSV), Sida mosaic Sinaloa virus (SiMSiV), Tomato yellow leaf curl virus (TYLCV).
Figure 3. Venn diagram of predominant begomovirus detected. A total of 126 individual species were examined for the presence of predominant begomovirus in the five sampling regions. The number of begomovirus-positive plants (97) showing single, double, triple or quadruple infections is reported. The values are also reported in percentage in parenthesis. Virus acronyms: Pepper huasteco yellow vein virus (PHYVV), Rhynchosia golden mosaic virus, (RhGMV), Rhynchosia golden mosaic Sinaloa virus (RhGMSV), Sida mosaic Sinaloa virus (SiMSiV), Tomato yellow leaf curl virus (TYLCV).
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Table
Table 1. Begomovirus detection in plants collected in the agro-ecological interface from Northern-Pacific regions of Mexico. Total DNA from each individual specimen was used as a template for PCR-mediated begomovirus detection.
Table 1. Begomovirus detection in plants collected in the agro-ecological interface from Northern-Pacific regions of Mexico. Total DNA from each individual specimen was used as a template for PCR-mediated begomovirus detection.
Plant Family 1Sampling Region 2
Begomovirus PCR-Positive/Total Plants Collected
BCSOSICNCD
Amaranthaceae 4/51/41/45/6
Apiaceae 1/1
Asteraceae4/108/168/223/40/15
Boraginaceae0/14/5
Brassicaceae1/20/21/1
Caesalpiniaceae 1/1
Capparaceae 1/2
Chenopodiaceae4/46/10
Convolvulaceae0/22/23/41/4
Cucurbitaceae0/1 2/61/3
Euphorbiaceae 2/35/150/2
Fabaceae 1/249/720/1
Hydrophyllaceae 1/1 0/1
Malvaceae5/614/1821/339/139/9
Menispermaceae 1/1
Nyctaginaceae 2/24/51/1
Onagraceae 1/2
Papaveraceae 1/1
Pedaliaceae 1/1
Polygonaceae1/23/5
Portulacaceae 3/31/2
Primulaceae1/1
Rhamnaceae 1/11/1
Rubiaceae 2/2
Sapindaceae 1/1
Solanaceae1/211/1418/220/214/14
Sterculiaceae 1/2
Verbenaceae 1/22/2
Vitaceae 1/10/1
Total positives17601222429
1 Plant families PCR-negative for begomovirus detection: Apocynaceae, Asclepiadiaceae, Bignoniaceae, Commelinaceae and Malpiguiaceae. 2 Plant specimens belonging to 34 families were collected from five regions: Baja California (BC), Sonora (SO), Sinaloa (SI), Colima-Nayarit (CN), Coahuila-Durango (CD).
Table
Table 2. Next generation sequence (NGS) data summary of reads and contigs mapping to Geminivirus genomes.
Table 2. Next generation sequence (NGS) data summary of reads and contigs mapping to Geminivirus genomes.
Library NameTotal ReadsTotal Geminivirus-Related ReadsNumber of Geminivirus-Related ContigsSmallest/Largest Geminivirus-Related Contig 1
Baja California16,056,86661569278/2437
Sonora30,440,80223,54619578/2293
Sinaloa215,007,4564,685,42315,46578/2723
Colima-Nayarit33,159,6202,475,219836878/2775
Coahuila-Durango70,782,034349,76316978/2858
Total365,446,7787,540,10724,28978/2858
1 bp: Base pairs.
Table
Table 3. Begomovirus signatures obtained by de novo assembly from the metagenomics study in plants collected in the agro-ecological interface from five regions in Northern-Pacific Mexico. Geminivirus-related reads for each NGS library were used for de novo assembly and generation of signatures.
Table 3. Begomovirus signatures obtained by de novo assembly from the metagenomics study in plants collected in the agro-ecological interface from five regions in Northern-Pacific Mexico. Geminivirus-related reads for each NGS library were used for de novo assembly and generation of signatures.
Host AdaptedVirus Acronym 2Plant Family of First DetectionGeminivirus-Signatures of DNA-A/DNA-B 1 per Region
Baja CaliforniaSonoraSinaloaColima-NayaritCoahuila-Durango
CropsPHYVVSolanaceaeND 398.5/96.5
LN848858.1/LN848912.1
1462/2124		99.6/100
LN848873.1/KP890828.1
251/594		100/99.6
X70418.1/X70419.1
(583/1826)		96.9/98.5
LN848872.1/LN848922.1
955/1742
PepGMVND/98.4
ND/AY928515.1
ND/524		ND98.1/95.9
U57457.1/AY928515.1
1115/2388		88.2/84.3
AY905553.1/LN848829.1
136/147		99.4/95.9
LN848772.1/LN848841.1
1120/1562
PepLRVND88/ND
KC769819.1/ND
458/ND		NDNDND
ChiLCVNDND81.2/NA 4
JN555601.1/NA
559/NA		NDND
TYLCV98.4/NA
JQ354991.1/NA
131/NA		99.5/NA
KU836749.1/NA
2540/NA		99.5/NA
FJ012359.1/NA
1247/NA		99.7/NA
EF523478.1/NA
1524/NA		99.4/NA
FJ012358.1/NA
2048/NA
ToChLPVND89.1/NA
AY339618.1/NA
120/NA		ND81.9/NA
HM459852.1/NA
337/NA		ND
ToSLCVND87.2/NA
DQ347946.1/NA
359/NA		ND99.5/NA
KC479066.1/NA
411/NA		ND
TPCTVNDNDND82.3/NA
X84735.1/NA
385/NA		ND
ToYSVND84.2/ND
DQ336350.1/ND
470/ND		95.4/ND
KJ742419.1/ND
155/ND		84.9/ND
KX348173.1/ND
192/ND		ND
PYMVNDNDND78.8/ND
FR851299.1/ND
321/ND		ND
OYMMVMalvaceaeND/90.3
ND/GU972604.1
ND/2354		ND93.6/96.4
GU990612.1/JX219471.1
174/226		98.9/94.9
GU990614.1/JX219471.1
1455/336		ND/94.9
ND/JX219471.1
ND/236
CabLCVBrassicaceaeNDND97.4/ND
AJ228570.1/ND
119/ND		84.2/82.8
MH359394.1/DQ178613.1
1645/157		ND
BCaMVFabaceaeNDND/95
ND/AF110190.1
ND/2576		97.2/96.7
AF110189.1/AF110190.1
2058/1296		92.3/88.9
AF110189.1/AF110190.1
353/135		97.9/82.9
AF110189.1/AF110190.1
1005/587
BYMMVND85.3/ND
FJ944023.1/ND
677/ND		NDNDND
ViYMVNDND86.6/86.7
KC430936.1/KC430937.1
758/369		89.6/86
KC430936.1/KC430937.1
242/115		ND
WmCSVCucurbitaceaeNDNDNDND100/100
KY124280.1/KY124281.1
239/1025
SLCVND94.2/ND
KM595165.1/ND
104/ND		ND80.6/83
KM595183.1/DQ285017.1
155/124		79.8/95.3
KM595165.1/M38182.1
188/1649
SPLCVConvolvulaceaeNDND92.4/NA
KX611145.1/NA
1818/NA		80/NA
KJ013582.1/NA
261/NA		ND
BCTVAmaranthaceae99.8/NA
JX487184.1/NA
508/NA		NDNDNDND
Non-cultivated plantsSoMBoVSolanaceaeNDND/84.7
ND/HM585436.1
ND/518		NDND/82.3
ND/HM585436.1
ND/655		ND
SiMSiVMalvaceae96.3/ND
DQ520944.1/ND
854/ND		95.8/96.7
DQ520944.1/DQ356428.1
2581/1582		96.9/90.2
DQ520944.1/DQ356428.1
1003/2085		95.6/87.7
DQ520944.1/DQ356428.1
1584/245		94.2/98.9
DQ520944.1/DQ356428.1
572/289
SiGYSVND84.6/ND
KX348185.1/ND
637/ND		NDNDND
MaBYMV96.9/ND
KU058856.1/ND
1037/ND		ND97.2/84.9
KU058865.1/KU058860.1
403/153		ND94.8/94.5
KU058853.1/KU058859.1
1822/1282
RhGMVFabaceae95/90
EU339939.1/EU339937.1
1049/536		88.9/ND
EU021216.1/ND
253/ND		98.9/95.7
EU339939.1/EU339937.1
2086/675		92.2/85.4
EU339938.1/DQ356429.1
155/240		96.2/82.6
AF408199.1/EU339937.1
264/543
RhGMSVNDND93.4/96.2
DQ406672.1/DQ406673.1
1754/1794		91.2/89.2
DQ406672.1/DQ406673.1
727/353		ND
EuMVEuphorbiaceaeND86.8/ND
JN368145.1/ND
678/ND		ND87.9/86.5
DQ318937.1/DQ520942.1
158/104		ND/93.1
ND/HQ185235.1
ND/249
EuYMVNDNDND91.3/80.1
KY559516.1/KY559581.1
138/342		ND
BleICVAcanthaceaeNDNDND/79
ND/JX827488.1
ND/783		NDND
1 Best match in %, accession numbers and contig length aligned is shown. Contig alignments of ≥300 bp in length were selected regardless whether one or both viral components (DNA A and B) were detected. Contigs alignments of <300 bp are also reported if at least one signature of ≥300 bp for the corresponding virus was detected. 2 Virus acronyms: Monopartite Geminiviruses: Beet curly top virus (BCTV), Chilli leaf curl virus (ChiLCV), Sweet potato leaf curl virus (SPLCV), Tomato pseudo-curly top virus (TPCTV), Tomato yellow leaf curl virus (TYLCV); Bipartite Geminiviruses: Bean calico mosaic virus (BCaMV), Blechum interveinal chlorosis virus (BleICV), Bean yellow mosaic Mexico virus (BYMMV), Cabbage leaf curl virus (CabLCV), Euphorbia mosaic virus (EuMV), Euphorbia yellow mosaic virus (EuYMV), Malvastrum bright yellow mosaic virus (MaBYMV), Okra yellow mosaic Mexico virus (OYMMV), Pepper golden mosaic virus (PepGMV), Pepper huasteco yellow vein virus (PHYVV), Pepper leafroll virus (PepLRV), Potato yellow mosaic virus (PYMV), Rhynchosia golden mosaic Sinaloa virus (RhGMSV), Rhynchosia golden mosaic virus (RhGMV), Sida golden yellow vein virus (SiGYVV), Sida mosaic Sinaloa virus (SiMSiV), Squash leaf curl virus (SLCV), Solanum mosaic Bolivia virus (SoMBoV), Tomato chino la Paz virus (ToChLPV), Tomato severe leaf curl virus (ToSLCV), Tomato yellow spot virus (ToYSV), Vigna yellow mosaic virus (ViYMV), Watermelon chlorotic stunt virus (WmCSV). 3ND: No detected. 4 NA: Not applicable.
Table
Table 4. Nucleotide and amino acid sequence identities (%) between DNA-A genome of Begomovirus isolates identified in the present study with best match sequences available in the database.
Table 4. Nucleotide and amino acid sequence identities (%) between DNA-A genome of Begomovirus isolates identified in the present study with best match sequences available in the database.
Clon CodeLength (bp)Accession No.Virus Acronym 1Reference GenomeComplete GenomeVirus Gene 2
CPV2RepTrApREnC4
N 3na 4nanananana
LV15-Ng-042781MK643155TYLCVEF523478.199.999.610099.799.199.910099.810099.598.5100100
LV15-Sa-032611MK636866SiMSiVDQ520944.195.196.398.4NA 5NA94.995.896.593.795.693.294.898.4
LV17-Rm-022605MK634355RhGMVEU339939.198.698.8100NANA98.598.998.597.198.997.799.297.7
LV15-RM-022578MK618662RhGMSVDQ406672.191.99195.2NANA92.992.396.995.395.393.99286.6
1 TYLCV: Tomato yellow leaf curl virus, SiMSiV: Sida mosaic Sinaloa virus, RhGMV: Rhynchosia golden mosaic virus, RhGMSV: Rhynchosia golden mosaic Sinaloa virus. 2 CP (V1): Coat protein, V2: Precoat protein, Rep (C1): Replication associated protein, TrAp (C2): Transcriptional activator protein, REn (C3): Replication enhancer protein, C4: C4 protein. 3 n: Nucleotide homologies in %. 4 a: Aminoacidic homologies in %. 5 NA: Not applicable.
Table
Table 5. Nucleotide and amino acid sequence identities (%) between DNA-B genome of Begomovirus isolates identified in the present study and best match sequences available in the database.
Table 5. Nucleotide and amino acid sequence identities (%) between DNA-B genome of Begomovirus isolates identified in the present study and best match sequences available in the database.
Clon CodeLength (bp)Accession No.Virus Acronym 1Reference Genome Complete GenomeMP 1NSP 2
n 3na 4Na
LV15-Sa-022583MK643154SiMSiVDQ356428.191.392.795.690.392.2
LV17-Rm-062568MK634539RhGMVDQ356429.19194.899.391.791.6
LV15-Rm-082525MK618663RhGMSVDQ406673.185.990.498.383.386.7
1 MP: Movement protein. 2 NSP: Nuclear shuttle protein. 3 n: Nucleotide homologies in %. 4 a: Aminoacidic homologies in %.
Table
Table 6. Specific PCR detection of begomovirus in individual non-cultivated plants collected from different counties of the five Northern Pacific-regions of Mexico included in this study.
Table 6. Specific PCR detection of begomovirus in individual non-cultivated plants collected from different counties of the five Northern Pacific-regions of Mexico included in this study.
Sampling AreaPlant FamilyPlant SpeciesCollection YearVirus 1 Specific PCR-Positive SamplesNegative SamplesTotal Samples
PHYVVTYLCVSiMSiVRhGMV/RhGMSV
BAJA CALIFORNIA
EnsenadaMalvaceaeMalva parviflora2015ND 21NDND01
SolanaceaeNicotiana glauca2015ND11101
San QuintinMalvaceaeMalva parviflora2015ND31ND14
SONORA
HuatabampoMalvaceaeAbutilon palmeri2015ND11112
Abutilon trisulcatun2015ND11ND01
Anoda pedunculosa2015NDNDNDND11
SolanaceaeDatura stramonium2015111101
Nicotiana glauca2015ND11ND12
Nicotiana plumbanginifolia2015ND11ND02
Solanum. spp201511ND101
Solanum nigrescens2015NDND1ND01
Solanum. spp2015111101
Solanum verbascifolium201511NDND01
NavojoaFabaceaeMeliotus indica2015ND1ND101
MalvaceaeMalva parviflora2015ND23125
Malvella leprosa2015ND11ND01
ObregónMalvaceaeAbutilon palmeri2015NDNDNDND11
Sida rombifolia2015111101
SolanaceaeNicotiana glauca2015111101
Nicotina plumbanginifolia2015111101
Río ColoradoMalvaceaeMalva parviflora2015ND11ND01
SINALOA
Agua calienteFabaceaeRhynchosia minima2016441315
ConcordiaMalvaceaeAnoda pentaschista2012ND1NDND01
GuasaveFabaceaeCrotalaria juncea2016132303
Lonchocarpus lanceolatus2012111101
Melilotus indicus2015NDNDNDND11
201611ND101
MalvaceaeAbutilon palmeri2012ND1NDND01
Abutilon trisulcatun2014ND1ND123
201211ND101
Herissantia crispa2012ND1ND101
Kosteletzkya depressa2012222202
Melochia piramydata2014444404
SolanaceaeDatura reburra2012ND1NDND01
Datura stramonium2012NDNDNDND11
201433ND314
Nicotiana glauca2012ND1ND112
Solanum americanum2012NDNDNDND11
Solanum nigrescens2012ND1ND101
MocoritoMalvaceaeAbutilon trisulcatun201211NDND12
Sidastrum lodiegensis201211ND101
SolanaceaeDatura discolor2012NDNDNDND22
Solanum tridynamum2012NDNDNDND11
Playa CeutaFabaceaeRhynchosia minima2016222202
RosarioFabaceaeMacroptilium atropurpureum2016234404
Rhynchosia precatoria2016222202
Rhynchosia minima2016344404
Senna uniflora20161ND1101
2014111101
MalvaceaeAbutilon trisulcatun2014ND1ND101
Herissantia crispa2014111101
Melochia piramydata2014NDNDNDND11
Sida acuta2014222202
SolanaceaePhysalis acutifolia2014NDNDNDND11
SinaloaDatura inoxia2012ND1NDND01
Solanum tridynamum2012ND2NDND13
COLIMA/NAYARIT
TecománMalvaceaeHerissantia crispa2014155505
Malvastrum coromandelianum2014111213
Sida acuta2014NDND+ND
COAHUILA/DURANGO
La GomaMalvaceaeSida acuta2015NDNDNDND11
Sida rombifolia2015ND21ND02
SolanaceaeDatura stramonium2015ND11ND01
PoanasSolanum elaeagnifolium2016NDNDNDND22
Solanum rostrarum2016233303
TlahualiloMalvaceaeSida rombifolia2015NDND1123
SolanaceaeSolanum elaeagnifolium2015111ND01
TorreónMalvaceaeSphaeralcea angustifolia2015ND2NDND13
SolanaceaeDatura stramonium2015ND11ND01
Nicotiana glauca2015121ND13
Solanum elaeagnifolium2015ND33ND03
Total 4689636230126
1 Virus acronyms: PHYVV, Pepper huasteco yellow vein virus; TYLCV, Tomato yellow leaf virus; SiMSiV, Sida mosaic Sinaloa virus; RhGMV, Rhynchosia golden mosaic virus; RhGMSV, Rhynchosia golden mosaic Sinaloa virus. 2 ND: Not detected.

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Rodríguez-Negrete, E.A.; Morales-Aguilar, J.J.; Domínguez-Duran, G.; Torres-Devora, G.; Camacho-Beltrán, E.; Leyva-López, N.E.; Voloudakis, A.E.; Bejarano, E.R.; Méndez-Lozano, J. High-Throughput Sequencing Reveals Differential Begomovirus Species Diversity in Non-Cultivated Plants in Northern-Pacific Mexico. Viruses 2019, 11, 594. https://0-doi-org.brum.beds.ac.uk/10.3390/v11070594

AMA Style

Rodríguez-Negrete EA, Morales-Aguilar JJ, Domínguez-Duran G, Torres-Devora G, Camacho-Beltrán E, Leyva-López NE, Voloudakis AE, Bejarano ER, Méndez-Lozano J. High-Throughput Sequencing Reveals Differential Begomovirus Species Diversity in Non-Cultivated Plants in Northern-Pacific Mexico. Viruses. 2019; 11(7):594. https://0-doi-org.brum.beds.ac.uk/10.3390/v11070594

Chicago/Turabian Style

Rodríguez-Negrete, Edgar Antonio, Juan José Morales-Aguilar, Gustavo Domínguez-Duran, Gadiela Torres-Devora, Erika Camacho-Beltrán, Norma Elena Leyva-López, Andreas E. Voloudakis, Eduardo R. Bejarano, and Jesús Méndez-Lozano. 2019. "High-Throughput Sequencing Reveals Differential Begomovirus Species Diversity in Non-Cultivated Plants in Northern-Pacific Mexico" Viruses 11, no. 7: 594. https://0-doi-org.brum.beds.ac.uk/10.3390/v11070594

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