Leaf Bacteriome in Sugar Beet Shows Differential Response against Beet curly top virus during Resistant and Susceptible Interactions.

Beet curly top virus (BCTV) significantly reduces sugar beet yield in semi-arid production areas. Genetic resistance to BCTV is limited; therefore, identification of additional resistance-associated factors is highly desired. Using 16S rRNA sequencing and BCTV resistant (R) genotypes (KDH13, KDH4-9) along with a susceptible (S) genotype (KDH19-17), we investigated leaf bacteriome changes during BCTV post inoculation (pi). At day 6 (~6-week-old plants), Cyanobacteria were predominant (~90%); whereas, at week 4 (~10-week-old plants) Firmicutes (11-66%), Bacteroidetes (17-26%), and Verrucomicrobia (12-29%) were predominant phyla and genotype dependent. Both Bacteroidetes and Verrucomicrobia, increased post infection only in the R lines. The bacterial genera Brevibacillus increased at 6 dpi, and Akkermansia and Bacteroides at 4 wkpi in the R lines. Linear discriminant analysis effect size (LEfSe) identified potential biomarkers in the R vs. S lines. Functional profiling revealed bacterial enrichment associated with the TCA cycle, polyisoprenoid, and L-methionine biosynthesis pathways only in KDH4-9 at 6 dpi. At 4 wkpi, bacteria associated with tryptophan and palmitate biosynthesis in the R lines, and uridine monophosphate, phosphatidyl glycerol, and phospholipid biosynthesis in the S line, were enriched. Future characterization of bacterial genera with antiviral properties will help establish their use as biocontrol agents/biomarkers against BCTV.

Genome-based identification of beet curly top Iran virus infecting sugar beet in Turkey and investigation of its pathogenicity by agroinfection.

Beet curly top disease (BCTD) is a yield-limiting viral infection of sugar beet (Beta vulgaris) throughout the arid and semi-arid regions of the world. Two virus species, belonging to two different genera of the family Geminiviridae (Curtovirus and Becurtovirus) had been described as the disease's causative agents on sugar beet. Despite the detection of the BCTD in some sugar beet fields of Turkey sixty years ago, the genome based characterization of BCTD-associated viruses have not been studied previously. In this study, 628 sugar beet plants exhibiting BCTD symptoms were collected from fourteen cities in central Anatolia, the major sugar beet production areas in Turkey. PCR assays of these samples using the respective Curtovirus and Becurtovirus genus-specific primers indicated that the Turkish sugar beet samples' viral sequences belong only to the genus Becurtovirus. The results of sequencing and phylogenetic analysis of the partial genome of the virus obtained from fourteen cities confirmed that BCTD-associated virus in Turkish sugar beet fields is beet curly top Iran virus (BCTIV-Becurtovirus) species. The whole genome of the collected viruses from fourteen cities were amplified by the rolling circle amplification (RCA) and the five most phylogenetically diverse viruses obtained from Afyon, Ankara, Adapazari, Yozgat and Aksaray were sequenced. The results of whole genome sequence analysis indicated >98 % sequence identities with that of a BCTIV variants reported from Urmia province (bordering Turkey) of Iran. A virus genome from Yozgat city had a genomic sequence identity of >97 % with those of BCTIV isolated from cowpea, tomato, pepper and sugar beet in the northern part of Iran. These results suggested that the spread of BCTIV through the region could create a significant threat to the production of sugar beet as well as other agricultural crops. A tandem dimer of a BCTIV-Turkish variant isolated from Ankara city was cloned into Agrobacterium plasmid to be used for agro-infection studies. Agroinoculation of this construct on sugar beet leaves generated severe BCTD symptoms (84 %) which were also confirmed by RCA and qPCR analysis. These results constituted the first genome based characterization of BCTIV Turkish variants and the first report of BCTIV spreading out of Iran.

Identification of a Novel Geminivirus in Fraxinus rhynchophylla in Korea.

Fraxinus rhynchophylla, common name ash, belongs to the family Oleaceae and is found in China, Korea, North America, the Indian subcontinent, and eastern Russia. It has been used as a traditional herbal medicine in Korea and various parts of the world due to its chemical constituents. During a field survey in March 2019, mild vein thickening (almost negligible) was observed in a few ash trees. High-throughput sequencing of libraries of total DNA from ash trees, rolling-circle amplification (RCA), and polymerase chain reaction (PCR) allowed the identification of a Fraxinus symptomless virus. This virus has five confirmed open reading frames along with a possible sixth open reading frame that encodes the movement protein and is almost 2.7 kb in size, with a nonanucleotide and stem loop structure identical to begomoviruses. In terms of its size and structure, this virus strongly resembles begomoviruses, but does not show any significant sequence identity with them. To confirm movement of the virus within the trees, different parts of infected trees were examined, and viral movement was successfully observed. No satellite molecules or DNA B were identified. Two-step PCR confirmed the virion and complementary strands during replication in both freshly collected infected samples of ash tree and Nicotiana benthamiana samples agro-inoculated with infectious clones. This taxon is so distantly grouped from other known geminiviruses that it likely represents a new geminivirus genus.

Nightshade Curly Top Virus: A Possible New Virus of the Genus Topocuvirus Infecting Solanum nigrum in China.

Virus-like symptoms, including leaf deformation and curling, were observed on nightshade (Solanum nigrum) in Zhejiang Province, China. To identify possible pathogenic viruses or viroids, a symptomatic sample was subjected to deep sequencing of small interfering RNAs. Assembly of the resulting sequences led to identification of a novel geminivirus, provisionally designated nightshade curly top virus (NCTV). The complete genomic DNA sequence is 2,867 nucleotides and encodes seven open reading frames. NCTV shares 77.1% overall nucleotide sequence identity, 86.3% coat protein amino acid identity, and 78.9% replication-associated protein amino acid sequence identity with Tomato pseudo-curly top virus, a member of the genus Topocuvirus. PCR screening of nightshade field isolates indicated that NCTV is widely distributed in Zhejiang. Agrobacterium-mediated inoculation revealed that NCTV is highly infectious to Nicotiana benthamiana, S. nigrum, S. lycopersicum, and S. tuberosum. Based on pairwise comparisons and phylogenetic analyses, NCTV is proposed as a provisional member of the genus Topocuvirus.

Requirements for the Packaging of Geminivirus Circular Single-Stranded DNA: Effect of DNA Length and Coat Protein Sequence.

Geminivirus particles, consisting of a pair of twinned isometric structures, have one of the most distinctive capsids in the virological world. Until recently, there was little information as to how these structures are generated. To address this, we developed a system to produce capsid structures following the delivery of geminivirus coat protein and replicating circular single-stranded DNA (cssDNA) by the infiltration of gene constructs into plant leaves. The transencapsidation of cssDNA of the Begomovirus genus by coat protein of different geminivirus genera was shown to occur with full-length but not half-length molecules. Double capsid structures, distinct from geminate capsid structures, were also generated in this expression system. By increasing the length of the encapsidated cssDNA, triple geminate capsid structures, consisting of straight, bent and condensed forms were generated. The straight geminate triple structures generated were similar in morphology to those recorded for a potato-infecting virus from Peru. These finding demonstrate that the length of encapsidated DNA controls both the size and stability of geminivirus particles.

Identification and Characterization of Two Novel Geminiviruses Associated with Paper Mulberry (Broussonetia papyrifera) Leaf Curl Disease.

Paper mulberry (Broussonetia papyrifera) is a perennial woody plant used as source material for Cai Lun paper making, in traditional Chinese medicine, and as livestock feed. To identify the presence of viruses in paper mulberry plants affected by a disease with leaf curl symptoms, high-throughput sequencing of total RNA was performed. Analysis of transcriptome libraries allowed the reconstruction of two geminivirus-like genomes. Rolling-circle amplification and PCR with back-to-back primers confirmed the presence of two geminiviruses with monopartite genomes in these plants, with the names paper mulberry leaf curl virus 1 and 2 (PMLCV-1 and PMLCV-2) proposed. The genomes of PMLCV-1 (3,056 nt) and PMLCV-2 (3,757 to 3,763 nt) encode six proteins, with the V4 protein of PMLCV-1 and the V3 proteins of both viruses having low similarities to any known protein in databases. Alternative splicing of an intron, akin to that of mastre-, becurto-, capula-, and grabloviruses, was identified by small RNA (sRNA)-seq and RNA-seq reads mapping to PMLCV-1 and PMLCV-2 antisense transcripts. Phylogenetic analyses and pairwise comparisons showed that PMLCV-1 and PMLCV-2 are most closely related to, but distinct from, two unassigned geminiviruses, citrus chlorotic dwarf associated virus and mulberry mosaic dwarf associated virus, suggesting that they are two new members of the family Geminiviridae. Field investigation confirmed the close association of the two viruses with leaf curl symptoms in paper mulberry plants and that coinfection can aggravate the symptoms.

Complementary-sense gene regulation in beet curly top virus-SpCT.

A 278-bp region upstream of the beet curly top virus-SpCT (BCTV-SpCT) C2/C3 genes is necessary for promoter activity and exhibits significant sequence similarity to AL2/3 promoter sequences in tomato golden mosaic virus (TGMV). Maximal expression of the downstream C2/3 genes in BCTV-SpCT requires the presence of the C1 protein, which is supported by observations that mutation of the initiator codon for C1 results in decreased C2/C3 expression. This is similar to TGMV and cabbage leaf curl virus, where AL1 is required for maximal AL2/3 expression. Together, these data suggest a common strategy for complementary-sense gene regulation amongst curtoviruses and begomoviruses.

Virus-mediated export of chromosomal DNA in plants.

The propensity of viruses to acquire genetic material from relatives and possibly from infected hosts makes them excellent candidates as vectors for horizontal gene transfer. However, virus-mediated acquisition of host genetic material, as deduced from historical events, appears to be rare. Here, we report spontaneous and surprisingly efficient generation of hybrid virus/host DNA molecules in the form of minicircles during infection of Beta vulgaris by Beet curly top Iran virus (BCTIV), a single-stranded DNA virus. The hybrid minicircles replicate, become encapsidated into viral particles, and spread systemically throughout infected plants in parallel with the viral infection. Importantly, when co-infected with BCTIV, B. vulgaris DNA captured in minicircles replicates and is transcribed in other plant species that are sensitive to BCTIV infection. Thus, we have likely documented in real time the initial steps of a possible path of virus-mediated horizontal transfer of chromosomal DNA between plant species.

Causative Role of Grapevine Red Blotch Virus in Red Blotch Disease.

Grapevine red blotch virus (GRBV) has a monopartite single-stranded DNA genome and is the type species of the genus Grablovirus in the family Geminiviridae. To address the etiological role of GRBV in the recently recognized red blotch disease of grapevine, infectious GRBV clones were engineered from the genome of each of the two previously identified phylogenetic clades for Agrobacterium tumefaciens-mediated inoculations of tissue culture-grown Vitis spp. plants. Following agroinoculation and one or two dormancy cycles, systemic GRBV infection was detected by multiplex polymerase chain reaction (PCR) in Vitis vinifera exhibiting foliar disease symptoms but not in asymptomatic vines. Infected rootstock genotype SO4 (V. berlandieri × V. riparia) exhibited leaf chlorosis and cupping, while infection was asymptomatic in agroinoculated 110R (V. berlandieri × V. rupestris), 3309C (V. riparia × V. rupestris), and V. rupestris. Spliced GRBV transcripts of the replicase-associated protein coding region accumulated in leaves of agroinfected vines, as shown by reverse-transcription PCR; this was consistent with systemic infection resulting from virus replication. Additionally, a virus progeny identical in nucleotide sequence to the infectious GRBV clones was recovered from agroinfected vines by rolling circle amplification, cloning, and sequencing. Concomitantly, subjecting naturally infected grapevines to microshoot tip culture resulted in an asymptomatic plant progeny that tested negative for GRBV in multiplex PCR. Altogether, our agroinoculation and therapeutic experiments fulfilled Koch's postulates and revealed the causative role of GRBV in red blotch disease.

Prunus geminivirus A: A Novel Grablovirus Infecting Prunus spp.

Increased use of metagenomics for routine virus diagnosis has led to the characterization of several genus level geminiviruses from tree fruit long thought to exclusively host RNA viruses. In this study, the identification and molecular characterization of a novel geminivirus is reported for the first time in Prunus spp. The virus, provisionally named Prunus geminivirus A (PrGVA), was identified by Illumina sequencing from an asymptomatic plum tree. PrGVA was subsequently confirmed by rolling cycle amplification, cloning, and Sanger sequencing of its complete genome (3,174 to 3,176 nucleotides) from an additional 18 (9 apricot and 9 plum) field isolates. Apart from the nonanucleotide motif TAATATT↓AC present in its virion strand origin of replication, other conserved motifs of PrGVA support its geminiviral origin. PrGVA shared highest complete genome (73 to 74%), coat protein amino acid (83 to 85%) and rep-associated amino acid (74%) identities with Grapevine red blotch virus (GRBV). PrGVA was graft but not mechanically transmissible. Quantitative polymerase chain reaction screening of Prunus spp. in the National Clonal Germplasm Repository collection using newly designed primers and probes revealed 69.4% (apricot), 55.8% (plum), and 8.3% (cherry) incidences of PrGVA. PrGVA is proposed as a novel member of the genus Grablovirus based on its close genome and phylogenetic relationship with GRBV.

Seed Transmission of Beet Curly Top Virus and Beet Curly Top Iran Virus in a Local Cultivar of Petunia in Iran.

Beet curly top virus (BCTV) and beet curly top Iran virus (BCTIV) are known as the causal agents of curly top disease in beet and several other dicotyledonous plants in Iran. These viruses are transmitted by Circulifer species, and until now, there has been no confirmed report of their seed transmission. A percentage (38.2-78.0%) of the seedlings developed from the seeds of a petunia local cultivar under insect-free conditions showed stunting, interveinal chlorosis, leaf curling, and vein swelling symptoms, and were infected by BCTV when tested by PCR. Presence of BCTV in seed extracts of petunia local cultivar was confirmed by PCR and IC-PCR, followed by sequencing. Agroinoculation of curly top free petunia plants with a BCTV infectious clone resulted in BCTV infection of plants and their developed seeds. These results show the seed infection and transmission of BCTV in a local cultivar of petunia. Similar experiments performed with BCTIV showed that this virus is also seed transmissible in the same cultivar of petunia, although with a lower rate (8.8-18.5%). Seed transmission of curly top viruses may have significant implications in the epidemiology of these viruses.

Beet curly top virus Strains Associated with Sugar Beet in Idaho, Oregon, and a Western U.S. Collection.

Curly top of sugar beet is a serious, yield-limiting disease in semiarid production areas caused by Beet curly top virus (BCTV) and transmitted by the beet leafhopper. One of the primary means of control for BCTV in sugar beet is host resistance but effectiveness of resistance can vary among BCTV strains. Strain prevalence among BCTV populations was last investigated in Idaho and Oregon during a 2006-to-2007 collection but changes in disease severity suggested a need for reevaluation. Therefore, 406 leaf samples symptomatic for curly top were collected from sugar beet plants in commercial sugar beet fields in Idaho and Oregon from 2012 to 2015. DNA was isolated and BCTV strain composition was investigated based on polymerase chain reaction assays with strain-specific primers for the Severe (Svr) and California/Logan (CA/Logan) strains and primers that amplified a group of Worland (Wor)-like strains. The BCTV strain distribution averaged 2% Svr, 30% CA/Logan, and 87% Wor-like (16% had mixed infections), which differed from the previously published 2006-to-2007 collection (87% Svr, 7% CA/Logan, and 60% Wor-like; 59% mixed infections) based on a contingency test (P < 0.0001). Whole-genome sequencing (GenBank accessions KT276895 to KT276920 and KX867015 to KX867057) with overlapping primers found that the Wor-like strains included Wor, Colorado and a previously undescribed strain designated Kimberly1. Results confirm a shift from Svr being one of the dominant BCTV strains in commercial sugar beet fields in 2006 to 2007 to becoming undetectable at times during recent years.

Splicing features in the expression of the complementary-sense genes of Beet curly top Iran virus.

Beet curly top Iran virus (BCTIV) is a distinct geminivirus which has been reported from sugar-beet-growing farms in Iran. In this study, the role of the splicing in expression of complementary-sense genes of BCTIV was studied. Total RNA was extracted from BCTIV-infected tissue, and the predicted intron position of complementary-sense mRNA transcripts was amplified by RT-PCR followed by cloning of the amplicons. Sequence confirmed that both spliced and unspliced mRNAs are synthesized by the same transcription unit. Sequence comparison showed that a 155-nt segment (intron) corresponding to nucleotides 1890-2044 of the viral genome has been removed from the latter transcript and therefore fusion of the C1:C2 genes resulted creation of a continuous reading frame for potential production of intact replication initiator protein (Rep). BCTIV intron comprises of most consensus splicing signals required for splicing in eukaryotes and several plant viruses including mastre- and capulaviruses.

Novel vaccination approach for dengue infection based on recombinant immune complex universal platform.

Dengue infection is on the rise in many endemic areas of the tropics. Vaccination remains the most realistic strategy for prevention of this potentially fatal viral disease but there is currently no effective vaccine that could protect against all four known serotypes of the dengue virus. This study describes the generation and testing of a novel vaccination approach against dengue based on recombinant immune complexes (RIC). We modelled the dengue RIC on the existing Ebola RIC (Phoolcharoen, et al. Proc Natl Acad Sci USA 2011;108(Dec (51)):20695) but with a key modification that allowed formation of a universal RIC platform that can be easily adapted for use for other pathogens. This was achieved by retaining only the binding epitope of the 6D8 ant-Ebola mAb, which was then fused to the consensus dengue E3 domain (cEDIII), resulting in a hybrid dengue-Ebola RIC (DERIC). We expressed human and mouse versions of these molecules in tobacco plants using a geminivirus-based expression system. Following purification from the plant extracts by protein G affinity chromatography, DERIC bound to C1q component of complement, thus confirming functionality. Importantly, following immunization of mice, DERIC induced a potent, virus-neutralizing anti-cEDIII humoral immune response without exogenous adjuvants. We conclude that these self-adjuvanting immunogens have the potential to be developed as a novel vaccine candidate for dengue infection, and provide the basis for a universal RIC platform for use with other antigens.

Comparison and evaluation of three diagnostic methods for detection of beet curly top virus in sugar beet using different visualizing systems.

To diminish the time required for some diagnostic assays including polymerase chain reaction (PCR), loop-mediated isothermal amplification (LAMP; due to mainly DNA extraction step) and also triple antibody sandwich enzyme-linked immunosorbent assay (TAS-ELISA) into a minimum level, an innovative immunocapture LAMP (IC-LAMP) and immunocapture PCR (IC-PCR) protocol on the basis of beet curly top virus (BCTV) genome was used and optimized. TAS-ELISA was employed first to validate the existence of the virus. All six IC-LAMP primers (i.e. forward outer primer (F3), backward outer primer (B3), forward inner primer (FIP), backward inner primer (BIP), loop forward (LF) and loop backward (LB)) together with IC-PCR primers were designed on the basis of the replication-associated protein (rep) gene (GenBank accession AF379637.1) of BCTV genome. Also, a novel colorimetric IC-LAMP assay for rapid and easy detection of BCTV was developed here, its potential compared with TAS-ELISA and IC-PCR assays. The method, on the whole, had the following advantages over the two mentioned procedures: (i) fascinatingly, no need of DNA extraction; (ii) no requirement of expensive and sophisticated tools for amplification and detection; (iii) no post-amplification treatment of the amplicons and (iv) a flexible and easy detection approach, which is visually detected by naked eyes using diverse visual dyes.

Defective DNAs of beet curly top virus from long-term survivor sugar beet plants.

Long-term surviving sugar beet plants were investigated after beet curly top virus infection to characterize defective (D) viral DNAs as potential symptom attenuators. Twenty or 14 months after inoculation, 20 D-DNAs were cloned and sequenced. In contrast to known D-DNAs, they exhibited a large range of sizes. Deletions were present in most open reading frames except ORF C4, which encodes a pathogenicity factor. Direct repeats and inverted sequences were observed. Interestingly, the bidirectional terminator of transcription was retained in all D-DNAs. A model is presented to explain the deletion sites and sizes with reference to the viral minichromosome structure, and symptom attenuation by D-DNAs is discussed in relation to RNA interference.

Intergeneric recombination between a new, spinach-infecting curtovirus and a new geminivirus belonging to the genus Becurtovirus: first New World exemplar.

A novel curtovirus, spinach severe curly top virus (SSCTV), was associated with symptomatic spinach plants collected from a commercial field in south-central Arizona during 2009. In addition, a second viral molecule of about 2.9 kb from the same spinach plants was amplified, cloned and sequenced. The latter isolate, herein named spinach curly top Arizona virus (SCTAV), was found to share 77 % pairwise sequence identity with beet curly top Iran virus (BCTIV), a leafhopper-transmitted geminivirus that has been assigned to the new genus Becurtovirus. The SCTAV genome encodes three viral-sense genes, V1, V2, and V3, and two complementary-sense genes, C1 and C2. There was no evidence for the presence of either a C3 or C4 ORF in the genome sequence. The genome organization of SCTAV is not like that of New World curtoviruses but instead is similar to that of BCTIV, which, to date, is only known to be present in Iran. Consistent with this observation, SCTAV and BCTIV both contain the unusual nonanucleotide TAAGATT/CC and a replication-associated protein, Rep (or C1), that is more closely related to the mastrevirus Rep than to those of curtoviruses reported to date. Both SSCTV and SCTAV were found to have a recombinant genome containing sequences (AY548948) derived from ancestral SCTV sequences in the virion-sense portions of the genome. Agroinoculation of Nicotiana benthamiana (Domin) plants with the cloned genome of SCTAV resulted in infection of 95 % of the plants and the development of severe curling symptoms, whereas only 20 % of the SSCTV-inoculated plants were infected, developing only mild curling symptoms. When plants were co-inoculated with both viruses, the frequency of infection remained higher for SCTAV than for SSCTV (80 % vs. 20 %), indicating no evidence of synergistic effects between the two viruses with respect to efficiency of infection.

Fulfilling Koch's postulates for beet curly top Iran virus and proposal for consideration of new genus in the family Geminiviridae.

Beet curly top Iran virus (BCTIV) is a divergent geminivirus with biological properties similar to those of curtoviruses; however, the virus is distinct from curtoviruses phylogenetically and in its genome organisation. The replication-associated protein is phylogenetically more closely related to those of mastreviruses than to those of curtoviruses whereas the capsid protein shares high amino acid sequence identity (77-83 %) with those of curtoviruses. The 17 BCTIV genomes from Iran share ~77 % pairwise nucleotide sequence identity with spinach curly top Arizona virus (SCTAV) from Arizona, USA, which was characterised recently. To demonstrate the infectivity of the monopartite BCTIV genome and to fulfil Koch's postulates, an infectious clone was constructed using a dimer of the full-length genome of an isolate from this study - BCTIV-[IR:Neg:B33P:Sug:08]. Agroinoculation with the cloned DNA resulted in the efficient infection of 74 % of sugar beet plants, which resulted in curly top symptoms. The curly top infection of agroinoculated plants was successfully transmitted to 80 % of healthy sugar beet plants by the natural BCTIV vector, Circulifer haematoceps. Since BCTIV and SCTAV share <62 % pairwise nucleotide sequence identity with all other geminiviruses and have unique genome architectures and properties, and since this is coupled with phylogenetic support at the full-genome level and that of it proteins, we propose that they should be re-classified as members of a new genus, "Becurtovirus", in the family Geminiviridae.

Genetic diversity and host range studies of turnip curly top virus.

Turnip curly top virus (TCTV) is a unique geminivirus that has recently been characterised as infecting turnips in Iran. The genome of TCTV shares <68 % pairwise identity with other geminiviruses and has a genome organisation similar to that of curtoviruses and topocuvirus. The replication-associated protein (Rep) bears the highest similarity to curtovirus Reps (48.5-69.0 %); however, in the case of the capsid protein (CP), the extent of similarity is only 39.5-44.5 %. We constructed an agroinfectious clone of TCTV and undertook host range studies on ten plant species; in three species (turnip, sugar beet and cowpea), we detected infection which presents curly top symptoms in turnip and sugar beet. The efficiency of TCTV infection in agroinoculated turnip plants was 71.7 %, and the infection was successfully transmitted to 80 % of the healthy turnip plants used in the insect transmission studies by Circulifer haematoceps under greenhouse conditions. We also determined the genome sequence of 14 new TCTV isolates from southern Iran isolated from turnips. We observed ~13 % diversity amongst all the TCTV isolates and found evidence of recombination in the CP- and Rep-coding regions of the genomes.

C2 from Beet curly top virus promotes a cell environment suitable for efficient replication of geminiviruses, providing a novel mechanism of viral synergism.

• Geminiviruses are plant viruses with circular, single-stranded (ss) DNA genomes that infect a wide range of species and cause important losses in agriculture. Geminiviruses do not encode their own DNA polymerase, and rely on the host cell machinery for their replication. • Here, we identify a positive effect of the curtovirus Beet curly top virus (BCTV) on the begomovirus Tomato yellow leaf curl Sardinia virus (TYLCSV) infection in Nicotiana benthamiana plants. • Our results show that this positive effect is caused by the promotion of TYLCSV replication by BCTV C2. Transcriptomic analyses of plants expressing C2 unveil an up-regulation of cell cycle-related genes induced on cell cycle re-entry; experiments with two mutated versions of C2 indicate that this function resides in the N-terminal part of C2, which is also sufficient to enhance geminiviral replication. Moreover, C2 expression promotes the replication of other geminiviral species, but not of RNA viruses. • We conclude that BCTV C2 has a novel function in the promotion of viral replication, probably by restoring the DNA replication competency of the infected cells and thus creating a favourable cell environment for viral spread. Because C2 seems to have a broad impact on the replication of geminiviruses, this mechanism might have important epidemiological implications.