First Report of Tomato Zonate Spot Virus Infecting Tobacco in Guizhou Province, China

Iris tectorum Maxim, a very popular Chinese traditional medicinal perennial herb belonging to the Iridaceae family, is widely grown as a year-round ornamental in China. During May to August 2014, as part of a survey for tospoviruses (family Bunyaviridae) in flue-cured tobacco, symptoms suspected to be caused by tospoviruses were observed on I. tectorum around farmers' fields in Kunming, Yunnan province. Symptoms were chlorotic spots on younger leaves and necrosis on older leaves. Since Tomato spotted wilt virus (TSWV) and Tomato zonate spot virus (TZSV) are two common tospoviruses in flue-cured tobacco fields in Yunnan, ELISA with monoclonal TSWV antibody (provided by J. X. Wu, Zhejiang University, China) and polyclonal TZSV antiserum (provided by J. H. Dong, Yunnan Academy of Agriculture Science, China) was performed to identify the presence of virus. Positive extinction values (ODλ405nm 0.835 ± 0.121 and 1.024 ± 0.193, as compared with the negative 0.153 ± 0.076 and the positive control 0.510 ± 0.109 at a confidence interval of P ≤ 0.05) were obtained from two symptomatic samples with TZSV antibody but not with TSWV. The absence of TSWV was confirmed with a commercially available immune-strip (Agdia, Elkhart, IN), following the manufacturer's instructions. To further verify the causal agent of these symptoms, total RNA was isolated from two symptomatic and one asymptomatic samples and reverse transcribed using degenerate primer J13 (1). These cDNAs were then used as a template in a universal PCR assay using specific primers TZSVNF (5'-ATGTCTAACGTCCGGAGTTTAACAC-3') and TZSVNR (5'-TTAAAAAGACAGATCATTGCTG-3'), which amplify the complete nucleocapsid (N) protein. The PCR was carried out for denaturation at 94°C for 3 min, and subsequently 30 cycles were carried out, with each cycle consisting of 94°C for 45 s, 55°C for 45 s, and 72°C for 1 min, followed by a final extension step at 72°C for 10 min. An 0.8-Kb DNA fragment was amplified from symptomatic samples and cloned into a pGEM-T Easy (Promega, Madison, WI) vector. Three clones of each sample were selected and sequenced. BLAST analysis of the obtained sequences (Accession Nos. KM452916 and KM452917) revealed that the N sequences of these isolates have 96 to 99% nucleotide identity and 99 to 100% amino acid identity with the deposit TZSV sequence in NCBI from Yunnan (JN116580 to JN116583 and EF552433) (2). These combined results provide further confirmation of TZSV infection. It is known that perennial herb or ornamental plants may act as reservoirs for tospoviruses that can infect cultivated crops because tospoviruses have a very broad host range. Therefore, elaborate inspections for tospoviruses and appropriate management strategies to limit virus spread are necessary for production of crops. To our knowledge, this is the first report of TZSV in I. tectorum Maxim.

Cyclocodon lancifolius (Roxb.) Merr. (Campanulaceae), commonly known as Hong Guo Ginseng (Pan et al., 2024), is a perennial herb widely distributed in tropical and subtropical East Asia, mainly cultivated in Yunnan, Sichuan, and Guizhou provinces of China, and has potential therapeutic effects on diabetes, antioxidation, and gout (Ma et al., 2023). Tomato zonate spot virus (TZSV), Orthotospovirus tomatozonae in the genus Orthotospovirus of the family Tospoviridae, was first reported infecting tomato (Solanum lycopersicum) and pepper (Capsicum annuum) in Yunnan, China, in 2008 (Dong et al., 2008). In November 2024, a total of 10 symptomatic C. lancifolius plants showing virus-like symptoms, including leaf malformation, chlorosis, and necrosis, were collected from Malipo County, Wenshan Prefecture, Yunnan Province (n=5) and the Fangxiangliqi Garden of Yunnan University of Chinese Medicine (n=5) (Fig. S1). Spherical viral particles with a diameter of 80 to 100 nm were observed in the sap of diseased leaves by transmission electron microscopy (Fig. S2). Reverse transcription-polymerase chain reaction (RT-PCR) screening was performed using degenerate primers CIFor/CIRev for potyviruses and dTospo-F2/dTospo-R2 for orthotospoviruses (Huang et al., 2018), with total RNA extracted from diseased leaves as the template. Only the latter primers amplified 312-bp fragments, while CIFor/CIRev yielded none. Sequencing and BLASTn analysis of these 312-bp fragments showed 95.53% nucleotide identity with the TZSV tomato-YN isolate (accession no. OR603945.1). The virus isolate was named TZSV-HGG. Total RNA from a sample pool (n=4) was sequenced on the Illumina NovaSeq 6000 (150-bp paired-end) by Lingke Biotechnology Co., Ltd. (Kunming, China). After quality control and filtering, 238,829,666 clean reads were obtained and assembled into contigs using SPAdes v. 3.15.3. Through online BLASTn search, six contigs were found to have nucleotide identities of 94.13%, 94.13% to 99.38%, and 95.53% to 98.77% with partial sequences of the L, M, and S segments of TZSV, respectively. To characterize the genome of the virus, specific primers were used to amplify the full-length genome sequence by fragment amplification. Following PCR product purification, cloning, and Sanger sequencing analysis, the amplified sequences were assembled and confirmed to be identical to high throughput sequencing assembled contigs by BLASTn (Fig. S3). The L, M, and S segments of the TZSV-HGG genome are 8,917 nt, 4,696 nt, and 3,212 nt, respectively, and have been deposited in GenBank (accession numbers PV892472, PV892473, and PV892474). Blastn analysis showed that these segments share 96.31%, 97.31%, and 94.24% nucleotide sequence identity with the L (NC_010491.1), M (NC_010490.1), and S (NC_010489.1) segments of the TZSV tomato-YN isolate, respectively. Compared to the TZSV tomato-YN isolate, the M RNA intergenic region contains a 252 nt deletion, and the S RNA intergenic region contains a 67 nt deletion. Mechanical inoculation and RT-PCR confirmed TZSV-HGG systemically infects N. tabacum cv. K326, causing chlorosis and necrotic spots at 14 days post-inoculation. To our knowledge, this is the first report of natural infection of C. lancifolius by TZSV, expanding the host range of this virus and providing critical bases for disease epidemic surveillance and integrated control.

Mosaic disease caused by Cucumber mosaic virus (CMV) and Chilli veinal mottle Virus (ChiVMV) has been distributed widely in chilli in Indonesia and considered as important disease. A research was conducted to investigate the spread and incidence of CMV and ChiVMV in Rejang Lebong, Bengkulu and to identify its insect vector. Symptomatic and asymptomatic leaf samples were collected systematically from several chillipepper fields for further detection by DAS-ELISA (Double antibody sandwich-enzyme linked immunosorbant assay) using specific antibody for CMV and ChiVMV. The result showed that infection of both CMV and ChiVMV was found with disease incidence reached 20-50%, whereas infection only by ChiVMV or CMV were 50-80% and 20-50%, respectively. One species of aphid, i.e. Aphis gossypii was found from the fields.Key words: Aphis gossypii, CMV, ChiVMV, disease incidence

Tomato zonate spot virus (TZSV) is a phytopathogen of the genus Orthotospovirus (Bunyaviridae) that is widespread in many areas of Southwest China. TZSV is mainly transmitted by Frankliniella occidentalis, but its exact infection route remains unclear. To explore this issue, we detected the nucleocapsid protein of TZSV in the digestive systems of first-instar F. occidentalis nymphs fed with TZSV-infected pepper leaves. TZSV infection in the F. occidentalis digestive system begins within 4 h post-first access to diseased plants: The foregut is likely the primary site of infection, and primary salivary glands (PSGs) are the destination. There are three potential routes for TZSV transmission from the alimentary canal to the PSGs: (1) virus dissemination from the midgut to hemocoel followed by movement to the PSGs; (2) accumulation in midgut epithelial cells and arrival at PSGs via tubular salivary glands and efferent ducts; and (3) arrival at epitheliomuscular cells of the forepart of the midgut and movement along the ligament to the PSGs. We tested the transmission efficiency of F. occidentalis in second-instar nymphs and female and male adults. TZSV was transmitted in a persistent-propagative mode by both nymphs and adults, with adults appearing to show slightly higher transmission efficiency than nymphs. We confirmed the presence of all three routes for TZSV transmission in F. occidentalis and determined that like other Orthotospoviruses, TZSV is transmitted in a persistent-propagative manner. These results should facilitate the control of TZSV-related diseases and further our understanding of the transmission biology of Orthotospoviruses in general.

Tomato zonate spot virus (TZSV), a member of the genus orthotospovirus, causes severe damage to vegetables and ornamental crops in southwest China. The NSs protein is an RNA silencing suppressor in various orthotospovirus like TZSV, but its mechanism and role in virus infection are poorly understood. Here, we observed that an NSs-GFP fusion protein was transiently expressed on the plasma membrane and Golgi bodies in Nicotiana benthamiana plants. The TZSV NSs gene was silenced and infiltrated into N. benthamiana and N. tabacum cv. K326. RT-qPCR and Indirect enzyme-linked immunosorbent assay (ID-ELISA) showed that the transcription and the protein expression of the NSs gene were inhibited by more than 90.00%, and the symptoms on silenced plants were alleviated. We also found that the expression of the Zingipain-2-like gene significantly decreased when the NSs gene was silenced, resulting in co-localization of the NSs-GFP and the Zingipain-2-like-mCherry fusion protein. The findings of this study provide new insights into the mechanism of silencing suppression by NSs, as well as its effect on systemic virus infection, and also support the theory of disease resistance breeding and control and prevention of TZSV in the field.

Tomato zonate spot virus (TZSV), a member of the genus orthotospovirus, causes severe damage to vegetables and ornamental crops in southwest China. The NSs protein is an RNA silencing suppressor in various orthotospovirus like TZSV, but its mechanism and role in virus infection are poorly understood. Here, we observed that an NSs-GFP fusion protein was transiently expressed on the plasma membrane and Golgi bodies in Nicotiana benthamiana plants. The TZSV NSs gene was silenced and infiltrated into N. benthamiana and N. tabacum cv. K326. RT-qPCR and Indirect enzyme-linked immunosorbent assay (ID-ELISA) showed that the transcription and the protein expression of the NSs gene were inhibited by more than 90.00%, and the symptoms on silenced plants were alleviated. We also found that the expression of the Zingipain-2-like gene significantly decreased when the NSs gene was silenced, resulting in co-localization of the NSs-GFP and the Zingipain-2-like-mCherry fusion protein. The findings of this study provide new insights into the mechanism of silencing suppression by NSs, as well as its effect on systemic virus infection, and also support the theory of disease resistance breeding and control and prevention of TZSV in the field.

Chilli veinal mottle virus (ChiVMV), a potyvirus, is widespread over the world. In China, it was first reported in chili pepper (Capsicum annuum) in Hainan Province (south China) in 2006 (2). Subsequently, it was reported in tobacco (Nicotiana tabacum) in Yunnan Province (southwest China) in 2011 (1). Sichuan Province is one of the largest vegetable producing areas of China. In May 2012, tomatoes with leaves displaying virus-infected symptoms like mottling, mosaic, narrowing, or curling were observed in several fields of Chengdu, eastern Sichuan Province, southwest China. Of the 20 fields we investigated, four fields with 90% tomato plants were infected. During 2012 and 2013, six samples were collected from symptomatic tomato leaves based on different symptoms and locations. All six samples were assayed by western blotting using polyclonal antisera (Cucumber mosaic virus [CMV], Tobacco mosaic virus [TMV]) obtained from Agdia (Elkhart) and one antiserum to ChiVMV obtained from Yunnan Academy of Agricultural Science (China). Two samples from Pengzhou and one sample from Shuangliu exhibiting mosaic leaves were positive for TMV, one sample from Pixian exhibiting narrowing leaves was positive for CMV, and the other two samples from Shuangliu exhibiting mottle and leaf distortion were positive for ChiVMV. Total RNAs was extracted from all six samples and healthy tomato leaves using Trizol reagent (Invitrogen), First-strand cDNA synthesis primed with oligo(dT) by SuperScript III Reverse Transcriptase (Invitrogen). RT-PCR was performed using primer pairs ChiVMV-CP F (5'-GCAGGAGAGAGTGTTGATGCTG-3') and ChiVMV-CP R (5'-(T)16AACGCCAACTATTG-3'), which were designed to direct the amplification of the entire capsid protein (CP) gene and 3' untranslated region (3'-UTR) of ChiVMV (GenBank Accession No. KC711055). The expected 1,166-bp DNA fragment was amplified from the two tomato samples from Shuangliu that were positive for ChiVMV in the western blot tests, but not from the others. The obtained fragments were purified and cloned into the PMD18-T vector (TaKaRa) and sequenced. The sequencing results showed that the two ChiVMV isolates from tomato in Shuangliu were identical (KF738253). Nucleotide BLAST analysis revealed that this ChiVMV isolate shared ~84 to 99% nucleotide identities with other ChiVMV isolates available in GenBank (KC711055 to KF220408). To fulfill Koch's postulates, we isolated this virus by three cycle single lesion isolation in N. tabacum, and mechanically inoculated it onto tomato leaves. The same mottle and leaf distortion symptoms in systemic leaves were observed. Subsequent RT-PCR, fragment clone, and sequence determination tests were repeated and the results were the same. All the evidence from these tests revealed that the two tomato plants were infected by ChiVMV. To our knowledge, this is the first report of ChiVMV naturally infecting tomato in China. It shows that ChiVMV is spreading in China and is naturally infecting a new solanaceous crop in the southwest area, and the spread of the virus may affect tomato crop yields in China. Thus, it is very important to seek an effective way to control this virus.

Chilli pepper is an important economic crop and virus diseases are constraints on its production. In 2018, disease surveys were conducted at a 15-ha chilli pepper plantation in Dehong, southwest of Yunnan Province, China. Throughout the chilli pepper growing season from March to September, pepper plants developed three different virus-like symptoms on leaves, including mosaic, yellow mottle and shrinkage (Fig. S1). Based on observation of virus-like symptomatic phenotypes, the field surveys indicated that the disease incidence ranged from 30% in March to a peak 100% in July, resulting in a significant loss of pepper fruit from 30 to 100% depending on plot of the field. Potyvirus-like filamentous particles, around 11*760 nm, were observed under electron microscopy in the sap of symptomatic leaves (Fig. S1). To further determine the viral species in these samples, total RNA was extracted from three symptomatic samples using a Trans ZolUp Plus RNA Kit (Trans Gene, Beijing, China). Complementary DNA (cDNA) was synthesized using oligo (dT) and M-MLV reverse transcriptase (Promega, Madison, Wisconsin, USA) according to the manufacturer's instructions, and the polymerase chain reaction (PCR) was performed using degenerate primers specific to genus Potyvirus targeting HC-Pro region (HPFor: 5-TGYGAYAAYCARYTIGAYIIIAAYG-3; HPRev: 5-GAICCRWAIGARTCIAIIACRTG-3) (Ha et al. 2008) under the following conditions: an initial denaturation at 94°C for 4min, 30 cycles of denaturation at 94°C for 30 s, annealing at 56°C for 30 s, extension at 72°C for 30s, and a 10min final extension at 72°C. An expected 683-bp DNA fragment was amplified and cloned into the pMD 18-T Vector (Takara, Japan) for sequencing. Sequence analysis using BLAST revealed that the amplicons of phenotype I (Fig. S1a) shared highest nucleotide identity (85.6%) with wild tomato mosaic virus (WTMV) isolate from Vietnam (GenBank no. DQ851495) while the amplicons of phenotype III (Fig. S1c) showed the highest nucleotide identity (93%) with chilli veinal mottle virus (ChiVMV) isolate from Sichuan, China. (GenBank no. MK405594). Amplicons of phenotype II included both sequence of above WTMV and ChiVMV, indicating co-infection of phenotype II (Fig. S1b). Phenotype I sample was used for mechanical inoculation on chilli pepper as described previously (Yang et al.2013). After ten days, virus-like symptoms similar to phenotype I were observed on leaves, and WTMV infection, but not ChiVMV infection, was confirmed by RT-PCR tests on inoculated pepper plants (Fig. S1 e, f). To further ascertain the incidence of these two viruses in the field, primers WT-F: 5'-GTTGTTGAATGTGGTTTAGTT-3' and WT-R: 5'-AGATGTGCTTTGGAAGCGACC-3' were designed based on the WTMV sequence (GenBank no. DQ851495) to amplify a 476 bp product, and primers Ch-F/Ch-R (Ch-F: 5'-AAAGAAGAACAAGCGACAGAA-3', Ch-R: 5'-CATCACGCAAATATTCAAAGC-3') were designed based on ChiVMV sequence (GenBank no. MK405594.1) to amplify a 332 bp product. RT-PCR was conducted on 31 field-collected samples, and amplicons of expected sizes, 476bp and 332bp, corresponding to WTMV and ChiVMV, respectively, were obtained and sequenced to verify their identity. The results (Fig. S2) showed that 71% (22/31) of the samples tested positive for WTMV, 90% (28/31) tested positive for ChiVMV, and 65% (20/31) were co-infected with the two viruses. The WTMV was first reported infecting wild tomatoes in Vietnam in 2008 (Ha et al. 2008), and later reported in China in Nicotiana tabacum (Sun et al. 2015), Solanum nigrum (Zhang et al. 2019), and wild eggplant (Zhang et al. 2014). To our knowledge, this is the first report of WTMV infection on chilli pepper under natural conditions. Our study revealed that the chilli pepper disease in Dehong was caused by single or co-infection of WTMV and ChiVMV. It is necessary to find effective methods to control these two viruses.

In Guangxi Province of southwest China, diseases caused by Tospoviruses (family Bunyaviridae) pose a serious threat to tobacco (Nicotiana tobacum) production. During surveys conducted annually at Xinrong Village in Jingxi County from 2008 to 2010, more than 130 ha of fields were found to have 10 to 50% of plants exhibiting symptoms similar to spotted wilt caused by Tomato spotted wilt virus (TSWV). During this period, disease symptoms at similar prevalence and incidence were also found at Fushan, Debao County in most of the cultivars produced in these areas, including Yunyan 85, 87, 92, 97, and K326. Symptoms on tobacco varied but commonly included dwarfing, midrib browning, distorted apical buds, and concentric ringspots that coalesced to form large areas of dead leaf tissue. Mechanical inoculation from diseased tobacco leaves with concentric ringspots back to tobacco cv. Yunyan 85 or 87, resulted in 12 of 16 plants with symptoms similar to those observed in the field. No symptoms on plants developed following inoculation with buffer only. Symptoms found in the field resembled those caused by TSWV. However, testing using TSWV-specific antiserum was shown to be negative by double-antibody sandwich-ELISA (Agdia, Elkhart, IN). Total RNA was extracted from 27 diseased tobacco plants collected from different regions in Guangxi using Trizol reagent (Invitrogen, Carlsbad, CA) according to the manufacturer's instructions. RNA extracts were amplified by reverse transcription (RT)-PCR using the degenerate primers T2740 (ATGGGDATNTTTGATTTCATG) and T3920c (TCATGCTCATSAGRTAAATYTCTCT) designed to target the partial RNA-dependent RNA polymerase (RdRp) sequence of members in the genus Tospovirus (3). Amplification was performed at 42°C for 60 min, followed by 35 cycles of PCR (30 s denaturation at 94°C, 45 s annealing at 55°C, and 30 s extension at 72°C) and a 7-min final extension at 72°C. A PCR product of approximately 1.2 kb was amplified from 21 diseased plants. RT-PCR amplicons were cloned into the pUC19-T Simple Vector (TaKaRa, Dalian, China) and sequenced in both directions. Sequences were assembled and analyzed by DNAStar 5.01 (DNASTAR, Madison, WI). Sequences of representative isolates were deposited in GenBank (Accession Nos. JN020022 to JN020027). The 1.2-kb partial RdRp sequences of these isolates were shown to have 94.4 to 95.3% nucleotide identity and 96.5 to 97.5% amino acids identity to Tomato zonate spot virus (TZSV) (GenBank Accession No. NC_010491) (1). Among these TZSV isolates from Guangxi, the partial RdRp sequences have 98.0 to 99.4% nucleotide identity and 98.8 to 100% amino acids identity with each other. The presence of TZSV was further confirmed in diseased tobacco plants by indirect ELISA using antiserum of TZSV (provided by Prof. Zhongkai Zhang, Agricultural Academy of Yunnan, China). TZSV has been characterized as a novel tospovirus on various hosts including tobacco in Yunnan province (1,2). To our knowledge, this is the first report of TZSV-associated disease on tobacco in Guangxi Province, southwest China. Further work is necessary to study the epidemiology and management of the disease.

BackgroundEmerging tospoviruses cause significant yield losses and quality reduction in vegetables, ornamentals, and legumes throughout the world. So far, eight tospoviruses were reported in China. Tomato fruits displaying necrotic and concentric ringspot symptoms were found in Guizhou province of southwest China.FindingELISA experiments showed that crude saps of the diseased tomato fruit samples reacted with antiserum against Tomato zonate spot virus (TZSV). Electron microscopy detected presence of quasi-spherical, enveloped particles of 80–100 nm in such saps. The putative virus isolate was designated 2009-GZT. Mechanical back-inoculation showed that 2009-GZT could infect systemically some solanaceous crop and non-crop plants including Capiscum annuum, Datura stramonium, Nicotiana benthamiana, N. rustica, N. tabacum and Solanum lycopersicum. The 3012 nt full-length sequence of 2009-GZT S RNA shared 68.2% nt identity with that of Calla lily chlorotic spot virus (CCSV), the highest among all compared viruses. This RNA was predicted to encode a non-structural protein (NSs) (459 aa, 51.7 kDa) and a nucleocapsid protein (N) (278 aa, 30.3 kDa). The N protein shared 85.8% amino acid identity with that of CCSV. The NSs protein shared 82.7% amino acid identity with that of Tomato zonate spot virus(TZSV).ConclusionOur results indicate that the isolate 2009-GZT is a new species of Tospovirus, which is named Tomato necrotic spot virus (TNSV). This finding suggests that a detailed survey in China is warranted to further understand the occurrence and distribution of tospoviruses.

The European Commission requested EFSA’s Panel on Plant Health to perform the pest categorisation for the 24 viruses of the Tospovirus genus for the EU territory. The following tospoviruses were analysed: Tomato spotted wilt virus (TSWV), Impatiens necrotic spot virus (INSV), Iris yellow spot virus (IYSV), Polygonum ringspot virus (PolRSV), Groundnut ringspot virus (GRSV), Tomato chlorotic spot virus (TCSV), Alstroemeria necrotic streak virus (ANSV), Chrysanthemum stem necrosis virus (CSNV), Melon severe mosaic virus (MSMV), Tomato yellow (fruit) ring virus (TYRV), Tomato zonate spot virus (TZSV), Groundnut yellow spot virus (GYSV), Groundnut chlorotic fan-spot virus (GCFSV), Groundnut bud necrosis virus (GBNV), Zucchini lethal chlorosis virus (ZLCV), Capsicum chlorosis virus (CaCV), Watermelon bud necrosis virus (WBNV), Watermelon silver mottle virus (WSMoV), Tomato necrotic ringspot virus (TNRV), Calla lily chlorotic spot virus (CCSV), Melon yellow spot virus (MYSV), Soybean vein necrosis associated virus (SVNaV), Bean necrotic mosaic virus (BeNMV) and Pepper necrotic spot virus (PNSV). In reaching its conclusions, the Panel considered four parameters to be of critical importance in the risk assessment area: (i) the presence of a tospovirus, (ii) the existence of host plants, (iii) the existence of thrips vector species and (iv) the potential for damage to crops grown in Europe. Based on its analysis, the Panel concluded that the 24 viruses analysed could be allocated to four different risk groups. Seven viruses (GRSV, TCSV, ANSV, CSNV, MSMV, TYRV, TZSV) for which both thrips species vectors and natural or experimental hosts crops are present in the EU territory were considered by the Panel to represent the highest risk to the EU territory. In contrast, three viruses (INSV, IYSV and PolRSV) already present in the risk assessment area were not considered by the Panel to pose a risk justifying the development of full risk assessments.

Capsicum chlorosis virus (CaCV) is a negative sense ssRNA virus belonging to the genus Orthotospovirus in the family Tospoviridae. It was first discovered in Australia, and then reported in other places including Thailand, China, India, Greece, and United States (Zheng et al.2011; Melzer et al.2014; Chrysoula et al. 2018; Abudurexiti et al. 2019). CaCV infects plants of the families Amaranthaceae, Apocynaceae, Chenopodiaceae, Cucurbitaceae, Amaryllidaceae, Fabaceae and Solanaceae (Basavaraj et al. 2017; Basavaraj et al. 2020). Chromolaena odorata L. (commonly known as Feiji cao in China) is an invasive weedy herb that belongs to the genus Eupatorium (family Asteraceae), and is native to Central America. In May 2020, serrated chlorotic ring and chlorotic ringspots resembling symptoms of orthotospovirus infection (Supplementary Figure 1) was observed on the leaves of C. odorata plants in Honghe County, Yunnan. Three symptomatic leaf samples were collected and double antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA) was performed using antisera targeting Tomato spotted wilt virus (TSWV), Calla lily chlorotic spot virus (CCSV), Capsicum chlorosis virus (CaCV), and Tomato zonate spot virus (TZSV) (Proteintech Group, Inc., China). Buffer solution and healthy leaves were used as a blank and negative controls, respectively. All three symptomatic samples showed positive reactions with only CaCV antiserum (OD450 of 0.315-0.345 relative to 0.078 and 0.076 for healthy plants and the blank control, respectively. The total RNA extracted from the positive samples were further analyzed by reverse transcription-polymerase chain reaction (RT-PCR) using generic primers gL3637 (CCTTTAACAGTDGAAACAT) and gL4435c (CATDGCRCAAGARTGRTARACAGA) which were designed to amplify partial L segment encoding the RNA-dependent RNA polymerase (RdRP) of orthotospoviruses (Chu, et al. 2001). The expected ~800 bp DNA fragment was amplified from all three positive samples by RT-PCR. The amplified DNA was cloned and sequenced. BLAST search of the partial L RNA sequence (GenBank acc. nos. MW964378 to MW964380) revealed that they shared 86.2-97.4% nucleotide (nt) and 97.2-100% amino acid (aa) sequence identities with different isolates of CaCV available in GenBank with CaCV chili isolates (KU941834 to KU941836) from India sharing the highest aa identity of 100%. This confirmed the presence of CaCV in the symptomatic C. odorata plants. The 825 bp complete nucleocapsid protein (NP) of CaCV was also amplified from the samples using primers CaCV-F: ATGTCTAMCGTYAGGCAAC and CaCV-R: TYACACYTCWATAGAWGTACTAG) (Basavaraj et al. 2020), cloned, and sequenced to obtain complete S fragment-nucleocapsid protein (NP) with a size of 825 bp (MW964381 to MW964383). The pairwise comparisons of three fragments showed 85.1-98.3% nt and 87.6-99.6% aa sequence identities with different isolates of CaCV. Maximum-Likelihood phylogenetic trees inferred from the partial RdRP and complete NP aa sequences showed that the C. odorata isolates (CaCV-YN) clustered closely with CaCV tomato isolate from Taiwan and tomato (Yuxi-2013) isolate from China, respectively (Supplementary Figure 1). To our knowledge, this is the first time CaCV has been detected in C. odorata. This study will serve as an important reference for the study of host range of CaCV. Further studies will be required to determine whether thrips could transmit CaCV between C. odorata and other hosts of the virus.

Laterization and its control to gold occurrence in Laowanchang gold deposit, Guizhou Province, Southwest of China

Taking Sichuan, Guizhou and Yunnan provinces in Southwest China as the research object, this paper establishes a comprehensive measurement system for the degree of coordinated development of industrialization, agricultural modernization, urbanization, informatization and greenization, and uses the entropy evaluation method to assign the indicators and determine the types of coordination stages of each province. The results show that: (1) The comprehensive development index of the five modernization of the three provinces are rising continuously, the development of urbanization is the best, and the degree of informationization and greenization is significantly backward. (2) From 2010 to 2017, the overall coordination degree of the five modernization in the three provinces displays an upward trend, but there was a drop in Yunnan Province. In 2010, the coordination of the three provinces was in a state of imbalance, and the imbalance in Guizhou Province was the most serious. In 2017, Sichuan Province had reached a stage of high-quality coordination, while Yunnan and Guizhou provinces were in a good stage of coordination.

The Shazi large-scale anatase deposit in Qinglong County, Guizhou Province, has been discovered re- cently and now is under exploration. Investigations show that the orebodies mostly occur at the top of the karst un- conformity of the Middle Permian Maokou Formation strata and at the bottom of the Emeishan basalt. And the fol- lowing three prerequisites should be satisfied for the formation of the deposit: 1) there must be the material source of anatase; 2) there must be weekly alkaline media and low-tempeature and low-pressure conditions; 3) there was no high-temperature and high-pressure environment subsequently for the transformation of anatase into rutile. In the Emeishan basalt of western Guizhou, the element Ti mostly entered the silicon-oxygen tetrahedra of picrite in het- erovalent isomorphism (Ti 4+ +Al 3+ →Mg 2+ +Si 4+ ). When volcanic ejecta resultant from strong eruption of the Eme- ishan basalt magma fell into water, picrite was usually dissociated to chlorite. Thus, the element Ti 4+ in the picrite could be released from the silicon-oxygen tetrahedraa of picrite into water, and conbined with oxygen in the water to form TiO2. This paper has proved that this deposit, enriched in anatase, discovered recently at Shazi, Qinglong Country, Guizhou Province, is a residual-deluvial-type deposit. Its genesis can be explained as follows. Volcanic clastics formed at the early stage of strong eruption of the Emeishan basalt magma were chemically deposited to form anatase in the low-temperature, low-pressure and weekly alkaline waters in the karst depressions at the top of the Maokou Formation (limestone) strata. The anatase was then dissociated owing to weathering and leaching during the Quaternary and the anatase was further enriched to form the residual-deluvial-type anatase ore deposit.