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The cucurbit vegetable chieh-qua (Benincasa hispida var. chieh-qua How) is an important crop in South China and southeast Asian countries. Viral diseases cause substantial loss of chieh-qua yield. To identify the viruses that affect chieh-qua in China, ribosomal RNA-depleted total RNA sequencing was performed using chieh-qua leaf samples with typical viral symptoms. The virome of chieh-qua comprises four known viruses (melon yellow spot virus (MYSV), cucurbit chlorotic yellows virus (CCYV), papaya ringspot virus (PRSV) and watermelon silver mottle virus (WSMoV) and two novel viruses: cucurbit chlorotic virus (CuCV) in the genus Crinivirus and chieh-qua endornavirus (CqEV) in the genus Alphaendornavirus. The complete genomes of the two novel viruses in chieh-qua and three other isolates of CuCV in pumpkin, watermelon and cucumber were determined and the recombination signals of pumpkin and watermelon isolates of CuCV were detected. A reverse transcriptase PCR indicated that the dominant viruses of chieh-qua in Hainan are MYSV (66.67%) and CCYV (55.56%), followed by CuCV (27.41%), WSMoV (7.41%), cucumber mosaic virus (8.15%), zucchini yellow mosaic virus (6.67%), PRSV (6.67%) and CqEV (35.56%). Our findings support diagnostic and prevalence studies of viruses infecting chieh-qua in China, enabling sustainable control strategies for cucurbit viruses worldwide. [ABSTRACT FROM AUTHOR]

Areca palm, rubber tree, and coffee are always planted adjacent to each other or intercropped with each other, and Colletotrichum siamense was found to be the dominant species of Colletotrichum from these crops in Hainan, China. To study the population structure, pathogenicity, and fungicide sensitivity of C. siamense from these three crops in Hainan, isolates were first identified by their morphological characteristics and multilocus phylogeny. Then both permutational multivariate analysis of variance (PERMANOVA) and pairwise FST analysis showed that the original host and geographical origin (counties) had significant effects on genetic variation in the C. siamense population, explaining 5.6% and 16.3% of genetic variation, respectively. There were significant genetic differentiations between coffee and rubber tree populations but the genetic differentiation was small (FST = 0.04), while significant differentiations were observed among all populations from different counties except those between Haikou and Chengmai. Pathogenicity analysis using artificial inoculation showed that isolates had significantly higher disease incidence and diseased lesion diameter on wounded leaves than on nonwounded leaves. However, the original host of an isolate and geographical origin did not significantly affect the pathogenicity of the C. siamense populations from these three hosts. In vitro tests showed that there were no significant differences in the sensitivity of C. siamense populations to carbendazim, prochloraz, difenoconazole, and propiconazole from different original hosts and geographical origins. The present study suggests that host specialization of C. siamense has not occurred yet in coffee, rubber tree, and areca palm. [ABSTRACT FROM AUTHOR]

Black pepper (Piper nigrum L.) is one of the most ancient spice crops in the world. Hainan Province is the largest black pepper‐producing region in China. Viral disease is one of the main factors affecting black pepper. However, there are rarely studies on black pepper viral disease in China. We performed a survey in five major black pepper‐growing counties (Qionghai, Wenchang, Wanning, Haikou and Ding'an) of Hainan Province and collected 589 samples, with virus‐like symptoms from 2014 to 2019. Various viruses were identified including cucumber mosaic virus (CMV), piper yellow mottle virus (PYMoV), Piper DNA virus 1 (PDV‐1) and Piper DNA virus 2 (PDV‐2) using small RNA deep sequencing. The RT‐PCR screening of field samples indicated three types of infection: CMV alone (5.56%), PYMoV + PDV‐1 + PDV‐2 (25.00%) and PYMoV + CMV +PDV‐1 + PDV‐2 (69.44%). The data suggested that PYMoV might be the main pathogen causing virus diseases of black pepper. Two complete genome sequences of PYMoV were obtained. Phylogenetic analysis showed that PYMoV isolates from China were placed in a branch distinct from PYMoV isolates from India. To our knowledge, this is the first report of PYMoV, PDV‐1 and PDV‐2 naturally infecting black pepper, and the first genome sequence of PYMoV in China. [ABSTRACT FROM AUTHOR]

Powdery mildew is an important disease of rubber trees worldwide. To assess the effects of temperature and leaf age on conidial germination and disease development, conidia were inoculated onto rubber tree seedlings with leaves at three phenological stages (copper bronze, colour‐changing, and light green) and then incubated at six constant temperatures (10, 15, 20, 25, 30, and 35°C). Leaf age did not affect conidial germination (p =.296) whilst temperature did (p <.0001), although conidia were able to germinate at all tested temperatures. The estimated optimal temperature for conidial germination was 23.2°C. Leaf age, temperature, and their interactions had significant effects on conidial infection and hypha number (p <.0001). At 10 and 35°C, more than 2 and 4 days were needed for infection to complete, respectively, compared to <2, 1, 0.5, and 0.5 days for 15, 20, 25, and 35°C, respectively. Sporulation and mildew symptoms were only observed on those inoculated leaves of all stages at 20 and 25°C, and at the copper bronze stage only at 15°C. The latent period on the copper bronze leaves at 15°C was longer (9 days) than at 20 and 25°C (4 days). The latent period at 20 and 25°C increased from 4 to 7 days as the leaf development stage increased from copper bronze to light green. Therefore, temperature affected germination and postgermination growth of rubber tree powdery mildew, whereas leaf age primarily affected postgermination growth of the pathogen. [ABSTRACT FROM AUTHOR]

In July 2008, Stylosanthes guianensis with witches'-broom symptoms and Stylosanthes capitata with yellows symptoms were observed in Hainan Province, China. From the symptomatic S. guianensis, the phytoplasma-specific 16S rRNA gene fragment of 1.4 kb was amplified by a direct PCR, and a fragment of 16S rRNA gene of 1.2 kb was obtained from the symptomatic Stylosanthes capitata by a nested PCR. Sequencing results and NCBI BLASTn analysis showed that the two phytoplasmas belonged to group 16SrII. Polygenetic analysis and virtual RFLP classified the two phytoplasmas into subgroup 16SrII-A. [ABSTRACT FROM AUTHOR]

Phytoplasmas are a group of plant prokaryotic pathogens distributed worldwide. To comprehensively reveal the diversity of the pathogens and the diseases they cause on Hainan, a tropical island with abundant biodiversity in China, a survey of phytoplasmal diseases was performed from 2009 to 2022. Herein, molecular identification and genetic analysis were conducted based on the conserved genes of phytoplasmas. The results indicated that phytoplasmas could be detected in 138 samples from 18 host plants among 215 samples suspected to be infected by the pathogens. The phytoplasma strains from 27 diseased samples of 4 host plants belonged to the 16SrI group and the strains from 111 samples of 14 hosts belonged to the 16SrII group. Among them, 12 plants, including important tropical cash crops such as Phoenix dactylifera , cassava, sugarcane, and Piper nigrum , were first identified as hosts of phytoplasmas on Hainan Island. Based on BLAST and i PhyClassifier analyses, seven novel 16Sr subgroups were proposed to describe the relevant phytoplasma strains, comprising the 16SrI-AP, 16SrI-AQ, and 16SrI-AR subgroups within the 16SrI group and the 16SrII-Y, 16SrII-Z, 16SrII-AB, and 16SrII-AC subgroups within the 16SrII group. Genetic variation and phylogenetic analysis indicated that the phytoplasma strains identified in this study and those reported previously on Hainan Island mainly belong to four 16Sr groups (including I, II, V, and XXXII) and could infect 44 host plants, among which the 16SrI and 16SrII groups were the prevalent 16Sr groups associated with 43 host plant species. The diversity of host plants infected by the phytoplasmas made it difficult to monitor and control their related diseases. Therefore, strengthening inspection and quarantine during the introduction and transit of the related phytoplasmal host crops would effectively curb the spread and prevalence of the phytoplasmas and their related lethal diseases., Competing Interests: The author(s) declare no conflict of interest.

Colletotrichum siamense was demonstrated as the dominant species among Colletotrichum spp. that infected rubber tree, areca palm, and coffee in Hainan, China. However, the extent of genetic differentiation within the species C. siamense in relation to geographical regions and host species is not known. In this study, 112 C. siamense isolates were genotyped with 12 microsatellite markers. In total, there were 99 multilocus genotypes. Results from permutational multivariate analysis of variance and analysis of molecular variance indicated that there was no significant genetic differentiation between fungal populations with respect to host, location (county), and year. Discriminant analysis of principal components and STRUCTURE analysis showed that C. siamense isolates grouped into three clusters; further analysis confirmed that there were significant ( P < 0.001) genetic differences among the three clusters. However, each cluster had isolates from different hosts, counties, or years, supporting the lack of genetic differentiation with respect to host, county, and year. Statistical analyses of allelic associations indicated some evidence for recombination within the populations defined on the basis of host or county. The present findings provide insights into the genetic structure of C. siamense on the three perennial host species in Hainan and suggest that the disease on these three crops can be effectively considered as one disease and, hence, needs to be controlled simultaneously in mixed plantations.

Genus Colletotrichum is one of the most important genera of plant-pathogenic fungi affecting numerous species, particularly tropical and subtropical crops and fruit trees. In this study, 43 Colletotrichum strains were isolated from areca palm leaves with anthracnose symptoms in 11 areca palm plantations in eight counties of Hainan, China. Based on the morphology, phylogenetic analysis of six loci (internal transcribed spacer, actin, chitin synthase 1, glyceraldehyde-3-phosphate dehydrogenase, β-tubulin, and mating type locus MAT1-2), and pathogenicity tests, eight Colletotrichum species were distinguished, comprising five previously known species ( C. cordylinicola , C. fructicola , C. gloeosporioides , C. siamense , and C. tropicale ), one unidentified Colletotrichum species, a new species ( C. arecicola ) in the gloeosporioides species complex, and C. karstii in the boninense species complex. C. siamense was the most common species found in areca palm in Hainan, followed by C. arecicola . Pathogenicity tests showed that all eight species could cause anthracnose symptoms on areca palm leaves using a wound inoculation method and that the isolates from the gloeosporioides species complex caused larger lesions than the isolates from the boninense species complex. Further research is needed to understand the epidemiology of these pathogenic species on areca palm in order to develop management strategies.

Colletotrichum gloeosporioides and C. acutatum have been reported to be causal agents of anthracnose disease of rubber tree. Recent investigations have shown that both C. gloeosporioides and C. acutatum are species complexes. The identities of Colletotrichum species causing anthracnose disease of rubber tree in Hainan, China, are unknown. In this study, 106 isolates obtained from rubber tree with symptoms of anthracnose were collected from 12 counties of Hainan and identified at the species complex level based on the ITS sequences and colony morphologies. Seventy-four isolates were identified as C. gloeosporioides species complex and the other 32 isolates as C. acutatum species complex. Forty-two isolates were selected for further multilocus phylogenetic analyses in order to identify the isolates to the species level. Twenty-six isolates from the C. gloeosporioides species complex were characterized for partial sequences of seven gene regions (ACT, TUB2, CHS-1, GAPDH, ITS, ApMat, and GS), and the other 16 isolates from the C. acutatum species complex for five gene regions (ACT, TUB2, CHS-1, GAPDH, and ITS). Three species were identified: C. siamense and C. fructicola from the C. gloeosporioides species complex, and a new species C. wanningense from the C. acutatum species complex. Artificial inoculation of rubber tree leaves confirmed the pathogenicity of the three species. The present study improves the understanding of species causing anthracnose on rubber tree and provides useful information for the effective control of the disease.

Colletotrichum gloeosporioides and C. acutatum species complexes are causal agents of Colletotrichum leaf disease (CLD) of rubber trees worldwide. To determine the geographic distribution of Colletotrichum species complexes associated with CLD of rubber trees in Hainan, China, and their sensitivity to fungicides used in the region, a total of 275 Colletotrichum isolates were collected from 52 rubber tree plantations in 11 counties. These isolates were identified based jointly on morphological characteristics and PCR-based methodology. Of these isolates, 78 and 22% belonged to the C. gloeosporioides species complex (CGSC) and the C. acutatum complex (CASC), respectively. The incidence of CGSC isolates was greater than the CASC in all counties sampled. The incidence of CASC isolates appeared to be lower in the western and central south of Hainan than in other regions. There was no association in their presence at a given plantation between the two species complexes. The in vitro sensitivity of these two species complexes to carbendazim, chlorothalonil, and four demethylation inhibitor (DMI) fungicides (difenoconazole, propiconazole, myclobutanil, and prochloraz) was determined. Carbendazim was effective against CGSC but not against CASC with mean ED 50 values of 0.176 and 2.182 µg/ml, respectively. CASC isolates were more sensitive to difenoconazole, propiconazole, and myclobutanil (mean ED 50 values of 0.177, 0.129, and 1.424 µg/ml, respectively) than CGSC isolates (mean ED 50 values of 0.710, 0.348, and 3.496 µg/ml, respectively). Mean ED 50 values of CGSC against chlorothalonil and prochloraz were 173.341 and 0.035 µg/ml, respectively; corresponding values for CASC were 151.441 and 0.040 µg/ml. These results suggest that prochloraz, propiconazole, and difenoconazole are effective against both species complexes.

Papaya is an important fruit crop cultivated in tropical and subtropical regions. Papaya little leaf (PLL) disease was observed in China. The phytoplasma 16S rRNA gene was detected from symptomatic papaya trees via PCR using phytoplasma universal primers P1/P7 followed by R16F2n/R16R2. No amplification products were obtained from templates of asymptomatic papaya trees. These results indicated a direct association between phytoplasma infection and PLL disease. Comparative and phylogenetic analyses of 16S rRNA gene sequences indicated that the papaya-infecting phytoplasmas under study belonged to the peanut witches' broom phytoplasma group (16SrII). Genotyping through use of computer-simulated RFLP analysis of 16S rRNA genes and coefficients of RFLP pattern similarities (0.97) reveal that the PLL phytoplasma was placed in a new subgroup. In this article, we describe the molecular characterization of a new phytoplasma associated with PLL disease and propose that the PLL phytoplasma be considered as a novel subgroup, 16SrII-U.