Sinensis Plants Research Articles

Effectiveness of A Combination of Kitolod Leaves (Laurentia longiflora), Papaya Leaves (Carica papaya), and Lemongrass (Cymbopogon citratus) as Candidates for Biocides on Bactrocera Pests in Citrus sinensis Plants

Bactrocera is a pest that is detrimental to citrus farming. Bactrocera attacks cause developmental disorders in citrus fruit, including changes in fruit color (black spots appear from Bactrocera infection) and premature rot. Bactrocera also has a symbiotic relationship with Liberibacter asiaticus which causes Citrus Vein Phloem Degeneration (CVPD) disease which attacks phloem tissue in citrus plants. Bactrocera pest control was carried out by combining plants that have the potential to be used as biocides, namely kitolod, papaya, and lemongrass plants. This combination was used to switch the habits of fruit farmers who previously used synthetic pesticides to control pests. Natural biocides are expected to have the same effectiveness as synthetic pesticides, but leave no residue on cultivated plants. A combination biocide from the three plants was made using the maceration method to obtain an infusion from each plant. Before being tested on Citrus plantations, an assay was carried out using an olfactometer to determine the effectiveness of the biocides. The research revealed that the combination of these three plants has the potential to be used as a natural biocide without leaving residueand causing damage or other developmental disorders to citrus plants.

First Report of Didymella pomorum Causing Leaf Blight on Angelica sinensis in China.

Angelica sinensis (Oliv.) Diels, is a perennial herbaceous plant of the Umbelliferae family. It has a long history of cultivation and is highly valued as a traditional Chinese medicine in China (Zhang et al. 2012). In September 2023, leaf blight on A. sinensis with an average disease incidence of 56% was recorded in an approximately 6.7-ha production field in Lijiang, Yunnan province, China (26.8215°N, 100.2369°E). At first, small, chlorotic lesions appeared on the leaves. They subsequently increased in density and gradually merged, causing leaves to yellow and wither. Ultimately the blight casused death of the entire foliage. In order to identify the causal agent, cross-sectional segments (5×5 mm2) were cut from the edge of leaf lesions, surface disinfected with a 1% sodium hypochlorite solution for 3 min and rinsed three times with sterile distilled water. They were subsequently placed on potato dextrose agar (PDA) plates and incubated for 3 days under a 12-h photoperiod at 28℃. A total of ten isolates with similar morphological characteristics were obtained by single spore isolation. After 10 days of incubation on PDA, the colony morphology of these isolates was characterized by a brownish central area with a white edge. Aged colonies became wrinkled in the center of the colony. Conidia (n = 30) were elliptical and brown, with a size range of 4.11 to 6.55 μm (average 5.37±0.74 μm) × 3.17 to 4.62 μm (average 3.92±0.43 μm). Chlamydospores (n = 30) formed chains in series, spherical or elliptical in shape, ranging from yellow-brown to dark brown, with a size range of 12.30 to 13.70 μm (average 12.98±0.46 μm) × 4.20 to 5.30 μm (average 4.63±0.26 μm). The nuclear ribosomal internal transcribed spacer region (ITS), the second largest subunit of RNA polymerase II (RPB2), and the 28S nuclear ribosomal large subunit rRNA (LSU) region of two isolates were amplified with the primer pairs ITS1/ITS4 (White et al. 1990), fRPB2-5F/fRPB2-7cR (Liu et al. 1999), and LR0R/LR5 (Schoch et al. 2012), respectively. These amplicons were sequenced bidirectionally and assembled. The two isolates produced the same nucleotide sequences, and the sequences of a representative isolate (AsDp1) were deposited in GenBank. BLASTn analyses showed that the ITS (PP510616), RPB2 (PP526010), and LSU (PP550143) sequences of isolate AsDp1 were 100%, 99.66%, and 100% identical with those of Didymella pomorum ex-type isolate CBS 354.52 (MH857081, KT389616, and MH868616), respectively. A phylogenetic tree was constructed based on the ITS, RPB2, and LSU concatenated nucleotide sequences using the maximum likelihood method in MEGAX. Isolate AsDp1 was clustered with four D. pomorum isolates. According to the morphological and nucleotide sequences analyses, isolate AsDp1 was identified as D. pomorum (Chen et al. 2015). To determine pathogenicity, 1-year-old A. sinensis plants (approximately 20 cm tall) grown in 7-liter pots filled with sterilized field soil were sprayed until runoff with a 1×106 conidia/ml suspension of isolate AsDp1 onto the foliage, while control plants were sprayed with sterile water. All plants were cultivated under a 12-h photoperiod at 25℃. The pathogenicity tests were performed in triplicate with ten plants in each treatment. After fifteen days, numerous chlorotic lesions appeared on the leaves of all inoculated plants. The symptoms were similar to those found on naturally infected plants in the field, while the control plants remained asymptomatic. Subsequently, D. pomorum was reisolated from the diseased leaves, and the identity was confirmed based on its ITS sequence and morphological characteristics. D. pomorum causing stem canker on Rosa spp. was reported in Canada (Ilyukhin 2022). To our knowledge, this is the first report of D. pomorum causing leaf blight on A. sinensis in China. This etiological finding will potentially pave the way for the development of control strategies of this disease.

Fertilizer Effects on the Nitrogen Isotope Composition of Soil and Different Leaf Locations of Potted Camellia sinensis over a Growing Season.

The nitrogen-stable isotopes of plants can be used to verify the source of fertilizers, but the fertilizer uptake patterns in tea (Camellia sinensis) plants are unclear. In this study, potted tea plants were treated with three types of organic fertilizers (OFs), urea, and a control. The tea leaves were sampled over seven months from the top, middle, and base of the plants and analyzed for the δ15N and nitrogen content, along with the corresponding soil samples. The top tea leaves treated with the rapeseed cake OF had the highest δ15N values (up to 6.6‱), followed by the chicken manure, the cow manure, the control, and the urea fertilizer (6.5‱, 4.1‱, 2.2‱, and 0.6‱, respectively). The soil treated with cow manure had the highest δ15N values (6.0‱), followed by the chicken manure, rapeseed cake, control, and urea fertilizer (4.8‱, 4.0‱, 2.5‱, and 1.9‱, respectively). The tea leaves fertilized with rapeseed cake showed only slight δ15N value changes in autumn but increased significantly in early spring and then decreased in late spring, consistent with the delivery of a slow-release fertilizer. Meanwhile, the δ15N values of the top, middle, and basal leaves from the tea plants treated with the rapeseed cake treatment were consistently higher in early spring and lower in autumn and late spring, respectively. The urea and control samples had lower tea leaf δ15N values than the rapeseed cake-treated tea and showed a generalized decrease in the tea leaf δ15N values over time. The results clarify the temporal nitrogen patterns and isotope compositions of tea leaves treated with different fertilizer types and ensure that the δ15N tea leaf values can be used to authenticate the organic fertilizer methods across different harvest periods and leaf locations. The present results based on a pot experiment require further exploration in open agricultural soils in terms of the various potential fertilizer effects on the different variations of nitrogen isotope ratios in tea plants.

The Circadian Clock Gene PHYTOCLOCK1 Mediates the Diurnal Emission of the Anti-Insect Volatile Benzyl Nitrile from Damaged Tea (Camellia sinensis) Plants.

Benzyl nitrile from tea plants attacked by various pests displays a diurnal pattern, which may be closely regulated by the endogenous circadian clock. However, the molecular mechanism by the circadian clock of tea plants that regulates the biosynthesis and release of volatiles remains unclear. In this study, the circadian clock gene CsPCL1 can activate both the expression of the benzyl nitrile biosynthesis-related gene CsCYP79 and the jasmonic acid signaling-related transcription factor CsMYC2 involved in upregulating CsCYP79 gene, thereby resulting in the accumulation and release of benzyl nitrile. Therefore, the anti-insect function of benzyl nitrile was explored in the laboratory. The application of slow-release beads of benzyl nitrile in tea plantations significantly reduced the number of tea geometrids and had positive effects on the yield of fresh tea leaves. These findings reveal the potential utility of herbivore-induced plant volatiles for the green control of pests in tea plantations.

Integrated transcriptomic and metabolomic analysis reveals the regulation mechanism of early bolting and flowering in two cultivars of Angelica sinensis

The root of Angelica sinensis is utilized in Traditional Chinese medicine to enhance blood replenishment and facilitate blood circulation. The early bolting and flowering (EBF) of A. sinensis, however, compromises the quality of the roots and restricts the yield of medicinal substances. The study was conducted to compare the transcriptomic and metabolomic profiles between EBF plants and normal plants of two cultivars of A. sinensis, followed by validation of the transcriptome results using qRT-PCR. There were 3677 DEGs in EBF plants compared to normal plants of cultivar 2 (Mingui No.2), and cultivar 4 (Mingui No.4) was 3354. The main differential metabolites in the EBF and normal plants were phenolic acids, flavonoids, lignans, and coumarins. The analysis of 5 EBF-related pathways revealed 28 genes exhibiting differential expression and 5 metabolites showing differential accumulation. The expression of the Lhcb5, Lhcb2, Lhcb6, Lhcb1, Lhca4, ATPG1, EGLC, CELB, AMY, glgA, CYCD3, SnRK2, PYL, AHK2, AUX1, BSK, FabI/K, ACACA and FabV decreased and the expression of the PsbR, PsbA, LHY, FT, CO, malQ, HK, GPI and DELLA increased in EBF plants. In addition, the Abscisic acid, d-Glucose-6P, α-d-Glucose-1P, NADP+, and ADP were more significantly enriched in EBF plants. The findings offer novel perspectives on the EBF mechanisms in A. sinensis and other medicinal plants of the Apiaceae family.

Multi-Omics Research Accelerates the Clarification of the Formation Mechanism and the Influence of Leaf Color Variation in Tea (Camellia sinensis) Plants

Tea is a popular beverage with characteristic functional and flavor qualities, known to be rich in bioactive metabolites such as tea polyphenols and theanine. Recently, tea varieties with variations in leaf color have been widely used in agriculture production due to their potential advantages in terms of tea quality. Numerous studies have used genome, transcriptome, metabolome, proteome, and lipidome methods to uncover the causes of leaf color variations and investigate their impacts on the accumulation of crucial bioactive metabolites in tea plants. Through a comprehensive review of various omics investigations, we note that decreased expression levels of critical genes in the biosynthesis of chlorophyll and carotenoids, activated chlorophyll degradation, and an impaired photosynthetic chain function are related to the chlorina phenotype in tea plants. For purple-leaf tea, increased expression levels of late biosynthetic genes in the flavonoid synthesis pathway and anthocyanin transport genes are the major and common causes of purple coloration. We have also summarized the influence of leaf color variation on amino acid, polyphenol, and lipid contents and put forward possible causes of these metabolic changes. Finally, this review further proposes the research demands in this field in the future.

Insights into the functional role of Actinomycetia in promoting plant growth and biocontrol in tea (Camellia sinensis) plants.

Tea, a highly aromatic and globally consumed beverage, is derived from the aqueous infusion of dried leaves of Camellia sinensis (L.) O. Kuntze. Northeast India, encompassing an expansive geographical area between 24° and 27°N latitude and 88° and 95°E longitude, is a significant tea-producing region covering approximately 312,210 hectares. Despite its prominence, this region faces persistent challenges owing to a conducive climate that harbors the prevalence of pests, fungal pathogens, and weeds, necessitating agrochemicals. Helopeltis theivora, Oligonychus coffeae, and Biston suppressaria are prominent among the tea pests in this region. Concurrently, tea plants encounter fungal infections such as blister blight, brown root rot, and Fusarium dieback. The growing demand for safer tea production and the need to reduce pesticide and fertilizer usage has spurred interest in exploring biological control methods. This review focuses on Actinomycetia, which potentially safeguards plants from diseases and pest infestations by producing many bioactive substances. Actinomycetia, which resides in the tea rhizosphere and internal plant tissues, can produce antagonistic secondary metabolites and extracellular enzymes while promoting plant growth. Harnessing the biocontrol potential of Actinomycetia offers a promising solution to enhance tea production, while minimizing reliance on harmful agrochemicals, contributing to a more environmentally conscious and economically viable tea cultivation system.

Analysis of Carica papaya Informs Lineage-Specific Evolution of the Aquaporin (AQP) Family in Brassicales.

Aquaporins (AQPs), a type of intrinsic membrane proteins that transport water and small solutes across biological membranes, play crucial roles in plant growth and development. This study presents a first genome-wide identification and comparative analysis of the AQP gene family in papaya (Carica papaya L.), an economically and nutritionally important fruit tree of tropical and subtropical regions. A total of 29 CpAQP genes were identified, which represent five subfamilies, i.e., nine plasma intrinsic membrane proteins (PIPs), eight tonoplast intrinsic proteins (TIPs), seven NOD26-like intrinsic proteins (NIPs), two X intrinsic proteins (XIPs), and three small basic intrinsic proteins (SIPs). Although the family is smaller than the 35 members reported in Arabidopsis, it is highly diverse, and the presence of CpXIP genes as well as orthologs in Moringa oleifera and Bretschneidera sinensis implies that the complete loss of the XIP subfamily in Arabidopsis is lineage-specific, sometime after its split with papaya but before Brassicaceae-Cleomaceae divergence. Reciprocal best hit-based sequence comparison of 530 AQPs and synteny analyses revealed that CpAQP genes belong to 29 out of 61 identified orthogroups, and lineage-specific evolution was frequently observed in Brassicales. Significantly, the well-characterized NIP3 group was completely lost; lineage-specific loss of the NIP8 group in Brassicaceae occurred sometime before the divergence with Cleomaceae, and lineage-specific loss of NIP2 and SIP3 groups in Brassicaceae occurred sometime after the split with Cleomaceae. In contrast to a predominant role of recent whole-genome duplications (WGDs) on the family expansion in B. sinensis, Tarenaya hassleriana, and Brassicaceae plants, no recent AQP repeats were identified in papaya, and ancient WGD repeats are mainly confined to the PIP subfamily. Subfamily even group-specific evolution was uncovered via comparing exon-intron structures, conserved motifs, the aromatic/arginine selectivity filter, and gene expression profiles. Moreover, down-regulation during fruit ripening and expression divergence of duplicated CpAQP genes were frequently observed in papaya. These findings will not only improve our knowledge on lineage-specific family evolution in Brassicales, but also provide valuable information for further studies of AQP genes in papaya and species beyond.

Eco-friendly Production of Bacterial Cellulose with Komagataeibacter intermedius Strain by Using Jasminum sambac and Camellia sinensis Plants

Eco-friendly Production of Bacterial Cellulose with Komagataeibacter intermedius Strain by Using Jasminum sambac and Camellia sinensis Plants

First molecular analysis of wisteria vein mosaic virus in Italy: eight new variants reported in Wisteria sinensis

Wisteria vein mosaic virus (WVMV) is a potyvirus infecting Wisteria spp. worldwide, making these largely used ornamental plants unattractive and even unsalable. In 2021, nine Wisteria sinensis plants in Sarzana (Liguria, Italy) showing WVMV symptoms like vein mosaic with irregular patterns, mottling, deformation and twisting margin on leaves were reported. This work describes research on symptomatic and asymptomatic leaves to confirm the identity of the virus infection. All plants tested resulted positive to WVMV. Sequencing of NIb/CP genomic region and comparison on GenBank revealed the presence of eight new genetic variants named Sar 5–12. The eight nucleotide sequences alignments revealed identity ranging between 87.13 and 99.85%. Negative selection (dN/dS < 1) was detected suggesting well adaptation in the area here examined and stability in population. Through a phylogenetic tree, WVMV isolates were grouped in four clades with high bootstrap values, two of which included the eight Italian variants here identified. Since the late 1950 and 1960s, the present study represents the first additional report of WVMV in Italy, as well as its first molecular characterization ever. Further research is required to strengthen our understanding of the movement and transmission of WVMV. Examining how virus infected plants can disseminate this pathogen could help forestall potential risks that neighbouring species may face in their environments.

Transmission of isolates of Citrus tristeza virus by single aphids (Toxoptera citricida)

Citrus tristeza virus (CTV) can be transmitted by several aphid species in a semi-persistent mode, with Toxoptera citricida being the most efficient vector. In Brazil, mild CTV isolates are used for pre-immunization of citrus trees against severe isolates. We aimed to determine the capacity and efficiency of T. citricida in separating the viral complex into haplotypes from three well-characterized CTV isolates (PIAC, CS1, and Forte Rolândia). Single-aphid transmission assays were conducted to determine CTV transmission efficiency. The results showed that T. citricida transmitted only haplotypes from the PIAC and CS1 isolates, with efficiencies of 8 and 4%, respectively. Both isolates caused mild CTV symptoms in Brazil. However, isolate Forte Rolândia, which causes severe symptoms in citrus trees, was not transmitted by T. citricida. The detection of CTV haplotypes from PIAC and CS1 isolates in sweet orange (Citrus sinensis) plants after a single aphid transmission occurred at different time points. The first haplotype observed was from PIAC 150 days after citrus was challenged by a single aphid, followed by CS1 at day 210 after transmission. In addition, differences in the single-strand conformation polymorphism patterns between the CTV isolates and CTV haplotypes were determined, suggesting that an aphid can acquire and transmit only one CTV haplotype to citrus plants during phloem feeding. The study of the mechanism of transmission by the vector can increase our knowledge of the interactions among hosts, vectors, and pathogens, which are often neglected.

Light induces an increasing release of benzyl nitrile against diurnal herbivore Ectropis grisescens Warren attack in tea (Camellia sinensis) plants.

Herbivore-induced plant volatiles (HIPVs) are critical compounds that directly or indirectly regulate the tritrophic interactions among herbivores, natural enemies and plants. The synthesis and release of HIPVs are regulated by many biotic and abiotic factors. However, the mechanism by which multiple factors synergistically affect HIPVs release remains unclear. Tea plant (Camellia sinensis) is the object of this study because of its rich and varied volatile metabolites. In this study, benzyl nitrile was released from herbivore-attacked tea plants more in the daytime than at night, which was consistent with the feeding behaviour of tea geometrid (Ectropis grisescens Warren) larvae. The Y-tube olfactometer assay and insect resistance analysis revealed that benzyl nitrile can repel tea geometrid larvae and inhibit their growth. On the basis of enzyme activities in transiently transformed Nicotiana benthamiana plants, CsCYP79 was identified as a crucial regulator in the benzyl nitrile biosynthetic pathway. Light signalling-related transcription factor CsPIF1-like and the jasmonic acid (JA) signalling-related transcription factor CsMYC2 serve as the activator of CsCYP79 under light and damage conditions. Our study revealed that light (abiotic factor) and herbivore-induced damage (biotic stress) synergistically regulate the synthesis and release of benzyl nitrile to protect plants from diurnal herbivorous tea geometrid larvae.

First report of Colletotrichum siamense causing leaf spot on Celtis sinensis in China.

Celtis sinensis Pers., a deciduous tree, is widely cultivated in China for its ornamental value (Yang et al. 2022). In July 2020, leaf spot symptoms were observed on Ce. sinensis plants at the campus of Jiangxi Agricultural University (28°45'56″N, 115°50'21″E) in Nanchang city, Jiangxi province, China. The disease incidence was estimated to be above 15%. The early symptoms were small spots on the edge or tip of the leaves. The spots gradually expanded and became grayish brown, eventually developing large irregular lesions. Leaf pieces (5 × 5 mm) from the lesion borders were surfaced and sterilized in 70% ethanol for 30 s, followed by 2% NaOCl for 1 min, and then rinsed three times with sterile water. Tissues were placed on potato dextrose agar (PDA) and incubated at 25°C. Pure cultures were obtained by monosporic isolation, and the representative isolates, JPS-4, JPS-9, and JPS-13 were used for morphological studies and phylogenetic analyses. Colonies on PDA medium of the three isolates were white to gray with cottony mycelia and grayish-white on the undersides of the culture. Conidia were single-celled, straight, hyaline, cylindrical, clavate, and measured 14.3-18.2 ×4.3-6.9 µm (15.8 ± 1.1 × 5.3 ± 0.4 µm, n = 100). Appressoria were brown to dark brown, ovoid to clavate, slightly irregular to irregular, and ranged from 5.6-9.4 × 4.5-6.8 µm (7.6 ± 0.1 × 5.4 ± 0.2 µm, n=100). Morphological features were similar to Colletotrichum gloeosporioides species complex (Weir et al. 2012). The internal transcribed spacer (ITS) regions, actin (ACT), calmodulin (CAL), β-tubulin 2 (TUB2), chitin synthase (CHS-1), and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) were amplified using primers ITS1/ITS4, ACT-512F/ACT-783R, CL1/CL2, T1/Bt2b, CHS-79F/CHS-354R and GDF/GDR (Weir et al. 2012), respectively. All sequences were deposited into GenBank (ITS, ON207804 - ON207806; ACT, ON239113 - ON239115; GAPDH, ON239122 - ON239124; TUB2, ON239125 - ON239127; CHS-1, ON239119 - ON239121; CAL, ON239116 - ON239118). A maximum likelihood and Bayesian posterior probability analyses using IQtree v. 1.6.8 and Mr. Bayes v. 3.2.6 with the concatenated sequences placed JPS-4, JPS-9, and JPS-13 in the clade of C. siamense. Based on the multi-locus phylogeny and morphology, three isolates were identified as C. siamense. To confirm pathogenicity, nine 6-year-old Ce. sinensis plants (three leaves each, n=27) grown outdoors were pin-pricked with a sterile needle and inoculated with 100 μL spore suspension per leaf (106 conidia per mL). Another 27 healthy leaves were inoculated with sterile water as the control. All the inoculated leaves were covered with plastic bags to keep a high-humidity environment for 2 days. The experiment was repeated three times. All the inoculated leaves showed similar symptoms to those observed in the field, whereas control leaves were asymptomatic for 8 days. Colletotrichum siamense was reisolated from the lesions, whereas no fungus was isolated from control leaves. Colletotrichum siamense can cause leaf diseases in a variety of hosts, including Allamanda cathartica (Huang et al. 2022), Osmanthus fragrans (Liu et al. 2022), and Crinum asiaticum (Khoo et al. 2022). To our knowledge, this is the first report of C. siamense causing leaf spots on Ce. sinensis worldwide. This work provided crucial information for epidemiologic studies and appropriate control strategies for this newly emerging disease.

First Report of Colletotrichum gloeosporioides Causing Anthracnose on Wisteria sinensis in China.

Wisteria (Wisteria sinensis) is a well-known ornamental plant for environmental protection in the garden, which also has a high value for medicinal use. In December 2021, leaf spots were observed on W. sinensis plants growing on the campus of Jiangxi Agricultural University in Nanchang, Jiangxi Province (28.45° N, 115.49° E), with a incidence rate of 40% plants were infested (n = 100 investigated plants). Initially leaf spots were small and pale brown (Approx. 2 mm in diameter), which gradually expanded into round or irregular dark brown spots as disease progressed, and lesions developed greyish-white necrotic tissues in the center at later stages, eventually causing the leaves to rot. To isolate the pathogen, tissues (5 × 5 mm) at the margin of lesions were cut from ten symptomatic leaves, surface disinfected with 75% ethanol for 30 s followed by 2% sodium chloride (NaClO) for 1 min, rinsed three times with sterile distilled water, and the dried tissues were cultured on potato dextrose agar (PDA) at 28 ± 1℃ in darkness for 3 days. After culture purification, five isolates were obtained and the representative single spore isolate (ZTTJ1) was used for subsequent identification tests. After 10 days of incubation on PDA medium, colonies had dense aerial mycelium with a gray center and dark gray-green mycelium outward, with orange-red conidial masses distributed in a ring on the surface. The underside of the colonies was light gray to dark gray. Conidia were cylindrical, with ends obtuse-rounded, 11.83 to 20.74 × 3.34 to 5.33 μm (av=16.11 μm × 4.26 μm, n = 50) in size. These morphological characteristics were consistent with Colletotrichum gloeosporioides (Shi et al, 2019). Six conserved regions of isolate (ZTTJ1), internal transcribed spacer (ITS), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), calmodulin (CAL), β-tublin (TUB), actin (ACT), and chitin synthase 1 (CHS1) gene regions were amplified using ITS1/ITS4 (Gardes et al, 1993), GDF/GDR (Templeton et al, 1992), CL1C/CL2C (Li et al, 2018), Bt2a/Bt2b (Prihastuti et al, 2009), ACT-512F/ACT-783R and CHS-79F/CHS-345R (Carbone et al, 1999) primers, respectively. Using BLAST, ITS, GAPDH, CAL, TUB, ACT, and CHS1 gene sequences (GenBank Accession No. OP703312, OP713773, OP713775, OP713776, OP713772, OP713774, respectively) were over 99% identical to C. gloeosporioides (GenBank Accession No. MK967281, MH594288, MT449307, MN624110, MN107239 and MN908602, respectively). A maximum likelihood (ML) phylogenetic analysis based on ITS-ACT-GAPDH-CHS1-CAL-TUB2 sequences using MEGA7.0, placed isolate (ZTTJ1) within C. gloeosporioides. To complete Koch's postulates, 10 μL spore suspension (1.0 × 106 conidia/mL) of ZTTJ1 (7-day-old culture on PDA medium) was dropped onto 10 leaves wounded with a sterilized needle and 10 non-wounded leaves, respectively. Ten wounded leaves were inoculated with sterile water as controls. All leaves were incubated at 28 ± 1 °C and 90 % relative humidity (12 h/12 h light/dark). After 7 days, all wounded leaves inoculated with C. gloeosporioides developed symptoms as previously observed, while the control and non-wounded leaves remained healthy. The fungus re-isolated from the inoculated leaves were identified as C. gloeosporioides by morphological and molecular identification; the pathogen causing disease in W. sinensis was determined to be C. gloeosporioides. To our knowledge, this is the first report of C. gloeosporioides causing anthracnose on W. sinensis in China. This work has identified the pathogenic species of the disease, which helps to take targeted measures to control its spread, providing a basis for the prevention and treatment of the disease.

‘Candidatus Liberibacter asiaticus’ secretory protein SDE3 inhibits host autophagy to promote Huanglongbing disease in citrus

ABSTRACT Antimicrobial acroautophagy/autophagy plays a vital role in degrading intracellular pathogens or microbial molecules in host-microbe interactions. However, microbes evolved various mechanisms to hijack or modulate autophagy to escape elimination. Vector-transmitted phloem-limited bacteria, Candidatus Liberibacter (Ca. Liberibacter) species, cause Huanglongbing (HLB), one of the most catastrophic citrus diseases worldwide, yet contributions of autophagy to HLB disease proliferation remain poorly defined. Here, we report the identification of a virulence effector in “Ca. Liberibacter asiaticus” (Las), SDE3, which is highly conserved among the “Ca. Liberibacter”. SDE3 expression not only promotes the disease development of HLB and canker in sweet orange (Citrus sinensis) plants but also facilitates Phytophthora and viral infections in Arabidopsis, and Nicotiana benthamiana (N. benthamiana). SDE3 directly associates with citrus cytosolic glyceraldehyde-3-phosphate dehydrogenases (CsGAPCs), which negatively regulates plant immunity. Overexpression of CsGAPCs and SDE3 significantly inhibits autophagy in citrus, Arabidopsis, and N. benthamiana. Intriguingly, SDE3 undermines autophagy-mediated immunity by the specific degradation of CsATG8 family proteins in a CsGAPC1-dependent manner. CsATG8 degradation is largely rescued by treatment with an inhibitor of the late autophagic pathway, E64d. Furthermore, ectopic expression of CsATG8s enhances Phytophthora resistance. Collectively, these results suggest that SDE3-CsGAPC interactions modulate CsATG8-mediated autophagy to enhance Las progression in citrus. Abbreviations: ACP: asian citrus psyllid; ACD2: ACCELERATED CELL DEATH 2; ATG: autophagy related; Ca. Liberibacter: Candidatus Liberibacter; CaMV: cauliflower mosaic virus; CMV: cucumber mosaic virus; Cs: Citrus sinensis; EV: empty vector; GAPC: cytosolic glyceraldehyde-3-phosphate dehydrogenase; HLB: huanglongbing; H2O2: hydrogen peroxide; Las: liberibacter asiaticus; Laf: liberibacter africanus; Lam: liberibacter americanus; Pst: Pseudomonas syringae pv. tomato; PVX: potato virus X; ROS: reactive oxygen species; SDE3: sec-delivered effector 3; TEM: transmission electron microscopy; VIVE : virus-induced virulence effector; WT: wild-type; Xcc: Xanthomonas citri subsp. citri;

(Z)-3-Hexenol integrates drought and cold stress signaling by activating abscisic acid glucosylation in tea plants.

Cold and drought stresses severely limit crop production and can occur simultaneously. Although some transcription factors and hormones have been characterized in plants subjected each stress, the role of metabolites, especially volatiles, in response to cold and drought stress exposure is rarely studied due to lack of suitable models. Here, we established a model for studying the role of volatiles in tea (Camellia sinensis) plants experiencing cold and drought stresses simultaneously. Using this model, we showed that volatiles induced by cold stress promote drought tolerance in tea plants by mediating reactive oxygen species and stomatal conductance. Needle trap microextraction combined with GC-MS identified the volatiles involved in the crosstalk and showed that cold-induced (Z)-3-hexenol improved the drought tolerance of tea plants. In addition, silencing C. sinensis alcohol dehydrogenase 2 (CsADH2) led to reduced (Z)-3-hexenol production and significantly reduced drought tolerance in response to simultaneous cold and drought stress. Transcriptome and metabolite analyses, together with plant hormone comparison and abscisic acid (ABA) biosynthesis pathway inhibition experiments, further confirmed the roles of ABA in (Z)-3-hexenol-induced drought tolerance of tea plants. (Z)-3-Hexenol application and gene silencing results supported the hypothesis that (Z)-3-hexenol plays a role in the integration of cold and drought tolerance by stimulating the dual-function glucosyltransferase UGT85A53, thereby altering ABA homeostasis in tea plants. Overall, we present a model for studying the roles of metabolites in plants under multiple stresses and reveal the roles of volatiles in integrating cold and drought stresses in plants.

First Report of Diaporthe eres Causing Leaf Spot on Pseudocydonia sinensis in China.

Pseudocydonia sinensis is a Chinese ornamental plant with great landscaping value. Its fruit is additionally used for medicinal purposes (Lim 2012). In June 2020, a leaf spot disease was observed in the campus of Nanjing Forestry University (32°04'34.53″N 118°48'42.06″E). The symptoms began with irregular red-brown spots, which gradually enlarged, extended and overlapped, with an incidence of 85% (29/34 trees). Pieces of leaf tissue (3 to 4 mm²) from the lesion margins were surface-sterilized with 75% ethanol for 30 s and 1% NaClO for 90 s. Subsequently, the tissues were rinsed with sterile H2O, placed on potato dextrose agar (PDA) medium and incubated at 25℃ for 5 days. The same fungus was isolated from 90% of tissues. Pure cultures were obtained by monosporic isolation.The representative isolate NJMG 5-7 was used for morphological and molecular characterization. The growing fungal colony on PDA was initially white, but gradually turned grey. Pycnidia formation was observed on PDA supplemented with alfalfa stems. The pycnidia produced two different types of conidia, α and β, which ooze out in yellow creamy mucilaginous masses. Conidiophores were hyaline, cylindrical and smooth, 16.8 to 33.1 × 1.5 to 2.6 μm (n=30). Conidiogenous cells were 13.6 to 29.3 × 1.5 to 2.7 μm (n=30). The α-conidia were, unicellular, hyaline elliptical or fusiform, bi-guttulate, 6.5 to 9.2 × 1.8 to 3.3 µm (n = 50). The β-conidia were hyaline, aseptate, without guttules, filiform, curved, with obtuse ends, 12.5 to 25 × 1.0 to 1.8 µm (n = 50). To verify species identity, the partial sequences of the internal transcribed spacer (ITS) region, and calmodulin (CAL), translation elongation factor 1 alpha (EF1-a), and beta-tubulin genes (TUB) were amplified from isolate NJMG 5-7 with primers ITS1/ITS4 (White et al. 1990), CAL-228F/CAL-737R (Carbone & Kohn 1999), EF1-728F/EF1-986R (Carbone and Kohn 1999), and Bt2a/Bt2b (Glass and Donaldson 1995), respectively. The sequences were deposited in GenBank (OP223050 for ITS, OP252809 for CAL, OP252807 for EF1-a, and OP252808 for TUB). A BLAST search of GenBank showed that ITS, CAL, EF1-a and TUB sequences of NJMG 5-7 were similar to those of D. eres CBS 138594 (99% identity), AR5193 (99%), AR5193 (99%) and MG281193 (100%), respectively. The morphological and molecular results identified the isolate as D. eres (Feng et al. 2015). To fulfill Koch's postulates, a pathogenicity test was conducted using three P. sinensis plants. Six leaves from each tree were wounded and inoculated with mycelial plugs (about 4 mm in diameter) of D. eres from a 3-day-old culture grown on PDA. Inoculations with sterile PDA plugs on different leaves of the same tree were used as controls. All inoculated leaves were enclosed in plastic bags together with a wet cotton ball inside. Sterile H2O was sprayed into the plastic bags to keep moisture conditions. Five days later, all inoculated points showed lesions similar to those previously observed in the field, whereas controls were asymptomatic. The pathogen was successfully reisolated from the inoculated symptomatic parts on PDA and identified from its morphology, thus fulfilling Koch's postulates. This fungus can cause a variety of diseases. To our knowledge, this is the first report of D. eres causing leaf spots on P. sinensis in the world. These findings provide a foundation for future studies on the epidemiology and control of this newly emerging disease.

Aluminum and Fluoride Stresses Altered Organic Acid and Secondary Metabolism in Tea (Camellia sinensis) Plants: Influences on Plant Tolerance, Tea Quality and Safety.

Tea plants have adapted to grow in tropical acidic soils containing high concentrations of aluminum (Al) and fluoride (F) (as Al/F hyperaccumulators) and use secret organic acids (OAs) to acidify the rhizosphere for acquiring phosphorous and element nutrients. The self-enhanced rhizosphere acidification under Al/F stress and acid rain also render tea plants prone to accumulate more heavy metals and F, which raises significant food safety and health concerns. However, the mechanism behind this is not fully understood. Here, we report that tea plants responded to Al and F stresses by synthesizing and secreting OAs and altering profiles of amino acids, catechins, and caffeine in their roots. These organic compounds could form tea-plant mechanisms to tolerate lower pH and higher Al and F concentrations. Furthermore, high concentrations of Al and F stresses negatively affected the accumulation of tea secondary metabolites in young leaves, and thereby tea nutrient value. The young leaves of tea seedlings under Al and F stresses also tended to increase Al and F accumulation in young leaves but lower essential tea secondary metabolites, which challenged tea quality and safety. Comparisons of transcriptome data combined with metabolite profiling revealed that the corresponding metabolic gene expression supported and explained the metabolism changes in tea roots and young leaves via stresses from high concentrations of Al and F. The study provides new insight into Al- and F-stressed tea plants with regard to responsive metabolism changes and tolerance strategy establishment in tea plants and the impacts of Al/F stresses on metabolite compositions in young leaves used for making teas, which could influence tea nutritional value and food safety.

Effect of the biosynthesis of the volatile compound phenylacetaldehyde on chloroplast modifications in tea (Camellia sinensis) plants.

Plant volatile compounds have important physiological and ecological functions. Phenylacetaldehyde (PAld), a volatile phenylpropanoid/benzenoid, accumulates in the leaves of tea (Camellia sinensis) plants grown under continuous shading. This study was conducted to determine whether PAld production is correlated with light and to elucidate the physiological functions of PAld in tea plants. Specifically, the upstream mechanism modulating PAld biosynthesis in tea plants under different light conditions as well as the effects of PAld on chloroplast/chlorophyll were investigated. The biosynthesis of PAld was inhibited under light, whereas it was induced in darkness. The structural gene encoding aromatic amino acid aminotransferase 1 (CsAAAT1) was expressed at a high level in darkness, consistent with its importance for PAld accumulation. Additionally, the results of a transcriptional activation assay and an electrophoretic mobility shift assay indicated CsAAAT1 expression was slightly activated by phytochrome-interacting factor 3-2 (CsPIF3-2), which is a light-responsive transcription factor. Furthermore, PAld might promote the excitation of chlorophyll in dark-treated chloroplasts and mediate electron energy transfer in cells. However, the accumulated PAld can degrade chloroplasts and chlorophyll, with potentially detrimental effects on photosynthesis. Moreover, PAld biosynthesis is inhibited in tea leaves by red and blue light, thereby decreasing the adverse effects of PAld on chloroplasts during daytime. In conclusion, the regulated biosynthesis of PAld in tea plants under light and in darkness leads to chloroplast modifications. The results of this study have expanded our understanding of the biosynthesis and functions of volatile phenylpropanoids/benzenoids in tea leaves.

A Review on Phytochemical and Pharmacological Screening of Hibiscus rosa-sinensis L

Due to the rising disease burden, natural plant products are becoming popular today. The plant Hibiscus rosa sinensis Linn., which belongs to the Malvaceae family, is found all over the world. In India's traditional medicine, its leaves, barks, roots, and flowers have been used to treat a variety of illnesses. Numerous studies have demonstrated the antioxidant, antimicrobial, antidiabetic, antiulcer, hepatoprotective, antifertility, antigenotoxic, and antiinflammatory qualities that the various sections of Hibiscus rosa sinensis plants contain, which aid in the treatment of a variety of ailments. Numerous herbal concoctions and beverages contain hibiscus rosa sinensis. Numerous studies using animal models analyze the flowers and leaves of Hibiscus rosa sinensis for their anti-diabetic and antioxidant properties. This review makes an effort to highlight the medicinal uses of Hibiscus rosa sinensis.