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Salinity is one of the predominant abiotic stressors that reduce plant growth, yield, and productivity. Ameliorating salt tolerance through nanotechnology is an efficient and reliable methodology for enhancing agricultural crops yield and quality. Nanoparticles enhance plant tolerance to salinity stress by facilitating reactive oxygen species detoxification and by reducing the ionic and osmotic stress effects on plants. This experiment was conducted to study the effects of NaCl salinity stress (0, 100, and 200 mM), and foliar application of quantum dot-graphene oxide, nano-TiO2, and CeO2 (zero and 2 g/l) on the growth and physiological responses of Capsicum annum L. The results revealed that the interaction effects of treatments significantly affected plant and fruit fresh weight, chlorophyll a, total soluble solids, phenolics, malondialdehyde, H2O2, and proline content. Moreover, catalase activity and sodium, and phosphorus content were responded to the treatments. The highest fresh weight of plants and fruits, fruit diameter, and chlorophyll a content were recorded under no-salinity × quantum dot-graphene oxide foliar use. The highest data for total phenolics content was recorded at NaCl100 mM × quantum dot-graphene oxide. In contrast, the maximum flavonoids content belonged to NaCl100 mM × quantum dot-graphene oxide and NaCl100 mM × TiO2. The experimental treatments independently affected the number of fruits, chlorophyll b, carotenoids, and vitamin C content, as well as K/Na ratio. The foliar treatment of quantum dot-graphene oxide nanoparticles improved the carotenoids and vitamin C content, stem diameter, and fruit number. The overall results disclosed that, when plants were exposed to high salinity levels; the foliar treatments were unable to effectively mitigate the negative impacts of salt stress on the plant, except for certain traits such as total phenolics, flavonoids, and TSS levels. However, under the low and mild salinity depression, the foliar treatments were enough capable to overcome the salinity defects. [ABSTRACT FROM AUTHOR]

[Objective] This study seeks to identify and analyze the PIN gene family members in pepper (Capsicum annuum L.) and to investigate their expression patterns in the abscission zone of the fruit pedicel as well as during root development. The ultimate goal is to provide candidate genes for elucidating the function of CaPINs and for breeding new pepper varieties. [Methods] PIN genes were identified through whole-genome screening of pepper, followed by comprehensive bioinformatics analysis. The expression patterns of these genes in the fruit pedicel abscission zone and during root development were systematically analyzed. [Results] 9 members of the PIN gene family were identified in the pepper genome and designated as CaPIN1 through CaPIN9. These genes are distributed across seven chromosomes, with CaPIN9 remaining unanchored. The proteins encoded by these genes exhibit amino acid length ranging from 358 to 654 residues, molecular weight from 39 251.05 D to 71 170.91 D, and isoelectric point from 6.40 to 9.38. Most of these proteins are stable. Additionally, significant collinearity was observed among the CaPINs, indicating a close genetic relationship with the tomato genome. CaPINs encompass a multitude of elements associated with growth, environmental responses, and hormonal signaling. Transcriptome data analysis revealed that the expression of CaPINs in pepper was tissue-specific. Quantitative PCR results demonstrated that CaPINs exhibited differential expression levels across various fruit pedicel abscission zones and throughout root development. [Conclusion] CaPINs play a crucial role in the development of the pedicel abscission zone and the growth and development of the root system. These genes hold significant potential as candidates for breeding pepper varieties with improved ease of de-stemming and for enhancing genetic traits related to root systems.

Abstract [Objective] PEX (pollen extensin-like) gene is involved in controlling pollen development, which plays a crucial role in embryogenesis of pollen microspores and haploid culture. In this study, the authors isolated the CaPEX3 gene from pepper and analyzed its protein structure and expression pattern, which provides a theoretical foundation for further investigation into the function of the CaPEX3 gene. [Methods] CaPEX3 was cloned by PCR, and its protein structure, evolution, and gene regulation were analyzed by bioinformatics. Expression of CaPEX3 in different tissues and pollen of different varieties were detected by qRT-PCR. Pollen viability of different pepper varieties was detected by staining method and their relationship with CaPEX3 expression was analyzed. [Results] The coding region of CaPEX3 was 1 581 bp in length, encoding 526 amino acids, which belongs to the LRX (leucine-rich repeats/extensins) gene family. The protein had phosphorylation and glycosylation sites, signaling peptides, and transmembrane domains. The expression level of CaPEX3 was the highest in flowers, and the gene expression was higher in tissues with high pollen vitality. The regulatory network predicted that CaPEX3 might be involved in the regulation of pollen development through the action of genes related to flower development. [Conclusion] CaPEX3 was specifically expressed in flowers, and a correlation was observed between its expression level and the activity of pollen grains. The analysis of gene expression and the construction of related vectors provided a theoretical foundation and technical support for the subsequent investigation of gene function and mechanism of CaPEX3

Abstract Temperature is an important limiting factor in the counter-seasonal cultivation of pepper. Currently, there are no studies on transcriptomic analysis of ‘cold stress memory’ in pepper. In this study, in order to understand the mechanism of ‘cold stress memory’ in pepper (Capsicum annuum L.), seedlings were subjected to the following treatments: normal temperature treatment (P0), the first cold treatment for 3 days (P3), the recovery temperature treatment for 3 days (R3), and another cold treatment for 3 days (RP3). The results showed that P3 plants wilted the most, RP3 the second and R3 the least. Leaf reactive oxygen species (ROS) and electrolyte leakage were the most in P3, the second in RP3 and the least in R3. In addition, RP3 had the highest accumulation of zeaxanthin, violaxanthin and β-cryptoxanthin, followed by P3, and R3 had the least. These results suggest that pepper seedlings are characterized by ‘cold stress memory’. Transcriptomics was used to analyze the key genes and transcription factors involved in the biosynthesis of zeaxanthin, violaxanthin and β-cryptoxanthin during the formation of ‘cold stress memory’. This study provides candidate genes and transcription factors for an in-depth study of the cold tolerance mechanism in pepper.

Abstract Background The growth and yield of pepper, a typical temperature-loving vegetable, are limited by low-temperature environments. Using low-temperature sensitive ‘Hangjiao No. 4’ (Capsicum annuum L.) as experimental material, this study analyzed the changes in plant growth and photosynthesis under different treatments: normal control (NT), low-temperature stress alone (LT), low-temperature stress in strigolactone pretreated plants (SL_LT), and low-temperature stress in strigolactone biosynthesis inhibitor pretreated plants (Tis_LT). Results SL pretreatment increased the net photosynthetic rate (Pn) and PSII actual photochemical efficiency (φPSII), reducing the inhibition of LT on the growth of pepper by 17.44% (dry weight of shoot). Due to promoting the accumulation of carotenoids, such as lutein, and the de-epoxidation of the xanthophyll cycle [(Z + A)/(Z + A + V)] by strigolactone after long-term low-temperature stress (120 h), non-photochemical quenching (NPQ) of pepper was increased to reduce the excess excitation energy [(1-qP)/NPQ] and the photoinhibition degree (Fv/Fm) of pepper seedlings under long-term low-temperature stress was alleviated. Twelve cDNA libraries were constructed from pepper leaves by transcriptome sequencing. There were 8776 differentially expressed genes (DEGs), including 4473 (51.0%) upregulated and 4303 (49.0%) downregulated genes. Gene ontology pathway annotation showed that based on LT, the DEGs of SL_LT and Tis_LT were significantly enriched in the cellular component, which is mainly related to the photosystem and thylakoids. Further analysis of the porphyrin and chlorophyll biosynthesis, carotenoid biosynthesis, photosynthesis-antenna protein, and photosynthetic metabolic pathways and the Calvin cycle under low-temperature stress highlighted 18, 15, 21, 29, and 31 DEGs for further study, which were almost all highly expressed under SL_LT treatment and moderately expressed under LT treatment, whereas Tis_LT showed low expression. Conclusion The positive regulatory effect of SLs on the low-temperature tolerance of pepper seedlings was confirmed. This study provided new insights for the development of temperature-tolerant pepper lines through breeding programs.

Abstract [Objective] The study aims to explore the expression and function of the RIN gene, encoding a MADS-box, and analyze its impact on the metabolism of carotenoids in pepper. [Methods] Based on the fruit development transcriptome of pepper, the full-length CDS of the MADS-box transcription factor Ca- RIN gene related to chili ripening was cloned. The bioinformatics, expression pattern, subcellular localization, and transcriptional activation activity were examined, and the effects of VIGS-induced CaRIN gene silencing on carotenoid metabolism were investigated. [Results] (1) The CDS of CaRIN gene was 732 bp in length. CaRIN gene encoded a protein containing 243 amino acids with a molecular mass of 27.95 kD and an isoelectric point (pI) of 7.06. CaRIN protein had a typical MEF2_ like MADS domain, belonging to the MICK type transcription factor. (2) CaRIN gene was mainly expressed in flowers and fruits with tissue specificity. CaRIN was located in the nucleus and had transcriptional activation activity. (3) The promoter of CaRIN gene had several hormone response elements such as ABRE, and exogenous ABA and ethephon accelerated the fruit to turn red and induced the high expression of CaRIN and related genes. (4) After VIGS-induced gene silencing of CaRIN , the expression level of PSY1, CCS, PDS, CRTZ, LCYB, and NCED1, genes involved in carotenoid metabolic pathway, was decreased by 0.27-0.59 times, and the total carotenoid content in fruits (0.379 mg/g) was decreased compared with the control (0.650 mg/g). [Conclusion] CaRIN may be an important regulatory factor of carotenoid metabolism in capsicum fruits.

Abstract [Objective] Pepper is the largest vegetable crop planted in China. With the land salinization becoming more and more serious, it is urgent to accelerate the functional study of key genes related to pepper salt tolerance. It is of great significance to strengthen the study of mechanisms underlying salt tolerance in peper to promote sustainable development of the pepper industry. [Methods] In the early stage of research, CaNAC36, a NAC transcription factor family gene related to salt stress response in peppers, was discovered. Based on this, the full-length gDNA and cDNA sequences of CaNAC36 were cloned using salttolerant pepper accession PI201224 and salt-sensitive pepper accession PI438643. The expression of CaNAC36 and its interacting genes in different tissues under salt stress was quantitatively analyzed by fluorescence, and the potential relationship between CaNAC36 and its interacting genes was further explored by bioinformatics analysis. [Results] The gDNA and cDNA sequence homology of CaNAC36 in salt-tolerant and salt-sensitive materials were 99.86% and 100%, respectively. CaNAC36 showed an up-expression in roots and stems in the salt-tolerant pepper, and a down-expression in roots and leaves in the salt-sensitive peper. Throgh analyzing the annotation information of 48 genes that may interact with CaNAC36, it was found that 14 genes were belonging to transmembrane proteins, transporter proteins, water porins, chloride channel proteins, detoxifying proteins, and other genes, which may have functional interaction with CaNAC36. Further analysis revealed that PI201224 and PI438643 exhibited significant differences in the expression of five related genes, inclding Capana08g002748, Capana00g004514, Capana09g000275, Capana07g001450, Capana02g001031, at different time points and tissues under salt stress. Meanwhile, the promoter region of CaNAC36 and five associated genes was found to contain a large number of stressrelated cis-acting elements. [Conclusion] CaNAC36 was an important regulatory gene in pepper in response to salt stress, and might interact with other genes to improve salt tolerance.

Abstract Background The B3 gene family, one of the largest plant-specific transcription factors, plays important roles in plant growth, seed development, and hormones. However, the B3 gene family, especially the REM subfamily, has not been systematically and functionally studied. Results In this study, we performed genome-wide re-annotation of B3 genes in five Solanaceae plants, Arabidopsis thaliana, and Oryza sativa, and finally predicted 1,039 B3 genes, including 231 (22.2%) newly annotated genes. We found a striking abundance of REM genes in pepper species (Capsicum annuum, Capsicum baccatum, and Capsicum chinense). Comparative motif analysis revealed that REM and other subfamilies (ABI3/VP1, ARF, RAV, and HSI) consist of different amino acids. We verified that the large number of REM genes in pepper were included in the specific subgroup (G8) through the phylogenetic analysis. Chromosome location and evolutionary analyses suggested that the G8 subgroup genes evolved mainly via a pepper-specific recent tandem duplication on chromosomes 1 and 3 after speciation between pepper and other Solanaceae. RNA-seq analyses suggested the potential functions of REM genes under salt, heat, cold, and mannitol stress conditions in pepper (C. annuum). Conclusions Our study provides evolutionary and functional insights into the REM gene family in pepper.

Due to the vulnerability of pepper (Capsicum spp.) and the virulence of tomato spotted wilt virus (TSWV), seasonal shortages and surges of prices are a challenge and thus threaten household income. Traditional bioassays for detecting TSWV, such as observation for symptoms and reverse transcription-PCR, are time-consuming, labor-intensive, and sometimes lack precision, highlighting the need for a faster and more reliable approach to plant disease assessment. Here, two imaging techniques—Red–Green–Blue (RGB) and hyperspectral imaging (using NDVI and wavelength intensities)—were compared with a bioassay method to study the incidence and severity of TSWV in different pepper accessions. The bioassay results gave TSWV an incidence from 0 to 100% among the accessions, while severity ranged from 0 to 5.68% based on RGB analysis. The normalized difference vegetative index (NDVI) scored from 0.21 to 0.23 for healthy spots on the leaf but from 0.14 to 0.19 for disease spots, depending on the severity of the damage. The peak reflectance of the disease spots on the leaves was identified in the visible light spectrum (430–470 nm) when spectral bands were studied in the broad spectrum (400.93–1004.5 nm). For the selected wavelength in the visible light spectrum, a high reflectance intensity of 340 to 430 was identified for disease areas, but between 270 and 290 for healthy leaves. RGB and hyperspectral imaging techniques can be recommended for precise and accurate detection and quantification of TSWV infection.

Pepper is a globally cultivated vegetable known for its distinct pungent flavor, which is derived from the presence of capsaicinoids, a class of unique secondary metabolites that accumulate specifically in pepper fruits. Since the accumulation of capsaicinoids is influenced by various factors, it is imperative to comprehend the metabolic regulatory mechanisms governing capsaicinoids production. This review offers a thorough examination of the factors that govern the metabolism of capsaicinoids in pepper fruit, with a specific focus on three primary facets: (1) the impact of genotype and developmental stage on capsaicinoids metabolism, (2) the influence of environmental factors on capsaicinoids metabolism, and (3) exogenous substances like methyl jasmonate, chlorophenoxyacetic acid, gibberellic acid, and salicylic acid regulate capsaicinoid metabolism. The findings of this study are expected to enhance comprehension of capsaicinoids metabolism and aid in the improvement of breeding and cultivation practices for high-quality pepper in the future.

Abiotic stress is one of the key factors harming global agriculture today, seriously affecting the growth and yield of vegetables. Pepper is the most widely grown vegetable in the world, with both high nutritional and economic values. Currently, the increase in global extreme weather events has heightened the frequency of abiotic stresses, such as drought, high and low temperatures, waterlogging, and high salt levels, which impairs pepper growth and development, leading to its reduced yield and quality. In this review, we summarize the research progress on the responses of pepper to abiotic stress in recent years in terms of physiology, biochemistry, molecular level, and mitigation measures. We then explore the existing problems and propose future research directions. This work provides a reference for the cultivation and development of new pepper varieties resistant to abiotic stress.

Salt stress is a major abiotic factor that negatively impacts the growth, performance, and secondary metabolite production in pepper (Capsicum annuum L.) plants. Brassinosteroids (BRs) play a crucial role in enhancing plant tolerance to abiotic stress, yet their potential in mitigating salt stress in pepper plants, particularly by promoting sugar and glycine betaine accumulation, remains underexplored. In this study, we investigated the effects of the foliar application of 2,4-epibrassinolide (EBR) on salt-stressed pepper seedlings. Our findings revealed that EBR treatment significantly increased the levels of proline, sugar, and glycine betaine under salt stress compared to untreated controls. Moreover, EBR enhanced the antioxidant defense mechanisms in pepper seedlings by increasing sugar and glycine betaine levels, which contributed to the reduction of reactive oxygen species (ROS) and malondialdehyde (MDA) accumulation.

Cerium oxide nanoparticles (CeO2NPS) have been shown to improve the salt tolerance of plant due to the potential to eliminate free radicals and the antioxidation properties. However, the effects and mechanisms of seed priming with CeO2NPS on pepper plant are unclear. To reveal impacts of CeO2NPS seed priming on the germination and seedling growth of peeper plants under salt stress, using the Maoshu 360 variety of pepper（Capsicum annuum）as material, a total of seven concentrations of treatments (0, 0.05, 0.1, 0.2, 0.3, 0.4, 0.5 mmol·L-1) and a control of un-treatment were carried out to investigate effects of CeO2NPS seed priming on seed germination rate, seedling biomass, and physiological and biochemical indexes under salt stress. The results were as follows: (1) In the seed priming with 0.5 mmol·L-1 CeO2NPS, soluble protein, content of proline, activity of catalase (CAT), content of ascorbic acid (AsA) and ratio of AsA/DHA significantly increased and levels of the superoxide anion (O2-) significantly decreased. There was no loss of viability for the seeds primed with cerium oxide nanoparticles. Indeed their germination rate and germination potential were enhanced under salt stress, and their germination index, and vigor index were improved under salt stress. (2) Under salt stress, seedlings from seeds primed with 0.4 mmol·L-1 CeO2NPS showed the highest fresh weight, dry weight, and root length; additionally, soluble protein content, AsA content and AsA/DHA ratio were also improved. In conclusion, seed priming with CeO2NPS can not only enhance seed germination under salt stress by decreasing seed water potential, accelerating storage substances metabolism and improving antioxidant capacity but also promote seedling growth through boosting protein synthesis and ascorbic acid-glutathione cycle (AsA-GSH).

This study evaluated the side-cleft grafting in native species of the genus Piper (Piper aduncum L., Piper hispidum Sw., and Piper tuberculatum Jacq.) with BRS Kottanadan pepper cultivar as rootstocks (Experiment 1). The top-cleft grafting was evaluated in native species Piper arboreum Aubl., P. aduncum, and P. tuberculatum with Balankotta pepper cultivar as graft using black pepper cv. Bragantina (Experiment 2). Black pepper cv. Bragantina autograft was considered as control. The rootstock P. tuberculatum showed initial incompatibility with the cv. Bragantina, with survival of about 34.8% (side) and 62.5% (top) and total inhibition of graft shooting. The side and top graft on P. aduncum resulted in 84.0 and 47.5% of survival, respectively. The JIP-test parameters indicated better photochemical efficiency in the species grafted using the method of side-cleft grafting. Finally, the initial compatibility was associated with greater survival, shooting, and better energy flow through the electron transport chain.

Bacterial wilt caused by Ralstonia solanacearum is one of the most important diseases in solanaceous plants, including peppers. It generally tends to be more serious under warm−temperature and moist (WM) conditions than at moist room−temperature (RM) conditions. Although immunity mechanisms at room temperature have been intensively studied, the mechanisms underlying WM conditions remain poorly understood. Herein, the pepper cysteine protease CaZingipain2 was expressed and functionally characterized in pepper immunity against R. solanacearum at WM conditions and at room temperature. The results showed that CaZingipain2 localized to the nucleus and was upregulated at the transcript level in pepper plants upon R. solanacearum infection under WM conditions (RSWM). Virus−induced gene silencing of CaZingipain2 significantly increased the susceptibility of pepper plants to RSWM, and was coupled with the downregulation of CaPRP1 and CaMgst3, which are specifically related to pepper immunity against RSWM, according to our previous studies, while its overexpression significantly reduced the susceptibility of N. benethamiana plants to RSWM compared to that of wild−type plants. In addition, our data showed that CaZingipain2 also acts positively in pepper immunity against R. solanacearum infection at room temperature by upregulating the SA− and JA−responsive PR genes, including CaNPR1 and CaDEF1. All these results indicate that CaZingipain2 improves pepper immunity against R. solanacearum under WM conditions and at room temperature by regulating different PR genes.

Low temperature combined with low light (LL) is an important factor limiting pepper quality and yield. ‘Hang Jiao No. 2′ were used as experimental materials, and different concentrations of MeJA (T1 (0 μM), T2 (100 μM), T3 (150 μM), T4 (200 μM), T5 (250 μM) and T6 (300 μM)) were sprayed under LL stress to explore the positive effect of exogenous methyl jasmonate (MeJA) on peppers under LL stress. The photosynthetic properties, osmoregulatory substance, reactive oxygen species, antioxidant enzyme activities, and related gene expressions of the peppers were measured. Our results demonstrated that 200 μM MeJA treatment significantly increased chlorophyll content, light quantum flux per active RC electron transfer (Eto/RC), maximum captured photonic flux per active RC (TRo/RC), energy flux for electron transfer in the excitation cross section (Eto/CSm), energy flux captured by absorption in the excitation cross section (TRo/CSm), soluble protein, and soluble sugar content. Moreover, it significantly improved the maximum photochemical efficiency of PSII (Fv/Fm) and performance index based on absorbed light energy (PI (abs)) by 56.77% and 67.00%, respectively, and significantly decreased malondialdehyde (MDA) content and relative conductivity by 30.55% and 28.17%, respectively. Additionally, antioxidant enzyme activities were elevated, and the expression of the related genes was activated in pepper seedlings under stress, leading to a significant reduction in reactive oxygen species content. In conclusion, our findings confirmed that 200 μM MeJA could reduce the injury of LL to pepper leaves to the photosynthetic organs of pepper leaves, protect the integrity of the cell membrane, and further improve the tolerance of pepper seedlings to LL.

As important transcription factors, WRKYs play a vital role in the defense response of plants against the invasion of multiple pathogens. Though some WRKY members have been reported to participate in pepper immunity in response to Ralstonia solanacearum infection, the functions of the majority of WRKY members are still unknown. Herein, CaWRKY22b was cloned from the pepper genome and its function against R. solanacearum was analyzed. The transcript abundance of CaWRKY22b was significantly increased in response to the infection of R. solanacearum and the application of exogenous methyl jasmonate (MeJA). Subcellular localization assay in the leaves of Nicotiana benthamiana showed that CaWRKY22b protein was targeted to the nuclei. Agrobacterium-mediated transient expression in pepper leaves indicated that CaWRKY22b overexpression triggered intensive hypersensitive response-like cell death, H2O2 accumulation, and the up-regulation of defense- and JA-responsive genes, including CaHIR1, CaPO2, CaBPR1, and CaDEF1. Virus-induced gene silencing assay revealed that knock-down of CaWRKY22b attenuated pepper’s resistance against R. solanacearum and the up-regulation of the tested defense- and jasmonic acid (JA)-responsive genes. We further assessed the role of CaWRKY22b in modulating the expression of JA-responsive CaDEF1, and the result demonstrated that CaWRKY22b trans-activated CaDEF1 expression by directly binding to its upstream promoter. Collectively, our results suggest that CaWRKY22b positively regulated pepper immunity against R. solanacearum in a manner associated with JA signaling, probably by modulating the expression of JA-responsive CaDEF1.

Abstract The ribosomal protein contains complex structures that belong to polypeptide glycoprotein family, which are involved in plant growth and responses to various stresses. In this study, we found that capsicum annuum 40S ribosomal protein SA-like (CaSLP) was extensively accumulated in the cell nucleus and cell membrane, and the expression level of CaSLP was up-regulated by Salicylic acid (SA) and drought treatment. Significantly fewer peppers plants could withstand drought stress after CaSLP gene knockout. The transient expression of CaSLP leads to drought tolerance in pepper, and Arabidopsis’s ability to withstand drought stress was greatly improved by overexpressing the CaSLP gene. Exogenous application of SA during spraying season enhanced drought tolerance. CaSLP-knockdown pepper plants demonstrated a decreased resistance of Pseudomonas syringae PV.tomato (Pst) DC3000 (Pst.DC3000), whereas ectopic expression of CaSLP increased the Pst.DC3000 stress resistance in Arabidopsis. Yeast two-hybrid (Y2H) and bimolecular fluorescence complementation (BiFC) results showed that CaNAC035 physically interacts with CaSLP in the cell nucleus. CaNAC035 was identified as an upstream partner of the CaPR1 promoter and activated transcription. Collectively the findings demonstrated that CaSLP plays an essential role in the regulation of drought and Pst.DC3000 stress resistance. Graphical Abstract

Although 14-3-3 proteins have been implicated in plant growth, development, and stress response, their roles in pepper immunity against R. solanacearum remain poorly understood. In this study, a 14-3-3-encoding gene in pepper, Ca16R, was found to be upregulated by R. solanacearum inoculation (RSI), its silencing significantly reduced the resistance of pepper plants to RSI, and its overexpression significantly enhanced the resistance of Nicotiana benthamiana to RSI. Consistently, its transient overexpression in pepper leaves triggered HR cell death, indicating that it acts positively in pepper immunity against RSI, and it was further found to act positively in pepper immunity against RSI by promoting SA but repressing JA signaling. Ca16R was also found to interact with CaASR1, originally using pull-down combined with a spectrum assay, and then confirmed using bimolecular fluorescence complementation (BiFC) and a pull-down assay. Furthermore, we found that CaASR1 transient overexpression induced HR cell death and SA-dependent immunity while repressing JA signaling, although this induction and repression was blocked by Ca16R silencing. All these data indicate that Ca16R acts positively in pepper immunity against RSI by interacting with CaASR1, thereby promoting SA-mediated immunity while repressing JA signaling. These results provide new insight into mechanisms underlying pepper immunity against RSI.

Trichomes are specialized organs located in the plant epidermis that play important defense roles against biotic and abiotic stresses. However, the mechanisms regulating the development of pepper epidermal trichomes and the related regulatory genes at the molecular level are not clear. Therefore, we performed transcriptome analyses of A114 (less trichome) and A115 (more trichome) to dig deeper into the genes involved in the regulatory mechanisms of epidermal trichome development in peppers. In this study, the epidermal trichome density of A115 was found to be higher by phenotypic observation and was highest in the leaves at the flowering stage. A total of 39,261 genes were quantified by RNA-Seq, including 11,939 genes not annotated in the previous genome analysis and 18,833 differentially expressed genes. Based on KEGG functional enrichment, it was found that DEGs were mainly concentrated in three pathways: plant–pathogen interaction, MAPK signaling pathway-plant, and plant hormone signal transduction. We further screened the DEGs associated with the development of epidermal trichomes in peppers, and the expression of the plant signaling genes GID1B-like (Capana03g003488) and PR-6 (Capana09g001847), the transcription factors MYB108 (Capana05g002225) and ABR1-like (Capana04g001261), and the plant resistance genes PGIP-like (Capana09g002077) and At5g49770 (Capana08g001721) in the DEGs were higher at A115 compared to A114, and were highly expressed in leaves at the flowering stage. In addition, based on the WGCNA results and the establishment of co-expression networks showed that the above genes were highly positively correlated with each other. The transcriptomic data and analysis of this study provide a basis for the study of the regulatory mechanisms of pepper epidermal trichomes.

The development of pepper cultivars with durable resistance to the oomycete Phytophthora capsici has been challenging due to differential interactions between the species that allow certain pathogen isolates to cause disease on otherwise resistant host genotypes. Currently, little is known about the pathogen genes involved in these interactions. To investigate the genetic basis of P. capsici virulence on individual pepper genotypes, we inoculated sixteen pepper accessions, representing commercial varieties, sources of resistance, and host differentials, with 117 isolates of P. capsici, for a total of 1,864 host-pathogen combinations. Analysis of disease outcomes revealed a significant effect of inter-species genotype-by-genotype interactions, although these interactions were quantitative rather than qualitative in scale. Isolates were classified into five pathogen subpopulations, as determined by their genotypes at over 60,000 single-nucleotide polymorphisms (SNPs). While absolute virulence levels on certain pepper accessions significantly differed between subpopulations, a multivariate phenotype reflecting relative virulence levels on certain pepper genotypes compared with others showed the strongest association with pathogen subpopulation. A genome-wide association study (GWAS) identified four pathogen loci significantly associated with virulence, two of which colocalized with putative RXLR effector genes and another with a polygalacturonase gene cluster. All four loci appeared to represent broad-spectrum virulence genes, as significant SNPs demonstrated consistent effects regardless of the host genotype tested. Host genotype–specific virulence variants in P. capsici may be difficult to map via GWAS with all but excessively large sample sizes, perhaps controlled by genes of small effect or by multiple allelic variants that have arisen independently. [Graphic: see text] Copyright © 2022 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Pepper (Capsicum annuum L.) is a herbaceous plant species in the family Solanaceae. Capsicum anthracnose is caused by the genus Colletotrichum. spp., which decreases pepper production by about 50% each year due to anthracnose. In this study, we evaluated the resistance of red ripe fruits from 17 pepper varieties against anthracnose fungus Colletotrichum capsici. We assessed the size of the lesion diameter and conducted significance analysis to identify the resistant variety of B158 and susceptible variety of B161. We selected a resistant cultivar B158 and a susceptible cultivar B161 of pepper and used a transcription to investigate the molecular mechanisms underlying the plant’s resistance to C. capsici, of which little is known. The inoculated fruit from these two varieties were used for the comparative transcription analysis, which revealed the anthracnose-induced differential transcription in the resistant and susceptible pepper samples. In the environment of an anthrax infection, we found that there were more differentially expressed genes in resistant varieties compared to susceptible varieties. Moreover, the response to stimulus and stress ability was stronger in the KANG. The transcription analysis revealed the activation of plant hormone signaling pathways, phenylpropanoid synthesis, and metabolic processes in the defense response of peppers against anthracnose. In addition, ARR-B, AP2-EREBP, bHLH, WRKY, and NAC are associated with disease resistance to anthracnose. Notably, WRKY and NAC were found to have a potentially positive regulatory role in the defense response against anthracnose. These findings contribute to a more comprehensive understanding of the resistance mechanisms of red pepper fruit to anthracnose infection, providing valuable molecular insights for further research on the resistance mechanisms and genetic regulations during this developmental stage of pepper.

Capsicum baccatum is a close relative of edible chili peppers (Capsicum annuum) with high economic value. The CBF gene family plays an important role in plant stress resistance physiology. We detected a total of five CBF genes in the C. baccatum genome-wide sequencing data. These genes were scattered irregularly across four chromosomes. The genes were categorized into three groupings according to their evolutionary relationships, with genes in the same category showing comparable principles for motif composition. The 2000 bp upstream of CbCBF contains many resistance-responsive elements, hormone-responsive elements, and transcription factor binding sites. These findings emphasize the crucial functions of these genes in responding to challenging conditions and physiological regulation. Analysis of tissue-specific expression revealed that CbCBF3 exhibited the greatest level of expression among all tissues. Under conditions of low-temperature stress, all CbCBF genes exhibited different levels of responsiveness, with CbCBF3 showing a considerable up-regulation after 0.25 h of cold stress, indicating a high sensitivity to low-temperature response. The importance of the CbCBF3 gene in the cold response of C. baccatum was confirmed by the use of virus-induced gene silencing (VIGS) technology, as well as the prediction of its protein interaction network. To summarize, this study conducts a thorough bioinformatics investigation of the CbCBF gene family, showcases the practicality of employing VIGS technology in C. baccatum, and confirms the significance of the CbCBF3 gene in response to low temperatures. These findings provide significant references for future research on the adaptation of C. baccatum to low temperatures.

The vegetable plug seedling plays an important role in improving vegetable production. The process of plug seedling contributes to high-quality vegetable seedlings. The substrate composition and chemical fertilizer are widely studied to promote seedling growth. However, little is known about the effect of beneficial bacteria in the rhizosphere microbial community and vegetables’ growth during plug seedling. The use of beneficial microbes to promote vegetable seedling growth is of great potential. In this study, we showed that the Serratia marcescens strain LYGN1 enhanced the growth of cucumber and pepper seedlings in plug seedling cultivation. The treatment with LYGN1 significantly increased the biomass and the growth-related index of cucumber and pepper, improving the seedling quality index. Specifically, LYGN1 also improved the cucumber and pepper root system architecture and increased the root diameter. We applied high-throughput sequencing to analyze the microbial community of the seedlings’ rhizosphere, which showed LYGN1 to significantly change the composition and structure of the cucumber and pepper rhizosphere microbial communities. The correlation analysis showed that the Abditibacteriota and Bdellovibrionota had positive effects on seedling growth. The findings of this study provide evidence for the effects of Serratia marcescens LYGN1 on the cucumber and pepper rhizosphere microbial communities, which also promoted seedling quality in plug seedling cultivation.

The absence of an externally-imposed 24 h light/dark cycle in closed plant production systems allows setting the light environmental parameters in unconventional ways. Innovative lighting modes for energy-saving, high-quality, and yield production are widely discussed. This study aimed to evaluate the effects of the light/dark cycles of 16/8 h (control) and 24/12 h, 48/24 h, 96/48 h, 120/60 h (unconventional cycles) based on the same total light amount, and continuous lighting (360/0 h) on plant performance of some Solanaceae species. Responses of eggplant (Solanum melongena L.), sweet pepper (Capsicum annuum L.), tobacco (Nicotiana tabacum L.), and tomato (Solanum lycopersicum L.) plants to extended light/dark cycles and continuous lighting were studied under controlled climate conditions. Plants with two true leaves were exposed to different light/dark cycles for 15 days. Light intensity was 250 µmol m−2 s−1 PPFD, provided by light-emitting diodes (LEDs). After the experiment, tomato, sweet pepper, and eggplant transplants were planted in a greenhouse and grown under identical conditions of natural photoperiod for the estimation of the after-effect of light treatments on fruit yield. Extended light/dark cycles of 24/12 h, 48/24 h, 96/48 h, 120/60 h, and 360/0 h affected growth, development, photosynthetic pigment content, anthocyanin and flavonoid content, and redox state of plants. Effects varied with plant species and length of light/dark cycles. In some cases, measured parameters improved with increasing light/dark periods despite the same total sum of illumination received by plants. Treatments of tomato and pepper transplants with 48/24 h, 96/48 h, and 120/60 h resulted in higher fruit yield compared to conventional 16/8 h photoperiod. The conclusion was made that extended light/dark cycles can result in increased light use efficiency compared to conventional photoperiod and, therefore, reduced product cost, but for practical application, the effects need to be further explored for individual plant species or even cultivars.

Abstract Background The plant homeodomain (PHD)-finger gene family that belongs to zinc-finger genes, plays an important role in epigenetics by regulating gene expression in eukaryotes. However, inaccurate annotation of PHD-finger genes hinders further downstream comparative, evolutionary, and functional studies. Results We performed genome-wide re-annotation in Arabidopsis thaliana (Arabidopsis), Oryza sativa (rice), Capsicum annuum (pepper), Solanum tuberosum (potato), and Solanum lycopersicum (tomato) to better understand the role of PHD-finger genes in these species. Our investigation identified 875 PHD-finger genes, of which 225 (26% of total) were newly identified, including 57 (54%) novel PHD-finger genes in pepper. The PHD-finger genes of the five plant species have various integrated domains that may be responsible for the diversification of structures and functions of these genes. Evolutionary analyses suggest that PHD-finger genes were expanded recently by lineage-specific duplication, especially in pepper and potato, resulting in diverse repertoires of PHD-finger genes among the species. We validated the expression of six newly identified PHD-finger genes in pepper with qRT-PCR. Transcriptome analyses suggest potential functions of PHD-finger genes in response to various abiotic stresses in pepper. Conclusions Our data, including the updated annotation of PHD-finger genes, provide useful information for further evolutionary and functional analyses to better understand the roles of the PHD-finger gene family in pepper.

Abstract Background Anthocyanins have distinct biological functions in plant coloring, plant defense against strong light, UV irradiation, and pathogen infection. Aromatic hydroxyl groups and ortho-dihydroxyl groups in anthocyanins are able to inhibit free-radical chain reactions and hydroxyl radicals. Thus, anthocyanins play an antioxidative role by removing various types of ROS. Pepper is one of the solanaceous vegetables with the largest cultivation area in China. The purple-fruited pepper is rich in anthocyanins, which not only increases the ornamental nature of the pepper fruit but also benefits the human body. In this experiment, light-induced regulatory pathways and related specific regulators of anthocyanin biosynthesis were examined through integrative transcriptomic and metabolomic analysis. Results Results revealed that delphinium 3-O-glucoside significantly accumulated in light exposed surface of pepper fruit after 48 h as compared to shaded surface. Furthermore, through strand-specific sequencing technology, 1341 differentially expressed genes, 172 differentially expressed lncRNAs, 8 differentially expressed circRNAs, and 28 differentially expressed miRNAs were identified significantly different among both surfaces. The flavonoid synthesis pathway was significantly enriched by KEGG analysis including SHT (XM_016684802.1), AT-like (XM_016704776.1), CCoAOMT (XM_016698340.1, XM_016698341.1), CHI (XM_016697794.1, XM_016697793.1), CHS2 (XM_016718139.1), CHS1B (XM_016710598.1), CYP98A2-like (XM_016688489.1), DFR (XM_016705224.1), F3’5’H (XM_016693437.1), F3H (XM_016705025.1), F3’M (XM_016707872.1), LDOX (XM_016712446.1), TCM (XM_016722116.1) and TCM-like (XM_016722117.1). Most of these significantly enriched flavonoid synthesis pathway genes may be also regulated by lncRNA. Some differentially expressed genes encoding transcription factors were also identified including MYB4-like (XM_016725242.1), MYB113-like (XM_016689220.1), MYB308-like (XM_016696983.1, XM_016702244.1), and EGL1 (XM_016711673.1). Three ‘lncRNA-miRNA-mRNA’ regulatory networks with sly-miR5303, stu-miR5303g, stu-miR7997a, and stu-miR7997c were constructed, including 28 differentially expressed mRNAs and 6 differentially expressed lncRNAs. Conclusion Possible light regulated anthocyanin biosynthesis and transport genes were identified by transcriptome analysis, and confirmed by qRT-PCR. These results provide important data for further understanding of the anthocyanin metabolism in response to light in pepper.

Phytophthora capsici, a destructive fungal pathogen, poses a severe threat to pepper (Capsicum annuum L.) crops worldwide, causing blights that can result in substantial yield losses. Traditional control methods often come with environmental concerns or entail substantial time investments. In this research, we investigate an alternative approach involving ferrous sulfate (FeSO4) application to combat P. capsici and promote pepper growth. We found that FeSO4 effectively inhibits the growth of P. capsici in a dose-dependent manner, disrupting mycelial development and diminishing pathogenicity. Importantly, FeSO4 treatment enhances the biomass and resistance of pepper plants, mitigating P. capsici-induced damage. Microbiome analysis demonstrates that FeSO4 significantly influences soil microbial communities, particularly fungi, within the pepper root. Metabolomics data reveal extensive alterations in the redox metabolic processes of P. capsici under FeSO4 treatment, leading to compromised cell membrane permeability and oxidative stress in the pathogen. Our study presents FeSO4 as a promising and cost-effective solution for controlling P. capsici in pepper cultivation while simultaneously promoting plant growth. These findings contribute to a deeper understanding of the intricate interactions between iron, pathogen control, and plant health, offering a potential tool for sustainable pepper production.

Peppers (Capsicum annuum L.), as a horticultural crop with one of the highest ascorbic acid contents, are negatively affected by detrimental environmental conditions both in terms of quality and productivity. In peppers, the high level of ascorbic acid is not only a nutrient substance but also plays a role in environmental stress, i.e., drought stress. When suffering from drought stress, plants accumulate dehydrins, which play important roles in the stress response. Here, we isolated an SK3-type DHN gene CaDHN2 from peppers. CaDHN2 was located in the nucleus, cytoplasm, and cell membrane. In CaDHN2-silenced peppers, which are generated by virus-induced gene silencing (VIGS), the survival rate is much lower, the electrolytic leakage is higher, and the accumulation of reactive oxygen species (ROS) is greater when compared with the control under drought stress. Moreover, when CaDHN2 (CaDHN2-OE) is overexpressed in Arabidopsis, theoverexpressing plants show enhanced drought tolerance, increased antioxidant enzyme activities, and lower ROS content. Based on yeast two-hybrid (Y2H), GST-pull down, and bimolecular fluorescence complementation (BiFC) results, we found that CaDHN2 interacts with CaGGP1, the key enzyme in ascorbic acid (AsA) synthesis, in the cytoplasm. Accordingly, the level of ascorbic acid is highly reduced in CaDHN2-silenced peppers, indicating that CaDHN2 interacts with CaGGP1 to affect the synthesis of ascorbic acid under drought stress, thus improving the drought tolerance of peppers. Our research provides a basis for further study of the function of DHN genes.

To investigate the effect of fertilization on Pb content in vegetables, pepper was planted in L1645 (the 5 influencing factors are fertilizers (N, P, K), organic fertilizers (sheep manure) and Pb2+; the 4 levels are blank, low, medium and high; a total of 16 treatments) pot orthogonal experiment. The effects of fertilizers on the growth and Pb content in various parts of pepper under Pb stress were analyzed. The results showed that: (1) The Pb content in pepper fruit ranged from 0.011 mg·kg−1 to 0.085 mg·kg−1, which did not exceed the limit value (0.1 mg·kg−1) in the National Standard for Food Safety-Limit of Contaminants in Food (GB2762-2017); (2) The effect order of fertilization on pepper fruit weight was P2O5 > sheep manure > N > K2O; The horizontal combination of factors that promoted the maximum fruit weight of pepper was N (0.15 g·kg−1), P2O5 (0.225 g·kg−1), K2O (0.15 g·kg−1) and sheep manure (9 g·kg−1); (3) The order of fertilizer effects on Pb content in pepper fruit was Pb2+ > K2O > N = sheep manure > P2O5; the factor level combination that resulted in the maximum Pb content in pepper fruits was N (0.15 g·kg−1), P2O5 (0 g·kg−1), K2O (0.45 g·kg−1), sheep manure (6 g·kg−1) and Pb2+ (350 mg·kg−1); (4) Based on the soil fertility characteristics of Urumqi, the recommended optimal fertilizer application rate was: high phosphorus fertilizer P2O5 (495 kg·hm−2), low-level potassium fertilizer K2O (330 kg·hm−2), medium-level nitrogen fertilizer N (660 kg·hm−2) (or low-level nitrogen fertilizer N (330 kg·hm−2) + high-level organic manure sheep manure (19,800 kg·hm−2), which can achieve high yield while ensuring that the Pb content in the fruits does not exceed the standard. Strengthening control of effective and reasonable fertilization methods in Urumqi agricultural land is helpful to reduce the Pb content in vegetables.

Phosphorus utilization by crop plants is often limited, thereby resulting in large accumulations of residual phosphorus fertilizer in the soil. Trichoderma fungi function as natural decomposition agents that can contribute to increasing decomposition and promoting nutrient absorption in plants. In this study, we developed a novel fertilizer application strategy that reduces phosphate fertilizer and increases Trichoderma and examined its effects on the growth, nutrient absorption, and fruit quality of pepper (Capsicum annuum L.). We compared the efficacies of eight treatments: P100 = standard dose application of phosphorus fertilizer; P85 = 85% dose; P70 = 70% dose; P0 = no phosphorus fertilizer; and the TP100, TP85, TP70, and TP0 treatments, in which a Trichoderma mixture was added to the P100, P85, P70, and P0 treatments, respectively. The combined fertilizer application strategy stimulated plant growth, increased chlorophyll content, improved yield, and enhanced nutrient absorption. Additionally, the strategy improved pepper fruit quality by increasing the contents of soluble proteins, soluble sugars, vitamin C, capsaicin, and capsanthin. A comprehensive analysis indicated that the TP85 treatment was the optimal fertilization regime for pepper. This study provides a novel fertilizer application strategy for pepper that not only ensures good plant growth but also protects soil health.

Low temperature combined with low light (LL stress) is a typical environmental stress that limits peppers’ productivity, yield, and quality in northwestern China. Glycine betaine (GB), an osmoregulatory substance, has increasingly valuable effects on plant stress resistance. In this study, pepper seedlings were treated with different concentrations of GB under LL stress, and 20 mM of GB was the best treatment. To further explore the mechanism of GB in response to LL stress, four treatments, including CK (normal temperature and light, 28/18 °C, 300 μmol m−2 s−1), CB (normal temperature and light + 20 mM GB), LL (10/5 °C, 100 μmol m−2 s−1), and LB (10/5 °C, 100 μmol m−2 s−1 + 20 mM GB), were investigated in terms of pepper growth, biomass accumulation, photosynthetic capacity, expression levels of encoded proteins Capsb, cell membrane permeability, antioxidant enzyme gene expression and activity, and subcellular localization. The results showed that the pre-spraying of GB under LL stress significantly alleviated the growth inhibition of pepper seedlings; increased plant height by 4.64%; increased root activity by 63.53%; and decreased photoinhibition by increasing the chlorophyll content; upregulating the expression levels of encoded proteins Capsb A, Capsb B, Capsb C, Capsb D, Capsb S, Capsb P1, and Capsb P2 by 30.29%, 36.69%, 18.81%, 30.05%, 9.01%, 6.21%, and 16.45%, respectively; enhancing the fluorescence intensity (OJIP curves), the photochemical efficiency (Fv/Fm, Fv′/Fm′), qP, and NPQ; improving the light energy distribution of PSΠ (Y(II), Y(NPQ), and Y(NO)); and increasing the photochemical reaction fraction and reduced heat dissipation, thereby increasing plant height by 4.64% and shoot bioaccumulation by 13.55%. The pre-spraying of GB under LL stress also upregulated the gene expression of CaSOD, CaPOD, and CaCAT; increased the activity of the ROS-scavenging ability in the pepper leaves; and coordinately increased the SOD activity in the mitochondria, the POD activity in the mitochondria, chloroplasts, and cytosol, and the CAT activity in the cytosol, which improved the LL resistance of the pepper plants by reducing excess H2O2, O2−, MDA, and soluble protein levels in the leaf cells, leading to reduced biological membrane damage. Overall, pre-spraying with GB effectively alleviated the negative effects of LL stress in pepper seedlings.

Low temperature combined with low light (LL) is an adverse factor seriously affecting pepper productivity and quality. However, little is known about the molecular mechanisms related to LL stress responses. Therefore, transcriptome and physiological changes in Hangjiao No. 2 (H2) and Xiangtela No. 2 (X2) pepper were studied under normal conditions and LL. We found 8,392 and 8,028 differentially expressed genes in H2 and X2, respectively, significantly associated with photosynthesis, photosynthesis antenna proteins, and carotenoids, were enriched in 27 and 40 gene ontology terms in H2 and X2, respectively, and 14 and 16 Kyoto Encyclopedia of Gene and Genomes pathways. The accuracy and reliability of the RNA-Seq results were confirmed by qRT-PCR. Furthermore, carotenoid-related genes ZDS, CA1, CA2, NCED, LOC107840293, and LOC107850059 functioned in response to LL. Additionally, LL significantly decreased photosynthesis capacity, photosynthetic pigment contents, as well as maximum quantum efficiency, and changed carotenoid-related compounds, revealing photosynthesis and carotenoids were involved in LL stress response. Our findings provide insight into LL stress-induced transcriptional expression patterns.

Abstract Background The NAC (NAM, ATAF1/ATAF2, and CUC2) transcription factors belong to a large family of plant-specific transcription factors in monocot and dicot species. These transcription factors regulate the expression of stress tolerance-related genes that protect plants from various abiotic stresses, including drought, salinity, and low temperatures. Results In this study, we identified the CaNAC46 transcription factor gene in Capsicum annuum. Its open reading frame was revealed to comprise 921 bp, encoding a protein consisting of 306 amino acids, with an isoelectric point of 6.96. A phylogenetic analysis indicated that CaNAC46 belongs to the ATAF subfamily. The expression of CaNAC46 was induced by heat, cold, high salt, drought, abscisic acid, salicylic acid, and methyl jasmonate treatments. Thus, CaNAC46 may be important for the resistance of dry pepper to abiotic stresses. A subcellular localization analysis confirmed that CaNAC46 is localized in the nucleus. The overexpression of CaNAC46 improved the tolerance of transgenic Arabidopsis thaliana plants to drought and salt stresses. The CaNAC46-overexpressing lines had longer roots and more lateral roots than wild-type lines under prolonged drought and high salt stress conditions. Additionally, CaNAC46 affected the accumulation of reactive oxygen species (ROS). Moreover, CaNAC46 promoted the expression of SOD, POD, RD29B, RD20, LDB18, ABI, IAA4, and P5CS. The malondialdehyde contents were higher in TRV2-CaNAC46 lines than in wild-type plants in response to drought and salt stresses. Furthermore, the expression levels of stress-responsive genes, such as ABA2, P5CS, DREB, RD22, CAT, and POD, were down-regulated in TRV2-CaNAC46 plants. Conclusions Under saline and drought conditions, CaNAC46 is a positive regulator that activates ROS-scavenging enzymes and enhances root formation. The results of our study indicate CaNAC46 is a transcriptional regulator responsible for salinity and drought tolerance and suggest the abiotic stress-related gene regulatory mechanisms controlling this NAC transcription factor are conserved between A. thaliana and pepper.

Abstract Background Pepper mild mottle virus (PMMoV) is a member in the genus Tobamovirus and infects mainly solanaceous plants. However, the mechanism of virus-host interactions remains unclear. To explore the responses of pepper plants to PMMoV infection, we analyzed the transcriptomic changes in pepper plants after PMMoV infection using a high-throughput RNA sequencing approach and explored the roles of host autophagy in regulating PMMoV infection. Results A total of 197 differentially expressed genes (DEGs) were obtained after PMMoV infection, including 172 significantly up-regulated genes and 25 down-regulated genes. The Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses revealed that most up-regulated DEGs were involved in plant abiotic and biotic stresses. Further analyses showed the expressions of multiple autophagy-related genes (ATGs) were increased after PMMoV infection in pepper and Nicotiana benthamiana plants. Through confocal microscopy and transmission electron microscopy, we have found that PMMoV infection in plant can induce autophagy, evidenced by the increased number of GFP-ATG8a fluorescent punctate and the appearance of double membrane autophagic structures in cells of N. benthamiana. Additionally, inhibition of autophagy significantly increased PMMoV RNA accumulation and aggravated systemic PMMoV symptoms through autophagy inhibitor (3-MA and E64d) treatment and silencing of NbATG expressions by a Tobacco rattle virus-induced gene silencing assays. These results indicated that autophagy played a positive role in plant resistance to PMMoV infection. Conclusions Taken together, our results provide a transcriptomic insight into pepper responding to PMMoV infection and reveal that autophagy induced by PMMoV infection has an antiviral role in regulating PMMoV infection. These results also help us to better understand the mechanism controlling PMMoV infection in plants and to develop better strategies for breeding projects for virus-resistant crops.

Abstract Background The DnaJ proteins play critical roles in plant development and stress responses. Recently, seventy-six DnaJ genes were identified through a comprehensive bioinformatics analysis in the pepper genome. However, there were no reports on understanding of phylogenetic relationships and diverse expression profile of pepper DnaJ genes to date. Herein, we performed the systemic analysis of the phylogenetic relationships and expression profile of pepper DnaJ genes in different tissues and in response to both abiotic stress and plant hormones. Results Phylogenetic analysis showed that all the pepper DnaJ genes were grouped into 7 sub-families (sub-family I, II, III, IV, V, VI and VII) according to sequence homology. The expression of pepper DnaJs in different tissues revealed that about 38% (29/76) of pepper DnaJs were expressed in at least one tissue. The results demonstrate the potentially critical role of DnaJs in pepper growth and development. In addition, to gain insight into the expression difference of pepper DnaJ genes in placenta between pungent and non-pungent, their expression patterns were also analyzed using RNA-seq data and qRT-PCR. Comparison analysis revealed that eight genes presented distinct expression profiles in pungent and non-pungent pepper. The CaDnaJs co-expressed with genes involved in capsaicinoids synthesis during placenta development. What is more, our study exposed the fact that these eight DnaJ genes were probably regulated by stress (heat, drought and salt), and were also regulated by plant hormones (ABA, GA3, MeJA and SA). Conclusions In summary, these results showed that some DnaJ genes expressed in placenta may be involved in plant response to abiotic stress during biosynthesis of compounds related with pungency. The study provides wide insights to the expression profiles of pepper DanJ genes and contributes to our knowledge about the function of DnaJ genes in pepper.

Abstract Background Plant transcription factors (TFs) are key transcriptional regulators to manipulate the regulatory network of host immunity. However, the globally transcriptional reprogramming of plant TF families in response to pathogens, especially between the resistant and susceptible host plants, remains largely unknown. Results Here, we performed time-series RNA-seq from a resistant pepper line CM334 and a susceptible pepper line EC01 upon challenged with Phytophthora capsici, and enrichment analysis indicated that WRKY family most significantly enriched in both CM334 and EC01. Interestingly, we found that nearly half of the WRKY family members were significantly up-regulated, whereas none of them were down-regulated in the two lines. These induced WRKY genes were greatly overlapped between CM334 and EC01. More strikingly, most of these induced WRKY genes were expressed in time-order patterns, and could be mainly divided into three subgroups: early response (3 h-up), mid response (24 h-up) and mid-late response (ML-up) genes. Moreover, it was found that the responses of these ML-up genes were several hours delayed in EC01. Furthermore, a total of 19 induced WRKY genes were selected for functional identification by virus-induced gene silencing. The result revealed that silencing of CaWRKY03–6, CaWRKY03–7, CaWRKY06–5 or CaWRKY10–4 significantly increase the susceptibility to P. capsici both in CM334 and EC01, indicating that they might contribute to pepper’s basal defense against P. capsici; while silencing of CaWRKY08–4 and CaWRKY01–10 significantly impaired the disease resistance in CM334 but not in EC01, suggesting that these two WRKY genes are prominent modulators specifically in the resistant pepper plants. Conclusions These results considerably extend our understanding of WRKY gene family in pepper’s resistance against P. capsici and provide potential applications for genetic improvement against phytophthora blight.

Abstract Background Colletotrichum species are the causal agents of anthracnose, a major disease affecting the yield and quality of pepper (Capsicum spp.). Colletotrichum scovillei is widespread in China, has strong pathogenicity and drug resistance, and causes anthracnose disease in pepper fruits that severely reduces production. Previously, an anti-anthracnose locus AnR GO 5 was mapped to the P5 chromosome on the basis of analyses of fruit at the green mature stage. The aim of this study was to narrow down the interval of this locus and identify the gene responsible for conferring resistance. Results On the basis of results of re-sequencing of Capsicum chinense ‘PBC932’ and C. annuum ‘77013’, we developed Kompetitive allele-specific PCR (KASPar) markers and insertion–deletion (InDel) markers linked to AnR GO 5 at the green mature fruit stage and used them to construct a genetic linkage map (42 markers, 24.4 cM in length). Using data obtained in phenotypic and genotypic analyses of BC4S1, BC4S2, and BC4S3 populations, AnR GO 5 was located between the markers P5in-2266-404 and P5in-2268-978 within a physical distance of 164 kb. This region contained five genes, including CA05g17730. CA05g17730 encodes ‘R1C-3-like’ putative late blight resistance protein homologs. The transcript level of CA05g17730 differed between ‘PBC932’ and ‘77013’. The structure of the CA05g17730 gene also differed between ‘PBC932’ and ‘77013’. Conclusions We narrowed down the QTL interval to a region containing five genes. These results will be useful for further research on the mechanisms of resistance to anthracnose, and for marker assisted selection for anthracnose-resistant capsicum lines.

Chili anthracnose is one of the most common and destructive fungal pathogens that affects the yield and quality of pepper. Although WRKY proteins play crucial roles in pepper resistance to a variety of pathogens, the mechanism of their resistance to anthracnose is still unknown. In this study, we found that CaWRKY50 expression was obviously induced by Colletotrichum scovillei infection and salicylic acid (SA) treatments. CaWRKY50-silencing enhanced pepper resistance to C. scovillei, while transient overexpression of CaWRKY50 in pepper increased susceptibility to C. scovillei. We further found that overexpression of CaWRKY50 in tomatoes significantly decreased resistance to C. scovillei by SA and reactive oxygen species (ROS) signaling pathways. Moreover, CaWRKY50 suppressed the expression of two SA-related genes, CaEDS1 (enhanced disease susceptibility 1) and CaSAMT1 (salicylate carboxymethyltransferase 1), by directly binding to the W-box motif in their promoters. Additionally, we demonstrated that CaWRKY50 interacts with CaWRKY42 and CaMIEL1 in the nucleus. Thus, our findings revealed that CaWRKY50 plays a negative role in pepper resistance to C. scovillei through the SA-mediated signaling pathway and the antioxidant defense system. These results provide a theoretical foundation for molecular breeding of pepper varieties resistant to anthracnose.

Pepper, as a vegetable crop with a wide cultivation area worldwide, besides being a significant condiment and food, also has a momentous use for chemistry, medicine, and other industries. Pepper fruits are rich in various pigments, such as chlorophyll, carotenoids, anthocyanins, and capsanthin, which have important healthcare and economic value. Since various pigments are continuously metabolized during the development of pepper fruits, peppers exhibit an abundant fruit-colored phenotype in both the mature and immature periods. In recent years, great progress has been made in the study of pepper fruit color development, but the developmental mechanisms are still unclear systematically dissected in terms of pigment, biosynthesis, and regulatory genes. The article outlines the biosynthetic pathways of three important pigments: chlorophyll, anthocyanin, and carotenoid in pepper and the various enzymes involved in these pathways. The genetics and molecular regulation mechanisms of different fruit colors in immature and mature peppers were also systematically described. The objective of this review is to provide insights into the molecular mechanisms of pigments biosynthesis in pepper. This information will provide theoretical basis for the breeding of high-quality colored pepper varieties in the future.

Foliar fertilisation is an application technique that is increasingly being used in agriculture and offers the possibility of providing nutrients directly to the site of highest demand. Especially for phosphorus (P), foliar application is an interesting alternative to soil fertilisation, but foliar uptake mechanisms are poorly understood. To gain a better understanding of the importance of leaf surface features for foliar P uptake, we conducted a study with tomato (Solanum lycopersicum) and pepper (Capsicum annuum) plants, which have different leaf surface traits. For this purpose, drops of 200 mM KH2PO4 without surfactant were applied onto the adaxial or abaxial leaf side or to the leaf veins and the rate of foliar P absorption was evaluated after one day. Additionally, leaf surfaces were characterised in detail by transmission electron microscopy (TEM) and scanning electron microscopy (SEM), estimating also leaf surface wettability and free energy, among other parameters. While the leaves of pepper hardly contained any trichomes, the abaxial side and the leaf veins of tomato leaves were densely covered with trichomes. The cuticle of tomato leaves was thin (approximately 50 nm), while that of pepper was thick (approximately 150–200 nm) and impregnated with lignin. Due to the fact that trichomes were most abundant in the leaf veins of tomato, dry foliar fertiliser drop residues were observed to be anchored there, and the highest P uptake occurred via tomato leaf veins, resulting in 62% increased P concentration. However, in pepper, the highest rate of P absorption was recorded after abaxial-side P treatment (+66% P). Our results provide evidence that different leaf parts contribute unequally to the absorption of foliar-applied agrochemicals, which could potentially be useful for optimising foliar spray treatments in different crops.

Capsicum species grown for pepper production suffer severely from thrips damage, urging the identification of natural resistance. Resistance levels are commonly assessed on leaves. However, Capsicum plants are flower-bearing during most of the production season, and thrips also feed on pollen and flower tissues. In order to obtain a comprehensive estimate of elements contributing to thrips resistance, flower tissues should be considered as well. Therefore, we assessed resistance to Frankliniella occidentalis in flowers, leaves, and whole plants of ten Capsicum accessions. Using choice assays, we found that thrips prefer flowers of certain accessions over others. The preference of adult thrips for flowers was positively correlated to trehalose and fructose concentration in anthers as well as to pollen quantity. Resistance measured on leaf discs and thrips population development on whole plants was significantly and positively correlated. Leaf-based resistance thus translates to reduced thrips population development. Results of the flower assays were not significantly correlated with resistance in leaves or on whole plants. This suggests that both leaves and flowers represent a different part of the resistance spectrum and should both be considered for understanding whole plant resistance and the identification of resistant Capsicum varieties.

Small heat shock proteins (sHSPs) are usually upregulated in plants under diverse environmental stresses. These proteins have been suggested to function as molecular chaperones to safeguard other proteins from stress-induced damage. The ripening of pepper (Capsicum annuum L.) fruit involves important phenotypic, physiological, and biochemical changes, which have associated endogenous physiological nitro-oxidative stress, but they can also be significantly affected by environmental conditions, such as temperature. Based on the available pepper genome, a total of 41 sHSP genes were identified in this work, and their distributions in the 12 pepper chromosomes were determined. Among these genes, only 19 sHSP genes were found in the transcriptome (RNA-Seq) of sweet pepper fruits reported previously. This study aims to analyze how these 19 sHSP genes present in the transcriptome of sweet pepper fruits are modulated during ripening and after treatment of fruits with nitric oxide (NO) gas. The time-course expression analysis of these genes during fruit ripening showed that 6 genes were upregulated; another 7 genes were downregulated, whereas 6 genes were not significantly affected. Furthermore, NO treatment triggered the upregulation of 7 sHSP genes and the downregulation of 3 sHSP genes, whereas 9 genes were unchanged. These data indicate the diversification of sHSP genes in pepper plants and, considering that sHSPs are important in stress tolerance, the observed changes in sHSP expression support that pepper fruit ripening has an associated process of physiological nitro-oxidative stress, such as it was previously proposed.

Through the variation of experimental conditions, this paper examines advantages of enriched Pešter's peat used for a cultivation of pepper seedling. Peat is enriched with equal doses of complex mineral fertilizer NPK 15:15:15 and various doses of hydrated lime, zeolite, microelements and perlite. Different biological indicators of seedling have been observed.Best results are achieved by enriching Pester's peat with NPK mineral fertilizer or NPK mineral fertilizer and microelements. Adding different microelements didn't have a positive effect on plant length and number of leaves, but it affected a higher number of flower buds being developed.

Abstract Background The widely cultivated pepper (Capsicum spp.) is one of the most diverse vegetables; however, little research has focused on characterizing the genetic diversity and relatedness of commercial varieties grown in China. In this study, a panel of 92 perfect single-nucleotide polymorphisms (SNPs) was identified using re-sequencing data from 35 different C. annuum lines. Based on this panel, a Target SNP-seq genotyping method was designed, which combined multiplex amplification of perfect SNPs with Illumina sequencing, to detect polymorphisms across 271 commercial pepper varieties. Results The perfect SNPs panel had a high discriminating capacity due to the average value of polymorphism information content, observed heterozygosity, expected heterozygosity, and minor allele frequency, which were 0.31, 0.28, 0.4, and 0.31, respectively. Notably, the studied pepper varieties were morphologically categorized based on fruit shape as blocky-, long horn-, short horn-, and linear-fruited. The long horn-fruited population exhibited the most genetic diversity followed by the short horn-, linear-, and blocky-fruited populations. A set of 35 core SNPs were then used as kompetitive allele-specific PCR (KASPar) markers, another robust genotyping technique for variety identification. Analysis of genetic relatedness using principal component analysis and phylogenetic tree construction indicated that the four fruit shape populations clustered separately with limited overlaps. Based on STRUCTURE clustering, it was possible to divide the varieties into five subpopulations, which correlated with fruit shape. Further, the subpopulations were statistically different according to a randomization test and F st statistics. Nine loci, located on chromosomes 1, 2, 3, 4, 6, and 12, were identified to be significantly associated with the fruit shape index (p

Abstract Background The Bemisia tabaci is a major leaf feeding insect pest to pepper (Capsicum annuum), causing serious damage to pepper growth and yield. It is particularly important to study the mechanism of pepper resistance to B. tabaci, and to breed and promote the varieties of pepper resistant to B. tabaci. However, very limited molecular mechanism is available about how plants perceive and defend themselves from the destructive pest. Proteome technologies have provided an idea method for studying plant physiological processes in response to B. tabaci. Results Here, a highly resistant genotype and a highly susceptible genotype were exposed to B. tabaci feeding for 48 h to explore the defense mechanisms of pepper resistance to B. tabaci. The proteomic differences between both genotypes were compared using isobaric tag for relative and absolute quantification (iTRAQ). The quantitative data were validated by parallel reaction monitoring (PRM). The results showed that 37 differential abundance proteins (DAPs) were identified in the RG (resistant genotype), while 17 DAPs were identified in the SG (susceptible genotype) at 48 h after B. tabaci feeding. 77 DAPs were identified when comparing RG with SG without feeding. The DAP functions were determined for the classification of the pathways, mainly involved in redox regulation, stress response, protein metabolism, lipid metabolism and carbon metabolism. Some candidate DAPs are closely related to B. tabaci resistance such as annexin D4-like (ANN4), calreticulin-3 (CRT3), heme-binding protein 2-like (HBP1), acidic endochitinase pcht28-like (PR3) and lipoxygenase 2 (LOX2). Conclusions Taken together, this study indicates complex resistance-related events in B. tabaci interaction, provides novel insights into the molecular mechanism underlying the response of plant to B. tabaci, and identifies some candidate proteins against B. tabaci attack.

Research in plant pathology has increasingly focused on developing environmentally friendly, effective strategies for controlling plant diseases. Cyanobacteria, including Desmonostoc muscorum, Anabaena oryzae, and Arthrospiraplatensis, were applied to Capsicum annuum L. to induce immunity against Fusarium wilt. Soil irrigation and foliar shoots (FS) application were used in this investigation. The disease symptoms, disease index, osmotic contents, total phenol, Malondialdehyde (MDA), hydrogen peroxide (H2O2), antioxidant enzymes (activity and isozymes), endogenous hormone content, and response to stimulation of defense resistance in infected plants were assessed. Results demonstrated that using all cyanobacterial aqueous extracts significantly reduced the risk of infection with Fusarium oxysporum. One of the most effective ways to combat the disease was through foliar spraying with Arthrospira platensis, Desmonostoc muscorum, and Anabaena oryzae (which provided 95, 90, and 69% protection percent, respectively). All metabolic resistance indices increased significantly following the application of the cyanobacterial aqueous extracts. Growth, metabolic characteristics, and phenols increased due to the application of cyanobacteria. Polyphenol oxidase (PPO) and peroxidase (POD) expressions improved in response to cyanobacteria application. Furthermore, treatment by cyanobacteria enhanced salicylic acid (SA) and Indole-3-Acetic Acid (IAA) in the infected plants while decreasing Abscisic acid (ABA). The infected pepper plant recovered from Fusarium wilt because cyanobacterial extract contained many biologically active compounds. The application of cyanobacteria through foliar spraying seems to be an effective approach to relieve the toxic influences of F. oxysporum on infected pepper plants as green and alternative therapeutic nutrients of chemical fungicides.

Grafting is an important agricultural practice to control soil-borne diseases, alleviate continuous cropping problems and improve stress tolerance in vegetable industry, but it is relatively less applied in pepper production. A recent study has revealed the key roles of β-1, 4-glucanase in graft survival. We speculated that the GH9 family gene encoding glucanase may be involved in the obstacles of pepper grafting. Therefore, we performed a systematic analysis of the GH9 family in pepper, tomato and tobacco. A total of 25, 24 and 42 GH9 genes were identified from these three species. Compared with the orthologues of other solanaceous crops, the deduced pepper GH9B3 protein lacks a conserved motif (Motif 5). Promoter cis-element analysis revealed that a wound-responsive element exists in the promoter of tobacco NbGH9B3, but it is absent in the GH9B3 promoter of most solanaceous crops. The auxin-responsive related element is absent in CaGH9B3 promoter, but it presents in the promoter of tobacco, tomato, potato and petunia GH9B3. Tissue and induction expression profiles indicated that GH9 family genes are functionally differentiated. Nine GH9 genes, including CaGH9B3, were detected expressing in pepper stem. The expression patterns of NbGH9B3 and CaGH9B3 in grafting were different in our test condition, with obvious induction in tobacco but repression in pepper. Furthermore, weighted correlation network analysis (WGCNA) revealed 58 transcription factor genes highly co-expressed with NbGH9B3. Eight WRKY binding sites were detected in the promoter of NbGH9B3, and several NbWRKYs were highly co-expressed with NbGH9B3. In conclusion, the missing of Motif 5 in CaGH9B3, and lacking of wound- and auxin-responsive elements in the gene promoter are the potential causes of grafting-related problems in pepper. WRKY family transcription factors could be important regulator of NbGH9B3 in tobacco grafting. Our analysis points out the putative regulators of NbGH9B3, which would be helpful to the functional validation and the study of signal pathways related to grafting in the future.

Abstract Background Bacterial wilt caused by Ralstonia solanacearum is one of the most important diseases in pepper worldwide, however, the molecular mechanism underlying pepper resistance to bacterial wilt remains poorly understood. Results Herein, a novel RD leucine-rich repeat receptor-like kinase, CaLRR-RLK1, was functionally characterized in immunity against R. solanacearum. CaLRR-RLK1 was targeted exclusively to plasma membrane and was up-regulated by R. solanacearum inoculation (RSI) as well as by the exogenous application of salicylic acid (SA), methyl jasmonate (MeJA) or ethephon (ETH). The silencing of CaLRR-RLK1 led to enhanced susceptibility of pepper plants to RSI, accompanied by down-regulation of immunity-related genes including CaACO1, CaHIR1, CaPR4 and CaPO2. In contrast, transient overexpression of CaLRR-RLK1 triggered hypersensitive response (HR)-like cell death and H2O2 accumulation in pepper leaves, manifested by darker trypan blue and DAB staining respectively. In addition, the ectopic overexpression of CaLRR-RLK1 in tobacco plants enhanced resistance R. solanacearum, accompanied with the immunity associated marker genes including NtPR2, NtPR2, NtHSR203 and NtHSR515. Furthermore, it was found that CaHDZ27, a positive regulator in pepper response to RSI in our previous study, transcriptionally activated CaLRR-RLK1 by direct targeting its promoter probably in a CAATTATTG dependent manner. Conclusion The study revealed that CaLRR-RLK1 confers pepper resistance to R. solanacearum as the direct targeting of CaHDZ27.

Arbuscular mycorrhizal fungi (AMF) are well-known for their ability to improve plant growth and help plants withstand abiotic stress conditions. Unlike other fungi and bacteria, AMF cannot be stored, as they are obligate biotrophs. Long-term preservation of AMF spores is challenging and may lead to the loss of viability and efficiency. This study aimed to understand the effect of prolonged subculture of AMF species on the growth and glomalin-related soil protein (GRSP) from red pepper (Capsicum annuum L.). AMF spores were mass-produced using different techniques and subcultured in pots with sorghum sudangrass as the host plant for 3 years. Experimental soil samples were collected from natural grassland. Five different AMF inocula were used in triplicate as treatments. After 70 days of growth, red pepper plants were harvested and plant dry weight, plant nutrient content, mycorrhizal colonization, AMF spore count, and soil glomalin content were determined. AMF-treated plants displayed higher dry weight than controls, with only fruit dry weight being significantly different. Similarly, significant differences in phosphorous and potassium contents of the above-ground plant parts were observed between mycorrhizal and control treatments. In addition, soil GRSP content was significantly higher in plants inoculated with Rhizophagus sp. and Gigaspora margarita. The increased plant growth and GRSP content suggest that AMF can be maintained for 3 years without losing their efficiency if subcultured regularly with different symbiotic host plants.