Indian Institute of Spices Research

The genus Curcuma (family Zingiberaceae ) comprising over 80 species of rhizomatous herbs, is endowed with widespread adaptation from sea level to altitude as high as 2000 m in the Western Ghats and Himalayas. Having originated in the Indo-Malayan region, the genus is widely distributed in the tropics of Asia to Africa and Australia. Curcuma species exhibit inter- and intra-specific variation for the biologically active principles coupled with morphological variation with respect to the above-ground vegetative and floral characters as well as the below-ground rhizome features besides for curcumin, oleoresin and essential oil. Curcuma is gaining importance world over as a potential source of new drug(s) to combat a variety of ailments as the species contain molecules credited with anti-inflammatory, hypocholestraemic, choleratic, antimicrobial, insect repellent, antirheumatic, antifibrotic, antivenomous, antiviral, antidiabetic, antihepatotoxic as well as anticancerous properties. Turmeric oil is also used in aromatherapy and in the perfume industry. Though the traditional Indian Ayurvedic system of medicine and Chinese medicine long ago recognized the medicinal property of turmeric in its crude form, the last few decades have witnessed extensive research interests in the biological activity and pharmacological actions of Curcuma , especially the cultivated species. Turmeric powder obtained from rhizomes of Curcuma longa or related species is extensively used as a spice, food preservative and colouring material, in religious applications as well as a household remedy for bilary and hepatic disorders, anorexia, diabetic wounds, rheumatism and sinusitis in India, China and South-East Asia and in folk medicine. Cucuminoids, the biologically active principles from Curcuma , promise a potential role in the control of rheumatism, carcinogenesis and oxidative stress-related pathogenesis. Curcuma longa L. syn. Curcuma domestica Val., common turmeric, is the most economically valuable member of the genus having over 150,000 hectares under its cultivation in India. In addition to Curcuma longa , the other economically important species of the genus are C. aromatica , used in medicine and toiletry articles, C. kwangsiensis , C. ochrorhiza , C. pierreana , C. zedoaria , C. caesia etc. used in folk medicines of the South-East Asian nations; C. alismatifolia , C. roscoeana etc. with floricultural importance; Curcuma amada used as medicine, and in a variety of culinary preparations, pickles and salads, and C. zedoaria , C. malabarica , C. pseudomontana , C. montana , C. decipiens , C. angustifolia , C. rubescens , C. haritha , C. caulina etc. all used in arrowroot manufacturing. Crop improvement work has been attempted mainly in C. longa and to a little extent in C. amada . At present there are about 20 improved varieties of C. longa in India and one in C. amada , evolved through germplasm/clonal selection, mutation breeding or open-pollinated progeny (true turmeric seedlings) selection. Though work on morphological characterization of Curcuma species has been attempted, its molecular characterization is in a nascent stage except for some genetic fidelity studies of micropropagated plants and isozyme-based characterization. The genus has also been examined from the biochemical profiling and anatomical characterization angle. This article is intended to provide an overview of biological diversity in the genus Curcuma from a utilitarian and bio-prospection viewpoint.

AIM: To isolate and identify black pepper (Piper nigrum L) associated endophytic bacteria antagonistic to Phytophthora capsici causing foot rot disease. METHODS AND RESULTS: Endophytic bacteria (74) were isolated, characterized and evaluated against P. capsici. Six genera belong to Pseudomonas spp (20 strains), Serratia (1 strain), Bacillus spp. (22 strains), Arthrobacter spp. (15 strains), Micrococcus spp. (7 strains), Curtobacterium sp. (1 strain) and eight unidentified strains were isolated from internal tissues of root and stem. Three isolates, IISRBP 35, IISRBP 25 and IISRBP 17 were found effective for Phytophthora suppression in multilevel screening assays which recorded over 70% disease suppression in greenhouse trials. A species closest match (99% similarity) of IISRBP 35 was established as Pseudomonas aeruginosa (Pseudomonas EF568931), IISRBP 25 as P. putida (Pseudomonas EF568932), and IISRBP 17 as Bacillus megaterium (B. megaterium EU071712) based on 16S rDNA sequencing. CONCLUSION: Black pepper associated P. aeruginosa, P. putida and B. megaterium were identified as effective antagonistic endophytes for biological control of Phytophthora foot rot in black pepper. SIGNIFICANCE AND IMPACT OF THE STUDY: This work provides the first evidence for endophytic bacterial diversity in black pepper stem and roots, with biocontrol potential against P. capsici infection.

Badnaviruses (Family: Caulimoviridae; Genus: Badnavirus) are non-enveloped bacilliform DNA viruses with a monopartite genome containing about 7.2 to 9.2 kb of dsDNA with three to seven open reading frames. They are transmitted by mealybugs and a few species by aphids in a semi-persistent manner. They are one of the most important plant virus groups and have emerged as serious pathogens affecting the cultivation of several horticultural crops in the tropics, especially banana, black pepper, cocoa, citrus, sugarcane, taro, and yam. Some badnaviruses are also known as endogenous viruses integrated into their host genomes and a few such endogenous viruses can be awakened, e.g., through abiotic stress, giving rise to infective episomal forms. The presence of endogenous badnaviruses poses a new challenge for the fool-proof diagnosis, taxonomy, and management of the diseases. The present review aims to highlight emerging disease problems, virus characteristics, transmission, and diagnosis of badnaviruses.

Endophytic actinobacteria, which reside in the inner tissues of host plants, are gaining serious attention due to their capacity to produce a plethora of secondary metabolites (e.g. antibiotics) possessing a wide variety of biological activity with diverse functions. This review encompasses the recent reports on endophytic actinobacterial species diversity, in planta habitats and mechanisms underlying their mode of entry into plants. Besides, their metabolic potential, novel bioactive compounds they produce and mechanisms to unravel their hidden metabolic repertoire by activation of cryptic or silent biosynthetic gene clusters (BGCs) for eliciting novel secondary metabolite production are discussed. The study also reviews the classical conservative techniques (chemical/biological/physical elicitation, co-culturing) as well as modern microbiology tools (e.g. next generation sequencing) that are being gainfully employed to uncover the vast hidden scaffolds for novel secondary metabolites produced by these endophytes, which would subsequently herald a revolution in drug engineering. The potential role of these endophytes in the agro-environment as promising biological candidates for inhibition of phytopathogens and the way forward to thoroughly exploit this unique microbial community by inducing expression of cryptic BGCs for encoding unseen products with novel therapeutic properties are also discussed.

Abstract This article describes an efficient, relatively original method for the detection of extraneous Curcuma Sp. contamination in the powdered market samples of turmeric using molecular markers (RAPD), which are not easily discriminated by other analytical techniques routinely used for the identification of adulterants in powdered market samples of turmeric. Three market samples of turmeric powder studied revealed the presence of more Curcuma zedoaria (wild species) powder than Curcuma longa (the common culinary turmeric) powder, though the curcumin levels of the samples tallied with the quality standards prescribed for the commodity.

Plant diseases caused by viruses limit crop production and quality, resulting in significant losses. However, options for managing viruses are limited; for example, as systemic obligate parasites, they cannot be killed by chemicals. Sensitive, robust, affordable diagnostic assays are needed to detect the presence of viruses in plant materials such as seeds, vegetative parts, insect vectors, or alternative hosts and then prevent or limit their introduction into the field by destroying infected plant materials or controlling insect hosts. Diagnostics based on biological and physical properties are not very sensitive and are time-consuming, but assays based on viral proteins and nucleic acids are more specific, sensitive, and rapid. However, most such assays require laboratories with sophisticated equipment and technical skills. By contrast, isothermal-based assays such as loop-mediated isothermal amplification (LAMP) and recombinase polymerase amplification (RPA) are simple, easy to perform, reliable, specific, and rapid and do not require specialized equipment or skills. Isothermal amplification assays can be performed using lateral flow devices, making them suitable for onsite detection or testing in the field. To overcome non-specific amplification and cross-contamination issues, isothermal amplification assays can be coupled with CRISPR/Cas technology. Indeed, the collateral activity associated with some CRISPR/Cas systems has been successfully harnessed for visual detection of plant viruses. Here, we briefly describe traditional methods for detecting viruses and then examine the various isothermal assays that are being harnessed to detect viruses.

Value-added forms of black pepper (Piper nigrum L.) are an important item of trade globally. Adulteration by default or design of the commodity not only leads to economic loss and public health issues but also to self-respect of a nation. DNA barcoding is assuming significance as a quality assurance technique in many agri-food commodities. Three barcoding loci viz., psbA-trnH, rbcL, rpoC1 were used in the study to detect bio adulteration of traded black pepper powder. PCR amplification of P. nigrum and traded black pepper powder was performed for all the three loci. Sequence analysis and BLAST results revealed chilli adulteration in two out of nine market samples, originating probably from exhausted black pepper powder fortified with chilli. Of the three loci, psbA-trnH proved to be the best and ideal for detection of chilli adulteration in black pepper yielding amplicons of size 600 bp and 350 bp, respectively. Cloning and sequencing of the adulterant specific band of both market samples were done to confirm the results. It was further validated using simulated samples of chilli and black pepper powders in various proportions. The method proved efficient to detect adulteration even at very low levels (0.5% adulteration). HPLC analysis also supported the chilli adulteration of black pepper powder. The method is easy, reliable and efficient, and can be used by the regulatory agencies for quality assurance of black pepper powder.

Endophytic Pseudomonas aeruginosa strain BP35 was originally isolated from black pepper grown in the rain forest in Kerala, India. Strain PaBP35 was shown to provide significant protection to black pepper against infections by Phytophthora capsici and Radopholus similis. For registration and implementation in disease management programmes, several traits of PaBP35 were investigated including its endophytic behaviour, biocontrol activity, phylogeny and toxicity to mammals. The results showed that PaBP35 efficiently colonized black pepper shoots and displayed a typical spatiotemporal pattern in its endophytic movement with concomitant suppression of Phytophthora rot. Confocal laser scanning microscopy revealed high populations of PaBP35::gfp2 inside tomato plantlets, supporting its endophytic behaviour in other plant species. Polyphasic approaches to genotype PaBP35, including BOX-PCR, recN sequence analysis, multilocus sequence typing and comparative genome hybridization analysis, revealed its uniqueness among P. aeruginosa strains representing clinical habitats. However, like other P. aeruginosa strains, PaBP35 exhibited resistance to antibiotics, grew at 25-41°C and produced rhamnolipids and phenazines. PaBP35 displayed strong type II secretion effectors-mediated cytotoxicity on mammalian A549 cells. Coupled with pathogenicity in a murine airway infection model, we conclude that this plant endophytic strain is as virulent as clinical P. aeruginosa strains. Safety issues related to the selection of plant endophytic bacteria for crop protection are discussed.

Black pepper endophytic Pseudomonas putida BP25 produced diverse antimicrobial volatile organic compounds having potential for plant disease management. Chemically synthesised volatiles such as 2, 5-dimethyl pyrazine; 2-methyl pyrazine; dimethyl trisulphide; 2-ethyl 5-methyl pyrazine; and 2-ethyl 3, 6-dimethyl pyrazine showed inhibitory activity against oomycete pathogens, Phytophthora capsici & Pythium myriotylum; fungal pathogens, Rhizoctonia solani, Colletotrichum gloeosporioides, Athelia rolfsii, Gibberella moniliformis & Magnaporthe oryzae; bacterial pathogen, Ralstonia pseudosolanacearum and plant parasitic nematode, Radopholus similis. Among them, dimethyl trisulphide completely inhibited oomycete and fungal pathogens as well as R. similis at a concentration of 2.65 µg cm−3 under in vitro conditions. Pyrazines suppressed Phytophthora lesions on shoot cuttings of black pepper upon in planta volatile treatment. Dimethyl trisulphide was the only compound that exhibited soil fumigant activity against P. capsici, R. solani and A. rolfsii (6.25 µg cm−3), C. gloeosporioides and G. moniliformis (12.5 µg cm−3), and R. similis (50 µg cm−3). Altogether, endophytic Pseudomonas putida BP25 and its volatile organic compounds offer an alternative strategy for eco-friendly disease management in agriculture.

Ginger ( Zingiber officinale Rosc.), originated in the Indo-Malayan region, is now widely distributed across the tropics of Asia, Africa, America and Australia. It was domesticated in India and China, which represent the centre of origin of the species. Cultivated ginger though sterile, exhibits variations in rhizome and vegetative characters. The crop is gaining importance as a curative agent for a variety of ailments. Yield and quality traits (such as essential oil, fibre and oleoresin contents) along with volatile and non-volatile constituents are important determinants of the commodity's end product. Cultivar diversity for yield and morphological features is well known in ginger with few primitive types having excellent quality. However, the common name(s) of ginger cultivars are confusing and have resulted in geographical bias in its ex situ conservation. Most of the molecular/biochemical marker studies reported in ginger show low levels of polymorphism in contrast to plentiful phenotypic variability recorded in the species. The large influence of environmental factors on the content of key compounds, lack of seed set and confusion of the common names are the leading constraints to further varietal improvement of ginger. As tailor-made ginger varieties assume future significance these aspects will have relevance.

CONTEXT: In its powdered form, turmeric [Curcuma longa L. (Zingiberaceae)], a spice of medical importance, is often adulterated lowering its quality. OBJECTIVE: The study sought to detect plant-based adulterants in traded turmeric powder using DNA barcoding. MATERIALS AND METHODS: Accessions of Curcuma longa L., Curcuma zedoaria Rosc. (Zingiberaceae), and cassava starch served as reference samples. Three barcoding loci, namely ITS, rbcL, and matK, were used for PCR amplification of the reference samples and commercial samples representing 10 different companies. PCR success rate, sequencing efficiency, occurrence of SNPs, and BLAST analysis were used to assess the potential of the barcoding loci in authenticating the traded samples of turmeric. RESULTS: The PCR and sequencing success of the loci rbcL and ITS were found to be 100%, whereas matK showed no amplification. ITS proved to be the ideal locus because it showed greater variability than rbcL in discriminating the Curcuma species. The presence of C. zedoaria could be detected in one of the samples whereas cassava starch, wheat, barley, and rye in other two samples although the label claimed nothing other than turmeric powder in the samples. DISCUSSION AND CONCLUSION: Unlabeled materials in turmeric powder are considered as adulterants or fillers, added to increase the bulk weight and starch content of the commodity for economic gains. These adulterants pose potential health hazards to consumers who are allergic to these plants, lowering the product's medicinal value and belying the claim that the product is gluten free. The study proved DNA barcoding as an efficient tool for testing the integrity and the authenticity of commercial products of turmeric.

Microsatellite or Simple Sequence Repeat (SSR) markers have evolved to the status of a most versatile and popular genetic marker in a ubiquity of plant systems. Due to their co-dominant, hyper-variable and multiallelic nature, they are the prominent markers of choice for fingerprinting, conservation genetics, plant breeding and phylogenetic studies. Despite its development of a new set of SSR markers for a species remained time consuming and expensive for many years. However, with the recent advancement in genomics, new strategies/protocols are now available for the generation of SSR markers. This review presents an overview on microsatellite markers with a special emphasis on the various strategies used for the development of microsatellite markers

Spices are prime source for flavor, aroma, and taste in cuisines and play an active role as medicines due to their high antioxidant properties. As medicine or food, the importance of spices cannot be overemphasized. The medicinal values of spices are very well established in treating various ailments like cancer, fever, malaria, stomach offset, nausea, and many more. A spice may be available in several forms: fresh, whole dried, or pre-ground dried which requires further processing to be utilized in the form of value-added product. This review paper deals with the cultivation, postharvesting, chemical composition, uses, health, and medicinal benefits of the selected spice viz., black pepper, coriander, cinnamon, fenugreek, turmeric, and technological advances in processing of spices viz., super critical fluid extraction, cryogenic grinding, and microencapsulation etc. This paper also focuses on issues related to utilization of spices toward its high end-product development and characterization in pharmaceuticals and other medicinal purposes. The availability of different spices and their varietal differences and location have their pertinent characters, which are much demanding to refine postharvest and processing to assure its quality in the international market.

Bacterial wilt in ginger (Zingiber officinale Rosc.) caused by Ralstonia solanacearum is one of the most important production constraints in tropical, sub-tropical and warm temperature regions of the world. Lack of resistant genotype adds constraints to the crop management. However, mango ginger (Curcuma amada Roxb.), which is resistant to R. solanacearum, is a potential donor, if the exact mechanism of resistance is understood. To identify genes involved in resistance to R. solanacearum, we have sequenced the transcriptome from wilt-sensitive ginger and wilt-resistant mango ginger using Illumina sequencing technology. A total of 26387032 and 22268804 paired-end reads were obtained after quality filtering for C. amada and Z. officinale, respectively. A total of 36359 and 32312 assembled transcript sequences were obtained from both the species. The functions of the unigenes cover a diverse set of molecular functions and biological processes, among which we identified a large number of genes associated with resistance to stresses and response to biotic stimuli. Large scale expression profiling showed that many of the disease resistance related genes were expressed more in C. amada. Comparative analysis also identified genes belonging to different pathways of plant defense against biotic stresses that are differentially expressed in either ginger or mango ginger. The identification of many defense related genes differentially expressed provides many insights to the resistance mechanism to R. solanacearum and for studying potential pathways involved in responses to pathogen. Also, several candidate genes that may underline the difference in resistance to R. solanacearum between ginger and mango ginger were identified. Finally, we have developed a web resource, ginger transcriptome database, which provides public access to the data. Our study is among the first to demonstrate the use of Illumina short read sequencing for de novo transcriptome assembly and comparison in non-model species of Zingiberaceae.

The present study reports the development of a specific, sensitive, and reproducible Sequence Characterized Amplified Region (SCAR) marker to detect papaya seed powder adulteration in traded black pepper powder. A putative RAPD marker (449 bp) specific to papaya seed was identified, cloned, and sequenced to design the SCAR primers. This specific SCAR marker could detect the presence of papaya seed in all the analyzed simulated standards and in one of five branded market samples of black pepper powder tested. The analytical strategy being very simple could be used for large scale screening of powdered black pepper market samples intended for export and domestic uses.

Employing Illumina Hiseq whole genome metagenome sequencing approach, we studied the impact of Trichoderma harzianum on altering the microbial community and its functional dynamics in the rhizhosphere soil of black pepper (Piper nigrum L.). The metagenomic datasets from the rhizosphere with (treatment) and without (control) T. harzianum inoculation were annotated using dual approach, i.e., stand alone and MG-RAST. The probiotic application of T. harzianum in the rhizhosphere soil of black pepper impacted the population dynamics of rhizosphere bacteria, archae, eukaryote as reflected through the selective recruitment of bacteria [Acidobacteriaceae bacterium (p = 1.24e−12), Candidatus koribacter versatilis (p = 2.66e−10)] and fungi [(Fusarium oxysporum (p = 0.013), Talaromyces stipitatus (p = 0.219) and Pestalotiopsis fici (p = 0.443)] in terms of abundance in population and bacterial chemotaxis (p = 0.012), iron metabolism (p = 2.97e−5) with the reduction in abundance for pathogenicity islands (p = 7.30e−3), phages and prophages (p = 7.30e−3) with regard to functional abundance. Interestingly, it was found that the enriched functional metagenomic signatures on phytoremediation such as benzoate transport and degradation (p = 2.34e−4), and degradation of heterocyclic aromatic compounds (p = 3.59e−13) in the treatment influenced the rhizosphere micro ecosystem favoring growth and health of pepper plant. The population dynamics and functional richness of rhizosphere ecosystem in black pepper influenced by the treatment with T. harzianum provides the ecological importance of T. harzianum in the cultivation of black pepper.

Traded forms of spice and spice powders are often subjected to admixing with inferior substances by design or default, affecting public health and national prestige. Cinnamomum verum (true cinnamon), a high-value spice, is often adulterated with its inferior species such as C. cassia and C. malabatrum. The presence and detection of the spurious species in traded barks (whole or powder) of true cinnamon is posing problems. DNA markers are now used to detect such adulteration. Here we report the application of a DNA barcoding method to detect these adulterants in traded market samples of true cinnamon using the barcoding loci rbcL, matK and psbA-trnH. The PCR success rate, sequencing efficiency, inter and intra specific divergence, and occurrence of single nucleotide polymorphisms (SNPs) were utilized to assess the potential of each barcode loci to authenticate C. verum from its related adulterants. The amplification and sequencing success was 100% for rbcL and psbA-trnH while matK failed to amplify in the market samples. The locus of rbcL showed higher interspecific divergence while psbA-trnH exhibited lower interspecific divergence. SNPs specific to C. cassia were detected in rbcL locus in seven out of the ten market samples studied thereby confirming the presence of C. cassia adulteration in commercial samples of true cinnamon. Out of the three loci, rbcL locus proved to be efficient in tracing out adulterants in traded cinnamon. The SNP sites in this locus can be exploited in designing C. cassia specific primers, enabling kit development for easy detection of adulterants at the band level itself thereby bypassing the cost of sequencing.

Abstract The essential oil of the leaves of Artemisia nilagirica from South India was investigated by GC and GC/MS, which lead to the identification of 59 compounds including α-thujone (41.9%), borneol (10.8%), and β-thujone (9.1%). The leaf oil inhibited the growth of the plant pathogen Phytophthora capsici. This property has been attributed to the presence of thujones in the oil.

Abstract BACKGROUND Phytophthora infestans is a late blight‐causing oomycetes pathogen. It rapidly evolves and adapts to the host background and new fungicide molecules within a few years of their release, most likely because of the predominance of transposable elements in its genome. Frequent applications of fungicides cause environmental concerns. Here, we developed target‐specific RNA interference (RNAi)‐based molecules, along with nanoclay carriers, that when sprayed on plants are capable of effectively reducing late blight infection. RESULTS Targeted the genes unique to sporulation, early satge infection and the metabolism pathway stages based on in an our own microarray data. We used nanoclay as a carrier for sorbitol dehydrogenase, heat shock protein 90, translation elongation factor 1‐α, phospholipase‐D like 3 and glycosylphosphatidylinositol‐anchored acidic serine–threonine‐rich HAM34‐like protein double‐stranded (ds)RNAs, which were assessed by culture bioassay, detached leaf assay and spray methods, and revealed a reduction in growth, sporulation and symptom expression. Plants sprayed with multigene targeted dsRNA–nanoclay showed enhanced disease resistance (4% disease severity) and less sporulation (<1 × 10 3 ) compared with plants sprayed with dsRNA alone. CONCLUSION The use of nanoclay with multigene targeted dsRNA was assumed to be involved in effective delivery, protection and boosting the action of RNAi as a spray‐induced gene silencing approach (SIGS). A significant reduction in growth, sporulation, disease severity and decreased gene expression authenticates the effects of SIGS on late blight progression. This study demonstrated as a proof of concept the dsRNA–nanoclay SIGS approach, which could be used as an alternative to chemical fungicides and transgenic approaches to develop an environmentally friendly novel plant protection strategy for late blight. © 2022 Society of Chemical Industry.

Smut caused by Sporisorium scitamineum is one of the important diseases of sugarcane with global significance. Despite the intriguing nature of sugarcane, S. scitamineum interaction, several pertinent aspects remain unexplored. This study investigates the proteome level alterations occurring in the meristem of a S. scitamineum infected susceptible sugarcane cultivar at whip emergence stage. Differentially abundant proteins were identified by 2DE coupled with MALDI-TOF/TOF-MS. Comprehensively, 53 sugarcane proteins identified were related to defence, stress, metabolism, protein folding, energy, and cell division; in addition, a putative effector of S. scitamineum, chorismate mutase, was identified. Transcript expression vis-à-vis the activity of phenylalanine ammonia lyase was relatively higher in the infected meristem. Abundance of seven candidate proteins in 2D gel profiles was in correlation with its corresponding transcript expression levels as validated by qRT-PCR. Furthermore, this study has opened up new perspectives on the interaction between sugarcane and S. scitamineum.