Murali Gopal

Multifarious beneficial traits and plant growth promoting potential of Serratia marcescens KiSII and Enterobacter sp. RNF 267 isolated from the rhizosphere of coconut palms (Cocos nucifera L.).

Two plant growth promoting bacteria designated as KiSII and RNF 267 isolated from the rhizosphere of coconut palms were identified as Serratia marcescens and Enterobacter sp. based on their phenotypic features, BIOLOG studies and 16S rRNA gene sequence analysis. Both bacteria exhibited phosphate solubilization, ammonification, and production of indole acetic acid, β-1, 3 glucanase activities and 1-aminocyclopropane-1-carboxylate-deaminase activity. They could also tolerate a range of pH conditions, low temperature and salinity (NaCl). In addition, S. marcescens KiSII exhibited N- fixation potential, chitinase activity, siderophore production and antibiotics production. Seed bacterization with these bacteria increased the growth parameters of test plants such as paddy and cowpea over uninoculated control in green house assay. In coconut seedlings, significant increase in growth and nutrient uptake accompanied with higher populations of plant beneficial microorganisms in their rhizospheres were recorded on inoculation with both the PGPRs. The present study clearly revealed that PGPRs can aid in production of healthy and vigorous seedlings of coconut palm which are hardy perennial crops. They offer a scope to be developed into novel PGPR based bioinoculants for production of elite seedlings that can benefit the coconut farming community and the coconut based ecology.

Bespoke microbiome therapy to manage plant diseases

Information gathered with advanced nucleotide sequencing technologies, small molecule detection systems and computational biology is revealing that a community of microbes and their genes, now termed “the microbiome,” located in gut and rhizosphere, is responsible for maintaining the health of human beings and plants, respectively. Within the complete microbiome a “core-microbiome” exists that plays the pivotal role in well being of humans and plants. Recent studies in medicine have shown that an artificial mixture of bacteria representing the core gut microbiome of healthy person when transferred into gut of diseased person results in re-establishment of normal microflora in the latter leading to alleviation from diseased condition. In agriculture, though not exactly in similar manner as in medicine, success in plant disease management has been achieved through transfer of microbiome by mixing disease suppressive soils with disease conducive soils. A study more similar to artificial gut microbiome transfer in medical field has been recently reported in agriculture, in which transfer of microbiome via soil solutions (filtered and unfiltered) has shown ability to alleviate drought stress in Arabidopsis thaliana. However, the exact practice of transferring artificially cultivated core-microbiome as in medicine has not thus far been attempted in plant disease management. Nonetheless, as the gut and rhizosphere microbiome are known to share many common traits, there exists a good scope for accomplishing similar studies in agriculture. Based upon the information drawn from all recent works in microbiome studies of gut and rhizosphere, we propose that tailor-made core-microbiome transfer therapy can be a success in agriculture too and it could become a viable strategy for management of plant diseases in future.

Microbiome Selection Could Spur Next-Generation Plant Breeding Strategies

“No plant is an island too…” Plants, though sessile, have developed a unique strategy to counter biotic and abiotic stresses by symbiotically co-evolving with microorganisms and tapping into their genome for this purpose. Soil is the bank of microbial diversity from which a plant selectively sources its microbiome to suit its needs. Besides soil, seeds, which carry the genetic blueprint of plants during trans-generational propagation, are home to diverse microbiota that acts as the principal source of microbial inoculum in crop cultivation. Overall, a plant is ensconced both on the outside and inside with a diverse assemblage of microbiota. Together, the plant genome and the genes of the microbiota that the plant harbors in different plant tissues, i.e., the ‘plant microbiome,’ form the holobiome which is now considered as unit of selection: ‘the holobiont.’ The ‘plant microbiome’ not only helps plants to remain fit but also offers critical genetic variability, hitherto, not employed in the breeding strategy by plant breeders, who traditionally have exploited the genetic variability of the host for developing high yielding or disease tolerant or drought resistant varieties. This fresh knowledge of the microbiome, particularly of the rhizosphere, offering genetic variability to plants, opens up new horizons for breeding that could usher in cultivation of next-generation crops depending less on inorganic inputs, resistant to insect pest and diseases and resilient to climatic perturbations. We surmise, from ever increasing evidences, that plants and their microbial symbionts need to be co-propagated as life-long partners in future strategies for plant breeding. In this perspective, we propose bottom–up approach to co-propagate the co-evolved, the plant along with the target microbiome, through – (i) reciprocal soil transplantation method, or (ii) artificial ecosystem selection method of synthetic microbiome inocula, or (iii) by exploration of microRNA transfer method – for realizing this next-generation plant breeding approach. Our aim, thus, is to bring closer the information accrued through the advanced nucleotide sequencing and bioinformatics in conjunction with conventional culture-dependent isolation method for practical application in plant breeding and overall agriculture.

Aflatoxin production by Aspergillus flavus isolates pathogenic to coconut insect pests

Aspergillus flavus isolated from naturally infected leaf-eating caterpillar (Opisina arenosella W.), lace bug (Stephanitis typica D.) and plant hopper (Proutista moesta Westwood), insect pests of the coconut palm, were tested for aflatoxin (AT) production by employing various media formulations. These A. flavus isolates were earlier found to be entomopathogenic in laboratory bioassays. A laboratory contaminant and four standard aflatoxigenic A. flavus isolates were also included in this study as reference strains. All A. flavus isolates were tested on seven AT detection media: coconut extract agar, coconut extract-sodium desoxycholate agar, coconut extract-ascorbic acid agar, coconut extract-Czapek Dox agar, coconut extract-milk powder agar, 10% commercial coconut milk powder agar (CCMPA) and 20% CCMPA. Only two isolates of A. flavus, originally isolated from O. arenosella and P. moesta, produced ATs. AT production was detected within 48 h of incubation and was detected continually up to 1 month. These AT-producing A. flavus isolates also produced bright yellow pigmentation in the medium. Of all the seven media used for AT detection, CCMPA (10%) was found to be the best one, followed by 20% CCMPA, for direct and rapid AT detection. AT production was not necessary for pathogenicity in the insects.

Whole Genome Sequencing and Analysis of Plant Growth Promoting Bacteria Isolated from the Rhizosphere of Plantation Crops Coconut, Cocoa and Arecanut

Coconut, cocoa and arecanut are commercial plantation crops that play a vital role in the Indian economy while sustaining the livelihood of more than 10 million Indians. According to 2012 Food and Agricultural organizations report, India is the third largest producer of coconut and it dominates the production of arecanut worldwide. In this study, three Plant Growth Promoting Rhizobacteria (PGPR) from coconut (CPCRI-1), cocoa (CPCRI-2) and arecanut (CPCRI-3) characterized for the PGP activities have been sequenced. The draft genome sizes were 4.7 Mb (56% GC), 5.9 Mb (63.6% GC) and 5.1 Mb (54.8% GB) for CPCRI-1, CPCRI-2, CPCRI-3, respectively. These genomes encoded 4056 (CPCRI-1), 4637 (CPCRI-2) and 4286 (CPCRI-3) protein-coding genes. Phylogenetic analysis revealed that both CPCRI-1 and CPCRI-3 belonged to Enterobacteriaceae family, while, CPCRI-2 was a Pseudomonadaceae family member. Functional annotation of the genes predicted that all three bacteria encoded genes needed for mineral phosphate solubilization, siderophores, acetoin, butanediol, 1-aminocyclopropane-1-carboxylate (ACC) deaminase, chitinase, phenazine, 4-hydroxybenzoate, trehalose and quorum sensing molecules supportive of the plant growth promoting traits observed in the course of their isolation and characterization. Additionally, in all the three CPCRI PGPRs, we identified genes involved in synthesis of hydrogen sulfide (H2S), which recently has been proposed to aid plant growth. The PGPRs also carried genes for central carbohydrate metabolism indicating that the bacteria can efficiently utilize the root exudates and other organic materials as energy source. Genes for production of peroxidases, catalases and superoxide dismutases that confer resistance to oxidative stresses in plants were identified. Besides these, genes for heat shock tolerance, cold shock tolerance and glycine-betaine production that enable bacteria to survive abiotic stress were also identified.

Control of the coconut pest Oryctes rhinoceros L. using the Oryctes virus

The coconut palm is an important plantation crop in India, where it is cultivated on 1.796 million hectares. The rhinoceros beetle, Oryctes rhinoceros L. (Coleoptera: Scarabaeidae) is a serious pest of coconut throughout India and southeast Asia, causing an estimated 10% yield loss in the crop. Successful biological control of this pest could be achieved using the non-occluded Oryctes virus (syn. Baculovirus oryctes or Oryctes baculovirus). This review provides an account of this microbial agent, its biology, effects and impact, production and maintenance and alternative hosts, particularly in the context of the Indian situation. It also proposes future areas for investigation on the virus, in order to achieve its commercial viability and more widespread use.RésuméLe palmier cocotier est une importante culture de plantation en Inde, où il est cultivé sur 1,796 millions d’hectares. Le scarabée rhinocéros, Oryctes rhinoceros L. (Coleoptera: Scarabaeidae) est un sérieux ravageur du cocotier en Inde et en Asie du Sud Est, responsable de pertes de récolte estimées à 10%. Un contrôle biologique efficace de ce ravageur pourrait être obtenu en utilisant le virion non inclus du virus d’Oryctes (syn. Baculovirus oryctes ou baculovirus de l’Oryctes). Cette revue bibliographique fait le point sur cet agent microbien, sa biologie, son efficacité et son impact, sa production et sa maintenance ainsi que sur ses hôtes alternatifs, en particulier dans le contexte de l’Inde. Elle propose également de futurs domaines de recherches sur ce virus, afin d’aboutir à sa viabilité commerciale et à une plus grande utilisation.

Distribution of Arbuscular Mycorrhizae Associated with Coconut and Arecanut Based Cropping Systems

Mycorrhizal fungi are widespread in agricultural systems and are especially relevant for organic agriculture. A study was conducted to assess diversity in the arbuscular mycorrhizal (AM) fungi associated with coconut and arecanut intercropping systems of Kasaragod and Thiruvananthapuram districts of Kerala. Mycorrhizal parameters like spore density, root colonization, species richness, and relative occurrence of species were recorded. Coconut cropping system was found to be densely vegetated with diverse AM fungi, Glomus spp., Gigaspora spp., and Acaulospora spp. The diversity of fungal species was found to be maximum in the high density multiple species cropping system plot of CPCRI, Kasaragod with coconut as the main crop and banana, pepper as intercrops and least in farmers’ plot of Mogral Puthur, Kasaragod with arecanut as main crop and banana as the intercrop. A total of fourteen AM fungi were identified from coconut and arecanut intercropping systems. Glomusaggregatum, Funneliformis mosseae, and Rhizophagus fasiculatum were observed in both the cropping systems studied. Among the six edaphic factors analysed, an inverse relationship of mycorrhizal population with soil pH and soil phosphorous was observed. An abundance of P-solubilizers, fluorescent Pseudomonas spp., N-fixers, Bacillus spp., and Trichoderma spp. were observed in the rhizosphere, with Bacillus spp. showing the maximum association with AM fungi. The higher AMF colonization in coconut cropping system imply that the AM fungi colonization is primarily dependent on the host plant rather than the climatic/edaphic components of an area.

Diversity, richness and degree of colonization of arbuscular mycorrhizal fungi in coconut cultivated along with intercrops in high productive zone of Kerala, India

Degree of arbuscular mycorrhizal (AM) association and its diversity were investigated in the coconut palm (Cocos nucifera L.) cultivated in crop mixed system under rain-fed condition in a highly productive humid tropical zone in Malappuram district of Kerala, India. Forty AM species belonging to ten genera viz. Acaulospora, Claroideoglomus, Dentiscutata, Diversispora, Funneliformis, Gigaspora, Glomus, Redeckera, Scutellospora and Septoglomus were recorded indicating high level of AM richness in coconut rhizosphere. Of these, eighteen AM fungal species are being reported for first time from this palm. Claroideoglomus, Glomus and Gigaspora were the most commonly present genera and Claroideoglomus etunicatum and Glomus aggregatum the common species. The diversity indices (Shannon indices values) of AM fungi associated with coconut palm in a crop mixed system and mono crop varied significantly. The Shannon index, Simpson’s index and evenness ranged from 1.40 ± 0.10 to 2.70 ± 0.01, 0.65 ± 0.033 to 0.90 ± 0.004, 0.51 ± 0.01 to 0.82 ± 0.01, respectively. Correlation between soil physio chemical characters and mycorrhizal parameters were worked out. Soil pH had negative correlation with spore count, root colonization, species richness, Shannon H and Simpson indices but had positive correlation with species evenness. Electrical conductivity showed positive correlation with root mycorrhizal colonization which indicated active AMF mediated uptake of nutrients.

Microbial pathogens of the coconut pest Oryctes rhinoceros: influence of weather factors on their infectivity and study of their coincidental ecology in Kerala, India

The rhinoceros beetle, Oryctes rhinoceros L., is an economically important pest of the coconut palm. Management of this pest has been accomplished using microbial agents viz., Oryctes virus (OrV) and an entomofungal pathogen Metarhizium anisopliae. Recently an opportunistic bacterial pathogen Pseudomonas alcaligenes has also been noticed to cause septicaemia in the grubs when under stress. To unravel the influence of abiotic weather factors and the interactions amongst these microbial pathogens, a 3 year study was conducted from September 1996 to August 1999 in three of the southern districts of Kerala, India. Of the 6627 grubs and 307 adults collected from various breeding sites of the pest, 5% of the grubs and 22% of the adults had natural virus infection, 3% larvae died of M. anisopliae mycosis and 20% larvae succumbed to bacterial septicaemia. Oryctes virus infection in grubs and adults was negatively correlated to minimum temperature (correlation co-efficient, r = −0.4, and −0.6 respectively, sample size, n = 0). Increase in relative humidity increased the fungal activity (r = 0.8) whereas, maximum temperature had a negative impact (r = −0.7). Occurrence of virus infection in grubs and adults was positively correlated (r = 0.6), supporting the contention of active transmission of the virus pathogen between these two stages. The bacterial septicaemia in the grubs was marginally correlated with virus infection and P. alcaligenes undermined the efficiency of the virus pathogen.

An Opportunistic Bacterial Pathogen, Pseudomonas alcaligenes , May Limit the Perpetuation of Oryctes Virus, a Biocontrol Agent of Oryctes rhinoceros L.

Pseudomonas alcaligenes was detected at a high concentration (10 9 -10 10 cells mL -1 ) in the haemolymph of some dead Oryctes rhinoceros grubs collected from its breeding sites in the three southern districts, viz. Alleppey, Quilon and Kottayam of Kerala State, India. In a laboratory colony maintained for production of Oryctes virus, an important biocontrol agent of this major coconut pest, approximately 52% of the grubs succumbed to septicaemia with similar symptoms. The bacterium was found to be a component of the gut microflora of healthy grubs. Occurrence of the viral infection naturally or when induced in the laboratory in the O. rhinoceros grubs, appeared to be one of the biotic stress factors for P.alcaligenes to become an opportunistic pathogen. A preponderance of this bacterial infection in field populations during the periods when natural viral infection in grubs was above average, agrees with this observation. This finding becomes significant as infection by the opportunistic bacterial pathogen, P. alcaligenes , reduces the production of Oryctes virus inoculum in nature and limits the field-perpetuation of this viral biocontrol agent.