Red Ring Disease Research Articles

A multi-omics deconstruction of red ring disease in oil palm

A multi-omics deconstruction of red ring disease in oil palm

Molecular characterisation and diagnostics of the red ring nematode, Bursaphelenchus cocophilus, from Mexico

Summary The red ring nematode, Bursaphelenchus cocophilus, vectored by the South American palm weevil, Rhynchophorus palmarum, is the causal agent of red ring disease in coconut and other palms in countries of Central and South America. The populations of B. cocophilus collected in the states of Guerrero and Tabasco, Mexico, were molecularly characterised using the D2-D3 expansion segments of 28S rRNA, ITS rRNA and COI gene sequences. Mexican B. cocophilus populations were molecularly different from other Central and South American populations. Comparative analysis of available rRNA sequences obtained from several countries showed that B. cocophilus consists of molecularly different populations and its species structure is likely congruent with that of the beetle vector. Conventional PCR, real-time PCR and recombinase polymerase amplification (RPA) with lateral flow dipstick (LF) assays have been developed for the identification of the red ring nematode in this study. The specificity of the ITS rRNA primers in the assays were examined using B. cocophilus and other 15 species of family Aphelenchoididae. Detection sensitivity levels, determined by using a dilution series of B. cocophilus extracts, was 0.13 nematode per reaction tube for real-time PCR and 0.25 nematode per reaction tube for conventional PCR and LF-RPA assays. The application of the LF-RPA assay has great potential for diagnosing infestation of this species with a minimal laboratory infrastructure.

Molecular identification of Bursaphelenchus cocophilus associated to oil palm (Elaeis guineensis) crops in Tibu (North Santander, Colombia).

The red ring nematode (Bursaphelenchus cocophilus (Cobb) Baujard 1989) has been registered in oil palm crops in the North, Central and Eastern zones of Colombia. In Tibu (North Santander), there are doubts regarding the diagnostic and identity of the disease. Oil palm crops in Tibu with the external and internal symptoms were inspected, and tissue samples were taken from different parts of the palm. The refrigerated samples were carried to the laboratory of Oleoflores in Tibu for processing. The light microscopy was used for the quantification and morphometric identification of the nematodes. Specimens of the nematode were used for DNA extraction, to amplify the segment D2-D3 of the large subunit of ribosomal RNA (28S) and perform BLAST and a phylogeny study. The most frequently symptoms were chlorosis of the young leaves, thin leaflets, collapsed, and dry lower leaves, beginning of roughening, accumulation of arrows and short leaves. Bursaphelenchus, was recovered in most of the tissues from the samples analyzed: stem, petiole bases, inflorescences, peduncle of bunches, and base of arrows in variable populations. The morphometric data and sequences obtained for the segment D2-D3 confirms to B. cocophilus as the causal agent of red ring disease on oil palms in the study area. For our knowledge, this study reports the first partial sequences of segment D2-D3 of B. cocophilus in oil palm in Colombia.

Generating the Baseline in the Early Detection of Bud Rot and Red Ring Disease in Oil Palms by Geospatial Technologies

Oil palm cultivation in Ecuador is important for the agricultural sector. As a result of it, the country generates sources of employment in some of the most vulnerable zones; it contributes 0.89% of the gross domestic product and 4.35% of the agricultural gross domestic product. In 2017, a value of USD $252 million was generated by exports, and palm contributed 4.53% of the agricultural gross domestic product (GDP). It is estimated that 125,000 hectares of palm were lost in the Republic of Ecuador due to Red Ring Disease (RRD) and specifically Bud Rot (BR). The current study aimed to generate an early detection of BR and RRD in oil palm. Image acquisition has been performed using Remotely Piloted Aircraft System (RPAS) with Red, Green, and Blue (RGB) cannons, and multispectral cameras, in study areas with and without the presence of the given disease. Hereby, we proposed two phases. In phase A, a drone flight has been conducted for processing and georeferencing. This allowed to obtain an orthomosaic that serves as input for obtaining several vegetation indices of the healthy crop. The data and products obtained from this phase served as a baseline to perform comparisons with plantations affected by BR and RRD and to differentiate the palm varieties that are used by palm growers. In phase B, the same process has been applied three times with an interval of 15 days in an affected plot, in order to identify the symptoms and the progress of them. A validation for the diseases detection has been performed in the field, by taking Global Positioning System (GPS) points of the palms that presented symptoms of BR and RRD, through direct observation by field experts. The inputs obtained in each monitoring allowed to analyze the spatial behavior of the diseases. The values of the vegetation indices obtained from Phase A and B aimed to establish the differences between healthy and diseased palms, with the purpose of generating the baseline of early responses of BR and RRD conditions. However, the best vegetation index to detect the BR was the Visible Atmospherically Resistant Index (VARI).

The First Detection of Metamasius hemipterus (L.) (Coleoptera: Curculionidae) from Coastal Mississippi, USA

Specimens of Metamasius hemipterus (L.) were collected from a nursery in coastal Mississippi during invasive insect pest monitoring by the Mississippi State University Invasive Insect Trapping Team. Specimens were collected during a USDA CAPS palm commodity survey occurring at nurseries and natural areas across the southern half of the state. Within the United States, Metamasius hemipterus is recognized as a pest of sugarcanes, bananas, and palms in Florida. It is also a pest in a variety of economically important fruits in its native range in the Antilles, Central, and South America. It has been implicated as a minor vector of red ring disease in palm, caused by the nematode Bursaphelenchus cocophilus (Cobb). This exotic insect poses a threat to the horticulture industry in Mississippi by vectoring pathogens injurious to palms and damaging other horticultural products.

Dynamic Monitoring Method of Coconut Red Ring Disease based on Apriori Algorithm

Dynamic Monitoring Method of Coconut Red Ring Disease based on Apriori Algorithm

Aggregation Pheromone of the Bearded Weevil, Rhinostomus barbirostris (Coleoptera: Curculionidae): Identification, Synthesis, Absolute Configuration and Bioactivity.

The bearded weevil, Rhinostomus barbirostris (Coleoptera: Curculionidae: Dryophthorinae), attacks coconut trees, oil palms and other species of Arecaceae. Besides direct damage, R. barbirostris may be a vector of diseases in coconut and oil palms, such as stem bleeding (resinosis) and red ring disease. Currently, the only method to control this weevil is by visual observation of damage and removal of infected plants. Semiochemical-based trapping could improve the effectiveness of monitoring and management of R. barbirostris. In comparisons of volatiles released by R. barbirostris males and females by gas chromatography (GC) two male-specific compounds were observed. GC-mass spectrometry (MS) and GC-Fourier transform-infrared (FTIR) analyses of the natural compounds suggested these were diastereoisomers of 5-hydroxy-4-methylheptan-3-one, also known as sitophilure, a pheromone component of other dryophthorine species. Synthesis of the mixture of all four stereoisomers of sitophilure was performed in two steps, and the chemical structures were confirmed by comparing GC retention times and MS and FTIR spectra of natural and synthetic compounds. The absolute configurations of the two male-specific compounds were elucidated by enantioselective GC; the major component was the (4S,5R)-isomer, and the minor component (4S,5S)-sitophilure. In analyses by GC-electroantennography (EAG) the antennae of male and female R. barbirostris only responded to the (4S,5R)-isomer of the synthetic sitophilure. The stereoisomeric mixture of sitophilure was attractive to both sexes of R. barbirostris in laboratory experiments in the presence of sugar cane volatiles, and a similar result was obtained in a preliminary field trapping test.

Persistence of the red ring nematode in the roots and in the rhizosphere of recently eradicated coconut palms

Red ring disease is one of the primary diseases that attacks the coconut palm (Cocos nucifera L.) in Brazil. This lethal disease is caused by a nematode (Bursaphelenchus cocophilus) and can spread via contaminated tools or by the South American palm weevil (Rhynchophorus palmarum). The goal of this research was to determine the persistence of nematodes within the roots and the rhizosphere of recently eradicated coconut palms, cultivated in the coastal region of the state of Alagoas. Soil and root samples from coconut palms with advanced symptoms of red ring disease were collected up to 6 months after the eradication of the plants. Sample processing was conducted at the Ecology and Insect Behavior Laboratory of the Federal University of Alagoas. Sample collection was ended when B. cocophilus were no longer found in the laboratory tests. B. cocophilus were able to survive in the coconut palm roots for at least 118 and 133 days after felling (DAF), in Coruripe and São Miguel dos Milagres, respectively. B. cocophilus could migrated from the roots to the soil, where they were found until 76 and 88 DAF, in Coruripe and São Miguel dos Milagres, respectively.

Mark Hoddle: Smiting weevils

Profile Mark Hoddle: Smiting weevils flying insects — have shown that the weevils have the potential to fly tens of miles in a day. The weevil is also known to carry a pathogen known as the red ring nematode which also infects the Canary Island palm, making weevil infestations particularly deadly. The nematode hasn’t yet been found on the weevils trapped in the United States, but Hoddle says it’s probably only a matter of time until it appears. So far, the Hoddle family monitoring program hasn’t found any South American palm weevils north of San Diego County. But the insect appears to be well-entrenched in some areas, such as the Sweetwater River Trail, a natural area near Bonita in San Diego County where many palms grow wild and unmanaged, amidst thickets of willows. Areas like this provide reservoirs for the weevil population to grow and spread into new areas. As one part of their monitoring effort, the Hoddles have assigned GPS coordinates to 300 trees in and near the Sweetwater River Trail area, and are monitoring them to better understand the dynamics of the weevil’s spread. Hoddle will be using drone- mounted cameras to do regular monitoring of the trees in in- fested areas, many of which are difficult to reach. Hoddle has been working with the California Department of Food and Agriculture (CDFA) to draw attention to the infesta- tion, yielding media coverage all the way up to an article in the New York Times. People finding a South American palm weevil UC Riverside entomology specialist Mark Hoddle and his son, Nicholas, set weevil traps along the Sweetwater River Trail in San Diego County, a natural area where weevils have infested many unmanaged Canary Island palms. http://calag.ucanr.edu • JANUARY–MARCH 2017 21 Christina Hoddle Mike Lewis M ark Hoddle’s research article in this issue documents a major achievement in invasive species control — a successful eradication, in this case of the Asian palm weevil, Rhynchophorus vulneratus, from Laguna Beach. Hoddle, a UC Riverside-based entomology specialist and di- rector of the Center for Invasive Species Research (CISR), is now working to address another weevil infestation. This time, the invader is Rhynchophorus palmarum, the South American palm weevil. Over decades, the insect has steadily increased its range northward in Mexico, enabled by abundant plantings of Canary Island palms which the weevil infests in irrigated desert areas. These unnatural oases have provided step- ping stones for the weevil through inhos- pitable habitat all the way to California. The South American palm weevil, The South Rhynchophorus palmarum (actual American palm wee- size), infests the Canary Island palm, a vil was first found in common landscape tree in California. the United States in 2011, just north of the border near San Diego. After that detec- tion, a two-year, federally supported trapping program con- cluded that the infestation hadn’t spread beyond San Diego and Imperial counties. After 2013, funding for the trapping effort ran out, leaving no system for monitoring the weevil. After a visit to Tijuana last May, Hoddle found 125 weevil-killed palms in a day of driving around the city with a colleague from Tijuana. After this he de- cided to take matters into his own hands. The following month, he and his wife, Christina — also an entomology researcher at UC Riverside and a co-author on the research paper in this issue — began a family mission to assess the state of the infestation in California. “We are spending a significant amount of our spare time trapping in San Diego,” he said. They bring along their three- year-old son, Nicholas, who likes hunting for the satisfyingly large black weevils. “It was easy to get him on board, because we are looking for super cool big beetles,” said Hoddle, who grew up in New Zealand and has loved hunting for bugs since he was a small boy. Because Canary Island palms are such a ubiquitous part of the California landscape, they could provide food for weevil invasion all the way up to Northern California. Laboratory tests Hoddle has conducted using “flight mills” — treadmills for

Loop-mediated isothermal amplification (LAMP) for detection of the red ring nematode, Bursaphelenchus cocophilus

As a first step in developing a quick, accurate and simple method for the diagnosis of red ring disease, the loop-mediated isothermal amplification (LAMP)-based identification procedure was applied to the causative agent,Bursaphelenchuscocophilus. Two LAMP primer sets were designed using two loci of ribosomal RNA genes,i.e., D2-D3 expansion segments of the large subunit (D2-D3 LSU), and internal transcribed spacers (ITS). Within those two sets of primers, the D2-D3 LSU primer set successfully yielded amplicons fromB. cocophilusnematode lysate prepared from 3-year-old DESS-fixed specimens. The specificity of the primers was examined using 18 species of confamilial Aphelenchoididae nematodes and primer sensitivity was tested using a diluted series ofB. cocophiluslysate. The primer set did not amplify the DNA from other aphelenchoidids, and sensitivity was achieved by ‘1:100 diluted’B. cocophilusDNA (roughly 1/1500 of total DNA from a single third-stage juvenile).

Palm Weevil Pheromones - Discovery and Use.

Male-produced aggregation pheromones of seven major pest species of weevils in the subfamily Rhynchophorinae have been identified as a closely related set of methyl-branched secondary alcohols. Although the weevils produce only one stereoisomer of these alcohols, no instances of isomeric inhibition have been observed, enabling stereoisomeric mixtures to be used in traps. Addition of fermenting plant material to traps synergizes attraction of weevils to the pheromones. The weevils are large, have long life cycles, and are strong fliers. These characteristics make mass trapping a suitable tactic to add to existing management strategies. When coupled with good phytosanitary practices, mass trapping of Rhynchophorus palmarum at 1 trap/5-ha significantly lowered the incidence of red ring nematode infection vectored by the weevil in commercial oil palm plantations in the Americas. Similarly, trap densities of 1-10 traps/ha have significantly lowered R. ferrugineus infestation of date palm throughout the Middle East. Although management of R. ferrugineus in urban areas is more problematic, trapping is an integral part of most programs aimed at protection of ornamental Canary palms in Europe. Overall, semiochemically-based management of these large weevils is now a mature and usually economically feasible control technology.

Genetic diversity of Bursaphelenchus cocophilus in South America

Molecular characterisation ofBursaphelenchus cocophilus, the causal agent of ‘red ring disease’, is imperative for efficient identification procedures in Brazil and Colombia, because quarantine species such asB. xylophilusandB. mucronatusare already listed in both countries. ITS-1/2 region and D2-D3 segment of LSU rDNA were used to characterise isolates ofB. cocophilusobtained from coconut plantations in Brazil and Colombia. Results from ITS-1/2 and LSU rDNA regions showed that all isolates ofB. cocophilusfrom Brazil and Colombia formed a monophyletic group. The LSU rDNA region indicated that all isolates formed a single monophyletic group with high Bayesian posterior probability (100%). This is the first study on ITS-1/2 for the characterisation ofB. cocophiluspopulations. A species-specific primer was designed for identification ofB. cocophilus.

Detection ofRhynchophorus palmarum(Coleoptera: Curculionidae) and Identification of Associated Nematodes in South Texas

This study reports a survey conducted to find the South American palm weevil Rhynchophorus palmarum (L.) and the red palm weevil R. ferrugineus (Olivier) (Coleoptera: Curculionidae), 2 invasive species of palm trees. The study was performed in the Rio Grande Valley of south Texas and near the border with Tamaulipas state, Mexico. A total of 40 traps were inspected biweekly from 26 Sep 2011 to 20 Sep 2012 and 4 traps were inspected from 20 Sep 2012 to 4 Sep 2013. To attract R. palmarum and R. ferrugineus, the lures 2-methylhept-5-en- 4-ol and 4-methyl-5-nonanol were used, respectively. We used these lures in combination with sugar and the ethyl acetate aggregation kairomone for both species, plus an ethylene glycol kill solution. Two specimens of R. palmarum were found and identified next to a commercial palm plantation on 11 Mar and 5 May 2012 near the city of Alamo, Texas, but no R. ferrugineus was detected during the entire study. Nematodes found in the 2 R. palmarum specimens were dauer juveniles of the order Rhabditida, and in one of these weevils only 1 nematode of an undetermined species within the family Aphelenchoididae was found. It is of great importance that Bursaphelenchus cocophilus Cobb Baujard (Nematoda: Parasitaphelenchinae), the nematode causal agent of coconut red ring disease, was not found within these insects. This is the first detection of R. palmarum in Texas, and the second in the United States.

Nematodes that Ride Insects: Unforeseen Consequences of Arriving Species§

Insects that arrive in new regions can be hosts for a variety of unseen metazoans, including microscopic nematodes, which are carried phoretically as dauer juveniles or as internal/ external parasites in various stages. This includes insects that arrived by natural means, were purposefully introduced for biological control before strict APHIS/PPQ provisions, were inadvertently introduced as hitchhikers or stowaways, or were brought in as part of the pet trade or for food. In some cases, the host associations are so specific that they may pose little threat, but in other cases where host specificity is relatively wide and/or host transfer opportunities exist, the nematode associates can expand, colonize and establish associations with native insects causing various downstream environmental effects. Because nematodes are mostly microscopic, the consequences of such introductions are usually not considered in the pet trade or as an added consequence as introduced or invasive insects arrive in the state and establish themselves. These arrival scenarios are discussed with real-world examples, including one with damage potential for Florida and the southeastern U.S., i.e. the red ring nematode that is associated with palm weevils.

Rhyncophorus palmarum L. (Linnaeus, 1758): A morphological and histological study of the female reproductive system

Rhyncophorus palmarum (Coleoptera) is a pest of great economic importance because of the damage caused in the plants of the Palmae family. This study showed that the female reproductive system of this beetle is composed of two ovaries of the telotrophic meroistic type invariably containing two ovarioles per ovary. Each ovariole is home to about 50 oocytes at different stages of maturation. The proximal region of the ovary is rather peculiar because it houses inside it a large quantity of bacilliform structures immersed in a glycolipoprotein substance, which has never before been described in the literature. The function of these structures is still unknown. However, it is suggested that they can function as a method of neutralizing the resistance of the plant because these insects are responsible for the transmission of the red ring disease to the palm plants.

Disposição do nematóide Bursaphelenchus cocophilus (Cobb) baujard, em coqueiros portadores da doença anel-vermelho

Este trabalho teve o objetivo de conhecer como o nematóide Bursaphelenchus cocophilus tende a se distribuir no interior das plantas de coqueiros em estágios avançados da doença anel-vermelho. Nas amostras de raízes coletadas junto à base da estipe de coqueiros doentes, o número de nematóides foi consideravelmente maior que encontrado em raízes situadas entre um e três metros de distância da estipe. Observou-se que, à medida que se afasta da estipe, a possibilidade de encontrar nematóides na raiz é mínima, de forma que as chances de transmissão da doença, de uma planta para outra, através das raízes, devem ser muito pequenas. Na região do palmito, onde o tecido é mais tenro, é possível encontrar o nematóide tanto nas áreas avermelhadas quanto nas áreas aparentemente sadias. Nos tecidos do pecíolo, foram encontrados nematóides em pequena quantidade. Portanto, práticas profiláticas, visando à desinfecção do facão utilizado na colheita e despalma, devem ser realizadas com a finalidade de eliminar a transmissão da doença. Nenhuma das amostras obtidas dos tecidos da ráquis e dos folíolos estava contaminada. A população de nematóide é mais alta nos excrementos de túneis larvais das regiões apicais do coqueiro, o que confere maior chance de serem transportados para outras plantas, aderidos ao corpo do seu principal vetor, os adultos de Rhynchophorus palmarum.

The history of expansion of the genus Bursaphelenchus (Nematoda: Aphelenchida: Parasitaphelenchidae)

97 Because of globalization and removal of geographical barriers, frequent biological invasions of introduced species become an urgent environmental problem. According to the Convention on Biological Diversity (CBD), precise identification of dangerous aggressive species at the early stages of their invasion to new regions is the most important component of the environmental control and monitoring. To resist the potential environmental hazard, the precise data are required on the current distribution and history of expansion of pests that are of global economic importance. We performed biogeographical analysis of phytopathogenic nematodes of the genus Bursaphelenchus , which are of world plant quarantine importance. We analyzed the species groups erected previously on the basis of morphological data and the biotic relationships with the host plants and insect vectors [1]. A model of the genus dispersal based on all data on Bursaphelenchus records in natural biotopes was developed using the secondary Brooks parsimony approach (sBPA). Two species of the genus are included into the A1 list of especially dangerous plant quarantine organisms [2]. These are Bursaphelenchus xylophilus , the causative agent of the pine wilt disease in Asia and America, and Bursaphelenchus cocophilus , the pathogen of the palm red ring disease in the Caribbean region. The Bursaphelenchus genus includes 78 phytophathogenic species the life cycle of which is associated with woody plants (mostly coniferous trees from the family Pinaceae), insects (beetles from the families Scolytidae, Cerambycidae, Curculionidae, and Buprestidae), and basidium-producing fungi. The insect vectors have played an important role in the Bursaphelenchus evolution: the initial association, Scolytidae beetles‐Pinaceae conifers, has changed, giving rise to two natural species groups within the genus: the xylophilus group (the vectors were longhorn beetles of the family Cerambycidae) and the hunti group (the vectors belong to the order Hymenoptera) [1]. A modified sBPA was used to determine the specific features of the spread of these nematodes and their relationships with the host plants [3‐5]. In works by Brooks and his followers, matrices and dendrograms were obtained by the parsimony method on the basis of well-grounded phylogenies of both plant and animal taxa. The dendrogram branches of the phylogenetic tree of the studied taxon served as fields in matrices for the regions and host taxa. On the basis of these matrices, new dendrograms of the regions and hosts were obtained to develop the concepts of the evolution for the taxa and fauna of the areas under consideration, accurate to species. In this study, the matrices of plants and large regions were developed on the basis of the

Nematodes of Rhynchophorus palmarum, L. (Coleoptera: Curculionidae), vector of the Red Ring disease in coconut plantations from the north of the Rio de Janeiro State

Rhynchophorus palmarum, the palm weevil, has been reported as a pest of palms and sugarcane plants. The Red Ring Disease is an infectious plant disease caused by nematodes. The etiological agent, Bursaphelencus (Rhadinaphelencus) cocophilus (Nematoda: Aphelenchoididae), completes its life cycle within 9 to 10 days inside the palm tree. The main symptom is a permanent wilting of the plant aerial parts. Previous studies stated that B. cocophilus cohabits with other nematodes the gut of R. palmarum. The aim of this study is to identify nematodes collected from palm weevil found in coconut plantations from the north of the Rio de Janeiro State. Light (LM) and scanning electron microscopy (SEM) analyses were carried on samples of infected R. palmarum and fragments and fresh juice of infected plants with B. cocophilus. Observations of R. palmarum fecal material made by LM and SEM showed three species cohabiting these samples, being also present in fresh juice and fragments of infected coconut tree: B. cocophilus, Teratorhabditis palmarum (Nematoda: Rhabditidae) and Diplogasteritus sp (Nematoda: Diplogasteridae). These findings confirm previous studies, which related that R. palmarum own a varied nematode fauna. Nematodes associated to B. cocophilus probably could be co-participates of the etiology of the Red Ring disease.

Rhynchophorus palmarum

<i>Rhynchophorus palmarum</i>

Trapping-a major tactic of BIPM strategy of palm weevils

Several species of curculionid weevils such as Amerrhinus ynca Sahlberg, Cholus annulatus Linnaeus, C. martiniquensis Marshall, C. zonatus (Swederus), Diocalandra frumenti (Fabricius), Dynamis borassi Fabricius, Homalinotus coriaceus Gyllenhal, Metamasius hemipterus Linnaeus, Paramasius distortus (Gemminger &amp; Horold), Rhabdoscelus obscurus (Boisduval), Rhinostomus barbirostris (Fabricius), R. afzelii (Gyllenhal), Rhynchophorus bilineatus (Montrouzier), R. cruentatus Fabricius, R. ferrugineus (Olivier), R. palmarum (Linnaeus) and R. phoenicis (Fabricius) are associated with palms. Some of these have become a major constraint in the successful cultivation of coconut palm (Cocos nucifera L.), date palm (Phoenix dactylifera L.) and oil palm (Elaeis guineensis L.). R. ferrugineus is distributed in over 33 countries and attacks more than two dozen palm species. In the recent past, it has spread to Middle Eastern countries, Mediterranean region of Africa and southern Europe (Spain) causing tremendous economic losses. The yield of date palm has decreased from 10 to 0.7 tons/ha. Coconut palms in India are infested upto 6.9 per cent in Kerala and 11.65 per cent in Tamil Nadu. R. palmarum is a major pest of oil and coconut palms in the tropical Americas and, vectors the nematode, Bursaphelenchus cocophilus (Cobb) Baujard which causes red ring disease (RRD). Palm losses due to RRD are commonly between 0.1 to 15% which amounts to tens of millions dollars. The status of other species is briefed. The grubs of weevils that develop in the stems, bud, rachis of leaves and inflorescence of cultivated, ornamental or wild palms cause direct damage. Because of the cryptic habitat of the grubs, which act as tissue borer, the management becomes difficult.