Received: 25 June, 2011. Accepted: 22 February, 2012. Invited Review Functional Plant Science and Biotechnology ©2012 Global Science Books Recent Biotechnological Approaches in Diagnosis and Management of Sugarcane Phytoplasma Diseases Govind P. Rao 1* • Smriti Mall 2 • Carmine Marcone 3 1 Division of Plant Pathology, Indian Agricultural Research Institute, New Delhi 110012, India 2 Sugarcane Research Station, Kunraghat, Gorakhpur 273008, U.P., India 3 Dipartimento di Scienze Farmaceutiche, Università degli Studi di Salerno, Via Ponte Don Melillo, I-84084 Fisciano (Salerno), Italy Corresponding author: * gprao_gor@rediffmail.com; gprao_gor@hotmail.com ABSTRACT Phytoplasmas have been reported to be associated with two major sugarcane diseases viz., grassy shoot disease (GSD) and white leaf disease (WLD). Sugarcane yellow leaf syndrome is a new syndrome of sugarcane caused by a luteovirus and a phytoplasma. Sugarcane green grassy shoot (SCGGS) and ramu stunt are also associated with sugarcane from Thailand and Papua New Guinea, respectively. GSD and WLD are causing significant economic losses to sugarcane yield and sugar recovery in Asian countries. Both these phytoplasmas have been spreading very rapidly to newer locations with the help of infected seed material and leafhopper vectors. Hence it would be important to diagnose and manage these phytoplasmas at an early stage of sugarcane growth to avoid further spread and significant losses caused by them. Because of unreliable and unspecific symptoms, the identification and characterization of the associated phytoplasma at an early stage of plant growth is problematic and unreliable. The introduction of molecular genetic methods into plant mycoplasmology about 15 years ago greatly improved the diagnosis of phytoplasma infections in plant and insect hosts. PCR offers several advantages over other methods including versatility, relative simplicity, specificity and high sensitivity, which can be increased by a two-step PCR (nested PCR). It has also become possible to differentiate, characterize and classify the phytoplasmas on a phylogenetic basis, using mainly sequence analysis of ribosomal DNA (rDNA). The biotechnological and molecular assays developed in recent years have been utilized at a large scale to characterize these pathogens at an early stage of growth and for screening planting seed materials for possible elimination and management of theses phytoplasma at an early stage of their growth and propagation. We summarize research being conducted on characterization and management of these phytoplasma by novel biotechnological and molecular approaches. _____________________________________________________________________________________________________________ Keywords: characterization, detection, PCR assays, grassy shoot disease, white leaf disease Abbreviations: PCR, polymerase chain reaction; RFLP, restriction fragment length polymorphism; SCGS, sugarcane grassy shoot; SCWL, white leaf disease; YLS, yellow leaf syndrome CONTENTS INTRODUCTION........................................................................................................................................................................................ 19 PHYTOPLASMAS INFECTING SUGARCANE ....................................................................................................................................... 20 Grassy shoot ............................................................................................................................................................................................ 20 White leaf ................................................................................................................................................................................................ 21 Leaf yellows ............................................................................................................................................................................................ 24 Green grassy shoot disease ...................................................................................................................................................................... 25 Ramu stunt............................................................................................................................................................................................... 25 ALTERNATIVE HOSTS OF SUGARCANE PHYTOPLASMAS .............................................................................................................. 26 MANAGEMENT APPROACH ................................................................................................................................................................... 26 CONCLUSION ............................................................................................................................................................................................ 27 REFERENCES............................................................................................................................................................................................. 27 _____________________________________________________________________________________________________________ INTRODUCTION Phytoplasmas are phytopathogenic prokaryotes that were formerly named as mycoplasma-like organism (MLOs) and were discovered in Japan in 1967 (Doi et al. 1967). They were observed in ultrathin sections of plant phloem tissue and named MLOs because they physically resembled myco- plasmas. Later, these microorganisms were renamed phyto- plasmas in 1994 at the 10 th Congress of the International Organization of Mycoplasmology (1993). Phytoplasmas are wall-less pleomorphic bacteria with sizes ranging from 200 to 800 nm, which could survive and multiply only in plant phloem or insect haemolymph. They are strictly host- dependent, but could multiply in insect vectors and also survive in their eggs. Phylogenetic studies have shown that the common ancestor of phytoplasmas is Acholeplasma laidlawii. Also, phytoplasmas and acholeplasmas use the conventional UGG codon for tryptophan (Trp) and retain UGA as a stop codon, while most other mollicutes use UGA as a Trp codon. In infected plants, phytoplasmas live mainly or perhaps only, in the sieve cells of phloem tissue. The sieve cells are highly specialized cells that are living but do not contain nuclei when they are mature. Infection of a plant by a phytoplasma often results in disease. Symptoms that are characteristic of phytoplasma diseases include yel- lowing and reduced size of leaves, stunting of the plant, and proliferation of axillary buds. Such growth from axillary buds often results in a witches' broom appearance. Other ®