Physiological effect of chitinase purified from Bacillus subtilis against the tobacco cutworm Spodoptera litura Fab. Rajamanickam Chandrasekaran 1 , Kannan Revathi 1 , Selvamathiazhagan Nisha, Suyambulingam Arunachalam Kirubakaran, Subbiah Sathish-Narayanan, Sengottayan Senthil-Nathan ⇑ Division of Biopesticides and Environmental Toxicology, Sri Paramakalyani Center for Excellence in Environmental Sciences (SPKCES), Manonmaniam Sundaranar University, Alwarkurichi 627 412, Tirunelveli, Tamil Nadu, India article info Article history: Received 5 January 2012 Accepted 2 July 2012 Available online xxxx Keywords: Bacillus Chitinase Enzyme Molecular masses Cutworm Dietary utilization Mortality abstract An extracellular chitinase was purified from Bacillus subtilis. The lethal concentration (LC 50 ) was deter- mined by using chitinase in first, second, and third instars of Spodoptera litura Fab. Chitinase showed the highest insecticidal activity at 6 lM concentration within 48 h. The nutritional indices were also significantly affected by the 6 lM concentration (P < 0.05). Food consumption, efficiency of conversion of ingested and digested food, relative growth rate, and consumption values declined significantly while approximate digestibility was increased. Our study indicates that treatment of host plant leaves with the chitinase can regulate (reduce) larval growth and weight, and enhance the mortality. This may serve as an effective biocide and alternative to Bt toxin. Ó 2012 Elsevier Inc. All rights reserved. 1. Introduction Pesticides are applied in agricultural systems for protecting crops from damage by insects and disease. The use of pesticides has also resulted in significant benefits effects to public health and the environment. In general the amount of pesticides released into the environment has risen significantly in the last five decades [1,2]. However scientists constantly seek for an effective and environmentally friendly method of controlling pests and diseases [3–5]. Control of crop pests by the use of biological agents holds great promise as an alternative to the use of chemicals. Secondary metabolites and crude enzyme from microorganisms have been used to control crop pest population [6]. Chitin is a long, unbranched polysaccharide of an amino sugar N-acetyl-b-D-glucosamine linked together by b-1,4-glycosidic linkages [7]. It is abundant in nature as a structural compound in cuticle and integument of animals, especially in insects [8]. Chitin is metabolized by various chitinases that are found in insects, bac- teria, fungi and higher plants [9]. Chitinases belong to glycosyl hydrolase families 18 and 19 according to the classification made by Henrissat and Bairoch [10]. The family 18 chitinases have been determined to possess a common (a/b) 8-barrel domain, compris- ing of eight a-helices and eight b-strands. The catalytic reaction of the family 18 enzymes takes place through a retaining mechanism, in which b-anomer is generated by hydrolysis of b-l,4-glycosidic linkages. The chitinases of the family 19 have been described as being similar to lysozyme and chitosanase in its mode of action [11]. Insect growth and development are strongly dependent on the construction and remodeling of chitinous structures [12]. Chitinase induced damage to the peritrophic membrane in the insect gut causes a significant reduction in nutrient utilization and conse- quently in insect growth [13]. Due to this, chitinase present in the insect diet can decrease insect growth [14,15]. Chitinolytic microorganisms have many potential applications as biocontrol agents [16]. Over-expression of a chitinase in an entomopatho- genic organism can increase insect mortality [17]. Chitinase plays a vital role in the control of plant pathogenic fungi and raising plant disease tolerance. In the microbial degrada- tion of chitin using chitinase, chitooligosaccahrides are produced, which are further degraded to N-acetylglucosamine by chitobiase [18]. Consequently, microbial chitinases have been isolated for the production of chitooligosaccharides [19]. Bacillus species are well-known as producers of extra-cellular enzymes, including cellulase, glucanase, amylase, proteinase and chitinase, secondary metabolic products, and they are used 0048-3575/$ - see front matter Ó 2012 Elsevier Inc. All rights reserved. http://dx.doi.org/10.1016/j.pestbp.2012.07.002 ⇑ Corresponding author. Fax: +91 4634 283270. E-mail addresses: senthilkalaidr@hotmail.com, senthil@msuniv.ac.in (S. Senthil-Nathan). 1 These authors contributed equally to this work. Pesticide Biochemistry and Physiology xxx (2012) xxx–xxx Contents lists available at SciVerse ScienceDirect Pesticide Biochemistry and Physiology journal homepage: www.elsevier.com/locate/pest Please cite this article in press as: R. Chandrasekaran et al., Physiological effect of chitinase purified from Bacillus subtilis against the tobacco cutworm Spo- doptera litura Fab., Pestic. Biochem. Physiol. (2012), http://dx.doi.org/10.1016/j.pestbp.2012.07.002