Carboxymethylcellulase Produced by Facultative Bacteria from the Hind-gut of the Termite Reticulitermes hesperus

Abstract

13Bacillus cereus RWl and Serratia marcescens R W~, isolated from the hind-gut of the termite Reticulitermes hesperus, both grew well on mesquite wood and produced moderate amounts of carboxymethylcellulase. Carboxymethylcellulose (CMC) gels were depolymerized rapidly by 23. cereus Rwl and slowly by S. marcescens RW3. The depolymerization of CMC was pH and temperature sensitive. Depolymerization of gels by growing cultures of B. cereus Rw1 and the action of cell-free extracts of B. cereus Rw1 on CMC sols were optimum at pH 6-0 and 5.5, respectively. Glucose and cellobiose increased the rate of CMC gel depolymerization. Enzyme synthesis rather than growth was stimulated by the addition of glucose to a culture of Rwl growing on a non-cellulosic substrate. Bacillus cereus Rw1 produced both cell-free and cell-bound carboxymethylcellulase. I N T R O D U C T I O NThe isolation of bacteria from the hind-gut of the termite Reticuliterrnes hesperus, their growth on mesquite wood and the depolymerization of carboxymethylcellulose were reported by Thayer (1976). Since wood contains many potential substrates, growth did not necessarily indicate cellulolytic activity. During growth Bacillus cereus RW1 and Serratia marcescens RW3 solubilized 2 to 5 % more mesquite wood than could be achieved by autoclaving, which solubilized 23 %. Although B. cereus Rwl and S. marcescens R W~ attacked the insoluble components of the wood, they preferred the soluble substrates. Some cellulose hydrolysis and distinct carboxymethylcellulose depolymerization indicated weak cellulolytic activity. Since these organisms were isolated from the hind-gut of the termite and grew on wood, they might contribute to cellulose digestion in the termite. This study characterized the carboxymethylcellulase activity of these organisms. M E T H O D SOrgaitisms. Bacillus cereus RW 1 and Serratia marcescens R W~, obtained as described previously from the hind-gut of Reticulitermes hesperus (Thayer, 1976), were maintained on BBL Trypticase Soy Agar (TSA).Media. The basal medium used to study growth on wood contained (g 1-1 in distilled water): NaCI, 3.0; KH2P04, 1.0; K,HPOI, 1.0; (NH4),S04, 2.0; MgS04, 0.05; CaCl,, 0.05; Difco yeast extract, 0.50; and dormant mesquite wood sawdust, 10.0; the pH was adjusted to 6-5 after autoclaving. To determine the temperatures supporting maximum growth rates, cultures were grown in mineral salts medium containing 5 mwglucose at pH 6.5 (the medium was sterilized by filtration).Incubation and inoculation. Cultures were routinely grown at 35 "C in 100 ml medium in 500 ml baffled Erlenmeyer flasks shaken at 250rev. min-1 on a gyratory shaker. In some experiments, flasks with an attached 10 mm diam. cuvette were used; the design of these limited the volume of medium to 80 ml. Inocula were 0.1 ml per 100 ml medium.Measurement ofgrowth. Absorbance measurements were made at 420 nm in colourless media or at the

Assembly of Minicellulosomes on the Surface of Bacillus subtilis

“…Several studies have shown that B. subtilis can be used as a host to secrete heterologous cellulases, and naturally occurring strains have been identified that secrete cellulases (26,27,53,57,62,63). Cellulase enzymes have also been targeted to the membrane, enabling protoplasts of B. subtilis to degrade CMC (29).…”

Disruption of the Termite Gut Microbiota and Its Prolonged Consequences for Fitness

“…Second, the appearance of rare or nonnative bacterial members in the rifampin-treated guts may be affected by interactions with other bacteria. For example, the Serratia marcescens 16S rRNA gene sequence identified in our study is 99.9% identical to that of an opportunistic pathogen of termites that has been hypothesized to induce replication of normal termite gut bacteria by suppressing the host immunity, changing available oxygen, and producing bacterial growth-promoting enzymes such as carboxymethyl cellulase (1,19,68,75). A Serratia-induced proliferation of the symbiotic community could cause septicemia, which can result in early termite mortality (75).…”

Endo-beta-1,4-glucanase gene of Bacillus subtilis DLG

“…As cellulolytic enzyme systems of other organisms are generally repressed by glucose (12,30,53,55), the effect observed here is quite unusual. Thayer (58) has also reported increased ,B-1,4-glucanase activity by B. cereus in the presence of glucose. It remains to be determined whether glucose itself acts as an inducer of enzyme synthesis or whether the stimulatory effect on P-1,4-glucanase production by glucose is more indirect.…”