Cellulolytic potential of probiotic Bacillus Subtilis AMS6 isolated from traditional fermented soybean (Churpi): An in-vitro study with regards to application as an animal feed additive

Microbiol Res. 2016 May-Jun:186-187:62-70. doi: 10.1016/j.micres.2016.03.004. Epub 2016 Mar 23.


The aim of the present study is to evaluate the probiotic attributes of Bacillus subtilis AMS6 isolated from fermented soybean (Churpi). This isolate exhibited tolerance to low pH (pH 2.0) and bile salt (0.3%), capability to autoaggregate and coaggregate. AMS6 also showed highest antibacterial activity against the pathogenic indicator strain Salmonella enterica typhimurium (MTCC 1252) and susceptibility towards different antibiotics tested. The isolate was effective in inhibiting the adherence of food borne pathogens to Caco-2 epithelial cell lines, and was also found to be non-hemolytic which further strengthen the candidature of the isolate as a potential probiotic. Further studies revealed B. subtilis AMS6 showed cellulolytic activity (0.54±0.05 filter paper units mL(-1)) at 37°C. The isolate was found to hydrolyze carboxymethyl cellulose, filter paper and maize (Zea mays) straw. The maize straw digestion was confirmed by scanning electron microscopy studies. The isolate was able to degrade filter paper within 96h of incubation. A full length cellulase gene of AMS6 was amplified using degenerate primers consisting of 1499 nucleotides. The ORF encoded for a protein of 499 amino acids residues with a predicted molecular mass of 55.04kDa. The amino acids sequence consisted of a glycosyl hydrolase family 5 domain at N-terminal; Glycosyl hydrolase catalytic core and a CBM-3 cellulose binding domain at its C terminal. The study suggests potential probiotic B. subtilis AMS6 as a promising candidate envisaging its application as an animal feed additive for enhanced fiber digestion and gut health of animal.

Keywords: Animal feed; Bacillus subtilis; Caco-2; Cellulolytic; Probiotics.

MeSH terms

  • Animal Feed*
  • Animals
  • Antibiosis
  • Bacillus subtilis / drug effects
  • Bacillus subtilis / isolation & purification*
  • Bacillus subtilis / metabolism*
  • Bacillus subtilis / physiology
  • Bacterial Adhesion
  • Bile Acids and Salts / metabolism
  • Caco-2 Cells
  • Cellulase / genetics
  • Cellulose / metabolism*
  • Epithelial Cells / microbiology
  • Food Additives*
  • Glycine max / microbiology
  • Humans
  • Hydrogen-Ion Concentration
  • Hydrolysis
  • Paper
  • Plant Stems / metabolism
  • Polymerase Chain Reaction
  • Probiotics / isolation & purification*
  • Probiotics / metabolism*
  • Salmonella typhimurium / growth & development
  • Sequence Analysis, DNA
  • Temperature
  • Zea mays / metabolism


  • Bile Acids and Salts
  • Food Additives
  • Cellulose
  • Cellulase