Bacillus subtilis

3.1 | B. subtilis View in CoL isolation and identification

One isolate was taken from darkbarbel catfish. Physiological and biochemical identification results revealed that the isolate was gram-positive, aerobic bacteria with the same ability to utilize gelatin, trehalose, mannitol, and glucose, and to produce catalase and amylase (Table 2). Despite being able to use gelatin and citrate, it was unable to make indole. The strain could not decompose xylose, mannose, inositol, galactose, and arabinose. The Voges – Proskauer (V-P) test revealed a positive result for this bacterial isolate. The phenotypic characteristics of this isolate matched those of B. subtilis according to Bergey's manual and the Manual for the Identification of Medical Bacteria. These characteristics were also shared by the reference strain B. subtilis 168. Furthermore, a Basic Local Alignment Search Tool examination of the single isolate's 16S rRNA gene sequences demonstrated strong similarity (98% – 100%) with B. subtilis . According to phylogenetic trees created using the 16S rRNA gene sequences, the single isolate belonged to the same group as the reference strains of B. subtilis (Figure 1). This isolate was thus recognized as B. subtilis based on the behavioral traits and phylogenetic analyses. Overall, it is certain that the isolated bacteria from darkbarbel catfish are members of the B. subtilis species.

Physiobiochemical indicator Bacillus subtilis 168 Bacillus subtilis

Gram stain test + +

Catalase test + +

Aerobic test + +

Hydrothion

Starch hydrolysis test + +

Methyl red test

Gelatin liquidized test + +

Citrate utilization test + +

Indole production

Voges – Proskauer test + +

Arginine dihydrolase

Lysine decarboxylase

Ornithine decarboxylase

Arabinose

Galactose

Glucose + +

Inositol

Mannitol + +

Mannose

Xylose

Trehalose + +

Note: “+”indicates positive reaction; “ ”indicates negative reaction.