Phytase Production from a Novel Klebsiella sp. on Wheat Bran for Animal Feed Digestion

Dilruba Akter ^a, Md. Murad Khan ^b, Md. Mahmuduzzaman Mian ^a, Shakila Nargis Khan ^a, Md. Mozammel Hoq ^a

The present findings will be very useful for the development of a bioprocess of the enzyme for its large-scale production at the pilot and finally at the commercial level.

Abstract

Background: The present study was aimed to isolate phytase producing bacteria and optimize the physicochemical parameters of their phytase production. Materials and methods: Four bacterial isolates (Phs4, Phs5, Phs6, and Phs8), based on clear zone formation on phytase screening medium, were selected and tested for finding out the highest phytase producing strain. The production of phytase was then optimized and its biochemical properties were determined to judge the applicability of phytase as a digestive aid in animal feed.  Results: The 4 bacterial isolates (Phs4, Phs5, Phs6 and Phs8) were identified by morphological, cultural, biochemical and molecular characterization as Burkholderia cepacia, Escherichia coli, Klebsiella pneumoniae and Klebsiella sp. respectively. Of these isolates, Phs8 (Klebsiella sp.) was found to produce maximum phytase in shake culture in a basal medium containing Na-phytate at 37^oC and pH 5.5 after 72 hours of incubation. The omission of Na-phytate from the medium almost completely abolished the phytase production capacity of the isolate and thus signified its important role as an inducer. Among the different complex carbon sources, viz., glucose, wheat bran, rice bran and chickpea, maximum phytase production (94 unit/ ml) was obtained with wheat bran under comparable cultivation conditions. The phytase works best at a temperature of 37^oC and pH of 4.0 with a wide temperature stability (more than 80% activity up to 80^oC) and wide pH stability (more than 80% activity within a range of 3-8). Although Zn²⁺, Co²⁺, and Fe²⁺ slightly increased the phytase activity Cu²⁺ and Mg²⁺ strongly inhibited the enzyme. Conclusion: The present findings will be very useful for the development of a bioprocess of the enzyme for its large-scale production at the pilot and finally at the commercial level.

Keywords: Phytase; animal feed; Klebsiella sp.; enzyme activity.

Significance: Economic production of bacterial phytase as a poultry feed supplement

Abbreviations: PSM, phytase screening medium; PPM, phytase production medium; LB, Luria-Bertani; RPM, rotation per minute; PCR, polymerase chain reaction; dNTPs, deoxyribonucleotides; EDTA, ethylenediaminetetraacetic acid; EtBr, ethidium bromide; KIA, Kligler iron agar; MIU, motility indole urease test; VP, Voges–Proskauer test.

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