Organic-acid-producing, phytate-mineralizing rhizobacteria and their effect on growth of pigeon pea ( Cajanus cajan)

Abstract

Phytates represent a significant pool of organic phosphorus (Po) that is largely unavailable to plants. This study deals with phytate-mineralizing (PM), organic-acid-producing (OAP) rhizobacterial isolates, their characterization and their effect on plant growth. Their genetic diversity was assessed by 16S rRNA amplified ribosomal DNA restriction analysis (ARDRA) and selected isolates were identified by partial sequencing of 16S rRNA gene. Na-phytate and Po rich poultry farm manure (PFM) used as sources of phosphorus in semi-solid-agar (SSA) medium and soil respectively, for plant inoculation studies, where Cajanus cajan (pigeon pea) used as plant. Of thirty-nine rhizobacterial isolates, nineteen were proficient at releasing phosphate (Pi) (up to 85 μg/ml) from sparingly soluble calcium (Ca)-phytate and concomitantly decreasing the pH of minimal medium with 100 mM glucose from 8.0 to below 5. When the medium contained glycerol in place of glucose, Ca-phytate remained undissolved with no significant Pi released and no decline in pH. Genetic diversity of phytate-mineralizing (PM) rhizobacterial isolates suggests that the isolates mainly fall in two populations: acid-producing (AP) population (mainly represented by members of Enterobacteriaceae) and non-AP population. OAP-PM rhizobacterial isolates were identified as Citrobacter, Pantoea, Klebsiella and Enterobacter species. Organic acids (OAs) secreted by PM isolates were detected by HPLC, showed secretion of gluconic and acetic acids. Importance of OAs in Ca-phytate dephosphorylation was demonstrated in vitro using A. ficuum phytase. Gluconate and acetate additions enhanced phytase catalyzed dephosphorylation of Ca-phytate in vitro. Sonicated cell lysates of isolates showed significant Pi release from Ca-phytate compared to whole cells, indicating inaccessibility of Ca-phytate due to poor solubility. Selected isolates showed that they possess cell-associated acid phytase and modulators of phytase activity suggested that the enzymes are histidine acid phosphatase (HAP) type of phytase. OAP-PM isolates PP1 and DHRSS showed significant increase in dry shoot/root ratio and P content of shoot in Na-phytate containing semi-solid agar (SSA) medium, but isolate DHRSS did not increase dry shoot/root ratio in soil experiments containing poultry farm manure as source of P, although it significantly increased shoot P content of plants. The inoculation of isolates enhanced the shoot P content and dry shoot/root ratio, but did not increase the dry weight in SSA medium. It may be concluded that some OAP-PM rhizobacterial isolates that release P from Ca-phytate show increase in shoot P content in phytate containing SSA medium and in soils.