Research Article Open Access Biofertilizers & Biopesticides Habte and Osorio, J Biofertil Biopestici 2012, 3:5 http://dx.doi.org/10.4172/2155-6202.1000127 Volume 3 • Issue 5 • 1000127 J Biofertil Biopestici ISSN:2155-6202 JBFBP, an open access journal Keywords: Mortierella sp; Phosphorus; Nitrate; Ammonium Introduction Soil phosphate deiciency is globally a major constraint for agriculture, particularly in the tropical regions [1-3]. An alternative to overcome this limitation is the use of rock phosphate (RP) available in many countries; unfortunately this material is quite insoluble, which impairs its use [4,5]. here is an increasing interest in developing strategies to improve the efectiveness of RP as a direct source of Pi for plant growth in parts of the tropics endowed with local deposits of the material [4,6-10]. One of the biotechnological strategies involve the use of phosphate- solubilizing microorganisms (PSM) [2,11-17]. Microbial dissolution of RP is brought about by a number of mechanisms including (i) release of organic acids [18,19], (ii) formation of complexes between organic anions and cations such as Al 3+ and Ca 2+ [20,21], and (iii) excretion of protons due to NH 4 + uptake [12,22]. he participation of the last mechanism in the microbial solubilization of RP has not been fully investigated for fungal P solubilizers [18]. Given the limited N supply of most soils [23,24], the N applied either as NO 3 - or NH 4 + fertilizers can control the extent of acid production by PSM, which is crucial in the eforts to evaluate the suitability of Mortierella sp. as an efective RP solubilizer in the rhizosphere or for its use in the biotechnological production of bio-acidulated RP [25]. he hypothesis of this experiment is that the efectiveness of PSM in dissolving RP may be inluenced by the N form present in the growth medium. his efectiveness may be enhanced if the PSM is supplied with NH 4 + as the sole source of N. he objective of the current investigation was to asses the efects of N form (NH 4 + or NO 3 -) on RP solubilization activity of Mortierella sp. under in vitro conditions. Materials and Methods Mortierella sp. was originally isolated from an Andisol of Hawaii [13] and maintained on Yeast malt agar (YMA) slants at 4°C. For this study, the fungus was multiplied in petri dishes on YMA medium for three days at 28°C. Mycelia were removed from the surface of the agar with a sterile loop and suspended in sterile deionized water and shaken by hand until the clumps were dispersed. 1 mL of a Mortierella sp. suspension containing 5.9x10 5 CUF was aseptically transferred into 250 mL Erlenmeyer lasks containing 75 mL of an autoclaved (30 minute, at 120°C and 0.1 MPa) liquid MgSO 4 .7H 2 O, 28 mg Fe-EDTA, 28 mg Cu-EDTA, 28 mg Mn-EDTA, 14 mg Zn-EDTA, 10.0 g glucose, and 3.5 g of Huila RP per liter. he RP was passed through a 0.5-mm aperture sieve. he P content of the Huila RP was 130 g kg -1 , and its empirical formula is Ca 9.69 Na 0.22 Mg 0.0 9 (PO 4 ) 5.14 (CO 3 ) 0.86 F 2.34 [26]. he medium contained 0.35 g N/liter, the source of N was NH 4 NO 3 (1.0 g L -1 ), NH4Cl (1.34 g L -1 ), or KNO 3 (2.53 g L -1 ). he lasks containing NH 4 NO 3 and NH 4 Cl also received KCl (1.87 g L -1 ) in order to maintain similar amounts of potassium in all the treatments. he initial solution pH was adjusted with 0.1 M NaOH to pH 7.6. Flasks were continuously shaken at 150 rpm on an orbital shaker (model Innova 4400, New Brunswick Scientiic Co, Inc., Edison, NJ) at 25°C for seven days at the Soil Microbiology Laboratory of the University of Hawaii at Manoa (Honolulu, HI, USA). Ater the incubation period, 50 mL of the suspension was pippeted into plastic tubes for centrifugation at 5000xg for 15 minutes. he supernatant was iltered through a Whatman No. 42 ilter paper followed by iltration through a membrane ilter (0.45 μm). Solution pH was measured with a pH-meter. Solution P concentration was determined using the molybdate-blue method [27]. he fungal mats were transferred onto a ilter paper, oven-dried (60°C for 48 h), and weighted for fungal dry weight (FDW) determination ater removal of remaining RP particles. Fungal P concentration was determined by the *Corresponding author: Nelson Walter Osorio, Universidad Nacional de Colombia, Calle 59A No. 63-20, Of. 14-216, 050034, Medellín, Colombia, Tel: 574- 4309310; Fax: 574- 4309301; E-mail: nwosorio@unal.edu.co Received September 11, 2012; Accepted October 16, 2012; Published October 18, 2012 Citation: Habte M, Osorio NW (2012) Effect of Nitrogen Form on the Effectiveness of a Phosphate-Solubilizing Fungus to Dissolve Rock Phosphate. J Biofertil Biopestici 3:127. doi:10.4172/2155-6202.1000127 Copyright: © 2012 Habte M, et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Effect of Nitrogen Form on the Effectiveness of a Phosphate-Solubilizing Fungus to Dissolve Rock Phosphate Mitiku Habte 1 and Nelson Walter Osorio 2 * 1 University of Hawaii, St. John 102, 31390 Maile Way, 96822, Honolulu, HI, USA 2 Universidad Nacional de Colombia, Calle 59A No. 63-20, Of. 14-216, 050034, Medellín, Colombia Abstract An in vitro experiment was carried out to evaluate the effect of nitrogen (N) form (NH 4 + and/or NO 3 -) on the dissolution of rock phosphate (RP) by a phosphate solubilizing fungus (PSF) identiied as Mortierella sp. In the presence of NH 4 Cl or NH 4 NO 3 , the solution of pH signiicantly decreased from an initial value of 7.6 to 3.4 and 3.7 respectively. In the presence of KNO 3 , the pH went down only to 6.7. As a result, signiicantly more P was detected in solution in the presence of NH 4 Cl (129.65 mg/L) than in the presence of NH 4 NO 3 (109.25 mg/L). The concentration of P in solution in the presence of KNO3 was only 0.08 mg/L. The excess of NH 4 + adversely affected the growth of Mortierella sp. However, this may have promoted a more active H+-pumping that decreased solution’s pH. In the presence of NO 3 - as the only source of N, Mortierella sp. not only dissolved a small amount of Pi from the RP but also immobilized most of it into its mycelia. In contrast, in the presence of NH 4 Cl, Mortierella sp. was effective to dissolve RP and the Pi released remaining in solution while only a little portion was immobilized by the fungal mycelia. medium. he medium consisted of 1.0 g NaCl, 0.2 g CaCl 2 .2H 2 O, 0.4 g