Research Journal of Agriculture and Biological Sciences, 2(1): 5-11, 2006

2006, INSInet Publication

Corresponding Author: M.A. Badr, Plant Nutrition Department, National Research Center, Cairo, Egypt.
5
The Dissolution of K and P-bearing Minerals by Silicate Dissolving Bacteria and
Their Effect on Sorghum Growth
M.A. Badr, A.M. Shafei and S.H. Sharaf El-Deen
Plant Nutrition Department, Soils and Water use Department and Microbial Genetics Department,
National Research Center, Cairo, Egypt.
Abstract: Potassium and phosphorus are vital components of plant nutrition package limiting crop yield and
quality. In particular, the crop production in Egypt relies quietly on the world potash fertilizers. It thus becomes
urgent to exploit the indigenous mineral resources to alleviate the importation of fertilizers. Therefore,
dissolution of feldspar (orthoclase) and rock phosphate by silicate dissolving bacteria was monitored in pure
culture and soil pot experiments to determine the effect of these bacteria on releasing K and P as well as their
effect on plant growth. Six strains of bacteria capable of dissolving silicate minerals were isolated from feldspar
samples. The release of K and P from feldspar and rock phosphate was substantially enhanced in pure culture
inoculated with bacterial strains. The different strains however, had obvious abilities to release K and P from
the minerals. The strain SBS had a higher capacity to release K and P from the minerals than others did. The
released K by this strain (490 mg/l) increased by 21-112 % while the released of P (758 mg /l) increased by 25-75
% compared with that by any of the five strains. The release of K from the feldspar was obviously affected by
pH and aerobic condition. The strain SBS had a much higher potential to release K in the pH 6.5-8.0 than other
pHs. More aerobic condition produced more K and P than a less aerobic one. Soil pot experiments with sorghum
plants indicated that the silicate dissolving bacteria could grow in the tested soils and have a potential of
minerals dissolution. The mean effect of silicate bacteria on feldspar dissolution was appreciable greater under
the tested soils (62 % of total K) rather than under bacterial culture (53 % of total K). Bacterial inoculation
combined with K and P bearing minerals gave 48, 65 and 58 % increase in dry matter yield of sorghum plants
in clay, sandy and calcareous soil respectively compared to non-inoculated soils. The uptake of K by sorghum
plants also increased by 71, 110 and 116 % while the uptake of P increased by 41, 93 and 79 % in the same soils
respectively. Residual soil fertility estimated by K and P concentration after harvest also underwent an increase
through inoculation of silicate dissolving bacteria especially with mineral additions in the tested soils.
key words: dissolution, feldspar, rock phosphate, bacillus, sorghum
INTRODUCTION Although some of these organisms are free-living (plank
Most of the Egyptian desert soils are poor or mineral surfaces , where they can impact water-rock
marginal in nutrient status and require adequate interaction, mineral surface chemistry, dissolution and
fertilization to sustain high productivity. However, due to precipitation of minerals, the evolution of ground water
economic consideration, the cost of applying imported or geochemistry and soil formation . The interaction
locally produced water-soluble fertilizers is becoming between microbes and mineral substrate can involve the
more expensive. Thus, the use of alternative indigenous oxidation or reduction of mineral constituents from which
resources such as feldspar (orthoclase) and rock the organism derives energy for growth and
phosphate are gaining importance to alleviate the reproduction . However, other types of interaction are
dependence of imported or costly commercial fertilizers. also possible and these may have impact on the diagnosis
Several laboratory studies have shown that microbes of wider range of mineral including abundant classes of
can increase the dissolution rate of silicate and aluminum silicate and aluminum-silicate minerals .
silicate minerals in laboratory patches experiments, Microbes can enhance mineral dissolution rate by
primarily by generating organic and inorganic acids . producing and excreting metabolic by-products that
[1]
tonic) in solution, most of these bacteria are attached to
[2,3]
[4-9]
[10-12]
[13,14]
Res. J. Agr. & Bio. Sci., 2(1): 5-11, 2006
6
interact with the mineral surface. Complete microbial deionized water to remove any remaining culture medium
respiration and degradation of particulate and dissolved and added in concentration of (10 CFU/ml) to modified
organic carbon can elevate carbonic acid concentration at Bromfield liquid meduim (1954): (NaH PO - 0.5 g;
mineral surfaces, in soils and in ground water , which MgSO . 7H O - 0.5 g; (NH4) SO - 1.0; NaCl - 0.2;
[4,15]
can lead to an increase in the rates of mineral weathering molasses - 5.0 g; 1000 ml distilled water; pH 7.5).
by a proton-promoted dissolution mechanism. In addition
to carbonic acid, microbes can produce and excrete Pure culture experiments: Two parallel dissolution
organic ligands by a variety of processes such as experiments were performed under laboratory conditions
fermentation and degradation of organic macromolecules, in 500 ml-Erlenmeyer flasks containing 10 g of either
or as a response to nutrient stress . feldspar or rock phosphate in 200 ml of modified Bromfield
[13,16-20]
It is well known that many organic compounds medium inoculated with either active or deead cells of
produced by microorganisms, such as acetate, citrate and silicate bacteria (NaH PO was not added to the culture in
oxalate can increase mineral dissolution rate . Carboxylic the rock phosphate experiment). The medium pH was
[21]
acid groups which shown to promote dissolution of adjusted to 6.0, 6.5, 7.0, 7.5, 8.0, and 8.5 with diluted HCl or
silicates are also common in extracellular organic materials. NaOH solutions in the case of feldspar experiment or to
Moreover, some microorganisms in soil environment 7.5 in the case of rock phosphate to avoid any effect of
contain enzymes that function in ways analogous to pH medium on rock phosphate dissolution. A control
chitinase and celluloses, i.e. they specifically break down without inoculation was also made.
mineral structure and extract elements required for All cultures were sterilized at 80C with shaking to
metabolism or structure purposes (e.g., mineralizes) . This stop the microbial metabolism after 30 days incubation.
[1]
may be especially important for ions such as Fe and Al , The cultures were further treated with 6% H O and
3+ 3+
which are expected to be rather insoluble. filtered to separate mineral powder residues from the cells.
Some recent reports showed that silicate dissolving The filtered culture solutions were digested on a hot plate
bacteria could activate soil P, K, Si reserves and promote with 30 % H O until all organic matter was oxidized. The
plant growth . Styriakova et al , reported that the concentrations of K and P in the digested solutions were
[22,23] [24]
activity of silicate dissolving bacteria played a determined with the flame and spectrophotometer,
pronounced role in the release of Si, Fe and K from respectively.
feldspar and Fe oxyhydroxides. At present, little
information about the K and P releasing condition by the Cultivation experiment: A pot culture experiment was
silicate bacteria are available. The aim in this study was to conducted in clay, sandy and calcareous soil. The basic
investigate K and P releasing condition by silicate properties of the soils are presented in (Table 2). Five Kg
bacteria and their effects on plant growth. of each soil was transferred into plastic pots. Prior to
planting, the minerals of feldspar and rock phosphate
MATERIAL AND METHODS were added to the soils at 0 and 10 g each per pot. A
Mineral perpetration: Feldspar (orthoclase) was obtained to all treatments at the rate of 1.5 g per pot in two equal
from three locations represent the sediments of potash splits. The treatments were replicated four times in a
feldspar in Eastern Desert of Egypt between Ras Benas complete randomized block design.
and Quseir. Rock phosphate was obtained from New The seeds of sorghum plants sterilized with 6% H O
Valley Project. All samples were ground and sieved and and dipped in distilled water for two days at 30 C and
the 250-m-size fraction was collected for these then air-dried. The tested bacterium SBS was identified as
experiments. The mineral powders were rinsed with a strain of (Bacillus cereus). The seeds were soaked in
distilled water to remove the fine particles. The element either fermented liquid containing SBS strain or distilled
contents of the minerals are presented in (Table 1). water for control. It was measured that there were about
Bacteria and media: Six strains of silicate dissolving seeds were planted with 5 plants per pot after thinning.
bacteria were isolated from diffrents feldspar samples Soil moisture was maintained at field capacity throughout
according to the dillution-plate methods. Prior to the course of the experiment. The plants were cut after
experimental use, these strains were initially grown in 30 days incubation and the dry matter of each treatment
Nutrient broth No. 2 at 28 C for 2 days and then was recorded. Sorghum dry matter samples were digested
harvested by centrofugation at 4000 rpm. The cultures with triacid mixture and the concentration of K and P of
were then resusbended and rinsed several times with the above-ground parts were determined using standard
10
[25]
2 4
4 2 2 4
2 4
2 2
2 2
constant dose of nitrogen in the form of urea was applied
2 2
(2x10 CFU) SBS strain per seed of sorghum. The treated
4
0
100
200
300
400
500
SBS S2 S3 S4 S5 S6
Bacterial strains
K
i
n
s
o
l
u
t
i
o
n
(
m
g
/
l
)
Living cells
Dead cells
Control
0
200
400
600
800
SBS S2 S3 S4 S5 S6
Bacreial strains
P
i
n
s
o
l
u
t
i
o
n
(
m
g
/
l
)
Living cells
Dead cells
Control
Res. J. Agr. & Bio. Sci., 2(1): 5-11, 2006
7
Table 1: Chemical analysis of the minerals (%).
Minerals K O P O SiO Fe O Na O CaO Al O SO
2 2 5 2 2 3 2 2 3 4
Orthocalse 11.25 0.12 65.23 0.16 3.68 0.26 18.72 0.41
---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
Rock phosphate 0.33 30.0 8.5 4.2 1.0 45.0 0.5 4.3
Table 2: Some basic properties of soils used in the experiment.
Soil EC pH CaCO Avail.N Avail. P Avail. K
3
dS/m % mg/kg soil mg/kg soil mg/kg soil
Clay 0.35 7.6 2.7 35 14 115
---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
Sandy 0.78 8.2 1.5 12 9 68
---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
Calcareous 1.24 8.5 28.3 17 5 47
methods . Post harvest soil samples were collected and
[26]
analyzed for K and P. The bacterial number in the
rhizosphere soil was determined by the dilution plate
method. Microbial K-releasing power from applied
feldspar was estimated in the post harvest soils and
calculated as: K-released (mg/pot) = (K-uptake + K-
residual under treatment) (K-uptake + K-residual under
corresponding treatment without feldspar addition).
RESULTS AND DISCUSSIONS
Releasing capacity in pure culture: The releases of K and
P from feldspar and rock phosphate were used as an
overall indicator of mineral dissolution during these Fig. 1: The relaese of K by different strains of silicate
experiments (Fig. 1&2). Six strains of silicate dissolving dissolving bacteria.
bacteria were obtained from the samples of feldspar.
Higher K and P concentration was observed in the culture
inoculated with bacterial strains compared to controls due
to lower resistance of the tested minerals to activity of
silicate bacteria. However, the different strains showed
various abilities on dissolution minerals and their effect
were more pronounced with rock phosphate.
The SBS strain had a higher capacity to release K
(53 % of total K) and P (58 % of total P) from the minerals
than others did. The release of K by the SBS strain (490
mg /l) increased by 21-112 % compared with that by any
of the five strains, while the release of P (758 mg /l)
increased by 25-75 %. All dead cells also released a
significant amount of K and P from the minerals. Fig. 2: The release of P by different strarins of silicate
Previous studies however, confirmed that these dissolivig bacteria.
bacterial strains produced several organic acids such as
acetic, butyric, byruvic and formic acid which can which decreased solution pH to about 3.5. The dead
specifically break down mineral structure and extract cells also had a small but significant enhancement of
elements required for metabolism or structure purpose . dissolution rates, as microbial cell surfaces contain
[1,24]
The organic acids produced by microbial colonization sites that can complex with metal ions released from
on the mineral surfaces greatly accelerated the release of mineral cultures .
mineral elements to solution from feldspar sample . In
[27]
the present dissolution experiments with silicate Effect of pH on K releasing from feldspar: Acidity
dissolving bacteria, K and P released to solution were of culture significantly influenced the bacterial
greatly increased due to the production of organic acids, activity (SBS strain) in releasing K from feldspar (Fig. 3).
[28,8]
0
100
200
300
400
500
600
pH6.0 pH6.5 pH7.0 pH7.5 pH8.0 pH8.5
K
i
n
s
o
l
u
t
i
o
n
(
m
g
/
l
)
Living cells Dead cells
0
100
200
300
400
500
500 ml 400 ml 300 ml 200 ml
Culture volume (ml)
K
i
n
s
o
l
u
t
i
o
n
(
m
g
/
l
)
Living cells Dead cells
Res. J. Agr. & Bio. Sci., 2(1): 5-11, 2006
8
Fig. 3: The release of K by SBS strain under different
pHs.
Fig. 4: The release of K by SBS strain under various
volumes in 1000 ml flask.
This strain could dissolve much more K under the
pH from 6.5-8.0 while the maximum release was obtained
at neutral pH. In this respect, Vandeviver et al , reported
[29]
that several strain of bacteria enhanced dissolution at
neutral pH in a buffered glucose media by producing
organic ligends, primarily gluconate. The releasing by the
dead cells was not significantly affected by the culture
acidity.
Effect of culture volume: The SBS bacterium was a
facultative microorganism. It can survive in both aerobic
and anaerobic conditions. However, the releasing
capacity of the isolate was different under these
conditions (Fig. 4). A small culture volume, i.e. a more
aerobic condition, had a much higher K releasing capacity
than a large culture volume. The K releasing by the dead
cells was not significantly affected by the culture
volumes.
Response of sorghum plants: The dry matter of sorghum
plants inoculated with silicate dissolving bacteria (SBS
strain) and supplied with minerals (feldspar and rock
phosphate) increased by 48, 65 and 58 % for clay, sandy
and calcareous soil respectively compared to the plants
supplied with minerals alone (Table 3). Furthermore, the
dry weight of sorghum inoculated with bacteria alone
increased by 34, 28 and 26 % respectively compared to
non-inoculated treatment. Potassium and phosphorus
uptake improved markedly in both cases with inoculation
of bacteria in the tested soils compared to corresponding
controls.
However, these bacteria were found to develop in the
rhizosphere of the sorghum plants. The release of
dissolved K and P-bearing minerals indicates that the
silicate dissolving bacteria could grow under the soil
conditions and have a potential of mineral dissolution.
After 30 days inoculation, the cells in rhizosphere of
sorghum plants treated with minerals increased to 38.6 x
10 , 30.5 x 10 and 26.8 x 10 for clay, sandy and calcareous
4 4 4
soil respectively. In comparison to non-inoculated
treatments, the release of K and P from the added minerals
was substantially enhanced with bacterial treatments. Soil
available K increased to 135 (clay), 82 (sandy) and 73
(calcareous) mg/kg soil respectively.
Soil available P also increased to 37 (clay), 32 (sandy)
and 12 (calcareous) mg/kg soil respectively. The release
of K was about 129, 242 and 306 % much higher than that
of non-inoculated treatment while the release of P was
106, 146 and 71 % in the clay, sandy and calcareous soil
respectively. On the other hand, inoculation of silicate
dissolving bacteria without mineral additions appreciably
activates the initial K and P content in all soils. The
available K content in the rhizosphere soil of sorghum
was 69, 43 and 50 % much higher than that of non-
inoculated treatments for clay, sandy and calcareous soil
respectively, while the available P content increased by
about 56, 67 and 33 % in the same soil respectively. The
most possible reason was that the inoculation of silicate
dissolving bacteria accelerated the transformation of non-
available forms of K and P into an available one.
Microbial K-releasing power was appreciably
raised in the tested soils upon bacterial inoculation
(Table 4). The mean value of released K from the
feldspar reached to (62 % of total K) in the soil culture
experiments compared to (53 % of total K) in bacterial
culture experiment. This discrepancy can be attributed to
the limited extent of microbial attachment to mineral
surfaces, the limited contact time between microbes
and minerals, or to the accelerated mineral dissolution
rates found under soil conditions. Similar result has
been reported by .
[27]
Res. J. Agr. & Bio. Sci., 2(1): 5-11, 2006
9
Table 3: Effect of silicate dissolving bacteria (SBS strain) on sorghum growth and nutrient availability in different soils.
Treatments D.W. K-uptake P-uptake Avail. K Avail. P No. of cells
mg/pot mg/pot mg/kg mg/kg x10 /g
4
Clay soil
---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
Minerals A 26.2 878 96 135 37 38.6
---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
Minerals B 17.7 513 68 59 18 4.3
---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
No minerals A 20.6 562 75 76 25 31.2
---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
No minerals B 15.4 327 58 45 16 2.5
---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
Sandy soil
---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
Minerals A 21.8 740 73 82 32 30.5
---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
Minerals B 13.2 383 36 24 13 0.8
---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
No minerals A 14.7 422 43 30 15 22.3
---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
No minerals B 11.5 283 12 21 9 0.5
---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
Calcareous soil
---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
Minerals A 19.8 657 61 73 12 26.8
---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
Minerals B 12.5 304 34 18 7 0.6
---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
No minerals A 12.8 345 25 27 8 20.1
---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
No minerals B 10.2 232 24 18 6 0.3
A: Inoculated with bacteria; B: non- inoculated.
LSD at 5% 3.24, 2.47 and 2.15 for clay, sandy and calcareous soil respectively.
Table 4: The release of potassium from feldspar added to different soils inoculated by silicate dissolving bacteria.
Soil Feldspar Initial Uptake Residual Released
mg/pot mg/pot mg/pot mg/pot
Clay + 575 878 675 611(65%)
- 575 562 380 -
---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
Sandy + 340 740 410 578(62%)
- 340 422 150 -
---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
Calcareous + 235 657 365 542(58%)
- 235 345 135 -
- Figures in parenthesis indicate percent of total K.
- Feldspar containing 9.34 % total K applied at the rate of 10 g per pot.
In soils and aquifers, most bacteria are attached to ubiquitous in the soil, plays an important role in
surfaces , therefore any dissolution enhancing destruction of K and P-bearing minerals as well as in
[2,3]
metabolites (protons, ligands, oxidants) that are produced the cycle and transformation of rocks. Mounting
are most concentrated at or near the mineral surfaces evidence, however, suggested that bacteria attached
where dissolution occurs. In laboratory experiments, the to feldspar surfaces could greatly accelerate the rate of
bacteria are primarily in suspension and any metabolites feldspar desolation to gain access to trace or limiting
produced by the bacteria are diluted by the bulk solution. nutrients for metabolic process, primarily by the
Under soil conditions, the length of contact time between production of organic ligands. The result of these
the minerals, microbe and solution is also much greater ligands excretion is the accelerated weathering of
than which can be maintain in the laboratory with bacterial feldspar . The activity of these bacteria was
culture. However, silicate dissolving bacteria greatly restricted to soil condition, K containing minerals, pH
accelerated the release of mineral elements to soil solution and incubation temperature. Potassium concentration
from feldspar and rock phosphate. in the soil also influenced the dissolving process.
The dissolution of these minerals by silicate bacteria Sheng, et al reported that silicate bacteria had the
under soil condition suggested that this bacteria specie, maximum efficiency of releasing potassium in the K
[30,27]
[23]
Res. J. Agr. & Bio. Sci., 2(1): 5-11, 2006
10
concentration 26.5-57.8 mg/kg. In addition, other factors reductive dissolution of minerals. Rev. mineral., 23,
like plants, soil fertility, moisture, temperature and soil
organisms, particularly in the field, may largely affect the
activity of microorganisms.
Finally, from the forgoing discussion, it is possible to
use the locally available feldspar mineral in combination
with silicate dissolving bacteria as a bio-fertilizer to
replace chemical fertilizer and reducing the cost of crop
production.
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