Phytate mineralising bacteria increase in the rhizosphere of nodulated common bean ([i]Phaseolus vulgaris[/i]) under P deficiency

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Phytate mineralising bacteria increase in the

rhizosphere of nodulated common bean ([i]Phaseolus vulgaris[/i]) under P deficiency

Rim Maougal, Alain Brauman, Claude Plassard, Josiane Abadie, A. Djakoun, Jean-Jacques Drevon

To cite this version:

Rim Maougal, Alain Brauman, Claude Plassard, Josiane Abadie, A. Djakoun, et al.. Phytate mineral-ising bacteria increase in the rhizosphere of nodulated common bean ([i]Phaseolus vulgaris[/i]) under P deficiency. 2. IBEMPA Conference ”Microorganisms for future agriculture”, Sep 2013, Seville, Spain. Universitad de Sevilla, 527 p., 2013. �hal-01268671�

Phytate mineralising bacteria increase in the rhizosphere of nodulated

common bean (Phaseolus vulgaris) under P deficiency

Maougal

1, 2

_{R.T., Brauman}

3

_{A., Plassard}

1

_{C., Abadi J}

1

_{., Djakoun}

2

_{A. and Drevon}

1

_J-J.

maougalr@gmail.com

1 _{UMR Eco&Sols - Place Viala - 34000 Montpellier - 34 – France}

2 _{Laboratoire  de  Génétique,  Biochimie  et  Biotechnologies  Végétales,  Département  de  Biologie  et  d’Ecologie,  Université  Constantine 1, Route de Ain el Bey, Constantine, Algeria.}

3 _{IRD, Eco&Sols, Land Department Development, Bangkok, Thailande}

A more sustainable agriculture based on the reduction of mineral fertilizers as Inorganic phosphate

P is one of the most limiting nutrients for plant growth in soil.

Depletion in P

Eutrophication

An urgent need

to reduce P inputs

Phaseolus Vulgaris RIL 147:

chosen as model plant .

Sterilised seeds were grown in pots under greenhouse.

A comparison between P deficient soil (P-: 15mg/kg) and control soil (P+: 50 mg/kg)

Have a great P acquisition by its nitrogen fixation

Extraction of rhizospheric bacteria and determination of isolates able to mineralise phytate

Density: bacterial colony counts and isolation.

Culture on a specific medium containing phytate as a sole source of P. Bae and al 1999.

The capacity of the isolates to release Pi from Na-phytate in broth was assayed.

3

1

2

1-Density of the culturable bacterial

Bacteria able to grow on the modified Angle medium represented 1% of total number of bacteria that were able to grow on the rich medium (LB).

Under P15, the density of phytate-utilizing bacteria on Angle medium was 10 fold higher in rhizospheric soil than in bulk soil, whereas under P50 there was no significant difference

selection of phytate-mineralizing bacteria in the rhizosphere of P-deficient bean.

2-The capacity of the isolates to release Pi from Na-phytate in broth

Translucent medium around the bacteria = solubilisation of phytate by acidification.

Objective: The aim of this work is to characterize the functional microbial community involved in the mineralization of phytate in the rhizosphere of a leguminous plant (Phaseolus

vulgaris)

I- Context:

Conclusion:

our results indicate that nodulated-bean plants can increase in case of P deficiency, the phytase

activities of a diverse community of phytase hydrolysing bacteria within their rhizosphere.

References:

1- Cordell, Dranger and White.2009. The story of phosphorus: Global food security and food for thought

2- Huoqing Huang, Pengjun Shi, Yaru Wang, Huiying Luo, Na Shao, Guozeng Wang, Peilong Yang, and Bin Yao.2009.Diversity of Beta-Propeller

Phytase Genes in the Intestinal Contents of Grass Carp Provides Insight into the Release of Major Phosphorus from Phytate in Nature

3- H.D. Bae , L.J. Yanke, K.-J. Cheng , L.B. Selinger.1999.A novel staining method for detecting phytase activity

Bacterial communities Replace the mineral P by the

organic P wich constitute up to 80% of total P.

One of the dominant organic P is the phytate

Ability of certain microorganisms to increase the concentration of soluble P by degrading phytate.

B

C

44 0,0 0,4 0,8 1,2 0 24 48 72 96 120 144 168 OD ( 6 3 0 n m ) 0 100 200 300 400 500 Pi c o n c e n tr a ti o n ( µ M) 34 0,0 0,4 0,8 1,2 OD ( 6 3 0 n m ) 0 100 200 300 400 500 Pi c o n c e n tr a ti o n ( µ M) OD ( 6 3 0 n m ) Pi c o n c e n tr a ti o n ( µ M) 0,0 0,4 0,8 1,2 0 100 200 300 400 500 Bacillus subtilis 111b

Time (h)

A

B

C

44 0,0 0,4 0,8 1,2 0 24 48 72 96 120 144 168 OD ( 6 3 0 n m ) 0 100 200 300 400 500 Pi c o n c e n tr a ti o n ( µ M) 44 0,0 0,4 0,8 1,2 0 24 48 72 96 120 144 168 OD ( 6 3 0 n m ) 0 100 200 300 400 500 Pi c o n c e n tr a ti o n ( µ M) 34 0,0 0,4 0,8 1,2 OD ( 6 3 0 n m ) 0 100 200 300 400 500 Pi c o n c e n tr a ti o n ( µ M) 34 0,0 0,4 0,8 1,2 OD ( 6 3 0 n m ) 0 100 200 300 400 500 Pi c o n c e n tr a ti o n ( µ M) OD ( 6 3 0 n m ) Pi c o n c e n tr a ti o n ( µ M) 0,0 0,4 0,8 1,2 0 100 200 300 400 500 Bacillus subtilis 111b OD ( 6 3 0 n m ) Pi c o n c e n tr a ti o n ( µ M) 0,0 0,4 0,8 1,2 0 100 200 300 400 500 Bacillus subtilis 111b

Time (h)

A

II- Materials and methods:

III-Results

Phytate utilizing soil bacteria

ph y tate uti li sing bac ter ia

[Lo g CFU (g DW soi l) -1] a b a _a 3 4 5 P 50 P 15

Their phytase activity was measured on bacterial pellet and supernatant after a 72 h incubation period according to Shimizu (1992) with modifications. + 50% of total P Pi (P)  limiting nutriment P organique Phytase Phytate

Rhizosphere

Growth of isolates on screening phytate medim.

3-The phytase activity

Kinetics of growth ( ) and concentration of Pi ( ) in the growth medium for rhizobacterial isolates with Na-IHP as sole source of P. for A, Bacillus subtilis 111b as representative of group II; B, isolate 34 as representative of group II; C, isolate 44 as representative of group III. Data are means ± sd of 3 replicated cultures. 2 0 0,2 0,4 0,6 0,8 1 1,2 1,4 1,6 1,8 2 1 22 20 18 17 23 3 2 13 9 14 12 15 16 II III I Ph yta se ac tiv ity (nm ol Pi ) B 47 48 52 31 44 43 59 40 38 42 36 34 0 0,2 0,4 0,6 0,8 1 1,2 1,4 1,6 1,8 II III I Ph yta se ac tiv ity (nm ol Pi ) A 2 0 0,2 0,4 0,6 0,8 1 1,2 1,4 1,6 1,8 2 1 22 20 18 17 23 3 2 13 9 14 12 15 16 II III I Ph yta se ac tiv ity (nm ol Pi ) B 0 0,2 0,4 0,6 0,8 1 1,2 1,4 1,6 1,8 2 1 22 20 18 17 23 3 2 13 9 14 12 15 16 II III I Ph yta se ac tiv ity (nm ol Pi ) B 47 48 52 31 44 43 59 40 38 42 36 34 0 0,2 0,4 0,6 0,8 1 1,2 1,4 1,6 1,8 II III I Ph yta se ac tiv ity (nm ol Pi ) 47 48 52 31 44 43 59 40 38 42 36 34 0 0,2 0,4 0,6 0,8 1 1,2 1,4 1,6 1,8 II III I Ph yta se ac tiv ity (nm ol Pi ) A

Extracellular (left) and Cell-associated phytase activity (right) of rhizobacterial isolates. A: on P 50, B: on P15.

A significant accumulation of Pi in the culture medium was observed for 26 among 34 isolates

The phytase activity varied considerably among isolates of the 3 above groups of curves (figure 2). A majority of isolates produced extra-cellular phytase.

However, some isolates showed additional or exclusive cell-associated phytase.

phytate could be mineralized either externally or within the periplasm of the bacteria; these two conditions seem to involve different phytases.