287
patterns were grouped according to a cluster analysis (UPGMA) by locality. Based
on this, representative isolates from each site whose similarity was less than 60%
were selected for subsequent analyses using the Dice coefficient (Di Rienzo et al.
2012). In total 33 isolates were selected: 14 from Bolsa Palomo, 11 from Isla Cuba,
5 from Padre Lozano, 1 from San Miguel, and 2 from Colonia Benítez.
Tolerance to water deficit was evaluated by cultivating the selected isolates
in hyperosmotic solutions of polyethylene glycol (PEG) testing water potentials
of −0.6 MPa and − 2 MPa. It was observed that isolates that tolerated greater water
deficit, correlated with greater indole compounds (IAA) production. From this set
of isolates, the 16S rRNA gene was analyzed. Alignment, sequence analysis and
construction of a phylogenetic tree were performed using the 16S rRNA gene fragment (1163 bp) with the ClustalW alignment tool of the MEGA software and the
UPGMA method (Tamura et al. 2011). The best results were obtained with two
isolates from Bolsa Palomo (Mesorhizobium and Sinorhizobium (Ensifer)), one
from Colonia Benítez (Mesorhizobium), and two from Padre Lozano (Bradyrhizobium
and Sinorhizobium (Ensifer)) (Chávez Díaz et al. 2013); all these isolates were
found to promote growth in P. alba.
In another study, Pozzi Tay (2016) evaluated drought tolerance of P. alba in symbiosis with rhizobacteria under greenhouse conditions. Using three isolates [N° 2
and N° 63 from Bolsa Palomo, (KC-759691, Mesorhizobium spp. and KC-759695,
Sinorhizobium spp.) and N° 53 from Padre Lozano (KC-759699, Bradyrhizobium
spp.)], a biofertilizer was elaborated. Seeds from two contrasting P. alba provenances with respect to precipitations were used: Santiago (27° 52′ 44″ S, 64° 9′ 16″
O, 579 mm average annual precipitation) and Campo Durán (22° 12′ 01″ S, 63° 40′
33″ W, 1054 mm average annual precipitation). A factorial design with three factors
of two levels each was used: provenance (Santiago and Campo Durán), biofertilization (with and without) and water stress (control and drought). Growth variables and
physiological responses to stress were evaluated (Pozzi Tay 2016). Under drought
stress, plants of Santiago with biofertilization conserved significantly more leaves
(node with leaf/total node) than the uninoculated plants of both provenances
(p ≥ 0.05). Furthermore, the highest values of chlorophyll and proline content were
recorded in biofertilized plants of Santiago. Biofertilized plants of both provenances
have similar values in number and weight of nodules, but under drought conditions,
these parameters were reduced, without distinction of provenances.
Summing up, the analysis of growth parameters and physiological variables
allowed to discriminate the behavior of two P. alba provenances under drought conditions in symbiosis with specific rhizobacteria from the Chaco region. Both biofertilized provenances maintained higher levels of proline and chlorophylls under
drought compared to those not inoculated. This result suggests that the establishment of symbiosis also gives adaptive advantages to water restriction. Future studies
will seek to identify the selected rhizobacterial species by sequencing other housekeeping genes as well as to analyze a possible synergistic effect of the mix of microorganisms in the biofertilizer or the prevalence of any of them for the colonization
of nodules and biological nitrogen fixation.
10 Genetic Breeding of Prosopis Species from the “Great American Chaco”
patterns were grouped according to a cluster analysis (UPGMA) by locality. Based
on this, representative isolates from each site whose similarity was less than 60%
were selected for subsequent analyses using the Dice coefficient (Di Rienzo et al.
2012). In total 33 isolates were selected: 14 from Bolsa Palomo, 11 from Isla Cuba,
5 from Padre Lozano, 1 from San Miguel, and 2 from Colonia Benítez.
Tolerance to water deficit was evaluated by cultivating the selected isolates
in hyperosmotic solutions of polyethylene glycol (PEG) testing water potentials
of −0.6 MPa and − 2 MPa. It was observed that isolates that tolerated greater water
deficit, correlated with greater indole compounds (IAA) production. From this set
of isolates, the 16S rRNA gene was analyzed. Alignment, sequence analysis and
construction of a phylogenetic tree were performed using the 16S rRNA gene fragment (1163 bp) with the ClustalW alignment tool of the MEGA software and the
UPGMA method (Tamura et al. 2011). The best results were obtained with two
isolates from Bolsa Palomo (Mesorhizobium and Sinorhizobium (Ensifer)), one
from Colonia Benítez (Mesorhizobium), and two from Padre Lozano (Bradyrhizobium
and Sinorhizobium (Ensifer)) (Chávez Díaz et al. 2013); all these isolates were
found to promote growth in P. alba.
In another study, Pozzi Tay (2016) evaluated drought tolerance of P. alba in symbiosis with rhizobacteria under greenhouse conditions. Using three isolates [N° 2
and N° 63 from Bolsa Palomo, (KC-759691, Mesorhizobium spp. and KC-759695,
Sinorhizobium spp.) and N° 53 from Padre Lozano (KC-759699, Bradyrhizobium
spp.)], a biofertilizer was elaborated. Seeds from two contrasting P. alba provenances with respect to precipitations were used: Santiago (27° 52′ 44″ S, 64° 9′ 16″
O, 579 mm average annual precipitation) and Campo Durán (22° 12′ 01″ S, 63° 40′
33″ W, 1054 mm average annual precipitation). A factorial design with three factors
of two levels each was used: provenance (Santiago and Campo Durán), biofertilization (with and without) and water stress (control and drought). Growth variables and
physiological responses to stress were evaluated (Pozzi Tay 2016). Under drought
stress, plants of Santiago with biofertilization conserved significantly more leaves
(node with leaf/total node) than the uninoculated plants of both provenances
(p ≥ 0.05). Furthermore, the highest values of chlorophyll and proline content were
recorded in biofertilized plants of Santiago. Biofertilized plants of both provenances
have similar values in number and weight of nodules, but under drought conditions,
these parameters were reduced, without distinction of provenances.
Summing up, the analysis of growth parameters and physiological variables
allowed to discriminate the behavior of two P. alba provenances under drought conditions in symbiosis with specific rhizobacteria from the Chaco region. Both biofertilized provenances maintained higher levels of proline and chlorophylls under
drought compared to those not inoculated. This result suggests that the establishment of symbiosis also gives adaptive advantages to water restriction. Future studies
will seek to identify the selected rhizobacterial species by sequencing other housekeeping genes as well as to analyze a possible synergistic effect of the mix of microorganisms in the biofertilizer or the prevalence of any of them for the colonization
of nodules and biological nitrogen fixation.
10 Genetic Breeding of Prosopis Species from the “Great American Chaco”