ФИЗИОЛОГИЯ РАСТЕНИЙ, 2012, том 59, № 1, с. 129-133
ЭКСПЕРИМЕНТАЛЬНЫЕ СТАТЬИ
УДК 581.1
The Role of Rhizobacteria in Salinity Effects on Biochemical Constituents of the Halophyte Sesuvium portulacastrum1
© 2012 R. Anburaj, M. A. Nabeel, T. Sivakumar, K. Kathiresan
Center of Advanced Study in Marine Biology, Annamalai University, Parangipettai, 608502, Tamil Nadu, India
Received June 23, 2010
An experiment was conducted to understand the role of rhizospheric microorganisms in salinity effects on growth, antioxidants, pigments, and ion concentrations in the halophyte Sesuvium portulacastrum L. The plants grown in non-sterilized soil exhibited the enhanced growth rate, suppressed antioxidant enzymes, increased contents of chlorophylls and carotenoids, the greater accumulation of sodium and the reduction in the potassium ion concentration, as compared with the plants raised in microbe-free soil. The dominant microbes identified from the rhizophere soil of non-sterilized plant groups included Bacillus cereus, Aeromonas hydrophila, Pseudomonas aeruginosa, Corynebacteriumxerosis, and Escherichia coli. The work emphasizes the importance of the rhizobacteria that colonize the root at the interface with soil in preventing the deleterious effects caused by salinity through accumulation of sodium and pigments and reduction of antioxidants and potassium.
Keywords: Sesuvium portulacastrum - mangrove associate - halophyte - antioxidants - rhizobacteria - salinity
INTRODUCTION
Sesuvium portulacastrum belonging to the family Aizoaceae is a dicotyledonous facultative halophyte naturally growing in the subtropical, Mediterranean, coastal, and warmer zones of the world [1]. This plant has food and medicinal values and is also utilized as a wild vegetable, fodder crop for the cattle and domestic animals (goats, sheeps, and camels) and as bait in crab traps [2]. The plant has a remarkable ability to survive under stress conditions of salinity, drought, and heavy metal accumulation [1]. It is also considered as a pioneer species in the environmental protection, such as phyto-bioremediation sand dune fixation, desalination, desert greening, production of cheap biomass for renewable energy, climate improvement through CO2 sequestration, landscaping, and as an ornamental plant with its attractive pink-purplish flowers [1-3].
Soil salinity is a growing problem due to a variety of natural and man-made factors. Seawater intrusion into coastal aquifers is one of the causes for salinity problems in coastal regions in almost all the regions of the world. One of the broad criteria for reclamation of saline soil is growing salt-tolerant wild plants in the arid and semi-arid regions with the conventional crops, which can accumulate excessive amounts of salt from soil and will improve the fertility of soil. S. portulacas-
1 This text was submitted by the authors in English.
Corresponding author. Kandasamy Kathiresan. Center of Advanced Study in Marine Biology, Annamalai University, Parangipettai, 608502, Tamil Nadu, India. Fax. + 91-41-44-24-3555; e-mail. kathirsum@rediffmail.com
trum is among the most appropriate species for the usage on salt-affected soils in arid and semi-arid regions. In India, it grows at the eastern and western coastal sides as a mangrove associate [1].
The soil bacteria growing at the soil-root interface, which are beneficial for the plants, are termed as plant growth-promoting rhizobacteria (PGPR). They are particularly important in controlling the chemical environment of the ecosystems, and they are also considered as the main primary producers, as well as being secondary producers and consumers. They perform different functions in the coastal ecosystems, such as photosynthesis, nitrogen fixation, and methanogene-sis [3]. However, not many studies are currently available principally focusing on microbial aspects in relation to salt stress of S. portulacastrum. Hence, the present study was aimed at the study of the salinity-induced changes in antioxidant and pigment contents, and also sodium and potassium uptake by the plant in relation to rhizobacterial presence.
MATERIALS AND METHODS
Study area and experimental plants. Sesuvium portulacastrum L. plants were collected freshly along with their habitat soil from Pichavaram mangrove forest (11°29' to 11°30' N and 79°45' to 79°55' E), southeast coast of India. The plant materials were purified from rhizosphere soil; the roots were disinfected for 5 min in saturated calcium hypochlorite solution and rinsed thoroughly with distilled water. The rhizosphere soil samples were separately collected from the same
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Changes in mean relative growth rate of Sesuvium portula-castrum during 30 (1) and 60 (2) days of experiment.
habitat. The plant materials and soil samples were transported in sterile polythene bags to the laboratory.
Experimental groups. The healthy plants of apparently uniform size were selected and maintained in the sterile polythene bag with rhizosphere soil. The entry of atmospheric microbes was blocked by closing the mouth of the bags. The plant stocks were separated into six different groups of soil treatments as follows:
group I: non-sterilized rhizosphere soil watered with sterilized water;
group II: sterilized rhizosphere soil watered with sterilized water;
group III: non-sterilized rhizosphere soil watered with sterilized 50% seawater;
group IV: sterilized rhizosphere soil watered with sterilized 50% seawater;
group V: non-sterilized rhizosphere soil watered with sterilized 100% seawater;
group VI: sterilized rhizosphere soil watered with sterilized 100% seawater.
Each group consisted of five replicates. The experimental plants were maintained under the natural sunlight for a period of 60 days. Sterilized water was poured to plant stocks two times a day.
The plants were analyzed for various parameters on the 60th day of experiment, except mean relative growth rate (RGR), which was measured on 30th and 60th days of experiment.
Microbial analysis. For microbial analysis, a known weight of soil sample was aseptically removed from the roots using a sterilized spatula. It was then transferred to a sterile conical 150-ml flask containing 99 ml of sterile diluent, and serial dilution was performed to get 10-1, 10-2, 10-3, 10-4, and 10-5 suspension samples. The samples were used to isolate total Heterotrophic Bacteria by spread plate method on Zobell marine agar medium (Hi-media, Mumbai) and enumerated as Colony Forming Units (CFU) per gram of the sample. The dominant bacteria present in the soil were identified by using biochemical and culture characteristics according to the Bergey's Manual [4].
Concentrations of Na+ and K+ in leaf samples were determined using a flame photometer [5]. The total chlorophyll and carotenoids were extracted with ice-cold 80% acetone from the leaves and estimated as described in [6]. Antioxidants, such as ascorbic acid [7], a-tocopherol [8], reduced glutathione [9], superoxide dismutase [10], and ascorbate peroxidase [11] were also analyzed using standard procedures.
Statistical analysis was carried out using SPSS version 17.0. To test the statistical significance, two-way analysis of variance (ANOVA) was performed using Duncan's Multiple Range Test (DMRT).
RESULTS
The relative growth rate varied significantly between the groups during 30 and 60 days of experiment (P < 0.05) (figure). In general the non-sterilized plants showed the higher growth rates as compared with the sterilized plants. The increasing salinity also showed decreasing growth rates, and hence the lowest growth rate was observed when 100% seawater was applied.
The total heterotrophic bacterial counts reduced from 10.5 to 1.9 CFU/g when seawater was treated to the soil. There was 3.75-fold reduction in counts in soil treated with 50% seawater and 5.5-fold reduction due to 100% seawater. Thus, salinity reduced the mi-crobial counts in the soil. Microbes could not be enumerated in the sterilized soil throughout the experiment (table 1). The dominant microbes associated with rhizosphere soil were identified as Bacillus cereus, Aeromonas hydrophila, Pseudomonas aeruginosa, Corynebacterium xerosis, and Escherichia coli. The morphological and biochemical characteristics of the microbes used for identification are given in table 2.
There was a significant accumulation of Na+ in leaves of plants under non-sterile conditions as compared to microbe-free conditions. However, the trend was reverse in the case of K+ ion, since there was a reduction K+ ion content under non-sterile conditions. The accumulation of Na+ in leaves increased, while that of K+ ion decreased with increasing salinity. There was a significant increase in total chlorophylls and car-otenoid pigments in the leaves of plants raised in non-sterilized soil than in sterilized soil. The total chlorophylls and carotenoids showed a decreasing trend with an increase in salinity. The levels of both enzymatic and non-enzymatic antioxidants increased in the leaves of plants growing in microbe-free sterilized soil, but the non-sterilized soil resulted in a decreased antioxidant levels in plants. However antioxidants manifested an increasing trend with an increase in salinity (table 1).
DISCUSSION
Seawater intrusion into coastal ecosystems is one of the causes for salinity problems in almost all the re-
Table 1. Pigments, biochemical constituents, sodium and potassium ions in the leaves and microbial counts in soil of Sesuvium portulacastrum raised for 60 days in sterilized (S) and non-sterilized (NS) soil watered with freshwater, 50%, and 100% seawater
Variables 0% seawater 50% seawater 100% seawater
NS S NS S NS S
Total heterotrophic bacteria in soil, CFU/g 10.5 ± 2.4a 0b 2.8 ± t 0.8c 0b 1.9 ± 0.5c 0b
Total chlorophylls, mg/g fr wt 3.5
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