Halotolerant Plant Growth-Promoting Bacteria Colonization of Agronomic Crops Under Saline Stress
The ability of a plant growth-promoting bacteria (PGPB) strain to colonize the roots and tissues of inoculated plants is important for their successful use in agricultural practices. The purpose of this study was to determine how effective 15 indigenous halotolerant PGPB were at colonizing three different agronomic crops via seed inoculation. Using standard Hoagland’s media and Hoagland’s media amended with 100 mM NaCl, we tested 15 gfp-tagged halotolerant bacterial isolates for their ability to colonize rice, maize, and soybean seedlings. The quantitative dilution plating method and fluorescent microscopy were used to determine the colonization degree of gfp-tagged halotolerant PGPB isolates in the rhizoplane zone and in the inner tissue of the seedlings at 21 days after germination. All halotolerant PGPB isolates colonized the rhizoplane zone of all seedlings. In both standard and 100 mM NaCl amended Hoagland’s media, isolates E194-3, D183-4, and E101-1 showed the highest colonization in rice, maize, and soybean seedlings, respectively. The ability of halotolerant PGPB isolates to colonize agronomic crops was found to vary depending on bacterial isolates, plant species, plant tissues, and NaCl concentration.
Keywords: inoculation, colonization, halotolerant, endophyte, rhizoplane
 Das S, Ho A, Kim PJ. Editorial: Role of microbes in climate smart agriculture. Frontier in Microbiology. 2019;10 (2756): 1-3. https://doi.org/10.3389/fmicb.2019.02756
 Raza A, Ashraf F, Zou X, Zhang X, Tosif H. Plant ecophysiology and adaptation under climate change: Mechanisms and perspectives Hasanuzzaman MI, editor. Springer Nature Singapore Pte Ltd. 2020. https://doi.org/10.1007/978-981-15-2156-0_5
 Sangiorgio D, Cellini A, Donati I, Pastore C, Onofrietti C, Spinelli F. Facing climate change: Application of microbial biostimulants to mitigate stress in horticultural crops. Agronomy. 2020;10 (794): 1-24. https://doi.org/10.3390/agronomy10060794
 Mayak S, Tirosh T, Glick BR. Plant growth-promoting bacteria confer resistance in tomato plants to salt stress. Plant Physiology and Biochemistry. 2004;42:565-572.
 Zahir ZA, Ghani U, Naveed M, Nadeem SM, Asghar HN. Comparative effectiveness of Pseudomonas and Serratia sp. containing ACC-deaminase for improving growth and yield of wheat (Triticum aestivum L.) under salt-stressed conditions. Archives Microbioly. 2009;191:415-424.
 Rojas-Tapias D, Moreno-Galvan A, Pardo-Diaz S, Obando M, Rivera D, Bonilla R. Effect of inoculation with plant growth promoting bacteria (PGPB) on amelioration of saline stress in maize (Zea Mays). Applied Soil Ecology. 2012;61:264-272.
 Bal HB, Nayak L, Das S, Adhya T. Isolation of ACC deaminase producing PGPR from rice rhizosphere and evaluating their plant growth promoting activity under salt stress. Plant and Soil. 2013;366:93-105.
 Ansari FA, Ahmad I, Pichtel J. Growth stimulation and alleviation of salinity stress to wheat by the biofilm forming Bacillus pumilus strain FAB10. Applied Soil Ecology. 2019;3:45–54.
 El-Esawi MA, Alaraidh IA, Alsahli AA, Alamri SA, Ali HM, Alayafi AA. Bacillus firmus (SW5) augments salt tolerance in soybean (Glycine max L.) by modulating root system architecture, antioxidant defense systems and stress-responsive genes expression. Plant Physiology and Biochemistry. 2018;132:375–384.
 Atouei TM, Pourbabaee AA, Shorafa M. Alleviation of salinity stress on some growth parameters of wheat by exopolysaccharide-producing bacteria. Iranian Journal of Science and Technology, Transaction A, Science. 2019;43:2725–2733.
 Ilyas N, Mazhar R, Yasmin H et al. Rhizobacteria Isolated from saline soil induce systemic tolerance in wheat (Triticum aestivum L.) against salinity stress. Agronomy. 2020;10(989): 1-20.
 Shultana R, Zuan AT, Yusop MR, Saud HM. Characterization of salt-tolerant plant growth-promoting rhizobacteria and the effect on growth and yield of saline-affected rice. PLoS ONE. 2020;15(9): 1-16.
 Hallmann J, Quadt-Hallmann A, Mahaffee WF, Kloepper JW. Bacterial endophytes in agricultural crops. The Canadian Journal of Microbiology. 1997;43:895-914.
 Rosenblueth M, Martinez-Romero E. Rhizobium etli maize populations and their competitiveness for root colonization. Archives. Microbiology. 2004;181:337-344.
 Hallmann J, Berg G. Spectrum and population dynamics of bacterial root endophytes. In Microbial Root Endophytes; Schulz, B.J.E., Boyle, C.J.C., Sieber, T.N., Eds.; Springer. Dordercht, The Netherlands. 2006; 15–31.
 Lodewyckx C, Vangronsveld J, Porteus F et al. Endophytic bacteria and their potential applications. Critical Reviews in Plant Sciences. 2002;21:586-606.
 Sturz AV, Nowak J. Endophytic communities of rhizobacteria and the strategies required to create yield enhancing associations with crops. Applied Soil Ecology. 2000;15:183-190.
 Naveed M, Mitter B, Yousaf S, Pastar M, Afzal M, Sessitsch A. The endophyte Enterobacter sp. FD17: A maize growth enhancer selected based on rigorous testing of plant beneficial traits and colonization characteristics. Biology and Fertility of Soils. 2013;50:249-262.
 Timmusk S, Behers L, Muthoni J, Muraya A, Aronsson AC. Perspectives and challenges of microbial application for crop improvement. Frontiers in Plant Science. 2017;8 (49):1-10.
 Meena KK, Bitla UM, Sorty AM et al. Mitigation of salinity stress in wheat seedlings due to the application of phytohormone-rich culture filtrate extract of methylotrophic Actinobacterium nocardioides sp. NIMMe6. Frontiers in Microbiology. 2019;11(2091): 1-16.
 Zakria M, Udonishi K, Ogawa T, Yamamoto A, Saeki Y, Akao S. Influence of inoculation technique on the endophytic colonization of rice by Pantoea sp. isolated from sweet potato and by Enterobacter sp. isolated from sugarcane. Soil Science and Plant Nutrition. 2008; 54: 224–236.
 Lacava PT, Azevedo JL. Endophytic Bacteria: A Biotechnological Potential in Agrobiology System. In: Maheshwari, D., Saraf, M., Aeron, A. (eds) Bacteria in Agrobiology: Crop Productivity. Springer, Berlin, Heidelberg. 2013; 1-44. https://doi.org/10.1007/978-3-642-37241-4_1
 Compant S, Reiter B, Sessitsch A, Nowak J, Clément C, Barka E. Endophytic colonization of Vitis vinifera L. by plant growth-promoting bacterium Burkholderia sp. strain PsJN. Applied and Environmental Microbiology Journal. 2005;71:1685-1693.
 Maa Y, Rajkumar M, Luoa YM, Helena F. Inoculation of endophytic bacteria on host and non-host plants—Effects on plant growth and Ni uptake. Journal of Hazardous Materials. 2011;195(2011):230–237.
 Schulz B, Römmert AK, Dammann U, Aust HJ, Strack D. The endophyte-host interaction: A balanced antagonism. Mycological Research. 1999;103:1275-1283.
 Rosenblueth M, Martínez-Romero E. Bacterial endophytes and their interaction with hosts. Molecular Plant-Microbe Interaction. 2006;19:827-837.
 de Zelicourt A, Al-Yousif M, Hirt H. Rhizosphere microbes as essential partners for plant stress tolerance. Molecular Plant. 2013;6:242-245.
 Hallmann J. Plant Interactions with Endophytic Bacteria. In: Jeger, M.J. & N.J. Spence (Eds) Biotic Interactions in Plant–Pathogen Associations, CABI Publishing, Wallingford, United Kingdom. 2001; 87-119.
 Verma SC, Ladha JK, Tripathi AK. Evaluation of plant growth promoting and colonization ability of endophytic diazotrophs from deep water rice. Journal of Biotechnology. 2001;91:127-141.
 Truyens S, Weyens N, Cuypers A, Vangronsveld J. Bacterial seed endophytes: Genera, vertical transmission and interaction with plants. Environmental Microbiology Reports. 2014;7:40-50.
 Sambrook J, Fritsch EF, Maniatis T. Molecular cloning. 2nd ed. New York: Cold Springs Harbor Laboratory; 1999
 Yanni YG, Rizk RY, Corich V et al. Natural endophytic association between Rhizobium leguminosarum bv. trifolii and rice roots and assessment of its potential to promote rice growth. Plant Soil. 1997;194:99-114.
 O’Callaghan M. Microbial inoculation of seed for improved crop performance: Issues and opportunities. Applied Microbiology and Biotechnology. 2016;100:5729–5746.
 Rocha I, Ma Y, Souza-Alonso P, Vosátka M, Freitas H, Oliveira RS. Seed coating: A tool for delivering beneficial microbes to agricultural crops. Frontiers in Plant Science. 2019;10:1357.
 Compant S, Mitter B, Colli-Mull JG, Gang H, Sessitsch A. Endophytes of grapevine flowers, berries, and seeds: Identification of cultivable bacteria, comparison with other plant parts, and visualization of niches of colonization. Microbial Ecology. 2011;62:188-197.
 Hallmann J, Berg G. Microbial root endophytes. Schulz BJE, Boyle CJC, Sieber TN, eds. Dordercht: Springer; 2006.
 Rosenblueth M, Martinez-Romero E. Rhizobium etli maize populations and their competitiveness for root colonization. Archives Microbiology. 2004;181:337-344.
 Kandel SL, Joubert PM, Doty SL. Bacterial endophyte colonization and distribution within plants. Microorganisms. 2017;5(77): 1-26.
 Compant S, Kaplan H, Sessitsch A, Nowak J, Barka E, Clement C. Endophytic colonization of Vitis vinifera L.by Burkholderia phytormans strain PsJN: From the rhizosphere to inflorescence tissue. FEMS Microbiology Ecology. 2007;63:84-93.
 Hansen ML, Kregelund L, Nybroe O, Sorensen J. Early colonization of barley roots by Pseudomonas fluorescens studied by immunofluorescence technique and confocal laser scanning microscopy. FEMS Microbiology Ecology. 1997;23:353-360.
 Soldana R, Mapellia F, Crottia E et al. Bacterial endophytes of mangrove propagules elicit early establishment of the natural host and promote growth of cereal crops under salt stress. Microbiological Research. 2019;223-225:33-43.