Pseudomonas fluorescens, PGPR, Inoculation, Siderophore.

Increasing agricultural production is one of the biggest challenges we face in the twenty-first century. The environment has also been eroded by an increase in the overuse of chemical fertilizers and pesticides. In order to reduce crop damage brought on by plant pathogens, the biological control measure thus emerges as a nonhazardous tactic (Weller 1998; Cook et al., 1995). Biologicals are readily available models that are an optional means of combating plant diseases. Research on the relationship between plants and pathogens has advanced significantly during the previous 20 years (Huang et al., 2013).

According to Shurtleff and Averre (1997), "Biological control" or "biocontrol" refers to an organism's direct or indirect involvement in limiting the spread of a pathogen and lessening the severity of disease. Microbes associated with plants play a dual role in stress tolerance, both biotic and abiotic, in addition to stimulating plant development (Welbaum et al., 2004; Jain, 2012). Worldwide, plant diseases are the main reason for yield loss (Alzandi and Naguib 2019). Utilizing biocontrol agents helps to reduce the use of chemicals while increasing the likelihood of disease resistance. The use of Pseudomonas fluorescens as a biocontrol agent and plant growth promoter necessitates a thorough understanding of how these bacteria interact with one another and function in the rhizosphere (Kloepper et al., 1989; Gotz et al., 2006; Panpatte et al., 2016). 

Among the reported genera of Gram-negative bacteria, the Pseudomonas genus is the most significant (Gomila et al., 2015). According to (Palleroni, 2008), the Pseudomonas belongs to the Order Pseudomonadales and Family Pseudomonadaceae, which are motile rods with a Gram-negative reaction that do not sporulate and contain one or more polar flagella. The excellent plant growth-promoting rhizobacteria (PGPR) Pseudomonads fluorescence acts as a biocontrol agent for a variety of phytopathogens and pests. In addition to its extensive use in the agricultural sector, Fig. 1 (Silby et al., 2011; Scales et al., 2014) clearly illustrates its numerous uses in the medical, industrial, bioremediation, environmental, and commercial sectors. By supplying minor or major phytopathogens, these bacterial biocontrol agents promote plant growth by causing the production of growth-promoting metabolites like auxins and gibberellins (Salt 1979; Fravel 2005). This review discusses bacterial antagonist-controlled in various agricultural crops as well as effects of Pseudomonas flourescens on growth parameters.

Mechanisms of bio-control agent Pseudomonas fluorescens against plant pathogens. For biological control to be effective, antagonists must be able to successfully colonize the roots (Weller 1998; Parke 1990). They also produce some toxic secondary metabolites that are antagonistic (Defago and Haas 1990). By using a variety of mechanisms and producing a variety of metabolites, such as siderophores (Kloepper et al., 1980; Leong, 1986; Schippers et al., 1987), antibiotics (Fravel, 1988; Thamashow and Weller 1990; Keel et al., 1992), and other compounds, these helpful rhizobacteria inhibit the growth of plant pathogens.

Fig. 1. Multidimensional use of Pseudomonas spp. in various sectors highlighting its versatility as a successful colonizer due to its various functional abilities (Source: Silby et al., 2011; Scales et al., 2014).

A Pseudomonas fluorescens's biological control mechanism, a wide variety of rhizosphere microorganisms have been described, examined, and studied recently for their potential to act as biocontrol agents against soil-borne phyto pathogens. By producing antibiotics, a variety of enzymes, side rophores, and occasionally by inducing systemic resistance that stimulates general defense in the host plants, microorganisms can limit or reduce the damage caused by phyto pathogens. Additionally, it has been discovered that the microbes serve as pathogens' competitors for vital nutrients and ecological niches (Panpatte et al., 2016). The fluorescent Pseudomonas exerts its bio-controlling effects by using the following mechanisms:

Antibiosis through production of Antibiotics and secondary metabolites. Since they are simple to isolate from the soil, bacterial bio-control agents have been found to produce antibiotics and are crucial in the management of a variety of plant diseases. Numerous variables, including pH, temperature, and the concentrations of various metal ions, particularly zinc Zn2+, can have an impact on the production of antibiotics (Duffy and Defago 1997). The potential to control soil- and seed-borne pathogenic fungi, as well as oomycetes, has drawn attention to some fluorescent pseudomonad strains (Keel et al., 1992; Keel et al., 1996).

Fewer Pseudomonas spp. biocontrol strains produce diffusible or/and volatile antibiotics that can inhibit pathogens in vitro than effective biocontrol strains (Haas et al., 2002). The peculiar ability of fluorescent pseudomonas to synthesize a variety of antibiotics helps to maintain plant pathogens and boosts the organism's own vigor (Mazzola et al., 1992; Gaur et al., 2004). However, strain-to-strain variations are also seen to exist (Raaijmakers et al., 2002). A number of these antibiotics have broad-spectrum activity. Secondary metabolite development is greatly aided by the moderate rate of bacterial growth in the rhizosphere (Haas and Defago 2005).

Table 1: Volatile antibiotics released by Pseudomonas fluorescens that are effective against diseases.

Competition for space and nutrients. The plants are the main source of nutrients for the soil microbes, which rely heavily on it. The surface of the rhizosphere acts as an extraordinary carbon sink and collects a wide range of nutrients, including water, hydrogen ions, iron, oxygen, enzymes, vitamins, and other significant secondary metabolites. Because the root contains a lot of nutrients, the various types of microorganisms it draws into it compete with one another for nutrients and space. 

By increasing competition for nutrients like carbon, nitrogen, oxygen, iron, or water, which hinders the ability of the fungal pathogens to disperse in the soil rhizosphere, fungal pathogens can be eliminated from the soil (Leong 1986; Loper and Buyer 1991). In these circumstances, Pseudomonas fluorescens can adapt and compete with the pathogens, reducing their chances of surviving (Rovira, 1969).

Therefore, it is important to evaluate the bacterium's capacity for colonizing roots, which in turn offers some defense against soil-borne pathogens. Root colonization is the process by which rhizobacteria introduced into seeds, vegetative plant parts, or soil are dispersed among roots growing in raw soil, multiply, and survive for a period of time in the presence of native soil macroflora, according to Weller et al. (2002).

Iron limitation is an illustration of nutrient competition because iron is a growth factor for all organisms. In neutral and alkaline soils, there is an iron limitation. The ferrous ion can be used by P. fluorescens to produce siderophores. It gives an advantage over other phyto pathogens that lack effective systems for iron binding and uptake. In comparison to their wild type parental strains, which were able to produce siderophores, siderophore deficient mutants were found to be less effective against phyto pathogens (Bakker et al., 1986).

Fig. 2. Pseudomonas fluorescens adapting and competing with the pathogens, reducing their chances of surviving (Rovira, 1969) & Competition for space and nutrients.

Table 2: Induced systemic resistance determinants by various Pseudomonas fluorescens strain in different plants.

Induced Systemic Resistance. The host defense mechanism of the host plants is activated by biochemical changes caused by biocontrol agents, which confers resistance to pathogen infections (Nega, 2014; Upadhyay et al., 2021). Microbe associated molecular pattern (MAMP) refers to the stimulus produced by microorganisms; whereas pathogen associated molecular pattern (PAMP) refers to the stimulus produced by plants (Kohl et al., 2019; Upadhyay et al., 2021). According to (Van et al., 1998), induced resistance is the improved defensive capacity that a plant develops in response to a specific biotic or chemical stimulus. ISR, also known as induced systemic resistance, is a generalized plant immune response that is triggered by beneficial plant bacteria that coexist with plant roots. According to research by (Nguyen et al., 2020), Pseudomonas spp. have been shown to prime various plant species to produce ISR against different pathogens.       

Table 2 lists a small number of fluorescent Pseudomonads that can cause the ISR response in plants to protect them from a variety of phytopathogen infections (Van Wees et al., 1997; Kamilova et al., 2005). After such a response, immunized plants exhibit defense responses that are quicker and stronger when a pathogen attacks, resulting in an improved level of protection. ISR is a mode of action of plant growth promoting rhizobacteria (PGPR), primarily of the fluorescent Pseudomonads group that aids in the suppression of diseases by inducing ISR response in the plants, as independently discovered in 1991 by Van Peer and his research team in the Netherlands (Van Peer et al., 1991).

Plants may develop defense mechanisms in response to flagellum, a protein found in bacterial flagella. Flagella's role in ISR in Arabidopsis thaliana was investigated for P. putida strain WCS358 and it was discovered that it induced an ISR response against P. Syringae tomato. According to Gomez-Gomez and Boller (2000); Zipfel et al. (2004); Meziane et al. (2005), the mutant of WCS358 that was devoid of flagella was also responsible for inducing ISR. This suggests that the mutant may have also contained other determinants that can trigger ISR in the Arabidopsis thaliana plant. 

LPS from the inradish, P. fluorescens WCS374 strain was crucial in the ISR against the pathogen F. oxysporum that causes Fusarium wilt. Radish seeds were treated with strain WCS374 as a seed treatment, which reduced Fusarium wilt by an average of 42% and increased yield by an average of 45%. A study that applied isolated LPS along with the use of mutant strains that were deficient in their LPS's O-antigenic side chain offered additional proof of the involvement of LPS in ISR. 

Application of LPS helped in triggering ISR in the radish plant, which helped in the significant reduction of fusarium wilt of radish (Leeman et al., 1995). The mutants lacking O-antigen were unable to elicit ISR response in radish for reduction of disease incidence. In a different study on bean and tomato plants, the WCS358 strain mutant lacking the O-antigen was unable to activate the ISR in bean and tomato plants, whereas application of LPS was successful in doing so (Meziane et al., 2005). The Pseudobactin siderophore of P. putida WCS358 strain was in charge of ISR in the management of bacterial wilt in eucalyptus. In contrast to the mutant lacking the pseudobactin siderophore, penetration of leaves with the bacterium WCS358 or purified pseudobactin was beneficial in reducing bacterial wilt by inducing ISR response in eucalyptus (Ran et al., 2005).

Additionally, purified pseudobactin of WCS374 was found in another study to induce ISR in radish against the pathogen that causes Fusarium wilt, whereas pseudobacterium isolated from other strains was ineffective (Leeman et al., 1996). When grown in the field under conditions of low iron availability, P. putida BTP1 is observed to induce ISR in beans to fend off Botrytis cineria. Nalkylated benzylamine derivative was the substance that caused the ISR response in beans to be triggered in response to Botrytis cineria; however, its mode of action appears to involve lipoxygenase pathway stimulation (Ongena et al., 2002; Ongena et al., 2004; Ongena et al., 2005). Pyochelin and pyocyanin's combined action caused P. aeruginosa 7NSK2 to cause ISR in tomato against Botrytis cineria.

Pyochelin and pyocyanin together significantly suppressed B. cineria, indicating that both compounds are necessary for inducing ISR in mutants of 7NSK2 lacking either one of them (Audenaert et al., 2002). Salicylic acid synthesis genes were extracted from the P. aeruginosa strain PAO1 and expressed in the P. fluorescens strain P3, which does not produce salicylic acid but was able to increase the ISR in tobacco against TNV (tobacco necrosis virus), indicating that salicylic acid can also induce the ISR response in tobacco against TNV (Maurhofer et al., 1998). P. aeruginosa strain PM12 was discovered to produce the potential elicitor 3-hydroxy-5-methoxy benzene methanol (HMB), which aided in the induction of ISR in tomato plants against the pathogen responsible for Fusarium wilt.

So, potential bacterial determinants in the induction of ISR response in plants include siderophores, lipopolysaccharides, salicylic acid, pyochelin, pyocyanin, and other cell envelope components like flagella (Audenaert et al., 2002).

In the last few years, significant progress has been made in our understanding of how Pseudomonads fluorescens colonizes roots and the various ways in which it can suppress soil-borne phytopathogens. In the 21st century, remarkable progress has been made in the biotechnology of P. fluorescens as a biocontrol agent for the defense of crops. Chemical pesticides do not need assistance for proper establishment in the soil rhizosphere, while biocontrol agents do even after application.

Eco-friendly practices must be used in the management of agricultural practices in the current environment. The health and productivity of the soil have been threatened by anthropogenic activities. Chemicals like pesticides, insecticides, and fertilizers should not be used indiscriminately because they harm the environment and ecological balance. Our reliance on the use of chemicals in agriculture has grown too much.

In order to maintain the health of the soil and the ecological balance, we must change to sustainable farming practices. We must take into account the use of biocontrol agents for managing disease and promoting plant growth in order to practice sustainable agriculture. A PGPR that can aid in both plant growth and the control of various plant diseases is pseudomonas fluorescens. Since many years ago, it has been used in agriculture as a biofertilizer and a biocontrol agent. It has enormous potential for supplying nutrients to plants, controlling various phytopathogens, bioremediation, etc. 

As a result, it has enormous potential as a substitute for agrochemicals because it is less expensive, more effective, environmentally friendly, and a good PGPR for increasing crop yield. In addition to this, it lowers input costs and pollution. Future prospects could include simple methods for reproducing organisms, extensions to shelf life, the discovery of potential biocontrol strains, and increased knowledge of the genetic, proteomic, and transcriptional up- and down-regulation that are crucial to plants' defense against a variety of biotic and abiotic stresses.

Aishwarya, Manjula, Ajay Kumar Gautam and Anupam Kumar  (2024). Biological Control Potential of Pseudomonas fluorescence in Managing Plant Diseases: A Comprehensive Review. Biological Forum – An International Journal, 16(3): 242-247.