Antimicrobial Resistance with Special Emphasis on Pathogens in Agriculture

*Nitya Meenakshi Raman, Murugesh Easwaran, Rashmi Kaul, Jyotsna Bharti, Khaled Fathy Abdel Motelb and Tanushri Kaul*

### **Abstract**

Antibiotics have been used globally to manage the bacterial plant diseases irrespective of the expense involved. Although plant pathogenesis by bacteria is far lower than fungal counterparts, disrupted monitoring and surveillance for drug resistance with respect to human health raise serious concerns. The resistance derived by the plant as the host by the antibiotics used for many generations has now posed as a problem in phyto-systems. Although we currently lack the molecular understanding of the pathogens rendering antibiotic resistance to plants, robust resistance management strategies are critical to ensure management of critically important diseases that specifically target crops of high value and/or global agrarian importance. This chapter discusses evolution of plant-pathogenic bacteria, application of antibiotics and its repercussions on the microbiome of plant agricultural systems, and sustainable crop disease management by genetic engineering.

**Keywords:** agriculture, bacteria, fruit, genetic engineering, host, molecular biology

#### **1. Introduction**

Antibiotic resistance most commonly evolves in bacteria either through mutation of a target site protein, through the acquisition of an antibiotic-resistant gene that confers resistance through efflux or inactivation of the antibiotic, or through synthesis of a new target protein that is insensitive to the antibiotic [1]. An extensive body of knowledge has been gained from studies of antibiotic resistance in human pathogens and in animal agriculture. The ability of bacterial pathogens to acquire antibiotic-resistant genes and to assemble them into blocks of transferable DNA encoding multiple antibiotic-resistant genes has resulted in significant issues that affect successful treatment interventions targeting some specific human infections. The current global antibiotic resistance crisis in bacterial populations has been fuelled by basic processes in microbial ecology and population dynamics, engendering a rapid evolutionary response to the global deployment of antibiotics by humans in the millions of kilograms per year. What was not anticipated when antibiotics were discovered and introduced into clinical medicine is that antibioticresistant genes pre-existed in bacterial populations [2–4]. Furthermore, the extent to which antibiotic-resistant genes could be transferred between bacteria, and

even between phylogenetically distinct bacteria, was not understood 70 years ago but is becoming more apparent through a number of elegant studies identifying the microbial antibiotic resistome. The collection of all known antibiotic-resistant genes in the full-microbial pan-genome is defined as the antibiotic resistome [5].

## **2. Use of antibiotics in agriculture**

Effective management of bacterial plant diseases is difficult and is exacerbated by factors such as the large size of bacterial pathogen populations on susceptible plant hosts and the few available bactericides. In the absence of durable and robust host disease resistance, antibiotics have represented the best option for bacterial disease control in many pathosystems because these materials provide the most efficacious means of reducing bacterial population size and limiting disease outbreaks. Although many new types of antibiotics were rapidly tested and then deployed in animal agriculture starting in the 1950s, antibiotic use for plant disease control was tempered by several factors, including lack of efficacy at lower doses, phytotoxicity problems at higher doses, and expense compared to other existing methods of disease control. Thus, although penicillin, streptomycin, aureomycin, chloramphenicol, and oxytetracycline were tested for plant disease control in the late 1940s [6, 7], only streptomycin and oxytetracycline were ultimately deployed in plant agriculture and only in specific disease pathosystems. Streptomycin is the main antibiotic currently in use for plant disease control around the world, targeting pathogens such as *Erwinia amylovora*, which causes fire blight of apple and pear; *Pseudomonas syringae*, which causes flower and fruit infection of apple and pear trees; and *Xanthomonas campestris*, which causes bacterial spot of tomato and pepper [8]. Oxytetracycline has been used as the primary antibiotic in specific disease control situations, including the control of *Xanthomonas arboricola* pv. *pruni*, the causal agent of bacterial spot of peach and nectarine [8]. In addition, oxytetracycline has been used as a secondary antibiotic for fire blight management in the United States, most prominently in situations in which streptomycin resistance has become a problem [9, 10].

The problem of antibiotic resistance is not limited to the Indian subcontinent only, but is a global problem. To date, no known method is available to reverse antibiotic resistance in bacteria. The discovery and development of the antibiotic penicillin during the 1900s gave a certain hope to medical science, but this antibiotic soon became ineffective against most of the susceptible bacteria. The antibiotic resistance in bacteria is generally a natural phenomenon for adaptation to antimicrobial agents. Once bacteria become resistant to some antibiotic, they pass on this characteristic to their progeny through horizontal or vertical transfer. The indiscriminate and irrational use of antibiotics these days has led to the evolution of new resistant strains of bacteria that are somewhat more lethal than the parent strain. More recently, in 2016, a Section 18 emergency exemption was granted by the US Environmental Protection Agency for the use of streptomycin and oxytetracycline on citrus trees in Florida for management of citrus Huanglongbing (HLB) disease [11–13]. Regarding other antibiotics, gentamicin has been used in Mexico for fire blight control and in Chile, Mexico, and Central American countries for vegetable disease control, while oxolinic acid (OA) has been used only in Israel for fire blight management [14, 15]. Lastly, kasugamycin is used in Japan and other Asian countries to control the fungal disease rice blast and bacterial seedling diseases of rice [16] and has recently been registered for use in the United States and Canada for managing fire blight [17]. Concerns regarding the use of antibiotics in plant disease control and potential impacts on human health have led to the banning of antibiotic

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*Antimicrobial Resistance with Special Emphasis on Pathogens in Agriculture*

use by the European Union. However, streptomycin is still utilized for fire blight management in Austria, Germany, and Switzerland under strict control parameters.

The lack of effective bactericide alternatives in several plant disease systems has resulted in a decade-long dependence or overdependence on streptomycin. As streptomycin has been used the longest, over the largest geographic area, and for treatment of the largest variety of crops, streptomycin resistance is relatively widespread among plant-pathogenic bacteria. Although the first streptomycin-resistant (SmR) plant-pathogenic bacteria detected were strains of *E. amylovora* harboring a chromosomal resistance mutation, the majority of SmR plant pathogens encode the transmissible SmR transposon Tn5393 [8]. Tn5393 is a Tn3-type transposon originally isolated from *E. amylovora* that harbors *strAB*, a tandem resistance gene pair that confers streptomycin resistance through covalent modification of the streptomycin molecule [18]. The Tn5393 transposon is composed of genes required for the transposition process (*tnpA* and *tnpR*), a central site that contains outwardly directed promoters for expression of both *tnpA* and *tnpR* as well as the *strAB SmR* genes. Expression of the *strAB* genes from Tn5393 in *E. amylovora* is driven by a promoter present in the 3 prime end of the insertion sequence IS1133 that is inserted directly upstream of the *strA* gene [19]. Two closely related variants of Tn5393 have also been found in plant pathogens: Tn5393a, an element that does not contain IS1133, has been detected in *P. syringae* and in a group of *E. amylovora* strains from California exhibiting a moderate level of resistance, and Tn5393b, an element that does not contain IS1133 but instead contains an insertion of IS6100 within the *tnpR*

There are two other reports of additional genetic mechanisms of streptomycin resistance in plant pathogens; these include the occurrence of the small, nonconjugative but mobilizable broad-host-range plasmid RSF1010 in some strains of *E. amylovora* isolated in California [21]. This observation carries further significance because RSF1010 has been distributed globally among a number of bacterial genera and also occurs in some human-pathogenic bacteria [22]. A recent report detailing an analysis of streptomycin-resistant *X. oryzae* subsp. *oryzae* from China indicated that four strains harbored the *aadA1* gene associated with class 1 integron sequences [23]. This observation is significant because of the importance of integrons in both the transfer of antibiotic resistance in human and animal pathogens and the accumulation of antibiotic resistance genes within one multiresistance element. To date, streptomycin resistance mediated by Tn5393 or the closely related variants has been reported in *E. amylovora*, *P. syringae*, and *X. campestris* isolated from North and South America and Asia [19, 20, 24–30]. The location of essentially the same genetic element in different genera of plant pathogens isolated from distinct crop hosts and from different continents is confirmatory evidence of the role of horizontal gene transfer (HGT) in the dissemination of antibiotic resistance in these pathosystems. The source of Tn5393 to the plant pathogens was likely not from the antibiotic preparations themselves as a study of 18 available agricultural streptomycin formulations revealed no contamination with the *strA SmR* gene [31]. Instead, the acquisition of Tn5393 by bacterial plant pathogens was likely from commensal co-occurring epiphytic bacteria via HGT. For example, Tn5393 was thought to have been acquired by *E. amylovora* on the plasmid pEa34 from *Pantoea agglomerans*, a common orchard epiphyte [18]. The transfer event most likely occurred on the

*DOI: http://dx.doi.org/10.5772/intechopen.88440*

**3. Evolution of plant-pathogenic bacteria**

gene, has been characterized in *X. campestris* [19, 20].

**3.1 Resistance to streptomycin**

use by the European Union. However, streptomycin is still utilized for fire blight management in Austria, Germany, and Switzerland under strict control parameters.
