Agrobacterium: A Biological Syringe

Plants, just like humans, fall victim to bacterial infections. Dr. Ann Matthysse, a researcher in the Department of Biology at the University of North Carolina, has studied interactions between plants and pathogens since 1970, when she thought that Agrobacterium tumefaciens might lead to advances with cases of human cancer.

Matthysse initially thought that A. tumefaciens, a Gram-negative, rod-shaped bacterium found in upper layers of the soil, could be a model for cancer because it causes tumors in plants (Figure 1)

She found instead that the cancer-causing mechanism utilized by A. tumefaciens has virtually nothing to do with human cancer. However, continuing studies with the bacterium is still very beneficial due to its unique initial surface reactions with wounded plants as it binds them to begin infection.

Matthysse describes A. tumefaciens as “a biological syringe” because its virulence comes from a transfer of DNA upon infection of a plant wound, a process unique to this specific plant bacteria2. The transferred DNA integrates into the host cell chromosome and transforms the plant’s cells into tumor cells. These transformed cells then make metabolites that only A. tumefaciens is able to utilize as an energy source. The virus essentially taps into the host plant’s energy source in the same way a cell phone charger would pull energy from your car battery. This results in smaller fruit than normal being produced by the plant host, but it is usually not fatal to the plant unless the tumor blocks its main vascular tissue. Additionally, these initial surface interactions involved in DNA transfer will function the same even if non-natural, specifically-selected genes are inserted into the bacterium for transfer into a plant.

Now, Matthysse is interested in manipulating this mechanism to more efficiently develop genetically engineered crops. Some crops have been difficult to engineer, but these problems can be alleviated by identifying restrictions on the host range for agrobacterium. “Because if we knew what [these factors] were,” she proclaims, “it might be possible to counteract them.”2 If A. tumefaciens can be manipulated to bind to these plants like it does to other plant hosts, one could engineer some of these crops. For example, one could transfer genes that resist pathogenic fungi, and there is opportunity to improve nutrient levels in certain foods. “For example, rice that contains a lot of vitamin A, which would be good for people in India that don’t have a lot of vitamin A in their diet, has been made by putting the genes for vitamin A biosynthesis into rice.”

In 2006, the Centers for Disease Control and Prevention (CDC) reported that an Escherichia coli outbreak occurred from the bacteria infecting salad vegetables and causing disease in those eating the vegetables raw (Figure 2). In the numerous studies that were based on the results reported, Salmonella was also identified as a cause of disease through a similar route. E. coli was traditionally used in the lab as a control for the A. tumefaciens experiments because it did not bind to the plant host, so E. coli studies quickly began. Matthysse found that salad leaves and sprouts encountered bacteria in multiple situations:  contamination of irrigation water or equipment, improperly prepared manure fertilizer, and in post-harvest situations. Once the bacteria are bound, they cannot be removed simply by washing; infected sprouts and fruits that are not cooked prior to eating pose the greatest risk for transferring the disease to humans.

It turns out that the signals produced by the plant cell stimulate bacterial binding for A. tumefaciens, E. coli, and Salmonella. There are multiple ways that A. tumefaciens,  E. coli, and Salmonella appear to be binding to alfalfa sprouts and other salad vegetables. However, studies show that there might be a single sensory pathway that could be blocked so the bacteria are unaware of the presence of a plant in their vicinity. Thus, although the bacteria are ultimately unable to be removed once bound to the plant tissue, the sensory pathway approach could bypass this problem and prevent the bacteria from ever binding in the first place (Figure 3). Currently, Matthysse is looking at multiple methods of blocking/changing signals from plants and/or altering signal receptors on pathogenic bacteria. It might be possible to manipulate the environment so that sensory genes are turned on too soon or too late, and thus attachment and infection are not as effective.

The future of these studies remains promising, and Matthysse acknowledges the difficulty and importance of designing the most effective experiments:  identifying which factors matter the most, pinpointing the best incubation time, appropriating growth temperatures, and other conditions are a serious time investment. Practicality also has to be considered in this situation; increasing costs to customers is not a helpful option when considering long-term reduction in E. coli and Salmonella infections in raw salad vegetables. Eating leafy greens has always seemed very healthy and beneficial to the human diet, but these foods are just as prone to contamination as others. It is probably not common to consider the conditions of our salad, given that it is purchased from a seemingly safe grocery store. “We have all gotten so far away from where our food actually comes from,” Matthysse says.

Ultimately, Matthysse’s studies could lead to revolutionary improvements in genetically modified foods, and the possibilities for utilizing the A. tumefaciens gene transfer mechanism are endless. Her experiments to prevent the binding of pathogens like E. coli and Salmonella to salad vegetables could significantly reduce the number of outbreaks of these pathogens. Understanding these complicated interactions will continue to provide a strong foundation for future studies of plant pathogens.

Escherichia coli, a Gram-negative bacterium widely studied in the lab as a model organism and also found in cases of food poisoning
Figure 2: Escherichia coli, a Gram-negative bacterium widely studied in the lab as a model organism and also found in cases of food poisoning
Attachment of Agrobacterium tumefaciens to tomato root hairs. The decrease in binding visible from (A) to (B) is a comparison of wildtype A. tumefaciens (A) to A. tumefaciens without the attachment mediating gene, UPP(B).
Figure 3: Attachment of Agrobacterium tumefaciens to tomato root hairs. The decrease in binding visible from (A) to (B) is a comparison of wildtype A. tumefaciens (A) to A. tumefaciens without the attachment mediating gene, UPP(B).
Dr. Ann Matthysse, Professor of Biology, University of North Carolina at Chapel Hill.
Dr. Ann Matthysse, Professor of Biology, University of North Carolina at Chapel Hill.

References:

  1. Ann G. Matthysse; Frontiers in Plant Science 2014, 5, 1-8.
  2. Interview with Ann G. Matthysse, Ph.D. 9/18/2014.
  3. Update on Multi-State Outbreak of E. coli O157:H7 Infections From Fresh Spinach, October 6, 2006. http://www.cdc.gov/ecoli/2006/september/updates/100606.htm. (accessed September 22, 2014).

Image Sources:

  1. A. tumefaciens and E. coli: Public Domain
  2. Figure 3: Ann Matthysse

This article was previously published in Carolina Scientific Magazine, Fall 2014. 

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