Day: 06/16/2015

Programming DNA to reverse antibiotic resistance in bacteria

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Growing bacteria in petri dishes.

At its annual assembly in Geneva last week, the World Health Organization approved a radical and far-reaching plan to slow the rapid, extensive spread of antibiotic resistance around the world. The plan hopes to curb the rise caused by an unchecked use of antibiotics and lack of new antibiotics on the market.

New Tel Aviv University research published in PNAS introduces a promising new tool: a two-pronged system to combat this dangerous situation. It nukes antibiotic resistance in selected bacteria, and renders other bacteria more sensitive to antibiotics. The research, led by Prof. Udi Qimron of the Department of Clinical Microbiology and Immunology at TAU’s Sackler Faculty of Medicine, is based on bacterial viruses called phages, which transfer “edited” DNA into resistant bacteria to kill off resistant strains and make others more sensitive to antibiotics.

According to the researchers, the system, if ultimately applied to pathogens on hospital surfaces or medical personnel’s hands, could turn the tide on untreatable, often lethal bacterial infections. “Since there are only a few pathogens in hospitals that cause most of the antibiotic-resistance infections, we wish to specifically design appropriate sensitization treatments for each one of them,” Prof. Qimron says. “We will have to choose suitable combinations of DNA-delivering phages that would deliver the DNA into pathogens, and the suitable combination of ‘killing’ phages that could select the re-sensitized pathogens.”

Reprogramming the system

“Antibiotic-resistant pathogens constitute an increasing threat because antibiotics are designed to select resistant pathogens over sensitive ones,” Prof. Qimron says. “The injected DNA does two things: It eliminates the genes that cause resistance to antibiotics, and it confers protection against lethal phages.

“We managed to devise a way to restore antibiotic sensitivity to drug-resistant bacteria, and also prevent the transfer of genes that create that resistance among bacteria,” he continues.

Earlier research by Prof. Qimron revealed that bacteria could be sensitized to certain antibiotics — and that specific chemical agents could “choose” those bacteria more susceptible to antibiotics. His strategy harnesses the CRISPR-Cas system — a bacterial DNA-reprogramming system Prof. Qimron pioneered — as a tool to expand on established principles.

According to the researchers, “selective pressure” exerted by antibiotics renders most bacteria resistant to them — hence the epidemic of lethal resistant infections in hospitals. No counter-selection pressure for sensitization of antibiotics is currently available. Prof. Qimron’s strategy actually combats this pressure — selecting for the population of pathogens exhibiting antibiotic sensitivity.

“We believe that this strategy, in addition to disinfection, could significantly render infections once again treatable by antibiotics,” said Prof. Qimron.

Prof. Qimron and his team are now poised to apply the CRISPR/phage system on pseudomonas aeruginosa — one of the world’s most prevalent antibiotic-resistant pathogens involved in hospital-acquired infections — and to test whether bacterial sensitization works in a more complex microbial environment: the mouse cage.

Missing link found between brain, immune system; major disease implications

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Maps of the lymphatic system: old (left) and updated to reflect UVA’s discovery.

In a stunning discovery that overturns decades of textbook teaching, researchers at the University of Virginia School of Medicine have determined that the brain is directly connected to the immune system by vessels previously thought not to exist. That such vessels could have escaped detection when the lymphatic system has been so thoroughly mapped throughout the body is surprising on its own, but the true significance of the discovery lies in the effects it could have on the study and treatment of neurological diseases ranging from autism to Alzheimer’s disease to multiple sclerosis.

“Instead of asking, ‘How do we study the immune response of the brain?’ ‘Why do multiple sclerosis patients have the immune attacks?’ now we can approach this mechanistically. Because the brain is like every other tissue connected to the peripheral immune system through meningeal lymphatic vessels,” said Jonathan Kipnis, PhD, professor in the UVA Department of Neuroscience and director of UVA’s Center for Brain Immunology and Glia (BIG). “It changes entirely the way we perceive the neuro-immune interaction. We always perceived it before as something esoteric that can’t be studied. But now we can ask mechanistic questions.”

“We believe that for every neurological disease that has an immune component to it, these vessels may play a major role,” Kipnis said. “Hard to imagine that these vessels would not be involved in a [neurological] disease with an immune component.”

New Discovery in Human Body

Kevin Lee, PhD, chairman of the UVA Department of Neuroscience, described his reaction to the discovery by Kipnis’ lab: “The first time these guys showed me the basic result, I just said one sentence: ‘They’ll have to change the textbooks.’ There has never been a lymphatic system for the central nervous system, and it was very clear from that first singular observation — and they’ve done many studies since then to bolster the finding — that it will fundamentally change the way people look at the central nervous system’s relationship with the immune system.”

Even Kipnis was skeptical initially. “I really did not believe there are structures in the body that we are not aware of. I thought the body was mapped,” he said. “I thought that these discoveries ended somewhere around the middle of the last century. But apparently they have not.”

‘Very Well Hidden’

The discovery was made possible by the work of Antoine Louveau, PhD, a postdoctoral fellow in Kipnis’ lab. The vessels were detected after Louveau developed a method to mount a mouse’s meninges — the membranes covering the brain — on a single slide so that they could be examined as a whole. “It was fairly easy, actually,” he said. “There was one trick: We fixed the meninges within the skullcap, so that the tissue is secured in its physiological condition, and then we dissected it. If we had done it the other way around, it wouldn’t have worked.”

After noticing vessel-like patterns in the distribution of immune cells on his slides, he tested for lymphatic vessels and there they were. The impossible existed. The soft-spoken Louveau recalled the moment: “I called Jony [Kipnis] to the microscope and I said, ‘I think we have something.'”

As to how the brain’s lymphatic vessels managed to escape notice all this time, Kipnis described them as “very well hidden” and noted that they follow a major blood vessel down into the sinuses, an area difficult to image. “It’s so close to the blood vessel, you just miss it,” he said. “If you don’t know what you’re after, you just miss it.”

“Live imaging of these vessels was crucial to demonstrate their function, and it would not be possible without collaboration with Tajie Harris,” Kipnis noted. Harris, a PhD, is an assistant professor of neuroscience and a member of the BIG center. Kipnis also saluted the “phenomenal” surgical skills of Igor Smirnov, a research associate in the Kipnis lab whose work was critical to the imaging success of the study.

Alzheimer’s, Autism, MS and Beyond

The unexpected presence of the lymphatic vessels raises a tremendous number of questions that now need answers, both about the workings of the brain and the diseases that plague it. For example, take Alzheimer’s disease. “In Alzheimer’s, there are accumulations of big protein chunks in the brain,” Kipnis said. “We think they may be accumulating in the brain because they’re not being efficiently removed by these vessels.” He noted that the vessels look different with age, so the role they play in aging is another avenue to explore. And there’s an enormous array of other neurological diseases, from autism to multiple sclerosis, that must be reconsidered in light of the presence of something science insisted did not exist.