A new mechanism has been discovered by which cells infected with the foodborne pathogen Listeria monocytogenes are able to escape the immune response, and which at the same time confers a higher probability that infected cells will adhere to and infect cerebral vessels and cross the blood-brain barrier.

The finding builds on an earlier observation that L. monocytogenes primarily infects inflammatory monocytes circulating in the blood stream, which then provide a protective niche for the bacteria.

Researchers at Institut Pasteur used hypervirulent strains of L. monocytogenes isolated from patients who suffered brain infections or neurolistiosis, to inoculate a humanized mouse model of the infection, which in humans is fatal in 30% of cases.

That enabled them to demonstrate in intricate detail how a cell surface protein, internalin B (InlB), which has long been known to be a key factor in the virulence of L. monocytogenes, not only enables the bacteria to evade the immune system, but also is involved in prolonging the life of infected monocytes by preventing cell death mediated by cytotoxic T cells.

"We discovered a specific, unexpected mechanism by which a pathogen increases the life span of the cells it infects by specifically blocking an immune system function that is crucial for controlling infection," said Marc Lecuit, head of the biology of infection unit at the Institut Pasteur and co-author of a paper in the March 16, 2022, issue of Nature describing the research.

In contrast to the reference strain of L. monocytogenes, hypervirulent stains systematically induce neuroinvasion. The researchers showed that only intracellular bacteria can cross the blood-brain barrier by treating infected mice with gentamicin, an antibiotic that kills extracellular, but not intracellular bacteria.

In mice infected with hypervirulent strain, the bacterial load in the brain was the same, with or without administration of gentamycin. As a further demonstration that only intracellular L. monocytogenes can cross the blood-brain barrier, in a mouse model in which monocytes are retained in the bone marrow, fewer bacteria are found in the brain after infection with hypervirulent L. monocytogenes, compared to wild type.

In addition, when infected monocytes from donor mice were transferred into gentamycin-treated, noninfected mice, these mice suffered neuroinvasion as early as day 2 after transfer.

Infected monocytes were observed adhering to the endothelium of blood vessels in brain sections of infected mice postmortem.

Taken together, the researchers say the data from their model indicate infected monocytes are necessary and sufficient to induce neuroinvasion.

Having identified infected monocytes as being critically involved in the onset of neurolisteriosis, the researchers looked for factors in L. monocytogenes that mediate its neuroinvasiveness.

They turned to previous research showing the bacterial cell surface proteins InlA and InlB play a part in L. monocytogenes' ability to cross intestinal and placental barriers, discounting involvement of InlA in crossing the blood-brain barrier, but showing InlB has a critical role.

That is notable because InlB is part of the core genome of L. monocytogenes and is therefore present in all strains of the bacterium, including those that are not neuroinvasive.

However, neuroinvasive strains of L. monocytogenes strongly upregulate the InlB operon, suggesting high expression of InlB is required for the pathogen to cross the blood-brain barrier.

This was confirmed by increasing expression of InlB in non-neuroinvasive strains, which prompted them to become as neuroinvasive as hypervirulent strains.

However, a series of subsequent experiments showed InlB has no direct effect on the ability of L. monocytogenes to adhere to blood brain vessels, but rather that it blocks the Fas-mediated CD95 tumor necrosis factor receptor superfamily member 6 killing of infected monocytes by CD8+ T cells.

Specifically, InlB acts through a c-Met and PI3K-alpha dependent pathway to upregulate FLIP, which in turn inhibits the cleavage of caspase-8, thereby blocking Fas-mediated cell death.

As a result, the lifespan of the monocytes is extended, providing them with the time to adhere to brain vessels and transfer intracellular L. monocytogenes to the brain parenchyma.

"It was known that immunosuppression was a risk factor for neurolisteriosis and therefore when testing the effect of InlB in neuroinvasion in immunosuppressed mice, we expected to see a larger effect, whereas it was actually abolished," Lecuit said. "That gave us a hint that neuroinvasion had something to do with the immune system," he told BioWorld.

The unanticipated mechanism whereby L. monocytogenes renders infected cells resistant to CD8+ T cell-mediated killing, creating a protective niche, also is involved in its persistence in intestinal tissue, its release into feces, and from there to the environment.

Other neuroinvasive intracellular pathogens, such as Mycobaterium tuberculosis and Toxoplasma gondii also stimulate PI3K and survive in myeloid cells, suggesting they may also protect infected cells from cell death.

The researchers also point to similarities with how tumor cells evade immune responses. "A detailed understanding of how microorganisms have selected mechanisms that interfere with the immune system may help in the rational design of anti-infective and antitumor therapies," they say.

Similarly, given the immunomodulatory mechanism of InlB is specific to and restricted to infected cells, the findings may also help in the development of immunosuppressive therapies that protect specific cells from cell death, as opposed to current drugs with a nonspecific action that can lead on to infections and other complications.