TUBERCULOSIS: How Tuberculosis Bacteria Manage to Survive Inside Body’s Macrophage Cells

ScienceDaily (Mar. 9, 2011)
Tuberculosis kills two million people each year. Researchers at Linköping University in Sweden are now presenting new findings that show how the bacterium that causes the disease manages to survive inside the body's macrophage cells in order eventually to blow them up and spread their infection.
The bacterium Mycobacterium tuberculosis is a successful organism that lives in an estimated one third of the world's population. But only about five percent of those infected develop the disease.
"We also know that many people do not become infected despite exposure to the infection. This is a question we are looking for an answer to," says Amanda Welin, who is now presenting her doctoral dissertation in medical microbiology.
The research group has studied phenomena in both the bacterium and the macrophage, whose task is to knock out infectious substances that get into the body.
One weapon is enzymes, which make the ingested bacteria feel sickly. Enzymes work best in acidic environments, with a pH level under 6. For their part, the bacteria can strike back by releasing substances that prevent the pH level from going down. Amanda Welin has shown that this warfare is directly reflected in the growth or reduction of bacteria.
These bacteria also have a capacity to kill macrophages and spread to new cells. Welin shows that this is done by having a tiny protein cause cell death, necrosis, which in turn leads to inflammation of the tissue.
To carry out these studies, Amanda Welin and her colleagues developed a new method for determining the number of bacteria inside a cell. They use a gene from sea-fire organisms, which cause strange lights in seawater at night. When this gene is added to the genes of the bacterium, the bacterium begins to produce the same luminescent substance, luciferase, as the sea-fire organism does. Thanks to this, it's possible to monitor developments inside the macrophage -- the intensity of the light radiating outward corresponds to the number of bacteria inside. If their number grows, this indicates that they have begun to multiply inside the human cell.
The method can be used to search for plausible drug candidates. In that field, the Linköping scientists are collaborating with a group of colleagues in Sudan, who are testing, among other things, various medicinal plants with substances that could possibly be used as active ingredients to combat tuberculosis.
http://www.sciencedaily.com/releases/2011/03/110309073940.htm

TUBERCULOSIS: New dual recognition mechanism discovered in tuberculosis

by bjs
09. Sep, 2010
One third of the world’s population is infected with Mycobacterium tuberculosis (MTB), which leads to tuberculosis (TB), a leading cause of death world-wide. A new discovery, led by a team of researchers from Case Western Reserve University School of Medicine, offers hope for new approaches to the prevention and treatment of TB. The team’s discovery of a novel mechanism that may contribute to immune recognition of MTB is published in the September issue of Nature Structural and Molecular Biology.
Most individuals with TB recover from the initial infection and become asymptomatic, but the bacterium persists for years, surviving largely inside macrophages, a type of cell that resides in the immune system. This presents a public health problem in that TB can reactivate and cause serious disease or death. Researchers and physicians know the body’s immune system is capable of containing the infection but not curing it completely. It begs the question: “How does the organism survive in the human immune system for so many years?”
For the past 15 years, Drs. Clifford Harding and W. Henry Boom of Case Western Reserve have been seeking the answer to this question. Their work indicated that MTB can inhibit the ability of macrophages to stimulate infection-fighting immune responses, and they identified that a protein on macrophages called Toll-like receptor 2 (TLR2) is involved in this immune evasion mechanism. TLR2 seems to be a two-edged sword in the complex immune response to MTB, as it helps some immunity mechanisms and inhibits others. Understanding the balance of these effects and the role of TLR2 may provide insights to design therapies for TB.
“Understanding how MTB interacts with the immune system and how it can both activate and inhibit the immune response is critically important for the design of the next generation of TB vaccines. The persistence of infection is dependent on MTB’s ability to manipulate our immune system to its advantage. The paradox here is that the MTB molecule, LprG, stimulates TLR2, one of the major receptors we have to identify disease-causing microorganisms. In this case, too much stimulation through TLR2 actually favors MTB by causing parts of the immune response to shut down,” explains W. Henry Boom, MD, professor of medicine and director of the Tuberculosis Research Unit at Case Western Reserve School of Medicine.
The new studies show that the potency of LprG to induce these responses is explained by its combination of two mechanisms to activate TLR2: first, by directly stimulating TLR2 and, second, by serving as a carrier to deliver other molecules that stimulate TLR2. This dual mechanism may drive stronger regulation of immune responses by MTB, and future vaccine development may be enhanced by designing approaches to use such mechanisms. Furthermore, the work indicates that LprG contributes to the assembly of the bacterial cell wall, suggesting that it may be possible to develop molecules to interfere with LprG function and potentially serve as new antibiotics to fight TB. The development of new antibiotics is an increasingly important goal, since resistance to existing antibiotics is becoming widespread.
A multi-institutional partnership contributed to the overall success of this research initiative. Two important collaborative groups were led by James C. Sacchettini, PhD, Texas A&M University and D. Branch Moody, MD, Harvard Medical School. In addition, the project was spearheaded by Michael G. Drage and Nicole D. Pecora, two Case Western Reserve students in the MSTP Program, granting dual MD and PhD degrees, in collaboration with Jennifer Tsai, a graduate student in Dr. Sacchettini’s group.
“Our research team is composed of several collaborative groups that each contributed key components to this project. The synergistic way in which the team interacted was a perfect example of scientists working together to advance the study of a disease that detrimentally impacts the lives of so many across the globe. We look forward to continuing to advance this research together,” says Clifford V. Harding, MD, PhD, professor and interim chair of pathology at Case Western Reserve School of Medicine.
As they look to the future, the research team will work to gain a better understanding of immune responses in TB and hopefully design approaches to treat the deadly disease, including antibiotics or immunotherapies. Continued work will include study of the mechanism of immune-evasion by MTB with the hope of finding ways to reverse this mechanism so that it no longer causes a persistent infection.

http://scienceblog.com/38378/new-dual-recognition-mechanism-discovered-in-tuberculosis/

TUBERCULOSIS: How mycobacteria avoid destruction inside human cells

Tuberculosis, or TB, is a dreaded contagious disease of the lungs and other organs. The causative agent, Mycobacterium tuberculosis (or M. tuberculosis), infects roughly a third of the world's population and one-in-ten to one-in-twenty of the infected population becomes sick or infectious at some point during their lifetime.
The mycobacteria survive, and even thrive, inside host macrophages – cells that are part of the human immune system and that usually engulf and destroy bacteria in structures called phagosomes. M. tuberculosis is taken into phagosomes but it somehow blocks phagosome maturation, and hence survives. Figuring out how could open up new therapeutic targets for the treatment of TB as well as shedding light on the mechanism of intracellular parasitism.
Researchers at the Pasteur Institutes in Seoul and Paris and Institute of Pharmacology and Structural Biology (IPBS) in Toulouse joined forces to systematically search for mycobacterial genes that block phagosome maturation. To do this, they generated 11,000 different mutants of the M. tuberculosis Beijing strain, which has been associated with large outbreaks of TB, increased virulence, and multidrug resistance.
Using a high-throughput visual assay, the researchers screened for mutant mycobacteria that had lost the ability to arrest phagosomal maturation. Lead author Dr. Priscille Brodin, heading the Inserm Avenir Unit at Institut Pasteur Korea describes the screen as "enabling stringent selection of mutants that have the most pronounced subcellular localization within intracellular acidic compartments through the use of automated confocal quantitative imaging. Our approach", she adds, "may be useful to identify virulence genes in other intracellular pathogens".
The team identified ten distinct mutants, only one of which had previously been shown to play a part in phagosome maturation arrest. Finding that two independent mutants mapped to the same region, they studied this locus in more detail. The work revealed that the biosynthesis of particular glycolipids containing acyltrehalose was perturbed, suggesting to the researchers that these glycolipids play a critical role in the early intracellular protection of mycobacteria.
"Our study unravels the role of novel lipid molecules in mycobacterial intracellular parasitism" says Dr. Olivier Neyrolles leading a CNRS Unit at IPBS in Toulouse France. "This establishes potential new drug targets", especially important given the emergence of multidrug-resistant and extensively drug-resistant TB. "In addition", Dr Brodin points out, "the assay that have we developed can be readily adapted for the screening of novel antimicrobials".
http://www.eurekalert.org/pub_releases/2010-09/ipk-hma090610.php

Jul. 29, 2010
Often causing no symptoms in carriers of the disease, worldwide tuberculosis (TB) infects eight to ten million people every year, kills two million, and it is highly contagious as it is spread through coughing and sneezing. “It’s a global health disaster waiting to happen, even here in Canada, but this new paradigm in TB research may offer an immediate opportunity to improve vaccination and treatment initiatives,” explains Dr. Maziar Divangahi of McGill University and of the Research Institute of the McGill University Health Centre.
The ability of TB bacteria to persist in individuals with apparently normal immune systems implies that they have developed strategies to avoid, evade, and even subvert immunity. The bacteria mainly enter the body through inhalation into the respiratory tract. Alveolar macrophages, a type of white blood cell residing in our lungs, initially recognize the bacteria and engulf them. This process is one of our immune system’s defense mechanisms. However, TB has evolved into a parasite that can survive and replicate inside the macrophages until they burst out, spreading the infection.
The way infected macrophages die is a determining factor in the development of immunity to TB. Macrophages can induce apoptosis, a type of cell death which keeps their membrane intact, trapping and reducing bacterial viability. However, TB bacteria induce another type of cell death called necrosis. Necrosis causes cell death by disrupting the cell membranes, which enables the bacteria to escape the cell. It may help to visualize a box with broken walls.
The key to the fate of the macrophages is the balance between two kinds of eicosanoids. Eicosanoids are molecules that contribute to the control of our immune system. The genetic code of TB bacteria enables it to tip this balance in favor of necrosis, and human genetic analysis revealed that modification in eicosanoids production is associated with susceptibility or resistance to TB. Fortunately, drugs that target the production of eicosanoids are already in use for treating other inflammatory diseases, such as rheumatoid arthritis.
“The next steps will be to see exactly how these drugs can be used to treat TB,” said Divangahi. The research received funding from the Fonds de la Recherche en Santé du Québec and was published in Nature Immunology. Divangahi is affiliated with the Departments of Medicine and Microbiology/Immunology of McGill’s Faculty of Medicine, with the Research Institute of the McGill University Health Centre, and with the Meakins-Christie Laboratory.
http://www.mcgill.ca/newsroom/news/item/?item_id=165121