European researchers develop test to pinpoint TB

A group of European researchers have developed a test that can diagnose active tuberculosis (TB) in people even when traditional tests have failed. the findings are published in the American Journal of Respiratory and Critical Care Medicine.

The research team, the Tuberculosis Network European Trialsgroup (TBNET), a scientific consortium founded in 2006 to research new methods of TB diagnosis, used the ELISPOT (enzyme-linked immunospot assay) test to develop the new method. they found that immune cells that are specific to TB bacilli are concentrated in the airways of patients who have active tuberculosis. these cells can be identified quickly using the ELISPOT test. the test can distinguish between latent TB infection, known as lasting tuberculosis immune responses (LTBI), and active TB by comparing the frequencies of TB-specific lymphocytes in the blood with those in the lung.

About a third of the world’s population is infected with the mycobacterium that causes TB but only about 10% to 20% of those will go on to develop the disease. the others have LTBI which can develop into TB at any time.

Sputum testing has traditionally been used for TB diagnosis: when TB bacilli show up on sputum tests, a positive diagnosis of TB can be established quickly. unfortunately, in about 50% of these tests the TB bacilli cannot be identified so a more accurate method is needed.

Dr Christoph Lange from the Research Center Borstel Leibniz-Center for Medicine and Biosciences in Germany, who led the TBNET research, said: in this study we showed that a differentiation between active pulmonary tuberculosis and LTBI is possible by using the ELISPOT test.

TB spreads quickly because it is disseminated through the air when infected people sneeze, spit or talk. It is the seventh leading cause of death worldwide and its incidence is rising again after a period of decline from the 1960s when the successful use of the Bacille Calmette-Guérin (BCG) vaccine led to a decline in the disease. TB usually attacks the lungs but other parts of the body can be affected too including the spine and the circulatory system.

To test the ELISPOT method, the TBNET researchers recruited 347 patients who were believed to have TB, but were either unable to produce sputum or had had 3 negative sputum tests. the ELISPOT test was performed using bronchoaveolar lavage (BAL, a method in which fluid is squirted into the lung and then withdrawn for analysis).

Of the 347 patients, 71 were diagnosed with active pulmonary TB and the results for patients with active TB were accurate in 91.5% of cases.

‘these findings show us that positive results in the BAL ELISPOT were highly indicative of actual cases of active TB,’ Dr Lange said. ‘A negative BAL ELISPOT result almost excludes active tuberculosis.

‘Future research about the immunodiagnosis of tuberculosis will need to identify indicators of treatment success that will allow safe discontinuation of anti-tuberculosis therapy without an increased risk of the reactivation,’ he pointed out. ‘This will be of great clinical importance to guide the treatment of individuals with LTBI and active tuberculosis, especially in cases of drug-resistant strains of mycobacterium tuberculosis.’

Dr Lange also pointed out that in 2008 a 27-institution consortium was established in Europe for the study of drug resistant tuberculosis (tbpannet.eu/>) and was granted EUR 12 million by the EU. the result is the TB PAN-NET (Pan-European network for the study and clinical management of drug resistant tuberculosis) project, supported by the Health Thematic area of the EUs Seventh Framework Programme (FP7).

by VincentRacanielloFriday, February 18, 2011 at 06:45 AMEST

The first detailed study ofinfection of nonhuman primates with the retrovirus XMRV reveals that the virusestablishes a persistent infection characterized by infection of multipletissues. Viremia (virus in the blood) is low and transient, with proviral DNAdetectable in blood lymphocytes. The results show that the Rhesus macaque canbe used to study XMRV infection, transmission, vaccines, and antiviraldrugs.

The subject of this study, the Rhesus macaque (Macaca mulatta), wasselected because of its evolutionary proximity to humans and a comparableimmune system. The monkeys used did not have antibodies to the capsid proteinp30 of XMRV, indicating that they were not previously infected. Animals wereinoculated intravenously with 3.6 million TCID50 of purified XMRV — a good amount of virus, toensure infection. The virus used, VP62, was produced by transfecting cells with cloned viral DNA isolated fromhuman prostate.

Virus in the plasma fraction of blood was assayed by quantitative RT-PCR. Ofthree animals infected, virus was detected in one animal at day 4 and not afterday 14; and in a second animal from days 14-20. The third animal did notdevelop detectable viremia. Proviral DNA was found in peripheral bloodmononuclear cells (PBMC) of all three monkeys for 3-4 weeks, indicatingsuccessful infection. at one month post-infection proviral DNA was no longerdetected. Plasma virus was again detected in one of the positive animals on day291, 16 days after being immunized with a mixture of XMRV proteins. This meansthat viral DNA had been present in this animal but was not detected. XMRV wasdetected in CD4+ and CD8+ T cells and NK cells, but not in B cells ormonocytes.

Rhesus macaques infected with XMRV did not display obvious clinicalsymptoms. Analysis of peripheral blood revealed increases in the number ofcirculating B and NK cells. Anti-viral antibody titers were detected afterinfection and re-infection of animals but soon decreased.

Other infected animals were sacrificed during the acute phase of infectionto identify pathological changes and sites of virus replication. No pathogenicconsequences were observed except for the formation of germinal centers inspleen and lymphoid organs, changes that are expected after immune stimulation.Virus was detected in a wide variety of tissues, including spleen, lymph nodes,the lining of the gastrointestinal tract, prostate, testis, cervix, vagina, andpancreas, but not in others including brain, heart, kidney, and bladder.different types of cells were infected in different tissues: lymphocytes inlymphoid organs, macrophages in lung, epithelial or interstitial cells in otherorgans. The authors note that “this viral behavior appears specific to thisvirus”.

Here are some other comments and conclusions drawn from this study:

Because the study involved only a small number of monkeys (8), theexperiments should be repeated with additional animals, and in differentlaboratories, to verify the findings. I also wonder if the choice of theintravenous inoculation route had an effect on the pattern of infection andtropism. it is well known that viral pathogenesis can be determined by how thevirus enters the host. for example, the same virus may replicate in differenttissues, or have different virulence, when inoculated in different ways. Thisquestion can be readily addressed by inoculating rhesus macaques via differentroutes.

Studying viral pathogenesis (the series of events that occur during viralinfection of a host) in animals is essential for understanding how virusescause disease in humans. however, the results of such studies must always beinterpreted with caution, because what is true in an animal is not always truefor a human. for example, simple differences in size, metabolism, anddevelopment can have substantial effects on pathogenesis. In interpreting theresults of animal studies, we must keep in mind the adage, ‘Mice lie, monkeys exaggerate‘.

Onlamoon,N, DasGupta, J, Sharma, P, Rogers, K, Suppiah, S, Rhea, J, Molinaro, RJ,Gaughan, C, Dong, B, Klein, E, Qui, X, Devare, S, Schochetman, G, Hackett, J,Silverman, R, & Villinger, F (2011). Infection, viral dissemination andantibody responses of Rhesus macaques exposed to the human gammaretrovirus XMRVJournal of Virology

This article originally appeared on virology blog.