Advances in Clinical Diagnosis of Tuberculosis

Tuberculosis (TB) holds a central and deadly platform around the globe, affecting mankind with around one-third of the world being affected by latent TB. TB progresses in the body through inhalation process and has a critical discrimination in terms of affecting individuals depending upon age, sex, socio-economic status, and even the stature of nation (developed or developing). The biggest challenge in TB management is accurate, direct, early diagnosis, and an ability to differentiate the type of mycobacterium. The most common and reliable direct methods include tuberculosis skin test (TST), smear microscopy, nucleic acid amplification tests (NAAT), and immuno-chromatographic-based methods. However, culturing the specimen on a mycobacterium specific media is considered the ‘gold standard' for diagnosis of TB by the WHO. Mycobacterium cultures are used extensively for bacilli differentiation and also for predicting drug susceptibility testing in multi-drug-resistant TB. This chapter discusses the merits and demerits of many approaches to distinguish and identify the type of mycobacterium.

Exploring the Risk Posed by Animals with an Inconclusive Reaction to the Bovine Tuberculosis Skin Test in England and Wales

The single intradermal comparative cervical tuberculin (SICCT) test is the primary test for ante-mortem diagnosis of bovine tuberculosis (TB) in England and Wales. When an animal is first classified as an inconclusive reactor (IR) using this test, it is not subject to compulsory slaughter, but it must be isolated from the rest of the herd. To understand the risk posed by these animals, a case-control study was conducted to measure the association between IR status of animals and the odds of them becoming a reactor to the SICCT at a subsequent test. The study included all animals from herds in which only IR animals were found at the first whole herd test in 2012 and used data from subsequent tests up until the end of 2016. Separate mixed-effects logistic regression models were developed to examine the relationship between IR status and subsequent reactor status for each risk area of England and for Wales, adjusting for other explanatory variables. The odds of an animal becoming a subsequent reactor during the study period were greater for IR animals than for negative animals in the high-risk area (odds ratio (OR): 6.85 (5.98–7.86)) and edge area (OR: 8.79 (5.92–13.04)) of England and in Wales (OR: 6.87 (5.75–8.22)). In the low-risk area of England, the odds were 23 times greater, although the confidence interval around this estimate was larger due to the smaller sample size (11–48, p < 0.001). These findings support the need to explore differential controls for IR animals to reduce the spread of TB, and they highlight the importance of area-specific policies.

Display of Antigens on Polyester Inclusions Lowers the Antigen Concentration Required for a Bovine Tuberculosis Skin Test

ABSTRACTThe tuberculin skin test is the primary screening test for the diagnosis of bovine tuberculosis (TB), and use of this test has been very valuable in the control of this disease in many countries. However, the test lacks specificity when cattle have been exposed to environmental mycobacteria or vaccinated withMycobacterium bovisbacille Calmette-Guérin (BCG). Recent studies showed that the use of three or four recombinant mycobacterial proteins, including 6-kDa early secretory antigenic target (ESAT6), 10-kDa culture filtrate protein (CFP10), Rv3615c, and Rv3020c, or a peptide cocktail derived from those proteins, in the skin test greatly enhanced test specificity, with minimal loss of test sensitivity. The proteins are present in members of the pathogenicMycobacterium tuberculosiscomplex but are absent in or not expressed by the majority of environmental mycobacteria and the BCG vaccine strain. To produce a low-cost skin test reagent, the proteins were displayed at high density on polyester beads through translational fusion to a polyhydroxyalkanoate synthase that mediates the formation of antigen-displaying inclusions in recombinantEscherichia coli. Display of the proteins on the polyester beads greatly increased their immunogenicity, allowing for the use of very low concentrations of proteins (0.1 to 3 μg of mycobacterial protein/inoculum) in the skin test. Polyester beads simultaneously displaying all four proteins were produced in a single fermentation process. The polyester beads displaying three or four mycobacterial proteins were shown to have high sensitivity for detection ofM. bovis-infected cattle and induced minimal responses in animals exposed to environmental mycobacteria or vaccinated with BCG.