Use of amino acid sequences from
The present invention refers to the use of gene sequences or portions thereof characterized in that the same belong to the classes of in vitro and ex vivo induced, repressed or conserved genes in Mycobacterium tuberculosis currently infected human macrophages and to corresponding peptides or consensus peptides or proteins for the preparation of specific bio-markers for the diagnosis and prevention of active or latent disease.
1. A method for detecting a Mycobacterium species infection in a test subject, comprising:
(a) incubating in vitro a blood sample or a lavage sample that comprises lymphocytes from the test subject with at least one isolated protein or peptide that comprises SEQ ID NO: 43, 44, 45, 46, or 47,
wherein said step of incubating is for a time and under conditions sufficient to stimulate the lymphocytes to produce an effector molecule; and
(b) detecting a test level of the effector molecule, wherein detection of a test level that is higher than a control level that is detected following incubation of control lymphocytes from a healthy control subject, indicates that the test subject is infected with, or has been previously exposed to, the Mycobacterium species.
2. The method of claim 1 wherein the Mycobacterium species is selected from M. tuberculosis, M. bovis, M. bovis BCG, M. africanum, M. canetti, M. caprae, M. microti, M. pinnipedii, M. avium, M. avium paratuberculosis, M. avium silvaticum, M. avium “hominissuis”, M. colombiense, M. asiaticum, M. gordonae, M. gastri, M. kansasii, M. hiberniae, M. nonchromogenicum, M. terrae, M. triviale, M. ulcerans, M. pseudoshottsii, M. shottsii, M. triplex, M. genavense, M. florentinum, M. lentiflavum, M. palustre, M. kubicae, M. parascrofulaceum, M. heidelbergense, M. interjectum, M. simiae, M. branderi, M. cookii, M. celatum, M. bohemicum, M. haemophilus, M. malmoense, M. szulgai, M. leprae, M. lepraemurium, M. lepromatosis, M. africanum, M. botniense, M. chimaera, M. conspicuum, M. doricum, M. farcinogenes, M. heckeshornense, M. intracellulare, M. lacus, M. marinum, M. monacense, M. montefiorense, M. murale, M. nebraskense, M. saskatchewanense, M. scrofulaceum, M. shimoidei, M. tusciae, M. xenopi, M. intermedium, M. abscessus, M. chelonae, M. bolletii, M. fortuitum, M. fortuitum subsp. acetamidolyticum, M. boenickei, M. peregrinum, M. porcinum, M. senegalense, M. septicum, M. neworleansense, M. houstonense, M. mucogenicum, M. mageritense, M. brisbanense, M. cosmeticum, M. parafortuitum, M. austroafricanum, M. diernhoferi, M. hodleri, M. neoaurum, M. frederiksbergense, M. aurum, M. vaccae, M. chitae, M. fallax, M. confluentis, M. flavescens, M. madagascariense, M. phlei, M. smegmatis, M. goodii, M. wolinskyi, M. thermoresistibile, M. gadium, M. komossense, M. obuense, M. sphagni, M. agri, M. aichiense, M. alvei, M. arupense, M. brumae, M. canariasense, M. chubuense, M. conceptionense, M. duvalii, M. elephantis, M. gilvum, M. hassiacum, M. holsaticum, M. immunogenum, M. massiliense, M. moriokaense, M. psychrotolerans, M. pyrenivorans, M. vanbaalenii, M. pulveris, M. arosiense, M. aubagnense, M. caprae, M. chlorophenolicum, M. fluoroanthenivorans, M. kumamotonense, M. novocastrense, M. parmense, M. phocaicum, M. poriferae, M. rhodesiae, M. seoulense and M. tokaiense.
3. The method of claim 1 wherein
the Mycobacterium species is Mycobacterium tuberculosis.
4. The method of claim 1 wherein at least one of:
(a) heparin is present in the step of incubating,
(b) heparin and an added carbohydrate are present in the step of incubating, and
(c) at least one Mycobacterium protein, or a peptide fragment thereof or a chemical analog thereof, is present in the step of incubating, said Mycobacterium protein being selected from ESAT6, CFP10, TB7.7 and PPD.
5. The method of claim 1 wherein the effector molecule is selected from interferon-γ, TNF-α, and an interleukin.
6. The method of claim 1 wherein the effector molecule is interferon-γ.
7. The method of claim 1 , wherein the subject is a human subject.
8. The method of claim 1 , wherein the subject is a non-human animal.
9. The method of claim 1 , wherein in the step of incubating there is present one or more Mycobacterium proteins, wherein the one or more Mycobacterium proteins are selected from the group consisting of ESAT6, CFP10, TB7.7 and PPD, and wherein the detected test level of the effector molecule is greater than a detectable effector molecule level that can be produced by said lymphocytes incubated only with said one or more Mycobacterium proteins selected from the group consisting of ESAT6, CFP10, TB7.7 and PPD.
10. The method of claim 1 , wherein in the step of incubating there is further present one or more isolated proteins or peptides comprising at least one of the amino acid sequences selected from the group consisting of SEQ ID NOS: 1-42 and 48-74.
11. The method of claim 1 , wherein in the step of incubating there is further present an isolated protein or peptide comprising SEQ ID NO: 48, 49, or 50.
12. The method of claim 1 , wherein in the step of incubating there is further present an isolated protein or peptide comprising SEQ ID NO: 64, 65, or 66.
13. The method of claim 1 , wherein in the step of incubating there is further present at least 5 isolated proteins or peptides comprising SEQ ID NOS: 43-47, respectively.
14. The method of claim 1 , wherein in the step of incubating there is further present at least 3 isolated proteins or peptides comprising SEQ ID NOS: 48-50, respectively.
15. The method of claim 1 , wherein in the step of incubating there is further present at least 3 isolated proteins or peptides comprising SEQ ID NOS: 64-66, respectively.