Posts Tagged ‘Mycobacteriophages’

Killing of Mycobacterium avium and Mycobacterium tuberculosis by a Mycobacteriophage Delivered by a Nonvirulent Mycobacterium: A Model for Phage Therapy of Intracellular Bacterial Pathogens

January 12, 2017
Lawrence Broxmeyer, Danuta Sosnowska, Elizabeth Miltner, Ofelia Chacon, Dirk Wagner, Jeffery McGarvey, Raul G. Barletta, and Luiz E. Bermudez

Killing of Mycobacterium avium and Mycobacterium tuberculosis by a Mycobacteriophage

The Journal of Infectious Diseases

View this document on Scribd



1. Bloom B. Tuberculosis: pathogenesis, protection and control. Washington,DC: American Society for Microbiology Press, 1995.
2. Surveillance TWIGP. Anti-tuberculosis drug resistance in the world. Geneva:World Health Organization Global Tuberculosis Programme, 1997.
3. Inderlied CB, Kemper CA, Bermudez LE. The Mycobacterium avium complex. Clin Microbiol Rev 1993; 6:266–310.
4. Palella FJ Jr, Delaney KM, Moorman AC, et al. Declining morbidity and mortality among patients with advanced human immunodeficiency virus infection. HIV Outpatient Study Investigators. N Engl J Med 1998; 338: 853–60.
5. Kaplan JE, Hanson D, Dworkin MS, et al. Epidemiology of human immunodeficiency virus–associated opportunistic infections in the United
States in the era of highly active antiretroviral therapy. Clin Infect Dis 2000 ; 30(Suppl 1):S5–14.
6. Falkinham JO 3rd. Epidemiology of infection by nontuberculous mycobacteria. Clin Microbiol Rev 1996; 9:177–215.
7. Guthertz LS, Damsker B, Bottone EJ, Ford EG, Midura TF, Janda JM. Mycobacterium avium and Mycobacterium intracellulare infections in patients with and without AIDS. J Infect Dis 1989; 160:1037–41.
8. Heifets L. Susceptibility testing of Mycobacterium avium complex isolates. Antimicrob Agents Chemother 1996; 40:1759–67.
9. Horsburgh CR Jr. Mycobacterium avium complex infection in the acquired immunodeficiency syndrome. N Engl J Med 1991; 324:1332–8.
10. Chaisson RE, Benson CA, Dube MP, et al. Clarithromycin therapy for bacteremic Mycobacterium avium complex disease: a randomized, double-blind, dose-ranging study in patients with AIDS. AIDS Clinical Trials Group Protocol 157 Study Team. Ann Intern Med 1994; 121:905–11.
11. Young LS, Wiviott L, Wu M, Kolonoski P, Bolan R, Inderlied CB. Azithromycin for treatment of Mycobacterium avium–intracellulare complex infection in patients with AIDS. Lancet 1991; 338:1107–9.
12. Bermudez LE, Kolonoski P, Young LS. Roxithromycin alone and in combination with either ethambutol or levofloxacin for disseminated Mycobacterium avium infections in beige mice. Antimicrob Agents Chemother 1996; 40:1033–5.
13. Dube MP, Sattler FR, Torriani FJ, et al. A randomized evaluation of ethambutol for prevention of relapse and drug resistance during treatment of Mycobacterium avium complex bacteremia with clarithromycin-based combination therapy. California Collaborative Treatment Group. J Infect Dis 1997; 176:1225–32.
14. Holzman D. Phage as antibacterial tool. Genetic Engineering News 1998; 18:11–16.
15. Ford ME, Stenstrom C, Hendrix RW, Hatfull GF. Mycobacteriophage TM4: genome structure and gene expression. Tuber Lung Dis 1998; 79:63–73.
16. Foley-Thomas EM, Whipple DL, Bermudez LE, Barletta RG. Phage infection,transfection and transformation of Mycobacterium avium complex and Mycobacterium paratuberculosis. Microbiology 1995; 141:1173–81.
17. Bermudez LE, Parker A, Goodman JR. Growth within macrophages increases the efficiency of Mycobacterium avium in invading other macrophages by a complement receptor–independent pathway. Infect Immun 1997; 65:1916–25.
18. Jacobs WR Jr, Kalpana GV, Cirillo JD, et al. Genetic systems for mycobacteria. Methods Enzymol 1991; 204:537–55.
19. Black CM, Bermudez LE, Young LS, Remington JS. Coinfection of macrophages modulates interferon gamma and tumor necrosis factor–induced activation against intracellular pathogens. J Exp Med 1990; 172:977–80.
20. Hafner R, Inderlied CB, Peterson DM, et al. Correlation of quantitative bone marrow and blood cultures in AIDS patients with disseminated Mycobacterium avium complex infection. J Infect Dis 1999; 180:438–47.
21. Sula L, Sulova J, Stolcpartova M. Therapy of experimental tuberculosis in guinea pigs with mycobacterial phages DS-6A, GR-21 T, My-327. Czech Med 1981; 4:209–14.
22. Rastogi N, Labrousse V. Extracellular and intracellular activities of clarithromycin used alone and in association with ethambutol and rifampin against Mycobacterium avium complex. Antimicrob Agents Chemother 1991; 35:462–70.
23. Bermudez LE, Young LS. New drugs for the therapy of mycobacterial infections. Curr Opinion Infect Dis 1995; 8:428–38.
24. Sturgill-Koszycki S, Schlesinger PH, Chakraborty P, et al. Lack of acidification in Mycobacterium phagosomes produced by exclusion of the vesicular proton-ATPase. Science 1994; 263:678–81.
25. Clemens DL, Horwitz MA. Characterization of the Mycobacterium tuberculosis phagosome and evidence that phagosomal maturation is inhibited. J Exp Med 1995
; 181:257–70.
26. de Chastellier C, Lang T, Thilo L. Phagocytic processing of the macrophage endoparasite, Mycobacterium avium, in comparison to phagosomes which contain Bacillus subtilis or latex beads. Eur J Cell Biol 1995; 68:167–82.
27. Gomes MS, Paul S, Moreira AL, Appelberg R, Rabinovitch M, Kaplan G. Survival of Mycobacterium avium and Mycobacterium tuberculosis in acidified vacuoles of murine macrophages. Infect Immun 1999; 67:3199–206.


Further Evidence for Cancer as a Cell-Wall-Deficient Mycobacterial Disease

December 5, 2016

A.P. Lysenko PhD, L. Broxmeyer MD, V.V. Vlasenko PhD, P.A. Krasochko PhD, A.P.Lemish PhD, and E.A. Krasnikova

Further Evidence for Cancer as a Cell-Wall-Deficient Mycobacterial Disease.pdf

Corresponding author:
Lawrence Broxmeyer, M.D

© Under License of Creative Commons Attribution 3.0 License


Received: October 07, 2016; Accepted: November 03, 2016; Published: November
14, 2016



In 2014, Buehring reported that Bovine Leukemic Virus (BLV), a common oncogenic retrovirus of cattle, was present in some humans, primarily localized to the breast epithelium  ―  the  very  cell  type  from  which  most  breast  malignancies  arise.  By 2015, there appeared data (Buehring, 2015) supporting that as many as 37% of human breast cancer cases could be attributable to BLV exposure. But if recent estimates suggest over 83% of U.S. dairy operations are currently positive for BLV, they also show that approximately 68% are positive for cell-wall-deficient Mycobacterium avium subspecies paratuberculosis (MAP). Although tubercular lung infection has been said to cause 11 times the incidence of lung cancer as normal control subjects, it is its cell-wall-deficient (CWD) forms (also called L-forms) that have recently repeatedly been found through genetic analysis and appropriate stains in such cancer tissue ― suggesting that CWD tuberculosis or atypical tuberculosis “is likely to be involved in the occurrence or development of lung carcinoma”. A similar relationship between tubercular L-forms and the genesis of the very breast cancer addressed in the aforementioned BLV viral trials. This is not a coincidence. L-forms (CWD forms) predominate and are crucial to the survival of mycobacteria in vivo and they have been documented by fluorescence microscopy in all intracellular macrophage-grown M. tuberculosis observed. From its origin, the very concept of the “BLV leukemic virus” has been on shaky, unstable ground. In 1969, veterinarians Janice and Lyle Miller from the University of Wisconsin-Madison spotted C-shaped “virus-like” particles in cattle lymphosarcoma insisting that these were similar to other C-type viruses “regarded as the cause of leukemia in other species.” But by 1978, scientists at Downstate reported atypical mycobacterial forms, including its preferred filterable virus-sized “L” or cell-wall-deficient (CWD) forms in not only leukemia but all other malignancies ― all having, as their common denominator the continuous presence of mycobacterial C-shaped forms.

Tracing back to techniques similar to Miller and Millers original BLV study we found in the very lyophilized antigens present in commercial kits for the diagnosis of BLV (AgBLV), these very same CWD (cell-wall-deficient) mycobacteria and mycobacterial DNA in all BLV samples ― which when introduced into guinea pigs stimulated the same antibody as occurred when mycobacteria-infected internal organ homogenates themselves were injected into other guinea pigs. It is therefore assumed that the Bovine Leukemic Virus (BLV) is being mistaken for viral-like forms of cell-wall-deficient (CWD) atypical tubercular mycobacteria. Since latent tubercular infection, as well as the administration of BCG and tuberculin also results in persistent CWD forms, their possible role in carcinogenesis is also considered.

KEYWORDS: Cancer; Mycobacterium tuberculosis; Bovine Leukemic Virus; BLV; Mycobacteriophages



1   Rous PA (1911) Sarcoma of the fowl transmissible by an agent separable from the tumour cells. J Exp Med 13: 397-411.

2    Bittner JJ (1936) Some possible effects of nursing on the mammary gland tumour incidence of mice. Science 84: 162.

3    Van Helvoort T (1994) History of virus research in the 20th century: the problem of conceptual continuity. History of Science 32: 185-235.

4    Livingston V (1972) Cancer: A New Breakthrough. Nash Publishing p: 269.

5  Livingston V (1970) Specific type of organism cultured from malignancy: bacteriology and proposed classification.  Ann NY Acad Sci 174: 636-654.

6    Duran-Reynals F (1950) Neoplastic infection and cancer. Am J Med 8: 440-511.

7    Glover T, Scott MA (1926) Study of the Rous chicken sarcoma no 1. Canada Lancet Pract 66: 49-62.

8    Diller I (1970) Experiments with mammalian tumor isolates. Ann NY Acad Sci 174: 655-674.

9    Miller JM, Miller LD, Olson C, Gillette KG (1969) Virus-like particles in phytohemagglutinin-stimulated lymphocyte cultures with reference to bovine lymphosarcoma. J Natl Cancer Inst 43: 1297-1305

10 Miller JM (1974) Animal model of human disease. Malignant lymphoma. Am J Pathol 75: 417-420.

11  Sorensen DK, Dutta SK, Hammer RF, Larson VL, Perman V, et al. (1970) Bovine lymphocytic leukemia: studies of etiology, pathogenesis and mode of transmission. Progress Report no. 10 to the U.S. Atomic Energy Commission, 1969-1970 p: 38

12  Alexander-Jackson EA (1954) specific type of microorganism isolated from animal and human cancer: bacteriology of the organism. Growth 18: 37-51.

13  Klieneberger-Nobel E (1949) Origin, development and significance of L-forms in bacterial cultures. J Gen Microbiol 3: 434-442.

14  Mattman LH (2000) Cell wall deficient forms: stealth pathogens. CRC Press.

15  Kashala O, Marlink R, Ilunga M, Diese M, Gormus B, et al. ( 1994) Infection with human immunodeficiency virus type 1 (HIV-1) and human T cell lymphotropic viruses among leprosy patients and contacts: correlation between HIV-1 cross-reactivity and antibodies to lipoarabinomannan. J Infect Dis 169: 296-304.

16  Glover T (1930) The bacteriology of cancer. Canada Lancet Pract 75: 92-111.

17  Mazet G (1941) Etude Bacteriologique sur la Maladie d’ Hodgkin. Montpellier Med pp: 1-6.

18  Livingston V, Allen R (1948) Presence of consistently recurring invasive myco- bacterial forms in tumor cells. Microscop Soc Bull 2: 5-18.

19  Wuerthele-Caspe V (1949) Mycobacterial forms observed in tumors. J Am Med. Womens Assoc 4: 135-141.

20  Alexander-Jackson  E  (1976)  Progenitor  Cryptocides,  The  Specific Pleomorphic Microorganism Isolated From Cancer. J Int Acad Metab 5: 31-39.

21  Alexander-Jackson E (1978) Microscopic and Submicroscopic Phases of P. Cryptocides from Fresh Lymphocytic leukemia. J Int Acad Metab 1: 9-18.

22  Diller I, Diller W (1965) Intracellular acid-fast organisms isolated from malignant tissues. Trans Am Micr Soc 84: 138-148.

23  Diller I, Donnelly A, Fisher M (1967) Isolation of pleomorphic, acid- fast organisms from several strains of mice. Cancer Res 27: 1402-1408.

24 Seibert F, Feldmann F, Davis R, Richmond I (1970) Morphological, biological, and immunological studies on isolates from tumors and leukemic bloods. Ann N Y Acad Sci 174: 690-728.

25  Wang A, Xie J (1998) Infection of mycobacterium tuberculosis in lung cancer. Zhongguo Fei Ai Za Zhi 1: 92-94.

26  Guliang  H,  Tefu  L  (1999)  Mycobacterium  tuberculosis  L-forms. Microb Ecol Health Dis 10: 129-133.

27  Xie J, Anchao W, Xiazhi Z (1999) Isolation of acid fast bacillus L- forms from carcinoma of Lung. Acta Academiae Medicinae Bengbu 24: 145-146.

28 Song LY, Yan WS, Zhao T (2002) Detection of Mycobacterium tuberculosis in lung cancer tissue by indirect in situ nested PCR. Di Yi Jun Yi Da Xue Xue Bao 22: 992-993.

29 Yesong WXQ , Lifa X (2004)   A case report on pneumoconiotu- berculosis complicated with lung cancer and   Mycobacterium tuberculosis- L form infection. Chin J Industrial Med.

30 Zhang  S,  Guang-ling  Z,  Yan-sheng  T  (2009)  Detection  of Mycobacterium tuberculosis L forms infection in tissues of lung carcinoma. Chin J Public Health 25: 1317-1318.

31 Yang B, Tian Y, Cui X, Zhang W, Ma Y et al. Detection of Mycobacterium tuberculosis L-forms and MPB64 in breast cancer tissues The Journal of Practical Medicine. 2013; 29(15) p2552-2555.

32  Sheng TY, Kun CX, Tong H, Guang LH, Wei Z, et al. (2009) Study on the relationship between Mycobacterium tuberculosis L infection and lung cancer. Tumor 29: 1085-1089.

33  Tian Y, Hao T, Cao B, Zhang W, Ma Y, et al. (2015) Clinical End-Points Associated with Mycobacterium tuberculosis and Lung Cancer: Implications into Host- Pathogen Interaction and    Coevolution. Bio Med Research Intern p: 9.

34 Alexander-Jackson E (1970) Ultraviolet spectrogramic microscope studies of Rous sarcoma virus cultured in cell-free medium. Ann N Y Acad Sci 174: 765-781.

35  Van der Maaten M, Miller J (1976) Replication of bovine leukemia virus in monolayer cell cultures. Bibl Haemat 43: 360-362.

36  Lysenko  AP,  Drogun  AG,  Shurinova  (1998)  Studies  of  influence atypical mycobacterial  infection on AGID results with sera of cattle infected BLV (in Russian). Vet. nauka – proizvodstvu 33: 56-54.

37  ShivRaj L, Patil SA, Girdhar A, Sengupta U, Desikan KV, et al. (1988) Antibodies to HIV-1 in sera from patients with mycobacterial infections. Int J Leprosy 56: 546-551.

38 Lysenko AP, Vlasenko AP, Broxmeyer L (2014) Phenomenon of variability of mycobacteria and its use for detection of a tuberculosis infection.

39 Lysenko AP, Vlasenko VV, Broxmeyer L, Lemish AP, Novik TP, et al. (2014) The tuberculin skin test: how safe is safe?  The tuberculins contain unknown forms capable of reverting to cell-wall deficient mycobacteria. Clin Exp Med Sci 2: 55-73.

40  Lysenko AP, Vlasenko VV, Lemish AP (2014) Detection of mycobacteria in tissues by means of the differentiating immunoperoxidase staining. Tuberculos i bolezni legkhih 10: 55-58.

41  Duesberg PH (1987)  Retroviruses  as carcinogens and pathogens: expectations and reality. Cancer Res 47: 1199-220.

42  Demochowski L, Grey CE (1957) Subcellular Structures of Possible Viral Origin in Some Mammalian Tumors. Ann NY Acad Sci : pp 559-615

43 Laemmli UK (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227: 680-685.

44  Dameshek W and Gunz (1965) Leukemia. Am J Med Sci 249: 115.

45 Seibert  FB,  Feldmann  PM,  Davis  RL,  Richmond  IS  (1970) Morphological, Biological, and Immunological Studies on Isolates from Tumors and Leukemic Bloods. Ann NY Acad Sci 174: 690-728.

46 Mankiewicz E (1965) Bacteriophages that lyse mycobacteria and corynebacteria, and show cytopathogenic effect on tissue cultures of renal cells of cercopithecus aethiops: a preliminary communication. Can Med Assoc J 92: 31-33.

47  Seibert FB (1968) Pebbles on the hill of a scientist. St Petersburg Printing Company 1: 162.

48 Dobrindt U, Reidl J (2000) Pathogenicity islands and phage conversion: evolutionary aspects of bacterial pathogenesis. Int J Med Microbiol 290: 519-27.

49 Landman OE, Burchard WK, Angelety LH (1962) Lysogeny and bacteriophage adsorption in stable and reverting L-forms of Salmonella paratyphi B and Escherichia coli. Bacteriol Proc p: 53.

50  Falagas ME, Kouranos VD, Athanassa Z, Kopterides P (2010) Review ―Tuberculosis and malignancy. Q J Med 103: 461-487.

51  Nelson EL, Pickett MJ (1951) The Recovery of L Forms of Brucella and their Relation to Brucella Phage. J Infect Dis 89: 226-32.

52  Kruegar AP, Cohn T, Smith PN, McGuire CD (1948) J Gen Physiol 31: 477-488.

53 Takahashi S (1979) L phase growth of Mycobacteria. 1. Cell wall deficient form of Mycobacteria. Kekkaku 54: 63-70.

54  Broxmeyer L, Sosnowska  D,  Miltner E, Chacón O, Wagner D, et al. (2002) Killing of Mycobacterium avium and Mycobacterium tuberculosis by a mycobacteriophage delivered by a nonvirulent mycobacterium: a model for phage therapy of intracellular bacterial pathogens. J Infect Dis 186: 1155-60.

55 Devadoss PO, Klegerman ME, Groves MJ (1993) Phagocytosis of Mycobacterium bovis BCG organisms by murine S180 sarcoma cells. Cytobios 74: 49-58.

56 Marcova N, Michailova L, Kussovsski V, Jordanova M (2008) Formation of persisting Cell Wall Deficient Forms M. bovis BCG during interaction with peritoneal macrophages in guinea pigs. Electronic J. of Biology 4: 1-10.

57  Chauhan A, Madiraju MV, Fol M, Lofton H, Maloney E, et al. (2006) Mycobacterium tuberculosis Cells Growing in Macrophages Are Filamentous and Deficient in FtsZ Rings. J Bacteriol 188: 1856-1865

58  Wagner PL, Waldor MK (2002) Bacteriophage Control of Bacterial Virulence. Infect Immun 70: 3985-3993.