The contributing authors to this article have declared no conflict of interest.
Review Article
Killer applications: Toward affordable rapid cell-based diagnostics for malaria and tuberculosis†‡
Article first published online: 28 JAN 2008
DOI: 10.1002/cyto.b.20401
Copyright © 2008 Clinical Cytometry Society
Issue

Cytometry Part B: Clinical Cytometry
Supplement: The Global Health and Diagnostic (Flow) Cytometry-Breakthrough in HIV and Tuberculosis
Volume 74B, Issue S1, pages S152–S164, 2008
Additional Information
How to Cite
Shapiro, H. M. and Perlmutter, N. G. (2008), Killer applications: Toward affordable rapid cell-based diagnostics for malaria and tuberculosis. Cytometry, 74B: S152–S164. doi: 10.1002/cyto.b.20401
- †
- ‡
How to cite this article: Shapiro HM, Perlmutter NG. Killer applications: Toward affordable rapid cell-based diagnostics for malaria and tuberculosis. Cytometry Part B 2008; 74B (Suppl. 1): S152–S164, 2008.
Publication History
- Issue published online: 19 MAR 2008
- Article first published online: 28 JAN 2008
- Manuscript Accepted: 28 NOV 2007
- Manuscript Received: 17 OCT 2007
Funded by
- NIH. Grant Numbers: AI 060272, AI 063833, HL 080898, AI074471
LITERATURE CITED
- 1,,,,,,. Pneumocystis carinii pneumonia and mucosal candidiasis in previously healthy homosexual men: Evidence of a new acquired cellular immunodeficiency. New Engl J Med 1981; 305: 1425–1431.
- 2,,,,,,. The prognostic value of cellular and serologic markers in infection with human immunodeficiency virus type 1. N Engl J Med 1990; 322: 166–172.
- 3,,,. T-cell subset counting and the fight against AIDS: Reflections over a 20-year struggle. Cytometry 2002; 50: 39–45.Direct Link:
- 4,,,,,. The origins of acquired immune deficiency syndrome viruses: Where and when? Philos Trans R Soc Lond B Biol Sci 2001; 356: 867–876.
- 5. The Malaria Capers. Tales of Parasites and People. New York: Norton; 1991. 288 p.
- 6. The Forgotten Plague: How the Battle Against Tuberculosis was Won—and Lost. Boston: Little, Brown and Company; 1993. xix + 460 pp.
- 7,,. Dual infection with HIV and malaria fuels the spread of both diseases in sub-Saharan Africa. Science 2006; 314: 1603–1606. Erratum in:Science2007;315:598.
- 8
- 9,,. Tuberculosis and HIV coinfection: Current state of knowledge and research priorities. J Infect Dis 2007; 196( Suppl 1): S1–S3.
- 10,,,,,,,. HIV infection and multidrug-resistant tuberculosis: The perfect storm. J Infect Dis 2007; 196( Suppl 1): S86–S107.
- 11. “Cellular astronomy”—A foreseeable future in cytometry. Cytometry Part A 2004; 60A: 115–124.Direct Link:
- 12,. Personal cytometers—Slow flow or no flow? Cytometry Part A 2009; 69A: 620–630.Direct Link:
- 13. Microbe Hunters (1926; republished). San Diego: Harcourt Brace; 1996. 357 p.
- 14. Plagues and Peoples. New York: Doubleday; 1976. 340 p.
- 15,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,. Genome sequence of the human malaria parasite Plasmodium falciparum. Nature 2002; 419: 498–511.
- 16,,,. Evolutionary relatedness of Plasmodium species as determined by the structure of DNA. Science 1984; 225: 808–811.
- 17,,,. Karyotype and synteny among the chromosomes of all four species of human malaria parasite. Mol Biochem Parasitol 1999; 101: 23–32.
- 18,,. The cell cycle of parasitic protozoa: Potential for chemotherapeutic exploitation. Prog Cell Cycle Res 2003; 5: 91–101.
- 19,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,. Deciphering the biology of Mycobacterium tuberculosis from the complete genome sequence. Nature 1998; 393: 537–544.
- 20World Health Organization. Global tuberculosis control—Surveillance, planning, financing. WHO Report 2007 (WHO/HTM/TB/2007.376). Available at http://www.who.int/tb/publications/global_report/en.
- 21,,,,,,,. Global incidence of multidrug-resistant tuberculosis. J Infect Dis 2006; 194: 479–485.
- 22Centers for Disease Control and Prevention (CDC). Emergence of Mycobacterium tuberculosis with extensive resistance to second-line drugs worldwide, 2000–2004. MMWR Morb Mortal Wkly Rep 2006; 55: 301–305.
- 23,,,,,,,,,,,,,,,,,,,. Worldwide emergence of extensively drug-resistant tuberculosis. Emerg Infect Dis 2007; 13: 380–387.
- 24. Legal power and legal rights—Isolation and quarantine in the case of drug-resistant tuberculosis. N Engl J Med 2007; 357: 433–435.
- 25. Micrographia. London: The Royal Society; 1665.Mineola, NY: Dover Publications;2003 (reprinted). 273 p.
- 26,. History of Staining, 3rd ed. Baltimore: Williams & Wilkins; 1983. x + 304 p.
- 27. Mauve. New York: Norton; 2001. 222 p.
- 28,,. The chemistry of the lipids of tubercle bacilli. LIV. The isolation and properties of mycolic acid. J Biol Chem 1938; 126: 505–513.
- 29,. Structure of the mycolic acids of mycobacteria. Nature 1950; 166: 782–783.
- 30,,,,,,. Mycolic acids: Structure, biosynthesis and physiological functions. Prog Lipid Res 1998; 37: 143–179.
- 31,,,. Separation and characterization of individual mycolic acids in representative mycobacteria. Microbiology 2001; 147: 1825–1837.
- 32,,. Pathway to synthesis and processing of mycolic acids in Mycobacterium tuberculosis. Clin Microbiol Rev 2005; 18: 81–101.
- 33. Fluoreszenzfärbung von Tuberkelbakterien mit Auramin. Münch Med Wschr 1938; 85: 1066–1068.
- 34,,,,,,,,,. Fluorescence versus conventional sputum smear microscopy for tuberculosis: A systematic review. Lancet Infect Dis 2006; 6: 570–581.
- 35,,,. Light emitting diodes for auramine O fluorescence microscopic screening of Mycobacterium tuberculosis. Int J Tuberc Lung Dis 2006; 10: 1060–1062.
- 36,,,. Fluorescence microscopy for tuberculosis diagnosis. Lancet Infect Dis 2007; 7: 236–237.
- 37,,,,. Fluorescence microscopy for tuberculosis diagnosis. Lancet Infect Dis 2007; 7: 238–239.
- 38. 100 years ago: Giemsa's solution for staining of plasmodia. Trop Med Int Health 2004; 9: 755–756.Direct Link:
- 39
- 40,. Identification of malaria parasites by fluorescence microscopy and acridine orange staining. Bull World Health Organ 1973; 48: 591–596.
- 41,,,,,. Malaria diagnosis by direct observation of centrifuged samples of blood. Am J Trop Med Hyg 1988; 39: 337–342.
- 42,,,,. Detection of Plasmodium falciparum infection with the fluorescent dye, benzothiocarboxypurine. Am J Trop Med Hyg 1991; 44: 11–16.
- 43,. Rapid microscopic detection of malaria parasites permanently fluorochrome stained in blood smears with aluminum and morin. Am J Clin Pathol 1991; 95: 424–427.
- 44,,,. Rapid diagnosis of malaria by use of fluorescent probes. Diagn Microbiol Infect Dis 1993; 17: 293–297.
- 45,,,,. The reliability of diagnostic techniques in the diagnosis and management of malaria in the absence of a gold standard. Lancet Infect Dis. 2006; 6: 582–588.
- 46“Student” [ ]. On the error of counting with a haemacytometer. Biometrika 1907; 5: 351–360.
- 47. Rapid diagnostic tests for malaria parasites. Clin Microbiol Rev 2002; 15: 66–78.
- 48. Diagnosis of malaria: A review of alternatives to conventional microscopy. Clin Lab Haematol 1999; 21: 235–245.Direct Link:
- 49,,,,,,,. Sources of variability in determining malaria parasite density by microscopy. Am J Trop Med Hyg 2005; 73: 593–598.
- 50,,. Malaria vaccine efficacy: The difficulty of detecting and diagnosing malaria. Malaria J 2007; 6: 36.
- 51. On the nature of Romanowsky-Giemsa staining and its significance for cytochemistry and histochemistry: An overall view. Histochem J 1983; 15: 1029–1047.
- 52,,,,,. [Romanowsky dyes and the Romanowsky-Giemsa effect. 3. Microspectrophotometric studies of Romanowsky-Giemsa staining. Spectroscopic evidence of a DNA-azure B-eosin Y complex producing the Romanowsky-Giemsa effect.] Histochemistry 1984; 81: 337–351 (in German).
- 53,. Understanding Romanowsky staining. I. The Romanowsky-Giemsa effect in blood smears. Histochemistry 1987; 86: 331–336.
- 54,. Cytometry in malaria: Moving beyond Giemsa. Cytometry Part A 2007; 71A: 643–645.Direct Link:
- 55,,,. Cytofluorograf detection of Plasmodium yoelii, Trypanosoma gambiense, and Trypanosoma equiperdum by laser excited fluorescence of stained rodent blood. J Parasitol 1977; 63: 593–598.
- 56,,. Plasmodium-infected blood cells analyzed and sorted by flow fluorimetry with the deoxyribonucleic acid binding dye 33258 Hoechst. J Histochem Cytochem 1979; 27: 803–813.
- 57,,,,,. Development and validation of flow cytometric measurement for parasitaemia using autofluorescence and YOYO-1 in rodent malaria. Parasitology 2007; 134: 1151–1162.
- 58,,,,,,,. Re-evaluating acridine orange for rapid flow cytometric enumeration of parasitemia in malaria-infected rodents. Cytometry Part A 2007; 71A: 662–667.Direct Link:
- 59,. Analysis of Plasmodium falciparum growth in culture using acridine orange and flow cytometry. J Histochem Cytochem 1986; 34: 215–220.
- 60. Two-color flow-cytometric analysis of the growth cycle of Plasmodium falciparum in vitro: Identification of cell cycle compartments. J Histochem Cytochem 1986; 34: 1651–1658.
- 61,,. Haemozoin: From melatonin pigment to drug target, diagnostic tool, and immune modulator. Lancet Infect Dis 2007; 7: 675–685.
- 62,,,,,,,,,. Automated malaria detection by depolarization of laser light. Br J Haematol 1999; 104: 499–503.Direct Link:
- 63,,,,. Relative frequency of malaria pigment-carrying monocytes of nonimmune and semi-immune patients from flow cytometric depolarized side scatter. Cytometry 2001; 45: 133–140.Direct Link:
- 64,,,,,,,. Sensitivity of hemozoin detection by automated flow cytometry in non- and semi-immune malaria patients. Cytometry Part B Clin Cytom 2003; 55B: 46–51.Direct Link:
- 65,,,,,,,,,. Sputum processing methods to improve the sensitivity of smear microscopy for tuberculosis: A systematic review. Lancet Infect Dis 2006; 6: 664–674.
- 66MurrayPR, BaronEJ, JorgensenJH, LandryML, PfallerMA, editors. Manual of Clinical Microbiology, 9th ed. Washington: ASM Press; 2007. 2488 p.
- 67
- 68. Fluorescence of auramine O in the presence of nucleic acid. C R Hebd Seances Acad Sci 1951; 232: 1708–1710 (in French).
- 69,,,,. An automated optoelectronic reticulocyte counter. Am J Clin Pathol 1989; 92: 57–61.
- 70,,. SYTO 16 labelling and flow cytometry of Mycobacterium avium. Lett Appl Microbiol 1997; 25: 437–441.Direct Link:
- 71,,. Safe susceptibility testing of Mycobacterium tuberculosis by flow cytometry with the fluorescent nucleic acid stain SYTO 16. J Med Microbiol 2005; 54: 77–81.
- 72,,,. Novel method using a laser scanning cytometer for detection of mycobacteria in clinical samples. J Clin Microbiol 2004; 42: 906–908.
- 73,,. Some observations of mycobacterial acid-fastness. Am Rev Respir Dis 1978; 118: 151–154.
- 74
- 75,,,,,,. Evaluation of four commercially available auramine O stain sets. Presented at the American Society for Microbiology General Meeting, Orlando, FL, May 2006. Poster C-324.
- 76,,,,. Automated identification of tubercle bacilli in sputum. A preliminary investigation. Anal Quant Cytol Histol 1999; 21: 277–282.
- 77,,. Automatic identificationof Mycobacterium tuberculosis by Gaussian mixture models. J Microsc 2006; 223(Part 2): 120–132.Direct Link:
- 78,,,. Diagnosis of smear-negative pulmonary tuberculosis in people with HIV infection or AIDS in resource-constrained settings: Informing urgent policy changes. Lancet 2007; 369: 2042–2049.
- 79,. A review of the diagnosis and treatment of smear-negative pulmonary tuberculosis. Int J Tuberc Lung Dis 2000; 4: 97–107.
- 80,,. Transmission of tubercle bacilli: The effects of chemotherapy. Tubercle 1976; 57: 275–299.
- 81FriedenT, editor. Toman's Tuberculosis. Case Detection, Treatment, and Monitoring—Questions and Answers,2nd ed. Geneva: World Health Organization; 2004. pp 11–21. Available at http://www.who.int/tb/publications/toman/en/index.html (online free).
- 82,,,. Enumeration of tubercle bacilli in sputum of patients with pulmonary tuberculosis. Antimicrob Agents Chemother 1973; 4: 94–104.
- 83,. Novel multipurpose methodology for detection of mycobacteria in pulmonary and extrapulmonary specimens by smear microscopy, culture, and PCR. J Clin Microbiol 2005; 43: 2697–2702.
- 84,,,,. Simplified detection of Mycobacterium tuberculosis in sputum using smear microscopy and PCR with molecular beacons. J Med Microbiol 2007; 56: 1356–1362.
- 85,,,,. Cytometric detection of mycobacterial surface antigens: Exposure of mannosyl epitopes and of the arabinan segment of arabinomannans. J Bacteriol 1996; 178: 7254–7259.
- 86,,,,. Analysis of bacteria in environmental and medical microbiology by flow cytometry. In: VeheriA, TiltonRC, BalowsA, editors. Rapid Methods and Automation in Microbiology and Immunology. Berlin: Springer; 1991. pp 131–140.
- 87,,,,,. Direct detection and identification of Mycobacterium tuberculosis in smear-positive sputum samples by fluorescence in situ hybridization (FISH) using peptide nucleic acid (PNA) probes. Int J Tuberc Lung Dis 1999; 3: 830–837.
- 88,,,,,,. Fluorescence in situ hybridization assay using peptide nucleic acid probes for differentiation between tuberculous and nontuberculous mycobacterium species in smears of mycobacterium cultures. J Clin Microbiol 1999; 37: 2760–2765.
- 89,,. Evaluation of a fluorescence in situ hybridization assay for differentiation between tuberculous and nontuberculous Mycobacterium species in smears of Lowenstein-Jensen and Mycobacteria growth indicator tube cultures using peptide nucleic acid probes. J Clin Microbiol 2001; 39: 1032–1035.
- 90,,,,,,,,,,,,,,. Microscopic-observation drug-susceptibility assay for the diagnosis of TB. N Engl J Med 2006; 355: 1539–1550.
- 91,,. Bacterial membrane potential analyzed by spectrofluorocytometry. Curr Microbiol 1985; 12: 183–186.
- 92,,,. Multiparameter flow cytometric analysis of antibiotic effects on membrane potential, membrane permeability, and bacterial counts of Staphylococcus aureus and Micrococcus luteus. Antimicrob Agents Chemother 2000; 44: 827–834.
- 93,,. Correlation of daptomycin bactericidal activity and membrane depolarization in Staphylococcus aureus. Antimicrob Agents Chemother 2003; 47: 2538–2544.
- 94,,,,. Rapid susceptibility testing of Mycobacterium tuberculosis (H37Ra) by flow cytometry. J Clin Microbiol 1995; 33: 1231–1237.
- 95,,,,. Rapid susceptibility testing for nontuberculosis mycobacteria using flow cytometry. J Clin Microbiol 1996; 34: 1386–1390.
- 96,,,,. Flow cytometric testing of susceptibilities of Mycobacterium tuberculosis isolates to ethambutol, isoniazid, and rifampin in 24 hours. J Clin Microbiol 1998; 36: 1568–1573.
- 97,,,,. Safe determination of susceptibility of Mycobacterium tuberculosis to antimycobacterial agents by flow cytometry. J Clin Microbiol 1999; 37: 479–483.
- 98,,,,,,. Flow cytometric testing of susceptibilities of Mycobacterium avium to amikacin, ciprofloxacin, clarithromycin and rifabutin in 24 hours. Clin Microbiol Infect 2000; 6: 368–375.Direct Link:
- 99,,,,. Mycobacterium tuberculosis susceptibility testing by flow cytometry. Unit 11.7. In: RobinsonJP, DarzynkiewiczZ, DeanP,HibbsAR, OrfaoA, RabinovitchP, WheelessL, editors. Current Protocols in Cytometry. New York: Wiley; 2004. pp 11.7.1–11.7.8.
- 100,,,. Susceptibility testing of Mycobacterium tuberculosis: Comparison of the BACTEC TB-460 method and flow cytometric assay with the proportion method. Clin Microbiol Infect 2005; 11: 372–378.Direct Link:
- 101,,,,,. Rapid pyrazinamide susceptibility testing of Mycobacterium tuberculosis by flow cytometry. J Microbiol Methods 2006; 67: 266–272.
- 102,,. Rapid assay for mycobacterial growth and antibiotic susceptibility using gel microdrop encapsulation. J Clin Microbiol 1995; 33: 1720–1726.
- 103,,,,. Comparison of flow cytometric and Alamar Blue tests with the proportional method for testing susceptibility of Mycobacterium tuberculosis to rifampin and isoniazid. J Clin Microbiol 2004; 42: 2247–2248.
- 104,,,. Rapid mycobacteria drug susceptibility testing using Gel Microdrop (GMD) Growth Assay and flow cytometry. J Microbiol Methods 2005; 62: 181–197.
- 105,,. Spectre II. General-purpose microscope input for a computer. Science 1969; 166: 328–333.
- 106,,,,. Computer-aided microspectrophotometry of biological specimens. Exp Cell Res 1971; 67: 81–89.
- 107,,,,. Improvements in fluorescence microscopy allowed by high power light emitting diodes. In: Méndez-VilasA, Labajos-BroncanoL, editors. Current Issues on Multidisciplinary Microscopy Research and Education, Vol. 2. Badajoz, Spain: FORMATEX; 2005. pp 181–188. Available at http://www.formatex.org/micro2003/papers/181–188. pdf.
- 108,,. Fluorescence array detector for large-field quantitative fluorescence cytometry. Cytometry 1994; 16: 206–213.Direct Link:
- 109,,,,,,,. Rapid, efficient detection and drug susceptibility testing of Mycobacterium tuberculosis in sputum by microscopic observation of broth cultures. J Clin Microbiol 2000; 38: 1203–1208.
- 110,,,,,. MODS—A user guide. 2006. Available at http://www.upch.edu.pe/facien/dbmbqf/MODS_user_guide.pdf.
- 111,,. Virulence and morphologic characteristics of mammalian tubercle bacilli. J Exp Med 1947; 86: 175–184.
- 112,. Cytochemical reaction of virulent tubercle bacilli. Am Rev Tuberc 1948; 58: 698–699.
- 113,,,,. Mycobacterium smegmatis displays the Mycobacterium tuberculosis virulence-related neutral red character when expressing the Rv0577 gene. FEMS Microbiol Lett 2004; 231: 283–289.Direct Link:
- 114,,,,,,,,,. Neutral-red reaction is related to virulence and cell wall methyl-branched lipids in Mycobacterium tuberculosis. Microbes Infect 2006; 8: 183–190.
- 115. Neutral red as a hydrophobic probe for monitoring neuronal activity. J Neurosci Methods 2000; 101: 85–92.
- 116,,,,,. The role of clinical flow cytometry in the interferon-gamma based diagnosis of active tuberculosis and its co-infection with HIV-1—A technically oriented review. Cytometry Part B Clin Cytom 2008; 74B ( Suppl): in press. doi://10.1002/cyto.b.20381 (this issue).
- 117,,,,,,,,,,,. Loss of receptor on tuberculin-reactive T-cells marks active pulmonary tuberculosis. PLoS ONE 2007; 2: e735.
