International Journal of Medical and Dental Sciences

Open Access Open Access  Restricted Access Subscription Access

Effect of Relative Centrifugal Forces and Centrifugation Times on Recovery of Mycobacterium Tuberculosis


Affiliations
1 Department of Microbiology, M.M.I.M.S.R., MM University, Mullana, Ambala-133207, India
 

Background: Tuberculosis (TB) is one of the major cause of morbidity and mortality worldwide, affecting one third of world's population and the incidence is much higher in South-East Asia (India and China together account for nearly 40% of the global TB cases)

Objectives: The current study was designed to evaluate the efficacy of relative centrifugal forces (RCFs) and various centrifugations times (CTs) on recovery of Mycobacterium tuberculosis (MTB) from sputum specimens pre-inoculated with MTB.

Materials & Methods: For this study, direct acid fast bacilli (AFB) smear negative sputum specimens were randomly collected, which were further subjected to sterilization ( by autoclaving at 121°C for 15 min ) and then seeded with MTB. Afterwards, these sputum specimens were subjected to centrifugation (in a cold centrifuge) at various RCFs and CTs. After centrifugation, supernatants and sediments were inoculated on LJ media and incubated at 37°C with daily monitoring for 8 weeks.

Results: In the current study, as the RCFs (6000xg) and CTs (25 min and 20 min) were increased, the growth detection time exponentially decreased (9 and 11 days) and culture grades (4+) were increased when cultured from the sediments. However, the recovery rates of MTB from the supernatant fluids were constantly decreased as RCFs and CTs increased.

Conclusion: The current study concluded that a higher RCF of 6000xg and CTs of 20 and 25 min could effectively detect the tubercle bacilli from the sputum specimens more efficiently than other lower RCFs and CTs.


Keywords

Pulmonary Tuberculosis, Mycobacterium Tuberculosis, Acid Fast Bacilli, Relative Centrifugal Force, Centrifugation Time.
User
Notifications
Font Size


  • Global tuberculosis report 2012; WHO [Internet]. Available at: www.who.int/tb/ publications/global report/gtbr12_main.pdf

  • Saceanu CA, Pfeiffer NC, McLean T. Evaluation of sputum smears concentrated by cytocentrifugation for detection of acidfast bacilli. J Clin Microbiol 1993;31(9): 2371-4.

  • WHO - World Health Organization 2012. Tuberculosis. [Internet] Available at: who.int/mediacentre/factsheets/fs104/ en/ index.html

  • Shinu P, Nair A, Singh V, Kumar S, Bareja R. Evaluation of rapid techniques for the detection of mycobacteria in sputum with scanty bacilli or clinically evident, smear negative cases of pulmonary and extrapulmonary tuberculosis. Mem Inst Oswaldo Cruz 2011;106(5):620-4.

  • Biotec Laboratories 2000a. Biotec FASTPlaque TB™ 2000. A rapid bacteriophage assay for detection of Mycobacterium complex in clinical samples. Instruction Manual, Biotec, Ipswich;2000.p. 15.

  • Biotec Laboratories 2000b. Biotec FASTPlaque TB™ 2000. An innovative breakthrough for the detection of tuberculosis. Technical data report, Biotec, Ipswich; 2000.p.9.

  • Ratnam S, March SB. Effect of relative centrifugal force and centrifugation time on sedimentation of mycobacteria in clinical specimens. J Clin Microbiol 1986;23(3):5825.

  • Ganoza CA, Ricaldi JN, Chauca J, Rojas G, Munayco C, Agapito J et al. Novel hypertonic saline-sodium hydroxide (HS-SH) method for decontamination and concentration of sputum samples for MTB microscopy and culture. J Med Microbiol 2008;57:1094-8.

  • Chakravorty S, Tyagi JS. Novel multipurpose methodology for detection of Mycobacteria in pulmonary and extrapulmonary specimens by smear microscopy, culture and PCR. J Clin Microbiol 2005;43:2697-702.

  • Carricajo A, Fonsale N, Vautrin AC, Aubert G. Evaluation of BacT/Alert 3D Liquid Culture System for Recovery of Mycobacteria from Clinical Specimens Using Sodium Dodecyl (Lauryl) Sulfate-NaOH Decontamination. J Clin Microbiol 2001;39: 3799.

  • Thornton CG, MacLellan KM, Brink TL Jr, Passen S. In vitro comparison of NALCNaOH, tween 80, and C18- carboxypropylbetaine for processing of specimens for recovery of mycobacteria. J Clin Microbiol 1998;36:3558-66.

  • Farnia P, Mohammadi F, Zarifi Z, Tabatabee DJ, Ganavi J, Ghazisaeedi K et al. Improving sensitivity of direct microscopy for detection of acid-fast bacilli in sputum: use of chitin in mucus digestion. J Clin Microbiol 2002;40(2):508-11.

  • Nadia Boujtita, Concentration of Mycobacteria in Clinical Samples using the Thermo Scientific General Purpose Centrifuge with 8x50 mL Individually Sealed Rotor [Internet] Available at: https:// static.thermoscientific.com/images/D01739

  • Rickman TW, Moyer NP. Increased sensitivity of acid-fast smears. J Clin Microbiol 1980;11(6):618-20.

  • Selvakumar N, Govindan D, Chandu NA, Frieden TR, Narayanan PR. Processing sputum specimens in a refrigerated centrifuge does not increase the rate of isolation of Mycobacterium tuberculosis. J Clin Microbiol 2003;41(1):469-71.


Abstract Views: 282

PDF Views: 168




  • Effect of Relative Centrifugal Forces and Centrifugation Times on Recovery of Mycobacterium Tuberculosis

Abstract Views: 282  |  PDF Views: 168

Authors

Amit Sharma
Department of Microbiology, M.M.I.M.S.R., MM University, Mullana, Ambala-133207, India
Isha Madaan
Department of Microbiology, M.M.I.M.S.R., MM University, Mullana, Ambala-133207, India
Pottathil Shinu
Department of Microbiology, M.M.I.M.S.R., MM University, Mullana, Ambala-133207, India
Varsha A. Singh
Department of Microbiology, M.M.I.M.S.R., MM University, Mullana, Ambala-133207, India
Anu Garg
Department of Microbiology, M.M.I.M.S.R., MM University, Mullana, Ambala-133207, India

Abstract


Background: Tuberculosis (TB) is one of the major cause of morbidity and mortality worldwide, affecting one third of world's population and the incidence is much higher in South-East Asia (India and China together account for nearly 40% of the global TB cases)

Objectives: The current study was designed to evaluate the efficacy of relative centrifugal forces (RCFs) and various centrifugations times (CTs) on recovery of Mycobacterium tuberculosis (MTB) from sputum specimens pre-inoculated with MTB.

Materials & Methods: For this study, direct acid fast bacilli (AFB) smear negative sputum specimens were randomly collected, which were further subjected to sterilization ( by autoclaving at 121°C for 15 min ) and then seeded with MTB. Afterwards, these sputum specimens were subjected to centrifugation (in a cold centrifuge) at various RCFs and CTs. After centrifugation, supernatants and sediments were inoculated on LJ media and incubated at 37°C with daily monitoring for 8 weeks.

Results: In the current study, as the RCFs (6000xg) and CTs (25 min and 20 min) were increased, the growth detection time exponentially decreased (9 and 11 days) and culture grades (4+) were increased when cultured from the sediments. However, the recovery rates of MTB from the supernatant fluids were constantly decreased as RCFs and CTs increased.

Conclusion: The current study concluded that a higher RCF of 6000xg and CTs of 20 and 25 min could effectively detect the tubercle bacilli from the sputum specimens more efficiently than other lower RCFs and CTs.


Keywords


Pulmonary Tuberculosis, Mycobacterium Tuberculosis, Acid Fast Bacilli, Relative Centrifugal Force, Centrifugation Time.

References





DOI: https://doi.org/10.18311/ijmds%2F2014%2F80643