3. BBL™ MGIT™ Tubes (4mL ; 7 mL) Modified Middlebrook 7H9 Broth Base Fluorescent indicator untukdeteksioxygen Fluorochrom, tris4,7-difenil-1, 10-phenonthroline ruteniumpentahydrateklorida yang ditanamdalamsilikonpadabagianbawahtabung Di produksidengan 10% CO2atmosfer.
4. MGIT™ Growth Supplement Sebagaisuplemenpertumbuhanmikobakterium Oleic Acid – pentinguntukmetabolismemikobakterium Albumin (bovine) – mengikatfree fatty acids yang dapatmeracunimikobakterium Dextrose – sumberenergiuntukmikobakterium Catalase– menghancurkanperoksida Polyoxyethylenestearate (POES) -- meningkatkanpertumbuhan of M. tuberculosisdanmembantuuniforminokulasi 4
5. BBL™ MGIT™ PANTA ™ Antimicrobial mixture: Polymyxin B Amphotericin B Naladixic Acid Trimethoprim Azlocillin MENEKAN pertumbuhannon-mikobakterium 5
6. O2 O2 O2 Bagianatas O2 CO2 O2 O2 O2 O2 O2 O2 CO2 Broth O2 CO2 O2 O2 CO2 O2 FO2 FO2 F F FO2 F F FO2 FO2 F F F Sensor F FO2 FO2 F F F FO2 FO2 Fluoresensikuat SedikitatauTidakadaFluoresensi SistemDeteksi MGIT KulturNegatif KulturPositif 6
58. SistemDeteksi BACTECTM MGITTM 960 Detektoradadisetiapbagianbawahrak: Data dikumpulkandarisetiaptabung, setiap jam sekali. Data dievaluasidantube positivityditentukanberdasarkansoftware algorithms.
59. BACTEC™ MGIT™ 960 System Di desainuntukmemenuhikebutuhanpemeriksaanberjumlahbanyak. Space-saving 960 tube capacity. Satuinstrumenbisamemproseshingga 8000 specimen per Tahun. LamanyaProtokoluntukpertumbuhanmulai 1 sampai 56 hari. Lama protokolstandar: 42 days 24
74. BACTEC™ MGIT™ 960 Pilihantessensitivitasantibiotikaterdiridari: Streptomycin, Isoniazid, Rifampin and Ethambutol (SIRE) testing against M. tuberculosis SIRE Kit to perform AST at the Critical Concentrations Separate kits are available to perform a 2nd test with higher concentrations of S and I if needed PZA testing against M. tuberculosis 36
76. 2. Scan AST set into instrument 1. Prepare Inoculum and inoculate SIRE tubes 3. When complete, scan out the set 4. Print the result ProsedurAST Instrument PZA test is performed in the same way, using just the 2-tube AST Set Carrier 38
77. Kelebihan BACTEC™ MGIT™ 960 System Non-radiometric. Menggunakan MGIT media and sensor yang menetap. EffisiendenganTeknologifluorometric Akurat: deteksipertumbuhanmelaluideteksipenggunaan O2olehMycobacteria. Otomatis Quality Control Untukmemastikanalatberjalansecarapresisidanlayak. 39
96. M. avium* M. abscessus M. bovis M. celatum M. fortuitum* M. gastri M. gordonae* M. haemophilum** M. intracellulare M. kansasii* M. malmoense M. marinum M. nonchromogenicum M. phlei M. simiae* M. scrofulaceum M. smegmatis M. szulgai* M. terrae M. trivale M. tuberculosis* M. xenopi* Species Detected as Positive in the BACTEC™ MGIT™ 960 System * Species recovered in clinical evaluation ** With addition of hemin 45
97. Isolation of Mycobacteria BBL™ MGIT™ Tubes (7 mL) Modified Middlebrook 7H9 Broth Base Fluorescent indicator untuk oxygen deteksi Di produksi dengan 10% CO2 atmosphere.
98. Isolation of Mycobacteria MGIT™ 960 Growth Supplement Sebagai supplemen pertumbuhan mycobacterial Oleic Acid – penting untuk metabolisma mycobacterial Albumin (bovine) – mengikat free fatty acids yang dapat meracuni mycobacteria Dextrose – sumber energi untuk mycobacteria Catalase– menghancurkan peroxida Polyoxyethylene stearate (POES) -- meningkatkan pertumbuhan of M. tuberculosis dan membantu uniform inokulasi 47
99. Isolation of Mycobacteria BBL™ MGIT™ PANTA ™ Antimicrobial mixture: Polymyxin B Amphotericin B Naladixic Acid Trimethoprim Azlocillin MENEKAN pertumbuhan non-mycobacterial 48
100. Specimen Collection & Handling Collect aseptically. If transport delayed > 1 hour, refrigerate specimen to minimize growth of resident flora. Refrigerate specimens in laboratory until processing can begin. 49
101. Specimen Processing Process specimens using NALC/NaOH method as recommended in CDC Manual: Public Health Mycobacteriology: A Guide for the Level III Laboratory. BBL™ MycoPrep™ may be used. 50
102. Specimen PreparationNALC/NaOH Digestion Procedure - Sputum Add the specimen to a 50 mL plastic centrifuge tube. The specimen should not exceed 10 mL. Add NALC / NaOH solution in volume equal to that of the specimen. Invert the tube to ensure that the NALC solution contacts all surfaces of tube and cap. Tighten cap securely and vortex tube for 15 - 20 seconds. Do not overmix - inactivation of NALC may occur. Allow specimen to stand for 15 – 20 minutes at room temperature For thick specimens, vortex occasionally. 51
103. Specimen PreparationNALC/NaOH Digestion Procedure - Sputum Fill tube to 50 mL mark with sterile phosphate buffer, pH 6.8. Swirl gently by hand to mix. Concentrate by centrifugation at 3,000 x g for 15 minutes. Insure that centrifuge tubes can withstand these high RCF’s. Carefully decant and discard supernatant fluid from pellet. Resuspend pellet with 1 - 3 mL sterile phosphate buffer. 52
104. MGIT™ Tube Preparation Prepare tubes while specimens are in centrifuge: Label MGIT™ tube with specimen number. Prepare PANTA. Reconstitute with 15 mL Growth Supplement For best results, add PANTA / Growth Supplement solution just prior to specimen inoculation. 53
105. MGIT™ Tube Preparation Unscrew cap and aseptically add 0.8 mL MGIT™ PANTA / Growth Supplement solution. 54
106.
107. MGIT™ Tube Preparation Tightly recap tube and mix well. Wipe tubes and caps with tuberculocidal disinfectant. Enter MGIT tubes into MGIT 960 Instrument. 56
108. KulturPositif Kontaminasi: Microorganisms other than AFB can grow and be detected as positive. May also contain AFB. Contaminated tubes may be re-processed and inoculated into fresh MGIT™ tube. Refer to User’s Manual for details. 57
110. BACTEC™ MGIT™ 960 AST Simple workflow Instrument knows from the barcode which AST set is being entered (i.e., 2 tube vs. 8 tube) Instrument automatically directs placement of AST set into instrument 59
111. BACTEC™ MGIT™ 960 AST Capacity and Flexibility Designed to meet the needs of laboratories performing mycobacteria growth and detection as well as susceptibility testing 5 AST tests (SIRE and PZA) per week will utilize approximately one third of a drawer Allows flexibility in the definition of an “AST set” AST sets can consist of 2, 3, 4, 5 or 8 tubes, one of which is the Growth Control tube 60
112. BACTEC™ MGIT™ 960 SIRETesting Options SIRE Susceptibility test options: SIRE Kit to perform AST at the critical concentrations Separate kits are available to perform a “2nd Tier” test with higher concentrations of S, I, and E. Recommended if the organism shows resistance to the critical concentration of the drug Testing can be done in any combination of drugs and concentrations 61
113. ProsedurAST Inoculation Add appropriate Supplement (SIRE or PZA) and drugs to appropriate tubes in AST Set Prepare standardized inoculum Positive MGIT tube: Use direct if Day 1 – 2 or dilute if Day 3 – 5 Isolate on solid media : Prepare 0.5 McFarland Inoculate drug tubes Prepare dilution and inoculate Growth Control tube Place AST set into instrument 62
114. Summary of Clinical Trial Study: BACTEC™ MGIT™ 960 System: False Positive rate* = 0.8% False Negative rate = 0.5% Average Breakthrough Contamination: 8.1% range = 1.8% - 14.6% *Based on the total number of specimens 63
116. PROTOCOL LENGTH OF MGIT Vs LJ CONVENTIONAL METHOD (DAYS) LJ METHOD Average (+) 29.6 4.4 Normal range(+) 28-56 4 – 14 Negatives 56 – 63 42 – 56 MGIT 65
117. RESULTS OF MGIT Vs. LJ FROM OTHER RESEARCHERS CONVENTIONAL BACTEC STUDY CULTURE SYSTEM Van Griethiuysen et al,1996 79.9% 95.9% Zanetti et al, 1997 69.3 % 95.0 % Jesus et al , 2001 46.6% 86.6% Lu et al , 2002 45.5 % 71.1 % * Recovery rates from positive samples 66
118.
119. Zannetti et al (1997) showed no significant difference between the radiometric BACTEC 460 TB and the fluorometric BACTEC 960 thus 91.9% positivity and 95.1% positivity respectively. But use of radioisotopically labeled substrates has inhibiteduniversal applicationof the radiometric BACTEC 460 TB system.67
120.
121. In poor economies MGIT is relatively expensive to acquire and sustain.68
