Effect of time duration of digestion/decontamination technique on yield of mycobacteria and contamination rates from sterile body fluids (pleural and ascitic fluid) and pus specimens

Abstract

Background: Duration of digestion/decontamination has a considerable impact on yield of mycobacteria especially from sterile body fluids and pus specimens. Additionally, duration of digestion/decontamination affects the contamination rates. This study evaluates the effect of digestion/decontamination protocol for 15 and 20min versus inoculation of media directly from the sample on contamination rates as well as the yield of mycobacteria from pus and sterile fluids other than cerebrospinal fluids. Methods: Pleural fluid (n = 60), pus (n = 48) and ascitic fluid (n = 12) specimens were cultured for mycobacteria and evaluated for contamination and mycobacterial yield using three different processing methodologies: without digestion/decontamination with 5% NaOH-NALC (D/D), D/D for 15min and D/D for 20min. All samples >3 mL in volume were spun at 3000 RCF for 15min, whereas those less than 3 mL were used as is. They were simultaneously processed using the three different methods as mentioned above, and inoculated on LJ media and MGIT. Smear was made from samples treated for 20min and stained with fluorescent stain. Kinyoun staining was done on smears with dubious findings. Mycobacterial culture yield and contamination rates were recorded at 6 weeks as recommended by the Global Laboratory Initiative (GLI) laboratory manual 2014. Results: Pleural fluid and pus contamination rates were substantially lowered by increasing decontamination time from 15 to 20min, but it did not have any effect for ascitic fluid (Table 1). The 5-min difference in the decontamination procedure improved mycobacterial culture yield for pus samples by 10%, but there was no substantial effect on pleural and ascitic fluids. Prolonged decontamination did not compromise the culture yield in any of the mentioned specimens. Conclusion: In areas where specimen delay is common and sterility of collection procedure cannot be ensured, digestion/decontamination with NaOH-NALC for up to 20min can reduce contamination rates without considerably compromising mycobacterial culture yield. However, one should be alert to the possibility of decreased viability, and culture should be supplemented with molecular methods.

Keywords

• Digestion/decontamination
• TB culture contamination
• TB culture yield