Something from nothing: Sensitivity and specificity of Xpert MTB/RIF Ultra on contaminated liquid cultures for tuberculosis and rifampicin-resistance detection
Authors: Ghebrekristos YT , Beylis N , Centner CM , Venter R , Derendinger B , Tshivhula H , Naidoo S , Alberts R , Prins B , Tokota A , Dolby T , Marx FM , Omar SV , Warren R , Theron G 
Published: December 2022

AbstractBackgroundXpert MTB/RIF Ultra (Ultra) (Cepheid, Sunnyvale, USA) is a widely-used rapid front-line TB and rifampicin susceptibility test. Mycobacterium Growth Indicator Tube 960 (MGIT960) culture is still used as an adjunct for tuberculosis (TB) and drug susceptibility diagnosis but is vulnerable to contamination. Whether Ultra can be used on to-be-discarded contaminated cultures is uninvestigated.MethodsWe stored contaminated MGIT960 tubes (growth-positive, acid-fast-bacilli-negative) inoculated to diagnose pulmonary TB in a routine high-volume laboratory in Cape Town, South Africa. Patients who had, at contamination-detection, no positive TB results (smear, Ultra, culture) and another specimen submitted three months post-contaminated specimen submission were selected. We evaluated the sensitivity and specificity of Ultra on contaminated growth from the first culture for 1) TB (next-available non-contaminated culture result reference standard), and 2) rifampicin resistance (vs. MTBDRpluson the later isolate). We calculated potential time-to-diagnosis improvements. We also evaluated MPT64 TBc (TBc).Findings2186 patients had a contaminated diagnostic culture. 49% (1068/2186) had no other specimen submitted, despite guidance to the contrary. After 319 ineligible patients were excluded, 799 with at least one repeat specimen submitted remained: 31% (n=246), 54% (n=429) and 16% (n=124) were repeat-specimen culture-positive, -negative, and -contaminated, respectively. When Ultra was done on the initial contaminated growth, sensitivity and specificity were 89% (95% CI 84-94) and 95% (90-98) for TB and 95% (75-100) and 98% (93-100) for rifampicin-resistance. If our approach were performed the day after initial contamination detection, time-to-TB-detection would improve a median (IQR) of 23 (13-45) days and, importantly, provide a result in many patients who had none. TBc had poor accuracy.ConclusionUltra on acid-fast-negative growth from contaminated MGIT960 tubes had high sensitivity and specificity; approximating World Health Organization-target product performance sputum test and exceeding drug susceptibility testing (DST) criteria. Our approach could mitigate contamination’s negative effects, especially when repeat specimens are not submitted.Research in contextEvidence before this studyImproving the diagnosis of tuberculosis (TB) and drug-resistance through strengthening the laboratory care cascade is a public health priority. Scale-up of molecular tests like Xpert MTB/RIF Ultra (Ultra) (Cepheid, Sunnyvale, USA), for the upfront diagnosis of TB and rifampicin-resistance has doubtlessly improved the care cascade, however, culture, despite several limitations, continues to be used for the diagnosis and susceptibility testing for technical, historic, and cost reasons (the most common TB culture platform is the MGIT960 liquid culture system). The fact that global TB diagnosis still, in part, relies on culture means that culture-contamination, which represents a failed attempt at testing, worsens care cascade gaps. Contamination requires another specimen to be collected from patients, however, this causes delays or complete care cascade drop out of patients. Contaminated cultures are traditionally checked with microscopy to see if they contain acid-fast bacilli (AFB), however, the use of Ultra on contaminated cultures, especially those who are AFB-negative, is unexplored. If performance is high, the negative impact of culture-contamination, which is frequent in many settings, could be drastically mitigated as Ultra is widely-available.Added value of this studyWe showed that Ultra on to-be discarded contaminated MGIT960 cultures can detect TB in a highly sensitive and specific manner (89% sensitivity, 95% specificity). It also had excellent sensitivity and specificity for rifampicin resistance (95% sensitivity, 98% specificity). Performance levels exceeded those accepted by the World Health Organization for Ultra done directly on respiratory specimens. In patients who, after initial culture contamination had another specimen submitted for culture, our approach could reduce time to diagnosis by approximately 23 days. Critically, many patients with contamination had, despite programmatic guidance, no record of a further attempt to diagnose TB (44%), and in these patients our Ultra on contaminated cultures approach would result in an accurate TB and rifampicin-resistance result where none would ordinarily occur.Implications of all available evidenceWhen done on contaminated MGIT960 culture growth resulting from a failed attempt to diagnose TB, Ultra has excellent performance for TB and rifampicin-resistance detection and would likely reduce the impact of culture-contamination on the diagnostic care cascade. Laboratories should consider evaluating and potentially implementing this approach wherever TB culture is done for diagnostic purposes.