Detection of Mycobacterium tuberculosis complex DNA in oronasal swabs from infected African buffaloes (Syncerus caffer)

Abstract

Mycobacterium bovis (M. bovis), a member of the Mycobacterium tuberculosis complex (MTBC), is the causative agent of bovine TB (bTB) in animals. Spread occurs through inhalation or ingestion of bacilli transmitted from infected individuals. Early and accurate detection of infected African buffaloes shedding M. bovis is essential for interrupting transmission. In this pilot study, we determined if MTBC DNA could be detected in M. bovis infected buffalo oronasal secretions using a molecular transport media (PrimeStore MTM) with oronasal swabs and a rapid qPCR assay (Xpert MTB/RIF Ultra). Bovine TB test-positive buffaloes were culled, then tissue samples and oronasal swabs collected post-mortem for mycobacterial culture and Ultra testing, respectively. The Ultra detected MTBC DNA in 5/12 swabs from M. bovis culture-confirmed buffaloes. Oronasal swabs from M. bovis negative buffaloes (n = 20) were negative on Ultra, indicating the high specificity of this test. This study showed that MTM can successfully preserve MTBC DNA in oronasal swabs. The proportion of MTBC positive oronasal swabs was higher than expected and suggests that the Ultra may be an additional method for identifying infected buffaloes. Further studies are needed to confirm the utility of the Ultra assay with oronasal swabs as an assay to evaluate possible MTBC shedding in buffaloes.

Identification and molecular characterization of Mycobacterium bovis DNA in GeneXpert® MTB/RIF ultra-positive, culture-negative sputum from a rural community in South Africa

“…Through spoligotyping, we further identified two M. bovis strains (SB 0130 and SB 1474 ) in 4/10 M. bovis DNA positive samples ( Table 2 , Table 3 ) [ [45] , [46] , [47] ]. These results further emphasized the constraints of relying solely on traditional mycobacterial culture and stress the significance of integrating multiple diverse molecular techniques with different sensitivities and suitable clinical specimens for a thorough comprehension of zoonotic TB transmission [ 13 , 18 , 20 , 21 , 48 ].…”

“…Despite available health data, the absence of host cell-mediated immunological responses, such as those assessed using the QuantiFERON TB Gold Plus test, complicates the assessment of true infection status [ 4 , 49 ]. Heat-inactivation limits all culture methods [ 48 ]. The limited sample size prohibits a comprehensive investigation into M. bovis strain diversity.…”

Detection of Mycobacterium bovis in nasal swabs from communal goats (Capra hircus) in rural KwaZulu-Natal, South Africa

“…Furthermore, a positive Ultra result does not differentiate between dead or non-viable mycobacteria, or residual DNA within the sample, as has been observed in human patients ( Miotto et al, 2012 ; Theron et al, 2018 ). Despite this, the Ultra’s diagnostic appeal, for use on animal specimens suspected of having MTBC infections, is increasing ( Chakravorty et al, 2017 ), and has provided rapid detection of MTBC DNA in tissue and respiratory samples collected from infected African buffaloes ( Clarke et al, 2021 , 2022a ), African elephants ( Goosen et al, 2020 ), and white rhinoceros ( Goosen et al, 2020 ). The positive Ultra results in the 19 communal goats were likely true positives, particularly since they share grazing and water sources with M. bovis infected cattle ( Sichewo et al, 2020 ).…”

“…Recently, culture-independent PCR-based sequencing methods have been investigated for direct detection and identification of important mycobacterial organisms in antemortem respiratory samples and postmortem tissues ( Clarke et al, 2022a ; Goosen et al, 2022a ). Studies have shown that the GeneXpert® MTB/RIF Ultra assay (Ultra) can detect MTBC DNA in animal samples and provides a rapid sensitive screening test ( Goosen et al, 2020 ; Clarke et al, 2021 ).…”

“…Studies have shown that the GeneXpert® MTB/RIF Ultra assay (Ultra) can detect MTBC DNA in animal samples and provides a rapid sensitive screening test ( Goosen et al, 2020 ; Clarke et al, 2021 ). Furthermore, conventional, and real-time PCR assays, followed by amplicon sequencing, have also shown promise for detecting and characterizing both MTBC and NTM species present in cultures, as well as directly from raw specimens ( Warren et al, 2006 ; Deggim-Messmer et al, 2016 ; Jung et al, 2016 ; Goosen et al, 2020 , 2022a ; Clarke et al, 2022a , b ). Amplicons from these PCRs can be used for targeted next generation sequencing, which facilitates the accurate detection and characterization of multiple mycobacterial species present, especially in paucibacillary oronasal swabs, other respiratory samples, as well as fecal samples ( Adékambi et al, 2003 ; Lee et al, 2020 ; Goosen et al, 2022a ).…”