Abstract
The high incidence of Mycobacterium infection, notably multidrug-resistant M. tuberculosis infection, has become a significant public health concern worldwide. In this study, we isolate and analyze a mycobacteriophage, BTCU-1, and a foundational study was performed to evaluate the antimycobacterial activity of BTCU-1 and its cloned lytic endolysins. Using Mycobacterium smegmatis as host, a mycobacteriophage, BTCU-1, was isolated from soil in eastern Taiwan. The electron microscopy images revealed that BTCU-1 displayed morphology resembling the Siphoviridae family. In the genome of BTCU-1, two putative lytic genes, BTCU-1_ORF7 and BTCU-1_ORF8 (termed lysA and lysB, respectively), were identified, and further subcloned and expressed in Escherichia coli. When applied exogenously, both LysA and LysB were active against M. smegmatis tested. Scanning electron microscopy revealed that LysA and LysB caused a remarkable modification of the cell shape of M. smegmatis. Intracellular bactericidal activity assay showed that treatment of M. smegmatis—infected RAW 264.7 macrophages with LysA or LysB resulted in a significant reduction in the number of viable intracellular bacilli. These results indicate that the endolysins derived from BTCU-1 have antimycobacterial activity, and suggest that they are good candidates for therapeutic/disinfectant agents to control mycobacterial infections.
Highlights
Mycobacterium tuberculosis (MTB) is the leading cause of tuberculosis which is a serious public health problem that results in millions of deaths around the world each year [1]
To classify mycobacteriophage BTCU-1 into a morphotype-specific group, the phage particles were examined by TEM
The electron microscopy images revealed that BTCU-1 possessed an icosahedral head (65 nm) and a non-contractile long tail, displaying morphology resembling the Siphoviridae family (Figure 1)
Summary
Mycobacterium tuberculosis (MTB) is the leading cause of tuberculosis which is a serious public health problem that results in millions of deaths around the world each year [1]. Phage therapy is being considered as a possible therapeutic alternative for the treatment of infections caused by MDR strains [5]. Mycobacteriophages were considered as an alternative therapy for Mycobacterium infection control [9]. Mycobacteriophage-encoded lytic endolysins have considerable potential to be effective antimicrobial agents-or enzybiotics-against a number of MDR and XDR MTB strains [10,11]. We used Mycobacterium smegmatis as a rapidly growing host, and a mycobacteriophage was isolated from the soil near Buddhist Tzu Chi University and named BTCU-1. To utilize mycobacteriophage lytic endolysins as therapeutic alternatives to antibiotics, we surveyed the genomic sequence of BTCU-1 and successfully identified two lytic-associated genes. Following cloning and expression/purification, various antimycobacterial activities of these two lytic proteins were determined in vitro
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