A novel lytic phage potentially effective for phage therapy against Burkholderia pseudomallei in the tropics

Abstract

Background Burkholderia pseudomallei is a tropical pathogen that causes melioidosis. Its intrinsic drug-resistance is a leading cause of treatment failure, and the few available antibiotics require prolonged use to be effective. This study aimed to assess the clinical potential of B. pseudomallei phages isolated from Hainan, China. Methods Burkholderia pseudomallei strain (HNBP001) was used as the isolation host, and phages were recovered from domestic environmental sources, which were submitted to the host range determination, lytic property assays, and stability tests. The best candidate was examined via the transmission electron microscope for classification. With its genome sequenced and analyzed, its protective efficacy against B. pseudomallei infection in A549 cells and Caenorhabditis elegans was evaluated, in which cell viability and survival rates were compared using the one-way ANOVA method and the log-rank test. Results A phage able to lyse 24/25 clinical isolates was recovered. It was classified in the Podoviridae family and was found to be amenable to propagation. Under the optimal multiplicity of infection (MOI) of 0.1, an eclipse period of around 20 min and a high titer (1012 PFU/ml) produced within 1 h were demonstrated. This phage was found stabile at a wide range of temperatures (24, 37, 40, 50, and 60 °C) and pH values (3–12). After being designated as vB_BpP_HN01, it was fully sequenced, and the 71,398 bp linear genome, containing 93 open reading frames and a tRNA-Asn, displayed a low sequence similarity with known viruses. Additionally, protective effects of applications of vB_BpP_HN01 (MOI = 0.1 and MOI = 1) alone or in combination with antibiotics were found to improve viability of infected cells (70.6 ± 6.8%, 85.8 ± 5.7%, 91.9 ± 1.8%, and 96.8 ± 1.8%, respectively). A significantly reduced mortality (10%) and a decreased pathogen load were demonstrated in infected C. elegans following the addition of this phage. Conclusions As the first B. pseudomallei phage was isolated in Hainan, China, phage vB_BpP_HN01 was characterized by promising lytic property, stability, and efficiency of bacterial elimination during the in vitro/vivo experiments. Therefore, we can conclude that it is a potential alternative agent for combating melioidosis. Graphical Abstract

Effective Therapeutic Options for Melioidosis: Antibiotics versus Phage Therapy

“…In this article, we discuss six phages with different B. pseudomallei strains (P37, K96243, E0237, CMS, 365A, and HNBP001) as hosts ( Table 2 ) [ 101 , 103 , 104 , 105 , 106 , 107 ]. Based on their morphological characteristics, four of the phages had contractile tails and were classified into the Myoviridae family [ 103 , 104 , 105 , 106 ].…”

“…Based on their morphological characteristics, four of the phages had contractile tails and were classified into the Myoviridae family [ 103 , 104 , 105 , 106 ]. Two of the phages had short tails and were classified as the Podoviridae family [ 101 , 107 ]. In the cases of staphylophages, Myoviridae phages were more common than Podoviridae phages in commercial preparations such as therapeutic cocktails [ 108 ].…”

“…Some phages were able to kill both biofilm-forming and non-biofilm-forming B. pseudomallei [ 106 , 109 ]. The efficiency of B. pseudomallei phages has been evaluated in vitro with A549 cells and in vivo with BALB/c mice and C. elegans , resulting in the rescue of infected organisms and reduction in bacterial burden [ 105 , 107 ]. The therapeutic efficiency can be further increased by introducing a combination therapy of phages and antibiotics [ 107 ].…”

“…Phage vB_BpP_HN01, the first B. pseudomallei phage found in Hainan, China, was among over 20 phages isolated from samples obtained near households afflicted by a melioidosis outbreak using the representative Hainan clinical strain HNBP001 [ 107 ]. Based on its icosahedral head size of 62.0 ± 1.3 nm and a short tail length of 20.4 ± 0.7 nm, the phage was assigned to the Podoviridae family.…”

“…New phages against B. pseudomallei can be discovered in days or weeks as they are easily isolated from the environment, especially water and soil. Several phages were found to be effective against B. pseudomallei , including ST79 and ST96 [ 103 ], ΦBp-AMP1 [ 101 ], φX216 [ 104 ], C34 [ 105 ], 365A [ 106 ], and vB_BpP_HN01 [ 107 ]. However, PT for melioidosis is currently being reviewed in clinical trials and has not yet been licensed for public use.…”