Isolation and Identification of Bioactive Compounds from Streptomyces actinomycinicus PJ85 and Their In Vitro Antimicrobial Activities against Methicillin-Resistant Staphylococcus aureus

Abstract

Antibiotic-resistant strains are a global health-threatening problem. Drug-resistant microbes have compromised the control of infectious diseases. Therefore, the search for a novel class of antibiotic drugs is necessary. Streptomycetes have been described as the richest source of bioactive compounds, including antibiotics. This study was aimed to characterize the antibacterial compounds of Streptomyces sp. PJ85 isolated from dry dipterocarp forest soil in Northeast Thailand. The 16S rRNA gene sequence and phylogenetic analysis showed that PJ85 possessed a high similarity to Streptomyces actinomycinicus RCU-197^T of 98.90%. The PJ85 strain was shown to produce antibacterial compounds that were active against Gram-positive bacteria including methicillin-resistant Staphylococcus aureus (MRSA). The active compounds of PJ85 were extracted and purified using silica gel column chromatography. Two active antibacterial compounds, compound 1 and compound PJ85_F39, were purified and characterized with spectroscopy, including liquid chromatography and mass spectrometry (LC-MS). Compound 1 was identified as actinomycin D, and compound PJ85_F39 was identified as dihomo-γ-linolenic acid (DGLA). To the best of our knowledge, this is the first report of the purification and characterization of the antibacterial compounds of S. actinomycinicus.

Keywords: Streptomyces actinomycinicus; Streptomyces sp.; antibacterial activity; dihomo-γ-linolenic acid; drug-resistant microorganisms.

Figure 1

Neighbor-joining phylogenetic tree based on relative 16S rRNA gene sequences showing the phylogenetic relationships among Streptomyces sp. PJ85 and its closest strains. Numbers at nodes indicate bootstrap percentages (1000 replicates), and only values greater than 50% are shown at the nodes. The scale bar indicates 0.01 substitutions per nucleotide position.

Figure 2

Effect of incubation temperature on the growth of Streptomyces sp. PJ85 cultured in ISP-2 medium. Incubation temperatures: (□) 30 °C and (●) 37 °C. Error bars represent the standard deviation of the mean (n = 3). The statistical significance of the differences between the cell biomass of PJ85 cultured at 30 °C and 37 °C was estimated using a two-way ANOVA with a Bonferroni multiple comparison test, ** p < 0.0001, compared on the same incubation day.

Figure 3

Effect of incubation temperature on the activity of antibacterial agents of Streptomyces sp. PJ85 cultured in ISP-2 medium against (A) MRSA DMST20651, (B) S. aureus ATCC29213, (C) S. epidermidis TISTR518, (D) B. subtilis TISTR008, and (E) B. cereus TISTR687. Incubation temperatures: (□) 30 °C and (●) 37 °C. Error bars represent the standard deviation of the mean (n = 3). The statistical significance of the differences between antibacterial activity of the crude extract of PJ85 cultured at 30 °C and 37 °C was determined using a two-way ANOVA with a Bonferroni multiple comparison test, ** p < 0.0001, compared on the same incubation day.

Figure 4

TLC-based bioautography of the ethyl acetate crude extract of PJ85 exhibiting antibacterial activity against MRSA DMST20651.

Figure 5

Liquid chromatography–mass spectrometry (LC–MS) analysis of compound PJ85_F39 in positive ion mode. MS spectrum showing the ion clusters for [M + H]⁺ at m/z 307.2171 correlated to dihomo-γ-linolenic acid (DGLA).