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Anti-methicillin-resistant Staphylococcus aureus (MRSA) substance from the marine bacterium Pseudomonas sp. UJ-6

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Abstract

A multivalent approach to discover a novel antibiotic substance against methicillin-resistant Staphylococcus aureus (MRSA), a marine bacterium, UJ-6, exhibiting an antibacterial activity against MRSA was isolated from seawater. The isolated strain was identified to be Pseudomonas sp. by the morphology, biochemical, and genetical analyses. The ethyl acetate extract of Pseudomonas sp. UJ-6 culture showed significant ant-MRSA activity. Bioassay-guided isolation of the extract using a growth inhibitory assay led to the isolation and identification of an active compound exhibiting anti-MRSA activity. Based on the analyses of the physicochemical and spectroscopic data including nuclear magnetic resonance and mass, the compound was identified to be 1-acetyl-beta-carboline. The minimum inhibitory concentration (MIC) of the compound was determined to be in a range of 32–128 μg/ml against MRSA strains. The MIC values against MRSA were superior or equal to those of other natural compounds such as catechins, suggesting that 1-acetyl-beta-carboline would be a good candidate in applications of the treatment of MRSA infection.

Introduction

The resistance of bacteria to clinically important antibiotics is a major factor throughout the world. According to other researchers, since methicillin and other similar penicillins are used for long periods of time, the incidence and prevalence of penicillin and other antibiotics resistance in Staphylococcus aureus has increased (Kaplan and Mason, 1998, Eom et al., in press). The first antibiotic such as penicillin was effective in infectious diseases, but the increasing use of antibiotic has become a problem of resistance of S. aureus (Finland, 1979). Since methicillin-resistance S. aureus (MRSA) were first reported in England (Jevons, 1961), MRSA is recently a serious problem because it exhibits multidrug resistant to almost all commercial antibiotics except vancomycin and teicoplanin (Isnansetyo and Kamei, 2003, Witte, 1999). Vancomycin has been used the only effective antibiotics for the treatment of MRSA infections. However, vancomycin-intermediate and -resistant S. aureus recently have been reported in several countries as the application of vancomycin has become more widespread. As an alternative to vancomycin for treatment of S. aureus infection, the new antimicrobial agents such as linezolid, quinupristin/dalfopristin, daptomycin, tigecycline, and fidaxomicin are being used for the most severe infections (Bush, 2011). Therefore, the development of new and potent antimicrobial agents against MRSA is required (Eom et al., 2011b, Hanaki et al., 1998, Hiramatsu et al., 1997, Lee et al., 2008, Witte, 1999).

It has been believed that marine organisms possess novel metabolites with unique structural features compared to metabolites produced by terrestrial organisms (Cha et al., 2011, Eom et al., 2011a, Larsen et al., 2005). Although the chemical compounds of marine microorganisms are less well known than those of their terrestrial counterparts, several novel antibiotics from marine microorganisms have been reported, including loloatins from Bacillus (Gerard et al., 1999), agrochelin and sesbanimides from Agrobacterium (Acebal et al., 1998, Acebal et al., 1999), pelagiomicins from Pelagiobacter variabilis (Imamura et al., 1997), δ-indomycinone from a Streptomyces sp. (Biabani et al., 1997), and dihydrophencomycin methyl ester from Streptomyces (Pusecker et al., 1997). Thus, it will be an attractive strategy to screen metabolites exhibiting anti-MRSA activity from marine bacteria to develop an alternative therapeutic agent against MRSA. However, there is less information available on marine bacteria producing anti-MRSA substances (Isnansetyo and Kamei, 2009, Kamei and Isnansetyo, 2003). In this paper, we report the isolation and identification of an anti-MRSA substance from a marine bacterium exhibiting antibacterial activity against MRSA.

Section snippets

Bacterial strains

Two MRSA strains purchased from the Korea Culture Center of Microorganisms (KCCM, Seoul, Korea) and ten MRSA strains isolated from clinical sources (Lee et al., 2008) were used. Other bacteria tested in this study were purchased from the Korean Collection for Type Cultures (KCTC, Daejeon, Korea). All of the bacterial strains were cultivated at 37 °C in Mueller–Hinton broth (MHB, Difco Laboratories, Detroit, USA) or in Mueller–Hinton agar (MHA, Difco Laboratories, Detroit, USA) to measure

Identification of a marine bacterium producing anti-MRSA substance

A bacterial strain UJ-6, which exhibits an anti-MRSA activity, was isolated from sea water collected at Uljin, Gyungbukdo, Korea. The cell-free culture of the UJ-6 was able to make a clear zone against MRSA, indicating that the UJ-6 produces an anti-MRSA substance. In order to identify this strain, morphological, biochemical and genetical analyses were performed as described in Section 2. Gram staining revealed that this strain was a Gram negative rod (data not shown). TEM indicated that the

Conclusion

To discover antimicrobial agents from various natural resources, we isolated a marine bacterium UJ-6 producing anti-MRSA substance. The isolated strain was identified to be Pseudomonas sp. by the morphology, biochemical, and genetical analyses. An active compound exhibiting anti-MRSA activity was purified from Pseudomonas sp. UJ-6 culture through a successive chromatographic purification procedure. The compound 2 was identified to be 1-acetyl-beta-carboline based on the analyses of NMR

Conflict of interest statement

There are no conflicts of interest in this study.

Acknowledgements

This work was supported by a grant from National Fisheries Research and Development Institute (NFRDI; Korea, RF-2011-FS-002). This research was also supported by the special fund of Pukyong National University donated by the SKS Trading Co. in Lynnwood, Washington, USA in memory of late Mr. Young Hwan Kang, who had a deep concerns and inspiration in fishery science.

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