Two new xanthone antibiotics, citreamicin delta (1) and epsilon (2), with potent activity against Gram-positive pathogens including multidrug-resistant Staphylococcus aureus (MDRSA) were discovered. Compounds 1 and 2 exhibited MIC values < 1 microg/mL versus a number of resistant strains. The compounds were obtained from EtOAc extracts of Streptomyces vinaceus and were purified by countercurrent chromatography and reversed-phase HPLC. Their structures were elucidated using primarily NMR and mass spectroscopy.
Resistance to currently available antibiotics has become a widely recognized crisis in the medical community. To address this, many companies and researchers are refocusing their attention towards natural products, which have an excellent track record of producing effective antibacterial drugs. The AMRI natural product library was screened for activity against multi-drug resistant Staphylococcus aureus (MDRSA). The active samples were counter screened for cytotoxicity against the human hepatocellular carcinoma HepG2 cell line to determine an in vitro therapeutic index (in vitro TI). Those samples with a high in vitro TI were selected for fractionation and dereplication. This led to the discovery of a new anthracycline structure. This metabolite, named mutactimycin E (1), exhibited moderate activity against several gram positive organisms. Here we report the isolation, structure elucidation and biological activities of this new compound. Keywords mutactimycin, anthracycline, antibiotic, AmycolatopsisIt has been known for some time that resistance to existing antibiotics is increasing. This is particularly true in hospital settings, but is also found in community-acquired infections. This development, when coupled with the decision by many pharmaceutical companies to abandon antibacterial research in the 1980's and 1990's, resulted in the absence of new antibiotics to combat current resistance mechanisms. In an effort to address this urgent need, we began a screening campaign to identify novel antibiotics from our natural product library. Natural products, particularly those produced by microbial fermentation, were the direct source or inspiration for almost all antibiotics used today and remain the richest source for new antibacterial compound series. Our extensive library consisting of over 280,000 samples was screened for activity against a multi-drug resistant strain of Staphylococcus aureus (ATCC 43300). The hits arising out of this assay were then tested against the human hepatocellular carcinoma cell line HepG2 to filter out those samples where activity was the result of general cytotoxicity. The resulting subset of samples possessing selectivity for the bacterial target were then fractionated on an HPLC system employing UV, evaporative light scattering, and MS detectors. The eluted fractions were collected into 96-well microtiter plates and submitted for bioassay. Active compounds were subsequently dereplicated on the basis of MS and UV data.One of the hits resulting from our efforts originated from the EtOAc extract of Amycolatopsis strain 17128. A scaledup fermentation of the active strain and subsequent purification of the active compound by reverse phase HPLC yielded mutactimycin E (1) as an orange solid (Fig. 1). LC/MS data for 1 indicated a molecular weight of 560 Daltons. This piece of information along with the UV spectrum was used to search internal and external databases [1]. Analysis of the hits from this search indicated
The departure of many pharmaceutical companies from antibiotic research starting in the 1980s resulted in the absence of new antibiotics to combat the current crisis of increasing microbial resistance to currently available antibiotics. In an effort to address the increasing need for novel antibiotics, AMRI began a screening campaign to identify potent compounds from our natural product library. Natural products, particularly those produced by microbial fermentation, were the direct source or inspiration for almost all antibiotics used today and remain the richest source for new antibacterial compound series. AMRI's extensive library consisting of over 280 000 samples was screened for activity against a multi-drug resistant strain of Staphylococcus aureus (ATCC 43 300, Rockville, MD, USA). The hits arising out of this assay were then tested against the human hepatocellular carcinoma cell line HepG2 to filter out those samples where activity was the result of general cytotoxicity and determine an in vitro therapeutic index (in vitro TI). Samples with a high in vitro TI were selected for fractionation and dereplication. The resulting subset of samples possessing selectivity for the bacterial target were then fractionated on an HPLC system employing UV, ELSD (evaporative light scattering) and MS detectors. The eluted fractions were collected into 96-well microtiter plates and submitted for bioassay. LC/MS data for the active fractions generated UV spectra and molecular weights, which were used to search internal and external databases. 1 The analysis of the extract from strain 4731, which exhibited excellent activity against several Gram-positive organisms, resulted in the discovery of a new anthraquinone, which was closely related to the known antibiotic ericamycin. 2,3 Here we report the isolation, structure elucidation and biological activities of 1.The Actinoplanes sp. strain 4731 was isolated from a soil sample collected from grassland near Nanton, in Alberta, Canada. The culture was isolated by a previously described capillary chemotaxis technique, 4 using 0.18% mannitol as a chemo-attractant, spread-plating on water-yeast extract agar plates 5 and incubating in the dark at 28 1C for 10 days. All colonies visible under a dissecting scope were transferred to and purified on starch casein agar plates. Pure cultures were macerated and stored in a 10% glycerol/5% lactose solution at À80 1C. Before fermentation, the culture was streaked from a cryovial onto starch casein agar to verify purity. Fresh culture macerate was prepared from these agar plates after 14 days and used to inoculate the fermentation. Strain 4731 was identified by sequencing of the 16S rRNA gene. Genomic DNA was isolated from a culture grown in tryptic soy broth for 7 days at 28 1C by a phenol:chloroform extraction method. 6 The 16S rRNA gene was amplified using Taq polymerase (Promega, Madison, WI, USA) and the following two primer pairs: 27F, 5 0 -AGA GTTTGATCMTGGCTCAG-3 0 ; 1115R, 5 0 -AGGGTTGCGCTCGTTG-3 0 ; and 339F, 5 0 -CTCCTACGGGAGGCAGCA...
Three new antibiotics, neopyrrolomycins B (1), C (2), and D (3), with potent activity against Gram-positive pathogens were discovered. They exhibited MIC values < 1 microg/mL versus a number of resistant strains. The compounds were obtained from the ethyl acetate extracts of a Streptomyces sp. after purification by column chromatography and RP-HPLC. Their structures were elucidated using X-ray crystallography (1) and NMR spectroscopy (2 and 3).
suggestive of a¯uid collection (Figure 1). Repeat ultrasound of the swelling disclosed that the left paravertebral muscles were bulky and more echogenic than previously, with a suspicion of an echogenic area adjacent to the L3 spinous process. Immunological investigations showed a normal nitroblue tetrazolium test, negative antistaphylococcal and antinuclease antibodies and normal total immunoglobulin levels. Initial IgG2 level was low (1.14 g/L, reference range 1.4±4.5) but normal on repeat testing two weeks later. After six weeks of antibiotic treatment the swelling had disappeared and radiological images and in¯ammatory markers were normal. COMMENT Pyomyositis is very rare in temperate climates. S. aureus is responsible for about 90% of cases, other pathogens being streptococci and anaerobic bacteria. In tropical climates there are two peaks in prevalenceÐearly childhood (age 2±5) and middle age (35±40). The aetiology is uncertain. Previous trauma has sometimes been noted but other reported associations are concurrent skin or respiratory infections, muscle injury through exercise, diabetes mellitus, AIDS, steroid use and induction therapy for acute lymphocytic leukaemia. The condition tends to occur in the muscles of the trunk and upper thigh and can lead to a misdiagnosis of septic arthritis (described with pyomyositis in the adductor, iliacus and psoas muscles) 1. Diagnosis is often delayed but ultrasound, gallium-67 scans, computed tomography and MRI have all been proved effective tools for con®rming the cause 2,3. Blood cultures are positive in less than 5% of patients 4. Treatment in most cases includes surgical incision and drainage but resolution without surgical intervention is well recognized 5. Rare complications include toxic shock syndrome and staphylococcal scalded skin syndrome 5. The diagnosis should be suspected in any child with the triad of pyrexia,¯ank pain and hip symptoms 2 .
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