Daptomycin and vancomycin heteroresistance revealed among CC5-SCCmecII MRSA clone and in vitro evaluation of treatment alternatives.

OBJECTIVES: Methicillin-resistant Staphylococcus aureus (MRSA) is a threat to the success of clinical treatment. Besides high antimicrobial resistance rates, the presence of heterogeneous vancomycin-intermediate S. aureus (hVISA) and heterogeneous daptomycin-non-susceptible S. aureus (hDNSSA) in the hospital environment is underestimated and is associated with treatment failure. The aim of this study was to investigate MRSA dissemination in a Brazilian hospital and to evaluate the efficacy of various treatment options in vitro. METHODS: MRSA strains were typed by MLST, PFGE and SCCmec typing. Minimum inhibitory concentrations (MICs) to daptomycin, linezolid, quinupristin/dalfopristin, teicoplanin, tetracycline, tigecycline, vancomycin and tedizolid were determined by broth microdilution. The presence of a heterogeneous population was detected by population analysis profile (PAP). Regarding hVISA and hDNSSA strains, the sequences and expression levels of genes involved in resistance to daptomycin and vancomycin were determined as well as cell wall thickness and autolysis. RESULTS: ST5/ST105-SCCmecII lineage was prevalent amongst 27 clinical MRSA characterised in this study. Two hDNSSA strains (one also hVISA) were detected and were confirmed by PAP. Isolate SCMSC29 (hVISA and hDNSSA) showed increased expression of genes involved in cell wall metabolism, slight cell wall thickening, reduction of autolysis, and single nucleotide polymorphisms (SNPs) in the rpoB and mprF genes compared with the susceptible strain SCMSC31. SCMSC35 (hDNSSA) presented SNPs in the rpoB and mprF genes as well as a thickened cell wall. CONCLUSIONS: Despite this worrying and hard to detect phenotype, treatment alternatives such as teicoplanin, linezolid, tetracycline, tigecycline, quinupristin/dalfopristin and tedizolid were all active against these isolates.

Mitochondria, endoplasmic reticulum and actin filament behavior after PDT with chloroaluminum phthalocyanine liposomal in HeLa cells.

Photodynamic therapy (PDT) for cancer is a therapeutic modality in the treatment of tumors in which visible light is used to activate a photosensitizer. Cell membranes have been identified as an important intracellular target for singlet oxygen produced during the photochemical pathway. This study analyzed the cytotoxicity in specific cellular targets of a photosensitizer used in PDT in vitro. The photosensitizing effects of chloroaluminum phthalocyanine liposomal were studied on the mitochondria, cytoskeleton and endoplasmic reticulum of HeLa cells. Cells were irradiated with a diode laser working at 670 nm, energy density of 4.5 J/cm2 and power density of 45 mW/cm2. Fluorescence microscopic analysis of the mitochondria showed changes in membrane potential. After PDT treatment, the cytoskeleton and endoplasmic reticulum presented basic alterations in distribution. The combined effect of AlPHCl liposomal and red light in the HeLa cell line induced photodamage to the mitochondria, endoplasmic reticulum and actin filaments in the cytoskeleton.