A silent carbapenemase gene in strains of Bacteroides fragilis can be expressed after a one-step mutation

High-level carbapenem-resistant (CpmR) mutants, with MICs for imipenem and carbapenem of greater than 128 micrograms/ml, were selected in vitro from four carbapenem-susceptible (CpmS) clinical isolates of Bacteroides fragilis. The CpmS strains produced very low levels of beta-lactamase activity, which was increased approx. 50- to 100-fold in the CpmR mutants. Isoelectric focussing and enzyme kinetic analysis (Km and Vrel) of the 'carbapenemases' from the CpmR mutants and similarly resistant clinical isolates suggested a close relatedness of the enzymes. A probe covering most of the cfiA gene encoding such an enzyme (Thompson, J.S. and Malamy, M.H. (1990) J. Bacteriol. 172, 2584-2593) hybridized with DNA from the CpmR mutants, their CpmS parental strains as well as clinical CpmR isolates, but not from randomly chosen carbapenem-susceptible strains. The possibility is considered that mutations leading to expression of the silent carbapenemase gene, and thereby to clinically relevant carbapenem resistance, may also occur in the clinical setting.     

Enumeration of Carnobacterium divergens V41, Carnobacterium piscicola V1 and Lactobacillus brevis LB62 by in situ hybridizationflow cytometry

The specific detection and enumeration of Lactobacillus brevis LB62, Carnobacterium divergens V14 and Carnobacterium piscicola VI were studied by in situ hybridizationflow cytometry. The method was performed on the exponential growth phase with three probes targeting 16S rRNA labelled with fluorescein isothicyanate (FITC) : EUB338 probe universal for Eubacteria, Lb probe specific for Lact. brevis and Cb probe specific for the genus Carnobacterium . EUB338 was used to determine the permeabilization and hybridization conditions for the cells. The Lb probe gave no hybridization signal whereas the Cb probe allowed the detection and quantification by flow cytometry at 520 nm of the two Carnobacterium strains in pure culture or in mixtures with Listeria innocua F.     

Identification of pathogenic fungi in surgical and autopsy specimens by immunofluorescence

Summary A method is presented for the preparation of immune sera and detection by immunofluorescence ofC. immitis, S. schenckii, B. dermatitidis, C. neoformans, andC. albicans in surgical and autopsy material. Formalin fixation does not affect the antigens of the mycotic agents. There are no cross reactions except withC. immitis andC. neoformans, which can be differentiated by the site of the specific fluorescence in each organism.     

EDC3 phosphorylation regulates growth and invasion through controlling P‐body formation and dynamics

Regulation of mRNA stability and translation plays a critical role in determining protein abundance within cells. Processing bodies (P‐bodies) are critical regulators of these processes. Here, we report that the Pim1 and 3 protein kinases bind to the P‐body protein enhancer of mRNA decapping 3 (EDC3) and phosphorylate EDC3 on serine (S)161, thereby modifying P‐body assembly. EDC3 phosphorylation is highly elevated in many tumor types, is reduced upon treatment of cells with kinase inhibitors, and blocks the localization of EDC3 to P‐bodies. Prostate cancer cells harboring an EDC3 S161A mutation show markedly decreased growth, migration, and invasion in tissue culture and in xenograft models. Consistent with these phenotypic changes, the expression of integrin β1 and α6 mRNA and protein is reduced in these mutated cells. These results demonstrate that EDC3 phosphorylation regulates multiple cancer‐relevant functions and suggest that modulation of P‐body activity may represent a new paradigm for cancer treatment.