Identification of a gene (arpU) controlling muramidase-2 export in Enterococcus hirae

Muramidase-2 of Enterococcus hirae is a 74-kDa peptidoglycan hydrolase that plays a role in cell wall growth and division. To study its regulation, we isolated a mutant defective in muramidase-2 release under certain growth conditions. This mutant had cell walls which apparently lacked 74-kDa muramidase-2 but which accumulated two proteolytic fragments of 32 and 43 kDa, which exhibited muramidase-2 activity in the membrane fraction. By complementation cloning, we identified a 2.6-kb fragment of the E. hirae chromosome containing a gene cluster coding for proteins of 58 to 137 amino acids. One of these genes (arpU), which encoded a 15.9-kDa protein, was shown to complement the defect of the A9 mutant in trans. We propose that this gene may be involved in the regulation of muramidase-2 export

Streptococcus faecium mutants that are temperature sensitive for cell growth and show alterations in penicillin-binding proteins

The penicillin-binding proteins (PBPs) of 209 cell division (or growth) temperature-sensitive mutants of Streptococcus faecium were analyzed in this study. A total of nine strains showed either constitutive or temperature-sensitive conditional damage in the PBPs. Analysis of these nine strains yielded the following results: one carried a PBP 1 constitutively showing a lower molecular weight; one constitutively lacked PBP 2; two lacked PBP 3 at 42 degrees C, but not at 30 degrees C; one was normal at 30 degrees C but at 42 degrees C lacked PBP 3 and overproduced PBP 5; two were normal at 42 degrees C and lacked PBP 5 at 30 degrees C; one constitutively lacked PBP 5; and one carried a PBP 6 constitutively split in two bands. The mutant lacking PBP 3 and overproducing PBP 5 continued to grow at 42 degrees C for 150 min and then lysed. Revertants selected for growth capability at 42 degrees C from the mutants altered in PBPs 5 and 6 maintained the same PBP alterations, while those isolated from the strains with altered PBP 1 or lacking PBP 2 or PBP 3 showed a normal PBP pattern. Penicillin-resistant derivatives were isolated at 30 degrees C from the mutants lacking PBP 2 and from that lacking PBP 3. All these derivatives continued to show the same PBP damage as the parents, but overproduced PBP 5 and grew at 42 degrees C. These findings indicate that high-molecular-weight, but not low-molecular-weight, PBPs are essential for cell growth in S. faecium.(ABSTRACT TRUNCATED AT 250 WORDS

Rapid detection of PCR products of hepatitis C virus (HCV) with a non-radioactive oligoprobe

Dot-blot for rapid detection of PCR amplification products of hepatitis C virus (HCV) using a digoxigenin (DIG)-labelled oligoprobe was developed and its sensitivity compared with Southern blot hybridization. The specificity and sensitivity of the DIG-labelled probe were identical to those of the 32P-labelled when the DIG-labelled nucleic acids were detected by enzyme-catalyzed chemiluminescent reaction. The lack of radioactivity makes this procedure suitable for routine use in diagnostic laboratories

Influenza del fattore sigma RpoS sulla sopravvivenza di Escherichia coli in stato vitale ma non coltivabile

Valutazione dell'influenza di Rpos o fattore sigma alternativo sula capacit\ue0 di sopravvivenza di Escserichia coli in presenza di situazioni di stress ambientale. E' stato indotto lo stato VBNC in vari mutanti di E.coli difettivi in rpoS: si \ue8 valutata la cinetica di acquisizione delo stato VBNC e la possibilit\ue0 di "rivitalizzazione" rispetto ai ceppi selvaggi. I mutanti RpoS-difettivi muoiono pi\uf9 rapidamente e presentano una capacit\ue0 di ricrescita molto pi\uf9 scarsa rispetto ai ceppi progentori, indicando un probabile ruolo di rpoS nella fase di entrata e mantenimento dello stato VBN