Tracking the evolutionary origin of the methicillin resistance gene: cloning and sequencing of a homologue of mecA from a methicillin susceptible strain of Staphylococcus sciuri.

The gene mecA, a central genetic determinant of methicillin resistance in Staphylococcus aureus and coagulase-negative staphylococci of human origin, has an unknown extra species origin in these human pathogens. After screening isolates representing over 15 different species within the genus Staphylococcus we could identify only one--S. sciuri--in which each of over 150 independent isolates showed positive hybridization with a mecA-specific DNA probe isolated from a methicillin-resistant strain of S. aureus, (MRSA). Yet, the majority of these isolates showed no resistance to penicillin or methicillin. The mecA gene homologue was cloned and sequenced from a S. sciuri strain and the sequence of mecA was compared to that of the mecA of prototype strains of methicillin resistant S. aureus and S. epidermidis. Similarly to mecA of MRSA, the mecA homologue of S. sciuri was composed of a putative transglycosylase and a transpeptidase domain the latter showing all the conserved motifs typical of the active sites of the penicillin binding domain of transpeptidases. Overall similarity between the deduced amino acid sequences of mecA of MRSA and the mecA homologue of S. sciuri was 88%. On the other hand, comparison of the transpeptidase domain of the S. sciuri mecA to the corresponding domain alone of the MRSA mecA showed a similarity of 96% and an identity of 91%, while comparison of the putative transglycosylase domains of the two bacteria showed only a 80% similarity and 68% identity of amino acid sequences. Our data suggests that mecA of methicillin-resistant strains of staphylococci pathogenic to Man originated within the genus of Staphylococcus from an evolutionary relative of the mecA homologue that we have identified in S. sciuri and which may perform a normal physiological function in this bacterium unrelated to beta-lactam resistance.

A light-sensitive yellow ommochrome pigment from the house fly.

A light-sensitive pigment (lambda max at 430 and 340 nm), extracted in acidic methanol from the Musca domestica heads, showed, in the absorption curve, a plateau at 480-500 nm and a new maximum at 400 nm, after visible light irradiation. The light-sensitive house fly pigment showed spectroscopic and chemical properties of the ommochrome pigments (Butenandt and Schäfer: Recent Progress in the Chemistry of Natural and Synthetic, Colouring Matters and Related Fields, Academic Press, New York, pp 13-33, 1962; Bolognese and Scherillo: Experientia 30:225-226, 1974). The treatment of the extracted pigment with a methanol-HClsat. mixture afforded some coloured compounds; two main products were identified by comparison of their chromatographic and spectral properties with authentic samples of 1-oxo-2H-3-carbomethoxy-5-methoxy-11-(fumaroyl-methylester)-pyrido [3,2-a] phenoxazine (compound 7) and 1-oxo-2H-3-carbomethoxy-5-methoxy-9-chlorine-11-(fumaroyl-methylester )-pyrido [3,2-a] phenoxazine (compound 8), obtained from the oxidation mixture of 3-hydroxykynurenine methylester (compound 9).

Hysterosalpingographic abnormalities in infertile women: radiological and clinical interpretation.

A review of 1035 hysterosalpingographies (HSG) has shown a frequency of acquired and congenital morphological alteration in 272 patients with spontaneous abortion and in 763 infertile couples. The frequency of congenital anomalies was 25.5% in spontaneous abortions and 5.3% in infertility. Acquired morphological anomalies were 11.7% in spontaneous abortion and 34.6% in infertility. Tubal problems represent 25% of lesions in infertility. Furthermore couples have been studied by correlating the male factor i.e. the sperm quality to various types of morphological anomalies observed in order to better evaluate the actual damage evidence by HSG.

A cytosine- over guanosine-rich sequence in RNA activates rho-dependent transcription termination.

We have constructed an expression vector carrying the Escherichia coli his operon control region to study the ability of defined segments of DNA to cause rho factor-mediated transcription termination both in vivo and in vitro. We have previously identified a consensus motif consisting of a region of high cytosine over guanosine content common to several cryptic intracistronic transcription termination elements unmasked by polar mutations. We show that a DNA fragment possessing features similar to the ones previously identified is capable of causing rho-mediated mediated release of transcripts in vivo and in vitro. The efficiency of termination depends on the length and efficiency of termination depends on the length and relative cytosine over guanosine ratio of the element.

Processing of a polycistronic mRNA requires a 5' cis element and active translation.

We have characterized a major processed species of mRNA in the his operon of Salmonella typhimurium. In vivo and in vitro analyses of the his transcripts from wild-type and mutant strains using S1 nuclease protection assays, measurements of RNA stability, deletion mapping, gel retardation, and in vitro translation assays demonstrate that the distal portion of the polycistronic his mRNA is processed, resulting in increased stability. The processing event requires an upstream cis-acting element and translation of the cistron immediately downstream of the 5' end of the processed species. The cistrons contained in this segment are also independently transcribed from an internal promoter which is maximally active in the absence of readthrough transcription from the primary promoter.

Ribonuclease E provides substrates for ribonuclease P-dependent processing of a polycistronic mRNA.

The polycistronic mRNA of the histidine operon is subject to a processing event that generates a rather stable transcript encompassing the five distal cistrons. The molecular mechanisms by which such a transcript is produced were investigated in Escherichia coli strains carrying mutations in several genes for exo- and endonucleases. The experimental approach made use of S1 nuclease protection assays on in vivo synthesized transcripts, site-directed mutagenesis and construction of chimeric plasmids, dissection of the processing reaction by RNA mobility retardation experiments, and in vitro RNA degradation assays with cellular extracts. We have found that processing requires (1) a functional endonuclease E; (2) target site(s) for this activity in the RNA region upstream of the 5' end of the processed transcript that can be substituted by another well-characterized rne-dependent cleavage site; (3) efficient translation initiation of the first cistron immediately downstream of the 5' end; and (4) a functional endonuclease P that seems to act on the processing products generated by ribonuclease E. This is the first evidence that ribonuclease P, an essential ribozyme required for the biosynthesis of tRNA, may also be involved in the segmental stabilization of a mRNA.