R. A. de Torres

Accuracy of Cefoxitin Disk Testing for Characterization of Oxacillin Resistance Mediated by Penicillin-Binding Protein 2a in Coagulase-Negative Staphylococci

ABSTRACT The Clinical and Laboratory Standards Institute (CLSI) proposed, beginning in 2004, the use of cefoxitin disks to predict resistance mediated by the mecA gene in all species of coagulase-negative staphylococci (CoNS). The aim of this work was to evaluate the efficiency of the cefoxitin disk and of oxacillin-salt agar screening (MHOX) to characterize the oxacillin resistance mediated by the mecA gene in CoNS. One hundred seven CoNS isolates from different clinical samples were studied. Detection of the mecA gene by PCR was considered the “gold standard.” The susceptibility to oxacillin and cefoxitin was detected by the disk diffusion and agar dilution tests, as described by the CLSI. MHOX was also performed with 6 μg/ml of oxacillin and 4% NaCl. The sensitivities of the oxacillin and cefoxitin disks for all CoNS species were 88% and 80%, respectively, whereas the specificities were 63% and 100%, respectively. The sensitivities of the agar dilution test for oxacillin and cefoxitin (for proposed breakpoints of ≥4 μg/ml for resistance and ≤2 μg/ml for susceptibility) were 90% and 85%, respectively, whereas the specificities were 76% and 98%, respectively. MHOX showed a sensitivity of 90% and a specificity of 95% for all CoNS species. Both the MHOX and the cefoxitin disk results indicate that these are appropriate methods for the evaluation of oxacillin resistance mediated by the mecA gene in all CoNS species.

Flagellin, a major protein present in SDS-PAGE profiles of Sarkosyl-OMP-enriched fractions from Bordetella bronchiseptica Bvg − or modulated Bvg + strains

The bvg or vir locus positively regulates the expression of many Bordetella virulence-associated determinants (encoded by vag genes), including cell envelope proteins, in response to environmental stimuli. On the other hand, several genes named vrg genes are negatively controlled by the bvg regulon (Knapp and Mekalanos, 1988). Flagellin is encoded by a vrg gene, which is expressed when the principal virulence factors are eliminated during antigenic modulation or in phase variants (Akerley et al., 1992). We have previously analyzed SDS-PAGE profiles of Sarkosyl-outer membrane protein (OMP)-enriched fractions from B. bronchiseptica Bvg- and modulated Bvg+ strains and reported a major band associated with the avirulent phenotype (Passerini de Rossi et al., 1995). In order to characterize this band we have purified flagellar filaments from Bvg- and modulated Bvg+ strains, and analyzed them by SDS-PAGE. These profiles revealed a single major band of 40 or 45 kDa depending on the strain. The N-terminal amino acid sequence of the putative flagellin expressed by BB7200a was identical over the first 21 residues analyzed to that of the flagellin from the modulated strain BB7865 reported by Akerley et al. (1992). Comparison of the SDS-PAGE profile of flagellar filaments with that of the OMP-enriched fraction of the corresponding strain showed that the flagellum-associated polypeptide had the same electrophoretic mobility as that of the characteristic band of the avirulent phenotype. Furthermore, this band was absent in the OMP-enriched fraction profile from a Bvg- strain subjected to a treatment that removes flagella. Our results indicate that the major protein observed in SDS-PAGE profiles of Sarkosyl-OMP-enriched fractions from B. bronchiseptica Bvg- and modulated Bvg+ strains corresponds to flagellin.

Evolution of Neisseria gonorrhoeae drug susceptibility in Buenos Aires, Argentina, 1985–99

Buenos Aires (Argentina) is an international port city of three million people surrounded by greater Buenos Aires, with a core population of 10 million people. The STD service of the Hospital Nacional de Cllinicas, University of Buenos Aires (HNC) deals with 80% patients from the city and 20% from the suburban areas. A total of 33 316 symptomatic patients (27 458 men and 5858 women) were studied from 1985–99.1 Patients attended under the standard regulations of HNC primary care protocol, and so no additional written informed consent was required. Neisseria gonorrhoeae (NG) was investigated by direct Gram stain microscopy and by culture in Columbia agar supplemented with 5% blood and …

Natural resistance of BALB/c mice to herpes simplex virus-type 1 intraperitoneal infection is abrogated by a plant extract with in vitro anti-herpes activity

Adult BALB/c mice, genetically resistant to HSV‐1 i.p. infection, were inoculated intraperitoneally with 1000 PFU of HSV‐1 and treated by the same route with four consecutive doses of Melia azedarach L. aqueous green leaves extract (MA) each containing 80 antiviral units against HSV‐1 replication in Vero cells. Mice were injected 1 day and 6 h before infection and 1 and 2 days after infection. Mortality among untreated 45 day old female and male mice was around 35% for both sexes, whereas MA treatment increased mortality to 95% in females and to 50% in males. Virus was isolated from brains of treated animals. Enhanced susceptibility to HSV‐1 infection upon treatment with MA extract was also observed in male mice 25 days old, before hormonal maturation, indicating that the impairment of natural resistance to HSV‐1 infection is sex‐linked. Treatment with MA extracts did not interfere with established specific immune protection nor with the induction of immune response to HSV‐1. Virus was recovered from peritoneal exudate cells only in MA‐treated animals. The abrogation of natural resistance of inbred mice to i.p. HSV‐1 infection by pretreatment and early administration of the plant extract indicates that the in vivo expression of immunomodulating activities on receptor cells interfered with its potent antiviral activity displayed in vitro against HSV‐1 replication.