Penicillinase testing is required for Staphylococcus aureus isolates with a penicillin MIC of =0.12 mg/L. This study compared five phenotypic assays with a PCR for blaZ when testing 197 S. aureus isolates. The starch-iodine plate method and nitrocefin tests had low sensitivities of 42.9% and 35.7%, respectively. The cloverleaf assay and the penicillin zone-edge determination method had sensitivities of 67.8% and 71.4%, respectively, and these methods might be appropriate in many circumstances, but were not as sensitive as blaZ PCR.
The International Space Station (ISS) and the Antarctic Research Station Concordia are confined and isolated habitats in extreme and hostile environments. The human and habitat microflora can alter due to the special environmental conditions resulting in microbial contamination and health risk for the crew. In this study, 29 isolates from the ISS and 55 from the Antarctic Research Station Concordia belonging to the genera Staphylococcus and Enterococcus were investigated. Resistance to one or more antibiotics was detected in 75.8 % of the ISS and in 43.6 % of the Concordia strains. The corresponding resistance genes were identified by polymerase chain reaction in 86 % of the resistant ISS strains and in 18.2 % of the resistant Concordia strains. Plasmids are present in 86.2 % of the ISS and in 78.2 % of the Concordia strains. Eight Enterococcus faecalis strains (ISS) harbor plasmids of about 130 kb. Relaxase and/or transfer genes encoded on plasmids from gram-positive bacteria like pIP501, pRE25, pSK41, pGO1 and pT181 were detected in 86.2 % of the ISS and in 52.7 % of the Concordia strains. Most pSK41-homologous transfer genes were detected in ISS isolates belonging to coagulase-negative staphylococci. We demonstrated through mating experiments that Staphylococcus haemolyticus F2 (ISS) and the Concordia strain Staphylococcus hominis subsp. hominis G2 can transfer resistance genes to E. faecalis and Staphylococcus aureus, respectively. Biofilm formation was observed in 83 % of the ISS and in 92.7 % of the Concordia strains. In conclusion, the ISS isolates were shown to encode more resistance genes and possess a higher gene transfer capacity due to the presence of three vir signature genes, virB1, virB4 and virD4 than the Concordia isolates.
A gene encoding a serine-aspartate repeat protein of Staphylococcus saprophyticus, an important cause of urinary tract infections in young women, has been cloned and sequenced. In contrast to other SD repeat proteins, SdrI carries 21 additional N-terminal repeats with a consensus sequence of (P/A)ATKE(K/E)A(A/V )(T/I)(A/T/S)EE and has the longest SD(AD) (1-5) repetitive region (854 amino acids) described so far. This highly repetitive sequence contains only the amino acids serine, asparagine, and a distinctly greater amount of alanine (37%) than all other known SD repeat proteins (2.3 to 4.4%). In addition, it is a collagen-binding protein of S. saprophyticus and the second example in this organism of a surface protein carrying the LPXTG motif. We constructed an isogenic sdrI knockout mutant that showed decreased binding to immobilized collagen compared with wild-type S. saprophyticus strain 7108. Binding could be reconstituted by complementation. Collagen binding is specifically caused by SdrI, and the recently described UafA protein, the only LPXTG-containing protein in the genome sequence of the type strain, is not involved in this trait. Our experiments suggest that, as in other staphylococci, the presence of different LPXTG-anchored cell wall proteins is common in S. saprophyticus and support the notion that the presence of matrix-binding surface proteins is common in staphylococci.
Staphylococcus saprophyticus surface-associated protein (Ssp) was the first surface protein described for this organism. Ssp-positive strains display a fuzzy layer of surface-associated material in electron micrographs, whereas Ssp-negative strains appear to be smooth. The physiologic function of Ssp, however, has remained elusive. To clone the associated gene, we determined the N-terminal sequence, as well as an internal amino acid sequence, of the purified protein. We derived two degenerate primers from these peptide sequences, which we used to identify the ssp gene from genomic DNA of S. saprophyticus 7108. The gene was cloned by PCR techniques and was found to be homologous to genes encoding staphylococcal lipases. In keeping with this finding, strains 7108 and 9325, which are Ssp positive, showed lipase activity on tributyrylglycerol agar plates, whereas the Ssp-negative strain CCM883 did not. Association of enzyme activity with the cloned DNA was proven by introducing the gene into Staphylococcus carnosus TM300. When wild-type strain 7108 and an isogenic mutant were analyzed by transmission electron microscopy, strain 7108 exhibited the fuzzy surface layer, whereas the mutant appeared to be smooth. Lipase activity and the surface appendages could be restored by reintroduction of the cloned gene into the mutant. Experiments using immobilized collagen type I did not provide evidence for the involvement of Ssp in adherence to this matrix protein. Our experiments thus provided evidence that Ssp is a surface-associated lipase of S. saprophyticus.
Staphylococcus saprophyticus, an obligate human pathogen, is the most common Gram-positive causative agent of urinary tract infection (UTI) in young, healthy women. Despite the clinical importance of S. saprophyticus, little is known about how it causes disease in the urinary tract or how the host responds to the infection. Here we established an in vivo model to study both host and bacterial factors contributing to S. saprophyticus UTI. Using this model, we show that S. saprophyticus preferentially infects C3H/HeN murine kidneys instead of the bladder, a trait observed for multiple clinical isolates. Bacterial persistence in the kidneys was observed in C3H/HeN mice but not in C57BL/6 mice, indicating that host factors strongly contribute to the ability of S. saprophyticus to cause UTI. Using C3H/HeN mice as a model, histologic and immunofluorescence analyses of infected tissues revealed that S. saprophyticus induced epithelial cell shedding in the bladder and an inflammatory response characterized by macrophage and neutrophil infiltration in the bladder and kidneys. The inflammatory response correlated with increased production of proinflammatory cytokines and chemokines in both the bladder and the kidneys. Finally, we observed that the putative S. saprophyticus virulence factors Ssp and SdrI were important for persistence, but not for initial colonization, in the murine urinary tract. Thus, we characterized both host and bacterial factors involved in progression of S. saprophyticus UTI, and we describe a useful model system for studying factors involved in the pathogenesis of this Gram-positive uropathogen.Urinary tract infections (UTI) affect over 11 million women annually in the United States (11). The primary cause of UTI is the Gram-negative bacterium Escherichia coli. However, the Gram-positive bacterium Staphylococcus saprophyticus can cause up to 10 to 15% of uncomplicated UTI (33). Thus, it is estimated that S. saprophyticus causes up to 1 million UTI each year and is the second most common cause of UTI in sexually active women (41). Moreover, Gram-positive bacteria, such as S. saprophyticus, often coexist with dominant uropathogens in the urine of infected patients, although the concentrations are lower, and therefore tend to be overlooked by routine laboratory diagnostics (35). Thus, the reported estimates of the incidence of this organism may be artificially low. Interestingly, there is a seasonal pattern for S. saprophyticus UTI; such infections peak during late summer and fall, a pattern similar to that observed for sexually transmitted diseases (23). Other Gram-positive bacteria that cause UTI include Enterococcus faecalis and Enterococcus faecium (3). In contrast to S. saprophyticus, Enterococcus spp. cause UTI in healthy young women infrequently, but they contribute to ϳ19% of complicated UTI and are often nosocomially acquired (31). Despite the fact that S. saprophyticus is the predominant cause of Gram-positive UTI, relatively little is known about how this organism causes disease in the urin...
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