Indications for both host-specific and introduced genotypes of Staphylococcus aureus in marine mammals.

Staphylococcus aureus is present in the marine environment and causes disease in marine mammals. To determine whether marine mammals are colonized by host-specific strains or by strains originating from other species, we performed multi-locus sequence typing on ten S. aureus strains isolated from marine mammals in the U.K., the Netherlands, and the Antarctic. Four new sequence types of S. aureus were discovered. S. aureus strains from a southern elephant seal (n=1) and harbour porpoises (n=2) did not cluster with known S. aureus strains, suggesting that they may be host species-specific. In contrast, S. aureus strains from harbour seals (n=3), other harbour porpoises (n=3), and a grey seal (n=1) clustered with S. aureus strains previously isolated from domestic ruminants, humans, or birds, suggesting that these S. aureus strains in marine mammals were introduced from terrestrial species.

Rhesus macaques (Macaca mulatta) are natural hosts of specific Staphylococcus aureus lineages.

Currently, there is no animal model known that mimics natural nasal colonization by Staphylococcus aureus in humans. We investigated whether rhesus macaques are natural nasal carriers of S. aureus. Nasal swabs were taken from 731 macaques. S. aureus isolates were typed by pulsed-field gel electrophoresis (PFGE), spa repeat sequencing and multi-locus sequence typing (MLST), and compared with human strains. Furthermore, the isolates were characterized by several PCRs. Thirty-nine percent of 731 macaques were positive for S. aureus. In general, the macaque S. aureus isolates differed from human strains as they formed separate PFGE clusters, 50% of the isolates were untypeable by agr genotyping, 17 new spa types were identified, which all belonged to new sequence types (STs). Furthermore, 66% of macaque isolates were negative for all superantigen genes. To determine S. aureus nasal colonization, three nasal swabs from 48 duo-housed macaques were taken during a 5 month period. In addition, sera were analyzed for immunoglobulin G and A levels directed against 40 staphylococcal proteins using a bead-based flow cytometry technique. Nineteen percent of the animals were negative for S. aureus, and 17% were three times positive. S. aureus strains were easily exchanged between macaques. The antibody response was less pronounced in macaques compared to humans, and nasal carrier status was not associated with differences in serum anti-staphylococcal antibody levels. In conclusion, rhesus macaques are natural hosts of S. aureus, carrying host-specific lineages. Our data indicate that rhesus macaques are useful as an autologous model for studying S. aureus nasal colonization and infection prevention.

Reclassification of Staphylococcus aureus nasal carriage types.

BACKGROUND: Persistent nasal carriers have an increased risk of Staphylococcus aureus infection, whereas intermittent carriers and noncarriers share the same low risk. This study was performed to provide additional insight into staphylococcal carriage types. METHODS: Fifty-one volunteers who had been decolonized with mupirocin treatment and whose carriage state was known were colonized artificially with a mixture of S. aureus strains, and intranasal survival of S. aureus was compared between carriage groups. Antistaphylococcal antibody levels were also compared among 83 carriage-classified volunteers. RESULTS: Persistent carriers preferentially reselected their autologous strain from the inoculum mixture (P=.02). They could be distinguished from intermittent carriers and noncarriers on the basis of the duration of postinoculation carriage (154 vs. 14 and 4 days, respectively; P=.017, by log-rank test). Cultures of swab samples from persistent carriers contained significantly more colony-forming units per sample than did cultures of swab samples from intermittent carriers and noncarriers (P=.004). Analysis of serum samples showed that levels of immunoglobulin G and immunoglobulin A to 17 S. aureus antigens were equal in intermittent carriers and noncarriers but not in persistent carriers. CONCLUSIONS: Along with the previously described low risk of infection, intermittent carriers and noncarriers share similar S. aureus nasal elimination kinetics and antistaphylococcal antibody profiles. This implies a paradigm shift; apparently, there are only 2 types of nasal carriers: persistent carriers and others. This knowledge may increase our understanding of susceptibility to S. aureus infection.

Anti-staphylococcal humoral immune response in persistent nasal carriers and noncarriers of Staphylococcus aureus.

BACKGROUND: Persistent carriers have a higher risk of Staphylococcus aureus infections than noncarriers but a lower risk of bacteremia-related death. Here, the role played by anti-staphylococcal antibodies was studied. METHODS: Serum samples from 15 persistent carriers and 19 noncarriers were analyzed for immunoglobulin (Ig) G, IgA, and IgM binding to 19 S. aureus antigens, by means of Luminex technology. Nasal secretions and serum samples obtained after 6 months were also analyzed. RESULTS: Median serum IgG levels were significantly higher in persistent carriers than in noncarriers for toxic shock syndrome toxin (TSST)-1 (median fluorescence intensity [MFI] value, 11,554 vs. 4291; P < .001) and staphylococcal enterotoxin (SE) A (742 vs. 218; P < .05); median IgA levels were higher for TSST-1 (P < .01), SEA, and clumping factor (Clf) A and B (P < .05). The in vitro neutralizing capacity of anti-TSST-1 antibodies was correlated with the MFI value (R(2) = 0.93) and was higher in persistent carriers (90.6% vs. 70.6%; P < .05). Antibody levels were stable over time and correlated with levels in nasal secretions (for IgG, R(2) = 0.87; for IgA, R(2) = 0.77). CONCLUSIONS: Antibodies to TSST-1 have a neutralizing capacity, and median levels of antibodies to TSST-1, SEA, ClfA, and ClfB are higher in persistent carriers than in noncarriers. These antibodies might be associated with the differences in the risk and outcome of S. aureus infections between nasal carriers and noncarriers.

Host-microbe interplay in persistent Staphylococcus aureus nasal carriage in HIV patients.

It has been shown that persistent Staphylococcus aureus nasal carriage results in increased bacterial dispersal and a higher risk of infection compared to non-or-intermittent S. aureus carriage. Although many studies investigated S. aureus nasal carriage in HIV patients, none compared persistent carriage to non-persistent carriage nor were studies performed in the HAART era. We investigated the host-microbe interplay of persistent S. aureus nasal carriage in HIV-infected patients by studying host determinants of persistent carriage as well as the genetic structure of S. aureus strains isolated. We compared this genetic structure with the previously determined population structure of S. aureus isolates obtained from healthy individuals. Between February 2004 and June 2005 all HIV patients visiting the outpatient department of Erasmus MC (Rotterdam, The Netherlands) were asked to participate in this study. Participants were interviewed and screened for persistent S. aureus carriage using two semi-quantitative nasal swab cultures. For 443 patients two cultures were available, 131 (29.6%) were persistent carriers, which is significantly higher as compared to healthy individuals from the same geographic region (17.6%; P<0.0001). Male sex (odds ratio [OR], 2.22; 95% confidence interval [CI], 1.32-3.73), current smoking (OR, 0.58; 95% CI, 0.38-0.90), Pneumocystis jiroveci pneumonia (PCP) prophylaxis (OR, 0.39; 95% CI, 0.16-0.97) and antiretroviral therapy (OR, 0.61; 95% CI, 0.38-0.98) were independent determinants of persistent carriage. Only two strains were mecA positive (1.2%) and no PVL positive strains were detected. The population structure of S. aureus strains isolated from HIV patients appeared to be strongly overlapping with that of S. aureus isolates from healthy individuals.

Key role for clumping factor B in Staphylococcus aureus nasal colonization of humans.

BACKGROUND: Staphylococcus aureus permanently colonizes the vestibulum nasi of one-fifth of the human population, which is a risk factor for autoinfection. The precise mechanisms whereby S. aureus colonizes the nose are still unknown. The staphylococcal cell-wall protein clumping factor B (ClfB) promotes adhesion to squamous epithelial cells in vitro and might be a physiologically relevant colonization factor. METHODS AND FINDINGS: We define the role of the staphylococcal cytokeratin-binding protein ClfB in the colonization process by artificial inoculation of human volunteers with a wild-type strain and its single locus ClfB knock-out mutant. The wild-type strain adhered to immobilized recombinant human cytokeratin 10 (CK10) in a dose-dependent manner, whereas the ClfB(-) mutant did not. The wild-type strain, when grown to the stationary phase in a poor growth medium, adhered better to CK10, than when the same strain was grown in a nutrient-rich environment. Nasal cultures show that the mutant strain is eliminated from the nares significantly faster than the wild-type strain, with a median of 3 +/- 1 d versus 7 +/- 4 d (p = 0.006). Furthermore, the wild-type strain was still present in the nares of 3/16 volunteers at the end of follow-up, and the mutant strain was not. CONCLUSIONS: The human colonization model, in combination with in vitro data, shows that the ClfB protein is a major determinant of nasal-persistent S. aureus carriage and is a candidate target molecule for decolonization strategies.

Host- and tissue-specific pathogenic traits of Staphylococcus aureus.

Comparative genomics were used to assess genetic differences between Staphylococcus aureus strains derived from infected animals versus colonized or infected humans. A total of 77 veterinary isolates were genetically characterized by high-throughput amplified fragment length polymorphism (AFLP). Bacterial genotypes were introduced in a large AFLP database containing similar information for 1,056 human S. aureus strains. All S. aureus strains isolated from animals in close contact with humans (e.g., pet animals) were predominantly classified in one of the five main clusters of the AFLP database (cluster I). In essence, mastitis-associated strains from animals were categorized separately (cluster IVa) and cosegregated with bacteremia-associated strains from humans. Distribution of only 2 out of 10 different virulence genes differed across the clusters. The gene encoding the toxic shock syndrome protein (tst) was more often encountered among veterinary strains (P < 0.0001) and even more in the mastitis-related strains (P<0.0001) compared to human isolate results. The gene encoding the collagen binding protein (cna) was rarely detected among invasive human strains. The virulence potential, as indicated by the number of virulence genes per strain, did not differ significantly between the human- and animal-related strains. Our data show that invasive infections in pets and humans are usually due to S. aureus strains with the same genetic background. Mastitis-associated S. aureus isolated in diverse farm animal species form a distinct genetic cluster, characterized by an overrepresentation of the toxic shock syndrome toxin superantigen-encoding gene.

Effect of mupirocin treatment on nasal, pharyngeal, and perineal carriage of Staphylococcus aureus in healthy adults.

Nasal carriage of Staphylococcus aureus is an important risk factor for S. aureus infections. Mupirocin nasal ointment is presently the treatment of choice for decolonizing the anterior nares. However, recent clinical trials show limited benefit from mupirocin prophylaxis in preventing nosocomial S. aureus infections, probably due to (re)colonization from extranasal carriage sites. Therefore, we studied the effectiveness of mupirocin nasal ointment treatment on the dynamics of S. aureus nasal and extranasal carriage. Twenty noncarriers, 26 intermittent carriers, and 16 persistent carriers had nasal, throat, and perineum samples taken 1 day before and 5 weeks after mupirocin treatment (twice daily for 5 days) and assessed for growth of S. aureus. The identities of cultured strains were assessed by restriction fragment length polymorphisms of the coagulase and protein A genes. The overall carriage rate (either nasal, pharyngeal, or perineal carrier or a combination) was significantly reduced after mupirocin treatment from 30 to 17 carriers (P = 0.003). Of the 17 carriers, 10 (60%) were still colonized with their old strain, 6 (35%) were colonized with an exogenous strain, and 1 (5%) was colonized with both. Two noncarriers became carriers after treatment. The acquisition of exogenous strains after mupirocin treatment is a common phenomenon. The finding warrants the use of mupirocin only in proven carriers for decolonization purposes. Mupirocin is effective overall in decolonizing nasal carriers but less effective in decolonizing extranasal sites.

Natural population dynamics and expansion of pathogenic clones of Staphylococcus aureus.

The population structure of Staphylococcus aureus carried by healthy humans was determined using a large strain collection of nonclinical origin (n = 829). High-throughput amplified fragment length polymorphism (AFLP) analysis revealed 3 major and 2 minor genetic clusters of S. aureus, which were corroborated by multilocus sequence typing. Major AFLP cluster I comprised 44.4% of the carriage isolates and showed additional heterogeneity whereas major AFLP groups II and III presented 2 homogeneous clusters, including 47.3% of all carriage isolates. Coanalysis of invasive S. aureus strains and epidemic methicillin-resistant S. aureus (MRSA) revealed that all major clusters contained invasive and multiresistant isolates. However, clusters and subclusters with overrepresentation of invasive isolates were also identified. Bacteremia in elderly adults, for instance, was caused by a IVa cluster-derived strain significantly more often than by strains from other AFLP clusters. Furthermore, expansion of multiresistant clones or clones associated with skin disease (impetigo) was detected, which suggests that epidemic potential is present in pathogenic strains of S. aureus. In addition, the virulence gene encoding Panton-Valentine leukocidin was significantly enriched in S. aureus strains causing abscesses and arthritis in comparison with the carriage group. We provide evidence that essentially any S. aureus genotype carried by humans can transform into a life-threatening human pathogen but that certain clones are more virulent than others.

Predicting the Staphylococcus aureus nasal carrier state: derivation and validation of a "culture rule".

BACKGROUND: To study determinants and risks of Staphylococcus aureus nasal carriage, adequate differentiation between the different S. aureus carrier states is obligatory. We set out to develop a "culture rule" capable of differentiating between persistent and intermittent or noncarriers that uses a minimum of nasal swab cultures. METHODS: In 51 healthy volunteers (derivation cohort), 12 quantitative nasal cultures were performed to establish S. aureus nasal carriage states. Persons with 11 or 12 cultures positive for S. aureus were classified as persistent carriers, and those with negative results of all cultures were classified as noncarriers. All other persons were classified as intermittent carriers. By means of logistic regression and receiver operating characteristic (ROC) curves, a culture rule was derived. This culture rule was subsequently validated in 106 participants of an ongoing study in 3882 elderly persons, again with the use of 12 quantitative nasal cultures. RESULTS: In both cohorts, the positive predictive value of 2 consecutive positive culture results for persistent carriage was 79%. The model best differentiating between persistent and intermittent or noncarriers used the number of positive culture results combined with the amount of S. aureus in these cultures. By using the outcome of 2 cultures, the areas under the ROC curves were 0.981 (95% confidence interval [CI], 0.949-1.0) for the derivation cohort and 0.936 (95% CI, 0.881-0.990) for the validation cohort. CONCLUSIONS: Combining qualitative and quantitative results of 2 nasal swab cultures accurately predicted the persistent S. aureus carriage state with a reliability of 93.6%. Thus, this culture rule can be used in studies of determinants and risks of S. aureus nasal carriage.