Role of teichoic acids in Staphylococcus aureus nasal colonization, a major risk factor in nosocomial infections

Colonization of the anterior nares in approximately 37% of the population is a major risk factor for severe Staphylococcus aureus infections. Here we show that wall teichoic acid (WTA), a surface-exposed staphylococcal polymer, is essential for nasal colonization and mediates interaction with human nasal epithelial cells. WTA-deficient mutants were impaired in their adherence to nasal cells, and were completely unable to colonize cotton rat nares. This study describes the first essential factor for S. aureus nasal colonization.     

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.     

Regulatory Adaptation of Staphylococcus aureus during Nasal Colonization of Humans

The nasopharynx is the main ecological niche of the human pathogen Staphylococcus aureus. Although colonization of the nares is asymptomatic, nasal carriage is a known risk factor for endogenous staphylococcal infection. We quantified S. aureus mRNA levels in nose swabs of persistent carriers to gain insight into the regulatory adaptation of the bacterium to the nasal environment. We could elucidate a general response of the pathogen to the surrounding milieu independent of the strain background or the human host. Colonizing bacteria preferentially express molecules necessary for tissue adherence or immune-evasion whereas toxins are down regulated. From the analysis of regulatory loci we found evidence for a predominate role of the essential two-component system WalKR of S. aureus. The results suggest that during persistent colonization the bacteria are metabolically active with a high cell surface turnover. The increased understanding of bacterial factors that maintain the colonization state can open new therapeutic options to control nasal carriage and subsequent infections.     

Characterisation of Staphylococcus aureus nasal and skin carriage among patients undergoing haemodialysis treatment

The aim of the study was to investigate the rate of Staphylococcus aureus nasal and skin carriage in patients undergoing haemodialysis. The cultured staphylococcal isolates were subsequently characterized by molecular methods. The study group comprised 43 haemodialysed patients from whom nasal and skin swabs from the vascular access sites were collected. The identification of staphylococcal isolates and antibiotic susceptibility testing were performed on the basis of conventional diagnostic procedures. The staphylococci were further characterized using Pulsed-Field Gel Electrophoresis (PFGE). S. aureus was cultured from 12 (27.9%) patients. Only one (8.3%) patient was colonized with the microorganism both in the anterior nares and the vascular access site representing a single strain, as evidenced by PFGE analysis. Antibiotic susceptibility testing identified one (7.6%) methicillin-resistant S. aureus (MRSA) strain. PFGE typing identified several S. aureus genotypes with the lack of one specific strain responsible for colonization. However, it should be noted that among two (A and D) PFGE patterns genetically indistinguishable and closely related isolates (two isolates for each pattern) were identified. The obtained results revealed a relatively low rate of S. aureus carriage accompanied by low methicillin resistance rate and a significant genetic diversity of cultured isolates with the lack of one predominant strain responsible for colonization.     

Prevention of Staphylococcal Bacteriophage Activity by Antigen A Precipitins in Human Sera

Antigen A precipitins in human sera prevented plaque formation and propagation of staphylococcal bacteriophages. Over 20% of total IgG was removed from human sera by absorption with staphylococci containing antigen A. The specific precipitating antibody in rabbit antisera formed lines of idenity with antigen A precipitins in lower dilutions of human sera but formed lines of nonidenity with antigen A precipitins in higher dilutions of the same sera, suggesting both specific and nonspecific antigen A precipitins in human sera. The specific and nonspecific antigen A precipitins in human sera may prevent the in vivo activity of staphylococcal bacteriophages which have been demonstrated previously in animals whose sera do not contain either specific or nonspecific antigen A precipitins.     

Bacteriophage conversion as a factor modifying the intensity of phagocytosis of Staphylococcus aureus by human leukocytes

Staphylococcus aureus NCTC 8325-4 and its eight variants lysogenized with phages responsible for the synthesis of staphylococcal staphylokinase were used for the study. Influence of phage conversion of S. aureus on its interaction with human leucocytes and influence of prophage on strain susceptibility to intracellular killing by human granulocytes without opsonins were evaluated. It was found that lysogenization of the strain with the bacteriophages decreased in each case reactivity of human leucocytes for staphylococcal strain what was expressed by lower bioluminescence values and by lower percentage of intracellular killing of bacterial cells carrying prophage.     

Changes in the nasal colonization with methicillin-resistant Staphylococcus aureus in children: 2004-2009

Background

Staphylococcus aureus is an important cause of infection, particularly in persons colonized with this organism. This study compared the annual prevalence and microbiological characteristics of methicillin-resistant S. aureus (MRSA) nasal colonization in Taiwanese children from 2004 through 2009. Risk factors for MRSA were determined for the overall study period.

Methods

Children from birth to ≤14 years of age presenting for health maintenance visits or attending 1 of 57 kindergartens were recruited. Nasal swabs were obtained, and a questionnaire was administered. The prevalence and microbiological characteristics of MRSA colonization were also calculated for two 3-year periods: 2004–2006 and 2007–2009.

Results

Cultures of the anterior nares were positive for S. aureus in 824 (25.8%) of the 3,200 children, and MRSA colonization was found in 371 (11.6%) children. The prevalence of S. aureus colonization decreased from 28.1% in 2004–2006 to 23.3% in 2007–2009 (p<0.01), whereas the prevalence of MRSA colonization increased from 8.1% to 15.1% during this period (p<0.0001). Multivariate analysis revealed that the independent risk factors for MRSA carriage were different for male and female children, and also among age groups. Most MRSA isolates belonged to sequence type 59 (ST59) (86.3%); however, a multiresistant MRSA clone with ST338 background emerged in 2007–2009. Ten (62.5%) of the 16 MRSA isolates expressed the genotypic profile ST338/staphylococcal cassette chromosome mec V_T/Panton-Valentine leukocidin-positive/staphylococcal enterotoxin B-positive, and differed only in their antimicrobial susceptibility patterns.

Conclusion

The prevalence of nasal colonization by MRSA increased among healthy Taiwanese children from 2004–2006 to 2007–2009, despite an overall decrease in the prevalence of nasal colonization by S. aureus. A multiresistant MRSA clone characterized as ST338 was identified from these children.     

Isolation and characterization of methicillin-resistant Staphylococcus aureus strains from nares of nurses and their gowns

The isolation and characterization of methicillin-resistant Staphylococcus aures (MRSA) strains from the bilateral nares of nurses and their gowns are described. MRSA strains could be isolated from eigth of fifty bilateral nares of nurses and two of their gowns. Ten MRSA strains were typed using coagulase typing, and divided into two types, coagulase II and III. In this study, we found a new group (producing toxic shock syndrome toxin -1, coagulase III and staphylococcal enterotoxin C) in Japanese MRSA. Furthermore, we confirmed that MRSA strains originating from bilateral nares of three nurses were identical and two strains isolated from the left naris of one nurse and her gown were also identical by pulsed-field gel electrophoresis.     

Persistent Staphylococcus aureus colonization is not a strongly heritable trait in Amish families

About 20% of adults are persistently colonized with S. aureus in the anterior nares. Host genetic factors could contribute susceptibility to this phenotype. The objective of this study was to determine whether the phenotype of persistent S. aureus colonization aggregates in family members who live in different households. Healthy adults and their eligible same sex siblings who lived in different households were recruited from the Old Order Amish of Lancaster, Pennsylvania. All participants had two cultures of the anterior nares to determine if they were persistently colonized with S. aureus. Three hundred and ninety eight participants finished the study, of whom 166 were index cases and 232 were siblings of index cases. Eighteen per cent (71/398) of all participants and 17% (29/166) of index cases were persistently colonized with S. aureus. Twenty two per cent (8/36) of siblings of persistently colonized index cases were persistently colonized with S. aureus compared to 17% (34/196) of siblings of non-persistently colonized index cases, yielding a prevalence rate ratio of 1.28 (95% CI: 0.65–2.54, p = 0.64) and sibling relative risk of 1.25 (95% CI: 0.65–2.38, p = 0.51). The heritability of persistent colonization was 0.19±0.21 (p = 0.31). Persistent S. aureus colonization does not strongly aggregate in Amish family members in different households and heritability is low, suggesting that environmental factors or acquired host factors are more important than host genetic factors in determining persistent S. aureus colonization in this community.     

Typing of Staphylococcus epidermidis Colonizing in Human Nares by Pulsed-Field Gel Electrophoresis

From the nares of 11 healthy adults, 253 strains of coagulase negative staphylococcus were isolated and 88% of them were identified as Staphylococcus epidermidis using the API STAPH system. Chromosomal DNA fingerprinting of the isolated strains revealed that each person carried multiple types of S. epidermidis in his or her nares. The colonization of the strains was not stable; the types of the isolates changed in the first and the second examinations 5 months apart. The results contrasted with previous findings in which only one strain of S. aureus colonized persistently in the nares of healthy adults.     

Effect of Protein A on Adsorption of Bacteriophages to Staphylococcus aureus

Experiments were performed to determine if protein A influenced the association of bacteriophages with Staphylococcus aureus. Bacteriophage adsorption was compared in a S. aureus strain rich in protein A and mutants of this strain with very little protein A, in a strain with little protein A, and in mutants of this strain with increased protein A. In addition, the effect of growth in mannitol-salt broth and trypsin digestion (known to reduce protein A) on bacteriophage absorption was measured. There was an inverse relationship between protein A content of strains and the quantity of bacteriophage absorbed. However, no inhibition of staphylococcal phages was obtained with purified soluble protein A. Protein A as a surface component rendered the bacteria more resistant to adsorption of staphylococcal typing phages presumably by masking the phage receptor sites. When protein A-deficient mutants were incubated with bacteriophages, there was survival of staphylococci with increased protein A content probably due to a selective action.     

Bacteriophage isolation from human saliva

AIMS: To detect bacteriophages for Gram-positive oral pathogens in human saliva. METHODS AND RESULTS: Saliva samples from 31 donors were screened for the presence of bacteriophages for Streptococcus sobrinus, Streptococcus mutans, Streptococcus salivarius, Actinomyces viscosus and Enterococcus faecalis. Bacteriophages for Enterococcus faecalis were found in seven samples. Enterococcus faecalis phages were still present in saliva re-collected from one donor one month, and one year after initial saliva collection. CONCLUSIONS: The presence and stability of the Enterococcus faecalis bacteriophages in human saliva suggests a possible role of these bacteriophages in the oral ecosystem. SIGNIFICANCE AND IMPACT OF THE STUDY: Phage therapy as a way to control oral bacteria might be considered.     

Role of lysosomes during the development of a fibroblast staphylococcal infection

The study of staphylococcal infection in chick embryo fibroblasts, carried out by electron-histochemical and morphometric methods, has revealed the presence of correlation between the degree of fibroblast destruction and the permeability of lysosomal membranes. The development of bacterial infection, depending on the presence of homologous bacteriophages in the medium, has also been studied.     

Unravelling bacteriophage ϕ11 requirements for packaging and transfer of mobile genetic elements in Staphylococcus aureus

Bacteriophages play a major role in spreading mobile genetic elements (MGEs)‐encoded genes among bacterial populations. In spite of this, the molecular requirements for building phage transducing particles have not been completely deciphered. Here, we systematically inactivated each ORF from the packaging and lysis modules of the staphylococcal phage ?11, used as a model for the Siphoviridae phages infecting Gram‐positive bacteria, and determined their functional role in transferring different MGEs including plasmids, staphylococcal pathogenicity islands (SaPIs) and the phage itself. In a previous report, we identified seven of these ORFs as being required for the production of functional phage or SaPI particles. In this report, we have completed the mutational analysis and have identified and characterized 15 additional phage‐encoded proteins required for the production of mature phage, SaPI, or transducing particles. Apart from these, we have not yet ascertained any specific function for the six remaining ?11 genes, though they are highly conserved among the staphylococcal bacteriophages. To the best of our knowledge, this study represents the first systematic deletion analysis of all the ORFs comprising the morphogenetic and lysis modules of a phage, clearly defining the molecular requirements involved in phage‐mediated MGEs transfer.     

Characterization of Staphylococcus aureus in a Pediatric Burn Unit

A one-year study on an endemic strain of Staphylococcus aureus phage type 84/85 in a children's burn unit is described. The endemic strain rapidly colonized the burns and nares of acute patients after admission but was not isolated from a patient on admission. Nonendemic strains of S. aureus found on some new patients were mostly non-phage typable and did not prevail in burns. The endemic strain was rarely isolated from the nares and skin of reconstructive patients or from the nares of hospital personnel. The endemic strain did colonize the oral cavity, normal skin, and intestinal tract of some acute patients. Endemic and nonendemic strains of S. aureus from the burned children were compared in their biochemical activities and antibiotic sensitivities to two groups of S. aureus from one other local and one Danish burns unit. The latter groups of strains represented different combinations of staphylococcal phage group III strains. Each of the four groups of strains differed in production of hemolysins, Tween 80 hydrolysis, egg yolk reaction, and proteolysis of casein and gelatin. All of the strains were uniformly sensitive to gentamicin, oxacillin, and cephalothin. Only 4 of 162 strains tested were methicillin resistant. The endemic S. aureus strains of phage type 84/85 were uniformly resistant to eight other antibiotics including lincomycin and clindamycin. The endemic strain was not the known cause of a clinically documented infection in a group of 82 acute patients studied. The possible role of S. aureus strains of phage group III in burn grafting problems is discussed.     

Defense against own arms: staphylococcal cysteine proteases and their inhibitors

Staphylococcus aureus is a human pathogen causing a wide range of diseases. Most staphylococcal infections, unlike those caused by other bacteria are not toxigenic and very little is known about their pathogenesis. It has been proposed that a core of secreted proteins common to many infectious strains is responsible for colonization and infection. Among those proteins several proteases are present and over the years many different functions in the infection process have been attributed to them. However, little direct, in vivo data has been presented. Two cysteine proteases, staphopain A (ScpA) and staphopain B (SspB) are important members of this group of enzymes. Recently, two cysteine protease inhibitors, staphostatin A and staphostatin B (ScpB and SspC, respectively) were described in S. aureus shedding new light on the complexity of the processes involving the two proteases. The scope of this review is to summarize current knowledge on the network of staphylococcal cysteine proteases and their inhibitors in view of their possible role as virulence factors.     

Bacteriophage therapy to reduce salmonella colonization of broiler chickens

Acute enteric infections caused by salmonellas remain a major public health burden worldwide. Poultry, particularly chickens, are known to be the main reservoir for this zoonotic pathogen. Although some progress has been made in reducing Salmonella colonization of broiler chickens by using biosecurity and antimicrobials, it still remains a considerable problem. The use of host-specific bacteriophages as a biocontrol is one possible intervention by which Salmonella colonization could be reduced. A total of 232 Salmonella bacteriophages were isolated from poultry farms, abattoirs, and wastewater in 2004 and 2005. Three phages exhibiting the broadest host ranges against Salmonella enterica serotypes Enteritidis, Hadar, and Typhimurium were characterized further by determining their morphology and lytic activity in vitro. These phages were then administered in antacid suspension to birds experimentally colonized with specific Salmonella host strains. The first phage reduced S. enterica serotype Enteritidis cecal colonization by > or = 4.2 log10 CFU within 24 h compared with controls. Administration of the second phage reduced S. enterica serotype Typhimurium by > or = 2.19 log10 CFU within 24 h. The third bacteriophage was ineffective at reducing S. enterica serotype Hadar colonization. Bacteriophage resistance occurred at a frequency commensurate with the titer of phage being administered, with larger phage titers resulting in a greater proportion of resistant salmonellas. The selection of appropriate bacteriophages and optimization of both the timing and method of phage delivery are key factors in the successful phage-mediated control of salmonellas in broiler chickens.     

Characteristics of Staphylococcus aureus associated with meat products and meat workers

Staphylococcus aureus strains with similar biotype characteristics occur in meat products and in the anterior nares of workers in slaughterhouses and meat-processing plants. Such strains are rare in humans not in frequent contact with meats. The biotype is staphylokinase-negative, β-haemolysin-negative, bovine plasma coagulase-negative, crystal violet type A and Protein A-positive. Most strains of this biotype belong to phage group II or are not typable with the international set for typing Staph. aureus of human origin.     

Diversity of toxic shock syndrome toxin 1-positive Staphylococcus aureus isolates

Staphylococcus aureus isolates from women with nasal, anal, or vaginal colonization were evaluated for population diversity by pulsed-field gel electrophoresis. Cluster analysis of restriction patterns revealed diversity indices of 0.89 and 0.99 for toxic shock syndrome toxin 1-positive and -negative isolates, respectively. Toxin-producing strains were isolated more frequently from the nares than from other sites.     

Intestinal colonization by enteroaggregative Escherichia coli supports long-term bacteriophage replication in mice

Bacteriophages have been known to be present in the gut for many years, but studies of relationships between these viruses and their hosts in the intestine are still in their infancy. We isolated three bacteriophages specific for an enteroaggregative O104:H4 Escherichia coli (EAEC) strain responsible for diarrhoeal diseases in humans. We studied the replication of these bacteriophages in vitro and in vivo in a mouse model of gut colonization. Each bacteriophage was able to replicate in vitro in both aerobic and anaerobic conditions. Each bacteriophage individually reduced biofilms formed on plastic pegs and a cocktail of the three bacteriophages was found to be more efficient. The cocktail was also able to infect bacterial aggregates formed on the surface of epithelial cells. In the mouse intestine, bacteriophages replicated for at least 3 weeks, provided the host was present, with no change in host levels in the faeces. This model of stable and continuous viral replication provides opportunities for studying the long-term coevolution of virulent bacteriophages with their hosts within a mammalian polymicrobial ecosystem.