Characterisation of neonatal Staphylococcus capitis NRCS-A isolates compared with non NRCS-A Staphylococcus capitis from neonates and adults.

Staphylococcus capitis is a frequent cause of late-onset sepsis in neonates admitted to Neonatal Intensive Care Units (NICU). One clone of S. capitis, NRCS-A has been isolated from NICUs globally although the reasons for the global success of this clone are not well understood.We analysed a collection of S. capitis colonising babies admitted to two NICUs, one in the UK and one in Germany as well as corresponding pathological clinical isolates. Genome analysis identified a population structure of three groups; non-NRCS-A isolates, NRCS-A isolates, and a group of 'proto NRCS-A' - isolates closely related to NRCS-A but not associated with neonatal infection. All bloodstream isolates belonged to the NRCS-A group and were indistinguishable from strains carried on the skin or in the gut. NRCS-A isolates showed increased tolerance to chlorhexidine and antibiotics relative to the other S. capitis as well as enhanced ability to grow at higher pH values. Analysis of the pangenome of 138 isolates identified characteristic nsr and tarJ genes in both the NRCS-A and proto groups. A CRISPR-cas system was only seen in NRCS-A isolates which also showed enrichment of genes for metal acquisition and transport.We found evidence for transmission of S. capitis NRCS-A within NICU, with related isolates shared between babies and multiple acquisitions by some babies. Our data show NRCS-A strains commonly colonise uninfected babies in NICU representing a potential reservoir for potential infection. This work provides more evidence that adaptation to survive in the gut and on skin facilitates spread of NRCS-A, and that metal acquisition and tolerance may be important to the biology of NRCS-A. Understanding how NRCS-A survives in NICUs can help develop infection control procedures against this clone.

Whole-genome sequencing reveals widespread presence of Staphylococcus capitis NRCS-A clone in neonatal units across the United Kingdom.

OBJECTIVE: Increased incidence of neonatal Staphylococcus capitis bacteraemia in summer 2020, London, raised suspicion of widespread multidrug-resistant clone NRCS-A. We set out to investigate the molecular epidemiology of this clone in neonatal units (NNUs) across the UK. METHODS: We conducted whole-genome sequencing (WGS) on presumptive S. capitis NRCS-A isolates collected from infants admitted to nationwide NNUs and from environmental sampling in two distinct NNUs in 2021. Previously published S. capitis genomes were added for comparison. Genetic clusters of NRCS-A isolates were defined based on core-genome single-nucleotide polymorphisms. RESULTS: We analysed WGS data of 838 S. capitis isolates and identified 750 NRCS-A isolates. We discovered a possible UK-specific NRCS-A lineage consisting of 611 isolates collected between 2005 and 2021. We determined 28 genetic clusters of NRCS-A isolates, which covered all geographical regions in the UK, and isolates of 19 genetic clusters were found in ≥2 regions, suggesting inter-regional spread. Within the NRCS-A clone, strong genetic relatedness was identified between contemporary clinical and incubator-associated fomite isolates and between clinical isolates associated with inter-hospital infant transfer. CONCLUSIONS: This WGS-based study confirms the dispersion of S. capitis NRCS-A clone amongst NNUs across the UK and urges research on improving clinical management of neonatal S. capitis infection.

Prevalence, biofilm formation, and mass spectrometric characterization of linezolid-resistant Staphylococcus capitis isolated from a tertiary hospital in China.

OBJECTIVES: Linezolid-resistant Staphylococcus capitis (LRSC) has become a new challenge for clinical anti-infective therapy. The present study aimed to investigate the trends of LRSC prevalence in a tertiary hospital of China 2017-2020. The resistance mechanisms, virulence genes, biofilm formation, and mass spectrometric characteristics of LRSC isolates were also analysed. METHODS: This study retrospectively analysed the antibiotic resistance trends of coagulase negative staphylococci (CoNS) isolated from clinical samples collected between 2017-2020. Antimicrobial resistance profiles were tested by micro-broth dilution and the E-test method. Antimicrobial resistance genes and virulence genes were detected by polymerase chain reaction, and dru-typing sequences were obtained by Sanger sequencing. Crystal violet staining in 96-well plates was used to detect biofilm formation ability. Mass spectrometric characterization of LRSC was analysed by matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) coupled with ClinProTools. RESULTS: The linezolid resistance rate in 3575 CoNS clinical strains was 1.6%, wherein the great majority of was LRSC (91.1%, n = 51/56), with a resistant rate of 15.5% (n = 51/328) in all S. capitis isolates. In this study, 48 out of the 51 LRSC strains and 54 of 277 linezolid-susceptible S. capitis (LSSC) strains were enrolled. G2576T, C2104T, T2130A, C2163T, and T2319C mutations in the 23S rRNA V region and acquisition of cfr were the main linezolid resistant mechanisms in LRSC. The biofilm-forming ability of LRSC was more potent than LSSC, with a higher detection rate of bap (P < 0.05). Eleven mass spectrometric peaks of interest were identified by using MALDI-TOF MS and ClinProTools, which were differently distributed between LRSC and LSSC strains, with the area under the receiver operating characteristic curve of more than 0.8, especially for 5465.37 m/z. CONCLUSIONS: Linezolid resistance was mediated by mutations in the 23S rRNA V region and presence of the cfr gene in LRSC strains. LRSC strains have stronger biofilm-forming ability than LSSC strains, which maybe associated with the adhesion-related gene of bap. Further, linezolid-resistant and linezolid-susceptible S. capitis could be rapidly identified with mass spectrometric characterization. To the best of our knowledge, this study is the first to document the biofilm formation ability of LRSC and the potential usefulness of MALDI-TOF MS for the discrimination of LRSC and LSSC.

Mechanisms of linezolid resistance in Staphylococcus capitis with the novel mutation C2128T in the 23S rRNA gene in China.

PURPOSE: The objective of this study was to investigate the molecular characteristics and potential resistance mechanisms of linezolid-resistant (LZR) Staphylococcus capitis isolates from a tertiary hospital in China. METHODS: S. capitis isolates were obtained from clinical patient specimens; three of the isolates came from blood cultures and one from the hydrothorax. The agar dilution and E-test methods were used to identify antibiotic resistance. The chloramphenicol-florfenicol resistance (cfr) gene carrier status of the strains was determined by PCR. Whole-genome sequencing (WGS) was used to identify point mutations and L3, L4, and L22 mutations and to study the genetic environment of the cfr gene and the relationships between strains. RESULTS: The 4 isolates obtained in this study were all linezolid-resistant Staphylococcus strains. A similar of susceptibility profile pattern was observed in all four S. capitis strains, each of which exhibited a multidrug-resistant phenotype. A potentially novel mutation, C2128T, was identified, and the cfr genes of S. capitis strains were all positive. Additionally, the same mutations (C2128T and G2600T) were identified in all 23S rRNA sequences of the isolates, whereas mutations were lacking in the L3, L4, and L22 ribosomal proteins. The genetic environments surrounding cfr were identical in all four isolates. A schematic diagram of the phylogenetic tree showed that they were closely related to AYP1020, CR01, and TW2795, and a total of seven drug resistance genes were identified in these strains. CONCLUSIONS: The study indicated that the resistance of the Staphylococcus capitis strains to linezolid was caused by multiple mechanisms, and a potential novel mutation, C2128T, that may have an impact on bacterial resistance was identified.

Development of a novel core genome MLST scheme for tracing multidrug resistant Staphylococcus capitis.

Staphylococcus capitis, which causes bloodstream infections in neonatal intensive care units, is a common cause of healthcare-associated infections. Thus, a standardized high-resolution typing method to document the transmission and dissemination of multidrug-resistant S. capitis isolates is required. We aimed to establish a core genome multilocus sequence typing (cgMLST) scheme to surveil S. capitis. The cgMLST scheme was defined based on primary and validation genome sets and tested with outbreaks of linezolid-resistant isolates and a validation set. Phylogenetic analysis was performed to investigate the population structure and compare it with the result of cgMLST analysis. The S. capitis population consists of 1 dominant, NRCS-A, and 4 less common clones. In this work, a multidrug-resistant clone (L clone) with linezolid resistance is identified. With the features of type III SCCmec and multiple copies of mutations of G2576T and C2104T in the 23S rRNA, the L clone has been spreading silently across China.

The epidemiology and molecular characteristics of linezolid-resistant Staphylococcus capitis in Huashan Hospital, Shanghai.

Introduction. Linezolid-resistant (LZR) Staphylococcus capitis has recently emerged in our hospital, and its potential resistance mechanisms are still not clear.Aim. This study aimed to investigate the epidemiology, clinical and genetic characteristics, resistance mechanisms and biofilm formation capacity of LZR S. capitis isolated from patients at Huashan Hospital, Shanghai, PR China between 2012 and 2018.Methodology. Strains were subjected to antimicrobial susceptibility testing (AST) with antibiotics using the broth microdilution method according to the Clinical and Laboratory Standards Institute (CLSI) guidelines. The presence of cfr, optrA and poxtA, as well as mutations in the 23S ribosomal (r)RNA and ribosomal proteins, was investigated using PCR and sequencing techniques. The genetic relationship between isolates was analysed using pulsed-field gel electrophoresis (PFGE) and whole-genome sequencing (WGS). Biofilm biomasses were detected by using crystal violet staining.Results. Twenty-one LZR S. capitis strains displayed MICs of 32-512 µg ml-1. All LZR strains showed G2576T and C2104T mutations in the 23S rRNA V region. Besides G2576T and C2104T, no base mutations were detected in the V region. The cfr was detected in 12 strains, while optrA and poxtA were not amplified in 21 S. capitis strains. PFGE showed that the LZR S. capitis strains belonged to a single clone. The phylogenetic tree showed that 20 LZR S. capitis strains were highly similar to LNZR-1, isolated from Harbin (located in the north of China) in 2013, which showed resistance to linezolid.Conclusions. In this research, cfr-negative strains displayed linezolid MICs of 32 µg ml-1. In comparison, cfr-positive strains exhibited linezolid MICs of 128-512 µg ml-1, indicating that high levels of linezolid resistance appear to be related to the presence of cfr. The outbreak of LZR S. capitis in our hospital needs to be monitored closely.

Detection of sasX Gene and Distribution of SCCmec Types in Invasive and Non-invasive Coagulase-negative Staphylococci

Background: Coagulase-negative staphylococci, which belong to the normal microbiota of the skin and mucous membranes, are opportunistic pathogens. sasX, a newly described protein, is thought to play an important role in nasal colonization and methicillin-resistant Staphylococcus aureus virulence, and it may be acquired from coagulase-negative staphylococci by horizontal gene transfer. It has been considered that understanding the function of sasX gene may help clarify the relevance of the different adhesion mechanisms in the pathogenesis of infections associated with biofilm. Aims: To investigate the sasX gene presence, staphylococcal cassette chromosome mec types, and antimicrobial resistance patterns of invasive and noninvasive coagulase-negative staphylococci isolates. Study Design: Cross-sectional study. Methods: The study included a total of 180 coagulase-negative staphylococci strains. Non-invasive isolates (n=91) were obtained from the hands of healthy volunteers who do not work at the hospital (n=30), the nasal vestibule of healthy volunteer hospital workers (n=26), and central venous catheter (n=35). Invasive isolates (n=89) were isolated from peripheral blood cultures of inpatients who do not have catheters. All isolates were identified by conventional microbiological methods, automated systems, and, if needed, with matrix-assisted laser desorption/ionization-time of flight. Staphylococcal cassette chromosome mec typing, sasX and mec gene detection, antibiotic susceptibility, and sasX gene sequence analysis were performed. Results: Peripheral blood, central venous catheter colonization, and nasal vestibule isolates were positive for the sasX gene, whereas hand isolates were negative. sasX gene was present in 17 isolates, and no statistical significance was found between invasive and noninvasive isolates (p=0.173). Sequence analysis of the sasX genes showed high homology to related proteins of Staphylococcus phage SPbeta-like and Staphylococcus epidermidis RP62A. staphylococcal cassette chromosome mec type V was the most prevalent regardless of species. staphylococcal cassette chromosome mec type II was more frequent in invasive isolates and found to be statistically important for invasive and noninvasive S. epidermidis isolates (p=0.029). Staphylococcus haemolyticus isolates had the overall highest resistance rates. Resistance to ciprofloxacin, trimethoprim-sulfamethoxazole, and erythromycin was found to be higher in isolates from catheter and blood culture. Staphylococcus hominis isolates had the highest rate for inducible clindamycin resistance. None of the isolates were resistant to vancomycin, teicoplanin, and linezolid. Conclusion: The sasX gene is detected in 9.44% of the isolates. There is no statistical difference between the sasX-positive and -negative isolates in terms of antibacterial resistance and the presence of sasX and SCCmec types. Further studies about the role of sasX at virulence in coagulase-negative staphylococci, especially from clinical samples such as tracheal aspirate and abscess isolates, and distribution of staphylococcal cassette chromosome mec types are needed.

Niche specialization and spread of Staphylococcus capitis involved in neonatal sepsis.

The multidrug-resistant Staphylococcus capitis NRCS-A clone is responsible for sepsis in preterm infants in neonatal intensive care units (NICUs) worldwide. Here, to retrace the spread of this clone and to identify drivers of its specific success, we investigated a representative collection of 250 S. capitis isolates from adults and newborns. Bayesian analyses confirmed the spread of the NRCS-A clone and enabled us to date its emergence in the late 1960s and its expansion during the 1980s, coinciding with the establishment of NICUs and the increasing use of vancomycin in these units, respectively. This dynamic was accompanied by the acquisition of mutations in antimicrobial resistance- and bacteriocin-encoding genes. Furthermore, combined statistical tools and a genome-wide association study convergently point to vancomycin resistance as a major driver of NRCS-A success. We also identified another S. capitis subclade (alpha clade) that emerged independently, showing parallel evolution towards NICU specialization and non-susceptibility to vancomycin, indicating convergent evolution in NICU-associated pathogens. These findings illustrate how the broad use of antibiotics can repeatedly lead initially commensal drug-susceptible bacteria to evolve into multidrug-resistant clones that are able to successfully spread worldwide and become pathogenic for highly vulnerable patients.

Antimicrobial Resistance Profiles and Molecular Characteristics of Coagulase-Negative Staphylococci Isolated from Two Tertiary Hospitals Before and 15 Years After Implementation of the Separation of Drug Prescribing and Dispensing Policy of Korea.

This study compared changes in antimicrobial susceptibilities and molecular characteristics of coagulase-negative staphylococci (CNS) between the year 2000 and the year 2014-2015 to evaluate the policy of separating drug prescribing and dispensing in Korea. We obtained 68 CNS clinical isolates from two tertiary general hospitals before (the year 2000; n = 25) and after (the year 2014 - 2015; n = 43) implementation of the separation. Isolates were identified as Staphylococcus capitis, Staphylococcus epidermidis, Staphylococcus haemolyticus, Staphylococcus hominis, Staphylococcus saprophyticus, and Staphylococcus warneri. When minimal inhibitory concentrations of 14 antimicrobials were applied to isolates, resistance rates to gentamicin and oxacillin in 2000 were significantly higher than in 2014-2015 (p < 0.05). Fifty-seven isolates were methicillin-resistant CNS (MR-CNS), 42 of which were also multidrug resistant; overall, multidrug resistance decreased from 72% in the year 2000 to 55.8% in 2014-2015. Staphylococcal cassette chromosome mec (SCCmec) type III was the dominant type of MR-CNS in the year 2000, while SCCmec type IV was the dominant type in 2014-2015. Twenty-five sequence types (STs) were identified; ST2 appeared most frequently in both periods. After 15 years of implementation of this policy, multidrug resistance as well as methicillin and gentamicin resistance in CNS decreased, but not resistance to other antibiotics. Long-term surveillance at both genotypic and phenotypic levels of various species is necessary for further evaluation of this policy.

Nisin J, a Novel Natural Nisin Variant, Is Produced by Staphylococcus capitis Sourced from the Human Skin Microbiota.

The skin microbiota is thought to play a key role in host protection from infection. Nisin J is a novel nisin variant produced by Staphylococcus capitis APC 2923, a strain isolated from the toe web space area in a screening study performed on the human skin microbiota. Whole-genome sequencing and mass spectrometry of the purified peptide confirmed that S. capitis APC 2923 produces a 3,458-Da bacteriocin, designated nisin J, which exhibited antimicrobial activity against a range of Gram-positive pathogens, including methicillin-resistant Staphylococcus aureus (MRSA) and Cutibacterium acnes The gene order in the nisin J gene cluster (nsjFEGBTCJP) differs from that of other nisin variants in that it is lacking the nisin regulatory genes, nisRK, as well as the nisin immunity gene nisI Nisin J has 9 amino acid changes compared to prototypical nisin A, with 8 amino acid substitutions, 6 of which are not present in other nisin variants (Ile4Lys, Met17Gln, Gly18Thr, Asn20Phe, Met21Ala, Ile30Gly, Val33His, and Lys34Thr), and an extra amino acid close to the C terminus, rendering nisin J the only nisin variant to contain 35 amino acids. This is the first report of a nisin variant produced by a Staphylococcus species and the first nisin producer isolated from human skin.IMPORTANCE This study describes the characterization of nisin J, the first example of a natural nisin variant, produced by a human skin isolate of staphylococcal origin. Nisin J displays inhibitory activity against a wide range of bacterial targets, including MRSA. This work demonstrates the potential of human commensals as a source for novel antimicrobials that could form part of the solution to antibiotic resistance across a broad range of bacterial pathogens.

Identification and characterisation of capidermicin, a novel bacteriocin produced by Staphylococcus capitis.

One hundred human-derived coagulase negative staphylococci (CoNS) were screened for antimicrobial activity using agar-based deferred antagonism assays with a range of indicator bacteria. Based on the findings of the screen and subsequent well assays with cell free supernatants and whole cell extracts, one strain, designated CIT060, was selected for further investigation. It was identified as Staphylococcus capitis and herein we describe the purification and characterisation of the novel bacteriocin that the strain produces. This bacteriocin which we have named capidermicin was extracted from the cell-free supernatant of S. capitis CIT060 and purified to homogeneity using reversed-phase high performance liquid chromatography (RP-HPLC). Matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometric (MS) analysis revealed that the capidermicin peptide has a mass of 5,464 Da. Minimal inhibitory concentration (MIC) experiments showed that capidermicin was active in the micro-molar range against all the Gram-positive bacteria that were tested. Antimicrobial activity was retained over a range of pHs (2-11) and temperatures (10-121°C x 15 mins). The draft genome sequence of S. capitis CIT060 was determined and the genes predicted to be involved in the biosynthesis of capidermicin were identified. These genes included the predicted capidermicin precursor gene, and genes that are predicted to encode a membrane transporter, an immunity protein and a transcriptional regulator. Homology searches suggest that capidermicin is a novel member of the family of class II leaderless bacteriocins.

Genomic analysis of Staphylococcus capitis isolated from blood cultures in neonates at a neonatal intensive care unit in Sweden.

Emergence of a genetically distinct, multidrug-resistant Staphylococcus capitis clone (NRCS-A) present in neonatal intensive care units has recently been extensively reported. The aims of the present study were to investigate which clones of S. capitis isolated from blood in a Swedish neonatal intensive care unit (NICU) have been present since 1987 and to investigate whether the NRCS-A clone has disseminated in Sweden. All S. capitis isolates from blood cultures of neonates (≤ 28 days of age) between 1987 and 2017 (n = 46) were whole-genome sequenced, and core genome multilocus sequence typing (cgMLST) was performed. Single-nucleotide polymorphism (SNP)-based phylogenetic relationships between the S. capitis isolates and in silico predictions of presence of genetic traits specific to the NRCS-A clone were identified. Furthermore, antibiotic susceptibility testing, including screening for heterogeneous glycopeptide-intermediate resistance, was performed. Thirty-five isolates clustered closely to the isolates previously determined as belonging to the NRCS-A clone and had fewer than 81 core genome loci differences out of 1063. Twenty-one of these isolates were multidrug resistant. The NRCS-A clone was found in 2001. Six pairs of isolates had differences of fewer than two SNPs. Genetic traits associated with the NRCS-A clone such as nsr, ebh, tarJ, and CRISPR were found in all 35 isolates. The increasing incidence of S. capitis blood cultures of neonates is predominantly represented by the NRSC-A clone at our NICU in Sweden. Furthermore, there were indications of transmission between cases; adherence to basic hygiene procedures and surveillance measures are thus warranted.

Molecular characterization of multi-drug resistant coagulase negative cocci in non-hospital environment.

Antibiotic resistance crisis occasioned by sporadic appearance of multi-drug resistance (MDR) in human pathogens to clinically applied antimicrobials is a serious threat to global health. In this study, we investigated the drug resistant phenotype of Gram-positive cocci isolates from environment. Staphylococcus capitis and Staphylococcus haemolyticus colonies were isolated on mannitol-salt agar plates supplemented with tetracycline. Antibiotic susceptibility profile of the isolates via minimum inhibitory concentration (MIC) determination was examined. Isolates showed decreased sensitivity to clinically applied antimicrobial agents: tetracycline, kanamycin, erythromycin, norfloxacin, teicoplanin, and ampicillin. Genomic analysis demonstrated the presence of multiple antibiotic resistant genes in these bacteria, suggesting the origin of the multiple antimicrobials resistant phenotype. Tetracycline resistance of these isolates was transduced to Staphylococcus aureus-RN4220 strain. These findings indicate the presence of multiple antimicrobials resistant S. capitis and S. haemolyticus strain in a non-hospital setting. Moreover, the presence of plethora of genes responsible for MDR suggest that these strains could present potential threat to human health by serving as reservoir for lateral transference of antimicrobial resistance conferring foreign genetic elements to other clinically relevant pathogens.

Staphylococcus capitis and NRCS-A clone: the story of an unrecognized pathogen in neonatal intensive care units.

BACKGROUND: In neonatal intensive care units (NICUs), nosocomial late-onset sepsis (LOS), mostly due to coagulase negative staphylococci, constitute a major cause of death or impairment. Staphylococcus capitis, usually considered as a poorly virulent species, has been reported as a cause of LOS. OBJECTIVES: To review data regarding S. capitis neonatal LOS and the features of isolates involved. SOURCES: PubMed was searched up to August 2018 to retrieve studies on the topic; the keywords used were 'S. capitis', 'neonate', 'neonatal ICU', 'bloodstream infection' and 'late onset sepsis'. CONTENT: Published data highlight the worldwide endemicity of a single S. capitis clone, named NRCS-A, specifically involved in LOS. NRCS-A harbours a multidrug resistance profile (including resistance to the usual first-line antibiotics used in NICUs). It is also able to adapt under vancomycin selective pressure that could confer an advantage for its implantation and dissemination in NICUs where this selective pressure is high. Moreover, a severe morbidity has been observed in NRCS-A-related LOS. The NICU environment, and especially incubators, constitute reservoirs of NRCS-A from which it could diffuse inside the setting. Finally, the virulome and resistome of S. capitis NRCS-A contain many genes potentially implicated in its specific epidemiology and pathophysiology, including the gene nsr that may be involved in its fitness and implantation in neonatal gut flora. IMPLICATIONS: S. capitis must be considered as a true pathogen in neonates. The decreased susceptibility to vancomycin may be involved in failure of vancomycin therapy. Further studies are needed to better manage its diffusion inside each NICU but also worldwide.

Evolution of Daptomycin Resistance in Coagulase-Negative Staphylococci Involves Mutations of the Essential Two-Component Regulator WalKR.

Coagulase-negative staphylococci (CoNS) represent one of the major causes of health care- and medical device-associated infections. Emerging antimicrobial resistance has complicated the treatment of systemic infections caused by CoNS. Here, we describe the prevalence of antimicrobial resistance in clinical CoNS strains from a tertiary care hospital over a 4-year period, and we observed a significant increase in resistance to daptomycin. Notably, Staphylococcus capitis accounted for the majority of these daptomycin-resistant (DAP-R) CoNS. To further investigate the mechanisms of daptomycin resistance in CoNS, daptomycin-susceptible clinical strains of S. capitis and Staphylococcus epidermidis underwent in vitro daptomycin exposure to generate DAP-R CoNS mutants. Unlike that seen with Staphylococcus aureus, alteration of cell surface charge was not observed in the DAP-R CoNS strains, but biofilm formation was compromised. Whole-genome sequencing analysis of the DAP-R CoNS strains identified single nucleotide polymorphisms (SNPs) in walKR, the essential two-component regulatory system controlling cell wall biogenesis. PCR and sequencing of walK and walR from 17 DAP-R CoNS clinical isolates identified seven nonsynonymous mutations. The results were confirmed by the recreation of the walK SNP in S. epidermidis, which resulted in reduced susceptibility to daptomycin and vancomycin. This study highlights the significance of CoNS in evolving daptomycin resistance and showed that walKR is shared among the staphylococcal species and is involved in antibiotic resistance development. Notably, we did not observe mutations in genes responsible for phospholipid biosynthesis or an altered cell surface charge, suggesting that reduced daptomycin susceptibility in CoNS may emerge in a fashion distinct from that in S. aureus.

Complete genome sequencing of three human clinical isolates of Staphylococcus caprae reveals virulence factors similar to those of S. epidermidis and S. capitis.

BACKGROUND: Staphylococcus caprae is an animal-associated bacterium regarded as part of goats' microflora. Recently, S. caprae has been reported to cause human nosocomial infections such as bacteremia and bone and joint infections. However, the mechanisms responsible for the development of nosocomial infections remain largely unknown. Moreover, the complete genome sequence of S. caprae has not been determined. RESULTS: We determined the complete genome sequences of three methicillin-resistant S. caprae strains isolated from humans and compared these sequences with the genomes of S. epidermidis and S. capitis, both of which are closely related to S. caprae and are inhabitants of human skin capable of causing opportunistic infections. The genomes showed that S. caprae JMUB145, JMUB590, and JMUB898 strains contained circular chromosomes of 2,618,380, 2,629,173, and 2,598,513 bp, respectively. JMUB145 carried type V SCCmec, while JMUB590 and JMUB898 had type IVa SCCmec. A genome-wide phylogenetic SNP tree constructed using 83 complete genome sequences of 24 Staphylococcus species and 2 S. caprae draft genome sequences confirmed that S. caprae is most closely related to S. epidermidis and S. capitis. Comparative complete genome analysis of eight S. epidermidis, three S. capitis and three S. caprae strains revealed that they shared similar virulence factors represented by biofilm formation genes. These factors include wall teichoic acid synthesis genes, poly-gamma-DL-glutamic acid capsule synthesis genes, and other genes encoding nonproteinaceous adhesins. The 17 proteinases/adhesins and extracellular proteins known to be associated with biofilm formation in S. epidermidis were also conserved in these three species, and their biofilm formation could be detected in vitro. Moreover, two virulence-associated gene clusters, the type VII secretion system and capsular polysaccharide biosynthesis gene clusters, identified in S. aureus were present in S. caprae but not in S. epidermidis and S. capitis genomes. CONCLUSION: The complete genome sequences of three methicillin-resistant S. caprae isolates from humans were determined for the first time. Comparative genome analysis revealed that S. caprae is closely related to S. epidermidis and S. capitis at the species level, especially in the ability to form biofilms, which may lead to increased virulence during the development of S. caprae infections.

Genomic Analysis of Multiresistant Staphylococcus capitis Associated with Neonatal Sepsis.

Coagulase-negative staphylococci (CoNS), such as Staphylococcus capitis, are major causes of bloodstream infections in neonatal intensive care units (NICUs). Recently, a distinct clone of S. capitis (designated S. capitis NRCS-A) has emerged as an important pathogen in NICUs internationally. Here, 122 S. capitis isolates from New Zealand (NZ) underwent whole-genome sequencing (WGS), and these data were supplemented with publicly available S. capitis sequence reads. Phylogenetic and comparative genomic analyses were performed, as were phenotypic assessments of antimicrobial resistance, biofilm formation, and plasmid segregational stability on representative isolates. A distinct lineage of S. capitis was identified in NZ associated with neonates and the NICU environment. Isolates from this lineage produced increased levels of biofilm, displayed higher levels of tolerance to chlorhexidine, and were multidrug resistant. Although similar to globally circulating NICU-associated S. capitis strains at a core-genome level, NZ NICU S. capitis isolates carried a novel stably maintained multidrug-resistant plasmid that was not present in non-NICU isolates. Neonatal blood culture isolates were indistinguishable from environmental S. capitis isolates found on fomites, such as stethoscopes and neonatal incubators, but were generally distinct from those isolates carried by NICU staff. This work implicates the NICU environment as a potential reservoir for neonatal sepsis caused by S. capitis and highlights the capacity of genomics-based tracking and surveillance to inform future hospital infection control practices aimed at containing the spread of this important neonatal pathogen.

Wide dissemination of SCCfusC in fusidic acid-resistant coagulase-negative staphylococci and implication for its spread to methicillin-resistant staphylococcus aureus in Taiwan.

The fusidic acid (FUS) resistance determinants fusB, fusC, fusD and fusF in coagulase-negative staphylococci (CoNS) clinical isolates were examined. Among 208 FUS-resistant isolates, the fusB gene was the most common resistance determinant in each species, except in Staphylococcus hominis subsp. hominis or in species carrying intrinsic fusD or fusF. In S. hominis subsp. hominis, the fusC gene was the major determinant responsible for FUS resistance. To understand the genetic context of fusC in S. hominis subsp. hominis, 31 fusC-positive S. hominis subsp. hominis isolates were examined. Among these isolates, 14 carried SCCfusC, 3 carried an SCC476-like element and 7 carried a new SCC structure (SCC3390). As shown by pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST) analyses, the S. hominis subsp. hominis clinical isolates showed limited clonality. Taken together, SCCfusC has been found in S. hominis subsp. hominis, Staphylococcus epidermidis, Staphylococcus haemolyticus, Staphylococcus capitis subsp. ureolyticus and Staphylococcus aureus, suggesting its wide distribution and spread among different species of staphylococci.

Phenotypic characterisation of coagulase-negative staphylococci isolated from blood cultures in newborn infants, with a special focus on Staphylococcus capitis.

AIM: This Swedish study determined which species of coagulase-negative staphylococci (CoNS) were found in neonatal blood cultures and whether they included Staphylococcus capitis clones with decreased susceptibility to vancomycin. METHODS: CoNS isolates (n = 332) from neonatal blood cultures collected at Örebro University Hospital during 1987-2014 were identified to species level with matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOF MS). The antibiotic susceptibility pattern of S. capitis isolates was determined by the disc diffusion test and Etest, and the presence of heterogeneous glycopeptide-intermediate S. capitis (hGISC) was evaluated. RESULTS: Staphylococcus epidermidis (67.4%), Staphylococcus haemolyticus (10.5%) and S. capitis (9.6%) were the most common CoNS species. Of the S. capitis isolates, 75% were methicillin-resistant and 44% were multidrug-resistant. No isolate showed decreased susceptibility to vancomycin, but at least 59% displayed the hGISC phenotype. Staphylococcus capitis isolates related to the strain CR01 displaying pulsotype NRCS-A were found. CONCLUSION: Staphylococcus epidermidis, S. haemolyticus and S. capitis were the predominant species detected in neonatal blood cultures by MALDI-TOF MS. The number of episodes caused by S. capitis increased during the study period, but no isolates with decreased susceptibility to vancomycin were identified. However, S. capitis isolates related to the strain CR01 displaying pulsotype NRCS-A were found.