Coagulase-Negative Staphylococci and Micrococcaceae

Coagulase-Negative Staphylococci

Staphylococcus epidermidis and other coagulase-negative staphylococci (CoNS), once considered nonpathogenic commensal organisms, now are recognized clearly as potential true pathogens, particularly in hospitalized patients and those with indwelling foreign bodies. Multiple surveys have shown that CoNS are the most frequent cause of nosocomial bloodstream infections (BSIs) around the globe. These organisms are implicated particularly in bacteremic disease in patients in the intensive care unit (ICU) and in patients immunocompromised by cancer or solid-organ transplantation. ^, CoNS account for 20%–50% of all nosocomial BSIs in these patient groups. ^,

Description of The Pathogen

Staphylococci are members of the family Staphylococcaceae. They are nonmotile, non–spore-forming, catalase-positive bacteria that stain as gram-positive cocci and grow in irregular clusters. Staphylococci are among the hardiest of non–spore-forming bacteria and can survive many nonphysiologic environmental conditions. Colonies are round, smooth, raised, and glistening and can be surrounded by a zone of hemolysis when grown on blood-containing media.

In the clinical microbiology laboratory, CoNS are distinguished from Staphylococcus aureus by their failure to produce coagulase, an enzyme that coagulates rabbit plasma. Originally, all CoNS were grouped together under the designation Staphylococcus albus. By 2015, however, nearly 40 species and numerous subspecies of CoNS had been recognized based on biochemical and genetic differences. New species are regularly characterized and others are reclassified, resulting in a fluid and complex taxonomy. Fifteen species are normal human flora: S. epidermidis, S. haemolyticus, S. hominis, S. saccharolyticus, S. capitis, S. warneri, S. caprae, S. pasteuri, S. saprophyticus, S. cohnii, S. xylosus, S. simulans, S. auricularis, S. lugdunensis, and S. schleiferi . All CoNS primarily are skin commensals, but they can colonize the upper respiratory tract, gastrointestinal tract, genitourinary tract, and mammary glands. Some species exhibit preferential niches for certain areas of the skin, such as S. capitis for the sebaceous gland-rich scalp, S. auricularis for the external auditory canal, S. hominis and S. haemolyticus for the apocrine gland-rich axillae and pubis, and S. saprophyticus for the female genitourinary tract.

Many clinical microbiology laboratories continue to characterize isolates simply as CoNS. However, commercially available kits can rapidly distinguish between CoNS species with an accuracy of 70% to >90%. Most kits include a series of biochemical reactions, although some systems are based on polymerase chain reaction amplification of ribosomal ribonucleic acid (rRNA) and other conserved genes. More recently, investigators have identified CoNS species using matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). Species identification of S. lugdunensis and S. schleiferi is particularly important because these species cause aggressive disease more typical of S. aureus . In the laboratory they can be distinguished from most other CoNS because they produce a positive result on a rapid coagulase slide test, reflecting the presence of the bound coagulase (clumping factor) rather than free coagulase; similar to other CoNS, however, they test negative with the more definitive coagulase tube assay. Both S. lugdunensis and S. schleiferi are relatively uncommon on normal human skin, but they can colonize implanted materials, such as catheters and drains. S. lugdunensis has been implicated in native and prosthetic valve endocarditis, septicemia, brain abscess, peritonitis, soft tissue infection, vascular graft infection, prosthetic joint infection, and catheter-related infection. S. schleiferi rarely has been reported as a pathogen in the US, but in some European countries S. schleiferi has caused brain empyema, wound infection, osteomyelitis, and BSI. ^{, ,}

Pathogenesis

Colonization is a prerequisite for CoNS disease. CoNS can be isolated from multiple skin surfaces, which suggests that most infections originate from organisms on the skin. Patients who develop a CoNS infection typically have an obvious disruption of the skin barrier, caused by surgery, placement of a catheter, or insertion of a prosthesis. Systemic infection also can originate from colonized mucous membranes of the pharynx and gastrointestinal tract when organisms enter the bloodstream, either through breaks in the mucosa or by translocation through mesenteric lymph nodes.

Unlike S. aureus, CoNS do not elaborate virulent exotoxins. Consequently, infections caused by most CoNS characteristically are indolent and prolonged, even in compromised patients. The prominence of CoNS as nosocomial pathogens stems largely from their propensity to adhere to the inanimate surfaces of catheters and prostheses and also in their ability, once established, to evade host defense mechanisms and antibiotic activity. Both characteristics are mediated by the production of biofilm, a thick accumulation of bacteria arranged in multilayered clusters that are covered by an extracellular matrix. Indeed, CoNS isolated from the hospital environment typically are biofilm producers, whereas the less disease-associated CoNS isolated from the community are not. The importance of biofilm in CoNS pathogenesis has been confirmed repeatedly by demonstrations that mutants deficient in biofilm-related molecules are less likely to cause disease than their isogenic counterparts. ^, The formation of biofilm occurs in three overlapping stages: attachment of the CoNS to polymer surfaces; maturation of the organisms within the biofilm to form a complex, multilayered structure; and detachment, which allows the organisms within the biofilm to disseminate to distant sites ( Fig. 116.1 ). ^, In the first stage, nonspecific factors, including long-range electromagnetic forces and surface hydrophobicity, promote attachment. ^, Several bacteria-specific products mediate this seminal stage of attachment. Chief among these is the surface-associated autolysin AtlE, which shows significant homology to the major autolysin of S. aureus . AtlE mediates binding to unmodified plastic surfaces indirectly, by altering the hydrophobicity of the bacterial cell wall. ^, Two related staphylococcal surface proteins, SSP-1 and SSP-2, are fimbria-like polymers encoded by S. epidermidis that further mediate adherence to polymers. ^, Once inserted into the body, plastic surfaces become coated with plasma and extracellular host proteins such as fibronectin, fibrinogen, vitronectin, thrombospondin, and von Willebrand factor, and bacterial molecules also bind these host molecules ( Fig. 116.1 ). Several such molecules have been characterized; they are known collectively as microbial surface components recognizing adhesive matrix molecules (MSCRAMMs). ^{, ,}

After attachment and during the maturation phase, organisms accumulate and form multilayered aggregates; this is the critical stage of biofilm formation ( Fig. 116.1 ). In 1994, Mack and coworkers found that an S. epidermidis transposon mutant that lacked a polysaccharide antigen, called polysaccharide intercellular adhesin (PIA), was unable to form multilayered cell clusters. PIA since has been confirmed as the central molecule in CoNS biofilm production. PIA is encoded by the icaADBC gene cluster, which is responsible for its synthesis and modification; mutation of any of the four open reading frames in the ica gene locus interferes with maturation of the CoNS biofilm. ^, The control of ica expression is incompletely understood, but is influenced by several negative and positive regulators, which presumably promote the production of biofilm under specific environmental conditions. ^{, ,} However, ica -negative CoNS that produce biofilm have been identified, indicating the existence of PIA-independent mechanisms of biofilm formation. Three candidates are a 140-kd extracellular protein called accumulation-associated protein (Aap), ^, extracellular matrix binding protein (Embp), and extracellular deoxyribonucleic acid (eDNA) released from lysed bacteria. All three of these also may participate in the first (attachment) phase of biofilm formation. A fourth molecule, termed biofilm associated protein (Bap), has been associated with biofilm production among some CoNS species that lack the ica operon, but its importance is less well established. These additional molecules add redundancy to the synthetic mechanisms of CoNS, mediating accumulation in organisms with a compromised ability to express PIA.

After attachment and maturation, most biofilm-producing isolates elaborate copious amounts of a complex extracellular material previously referred to as slime. In a hospital-wide survey, Ishak and associates found slime production in 13 of 14 clinically significant bloodstream isolates of S. epidermidis, but in only 3 of 13 blood culture contaminants and 4 of 27 skin isolates. Studies of infants with CoNS infection have yielded similar results. In independent studies, slime production was noted in 82% of isolates from infants with invasive disease.

The third phase of biofilm formation, detachment, is mediated by the formation of channels within the structure that allow the resident bacteria access to nutrients. Quorum-sensing mechanisms in CoNS recognize the bacterial density in the biofilm and convert the replicating populations to a more stationary phase. Additionally, these mechanisms activate the expression of a family of surfactants called phenol-soluble modulins (PSMs), which affect the volume of biofilm and the formation of channels. Under flow conditions, such biofilms detach, allowing dissemination of organisms to distant sites. CoNS with disrupted PSM function form biofilms that are more compact and tightly adherent to the plastic surface than the biofilms produced by CoNS with functional PSMs; however, the ability of the former to spread is diminished.

Several biofilm-related molecules are implicated in immune evasion. PIA may render the organism more resistant to neutrophil clearance; S. epidermidis organisms that contain ica gene mutations and deficient PIA are phagocytosed more readily in vitro and are more susceptible to the toxic effects of neutrophil granules than their isogenic correlates. ^, These antiphagocytic properties may make the organism particularly pathogenic in premature infants, who have depressed neutrophil function compared with that of term infants and adults. ^, The biofilm further functions as a nonspecific physical barrier to cellular and humoral defense mechanisms. In addition, biofilm inhibits neutrophil chemotaxis and phagocytosis and suppresses lymphocyte blastogenesis. ^, Crude extracts of biofilm can inhibit the antimicrobial action of the glycopeptide antibiotics vancomycin and teicoplanin. This effect appears to be limited to the glycopeptide class of antibiotics because extracts produce no change in the minimal inhibitory concentration (MIC) of cefazolin, clindamycin, or rifampin.

CoNS also elaborate a variety of exoproteins. Examples include urease, lipase/esterase, fibrinolysin, DNase, and a number of proteases. For the most part the specific function of these bacterial products remains undetermined. In addition, some isolates produce δ-like toxin, an extracellular hemolysin that resembles the enteropathic δ-toxin of S. aureus in size, biologic properties, and antigenicity. This δ-like toxin has cytotoxic effects on intestinal epithelium, and investigators have correlated free toxin in the stool of premature infants with the development of necrotizing enterocolitis (NEC). S. epidermidis further produces a number of lantibiotics, which are antibiotic peptides containing the unusual amino acids lanthionine or methyllanthionine that can suppress the growth of competing bacteria on skin and mucous membrane surfaces.

S. saprophyticus elaborates a number of factors that may contribute to its ability to cause urinary tract infections (UTIs). A hemagglutinin/adhesin expressed on the surface of S. saprophyticus and the surface fibrillar protein SSP both bind to uroepithelial cells, which may explain the peculiar tropism of this species for the urinary tract. S. saprophyticus further elaborates a urease, which damages bladder tissue once infection has been established.

Epidemiology

CoNS are acquired early in life; virtually all infants carry CoNS at multiple sites by 2–4 days of age. Colonization among healthy people remains ubiquitous beyond the newborn period. Many CoNS responsible for community-associated infections previously were methicillin susceptible, but community strains have become increasingly resistant. ^, In one Japanese study, nearly one-third of healthy children had nasal colonization with methicillin-resistant CoNS. Although community CoNS have a diverse lineage, a substantial proportion carry the same staphylococcal cassette chromosome mec (SCC mec ) IV genetic element, which is responsible for methicillin resistance in community-associated methicillin-resistant S. aureus . ^,

Accumulating evidence suggests that, in contrast to the diversity of community-derived CoNS strains, one or more CoNS strains in an individual hospital unit can become predominant over a protracted period. Genetically identical or strongly related organisms have been isolated from a number of patients in an ICU, a hematology-oncology unit, a dialysis unit, and even among several wards in a particular hospital. ^, Dominant strains can circulate in affected units for many years. Remarkably, a recent study employing whole genome analysis has determined that some lineages of CoNS have spread to neonatal units across the globe. The hands of nurses become colonized by hospital-associated CoNS strains soon after employment, ^, a finding highlighting the importance of personnel in patient-to-patient transmission. The biologic properties that confer strain predominance are not known, but interference with the growth of surrounding organisms and superior adherence to colonizing surfaces have been speculated to provide a relative survival advantage.

Clinical Manifestations