Enterococci

Enterococci (from Greek enteron = intestine, small intestine) are gram-positive, catalase-negative, facultatively anaerobic bacteria with a spherical morphology. In the microscopic image they usually appear as short chains, more rarely in pairs. They belong to Lancefield serogroup D, but are listed as a separate genus(Enterococcaceae). More than 30 species are described. Their name is derived from their natural habitat; this is primarily the intestinal tract of numerous mammals, including humans, as well as the digestive systems of many species of birds, reptiles, fish, and insects. However, most enterococci can also survive in the environment. For example, enterococci are frequently isolated from soils where livestock are raised and from sewage (Klare I et al.2003; Iversen A et al. 2002).

The most important representatives are:

• Enterococcus faecalis
• Enterococcus durans
• Enterococcus faecium
• Enterococcus casseliflavus
• Enterococcus gallinarum

Enterococcus faecalis and Enterococcus faecium account for up to 50% of the aerobic intestinal flora on diets high in fiber and carbohydrates and low in fat and protein.

Enterococci show no or incomplete hemolysis (α-hemolysis) on solid blood-containing culture media and, with few exceptions, possess a common antigen that belongs to group D according to the so-called Lancefield serology. Although enterococci are generally considered to be rather weakly virulent, they are among the most clinically significant pathogens. This is due on the one hand to their marked resistance to changing environmental factors (enterococci are able to grow between 10 and 45 0 C and are insensitive to bile secretions, alkaline pH values (growth still occurs at pH values of up to 9.6) as well as high saline concentrations (Werner G et al.2008). These allow selective enrichment of these bacteria in the presence of 6.5% sodium chloride. In addition, enterococci are relatively thermotolerant and, unlike many other bacterial pathogens, survive thermal treatment at 60 0C (for 30 minutes). On the other hand, the medical importance of enterococci stems from the resistance properties of these microbes to antibacterial anti-infectives. All enterococci possess numerous natural anti-infective resistances, some of which differ from species to species. In addition to this species-dependent natural multi-resistance, many strains also possess a number of acquired resistance properties.

With few exceptions, enterococcal diseases arise endogenously. They arise from intestinal colonization of humans with these pathogens. Antibacterial therapy of other diseases, in particular the oral use of cephalosporins, carbapenems and quinolones, is an important predisposing factor, since these anti-infectives select enterococci particularly frequently (Tacconelli E et al.2008).

In intensive care units of German hospitals, enterococci are considered to be the fourth most frequent bacterial pathogen of nosocomial diseases , and in German hospitals without taking the ward into account, even the second to third most frequent (Miro JM et al.2008; Klare I et al.2008; Geffers C et al.2004). In recent years, a further increasing incidence of hospital-associated severe enterococcal disease has been observed in many European countries. The main reasons for the increasing incidence of these diseases include the steadily growing proportion of elderly and/or immunosuppressed patients and thus the increasing number of patients requiring intensive care, which usually require broad and frequent administration of anti-infectives. However, the most commonly used agents have little or no efficacy against enterococci. The use of many antibacterial anti-infectives may result in selection of enterococci, as they have natural and acquired resistance properties to numerous agents. In particular, antibacterial therapy of severe nosocomial diseases caused by multidrug-resistant Enterococcus faecium strains is a special and increasingly important medical challenge in clinical practice.

E. faecalis and E. faecium are responsible for a number of different diseases and infections. These primarily include urinary tract infections, sepsis, endocarditis, peritonitis, intra-abdominal abscesses, catheter-associated infections, and wound infections.

Most illnesses are acquired exclusively or primarily in the hospital(nosocomial infection) and often affect patients with immunosuppression, severe underlying diseases, intra-abdominal or cardiothoracic surgery, permanent catheters, and long hospital stays (Robert Koch Institute 2008). Individuals on multilayered antibacterial therapy and the elderly are also frequently affected Enterococcaceae.Of particular clinical importance are enterococci as pathogens of urinary tract infections. After Escherichia coli, they are the most common pathogens of nosocomially acquired urinary tract infections. It is estimated that 10 to 20% of all acute ur inary tract infections and in many cases more than 50% of all such chronic infections are caused by these Enterococcaceae.

It is also estimated that 3 to 15% of all endocarditis and up to 30% of septic infections in adults are caused by enterococci. Endocarditis caused by enterococci is associated with a high lethality, but this is partly due to the severe underlying diseases occurring simultaneously in affected individuals. Medically, sepsis caused by E. faecium is particularly significant because it is associated with a higher mortality rate than corresponding E. faecalis sepsis(Ghanem G et al.2007).

Diseases caused by enterococci should always be treated with antibacterial therapy. Due to the numerous natural resistances to antibacterial agents and the worrying epidemiological situation of acquired anti-infective resistance, the therapy of severe nosocomial enterococcal diseases in particular, which are often associated with a high mortality rate, should be strictly based on the antibiogram and an identification of the pathogen should be carried out. If necessary, the initial therapy, which must be carefully calculated, must be adapted as quickly as possible to the results obtained from the susceptibility tests.

Aminopenicillins are considered standard therapeutic agents for mild urinary tract infections with E. faecalis. For severe disease caused by E. faecalis, especially endocarditis, administration of an aminopenicillin or ureidopenicillin (especially mezlocillin) in combination with an aminoglycoside (usually gentamicin) is considered first-line therapy.

Vancomycin-resistant strains: Extremely problematic is the therapy of severe diseases caused by vancomycin-resistant E. faecium strains, since such pathogens are usually also aminopenicillin-resistant and often also show "high resistance" to aminoglycosides and other secondary resistances. For antibacterial therapy of such diseases, linezolid and tigecycline appear to have good efficacy in severe disease caused by vancomycin-resistant E. faecium strains. Daptomycin (lipopeptide), also shows good in vitro efficacy on vancomycin-resistant enterococci (Sader HS et al. 2007).

Numerous pathogenicity factors are involved in the pathogenesis of diseases caused by enterococci, the formation of which depends in part on the individual strain. Most of these factors serve to promote adhesion, establishment, and spread of the pathogens in tissues. Similar to staphylococci and some streptococci, many strains of enterococci are capable of forming biofilms. Pathogenicity factors include:

• the so-called aggregation substance
• an accessory colonization factor
• an enterococcal surface protein (ES protein, ESP)
• various enzymes (gelatinases, serine proteases and hyaluronidases, cytolysins/hemolysins) collagen-binding proteins
• bacteriocins
• As well as extracellularly formed superoxides (Werner G et al.2008).

Natural anti-infective resistance: When treating diseases caused by E. faecalis and E. faecium, the natural resistance of these bacteria to numerous anti-infective agents must be taken into account. In contrast to acquired resistance, natural resistance is a species-dependent trait and consequently occurs in (almost) all strains of a species.

Resistance to cephalosporins and some other beta-lactam antibiotics: Of particular clinical and therapeutic importance are the natural resistances of E. faecalis and E. faecium to cephalosporins and some other beta-lactams such as isoxazolylpenicillins (e.g. oxacillin, flucloxacillin) and monobactams (aztreonam) (Werner G et al.2008). Cephalosporins are not effective due to the so-called penicillin binding protein (PBP) 5 - a specific protein formed by enterococci, which has only a reduced affinity to these beta-lactams. This results in mostly "high-grade" resistance of the pathogens.

Resistance to aminoglycosides: Furthermore, enterococci show a natural resistance to aminoglycosides. These anti-infectives can only pass through the cell wall of enterococci to a small extent, resulting in "low grade" resistance. In addition, some Enterococcus species produce a chromosomally encoded acetyltransferase that also contributes to "low grade" resistance to certain aminoglycosides (Werner G et al.2008).

Resistance to quinolones: Many enterococcal species also show reduced susceptibility to quinolones, although E. faecium, in contrast to E. faecalis, should already be considered naturally resistant to most quinolones. The main reason for the weak activity of quinolones on enterococci is probably the natural occurrence of resistance genes encoding proteins that can protect topoisomerase II (topoisomerase II is an important target protein of quinolones), from inhibitory action by these anti-infectives (Arsene S et al.2007). Since such resistance genes have been detected in E. faecalis, this species should also be designated as naturally quinolone resistant.

Resistance to polymyxins: Enterococci are naturally resistant to polymyxins such as polymyxin B or colistin. Such anti-infectives generally show little or no activity on most gram-positive bacteria.

Resistance to lincosamides and streptogramins: E. faecalis, but not E. faecium, also has natural lincosamide and streptogramin resistance. The lack of effect of lincosamides and streptogramins on E. faecalis is due to the natural expression of a species-specific efflux protein called the ABC transporter LSA (Singh KV et al.2001).

'Low-grade' vancomycin resistance of E. gallinarum and E. casseliflavus: The natural 'low-grade' vancomycin resistance of E. gallinarum and E. casseliflavus, the so-called VanC phenotype, has been of secondary medical importance due to the rare occurrence of these bacteria in the clinic. The natural and acquired glycopeptide resistance of enterococci is due to a modification of the site of attack for these anti-infectives.

Acquired anti-infective resistance: Acquired (secondary) resistance to antibacterial anti-infective agents also plays an important role in the therapy of diseases caused by E. faecalis and E. faecium. While many secondary resistances such as aminoglycoside "high resistance" are common in both species, others such as aminopenicillin or glycopeptide resistance occur mainly in E. faecium. In all other respects, the particular resistance situation should be assessed selectively from pat. to pat.

Aminopenicillins: The greatest difference in the frequency of secondary resistance between strains of E. faecalis and E. faecium concerns aminopenicillin resistance. Although the proportion of aminopenicillin-resistant E. faecalis strains may vary by region, worldwide, typically less than 5% of all clinical E. faecalis isolates show ampicillin or amoxicillin resistance. Studies from the United States, for example, document a 1- to 3% ampicillin resistance rate in E. faecalis (Tsigrelis C et al.2007). In contrast, particularly as a result of the spread of the ampicillin-resistant CC17 E. faecium clone, more than 80% of clinical E. faecium strains are currently aminopenicillin-resistant worldwide (Top J et al. 2008). According to PEG data, the ampicillin resistance rate in E. faecium in Germany and Central Europe was still 49% in 1995. In subsequent years, a rapid increase in the resistance rate was documented up to 89% in 2004 (Kresken M et al. 2007). Since then, a roughly constant rate of ampicillin resistance in E. faecium has been observed in our country (Kresken M et al. 2007). Various mechanisms have been described as the cause of acquired aminopenicillin resistance in enterococci. In addition to the expression of penicillinases, i.e., penicillin-inactivating beta-lactamases, these include a reduced affinity of certain penicillin-binding proteins for beta-lactams. In Europe, mutations or recombinations in penicillin-binding protein 5 in particular appear to be responsible for the aminopenicillin resistance of clinical strains of E. faecium.

Aminoglycosides: Of particular therapeutic interest is the secondary resistance of E. faecium and E. faecalis to aminoglycosides. Aminoglycoside "highly resistant" enterococci were first described in the 1980s and are now widely distributed worldwide. According to PEG data, 30% of all E. faecalis strains and as many as 35% of all E. faecium strains in Central Europe currently show a gentamic resistance to aminoglycosides.faecium strains show a gentamicin "high resistance" (Kresken M et al. 2007).The cause of the aminoglycoside "high resistance" of enterococci are the so-called aminoglycoside-modifying enzymes, which inactivate certain aminoglycosides by transferring acetyl, nucleotidyl or phosphate groups and are accordingly called acetyltransferases (AAC), nucleotidyltransferases (ANT) or phosphotransferases (APH).

Glycopeptides: The acquired and widespread resistance of E. faecium to vancomycin and, in some cases, teicoplanin poses a particular therapeutic problem. This is mainly because corresponding strains are usually almost always also secondarily resistant to aminopenicillins and often to a number of other anti-infective agents. Fortunately, in most countries, only a small proportion of clinical E. faecalis strains - usually less than 2% of isolates - continue to exhibit glycopeptide resistance (Kresken M et al. 2007). Glycopeptide-resistant enterococci (GRE) or vancomycin-resistant enterococci (VRE) were first observed in hospitals in France and the United Kingdom in 1986 [51] and are now present in almost all hospitals worldwide. The original reservoir of these strains is thought to be mammals raised in factory farms, in which selection and multiplication of vancomycin-resistant strains occurred as a result of intensive use of the vancomycin analog avoparcin, a growth-promoting substance used in animal husbandry in part until the 1990s, which then passed to humans through the food chain [5, 22].

Acquired resistance to other anti-infectives: Acquired resistance of enterococci is known to all therapeutically used anti-infectives with potential enterococcal activity. For example, in addition to the secondary resistances already described, "high resistance" to quinolones is also common in enterococci. The cause of ciprofloxacin "high resistance" has been shown to be mutations in the genes encoding topoisomerase II and topoisomerase IV in E. faecium (Leavis HL et al.2006). (Like topoisomerase II, topoisomerase IV is an important target of quinolones in Gram-positive bacteria). In addition, acquired resistance to macrolides, trimethoprim/sulfamethoxazole (co-trimoxazole), rifampicin, fusidic acid , and fosfomycin occurs frequently in E. faecalis and E. faecium strains (often in more than 30% of strains). Particularly high rates of resistance to these anti-infectives are observed in glycopeptide-resistant E. faecium strains

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