Methicillin resistance in staphylococci is determined by mec, composed of 50 kb or more of DNA found only in methicillin-resistant strains. mec contains mecA, the gene for penicillin-binding protein 2a (PBP 2a); mecI and mecR1, regulatory genes controlling mecA expression; and numerous other elements and resistance determinants. A distinctive feature of methicillin resistance is its heterogeneous expression. Borderline resistance, a low-level type of resistance to methicillin exhibited by strains lacking mecA, is associated with modifications in native PBPs, beta-lactamase hyperproduction, or possibly a methicillinase. The resistance phenotype is influenced by numerous factors, including mec and beta-lactamase (bla) regulatory elements, fem factors, and yet to be identified chromosomal loci. The heterogeneous nature of methicillin resistance confounds susceptibility testing. Methodologies based on the detection of mecA are the most accurate. Vancomycin is the drug of choice for treatment of infection caused by methicillin-resistant strains. PBP 2a confers cross-resistance to most currently available beta-lactam antibiotics. Investigational agents that bind PBP 2a at low concentrations appear promising but have not been tested in humans. Alternatives to vancomycin are few due to the multiple drug resistances typical of methicillin-resistant staphylococci.

Methicillin resistance in staphylococci : Molecular and biochemical basis and clinical implications

The majority of infections with glycopeptide intermediate-level resistant Staphylococcus aureus (GISA) originate in biomedical devices, suggesting a possible increased ability of these strains to produce biofilm. Loss of function of the accessory gene regulator (agr) of S. aureus has been suggested to confer an enhanced ability to bind to polystyrene. We studied agr in GISA, hetero-GISA, and related glycopeptide-susceptible S. aureus isolates. All GISA strains from diverse geographic origins belong to agr group II. All GISA strains were defective in agr function, as demonstrated by their inability to produce delta-hemolysin. Hetero-GISA isolate A5940 demonstrated a nonsense mutation in agrA that was not present in a pulsed-field gel electrophoresis-indistinguishable vancomycin-susceptible isolate from the same patient. Various other agr point mutations were noted in several clinical GISA and hetero-GISA isolates. A laboratory-generated agr-null strain demonstrated a small but reproducible increase in vancomycin heteroresistance after growth in vitro in subinhibitory concentrations of vancomycin. This was not seen in the isogenic agr group II parent strain in which agr was intact. The in vitro bactericidal activity of vancomycin was attenuated in the agr-null strain compared to the parent strain. These findings imply that compromised agr function is advantageous to clinical isolates of S. aureus toward the development of vancomycin heteroresistance, perhaps through the development of vancomycin tolerance.

Accessory Gene Regulator (agr) Locus in Geographically Diverse Staphylococcus aureus Isolates with Reduced Susceptibility to Vancomycin

SUMMARY “Heteroresistance” describes a phenomenon where subpopulations of seemingly isogenic bacteria exhibit a range of susceptibilities to a particular antibiotic. Unfortunately, a lack of standard methods to determine heteroresistance has led to inappropriate use of this term. Heteroresistance has been recognized since at least 1947 and occurs in Gram-positive and Gram-negative bacteria. Its clinical relevance may be considerable, since more resistant subpopulations may be selected during antimicrobial therapy. However, the use of nonstandard methods to define heteroresistance, which are costly and involve considerable labor and resources, precludes evaluating the clinical magnitude and severity of this phenomenon. We review the available literature on antibiotic heteroresistance and propose recommendations for definitions and determination criteria for heteroresistant bacteria. This will help in assessing the global clinical impact of heteroresistance and developing uniform guidelines for improved therapeutic outcomes.

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Antimicrobial Heteroresistance: an Emerging Field in Need of Clarity

Methicillin resistance in staphylococci is due to an acquired penicillin-binding protein, PBP2' (PBP2a). This additional PBP, encoded by mecA, confers an intrinsic resistance to all beta-lactams and their derivatives. Resistance levels in methicillin-resistant Staphylococcus aureus (MRSA) depend on efficient PBP2' production and are modulated by chromosomal factors. Depending on the genetic background of the strain that acquired mecA, resistance levels range from phenotypically susceptible to highly resistant. Characteristic for most MRSA is the heterogeneous expression of resistance, which is due to the segregation of a more highly resistant subpopulation upon challenge with methicillin. Maximal expression of resistance by PBP2' requires the efficient and correct synthesis of the peptidoglycan precursor. Genes involved in cell-wall precursor formation and turnover, regulation, transport, and signal transduction may determine the level of resistance that is expressed. At this stage, however, there is no information available on the functionality or efficacy of such factors in clinical isolates in relation to methicillin resistance levels.

Factors influencing methicillin resistance in staphylococci.

Fusobacterium necrophorum, a Gram-negative, non-spore-forming anaerobe, is a normal inhabitant of the alimentary tract of animals and humans. Two types of F. necrophorum, subspecies necrophorum (biotype A) and funduliforme (biotype B), have been recognized, which differ morphologically, biochemically, and biologically. The organism is an opportunistic pathogen that causes numerous necrotic conditions (necrobacillosis) such as bovine hepatic abscesses, ruminant foot abscesses and human oral infections. The pathogenic mechanism of F. necrophorum is complex and not well defined. Several toxins, such as leukotoxin, endotoxin, haemolysin, haemagglutinin and adhesin, have been implicated as virulence factors. Among these, leukotoxin and endotoxin are believed to be more important than other toxins in overcoming the host's defence mechanisms to establish the infection. F. necrophorum is encountered frequently in mixed infections and, therefore, synergisms between F. necrophorum and other pathogens may play an important role in infection. Several investigators have attempted to induce protective immunity against F. necrophorum using bacterins, toxoids, and other cytoplasmic components. Generally, none of the immunogens has afforded satisfactory protection against Fusobacterium infections. Because of the unavailability of suitable immunoprophylaxis, the control of F. necrophorum infection has depended mainly on the use of antimicrobial compounds.

Fusobacterium necrophorum infections: Virulence factors, pathogenic mechanism and control measures

The global regulators agr and sar control expression of cell wall and extracellular proteins. Inactivation of either sar and/or agr in a typical heterogeneously methicillin-resistant Staphylococcus aureus resulted in a small but reproducible decrease in the number of cells in the subpopulation expressing high methicillin resistance. The amount of low affinity penicillin-binding protein PBP2′, the prerequisite for methicillin resistance, was apparently not affected, however, a reduction in PBP1 and PBP3 production was observed, suggesting that these resident PBPs of the cells might be involved somehow together with PBP2′ in high level methicillin resistance.

https://www.zora.uzh.ch/id/eprint/155492/1/ZORA_NL_155492.pdf

Impact of sar and agr on methicillin resistance in Staphylococcus aureus.

Obligately anaerobic bacterial species isolated from foot-rot lesions in goats.

In-vitro antimicrobial susceptibility of Bacteroides and Fusobacterium isolated from footrot in goats.

The agar dilution method was used to determine the bacteriostatic activities of 10 beta-lactam antibiotics against 132 strains belonging to the genus Bacteroides and 25 strains belonging to the genus Fusobacterium, all isolated from clinical cases of caprine foot rot. The three ureidopenicillins studied proved to be the most effective antimicrobial agents.

S.Piriz Duran

Papers

Impact of sar and agr on methicillin resistance in Staphylococcus aureus.

Obligately anaerobic bacterial species isolated from foot-rot lesions in goats.

In-vitro antimicrobial susceptibility of Bacteroides and Fusobacterium isolated from footrot in goats.

Susceptibilities of Bacteroides and Fusobacterium spp. from foot rot in goats to 10 beta-lactam antibiotics.

Isolation and identification of anaerobic bacteria from ovine foot rot in Spain.