Streptococcal Macrolide Resistance Mechanisms


Importance of Macrolide Resistance of Streptococci

Macrolide resistance of S. pneumoniae is a major problem worldwide, with very high rates in some European countries and Asia, and somewhat lower rates in the Americas. The practical implication of this for the treatment of pneumococcal pneumonia is debated in the literature, but there are several case reports of failures of macrolide antimicrobials in the treatment of such infections. Treatment of upper respiratory tract infections, especially of anatomic sites where drug concentrations may be low, such as the respiratory sinuses and middle ear, may be more affected by this resistance than pneumococcal pneumonia.

Prevalence of Erythromycin Resistance of S. pneumoniae in Various Regions

Country or Region Years # isol % Erythromycin Resistance Reference
USA - Philadelphia, UPHS 2000 135 22 This web site
USA - Nationally 1997-1999 4193 18 CID 2001;32:S82
China - Hong Kong 1994-1998 197 80 AAC 2001;45:1578
Singapore 1997-1999 180 68 AAC 2000;44:2193
Italy 1997 302 33 AAC 1999;43:2510
Latin America 1999 257 13 AAC 2001;45:1463
Canada 1997-1999 887 11 CID 2001;32:S82
Europe 1997-1999 1478 20 CID 2001;32:S82
Spain 1994-1995 119 52 JAC 1998;41:207
Asia-Pacific 1997-1999 746 39 CID 2001;32:S82

 

 

Macrolide resistance of S. pyogenes appears to be relatively infrequent in North America, but emergence of more widespread resistance has been reported in Europe and other regions. The implications of this for the eradication of this bacterium from the pharynx are not clear cut; in one study the eradication rate for erythromycin treatment was 80% if the bacterium was erythromycin susceptible and 60% if it was not (Clin Infect Dis 1999;29:869).

Erythromycin susceptibility of S. pyogenes in various geographic regions

Country or Region Year # Isol. % Erythromycin Resistance Citation
USA 1994-1997   2.6 Ped Inf Dis J 1999;18:1069
San Francisco, USA 1994-1997 157-invasive strains 32 JCM 1999;37:1727
San Francisco 1994-1997 149 -pharyngitis strains 9 same as above
Pittsburgh, USA 2000-2001 100- non-invasive 38 NEJM 2002;346:1200
Philadelphia, USA 2002 (April) 100 - pharyngitis 1 unpublished, Univ Penn Med System
Canada 1997 3205 2.1 AAC 1999;43:2144
Chile 1990-1998 594 7.2 AAC 2001;45:339
Berlin   212 12.7 JAC 2000;46:621
Spain 1998 486 23.5 JAC 2000;45:605
France 1996-1999 1500 6.2 AAC 2000;44:1453
Italy   180 38 Emerg Inf Dis 2000;6:180
Sweden 1998-1999   3.7 Int J. Clin Pract 2000;54:585
Italy same   31 same
Turkey same   4.8 same
France same   3.8 same
Spain same   3.8 same

 

 


The resistance mechanisms can be desribed phenotypically and by genotype

Phenotypic Description

There are two major recognized resistance phenotypes, "MLSB" and "M"

MLSB - Resistant to macrolides, lincosamides, and streptogramin B. Conferred by methylation of a single adenine in the bacterial 50s ribosome that binds to erythryomycin, erm. Can be either inducible (iMLSB) or constitutive (cMLSB)

M -Resistant to macrolides, but not lincosamides or streptograminB. Conferred by macrolide efflux pump, mef.

Very recently, two novel mechanisms of macrolide resistance of pneumococci have been described, which complicate the inference of the genetic mechanism of resistance based on the phenotype. These two mechanisms are mutations in Domain V (Roman numeral V, not the letter "V") of the 23s rRNA, and mutations of two different ribosomal proteins that constitute part of the assembled 50s rRNA, ribosomal protein L4 and ribosomal protein L22. These mutations appear to result in two different phenotypes, ML and MSB, which can be confused with the phenotypes produced by erm and mef, respectively. These phenotypes can be differentiated from the M and MLSB phenotypes by inclusion of a streptogramin B and clindamycin in the testing.

The table below gives an example of typical susceptibility patterns for the various phenotypes


Genes Encoding for Macrolide Resistance

 

The genes classes that encode for the phenotypes are also of two main types, "erm" and "mef"

erm - The bacterial gene class coding for erythromycin ribosomal methylase, which methylates a single adenine in 23S rRNA, itself a component of 50S rRNA. Multiple erm gene types are recognized, and the nomenclature is very confusing. However a new nomenclature system may help (Antimicrob. Agents Chemother 1999;43:2823-2830)

The erm classes that are important for resistance of streptococci are class A and class B. Most authors refer to erm(TR) and erm(AM) for designating streptococcal erm

Class Proposed new gene name Includes these genes
A erm(A) erm(A), erm(TR)
B erm(B) erm(AM), erm(B), erm(AMR), erm(BC), erm(P), erm(BP), erm(IP), erm(Z), erm, erm(2)


To make matters even more confusing there can be both constitutive (always on) and inducible variants of erm, the phenotypes of which are designated as either cMLSB or iMLSB, respectively. erm gene regulation is complex and may involve more than one mechanism.

mef macrolide efflux pump. There are two recognized streptococcal mef types, mef(A) and mef(E), the former described in S. pyogenes and the latter in S. pneumoniae. mef(A) and mef(E) are 90% homologous, and it has been proposed that both be referred to as mef(A). mef actively pumps out macrolides (14 and 15 ring , but not 16 ring), but not lincosamides, streptogramins, or ketolides.

rRNA and ribosomal protein mutations The rRNA mutations are all in Domain 5 ("V") of 23S rRNA, very close to, or at the same location as the adenine that gets methylated by ermA ("A2058"), either at position 2058, 2059, or 2611, and are all adenine to guanine mutations. This region of Domain V is at the active translation site (peptidyl transferase region) of the 50S rRNA subunit (see Science 2000;289:905, or this link for more on ribosomal structure). There are four copies of rRNA in pneumococci, and these mutations have to occur in at least two of the ribsomes; the degree of resistance probably depends on whether the mutations are in 2, 3, or all 4 rRNAs. The ribosomal protein mutations have been mutations in either the L4 or L22 proteins, with a variety of changes, including substitutions, deletions, and additions (AAC 2002;46:371; AAC 2000;44:3395; AAC 2000;44:2118).

macrolide - erythromycin (more properly erythromycin A), clarithromycin, azithromycin (more properly an azalide) [all of these are either 14 or 15 ring compounds]. Rokitamycin is a 16 ring macrolide.

lincosamide - lincomycin and clindamycin

streptogramin B - quinupristin, a component of dalfopristin/quinupristin, or Synercid. Dalfopristin is a streptogramin A compound. Also pristinimycin IA is a streptogramin B compound, while pristinamycin IIA is a streptogramin A compound, both of which are components of the combination antimicrobial agent, pristinamycin .

ketolide - a macrolide which has been chemically modified. Telithromycin is one example of a ketolide.


Phenotypic Characterization of Macrolide Resistance Mechanisms

 

-By MIC Determinations-

Examples of macrolide resistance phenotypes of S. pyogenes**

MIC (ug/ml)

phenotype wild type M iMLSB cMLSB ML MSB
genotype mef(A) erm(TR) erm(B) mutations in 23S rRNA, or in L4 or L22 ribosomal proteins
erythromycin (14 ring) 0.03 4 1 >32 >32 >32
azithromycin (15 ring, azalide) 0.125 4 4 >32 >32 >32
clarithromycin (14 ring) 0.03 4 0.25 >32 >32 >32
roxithromycin (15 ring) 0.06 8 4 >32 >32 >32
clindamycin (lincosamide) 0.03 0.03 0.06 >32 0.8 0.1
telithromycin (ketolide) 0.03 0.25 0.06 1 to 16 0.01 0.1
clindamycin after exposure of bacteria to erythromycin 0.03 0.03 >32 >32 ? ?
rokitamycin (16 ring macrolide) 0.25 0.25 4 >32 >32 >32
streptogramin B 3 3 25 25 3 25

Portions of this table modified very liberally from Antimicrob. Agents Chemother 2000;44:2999-3002, and from AAC 2000;44:3395
** Note that in the case of S. pneumoniae the ketolides retain activity for the vast majority of cMLSB producers

-By Double Disk Susceptibility Testing-

Testing of streptococcal isolates with erythromycin and clindamycin disks applied close together can often yield phenotypic information, although it is not always possible to differentiate between phenotypes using this method. See J. Clin. Microbiol 2001;39:1311-1315 for details of this method for phenotypic analysis of S. pneumoniae. The figures below are taken from that paper, with some major modifications made to simplify this discussion; see the paper for the real and more complicated story.

 

iMLSB cMLSB M

iMLSB- the clindamycin zone is blunted towards the erythromycin because the erythromycin induces clindamycin resistance

cMLSB- no zone around either erythromycin or clindamycin because erm is fully expressed at all times

M- no change in the clindamycin zone induced by erythromycin because mef does not pump out clindamycin regardless of erythromycin presence

A= clindamycin and B=erythromycin

 

Examples of triple disk testing of rRNA or ribosomal protein mutants. Disk on left is clindamycin, middle is erythromycin, and right is streptogramin B. All bacteria are S. pneumoniae. Plate A shows ML (rRNA mutation), plates B and C show MSB (Both L4 mutations). Taken from AAC 2000;44:3395.

 


Prevalence of the Various Phenotypes/Genotypes By Geographic Region for S. pneumoniae

 

 

Percent Prevalence of Macrolide Resistance Phenotypes/Genotypes
Country Years # isol M iMLSB cMLSB cMLSB & M Other rRNA, L22 or L4 mutation Ref.
USA 1994-1995 114 61 0 32 4 3   CID 1999;29:1186
Canada 1993-1996
147
56 0 42.5 0.7 0.7   AAC 1998;42:2425
France 1987-1997 48 0 0 100 0 0   AAC 2001;45:636
Italy 1998-2000 85 77 0 23 0 0   JCM 2001;39:1311
UK, Ireland 1997-1998 83 62   35 2 1   JAC 2001;48:541
Slovak Rep 1999-2000 28 0   39     61 AAC 2002;46:371
Romania 1999-2000 31 0   84     16 AAC 2002;46:371
Poland 1999-2000 27 11   85     4 AAC 2002;46:371
Eastern, Central Europe 1999-2000 180 14   70     16 AAC 2002;46:371

 


 


Practical Implications of the S. pneumoniae Macrolide Resistance Phenotype


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Written by Paul H. Edelstein 1/25/02.Modified 4/29/03. If you spot any errors, please let me know.