Genetics of resistance to macrolide antibiotics and lincomycin in natural isolates of Streptococcus pyogenes

Of 5 clinically isolated strains of Streptococcus pyogenes, 3 showed high-level resistance to erythromycin and lincomycin that was inducible by subinhibitory concentrations of these drugs (IR strains) while 2 strains exhibited constitutive erythromycin and lincomycin resistance (CR strains) which was expressed without prior exposure to low drug concentrations. The CR strain 15346 showed spontaneous loss of resistance whereas resistance in the other strains was quite stable even under curing conditions. The IR strain 13234 was found to be polylysogenic for at least 4 different phages designated P13234ma, mi, mu, and mo. Phage mo, antigenically distinct from the other three, was shown to mediate the transfer of the resistance determinant ERL1 of strain 13234. ERL1 if borne by appropriate strains was also transducible by the virulent phage A25. ERL1 behaved as a discrete genetic unit in transduction experiments, was not linked to either of two chromosomal regions governing resistance to antibiotics that affect the ribosome, could be transferred to recombination deficient hosts, represented a relatively large UV inactivation target, and showed no stimulation of transduction by low UV doses. These findings suggest that resistance to erythromycin and lincomycin in certain natural isolates of S. pyogenes is specified by, or under the control of, a plasmid.     

23S ribosomal ribonucleic acid of macrolide-producing streptomycetes contains methylated adenine.

Coresistance to macrolide, lincosamide, and streptogramin B-type (MLS) antibiotics by a common biochemical mechanism characterizes clinically resistant pathogens. Of 10 streptomycetes tested for resistance to macrolide, lincosamide, and streptogramin B-type antibiotics, only 1, Streptomyces erythreus, the organism used for production of erythromycin, was found resistant to all three classes; moreover, it was the only streptomycete in the series tested found to contain N6-dimethyladenine (m62A) in 23S ribosomal ribonucleic acid, the structural alteration of ribosomal ribonucleic acid associated with clinical resistance. Of the seven streptomycetes tested for the presence of m62A and N6-methyladenine (m6A), two, S. fradiae and S. cirratus, which produce the macrolide antibiotics tylosin and cirramycin, respectively, were found to contain m6A, but not m62A. The remaining strains tested, including strains which produce lincomycin and streptogramins, contained neither m6A nor m62A.     

Pneumococci resistant to erythromycin.

Susceptibility to erythromycin was determined for all pneumococci isolated in one laboratory from clinical specimens between 1969 and 1977. All 4724 isolates examined prior to October 1973 were susceptible to erythromycin. From October 1973 to December 1977, 64 (0.71%) of 8995 pneumococcus isolates were resistant to erythromycin. The resistant strains were isolated from 38 patients living in six widely separated communities in Alberta. The erythromycin-resistant strains were of nine capsular types, including six that often cause bacteremic disease and five for which resistance to erythromycin has not been reported hitherto. Certain strains of type 33 and of type 15 were highly resistant, the minimum inhibitory concentration (MIC) of erythromycin being 2000 microgram/mL; these strains were also highly resistant to lincomycin and clindamycin. Resistance in strains of other types was much lower, the MIC of erythromycin being 0.6 to 20 microgram/mL, and all but one of these strains were susceptible to lincomycin and clindamycin. All the erythromycin-resistant pneumococci were suspectible to penicillin.     

Drug Resistance of Staphylococci

Examination of cross resistance to macrolide antibiotics in erythromycin resistant staphylococcal strains isolated from clinical sources in the United States showed that there were two types of cross resistance, group A (13.4%) and group C (86.6%). Group A possessed multiple resistance to the macrolide antibiotics, erythromycin, oleandomycin, leucomycin and spiramycin, and was also resistant to lincomycin. In group C, resistance to erythromycin alone or to both erythromycin and oleandomycin could be induced by exposure to erythromycin.     

Action of Lincomycin on Staphylococci

On a solid medium, 0.1 to 1 μg/ml of lincomycin hydrochloride had a bacteriostatic effect upon 95 of 100 strains of staphylococci. Using cellophane transfers, we observed a bactericidal effect upon 54 of these strains after 3 to 14 hr of contact with 1 μg/ml. Five staphylococcal strains resistant to 100 μg/ml of lincomycin were also resistant to penicillin G, streptomycin, erythromycin, tetracycline, chloramphenicol (three strains), and rovamycin (three strains). Other staphylococcal strains resistant to methicillin, ampicillin, tetracycline, streptomycin, chloramphenicol, and erythromycin were sensitive to lincomycin.     

Mutation to erythromycin dependence in Escherichia coli K-12

A nitrosoguanidine-induced mutant of Escherichia coli K-12 strain JC12 was absolutely dependent on erythromycin or related macrolide antibiotics for growth. The only other drugs which permitted growth (lincomycin and chloramphenicol) are, like the macrolides, inhibitors of the 50S ribosome. The order of relative effectiveness of these drugs was macrolides > lincomycin > chloramphenicol. Rates of growth with all drugs were concentration dependent. Erythromycin starvation was followed by normal rates of increase in cell mass and macromolecular synthesis for approximately one mass-doubling time, after which macromolecular synthesis abruptly ceased and cell lysis and death occurred. The dependent mutant gave rise spontaneously to revertants to independence with very high frequency (10(-4)). The gene (mac) for macrolide dependence is located near minute 25 on the E. coli chromosome; it does not result in increased resistance to these drugs. A separate gene for erythromycin resistance (eryA) is located in the cluster of ribosomal structural genes near spc, close to minute 63. Dependence on macrolides was most clearly evident in strains carrying mutations at both eryA and mac.     

Sensitivity of 56 strains of Streptococcus group B to 12 antibiotics: its determination by dilution and diffusion in agar

Sensitivity of 56 streptococcal strains of group B to 12 antibiotics was studied with the method of dilution in a special solid medium for cultivation of streptococci and with the method of agar diffusion in the same medium. All the strains were found to be sensitive to chloramphenicol, ristomycin and erythromycin. The predominating majority of the strains were sensitive to beta-lactam antibiotics and lincomycin. All the strains were resistant to aminoglycoside antibiotics, such as streptomycin and gentamicin. More than 85 per cent of the strains were resistant to tetracycline. Strains with multiple resistance to 2-7 antibiotics were detected. Satisfactory correlation between the two methods was observed. It was shown to be clinically advisable to determine the sensitivity of streptococci of group B to beta-lactam antibiotics, erythromycin and lincomycin.     

The sensitivity to 12 antibiotics of 125 strains of Streptococcus group B isolated in a maternity home

Sensitivity of 125 strains of group B streptococci isolated from newborns, their mothers and personnel in a maternity home was studied with respect to 12 antibiotics: benzylpenicillin, ampicillin, methicillin, cephalotin, erythromycin, lincomycin, levomycetin (chloramphenicol), oxacillin, tetracycline, streptomycin, gentamicin and ristomycin. The method of serial dilutions in a solid medium was applied. All the strains were sensitive to ristomycin and erythromycin. The predominating number of the strains were sensitive to lincomycin, levomycetin and the beta-lactam antibiotics. Strains resistant or moderately resistant to benzylpenicillin, ampicillin, oxacillin, methicillin and cephalotin were detected. The majority of the strains were resistant to streptomycin, tetracycline and gentamicin. Multiple antibiotic resistance with 2-7 determinants was revealed in 11.2 per cent of the strains. The antibiotic sensitivity of the strains isolated from the newborns, their mothers and the personnel in the maternity home was on the whole similar or insignificantly differed.     

Ketolide resistance in Streptococcus pyogenes correlates with the degree of rRNA dimethylation by Erm

Macrolide and ketolide antibiotics inhibit protein synthesis on the bacterial ribosome. Resistance to these antibiotics is conferred by dimethylation at 23S rRNA nucleotide A2058 within the ribosomal binding site. This form of resistance is encoded by erm dimethyltransferase genes, and is found in many pathogenic bacteria. Clinical isolates of Streptococcus pneumoniae with constitutive erm(B) and Streptococcus pyogenes with constitutive erm(A) subtype (TR) are resistant to macrolides, but remain susceptible to ketolides such as telithromycin. Paradoxically, some strains of S. pyogenes that possess an identical erm(B) gene are clinically resistant to ketolides as well as macrolides. Here we explore the molecular basis for the differences in these streptococcal strains using mass spectrometry to determine the methylation status of their rRNAs. We find a correlation between the levels of A2058-dimethylation and ketolide resistance, and dimethylation is greatest in S. pyogenes strains expressing erm(B). In constitutive erm strains that are ketolide-sensitive, appreciable proportions of the rRNA remain monomethylated. Incubation of these strains with subinhibitory amounts of the macrolide erythromycin increases the proportion of dimethylated A2058 (in a manner comparable with inducible erm strains) and reduces ketolide susceptibility. The designation 'constitutive' should thus be applied with some reservation for most streptococcal erm strains. One strain worthy of the constitutive designation is S. pyogenes isolate KuoR21, which has lost part of the regulatory region upstream of erm(B). In S. pyogenes KuoR21, nucleotide A2058 is fully dimethylated under all growth conditions, and this strain displays the highest resistance to telithromycin (MIC > 64 microg ml-1).     

Antimicrobial Susceptibility of Clostridium difficile from Different Sources

A total of 79 Clostridium difficile strains from healthy young and elderly adults, elderly patients without gastrointestinal disease, elderly patients receiving antibiotics without gastrointestinal complications, and elderly patients with antibiotic-associated diarrhea or pseudomembranous colitis were tested for their susceptibilities to 24 antimicrobial agents. All of the 79 strains were inhibited by low concentrations of rifampicin, metronidazole, fusidic acid, vancomycin, ampicillin, and penicillin G. The strains were highly resistant to aminoglycosides, trimethoprim, sulfamethoxazole, nalidixic acid, and cycloserine and often resistant to neomycin, cefoxitin, and cefalexin. Wide variations in the susceptibility of C. difficile strains to erythromycin, clindamycin, lincomycin, chloramphenicol, and tetracycline were found. Strains resistant to erythromycin, clindamycin, and lincomycin were more frequently found among strains isolated from elderly adults than those isolated from young adults, with particularly high frequency among strains isolated from elderly patients receiving antibiotics. None of the 23 strains isolated from healthy young adults was resistant to chloramphenicol. All of the 14 strains resistant to erythromycin, clindamycin, lincomycin, and chloramphenicol were sensitive to tetracycline and all of the 15 strains resistant to erythromycin, clindamycin, lincomycin, and tetracycline were sensitive to chloramphenicol. Only one out of 19 tetracycline-resistant strains was highly toxigenic, whereas 42 (70%) of 60 sensitive strains were highly toxigenic.     

Sensitivity of the causative agent of tularemia to antibiotics

Sensitivity of the tularemia causative agent of different geographical races to antibiotics such as streptomycin, tetracycline, gentamicin, rifampicin (20 strains), ampicillin, polymyxin M, erythromycin, oleandomycin (361 strains) and lincomycin (294 strains) was studied. High sensitivity of the tularemi a microbe to streptomycin, tetracycline, rifampicin (MIC of 10 gamma/ml), gentamicin (MIC of 1 gamma/ml) and resistance to 50 gamma of ampicillin and 1000 gamma/ml of polymyxin M were found. Combined use of 50 gamma of ampicillin and 100 gamma/ml of polymyxin M added to the nutrient medium for growth inhibition of the foreign flora on isolation of the tularemia causative agent from the infected material including stable laboratory animal carcases was recommended. Marked differences in sensitivity of the strains of different geographical races to the macrolides and lincomycin were observed. The strains of the non-Arctic and Central Asiatic races were of low resistance to the above drugs (the MIC of erythromycin, oleandomycin and lincomycin were 10--50, 50--400 and 25--100 gamma/ml respectively. Within the holarctic race 40 per cent were low resistant and 60 per cent were highly resistant to these drugs. The above drugs should not be used for treatment of tularemia cases.     

Susceptibility to mercurials of clinical Pseudomonas aeruginosa isolated in México

Susceptibility to inorganic mercuric ions and to organomercurials of 237 Pseudomonas aeruginosa clinical strains isolated in Mexico was determined by agar dilution tests. Resistant strains fell into two classes: i) narrow-spectrum resistant strains (27% of total isolates) resistant only to mercuric ions and to merbromin, and most grouped in pyocin type 1; and ii) broad-spectrum resistant strains (5%) with additional resistances to thimerosal, phenylmercury, methylmercury and p-hydroxymercuribenzoate, that belonged mostly to pyocin type 10. Mercurial resistant isolates showed a higher proportion of resistance to antibiotics and metals than did mercurial sensitive isolates, and broad-spectrum resistant strains had the highest frequency of resistance to antibiotics and to tellurite and arsenate.     

Sensitivity of Corynebacterium diphtheriae isolated in Saint-Petersburg to antibacterial drugs]

One hundred and thirty strains of Corynebacterium diphtheriae isolated in St. Petersburg (42 toxigenic and 88 nontoxigenic) were tested with the method of serial dilutions in solid media for their susceptibility to 20 antibacterial drugs. The MICs of the drugs for the isolates ranged from < or = 0.015 to > or = 32.0 micrograms/ml. 13 per cent of the isolates was resistant at least to one antibacterial drug. The isolates resistant to erythromycin (11.5 per cent), lincomycin (11.5 per cent) and trimethoprim (8.5 per cent) were most frequent. 3 isolates (2.3 per cent) had multiple resistance to 8 drugs: benzylpenicillin, ampicillin, oxacillin, chloramphenicol, erythromycin, lincomycin, trimethoprim, and nitroxolin. No significant differences in the susceptibility of the toxigenic and nontoxigenic strains of C. diphtheriae were detected. Gentamicin, rifampicin, tetracycline, doxycycline and pefloxacin were the most active antibiotics.     

Characterization of multiple changes of antibiotic resistance characters in Streptomyces coelicolor A3(2)

Among mutants of Streptomyces coelicolor A3(2) studied which were sensitive to chloramphenicol (Cmls), strains sensitive to a number of antibiotics (ristomycin, tetracycline, polymyxin, lincomycin) amount of 46%. Antibiotic-sensitive mutants are capable to form different classes of resistant revertants with frequency varying from 10(-2) to 10(-6) in independent strains. Ristomycin-sensitive clones (Rims) have been found to occur with high frequency in Cmls strains and Cmlr revertants. Mutations mediating the Rims phenotype are mapped in a locus linked to the gene for resistance to chloramphenicol. The results obtained are discussed, in accordance with the notion about possible role of cml mutation in induction of secondary mutational changes in the genome of S. coelicolor A3(2).     

On the Origin of Mitochondrial Mutants: Evidence for Intracellular Selection of Mitochondria in the Origin of Antibiotic-Resistant Cells in Yeast

In wild-type Saccharomyces cerevisiae, erythromycin and certain other antibacterial antibiotics inhibit the formation of respiratory enzymes in mitochondria by inhibiting translation on mitochondrial ribosomes. This paper is concerned with the origin of mutant cells, resistant to erythromycin by virtue of having a homogeneous population of mutant mitochondrial DNA molecules. Such mutant cells are obtained by plating wild-type (sensitive) cells on a nonfermentable substrate plus the antibiotic. Colonies of mutant cells appear first about four days after the time of appearance of established mutant cells; new colonies continue to appear, often at a constant rate, for many days. Application of the Newcombe respreading experiment demonstrates that most or all of the mutant cells which form the resistant colonies on selective medium arise only after exposure of the population to erythromycin. It is suggested that this result is most probably due to intracellular selection for mitochondrial genomes. Resistant mitochondria arising from spontaneous mutation are postulated to be at a selective disadvantage in the absence of erythromycin; reproducing more slowly than wild-type sensitive mitochondria, they cannot easily accumulate in sufficient numbers in a cell to render it resistant as a whole. In the presence of erythromycin, resistant mitochondria can continue to reproduce while sensitive mitochondria cannot, until there is a sufficient number to make the cell resistant, i.e. to permit normal cell growth. The same phenomenon is seen with respect to chloramphenicol resistance. Intracellular selection is considered more likely than direct induction of mutation by the antibiotic, since mutant cells do not accumulate in the presence of erythromycin if the mitochondrial genome is rendered non-essential by growth on glucose or nontranslatable by chloramphenicol. Intra-cellular selection provides a mechanism for direct adaptation at the cell level, compatible with currently acceptable ideas of spontaneous mutation and selection at the organelle level.     

Transfer of plasmid-mediated antibiotic resistance from streptococci to lactobacilli.

The transmissible plasmid pAMbeta1, which codes for erythromycin and lincomycin resistance, was conjugally transferred from a Lancefield group F Streptococcus to a strain of Streptococcus avium. Both organisms served as pAMbeta1 donors for three strains of Lactobacillus casei. Introduction of pAMbeta1 into one of the L. casei strains caused the organism to lose its native 6.7 X 10(6)-dalton plasmid. Loss of the native plasmid produced no alterations in the organism's growth characteristics or fermentation pattern.     

Antibiotic sensitivity of Staphylococci populating the breast skin of nursing women

Sensitivity to penicillins, tetracycline, erythromycin and lincomycin was tested in 1513 staphylococcal strains isolated from the skin of various anatomic areas of the breast of nursing mothers. It was shown that 82.9, 78.4 and 33.9 per cent of the isolated cultures were highly sensitive to lincomycin, erythromycin and methicillin respectively. Sensitivity to tetracycline and benzylpenicillin was detected in 33.1 and 19.8 per cent of the cultures respectively. The study of the resistance spectra of 1343 strains resistant to certain antibiotics revealed that 498 cultures (37.1 per cent) were polyresistant. None of the tested antibiotics could be used for selective decontamination of the breast skin with respect to S. aureus in prophylaxis of lactic mastitis.     

Large scale isolation of maternally inherited lincomycin resistance mutations, in diploid Nicotiana plumbaginifolia protoplast cultures

Summary Lincomycin-resistant clones were isolated in diploid protoplast cultures of Nicotiana plumbaginifolia. Selection of the resistant clones was based on the ability of resistant calli to green in the presence of the antibiotic (1,000 mg l^-1). Sensitive colonies formed white calli under the same conditions. In the absence of mutagenic treatment the frequency of the resistant clones was 1.0×10^-4. This frequency could be increased up to 5.8×10^-4 and 7.2×10^-4 by treatment with 0.1 mM and 0.3 mM N-ethyl-N-nitrosourea (NEU), respectively.Regenerated plants of 56 clones were tested for lincomycin resistance. Regenerates from all but seven clones were resistant to lincomycin, as demonstrated by leaf assay. The lincomycin-resistant regenerates tested were also resistant to clindamycin (a lincomycin derivative), but sensitive to streptomycin.Regenerated plants in 17 clones were fully fertile and inherited lincomycin resistance maternally. Segregation for lincomycin resistance was observed in the seed progeny of five clones, which indicated maintenance of mixed cytoplasmic determinants after plant regeneration. Seed transmission of lincomycin resistance was confirmed in an additional 17 clones but the mode of inheritance (maternal or Mendelian) was not determined because of pollen sterility or reduced seed germination ability. These defects first appeared when the higher concentration of NEU was used. Various pigment deficiencies were also observed in a few clones.     

The correlation between phenotypes and genotypes of macrolides, lincosamides and streptogramins B resistance in Staphylococcus aureus

For 31 clinical strains of S. aureus the correlation between phenotype and genotype of resistance to macrolides, lincosamides and streptogramins B (MLSB) was established.. Phenotypes were determined on the basis of: susceptibility to erythromycin and clindamycin and the ability to an induction of the resistance (phenotypes S, susceptible; R , constitutive resistant, D, resistant after induction with erythromycin, D+, resistant after induction with erythromycin and with a presence of the small colonies inside inhibition zone between erythromycin and clindamycin discs), and on the basis of the resistance to spectinomycin (spR, resistant, spS, susceptible). Among examined S. aureus strains eight phenotypes of resistance to MLSB were recognized (the corresponding genotypes are given in brackets). Six phenotypes were typical: SspS (lack of MLS-B resistance genes), NEGspS (msrA/B, 1 strain), D+spS (ermCi, 4 strains),. DspR (ermAi, 11 strains and ermAi + msrA/B, 2 strains), RspR (ermAc, 4 strains and ermA + msrA/B,1 strain and ermA + ermC, 1 strain) and RspS (ermCc, 6 strains and ermB, 1 strain). Two rare phenotypes in two single strains were observed: SspR (ermAi, the strain with altered inducibility, inductor other than erythromycin) and DspS (ermAi, presumably mutation or lack of spc in Tn554).     

Susceptibility of sixty-five non-oral clinical isolates of the Streptococcus milleri group to seven antimicrobial agents.

Antimicrobial susceptibilities of sixty-five non-oral Streptococcus milleri group clinical isolates to penicillin, gentamicin, lincomycin, ampicillin, chloramphenicol, tetracycline and erythromycin were determined by an agar dilution method. All strains were penicillin-sensitive (MIC < or = 0.031 microgram/ml) and the majority (64/65) were susceptible to erythromycin (MIC < or = 0.125 microgram/ml). Low-level resistance to gentamicin was observed, and the majority of strains possessed an MIC of 8 micrograms/ml. Lincomycin and ampicillin at 0.5 microgram/ml inhibited 52/65 and 61/65 strains, respectively. Of the isolates 92% were inhibited by chloramphenicol at < or = 2 micrograms/ml. Twenty-two S. milleri group strains (of which thirteen were vaginal isolates) were resistant to tetracycline (MIC > or = 8 micrograms/ml).