Characterization of Streptococcus pyogenes from Animal Clinical Specimens, Spain.
We conducted a study to provide a detailed characterization of animal S. pyogenes isolates using emm typing, pulsed-field gel electrophoresis (PFGE), and multilocus sequence typing (MLST). We also investigated erythromycin resistance mechanisms and phenotypes, as well as virulence genes.
Materials and Methods
Origin and Identification of Bacterial Isolates
We analyzed 15 isolates of S. pyogenes obtained from rabbits (n = 14) and sheep (n = 1) in Spain during 2006-2014 (Table 1). Most rabbit isolates were from unrelated animals, located in different commercial farms (n = 14) and locations throughout Spain. Links between rabbit farms were not identified. The sheep included in this study was from a farm that had no rabbits. Human contact with animals was restricted to the personnel working in the rabbit farms and sheep flocks.
We recovered isolates from different clinical backgrounds: 8 from skin infections, 4 from genital tract infections, and 1 each from respiratory infections, mastitis, and otitis. We collected samples from skin and ear infections with sterile cotton swabs and collected the milk sample from the mastitis case aseptically in a sterile tube. Rabbits with genital tract or lung infections were euthanized, at farms or laboratories, and necropsied under aseptic conditions; clinical specimens were collected with forceps and scissors scrubbed in 70% ethanol. Samples taken at farms were transported to the laboratory in refrigerated polyethylene bags and processed within 24 hours after sampling.
Clinical specimens were sampled onto blood agar plates that were incubated at 37[degrees]C for 24-48 hours. Identification of isolates as S. pyogenes was based on colony morphology, P-hemolysis, and biochemical characteristics using the commercial identification system rapid ID 32 STREP (BioMerieux, Marcy L'Etoile, France). Biochemical identification was also confirmed by sequencing the 16S rRNA gene (8).
Antimicrobial Drug Susceptibility Tests
We performed drug susceptibility testing using the Clinical and Laboratory Standards Institute broth microdilution method (9) in Mueller-Hinton broth supplemented with 5% lysed horse blood. We determined the susceptibilities of the isolates with a commercially available susceptibility test (CMV3AGPF Sensititer standard panel; Trek Diagnostics, West Essex, UK) performed according to the manufacturer's instructions. The agents we tested were penicillin (0.25-16 [micro]g/mL), erythromycin (0.25-8 [micro]g/mL), vancomycin (0.25-32 [micro]g/mL), daptomycin (0.25-16 [micro]g/ mL), chloramphenicol (2-32 [micro]g/mL), linezolid (0.5-8 [micro]g/ mL), tetracycline (1-32 [micro]g/mL), quinupristin (0.5-32 [micro]g/ mL), tigecycline (0.05-0,5 [micro]g/mL), streptomycin (512-2048 [micro]g/mL), kanamycin (128-1024 [micro]g/mL), lincomycin (1-8 [micro]g/mL), and gentamicin (128-1024 [micro]g/mL). In addition, we determined MICs of clindamycin, erythromycin, and tetracycline by Etest (AB Biodisk, Solna, Sweden). We interpreted the results using the Clinical and Laboratory Standards Institute breakpoints for streptococci (9) for penicillin, erythromycin, vancomycin, daptomycin, chloramphenicol, tetracycline, and quinupristin; the European Committee on Antimicrobial Susceptibility Testing breakpoints for tigecycline and linezolid (http://www.eucast.org/ clinical_breakpoints); and the Comite de l'Antibiogramme de la Societe Francaise de Microbiologie breakpoints (10) for streptomycin, kanamycin, lincomycin, and gentamicin.
Macrolide Resistance Phenotype
To identify macrolide resistance phenotypes, we used a double-disk diffusion test (D-zone test) using erythromycin (15 pg) and clindamycin (2 pg) disks, as described by Hasenbein et al. (11). Isolates with blunting of the clindamycin inhibition zone around the disk adjacent to the erythromycin disk were considered to have an [iMLS.sub.B] phenotype (erythromycin resistant and clindamycin inducible). Clindamycin-susceptible isolates without blunting indicated an M phenotype (erythromycin resistant and clindamycin susceptible). Isolates that were resistant to both antimicrobial drugs were considered to have a [cMLS.sub.B] phenotype (constitutive erythromycin and clindamycin resistant).
Detection of Macrolides and Tetracycline
We extracted DNA according to the protocol in the US Centers for Disease Control and Prevention (CDC) S. pyogenes sequence database (http://www.cdc.gov/ncidod/biotech/strep/protocols.htm). We screened all erythromycinresistant isolates by PCR for the erythromycin resistance genes ermB (12), ermA (13), mefA (14), and msrD (15). We tested tetracycline-resistant isolates for the tetracycline resistance genes tetM and tetO (14).
Detection of Virulence Genes
We tested the S. pyogenes isolates for the presence of the virulence genes speA, speB, speC, speF, speG, speH, speJ, speM, ssa, and smeZ by PCR. We used primers and conditions described previously (16,17).
PFGE Analysis, MLST, and emm Typing
For PFGE analysis, genomic DNAs of the S. pyogenes isolates were prepared and digested with SmaI restriction enzyme (MBI Fermentas, Vilnius, Lithuania) following a previously published protocol (18). We performed MLST following the method established by Enright et al. (19) and assigned the allele and sequence type (ST) according to the PubMLST website (http://pubmlst.org/spyogenes). We amplified and sequenced the emm gene according to the protocol of the CDC International Streptococcal Reference Laboratory (http://www.cdc.gov/streplab/protocol-emmtype.html). We compared the sequences of the emm genes with those in the CDC database using BLAST analysis (http://www.cdc.gov/ncidod/biotech/strep/strepblast.htm) for type assignment.
We observed 2 emm types (Table 2): emm 12 was the most frequent (11 isolates), followed by emm77 (4 isolates). Two pulsotypes (A and B) were generated after typing the isolates by PFGE with the restriction enzyme SmaI; 11 isolates were pulsotype A and 4 isolates pulsotype B (Figure). Similarly, we observed 2 genetic linages (ST26 and ST63) after MLST analysis.
All 15 S. pyogenes isolates were susceptible to penicillin (MIC [less than or equal to]0.25 mg/L), vancomycin (MICs [less than or equal to]0.25 to 0.5 mg/L), daptomycin (MIC [less than or equal to]0.25 mg/L), chloramphenicol (MICs [less than or equal to]2 to 4 mg/L), tigecycline (MICs [less than or equal to]0.015 to 0.12 mg/L), and gentamicin (MIC [less than or equal to]128 mg/L). Additionally, all isolates but 1 were susceptible to kanamycin (MIC [less than or equal to]128 mg/L), and 12 isolates showed susceptibility to linezolid (MICs <2 mg/L), streptomycin (MICs [greater than or equal to]2,048 mg/L), and lincomycin ([greater than or equal to]8 mg/L). On the other hand, all isolates were resistant to tetracycline, with MICs ranging from 24 to 96 mg/L using Etest (Table 2). Eleven isolates showed tetracycline-resistant genotype tetM/tetO, 2 isolates tetO, and 1 isolate tetM (Table 2).
Most isolates (7/15) exhibited the M phenotype, 2 isolates the phenotype [cMLS.sub.B], and 1 the phenotype [iMLS.sub.B] (Table 2). The macrolide-resistant genotype mefA/ermB was the most frequently observed, seen in all isolates but 1 with the M phenotype and in the isolate with phenotype [cMLS.sub.B]. The genotype ermB was observed alone in 1 isolate of each phenotype. No isolate carried the msrD or ermA macrolide-resistant determinants.
We detected the chromosomal-encoded speB and speF genes in all isolates. We observed 2 different virulence gene profiles based on the presence/absence of the speG and speC genes. We detected the genotype speG in 11 isolates and the genotype speC in 4 isolates (Table 2).
We grouped the 15 S. pyogenes isolates into 2 different clones on the basis of emm-MLST-PFGE-virulence genes profile combinations. Clone 1 grouped isolates characterized by the combined genotype emm 12-ST36-pulsotype A-speB/speF/speG, whereas isolates of clone 2 were characterized by the genotype emm77-ST63-pulsotype B-speB/ speF/speC (Table 2). In addition, isolates of clone 1 were erythromycin resistant, mainly exhibiting an M phenotype, and isolates of clone 2 were erythromycin susceptible.
S. pyogenes is a human pathogen that has rarely been isolated from animals. It has been isolated from abscesses in cervical and mesenteric lymph nodes and liver of a free-living European hedgehog (E. europaeus) and from 2 dogs with severe colonic disease and conjunctivitis (5-7). Here we describe the detailed characterization of animal S. pyogenes isolates from different clinical specimens obtained from rabbits (n = 14) and sheep (n = 1) in Spain during 2006-2014. This pathogen was recovered mainly from noninvasive cases, with skin infections being the most common clinical presentation (n = 6), followed by genital tract infections (n = 4) (Table 1). S. pyogenes was isolated from all skin clinical samples together with Staphylococcus aureus, a well-recognized pathogen associated with different skin diseases in animals (20). These results indicate that although S. pyogenes should be able to colonize the skin of animals, it is difficult to ascertain its etiologic significance in skin infections. However, S. pyogenes was isolated in pure culture from clinical specimens of the genital tract, ears, mammary glands, and lungs in rabbits, indicating the potential role of S. pyogenes in these infections.
Most of the S. pyogenes isolates we tested (n = 11) exhibited the genotype emm12-ST36, which has been isolated repeatedly from humans in different countries (21-27), including Spain (28-30). This genotype can exhibit an M phenotype (31) and has been associated with skin and soft tissue infections (32), data that fit with our results, as more than half of the isolates with this genotype were isolated from abscesses and dermatitis (Table 2). The genotype emm77-ST63 that we identified in 4 animal isolates has also been detected in human S. pyogenes isolates (21,25,33), but unlike human isolates, the isolates in our study were erythromycin and clindamycin susceptible (Table 2).
All 11 isolates in clone 1 (pulsotype A) exhibited PFGE profiles that were indistinguishable from each other, and all 4 isolates in clone 2 also exhibited PFGE profiles that were indistinguishable PGFE from each other (pulsotype B; Figure). Isolates of S. pyogenes usually exhibit high levels of genetic diversity (4). Thus, the fact that we identified only 2 clones in different isolates collected over a period of 8 years was unexpected. The possibility of a common source of infection is very unlikely because all isolates were recovered at different times from different animals in farms located at geographically distant locations spread throughout Spain, without any epidemiologic relationship (Table 1). In addition, clinical specimens were processed independently in the same laboratory by highly qualified and trained personnel, which makes the possibility of a cross-contamination in the laboratory unlikely.
Under these conditions, multiple human-to-animal transmission events should be the most likely origin of these genotypes in sheep and rabbits. Another possible explanation could be that genotypes ST36 and ST63, although originating from humans, represent genetic linages with a specific host tropism, mainly for rabbits, which contributed to their successful dissemination in these animals, as observed with other streptococci (34). Cases of S. pyogenes infection were not recorded among the personnel working in the rabbit farms and sheep flock from which S. pyogenes was isolated. Asymptomatic human carriers have a key role in S. pyogenes transmission (35). For these reasons and even though screenings to identify asymptomatic S. pyogenes carriers were not carried out, we can speculate that asymptomatic employees were the most probable source of S. pyogenes in the animals included in the study. Although we cannot infer from the results of this study that animals, mainly rabbits, may represent a new reservoir of S. pyogenes, the results clearly indicate the ability of human-derived S. pyogenes isolates to colonize and infect animals, which could be more frequent than has been recognized until now.
Isolates with the genotype mefA/ermB usually correlate with the [cMLS.sub.B] phenotype, but 5 of the 6 S. pyogenes isolates with the mefA/ermB genotype in our study exhibited M phenotype (Table 2), which agrees with previous observations (29). The erm gene usually confers co-resistance to macrolides, lincosamides, and streptogramins. Curiously, all M phenotype isolates in our study showed susceptibility to clindamycin and were positive for the emrB gene. This result, although unusual, has also been observed previously in S. pyogenes isolates from different countries (26,36-38). A possible explanation could be that the ermB gene was nonfunctional in the isolates with clindamycin-susceptible phenotypes. The isolate M72193 exhibited the [iMLS.sub.B] phenotype but was ermA-negative (Table 2). This result, although infrequent, has also been observed in previous studies (39). Isolates with the [iMLS.sub.B] phenotype have been further subdivided into 3 distinct types: type A, associated with the presence of the ermB gene; and types B and C, associated with the presence of the ermA gene (40,41). This isolate carried the ermB gene (Table 2), suggesting therefore an [iMLS.sub.B]-A phenotype.
Unlike most human S. pyogenes isolates, which usually carry either tetM or tetO genes, most of the isolates in this study (n = 11) carried both genes (Table 2). Human isolates with the combination of tetM and tetO tetracycline-resistance genes have been identified previously in Spain (29). Another uncommon result was the identification of 1 isolate (83553) that was resistant to tetracycline (MIC 64 mg/L) but lacked resistance tetM and tetO genes (Table 2) commonly associated with tetracycline resistance in S. pyogenes (42). However, tetracycline-resistant strains and negativity to these genes have also been reported (43). Further studies will be necessary to elucidate the precise mechanism of resistance to tetracycline in this strain.
In summary, this study provides a detailed characterization of animal S. pyogenes isolates associated with different clinical backgrounds. This pathogen should be considered by veterinary microbiologists when processing clinical material from animals.
Dr. Vela is an associate professor at the Animal Health Department, Veterinary Faculty, Complutense University, Madrid, Spain. Her research focuses on the characterization of relevant animal bacterial pathogens.
(1.) Bisno AL, Stevens D. Streptococcus pyogenes (including streptococcal toxic shock syndrome and necrotizing fasciitis). In: Mandell GL, Douglas RG, Dolin R, editors. Principles and practice of infectious diseases, 5th ed., vol. 2. Philadelphia: Churchill Livingstone; 2000. p. 2101-17.
(2.) Ralph AP, Carapetis JR. Group A streptococcal diseases and their global burden. Curr Top Microbiol Immunol. 2013;368:1-27.
(3.) Walker MJ, Barnett TC, McArthur JD, Cole JN, Gillen CM, Henningham A, et al. Disease manifestations and pathogenic mechanisms of group A Streptococcus. Clin Microbiol Rev. 2014;27:264-301. http://dx.doi.org/10.1128/CMR.00101-13
(4.) Bessen DE. Population biology of the human restricted pathogen, Streptococcus pyogenes. Infect Genet Evol. 2009;9:581-93. http://dx.doi.org/10.1016/j.meegid.2009.03.002
(5.) Franklinos LH, Efstratiou A, Macgregor SK, John SK, Hopkins T, Cunningham AA, et al. Streptococcus pyogenes infection in a free-living European hedgehog (Erinaceus europaeus). EcoHealth. 2015;12:689-92. http://dx.doi.org/10.1007/s10393-015-1051-2
(6.) Willard MD, Berridge B, Braniecki A, Bouley D. Possible antibiotic-associated colitis in a dog. J Am Vet Med Assoc. 1998; 213:1775-9,1753-4.
(7.) Sprot H, Efstratiou A, Hubble M, Morgan M. Man's best friend?--first report of prosthetic joint infection with Streptococcus pyogenes from a canine source. J Infect. 2012;64:625-7. http://dx.doi.org/10.1016/jjinf.2012.02.006
(8.) Vela AI, Fernandez E, Lawson PA, Latre MV, Falsen E, Dominguez L, et al. Streptococcus entericus sp. nov., isolated from cattle intestine. Int J Syst Evol Microbiol. 2002;52:665-9. http://dx.doi.org/10.1099/00207713-52-2-665
(9.) Clinical and Laboratory Standards Institute. Performance standards for antimicrobial susceptibility testing; twenty-second informational supplement (M100-S22). Wayne (PA): The Institute; 2012.
(10.) Comite de l'antibiogramme de la Societe Francaise de Microbiologie. Recomendations 2012. Paris: Societe Francaise de Microbiologie; 2012 [cited 2015 Jul 8]. http://www.sfm-microbiologie.org/
(11.) Hasenbein ME, Warner JE, Lambert KG, Cole SE, Onderdonk AB, McAdam AJ. Detection of multiple macrolide- and lincosamide-resistant strains of Streptococcus pyogenes from patients in the Boston area. J Clin Microbiol. 2004;42:1559-63. http://dx.doi.org/10.1128/JCM.42.4T559-1563.2004
(12.) Sutcliffe J, Grebe T, Tait-Kamradt A, Wondrack L. Detection of erythromycin-resistant determinants by PCR. Antimicrob Agents Chemother. 1996;40:2562-6.
(13.) Seppala H, Skurnik M, Soini H, Roberts MC, Huovinen P. A novel erythromycin resistance methylase gene (ermTR) in Streptococcus pyogenes. Antimicrob Agents Chemother. 1998;42:257-62.
(14.) Malhotra-Kumar S, Lammens C, Piessens J, Goossens H. Multiplex PCR for simultaneous detection of macrolide and tetracycline resistance determinants in streptococci. Antimicrob Agents Chemother. 2005;49:4798-800. http://dx.doi.org/10.1128/ AAC.49.11.4798-4800.2005
(15.) Luthje P, Schwarz S. Molecular basis of resistance to macrolides and lincosamides among staphylococci and streptococci from various animal sources collected in the resistance monitoring program BfT-GermVet. Int J Antimicrob Agents. 2007;29:528-35. http://dx.doi.org/10.1016/j.ijantimicag.2006.12.016
(16.) Commons R, Rogers S, Gooding T, Danchin M, Carapetis J, Robins-Browne R, et al. Superantigen genes in group A streptococcal isolates and their relationship with emm types. J Med Microbiol. 2008;57:1238-46. http://dx.doi.org/10.1099/jmm.0.2008/001156-0
(17.) Rivera A, Rebollo M, Miro E, Mateo M, Navarro F, Gurgui M, et al. Superantigen gene profile, emm type and antibiotic resistance genes among group A streptococcal isolates from Barcelona, Spain. J Med Microbiol. 2006;55:1115-23. http://dx.doi.org/10.1099/ jmm.0.46481-0
(18.) Stanley J, Linton D, Desai M, Efstratiou A, George R. Molecular subtyping of prevalent M serotypes of Streptococcus pyogenes causing invasive disease. J Clin Microbiol. 1995;33:2850-5.
(19.) Enright MC, Spratt BG, Kalia A, Cross JH, Bessen DE. Multilocus sequence typing of Streptococcus pyogenes and the relationships between emm type and clone. Infect Immun. 2001;69:2416-27. http://dx.doi.org/10.1128/IAI.69.4.2416-2427.2001
(20.) Foster AP. Staphylococcal skin disease in livestock. Vet Dermatol. 2012;23:342-51, e63. http://dx.doi.org/10T111/jT365-3164.2012. 01093.x
(21.) Enright MC, Spratt BG, Kalia A, Cross JH, Bessen DE. Multilocus sequence typing of Streptococcus pyogenes and the relationships between emm type and clone. Infect Immun. 2001;69:2416-27. http://dx.doi.org/10.1128/IAI.69.4.2416-2427.2001
(22.) Steer AC, Law I, Matatolu L, Beall BW, Carapetis JR. Global emm type distribution of group A streptococci: systematic review and implications for vaccine development. Lancet Infect Dis. 2009;9:611-6. http://dx.doi.org/10.1016/S1473-3099(09)70178-1
(23.) Luca-Harari B, Ekelund K, van der Linden M, Staum-Kaltoft M, Hammerum AM, Jasir A. Clinical and epidemiological aspects of invasive Streptococcus pyogenes infections in Denmark during 2003 and 2004. J Clin Microbiol. 2008;46:79-86. http://dx.doi.org/10.1128/JCM.01626-07
(24.) Silva-Costa C, Pinto FR, Ramirez M, Melo-Cristino J; Portuguese Surveillance Group for the Study of Respiratory Pathogens. Decrease in macrolide resistance and clonal instability among Streptococcus pyogenes in Portugal. Clin Microbiol Infect. 2008;14:1152-9. http://dx.doi.org/10.1111/j.1469-0691.2008.02104.x
(25.) McGregor KF, Spratt BG. Identity and prevalence of multilocus sequence typing-defined clones of group A streptococci within a hospital setting. J Clin Microbiol. 2005;43:1963-7. http://dx.doi.org/ 10.1128/JCM.43.4.1963-1967.2005
(26.) Reinert RR, Lutticken R, Sutcliffe JA, Tait-Kamradt A, Cil MY, Schorn HM, et al. Clonal relatedness of erythromycin-resistant Streptococcus pyogenes isolates in Germany. Antimicrob Agents Chemother. 2004;48:1369-73. http://dx.doi.org/10.1128/ AAC.48.4.1369-1373.2004
(27.) Szczypa K, Sadowy E, Izdebski R, Hryniewicz W. A rapid increase in macrolide resistance in Streptococcus pyogenes isolated in Poland during 1996-2002. J Antimicrob Chemother. 2004;54:828-31. http://dx.doi.org/10.1093/jac/dkh420
(28.) Ardanuy C, Domenech A, Rolo D, Calatayud L, Tubau F, Ayats J, et al. Molecular characterization of macrolide- and multidrug-resistant Streptococcus pyogenes isolated from adult patients in Barcelona, Spain (1993-2008). J Antimicrob Chemother. 2010;65:634-43. http://dx.doi.org/10.1093/jac/dkq006
(29.) Rubio-Lopez V, Valdezate S, Alvarez D, Villalon P, Medina MJ, Salcedo C, et al. Molecular epidemiology, antimicrobial susceptibilities and resistance mechanisms of Streptococcus pyogenes isolates resistant to erythromycin and tetracycline in Spain (1994-2006). BMC Microbiol. 2012;12:215. http://dx.doi.org/ 10.1186/1471-2180-12-215
(30.) Rivera A, Rebollo M, Miro E, Mateo M, Navarro F, Gurgui M, et al. Superantigen gene profile, emm type and antibiotic resistance genes among group A streptococcal isolates from Barcelona, Spain. J Med Microbiol. 2006;55:1115-23. http://dx.doi.org/10.1099/ jmm.0.46481-0
(31.) Opavski N, Gajic I, Borek AL, Obszanska K, Stanojevic M, Lazarevic I, et al. Molecular characterization of macrolide resistant Streptococcus pyogenes isolates from pharyngitis patients in Serbia. Infect Genet Evol. 2015;33:246-52. http://dx.doi. org/10.1016/j.meegid.2015.05.011
(32.) Lin JN, Chang LL, Lai CH, Lin HH, Chen YH. Clinical and molecular characteristics of invasive and noninvasive skin and soft tissue infections caused by group A Streptococcus. J Clin Microbiol. 2011;49:3632-7. http://dx.doi.org/10.1128/ JCM.00531-11
(33.) Perez-Trallero E, Montes M, Orden B, Tamayo E, Garcia-Arenzana JM, Marimon JM. Phenotypic and genotypic characterization of Streptococcus pyogenes isolates displaying the MLSB phenotype of macrolide resistance in Spain, 1999 to 2005. Antimicrob Agents Chemother. 2007;51:1228-33. http://dx.doi.org/ 10.1128/AAC.01054-06
(34.) Sorensen UB, Poulsen K, Ghezzo C, Margarit I, Kilian M. Emergence and global dissemination of host-specific Streptococcus agalactiae clones. MBiol. 2010;1:e00178-10. http://dx.doi.org/ 10.1128/mBio.00178-10
(35.) Jordan HT, Richards CL Jr, Burton DC, Thigpen MC, Van Beneden CA. Group A streptococcal disease in long-term care facilities: descriptive epidemiology and potential control measures. Clin Infect Dis. 2007;45:742-52. http://dx.doi.org/ 10.1086/520992
(36.) Acikgoz ZC, Gocer S, Tuncer S. Macrolide resistance determinants of group A streptococci in Ankara, Turkey. J Antimicrob Chemother. 2003;52:110-2. http://dx.doi.org/10.1093/jac/dkg300
(37.) Szczypa K, Sadowy E, Izdebski R, Hryniewicz W. A rapid increase in macrolide resistance in Streptococcus pyogenes isolated in Poland during 1996-2002. J Antimicrob Chemother. 2004;54:828-31. http://dx.doi.org/10.1093/jac/dkh420
(38.) Richter SS, Heilmann KP, Beekmann SE, Miller NJ, Miller AL, Rice CL, et al. Macrolide-resistant Streptococcus pyogenes in the United States, 2002-2003. Clin Infect Dis. 2005;41:599-608. http://dx.doi.org/10.1086/432473
(39.) Pavlovic L, Grego E, Sipetic-Grujicic S. Prevalence of macrolide resistance in Streptococcus pyogenes collected in Serbia. Jpn J Infect Dis. 2010;63:275-6.
(40.) Brenciani A, Bacciaglia A, Vecchi M, Vitali LA, Varaldo PE, Giovanetti E. Genetic elements carrying erm (B) in Streptococcus pyogenes and association with tet(M) tetracycline resistance gene. Antimicrob Agents Chemother. 2007;51:1209-16. http://dx.doi.org/ 10.1128/AAC.01484-06
(41.) Giovanetti E, Montanari MP, Mingoia M, Varaldo PE. Phenotypes and genotypes of erythromycin-resistant Streptococcus pyogenes strains in Italy and heterogeneity of inducibly resistant strains. Antimicrob Agents Chemother. 1999;43:1935-40.
(42.) Giovanetti E, Brenciani A, Lupidi R, Roberts MC, Varaldo PE. Presence of the tet(O) gene in erythromycin- and tetracycline-resistant strains of Streptococcus pyogenes and linkage with either the mef(A) or the erm (A) gene. Antimicrob Agents Chemother. 2003;47:2844-9. http://dx.doi.org/10.1128/AAC.47.9.2844-2849.2003
(43.) Dundar D, Sayan M, Tamer GS. Macrolide and tetracycline resistance and emm type distribution of Streptococcus pyogenes isolates recovered from Turkish patients. Microb Drug Resist. 2010;16:279-84.
Address for correspondence: Jose F. Fernandez-Garayzabal, Universidad Complutense de Madrid, Ftad. de Veterinaria-Patologia Animal I (Sanidad Animal), Avda. Puerta de Hierro s/n n/a, Madrid, Madrid 28040 Spain; email: email@example.com
Ana Isabel Vela, Pilar Villalon, Juan Antonio Saez-Nieto, Gema Chacon, Lucas Dominguez, Jose Francisco Fernandez-Garayzabal
Author affiliations: Complutense University, Madrid, Spain (A.I. Vela, L. Dominguez, J.F. Fernandez-Garayzabal); Instituto de Salud Carlos III, Majadahonda, Madrid (P. Villalon, J.A. Saez-Nieto); Laboratorio Exopol San Mateo, Zaragoza, Spain (G. Chacon)
Caption: Figure. Pulsed-field gel electrophoresis patterns of Smal-digested DNA of clinical isolates of Streptococcus pyogenes from animal specimens, Spain, 2006-2014. Lanes 1 and 17, DNA molecular size marker; lanes 3, 4, 5, 7, 8, 9, 10, 11, 12, 13, 15, and 16, isolates M50163, M79144, M78761, M75791, M75539, M75533, M75123, M73512, M72636, M72193, and M83639, respectively (pulsotype A); lanes 2, 6, 14, and 16, isolates 83553, 85374, M75768, and M82209, respectively (pulsotype B).
Table 1. Features and disease manifestations of 15 animals from which Streptococcus pyogenes isolates were collected, Spain, 2006-2014 Isolate * Animal Clinical background Specimen M50163 Rabbit Metritis Uterus M79144 Rabbit Abscesses and dermatitis Skin M78761 Rabbit Dermatitis Skin M75791 Rabbit Abscesses Skin M75539 Sheep Abscesses Skin M75533 Rabbit Otitis Ear M75123 Rabbit Metritis Uterus M73512 Rabbit Abortion Uterus M72636 Rabbit Metritis Uterus M72193 Rabbit Abscesses Skin 83639 Rabbit Abscesses and dermatitis Skin 83553 Rabbit Pneumonia Lung M82209 Rabbit Abscesses Skin M75768 Rabbit Mastitis Milk 85374 Rabbit Skin infection Skin Isolate * Geographic region Isolation date ([dagger]) M50163 Valencia 2006 Jan M79144 Valladolid 2013 Mar M78761 Valladolid 2013 Feb M75791 Valencia 2012 Apr M75539 Zaragoza 2012 Mar M75533 Valencia 2012 Mar M75123 Castellon 2012 Feb M73512 Zaragoza 2011 Aug M72636 Zaragoza 2011 May M72193 Valencia 2011 Apr 83639 Valladolid 2014 Mar 83553 Zaragoza 2014 Mar M82209 Valladolid 2013 Dec M75768 Zaragoza 2012 Mar 85374 Valladolid 2014 Aug * Isolates M50163 and M73512 were recovered in pure culture. The remaining isolates were recovered together with Staphylococcus aureus. ([dagger]) Except for isolates M79144 and M78761, which were isolated in the same farm but at different times, all other isolates were recovered from animals at different farms. Table 2. Testing results for the 15 isolates characterized in study of Streptococcus pyogenes from animal specimens, Spain * Isolate MIC, mg/L emm PFGE MLST type profile type ERY CLIN TET M50163 12 A ST36 >256 32 96 M79144 12 A ST36 >256 0.75 48 M78761 12 A ST36 >256 0.75 32 M75791 12 A ST36 6 0.09 32 M75539 12 A ST36 8 0.19 24 M75533 12 A ST36 16 0.19 32 M75123 12 A ST36 12 0.19 48 M73512 12 A ST36 0.25 0.12 32 M72636 12 A ST36 >256 >256 48 M72193 12 A ST36 >256 1.5 96 83639 12 A ST36 >256 0.38 48 83553 77 B ST63 0.19 0.12 64 85374 77 B ST63 0.12 0.09 64 M75768 77 B ST63 0.12 0.09 32 M82209 77 B ST63 0.19 0.12 64 Isolate Macrolide resistance TET resistance Phenotype Genotype genes M50163 [cMLS.sub.B] mefA/ermB tetM/tetO M79144 M mefA/ermB tetM/tetO M78761 M mefA/ermB tetM/tetO M75791 M mefA/ermB tetM/tetO M75539 M mefA/ermB tetM/tetO M75533 M mefA/ermB tetM/tetO M75123 M ermB tetM/tetO M73512 tetM/tetO M72636 [cMLS.sub.B] ermB tetM/tetO M72193 [iMLS.sub.B] ermB tetM/tetO 83639 M ermB tetM 83553 85374 tetO M75768 tetM/tetO M82209 tetO Isolate Virulence genes M50163 speB/speF/speG M79144 speB/speF/speG M78761 speB/speF/speG M75791 speB/speF/speG M75539 speB/speF/speG M75533 speB/speF/speG M75123 speB/speF/speG M73512 speB/speF/speG M72636 speB/speF/speG M72193 speB/speF/speG 83639 speB/speF/speG 83553 speB/speF/spec 85374 speB/speF/spec M75768 speB/speF/speC M82209 speB/speF/speC * ERY, erythromycin; CLIN, clindamycin; MLST, multilocus sequence typing; PFGE, pulsed-field gel electrophoresis; ST, sequence type; TET, tetracycline.
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|Author:||Vela, Ana Isabel; Villalon, Pilar; Saez-Nieto, Juan Antonio; Chacon, Gema; Dominguez, Lucas; Fernand|
|Publication:||Emerging Infectious Diseases|
|Date:||Dec 1, 2017|
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|Next Article:||July 2016: Zoonoses.|