Genotypic identification and antimicrobial susceptibility profiles of coagulase-negative staphylococci isolated from sheep’s milk
Identificação genotípica de Staphylococcus coagulase negativos (SCN) isolados de leite ovino e perfil de suscetibilidade a agentes antimicrobianos
Palavras-chave:
Genotypc identification, Coagulase negative Staphylococcus, Sheep milk, susceptibility profileResumo
Mastitis affect milk production and dairy products in sheep farming. Its main etiological agents are coagulase-negative staphylococci (CoNS). Identification based on DNA sequencing is considered the gold-standard method for these organisms. The present study identify CoNS species from 32 sheep’s milk isolates through polymerase chain reaction (PCR) with restriction fragment length polymorphism (RFLP) of the gap gene and to ascertain their susceptibility to nine selected antimicrobial agents by the agar disk diffusion technique. Four PCR-RFLP profiles were obtained, corresponding to three CoNS species: S. epidermidis (59.4%), S. chromogenes (34.4%), and S. devriesei (6.2%). All isolates were susceptible to cefalotin, ceftiofur, and penicillin/novobiocin. Twenty-one isolates (65.6%) were resistant to at least one antimicrobial agent, and 11 isolates (34.4%) were susceptible to all tested agents. Four isolates (12.5%) were multidrug-resistant, including an S. epidermidis isolate which exhibited resistance to oxacillin, confirmed by amplification of the mecA gene. In addition to constituting an animal health problem and causing economic losses to sheep farmers, CoNS have zoonotic potential
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Referências
AARESTRUP, F.M. Veterinary drug usage and antimicrobial resistance in bacteria of animal origin. Basic Clin Pharmacol Toxicol, v. 96, p. 271–281, 2005. Disonível em: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3291075/. Acessado em: 10 Ago. 2023.
BARILLET, F.; RUPP, R.; MIGNON-GRASTEAU, S.; ASTRUC, J.M.; JAQUIN, M. Genetic analysis for mastitis resistance and milk somatic cell score in French Lacaune dairy sheep. Genet Se Evol, v. 33, p. 397-415, 2011. Disponível em: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2705413/. Acessado em: 10 Ago. 2023.
BERGONIER, D.; BERTHELOT, X. New advances in epizootiology and control of ewe mastitis. Livest Prod Sci, v. 79, p. 1–16, 2003. Disponível em: https://www.sciencedirect.com/science/article/pii/S0301622602001458. Acessado em: 10 Ago. 2023.
CLSI - CLINICAL AND LABORATORY STANDARDS INSTITUTE STANDARDS DEVELOPMENT POLICIES AND PROCESS. Performance Standards for Antimicrobial Susceptibility Testing: Twenty-third Informational Supplement M100-S23., Wayne, PA: USACLSI, 2013. Disponível em: https://clsi.org/media/3481/m100ed30_sample.pdf. Acessado em: 10 Ago. 2023.
CONTRERAS, A.; CORRALES, J.C.; SIERRA, D.; MARCO, J. Prevalence and aetiology of non-clinical intramammary infection in Murciano-Granadina goats. Small Rumin Res, v. 17, n. 1, p. 71-78, 1995. https://www.sciencedirect.com/science/article/pii/092144889500651Z. Acessado em: 10 Ago. 2023.
DALL AGNOL, A.M.; CAVALCANTE, M.B.; DE FRANÇA, C.A.; DA COSTA, C.; KREWER, C.C.; DE QUEIROS, A.A.; DA COSTA, M.M.; BRAGANÇA, J.F.M.; GIRARDINI, L.K. Caracterização fenotípica e molecular de isolados de Staphylococcus spp. obtidos de leite de ovelhas do município de Chapecó-SC. Semina: Ciências Agrárias, v. 34, n. 1, p. 311-322, 2013. Disponível em: https://ojs.uel.br/revistas/uel/index.php/semagrarias/article/view/11516/pdf. Acessado em: 10 Ago. 2023.
DAVENPORT, K.W.; DALIGAULT, H.E.; MINOGUE, T.D.; BISHOP-LILLY, K.A.; BROOMALL, S.M.; BRUCE, D.C.; CHAIN, P.S.; COYNE, S.R. Complete Genome Assembly of Staphylococcus epidermidis AmMS 205. Genome Announc, v. 2, p. 5–6, 2014. Disponível em: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4223448/. Acessado em: 10 Ago. 2023.
EL-JAKEE, J.K.; AREF, N.E.; GOMAA, A.; EL-HARIRI, M.D.; GALAL, H.M.; OMAR, S.A.; SAMIR, A. Emerging of coagulase negative staphylococci as a cause of mastitis in dairy animals: An environmental hazard. Int J Vet Sci Med, v. 1, p.74–78, 2013. Disponível em: https://www.sciencedirect.com/science/article/pii/S2314459913000215. Acessado em: 10 Ago. 2023.
FESSLER, A.T.; BILLERBECK, C.; KADLEC, K.; SCHWARZ, S. Identification and characterization of methicillin-resistant coagulase-negative staphylococci from bovine mastitis. J Antimicrob Chemother, v. 65, p. 1576–1582, 2010. Disponível em: https://academic.oup.com/jac/article/65/8/1576/736880. Acessado em: 10 Ago. 2023.
GHEBREMEDHIN, B.; LAYER, F.; KÖNIG, W.; KÖNIG, B. Genetic classification and distinguishing of Staphylococcus species based on different partial gap, 16S rRNA, hsp60, rpoB, sodA, and tuf gene sequences. J Clin Microbiol, v. 46, p. 1019–1025, 2008. Disponível em: https://journals.asm.org/doi/full/10.1128/jcm.02058-07. Acessado em: 10 Ago. 2023.
HOSSEINZADEH, S.; DASTMALCHI SAEI, H. Staphylococcal species associated with bovine mastitis in the North West of Iran: Emerging of coagulase-negative staphylococci. Int J Vet Sci Med, v. 2, p. 27–34, 2014. Disponível em:
https://www.sciencedirect.com/science/article/pii/S2314459914000040. Acessado em: 10 Ago. 2023.
KIOSSIS, E.; BROZOS, C.N.; PETRIDOU, E.; ZDRAGAS, A.; PAPADOPOULOS, T.; BOSCOS, C. Study on the possible survival of Staphylococcus chromogenes through the dry period in dairy ewes. Small Rumin Res, v. 115, p. 124–129, 2013. Disponível em: https://www.sciencedirect.com/science/article/pii/S0921448813002964, Acessado em: 10 Ago. 2023.
LANGE, C.C.; BRITO, M.A.V.P.; REIS, D.R.L.; MACHADO, M.A.; GUIMARÃES, A.S.; AZEVEDO, A.L.S.; SALLES, É.B.; ALVIM, M.C.T.; SILVA, F.S.; MEURER, I.R. Species-level identification of staphylococci isolated from bovine mastitis in Brazil using partial 16S rRNA sequencing. Vet Microbiol, v. 176, p. 382–388, 2015. Disponível em: https://www.sciencedirect.com/science/article/pii/S037811351500053X. Acessado em: 10 Ago. 2023.
MARKEY, B.; LEONARD, F.; ARCHAMBAULT, M.; CULLINAME, A.; MAGUIRE, D. Clinical Veterinary Microbiology. Mosby-Elsevier, Philadelphia, 2013.
MAROGNA, G.; ROLESU, S.; LOLLAI, S.; TOLA, S.; LEORI, G. Clinical findings in sheep farms affected by recurrent bacterial mastitis. Small Rumin Res, v. 88, p. 19–125, 2010. Disponível em: https://www.sciencedirect.com/science/article/pii/S0921448809002910. Acessado em: 10 Ago. 2023.
MARTINS, K.B. Caracterização do perfil clonal, fatores de virulência e determinação da resistência em Staphylococcus spp. isolados de leite ovino. 96 f. Dissertação (Mestrado em Biologia Geral e Aplicada) – UNESP, Batucatu, Brasil, 2013. Disponível em: https://repositorio.unesp.br/handle/11449/87784. Acessado em: 10 Ago. 2023.
MURAKAMI, K.; MINAMIDE, W.; NAKAMURA, E.; TERAOKA, H.; WATANABE, S. Identification of methicillin-resistant strains of staphylococci by polymerase chain reaction. J Clin Micrbiol, v. 29, n. 10, p. 2240-2244, 1991.
OLIVES, A.M.; DIAZ, J.R.; MOLINA, M.P.; PERIS, C. Quantification of milk yield and composition changes as affected by subclinical mastitis during the current lactation in sheep. J Dairy Sci, v. 96, p. 7698–7708, 2013. Disponível em: https://www.sciencedirect.com/science/article/pii/S0022030213006917. Acessado em: 10 Ago. 2023.
ONNI, T.; SANNA, G.; CUBEDDU, G.P.; MAROGNA, G.; LOLLAI, S.; LEORI, G.; TOLA, S. Identification of coagulase-negative staphylococci isolated from ovine milk samples by PCR-RFLP of 16S rRNA and gap genes. Vet Microbiol, v. 144, p. 347–352, 2010. Disponível em: https://www.sciencedirect.com/science/article/pii/S0378113510000520. Acessado em: 10 Ago. 2023.
ONNI, T.; SANNA, G.; LARSEN, J.; TOLA, S. Antimicrobial susceptibilities and population structure of Staphylococcus epidermidis associated with ovine mastitis. Vet Microbiol, v. 148, p. 45–50, 2011. Disponível em: https://www.sciencedirect.com/science/article/pii/S037811351000372X, Acessado em: 10 Ago. 2023.
ONNI, T.; VIDILI, A.; BANDINO, E.; MAROGNA, G.; SCHIANCHI, S.; TOLA, S. Identification of coagulase-negative staphylococci isolated from caprine milk samples by PCR-RFLP of groEL gene. Small Rumin Res, v. 104, p. 185–190, 2012. Disponível em: https://www.sciencedirect.com/science/article/pii/S0921448811004068. Acessado em: 10 Ago. 2023.
PEIXOTO, R.D.M.; MOTA, R.A.; DA COSTA, M.M. Mastite em pequenos ruminantes no Brasil. Pesq Vet Bras, v. 30, p. 754–762, 2010a. Disponível em: https://www.scielo.br/j/pvb/a/YbNK3hKRRp9PXRBx7Phvh9C/, Acessado em: 10 Ago. 2023.
PEIXOTO, R.D.M.; DE FRANÇA, C.A.; DE SOUZA JÚNIOR, A.F.; VESCHI, J.L.A.; DA COSTA, M.M. Etiologia e perfil de sensibilidade antimicrobiana dos isolados bacterianos da mastite em pequenos ruminantes e concordância de técnicas empregadas no diagnóstico. Pesq Vet Bras, v. 30, p. 735–740, 2010b. Disponível em: https://www.scielo.br/j/pvb/a/6MfZW5BPT63fLCnxmfC3CYH/. Acessado em: 10 Ago. 2023.
SAMPIMON, O.C.; LAM, T.J.G.M.; MEVIUS, D.; SCHUKKEN, Y.H.; ZADOKS, R.N. Antimicrobial susceptibility of coagulase-negative staphylococci isolated from bovine milk samples. Vet Microbiol, v. 150, p. 173–179, 2011.
SANTANA, R.C.M.; ZAFALON, L.F.; ESTEVES, S.N.; TANAKA, E.V.; PILON, L.E.; MASSA, R. Occurrence of Etiologic Agents Causing Subclinical Mastitis in Morada Nova and Santa Ines Ewes. Ars Vet, v. 29, p. 148–152, 2013. Disponível em: https://arsveterinaria.org.br/index.php/ars/article/view/689/869. Acessado em: 10 Ago. 2023.
SANTOS, O.C.D.S.; BARROS, E.M.; BRITO, M.A.V.P.; BASTOS, M.D.C.D.F.; DOS SANTOS, K.R.N.; GIAMBIAGI-DEMARVAL, M. Identification of coagulase-negative staphylococci from bovine mastitis using RFLP-PCR of the groEL gene. Vet Microbiol, v. 130, p. 134–140, 2008. Disponível em: https://www.sciencedirect.com/science/article/pii/S0378113507006402. Acessado em: 10 Ago. 2023.
SILVA, N.C.C.; GUIMARÃES, F.F.; MANZI, M.P.; BUDRI, P.E.; GÓMEZ-SANZ, E.; BENITO, D.; LANGONI, H.; RALL, V.L.M.; TORRES, C. Molecular characterization and clonal diversity of methicillin-susceptible Staphylococcus aureus in milk of cows with mastitis in Brazil. J Dairy Sci, v. 96, p. 6856–62, 2013. Disponível em: https://www.journalofdairyscience.org/article/S0022-0302(13)00641-3/pdf. Acessado em: 10 Ago. 2023.
SUPRÉ, K.; DE VLIEGHER, S.; CLEENWERCK, I.; ENGELBEEN, K.; VAN TRAPPEN, S.; PIEPERS, S.; SAMPIMON, O.C.; ZADOKS, R.N.; DE VOS, P.; HAESEBROUCK, F. Staphylococcus devriesei sp. nov., isolated from teat apices and milk of dairy cows. Int J Syst Evol Microbiol, v. 60, p. 2739–2744, 2010. Disponível em: https://www.microbiologyresearch.org/content/journal/ijsem/10.1099/ijs.0.015982-0, Acessado em: 10 Ago. 2023.
TEJADA, T.S.; SILVA, D.T.; DIAS, P.A.; CONCEIÇÃO, R.C.S.; MÜLLER NETO, H.; TIMM, C.D. Mastite subclínica por Staphylococcus coagulase negativa em ovinos de corte. Arq Bras Med Vet e Zootec, v. 64, p. 1074–1076, 2012. Disponível em: https://www.scielo.br/j/abmvz/a/8qCxbLVzZycYkhdD4HFgjWp/?lang=pt. Acessado em: 10 Ago. 2023.
VIRDIS, S.; SCARANO, C.; COSSU, F.; SPANU, V.; SPANU, C.; DE SANTIS, E.P.L. Antibiotic resistance in Staphylococcus aureus and coagulase negative Staphylococci isolated from goats with subclinical mastitis. Vet Med Int, 517060, 2010. Disponível em: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2860459/. Acessado em: 10 Ago. 2023.
ZADOKS, R.N.; TASSI, R.; MARTIN, E.; HOLOPAINEN, J.; MCCALLUM, S.; GIBBONS, J.; BALLIGALL, K.T. Comparison of bacteriological culture and PCR for detection of bacteria in ovine milk — Sheep are not small cows. J Dairy Sci, v. 97, p. 6326–6333, 2014. Disponível em: https://www.sciencedirect.com/science/article/pii/S0022030214005499. Acessado em: 10 Ago. 2023.
ZAFALON, L.F.; CUNHA, M.L.R.S.; RIBOLDI, D.F.M.; PILON, L.E. Persistência de Staphylococcus coagulase-negativos em glândulas mamárias de ovelhas com mastite subclínica após o tratamento antimicrobiano à secagem. Cienc anim bras, v. 18, p. 1-11, e-44328, 2017. Disponível em: https://www.scielo.br/j/cab/a/n9WLMNTgQDNBZTsd8g5NRgB/abstract/?lang=pt. Acessado em: 10 Ago. 2023.
