Coagulase-Positive and Coagulase-Negative Staphylococci Animal Diseases

Chapter 4

Coagulase-Positive and Coagulase-Negative Staphylococci Animal Diseases Fulvio Marsilio, Cristina E. Di Francesco, Barbara Di Martino Faculty of Veterinary Medicine, University of Teramo, Teramo, Italy

4.1 INTRODUCTION Staphylococci are among the major groups of bacterial commensals isolated from skin, skin glands, and mucous membranes of mammals. Although staphylococci may colonize inner and/or external surfaces of healthy individuals, they may also behave as opportunistic pathogens as well as leading causes of community-associated and hospital-acquired disease in humans and animals worldwide. Coagulase positive staphylococci (CoPS) (i.e., Staphylococcus aureus in humans, S. aureus and Staphylococcus (pseudo)intermedius in animals) are most commonly implicated in pathologic processes. Coagulase-negative staphylococci (CoNS—mostly Staphylococcus epidermidis in humans and dogs), instead, are considered to be less common causes of disease in animals. Although their role as nosocomial pathogens in humans is becoming increasingly important, their zoonotic potential and importance in veterinary medicine is still unclear [1].

4.2  COAGULASE-POSITIVE STAPHYLOCOCCI S. aureus, along with Escherichia coli and Streptococcus uberis, are considered the three most important pathogens of the udder, and sometimes their high prevalence in a dairy herd is looked at as a threat to the economic sustainability of the livestock. Mastitis is the leading cause of economic losses in dairy cattle herds, because of poor yields in the infected udder, need for veterinary treatments, the amount of milk to be discarded (due to contamination with pathogens and/or antibiotic residues), and anticipated culling. Particularly if leukocytes counts in bulk milk are higher than 400,000 cells/mL, the product is considered unfit for human consumption (EEC directive 94/71) [2]. Pet-to-Man Travelling Staphylococci: A World in Progress. https://doi.org/10.1016/B978-0-12-813547-1.00004-2 © 2018 Elsevier Inc. All rights reserved.

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44  Pet-to-Man Travelling Staphylococci: A World in Progress

S. aureus remains the most important agent of bovine mastitis. The prevalence of the staphylococcal infection of mammary glands in the herds may differ among countries. It varies from 5% to 30% of clinical mastitis and from 5% to 10% of subclinical diseases [3]. S. aureus may be a potential problem from the public health point of view, because some strains are able to produce enterotoxins. The shedding from the infected udder is low, but contamination of bulk milk is pivotal for staphylococcal poisoning in fermented raw milk food [4]. Cows may be silent carriers of S. aureus in their udder, teat skin, nasal cavity, and rectum. However, the infected udder is the most important staphylococcal reservoir. Even if S. aureus needs to infect animals in order to multiply and survive, it may inhabit the environment for a long time, then the most important route of transmission “udder to udder” are the milking machine or the farmer’s hands [2]. S. aureus is able to cross the teat canal and may establish infection in the lumen of the mammary gland where it can multiply very rapidly, provided that oxygen is present (as it occurs in healthy milk). Furthermore, S. aureus adheres at the mammary epithelial cells at cisterns and duct levels, mainly in the presence of microdamage of the epithelium that exposes tissues and the extracellular matrix protein for which S. aureus has specific adhesins [2]. The next step is the internalization of the pathogens in the mammary epithelial cells that support chronicization of mastitis via immune system escape. The inflammatory response induced by the invasion of the mammary gland attracts a large number of neutrophils, together with monocytes and lymphocytes, that are able to produce specific mediators that are in turn responsible for the mastitis clinical picture [2]. Several upshots of staphylococcal mastitis are described that can range from subclinical to gangrenous processes, depending on the virulence of the strain involved [5]. Differences in virulence potential correlate with gene content and levels of cytolytic toxin expression [6]. α-Hemolysin (or α-toxin) plays an important role in the severity of infection [7] and producing strains are associated with gangrenous mastitis [2]. Some studies have shown that strains isolated from clinical mastitis may also differ from mild isolates with regard to toxins overproduction and proteases involved in blood clot formation. Furthermore, some strains that appear to be associated with severe mastitis show propensity for iron acquisition [5]. In order to avoid mastitis-related economic losses in dairy herds, it is necessary to introduce specific control measures such as milking procedures, post-milking teat disinfection, reform of infected animals, and preventing the introduction of infected animals during the herd turnover [2]. Furthermore, in order to prevent the infection spread within the herd, the S. aureus infected quarters have to be diagnosed as soon as possible to permit treatment and to allow infected animals to be rapidly isolated or reformed [2]. Staphylococcal mastitis is very difficult to treat. Antibiotics are administered via the teat canal, sometimes in combination with the parenteral therapy. The relevant frequency

CoPS and CoNS Animal Diseases  Chapter | 4  45

of treatment failure, however, may be explained by the intracellular localization of S. aureus or the formation of microabscesses and intramammary biofilms [2]. In order to reduce the impact of the extensive use of antibiotics in livestock, and then to prevent the emergence of resistant strains, several alternative methods for mastitis control are under investigation. The vaccination would be particularly useful, in this context, but currently available vaccines show only limited efficacy when used in the field [8]. The use of lactic acid bacteria as a mammary probiotic is a promising alternative strategy. For example, intrammamary infusions with Lactococcus lactis in cows with chronic subclinical or clinical mastitis were shown to be as effective as antibiotics in preventing the onset of mastitis [9]. Furthermore, it was demonstrated that Lactobacillus casei is able to inhibit S. aureus adhesion to and invasion of bovine mammary epithelial cells, in vitro [10]. These strategies appear promising, but the protective effect still has to be proven experimentally. The genetic selection of resistance to mastitis in ruminants, based on low somatic cell counts and low mastitis incidence, may be considered as a possible strategy for large and small ruminants [11]. Another direct strategy is to breed transgenic cows that are able to produce the lysostaphin in their milk. As a consequence, milk is able to lyse S. aureus and, in a context of experimental mastitis, the recombinant cows were fully protected against different S. aureus strains [12]. However, the ethical acceptability of using transgenic animals has to be considered. Since description of S. intermedius and, later, S. pseudintermedius, [13,14], more than one isolate previously identified as the former were reclassified as belonging to the second, based on nucleotide sequence analysis of the sodA and hsp60 genes [13,14]. Nowadays, these two species, together with S. delphini [15], are considered to form the so called S. intermedius Group (SIG) [16,17]. Based on findings by Authors [15,18] that investigated SIG strains from different countries by multilocus sequence typing, it has been proposed currently, that all canine strains be reported as S. pseudintermedius, unless genomic analyses prove they belong to another species [19]. S. pseudintermedius has been recognized as an opportunistic pathogen in several animal species, especially dogs and cats. It is mostly associated with canine pyoderma, otitis externa, wound and urinary tract infections, as well as other kinds of infections in pets [20]. To date, in birds, S. pseudintermedius has been isolated from healthy pigeons [15]. Some recent reports indicated that it could occasionally cause human colonization and, sporadically, infection suggesting that S. pseudintermedius may be a zoonotic pathogen of public health concern [21]. Unlike S. aureus, S. pseudintermedius colonization is unusual in humans, even among individuals with frequent contact with animals. When studying 144 healthy veterinary college staff members, the authors found one only person to harbor the organism [22]. Again, among 3397 CoPS isolates from hospitalized patients, only two S. pseudintermedius isolates were identified [23]. However,

46  Pet-to-Man Travelling Staphylococci: A World in Progress

by reanalyzing 14 isolates from human dog-bite wounds that were originally identified as S. aureus, three strains (22%) could be reclassified as S. pseudintermedius [24], highlighting that the two species may be misidentified as each other, and the real incidence in humans might be consequently underestimated. A study on S. pseudintermedius prevalence in 13 dogs affected by deep pyoderma, their owners, and 13 individuals without daily dog contact [25] showed that the occurrence in owners of ill dogs was significantly higher (6/13) than in the control group (1/13) and that owners often carried the same S. pseudintermedius strains as their pets. Interestingly, all individuals were sampled a second time and were found not to be carriers when the dogs no longer had purulent lesions, so it may be argued that contact with animal lesions (rather than healthy surfaces) is the true risk factor for transmission to humans [25]. S. pseudintermedius is a potentially invasive pathogen in the case of dog bite-related wounds in humans, and was identified in 18% of such infected lesions in Great Britain [26]. In another case, S. pseudintermedius was cultured from the ear fluid of a patient with otitis externa as well as from her pet dog [27]. A methicillin-resistant S. pseudintermedius (MSP) was isolated from 17 dogs and one staff member at a veterinary teaching hospital in Japan. The human isolate shared susceptibility and a genotype profile with some of the dogs, suggesting animal-to-man transmission [28]. Nosocomial spread of MRSP was also described in a veterinary clinic in the Netherlands [29]. The strain was isolated from infected surgical wounds of five dogs and one cat, and all patients had undergone surgery at the same facility. Again, four of 22 environmental samples and four of seven persons working at the clinic were found to be MRSP carriers. The genomic profiles of the isolates were indistinguishable, and it was concluded the isolates were genotypically related, and that a nosocomial epidemic had occurred. As dogs and cats had had no contact with each other, it seemed likely that the veterinary operators (surgeons, nurses) were the indirect source for the infections. Good practice and careful hygiene are therefore necessary to prevent multidrug-resistant isolates’ spread in veterinary facilities.

4.3  COAGULASE NEGATIVE STAPHYLOCOCCI CoNS have become the most common bacterial pathogens isolated from milk samples in many countries and are responsible for bovine intramammary infections; therefore, they might be considered as emerging agents of mastitis [30]. They are opportunistic bacteria able to adhere to metal devices, thus producing a protective biofilm. This ability enables CoNS to persist on milking equipment as well as on the milker’s hands, which may be a major source of staphylococcal spread. CoNS have traditionally been considered to be part of the normal skin microbiota; in this context, as opportunistic bacteria, they may adopt a pathogenic behavior and cause mastitis. In dairy farms that have successfully controlled mammary gland infections S. aureus, CoNS have become frequently encountered causes of bovine mastitis [31].

CoPS and CoNS Animal Diseases  Chapter | 4  47

To date, several CoNS species have been described. Staphylococcus chromogenes belong to the bovine skin microbiota and is the most commonly isolated CoNS species found in bovine milk, especially in heifers around calving and in first lactation. Staphylococcus simulans, a commonly isolated organism in mastitic milk samples, has been reported to show propensity to cause a stronger inflammatory reaction than other CoNS species. Staphylococcus agnetis, again, is a recently described bovine-associated coagulase-variable species; it is closely related to and was initially classified as Staphylococcus hyicus, coaulase expression of which is strain-dependent [16], and that has been reported to cause a particularly strong inflammatory reaction in bovine mammary glands [30]. Furthermore, some CoNS colonizing animal skin and mucous membrane as harmless inhabitants are now recognized to be implicated in human skin and soft tissue infections as well as bacteremia. Among them, Staphylococcus haemolyticus, Staphylococcus capitis, and Staphylococcus xylosus are commensals of farm animals, but can cause opportunistic infections in man [32]. CoNS pathogenicity has long been underestimated as they were associated with chronic or subacute infections that were milder when compared with those by CoPS species. Nonetheless, the etiological role of CoNS in prosthesis and foreign body infections is increasingly being recognized in human medicine [33]. In pets, the pathogenic potential of these microorganisms has not yet been clearly defined, although there have been some reports of infections related to methicillin-resistant CoNS in these hosts. To date, CoNS strains in dogs and cats have been neglected, although the recent development of new molecular techniques has allowed accurate CoNS identification [34], leading to a better understanding of them. Further knowledge of CoNS carriage in animals will be of benefit as these bacteria might represent a reservoir of antibiotic resistance traits that might be transmitted to CoPS, especially the mecA element. This assumption was mainly based on studies reporting antibiotic resistance in clinical CoPS isolates from dogs and humans living in close contact with pets [35,36]. However, a clear picture of CoPS and CoNS distribution, diversity, and multidrug resistance in pets as well as the role of dogs and cats as drug resistance reservoirs is still lacking nowadays.

4.4  CONCLUDING REMARKS Staphylococci are pathogens whose versatility in terms of infections and hosts makes them a serious threat to animal and human health, food security, and consequently, public health. Furthermore, they have a notable ability to develop resistance to antimicrobial agents, and the issue of antibiotic resistance in this group of microorganisms has to be addressed by the existing antibiotics, as well as the development of novel molecules. The genomic analysis of staphylococci isolated from animals (from cows and poultry, mainly) may be used to ­demonstrate the evolutionary origin of some clones from human beings and explain the molecular mechanisms of host adaptation [2]. These approaches have to be extended to other livestock isolates, and population genomic analysis may

48  Pet-to-Man Travelling Staphylococci: A World in Progress

contribute to provide a better understanding as to how and why some clones spread so efficiently in a given host population. Despite host adaptation and specialization, some strains seem instead to lack specific host tropism and can easily be transmitted from animals to humans and vice versa [36]. This raises the question of whether staphylococcal infections should be considered zoonoses or humanoses [2]; nevertheless, this also means that staphylococci may be used for studies in the framework of the One Health concept, involving cooperation among experts in animal, human, and public health sciences.

CONFLICT OF INTEREST None.

REFERENCES [1] Weese JS, van Duijkeren E. Methicillin-resistant Staphylococcus aureus and Staphylococcus pseudintermedius in veterinary medicine. Vet Microbiol 2010;140(3–4):418–29. [2] Peton  V, Le Loir  Y. Staphylococcus aureus in veterinary medicine. Infect Genet Evol 2014;21:602–15. [3] Bergonier D, de Crémoux R, Rupp R, Lagriffoul G, Berthelot X. Mastitis of dairy small ruminants. Vet Res 2003;34(5):689–716. [4] Le Loir Y, Baron F, Gautier M. Staphylococcus aureus and food poisoning. Genet Mol Res 2003;2(1):63–76. [5] Le Maréchal  C, Seyffert  N, Jardin  J, et  al. Molecular basis of virulence in Staphylococcus aureus mastitis. PLoS ONE 2011;6(11):e27354. [6] Guinane CM, Sturdevant DE, Herron-Olson L, et al. Pathogenomic analysis of the common bovine Staphylococcus aureus clone (ET3): emergence of a virulent subtype with potential risk to public health. J Infect Dis 2008;197(2):205–13. [7] Zhao  X, Lacasse  P. Mammary tissue damage during bovine mastitis: causes and control. J Anim Sci 2008;86(Suppl 13):57–65. [ 8] Pereira  UP, Oliveira  DG, Mesquita  LR, Costa  GM, Pereira  LJ. Efficacy of Staphylococcus aureus vaccines for bovine mastitis: A systematic review. Vet Microbiol 2011;148(2-4):117–24. [9] Klostermann K, Crispie F, Flynn J, Ross RP, Hill C, Meaney W. Intramammary infusion of a live culture of Lactococcus lactis for treatment of bovine mastitis: comparison with antibiotic treatment in field trials. J Dairy Res 2008;75(3):365–73. [10] Bouchard DS, Rault L, Berkova N, Le Loir Y, Evan S. Inhibition of Staphylococcus aureus invasion into bovine mammary epithelial cells by contact with live Lactobacillus casei. Appl Environ Microbiol 2013;79(3):877–85. [11] Rupp R, Bergonier D, Dion S, et al. Response to somatic cell count-based selection for mastitis resistance in a divergent selection experiment in sheep. J Dairy Sci 2009;92(3):1203–19. [12] Wall RJ, Powell AM, Paape MJ, et al. Genetically enhanced cows resist intramammary Staphylococcus aureus infection. Nat Biotechnol 2005;23(4):445–51. [13] Hajek V. Staphylococcus intermedius, a new species isolated from animals. Int J Syst Bacteriol 1976;26(4):401–8. [14] Devriese  LA, Vancanneyt  M, Baele  M, et  al. Staphylococcus pseudintermedius sp. nov., a coagulase-positive species from animals. Int J Syst Evol Microbiol 2005;55(4):1569–73.

CoPS and CoNS Animal Diseases  Chapter | 4  49 [15] Sasaki  T, Kikuchi  K, Tanaka  Y, Takahashi  N, Kamata  S, Hiramatsu  K. Reclassification of phenotypically identified Staphylococcus intermedius strains. J Clin Microbiol 2007;45(9):2770–8. [16] Savini V, Passeri C, Mancini G, et al. Coagulase-positive staphylococci: my peťs two faces. Res Microbiol 2013;164(5):371–4. [17] Savini V, Barbarini D, Polakowska K, et al. Methicillin-resistant Staphylococcus pseudintermedius infection in a bone marrow transplant recipient. J Clin Microbiol 2013;51(5):1636–8. [18] Bannoehr J, Ben Zakour NL, Waller AS, et al. Population genetic structure of the Staphylococcus intermedius group: insights into agr diversification and the emergence of methicillinresistant strains. J Bacteriol 2007;189(23):8685–92. [19] Devriese LA, Hermans K, Baele M, Haesebrouck F. Staphylococcus pseudintermedius versus Staphylococcus intermedius. Vet Microbiol 2009;133(1–2):206–7. [20] Ruscher C, Lübke-Becker A, Wleklinski CG, Soba A, Wieler LH, Walther B. Prevalence of methicillin-resistant Staphylococcus pseudintermedius isolated from clinical samples of companion animals and equidaes. Vet Microbiol 2009;136(1-2):197–201. [21] van Duijkeren  E, Kamphuis  M, van der Mije  IC, et  al. Transmission of methicillin-­ resistant Staphylococcus pseudintermedius between infected dogs and cats and contact pets, humans and the environment in households and veterinary clinics. Vet Microbiol 2011;150(3-4):338–43. [22] Talan DA, Staatz D, Staatz A, Overturf GD. Frequency of Staphylococcus intermedius as human nasopharyngeal flora. J Clin Microbiol 1989;27(10):2393. [23] Mahoudeau I, Delabranche X, Prevost G, Monteil H, Piemont Y. Frequency of isolation of Staphylococcus intermedius from humans. J Clin Microbiol 1997;35(8):2153–4. [24] Talan DA, Goldstein EJ, Staatz D, Overturf GD. Staphylococcus intermedius: clinical presentation of a new human dog bite pathogen. Ann Emerg Med 1989;18(4):410–3. [25] Guardabassi  L, Loeber  ME, Jacobson  A. Transmission of multiple antimicrobial-resistant Staphylococcus intermedius between dogs affected by deep pyoderma and their owners. Vet Microbiol 2004;98(1):23–7. [26] Lee J. Staphylococcus intermedius isolated from dog-bite wounds. J Infect 1994;29(1):105. [27] Tanner MA, Everett CL, Youvan DC. Molecular phylogenetic evidence for noninvasive zoonotic transmission of Staphylococcus intermedius from a canine pet to a human. J Clin Microbiol 2000;38(4):1628–31. [28] Sasaki T, Kikuchi K, Tanaka Y, Takahashi N, Kamata S, Hiramatsu K. Methicillin-­resistant Staphylococcus pseudintermedius in a veterinary teaching hospital. J Clin Microbiol 2007;45:1118–25. [29] van Duijkeren  E, Houwers  DJ, Schoormans  A, et  al. Transmission of methicillin-resistant Staphylococcus intermedius between humans and animals. Vet Microbiol 2008;128:213–5. [30] Simojoki H, Orro T, Taponen S, Pyörälä S. Host response in bovine mastitis experimentally induced with Staphylococcus chromogenes. Vet Microbiol 2009;134:95–9. [31] Ruegg  PL. The quest for the perfect test: phenotypic versus genotypic identification of ­coagulase-negative staphylococci associated with bovine mastitis. Vet Microbiol 2009;134:15–9. [32] Åvall-Jääskeläinen S, Koort J, Simojoki H, Taponen S. Bovine-associated CNS species resist phagocytosis differently. BMC Vet Res 2013;9:227. [33] Piette  A, Verschraegen  G. Role of coagulase-negative staphylococci in human disease. Vet Microbiol 2009;134:45–54. [34] Gandolfi-Decristophoris P, Regula G, Petrini O, Zinsstag J, Schelling E. Prevalence and risk factors for carriage of multi-drug resistant staphylococci in healthy cats and dogs. J Vet Sci 2013;14:449–56.

50  Pet-to-Man Travelling Staphylococci: A World in Progress [35] Weese JS, Dick H, Willey BM, et al. Suspected transmission of methicillin-resistant Staphylococcus aureus between domestic pets and humans in veterinary clinics and in the household. Vet Microbiol 2006;115:148–55. [36] Somavaji R, Privantha MA, Rubin JE, et al. Human infections due to Staphylococcus pseudintermedius, an emerging zoonosis of canine origin: report of 24 cases. Diagn Microbiol Infect Dis 2016;85:471–6.