Methicillin-resistant Staphylococcus aureus ST398 in humans and animals, Central Europe.Methicillin-resistant Staphylococcus aureus methicillin-resistant Staphylococcus aureus Methicillin-aminoglycoside resistant Staphylococcus aureus, MRSA An organism with multiple antibiotic resistances–eg, aminoglycosides, chloramphenicol, clindamycin, erythromycin, rifampin, tetracycline, of clonal lineage ST398 that exhibits related spa types and contains SCCmec elements of types IVa or V has been isolated from colonized Colonized This occurs when a microorganism is found on or in a person without causing a disease. Mentioned in: Isolation and infected humans and companion animals (e.g., dog, pig, horse) in Germany and Austria. Of particular concern is the association of these cases with cases of nosocomial nosocomial /noso·co·mi·al/ (nos?o-ko´me-il) pertaining to or originating in a hospital. nos·o·co·mi·al adj. 1. Of or relating to a hospital. 2. ventilator-associated pneumonia Ventilator-associated pneumonia (VAP) is a sub-type of hospital-acquired pneumonia (HAP) which occurs in people who are on mechanical ventilation through an endotracheal or tracheostomy tube for at least 48 hours. . ********** Methicillin-resistant Staphylococcus aureus (MRSA MRSA Methicillin-resistant Staphylococcus aureus. See MARSA. ) has become an infection control problem in hospitals worldwide, mainly associated with intrahospital and interhospital dissemination of particular epidemic clonal lineages of the S. aureus The aureus (pl. aurei) was a gold coin of ancient Rome valued at 25 silver denarii. The aureus was regularly issued from the 1st century BC to the beginning of the 4th century AD, when it was replaced by the solidus. population (hMRSA; [1]). MRSA primarily associated with healthcare facilities may also be disseminated to the community through colonized medical staff or discharged patients. The emergence and spread of MRSA in the community during the past 5 years, independent of the healthcare setting and in the absence of typical risk factors for nosocomial MRSA infections, are matters of further concern. These community-acquired MRSA infections are less broadly resistant to antimicrobial agents Antimicrobial agents Chemical compounds biosynthetically or synthetically produced which either destroy or usefully suppress the growth or metabolism of a variety of microscopic or submicroscopic forms of life. than are healthcare-associated MRSA and often contain the determinants lukS-lukF, which code for Panton-Valentine leukocidin Panton-Valentine leukocidin a nonhemolytic toxin produced by Staphylococcus aureus which kills segmented neutrophils and macrophages. (2). Even though MRSA has been known as a nosocomial pathogen for >30 years, its development in companion animals and livestock has been rare (3). Recent reports, however, have documented MRSA infections in animals such as horses from Canada (4) and Europe (3) and pets (5,6). Of particular interest is whether MRSA may be transmitted between animals and humans. MRSA of clonal lineage sequence type (ST) 22 is widely disseminated in human hospitals in the United Kingdom  This article has recently been split into multiple articles entitled:
We report on molecular characterization of MRSA, from sporadic infections in humans and in various animal species, that belong to clonal lineage ST398 according to according to prep. 1. As stated or indicated by; on the authority of: according to historians. 2. In keeping with: according to instructions. 3. multilocus sequence typing Multilocus sequence typing (MLST) is a technique in molecular biology for the typing of multiple loci. The procedure characterizes isolates of bacterial species using the DNA sequences of internal fragments of multiple (usually seven) housekeeping genes. (MLST MLST Multi Locus Sequence Typing MLST Medical Logistics Support Team MLST Mini Losi Super Truck (1/18th scale radio control vehicle) ). These isolates were further characterized by spa-sequence typing (repeat polymorphism polymorphism, of minerals, property of crystallizing in two or more distinct forms. Calcium carbonate is dimorphous (two forms), crystallizing as calcite or aragonite. Titanium dioxide is trimorphous; its three forms are brookite, anatase (or octahedrite), and rutile. of the X-region of the spa gene) and by PCR PCR polymerase chain reaction. PCR abbr. polymerase chain reaction Polymerase chain reaction (PCR) for grouping of staphylococcal staphylococcal pertaining to Staphylococcus spp. staphylococcal clumping test used as a means of measuring the quantity of fibrinogen-split products in a sample of blood. cassette chromosome mec (SCCmec) elements, which contain the mecA gene and of which at least 5 basic types have been described. Methods MRSA isolates from infections in humans and in animals were sent to the National Reference Center for Staphylococci staph·y·lo·coc·cus n. pl. staph·y·lo·coc·ci A spherical gram-positive parasitic bacterium of the genus Staphylococcus, usually occurring in grapelike clusters and causing boils, septicemia, and other infections. at the Robert Koch Institute, Wernigerode Branch, in Germany, for typing by means of SmaI-macrorestriction pattern as well as spa typing. Selected isolates also underwent MLST. Four human isolates were grown from nasal swabs taken from the staff of a veterinary practice at Veterinary Analytical Center, Geesthacht, Germany. All isolates were primarily grown on sheep blood agar blood agar n. A nutrient culture medium that is enriched with whole blood and used for the growth of certain strains of bacteria. and confirmed by standard procedures as S. aureus. Eleven additional MRSA specimens of lineage ST398 (1 isolate per patient affected) were found among 4,370 MRSA isolates from patients with recognized infections. These isolates were identified by indigestibility in·di·gest·i·ble adj. Difficult or impossible to digest: an indigestible meal. in of their whole cellular DNA DNA: see nucleic acid. DNA or deoxyribonucleic acid One of two types of nucleic acid (the other is RNA); a complex organic compound found in all living cells and many viruses. It is the chemical substance of genes. when subjected to SmaI-macrorestriction analysis. Animal isolates were collected from 1 dog and 1 foal foal a junior horse from birth to one year. May be filly foal, colt foal. foal ataxia see enzootic equine incoordination. at the Veterinary Analytical Center, Geesthacht, Germany; from 1 pig at the diagnostic laboratory of the Institute for Microbiology and Infectious Diseases infectious diseases: see communicable diseases. , School of Veterinary Medicine veterinary medicine, diagnosis and treatment of diseases of animals. An early interest in animal diseases is found in ancient Greek writings on medicine. Veterinary medicine began to achieve the stature of a science with the organization of the first school in the , Hannover, Germany; and from 2 horses at the Department of Orthopaedics, Veterinary University, Vienna, Austria. Procedures and primers for DNA extraction DNA extraction is a routine procedure to collect DNA for subsequent molecular or forensic analysis. Outline of a DNA extraction There are three basic steps in a DNA extraction, the details of which may vary depending on the type of sample and any substances that may and PCR detection of resistance genes were as described previously (6). Macrorestriction patterns were determined by using lysis lysis /ly·sis/ (li´sis) 1. destruction or decomposition, as of a cell or other substance, under influence of a specific agent. 2. mobilization of an organ by division of restraining adhesions. 3. of cells, deproteinization and digestion of DNA (here by SmaI and ApaI), and pulsed-field gel electrophoresis (7). The polymorphic polymorphic - polymorphism X-region of the protein A gene (spa) was amplified and sequenced according to the Ridom StaphType standard protocol (www.ridom.org). The resulting spa-types were assigned by using the Ridom StaphType software package (Ridom GmbH, Wurzburg, Germany). The BURP algorithm, implemented in the most recent Ridom StaphType software version, was used for cluster analysis Cluster analysis A statistical technique that identifies clusters of stocks whose returns are highly correlated within each cluster and relatively uncorrelated across clusters. Cluster analysis has identified groupings such as growth, cyclical, stable, and energy stocks. of spa types (7). Primers used for MLST correspond to the protocol as described previously (8), with the exception of the forward primer for tpi; we used the sequence tpif 5'-GCAT TAGCAGATTTAGGCGT-3'. Antimicrobial susceptibility testing was performed by broth microdilution, performed according to DIN 58940, Deutsches Institut fur Normung (9). SCCmec elements of types I to IV were characterized by using a PCR approach, including a combination of different PCRs (6). To demonstrate SCCmec-elements of type V, we used primers type VF/type VR, as described by Zhang et al. (10), as well primer pair ccrC9f 5'-CACTTAATCCATGTACACAG-3' and ccrC-R (10). The following set of primers was used for PCR for virulence-associated genes: tst, sea, seb, sec, sed, see, as described by Johnson et al. (11); for lukS-lukF, forward 5'ATCATTAGGTAAAATGTCTGGACATGATCCA-3', reverse 5'-GCATCAAGTGTATTGGATAGCAAAAGC3'; for cna, forward 5'-CGGTTCCCCCATAAAAGT GAAG-3", reverse 5'-CCCATAGCCTTGTGGATTTG-3'. Annealing annealing (ənēl`ĭng), process in which glass, metals, and other materials are treated to render them less brittle and more workable. temperature was 55[degrees]C; cyclic scheme and further conditions were as reported previously (6). Specimen collection, characterization of the isolates, data processing, and exchange of data were performed within the framework of German public health activities for infection control and prevention of MRSA dissemination. Ethical approval was obtained within this framework as well. Results Characteristics of the 20 MRSA isolates investigated are shown in the Table. All isolates share MLST ST398 with the allelic al·lele n. One member of a pair or series of genes that occupy a specific position on a specific chromosome. [German Allel, short for Allelomorph, allelomorph, from English profile 3-35-19-2-20-26-39. Three different spa-types are obviously related (Figure). Types t11 and t34 may have been derived from each other by either deletion or duplication of 2 repeats; t1197 and t11 differ by a single nucleotide polymorphism Noun 1. single nucleotide polymorphism - (genetics) genetic variation in a DNA sequence that occurs when a single nucleotide in a genome is altered; SNPs are usually considered to be point mutations that have been evolutionarily successful enough to recur in a . BURP analysis of these spa-types groups them as a separate cluster unrelated to other BURP clusters (7). A peculiarity of S. aureus of clonal lineage ST398 is the indigestibility of whole cellular DNA by restriction enzyme restriction enzyme Protein (more specifically, an endonuclease) produced by bacteria that cleaves DNA at specific sites along its length. Thousands have been found, from many different bacteria; each recognizes a specific nucleotide sequence. SmaI. Therefore, SmaI macrorestriction patterns generate only 1 large fragment because of protection by a novel DNA methylation enzyme (12). We also found poor digestion by the isoschizomeric enzyme XmaI. However, digestion by enzyme ApaI generated similar fragment patterns that differed at most by 3 fragments independent of spa types. [FIGURE OMITTED] The 2 horse isolates from the Vienna veterinary university contained SCCmec elements of group IVa. For all other isolates investigated, PCR indicated SCCmecV. These findings suggest that MRSA of ST398 from horses are unrelated to the other isolates and probably have evolved independently by acquisition of a different SCCmec element. In addition to mecA, all investigated isolates contained tetM; isolates from animals and humans from Lower Saxony also contained ermA. The nosocomial human and horse isolates contained ermC; in the horse isolates, aph2"aac6'--mediating aminoglycoside aminoglycoside /ami·no·gly·co·side/ (-gli´ko-sid) any of a group of antibacterial antibiotics (e.g., streptomycin, gentamicin) derived from various species of Streptomyces resistance was demonstrated. PCR was negative for virulence-associated genes and for lukS-lukF (coding for Panton-Valentine leukocidin), tst, sea, seb, seb, and sed, as well as for cna (collagen-binding protein). Discussion Isolates of clonal lineage ST398 seem not to be frequently represented among the S. aureus population. They were not recorded by Grundmann et al. (13) among a population sample of nasal colonizers in the Nottingham area in the United Kingdom and were not found among 108 isolates from carriers in a rural territory in northern Germany (S. Holtfreter et al., unpub, data). Only 2 notations of ST398 are found in the S. aureus MLST database (www.mlst.net), 1 from the Netherlands and 1 from the Cape Verde Islands Noun 1. Cape Verde Islands - a group of islands in the Atlantic off of the coast of Senegal Cape Verde, Republic of Cape Verde - an island country in the Atlantic off the coast of Senegal . Among 11,250 isolates of various origin (colonization and infections in hospitals as well in the community in humans from all Germany) typed from 1992 through 2003, no isolates refractory to SmaI macrorestrition analysis were seen. Therefore, a rather recent emergence of MRSA ST398 among humans seems likely. However, MRSA of lineage ST398 had been reported from infections in pigs and from nasal colonization in pig farmers in France (14). A more recent report from the Netherlands describes MRSA of ST398 (spa t108, which is in the same BURP cluster as t11 and t34) in pigs and in humans who had contact with pigs (15). A comparison of the allelic profile of ST398 by means of the MLST database does not indicate any relationship to profiles of prevalent clonal complexes of methicillin-susceptible S. aureus (13), of epidemic healthcare-associated MRSA, or of lukS-lukF--containing community-associated MRSA from Europe. Conclusions MRSA exhibiting ST398 may colonize col·o·nize v. col·o·nized, col·o·niz·ing, col·o·niz·es v.tr. 1. To form or establish a colony or colonies in. 2. To migrate to and settle in; occupy as a colony. 3. and cause infections in humans and in certain animal species such as dogs, horses, and pigs. The isolation of MRSA ST398 showing the same characteristics from a wound infection in a dog and from nasal colonization of the staff of a veterinary practice where this dog had been treated suggests that interspecies transmission may occur. The differences in spa types between the isolates containing the same PCR results for SCCmec can be explained by a single genetic event. Because isolates taken at the same time from nasal colonization in veterinary staff of the same practice exhibit either spa-type t011 or t034, this difference does not justify discrimination between the two types. Of particular concern was the subsequent detection of MRSA ST398 not only in outpatients but also in inpatients with ventilator-associated pneumonia in the same hospital unit at about the same time (Table). Future recording of MRSA ST398 from infected and colonized humans (especially when detected by screening at admission to hospitals) will require a thorough analysis with respect to association with animals and routes of transmission. Tracing MRSA carriers among contacts should also include pet animals, horses, and other livestock. Because of the time and labor needed to complete MLST, spa-typing combined with BURP analysis of types is an efficient tool for recognizing this clonal lineage. Furthermore, detection of MRSA by appropriate methods should be implemented into antimicrobial resistance surveillance programs in veterinary medicine. References (1.) Enright MC, Robinson DA, Randle G, Feil E J, Grundmann H, Spratt BG. The evolutionary history of methicillin-resistant Staphylococcus aureus (MRSA). Proc Natl Acad Sei (Software Engineering Institute, Pittsburgh, PA, www.sei.cmu.edu) A federally funded research and development center that is under contract to Carnegie Mellon University and is devoted to the advancement of software engineering and the quality of software support systems. U S A. 2002;99:7687-92. (2.) Vandenesch F, Naimi T, Enright MC, Lina G, Nimmo GR, Heffeman H, et al. Community acquired methicillin-resistant Staphylococcus staphylococcus (stăf'ələkŏk`əs), any of the pathogenic bacteria, parasitic to humans, that belong to the genus Staphylococcus. The spherical bacterial cells (cocci) typically occur in irregular clusters [Gr. anreus carrying Panton-Valentin leukocidin genes: worldwide emergence. Emerg Infect Dis. 2003;9:978-84. (3.) Cuny C, Kuemmerle J, Stanek C, Willey B, Strommenger B, Witte W. Emergence of MRSA infections in horses in a veterinary hospital: strain characterization and comparison with MRSA from humans. Euro Surveill. 2006;11:44-7. (4.) Weese JS, Archambault M, Willey BM, Hearn P, Kreiswirth BN, Said-Salim B, et al. Methicillin-resistant Staphylococcus aureus in horses and horse personnel, 2000-2002. Emerg Infect Dis. 2005; 11:430-5. (5.) Loeffler A, Boag AK, Sung J, Lindsay JA, Guardabassi L, Dalsgaard A, et al. Prevalence of methicillin-resistant Staphylococcus aureus among staff and pets in a small animal referral hospital in the UK. J Antimicrob Chemother. 2005;56:692-7. (6.) Strommenger B, Kehrenberg C, Kettlitz C, Cuny C, Verspohl J, Witte W, et al. Molecular characterization of methicillin-resistant Staphylococcus aureus strains from pet animals and their relationship to human isolates. J Antimicrob Chemother. 2006;57:461-5. (7.) Strommenger B, Kettlitz C, Weniger T, Harmsen D, Friedrich AW, Witte W. Assignment of Staphylococcus aureus Staphylococcus au·re·us n. A bacterium that causes furunculosis, pyemia, osteomyelitis, suppuration of wounds, and food poisoning. Staphylococcus aureus Staphylococcus pyogenes isolates to groups by spa-typing, SmaI-macrorestriction analysis, and multilocus sequence typing. J Clin Microbiol. 2006;44:2533-40. (8.) Enright MC, Day NP, Davies CE, Peacock SJ, Spratt BG. Multilocus sequence typing for characterization of methicillin-resistant and methicillin-susceptible clones of Staphylococcus aureus. J Clin Microbiol. 2000;38:1008-15. (9.) German Institute for Standardization. Methods for susceptibility testing of bacterial pathogens (besides mycobacteria mycobacteria members of the genus Mycobacterium. anonymous mycobacteria see opportunist (atypical) mycobacteria (below). nontubercular mycobacteria see opportunist (atypical) mycobacteria (below). ) against chemotherapeutics. DIN 58940, Part 8. Microdilution. In: Medical microbiology and immunology: diagnostic procedures. Vienna: Barth Publishing; 2000. p. 215-27. (10.) Zhang K, McClure JA, Elsayed S, Louie T, Conley JM. Novel multiplex PCR assay for characterization and concomitant subtyping of staphylococcal cassette chromosome mec types I to V in methicillin-resistant Staphylococcus aureus. J Clin Microbiol. 2005;43:5026-33. (11.) Johnson WM, Tyler SD, Ewan EP, Aston FE, Pollard DR, Kozee KR. Detection of genes for enterotoxins, exfoliative ex·fo·li·a·tive adj. Marked by exfoliation, desquamation, or profuse scaling. toxins, and toxic shock syndrome toxic shock syndrome (TSS). acute, sometimes fatal, disease characterized by high fever, nausea, diarrhea, lethargy, blotchy rash, and sudden drop in blood pressure. It is caused by Staphylococcus aureus, an exotoxin-producing bacteria (see toxin). toxin I in Staphylococcus aureus by the polymerase chain reaction polymerase chain reaction (pŏl`ĭmərās') (PCR), laboratory process in which a particular DNA segment from a mixture of DNA chains is rapidly replicated, producing a large, readily analyzed sample of a piece of DNA; the process is . J Clin Microbiol. 1991;29:426-30. (12.) Bens CC, Voss A, Klaassen CH. Presence of a novel DNA methylation enzyme in methicillin-resistant Staphylococcus attreus associated with pig farming leads to uninterpretable results in standard pulsed-field gel electrophoresis analysis. J Clin Microbiol. 2006;44:1875-6. (13.) Grundmann H, Hori S, Enright MC, Webster C, Tami A, Feil EJ, et al. Determining the genetic structure of the natural population of Staphylococcus aureus: a comparison of multilocus sequence typing with pulsed-field gel etectrophoresis, randomly amplified polymorphic DNA analysis DNA analysis Any technique used to analyze genes and DNA. See Chromosome walking, DNA fingerprinting, Footprinting, In situ hybridization, Jeffries' probe, Jumping libraries, PCR, RFLP analysis, Southern blot hybridization. , and phage phage: see bacteriophage. phage - A program that modifies other programs or databases in unauthorised ways; especially one that propagates a virus or Trojan horse. See also worm, mockingbird. The analogy, of course, is with phage viruses in biology. typing. J Clin Microbiol. 2002;40:4544-6. (14.) Armand-Lefevre L. Ruimy R, Andremont A. Clonal comparison of Staphylococcus aureus isolates from healthy pig farmers, human controls, and pigs. Emerg Infect Dis. 2005; 11: 711-4. (15.) van Diijke B, Kopgen H, Wannet W, Huisdens X, Neeling H, Voss A. Methicillin-resistant Staphylococcus aureus and pig fanning [abstract]. 16th European Congress of Clinical Microbiology and Infectious Diseases. 2006 April 1-4; Nice, France [cited 2007 Jan 10]. Available from http://www.blackwellpublishing.com/ eccmid16/abstract.asp?id=49270 Address for correspondence: Wolfgang Witte, Robert Koch Institute, Wernigerode Br, BurgstraBe 37, 38855 Wernigerode, Germany; email: wittew@rki.de Wolfgang Witte, * Birgit Strommenger, * Christian Stanek, ([dagger]) and Christiane Cuny ([dagger]) * Robert Koch Institute, Wernigerode, Germany; and ([dagger]) Veterinary University, Vienna, Austria Prof Witte is head of the laboratory for nosocomial infections Nosocomial infections Infections that were not present before the patient came to a hospital, but were acquired by a patient while in the hospital. Mentioned in: Enterobacterial Infections, Staphylococcal Infections at the Robert Koch Institute, Wernigerode Branch. He also is a professor on the medical faculty of Magdeburg University.
Table. Typing characteristics of methicillin-resistant Staphylococcus
aureus of clonal lineage ST398, Central Europe
Carrier; type No.
Country/area of infection isolates
Germany
Lower Saxony ([dagger]) Dog; skin infection 1
Human ([double 1
dagger]) (n = 4);
nasal
carriage 3
Pig; colonization 1
Foal; sinusitis 1
Schleswig-Holstein Human, infection of 1
skin (outpatient)
Lower Saxony ([dagger]) Human; wound 1
infection
Human; nasal 1
colonization
([section])
Hesse Human; nasal 1
colonization
([section])
Saxony-Anhalt Human; ventilator- 1
associated pneumonia
Saxony Human (n = 2); 2
ventilator-
associated
nosocomial
pneumonia
([paragraph])
Baden-Wurttemberg Human (n = 4); 4
ventilator-
associated
nosocomial
pneumonia (#)
Austria
Vienna Horse (n = 2); wound 2
infection
SCCmec
Country/area spa-type group
Germany
Lower Saxony ([dagger]) t034 V
t011 V
t034
t034 V
t1197 V
Schleswig-Holstein t011
Lower Saxony ([dagger]) t011 V
t034 V
Hesse t034 V
Saxony-Anhalt t034 V
Saxony t011 V
Baden-Wurttemberg t011 V
Austria
Vienna t011 IVa
Resistance
Country/area phenotype * Resistance genes
Germany
Lower Saxony ([dagger]) PEN, OXA, ERY, mecA, ermA, tetM
CLI, OTE
PEN, OXA, ERY, mecA, ermA, tetM
CLI, OTE
PEN, OXA, ERY, mecA, ermA, tetM
CLI, OTE
PEN, OXA, OTE mecA, tetM
Schleswig-Holstein PEN, OXA, OTE mecA, tetM
Lower Saxony ([dagger]) PEN, OXA, OTE, mecA, tetM
CIP
PEN, OXA, ERY, mecA, ermA, tetM
CLI, OTE
Hesse PEN, OXA, OTE mecA, ermA, tetM
Saxony-Anhalt PEN, OXA, ERY, mecA, ermC, tetM
CLI, OTE
Saxony PEN, OXA, ERY, mecA, ermC, tetM
CLI, OTE, CIP
Baden-Wurttemberg PEN, OXA, GEN, mecA, ermC, tetM
ERY, CLI, OTE
Austria
Vienna PEN, OXA, GEN, mecA, ermC, tetM,
ERY, CLI, OTE aph2"-aac6'
* PEN, penicillin, GEN, gentamicin; ERY, erythromycin; CLI,
clindamycin; OTE, oxytetracycline; CIP, ciprofloxacin.
([dagger]) Two Lower Saxony regions are listed separately because the
isolates originated from different locations within the region and,
although belonging to the same clonal lineage, exhibited different
spa-sequence types and resistance traits with regard to ermA and
erythromycin resistance.
([double dagger]) Staff members who worked in the practice where the
dog was treated.
([section]) Colonization detected upon hospital admission.
([paragraph]) Infections at the same ward within a 2-week period.
(#) Infections at the same ward within a 10-day period.
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