Summary: Staphylococcus aureus is an important cause of skin and soft-tissue infections (SSTIs), endovascular infections, pneumonia, septic arthritis, endocarditis, osteomyelitis, foreign-body infections, and sepsis. Methicillin-resistant S. aureus (MRSA) isolates were once confined largely to hospitals, other health care environments, and patients frequenting these facilities. Since the mid-1990s, however, there has been an explosion in the number of MRSA infections reported in populations lacking risk factors for exposure to the health care system. This increase in the incidence of MRSA infection has been associated with the recognition of new MRSA clones known as community-associated MRSA (CA-MRSA). CA-MRSA strains differ from the older, health care-associated MRSA strains; they infect a different group of patients, they cause different clinical syndromes, they differ in antimicrobial susceptibility patterns, they spread rapidly among healthy people in the community, and they frequently cause infections in health care environments as well. This review details what is known about the epidemiology of CA-MRSA strains and the clinical spectrum of infectious syndromes associated with them that ranges from a commensal state to severe, overwhelming infection. It also addresses the therapy of these infections and strategies for their prevention.

Community-associated methicillin-resistant Staphylococcus aureus: epidemiology and clinical consequences of an emerging epidemic.

The definition of the heterogeneous group of coagulase-negative staphylococci (CoNS) is still based on diagnostic procedures that fulfill the clinical need to differentiate between Staphylococcus aureus and those staphylococci classified historically as being less or nonpathogenic. Due to patient- and procedure-related changes, CoNS now represent one of the major nosocomial pathogens, with S. epidermidis and S. haemolyticus being the most significant species. They account substantially for foreign body-related infections and infections in preterm newborns. While S. saprophyticus has been associated with acute urethritis, S. lugdunensis has a unique status, in some aspects resembling S. aureus in causing infectious endocarditis. In addition to CoNS found as food-associated saprophytes, many other CoNS species colonize the skin and mucous membranes of humans and animals and are less frequently involved in clinically manifested infections. This blurred gradation in terms of pathogenicity is reflected by species- and strain-specific virulence factors and the development of different host-defending strategies. Clearly, CoNS possess fewer virulence properties than S. aureus, with a respectively different disease spectrum. In this regard, host susceptibility is much more important. Therapeutically, CoNS are challenging due to the large proportion of methicillin-resistant strains and increasing numbers of isolates with less susceptibility to glycopeptides.

Coagulase-Negative Staphylococci

In recent years, methicillin-resistant Staphylococcus aureus (MRSA) have become a truly global challenge. In addition to the long-known healthcare-associated clones, novel strains have also emerged outside of the hospital settings, in the community as well as in livestock. The emergence and spread of virulent clones expressing Panton-Valentine leukocidin (PVL) is an additional cause for concern. In order to provide an overview of pandemic, epidemic and sporadic strains, more than 3,000 clinical and veterinary isolates of MRSA mainly from Germany, the United Kingdom, Ireland, France, Malta, Abu Dhabi, Hong Kong, Australia, Trinidad & Tobago as well as some reference strains from the United States have been genotyped by DNA microarray analysis. This technique allowed the assignment of the MRSA isolates to 34 distinct lineages which can be clearly defined based on non-mobile genes. The results were in accordance with data from multilocus sequence typing. More than 100 different strains were distinguished based on affiliation to these lineages, SCCmec type and the presence or absence of PVL. These strains are described here mainly with regard to clinically relevant antimicrobial resistance- and virulence-associated markers, but also in relation to epidemiology and geographic distribution. The findings of the study show a high level of biodiversity among MRSA, especially among strains harbouring SCCmec IV and V elements. The data also indicate a high rate of genetic recombination in MRSA involving SCC elements, bacteriophages or other mobile genetic elements and large-scale chromosomal replacements.

/pdf/a-field-guide-to-pandemic-epidemic-and-sporadic-clones-of-2fj5koe4iu.pdf

A field guide to pandemic, epidemic and sporadic clones of methicillin-resistant Staphylococcus aureus

Meticillin-resistant Staphylococcus aureus (MRSA): global epidemiology and harmonisation of typing methods

https://dspace.library.uu.nl/bitstream/handle/1874/315400/Weese%20et%20al%202009.pdf?sequence=1

Methicillin-resistant Staphylococcus aureus and Staphylococcus pseudintermedius in veterinary medicine.

Reverse transcription‑quantitative polymerase chain reaction (RT‑qPCR) is the gold standard method for the diagnosis of COVID‑19 infection. Due to pre‑analytical and technical limitations, samples with low viral load are often misdiagnosed as false‑negative samples. Therefore, it is important to evaluate other strategies able to overcome the limits of RT‑qPCR. Blinded swab samples from two individuals diagnosed positive and negative for COVID‑19 were analyzed by droplet digital PCR (ddPCR) and RT‑qPCR in order to assess the sensitivity of both methods. Intercalation chemistries and a World Health Organization (WHO)/Center for Disease Control and Prevention (CDC)‑approved probe for the SARS‑CoV‑2 N gene were used. SYBR‑Green RT‑qPCR is not able to diagnose as positive samples with low viral load, while, TaqMan Probe RT‑qPCR gave positive signals at very late Ct values. On the contrary, ddPCR showed higher sensitivity rate compared to RT‑qPCR and both EvaGreen and probe ddPCR were able to recognize the sample with low viral load as positive even at 10‑fold diluted concentration. In conclusion, ddPCR shows higher sensitivity and specificity compared to RT‑qPCR for the diagnosis of COVID‑19 infection in false‑negative samples with low viral load. Therefore, ddPCR is strongly recommended in clinical practice for the diagnosis of COVID‑19 and the follow‑up of positive patients until complete remission.

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Sensitivity assessment of droplet digital PCR for SARS-CoV-2 detection.

The aim of our study was to trace the dynamic changes of hospital-associated methicillin-resistant Staphylococcus aureus (HA-MRSA) lineages in Italy, comparing the genotypic backgrounds of contemporary isolates over a period of 17 years, with those of a sample of early MRSA strains from 1980. In total, 301 non-repetitive MRSA clinical isolates, recovered from 19 Italian hospitals between 1990 and 2007 were selected and analyzed for their antibiotic resistance, typed by PFGE and SCCmec, grouped into clonal-types and further characterized using Multi Locus Sequence Typing (MLST). A sample of fifteen early MRSA strains from 1980 was also used for comparison. The most interesting feature was the recent increase of ST228-MRSA-I (formerly the Italian clone; PFGE E) over the period 2000–2007 (57%), when compared to the period 1990–1999 (29%), and its stability to date, associated with a decrease of the highly epidemic ST247-MRSA-IA (formerly the Iberian clone; PFGE A), (23% from 1990 to 1999, 6% from 2000 to 2007). ST1-MRSA-I (1 out of 2 strains carrying ccr A2B2), ST8-MRSA-I (4 strains), ST15-MRSA-I (1 out of 4 carrying ccr A2B2) and ST30-MRSA-I (2 out of 5 carrying no ccr AB-types and ccr C) were the predominant earliest STs among the MRSA strains in 1980. A temporal shift in the susceptibility levels to glycopeptides was observed: strains with vancomycin MIC of ≥ 2 mg/L increased from 19.4% to 35.5%. In conclusion, we describe the alternation of MRSA clones that occurred in hospitals from 1990 to 2007 and the increase of the glycopeptide MIC levels, reflecting a worldwide trend. We document the detection of ST1, ST8, ST15 and ST30 in the 1980 isolates; we hypothesize their possible latency and their appearance as the current CA-MRSA clones.

/pdf/hospital-associated-methicillin-resistant-staphylococcus-iuqtz5co56.pdf

Hospital-associated methicillin-resistant Staphylococcus aureus (HA-MRSA) in Italy

Linezolid, the first oxazolidinone to be used clinically, is effective in the treatment of infections caused by various Gram-positive pathogens, including multidrug resistant enterococci and methicillin-resistant Staphylococus aureus. It has been used successfully for the treatment of patients with endocarditis and bacteraemia, osteomyelitis, joint infections and tuberculosis and it is often used for treatment of complicated infections when other therapies have failed. Linezolid resistance in Gram-positive cocci has been encountered clinically as well as in vitro, but it is still a rare phenomenon. The resistance to this antibiotic has been, until now, entirely associated with distinct nucleotide substitutions in domain V of the 23S rRNA genes. The number of mutated rRNA genes depends on the dose and duration of linezolid exposure and has been shown to influence the level of linezolid resistance. Mutations in associated ribosomal proteins also affect linezolid activity. A new phenicol and clindamycin resistance phenotype has recently been found to be caused by an RNA methyltransferase designated Cfr. This gene confers resistance to lincosamides, oxazolidinones, streptogramin A, phenicols and pleuromutilins, decrease the susceptibility of S. aureus to tylosin, to josamycin and spiramycin and thus differs from erm rRNA methylase genes. Research into new oxazolidinones with improved characteristics is ongoing. Data reported in patent applications demonstrated that some oxazolidinone derivatives, also with improved characteristics with respect to linezolid, are presently under study: at least three of them are in an advanced phase of development.

Linezolid Resistance in Staphylococci

Background: Despite 10 years of clinical use, linezolid resistance in Staphylococcus aureus and coagulase-negative staphylococci (CoNS) is still a rare phenomenon. This study reports the mechanisms of resistance and strain types seen in clusters of linezolid-resistant CoNS from two different hospitals in Italy during the period 2008-09. Methods: Genes associated with linezolid resistance were subjected to molecular analysis and isolates were characterized by PFGE macrorestriction analysis using SmaI. Results: Thirty-three linezolid-resistant isolates of methicillin-resistant CoNS comprising Staphylococcus epidermidis (24), Staphylococcus hominis (5) and Staphylococcus simulans (4) were studied. The isolates showed varying levels of linezolid resistance. Almost all isolates for which linezolid MICs were 64 mg/L possessed point mutations in domain V of 23S rRNA, while isolates for which the MICs were 256 mg/L expressed methylase activity at position A2503 mediated by the cfr gene. Overall, the isolates showed reduced susceptibility to vancomycin (MICs 1-2 mg/L) and 11 of the 33 isolates showed no susceptibility to teicoplanin. These strains were also resistant to chloramphenicol (28 of 33), lincomycin (24 of 33), erythromycin (17 of 33) and quinupristin/dalfopristin (13 of 33). S. epidermidis isolates, showing mutations or methylase modifcations, belonged to different PFGE profiles and to two different sequence types (ST2 and ST23), in which the cfr gene was carried on a plasmid of ∼50 kb. Conclusions: Clinical CoNS strains with resistance to linezolid and other second-line antibiotics, as well as reduced susceptibility to glycopeptides, have emerged in Italy.

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DNA methylase modifications and other linezolid resistance mutations in coagulase-negative staphylococci in Italy

Here we report for the first time, a detailed characterization of a variant of the SCCmec element, in a methicillin-resistant Staphylococcus intermedius human isolate. S. intermedius is a coagulase-positive zoonotic microrganism, recently classified as a separate species. In routine clinical laboratory practice, the coagulase production is used as criterion of pathogenicity related to S. aureus, but S. intermedius is frequently misidentified—being mistaken for S. aureus—and consequently its real incidence underestimated. S. intermedius have been found only occasionally in human beings, and methicillin-resistance is very rare for this organism. Even if the genetic element responsible for methicillin-resistance—the mecA gene carried by diverse staphylococcal chromosomal cassettes—has been described in various staphylococcal species, the current literature doesn't report any case of S. intermedius isolate carrying SCCmec-like elements. Our study could be useful to explain the mechanism and routes of transfer...

Dafne Bongiorno

Papers

Sensitivity assessment of droplet digital PCR for SARS-CoV-2 detection.

Hospital-associated methicillin-resistant Staphylococcus aureus (HA-MRSA) in Italy

Linezolid Resistance in Staphylococci

DNA methylase modifications and other linezolid resistance mutations in coagulase-negative staphylococci in Italy

Characterization of a variant of the SCCmec element in a bloodstream isolate of Staphylococcus intermedius.