Emergence of a mupirocin-resistant, methicillin-susceptible Staphylococcus aureus clone associated with skin and soft tissue infections in Greece

Background Staphylococcus aureus causes various infections, including skin and soft tissue infections (SSTIs). In this study, methicillin-susceptible S. aureus (MSSA) from SSTIs among patients in three tertiary-care hospitals in Greece were studied in terms of antimicrobial resistance, clonal distribution, toxin and adhesin genes carriage. Results During a five-year period (2014–2018), 6145 S. aureus were recovered from 13,244 patients with SSTIs and tested for antimicrobial susceptibility. MSSA were 4806 (78.21 %) including 1484 isolates with mupirocin minimum inhibitory concentration (MIC) > 64 mg/L (30.88 %). Two hundred and sixty representative mupirocin-resistant MSSA were analyzed for genes encoding Panton-Valentine leukocidin (PVL, lukS/lukF-PV), exfoliative toxins (eta, etb), adhesin FnbA (fnbA) and resistance genes mupA (high-level resistance to mupirocin), fusB (fusidic acid), aminoglycosides’ modifying enzymes, ermA, ermC and msrA (macrolides/lincosamides) by PCRs. Strains were classified into clones by PFGE and MLST. All mupirocin-resistant MSSA were penicillin-resistant; 92.7 % expressed resistance to fusidic acid and 88.9 % to tobramycin. All 260 molecularly analyzed isolates were mupA-positive; all fusidic acid-resistant (241/260) carried fusB whereas, the tobramycin-resistant ones (230), ant(4′)-Ia. The majority carried eta (93.85 %), etb (98.08 %) and fnbA (88.85 %). PFGE typing revealed a mostly unvarying population; 260 MSSA were grouped into three types. One major eta/etb-positive clone comprising of 258/260 strains (99.2 %), PFGE type 1, was classified as ST121, including nine strains co-carrying PVL. Another PVL-positive strain was identified as ST1, and one toxins-negative as ST21. Conclusions A mupirocin-resistant MSSA clone, ST121, carrying resistance, exfoliative toxins and adhesin genes, was spread and predominated in SSTIs from patients in Greece during the five-year studied period.

Antimicrobial resistance and genotyping of Staphylococcus aureus obtained from food animals in Sichuan Province, China

Background Staphylococcus aureus (S. aureus), especially methicillin-resistant Staphylococcus aureus (MRSA), is considered a common zoonotic pathogen, causing severe infections. The objective of this study was to investigate the antimicrobial susceptibility, resistance genes and molecular epidemiology among MRSA and methicillin-susceptible Staphylococcus aureus (MSSA) isolated from food animals in Sichuan Province, China. Methods This study was conducted on 236 S. aureus isolates. All isolates were subjected to antimicrobial susceptibility testing by using a standard microbroth dilution method. The Polymerase Chain Reaction (PCR) was performed to identify genes encoding the β-lactams resistance (blaZ, mecA), macrolides (ermA, ermB, ermC) and aminoglycosides (aacA-aphD). The molecular structures and genomic relatedness of MRSA isolates were determined by staphylococcal chromosome cassette mec (SCCmec) typing and pulsed-field gel electrophoresis (PFGE), respectively. Results Among 236 isolates, 24 (10.17 %) were recognized as MRSA. MRSA isolates showed different resistance rates to 11 antimicrobials ranging from 33.33 to 100 %, while for MSSA isolates the rates varied from 8.02 to 91.51 %. Multi-drug resistance phenotype was found in all MRSA isolates. The ermC gene encoding macrolides-lincosamides-streptogramin B was the most prevalent gene detected in 87.29 % of the S. aureus isolates, followed by ermB (83.05 %), blaZ (63.98 %), aacA-aphD (44.07 %), ermA (11.44 %) and mecA (11.02 %) genes. The prevalence of resistance genes in MRSA isolates was significantly higher than that of MSSA. Regarding the molecular morphology, SCCmec III (12/24, 50 %) was the most common SCCmec type. Furthermore, the PFGE typing showed that 24 MRSA were divided into 15 cluster groups (A to O), the major pulsotype J encompassed 25 % of MRSA isolates. Conclusions The S. aureus isolates from food animals in Sichuan province of China have severe antimicrobials resistance with various resistance genes, especially MRSA isolates. Additionally, the genetic pool of MRSA isolates is diverse and complex, and further investigation is necessary.

The Changing Landscape of Molecular Epidemiology of Staphylococcus Aureus Infections in Children.

Background: Staphylococcus aureus infections cause significant morbidity and mortality in children and adolescents. Aim of this study was to investigate the molecular epidemiology and antibiotic resistance of Staphylococcus aureus clinical isolates from children and adolescents.Methods: All S. aureus isolates recovered from patients aged < 18 years, admitted to a referral hospital, with culture-proven invasive or non-invasive, community-associated or community-onset healthcare-associated or hospital-associated infections during the 4-year period from January 2015 to December 2018 were analyzed for antimicrobial resistance, virulence genes, pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST).Results: Among 139 S. aureus clinical isolates, 16 (11.5%) were methicillin-resistant (MRSA) and 123 (88.5%) methicillin-susceptible (MSSA). MSSA infections increased significantly over time (2017-2018 vs. 2015-2016, 0R 3.32; 95%CI 1.18-8.96; p 0.03) along with increasing resistance to fusidic acid (OR 2.38; 95%CI 1.14-5.12; p 0.02) and in staphylococcal scalded skin syndrome prevalence (OR 3.24; 95% CI 1.10-8.36; p 0.03). A total of five sequence types (ST) were identified among 58 isolates that were analyzed by MLST. By PFGE typing, 22 pulsotypes were identified, whereas, PFGE type 1 classified as ST121 clone was the predominant (40/58,68.9%). MRSA were distributed into four pulsotypes and PFGE type C- ST80 was the most frequent. ST121 strains carried fnbA (40/40), eta/etb genes (29/40) and lukF/S-PVL genes in 3/40 cases. All ST121 exhibited high resistance percentage to fusidic acid and were increasingly resistant to mupirocin. Conclusion: In our population, a CA-MSSA clone emerged, resistant to fusidic acid and increasingly resistant to mupirocin which belonged to the PFGE type 1, ST121 clone, harbored exfoliative toxins genes and was associated with rising trends of SSSS.

Co-Occurrence of Listeria spp. and Spoilage Associated Microbiota During Meat Processing Due to Cross-Contamination Events

A large part of foodborne outbreaks related to Listeria monocytogenes are linked to meat and meat products. Especially, recontamination of meat products and deli-meat during slicing, packaging, and repackaging is in the focus of food authorities. In that regard, L. monocytogenes persistence in multi-species biofilms is one major issue, since they survive elaborate cleaning and disinfection measures. Here, we analyzed the microbial community structure throughout a meat processing facility using a combination of high-throughput full-length 16S ribosomal RNA (rRNA) gene sequencing and traditional microbiological methods. Samples were taken at different stages during meat cutting as well as from multiple sites throughout the facility environment to capture the product and the environmental associated microbiota co-occurring with Listeria spp. and L. monocytogenes. The listeria testing revealed a widely disseminated contamination (50%; 88 of 176 samples were positive for Listeria spp. and 13.6%; 24 of 176 samples were positive for L. monocytogenes). The pulsed-field gel electrophoresis (PFGE) typing evidenced 14 heterogeneous L. monocytogenes profiles with PCR-serogroup 1/2a, 3a as most dominant. PFGE type MA3-17 contributed to the resilient microbiota of the facility environment and was related to environmental persistence. The core in-house microbiota consisted mainly of the genera Acinetobacter, Pseudomonas, Psychrobacter (Proteobacteria), Anaerobacillus, Bacillus (Firmicutes), and Chryseobacterium (Bacteroidota). While the overall microbial community structure clearly differed between product and environmental samples, we were able to discern correlation patterns regarding the presence/absence of Listeria spp. in both sample groups. Specifically, our longitudinal analysis revealed association of Listeria spp. with known biofilm-producing Pseudomonas, Acinetobacter, and Janthinobacterium species on the meat samples. Similar patterns were also observed on the surface, indicating dispersal of microorganisms from this multispecies biofilm. Our data provided a better understanding of the built environment microbiome in the meat processing context and promoted more effective options for targeted disinfection in the analyzed facility.

Screening of Antimicrobial Resistance Genes and Epidemiological Features in Hospital and Community-Associated Carbapenem Resistant Pseudomonas aeruginosa Infections

Background: Researching carbapenem-resistant isolates and the use of antibiotics and following infection control policies enable the identification of carbapenemase-producing bacteria and prevent their spread.Methods: P. aeruginosa isolates were recovered from Medicine Faculty of Recep Tayyip Erdoğan University between April 2015 and October 2016 and identified by conventional methods and the automated Vitek 2 Compact (BioMerieux, France) system. Antimicrobial susceptibility experiments were performed in accordance with CLSI criteria and the automated Vitek 2 Compact system. The PCR method was investigated for the presence of β-lactamase resistance genes. PFGE typing was performed to show clonal relation among samples.Results: Seventy P. aeruginosa strains were isolated from seventy patients. The median age of 70 cases was found 66 with minimum 17 and maximum 92 years old. 67.1% of the patients had contact with the health service in the last 90 days and 75.7% of the patients had received antimicrobial therapy in the previous 90 days. The most common comorbidity was cardiovascular diseases. Twenty-four (34.3%) strains were carbapenem resistant, 2 strains were multidrug-resistant except colistin, and none of the samples had colistin resistance. The gene encoding β-lactamase or metallo-β-lactamase was found in a total of 36 strains. The bla VEB gene was identified in only 1 strain alone, but in combination with other resistance genes in a total of 17 strains. While the bla PER gene was detected in 5 samples alone, it was found in 13 samples in combination with other genes. Among the genes encoding metallo-β-lactamase, the most bla NDM positive was detected (n=22), followed by 14 positive samples of bla KPC . bla IMP and bla VIM were detected in 5 and 1 samples, respectively. Also, the association of bla VEB - bla PER and bla VEB - bla KPC - bla NDM was found to be very high. Much more resistance genes and associations were detected in hospital-acquired samples than community-acquired samples, both proportionally and in terms of co-occurrence. Most of the community-associated strains were collected in the F2 clade, while most of the hospital-associated strains were collected in the G1 clade. However, no difference was found between the community and hospital-associated strains according to PFGE results. Simultaneously, other microorganisms were also isolated from patients from which these 6 P. aeruginosa strains were isolated. Of these patients, 5 patients died, except the number 70.Conclusions: The median length of stay (days) was found to be significantly higher in the group with HAI than in the group with CAI. Compared to sample 28 and 37, which carried 5 β-lactamase coding genes, the death of these 5 patients with fewer or no resistance genes showed that the coexistence of other factors - especially other microorganisms in addition to resistance genes, was important.

Genetic relationship of Salmonella isolates found in subcutaneous abscesses in leopard geckos (Eublepharis macularius)

IntroductionThe article describes the occurrence and phylogenetic relationship of Salmonella isolates found in subcutaneous abscesses of leopard geckos. The aim of the study was to determine the cause of the abscesses and to characterise isolated Salmonella strains.Material and MethodsSamples of abscesses from five animals and internal organs (lungs, liver, and gut) of three of them were tested for Salmonella according to the PN-EN ISO 6579:2002/A1:2007 standard. The antimicrobial resistance was evaluated by minimal inhibitory concentrations and the genetic similarity of the isolates was assessed with pulsed field gel electrophoresis (PFGE).ResultsIn total, seventeen Salmonella isolates belonging to five different serovars were found to be susceptible to all tested antimicrobials except streptomycin. The serovars were S. Hadar, S. Fluntern, S. Tennessee, S. enterica subsp. salamae 55:k:z39, and S. Kentucky. Up to three serovars from different organs were isolated from the same individual. In two geckos, Salmonella were detected in the lungs. In three serovars, XbaI-PFGE typing revealed indistinguishable isolates from organs and abscesses.ConclusionMultiple Salmonella serovars might be involved in abscess formation and infections. The occurrence of the same PFGE profiles of the isolates may testify to the role of opportunistic organisms in causing infection.

Predominance of Distinct Listeria Innocua and Listeria Monocytogenes in Recurrent Contamination Events at Dairy Processing Facilities

The genus Listeria now comprises up to now 21 recognized species and six subspecies, with L. monocytogenes and L. innocua as the most prevalent sensu stricto associated species. Reports focusing on the challenges in Listeria detection and confirmation are available, especially from food-associated environmental samples. L. innocua is more prevalent in the food processing environment (FPE) than L. monocytogenes and has been shown to have a growth advantage in selective enrichment and agar media. Until now, the adaptive nature of L. innocua in FPEs has not been fully elucidated and potential persistence in the FPE has not been observed. Therefore, the aim of this study is to characterize L. innocua (n = 139) and L. monocytogenes (n = 81) isolated from FPEs and cheese products collected at five dairy processing facilities (A–E) at geno- and phenotypic levels. Biochemical profiling was conducted for all L. monocytogenes and the majority of L. innocua (n = 124) isolates and included a rhamnose positive reaction. L. monocytogenes isolates were most frequently confirmed as PCR-serogroups 1/2a, 3a (95%). Pulsed-field gel electrophoresis (PFGE)-typing, applying the restriction enzymes AscI, revealed 33 distinct Listeria PFGE profiles with a Simpson’s Index of Diversity of 0.75. Multi-locus sequence typing (MLST) resulted in 27 STs with seven new L. innocua local STs (ST1595 to ST1601). L. innocua ST1597 and ST603 and L. monocytogenes ST121 and ST14 were the most abundant genotypes in dairy processing facilities A–E over time. Either SSI-1 (ST14) or SSI-2 (ST121, all L. innocua) were present in successfully FPE-adapted strains. We identified housekeeping genes common in Listeria isolates and L. monocytogenes genetic lineage III. Wherever there are long-term contamination events of L. monocytogenes and other Listeria species, subtyping methods are helpful tools to identify niches of high risk.

Streptococcus suis in Brazil: Genotypic, Virulence, and Resistance Profiling of Strains Isolated from Pigs between 2001 and 2016

Streptococcus suis remains an important challenge for the worldwide swine industry. Considering that Brazil is a major pork producer and exporter, proper monitoring of the pathogen and resistance rates are required. We present here the characterization of Brazilian S. suis strains isolated over a 15 year period by pulsed-field gel electrophoresis (PFGE) typing, capsular, virulence, and antimicrobial resistance profiling. Serotype prevalence revealed a predominance of serotype 2/½ followed by 3, 7, 1/14, 6, 8, 18, 28, and 27; the latter had not yet been reported in Brazil. Resistance profiling enabled the differentiation of nine profiles presenting resistance to three and up to eight antimicrobial classes. Even though an association between the most resistant strains and isolation year starting from 2009 was observed, a high frequency of multidrug-resistant strains isolated from 2001 to 2003 was also detected. This suggests that despite the isolation period, S. suis strains already presented high resistance selection pressure. A slight association of serotype 2/½ with some virulence profiles and PFGE pulsotypes was also identified. Nevertheless, no clonal dispersion or persistency of clones over the analyzed years and herds was detected.

High Prevalence of 16S rRNA methylase genes High Prevalence of 16S rRNA methylase genes Among Carbapenem-resistant Hypervirulent Klebsiella pneumoniae Isolates in China

Background : the existence of 16S rRNA methylase genes would increase treatment difficulty of patients infected with CR-hvKP strains, this study was aimed to testify the prevalence of the 16S rRNA methylase genes genes in the CR-hvKP strains in China.Methods : Thirty-nine carbapenem-resistant hypervirulent Klebsiella pneumoniae (CR-hvKP) isolates collected from a Chinese hospital during the whole year of 2018 were evaluated to characterize the prevalence of 16S rRNA methylase genes. Results : In tatal 66.7% (26/39) of the CR-hvKP isolates were found to carry 16S rRNA methylase genes, and the most frequently detected gene was armA (11,42.3%), followed by rmtB (8,30.8%),and 7 CR-hvKP strains were found to carry both armA and rmtB (26.9%). All the clinical isolates were found to carry at least one carbapenemase gene,with KPC-2 (79.5%,31/39), NDM-1 (10.3%,4/39), and cocarrying KPC-2 and NDM-1 (10.3%,4/39). A total of 89.7% (35/39) isolates carried ESBL genes, including 61.5% (24/39) blaSHV-1 ,71.8% (28/39) blaTEM-1 and 89.7% (35/39) blaCTX-M-1 4. All except four isolates (89.7%,35/39) harbored PMQR genes,with qnrS (82.1%,32/39), aac(6’)-Ib-cr (79.5%,31/39), qnrB (2.6%,1/39).All the 16S rRNA methylase genes-positive CR-hvKP strains were firstly found to cocarry carbapenemase genes, ESBL genes and PMQR genes simultaneously. The most prevalent virulence genes were rmpA2 and entB (100%, 39/39),followed by silS (97.4%, 38/39), ybtS (94.9%, 37/39), iutA (92.3%, 36/39), kpn (92.3%, 36/39), rmpA (87.2%, 34/39), terW (84.6%, 33/39), aerobactin (23.1%, 9/39), repA (17.9%, 7/39), magA (10.3%, 4/39), kfuB C (10.3%, 4/39), w ca G (10.3%, 4/39), allS (10.3%, 4/39). Multilocus sequence typing (MLST) analysis assigned the 39 CR-hvKP isolates into 4 sequence types (STs), with ST11 encompassing 79.5% of the strains. Pulsed field gel electrophoresis (PFGE) typing showed that strains closely related by MLST clustered in major PFGE clusters, of which cluster A accounts for 31 ST11 isolates.The analysis of the transconjugants showed a high-level aminoglycoside resistance and a popular cotransfer of bla KPC-2 with the 16S rRNA methylase genes.Conclusions : 16S rRNA methylase genes are highly prevalent in CR-hvKP clinical isolates especially for ST11, it is therefore critical to continuously monitor the 16S rRNA methylase-producing CR-hvKP epidemiology and minimize potential risks from aminoglycoside -resistant CR-hvKP.