Genetic background of aminoglycoside-modifying enzymes in various genetic lineages of clinical aminoglycosides-resistant E. coli and K. pneumoniae isolates in Tunisia.

AIMS: This study was conducted to evaluate the in vitro activity of clinically relevant aminoglycosides and to determine the prevalence of genes encoding aminoglycoside modifying enzymes (AMEs) and 16S ribosomal RNA (rRNA) methyltransferases among aminoglycoside-resistant E. coli (n = 61) and K. pneumoniae (n = 44) clinical isolates. Associated resistances to beta-lactams and their bla genes as well as the genetic relatedness of isolates were also investigated. MATERIALS AND METHODS: A total of 105 aminoglycoside-resistant E. coli (n = 61) and K. pneumoniae (n = 44) isolates recovered between March and May 2017 from 100 patients hospitalized in different wards of Charles Nicolle Hospital of Tunis, Tunisia, were studied. Minimal inhibitory concentrations of aminoglycoside compounds were determined by broth microdilution method. Aminoglycosides resistance encoding genes [aph(3´)-Ia, aph(3') IIa, aph(3´)-VIa, ant(2″)-Ia, aac(3)-IIa, aac(3)-IVa, aac(6')-Ib, rmtA, rmtB, rmtC, armA, and npmA] and bla genes were investigated by PCR and sequencing. Genetic relatedness was examined by multilocus sequence typing (MLST) for representative isolates. RESULTS: High rates of aminoglycoside resistance were found: gentamicin (85.7%), tobramycin (87.6%), kanamycin (78.0%), netilmincin (74.3%), and amikcin (18.0%). Most common AME gene was aac(3)-IIa (42%), followed by aac(6')-Ib (36.2%) and aph(3')-VIa (32.4%). The majority of isolates were resistant to beta-lactams and blaCTX-M-15 was the most common ESBL. The blaNDM-1 and blaOXA-48 were also produced by 1 and 23 isolates, respectively. Novel sequence types have been reported among our isolates and high-risk clonal lineages have been detected, such as E. coli ST43 (ST131 in Achtman MLST scheme) and K. pneumoniae (ST11/ST13). CONCLUSIONS: The high prevalence of aminoglycoside resistance rates and the diversity of corresponding genes, with diverse ß-lactamase enzymes among genetically heterogeneous clinical isolates present a matter of concern.

Gene expression response under thermal stress in two Hawaiian corals is dominated by ploidy and genotype.

Transcriptome data are frequently used to investigate coral bleaching; however, the factors controlling gene expression in natural populations of these species are poorly understood. We studied two corals, Montipora capitata and Pocillopora acuta, that inhabit the sheltered Kane'ohe Bay, Hawai'i. M. capitata colonies in the bay are outbreeding diploids, whereas P. acuta is a mixture of clonal diploids and triploids. Populations were sampled from six reefs and subjected to either control (no stress), thermal stress, pH stress, or combined pH and thermal stress treatments. RNA-seq data were generated to test two competing hypotheses: (1) gene expression is largely independent of genotype, reflecting a shared treatment-driven response (TDE) or, (2) genotype dominates gene expression, regardless of treatment (GDE). Our results strongly support the GDE model, even under severe stress. We suggest that post-transcriptional processes (e.g., control of translation, protein turnover) modify the signal from the transcriptome, and may underlie the observed differences in coral bleaching sensitivity via the downstream proteome and metabolome.

Building the brain mosaic: an expanded view.

The complexity of the brain is closely tied to its nature as a genetic mosaic, wherein each cell is distinguished by a unique constellation of somatic variants that contribute to functional and phenotypic diversity. Postzygotic variation arising during neurogenesis is recognized as a key contributor to brain mosaicism; however, recent advances have broadened our understanding to include sources of neural genomic diversity that develop throughout the entire lifespan, from embryogenesis through aging. Moving beyond the traditional confines of neurodevelopment, in this review, we delve into the complex mechanisms that enable various origins of brain mosaicism.

Genetic basis of antimicrobial resistance, virulence features and phylogenomics of carbapenem-resistant Acinetobacter baumannii clinical isolates.

PURPOSE: The worldwide emergence and clonal spread of carbapenem-resistant Acinetobacter baumannii (CRAB) is of great concern. In the present study, we determined the mechanisms of antimicrobial resistance, virulence gene repertoire and genomic relatedness of CRAB isolates circulating in Serbian hospitals. METHODS: CRAB isolates were analyzed using whole-genome sequencing (WGS) for the presence of antimicrobial resistance-encoding genes, virulence factors-encoding genes, mobile genetic elements and genomic relatedness. Antimicrobial susceptibility testing was done by disk diffusion and broth microdilution methods. RESULTS: Eleven isolates exhibited an MDR resistance phenotype, while four of them were XDR. MIC90 for meropenem and imipenem were > 64 µg/mL and 32 µg/mL, respectively. While all CRABs harbored blaOXA-66 variant of blaOXA-51 gene, those assigned to STPas2, STPas636 and STPas492 had blaADC-73,blaADC-74 and blaADC-30 variants, respectively. The following acquired carbapenemases-encoding genes were found: blaOXA-72 (n = 12), blaOXA-23 (n = 3), and blaNDM-1(n = 5), and were mapped to defined mobile genetic elements. MLST analysis assigned the analyzed CRAB isolates to three Pasteur sequence types (STs): STPas2, STPas492, and STPas636. The Majority of strains belonged to International Clone II (ICII) and carried tested virulence-related genes liable for adherence, biofilm formation, iron uptake, heme biosynthesis, zinc utilization, serum resistance, stress adaptation, intracellular survival and toxin activity. CONCLUSION: WGS elucidated the resistance and virulence profiles of CRABs isolated from clinical samples in Serbian hospitals and genomic relatedness of CRAB isolates from Serbia and globally distributed CRABs.

Resistome, Virulome, and Clonal Variation in Methicillin-Resistant Staphylococcus aureus (MRSA) in Healthy Swine Populations: A Cross-Sectional Study.

This cross-sectional study investigates the methicillin-resistant Staphylococcus aureus (MRSA): its prevalence, antimicrobial resistance, and molecular characteristics in healthy swine populations in central Portugal. A total of 213 samples were collected from pigs on twelve farms, and MRSA prevalence was assessed using selective agar plates and confirmed via molecular methods. Antimicrobial susceptibility testing and whole genome sequencing (WGS) were performed to characterize resistance profiles and genetic determinants. Among the 107 MRSA-positive samples (83.1% prevalence), fattening pigs and breeding sows exhibited notably high carriage rates. The genome of 20 isolates revealed the predominance of the ST398 clonal complex, with diverse spa types identified. Antimicrobial susceptibility testing demonstrated resistance to multiple antimicrobial agents, including penicillin, cefoxitin, and tetracycline. WGS analysis identified a diverse array of resistance genes, highlighting the genetic basis of antimicrobial resistance. Moreover, virulence gene profiling revealed the presence of genes associated with pathogenicity. These findings underscore the significant prevalence of MRSA in swine populations and emphasize the need for enhanced surveillance and control measures to mitigate zoonotic transmission risks. Implementation of prudent antimicrobial use practices and targeted intervention strategies is essential to reducing MRSA prevalence and safeguarding public health. Continued research efforts are warranted to elucidate transmission dynamics and virulence potential, ultimately ensuring food safety and public health protection.

Clonal Flux and Spread of Staphylococcus aureus Isolated from Meat and Its Genetic Relatedness to Staphylococcus aureus Isolated from Patients in Saudi Arabia.

In this study, we investigated both meat-derived and methicillin-resistant Staphylococcus aureus (MRSA), exploring their genetic relatedness to patient-derived MRSA isolates in Saudi Arabia. We collected 250 meat samples and identified 53 S. aureus isolates, with 79% being methicillin-sensitive Staphylococcus aureus (MSSA) and 21% being MRSA. Moreover, we included 80 clinically confirmed patient-derived MRSA isolates. We identified the most common S. aureus clone in both patients and retail meat. In meat, ST6 and ST97 were the most common clones in 55% of the MRSA isolates, and ST1153 and ST672 were the most common in 21% and 17% of the MSSA isolates. In patients, ST5 and ST6 were the predominant clones in 46% of the S. aureus isolates. CC5/ST5-SCCmecVc-t311 and CC361/ST672-SCCmecV-t3841 were common MRSA clones in both meat and patients. CC97 and CC361 clones were the second most prevalent S. aureus clones in meat and were relatively common in patients. Furthermore, we sequenced and characterized novel S. aureus strains ST8109, ST8110, and ST8111. The genomic similarities between meat- and patient-derived S. aureus isolates suggest that retail meat might be a reservoir for S.aureus and MRSA transmission. Therefore, a structured One Health approach is recommended for S. aureus dissemination, genetic characterization, antibiotic resistance, and impact on human health.

Antimicrobial Resistance and Clonal Lineages of Escherichia coli from Food-Producing Animals.

Escherichia coli are one of the most important pathogenic bacteria readily found in the livestock and widely studied as an indicator that carries drug-resistant genes between humans, animals, and the environment. The use of antimicrobials in the food chain, particularly in food-producing animals, is recognized as a significant contributor to the development and spread of antimicrobial resistance (AMR) and resistance genes can be transferred from the farm through the food-chain. The objective of this review is to highlight the background of the antimicrobials use in food-producing animals, more specifically, to study clonal lineages and the resistance profiles observed in E. coli, as well as in extended spectrum beta-lactamases (ESBL) producing E. coli, in a set of food-production animals with greater relevance in food consumption, such as pigs, poultry, cattle, fish farming and rabbits. Regarding the prevalence of ESBL-producing E. coli among farm animals, high-to-moderate prevalence was observed, and the highest resistance rates to tetracycline and ampicillin was detected in different farms in all geographic regions. Worldwide pandemic clones and high-risk zoonotic E. coli clones have been identified in most food-producing animals, and some of these clones are already disseminated in different niches, such as the environment and humans. A better understanding of the epidemiology of E. coli and ESBL-producing E. coli in livestock is urgently needed. Animal production is one of the major causes of the antibiotic resistance problem worldwide and a One Health approach is needed.

Lineages, Virulence Gene Associated and Integrons among Extended Spectrum ß-Lactamase (ESBL) and CMY-2 Producing Enterobacteriaceae from Bovine Mastitis, in Tunisia.

Extended Spectrum Beta-Lactamase (ESBL) Enterobacteriaceae are becoming widespread enzymes in food-producing animals worldwide. Escherichia coli and Klebseilla pneumoniae are two of the most significant pathogens causing mastitis. Our study focused on the characterization of the genetic support of ESBL/pAmpC and antibiotic resistance mechanisms in cefotaxime-resistant (CTXR) and susceptible (CTXS) Enterobacteriaceae isolates, recovered from bovine mastitis in Tunisia, as well as the analyses of their clonal lineage and virulence-associated genes. The study was carried out on 17 ESBL/pAmpC E. coli and K. pneumoniae and 50 CTXS E. coli. Detection of resistance genes and clonal diversity was performed by PCR amplification and sequencing. The following ß-lactamase genes were detected: blaCTX-M-15 (n = 6), blaCTX-M-15 + blaOXA-1 (2), bla CTX-M-15 + blaOXA-1 + blaTEM-1b (2), blaCTX-M-15 + blaTEM-1b (4), blaCMY-2 (3). The MLST showed the following STs: ST405 (n = 4 strains); ST58 (n = 3); ST155 (n = 3); ST471 (n = 2); and ST101 (n = 2). ST399 (n = 1) and ST617 (n = 1) were identified in p(AmpC) E. coli producer strains. The phylogroups A and B1 were the most detected ones, followed by the pathogenic phylogroup B2 that harbored the shigatoxin genes stx1/stx2, associated with the cnf, fimA, and aer virulence factors. The qnrA/qnrB, aac(6')-Ib-cr genes and integrons class 1 with different gene cassettes were detected amongst these CTXR/S isolated strains. The presence of different genetic lineages, associated with resistance and virulence genes in pathogenic bacteria in dairy farms, may complicate antibiotic therapies and pose a potential risk to public health.

The Use of Galleria mellonella Larvae to Study the Pathogenicity and Clonal Lineage-Specific Behaviors of the Emerging Fungal Pathogen Candida auris.

Candida species are the most common fungal causes of disseminated infections in humans. Although such infections are associated with high morbidity and mortality, it is widely accepted that virulence, antifungal susceptibility, and disease outcome vary according to individual Candida species. In this respect, the emerging pathogen Candida auris has received much attention due to its propensity to cause widespread nosocomial outbreaks, to exhibit high virulence in several infection models, and to develop resistance to multiple classes of antifungal drugs. Although mammalian models of infection have long been viewed as the gold standard for studies on fungal virulence, comparative pathogenicity, and evaluation of antifungal drug efficacy, the larvae of the greater wax moth Galleria mellonella have shown considerable promise as an alternative invertebrate model of infection. Galleria larvae are inexpensive, are easily maintained in the laboratory, tolerate incubation at human physiological temperatures, possess cellular and humoral immune systems that share many features with mammals, and allow investigation of pathogenicity/virulence using multiple different reading endpoints. Here, I describe in detail the methods that can be used to study the virulence/pathogenicity of Candida auris in G. mellonella.

Livestock-associated methicillin-resistant Staphylococcus aureus in Czech retailed ready-to-eat meat products.

This study was aimed on the detection of methicillin resistant Staphylococcus aureus (MRSA) in different categories of retailed ready-to-eat (RTE) meat products from the Czech producers and determination of their genetic properties, antimicrobial resistance and virulence. In RTE meat products, 2% (4/181) of examined samples were MRSA positive. MRSA strains were detected only in durable fermented meat products made exclusively from pork meat. Detection of livestock-associated MRSA (LA-MRSA) clonal lineages (ST398 and ST4999), SCCmec cassette type V and tetracycline resistance indicate a source of contamination from raw pork. The study confirms the ability of these strains to survive the technological process rather than contamination of meat products from the food processing environment. MRSA strains did not carry any of the tested genes encoding staphylococcal enterotoxins or virulence genes (for Panton-Valentine leukocidin, exfoliative toxins A, B and toxic shock syndrome). Our results point out the spread of LA-MRSA through the meat processing chain.

Staphylococcus aureus Causing Skin and Soft Tissue Infections in Companion Animals: Antimicrobial Resistance Profiles and Clonal Lineages.

Staphylococcus aureus is a relevant agent of skin and soft tissue infections (SSTIs) in animals. Fifty-five S. aureus comprising all SSTI-related isolates in companion animals, collected between 1999 and 2018 (Lab 1) or 2017 and 2018 (Lab 2), were characterized regarding susceptibility to antibiotics and heavy metals and carriage of antimicrobial resistance determinants. Clonal lineages were established by PFGE, MLST and agr typing. Over half of the isolates (56.4%, 31/55) were methicillin-resistant S. aureus (MRSA), and 14.5% showed a multidrug resistance (MDR) phenotype. Resistance was most frequently observed for beta-lactams (81.8%, related to blaZ and/or mecA), fluoroquinolones (56.4%) and macrolides/lincosamides (14.5%, related to erm(A) or erm(C)). The distributions of heavy-metal MICs allowed the detection of non-wild-type populations associated with several resistance genes. The collection showed genetic diversity, with prevalence of clonal lineage ST22-agrI (45.5%, 25/55), comprising only MRSA isolates, and several less frequently detected clones, including ST5-agrII (14.6%, 8/55), ST398-agrI (9.1%, 5/55) and ST72-agrI (7.3%, 4/55). This work highlights the high frequency of SSTI-related MRSA strains that reflect the clonal lineages circulating both in companion animals and humans in Portugal, reinforcing the need for a One Health approach when studying staphylococci causing infections in companion animals.

Epidemiological Studies of Pan-Azole Resistant Aspergillus fumigatus Populations Sampled during Tulip Cultivation Show Clonal Expansion with Acquisition of Multi-Fungicide Resistance as Potential Driver.

Pan-azole resistant isolates are found in clinical and environmental Aspergillus fumigatus (Af) populations. Azole resistance can evolve in both settings, with Af directly targeted by antifungals in patients and, in the environment, Af unintendedly exposed to fungicides used for material preservation and plant disease control. Resistance to non-azole fungicides, including methyl benzimidazole carbamates (MBCs), quinone outside inhibitors (QoIs) and succinate dehydrogenase inhibitors (SDHIs), has recently been reported. These fungicide groups are not used in medicine but can play an important role in the further spread of pan-azole resistant genotypes. We investigated the multi-fungicide resistance status and the genetic diversity of Af populations sampled from tulip field soils, tulip peel waste and flower compost heaps using fungicide sensitivity testing and a range of genotyping tools, including STRAf typing and sequencing of fungicide resistant alleles. Two major clones were present in the tulip bulb population. Comparisons with clinical isolates and literature data revealed that several common clonal lineages of TR34/L98H and TR46/Y121F/T289A strains that have expanded successfully in the environment have also acquired resistance to MBC, QoI and/or SDHI fungicides. Strains carrying multiple fungicide resistant alleles have a competitive advantage in environments where residues of multiple fungicides belonging to different modes of action are present.

First Detection of Human ST131-CTX-M-15-O25-B2 Clone and High-Risk Clonal Lineages of ESBL/pAmpC-Producing E. coli Isolates from Diarrheic Poultry in Tunisia.

Circulation of a multi-resistance clone of bacteria associated with genetic elements in diseased animals constitutes a global public health problem. Our study focused on the characterization of the support of ESBL in cefotaxime resistant E. coli (CTXR) isolates recovered from poultry with diarrhea, analysis of their clonal lineage, and virulence-associated genes. The study was carried out on 130 samples of chickens with diarrhea, collected in 2015 from poultry farms in Tunisia. Isolates of 20 CTXR E. coli strains were identified as ESBL and AmpC ß- lactamase producers. The following ß-lactamase genes (number of isolates) were detected: blaCTX-M-15+ blaOXA1 (4), blaCTX-M-15 + blaOXA1 + blaTEM-1b (2), blaCTX-M-1 + blaTEM-1b (9), blaCTX-M-1 (2), blaCMY2 + blaTEM-1b (3). Six E. coli harboring blaCTXM-15 were allocated to ST131-B2-O25b-; six and three blaCTX-M-1 were grouped in ST155, ST10, and ST58, respectively, related to the phylogroup D and A. The qnrB gene, the variant aac(6')-Ib-cr, and the class 1 integrons with different gene cassettes, were detected amongst our 20 isolated strains, which were classified as ExPEC and aEPEC. Our findings highlighted the emergence of the human pandemic ST131-CTX-M-15-O25-B2 clone and the high risk of such clonal lineage strains in diarrheic poultry, in Tunisia, which could constitute a risk of their transfer to healthy animals and humans.

Water distribution systems in Sardinian hospitals host invasive clonal lineages of the Fusarium oxysporum and Fusarium solani species complexes.

Several Fusarium species cause disease on human hosts, including commonly fatal infections in immunocompromised individuals. Recently, cases of hospitalized patients affected by fusaria were reported in the Tyrrhenian Island of Sardinia, Italy. To precisely characterize the Fusarium species and haplotypes present in hospitals of the region, a multilocus DNA sequence typing (MLST) approach was applied. Water distribution systems in four departments belonging to four Sardinian hospitals were sampled. Fusarium species and sequence types (STs) were identified using MLST based on sequences of the elongation factor 1-alpha (EF-1α) gene, the nuclear ribosomal DNA intergenic spacer region (IGS rDNA), and/or a portion of the second-largest subunit of RNA polymerase (RPB2) gene. The majority of isolates obtained from Sardinian hospitals (90.7%) were identified as representatives of the Fusarium oxysporum species complex (FOSC), followed by those of the F. solani species complex (FSSC) (8.2%), and F. dimerum (1.1% of all isolates). Ten STs were found among the FOSC and FSSC, with more than 60% of the isolates identified as either FOSC ST 33 or FSSC 1 (F. petroliphilum). More than half of the FOSC isolates obtained from the water systems in all four hospitals belonged to the worldwide distributed clonal lineage ST 33. This haplotype is the most prevalent among the FOSC in different countries, being responsible for the vast majority of cases of human fusariosis.

Clonal Lineages, Antimicrobial Resistance, and PVL Carriage of Staphylococcus aureus Associated to Skin and Soft-Tissue Infections from Ambulatory Patients in Portugal.

Staphylococcus aureus (S. aureus) is a leading cause of skin and soft-tissue infections (SSTIs) in the community. In this study, we characterized a collection of 34 S. aureus from SSTIs in ambulatory patients in Portugal and analyzed the presence of Panton-Valentine leucocidin (PVL)-encoding genes and antibiotic-resistance profile, which was correlated with genetic determinants, plasmid carriage, and clonal lineage. Nearly half of the isolates (15, 44.1%) were methicillin-resistant Staphylococcus aureus (MRSA) and/or multidrug resistant (MDR). We also detected resistance to penicillin (33/34, 97.1%), fluoroquinolones (17/34, 50.0%), macrolides and lincosamides (15/34, 44.1%), aminoglycosides (6/34, 17.6%), and fusidic acid (2/34, 5.9%), associated with several combinations of resistance determinants (blaZ, erm(A), erm(C), msr(A), mph(C), aacA-aphD, aadD, aph(3')-IIIa, fusC), or mutations in target genes (fusA, grlA/gyrA). The collection presented a high genetic diversity (Simpson's index of 0.92) with prevalence of clonal lineages CC5, CC22, and CC8, which included the MRSA and also most MDR isolates (CC5 and CC22). PVL-encoding genes were found in seven isolates (20.6%), three methicillin-susceptible Staphylococcus aureus (MSSA) (ST152-agrI and ST30-agrIII), and four MRSA (ST8-agrI). Plasmid profiling revealed seventeen distinct plasmid profiles. This work highlights the high frequency of antimicrobial resistance and PVL carriage in SSTIs-related S. aureus outside of the hospital environment.

Clonal Clusters, Molecular Resistance Mechanisms and Virulence Factors of Gram-Negative Bacteria Isolated from Chronic Wounds in Ghana.

Wound infections are common medical problems in sub-Saharan Africa but data on the molecular epidemiology are rare. Within this study we assessed the clonal lineages, resistance genes and virulence factors of Gram-negative bacteria isolated from Ghanaian patients with chronic wounds. From a previous study, 49 Pseudomonas aeruginosa, 21 Klebsiellapneumoniae complex members and 12 Escherichia coli were subjected to whole genome sequencing. Sequence analysis indicated high clonal diversity with only nine P. aeruginosa clusters comprising two strains each and one E. coli cluster comprising three strains with high phylogenetic relationship suggesting nosocomial transmission. Acquired beta-lactamase genes were observed in some isolates next to a broad spectrum of additional genetic resistance determinants. Phenotypical expression of extended-spectrum beta-lactamase activity in the Enterobacterales was associated with blaCTX-M-15 genes, which are frequent in Ghana. Frequently recorded virulence genes comprised genes related to invasion and iron-uptake in E. coli, genes related to adherence, iron-uptake, secretion systems and antiphagocytosis in P. aeruginosa and genes related to adherence, biofilm formation, immune evasion, iron-uptake and secretion systems in K. pneumonia complex. In summary, the study provides a piece in the puzzle of the molecular epidemiology of Gram-negative bacteria in chronic wounds in rural Ghana.

The Epidome - a species-specific approach to assess the population structure and heterogeneity of Staphylococcus epidermidis colonization and infection.

BACKGROUND: Although generally known as a human commensal, Staphylococcus epidermidis is also an opportunistic pathogen that can cause nosocomial infections related to foreign body materials and immunocompromized patients. Infections are often caused by multidrug-resistant (MDR) lineages that are difficult and costly to treat, and can have a major adverse impact on patients' quality of life. Heterogeneity is a common phenomenon in both carriage and infection, but present methodology for detection of this is laborious or expensive. In this study, we present a culture-independent method, labelled Epidome, based on an amplicon sequencing-approach to deliver information beyond species level on primary samples and to elucidate clonality, population structure and temporal stability or niche selection of S. epidermidis communities. RESULTS: Based on an assessment of > 800 genes from the S. epidermidis core genome, we identified genes with variable regions, which in combination facilitated the differentiation of phylogenetic clusters observed in silico, and allowed classification down to lineage level. A duplex PCR, combined with an amplicon sequencing protocol, and a downstream analysis pipeline were designed to provide subspecies information from primary samples. Additionally, a probe-based qPCR was designed to provide valuable absolute abundance quantification of S. epidermidis. The approach was validated on isolates representing skin commensals and on genomic mock communities with a sensitivity of < 10 copies/µL. The method was furthermore applied to a sample set of primary skin and nasal samples, revealing a high degree of heterogeneity in the S. epidermidis populations. Additionally, the qPCR showed a high degree of variation in absolute abundance of S. epidermidis. CONCLUSIONS: The Epidome method is designed for use on primary samples to obtain important information on S. epidermidis abundance and diversity beyond species-level to answer questions regarding the emergence and dissemination of nosocomial lineages, investigating clonality of S. epidermidis communities, population dynamics, and niche selection. Our targeted-sequencing method allows rapid differentiation and identification of clinically important nosocomial lineages in low-biomass samples such as skin samples.

Antibiotic Treatment and Age Are Associated With Staphylococcus aureus Carriage Profiles During Persistence in the Airways of Cystic Fibrosis Patients.

BACKGROUND: Staphylococcus aureus is one of the most isolated pathogens from the airways of cystic fibrosis (CF) patients. There is a lack of information about the clonal nature of S. aureus cultured from CF patients and their impact on disease. We hypothesized that patients would differ in their clinical status depending on S. aureus clonal carriage profiles during persistence. METHODS: During a 21-months prospective observational multicenter study (Junge et al., 2016), 3893 S. aureus isolates (nose, oropharynx, and sputa) were cultured from 183 CF patients (16 German centers, 1 Austrian center) and subjected to spa-sequence typing to assess clonality. Data were associated to lung function, age, gender, and antibiotic treatment by multivariate regression analysis. RESULTS: Two hundred and sixty-five different spa-types were determined with eight prevalent spa-types (isolated from more than 10 patients): t084, t091, t008, t015, t002 t012, t364, and t056. We observed different carriage profiles of spa-types during the study period: patients being positive with a prevalent spa-type, only one, a dominant or related spa-type/s. Patients with more antibiotic cycles were more likely to be positive for only one spa-type (p = 0.005), while older patients were more likely to have related (p = 0.006), or dominant spa-types (p = 0.026). Two percent of isolates were identified as methicillin-resistant S. aureus (MRSA) and evidence of transmission of clones within centers was low. CONCLUSION: There was a significant association of antibiotic therapy and age on S. aureus carriage profiles in CF patients indicating that antibiotic therapy prevents acquisition of new clones, while during aging of patients with persisting S. aureus, dominant clones were selected and mutations in the spa-repeat region accumulated.

Extensively drug-resistant Acinetobacter baumannii isolated from intensive care units in northern Italy: a genomic approach to characterize new sequence types.

Aim: This study aims to characterize clinical strains of Acinetobacter baumannii with an extensively drug-resistant phenotype. Methods: VITEK® 2, Etest® method and broth microdilution method for colistin were used. PCR analysis and multilocus sequence typing Pasteur scheme were performed to identify bla-OXA genes and genetic relatedness, respectively. Whole-genome sequencing analysis was used to characterize three isolates. Results: All the isolates were susceptible only to polymyxins. blaOXA-23-like gene was the only acquired carbapenemase gene in 88.2% of the isolates. Multilocus sequence typing identified various sequence types: ST2, ST19, ST195, ST577 and ST632. Two new sequence types, namely, ST1279 and ST1280, were detected by whole-genome sequencing. Conclusion: This study showed that carbapenem-resistant A. baumannii isolates causing infections in intensive care units almost exclusively produce OXA-23, underlining their frequent spread in Italy.

Distinct Phenotypic and Genomic Signatures Underlie Contrasting Pathogenic Potential of Staphylococcus epidermidis Clonal Lineages.

Background: Staphylococcus epidermidis is a common skin commensal that has emerged as a pathogen in hospitals, mainly related to medical devices-associated infections. Noteworthy, infection rates by S. epidermidis have the tendency to rise steeply in next decades together with medical devices use and immunocompromized population growth. Staphylococcus epidermidis population structure includes two major clonal lineages (A/C and B) that present contrasting pathogenic potentials. To address this distinction and explore the basis of increased pathogenicity of A/C lineage, we performed a detailed comparative analysis using phylogenetic and integrated pangenome-wide-association study (panGWAS) approaches and compared the lineages's phenotypes in in vitro conditions mimicking carriage and infection. Results: Each S. epidermidis lineage had distinct phenotypic signatures in skin and infection conditions and differed in genomic content. Combination of phenotypic and genotypic data revealed that both lineages were well adapted to skin environmental cues. However, they appear to occupy different skin niches, perform distinct biological functions in the skin and use different mechanisms to complete the same function: lineage B strains showed evidence of specialization to survival in microaerobic and lipid rich environment, characteristic of hair follicle and sebaceous glands; lineage A/C strains showed evidence for adaption to diverse osmotic and pH conditions, potentially allowing them to occupy a broader and more superficial skin niche. In infection conditions, A/C strains had an advantage, having the potential to bind blood-associated host matrix proteins, form biofilms at blood pH, resist antibiotics and macrophage acidity and to produce proteases. These features were observed to be rare in the lineage B strains. PanGWAS analysis produced a catalog of putative S. epidermidis virulence factors and identified an epidemiological molecular marker for the more pathogenic lineage. Conclusion: The prevalence of A/C lineage in infection is probably related to a higher metabolic and genomic versatility that allows rapid adaptation during transition from a commensal to a pathogenic lifestyle. The putative virulence and phenotypic factors associated to A/C lineage constitute a reliable framework for future studies on S. epidermidis pathogenesis and the finding of an epidemiological marker for the more pathogenic lineage is an asset for the management of S. epidermidis infections.