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Methicillin-resistant staphylococci (MRS) are important antimicrobial-resistant pathogens in sepsis. Conventional blood cultures take 24-72 h. The polymerase chain reaction (PCR)-based methods give faster results (2-3 h) but need expensive thermal cyclers. We therefore developed an isothermal recombinase polymerase amplification (RPA) combined with lateral flow dipstick (LFD) assay for rapid detection of MRS in spiked blood culture samples. Fifty-six clinical isolates including 38 mecA-carrying staphylococci and 18 non-mecA-carrying organisms as confirmed by PCR methods were studied. RPA primer set and probe specific for mecA gene (encoding penicillin-binding protein 2a) were designed. RPA reaction was carried out under isothermal condition (45 °C) within 20 min and read by LFD in 5 min. The RPA-LFD provided 92.1% (35/38) sensitivity for identifying MRS in positive blood culture samples, and no cross-amplification was found (100% specificity). This test failed to detect three mecA-carrying S.sciuri isolates. The detection limits of RPA-LFD method for identifying MRS were equal to those of PCR method. The RPA-LFD is simple, fast, and user-friendly. This method could detect the mecA gene directly from the positive blood culture samples without requirement for special equipment. This method would be useful for appropriate antibiotic therapy and infection control, particularly in a low-resource setting.

Both pathogenic as well as nonpathogenic species of staphylococci have been reported in poultry, but these studies have not compared staphylococcal flora of both farmed and household broiler chickens. Staphylococci from farmed (n = 51) and household chicken intestines (n = 43) were isolated and tested for resistance to antimicrobials, presence of resistance genes, and inhibitory activity against other bacteria; correlation of resistance phenotype and genotype was also evaluated. At least 12 staphylococcal species were identified; Staphylococcus carnosus subspecies carnosus was the predominant species from both sources. Most farmed chicken staphylococci were resistant to tigecycline (38/51; 74.8%) while the highest level of resistance among the household chicken staphylococci was to clindamycin (31/43; 72.1%). The mecA gene was only detected in staphylococci from household chickens, whereas ermC and tetK or tetM were found in staphylococci from both groups of birds. Multidrug resistance (resistance ≥ 2 antimicrobial classes) was observed in 88% of resistant staphylococci ranging from 2 to 8 classes and up to 10 antimicrobials. Isolates produced inhibitory activity against 7 clinical bacterial strains primarily Enterococcus faecalis (25/88; 28.4%) and Escherichia coli (22/88; 25%). This study demonstrated that the staphylococcal population among farmed and household chickens varies by species and resistance to antimicrobials. These results may reflect the influence of the environment or habitat of each bird type on the intestinal microflora. As resistance in the staphylococci to antimicrobials used to treat human infections was detected, further study is warranted to determine strategies to prevent transfer of these resistant populations to humans via contamination of the poultry meat., (Copyright © 2020. Published by Elsevier Inc.)

Background: HIV causes a decrease in CD4+ lymphocyte cells count, exposing the individual to infections (urinary tract infections). This study was carried out to determine the prevalence of bacteriuria and antimicrobial susceptibility pattern of bacteria isolates among HIV patients., Methods: Clean catch mid-stream urine samples were collected from 135 HIV- seropositive patients, cultured on Cystein lactose electrolyte deficient (CLED) agar and incubated at 370C for 24 hours. The modified Kirby-Bauer's disc diffusion method was used to assess susceptibility to antimicrobial agents., Results: The prevalence of bacteriuria was 67.4% (91/135). Staphylococcus aureus was the most predominant (42.9%) isolate, followed by Escherichia. coli (24.2%), then Coagulase negative Staphylococci (10.9%). The highest proportion of bacteria was isolated from patients having a CD4+ T-cell count of less than 300 cells/mm3 (39.6%). There was an association between the level of CD4+cell count and bacterial urinary tract infection (P= 0.001). Most sensitive drugs were gentamycin, vancomycin and amoxicillin-clavulanic acid while the drug with the greatest resistance was sulphamethoxazole-trimethoprim, with Enterococcus and Proteus showing 100% resistance to this drug., Conclusion: Bacteriuria and resistance to commonly used antibiotics is prevalent among HIV/AIDS patients attending the Bamenda Regional Hospital. Therapy based on antimicrobial susceptibility test is encouraged., (© 2020 Samje M et al.)

Objectives: To investigate the prevalence and characteristics of methicillin-resistant staphylococci on dairy farms in England and Wales including zoonotic MRSA., Methods: Bulk tank milk was sampled from 363 dairy farms in 2015-2016 and methicillin-resistant staphylococci were isolated by salt broth enrichment and plating on MRSA Brilliance selective agar. Isolates were characterised through antimicrobial susceptibility testing and whole-genome sequencing., Results: Methicillin-resistant staphylococci were isolated from ∼5% of dairy farms and belonged to six different species, Staphylococcus aureus, Staphylococcus epidermidis, Staphylococcus lentus, Staphylococcus saprophyticus, Staphylococcus fleurettii and Staphylococcus sciuri. Whole-genome sequencing revealed a large variety of antimicrobial resistance genes and SCCmec elements were present, including mecA and mecC alleles. Potentially zoonotic methicillin-resistance S. aureus were found at a low prevalence (0.83% of sampled dairy farms). Whole-genome sequencing also provided evidence for the mobility of a primordial mec gene complex, independently of a SCCmec element, which appears to have been acquired by S. saprophyticus from S. fleurettii., Conclusions: These data give new insight into the epidemiology of veterinary methicillin-resistant staphylococci to inform future surveillance and zoonotic risk evaluation. Our data indicate that MRSA has likely decreased in prevalence since earlier survey work in England and Wales during 2011-12 and highlights the diversity of methicillin resistance and other resistance determinants among bovine-associated staphylococci with implications for veterinary and human medicine., (Copyright © 2020 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

The most clinically relevant staphylococci in veterinary medicine are those that are coagulase-positive, namely Staphylococcus aureus. During microbiological udder health monitoring (2009-2018), a new S. aureus strain (coagulase-positive and maltose-negative) was discovered as an emerging udder pathogen during routine examinations of South African dairy herds. This study challenged the conventional microbiological diagnosis of staphylococci by comparing its results to those of the MALDI-TOF mass spectrometry and 16S rRNA sequencing. Both of these tests confirmed that the maltose-negative staphylococcus (MNS), identified as Staphylococcus pseudintermedius by conventional microbiology, was S. aureus ST2992. Multi locus sequence typing was performed on 3 of the MNS isolates and indicated that these isolates were of single origin. These strains tested positive for both MALA and MALR genes (control: S. aureus ATCC 25923). Although the α-glucosidase gene was present, it was not expressed phenotypically. The latter may be attributed to the abnormal stop codon identified in the MALA gene sequence of S. aureus ST2992 (GenBank accession number, MN531305). The newly identified MNS has a field behavior different to that of maltose-positive S. aureus, and more similar to the low virulence of non-aureus staphylococci., (Copyright © 2020 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.)

Six Staphylococcus strains were isolated from healthy black bears ( Ursus americanus ) in the Great Smoky Mountains National Park, Tennessee, USA. Phylogenetic analysis based on complete genome, 16S rRNA, dnaJ , hsp60 , rpoB and sodA genes, and MALDI-TOF-MS main spectral profiles revealed that the strains belonged to one species and showed the closest relatedness to members of the ' Staphylococcus intermedius group' (SIG), which include Staphylococcus intermedius , Staphylococcus pseudintermedius, Staphylococcus delphini and Staphyloccoccus cornubiensis . The strains were positive in SIG-specific and negative in individual species-specific PCR assays for the nuc gene. The strains can be differentiated from the other SIG species by the absence of sucrose fermentation, from S. intermedius DSM 20373 T , S. pseudintermedius CCUG 49543 T and S. cornubiensis DSM 105366 T by the absence of methyl β-d-glucopyranoside fermentation and from S. delphini DSM 20771 T by fermentation of trehalose. DNA relatedness of the type strain MI 10-1553 T with the type strains of S. delphini , S. pseudintermedius , S. intermedius and S. cornubiensis was ≤48.2 % by digital DNA-DNA hybridization and ≤92.3 % by average nucleotide identity calculations. Iso- C15:0 , anteiso-C 15 : 0 and anteiso-C 17 : 0 were the most common fatty acids. Polar lipids consisted of phosphadidylglycerols, phospholipids, glycolipid, diphosphatidylglycerol and aminophospholipid. Cell-wall peptidoglycan was of type A3α l-Lys-Gly 3 (Ser; similar to A11.2 and A11.3). The respiratory quinone belonged to menaquinone 7 (MK-7). The G+C content of MI 10-1553 T was 39.3 mol%. The isolated strains represent a novel species of the genus Staphylococcus , for which we propose the name Staphylococcus ursi sp. nov. The type strain is MI 10-1553 T (=ATCC TSD-55 T =CCOS 1900 T ).

Background: Staphylococci species are the major constituents of infectious bioaerosols, particularly methicillin-resistant Staphylococci (MRS) have serious health impacts. Here, the bacterial burden was quantified, especially prevalence of MRS in bioaerosols collected from indoors of Dr. B.R. Ambedkar Central Library (DBRACL) and Central Laboratory Animal Resources (CLAR) of Jawaharlal Nehru University, New Delhi, India. Air samplings from DBRACL and CLAR were done using the settle plate method and SKC biosampler, respectively., Results: This study showed a maximum 6757 CFU/m 2 /hr of bacterial load in the DBRACL reading room, while unacceptable bacterial loads (> 1000 CFU/m 3 of air) at different sites of CLAR. Further, at both the sampling sites the predominance of coagulase negative Staphylococci (CNS) was observed. A total 22 and 35 Staphylococci isolates were isolated from DBRACL and CLAR bioaerosols, respectively. Majority (16/22) of the Staphylococcal isolates from DBRACL belonged to human-associated Staphylococci where S. haemolyticus (5/22) was the most dominating species. However, in CLAR facility centre, animal-associated Staphylococci (19/35) were dominating, where S. xylosus (12/35) was the most dominating species. Further, antibiotic sensitivity tests revealed 41% MRS and 73% multidrug resistant (MDR) among airborne Staphylococci from DBRACL indoor bioaerosols. Similarly, in CLAR facility, approximately, 66% Staphylococci isolates were methicillin resistant, out of which 2 isolates showed high MIC value ≥ 16 μg/mL. Further, we confirmed the presence of 49% multidrug resistant Staphylococci in the indoor air of CLAR facility., Conclusions: This study suggested that the exposure of workers and students in CLAR to such a high concentration of drug-resistant Staphylococci should not be undermined, as these bacterial concentrations are the direct representative of inhalable particulate matter (PM 2.5 ) as per collection procedure. Simultaneously, passive sampling from DBRACL assessed the risks due to microbial contamination in particle agglomerates, which may deposit on the crucial surfaces such as wounds/ cuts or on the frequently used items.

Objectives: This study aims to investigate the effects of various mouthwashes on bacteremia development following a debonding process, which is performed after orthodontic treatment., Subjects and Methods: The study included patients who received fixed orthodontic treatment and were indicated for debonding. A total of 40 patients in four groups were selected for the study; no mouthwash (Group 1), mouthwash containing 0.12% chlorhexidine-gluconate (Group 2), mouthwash containing essential-oils (Group 3), and mouthwash containing 7.5% povidone-iodine (Group 4). Before (T 0 ) and following (T 1 ) the debonding procedure, blood samples were obtained from the patients. Then, the blood samples were placed in blood culture bottles to investigate bacterial growth., Results: Based on the results of the study, it was determined that the blood samples obtained at T 0 did not indicate any bacterial growth. Furthermore, it was observed that the blood samples obtained at T 1 included Streptococcus viridans, Streptococcus oralis, Streptococcus mutans, and Staphylococcus aereus growth, respectively, in 4 patients from Group 1 while Streptococcus salivarius growth was observed in 1 patient from Group 3 in addition to Streptococcus mitis growth in 1 patient from Group 4. No bacterial growth was observed in Group 2. While the results obtained between Group 1 and Group 2 were statistically significant, no statistically significant difference was observed between other groups., Conclusions: Finally, it was determined that the mouthwash 0.12% chlorhexidine-gluconate was statistically significant in comparison to the control group. It can be concluded that this mouthwash can be used to decrease bacterial density in oral flora before debonding procedures., Competing Interests: None

Staphylococcus agnetis has been previously associated with subclinical or clinically mild cases of mastitis in dairy cattle and is one of several staphylococcal species that have been isolated from the bones and blood of lame broilers. We reported that S. agnetis could be obtained frequently from bacterial chondronecrosis with osteomyelitis (BCO) lesions of lame broilers (A. Al-Rubaye et al., PLoS One 10:e0143336, 2015 [https://doi.org/10.1371/journal.pone.0143336]). A particular isolate, S. agnetis 908, can induce lameness in over 50% of exposed chickens, exceeding normal BCO incidences in broiler operations. We reported the assembly and annotation of the genome of isolate 908. To better understand the relationship between dairy cattle and broiler isolates, we assembled 11 additional genomes for S. agnetis isolates, an additional chicken BCO strain, and ten isolates from cattle milk, mammary gland secretions, or udder skin from the collection at the University of Missouri. To trace phylogenetic relationships, we constructed phylogenetic trees based on multilocus sequence typing and genome-to-genome distance comparisons. Chicken isolate 908 clustered with two of the cattle isolates, along with three isolates from chickens in Denmark and an isolate of S. agnetis we isolated from a BCO lesion on a commercial broiler farm in Arkansas. We used a number of BLAST tools to compare the chicken isolates to those from cattle and identified 98 coding sequences distinguishing isolate 908 from the cattle isolates. None of the identified genes explain the differences in host or tissue tropism. These analyses are critical to understanding how staphylococci colonize and infect different hosts and potentially how they can transition to alternative niches (bone versus dermis). IMPORTANCE Staphylococcus agnetis has been recently recognized as associated with disease in dairy cattle and meat-type chickens. The infections appear to be limited in cattle and systemic in broilers. This report details the molecular relationships between cattle and chicken isolates in order to understand how this recently recognized species infects different hosts with different disease manifestations. The data show that the chicken and cattle isolates are very closely related, but the chicken isolates all cluster together, suggesting a single jump from cattle to chickens., (Copyright © 2020 American Society for Microbiology.)

Mortality of mink kits represents a significant loss to production. However, causes of post-weaning mortality in mink kits in modern Danish mink production systems are still relatively poorly documented. We performed a cross-sectional mortality study on eight Danish mink farms including 1893 post mortem examinations of mink kits found dead or euthanized. We assessed the prevalence of cystitis and urolithiasis leading to mortality. Gross pathological findings as well as animal characteristics were recorded and associations with post mortem microbiology (using culture and MaldiTof-MS Vitek MS system) were investigated. Cystitis and/or urolithiasis were associated with death in 33 % (n = 476) and 37 % (n = 166) of the examined mink kits in 2015 and 2017. On farm level, the prevalence of cystitis and/or urolithiasis leading to mortality varied from 0.25 % to 1.27 % with a low overall mortality of 0.9-4.5 %. The bacterial agent most frequently isolated in post mortem bladder swabs from mink with a post mortem diagnosis of urolithiasis and cystitis was Staphylococcus delphini group A (51/283) with a significant (p < 0.0001, CI = [19.5;4745.7]) association to gross pathological findings in the urinary tract. Staphylococcus delphini group A was cultured from 70 % of the skin swabs obtained from apparently healthy mink euthanized at pelting (n = 222). In conclusion urinary tract disease (cystitis and urolithiasis) was the most prevalent post mortem diagnosis during the growth period and was associated with Staphylococcus delphini group A., (Copyright © 2020 The Authors. Published by Elsevier B.V. All rights reserved.)

The study was carried out in Polish goat population to estimate the prevalence of the nasal cavity infection with various staphylococcal species including methicillin-resistant Staphylococcus aureus(MRSA), investigate the potential permissive role of small ruminant lentivirus (SRLV) infection and determine the level of clonality of S. aureus nasal isolates. Nasal swabs and blood samples were collec-ted from 1300 clinically healthy adult goats from 21 Polish goat herds. Blood samples were serological-ly screened for SRLV. Staphylococci were isolated from nasal swabs and identified using classical microbiological methods, MALDI-TOF, multiplex-PCR, and their clonality was assessed using PFGE. Antimicrobial resistance was determined on the basis of minimum inhibitory concentration and by demonstration of the presence of the mecA gene encoding the multiplex-PCR PBP2a protein and of the five main types of staphylococcal cassette chromosome mec. The apparent prevalence of staphylococ-cal and S. aureus infection of the nasal cavity was 29.1% (CI 95%: 26.9%, 31.5%) and 7.3% (CI 95%: 6.1%, 8.8%), respectively. No relationship was found between the SRLV-infection and the presence of any staphylococcal species including S. aureus (p=0.143). Only 9.8% of S. aureus isolates were resistant to amoxicillin/clavulanic acid and 5.9% to chloramphenicol and ciprofloxacin. All tested isolates proved to be phenotypically and genotypically sensitive to methicillin, which yielded the appar-ent prevalence of MRSA of 0% (CI 95%: 0%, 7.0%). S. aureus isolates show high genetic similarity within goat herds, however vary considerably between herds. Goats do not appear to be an important source of S. aureus for humans in Poland., (Copyright© by the Polish Academy of Sciences.)

Methicillin-resistant Staphylococcus pseudintermedius (MRSP) have recently emerged as a major therapeutic challenge in small animal medicine because of their antimicrobial multidrug resistance and their role as nosocomial pathogens. This study focused on the prevalence, molecular characteristics and antimicrobial resistance pheno- and genotypes of MRSP isolated from conjunctival swabs of dogs and cats. Conjunctival swabs were collected from 72 dogs and 24 cats suffering from conjunctivitis/blepharitis, keratitis or uveitis and screened for the presence of MRSP. S. pseudintermedius was isolated from 38 (39.6 %) of all samples. Three (7.9 %) S. pseudintermedius isolates were confirmed as MRSP. They harboured the mecA gene and originated from dogs. One MRSP isolate was from a case of uveitis while the other two MRSP isolates originated from cases of conjunctivitis/blepharitis. All MRSP isolates were subjected to broth microdilution and whole genome sequencing (WGS). Resistance and virulence genes, multilocus sequence (MLS), spa, dru and SCCmec types were deduced from WGS data. Two of the three MRSP isolates, IMT360/16 and IMT515/16, shared the same MLS type (ST71), spa type (t02), dru type (dt9a), SCCmec type (II-III), and indistinguishable multidrug resistance pheno- and genotypes, including resistance to β-lactams (blaZ, mecA), erythromycin and clindamycin (erm(B)), streptomycin (aphA3), gentamicin (aacA-aphD), enrofloxacin (mutations in grlA and gyrA), tetracycline (tet(K)), and trimethoprim (dfrG)/sulfamethoxazole. The third isolate, IMT1670/16, differed in all those characteristics (MLST (ST1403), dru type (dt10h), SCCmec type (IVg), except the spa type (t02). In addition, isolate IMT1670/16 carried a different tetracycline resistance gene (tet(M)) and was susceptible to erythromycin and clindamycin., Competing Interests: Declaration of Competing Interest None to declare., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Food poisoning risk related to the consumption of contaminated food with known foodborne pathogens or antibiotic-resistant bacteria is currently a serious threat for public health. Thus, pathogenic methicillin-resistant Staphylococcus strains are considered as one of the major cause of foodborne diseases in hospitals. The present study aims to determine the prevalence and the antibiotic resistance patterns of Staphylococcus in various types of hospital food samples, work surfaces and its carriage by food handlers. A total of 608 collected samples including 300 food samples, 238 food contact surfaces and 70 nasal and hand samples were tested. The identified Staphylococcus and their antibiotic resistance patterns were analysed using the agar disk-diffusion and PCR method was used for mecA resistance gene amplification. The prevalence of S. aureus and the coagulase-negative staphylococci were 17·33 and 23·33%, respectively. The antibiotic resistance reached 100% towards oxacillin and Penicillin G for both S. aureus and CoNs. The mecA gene was detected in 5·71% (4/70) and 7·69% (4/52) of S. aureus and CoNs strains, respectively. The outcome of this study enlightens isolation of MRSA strains and resistant CoNs from food, food contact surfaces and food handlers. The presence of this resistant species in this critical setting, where products were intended to vulnerable and immunocompromised patients, represents a serious threat to this community. It can be a source of nosocomial infection and more precautions must be taken to prevent staphylococci food contamination mainly in hospitals. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first study describing the antibiotic resistance patterns of Staphylococcus aureus and coagulase-negative Staphylococcus isolated from hospital food, food contact surfaces and food handlers samples in a Moroccan hospital kitchen. High levels of multi-resistance were reported. The alarming outcome of this study emphasizes the crucial need of implementing an approach to fight multidrug-resistant staphylococci mainly in healthcare settings, where the community have already compromised health issues., (© 2020 The Society for Applied Microbiology.)

Background: Antimicrobial resistance is a problem in human and animal healthcare. Honey may be used for its wound healing properties and antimicrobial effects., Objective: To investigate the antimicrobial activity of two commercially available medical grade honeys (MGHs) against Staphylococcus spp. and Pseudomonas spp. isolates., Methods and Materials: Two formulations, MGH1 (40% w/v honey) and MGH2 (80% w/v Manuka honey), were tested in vitro for minimal inhibitory concentrations (MIC) and minimal bactericidal concentrations (MBC) against 11 Staphylococcus and 11 Pseudomonas isolates at low [1.5 × 10 4  colony forming units (cfu)/well] and high (1.5 × 10 6  cfu/well) concentrations of inoculum, representing systemic and cutaneous bacterial loads during infection, respectively., Results: MGH2 showed a lower MIC against staphylococci than MGH1, although this was not statistically significant. MGH1 had stronger bactericidal effects against staphylococci than MGH2, although this effect was statistically significant only at the higher bacterial concentration (P < 0.01). For Pseudomonas spp., MGH1 had significantly higher antimicrobial activity (both MIC and MBC) than MGH2 against all isolates tested and at both bacterial concentrations (P < 0.05)., Conclusions and Clinical Importance: Both MGHs were effective in vitro against common cutaneous pathogens including meticillin-resistant staphylococci and Pseudomonas species. The higher efficacy of the MGH1 formulation against Pseudomonas and its consistent effects against staphylococci, while containing only half of the amount of honey compared to MGH2, invites further investigation of the mechanisms and clinical applications of MGH1., (© 2019 ESVD and ACVD.)

The recognition in 2011 of the methicillin resistance determinate mecC among staphylococci has raised many questions over its evolution and epidemiology. While mecC has been best studied in Staphylococcus aureus it has also been described in at least nine other species of staphylococci. In most cases these studies are limited to single isolates. In the widespread animal commensal Staphylococcus sciuri mecC has been described in two isolates and is located within a distinct SCCmec-mecC composite element. In this study, a further 11 mecA/mecC S. sciuri isolated from dairy farms in England and Wales in 2015 and 2016 were genome sequenced and characterised. The results show that two variants of the SCCmec-mecC element are present in S. sciuri, differentiated by different ccr alleles and likely to have arisen by homologous recombination. A phylogeny of sixty genome-sequenced S. sciuri isolates was made using core genome multi-locus sequence typing and reveals a diverse population with the SCCmec-mecC element present in four distinct branches, indicative of four independent acquisitions by S. sciuri. Finally, the study identified the rapid clonal expansion of a mecA/mecC lineage of S. sciuri among dairy farms across a wide geographical area which may contribute to the future dissemination of this methicillin resistance cassette., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Introduction : Compared to Staphylococcus aureus , coagulase-negative staphylococci (CoNS) are characterized by a lower capacity to cause acute, live-threatened infections. CoNS are, however, of ever increasing importance as pathogens causing infections in immunocompromised patients and after foreign-material implantation. Typically, antibiotics fail to cure foreign body-related infections and removal of the implanted device is inevitable. Areas covered : This review focuses on the emergence of CoNS species, their pathogenic potential in particular due to their ability to form therapy-refractory biofilms on biotic and abiotic surfaces and evasion strategies to resist host response and antibiotic treatment. Their medical significance and proven and novel therapy strategies are discussed. Expert opinion : CoNS contribute significantly to morbidity and socio-economic costs. The anticipated developments in modern medicine, in particular the increasing use of foreign materials and the rising numbers of immunocompromised patients, as well as the changing demographic and hospital-related factors will inevitably contribute to further emergence of CoNS infections. Increasing rates of (multi-)resistant CoNS strains will limit the therapeutic armamentarium and aggravate treatment strategies. Increased research is necessary to understand their role as resistance and virulence gene reservoir and to reduce CoNS infections by the development of innovative colonization-preventing materials and other CoNS-tailored treatment strategies.

Staphylococcus spp. is one of the pathogens that cause bovine mastitis and may present multiple resistance to different antimicrobial groups. The aim of this study was to phenotypically identify Staphylococcus spp. isolates obtained from bovine milk and to characterize their antimicrobial resistance profile. The 101 strains were classified by phenotypic tests, their resistance to oxacillin, cefoxitin, penicillin, ampicillin, tetracycline, kanamycin, sulfamethoxazole / trimethoprim, clindamycin and erythromycin was determined by the Kirby-Bauer technique and the presence of resistance genes by PCR. A total of 65 strains was S. aureus and 36 strains were coagulase-negative staphylococci (CoNS). We found different patterns of resistance to antibiotics evaluated in strains of S. aureus and CoNS, only the resistance to ampicillin was found associated with the species (p<0.005). In the 101 strains, the mecA gene was detected in 27%, aph(3')-IIIa in 75.2%, aac(6')/aph(2")-3 in 47.4%, ant(4')-Ia in 32.7%, tetM in 63% and tetK in 43.6%; however, no association was found with the resistance to penicillin, ampicillin, cefoxitin, kanamycin and tetracycline, respectively (p>0.05). On the other hand, the blaZ gene was found in 59.4% of the 101 strains and the ermCgene in 62.3%, which was associated with resistance to β-lactams and macrolides, respectively (p<0.001). In this study, antimicrobial multiresistance was found in S. aureus and CoNS strains. This finding impacts on the dairy industry, representing a risk to public health., (Copyright © 2019 Asociación Argentina de Microbiología. Publicado por Elsevier España, S.L.U. All rights reserved.)

The presence of non-aureus staphylococci (NAS) in bovine rectal feces has recently been described. Similar to other mastitis causing pathogens, shedding of NAS in the environment could result in intramammary infection. The objective of this study was to investigate whether NAS strains present in feces can cause intramammary infection, likely via teat apex colonization. During a cross-sectional study in 5 dairy herds, samples were collected from the habitats quarter milk, teat apices, and rectal feces from 25%, 10%, and 25% of the lactating cows, respectively, with a cow serving as the source of one type of sample only. Samples from clinical mastitis cases were continuously collected during the 1-year study period as well. The 6 most prevalent NAS species, Staphylococcus (S.) chromogenes, S. cohnii, S. devriesei, S. equorum, S. haemolyticus, and S. hominis, were further subtyped by random amplification of polymorphic deoxyribonucleic acid polymerase chain reaction (RAPD-PCR), when the same NAS species was present in the same herd in the three habitats. For S. chromogenes, S. cohnii, S. devriesei, and S. haemolyticus, the same RAPD type was found in rectal feces, teat apices, and quarter milk, indicating that fecal NAS can infect the mammary gland. For S. hominis and S. equorum, we were unable to confirm the presence of the same RAPD types in the three habitats.

Non-S. aureus staphylococci (NAS) are the most frequently isolated pathogens from bovine milk and can cause intramammary infections (IMI). They can also be found in teat canals, on bovine skin and in cows' environment, which may lead to unnoticed contamination of milk samples. The aim of this study was to investigate the role of NAS species as mastitis-causing pathogens or contaminants, and to identify possible differences between NAS species. A longitudinal study was conducted with consecutive milk sampling in five German dairy herds. Species identification was performed using matrix-assisted laser desorption ionization-time of flight mass spectrometry. Infections were distinguished from contaminations using two different definitions based on the repeated detection of an NAS species. Of 15 NAS species found, eight and ten, respectively, were associated with an IMI. Staphylococcus simulans and S. chromogenes were associated with IMI in more than 90 % of the findings. S. warneri, S. xylosus, S. microti, S. haemolyticus, and S. succinus seem to be frequent causes of IMI as well as contaminants. If a species-differentiation is available after cultivating NAS, the findings should be interpreted in consideration of the observations made in this study, whether it is more likely a question of a contaminant or a cause of intramammary infection. The bacteria shedding intensity of the NAS species with a more substantially adverse effect on udder health seems to be higher than that of the less important NAS pathogens., Competing Interests: Declaration of Competing Interest The authors declare no conflicts of interest., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Objectives: Enterococcus pRE25 is a conjugative and mobilising multiresistance plasmid from Enterococcus faecalis RE25. pRE25-like enterococcal plasmid pWZ909 mediates the delivery of vancomycin resistance to methicillin-resistant Staphylococcus aureus via a Tn1546-like transposon. However, whether pRE25-like elements contribute to multidrug resistance in Staphylococcus spp. has not yet been investigated. Here we describe the first detection of multiresistance pRE25-like elements in the chromosomal DNA of multidrug-resistant Staphylococcus pseudintermedius (MDRSP)., Methods: A total of 46 MDRSP clinical strains were isolated from canine pyoderma in Korea. Their genetic characteristics were analysed by multilocus sequence typing (MLST) and PCR targeting pRE25-like elements. Whole-genome sequencing (WGS) was performed on four isolates., Results: WGS detected that the chromosomal 22-kb pRE25-like elements contained five antimicrobial resistance genes [cat, erm(B), aphA-3, aadK and sat4], IS1252, IS256, and a toxin-antitoxin system within copies of IS1216. BLASTn alignment analysis revealed that 84% of the chromosomal 22-kb pRE25-like elements sequence is homologous (99.8% identity) to the enterococcal pRE25 plasmid sequence. PCR assay showed that 52.2% of MDRSP isolates were positive for pRE25-like elements and were presumed to contain pRE25-like elements (pRE25 group). The sequence types (STs) of the pRE25 group were diverse, with 18 STs identified, among which 12 STs were first reported in Korea., Conclusion: Enterococcal pRE25-like elements are suspected to be widespread in MDRSP isolated from companion dogs in Korea. Considering that companion dogs live in a closely shared environment with humans, continuous surveillance of pRE25-like elements is needed for other staphylococci commonly isolated from humans., (Copyright © 2019 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Introduction: Ventilator-associated pneumonia (VAP) is one of the common serious infectious diseases encountered in the intensive care unit (ICU), which highly affects the healthcare cost and patient prognosis. VAP is caused by various antimicrobial-resistant aetiological agents and the clinical manifestations lack sensitivity and specificity, making the prompt treatment is a challenge. This study aimed to investigate the microbial profile of VAP causing microorganisms among ICU patients in Egypt, antimicrobial susceptibility patterns and the genetic diversity among the frequently isolated organisms., Methodology: Throughout the period from August 2016 to August 2017, endotracheal aspirate (ETA) specimens were collected from ICU patients with clinically suspected VAP in two tertiary hospitals in Cairo. ETA specimens were investigated for the microbial content. The antimicrobial susceptibility was determined by the Kirby-Bauer method. ERIC-PCR was performed for genotyping., Results: Fifty microbiologically confirmed VAP cases were identified. The most frequently isolated microorganisms were Klebsiella spp., followed by Pseudomonas aeruginosa, Acinetobacter baumannii. Candida spp. was the most isolated fungi. A single isolate of each Cupriavidus pauculus and Aeromonas salmonicida was isolated. Antimicrobial susceptibility profiles indicated 40% of isolates were multidrug-resistant (MDR). ERIC-PCR revealed no genetic relatedness among K. pneumoniae isolates, the most frequently isolated microorganism., Conclusions: Gram-negative bacteria are the main causative agents of VAP cases, which mostly are MDR. Microorganisms like C. pauculus and A. salmonicida should be taken into consideration as VAP causative agents. There was no common source of infection suggesting likely endogenous sources of K. pneumoniae, the main causative agent of VAP in this study., Competing Interests: No Conflict of Interest is declared, (Copyright (c) 2020 Mahmoud Tawfick, Alaa Farag, Mostafa Abozeed, Emad Shaban, Maha Abo-Shadi.)

S. schleiferi is one of the main species isolated from canine otitis externa, pyoderma and from apparently healthy dogs. The species is divided into two subspecies, S. schleiferi schleiferi and S. schleiferi coagulans. MALDI-TOF MS does not distinguish correctly these two subspecies. This study aimed to identify biomarkers that could possibly discriminate Staphylococcus schleiferi subspecies by MALDI-TOF MS. Twelve strains (eight S. schleiferi schleiferi and four S. schleiferi coagulans) were firstly identified. Each isolate was submitted to a protein extraction protocol and subjected to spectrometry on Bruker Microflex LT mass spectrometer. Spectra were analyzed with the BioNumerics software v7.6. Our results showed that spectra clustered according to subspecies, and a set of five MALDI-TOF MS biomarkers were selected to enable the discrimination of S. schleiferi subspecies. In addition, these biomarkers were predicted to represent highly conserved proteins, which could contribute to the identification of subspecies-specific proteins that could be used for improved subspecies identification in clinical samples., (Copyright © 2019 Elsevier B.V. All rights reserved.)

MRSA in humans, pets and livestock have been widely investigated, nevertheless, there is still little information of MRSA in wild animals. Therefore, this study aimed to investigate the occurrence and antimicrobial resistance profiles of methicillin-resistant staphylococci (MRS) in wild Iberian hares and to characterize their genetic lineages. Samples from 83 wild hares (Lepus granatensis) were collected during the hunting season. Isolation of MRS was accomplish using Oxacillin Resistant Screening Agar medium with 2 mg/L of oxacillin. The susceptibility of the isolates was tested by the Kirby-Bauer disc diffusion method. The presence of resistance and virulence genes was studied by PCR. S. aureus strains were further characterized by multilocus sequence typing, agr, spa and SCCmec typing. From the 83 samples, 12 (14.45%) coagulase-negative staphylococci and 3 (3.6%) MRSA strains were isolated. Nine coagulase-negative isolates showed resistance to at least one antibiotic. One MRSA isolate showed a multidrug-resistant profile with resistances to ß-lactams, aminoglycosides, macrolides and lincosamides. All MRSA strains were ascribed to ST2855, t1190 and SCCmec type III. The frequency of MRSA strains in wild hares was low, nevertheless, the presence of MRSA in game animals is considered a public health problem and may represent a route of transmission between animals and humans., (© FEMS 2019.)

We present TaxPhlAn, a new method and bioinformatics pipeline for design and analysis of single-locus sequence typing (SLST) markers to type and profile bacteria beyond the species-level in a complex microbial community background. TaxPhlAn can be applied to any group of phylogenetically-related bacteria, provided reference genomes are available. As TaxPhlAn requires the SLST targets identified to fit the phylogenetic pattern as determined through comprehensive evolutionary reconstruction of input genomes, TaxPhlAn allows for the identification and phylogenetic inference of new biodiversity. Here, we present a clinically relevant case study of high-resolution Staphylococcus profiling on skin of atopic dermatitis (AD) patients. We demonstrate that SLST enables profiling of cutaneous Staphylococcus members at (sub)species level and provides higher resolution than current 16S-based techniques. With the higher discriminative ability provided by our approach, we further show that the presence of Staphylococcus capitis on the skin together with Staphylococcus aureus associates with AD disease.

Different samples of three products including Bikalga and Soumbala from Burkina Faso (West Africa) and Ntoba Mbodi from Congo-Brazzaville (Central Africa) were evaluated. The bacteria (400) were phenotyped and genotypically characterized by Rep-PCR, PFGE, 16S rRNA and rpoB gene sequencing and spa typing. Their PFGE profiles were compared with those of 12,000 isolates in the Center for Disease Control (CDC, USA) database. They were screened for the production of enterotoxins, susceptibility to 19 antimicrobials, presence of 12 staphylococcal toxin and 38 AMR genes and the ability to transfer erythromycin and tetracycline resistance genes to Enterococcus faecalis JH2-2. Fifteen coagulase negative (CoNS) and positive (CoPS) species characterized by 25 Rep-PCR/PFGE clusters were identified: Staphylococcus arlettae, S. aureus, S. cohnii, S. epidermidis, S. gallinarum, S. haemolyticus, S. hominis, S. pasteuri, S. condimenti, S. piscifermentans, S. saprophyticus, S. sciuri, S. simulans, S. warneri and Macrococcus caseolyticus. Five species were specific to Soumbala, four to Bikalga and four to Ntoba Mbodi. Two clusters of S. gallinarum and three of S. sciuri were particular to Burkina Faso. The S. aureus isolates exhibited a spa type t355 and their PFGE profiles did not match any in the CDC database. Bacteria from the same cluster displayed similar AMR and toxin phenotypes and genotypes, whereas clusters peculiar to a product or a location generated distinct profiles. The toxin genes screened were not detected and the bacteria did not produce the staphylococcal enterotoxins A, B, C and D. AMR genes including blazA, cat501, dfr(A), dfr(G), mecA, mecA1, msr(A) and tet(K) were identified in CoNS and CoPS. Conjugation experiments produced JH2-2 isolates that acquired resistance to erythromycin and tetracycline, but no gene transfer was revealed by PCR. The investigation of the heterogeneity of Staphylococcus species from alkaline fermented foods, their relationship with clinical and environmental isolates and their safety in relation to antimicrobial resistance (AMR) and toxin production is anticipated to contribute to determining the importance of staphylococci in alkaline fermented foods, especially in relation to the safety of the consumers., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Temperature and relative humidity are important conditions in paddy storage; however, their influence on microbial communities in stored paddy remains poorly understood. In this study, paddy was stored at different temperatures and relative humidity, and severe mildew was observed in samples stored at relative humidity of 97% and different temperatures (15, 28, and 37 °C) after 50 days. High-throughput 16S rRNA and ITS genes amplicon sequencing analyses showed that Proteobacteria, Firmicutes, and Ascomycota were the dominant phyla across the paddy samples. Moreover, Staphylococcus in bacteria and Aspergillus in fungi were predominant in the composition of microbial community among mildewed paddy samples. The maximum abundance of the two types of organisms appeared in samples stored at 37 °C and relative humidity of 97% (T37H97; 85.02 ± 9.77%), and 15 °C and relative humidity of 97% (T15H97; 93.60 ± 0.01%), respectively. In addition, the two genera were also found in mildew from spoiled paddy by using plate isolation method, revealing their important role in paddy spoilage. Statistical analysis indicated that relative humidity was the most important factor affecting the relative abundances of microbial communities and had the 38.98% and 15.74% contributions to the variation of bacterial and fungal communities, respectively. Hence these results could be very conducive to the promotion of safe storage of paddy., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Staphylococcus petrasii is recently described coagulase negative staphylococcal species and an opportunistic human pathogen, still often misidentified in clinical specimens. Four subspecies are distinguished in S. petrasii by polyphasic taxonomical analyses, however a comparative study has still not been done on the majority of isolates and their genome properties have not yet been thoroughly analysed. Here, we describe the phenotypic and genotypic characteristics of 65 isolates and the results of de novo sequencing, whole genome assembly and annotation of draft genomes of five strains. The strains were identified by MALDI-TOF mass spectrometry to the species level and the majority of the strains were identified to the subspecies level by fingerprinting methods, (GTG) 5 repetitive PCR and ribotyping. Macrorestriction profiling by pulsed-field gel electrophoresis was confirmed to be a suitable strain typing method. Comparative genomics revealed the presence of new mobile genetic elements carrying antimicrobial resistance factors such as staphylococcal cassette chromosome (SCC) mec, transposones, phage-inducible genomic islands, and plasmids. Their mosaic structure and similarity across coagulase-negative staphylococci and Staphylococcus aureus suggest the possible exchange of these elements. Numerous putative virulence factors such as adhesins, autolysins, exoenzymes, capsule formation genes, immunomodulators, the phage-associated sasX gene, and SCC-associated spermidine N-acetyltransferase gene, pseudouridine and sorbitol utilization operons might explain clinical manifestations of S. petrasii isolates. The increasing recovery of S. petrasii isolates from human clinical material, the multi-drug resistance including methicillin resistance of S. petrasii subsp. jettensis strains, and virulence factors homologous to other pathogenic staphylococci demonstrate the importance of the species in human disease., (Copyright © 2019 Elsevier GmbH. All rights reserved.)

Objectives: Methicillin-resistant coagulase-negative staphylococci (CoNS) have emerged as one of the major nosocomial pathogens, and antimicrobial resistance has aggravated the problem. Sixteen million public shared bicycles (PSBs) were launched in China, and 106 million users were recorded. However, no standard clean strategy has been applied to PSBs in China, and no formal surveillance has been carried out. The objective of this study was to understand the diversity and antimicrobial resistance of staphylococcal species from PSBs., Methods: Seventy-nine specimens and 128 isolates were collected from 79 PSBs in Tianjin city, China. Antimicrobial susceptibility and molecular testing were performed to understand the species, antimicrobial resistance, carriage of mecA gene, and clonal lineage., Results: Thirty-five staphylococcal isolates were identified, and 80% of staphylococci were resistant to at least one antibiotic. Seventeen (49%) staphylococci were mecA-positive. SCCmec type V (n = 5), SCCmec type 1A (n = 5), SCCmec type I (n = 3), SCCmec type III (n = 2), 1 SCCmec type II (n = 1), and SCCmec type IV (n = 1) were determined. In addition, Staphylococcus epidermidis ST5, ST35, ST234 and ST419 were determined, and a new ST (ST831) was also found., Conclusions: The Staphylococcus epidermidis group was prevalent among PSBs, and multiple resistant staphylococci were determined in this study. The diversity of SCCmec elements was observed, and PSBs may act as the reservoir for antimicrobial pathogenic bacteria. Moreover, additional studies are necessary to systematically understand the prevalence and molecular epidemiology of methicillin-resistant staphylococci in PSBs., (Copyright © 2019 International Society for Antimicrobial Chemotherapy. Published by Elsevier Ltd. All rights reserved.)

The only known elements encoding enterotoxins in coagulase-negative staphylococci are composite Staphylococcus epidermidis pathogenicity islands (SePIs), including SePI and S. epidermidis composite insertion (SeCI) regions. We investigated 1545 Staphylococcus spp. genomes using whole-genome MLST, and queried them for genes of staphylococcal enterotoxin family and for 29 ORFs identified in prototype SePI from S. epidermidis FRI909. Enterotoxin-encoding genes were identified in 97% of Staphylococcus aureus genomes, in one Staphylococcus argenteus genome and in nine S. epidermidis genomes. All enterotoxigenic S. epidermidis strains carried composite SePI, encoding sec and sel enterotoxin genes, and were assigned to a discrete wgMLST cluster also containing genomes with incomplete islands located in the same region as complete SePI in enterotoxigenic strains. Staphylococcus epidermidis strains without SeCI and SePI genes, and strains with complete SeCI and no SePI genes were identified but no strains were found to carry only SePI and not SeCI genes. The systematic differences between SePI and SeCI regions imply a lineage-specific pattern of inheritance and support independent acquisition of the two elements in S. epidermidis. We provided evidence of reticulate evolution of mobile elements that contain elements with different putative ancestry, including composite SePI that contains genes found in other coagulase-negative staphylococci (SeCI), as well as in S. aureus (SePI-like elements). We conclude that SePI-associated elements present in nonenterotoxigenic S. epidermidis represent a scaffold associated with acquisition of virulence-associated genes. Gene exchange between S. aureus and S. epidermidis may promote emergence of new pathogenic S. epidermidis clones., (© The Author(s) 2019. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.)

Multi-drug resistance traits of Staphylococcus species especially methicillin-resistant Staphylococcus aureus (MRSA) in the clinical settings are well established. Of environmental concern is hospital effluents discharging into wastewaters. This article investigated the prevalence and detection of antibiotic resistance genes in Staphylococcus species from clinical and environmental sources in Ile-Ife, Nigeria. Standard culture-based and molecular protocols were used. Seventy-six (27 clinical, 14 hospital effluent and 35 environmental) Staphylococcus isolates were recovered: 56.58% were coagulase-negative and 43.42% coagulase-positive (S. aureus). For the clinical isolates, 10, 6, 4, 4 and 1 were isolated from urine, skin, wounds, blood and pus, respectively. Isolates were resistant to methicillin and amoxycillin (91.7%), cloxacillin (88.0%), ciprofloxacin (84.0%), ofloxacin (83.3%), azithromycin (78.0%), ceftazidime (76.0%), gentamycin (75.0%), cefuroxime (75.0%) and erythromycin (72.0%). Nearly, all isolates (90.8%) had multiple antibiotic resistance (MAR) index >0.2. Overall MAR indices for Staphylococcus species isolated from the clinical, hospital effluent and environmental wastewaters were relatively similar (0.482; 0.500; 0.435). mecA, nuc and luk-pvl genes were detected in S. aureus, while mecA was detected in S. arlettae, S. sciuri, S. cohnii, S. epidermidis and S. saprophyticus. This study informs on the potential contamination of environmental waters downstream from hospitals and possible impacts that this could have on human and animal health.

Background: Increased prevalence of antimicrobial resistance, treatment failure, and financial losses have been reported in dairy cows with coagulase-negative Staphylococcus (CoNS) clinical mastitis, however, studies on CoNS infections are limited in South Africa. Therefore, the objectives of this study were to investigate the antimicrobial resistance patterns and biofilm formation in CoNS isolated from cow milk samples submitted to the Onderstepoort Milk Laboratory., Results: A total of 142 confirmed CoNS isolates were used for this study. Biofilm formation was identified in 18% of CoNS tested. Staphylococcus chromogenes (11%) had the highest proportion of biofilm formation followed by S. haemolyticus (4.0%), S. epidermidis, S. hominis, S. xylosus, and S. simulans with 1% respectively. Ninety percent (90%) of CoNS were resistant to at least one antimicrobial (AMR) and 51% were multidrug-resistant (MDR). Resistance among CoNS was the highest to ampicillin (90%) and penicillin (89%), few isolates resistant to cefoxitin and vancomycin, 9% respectively. Similarly, MDR-S. haemolyticus (44%), MDR-S. epidermidis (65%), and MDR-S. chromogenes (52%) were mainly resistant to penicillins. The most common resistance patterns observed were resistance to penicillin-ampicillin (16%) and penicillin-ampicillin-erythromycin (10%). Only 42% of biofilm positive CoNS were MDR., Conclusion: The majority of CoNS in this study were resistance to penicillins. In addition, most isolates were β-lactam resistant and MDR. Biofilm formation among the CoNS in this study was uncommon and there was no significant difference in the proportion of MDR-CoNS based on the ability to form a biofilm.

Ultrasound (US) has a high capacity to increase food safety. Although high and/or moderate temperature in combination with US has been studied, the knowledge about cooling/low temperatures as well as its combined effect with chemical preservation methods is scarce. Therefore, the aim of this study was to describe the inactivation of Staphylococcus spp. (SA) present in the natural microbiota of sliced Brazilian dry-cured loin (Socol, BDL) using US (40 kHz and 5.40 W/g) at 1.6-17.9 kJ/g, temperature (T) between 6.4 and 73.6 °C and peracetic acid (PA) between 5.5 and 274.5 mg/L employing the Central Composite Rotatable Design. The model fully describes how the combination of US, T, and PA affects SA inactivation. In BDL, an increase in US acoustic energy density (kJ/g) allows the reduction of T necessary to inactivate SA because of the occurrence of synergistic effect. However, US applied at low T was inefficient. On the other hand, PA was more efficient at low T, since high T degraded this compound at different rates according to the holding T. Therefore, the data indicates a relation between the technologies used in the combined decontamination of sliced BDL improving dry-cured meat safety., (Copyright © 2019 Elsevier B.V. All rights reserved.)

The intestinal microbiota has enormous impact on the health and performance of horses. Staphylococci belong in the phylum Firmicutes, and their occurrence, especially of methicillin-resistant strains and species, has been reported in horses previously. Moreover, biofilm formation is one of the virulence factors; it has been not completely studied in fecal coagulase-negative staphylococci (CoNS) from horses. Therefore, this study was focused on biofilm formation by various species of fecal CoNS from horses because it has been never reported before. In addition, their antibiotic profile was tested. Horses (42) of various breeds from Slovakia/Poland were sampled. Variability in the species of CoNS was detected in feces of horses. Thirty-two strains were identified by using the MALDI-TOF system and classified into nine species and three subspecies of CoNS: Staphylococcus capitis, S. cohnii subsp. cohnii, S. cohnii subsp. urealyticus, S. cohnii subsp. casei, S. epidermidis, S. haemolyticus, S. pasteuri, S. sciuri, S. vitulinus, S. warneri, and S. xylosus. The most frequent species was S. vitulinus. Twenty-two strains showed high biofilm production; 10 strains showed low-grade biofilm production. The highest biofilm formation was measured in the species S. xylosus. Eleven strains (of 32) were methicillin-resistant; the others were susceptible to methicillin.

Staphylococci, and in particular Staphylococcus aureus , cause an extensive variety of infections in a range of hosts. The comprehensive analysis of staphylococcal genomes reveals mechanisms controlling the organism's biology, pathobiology, and dissemination. Whole-genome sequencing technologies led to a quantum leap in our understanding of bacterial genomes. The recent cost reduction of sequencing has resulted in unprecedented volumes of genomic information about S. aureus , one of the most sequenced bacterial species. Collecting, comparing, and interpreting big data is challenging, but fascinating insights have emerged. For example, it is becoming clearer which selective pressures staphylococci face in their habitats and which mechanisms allow this pathogen to adapt, survive, and spread. A key theme is the constant evolution of staphylococci as they alter their genome, exchange DNA, and adapt to new environments, leading to the emergence of increasingly successful, antibiotic-resistant, immune-evading, and host-adapted colonizers and pathogens. This article introduces the structure of staphylococcal genomes, details how genomes vary between strains, outlines the mechanisms of genetic variation, and describes the features of successful clones.

Aims: This study was conducted to determine the occurrence of staphylococci from cows with subclinical mastitis from independent herds in Algeria, and to characterize Staphylococcus aureus isolates., Methods and Results: Quarter milk samples were collected separately, somatic cells were counted and samples with more than 200 000 somatic cells per ml were cultured on blood agar. Staphylococci isolates were identified by routine diagnostics, and S. aureus isolates were tested for antibiotic susceptibility by disk diffusion and microdilution. Congo red agar was used to detect biofilm formation and capsule synthesis was detected on serum soft agar (SSA). The S. aureus isolates were characterized by spa typing. DNA microarray analysis was performed to detect resistance and virulence genes. Overall, 40·0% (167/418) of the cows suffered from mastitis. In 63·5% (106/167) of the cows staphylococci were identified. Nine of the 106 Staphylococcus isolates (8·5%) were S. aureus. The coagulase-negative staphylococci belonged to 14 species. All S. aureus isolates were multiresistant and biofilm forming, with 66·67% of them showing diffuse colonies on SSA and belonged to CC97-agrI-cap5. Biofilm genes (icaA/C/D), 13 genes encoding for adhesion, six genes encoding proteases, 11 genes encoding superantigen like toxins were found. Genes conferring resistance to tetracycline (tet(K)), penicillin (blaZ/I/R) and macrolide-lincosamide-streptogramin B (erm(B), erm(A)) were also detected in the S. aureus from this study., Conclusions: The current investigation provides a detailed molecular and biofilm formation ability of S. aureus involved in subclinical mastitis in Algeria and shows the wide distribution of adhesion and enterotoxin(-like) genes among S. aureus responsible for causing subclinical bovine mastitis., Significance and Impact of the Study: These findings are valuable in tracking the evolution and genomic variation of S. aureus from bovine origin., (© 2019 The Society for Applied Microbiology.)

Background: Ear, nose and throat involvement in granulomatosis with polyangiitis (GPA) is frequently the initial disease manifestation. Previous investigations have observed a higher prevalence of Staphylococcus aureus in patients with GPA, and chronic nasal carriage has been linked with an increased risk of disease relapse. In this cross-sectional study, we investigated changes in the nasal microbiota including a detailed analysis of Staphylococcus spp. by shotgun metagenomics in patients with active and inactive granulomatosis with polyangiitis (GPA). Shotgun metagenomic sequence data were also used to identify protein-encoding genes within the SEED database, and the abundance of proteins then correlated with the presence of bacterial species on an annotated heatmap., Results: The presence of S. aureus in the nose as assessed by culture was more frequently detected in patients with active GPA (66.7%) compared with inactive GPA (34.1%). Beta diversity analysis of nasal microbiota by bacterial 16S rRNA profiling revealed a different composition between GPA patients and healthy controls (P = 0.039). Beta diversity analysis of shotgun metagenomic sequence data for Staphylococcus spp. revealed a different composition between active GPA patients and healthy controls and disease controls (P = 0.0007 and P = 0.0023, respectively), and between healthy controls and inactive GPA patients and household controls (P = 0.0168 and P = 0.0168, respectively). Patients with active GPA had a higher abundance of S. aureus, mirroring the culture data, while healthy controls had a higher abundance of S. epidermidis. Staphylococcus pseudintermedius, generally assumed to be a pathogen of cats and dogs, showed an abundance of 13% among the Staphylococcus spp. in our cohort. During long-term follow-up of patients with inactive GPA at baseline, a higher S. aureus abundance was not associated with an increased relapse risk. Functional analyses identified ten SEED protein subsystems that differed between the groups. Most significant associations were related to chorismate synthesis and involved in the vitamin B 12 pathway., Conclusion: Our data revealed a distinct dysbiosis of the nasal microbiota in GPA patients compared with disease and healthy controls. Metagenomic sequencing demonstrated that this dysbiosis in active GPA patients is manifested by increased abundance of S. aureus and a depletion of S. epidermidis, further demonstrating the antagonist relationships between these species. SEED functional protein subsystem analysis identified an association between the unique bacterial nasal microbiota clusters seen mainly in GPA patients and an elevated abundance of genes associated with chorismate synthesis and vitamin B 12 pathways. Further studies are required to further elucidate the relationship between the biosynthesis genes and the associated bacterial species.

A nonpathogenic endophytic bacterial isolate, recovered from Fragaria × ananassa stolon, and its antifungal activity against Colletotrichum nymphaeae was evaluated under in vitro, in vivo, and greenhouse conditions. Bacterial isolate was identified as Staphylococcus sciuri MarR44 (Strain ID: WDCM 891 = CCSM-B 00640) using phenotypic and biochemical properties and molecular phylogenetic analysis of the 16S rDNA gene sequences. The living cells of strain MarR44 inhibited mycelial growth of C. nymphaeae (52.46%) using dual-culture method. The volatile compounds (VOCs) produced by MarR44 inhibited mycelial growth and conidial germination of C. nymphaeae by 34.52% and 82.81%, respectively. However, inhibition percentage of mycelial growth of pathogen by culture filtrates of the strain MarR44 was lower (23.07%) than that for the two dual culture and volatile compounds assay tests. Moreover, the cell-free-culture filtrates of this strain reduced the biomass and conidial germination of pathogen by 91.89% and 41.10%, respectively. Also, the strain MarR44 was capable of producing protease, chitinase, HCN, siderophore, IAA, gibberellin, and biofilm. The living cells and volatile compounds of the strain MarR44 reduced anthracnose disease at post-harvest on fruit by 52.45% and 72.17%, respectively. Furthermore, disease severity of strawberry anthracnose was reduced using drenching soil and inoculated plants methods by 77.77 and 72.22%, respectively, 60 days after inoculation. The VOCs released by strain MarR44 were analyzed by Gas chromatography-mass spectroscopy (GC-MS). Out of 24 identified VOCs, Mesityl oxide (81.436%), Acetic acid, 2-methylpropyl ester (3.442%), 4-Methyldecane (1.837%), 4-Penten-2-one,4-methyl- (1.739%), Toluene (1.248%), and o-Xylene (1.24%) were the major components. The mode of action of S. sciuri MarR44 on the C. nymphaeae was through the production of antifungal volatile compounds (Antibiosis), which inhibited mycelial growth and conidial germination of pathogen in vitro and fruit decay development in vivo. To the best of our knowledge, this is the first report of S. sciuri having antifungal activity against causal agent strawberry anthracnose. These results indicated that the VOCs of S. sciuri strain MarR44 are promising biofumigant for management of strawberry anthracnose., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Bulk tank milk from 58 dairy goat and sheep flocks located in southern Spain was examined to determine the prevalence and distribution of Staphylococci . A total of 45 isolates were obtained and characterized to determine the species, antimicrobial resistance profile, and genetic similitude by pulse-field gel electrophoresis (PFGE) using Sma I. Staphylococcus aureus isolates were confirmed by polymerase chain reaction (PCR) analysis of nuc , and resistance to methicillin was determined by PCR analysis of mec A. A total of 10 different staphylococcal species were identified, 22.2% and 77.8% of which were coagulase positive and negative, respectively. Twenty-two (48.89%) isolates were resistant to at least one antimicrobial agent. Higher antimicrobial resistance values were obtained against tetracycline (28.9%) and penicillin (22.2%). Two isolates ( S. aureus and Staphylococcus lentus ) were resistant to cefoxitin; however, none of the 45 isolates harbored mec A. Thirty pulsotypes were detected by PFGE. Interestingly, some isolates of S. aureus , S. lentus , Staphylococcus simulans , and Staphylococcus caprae showed high genetic similarity (>80%). These data suggest that genetically similar staphylococcal isolates circulate among goat and sheep dairy herds, and their different resistance patterns could be influenced by the management systems used.

The aims of this study were to determine whether non-aureus staphylococci (NAS) are present in rectal feces of healthy dairy cows, and if so, to delineate species to which they belong and to study several phenotypic and genotypic traits as a first step toward determining the potential impact of fecal shedding of NAS on bovine udder health. Fecal samples were aseptically collected from the rectum of 25 randomly selected clinically healthy dairy cows in a commercial dairy herd using an automated milking system. Fecal NAS were isolated and then identified at the species level using transfer RNA-intergenic spacer PCR and sequencing of the 16S rRNA housekeeping gene. Strain typing was performed using random amplification of polymorphic DNA (RAPD)-PCR. The antimicrobial resistance profiles, biofilm formation, and growth and inhibitory characteristics of all NAS isolates were evaluated. Half of the cows were shedding NAS, resulting in 31 NAS isolates belonging to 11 different species. The most prevalent species were Staphylococcus rostri (23%, n = 7), Staphylococcus cohnii (16%, n = 5), and Staphylococcus haemolyticus (13%, n = 4) with all Staphylococcus agnetis, Staphylococcus chromogenes, and Staph. rostri isolates belonging to the same strain according to RAPD banding patterns. Acquired antimicrobial resistance was observed in 28 of the 31 NAS isolates, mainly due to β-lactamase production. Most of the isolates (84%, n = 27) had a weak biofilm-forming potential, but only 2 contained the bap gene. The ica and aap genes were not detected in any of the isolates. In vitro growth of Staphylococcus aureus and Streptococcus dysgalactiae was inhibited by Staph. agnetis isolates, and Staph. chromogenes isolates were able to inhibit the growth of Strep. dysgalactiae and Streptococcus uberis. All fecal isolates were able to grow when oxygen and iron were limitedly available, mimicking the growth conditions in the mammary gland., (Copyright © 2019 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.)

The antibiotic resistance among coagulase - negative Staphylococcus (CoNS) species towards methicillin is rarely reported in veterinary medicine. Under the aspect/concept of One Health, those strains pose a risk to human health due to the presence in canine pets where the transfer of resistant genetic markers might occur to other staphylococci species. The aim of this study was to investigate the antimicrobial resistance pattern among Coagulase Negative Staphylococci (CoNS) isolated from asymptomatic dogs and those affected by topic infections. Swabs from 254 dogs were first seeded in Mannitol Salt Agar. Species identification was conducted by Matrix Assisted Laser Desorption ionization - time of flight (MALDI-TOF ms) as previously described. The susceptibility test was performed by disk diffusion according to CLSI standards. Detection of mecA gene was performed. CoNS could be recovered from both groups of dogs and an alarming presence of methicillin-resistant coagulase negative staphylococci (MRCoNS) was confirmed, in 10.2% (17/166) of the samples. Eight of those methicillin resistant strains were isolated from asymptomatic dogs whereas nine were present in dogs affected by pyoderma and otitis externa., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

The microbiota of traditional dry-cured sausages and industrial environment was assessed to characterize the diversity of coagulase-negative staphylococci (CNS), and establish potential relationships with hygiene level or technological characteristics. Eight processing units from South Portugal were audited according to a checklist of requirements. Environmental and products' samples at different production stages were evaluated regarding hygiene and safety criteria. CNS were recovered, characterized, and their potential use as starters evaluated. Low genetic diversity was observed for Staphylococcus xylosus, whereas Staphylococcus equorum showed diverse genetic profiles. Staphylococcus xylosus predominated in products with a long period of cold smoking, Staphylococcus saprophyticus in products with a long period of hot smoking, Staphylococcus epidermidis in products with a short period of cold smoking, and S. equorum in nonsmoked products. Most S. xylosus were resistant to tetracycline, whereas S. equorum were susceptible. Antibioresistance restricted the selection of starters due to safety recommendations. PRACTICAL APPLICATION: The present manuscript highlighted a few staphylococci strains that could potentially be used as starter cultures in fermented meat products. These selected strains do not show resistance to antimicrobials, exhibit adequate technological features, and are well adapted to the industrial environments of meat processing industries using different processing technologies. Therefore, the selected strains ready to be used in the manufacturing of traditional fermented meat products to ensure safety, standardize product properties, and shorten ripening., (© 2019 Institute of Food Technologists®.)

Background: Resistance to methicillin and mupirocin in coagulase-negative staphylococci (CoNS) is considered as one of the most important issues. Meanwhile, use of phenotypic methods in detecting the resistance to methicillin and mupirocin has a lower accuracy and speed compared to other molecular methods, such as High-Resolution Melting Analysis (HRM). In this study, HRM technique was used to identify CoNS resistance stains., Methods: This experimental study was done on 86 isolates of CoNS strains isolated at Hamadan Hospital. Pheno-typic tests were used to identify Staphylococcus saprophyticus and S. epidermidis. Methicillin and Mupirocin resistant strains were tested using the MIC and HRM methods, and sensitivity and specificity of the primers were determined based on melting curve temperature range. In addition, data was analyzed by Applied Biosystems StepOne v 2.3 and Applied Biosystems HRM v 3.0.1 software., Results: Eighty-six (86) coagulase-negative isolates were isolated from different clinical specimens. Among these, 69 isolates of S. epidermidis and 17 isolates of S. saprophyticus were identified. Of the 69 S. epidermidis isolates, 19 isolates with oxacillin MIC ≥ 0.5 µg/mL and methicillin-resistant, and 11 isolates with mupirocin MIC ≥ 32 µg/mL and resistant to mupirocin. Of the 17 S. saprophyticus isolates, three isolates with oxacillin MIC ≥ 0.5 µg/mL were also methicillin-resistant, and one isolate with mupirocin MIC ≥ 32 µg/mL was resistant to mupirocin. Melting curve analysis for mecA and mupA primers was determined 81.7 ± 0.5°C and 74 ± 0.5°C, respectively. Sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of HRM assay for detection of methicillin and mupirocin strains testing were 100%., Conclusions: Identification of methicillin and mupirocin resistance in CoNSs using the HRM method has high sensitivity and specificity. Molecular methods are more accurate and faster than phenotypic methods and can identify a large number of resistant isolates in a short time.

Brining is an important step in cheese making, and using brine baths for this purpose is common practice in German dairies. Time of brining, brine concentration, and composition of the complex and heterogeneous microbiota, including coagulase-negative staphylococci (CNS), contribute to the ripening and taste of cheese. As well as producing staphylococcal enterotoxins, some CNS show antibiotic resistance; therefore, we isolated 52 strains of presumptive CNS from cheese brines from 13 factories in Germany. Species identification by sodA gene sequencing revealed that 50 isolates were CNS: 31 Staphylococcus saprophyticus, 4 Staphylococcus carnosus, 4 Staphylococcus equorum, 3 Staphylococcus sciuri, 2 Staphylococcus hominis, and 2 Staphylococcus warneri. One isolate each was identified as Staphylococcus epidermidis, Staphylococcus pasteurii, Staphylococcus succinus, and Staphylococcus xylosus. Further subtyping of the Staph. saprophyticus isolates to the subspecies level revealed the presence of 6 Staph. saprophyticus ssp. saprophyticus. Using pulsed-field gel electrophoresis with the identified Staph. saprophyticus strains, 12 independent clones were identified, resulting in the exclusion of 18 strains from further testing. In 19 of the remaining 32 CNS isolates, resistance to antibiotics was observed. Resistance was found against oxacillin (17), penicillin (5), and cefoxitin (1). Four isolates expressed resistance to both oxacillin and penicillin. No resistance was found to enrofloxacin, tetracycline, gentamicin, or erythromycin. Then, PCR analysis for antibiotic resistance genes was performed for 22 different genes. Only genes blaZ and bla TEM were found in 7 isolates. These isolates were selected for challenge tests with different concentrations of lactic acid and NaCl to examine whether expression of antibiotic resistance was influenced by these stressors. An increase in the minimal inhibitory concentration from 0 to 2.0 µg/mL was seen for trimethoprim/sulfamethoxazole only in one isolate of Staph. saprophyticus at an increased lactic acid concentration. Finally, all isolates were tested for genetic determinants (entA, entB, entC, entD, and entE) of the most common staphylococcal enterotoxins; none of these genes were detected. We found no indication for unacceptable risks originating from the isolated CNS., (Copyright © 2019 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.)

Objectives: Staphylococcus argenteus has been increasingly reported since the species was defined as a novel staphylococcal species in 2015. This study aims to investigate genetic epidemiological links and antimicrobial susceptibilities of methicillin-resistant S. argenteus isolates recovered in Stockholm., Methods: Sixteen methicillin-resistant S. argenteus isolates were identified from a collection of methicillin-resistant Staphylococcus aureus in Stockholm 2007-2018, by using whole-genome sequencing and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). The genomes of the isolates were investigated by pulsed-field gel electrophoresis, single-nucleotide polymorphism (SNP)-based phylogeny, k-mer analysis, core-genome multi-locus sequence typing (cgMLST), resistance traits and virulence factors. The MICs of 19 antimicrobial agents for each isolate were determined by using the broth microdilution method., Results: Of the 16 isolates, seven, seven and two isolates were assigned to ST1223, ST2250 and ST2793, respectively, with the S. aureus MLST-scheme. Analyses based on SNPs and cgMLST revealed a likely clonal spread of methicillin-resistant S. argenteus in 2007. Four isolates were found to be resistant to non-β-lactams in antimicrobial susceptibility testing., Conclusions: A transmission event of methicillin-resistant S. argenteus in family was identified by this study. Among our limited number of isolates, non-β-lactam resistance was detected, which highlights the necessity of a continued surveillance on this emerging pathogen. S. argenteus could be correctly identified by MALDI-TOF MS with the updated database, enabling its detection also in clinical laboratories., (Copyright © 2019 European Society of Clinical Microbiology and Infectious Diseases. Published by Elsevier Ltd. All rights reserved.)

The aim of this study was to assess the prevalence, the genotypic diversity, the antimicrobial resistance traits of canine and feline clinical methicillin-resistant Staphylococcus pseudintermedius (MRSP) isolates in a diagnostic laboratory in Italy during 2015-2016. All isolates were characterized by multilocus sequence typing (MLST), staphylococcal cassette chromosome (SCC)-mec typing and staphylococcal protein A (spa)-typing. The resistance profiles were assessed by antimicrobial susceptibility testing and confirmed genotypically by the detection of mecA gene and by microarray analyses. The prevalence of MRSP isolates was high (31.6%). All the strains were multidrug resistant and the most frequent clone was ST71-SCCmec type II-III. These results confirm a high prevalence of MRSP amongst clinical samples from pets in Italy. These isolates show multidrug resistance features that are of concern both in veterinary and human medicine for clinical and epidemiological reasons., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

The aim of the study was to analyze the reasons for removing implants after breast reconstruction in the course of treatment of breast cancer. The study involved 428 patients, who underwent a total of 648 breast reconstruction procedures using artificial implants. 47 out of 648 cases (7.3%) were identified in which the implant had to be removed. Of the 47 cases, 57.4% had undergone deferred reconstruction, and 42.6% immediate reconstruction; 27.7% had undergone pre-operative chemotherapy and radiotherapy, 27.7% pre-operative chemotherapy, and 2.1% pre-operative radiotherapy; 6.4% were diabetic, 4.3% active smokers, and more than 50.0% had BMI greater than 25 kg/m 2 . In 83.0% of the analyzed cases, the reason for removal of the implant was infection, in 8.5% it was local recurrence of breast cancer, in 4.3% it was damage (leakage) of the implant, and in 2.1% it was post-operative pain. About 87.0% of infections appeared within one year of implantation; however, less than a half developed within 90 days of the reconstructive surgery, and up to 30 days only about 13.0% had appeared. Among the etiological agents of infections were: coagulase-negative Staphylococcus (31.3%), Staphylococcus aureus (18.7%), Enterococcus faecalis (9.4%), Enterobacter cloacae (18.8%), Pseudomonas aeruginosa (12.5%), Acinetobacter lwoffii (3.1%), and other Gram-negative fermenting rods accounted for 6.2%. Infections were the most common reason for removing the implant after breast reconstruction. and occurred most often as late infections (>30 days after surgery). The time of observation for infectious complications should be at least 1 year., The aim of the study was to analyze the reasons for removing implants after breast reconstruction in the course of treatment of breast cancer. The study involved 428 patients, who underwent a total of 648 breast reconstruction procedures using artificial implants. 47 out of 648 cases (7.3%) were identified in which the implant had to be removed. Of the 47 cases, 57.4% had undergone deferred reconstruction, and 42.6% immediate reconstruction; 27.7% had undergone pre-operative chemotherapy and radiotherapy, 27.7% pre-operative chemotherapy, and 2.1% pre-operative radiotherapy; 6.4% were diabetic, 4.3% active smokers, and more than 50.0% had BMI greater than 25 kg/m 2 . In 83.0% of the analyzed cases, the reason for removal of the implant was infection, in 8.5% it was local recurrence of breast cancer, in 4.3% it was damage (leakage) of the implant, and in 2.1% it was post-operative pain. About 87.0% of infections appeared within one year of implantation; however, less than a half developed within 90 days of the reconstructive surgery, and up to 30 days only about 13.0% had appeared. Among the etiological agents of infections were: coagulase-negative Staphylococcus (31.3%), Staphylococcus aureus (18.7%), Enterococcus faecalis (9.4%), Enterobacter cloacae (18.8%), Pseudomonas aeruginosa (12.5%), Acinetobacter lwoffii (3.1%), and other Gram-negative fermenting rods accounted for 6.2%. Infections were the most common reason for removing the implant after breast reconstruction. and occurred most often as late infections (>30 days after surgery). The time of observation for infectious complications should be at least 1 year.

Objectives: Coagulase-negative staphylococci such as Staphylococcus cohnii are opportunistic pathogens of humans and animals. A multidrug-resistant isolate of S. cohnii subsp. urealyticus (073AN) was isolated from the nasal cavity of a healthy goat in Tanzania. This study produced and analysed a draft genome sequence of strain 073AN to investigate the genetic basis for antimicrobial resistance in this isolate., Methods: Strain 073AN was sequenced using HiSeq 2000 technology, sequencing reads were assembled using Velvet, and the genome was annotated using Prokka., Results: The draft genome of strain 073AN is 2677652bp in size with a GC content of 32.5%. The isolate was resistant to several classes of antimicrobials, which correlated with the presence of known antimicrobial resistance genes. Of particular note, the draft genome sequence of strain 073AN represents the first report of fosD in S. cohnii and the first descriptions of fosD and fusF in Africa., Conclusion: The draft genome sequence of S. cohnii subsp. urealyticus 073AN released here provides an insight into the antimicrobial resistance determinants found in this species and in Tanzania and offers a valuable resource for further studies on staphylococcal genomics and antimicrobial resistance., (Copyright © 2019 International Society for Antimicrobial Chemotherapy. Published by Elsevier Ltd. All rights reserved.)

Background: Progress in contemporary medicine is associated with an increasing number of immunocompromised individuals. In this vulnerable group, the underlying disease together with long-term hospitalization and the use of medical devices facilitate infections by opportunistic pathogens, of which coagulase-negative staphylococci (CoNS) represent a prime example., Objectives: The diversity of CoNS with species- and strain-specific differences concerning virulence and clinical impact is highlighted. A focus is on the ability of CoNS to generate biofilms on biotic and abiotic surfaces, which enables skin and mucosa colonization as well as establishment of CoNS on indwelling foreign bodies., Sources: Literature about the virulence of CoNS listed in PubMed was reviewed., Content: Most catheter-related and prosthetic joint infections as well as most other device-related infections are caused by CoNS, specifically by Staphylococcus epidermidis and Staphylococcus haemolyticus. A common theme of CoNS infections is a high antibiotic resistance rate, which often limits treatment options and contributes to the significant health and economic burden imposed by CoNS., Implications: Breaching the skin barrier along with the insertion of medical devices offers CoNS opportunities to gain access to host tissues and to sustain there by forming biofilms on foreign body surfaces. Biofilms represent the perfect niche to protect CoNS from both the host immune response and the action of antibiotics. Their particular lifestyle, combined with conditions that facilitate host colonization and infection, has led to the growing impact of CoNS as pathogens. Moreover, CoNS may serve as hidden reservoirs for antibiotic resistance and virulence traits., (Copyright © 2018 European Society of Clinical Microbiology and Infectious Diseases. Published by Elsevier Ltd. All rights reserved.)

A single-tube multiplex real-time PCR targeting the nuclease (nuc) gene and subsequent high-resolution melting analysis (HRMA) were used to identify 13 different Staphylococcus species. The nuc gene was targeted due to its low intraspecies variation and the greater interspecies variation than the 16S rRNA gene in Staphylococcus. We used HRMA software that can store and compare HRMA profiles from different runs as long as the runs contain the same reference reaction. Thus, we reduced the 14 PCRs to 2 different PCRs, one targeting the unknown sequence and the other targeting the reference sequences to screen 13 different Staphylococcus species. The specificity of the developed method was tested on 16 different Staphylococcus reference strains and 115 different field strains that were isolated from the milk of cattle with subclinical mastitis. We conclude that the method can be used to quickly and cost-effectively differentiate Staphylococcus aureus (S. aureus) from other Staphylococcus species (S. epidermidis, S. lugdunensis, S. schleiferi, S. hyicus, S. chromogenes, S. lentus, S. haemolyticus, S. xylosus, S. saprophyticus, S. warneri, S. simulans and S. hominis)., (Copyright © 2019 Elsevier B.V. All rights reserved.)