scholarly journals Culture-Independent Genotyping, Virulence and Antimicrobial Resistance Gene Identification of Staphylococcus aureus from Orthopaedic Implant-Associated Infections

2021 ◽  
Vol 9 (4) ◽  
pp. 707
Author(s):  
J. Christopher Noone ◽  
Fabienne Antunes Ferreira ◽  
Hege Vangstein Aamot
Keyword(s):  
Staphylococcus Aureus ◽  
Antimicrobial Resistance ◽  
Resistance Genes ◽  
Virulence Gene ◽  
Orthopaedic Implant ◽  
Metagenomic Sequencing ◽  
Gene Detection ◽  
Shotgun Metagenomic Sequencing ◽  
Antimicrobial Resistance Genes ◽  
Culture Independent

Our culture-independent nanopore shotgun metagenomic sequencing protocol on biopsies has the potential for same-day diagnostics of orthopaedic implant-associated infections (OIAI). As OIAI are frequently caused by Staphylococcus aureus, we included S. aureus genotyping and virulence gene detection to exploit the protocol to its fullest. The aim was to evaluate S. aureus genotyping, virulence and antimicrobial resistance genes detection using the shotgun metagenomic sequencing protocol. This proof of concept study included six patients with S. aureus-associated OIAI at Akershus University Hospital, Norway. Five tissue biopsies from each patient were divided in two: (1) conventional microbiological diagnostics and genotyping, and whole genome sequencing (WGS) of S. aureus isolates; (2) shotgun metagenomic sequencing of DNA from the biopsies. Consensus sequences were analysed using spaTyper, MLST, VirulenceFinder, and ResFinder from the Center for Genomic Epidemiology (CGE). MLST was also compared using krocus. All spa-types, one CGE and four krocus MLST results matched Sanger sequencing results. Virulence gene detection matched between WGS and shotgun metagenomic sequencing. ResFinder results corresponded to resistance phenotype. S. aureus spa-typing, and identification of virulence and antimicrobial resistance genes are possible using our shotgun metagenomics protocol. MLST requires further optimization. The protocol has potential application to other species and infection types.

Virulence and antimicrobial resistance genes associated with the in-vivo pathogenicity of avian pathogenic E. coli isolates

2021 ◽  
Vol 1 (1) ◽  
pp. 17-20
Author(s):  
Ahmed Abd El-Mawgoud ◽  
Azza El-Sawah ◽  
Soad Nasef ◽  
Al-Hussien Dahshan ◽  
Ahmed Ali
Keyword(s):  
Antimicrobial Resistance ◽  
Resistance Genes ◽  
Broiler Chickens ◽  
Virulence Genes ◽  
Virulence Gene ◽  
Health Concern ◽  
Pathogenicity Test ◽  
E Coli ◽  
Antimicrobial Resistance Genes

In the current study, ten avian pathogenic E. coli (APEC) isolates of the most predominant APEC serogroups isolated from broiler chickens in Egypt were screened for their virulence and antimicrobial resistance genes pattern using PCR. Five selected virulence gene patterns were further investigated for their in-vivo pathogenicity test. Results showed a 100% prevalence of the β-lactams and tetracyclines resistance genes. However, aminoglycoside and quinolone resistance genes were not detected. Also, 80% of the tested isolates harbored mcr-1 gene, colistin resistance gene. In-vivo pathogenic strains consistently harbored the virulence gene pattern of fimH, fimA, papC, iutA, and tsh. Additionally, the tsh gene was consistently detected with lethal APEC isolates in day-old chicks. These results highlighted the high prevalence of antimicrobial and virulence genes in APEC that potentially represent a public health concern. In this study, the virulence genes fimH, fimA, papC, iutA, and tsh were the most common virulence gene patterns associated with pathogenicity in day-old chicks.

scholarly journals Merging Metagenomics and Spatial Epidemiology To Understand the Distribution of Antimicrobial Resistance Genes from Enterobacteriaceae in Wild Owls

2020 ◽  
Vol 86 (20) ◽  
Author(s):  
Elizabeth A. Miller ◽  
Julia B. Ponder ◽  
Michelle Willette ◽  
Timothy J. Johnson ◽  
Kimberly L. VanderWaal
Keyword(s):  
Antimicrobial Resistance ◽  
Resistance Genes ◽  
Spatial Epidemiology ◽  
Anthropogenic Sources ◽  
Resistant Bacteria ◽  
Metagenomic Sequencing ◽  
Wild Animals ◽  
Natural Ecosystems ◽  
Content Type ◽  
Antimicrobial Resistance Genes

ABSTRACT Antimicrobial resistance (AMR) is a well-documented phenomenon in bacteria from many natural ecosystems, including wild animals. However, the specific determinants and spatial distribution of resistant bacteria and antimicrobial resistance genes (ARGs) in the environment remain incompletely understood. In particular, information regarding the importance of anthropogenic sources of AMR relative to that of other biological and ecological influences is lacking. We conducted a cross-sectional study of AMR in great horned owls (Bubo virginianus) and barred owls (Strix varia) admitted to a rehabilitation center in the midwestern United States. A combination of selective culture enrichment and shotgun metagenomic sequencing was used to identify ARGs from Enterobacteriaceae. Overall, the prevalence of AMR was comparable to that in past studies of resistant Enterobacteriaceae in raptors, with acquired ARGs being identified in 23% of samples. Multimodel regression analyses identified seasonality and owl age to be important predictors of the likelihood of the presence of ARGs, with birds sampled during warmer months being more likely to harbor ARGs than those sampled during cooler months and with birds in their hatch year being more likely to harbor β-lactam ARGs than adults. Beyond host-specific determinants, ARG-positive owls were also more likely to be recovered from areas of high agricultural land cover. Spatial clustering analyses identified a significant high-risk cluster of tetracycline resistance gene-positive owls in the southern sampling range, but this could not be explained by any predictor variables. Taken together, these results highlight the complex distribution of AMR in natural environments and suggest that both biological and anthropogenic factors play important roles in determining the emergence and persistence of AMR in wildlife. IMPORTANCE Antimicrobial resistance (AMR) is a multifaceted problem that poses a worldwide threat to human and animal health. Recent reports suggest that wildlife may play an important role in the emergence, dissemination, and persistence of AMR. As such, there have been calls for better integration of wildlife into current research on AMR, including the use of wild animals as biosentinels of AMR contamination in the environment. A One Health approach can be used to gain a better understanding of all AMR sources and pathways, particularly those at the human-animal-environment interface. Our study focuses on this interface in order to assess the effect of human-impacted landscapes on AMR in a wild animal. This work highlights the value of wildlife rehabilitation centers for environmental AMR surveillance and demonstrates how metagenomic sequencing within a spatial epidemiology framework can be used to address questions surrounding AMR complexity in natural ecosystems.

scholarly journals Rapid Diagnostics of Orthopaedic-Implant-Associated Infections Using Nanopore Shotgun Metagenomic Sequencing on Tissue Biopsies

2021 ◽  
Vol 9 (1) ◽  
pp. 97
Author(s):  
J. Christopher Noone ◽  
Karin Helmersen ◽  
Truls Michael Leegaard ◽  
Inge Skråmm ◽  
Hege Vangstein Aamot
Keyword(s):  
Antimicrobial Resistance ◽  
University Hospital ◽  
Orthopaedic Implant ◽  
Metagenomic Sequencing ◽  
Pathogen Identification ◽  
Shotgun Metagenomics ◽  
Microbial Identification ◽  
Conventional Culture ◽  
Shotgun Metagenomic Sequencing ◽  
Antimicrobial Resistance Gene

Conventional culture-based diagnostics of orthopaedic-implant-associated infections (OIAIs) are arduous. Hence, the aim of this study was to evaluate a culture-independent, rapid nanopore-based diagnostic protocol with regard to (a) pathogen identification, (b) time to pathogen identification, and (c) identification of antimicrobial resistance (AMR). This prospective proof-of-concept study included soft tissue biopsies from 32 patients with OIAIs undergoing first revision surgery at Akershus University Hospital, Norway. The biopsies were divided into two segments. Nanopore shotgun metagenomic sequencing and pathogen and antimicrobial resistance gene identification using the EPI2ME analysis platform (Oxford Nanopore Technologies) were performed on one segment. Conventional culture-based diagnostics were performed on the other. Microbial identification matched in 23/32 OIAI patients (72%). Sequencing detected additional microbes in 9/32 patients. Pathogens detected by culturing were identified by sequencing within a median of 1 h of sequencing start [range 1–18 h]. Phenotypic AMR was explained by the detection of resistance genes in 11/23 patients (48%). Diagnostics of OIAIs using shotgun metagenomics sequencing are possible within 24 h from biopsy using nanopore technology. Sequencing outperformed culturing with respect to speed and pathogen detection where pathogens were at sufficient concentration, whereas culture-based methods had an advantage at lower pathogen concentrations. Sequencing-based AMR detection may not yet be a suitable replacement for culture-based antibiotic susceptibility testing.

scholarly journals ResistoXplorer: a web-based tool for visual, statistical and exploratory data analysis of resistome data

2021 ◽  
Vol 3 (1) ◽  
Author(s):  
Achal Dhariwal ◽  
Roger Junges ◽  
Tsute Chen ◽  
Fernanda C Petersen
Keyword(s):  
Antimicrobial Resistance ◽  
Resistance Gene ◽  
High Throughput ◽  
Resistance Genes ◽  
Visual Analytics ◽  
Gene List ◽  
Metagenomic Sequencing ◽  
High Throughput Analysis ◽  
Antimicrobial Resistance Genes ◽  
Antimicrobial Resistance Gene

Abstract The study of resistomes using whole metagenomic sequencing enables high-throughput identification of resistance genes in complex microbial communities, such as the human microbiome. Over recent years, sophisticated and diverse pipelines have been established to facilitate raw data processing and annotation. Despite the progress, there are no easy-to-use tools for comprehensive visual, statistical and functional analysis of resistome data. Thus, exploration of the resulting large complex datasets remains a key bottleneck requiring robust computational resources and technical expertise, which creates a significant hurdle for advancements in the field. Here, we introduce ResistoXplorer, a user-friendly tool that integrates recent advancements in statistics and visualization, coupled with extensive functional annotations and phenotype collection, to enable high-throughput analysis of common outputs generated from metagenomic resistome studies. ResistoXplorer contains three modules—the ‘Antimicrobial Resistance Gene Table’ module offers various options for composition profiling, functional profiling and comparative analysis of resistome data; the ‘Integration’ module supports integrative exploratory analysis of resistome and microbiome abundance profiles derived from metagenomic samples; finally, the ‘Antimicrobial Resistance Gene List’ module enables users to intuitively explore the associations between antimicrobial resistance genes and the microbial hosts using network visual analytics to gain biological insights. ResistoXplorer is publicly available at http://www.resistoxplorer.no.

scholarly journals Investigation of Spaceflight Induced Changes to Astronaut Microbiomes

2021 ◽  
Vol 12 ◽  
Author(s):  
Michael D. Morrison ◽  
James B. Thissen ◽  
Fathi Karouia ◽  
Satish Mehta ◽  
Camilla Urbaniak ◽  
...  
Keyword(s):  
Antimicrobial Resistance ◽  
Resistance Gene ◽  
Relative Abundance ◽  
Human Microbiome ◽  
Metagenomic Sequencing ◽  
Beta Lactam ◽  
Gene Markers ◽  
Shotgun Metagenomic Sequencing ◽  
Antimicrobial Resistance Genes ◽  
Antimicrobial Resistance Gene

The International Space Station (ISS) is a uniquely enclosed environment that has been continuously occupied for the last two decades. Throughout its operation, protecting the health of the astronauts on-board has been a high priority. The human microbiome plays a significant role in maintaining human health, and disruptions in the microbiome have been linked to various diseases. To evaluate the effects of spaceflight on the human microbiome, body swabs and saliva samples were collected from four ISS astronauts on consecutive expeditions. Astronaut samples were analyzed using shotgun metagenomic sequencing and microarrays to characterize the microbial biodiversity before, during, and after the astronauts’ time onboard the ISS. Samples were evaluated at an individual and population level to identify changes in microbial diversity and abundance. No significant changes in the number or relative abundance of taxa were observed between collection time points when samples from all four astronauts were analyzed together. When the astronauts’ saliva samples were analyzed individually, the saliva samples of some astronauts showed significant changes in the relative abundance of taxa during and after spaceflight. The relative abundance of Prevotella in saliva samples increased during two astronauts’ time onboard the ISS while the relative abundance of other commensal taxa such as Neisseria, Rothia, and Haemophilus decreased. The abundance of some antimicrobial resistance genes within the saliva samples also showed significant changes. Most notably, elfamycin resistance gene significantly increased in all four astronauts post-flight and a CfxA6 beta-lactam marker significantly increased during spaceflight but returned to normal levels post-flight. The combination of both shotgun metagenomic sequencing and microarrays showed the benefit of both technologies in monitoring microbes on board the ISS. There were some changes in each astronaut’s microbiome during spaceflight, but these changes were not universal for all four astronauts. Two antimicrobial resistance gene markers did show a significant change in abundance in the saliva samples of all four astronauts across their collection times. These results provide insight for future ISS microbial monitoring studies and targets for antimicrobial resistance screenings.

scholarly journals Methicillin-Resistant Staphylococcus aureus Clonal Complex 398 as a Major MRSA Lineage in Dogs and Cats in Thailand

Antibiotics ◽  
2021 ◽  
Vol 10 (3) ◽  
pp. 243
Author(s):  
Surawit Chueahiran ◽  
Jitrapa Yindee ◽  
Pongthai Boonkham ◽  
Nipattra Suanpairintr ◽  
Pattrarat Chanchaithong
Keyword(s):  
Staphylococcus Aureus ◽  
Antimicrobial Resistance ◽  
Resistance Genes ◽  
Methicillin Resistant Staphylococcus Aureus ◽  
Clonal Complex ◽  
Clinical Samples ◽  
Methicillin Resistant ◽  
Class A ◽  
Antimicrobial Resistance Genes ◽  
Mrsa St398

The aim of this study was to present molecular and antimicrobial resistance characteristics of methicillin-resistant Staphylococcus aureus (MRSA) clonal complex (CC) 398 isolated from diseased dogs and cats in Thailand. A total of 20 MRSA isolates of 134 Staphylococcus aureus isolated from canine and feline clinical samples during 2017–2020 were CC398, consisting of sequence type (ST) 398 (18 isolates), ST5926 (1 isolate), and ST6563 (1 isolate) by multilocus sequence typing. spa t034 and staphylococcal cassette chromosome mec (SCCmec) V were predominantly associated with ST398. Intraclonal differentiation was present by additional spa (t1255, t4653), non-detectable spa, composite SCCmec with a hybrid of ccrA1B1+ccrC and class A mec complex, and DNA fingerprints by pulsed-field gel electrophoresis. The isolates essentially carried antimicrobial resistance genes, mediating multiple resistance to β-lactams (mecA, blaZ), tetracyclines [tet(M)], aminoglycosides [aac(6′)-Ie-aph(2′)-Ia], and trimethoprim (dfr). Livestock-associated MRSA ST398 resistance genes including lnu(B), lsa(E), spw, fexA, and tet(L) were heterogeneously found and lost in subpopulation, with the absence or presence of additional erm(A), erm(B), and ileS2 genes that corresponded to resistance phenotypes. As only a single CC398 was detected with the presence of intraclonal variation, CC398 seems to be the successful MRSA clone colonizing in small animals as a pet-associated MRSA in Thailand.

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