Successful identification of pathogens by polymerase chain reaction (PCR)-based electron spray ionization time-of-flight mass spectrometry (ESI-TOF-MS) in culture-negative periprosthetic joint infection.

BACKGROUND: The diagnosis of periprosthetic joint infection poses many challenges, one of which is the difficulty of isolating the infecting organism. Recently, a sophisticated modality (the Ibis Biosciences T5000 biosensor system) has been introduced that uses pan-domain primers in a series of polymerase chain reactions (PCRs) to identify and speciate essentially all bacteria and fungi as well as to identify key antibiotic resistance genes. We investigated the role of the Ibis in identifying infecting organisms in cases of known and suspected periprosthetic joint infection. METHODS: Synovial fluid specimens were collected prospectively from eighty-two patients undergoing eighty-seven arthroplasty procedures (sixty-five knee revisions, fifteen hip revisions, and seven primary knee arthroplasties) and were sent for both conventional culture and Ibis analysis. The surgeon\u27s clinical determination of the cause for revision arthroplasty was failure due to infection in twenty-three cases and noninfectious failure in fifty-seven cases. RESULTS: In the twenty-three cases that were considered on clinical grounds to involve a periprosthetic joint infection, the Ibis detected the same pathogen isolated by conventional culture in seventeen of eighteen cases and also detected one or more organisms in four of the five culture-negative cases. In addition, the Ibis detected organisms in fifty (88%) of the fifty-seven cases in which revision arthroplasty was performed for a presumed noninfectious failure. CONCLUSIONS: The Ibis technology was not only effective at detecting organisms in cases of suspected periprosthetic joint infection in which cultures were negative, but it also suggested that many of the revision arthroplasty cases that have previously been considered to be purely aseptic may have a component of unrecognized, subclinical infection

TRY plant trait database – enhanced coverage and open access

Plant traits - the morphological, anatomical, physiological, biochemical and phenological characteristics of plants - determine how plants respond to environmental factors, affect other trophic levels, and influence ecosystem properties and their benefits and detriments to people. Plant trait data thus represent the basis for a vast area of research spanning from evolutionary biology, community and functional ecology, to biodiversity conservation, ecosystem and landscape management, restoration, biogeography and earth system modelling. Since its foundation in 2007, the TRY database of plant traits has grown continuously. It now provides unprecedented data coverage under an open access data policy and is the main plant trait database used by the research community worldwide. Increasingly, the TRY database also supports new frontiers of trait‐based plant research, including the identification of data gaps and the subsequent mobilization or measurement of new data. To support this development, in this article we evaluate the extent of the trait data compiled in TRY and analyse emerging patterns of data coverage and representativeness. Best species coverage is achieved for categorical traits - almost complete coverage for ‘plant growth form’. However, most traits relevant for ecology and vegetation modelling are characterized by continuous intraspecific variation and trait–environmental relationships. These traits have to be measured on individual plants in their respective environment. Despite unprecedented data coverage, we observe a humbling lack of completeness and representativeness of these continuous traits in many aspects. We, therefore, conclude that reducing data gaps and biases in the TRY database remains a key challenge and requires a coordinated approach to data mobilization and trait measurements. This can only be achieved in collaboration with other initiatives

Direct demonstration of bacterial biofilms on prosthetic mesh after ventral herniorrhaphy

Background: Prosthetic mesh is employed routinely in the treatment of ventral and parastomal hernias, but its use can lead to major complications, including infection, extrusion, and fistula. Bacterial biofilms have been posited to play a role in mesh-related infection, but although bacteria have been noted to form biofilms on mesh surfaces in vitro, they have never been visualized directly in biofilms on mesh recovered from patients experiencing infectious complications.Methods: Five patients who developed complications after ventral hernia repair with prosthetic mesh were operated on again. Explanted mesh was examined for biofilm with confocal laser scanning microscopy (CLSM) and fluorescence in situ hybridization (FISH). In two cases, a novel molecular assay (the Ibis T5000) was used to characterize the biofilm-forming bacteria.Results: The CLSM examination demonstrated adherent biofilms on mesh surfaces in all five patients. Biofilms also were noted on investing fibrous tissue. The FISH study was able to discriminate between bacterial species in polymicrobial biofilms. In two patients the Ibis T5000 detected more species of constituent biofilm bacteria than did standard culture. Removal of the mesh and reconstruction with autologous tissues or biologic materials resolved the presenting complaints in all cases.Conclusion: Bacterial biofilms should be considered an important contributor to the pathology and complications associated with prosthetic mesh implanted in the abdominal wall. If biofilms are present, complete removal of the mesh and repair of the resulting defect without alloplastic materials is an effective intervention<br/

Resistance of Synthetic and Biologic Surgical Meshes to Methicillin-Resistant Staphylococcus aureus Biofilm: An In Vitro Investigation

Surgical meshes have become the standard procedure for a variety of surgical applications with 20 million meshes being implanted each year. The popularity of mesh usage among surgeons is backed by the multiple studies that support its functionality as a tool for improving surgical outcomes. However, their use has also been associated with infectious surgical complications and many surgeons have turned to biologic meshes. While there have been several studies investigating synthetic meshes, there is limited data comparing synthetic and biologic meshes in vitro in an infection model. This study evaluates the in vitro susceptibility of both synthetic and biologic meshes to single-species methicillin-resistant Staphylococcus aureus (MRSA) biofilms. This research compares biofilm biomass, average thickness, and coverage between the three meshes through florescent in situ hybridization (FISH), confocal scanning microscopy (CSLM), and image analysis. We also report the varying levels of planktonic and attached bacteria through sonication and cfu counts. While the data illustrates increased biofilm formation on biologic mesh in vitro, the study must further be investigated in vivo to confirm the study observations

[In Press] Trait filtering in island floras : a conceptual framework

Aim: Dispersal and environmental filtering processes affect plant species colonisation success on islands and can be identified by functional traits. However, the lack of synthesis about the different methodological approaches in functional ecology hampers generalisation of filtering processes across island systems. Location: Seventy islands of the Houtman Abrolhos archipelago, Western Australia. Major Taxa Studied: Angiosperms. Methods: We (i) apply a simple, conceptual framework based on the mean and variability of individual functional traits in plant assemblages to identify species filters on islands, (ii) illustrate how trait distributions of island assemblages change in relation to island area and their source pool, (iii) compare distributions of individual traits to multivariate functional diversity indices and trait spaces and (iv) provide guidelines to detect a signal of trait filtering in island floras. Results: The island assemblages showed evidence for selective filters operating on seed mass and marginally on leaf area but not on plant height. Mean and variability of seed mass differed to those of the source pool indicating selective forces operating between source pool and island assemblages, especially on smaller islands. Multivariate functional diversity indices and trait spaces failed to reveal filtering processes acting on the island assemblages and insights into the putative processes. Main Conclusions: Using the mean and variability of individual traits in plant assemblages provides direct information on the trait composition of island floras and the processes involved beyond what can be inferred from multivariate functional diversity indices or trait spaces. We used islands as their distinct boundaries and relatively simple sets of species provide good research models, but joint analyses of trait means and variability should also be applicable to understand filtering processes in isolates and habitat fragments on mainlands

Demonstration of bacillus cereus in orthopaedic-implant-related infection with use of a multi-primer polymerase chain reaction-mass spectrometric assay: report of two cases

The recent advent of a novel coupled PCR-mass spectrometric technology, the Ibis T5000(Abbott Laboratories, Chicago, Illinois) offers multiple potential advantages for molecular detection of periprosthetic joint infection or other implant-related infections. The Ibis assay simultaneously tests for &gt;3,000 species, including virtually all known pathogens, in a multiplex fashion, eliminating the need for an a priori choice regarding the likely infecting organism. For known pathogens, the Ibis is capable of yielding information down to the species level and can simultaneously assay for antibiotic resistance markers. It is semiquantitative and provides for a rapid turnaround, with clinically useful results available in as little as six hours. Although the Ibis T5000 is not yet approved by the Food and Drug Administration (FDA) for use in clinical diagnostics, we have used it to investigate clinical samples from multiple sources

Characterization of a mixed MRSA/MRSE biofilm in an explanted total ankle arthroplasty

Bacterial biofilms have been observed in many prosthesis-related infections, and this mode-of-growth renders the infection both difficult to treat, and especially difficult to detect and diagnose by standard culture methods. We (1) tested a novel coupled PCR-mass spectroscopic assay (the Ibis T5000) on an ankle arthroplasty that was culture negative on pre-operative aspiration, and then (2) confirmed that the Ibis assay had in fact detected viable multi-species biofilm by further micrographic and molecular examinations, including confocal microscopy using Live/Dead stain, bacterial fluorescent in situ hybridization (FISH), and reverse-transciptase-PCR (RT-PCR) assay for bacterial messenger RNA. The Ibis technology detected Staphylococcus aureus, Staphylococcus epidermidis, and the methicillin resistance gene mecA in soft tissues associated with the explanted hardware. Viable S. aureus were confirmed using RT-PCR, and viable cocci in biofilm configuration were detected microscopically on both tissue and hardware. Species-specific bacterial FISH confirmed a poly-microbial biofilm containing S. aureus. A novel culture method recovered S. aureus and S. epidermidis (both methicillin-resistant) from the tibial metal component. These observations suggest that molecular methods, particularly the new Ibis methodology, may be a useful adjunct to routine cultures in the detection of biofilm bacteria in prosthetic joint infectio

Comparing culture and molecular methods for the identification of microorganisms involved in necrotizing soft tissue infections

Abstract Background Necrotizing soft tissue infections (NSTIs) are a group of infections affecting all soft tissues. NSTI involves necrosis of the afflicted tissue and is potentially life threatening due to major and rapid destruction of tissue, which often leads to septic shock and organ failure. The gold standard for identification of pathogens is culture; however molecular methods for identification of microorganisms may provide a more rapid result and may be able to identify additional microorganisms that are not detected by culture. Methods In this study, tissue samples (n = 20) obtained after debridement of 10 patients with NSTI were analyzed by standard culture, fluorescence in situ hybridization (FISH) and multiple molecular methods. The molecular methods included analysis of microbial diversity by 1) direct 16S and D2LSU rRNA gene Microseq 2) construction of near full-length 16S rRNA gene clone libraries with subsequent Sanger sequencing for most samples, 3) the Ibis T5000 biosensor and 4) 454-based pyrosequencing. Furthermore, quantitative PCR (qPCR) was used to verify and determine the relative abundance of Streptococcus pyogenes in samples. Results For 70 % of the surgical samples it was possible to identify microorganisms by culture. Some samples did not result in growth (presumably due to administration of antimicrobial therapy prior to sampling). The molecular methods identified microorganisms in 90 % of the samples, and frequently detected additional microorganisms when compared to culture. Although the molecular methods generally gave concordant results, our results indicate that Microseq may misidentify or overlook microorganisms that can be detected by other molecular methods. Half of the patients were found to be infected with S. pyogenes, but several atypical findings were also made including infection by a) Acinetobacter baumannii, b) Streptococcus pneumoniae, and c) fungi, mycoplasma and Fusobacterium necrophorum. Conclusion The study emphasizes that many pathogens can be involved in NSTIs, and that no specific “NSTI causing” combination of species exists. This means that clinicians should be prepared to diagnose and treat any combination of microbial pathogens. Some of the tested molecular methods offer a faster turnaround time combined with a high specificity, which makes supplemental use of such methods attractive for identification of microorganisms, especially for fulminant life-threatening infections such as NSTI