K. Greenwood-Quaintance - Academia.edu (original) (raw)

Papers by K. Greenwood-Quaintance

Research paper thumbnail of Comparative evaluation of cDNA library construction approaches for RNA-Seq analysis from low RNA-content human specimens

Journal of Microbiological Methods, 2018

With the emergence of RNA sequencing technologies, metatranscriptomic studies are rapidly gaining... more With the emergence of RNA sequencing technologies, metatranscriptomic studies are rapidly gaining attention as they simultaneously provide insight into gene expression profiles and therefore disease association pathways of microbial pathogens and their hosts. This approach, therefore, holds promise for applicability in infectious disease diagnostics. A challenge of this approach in the clinical setting is the low amount and quality of RNA, especially microbial RNA in most clinically-infected specimens. Here, we compared two commercially available stranded cDNA library preparation kits, the NuGEN Ovation SoLo RNA-Seq System and the Illumina TruSeq Stranded Total RNA, using RNA extracted from synovial and sonicate fluids from a subject with periprosthetic joint infection. The Ovation SoLo RNA-Seq System provided more useful transcriptomic data for the infecting bacterium, whereas the TruSeq Stranded Total RNA kit provided more useful human transcriptomic data.

Research paper thumbnail of Ureaplasma parvum causes hyperammonemia in a pharmacologically immunocompromised murine model

European Journal of Clinical Microbiology & Infectious Diseases, 2016

A relationship between hyperammonemia and Ureaplasma infection has been shown in lung transplant ... more A relationship between hyperammonemia and Ureaplasma infection has been shown in lung transplant recipients. We have demonstrated that Ureaplasma urealyticum cases hyperammonemia in a novel immunocompromised murine model. Herein, we determined whether Ureaplasma parvum can do the same. Male C3H mice were given mycophenolate mofetil, tacrolimus and prednisone for seven days, and then challenged with U. parvum intratracheally (IT) and/or

Research paper thumbnail of Superantigens produced by catheter-associated Staphylococcus aureus elicit systemic inflammatory disease in the absence of bacteremia

Journal of Leukocyte Biology, 2015

SAgs, produced by Staphylococcus aureus, play a major role in the pathogenesis of invasive staphy... more SAgs, produced by Staphylococcus aureus, play a major role in the pathogenesis of invasive staphylococcal diseases by inducing potent activation of the immune system. However, the role of SAgs, produced by S. aureus, associated with indwelling devices or tissues, are not known. Given the prevalence of device-associated infection with toxigenic S. aureus in clinical settings and the potency of SAgs, we hypothesized that continuous exposure to SAgs produced by catheter-associated S. aureus could have systemic consequences. To investigate these effects, we established a murine in vivo catheter colonization model. One centimeter long intravenous catheters were colonized with a clinical S. aureus isolate producing SAgs or isogenic S. aureus strains, capable or incapable of producing SAg. Catheters were subcutaneously implanted in age-matched HLA-DR3, B6, and AE(o) mice lacking MHC class II molecules and euthanized 7 d later. There was no evidence of systemic infection. However, in HLA-DR3 transgenic mice, which respond robustly to SSAgs, the SSAg-producing, but not the nonproducing strains, caused a transient increase in serum cytokine levels and a protracted expansion of splenic CD4(+) T cells expressing SSAg-reactive TCR Vβ8. Lungs, livers, and kidneys from these mice showed infiltration with CD4(+) and CD11b(+) cells. These findings were absent in B6 and AE(o) mice, which are known to respond poorly to SSAgs. Overall, our novel findings suggest that systemic immune activation elicited by SAgs, produced by S. aureus colonizing foreign bodies, could have clinical consequences in humans.

Research paper thumbnail of Thymidine Auxotrophic Staphylococcus aureus Small-Colony Variant Endocarditis and Left Ventricular Assist Device Infection

Journal of Clinical Microbiology, 2011

We describe a thymidine-dependent small-colony variant of Staphylococcus aureus associated with l... more We describe a thymidine-dependent small-colony variant of Staphylococcus aureus associated with left ventricular assist device infection and prosthetic valve and pacemaker endocarditis.

Research paper thumbnail of Corynebacterium Prosthetic Joint Infection

Journal of Clinical Microbiology, 2012

Identification of Corynebacterium species may be challenging. Corynebacterium species are occasio... more Identification of Corynebacterium species may be challenging. Corynebacterium species are occasional causes of prosthetic joint infection (PJI), but few data are available on the subject. Based on the literature, C. amycolatum , C. aurimucosum , C. jeikeium , and C. striatum are the most common Corynebacterium species that cause PJI. We designed a rapid PCR assay to detect the most common human Corynebacterium species, with a specific focus on PJI. A polyphosphate kinase gene identified using whole-genome sequence was targeted. The assay differentiates the antibiotic-resistant species C. jeikeium and C. urealyticum from other species in a single assay. The assay was applied to a collection of human Corynebacterium isolates from multiple clinical sources, and clinically relevant species were detected. The assay was then tested on Corynebacterium isolates specifically associated with PJI; all were detected. We also describe the first case of C. simulans PJI.

Research paper thumbnail of Prosthetic Joint Infection Diagnosis Using Broad-Range PCR of Biofilms Dislodged from Knee and Hip Arthroplasty Surfaces Using Sonication

Journal of Clinical Microbiology, 2012

Periprosthetic tissue and/or synovial fluid PCR has been previously studied for prosthetic joint ... more Periprosthetic tissue and/or synovial fluid PCR has been previously studied for prosthetic joint infection (PJI) diagnosis; however, few studies have assessed the utility of PCR on biofilms dislodged from the surface of explanted arthroplasties using vortexing and sonication (i.e., sonicate fluid PCR). We compared sonicate fluid 16S rRNA gene real-time PCR and sequencing to culture of synovial fluid, tissue, and sonicate fluid for the microbiologic diagnosis of PJI. PCR sequences generating mixed chromatograms were decatenated using RipSeq Mixed. We studied sonicate fluids from 135 and 231 subjects with PJI and aseptic failure, respectively. Synovial fluid, tissue, and sonicate fluid culture and sonicate fluid PCR had similar sensitivities (64.7, 70.4, 72.6, and 70.4%, respectively; P > 0.05) and specificities (96.9, 98.7, 98.3, and 97.8%, respectively; P > 0.05). Combining sonicate fluid culture and PCR, the sensitivity was higher (78.5%, P < 0.05) than those of individual...

Research paper thumbnail of Detection of Prosthetic Joint Infection Using Synovial Fluid Polymerase Chain Reaction-Electrospray Ionization Mass Spectrometry

Journal of Clinical Microbiology, 2014

PCR coupled with electrospray ionization mass spectrometry applied to synovial fluid specimens ha... more PCR coupled with electrospray ionization mass spectrometry applied to synovial fluid specimens had an 81% sensitivity and a 95% specificity for the diagnosis of prosthetic joint infection.

Research paper thumbnail of Diagnosis of Prosthetic Joint Infection by Use of PCR-Electrospray Ionization Mass Spectrometry

Journal of Clinical Microbiology, 2013

We compared PCR-electrospray ionization mass spectrometry (PCR-ESI/MS) to culture using sonicate ... more We compared PCR-electrospray ionization mass spectrometry (PCR-ESI/MS) to culture using sonicate fluid from 431 subjects with explanted knee ( n = 270) or hip ( n = 161) prostheses. Of these, 152 and 279 subjects had prosthetic joint infection (PJI) and aseptic failure, respectively. The sensitivities for detecting PJI were 77.6% for PCR-ESI/MS and 69.7% for culture ( P = 0.0105). The specificities were 93.5 and 99.3%, respectively ( P = 0.0002).

Research paper thumbnail of Rapid Molecular Microbiologic Diagnosis of Prosthetic Joint Infection

Journal of Clinical Microbiology, 2013

We previously showed that culture of samples obtained by prosthesis vortexing and sonication was ... more We previously showed that culture of samples obtained by prosthesis vortexing and sonication was more sensitive than tissue culture for prosthetic joint infection (PJI) diagnosis. Despite improved sensitivity, culture-negative cases remained; furthermore, culture has a long turnaround time. We designed a genus-/group-specific rapid PCR assay panel targeting PJI bacteria and applied it to samples obtained by vortexing and sonicating explanted hip and knee prostheses, and we compared the results to those with sonicate fluid and periprosthetic tissue culture obtained at revision or resection arthroplasty. We studied 434 subjects with knee ( n = 272) or hip ( n = 162) prostheses; using a standardized definition, 144 had PJI. Sensitivities of tissue culture, of sonicate fluid culture, and of PCR were 70.1, 72.9, and 77.1%, respectively. Specificities were 97.9, 98.3, and 97.9%, respectively. Sonicate fluid PCR was more sensitive than tissue culture ( P = 0.04). PCR of prosthesis sonicati...

Research paper thumbnail of Combinations of cefoxitin plus other β-lactams are synergistic in vitro against community associated methicillin-resistant Staphylococcus aureus

European Journal of Clinical Microbiology & Infectious Diseases, 2013

In vitro studies demonstrate that oxacillin minimal inhibitory concentrations (MICs) of methicill... more In vitro studies demonstrate that oxacillin minimal inhibitory concentrations (MICs) of methicillin-resistant S. aureus (MRSA) strains USA300 and 400 decrease in the presence of cefoxitin. The aim of this study was to characterize the activity of cefoxitin plus β-lactams against a collection of MRSA isolates. We assessed the in vitro antimicrobial activity of a selection of β-lactams alone and together with subinhibitory concentrations of cefoxitin against a collection of MRSA, methicillin-susceptible S. aureus (MSSA), and vancomycin-intermediate S. aureus (VISA) isolates using MICs and time kill assays. For community-associated (CA) MRSA strains USA300 and USA400, MICs of nafcillin, cefazolin, cephalexin, cefuroxime, ceftriaxone and cefotaxime decreased by 8- to 64-times in the presence of 10 μg/ml cefoxitin. In contrast, for hospital-associated (HA) strains COLn, N315, and Mu50, there was no change in any β-lactam MIC in the presence of cefoxitin. When combined with cefoxitin, the cephalexin MIC decreased for eight CA-MRSA and five MSSA sequence types but did not change for seven HA-MRSA sequence types. β-lactam/cefoxitin combinations were synergistic against CA- but not HA-MRSA strains in time kill assays. Cefoxitin combined with a variety of β-lactams enhances their activity against CA-MRSA strains in vitro. Further studies of combination β-lactam therapy may provide insight into β-lactam biology, penicillin binding protein cooperativity, and novel therapeutic strategies against MRSA.

Research paper thumbnail of A Biofilm Approach to Detect Bacteria on Removed Spinal Implants

Spine, 2010

This is a prospective study comparing the diagnosis of spinal implant infection by conventional p... more This is a prospective study comparing the diagnosis of spinal implant infection by conventional peri-implant tissue culture with a technique which uses a combination of vortexing and bath sonication to dislodge bacteria growing as a biofilm on the surface of retrieved spinal implants. We hypothesized that the biofilm-sampling technique would be more sensitive than peri-implant tissue culture. Culture of peri-implant tissue is inaccurate for the diagnosis of orthopedic device-related infection; cultures taken from the implant may be more sensitive. We have developed a technique which uses vortexing-bath sonication to sample bacterial biofilms on the surface of retrieved hip and knee implants, and shown that it is more sensitive than peri-prosthetic tissue culture for the microbiologic diagnosis of prosthetic knee, hip, and shoulder infection. We compared peri-implant tissue culture to the vortexing-bath sonication technique which samples bacterial biofilm on the surface of retrieved spinal implants, for the diagnosis of spinal implant infection. In addition, we compared detection of Staphylococcus and Propionibacterium acnes by rapid cycle real-time polymerase chain reaction with culture of sonicate fluid. A total of 112 subjects were studied; 22 had spinal implant infection. The sensitivities of peri-implant tissue and sonicate fluid culture were 73% and 91% (P = 0.046), and the specificities were 93% and 97%, respectively. P. acnes and coagulase-negative staphylococci were the most frequent microorganisms detected among subjects with spinal implant infection, with P. acnes detected in 56 and 45%, and coagulase-negative staphylococci detected in 31 and 40% of peri-implant tissue and sonicate fluid cultures, respectively. Compared with the culture of sonicate fluid, polymerase chain reaction was 100 and 67% sensitive for the detection of culture-positive Staphylococcus and P. acnes spinal implant infection, respectively. Implant sonication followed by culture is more sensitive than peri-implant tissue culture for the microbiologic diagnosis of spinal implant infection.

Research paper thumbnail of Clinical Characteristics and Outcomes of Prosthetic Joint Infection Caused by Small Colony Variant Staphylococci

Small colony variants (SCVs) are naturally occurring subpopulations of bacteria. The clinical cha... more Small colony variants (SCVs) are naturally occurring subpopulations of bacteria. The clinical characteristics and treatment outcomes of patients with prosthetic joint infection (PJI) caused by staphylococcal SCVs are unknown. This study was a retrospective series of 113 patients with staphylococcal PJI, with prospective testing of archived sonicate fluid samples. SCVs were defined using two-investigator review. Treatment failure was defined as (i) subsequent revision surgery for any reason, (ii) PJI after the index surgery, (iii) prosthesis nonreimplantation due to ongoing infection, or (iv) amputation of the affected limb. There were 38 subjects (34%) with SCVs and 75 (66%) with only normal-phenotype (NP) bacteria. Subjects with SCVs were more likely to have been on chronic antimicrobials prior to surgery (P ‫؍‬ 0.048), have had prior surgery for PJI (P ‫؍‬ 0.03), have had a longer duration of symptoms (P ‫؍‬ 0.0003), and have had a longer time since joint implantation (P ‫؍‬ 0.007), compared to those with only NP bacteria. Over a median follow-up of 30.6 months, 9 subjects (24%) with SCVs and 23 (32%) with only NP bacteria experienced treatment failure (P ‫؍‬ 0.51). Subjects infected with Staphylococcus aureus were more likely to fail than were those infected with Staphylococcus epidermidis (hazard ratio [HR], 4.03; 95% confidence interval [CI], 1.80 to 9.04). While frequently identified in subjects with PJI and associated with several potential predisposing factors, SCVs were not associated with excess treatment failure compared to NP infections in this study, where they were primarily managed with two-stage arthroplasty exchange. IMPORTANCE Bacteria with the small colony variant (SCV) phenotype are described in small case series as causing persistent or relapsing infection, but there are insufficient data to suggest that they should be managed differently than infection with normal-phenotype bacteria. In an effort to investigate the clinical importance of this phenotype, we determined whether SCVs were present in biofilms dislodged from the surfaces of arthroplasties of patients with staphylococcal prosthetic joint infection and assessed the clinical outcomes associated with detection of SCVs. We found that prosthetic joint infection caused by SCV staphylococci was associated with a longer duration of symptoms and more prior treatment for infection but not with an increased rate of treatment failure, compared to infection caused by normal-phenotype staphylococci.

Research paper thumbnail of A Biofilm Approach to Detect Bacteria on Removed Spinal Implants

Spine, 2010

Study Design. This is a prospective study comparing the diagnosis of spinal implant infection by ... more Study Design. This is a prospective study comparing the diagnosis of spinal implant infection by conventional peri-implant tissue culture with a technique which uses a combination of vortexing and bath sonication to dislodge bacteria growing as a biofilm on the surface of retrieved spinal implants.

Research paper thumbnail of Rapid Molecular Microbiologic Diagnosis of Prosthetic Joint Infection

Journal of Clinical Microbiology, 2013

Research paper thumbnail of Comparative evaluation of cDNA library construction approaches for RNA-Seq analysis from low RNA-content human specimens

Journal of Microbiological Methods, 2018

With the emergence of RNA sequencing technologies, metatranscriptomic studies are rapidly gaining... more With the emergence of RNA sequencing technologies, metatranscriptomic studies are rapidly gaining attention as they simultaneously provide insight into gene expression profiles and therefore disease association pathways of microbial pathogens and their hosts. This approach, therefore, holds promise for applicability in infectious disease diagnostics. A challenge of this approach in the clinical setting is the low amount and quality of RNA, especially microbial RNA in most clinically-infected specimens. Here, we compared two commercially available stranded cDNA library preparation kits, the NuGEN Ovation SoLo RNA-Seq System and the Illumina TruSeq Stranded Total RNA, using RNA extracted from synovial and sonicate fluids from a subject with periprosthetic joint infection. The Ovation SoLo RNA-Seq System provided more useful transcriptomic data for the infecting bacterium, whereas the TruSeq Stranded Total RNA kit provided more useful human transcriptomic data.

Research paper thumbnail of Ureaplasma parvum causes hyperammonemia in a pharmacologically immunocompromised murine model

European Journal of Clinical Microbiology & Infectious Diseases, 2016

A relationship between hyperammonemia and Ureaplasma infection has been shown in lung transplant ... more A relationship between hyperammonemia and Ureaplasma infection has been shown in lung transplant recipients. We have demonstrated that Ureaplasma urealyticum cases hyperammonemia in a novel immunocompromised murine model. Herein, we determined whether Ureaplasma parvum can do the same. Male C3H mice were given mycophenolate mofetil, tacrolimus and prednisone for seven days, and then challenged with U. parvum intratracheally (IT) and/or

Research paper thumbnail of Superantigens produced by catheter-associated Staphylococcus aureus elicit systemic inflammatory disease in the absence of bacteremia

Journal of Leukocyte Biology, 2015

SAgs, produced by Staphylococcus aureus, play a major role in the pathogenesis of invasive staphy... more SAgs, produced by Staphylococcus aureus, play a major role in the pathogenesis of invasive staphylococcal diseases by inducing potent activation of the immune system. However, the role of SAgs, produced by S. aureus, associated with indwelling devices or tissues, are not known. Given the prevalence of device-associated infection with toxigenic S. aureus in clinical settings and the potency of SAgs, we hypothesized that continuous exposure to SAgs produced by catheter-associated S. aureus could have systemic consequences. To investigate these effects, we established a murine in vivo catheter colonization model. One centimeter long intravenous catheters were colonized with a clinical S. aureus isolate producing SAgs or isogenic S. aureus strains, capable or incapable of producing SAg. Catheters were subcutaneously implanted in age-matched HLA-DR3, B6, and AE(o) mice lacking MHC class II molecules and euthanized 7 d later. There was no evidence of systemic infection. However, in HLA-DR3 transgenic mice, which respond robustly to SSAgs, the SSAg-producing, but not the nonproducing strains, caused a transient increase in serum cytokine levels and a protracted expansion of splenic CD4(+) T cells expressing SSAg-reactive TCR Vβ8. Lungs, livers, and kidneys from these mice showed infiltration with CD4(+) and CD11b(+) cells. These findings were absent in B6 and AE(o) mice, which are known to respond poorly to SSAgs. Overall, our novel findings suggest that systemic immune activation elicited by SAgs, produced by S. aureus colonizing foreign bodies, could have clinical consequences in humans.

Research paper thumbnail of Thymidine Auxotrophic Staphylococcus aureus Small-Colony Variant Endocarditis and Left Ventricular Assist Device Infection

Journal of Clinical Microbiology, 2011

We describe a thymidine-dependent small-colony variant of Staphylococcus aureus associated with l... more We describe a thymidine-dependent small-colony variant of Staphylococcus aureus associated with left ventricular assist device infection and prosthetic valve and pacemaker endocarditis.

Research paper thumbnail of Corynebacterium Prosthetic Joint Infection

Journal of Clinical Microbiology, 2012

Identification of Corynebacterium species may be challenging. Corynebacterium species are occasio... more Identification of Corynebacterium species may be challenging. Corynebacterium species are occasional causes of prosthetic joint infection (PJI), but few data are available on the subject. Based on the literature, C. amycolatum , C. aurimucosum , C. jeikeium , and C. striatum are the most common Corynebacterium species that cause PJI. We designed a rapid PCR assay to detect the most common human Corynebacterium species, with a specific focus on PJI. A polyphosphate kinase gene identified using whole-genome sequence was targeted. The assay differentiates the antibiotic-resistant species C. jeikeium and C. urealyticum from other species in a single assay. The assay was applied to a collection of human Corynebacterium isolates from multiple clinical sources, and clinically relevant species were detected. The assay was then tested on Corynebacterium isolates specifically associated with PJI; all were detected. We also describe the first case of C. simulans PJI.

Research paper thumbnail of Prosthetic Joint Infection Diagnosis Using Broad-Range PCR of Biofilms Dislodged from Knee and Hip Arthroplasty Surfaces Using Sonication

Journal of Clinical Microbiology, 2012

Periprosthetic tissue and/or synovial fluid PCR has been previously studied for prosthetic joint ... more Periprosthetic tissue and/or synovial fluid PCR has been previously studied for prosthetic joint infection (PJI) diagnosis; however, few studies have assessed the utility of PCR on biofilms dislodged from the surface of explanted arthroplasties using vortexing and sonication (i.e., sonicate fluid PCR). We compared sonicate fluid 16S rRNA gene real-time PCR and sequencing to culture of synovial fluid, tissue, and sonicate fluid for the microbiologic diagnosis of PJI. PCR sequences generating mixed chromatograms were decatenated using RipSeq Mixed. We studied sonicate fluids from 135 and 231 subjects with PJI and aseptic failure, respectively. Synovial fluid, tissue, and sonicate fluid culture and sonicate fluid PCR had similar sensitivities (64.7, 70.4, 72.6, and 70.4%, respectively; P > 0.05) and specificities (96.9, 98.7, 98.3, and 97.8%, respectively; P > 0.05). Combining sonicate fluid culture and PCR, the sensitivity was higher (78.5%, P < 0.05) than those of individual...

Research paper thumbnail of Detection of Prosthetic Joint Infection Using Synovial Fluid Polymerase Chain Reaction-Electrospray Ionization Mass Spectrometry

Journal of Clinical Microbiology, 2014

PCR coupled with electrospray ionization mass spectrometry applied to synovial fluid specimens ha... more PCR coupled with electrospray ionization mass spectrometry applied to synovial fluid specimens had an 81% sensitivity and a 95% specificity for the diagnosis of prosthetic joint infection.

Research paper thumbnail of Diagnosis of Prosthetic Joint Infection by Use of PCR-Electrospray Ionization Mass Spectrometry

Journal of Clinical Microbiology, 2013

We compared PCR-electrospray ionization mass spectrometry (PCR-ESI/MS) to culture using sonicate ... more We compared PCR-electrospray ionization mass spectrometry (PCR-ESI/MS) to culture using sonicate fluid from 431 subjects with explanted knee ( n = 270) or hip ( n = 161) prostheses. Of these, 152 and 279 subjects had prosthetic joint infection (PJI) and aseptic failure, respectively. The sensitivities for detecting PJI were 77.6% for PCR-ESI/MS and 69.7% for culture ( P = 0.0105). The specificities were 93.5 and 99.3%, respectively ( P = 0.0002).

Research paper thumbnail of Rapid Molecular Microbiologic Diagnosis of Prosthetic Joint Infection

Journal of Clinical Microbiology, 2013

We previously showed that culture of samples obtained by prosthesis vortexing and sonication was ... more We previously showed that culture of samples obtained by prosthesis vortexing and sonication was more sensitive than tissue culture for prosthetic joint infection (PJI) diagnosis. Despite improved sensitivity, culture-negative cases remained; furthermore, culture has a long turnaround time. We designed a genus-/group-specific rapid PCR assay panel targeting PJI bacteria and applied it to samples obtained by vortexing and sonicating explanted hip and knee prostheses, and we compared the results to those with sonicate fluid and periprosthetic tissue culture obtained at revision or resection arthroplasty. We studied 434 subjects with knee ( n = 272) or hip ( n = 162) prostheses; using a standardized definition, 144 had PJI. Sensitivities of tissue culture, of sonicate fluid culture, and of PCR were 70.1, 72.9, and 77.1%, respectively. Specificities were 97.9, 98.3, and 97.9%, respectively. Sonicate fluid PCR was more sensitive than tissue culture ( P = 0.04). PCR of prosthesis sonicati...

Research paper thumbnail of Combinations of cefoxitin plus other β-lactams are synergistic in vitro against community associated methicillin-resistant Staphylococcus aureus

European Journal of Clinical Microbiology & Infectious Diseases, 2013

In vitro studies demonstrate that oxacillin minimal inhibitory concentrations (MICs) of methicill... more In vitro studies demonstrate that oxacillin minimal inhibitory concentrations (MICs) of methicillin-resistant S. aureus (MRSA) strains USA300 and 400 decrease in the presence of cefoxitin. The aim of this study was to characterize the activity of cefoxitin plus β-lactams against a collection of MRSA isolates. We assessed the in vitro antimicrobial activity of a selection of β-lactams alone and together with subinhibitory concentrations of cefoxitin against a collection of MRSA, methicillin-susceptible S. aureus (MSSA), and vancomycin-intermediate S. aureus (VISA) isolates using MICs and time kill assays. For community-associated (CA) MRSA strains USA300 and USA400, MICs of nafcillin, cefazolin, cephalexin, cefuroxime, ceftriaxone and cefotaxime decreased by 8- to 64-times in the presence of 10 μg/ml cefoxitin. In contrast, for hospital-associated (HA) strains COLn, N315, and Mu50, there was no change in any β-lactam MIC in the presence of cefoxitin. When combined with cefoxitin, the cephalexin MIC decreased for eight CA-MRSA and five MSSA sequence types but did not change for seven HA-MRSA sequence types. β-lactam/cefoxitin combinations were synergistic against CA- but not HA-MRSA strains in time kill assays. Cefoxitin combined with a variety of β-lactams enhances their activity against CA-MRSA strains in vitro. Further studies of combination β-lactam therapy may provide insight into β-lactam biology, penicillin binding protein cooperativity, and novel therapeutic strategies against MRSA.

Research paper thumbnail of A Biofilm Approach to Detect Bacteria on Removed Spinal Implants

Spine, 2010

This is a prospective study comparing the diagnosis of spinal implant infection by conventional p... more This is a prospective study comparing the diagnosis of spinal implant infection by conventional peri-implant tissue culture with a technique which uses a combination of vortexing and bath sonication to dislodge bacteria growing as a biofilm on the surface of retrieved spinal implants. We hypothesized that the biofilm-sampling technique would be more sensitive than peri-implant tissue culture. Culture of peri-implant tissue is inaccurate for the diagnosis of orthopedic device-related infection; cultures taken from the implant may be more sensitive. We have developed a technique which uses vortexing-bath sonication to sample bacterial biofilms on the surface of retrieved hip and knee implants, and shown that it is more sensitive than peri-prosthetic tissue culture for the microbiologic diagnosis of prosthetic knee, hip, and shoulder infection. We compared peri-implant tissue culture to the vortexing-bath sonication technique which samples bacterial biofilm on the surface of retrieved spinal implants, for the diagnosis of spinal implant infection. In addition, we compared detection of Staphylococcus and Propionibacterium acnes by rapid cycle real-time polymerase chain reaction with culture of sonicate fluid. A total of 112 subjects were studied; 22 had spinal implant infection. The sensitivities of peri-implant tissue and sonicate fluid culture were 73% and 91% (P = 0.046), and the specificities were 93% and 97%, respectively. P. acnes and coagulase-negative staphylococci were the most frequent microorganisms detected among subjects with spinal implant infection, with P. acnes detected in 56 and 45%, and coagulase-negative staphylococci detected in 31 and 40% of peri-implant tissue and sonicate fluid cultures, respectively. Compared with the culture of sonicate fluid, polymerase chain reaction was 100 and 67% sensitive for the detection of culture-positive Staphylococcus and P. acnes spinal implant infection, respectively. Implant sonication followed by culture is more sensitive than peri-implant tissue culture for the microbiologic diagnosis of spinal implant infection.

Research paper thumbnail of Clinical Characteristics and Outcomes of Prosthetic Joint Infection Caused by Small Colony Variant Staphylococci

Small colony variants (SCVs) are naturally occurring subpopulations of bacteria. The clinical cha... more Small colony variants (SCVs) are naturally occurring subpopulations of bacteria. The clinical characteristics and treatment outcomes of patients with prosthetic joint infection (PJI) caused by staphylococcal SCVs are unknown. This study was a retrospective series of 113 patients with staphylococcal PJI, with prospective testing of archived sonicate fluid samples. SCVs were defined using two-investigator review. Treatment failure was defined as (i) subsequent revision surgery for any reason, (ii) PJI after the index surgery, (iii) prosthesis nonreimplantation due to ongoing infection, or (iv) amputation of the affected limb. There were 38 subjects (34%) with SCVs and 75 (66%) with only normal-phenotype (NP) bacteria. Subjects with SCVs were more likely to have been on chronic antimicrobials prior to surgery (P ‫؍‬ 0.048), have had prior surgery for PJI (P ‫؍‬ 0.03), have had a longer duration of symptoms (P ‫؍‬ 0.0003), and have had a longer time since joint implantation (P ‫؍‬ 0.007), compared to those with only NP bacteria. Over a median follow-up of 30.6 months, 9 subjects (24%) with SCVs and 23 (32%) with only NP bacteria experienced treatment failure (P ‫؍‬ 0.51). Subjects infected with Staphylococcus aureus were more likely to fail than were those infected with Staphylococcus epidermidis (hazard ratio [HR], 4.03; 95% confidence interval [CI], 1.80 to 9.04). While frequently identified in subjects with PJI and associated with several potential predisposing factors, SCVs were not associated with excess treatment failure compared to NP infections in this study, where they were primarily managed with two-stage arthroplasty exchange. IMPORTANCE Bacteria with the small colony variant (SCV) phenotype are described in small case series as causing persistent or relapsing infection, but there are insufficient data to suggest that they should be managed differently than infection with normal-phenotype bacteria. In an effort to investigate the clinical importance of this phenotype, we determined whether SCVs were present in biofilms dislodged from the surfaces of arthroplasties of patients with staphylococcal prosthetic joint infection and assessed the clinical outcomes associated with detection of SCVs. We found that prosthetic joint infection caused by SCV staphylococci was associated with a longer duration of symptoms and more prior treatment for infection but not with an increased rate of treatment failure, compared to infection caused by normal-phenotype staphylococci.

Research paper thumbnail of A Biofilm Approach to Detect Bacteria on Removed Spinal Implants

Spine, 2010

Study Design. This is a prospective study comparing the diagnosis of spinal implant infection by ... more Study Design. This is a prospective study comparing the diagnosis of spinal implant infection by conventional peri-implant tissue culture with a technique which uses a combination of vortexing and bath sonication to dislodge bacteria growing as a biofilm on the surface of retrieved spinal implants.

Research paper thumbnail of Rapid Molecular Microbiologic Diagnosis of Prosthetic Joint Infection

Journal of Clinical Microbiology, 2013