Biomedical subjects

E Ingham

Publications and source records attributed to E Ingham.

At least 37 records · Page 2Linked to original sources

PubMed2002

Alumina-alumina artificial hip joints. Part II: characterisation of the wear debris from in vitro hip joint simulations.

Until recently it was not possible to reproduce clinically relevant wear rates and wear patterns in in vitro hip joint simulators for alumina ceramic-on-ceramic hip prostheses. The introduction of microseparation of the prosthesis components into in vitro wear simulations produced clinically relevant wear rates and wear patterns for the first time. The aim of this study was to characterise the wear particles generated from standard simulator testing and microseparation simulator testing of hot isostatically pressed (HIPed) and non-HIPed alumina ceramic-on-ceramic hip prostheses, and compare these particles to those generated in vivo. Standard simulation conditions produced wear rates of approximately 0.1 mm3 per million cycles for both material types. No change in surface roughness was detected and very few wear features were observed. In contrast, when microseparation was introduced into the wear simulation, wear rates of between 1.24 (HIPed) and 1.74 mm3 per million cycles (non-HIPed) were produced. Surface roughness increased and a wear stripe often observed clinically on retrieved femoral heads was also reproduced. Under standard simulation conditions only nanometre-sized wear particles (2-27.5 nm) were observed by TEM, and it was thought likely that these particles resulted from relief polishing of the alumina ceramic. However, when microseparation of the prosthesis components was introduced into the simulation, a bi-modal distribution of particle sizes was observed. The nanometre-sized particles produced by relief polishing were present (1-35nm). however, larger micrometre-sized particles were also observed by both transmission electron microscopy (TEM) (0.021 microm) and scanning electron microscopy (SEM) (0.05-->10 microm). These larger particles were thought to originate from the wear stripe and were produced by trans-granular fracture of the alumina ceramic. In Part I of this study, alumina ceramic wear particles were isolated from the periprosthetic tissues from around Mittelmeier ceramic-on-ceramic hip prostheses. Characterisation of the particles by TEM and SEM revealed a bi-modal size distribution. SEM analysis revealed particles in the 0.05-3.2 microm size range. and TEM revealed particles in the 5-90 nm size range, indicating that microseparation of the prosthesis components may be a common event in vivo. This study (Part II) has revealed that the introduction of microseparation of the prosthesis components during the swing phase of the wear simulation reproduced clinically relevant wear rates, wear patterns and wear particles in in vitro hip joint simulators.

Aluminum Oxide↗

PubMed2002

Propionibacterium acnes and inflammation in acne; P. acnes has T-cell mitogenic activity.

BACKGROUND: Circumstantial evidence suggests that Propionibacterium acnes has a role in the inflammation of acne. This could be effected by antigenic or superantigenic or mitogenic reactions. OBJECTIVES: The purpose of this investigation was to determine whether P. acnes had only antigenic activity or additional superantigenic and mitogenic activity. METHODS: A lymphocyte transformation assay was used to detect responses to a mixture of eight P. acnes whole cell isolates, and their supernatant culture fluids. In order to determine the nature of T-cell reactions to P. acnes cells a mouse-antihuman major histocompatibility complex class II monoclonal antibody was used in the lymphocyte transformation assay to inhibit the antigenic stimulation of lymphocytes. An analysis of the T-cell receptor (TCR) variable region beta (BV) repertoire was undertaken using flow cytometry of the unstimulated and stimulated cells. RESULTS: Peripheral blood mononuclear cells (PBMNC) from adults with no history of acne responded strongly to stationary growth phase cells of P. acnes, less strongly to cells in the exponential growth phase. No response was detected to supernatant culture fluids. PBMNC from five cord blood samples (CBMNC) responded maximally after 3 and 7 days of incubation with stationary growth phase cells of P. acnes. The reaction of CBMNC to P. acnes cells was not suppressed completely by the blocking antibody. The analysis of the TCRBV repertoire indicated that P. acnes induced no deletion or over-representation of certain BV element-bearing T cells. The TCRBV analysis was repeated after preincubation with the blocking antibody. Deletion of T cells bearing certain BV components occurred and there was no over-representation of T cells carrying certain BV components. CONCLUSIONS: Two mechanisms of lymphocyte activation by P. acnes cells are proposed, antigen and mitogen driven. These results are consistent with the histological evidence of inflammation in acne lesions.

Acne Vulgaris↗

PubMed2002

Differential invasion of respiratory epithelial cells by members of the Burkholderia cepacia complex.

To investigate whether there are differences between members of the Burkholderia cepacia complex in their ability to invade human respiratory epithelial cells, 11 strains belonging to genomovars I-V were studied in an antibiotic protection assay using the A549 cell line. Strains belonging to genomovars II and III were more invasive than those of genomovars I, IV and V. There was also intra-genomovar variation in invasiveness. No correlation between invasiveness and other putative virulence factors of importance in B. cepacia infection in individuals with cystic fibrosis, cable pilus and B. cepacia epidemic strain marker was identified.

Burkholderia cepacia↗

PubMed2002

Investigation of wear of knee prostheses in a new displacement/force-controlled simulator.

The performance of two knee simulators designed by ProSim (Manchester, UK) was evaluated by comparison of the wear seen in the press-fit condylar (PFC) Sigma (DePuy) knee prosthesis. Twelve specimens of the same design and manufacturing specification, were subjected to a wear test of 2 x 10(6) cycles duration using bovine serum as a lubricant. The anterior/posterior displacement and internal/external rotation inputs were based on the kinematics of the natural knee. International Standards Organization (ISO) standards were used for the flexion and axial load. The wear rates and wear scar areas were compared across all stations. The mean wear rates found were 17.6+/-5 mm3/10(6) cycles for stations 1 to 6 and 19.6+/-4 mm3/10(6) cycles for stations 7 to 12, resulting in an overall mean wear rate of 18.1+/-3 mm3/10(6) cycles. The differences between the two simulators were not significant. The average wear scar area seen on inserts from stations I to 6 was calculated at 32.4+/-1 per cent of the intended articulating surface. Similarly on stations 7 to 12 the average wear scar area was 30.7+/-3 per cent. The wear scars seen were a good physiological representation of those found from clinical explant data. This study has shown good repeatability from the simulator, both within and between the simulators.

Arthroplasty, Replacement, Knee↗

PubMed2002

Comparison of wear, wear debris and functional biological activity of moderately crosslinked and non-crosslinked polyethylenes in hip prostheses.

The wear, wear debris and functional biological activity of non-crosslinked and moderately crosslinked ultrahigh molecular weight polyethylene (UHMWPE) acetabular cups have been com pared when articulating against smooth and intentionally scratched femoral heads. Volumetric wear rates were determined in a hip joint simulator and the debris was isolated from the lubricant and characterized by the percentage number and volumetric concentration as a function of particle size. The volumetric concentration was integrated with the biological activity function determined from in vitro cell culture studies to predict an index of specific biological activity (SBA). The product of specific biological activity and volumetric wear rate was used to determine the index of functional biological activity (FBA). On smooth femoral heads the crosslinked UHMWPE had a 30 per cent lower wear rate, but it had a greater percentage volume of smaller, more biologically active particles, which resulted in a similar index of FBA compared with the non-crosslinked material. On the scratched femoral heads the volumetric wear rate was three times higher for the moderately crosslinked UHMWPE and two times higher for the non-crosslinked UHMWPE compared with the smooth femoral heads. This resulted in a higher wear rate for the moderately crosslinked material on the scratched femoral heads. All the differences in wear rate were statistically significant. There were only small differences in particle volume concentration distributions, and this resulted in similar indices of FBA which were approximately twice the values of those found on the smooth femoral heads. Both materials showed lower wear and FBA than for previously studied aged and oxidized UHMWPE gamma irradiated in air. However, this study did not reveal any advantage in terms of predicted FBA for moderately crosslinked UHMWPE compared with non-crosslinked UHMWPE.

Acetylene↗

PubMed2002

An in vitro study of the reduction in wear of metal-on-metal hip prostheses using surface-engineered femoral heads.

Although the wear of existing metal-on-metal (MOM) hip prostheses (1 mm3/10(6) cycles) is much lower than the more widely used polyethylene-on-metal bearings, there are concerns about the toxicity of metal wear particles and elevated metal ion levels, both locally and systemically, in the human body. The aim of this study was to investigate the possibility of reducing the volume of wear, the concentration of metal debris and the level of metal ion release through using surface-engineered femoral heads. Three thick (8-12 microm) coatings (TiN, CrN and CrCN) and one thin (2 microm) coating (diamond-like carbon, DLC), were evaluated on the femoral heads when articulating against high carbon content cobalt-chromium alloy acetabular inserts (HC CoCrMo) and compared with a clinically used MOM cobalt-chromium alloy bearing couple using a physiological anatomical hip joint simulator (Leeds Mark II). This study showed that CrN, CrCN and DLC coatings produced substantially lower wear volumes for both the coated femoral heads and the HC CoCrMo inserts. The TiN coating itself had little wear, but it caused relatively high wear of the HC CoCrMo inserts compared with the other coatings. The majority of the wear debris for all half-coated couples comprised small, 30 nm or less, CoCrMo metal particles. The Co, Cr and Mo ion concentrations released from the bearing couples of CrN-, CrCN- and DLC-coated heads articulating against HC CoCrMo inserts were at least 7 times lower than those released from the clinical MOM prostheses. These surface-engineered femoral heads articulating on HC CoCrMo acetabular inserts produced significantly lower wear volumes and rates, and hence lower volumetric concentrations of wear particles, compared with the clinical MOM prosthesis. The substantially lower ion concentration released by these surface-engineered components provides important evidence to support the clinical application of this technology.

Carbon↗

PubMed2002

Quantitative analysis of UHMWPE wear debris isolated from the periprosthetic femoral tissues from a series of Charnley total hip arthroplasties.

Submicrometer and micrometer-sized UHMWPE wear particles have been associated with osteolysis and failure of total hip replacements. A previous study by Tipper et al. examined the wear debris isolated from the acetabular periprosthetic tissues from 18 Charnley total hip replacements, and compared this data to the tribological variables of the prostheses. The present study aimed to isolate the UHMWPE wear debris from the femoral periprosthetic tissues from 10 of the same cohort of patients, and compare it with the debris isolated from the corresponding acetabular tissues. A variety of particle morphologies were observed, discrete submicrometer particles, along with flakes and fibrils. The particle size distributions ranged from 0.1 to >250 microm, however, the largest particles were only found in samples when the femoral head damage was characterised as low (R(pm) < 0.2 microm). The mode of the frequency distribution of particles was in the range of 0.1-0.5 microm for all the femoral tissues. Considerable variations were found in the mass distributions of the wear particles as a function of size for different patients. The net mass of debris isolated from the femoral tissues was significantly lower (p < 0.05, Student's t-test) than from the corresponding acetabular tissues. This along with considerable spatial variation in the net mass of debris isolated from the different regions of the same sample of acetabular tissue, indicates that the transportation of the debris has a marked effect on the net mass of debris accumulated in different tissues.

Acetabulum↗

PubMed2001

Quantitative characterization of polyethylene debris isolated from periprosthetic tissue in early failure knee implants and early and late failure Charnley hip implants.

This study isolated and characterized UHMWPE particles from 3 explant groups: early Charnley hip failures (ECE; < 10 years), late Charnley hip failures (LCE; > 10 years) and early knee failures (EKE; < 10 years). Debris isolated from the 3 groups had percentage particle number and percentage volumetric concentration distributions that were not significantly different. The greatest number of particles were found in the 0.1-0.5 microm size range and 19-20.6% of the volumetric concentration was below 1 microm in size in all groups. However, there were significant differences in the total volumetric concentration of debris isolated per g of tissue. LCE had significantly higher volumes of debris than ECE and EKE, there was no significant difference in the volume of debris from the EKE and ECE. The mean aspect ratio and mean irregularity ratio of the LCE group were also significantly higher than the ECE and EKE, suggesting that different wear mechanisms were occurring in the late Charnley group compared to the early Charnley and knee groups. These results also suggest that early knees, with normal surface wear, may have similar wear mechanisms to early Charnley hips and indicate that similar volumes of biologically active micrometer and sub-micrometer UHMWPE particles were produced. This may have important implications in the longer-term outcome of total knee arthroplasties, because it indicates a similar potential for osteolysis induced by wear debris.

Aged↗

PubMed2001

Invasion of human type II pneumocytes by Burkholderia cepacia.

Burkholderia cepacia is known to invade and survive within respiratory epithelial cells. Previous studies have employed transformed cell lines and it is not known whether the bacterium is capable of manifesting the same phenomena in primary cell culture. Two strains of B. cepacia of environmental (NCTC 10661) and clinical origin (C1359) were examined for their ability to invade and survive (over a 24 h period) within type II pneumocytes in primary culture using a gentamicin-ceftazidime antibiotic protection assay. Both strains of B. cepacia were capable of invasion of type II pneumocytes in primary culture. Strain C1359 was capable of multiplying intracellularly as indicated by a seven-fold increase in the numbers of bacteria from 4-24 h, whereas strain 10661, although unable to replicate intracellularly, was found to survive in the pneumocytes for at least 24 h. Future studies on the invasiveness of B. cepacia can employ A549 cells as a valid surrogate for primary cell culture assays which are time-consuming, labour-intensive and expensive to perform.

Adult↗

PubMed2001

A novel low wearing differential hardness, ceramic-on-metal hip joint prosthesis.

Osteolysis and loosening of artificial joints caused by polyethylene wear debris has prompted renewed interest in alternative bearing materials for hip prosthesis designs. Lower wearing metal-on-metal (MOM) and ceramic-on-ceramic prostheses are being used more extensively, and there is considerable interest in further improving on their performance. This study investigated the wear properties and debris morphology of a novel differential hardness ceramic-on-metal (COM) prosthesis, in comparison with MOM articulations in a physiological anatomical hip joint simulator. The COM pairings were found to have wear rates approximately 100-fold lower than the MOM pairings. The MOM pairings showed a higher "bedding in" wear rate (3.09+/-0.46mm(3)/10(6) cycles) in the first million cycles, which then reduced to a steady state wear rate of 1.23+/-0.5mm(3)/10(6) cycles. The wear rate of the COM pairings over the duration of the test was approximately 0.01mm(3)/10(6) cycles with very little wear detected on the surface of the prosthesis components. The wear particles from both articulations were oval to round in shape and in the nanometer size range. After one million cycles the mean maximum diameter of the MOM and COM wear particles were 30+/-2.25 and 17.57+/-1.37nm, respectively. After five million cycles the wear particles were statistically significantly smaller than at one million cycles, 13.9+/-0.72nm for the MOM pairings and 6.11+/-0.40nm for the COM pairings. The wear rates of the MOM prostheses were representative of clinical values. The use of differential hardness COM pairings dramatically reduced the wear rate compared to MOM hip prostheses. The wear particles from the MOM articulation were similar to particles found in retrieved tissues from around MOM prostheses. The extremely low wearing differential hardness COM bearings presented in this study produced far smaller volumetric particle loads compared to MOM prostheses currently used clinically.

Biocompatible Materials↗

PubMed2001

An evaluation of the capacity of differently prepared demineralised bone matrices (DBM) and toxic residuals of ethylene oxide (EtOx) to provoke an inflammatory response in vitro.

Demineralised bone matrix (DBM) is a form of allogeneic tissue graft widely used in oral and maxillofacial procedures. There is a long history of controversy relating to the suitability of ethylene oxide gas (EtOx) as a terminal sterilisation agent for this graft, relating to its effects on the clinical performance of the grafts. Furthermore, the generation of a toxic residual chemical (ethylene chlorohydrin, ECl) during the ethylene oxide sterilisation of patellar tendon allografts has been implicated in the failure of these grafts owing to the induction of a localised inflammatory response. In this study we have investigated the capacity of a range of different DBM preparations, and ECl dilutions, to induce the production of three pro-inflammatory cytokines, interleukin-6 (IL-6), interleukin-1beta (IL-1beta), and tumour necrosis factor alpha (TNF-alpha) from human peripheral blood mononuclear cells (PBMNCs). The levels of EtOx and ECl in EtOx terminally sterilised DBM and mineralised bone grafts were measured by gas chromatography. It was found that the only factor capable of rendering DBM pro-inflammatory was the presence of small (<20 micrometre diameter) DBM particles. No other processing or sterilisation technique resulted in the DBM becoming pro-inflammatory. Although it was also found that DBM, when EtOx-sterilised, retained more ECI than mineralised bone grafts following a standard EtOx sterilisation protocol, ECl did not provoke an inflammatory response in vitro at levels up to and including those which are cytotoxic to PBMNCs.

Bone Demineralization Technique↗

PubMed2001

Wear of HIPed and non-HIPed alumina-alumina hip joints under standard and severe simulator testing conditions.

Wear and the biological response to wear debris of artificial joints remain major concerns in total hip arthroplasty (THA). The long-term effects of UHMWPE wear debris are well documented and these have led to interest in alternate bearing materials for THA. Alumina ceramic-ceramic hip joints have been successfully used for more than 30 years with low wear and little incidence of osteolysis. The most common wear pattern observed on retrieved components is an elliptical wear 'stripe' on the heads and a corresponding worn area on the cup with an approximated wear rate of 1-5 mm3 pa. More severe wear has also occasionally occurred, usually in association with an abnormal clinical history. Modern alumina-alumina THAs use an improved HIPed (hot isostatically pressed) alumina ceramic-bearing material which may be more resistant to severe wear. Previous in vitro simulator studies have not replicated in vivo wear rates or mechanisms. The aim of this study was to compare previous generation non-HIPed alumina and modern HIPed alumina in a hip joint simulator under 'normal' and 'harsh' testing conditions. HIPed alumina was found to have a lower wear rate than non-HIPed alumina, although the difference was not statistically significant at the 95% confidence level. Testing in Gelofusine and water lubricants did not elevate the wear rates of either material. Elevated swing phase load testing also had no significant effect on the wear rates of either material. Testing in the absence of any lubricant produced very severe wear of the non-HIPed material in one specimen only.

Aluminum Oxide↗

PubMed2001

Comparison of the response of three human monocytic cell lines to challenge with polyethylene particles of known size and dose.

The response of three human monocytic cell lines (Monomac-1, U937 and THP-1) to challenge with polyethylene particles of known size and dose was evaluated. Particles with a mean size of 0.21, 0.49, 4.3, 7.2, and 88 microm were co-cultured with the cells for 24 hours prior to the assessment of cell viability and production of the pro-inflammatory cytokines, IL-1beta, IL-6 and TNFalpha. Additionally, GM-CSF and prostaglandin E2 were measured in culture supernatants from particle stimulated U937 cells. All particle fractions were evaluated at particle volume (microm3) to cell number ratios of 100 : 1, 10 : 1, 1 : 1 and 0.1 : 1. None of the test fractions had any effect on cell viability. Only the response of the U937 cell line was demonstrated to be comparable to that of primary macrophages as determined in a previous study. Furthermore only particle volume to cell number ratios of 10 : 1 or greater consistently stimulated significantly enhanced levels of cytokine secretion with particles within the phagocytosable size range (0.1 to 10 microm) being the most biologically active. No response was observed when U937 macrophages were stimulated with the largest (88 microm) particles at any of the volume ratios tested in this study. These results suggest that the size and volume of polyethylene particles are critical factors in macrophage activation. In addition, the U937 cell line has been shown to be a suitable model for the in vitro study of macrophage-particle interactions.

Journal Article↗

PubMed2001

The influence of acetabular cup angle on the wear of "BIOLOX Forte" alumina ceramic bearing couples in a hip joint simulator.

The wear of "Biolox Forte" alumina ceramic bearing couples has been investigated at two different acetabular cup angles in a physiological hip joint simulator. All cups were set in the anatomical position of 45 degrees inclination in the M/L plane for the first two million cycles and then four of the six cups were re-aligned to 60 degrees for a further three million cycles. A "running-in" wear of 0.14 mm3 per million cycles was observed for the first million cycles, after which a steady state wear rate of 0.05 mm3 per million cycles was observed. Increasing the acetabular cup angle to 60 degrees did not significantly affect the wear rate.

Journal Article↗

PubMed2001

Cell-mediated immunity to the mycelial phase of Malassezia spp. in patients with pityriasis versicolor and controls.

BACKGROUND: Malassezia is the aetiological agent of pityriasis versicolor. The mycelial phase of the organism predominates in lesions of pityriasis versicolor. OBJECTIVES: To evaluate the cell-mediated immune (CMI) response to the mycelial phase of Malassezia in patients with this disease, which has not previously been studied. METHODS: The CMI status of 12 patients with pityriasis versicolor and 12 age- and sex-matched controls to mycelial antigen(s) of the organism was examined. The responses to the mycelial form of three strains of the organism were assessed using lymphocyte transformation and leucocyte migration inhibition assays. RESULTS: The transformation responses of the lymphocytes from both patients and controls gave transformation indices < or = 3, although the responses of lymphocytes from patients with pityriasis versicolor to the mycelial form of Malassezia strains were generally higher than those of the controls. In the leucocyte migration inhibition assay, leucocytes from patients with pityriasis versicolor and controls responded to the mycelial antigens of three different Malassezia strains; however, there was no significant difference in leucocyte response between patients with pityriasis versicolor and controls. CONCLUSIONS: Patients with pityriasis versicolor do not therefore have a CMI deficiency to Malassezia mycelial antigens but fail to generate a protective CMI response to mycelial antigens over and above that of control individuals during active disease.

Adult↗

PubMed2001

Production of the mycelial phase of Malassezia in vitro.

To study the pathogenicity of Malassezia, the agent of pityriasis versicolor, it is necessary to obtain the mycelial form in vitro. A range of different components and conditions were tested to induce yeast cells of the organism to produce mycelia in vitro using different culture media. A mycelial culture medium was developed that consisted of bacteriological peptone, glucose, yeast extract, ox bile, glycerol, glycerol monostearate, Tween 80, squalene, glycine, potassium nitrate, sodium chloride, ferrous sulphate and magnesium sulphate with or without agar. The liquid and solid medium had a pH of 5.6 and the temperature of incubation was 30 degrees C. Cultures were incubated in air. This medium was able to induce some strains of Malassezia to produce up to 40% mycelium in vitro. In total, 33 different strains of Malassezia obtained from the skin of the healthy individuals and patients with pityriasis versicolor were tested for mycelium production. The strains of Malassezia capable of producing mycelium in vitro all possessed the serovar A antigen.

Culture Media↗

PubMed2001

PerR controls oxidative stress resistance and iron storage proteins and is required for virulence in Staphylococcus aureus.

The Staphylococcus aureus genome encodes three ferric uptake regulator (Fur) homologues: Fur, PerR, and Zur. To determine the exact role of PerR, we inactivated the gene by allelic replacement using a kanamycin cassette, creating strain MJH001 (perR). PerR was found to control transcription of the genes encoding the oxidative stress resistance proteins catalase (KatA), alkyl hydroperoxide reductase (AhpCF), bacterioferritin comigratory protein (Bcp), and thioredoxin reductase (TrxB). Furthermore, PerR regulates transcription of the genes encoding the iron storage proteins ferritin (Ftn) and the ferritin-like Dps homologue, MrgA. Transcription of perR was autoregulated, and PerR repressed transcription of the iron homeostasis regulator Fur, which is a positive regulator of catalase expression. PerR functions as a manganese-dependent, transcriptional repressor of the identified regulon. Elevated iron concentrations produced induction of the PerR regulon. PerR may act as a peroxide sensor, since addition of external hydrogen peroxide to 8325-4 (wild type) resulted in increased transcription of most of the PerR regulon, except for fur and perR itself. The PerR-regulated katA gene encodes the sole catalase of S. aureus, which is an important starvation survival determinant but is surprisingly not required for pathogenicity in a murine skin abscess model of infection. In contrast, PerR is not necessary for starvation survival but is required for full virulence (P < 0.005) in this model of infection. PerR of S. aureus may act as a redox sentinel protein during infection, analogous to the in vitro activities of OxyR and PerR of Escherichia coli and Bacillus subtilis, respectively. However, it differs in its response to the metal balance within the cell and has the added capability of regulating iron uptake and storage.

Amino Acid Sequence↗

PubMed2001

In Staphylococcus aureus, fur is an interactive regulator with PerR, contributes to virulence, and Is necessary for oxidative stress resistance through positive regulation of catalase and iron homeostasis.

The Staphylococcus aureus genome encodes three ferric uptake repressor (Fur) homologues: Fur, PerR, and Zur. To determine the exact role of Fur in S. aureus, we inactivated the fur gene by allelic replacement using a tetracycline resistance cassette, creating strain MJH010 (fur). The mutant had a growth defect in rich medium, and this defect was exacerbated in metal-depleted CL medium. This growth defect was partially suppressed by manganous ion, a metal ion with known antioxidant properties. This suggests that the fur mutation leads to an oxidative stress condition. Indeed, MJH010 (fur) has reduced levels of catalase activity resulting from decreased katA transcription. Using a katA-lacZ fusion we have determined that Fur functions, either directly or indirectly, as an iron-dependent positive regulator of katA expression. Transcription of katA is coregulated by Fur and PerR, since in MJH010 (fur) transcription was still repressed by manganese while transcription in MJH201 (fur perR) was unresponsive to the presence of iron or manganese. Siderophore biosynthesis was repressed by iron in 8325-4 (wild-type) but in MJH010 (fur) was constitutive. A number of putative Fur-regulated genes were identified in the incomplete genome databases using known S. aureus Fur box sequences. Of those tested, the sstABCD and sirABC operons and the fhuD2 and orf4 genes were found to have Fur-regulated expression. MJH010 (fur) was attenuated (P<0.04) in a murine skin abscess model of infection, as was double-mutant MJH201 (fur perR) (P<0.03). This demonstrates the importance in vivo of iron homeostasis and oxidative stress resistance regulation in S. aureus.

Animals↗