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R C Matthews

Publications and source records attributed to R C Matthews.

At least 19 recordsLinked to original sources
PubMed2005

Differential growth of epidemic methicillin-resistant Staphylococcus aureus in vancomycin.

This study examines the ability of isolates representing the 17 epidemic methicillin-resistant strains of Staphylococcus aureus to grow in increasing levels of vancomycin. Only EMRSAs 1, 2, 8, 11, 12 and 15 showed any growth and were designated EMRSAs 1A, 2A, 8A, 11A, 12A and 15A. On population analysis, these strains all produced clones that grew on 32 microg/mL vancomycin, while EMRSA 12A and 15A grew at 128 microg/mL. This was associated with increased resistance to lysostaphin and teicoplanin, a loss of phage sensitivity and an increase in cell wall diameter. Typing by pulsed-field gel electrophoresis following SmaI digestion showed no change in EMRSA 8A and 15A, while the other EMRSAs all lost or gained at least one band. EMRSAs 1A, 2A and 15A became more resistant to methicillin, and EMRSAs 8A, 11A and 12A became less resistant to methicillin. These results suggest that more than one mechanism is responsible for this phenomenon.

Anti-Bacterial Agents↗
PubMed2005

Human recombinant antibody to HSP90: a natural partner in combination therapy.

Recent years have seen the development of the concept of combination therapy for treating severe fungal sepsis. The advantages of this approach are a potential improvement in patient survival and a reduction in the chance of resistance developing to each of the single agents. The disadvantage is that combining drugs may increase the chance of toxicity. Mycograb is a genetically recombinant antibody against fungal heat shock protein 90 (hsp90) which is poised to become the mainstay of combination therapy. This paper presents data on how hsp90 is important to fungi and what role it might play in human disease with possible interactions with interleukin 6 and nitric oxide. There is discussion of preclinical data demonstrating synergy in vitro between Mycograb and amphotericin B and caspofungin. The progress of Mycograb through a Phase II pharmacokinetic study when used in escalating doses with a liposomal amphotericin B preparation has also been reviewed. The concepts behind a Phase II pivotal study, where Mycograb or a placebo was given in combination with a liposomal amphotericin B drug for five days for the treatment of disseminated candidiasis are discussed.

Amphotericin B↗
PubMed2002

Identification of an immunodominant drug efflux pump in Burkholderia cepacia.

Burkholderia cepacia, a major pathogen amongst individuals with cystic fibrosis (CF), is intrinsically resistant to most clinically available antibiotics. We report the identification of an immunodominant antigen in CF patients infected with B. cepacia, a multidrug-resistance efflux pump called BcrA. The bcrA gene encodes a 46 kDa peptide with 14 potential alpha-helices that belongs to the major facilitator superfamily of drug transporters. A recombinant Escherichia coli strain was constructed containing the bcrA gene, which resulted in a four-fold increase in resistance to tetracycline and an eight-fold increase in resistance to nalidixic acid. These results demonstrate that the bcrA gene is part of a drug efflux system that is potentially a major contributor to the high-level antibiotic resistance observed in B. cepacia and thus a potential target for novel therapeutics.

ATP-Binding Cassette Transporters↗
PubMed2001

Quantification of Escherichia coli genomic DNA contamination in recombinant protein preparations by polymerase chain reaction and affinity-based collection.

This study describes the development of a novel assay for the quantification of Escherichia coli genomic DNA contamination in recombinant protein samples. The technique is based on PCR amplification and digoxygenin labeling of the genes encoding 5S ribosomal RNA followed by affinity-based collection and detection. Samples containing 1 pg x mL(-1) of extracted E. coli genomic DNA (gDNA) could be measured using this method. Using extracted E. coli gDNA as standards, a 35-cycle PCR reaction exhibited a linear response versus template concentration between 1 pg x mL(-1) and1 ng x mL(-1) genomic DNA even when diluted in a variety of buffering conditions. Comparison of the novel assay with a traditional filter binding and hybridization technique using recombinant protein samples confirmed that the procedure was accurate and sensitive. The assay described in this report is a safer and less expensive alternative to radioactive techniques employed for DNA quantification, utilizing readily available reagents and apparatus.

Base Sequence↗
PubMed2000

Bordetella pertussis infection: pathogenesis, diagnosis, management, and the role of protective immunity.

Whooping cough is presently one of the ten most common causes of death from infectious disease worldwide. Despite a high vaccine uptake, resurgences of this disease have been observed in several countries. Virulence factors of Bordetella pertussis include agglutinogens, fimbriae, P.69/pertactin, pertussis toxin, filamentous haemagglutinin, adenylate cyclase, tracheal cytotoxin, dermonecrotic toxin, lipopolysaccharide, tracheal colonisation factor, serum resistance factor, and type III secretion. Virulence factor expression is regulated by the bvgAS locus, a two-component signal transduction system. The pathophysiologic sequence consists of attachment (fimbriae, P.69/pertactin, tracheal colonisation factor, pertussis toxin, filamentous haemagglutinin), evasion of host defence (adenylate cyclase, pertussis toxin, serum resistance factor), local effects (tracheal cytotoxin), and systemic effects (pertussis toxin). Bordetella pertussis is transmitted by respiratory droplets and causes disease only in humans. Various diagnostic methods are available, including culture, serological methods, and the polymerase chain reaction. Serotyping of isolates to detect agglutinogens 2 and 3 is useful because serotype 1,2 may be associated with higher mortality, and antibodies to these antigens (agglutinins) may be protective in both animals and humans. Immunisation using whole-cell vaccine is effective but is reactogenic. Acellular vaccines containing one to five components are being used increasingly in various countries. Protective immunity to pertussis correlates with high levels of antibody to each of pertactin, fimbriae, and pertussis toxin; however, doubt remains as to the relationship between agglutinogen 3 and fimbria 3, making results of trials investigating these virulence factors difficult to interpret.

Animals↗
PubMed2000

Analysis of antibody response to Cryptococcus neoformans in five patients with AIDS and cryptococcosis by immunoblotting.

OBJECTIVES: The serological response of five patients with AIDS and cryptococcosis to non capsular antigens from Cryptococcus neoformans var. neoformans has been investigated. METHODS: Pressates of different isolates of C. neoformans were used as antigenic preparation for immunoblotting of patient samples. RESULTS: Multiple sera and cerebrospinal fluids sequentially collected from five AIDS patients with cryptococcosis showed a wide heterogeneity in antibody response with bands at 48. 43, 38 and 26 kD being present in all clinical samples of all five patients. The variation in banding patterns of the sequential samples from three patients was correlated with a decrease of the antigen titre and with an amelioration of the cryptococcal infection. CONCLUSIONS: We identified antibodies to four immunodominant non-capsular antigens, which might represent major target molecules of the humoral response of patients with cryptococcosis.

AIDS-Related Opportunistic Infections↗
PubMed2000

Identification of an immunodominant ABC transporter in methicillin-resistant Staphylococcus aureus infections.

Immunoblotting sera from 26 patients with septicemia due to an epidemic strain of methicillin-resistant Staphylococcus aureus (EMRSA-15), 6 of whom died, revealed an immunodominant EMRSA-15 antigen at 61 kDa. There was a statistically significant correlate (P < 0.001) between survival and immunoglobulin G to the 61-kDa band. The antigen was identified by sequencing positive clones obtained by screening a genomic expression library of EMRSA-15 with pooled sera from patients taken after the septicemic episode. Eluted antibody reacted with the 61-kDa antigen on immunoblots. The amino terminus was obtained by searching the S. aureus NCTC 8325 and MRSA strain COL databases, and the whole protein was expressed in Escherichia coli TOP 10F'. The derived amino acid sequence showed homology with ABC transporters, with paired Walker A and Walker B motifs and 73% homology to YkpA from Bacillus subtilis. Epitope mapping of the derived amino acid sequence with sera from patients who had recovered from EMRSA-15 septicemia delineated seven epitopes. Three of these epitopes, represented by peptides 1 (KIKVYVGNYDFWYQS), 2 (TVIVVSHDRHFLYNNV), and 3 (TETFLRGFLGRMLFS), were synthesized and used to isolate human recombinant antibodies from a phage antibody display library. Recombinant antibodies against peptides 1 and 2 gave logarithmic reductions in organ colony counts, compared with control groups, in a mouse model of the infection. This study suggests the potential role of an ABC transporter as a target for immunotherapy.

ATP-Binding Cassette Transporters↗
PubMed2000

The efficacy of acquired humoral and cellular immunity in the prevention and therapy of experimental fungal infections.

In the past two decades, numerous studies have documented the importance of acquired immunity for host defense against invasive fungal infections. There is widespread consensus in the field of medical mycology that cellular immunity is critical for successful host defense against fungi. However, in recent years several studies have established the potential efficacy of humoral immunity in host protection against two major fungal pathogens: Candida albicans and Cryptococcus neoformans. For C. albicans, antibodies to mannan, proteases and a heat shock proteins have been associated with protection against infection. Furthermore, anti-idiotypic antibodies to antibodies recognizing killer toxin from Pichia anomala and mimicking natural anti-killer toxin receptor antibodies can protect against C. albicans and other microorganisms. For C. neoformans, antibodies to the capsular glucuronoxylomannan have been shown to mediate protection in animal models of infection. Vaccines that induce protective antibodies have been shown to protect against experimental C. albicans and C. neoformans infection. In contrast, humoral immunity has not yet been demonstrated to mediate protection against Coccidioides immitis. For C. immitis, protection against infection is thought to rely on T cell mediated immunity, and the emphasis is on identifying the antigens that stimulate protective cellular immune responses and several candidate vaccines have been identified. These results provide encouragement for the view that acquired immune responses can be mobilized for the prevention and treatment of fungal infections.

Animals↗
PubMed1999

The Bpel locus encodes type III secretion machinery in Bordetella pertussis.

Type III secretory genes(Bscl, J, K, L, N and O) have recently been identified in Bordetella bronchiseptica and shown to be under the control of the BvgAS locus. We examined a 35 616 byte DNA sequence amplified from Bordetella pertussis Tohama I for homology with known type III secretory genes in Yersinia spp. and Pseudomonas sppand a total of 20 homologous open reading frames were detected. Putative type III secretion proteins in B. pertussis were designated according to their homology with type III secretion proteins in B. bronchiseptica, Yersinia and Pseudomonas. These ORFs were arranged in two putative operons, which together we have designated as the BpeI locus. The first spans nucleotides 23385-7888 and encodes the putative proteins LcrH1, BopD, BopB, LcfH2, BscI, BscJ, BscK, BscL, BscN, BscO, BscQ, BscR, BscS, BscT, BscU, and BscC, in this order. The second spans nucleotides 23580-29863 and encodes the putative proteins LcrE, LcrD, BscD and BscF, in this order. The homology of these proteins to type III secretory proteins was B. bronchiseptica (73-99%), Yersinia spp. (17-65%), Pseudomonas spp. (18-64%). The B. pertussis proteins were similar to their homologues in B. bronchiseptica, Yersinia and Pseudomonas in terms of length, molecular weight and isoelectric point. Coiled-coil domains were detected in putative translocation proteins, BopB and BopD. BopB and BopD were similar to each other, to the RTX toxin family and to cyaA, cyaB, cyaD and cyaE. The percentage G+C content of the sequence analysed was 66.16%, which is similar to the published percentage G+C (67-70%) for the B. pertussis chromosome.

Amino Acid Sequence↗
PubMed1999

Semiquantitative polymerase chain reaction enzyme immunoassay for the diagnosis of pertussis.

The two most commonly used targets for diagnosis of pertussis by the polymerase chain reaction have been the pertussis toxin promoter and the repeated insertion sequence IS481. A comparative assessment of these primers was performed on routinely collected nasopharyngeal swabs, stored at -20 C, using novel semiquantitative enzyme immunoassays. Both sets of primers behaved similarly with bacterial suspensions, and the 17 culture-positive nasopharyngeal swabs were also positive with the pertussis toxin promoter primers, with one exception, which had been subject to prolonged storage. Significantly more of the 69 culture-negative swabs were positive with the pertussis toxin promoter primers (n = 36) than with the IS481 primers (n = 18). To determine the effect of inhibitors, a comparative assessment of three primer pairs against human DNA (beta-globin and glyceraldehyde-3-phosphate dehydrogenase) was also performed.

Child↗
PubMed1999

Bordetella pertussis surveillance in England and Wales: 1995-7.

Available data sources on disease due to Bordetella pertussis, including notifications, hospital admissions, deaths, and an enhanced laboratory-based surveillance system commenced in January 1994, were reviewed for the period 1995-7. Pertussis notifications continued their approximately 3-year cycle although at historically reduced levels. A slight seasonal increase in late summer/early autumn existed over and above a relatively constant background rate. Over time, the proportion of pertussis cases in younger, unvaccinated children, and to a lesser extent, adolescents and young adults, is increasing. There is a continuing significant and underreported mortality associated with pertussis in the very young age group. Disease due to serotype 1,2 is on the increase despite persistent high vaccination levels and this serotype causes more severe disease. The provision of preventative antibiotics prior to disease onset reduced the severity of the disease but its use remains uncommon in England and Wales. While overall levels of pertussis notifications have declined in recent times, vaccination efficacy wanes with increasing age, and pertussis remains a significant cause of mortality and severe morbidity in the very young. This could be reduced by timely booster vaccination and increased recognition of mild disease in older cases followed by early antibiotic therapy for the very young household contacts.

Adolescent↗
PubMed1999

A polymerase chain reaction enzyme immunoassay for diagnosing infection caused by Aspergillus fumigatus.

AIM: To develop a polymerase chain reaction enzyme immunoassay (PCR-EIA) to measure levels of circulating aspergillus DNA in invasive aspergillosis caused by Aspergillus fumigatus. METHODS: The PCR reaction was based on primers from the 18s rRNA gene. Binding of the product to a streptavidin coated microtitration plate was mediated by a biotinylated capture probe. The product was digoxigenylated during PCR and this was the tag to which antibody was bound in the subsequent EIA. RESULTS: The optical density (OD) endpoint was < 0.1 in 10 sera from neutropenic patients with no evidence of invasive aspergillosis, and in 10 sera from nonneutropenic patients with bacterial pneumonia (group 1). The OD from five of 12 patients with allergic bronchopulmonary aspergillosis (ABPA) (group 2), three with an aspergilloma (group 3), and five with possible invasive aspergillosis (group 4) was > or = 0.1. In 63 sera from 33 cases of proven invasive aspergillosis (group 5) an OD > or = 0.1 was achieved in 48 sera from 30 patients. The maximum OD was 0.510. The level fell in survivors and gradually rose in fatal cases. CONCLUSIONS: This assay validated the concept of diagnosing invasive aspergillosis by measuring levels of circulating fungal DNA in serum.

Aspergillosis↗
PubMed1998

Stress proteins in fungal diseases.

Heat shock proteins (hsps) are ubiquitous families of proteins, found in all organisms studied so far. They are highly conserved across the species barrier and serve fundamental functions in cell physiology. The term 'heat shock' was adopted because of the early observation of the heat-inducible nature of these proteins, although, as it is now realized that they can be induced by a variety of stressful stimuli, it is probably more appropriate to call them 'stress proteins'. The nomenclature of many hsps, for example hsp90, hsp70 and hsp60, reflects the approximate molecular mass of hsps within each of these families. For many bacterial and parasitic infections, hsps were first recognized as immunodominant antigens on immunoblots of extracts from the organism probed with immune sera, or in T-cell proliferation assays. They have now been identified in a range of fungal pathogens, again often linked to an immune response. In this symposium, we review the association of hsps with humoral immunity to candidosis and aspergillosis, cellular immunity to histoplasmosis, and the identification of hsp70 in another dimorphic fungus, Paracoccidioides brasiliensis. Finally, the crucial role of the membrane in setting the temperature of the heat shock response in yeasts is discussed.

Animals↗
PubMed1997

Semiquantitative polymerase chain reaction enzyme immunoassay for diagnosis of disseminated candidiasis.

A polymerase chain reaction enzyme immunoassay (PCR-EIA) was developed for the semiquantitative of circulating candidal DNA in disseminated candidiasis due to Candida albicans. Polymerase chain reaction was based on primers from the internal transcribed ribosomal region. Binding of the product to a streptavidin-coated microtitration plate was mediated by a biotinylated capture probe. The product was digoxigenylated during PCR; this was the tag to which antibody was bound in the subsequent EIA. The optical density (OD) endpoint was < 0.1 in 15 sera from patients with no evidence of candidal infection (group 1) and in 13 of the 16 sera from colonized patients (group 2); it was > 0.1 in the other three sera from group 2 blood culture-negative patients who required intravenous amphotericin B for cure. The OD was positive in 28 patients with disseminated candidiasis (group 3), defined as positive blood cultures and successful treatment with amphotericin B (n = 11), positive blood culture confirmed at autopsy (n = 11), or negative blood culture first proven at necropsy (n = 6). In patients from whom multiple samples were available, recovery correlated with an optical density of < 0.1 by day 4 in four patients and by day 13 in the rest. In the five patients with fatal outcome from whom multiple samples were available, the mean OD rose from 0.174 to 0.668. Samples seeded with Candida albicans blastoconidia demonstrated that on OD of 0.220 was equivalent to 10 cfu. Assay of the group 3 sera by a commercial antigen detection test gave a corresponding sensitivity of 60% which rose to 67.9% when an in-house reverse passive latex agglutination test was used.

Base Sequence↗
PubMed1997

Epitope mapping Candida albicans proteinase (SAP 2).

The continuous epitopes of Candida albicans proteinase SAP 2 were derived by epitope mapping with sera from patients with oral candidiasis (n = 3), necropsy-proven disseminated candidiasis (n = 5), paired sera from patients who had recovered from blood culture-proven disseminated candidiasis (n = 3) and infection due to Candida parapsilosis (n = 2) and Candida tropicalis (n = 2). In C. albicans infection, IgM identified epitopes in amino acid positions 57-61 (QAVPV), 146-151 (SQGTLY) and 346-351 (PYDKCQ) and IgG at position 386-390 (VKYTS). For C. tropicalis IgM and IgG were positive for the same epitopes whilst IgG also detected epitopes at 78-83 (SNNQKL) and 159-164 (GVSIKN). For C. parapsilosis, IgM was positive for SNNQKL and IgG detected no epitopes. Reactivity of two of the epitopes as peptides KTSKRQAVPVTL and SLAQVKYTSASSI was confirmed in an indirect ELISA. At a cut-off optical density of 0.4, IgM against either peptide was associated with survival but present in only about half of the sera (n = 60) from patients who recovered from disseminated candidiasis whilst IgG levels were disappointing. Human recombinant antibodies from a patient who had recovered from disseminated candidiasis against either of these peptides had no activity in a lethal mouse model of candidal infection.

Animals↗