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Biomedical subjects

M D Curran

Publications and source records attributed to M D Curran.

At least 19 recordsLinked to original sources
PubMed2005

Molecular diagnosis of Fusobacterium necrophorum infection (Lemierre's syndrome).

Presented here is the case of a 27-year-old male with atypical features of Lemierre's syndrome in which a definitive diagnosis was achieved using molecular methods. While routine investigations, including bacterial cultures, were unhelpful, two real-time PCR assays demonstrated Fusobacterium necrophorum-specific DNA in aspirates from brain and renal abscesses. This is the first report demonstrating that a laboratory diagnosis can be made using molecular methods in suspected cases of Lemierre's syndrome. Use of these methods can thus resolve diagnostic confusion, prevent unnecessary investigation, and direct specific antimicrobial treatment.

Adult↗
PubMed2004

Real-time PCR investigation into the importance of Fusobacterium necrophorum as a cause of acute pharyngitis in general practice.

Fusobacterium necrophorum is recognized as the cause of a severe life-threatening illness characterized by bacteraemia with metastatic abscesses following an acute sore throat (Lemierre's disease). However, the importance of F. necrophorum as a cause of simple sore throat in the community is unknown. Using quantitative real-time PCR with primers targeting the rpoB gene, 100 routine throat swabs collected from patients presenting to general practitioners with pharyngitis were analysed for the presence of F. necrophorum-specific DNA. The results were compared with those obtained from throat swabs collected from 100 healthy subjects. Ten clinical samples were positive for F. necrophorum DNA, identified as F. necrophorum subspecies funduliforme, using a haemagglutinin-related protein gene-specific PCR assay. All the healthy controls were negative (two-tailed P value = 0.0015; Fisher exact test). These findings suggest that F. necrophorum may play a more important role as a cause of simple sore throat in the community than has been previously appreciated.

Acute Disease↗
PubMed2002

A common KIR2DS4 deletion variant in the human that predicts a soluble KIR molecule analogous to the KIR1D molecule observed in the rhesus monkey.

A KIR2DS4 deletion variant allele, previously identified through KIR PCR-SSOP typing studies, was characterized, alongside a normal KIR2DS4 allele, by cDNA cloning and sequencing and its prevalence in the population determined using a deletion specific probe. The KIR2DS4 deletion variant was found in 72 of the 90 individuals screened and differed from the normal KIR2DS4 sequence by a single 22 bp deletion in exon 5. The deletion causes a frameshift predicting a truncated KIR2DS4 protein with a significantly altered D2 domain that would be secreted due to the loss of the transmembrane/cytoplasmic domains. Parallels with a recent study in the rhesus monkey highlighting access to the same open reading frame as the deletion variant, also predicting a soluble KIR molecule, are drawn.

Amino Acid Sequence↗
PubMed2002

Natural killer cell immunoglobulin-like receptor (KIR) locus profiles in African and South Asian populations.

Natural killer (NK) and some T cells express killer cell immunoglobulin-like receptors (KIRs), which interact with HLA class I expressed by target cells and consequently regulate cytolytic activity. The number of KIR loci can vary and so a range of genetic profiles is observed. We have determined the KIR genetic profiles from one African (n = 62) and two South Asian (n = 108, n = 78) populations. Several of the KIRs are present at significantly different frequencies between the two major ethnic groups (eg KIR2DS4 gene frequency 0.82 African, 0.47 S Asian. Pc < 1 x 10(-6)) and this is due to uneven distribution of two KIR haplotype families 'A' and 'B'. All three populations described here displayed a greater degree of diversity of KIR genetic profiles than other populations investigated, which indicates further complexity of underlying haplotypes; in this respect we describe two individuals who appear homozygous for a large deletion including the previously ubiquitous 2DL4. We have also reanalysed three populations that we studied previously, for the presence of a KIR which is now known to be an indicator of the 'B' haplotype. South Asians had the highest overall frequencies of all KIR loci characteristic of 'B' haplotypes (Pc < 0.0001 to < 0.004). Furthermore, gene frequency independent deviances in the linkage disequilibrium were apparent between populations.

Africa, Western↗
PubMed2001

Association between polymorphism in regulatory region of gene encoding tumour necrosis factor alpha and risk of Alzheimer's disease and vascular dementia: a case-control study.

BACKGROUND: Deposition of beta-amyloid in the brains of patients with Alzheimer's disease is thought to precede a chain of events that leads to an inflammatory response by the brain. We postulated that genetic variation in the regulatory region of the gene for the proinflammatory cytokine tumour necrosis factor alpha (TNF-alpha) leads to increased risk of Alzheimer's disease and vascular dementia. METHODS: A polymorphism in the regulatory region of the TNF-alpha gene was analysed in a case-control study. The polymorphism (C-850T) was typed in 242 patients with sporadic Alzheimer's disease, 81 patients with vascular dementia, 61 stroke patients without dementia, and 235 normal controls. These groups of individuals were also genotyped for the apolipoprotein E polymorphism, and the vascular dementia and stroke groups were typed at the HLA-DR locus. FINDINGS: The distribution of TNF-alpha genotypes in the vascular dementia group differed significantly from that in the stroke and normal control groups, giving an odds ratio of 2.51 (95% CI 1.49-4.21) for the development of vascular dementia for individuals with a CT or TT genotype. Logistic regression analysis indicated that the possession of the T allele significantly increased the risk of Alzheimer's disease associated with carriage of the apolipoprotein E epsilon4 allele (odds ratio 2.73 [1.68-4.44] for those with apolipoprotein E epsilon4 but no TNF-alpha T, vs 4.62 [2.38-8.96] for those with apolipoprotein E epsilon4 and TNF-alpha T; p=0.03). INTERPRETATION: Possession of the TNF-alpha T allele significantly increases the risk of vascular dementia, and increases the risk of Alzheimer's disease associated with apolipoprotein E. Although further research is needed, these findings suggest a potential role for anti-inflammatory therapy in vascular dementia and Alzheimer's disease, and perhaps especially in patients who have had a stroke.

Aged↗
PubMed2001

Mitochondrial DNA polymorphism: its role in longevity of the Irish population.

The mtDNA genome has been implicated as playing a pivotal role in determining the longevity and success of the human lifespan. A PCR-RFLP methodology was used to identify polymorphic restriction enzyme sites within a 2643 bp region of the mtDNA genome and a table of genetic haplotypes for a healthy aged and a younger control cohort of patients was constructed. Forty-six different mtDNA haplotypes and 11 groups of related haplotypes were identified across the two age groups but statistical analysis failed to show any significant associations. The European J haplogroup, previously reported to be associated with longevity, was not found at an increased frequency within the Irish aged population (P=0.36). However, the haplotypes comprising the J haplogroup could be differentiated into two distinct branches by the presence or absence of the two polymorphic restriction sites, 16,389g and 16,000g. The branch of haplotypes defined by 16,389g displayed a significant increased frequency in the aged samples (8%) compared to the controls (1%), P=0.015. Inversely, the branch of haplotypes defined by 16,000g displayed a significant decreased frequency in the aged samples (4%) compared to the controls (13%), P=0.011. The polymorphism (mt5178A) associated with longevity in the Japanese was not found in the Irish population, while the polymorphism (mt9055A) associated with successful ageing in the French centenarians was found at an increased frequency in the Irish aged population (9%) compared to the younger control group (5%), but failed to reach a level of statistical significance, P=0.164.

Adult↗
PubMed2000

Parvovirus B19 in kidney transplant patients.

Renal transplant patients were screened for the presence of parvovirus B19, before transplantation and monthly for 4 months after transplantation, by means of a sensitive nested PCR assay. Upon screening plasma from 110 patients, we found that two asymptomatic patients were B19 DNA positive. One of these patients was PCR positive in the plasma sample taken 2 months after transplantation; the plasma contained anti-B19 IgG antibodies before transplant and throughout the follow-up period, with an increase in the IgG level in the second posttransplant sample coinciding with the detection of B19 DNA. IgM antibodies to B19 were not detected in this patient. Because, for this patient, the donor's spleen DNA was also B19 DNA positive, we suspect B19 transmission from the donor and limited B19 replication, inasmuch as this patient already had a primed immune response to B19. The other patient was PCR positive in the pretransplant and in the plasma sample taken 1 month after transplant and contained a strong anti-B19 IgG response in the pretransplant sample and throughout the follow-up period-and anti-B19 IgM antibodies were not detected before or after transplantation. By testing samples taken from this patient at 2 weeks, 2 months, and 3 months before transplantation, we were able to determine that the infection occurred shortly before transplantation. Unexpectedly, this graft failed and was removed 2 days after transplantation despite a negative cross-match. A pathological examination of the kidney indicated acute vascular rejection, suggesting a possible role for B19 in this complication.

Antibodies, Viral↗
PubMed2000

Effect of a dCTP:dTTP pool imbalance on DNA replication fidelity in Friend murine erythroleukemia cells.

Nucleotide pool imbalances have been reported to affect the fidelity of DNA replication and repair in prokaryotic and eukaryotic cells. We have reported previously that the mutagen-hypersensitive thymidine kinase (TK)-deficient Friend erythroleukemia (FEL) cells (subclones 707BUF and 707BUE), have a more than sixfold increase in the dCTP:dTTP pool ratio when compared to that of wild-type, TK-positive (TK(+)) clone 707 cells. In this study we present the results of an investigation of the effect of the dCTP:dTTP pool imbalance on the accuracy of DNA replication within 707BUF cells. We examined the spontaneous mutation spectra occurring at the adenine phosphoribosyltransferase (aprt) locus within clone 707 (TK(+)) and 707BUF (TK(-)) FEL cells. Mutations recovered at the aprt locus in FEL cells comprised: base substitutions (43:73), frameshifts (14:13.5), and deletions (43:13.5) [clone 707 (TK(+)):707BUF (TK(-)), respectively, expressed as percentages]. A comparison of the mutation spectra obtained for the two cell lines did not reveal any significant increase in misincorporation of dCTP, the nucleotide in excess, in 707BUF (TK(-)) cells, during DNA replication synthesis. These data suggest that the dCTP:dTTP pool imbalance does not alter the fidelity of DNA replication synthesis in 707BUF (TK(-)) FEL cells. Rather, the predominance of GC --> AT transitions (53%) in the 707BUF (TK(-)) spectrum may reflect a reduced efficiency of repair by uracil DNA glycosylase of uracil residues within these cells.

Adenine Phosphoribosyltransferase↗
PubMed2000

Development of a PCR-SSOP approach capable of defining the natural killer cell inhibitory receptor (KIR) gene sequence repertoires.

A molecular typing method based on polymerase chain reaction (PCR) amplification of three different target domains (immunoglobulin domains 1 and 3, and the transmembrane-cytoplasmic domain), followed by hybridisation with 26 digoxigenin-labelled sequence-specific oligonucleotide probes (SSOP) has been established for the polymorphic killer inhibitory receptor (KIR) genes. In addition to identifying the 12 KIR subfamilies, our PCR-SSOP typing approach could also distinguish the putative alleles, NKB1 and NKAT3, that comprise the KIR3DL1 subfamily. Ninety unrelated blood donors and 13 families (52 individuals), including both parents, were subjected to our KIR PCR-SSOP typing approach. All 12 KIR subfamilies, including a 2DS5 variant sequence, were present in the 90 individuals and displayed varied phenotype frequencies: 2DL1 (0.96), 2DL2 (0.31), 2DL3 (0.95), 2DS1 (0.56) 2DS2 (0.51), 2DS3 (0.27), 2DS4 (0.96), 2DS5v (0.35), 3DS1 (0.47), 3DL1 (0.96), 3DL2 (1.0) and 2DL4 (1.0). A total of 23 different KIR phenotypes were defined in this study, and 10 of these were only found on one occasion in one individual, indicating considerable diversity in the KIR phenotype profiles within the Irish population. Most individuals (93%) possessed the complement of inhibitory KIR specificities for the three well-defined HLA-B and -C ligands. An unusual probe pattern for 3DS1 was observed in 3 individuals indicating a variant 3DS1 gene sequence with changes at nucleotide positions 1185-1186, within the cytoplasmic domain. Sequencing analysis revealed a new single nucleotide polymorphism in exon 3 of 3DL1 NKB1(195, G-A) and a 22-bp deletion polymorphism in exon 5 of 2DS4 (nucleotides 777-798 deleted). A number of strong KIR associations were observed, namely 2DL1 with 2DL3, 2DS4 with 3DL1, 2DL2 with 2DS1/2DS2/2DS3, 2DS1 with 2DS3/2DS5v/3DS1, 2DS2 with 2DS3 and 2DS5v with 3DS1. Analysis of the KIR segregation observed in the 13 families confirmed these strong associations and permitted the definition of a number of partial KIR haplotypes, e.g. 2DL2-2DS1-2DS2-2DS3-3DL1. The segregation analysis concluded that at least 3 distinct gene loci encode 2DL1-4 and at least 4 gene loci encode the non-inhibitory KIR2DS1-2DS5. In the case of 3DL1-2 and 3DS1, our data suggests 3 gene loci, one for each subfamily.

Humans↗
PubMed1999

Risk for Alzheimer's disease in older late-onset cases is associated with HLA-DRB1*03.

The allele frequency of the HLA-DRB1 gene was compared between groups of 48 clinically diagnosed elderly Alzheimer's disease (AD) cases and 44 pathologically confirmed elderly control cases. Specific primers were used to PCR amplify the highly polymorphic second exon of HLA-DRB1 using DNA extracted from blood samples or frozen brain tissue. The allele type was identified using sequence specific oligonucleotide probes. The results showed an increased frequency of DRB1*03 (P < 0.006) and decreased frequency of DRB1*09 (P < 0.049) in the AD cases compared with the controls. The results suggest that DRB1*03 is associated with an increased risk and DRB1*09 a possible decreased risk for the development of late-onset AD with first detectable clinical symptoms occurring at age 75 years or greater.

Age of Onset↗
PubMed1999

Characterization of kinetoplast DNA minicircles of an Indian isolate of Leishmania donovani.

Characterization of the kinetoplast DNA minicircles in a human pathogenic Indian isolate of Leishmania donovani has not been reported previously. Using inverse PCR, we constructed a library of PCR-amplified minicircle variable region from the kinetoplast DNA of this isolate. A combination of restriction enzyme digestion and nucleotide sequence analysis revealed five minicircle DNA sequence classes within the library, one of which was predominant, representing 75% of the kDNA network. Another distinct sequence class represented 15% of the minicircle network. Other minor sequence classes collectively constituted the remaining 10% of the network. Apart from generating basic information on the organisation and distribution of the different sequence classes within the minicircles, the DNA sequence analysis also revealed unique attributes to our minicircles. One was the surprising homology of our isolate (an Old World sp.) with distantly related New World Leishmania species. Secondly, open reading frames were also identified, indicating the possibility that these minicircles may have more than a structural role to play within the kinetoplast network.

Amino Acid Sequence↗
PubMed1999

Frequency of HLA-A and B alleles in early and late-onset Alzheimer's disease.

The frequency of various allele types of the class I Major Histocompatibility Complex (MHC) genes HLA-A and HLA-B were compared between pathologically confirmed groups of late and early-onset Alzheimer's disease (AD) and a control group. DNA was extracted from frozen brain tissue and the highly polymorphic second and third exons of the HLA-A and HLA-B genes were independently PCR amplified using specific primers. Individual allele types were identified using sequence-specific oligonucleotide probes. The results showed that the main frequency differences occurred between the late-onset AD and the control group however none of these reached statistical significance.

Adult↗
PubMed1999

Implications of endotracheal tube biofilm for ventilator-associated pneumonia.

OBJECTIVE: To determine the relationship between, and antibiotic resistance of, endotracheal tube (ET) biofilm and pulmonary pathogens in ventilator-associated pneumonia (VAP). SETTING: General intensive care units in two university teaching hospitals. DESIGN: The microbiology of ET biofilm and tracheal samples from patients with and without VAP were compared. For individual patients, matching pairs of pathogens were confirmed as identical and characterised for antibiotic susceptibility. PATIENTS: 40 intensive care unit patients - 20 with VAP, 20 without VAP as control. The duration of intubation (median and range) was 6.5 days (3-17) and 5 days (2-10), respectively. MEASUREMENTS AND RESULTS: Samples of tracheal secretions were taken during ventilation for bacteriological culture. Following extubation, ETs were examined for the presence of biofilm. Isolates of high pathogenic potential included Staphylococcus aureus, enterococci, Enterobacteriaceae, pseudomonads and Candida spp. Where the same microorganism was found on tracheal and ET samples by phenotyping, these were confirmed as identical by genotyping and characterised for antibiotic susceptibility in both the free floating and biofilm forms. Seventy per cent of patients with VAP had identical pathogens isolated from both ET biofilm and tracheal secretions. No pairing of pathogens was observed in control patients (p < 0.005). Susceptibility data for these pairs show that the ET acts as a reservoir for infecting microorganisms which exhibit significantly greater antibiotic resistance than their tracheal counterparts. CONCLUSION: This investigation provides further evidence for the role of ET biofilm in VAP. The difficulty in eradicating an established microbial biofilm using antibiotics implies that increased attention must be directed towards modification of the ET to prevent or substantially reduce biofilm formation.

Adolescent↗