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M S Phillips

Publications and source records attributed to M S Phillips.

At least 19 recordsLinked to original sources
PubMed2006

Real-time PCR detection and quantification of vector trichodorid nematodes and Tobacco rattle virus.

This report describes a novel diagnostic method for virus-vector trichodorid nematodes and associated Tobacco rattle virus (TRV) based on a real-time fluorogenic 5' nuclease PCR assay (TaqMan). Two independent primer/probe sets were designed targeting the 18S gene of the ribosomal cistron for the trichodorid species, Paratrichodorus pachydermus and Trichodorus similis. Assays using purified plasmid DNA containing clones of the 18S region and genomic DNA extracted from individuals from both nematode species displayed high specificity as no cros s-reaction was observed between the species or with two non-target trichodorid species Paratrichodorus anemones and Trichodorus primitivus. Relative quantification of target DNA present in unknown samples was performed by comparison of the fluorescence signals of the samples to those obtained from plasmid standard dilutions. Three primer/probe sets were also used to target TRV; one set for RNA1 and the two other sets for RNA2 of specific isolates (TRV-PpK20 and TRV-TpO1). Detection of both trichodorid species and TRV RNA1 and RNA2 from a single sample was achieved and field samples were used to demonstrate the potential of this assay to provide rapid, accurate and sensitive molecular information in relation to risk assessment in the field.

Analysis of Variance↗
PubMed2003

Chromosome-wide distribution of haplotype blocks and the role of recombination hot spots.

Recent studies of human populations suggest that the genome consists of chromosome segments that are ancestrally conserved ('haplotype blocks'; refs. 1-3) and have discrete boundaries defined by recombination hot spots. Using publicly available genetic markers, we have constructed a first-generation haplotype map of chromosome 19. As expected for this marker density, approximately one-third of the chromosome is encompassed within haplotype blocks. Evolutionary modeling of the data indicates that recombination hot spots are not required to explain most of the observed blocks, providing that marker ascertainment and the observed marker spacing are considered. In contrast, several long blocks are inconsistent with our evolutionary models, and different mechanisms could explain their origins.

Alleles↗
PubMed2001

Nosocomial infections due to nontuberculous mycobacteria.

Nontuberculous mycobacteria (NTM) are ubiquitous in the environment and cause colonization, infection, and pseudo-outbreaks in health care settings. Data suggest that the frequency of nosocomial outbreaks due to NTM may be increasing, and reduced hot water temperatures may be partly responsible for this phenomenon. Attention to adequate high-level disinfection of medical devices and the use of sterile reagents and biologicals will prevent most outbreaks. Because NTM cannot be eliminated from the hospital environment, and because they present an ongoing potential for infection, NTM should be considered in all cases of nosocomial infection, and careful surveillance must be used to identify potential outbreaks. Analysis of the species of NTM and the specimen source may assist in determining the significance of a cluster of isolates. Once an outbreak or pseudo-outbreak is suspected, molecular techniques should be applied promptly to determine the source and identify appropriate control measures.

Cross Infection↗
PubMed2001

Cuticle heterogeneity as exhibited by Pasteuria spore attachment is not linked to the phylogeny of parthenogenetic root-knot nematodes (Meloidogyne spp.).

The cuticle is a major barrier prohibiting the infection of nematodes against micro-organisms. The attachment of bacterial spores of the nematode hyperparasite Pasteuria penetrans (PP1) to field populations of root-knot nematodes (RKN, Meloidogyne spp.) from Burkino Faso, Ecuador, Greece, Malawi, Senegal and Trinidad and Tobago were assayed in standard attachment tests. The attachment of spore population PP1 to different field populations of root-knot nematode showed that the rates of attachment differed between countries. Similar tests were also undertaken on P. penetrans spores from these countries against 2 species of RKN, M. incognita and M. arenaria. The results showed a high degree of variability in spore attachment with no clear distinction between the 2 species of nematode. It has been hypothesized that Pasteuria spore attachment is linked to nematode species designations and this study clearly shows that this is not the case. Further tests showed that variation in spore attachment was not linked to nematode phylogeny. The results therefore beg the question of how do parthenogenetic root-knot nematodes maintain cuticle variability in the face of such an aggressive hyperparasite.

Animals↗
PubMed2001

Mapping by genetic interaction: high-resolution congenic mapping of the type 1 diabetes loci Idd10 and Idd18 in the NOD mouse.

As many of the linked chromosome regions that predispose to type 1 diabetes in the NOD mouse have been dissected, it has become apparent that the initially observed effect is in fact attributable to several loci. One such cluster of loci on distal chromosome 3, originally described as Idd10, is now known to comprise three separate loci, Idd10, Idd17, and Idd18. Although these loci have a significant combined effect on diabetes development, their individual effects are barely detectable when diabetes is used as a read-out, which makes fine-mapping them by use of a conventional congenic approach impractical. In this study, we demonstrate that it is possible to map loci, with modest effects, to regions small enough for systematic gene identification by capitalizing on the fact that the combined loci provide more profound, measurable protection. We have mapped the Idd10 and Idd18 loci to 1.3- and 2.0-cM intervals, respectively, by holding the Idd3 allele constant. In addition, we have excluded Csf1 and Nras as candidates for both loci.

Animals↗
PubMed2000

The NOD Idd9 genetic interval influences the pathogenicity of insulitis and contains molecular variants of Cd30, Tnfr2, and Cd137.

Previous analyses of NOD mice have shown that some genes control the development of both insulitis and diabetes, while other loci influence diabetes without reducing insulitis. Evidence for the existence of a gene only influencing diabetes, Idd9 on mouse chromosome 4, is provided here by the development of a novel congenic mouse strain, NOD.B10 Idd9. NOD.B10 Idd9 mice display profound resistance to diabetes even though nearly all develop insulitis. Subcongenic analysis has demonstrated that alleles of at least three B10 genes, Idd9.1, Idd9.2, and Idd9.3 are required to produce Idd9-mediated diabetes resistance. Candidate genes with amino acid differences between the NOD and B10 strains have been localized to the 5.6 cM Idd9.2 interval (Tnfr2, Cd30) and to the 2.0 cM Idd9.3 interval (Cd137).

Alleles↗
PubMed2000

A multipartite mitochondrial genome in the potato cyst nematode Globodera pallida.

The mitochondrial genome (mtDNA) of the plant parasitic nematode Globodera pallida exists as a population of small, circular DNAs that, taken individually, are of insufficient length to encode the typical metazoan mitochondrial gene complement. As far as we are aware, this unusual structural organization is unique among higher metazoans, although interesting comparisons can be made with the multipartite mitochondrial genome organizations of plants and fungi. The variation in frequency between populations displayed by some components of the mtDNA is likely to have major implications for the way in which mtDNA can be used in population and evolutionary genetic studies of G. pallida.

Animals↗
PubMed2000

Evaluation of single nucleotide polymorphism typing with invader on PCR amplicons and its automation.

Large-scale pharmacogenetics and complex disease association studies will require typing of thousands of single-nucleotide polymorphisms (SNPs) in thousands of individuals. Such projects would benefit from a genotyping system with accuracy >99% and a failure rate <5% on a simple, reliable, and flexible platform. However, such a system is not yet available for routine laboratory use. We have evaluated a modification of the previously reported Invader SNP-typing chemistry for use in a genotyping laboratory and tested its automation. The Invader technology uses a Flap Endonuclease for allele discrimination and a universal fluorescence resonance energy transfer (FRET) reporter system. Three hundred and eighty-four individuals were genotyped across a panel of 36 SNPs and one insertion/deletion polymorphism with Invader assays using PCR product as template, a total of 14,208 genotypes. An average failure rate of 2.3% was recorded, mostly associated with PCR failure, and the typing was 99.2% accurate when compared with genotypes generated with established techniques. An average signal-to-noise ratio (9:1) was obtained. The high degree of discrimination for single base changes, coupled with homogeneous format, has allowed us to deploy liquid handling robots in a 384-well microtitre plate format and an automated end-point capture of fluorescent signal. Simple semiautomated data interpretation allows the generation of approximately 25,000 genotypes per person per week, which is 10-fold greater than gel-based SNP typing and microsatellite typing in our laboratory. Savings on labor costs are considerable. We conclude that Invader chemistry using PCR products as template represents a useful technology for typing large numbers of SNPs rapidly and efficiently.

Base Sequence↗
PubMed1999

Clinical research: ASHP guidelines and future directions for pharmacists.

ASHP guidelines on the pharmacist's role in clinical drug research and future directions for pharmacists in clinical drug research are described. Health-system pharmacists have a responsibility to the patient and to the institution to ensure that clinical research systems are sound, that patients are protected, and that research is conducted in a safe, effective way. ASHP guidelines list minimum standards that are essential for improving performance. The ASHP Guidelines on Clinical Drug Research, approved in November 1997, update information previously found in the ASHP Guidelines for the Use of Investigational Drugs in Organized Health-Care Settings, approved in 1990, but have an expanded focus. Additions include recognition of relevant business practices, implications of technology, and the expansion of clinical research beyond the academic health center. At a minimum, all pharmacists involved in clinical research should handle the record keeping for drug accountability, provide drug information to patients and to other health care providers, ensure the appropriate care of patients at sites not involved in the study, and provide accountability at nonpharmacy locations. Managing and coordinating clinical drug research is an area of growth that represents a great opportunity for clinical drug research. By providing baseline and higher-level pharmaceutical services for clinical research projects, pharmacists can help to ensure data accuracy and completeness and patient safety.

Guidelines as Topic↗
PubMed1998

Cloning of a novel member of the low-density lipoprotein receptor family.

A gene encoding a novel transmembrane protein was identified by DNA sequence analysis within the insulin-dependent diabetes mellitus (IDDM) locus IDDM4 on chromosome 11q13. Based on its chromosomal position, this gene is a candidate for conferring susceptibility to diabetes. The gene, termed low-density lipoprotein receptor related protein 5 (LRP5), encodes a protein of 1615 amino acids that contains conserved modules which are characteristic of the low-density lipoprotein (LDL) receptor family. These modules include a putative signal peptide for protein export, four epidermal growth factor (EGF) repeats with associated spacer domains, three LDL-receptor (LDLR) repeats, a single transmembrane spanning domain, and a cytoplasmic domain. The encoded protein has a unique organization of EGF and LDLR repeats; therefore, LRP5 likely represents a new category of the LDLR family. Both human and mouse LRP5 cDNAs have been isolated and the encoded mature proteins are 95% identical, indicating a high degree of evolutionary conservation.

Amino Acid Sequence↗
PubMed1997

Molecular analysis of rat pituitary and hypothalamic growth hormone secretagogue receptors.

GH release is thought to occur under the reciprocal regulation of two hypothalamic peptides, GH releasing hormone (GHRH) and somatostatin, via their engagement with specific cell surface receptors on the anterior pituitary somatotroph. In addition, GH-releasing peptides, such as GHRP-6 and the nonpeptide mimetics, L-692,429 and MK-0677, stimulate GH release through their activation of a distinct receptor, the GH secretagogue receptor (GHS-R). The recent cloning of the GHS-R from human and swine pituitary gland identifies yet a third G protein-coupled receptor (GPC-R) involved in the control of GH release and further supports the existence of an undiscovered hormone that may activate this receptor. Using the human GHS-R as a probe, we report the isolation of a rat pituitary GHS-R cDNA derived from an unspliced, precursor mRNA. The rat cDNA encodes a protein of 364 amino acids containing seven transmembrane domains (7-TM) with >90% sequence identity to both the human and swine GHS-Rs. A single intron of approximately 2 kb divides the open reading frame into two exons encoding TM 1-5 and TM 6-7, thus placing the GHS-R into the intron-containing class of GPC-Rs. The intron maps to the site of sequence divergence between the human and swine type 1a and 1b GHS-R mRNAs. In addition, determination of the nucleotide sequence for the human GHS-R gene confirmed the position of an intron in the human GHS-R gene at this position. A full-length contiguous cDNA from rat hypothalamus was isolated and shown to be identical in its nucleotide and deduced amino acid sequence to the rat pituitary GHS-R. The cloned rat GHS-R binds [35S]MK-0677 with high affinity [dissociation constant (K(D)) = 0.7 nM] and is functionally active when expressed in HEK-293 cells. Expression of the rat GHS-R was observed specifically in the pituitary and hypothalamus when compared with control tissues.

Amino Acid Sequence↗
PubMed1996

The structural organization of the human skeletal muscle ryanodine receptor (RYR1) gene.

The RYR1 gene encoding the Ca2+ release channel of human skeletal muscle sarcoplasmic reticulum has been cloned and exon/intron boundaries have been determined, together with a minimum of 30 bp of intron sequence flanking each splice junction. The gene contains 106 exons, of which two are alternatively spliced. The length of the gene, determined by the alignment of 16 genomic phage clones, a cosmid clone, and several long polymerase chain reaction products, is approximately 160 kb. Exons range from 15 to 813 bp, while introns range from 85 to about 16,000 bp. Analysis of the gene has confirmed published errors in the human RYR1 cDNA and confirmed the structure of two alternatively spliced exons. The numbering of the nucleotides comprising the RYR1 cDNA and the numbering of amino acids encoded by them were corrected to account for these earlier errors and omissions. Analysis of 2.4 kb of the 5' upstream sequence indicated the presence of a CCAAT box and several Sp1 binding sites between nucleotides -200 and -60 bp, flanking the proposed transcription start site at -130 bp. Several other potential transcription factor binding sites were identified throughout the 5' sequence. Knowledge of the structure of the RYR1 gene will provide an invaluable resource for the discovery of mutations in the gene that are causal of human malignant hyperthermia and central core disease.

Amino Acid Sequence↗
PubMed1996

Comparison of the segregation of the RYR1 C1840T mutation with segregation of the caffeine/halothane contracture test results for malignant hyperthermia susceptibility in a large Manitoba Mennonite family.

BACKGROUND: Malignant hyperthermia (MH) is an important cause of anesthesia-induced death. Malignant hyperthermia susceptibility is diagnosed using the in vitro caffeine/halothane contracture test (CHCT) in fresh muscle biopsy specimens. The CHCT test is highly invasive, expensive, and lacks 100% specificity. Genetic and biochemical evidence provide strong support for the view that the substitution of cysteine for arginine 614 (Arg614Cys) in the human ryanodine receptor gene is one of several mutations that are likely to cause human MH. DNA testing was compared with CHCT as a means of predicting MH susceptibility in a large MH family in which the Arg614Cys mutation was detected. METHODS: A comparison of CHCT and DNA-based diagnosis was conducted in a large Manitoba Mennonite MH kindred identified by an index patient who died at age 45 yr of an MH crisis after general anesthesia. The presence of the Arg614Cys mutation was detected through a combination of polymerase chain reaction and restriction endonuclease digestion. Blood samples for DNA analysis were obtained from 68 family members, including 19 who had undergone muscle biopsies and 1 who had a documented crisis but did not undergo biopsy. Family members were classified as MH-susceptible or MH-normal on the basis of the CHCT. RESULTS: Twenty-two persons were found to be heterozygous for the Arg614Cys mutation. Five of these persons had prior positive CHCT results and one had an MH crisis but did not undergo biopsy. On DNA testing, 44 persons were found to be homozygous for the normal allele. Of these, ten had been classified as MH-normal and five as MH-susceptible on the basis of the CHCT. On reevaluation of the data obtained in our earlier CHCT diagnoses, we found that the condition of the muscle was poor, with no twitch, for three of five individuals homozygous for the normal allele but originally classified as MH-susceptible and for one who was homozygous for the normal allele and originally classified as MH-normal. Caffeine/halothane contracture test results for these four persons were considered invalid. The twitch response was good for the two remaining persons who were homozygous for the normal allele but classified as MH- susceptible, because contracture was observed with appropriately low levels of both caffeine and halothane. CONCLUSIONS: An absolute correlation between DNA test results and CHCT assignment could not be made in this kindred. Possible explanations for discordance are that the Arg614Cys mutation is not linked to MH, that a second MH mutation is segregating in the family, or that there are errors in the CHCT. Because there is strong evidence supporting the causal nature of the Arg614Cys mutation, the discordant persons are not closely related within the pedigree as they would be if a second MH mutation were segregating, and the CHCT is not 100% accurate, we propose that the observed discordance between DNA test results and CHCT assignment in this kindred results from two false-positive diagnoses by the CHCT.

Anesthetics, Inhalation↗
PubMed1996

Functional STAT 1 and 3 signaling by the leptin receptor (OB-R); reduced expression of the rat fatty leptin receptor in transfected cells.

The leptin receptor (OB-R) bears homology to members of the class I cytokine receptor family. We demonstrate that leptin binding to OB-R stimulates formation of STAT-1 and STAT-3 complexes, thereby defining transcriptional motifs for genes that are under leptin control. Transfected fa OB-R bound leptin with equal affinity to that of wild type OB-R. fa OB-R abundance was about 7 fold reduced compared to control cells. Surprisingly, the low level of fa OB-R is fully capable of activating the STAT signal transduction pathway. We discuss plausible explanations for the obese phenotype in Zucker fatty rats.

Animals↗
PubMed1995

Characterization of cDNA and genomic DNA encoding SERCA1, the Ca(2+)-ATPase of human fast-twitch skeletal muscle sarcoplasmic reticulum, and its elimination as a candidate gene for Brody disease.

Genomic DNA and cDNA encoding human SERCA1, the Ca(2+)-ATPase of fast-twitch skeletal muscle sarcoplasmic reticulum (the ATP2A1 gene on chromosome 16p12), were isolated and characterized. The cDNA encodes 994 amino acids. The genomic DNA is 26 kb long and contains 23 exons, one of which can be alternatively spliced. The locations of each of the exon/intron boundaries are the same as those previously identified in the rabbit ATP2A1 gene. Brody disease is an inherited disorder of skeletal muscle, characterized by exercise-induced impairment of muscle relaxation. It has been postulated to result from a deficiency in SERCA1. In a search for the genetic basis of Brody disease, the coding sequence of the ATP2A1 gene in one Brody patient and the full-length sequences of two SERCA1 cDNAs in two other, unrelated Brody patients were compared with normal ATP2A1 sequences. In all three cases, the coding and splice junction sequences were normal, indicating that the forms of Brody disease manifested in these three patients are not caused by mutations in the coding or splice junction regions of the ATP2A1 gene.

Amino Acid Sequence↗
PubMed1995

Drugs and sedation for colonoscopy.

Sedation, with or without analgesia, is commonly used for colonoscopy procedures in the United States. Prudent drug product selection, careful titration of drug dosage to ensure use of the lowest effective dose (Table 1), and vigilant monitoring of medicated patients will optimize the value of conscious sedation in colonoscopy. When a close patient-physician relationship exists in the primary care setting, use of medications only "if needed" during the procedure may be a reasonable alternative that can minimize the exposure of patients to sedation-related side effects. Patient-controlled medication delivery may be one method used to address patient variability in the need for sedation.

Adult↗