Restriction enzymes.
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DNA Restriction Enzymes↗
Biomedical subjects
E D Spitzer
Publications and source records attributed to E D Spitzer.
A laboratory medicine residency training program that includes clinical consultation and research.
We describe a laboratory medicine residency training program that includes ongoing interaction with both clinical laboratories and clinical services as well as significant research experience. Laboratory medicine residents serve as on-call consultants in the interpretation of test results, design of testing strategies, and assurance of test quality. The consultative on-call beeper system was evaluated and is presented as an effective method of clinical pathology training that is well accepted by the clinical staff. The research component of the residency program is also described. Together, these components provide training in real-time clinical problem solving and prepare residents for the changing technological environment of the clinical laboratory. At the completion of the residency, the majority of the residents are qualified laboratory subspecialists and are also capable of running an independent research program.
Chemistry, Clinical↗
Antimicrobial susceptibility patterns of Haemophilus isolates from children in eleven developing nations. BOSTID Haemophilus Susceptibility Study Group.
The antimicrobial susceptibilities of 426 isolates of Haemophilus species, which were collected as part of a worldwide study of the etiology of acute respiratory disease in children in selected developing countries, were determined. Eleven antibiotics were tested using the recently described Haemophilus Test Medium. There was a low prevalence of antibiotic resistance; 6% of strains were resistant to ampicillin, and 1.6% were resistant to chloramphenicol. Strains resistant to both ampicillin and chloramphenicol were recovered only from Thailand. Susceptibility to penicillin G was also determined; the minimum inhibitory concentrations for penicillin and ampicillin were concordant within one 2-fold dilution in 97% of the isolates. Thus, Haemophilus isolates were as susceptible to penicillin G as they were to ampicillin, and penicillin resistance was infrequent overall. These data provide support for the current protocols for the management of acute respiratory infections in children in developing countries, in which penicillin G is a first-line agent.
Anti-Bacterial Agents↗
Testing of organisms for susceptibility to triazoles: is it justified?
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Antifungal Agents↗
Use of mitochondrial and ribosomal DNA polymorphisms to classify clinical and soil isolates of Histoplasma capsulatum.
We have developed an improved scheme for the classification of environmental and clinical isolates of Histoplasma capsulatum that is based on analysis of mitochondrial DNA (mtDNA) and ribosomal DNA (rDNA). Strains were initially divided into mtDNA groups according to restriction digests of whole-cell DNA and Southern hybridization with cloned mtDNA probes. Strains within a mtDNA class could be further grouped by polymorphisms in rDNA. The majority of soil and clinical isolates from the United States had identical mtDNA patterns; however, rDNA polymorphisms were common in both types of isolates. The combination of mtDNA and rDNA typing described in this report will be useful in resolving questions concerning the epidemiology of H. capsulatum infections.
Blotting, Southern↗
DNA probe for the identification of Histoplasma capsulatum.
A 1.85-kilobase HindIII nuclear DNA probe from Histoplasma capsulatum G217B detected polymorphic restriction fragments within whole-cell DNA from different clinical isolates of H. capsulatum, consistent with the previous system of classification. The probe failed to hybridize to DNA from Blastomyces dermatitidis, Candida spp., Saccharomyces cerevisiae, Sepedonium chrysospermum, and Chrysosporium keratinophilum under low-stringency conditions and therefore may have value as a diagnostic reagent to identify H. capsulatum.
DNA Probes↗
Comparative in vitro activity of LY121019 and amphotericin B against clinical isolates of Candida species.
LY121019 and amphotericin B were equally active in vitro against most clinical isolates of Candida albicans and Candida tropicalis. Higher concentrations of LY121019 were required for inhibition of Candida glabrata. Other Candida species were inhibited by amphotericin B but not by LY121019.
Amphotericin B↗
beta-Alanine auxotrophy associated with dfp, a locus affecting DNA synthesis in Escherichia coli.
Strains containing the conditional-lethal dfp-707 mutation, which have a defect in DNA synthesis at 42 degrees C, were found to require either pantothenate or its precursor, beta-alanine, for growth at 30 degrees C. The auxotrophy and conditional lethality were corevertible. Through localized mutagenesis of the dfp-pyrE region of Escherichia coli, another mutation, dfp-1, was obtained. It conferred the auxotrophy but not the conditional lethality of dfp-707. Complementation analysis, performed with a set of plasmid-borne deletion and insertion mutations, revealed a correspondence between the complementation of each mutant phenotype and the production of the dfp gene product, previously identified as a 45-kilodalton flavoprotein. The dfp mutants had a normal level of aspartate-1-decarboxylase, which is the only enzyme known to produce beta-alanine in E. coli and which is specified by the distant panD gene. A prototrophic pseudorevertant of a dfp-1 strain was found to have retained the dfp mutation, to be genetically unstable, and to have an elevated level of aspartate-1-decarboxylase, suggesting that it had acquired a duplication of panD. It is not known what steps in pantothenate or DNA metabolism are affected by the mutant dfp product or how its flavin moiety may be involved.
Alanine↗
dfp Gene of Escherichia coli K-12, a locus affecting DNA synthesis, codes for a flavoprotein.
The cloned dfp gene complements dna-707 (now designated dfp-707), a temperature-sensitive conditionally lethal mutation that results in a slow cessation of DNA synthesis while protein synthesis is maintained. In vitro and in vivo experiments failed to demonstrate a specific defect in the initiation of DNA replication, and turn-off of DNA synthesis at high temperature was slower than that of a typical initiation (dnaA) mutant. The gene was localized, and its product was identified through the construction and analysis of deletion and insertion mutants of dfp-containing plasmids. dfp is located between the rpmB and dut genes at 81 min on the linkage map of Escherichia coli K-12. It is transcribed clockwise, independently of dut. The ability of a plasmid to complement a chromosomal dfp-707 mutation was correlated with its ability to produce a 45-kilodalton polypeptide. The purified protein contained 1 mol of flavin mononucleotide per mol of polypeptide.
Bacterial Proteins↗
Rhodamine-auramine O versus Kinyoun-carbolfuchsin acid-fast stains for detection of Cryptosporidium oocysts.
The rhodamine-auramine O stain was compared with the Kinyoun carbolfuchsin acid-fast stain for detection of Cryptosporidium oocysts in samples from patients infected with the human immunodeficiency virus (HIV). A total of 283 fecal specimens from HIV-infected patients were examined for the presence of Cryptosporidium oocysts. Duplicate smears of the fecal concentrates, prepared by the formalin ethyl acetate procedure, were stained by the Kinyoun carbolfuchsin and fluorescent rhodamine-auramine O acid-fast methods. The Kinyoun stain detected 13 positive specimens, while the rhodamine-auramine O stain detected 14 positive specimens. The average time required to survey a stained smear was 2.5 minutes with the fluorescent method, compared with 6.0 minutes with the Kinyoun technique. The rhodamine-auramine O stain is a dependable and efficient method of examining fecal smears for the presence of Cryptosporidium oocysts in a high-risk population.
AIDS-Related Opportunistic Infections↗
Isolation of Cryptococcus neoformans chromosome-specific probes using expressed sequence tags.
Clinical and environmental isolates of Cryptococcus neoformans exhibit a high degree of karyotypic variability. Analysis of the molecular basis of karyotypic differences requires a large set of chromosome-specific probes. We have determined the chromosomal distribution of a set of randomly selected C. neoformans cDNA clones and have explored the feasibility of identifying these clones through partial DNA sequencing. Forty-four randomly selected cDNA clones were labelled and hybridized to PFGE blots of C. neoformans. Expressed sequence tags were generated by sequencing the 5'-end of each clone. Thirty-five clones hybridized to single bands on PFGE blots. At least seven chromosomes were recognized by these probes. Homology searches identified potential homologs of several groups of proteins not previously studied in C. neoformans. PFGE hybridization and sequencing of random cDNA clones is an efficient method for identifying chromosomal-specific probes in fungi that lack extensive sets of genetic markers.
Chromosomes, Fungal↗