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F L Macrina

Publications and source records attributed to F L Macrina.

89 records · Page 5Linked to original sources
PubMed1980

Chimeric streptococcal plasmids and their use as molecular cloning vehicles in Streptococcus sanguis (Challis).

Chimeric plasmids, which were useful as cloning vehicles in a Streptococcus sanguis (Challis) host vector system, have been constructed. By using three different strategies of restriction endonuclease digestion and ligation, a deoxyribonucleic acid (DNA) fragment bearing an erythromycin resistance determinant was ligated in vitro to a phenotypially cryptic plasmid from Streptococcus ferus. Recombinant plasmids could be recovered after transformation of S. sanguis (Challis) with these preparations. Three useful chimeras were constructed. pVA680, 5.5 megadaltons in size, contained a single KpnI site into which passenger DNA may be spliced. pVA736, 5.0 megadaltons in size, contained single EcoRI, HindIII, and KpnI sites into which passenger DNA may be spliced. The EcoRI and KpnI sites of pVA736 may be used in combination with one another when ligating DNA into this plasmid. pVA738, 3.7 megadaltons in size, contained single HindIII and AvaI sites into which passenger DNA may be spliced. pVA680, pVA736, and pVA738 were stably maintained as multicopy plasmids in S. sanguis (Challis). None of them continued to replicate (amplify) in chloramphenicol-treated cells. By using pVA736 as a vector, we have cloned a chloramphenicol resistance determinant obtained from a large, conjugative streptococcal R plasmid. In addition, chromosomal DNA sequences from Streptococcus mutans have been inserted into pVA736 by using the KpnI-EcoRI site combination.

Chloramphenicol↗
PubMed1978

Extrachromosomal gene systems in Streptococcus mutans.

1) Four of 86 S. mutans strains of human origin harbor a 3.6 Mdal multicopy plasmid while 2 of 13 strains of rat origin carry multiple copies of a 2.3 Mdal and 2.5 Mdal plasmid. 2) There is no evidence to support that any of these plasmids confers clinically or ecologically important phenotypes. 3) The restriction endonucleases Hind III and Bam HI cleave the 3.6 Mdal plasmid once, thus facilitating the use of this extrachromosomal element in gene cloning systems.

DNA Restriction Enzymes↗
PubMed1978

Evidence for a disseminated plasmid in Streptococcus mutans.

Based on a survey of 86 isolates, approximately 5% of all naturally occurring strains of Streptococcus mutans contains a 3.6 X 10(6)-dalton (3.6-megadalton) multicopy plasmid of unknown function. The amount of plasmid deoxyribonucleic acid per chromosome varies from 2 to 6% depending on the host strain. About 13% of the total covalently closed circular deoxyribonucleic acid in each of the four plasmid-containing strains consists of dimeric molecules, with interlocked circular forms predominating. Site-specific restriction endonucleases have been identified that cleave this 3.6-megadalton plasmid at single and at multiple sites. Each of the four plasmids is cleaved once by the HindIII and BamHI restriction enzymes. The HpaI, TaqI, and HhaI enzymes generate two, five, and six components, respectively, and the digestion products of each of the four plasmids are identical. Because the four plasmid-containing S. mutans strains are physiologically unique with respect to one another, we conclude this plasmid to be a disseminated extrachromosomal element in S. mutans.

DNA, Bacterial↗
PubMed1978

R-factor inheritance and plasmid content in mucoid Pseudomonas aeruginosa.

Eighteen strains of alginate-producing mucoid Pseudomonas aeruginosa were evaluated with respect to plasmid content and the ability to maintain well-characterized R plasmids. The spontaneous loss of alginate production in these strains varied from 0.01 to 0.7% and was not significantly increased by plasmid curing regimens. Examination of cleared lysates of these strains and their isogenic nonmucoid derivatives by agarose gel electrophoresis failed to reveal plasmid DNA. R-plasmid (P-incompatibility-group) transfer to mucoid P. aeruginosa was unaffected by the presence of the alginate capsule. Maintenance and expression of such plasmids in the mucoid strains were confirmed by agarose gel electrophoresis and by verification of plasmid-linked drug resistance and pilus-specific bacteriophage sensitivity. These studies demonstrate that alginate production does not appear to be plasmid linked and that mucoid P. aeruginosa are capable of receiving and donating certain drug resistance plasmids. Since some of the plasmids used here have been shown to mobilize chromosomal DNA, strains constructed in this study should afford the means for exploring the genetic basis of the mucoid phenotype.

Alginates↗
PubMed1977

Survey of the extrachromosomal gene pool of Streptococcus mutans.

Fifty strains of Streptococcus mutans independently isolated from human dental plaque were examined for the presence of covalently closed circular plasmid deoxyribonucleic acid (DNA). Cesium chloride-ethidium bromide centrifugation of [3H]thymidine-labeled, Sarkosyl-lysed cells revealed that 2 of the 50 strains contained plasmid DNA. The plasmid DNA from these strains was characterized by velocity and equilibrium centrifugation and by electron microscopy. The plasmids in these strains were virtually identical in size, with molecular weights of 3.6 X 10(6) and 3.7 X 10(6), Both were present to the extent of approximately 20 molecules per genome equivalent. Interlocked catenated dimeric molecules of each plasmid were readily detected by velocity sedimentation and electron microscopy. These plasmid-containing strains were compared with representative plasmid-free S. mutans strains by using such criteria as bacteriocin production, antibiotic susceptibility, and hemolysis of mammalian erythrocytes. Although no correlation of phenotype to plasmid content could be made, production of bacteriocin-like activity differed significantly between the two plasmid-containing S. mutans isolates. Thus, although the plasmids in these two isolates appeared identical by the criteria of molecular weight, presence of dimers, and copy number, they appeared to be harbored by two distinct S. mutans strains.

Anti-Bacterial Agents↗
PubMed1976

Extrachromosomal elements in a variety of strains representing the Bacteroides fragilis group of organisms.

Previous nucleic acid association studies have identified at least nine deoxyribonucleic acid (DNA) homology classes of the Bacteroides fragilis group of organisms. Using these classes as a taxonomic framework, we have screened representative strains of the B. fragilis group for the presence of extrachromosomal (plasmid) DNA. [3H]thymidine-labeled cell lysates were subjected to sodium dodecyl sulfate-salt precipitation, and supernatant fractions from such preparations were analyzed using cesium chloride-ethidium bromide equilibrium centrifugation. One strain from each group was examined in this fashion. Five of the strains were judged to contain no detectable plasmid DNA; however, four strains were observed to yield satellite bands corresponding to covalently closed circular plasmid DNA. Plasmid DNA from such gradients was subjected to velocity sedimentation through both neutral and alkaline sucrose gradients to determine molecular size. A 23 X 10(6)-molecular-weight plasmid was found in a B. fragilis strain representing one DNA homology group of this species, whereas a 3 X 10(6)-molecular-weight plasmid was found in a B. fragilis strain representing a second homology group. Similarly, a 31 X 10(6)-molecular-weight plasmid was found in a Bacteroides thetaiotaomicron strain representing one DNA homology group of this species, whereas a 3 X 10(6)-molecular-weight plasmid was found in a B. thetaiotaomicron strain representing a second homology group. In all instances, the small-molecular weight plasmids were present to the extent of about 15 copies per chromosomal equivalent, whereas the large plasmids were present to the extent of approximately 1 copy per chromosomal equivalent. The biological function of these plasmids is unknown.

Bacteroides fragilis↗
PubMed1974

R6K plasmid replication: influence of chromosomal genotype in minicell-producing strains of Escherichia coli K-12.

Alkaline sucrose velocity sedimentation and cesium chloride-ethidium bromide equilibrium centrifugation have been used to determine the number of copies per chromosomal equivalent of the relaxedly replicating R6K plasmid (a conjugative plasmid conferring ampicillin and streptomycin resistance) in two minicell-producing strains of Escherichia coli K-12. In one strain, the average number of covalently closed circular R6K molecules per chromosomal equivalent is 13 in log-phase and 35 in stationary-phase cells. In the other strain, there is an average of six covalently closed circular R6K molecules per chromosomal equivalent in both log- and stationary-phase cells. Selection from this strain of spontaneously occurring mutants resistant to high concentrations of ampicillin has been accomplished and such mutants show a two- to threefold increase in the number of R6K copies per chromosomal equivalent. Relative to the parental strain, mutants display the following properties: (i) elevated streptomycin resistance, (ii) a 10-fold increase in R6K conjugal transfer, (iii) a 10-fold increase in the amount of R6K plasmid deoxyribonucleic acid segregated into minicells, and (iv) a two- to threefold increase in R6K-specified beta-lactamase. The mutation(s) responsible for the increase in the number of R6K molecules per chromosomal equivalent is located on the bacterial chromosome. No R6K-linked mutations conferring the above phenotypes have been obtained. The mutations are presumed to be in chromosomal genes which play a role in the regulation of R6K replication in this strain.

Cell-Free System↗
PubMed1973

Plasmid-associated functions of a stable Flac.

Several functions associated with the stable plasmid, FlacS, have been examined. Our results indicate that the sex pili synthesized by Escherichia coli strains carrying FlacS are altered in some manner as evidenced by a very inefficient adsorption of male-specific phages. On the other hand, FlacS-mediated entry exclusion of related plasmids and plasmid incompatibility function(s) appear normal. The presence of covalently closed circular deoxyribonucleic acid in E. coli strains harboring FlacS indicates that it is an autonomously replicating plasmid. Based on beta-galactosidase levels and the percentage of covalently closed circular deoxyribonucleic acid, it appears that the stability of the FlacS is not the result of multiple copies of this plasmid. FlacS appears larger than its precursor, F(ts114)lac, in sedimentation through alkaline sucrose gradients.

Adsorption↗
PubMed1972

Genetic Characterization of a Stable F' lac Plasmid.

A mutant F' plasmid has been isolated in a strain of Salmonella typhimurium harboring F(ts114)lac. This mutant, designated FlacS, exhibits unique genetic stability in strains of S. typhimurium and Escherichia coli. It shows no thermolability and is lost at frequencies of 20 to 100 times less than the wild-type F'lac (F42) in the same genetic backgrounds. The FlacS is also insensitive to conventional plasmid curing agents, whereas both F(ts114)lac and F42 are readily cured. The nature of the mutation(s) conferring stability to the FlacS is unclear, but plasmid linkage has been established. The high frequency of conjugal transfer of the FlacS and its behavior in recombination-deficient strains of S. typhimurium and E. coli argue against its stability being due to stable chromosomal integration. The FlacS is also capable of transferring chromosomal markers in S. typhimurium and E. coli mating systems. No major differences in chromosomal mobilization have been observed among F42, F(ts114)lac, and FlacS donors of either genus.

Journal Article↗
PubMedDate not supplied

The case study approach to teaching scientific integrity in nursing and the biomedical sciences.

Scientific integrity courses for graduate trainees are emerging at many institutions. The content of such courses varies and is evolving as faculty strive to meet the needs of their specific student audiences. Typical course subject matters include responsible authorship, conflict of interest, handling of scientific misconduct, data management, and the use of humans and animals in biomedical experimentation. These and other topic areas are amenable to teaching methods that use the case study approach. In this article we discuss our experiences in teaching scientific integrity using the case study approach. Issues covered include underlying philosophy of the approach, preparation and use of cases, desired inclass preparation and teaching. Examples of short cases are presented and, where appropriate, discussed in the context of these issues.

Animal Experimentation↗
PubMedDate not supplied

Nosocomial klebsiella infection in a neonatal unit: identification of risk factors for gastrointestinal colonization.

Sequential outbreaks of infection due to gentamicin-resistant Klebsiella pneumoniae (GRKP) types 30 and 19 occurred in the neonatal intensive care unit (NICU) at the Medical College of Virginia in 1977 and 1978. The extensive epidemiologic investigation carried out included a case-control study, careful review of aseptic technique, and cultures from nursery staff and environment. The gastrointestinal (GI) tracts of the patients were the reservoirs for GRKP, and the epidemic strain was transmitted by hands of personnel. The case-control study showed a significant relationship between acquisition of GRKP by patients and oropharyngeal and GI instrumentation, including use of bag resuscitation, oropharyngeal suctioning, and use of nasogastric feeding tubes. The findings of the case-control study were supported by observation of the patient care techniques practiced by NICU staff. Institution of control measures based on results of the epidemiologic investigation of the first outbreak rapidly brought the second outbreak under control, even though cohorting or use of routine isolation was not possible. Whereas GI colonization and hand transmission have been described previously in outbreaks of K. pneumoniae infections in NICUs, this study is the first to document the mode of inoculation of patients' GI tracts by contaminated hands of personnel.

Cross Infection↗
PubMedDate not supplied

Tn5030: a conjugative transposon conferring clindamycin resistance in Bacteroides species.

Several examples of conjugal transfer of antibiotic resistance determinants in the absence of plasmid DNA have been described in gram-positive species. Previous work had suggested that transfer of clindamycin resistance among the gram-negative anaerobic Bacteroides species might occur by a similar mechanism. We report here the characterization of Tn5030, a 45-kb +/- 5-kb chromosomal, mobile genetic element devoid of the detectable terminal repeats typically associated with gram-negative transposons. Tn5030 transfers clindamycin resistance (ermFU) and sequences homologous to tetF in the absence of detectable plasmid DNA. This element represents a novel mechanism of genetic exchange among gram-negative organisms.

Bacteroides↗