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At least 379 records · Page 21Linked to original sources
PubMed1998

Epidemiological study of a food-borne outbreak of enterotoxigenic Escherichia coli O25:NM by pulsed-field gel electrophoresis and randomly amplified polymorphic DNA analysis.

This study investigated the applicability of molecular epidemiological techniques to the identification of the causal agent of an outbreak of diarrhea caused by ingestion of food contaminated with enterotoxigenic Escherichia coli (ETEC). The outbreak occurred at four elementary schools in July 1996 and affected more than 800 people. Illness was most strongly associated with eating tuna paste (relative risk, 1.79; 95% confidence interval = 1.16 to 2.79; P = 0.0001). To evaluate the epidemiological characteristics of the pathogen, the DNAs from numerous isolated ETEC strains were subjected to randomly amplified polymorphic DNA analysis, pulsed-field gel electrophoresis of nuclease S1-treated plasmid DNA, and analysis of genomic DNA restriction fragment length polymorphisms. All ETEC isolates were of the O25:NM (nonmotile) serotype, which carries a heat-stable enterotoxin Ib gene. Genotypic analysis demonstrated that the strains isolated from the patients at all four schools were identical. The isolates of ETEC O25:NM obtained from the tuna paste that had been served for lunch at these schools were genetically indistinguishable from those isolated from the patients. Results suggest that this outbreak was food borne. The molecular biology-based epidemiological techniques used in this study were useful in characterizing the causal agent in this food-borne epidemic.

Animals↗
PubMed2006

DNA analysis: what and when to request?

Over the last 15 years genetic testing by DNA analysis has expanded enormously both in volume and range due to advances in scientific knowledge and analytical technology. This type of analysis has the potential to provide rapid, cost effective, and accurate diagnostic information but also has its limitations. Some of the changes detected may be of ambiguous consequence and as the knowledge base expands so too does the recognition that other factors can influence the clinical picture. In many cases outcomes may be predicted only on a statistical basis rather than individually. Careful attention should therefore be given to the clinical question that is being addressed before such testing is requested.

DNA↗
PubMed1988

Flow cytometric DNA analysis of neuroblastoma and ganglioneuroma. A 10-year retrospective study.

Retrospective quantitative DNA analysis was done on 147 samples from 89 patients with neuroblastoma and ganglioneuroma using flow cytometry. In the neuroblastoma patients, nuclear DNA content was found to be a stable tumor marker irrespective of site (primary versus metastatic) and despite changes with time in tumor progression, maturation, or therapy. The occurrence of DNA aneuploidy, which was detected in 60% of the neuroblastoma patients, paralleled other favorable indicators and was highly associated with survival (P less than 0.001). Of clinical stage, age, primary site, sex, and DNA content, only stage and DNA content correlated with survival. Those patients with favorable stage and DNA aneuploidy had higher survival rates. Further, favorable stage and the presence of DNA aneuploidy were independent prognostic indicators. Abnormal DNA content was also detected in samples from ganglioneuromas in which significant numbers of ganglion cell nuclei were recovered. These results indicate a striking difference between neuroblastoma and adult tumors in which DNA aneuploidy is generally a poor prognostic sign and provide a molecular link between ganglioneuromas and their malignant counterparts.

Adolescent↗
PubMed1995

DNA analysis of in situ ductal carcinoma of the breast via flow cytometry.

Flow cytometric (FCM) DNA analysis of 41 clinical cases of ductal carcinoma in situ (DCIS) of the breast was performed on fine-needle aspirates of unfixed, frozen tissue. Based on analysis of a single tissue sample, abnormal DNA content was found in 35 (85%) of the cases, and 15 (37%) disclosed heterogeneity. If five or more samples were analyzed, then heterogeneity was found in eight of ten cases. No conclusive correlation could be found between DNA index (DI) and histopathology. Our results on DCIS demonstrate a very high concordance with invasive breast cancer with respect to the frequency of DNA nondiploid cases, distribution of DI, occurrence of multiple clones, and S-phase fraction. This indicates that major genetic alterations and DNA heterogeneity are early events in carcinogenesis that are already established at the preinvasive stage.

Breast Neoplasms↗
PubMed1999

Characterisation of Clostridium botulinum groups I and II by randomly amplified polymorphic DNA analysis and repetitive element sequence-based PCR.

Random amplified polymorphic DNA analysis (RAPD) and repetitive element sequence-based PCR (rep-PCR) were evaluated with respect to their applicability to characterise Clostridium botulinum group I and II strains, the species causing human botulism. Fifteen group I and 21 group II strains of various geographical and temporal origins were characterised with four single arbitrary RAPD primers at low stringency amplification conditions and with a degenerate REP primer pair at moderately stringent conditions. Ready-To-Go RAPD Analysis Beads and Ready-To-Go PCR Beads were used for PCR reactions with RAPD and rep-PCR, respectively. Arbitrary primer OPJ 6 yielded the most discriminating patterns, and distinguished group II C. botulinum serotypes at the strain level. Group I strains were mainly discriminated at the serotype level. The discriminatory power of rep-PCR was found to be inferior to that of RAPD. The REP1R-Dt and REP2R-Dt primer pair generated group I- and II-specific fragments and arbitrary primer OPJ 13 produced a serotype E-specific fragment. The use of pre-dispensed and pre-optimised beads attributed to highly reproducible results. As compared to more time-consuming typing methods, such as pulsed-field gel electrophoresis (PFGE), both RAPD and rep-PCR were characterised by rapid performance and a typeability of 100%.

Animals↗
PubMed1984

Flow DNA analysis of primary bone tumors. Relationship between cellular DNA content and histopathologic classification.

The cellular DNA content of 15 benign and 34 malignant primary bone tumors was analyzed by means of flow cytophotometry. All benign tumors except one of questionable histologic type exhibited a normal DNA content (diploid), whereas 23 of 34 malignant tumors showed an abnormal DNA content (aneuploid). Closer analysis revealed that all supposedly highly malignant tumors, i.e., 16 osteosarcomas and 1 Ewing sarcoma were aneuploid, while 8 of 13 chondrosarcomas, 2 periosteal osteosarcomas, and 1 of 2 adamantinomas were diploid. Interestingly, these diploid malignant tumors represent tumor entities which are known to include variants of low-grade malignancy. Cell distribution analysis showed that the aneuploid tumors exhibited a higher proportion of S-phase and G2 + M cells than the diploid tumors, indicating differences in proliferative activity. However, no significant difference in this respect could be demonstrated between diploid benign and diploid malignant tumors. The current study clearly shows that flow DNA cytophotometry can be applied to most primary bone tumors despite a substantial content of hard tissue. The results also indicate that DNA determinations as an adjunct to conventional histopathologic assessment may provide objective clinically relevant information with respect to the degree of malignancy. Thus, regardless of histogenetic origin, it appears that benign bone tumors as well as malignant bone tumors of low-grade malignancy in general, are diploid, whereas highly malignant bone tumors in general are aneuploid.

Adolescent↗
PubMed1996

Flow cytometric DNA-analysis of cells obtained with fine needle aspiration biopsy of the breast.

In 221 consecutive, unselected palpable breast lesions, evaluated by aspiration biopsy, flow cytometric DNA analysis was performed on cell material remaining in needle and syringe after smears had been prepared. DNA histograms were obtained in 186 cases, whereas in 44 samples the cell number was too low for detailed estimation of cell kinetics. Aneuploidy was observed in 71 of 90 samples where cancer cells were detected by cytology, in four of 20 samples where breast cancer could only be suspected and in 12 of those 76 where no cancer cells occurred in cytologic specimens. The average S-phase and S+G2M values were higher in aneuploid than in diploid cell samples. In the group of 40 cases (34 breast cancers, 6 benign lesions) comparative analysis of DNA histograms obtained on aspirates and tissue fragments revealed a concordance in 88.9%. Modified radical mastectomy was performed in 41 breast cancers and in 33 cases ductal infiltrating carcinomas were recognized by histology and scored according to Bloom-Richardson score. Cell features assessed by cytology in these cases did not correlate with histology. In most cases, however, nuclear polymorphism and numerous naked nuclei were present in cytologic specimens. These alterations, characterized as score 3, reflect values of DNA-index in compartment of 1.4-1.8.

Biopsy, Needle↗
PubMed1992

Diagnosis of heterozygous familial hypercholesterolemia. DNA analysis complements clinical examination and analysis of serum lipid levels.

The concordance of clinical and molecular genetic diagnoses of heterozygous familial hypercholesterolemia (FH) was studied in 65 subjects (10 propositi and 55 first-degree relatives) from 10 families with FH. Nine propositi were carriers of the FH-Helsinki deletion of the low density lipoprotein (LDL) receptor gene, prevalent in the Finnish population, while a new deletion, extending from intron 14 to intron 15 of the LDL receptor gene, was identified in one family. Serum LDL cholesterol levels used in the clinical diagnosis (less than 5.0 mmol/l, not FH; 5.0-5.9 mmol/l, possible FH; greater than or equal to 6.0 mmol/l, FH; limits are 1 mmol/l lower for those less than 18 years) were derived from an authoritative recommendation. Tendon xanthomas constituted an additional criterion. With the DNA analysis as the reference, 55 (85%) subjects could be correctly classified clinically as FH patients or subjects without FH. The remaining 10 subjects were misclassified or were in the "possible FH" category. When the age- and sex-specific 95th percentile LDL cholesterol levels were used instead of the rigid values for both adults and children, the percentage of correct diagnoses rose to 95%. Common genetic polymorphisms of apolipoproteins E and B did not markedly affect LDL cholesterol levels in FH patients, whereas increasing age and obesity were associated with elevated LDL levels. In conclusion, DNA analysis is a valuable adjunct to the diagnosis of FH that is applicable to families with a known mutation of the LDL receptor gene. If DNA methods are not available, age- and sex-specific LDL levels should be used as an aid in the clinical diagnosis of FH.

Adolescent↗
PubMed2003

Using hydrophilic adhesive tape for collection of evidence for forensic DNA analysis.

Known exemplar samples of human DNA have traditionally been body fluids, such as blood, saliva, and semen. In each case, the presence of water is a risk for the bacterial growth, which may degrade the DNA evidence. In this study, the authors have developed a method that employed a hydrophilic adhesive tape (HAT) for collecting DNA evidence. The HAT method was used to remove surface cells from relatively hairless areas on the body. The area examined were ankle, arm, behind the ear, between fingers and back of the neck. The HAT was then dissolved in the extraction buffer. DNA typing was performed at vWA, THo1, F13A1, and FES loci using the short tandem repeat (STR) analysis. Our results show that the samples collected from ear give the best results with a success rate of 100%. All subjects tested by this method had known STR genotypes established from buccal swabs. The authors' results suggest that the HAT method can be used as a less invasive method for collecting biological evidence for forensic DNA analysis. In addition, this collection method should reduce the risk of DNA degradation due to the moisture, which is encountered using conventional collecting methods.

Adhesives↗
PubMed1981

Flow cytometric DNA analysis of anal canal epithelium and ano-rectal tumours.

Flow cytometric DNA analysis was performed on mucosal biopsies from the various epithelial types normally present in the anal canal, i.e. colo-rectal epithelium, ATZ-epithelium and squamous epithelium as well as from rectal and anal adenocarcinomas and squamous carcinomas of the anal canal. Normal colo-rectal mucosa show homogeneous cell populations with diploid DNA values, while adenocarcinomas contain diploid as well as aneuploid cell populations, regardless of location in the rectum or in the anal canal. The ATZ-epithelium shows a dominating normoploid population with a small "hyperdiploid" peak. Polyploid nuclei were not found. The findings indicate that the ATZ-epithelium is related to metaplastic epithelium rather than to urothelium. The variants of squamous carcinoma in this study were rather poorly differentiated, but the FCM-analysis showed a near diploid population with a high proliferative activity. The number of cases, however, is too small to be conclusive.

Anal Canal↗
PubMed2003

Morphological convergence in Hippidion and Equus (Amerhippus) South American equids elucidated by ancient DNA analysis.

Unusual equids named hippidions inhabited South America for more than 2 MY (million years). Like many other animals they succumbed to the worldwide climatic change that occurred 10 KY (thousand years) ago and completely disappeared during the great late Pleistocene megafaunal extinction. According to fossil records and numerous dental, cranial, and postcranial characters, Hippidion and Equus lineages are known to have diverged prior to 10 MY. Some equid bones from Rio Verde and Ultima Esperanza (Patagonia, Chile) dating back to the late Pleistocene period (8-13 KY) have been identified as Hippidion saldiasi, while a few teeth have been assigned to Equus. Six samples of those remains have been obtained from the Zoological Museum of Amsterdam for ancient DNA analysis to try to place Hippidion in the evolutive tree of Perissodactyla. Two samples of Hippidion and one sample of Equus yielded 241-394 bp of the mtDNA control region and 172-296 bp of the cytochrome b gene. Unexpectedly, all the sequences clustered deep inside the Equus genus, casting doubt on the initial identification of the bones. For paleontologists, one of the striking and classical diagnostic characters of Hippidion is their extremely short and massive metapodials, a probable locomotory adaptation to the Andine steep slopes. However, our DNA analysis reveals that a very Hippidion-like metapod might also have been possessed by another South American equid, i.e., Equus (Amerhippus), an interpretation supported by complementary anatomical observations. This adaptive convergence between members of the two South American equid genera may lead paleontologists to limb bone misidentification.

Animals↗
PubMed2006

DNA analysis of digested tomato seeds in stomach contents.

Examination of stomach contents is one of the important steps in medical legal autopsy. Vegetative materials such as stems, roots, and seeds in stomach contents can be valuable evidence for providing investigative leads in death investigation. Currently, the identification of plant materials relies on microscopic and morphologic examination. We have found that many seeds are often protected from acid degradation during stomach digestion by their tough exterior seed coat. Tomato seeds were selected as a model system to assess DNA analysis and plant variety marker identification. The DNA-amplified fragment length polymorphism method was performed to determine if the DNA obtained from single seeds could be used for PCR analysis. From the amplified fragment length polymorphism results, some candidate markers for individualizing seeds from morphologically distinct tomatoes were identified. These data on DNA analysis of tomato seeds indicate amplified fragment length polymorphism is a viable procedure for the individualization of seeds from stomach contents in forensic investigations.

DNA, Plant↗
PubMed1983

Long-term storage of samples for flow cytometric DNA analysis.

A simple procedure for long-term storage of cells for flow cytometric DNA analysis was developed and tested. The cells were stored as single cells or fine-needle aspirates suspended in a citrate buffer with dimethylsulfoxide (DMSO), or as small blocks of tissue from solid tumors. The cells were stored for up to one year by freezing at -80 degrees C. Statistical analysis of the results showed no change in the fractions of cells in the cell cycle phases as determined by deconvolution of the DNA-histograms. It was found that in addition to the intrinsic sample variation from the parameter estimation by deconvolution, there was significant intraday and interday variation. Hence the most accurate results are obtained if different aliquots of a sample are measured on different days rather than on the same day. Use of the storage method thus has the potential of increasing the accuracy of the analysis. The storage method makes sample collection independent of immediate subsequent analysis. This has enabled us to perform large internally controlled experiments, involving more samples than can be analyzed in one day, to examine tumor samples from different hospitals and to utilize fully the capacity of our flow cytometer. The method was a prerequisite for developing an accurate standardization procedure for DNA content determination.

Analysis of Variance↗
PubMed1994

[Clinical significance of flow cytometric DNA analysis in metastatic lymph node of colorectal cancer].

Significance of flow cytometric DNA analysis in metastatic lymph nodes for assessing malignant potential of colorectal cancer was investigated using paraffin-embedded materials of primary lesions and metastatic lymph nodes from 65 patients who had been treated between 1975 and 1990. The DNA ploidy patterns of metastatic nodes were identical in 61.5% with those of primary lesions. Diploid cancers were significantly more frequent in metastatic nodes than in primary lesions. There were significantly more aneuploid cancers in proximal nodes than in distant nodes. There was no relation between ploidy patterns in primary lesions and survival. However, a significant relation was found between ploidy patterns in metastatic nodes and survival. Diploid cancer in metastatic nodes had a significantly better survival than aneuploid cancer, in all patients as well as those with curative resection. In patients with stage III and in those with the same depth of invasion, the survival rate of diploid cancer in metastatic nodes was significantly higher. There was no correlation between ploidy patterns in metastatic nodes and clinicopathological variables in primary lesions, such as histological type, depth of invasion, nodal involvement, peritoneal or hepatic involvement and stage. These results suggest that nuclear DNA content in metastatic lymph nodes may be a prognostic indicator in colorectal cancer with nodal involvement.

Colorectal Neoplasms↗
PubMed1996

Analysis of genetic relationships among strains of groups A, C and G streptococci by random amplified polymorphic DNA analysis.

Random amplified polymorphic DNA (RAPD) analysis was evaluated for its capacity to distinguish species and strains within species of groups A, C and G streptococci. The 99 strains tested, previously typed by multilocus enzyme electrophoresis (MLEE), included 41 group A streptococci (Streptococcus pyogenes), 25 group G Streptococcus spp. (GGS), seven S. dysgalactiae, 11 S. equisimilis, four S. canis, three S. equi and eight S. zooepidemicus. The combined data obtained with three single primers distinguished 82 types. RAPD analysis provided taxonomic results that were in general agreement with previous species classification based on DNA-DNA homology and MLEE. The intraspecies typing efficiency of the technique was significantly improved by the parallel use of several primers. RAPD analysis had greater discriminatory power than MLEE for GAS and GGS. There was not total agreement between the two techniques as RAPD distinguished strains with identical electrophoretic types, whereas MLEE differentiated strains with identical PCR types. RAPD analysis did not distinguish all GAS strains with different biotypes and its already high discriminatory power was further enhanced by concomitant biotyping.

Animals↗
PubMed1987

Hemophilia A: carrier detection and prenatal diagnosis by DNA analysis.

In this study, we used DNA polymorphisms for carrier detection and prenatal diagnosis of hemophilia A in a large group of Italian families. The restriction fragment length polymorphisms (RFLPs) investigated were the intragenic polymorphic Bc/I site within the factor VIII gene; the extragenic multiallelic Taq I system at the St14 locus; and the extragenic Bg/II site at the DX13 locus. The factor VIII probe was informative in 30%, St14 in 82%, and DX13 in 60% of obligate carriers. The combination of factor VIII-Bc/I and St14-Taq I showed that 91% of obligate carriers were heterozygotes for one or both; with all three probes, only 4% of obligate carriers were noninformative. In families clearly segregating for hemophilia A, RFLP analysis allowed us to define the carrier status for the hemophilia A gene in all 27 women tested. RFLP analysis allowed us to exclude the carrier status in 39 of 45 female relatives of sporadic patients. The combination of RFLP analysis and biological assay of factor VIII allowed us to identify a de novo mutation in the maternal grandfather in 7 of 12 of the families with sporadic cases, for which members of three generations were available for study. Nine of 10 couples requesting prenatal diagnosis provided informative RFLP DNA pattern. Carrier status was excluded in two women, two fetuses were shown to be female, and prenatal diagnosis was carried out in five pregnancies by DNA analysis. Prenatal testing was successful in three instances and failed in two because a sufficient amount of chorionic villous DNA was not obtained for the analysis.

DNA↗
PubMed1984

Flow cytometric DNA analysis as a diagnostic aid for cervical condyloma and cancer.

Flow cytometric DNA analysis data (FCDA) were obtained from 324 samples provided through the Gynecology-Oncology Clinic. These samples consisted of 294 combined endoectocervical and vaginal smears and 30 peritoneal washings. Using a conventional scheme for G0/G1, S + G2/M and the coefficient of variation with computer correction for the cell-cycle kinetics, it was possible to assign a diagnostic Class I, II, III or V similar to that used by the Cytology Laboratory. These data were then compared with the histopathologic and colposcopic diagnoses. The correlation between FCDA and cytologic results were essentially similar to the previous data obtained from only endocervical sampling. The most interesting finding in this study was the recognition of an FCDA pattern showing a higher DNA content in the G0/G1 and the early S regions in 70 of 94 (74.5%) of samples from patients with condyloma acuminata. All condyloma samples were diagnosed either by cytologic, histopathologic, or colposcopic examination, or a combination of two or three. All biopsy specimens were then reviewed by one pathologist to verify any discrepancies. The relationship of this pattern to the viral etiology of this disease is discussed with the three methods of diagnosis and electron microscopic observations. It is suggested that, based on this study. FCDA analysis of pap smears may also be useful in determining the presence of condyloma in a gynecology clinic. The potential value of FCDA analysis from peritoneal washings for the diagnosis of gynecologic cancer can not be ascertained in this preliminary investigation because of insufficient samples.

Cell Cycle↗
PubMedDate not supplied

Image cytometry DNA analysis of dysplastic squamous epithelial lesions in the larynx.

The Feulgen-DNA content of cell nuclei from the human larynx was assessed in 62 lesions from 14 patients with dysplastic and cancerous lesions and in 14 control patients with non-neoplastic chronic laryngitis. All the carcinomas displayed aneuploid cell nuclei, and the cellular DNA content was substantially altered in dysplasias which later progressed to cancer in situ or invasive cancer. Thus the process of laryngeal carcinogenesis can be monitored not only by histological changes, but also by cellular DNA aberrations. Quantitative DNA analysis appears to be a complement to the histopathological evaluation of laryngeal lesions in the search for neoplasia.

Biopsy↗