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PubMed2004

Quantitative DNA fingerprinting may distinguish new primary breast cancer from disease recurrence.

PURPOSE: Approximately 10% of women with breast cancer develop a second breast tumor, either a new primary or a recurrence. Differentiating between these entities using standard clinical and pathologic criteria remains challenging. Ambiguous cases arise, and misclassifications may occur. We investigated whether quantitative DNA fingerprinting, based on allele imbalance (AI) or loss of heterozygosity (LOH), could evaluate clonality and distinguish second primary breast cancer from recurrence. METHODS: We developed a scoring system based on the AI/LOH fingerprints of 20 independent breast tumors and generated a decision rule to classify any breast tumor pair as related or unrelated. We validated this approach on eight related tumors (cancers and synchronous positive lymph nodes). Finally, we analyzed paired tumors from 13 women (bilateral cancers, primary tumors and contralateral positive axillary lymph nodes, or two ipsilateral tumors). Each pair's genetic classification was compared with their clinical diagnosis and outcome. RESULTS: Each independent cancer had a unique fingerprint. Every tumor pair's relationship was quantifiable. Six of eight related tumor pairs were genetically classified correctly, two were indeterminate, and none were misclassified. Among the 13 women with two cancers, four of five clinically indeterminate pairs could be classified genetically. In three of 13 women, the pair's classification contradicted the clinical diagnosis. These women had bilateral cancers genetically classified as related and disease progression. This challenges the paradigm that bilateral cancers represent independent tumors. Overall, women with tumors genetically classified as related had poorer outcomes. CONCLUSION: Quantitative AI/LOH fingerprinting is a potentially valuable tool to improve diagnosis and optimize treatment for the growing number of second breast malignancies.

Alleles↗
PubMed1997

DNA fingerprints from Mycobacterium tuberculosis isolates of patients confined for therapy noncompliance show frequent clustering.

STUDY OBJECTIVE: To test the hypothesis that individuals chronically noncompliant with antituberculous chemotherapy are vectors for ongoing transmission of the disease in the community. DESIGN: Cohort study. SETTING: A large public hospital with a tuberculosis detention unit for patients with repeated and prolonged nonadherence to therapy. PATIENTS: Mycobacterium tuberculosis isolates from patients confined on the detention unit were obtained from the hospital's mycobacteriology laboratory. INTERVENTIONS: None. MEASUREMENTS AND RESULTS: A standardized IS6110-based Southern blot hybridization protocol was used to genotype M tuberculosis isolates recovered from patients confined on the detention unit at the hospital. Each DNA fingerprint pattern was compared with the IS6110-fingerprint database at the Public Health Research Institute Tuberculosis Center, which has archived fingerprint patterns from over 2,500 M tuberculosis isolates collected from New York City patients in the past 5 years. Eighty percent of available isolates from detained patients belonged to an identifiable DNA fingerprint cluster, suggesting an epidemiologic link between the detainees and other New York City tuberculosis patients. CONCLUSIONS: Chronic noncompliance with therapy is associated with ongoing spread of tuberculosis in the community. Aggressive measures, including detention, for the small number of recalcitrant, noncompliant patients may interrupt a chain of transmission and contribute to a decline in the spread of tuberculosis in urban areas.

Animals↗
PubMed2001

Value of Mycobacterium tuberculosis fingerprinting as a tool in a rural state surveillance program.

OBJECTIVE: This study demonstrates the value of Mycobacterium tuberculosis fingerprinting used in conjunction with traditional epidemiologic methods to identify smoldering outbreaks of tuberculosis in endemic areas where background rates of tuberculosis are high. METHODS: IS6110 DNA fingerprinting was performed on isolates of M tuberculosis from verified cases of tuberculosis in Alabama from 1994 to 1998. A statewide database groups isolates into "clusters" and tracks them cumulatively over time. A large cluster was identified and was secondarily investigated using traditional epidemiologic methods. RESULTS: Twenty-five isolates were found to be identical by fingerprinting analysis. Patients were living within 10 counties across the state, and 12 cases were localized to a single county. This represented an ongoing, statewide tuberculosis outbreak previously unrecognized by local and state health officials. Secondary investigation of the cases revealed the primary sites of transmission to be a correctional facility and two homeless shelters. CONCLUSIONS: Population surveillance using M tuberculosis fingerprinting was successfully utilized to detect a significant and smoldering tuberculosis outbreak. Measures are currently in place to identify and prevent further transmission in the involved locations.

Adult↗
PubMed1998

Denaturation fingerprinting: two related mutation detection methods especially advantageous for high G + C regions.

Two versions of denaturation fingerprinting (dnF2R and dnF1R are described for detecting mutations. DnF2R is a sensitive screening method in which fingerprints are generated by performing denaturing gel electrophoresis on bidirectional "cycle-sequencing" reactions with each of two dideoxy terminators, e.g. ddATP and ddCTP. When the fingerprints generated by ddATP and ddCTP are combined, all sequence changes are expected to result in one extra and one absent segment. DnF2R was performed on 246- and 318-bp segments of the human factor IX gene, and the products were electrophoresed through a 6% Long Ranger gel with 7 M urea. All 32 single-base mutations were detected in hemizygous males with hemophilia B. DnF2R has been applied to detect a total of seven heterozygous sequence changes in large-scale screening and was found to be especially suitable for high G+C regions. In a blinded analysis, all of twenty-four additional single-base mutations were detected, but 7 of 31 heterozygous mutations were missed (23%). To reduce the effort of dnF2R by almost twofold while retaining the ability to detect all types of single-base changes, one fingerprint (dnF1R) was generated by performing a single reaction with ddATP and a second chemically modified terminator (e.g., ROX-conjugated ddCTP), which retards the mobility of the same termination products. The sensitivity and specificity of dnF1R equaled that of dnF2R, with the exception that the blinded analysis of heterozygotes in the 318-bp segment, which revealed the presence of an additional mutation. DnF under partially denaturing conditions may have optimal sensitivity for the detection of heterozygotes.

DNA Fingerprinting↗
PubMedDate not supplied

Accommodating error analysis in comparison and clustering of molecular fingerprints.

Molecular epidemiologic studies of infectious diseases rely on pathogen genotype comparisons, which usually yield patterns comprising sets of DNA fragments (DNA fingerprints). We use a highly developed genotyping system, IS6110-based restriction fragment length polymorphism analysis of Mycobacterium tuberculosis, to develop a computational method that automates comparison of large numbers of fingerprints. Because error in fragment length measurements is proportional to fragment length and is positively correlated for fragments within a lane, an align-and-count method that compensates for relative scaling of lanes reliably counts matching fragments between lanes. Results of a two-step method we developed to cluster identical fingerprints agree closely with 5 years of computer-assisted visual matching among 1,335 M. tuberculosis fingerprints. Fully documented and validated methods of automated comparison and clustering will greatly expand the scope of molecular epidemiology.

Algorithms↗
PubMed1995

Interstrain polymorphisms of isoenzyme profiles and mitochondrial DNA fingerprints among seven strains assigned to Acanthamoeba polyphaga.

Interstrain polymorphisms of isoenzyme profiles and mitochondrial (Mt) DNA fingerprints were observed among seven strains of Acanthamoeba isolated from different sources and morphologically assigned to A. polyphaga. Mt DNA fingerprints by eight restriction endonucleases (Bgl II, Sca I, Cla I, EcoR I, Xba I, Kpn I, Sal I, and Sst I) revealed considerable interstrain polymorphisms. Isoenzyme profiles revealed considerable interstrain polymorphisms for acid phosphatase, lactate dehydrogenase, and glucose-6-phosphate dehydrogenase while those for glucose phosphate isomerase, leucine aminopeptidase, and malate dehydrogenase showed similarity. Despite of the interstrain polymorphisms, the isoenzyme profiles and Mt DNA fingerprints of the strain Ap were found to be identical with those of the strain Jones. Mt DNA fingerprinting was found to be highly applicable for the strain identification, characterization, and differentiation.

Acanthamoeba↗
PubMed2000

A prospective evaluation of the clinical value of nation-wide DNA fingerprinting of tuberculosis isolates in Denmark.

SETTING: Denmark, a country with a low incidence of tuberculosis (TB). OBJECTIVE: To evaluate the value of the nation-wide DNA fingerprinting of Mycobacterium tuberculosis isolates performed in Denmark since 1992. DESIGN: Prospective study of consecutive patients with culture-verified TB from five large TB Departments in Denmark during a 7-month period in 1998. Results of IS6110 RFLP and spoligotyping were compared to those in the nation-wide Danish DNA-fingerprint database which covers approximately 95% of all culture-verified TB cases from 1992 onwards. Questionnaires asking about contact tracing and epidemiological links were sent to the patients' treating physicians. RESULTS: Of the 177 patients included in the study, 57 were Danes, one was from Iceland, 111 were immigrants, and eight were from Greenland. Responses to the questionnaires were obtained from 163 patients (92%). Four cases of unsuspected transmission were detected: one of nosocomial spread of TB, one of occupational acquisition of TB and two of transmission in an international school, leading to further contact tracing among 75 schoolchildren. These four cases were all the result of short-term contacts. In 22 cases, contact with one or more TB patient(s) was reported. In six of these, the DNA-fingerprint result revealed that the presumed contact could not be the source of infection, even though in two of the cases the known TB contact was from the household. CONCLUSION: Nation-wide DNA fingerprinting of TB isolates provides information that could not have been obtained otherwise, and contribute to the understanding of TB transmission in Danish society. In some cases the results lead to further contact tracing. Short-term contact can apparently result in transmission of TB.

Adult↗
PubMedDate not supplied

DNA fingerprint patterns of Pasteurella multocida from the same turkey farm on the same and different years.

The DNA fingerprint profiles of 126 isolants of Pasteurella multocida from 41 turkey farms in Missouri were analyzed after digestion with the restriction endonuclease HhaI and compared with their somatic antigenic type. The goal was to determine if the same isolant of P. multocida was reisolated from the the same farm during the same and consecutive years and after an interval of one or more years. Of the 37 pairs of P. multocida collected during the same year from the same turkey farms, the DNA fingerprint profiles were the same with 26 pairs (70.3%) and different with 11 pairs (29.7%). Of the 33 pairs of P. multocida collected during consecutive years from the same 22 turkey farms, 21 pairs (63.6%) were the same and 12 pairs (36.4%) were different. Of the 15 pairs of P. multocida collected with an interval of one or more years between them from the same 14 turkey farms, only four pairs (26.7%) were the same and 11 pairs (73.3%) were different. There did not appear to be any relationship between the DNA fingerprint profiles and the typing of their somatic antigens because, although 44 pairs of isolants had the same DNA fingerprint profile and somatic antigenic type, 42 pairs differed in these parameters when all pairs were combined.

Animals↗
PubMed2002

[Establishment of comprehensive fingerprint database of traditional Chinese herbs].

Quality control of traditional Chinese herbs by fingerprint is being paid close attention in these years. A database software has been designed and built to store these fingerprints and other relevant information. The use of this database is very simple and convenient for ordinary analysts who have no experience on professional database software. This software could be used under many kinds of operation systems such as Windows 95, 98, 2000 and Windows NT 4.0, 5.0. By now, nearly 30 fingerprints and their relevant information of ten kinds of traditional Chinese herbs have been input into this database, and the number of fingerprints in this base will be continuously increased in the future. This database is helpful for the analysis and quality control of traditional Chinese herbs.

Chromatography, Gas↗
PubMed2003

[Study on molecular epidemiology of Mycobacterium tuberculosis in Chinese army with PCR amplified fingerprinting methods].

OBJECTIVE: Typing of Mycobacterium tuberculosis strains and epidemiological studies in the army of southern China to provide scientific basis for prevention of pulmonary tuberculosis. METHODS: A rapid fingerprinting of M. tuberculosis strains method by polymerase chain reaction (PCR) with outward-directed primers that designed to the ends of the insertion sequence IS6110 was developed, and to analyze the relationship between the polymorphism of DNA fingerprinting and epidemiology of M. tuberculosis. RESULTS: One hundred and fifty-four M. tuberculosis detected were classified into eight types according to their characters of PCR amplified fingerprints. The main types were type I (36.4%), type II (31.8%), and type III (21.4%), while other types were less than 4 percentage. In those main type groups, patients aged 20 to 29 and 30 to 39 took up 31.8% and 27.9% respectively. For those main types, the distribution of those types in the first treated patients showed significant difference compared with that in the retreated patients, and the rate of drug-resistance was also statistically different. However, the distribution was not statistically significant to history of BCG vaccination and patients living in urban or rural area. The main drug-resistant strains were only Isoniazid-resistant or Rifampin-resistant strains, while the drug-resistant strains were 44.4%, 29.6% and 14.8% respectively in type I, type II and type III. CONCLUSION: PCR fingerprinting was a rapid, precise, sensitive, specific method to type M. tuberculosis, and could be used to study the epidemiology of tuberculosis; The prevalence of tuberculosis was primarily due to the transmission of type I, type II and type III in the army being studied from Southern China, to suggest that surveillance needs to be strengthened.

Adult↗
PubMed1992

Oligonucleotide fingerprinting as a means to identify and survey long-term cultured B cell hybridomas and T cell lines.

Common problems encountered during cell culture are cross-contamination, instability, and inadvertent exchange of cells. Here we report on the application of oligonucleotide fingerprinting as a simple and efficient method to screen hybridomas and T cell lines. Among the fingerprint probes tested, the simple repetitive oligonucleotide (CAC)5/(GTG)5 proved to be most useful for obtaining many fragments specific for each cell line. Because of variable loss of chromosomes, cloned hybridoma cells from one fusion exhibit different fingerprint patterns. Thus, antibody-secreting B cell hybridomas can be distinguished easily even when they originate from the same fusion. Furthermore, we were able to monitor the genomic integrity of myelomas and hybridomas over a period of more than 2 years, thereby highlighting long-term stability. Another application of the method is the control of T cell lines requiring irradiated or mitomycin-treated feeder cells for continuous growth or cloning. There cell lines are always threatened to be overgrown by feeder cells that have escaped from the lethal pretreatment. T cell clones of one individual, known to display differently rearranged T cell receptors, did not show differences in their fingerprint patterns. However, in EBV-transformed cloned B cells, slight differences between clones from the same donor were identified.

B-Lymphocytes↗
PubMed1992

Differences in DNA fingerprints of continuous leukemia-lymphoma cell lines from different sources.

The genetic stability of human cell lines in long-term culture has been tested by DNA fingerprinting a panel of 31 different continuous cell lines from patients with leukemias or lymphomas. Duplicates of the same cell line obtained from different sources, subclones of cell lines, and samples of cell lines at different passage levels were studied. In most cases the fingerprints of duplicates of the same cell line remained perfectly preserved even after long-time passaging. However, in five cases there were notable differences between individual fragments of corresponding fingerprints. We have found four cases of mislabeled and/or cross-contaminated cell lines so far. Taken together, our results indicate that DNA fingerprinting qualifies as a very reliable means of cell line identification which allows the detection of mislabelling or contamination and of genetic variation among subclones.

Base Sequence↗
PubMed2003

The application of PCR fingerprinting to the differentiation of Yersinia enterocolitica strains isolated from humans and pigs.

The distribution of different genotypes of Yersinia enterocolitica strains recovered from humans and from healthy pigs was investigated using PCR fingerprinting. The thirty six strains of Y. enterocolitica from humans, thirty five strains from pigs and Y. enterocolitica ATCC 9610 strain were included in this study. The tested strains of Y. enterocolitica belonged to O3 and O9 serogroups. The PCR fingerprinting using EAE5 primer (5' CTT AAT CTC AGT AAT GCT GGC CTT GG) made it possible to form five groups among the tested Y. enterocolitica strains. Two groups were very numerously represented by the tested strains. The thirty of Y. enterocolitica O3 strains from humans (thirty one of tested) and eighteen of Y. enterocolitica O3 strains from pigs (twenty of tested) belonged to one group. This group also included Y. enterocolitica ATCC9610 strain and four Y. enterocolitica O9 strains from pigs. All investigated Y. enterocolitica O9 strains from humans and the majority of Y. enterocolitica O9 strains isolated from pigs created a second, numerous group. The third genotype was created by two strains O9 from pigs, and the remaining two strains, isolated from pigs, belonging to O3 and O9 serogroups showed different binding patterns revealed by gel electrophoresis and created two other genotypes. The tested Y. enterocolitica strains which were isolated from humans formed only two groups but Y. enterocolitica strains isolated from pigs were found in five groups but such as the Y. enterocolitica strains from humans, the majority of strains from pigs were in first and second group. The Y. enterocolitica O3 strains regardless of their origin mostly represented the same PCR fingerprinting profile. The tested Y. enterocolitica O9 strains were more genetically diverse and represented four PCR fingerprinting profiles.

Animals↗
PubMed1992

[The study of DNA fingerprint using probe directly labelled with horseradish peroxidase and enhanced chemiluminescence in forensic science application].

We describe here the technique of DNA probe directly labelled with horseradish peroxidase (HRP) and enhanced chemiluminescence (ECL) detection for determining DNA fingerprints. The patterns of DNA fingerprint were clear and with high distinction. The sensitivity of DNA fingerprint was 0.8 microgram, similar to the method of 32P labelling. The DNA fingerprints of 150 unrelated individuals were studied and showed that the probability of chance association of DNA fragments between random individuals was 3.7 x 10(-14). The method is stable, simple and quick, and can be used in individual identification and paternity test.

DNA Fingerprinting↗
PubMed2005

In silico simulation of fingerprinting techniques based on double endonuclease digestion of genomic DNA.

UNLABELLED: We have developed an online generic tool for simulation of fingerprinting techniques based on the double endonuclease digestion of DNA. This tool allows modelling and modifications of already existing techniques, as well as new theoretical approaches not yet tried in the lab. It allows the use of any combination of recognition patterns and discrimination of end types yielded by restriction with non palindromic recognition sizes. Re-creation of experimental conditions in silico saves time and reduces laboratory costs. This tool allows simulation of Amplified Fragment Length Polymorphism (AFLP-PCR), Subtracted Restriction Fingerprinting (SRF), and additional novel fingerprinting techniques. Simulation may be performed against custom sequences uploaded to the server, or against all sequenced bacterial genomes. Different endonuclease types may be selected from a list, or a recognition sequence may be introduced in the form. After double digestion of DNA, four fragment types are yielded, and the program allows their customised selection. Selective nucleotides may be used in the experiment. Scripts for specific simulation of AFLP-PCR and SRF techniques are available, and both include a suggestion tool for the selection of endonucleases. This is the first program available for the simulation of SRF fingerprinting. AVAILABILITY: This free online tool is available at http://www.in-silico.com/DDF/.

DNA↗
PubMedDate not supplied

DNA fingerprinting by ERIC-PCR for comparing Listeria spp. strains isolated from different sources in San Luis, Argentina.

In this study, a total of 24 Listeria spp. strains were analyzed. Twenty-two isolates were obtained in San Luis (Argentina) from human, animal, and food samples. Two types of strains, Listeria monocytogenes CLIP 22762 and Listeria innocua CLIP 74915, were included as reference strains. All isolates were biochemically identified and characterized by serotyping, phage typing, and amplification of the flaA gene by polymerase chain reaction (PCR). Repetitive intergenic consensus (ERIC) sequence-based PCR was used to generate DNA fingerprints. On the basis of ERIC-PCR fingerprints, Listeria spp. strains were divided into three major clusters matching origin of isolation. ERIC-PCR fingerprints of human and animal isolates were different from those of food isolates. In addition, groups I and II included ten L. monocytogenes strains, and only one Listeria seeligeri strain. Group III included nine L. innocua strains and four L. monocytogenes strains. Computer evaluation of ERIC-PCR fingerprints allowed discrimination between the tested serotypes 1/2b, 4b, 6a, and 6b within each major cluster. The index of discrimination calculated was 0.94. This study suggests that the ERIC-PCR technique provides an alternative method for the identification of Listeria species and the discrimination of strains within one species.

Animals↗
PubMed1995

Simultaneous DNA 'fingerprinting', diagnosis of sex and single-gene defect status from single cells.

Sex and cystic fibrosis status have been previously diagnosed separately at the single cell level. We have developed a sensitive, reliable, accurate and rapid (within 5-6 h) system for the simultaneous diagnosis of sex, cystic fibrosis and a DNA 'fingerprint' within a single reaction from a variety of single cells. As contamination cannot be totally excluded, particularly at the single cell level, DNA 'fingerprinting' can be used to assess the risk of contamination. High sensitivity with single cells is combined with very high specificity (estimated matching probability of 10(-7)-10(-8)), allowing the source of the amplified cell to be identified with a very high degree of probability. Fluorescent primers were multiplexed for six tetranucleotide microsatellite sequences to determine the DNA fingerprint; the amelogenin gene was used to diagnose sex, and primers for the CFTR region were used to determine cystic fibrosis (CF) status. Analysis of the fluorescent product was undertaken using an automated DNA sequencer with Genescan software. This technique has many applications such as prenatal and preimplantation diagnosis, forensic identification of small or degraded samples, and detection of contamination sources. DNA fingerprints of single haploid spermatozoa and other cells can be assessed, so ensuring the detection of both diploid and haploid contamination during preimplantation diagnosis.

Cystic Fibrosis↗
PubMed1993

Parentage determination on placental tissues through deoxyribonucleic acid fingerprints.

Investigation of genomic polymorphisms detected by a mini-satellite "Myo" probe gives distinct and different Deoxyribonucleic Acid (DNA) fingerprints of chorionic villus and decidual membrane in the same placenta. The chorionic villus, which is regarded as the extra-embryonal tissue, represents the essential embryonal DNA fingerprint pattern, while the decidual membrane reveals the maternal pattern. A comparison between the DNA fingerprints from the chorionic villus and from the blood sample of the suspected father provides the possibility of setting a paternity test in the early gestational stage. Twenty-eight cases of paternity test on aborted placental tissues by DNA fingerprints were analysed.

Abortion, Induced↗