Comparative evaluation of a new molecular method for typing Staphylococcus epidermidis.

DNA samples from different inbred strains and F1-hybrids, from two outbred strains and from transgenic animals of the species Mus musculus were tested according to the "DNA fingerprint" technique (Jeffreys et al., 1985) using the B.E.S.T.-probe MZ 1.3 (Fa. Biotest, Frankfurt) and the restriction endonuclease Hinf I. In addition, the same method was applied to two cell types i.e. BALB/3T3 clone A 31 and 3T3 B-SV40. The DNA fingerprinting technique with probe MZ 1.3 proved to be a reliable method for genetic monitoring of different strains of mice. All inbred strains tested as well as their substrains could be identified and distinguished from each other without any doubt. Congenic and transgenic individuals, however, were identical with their background strains. After several in vitro passages, cells from cultures showed the similar DNA configuration as the donor strains. Within outbred strains, it was possible to quantify heterozygosity by the configuration of the DNA-patterns. These results suggest that it might be appropriate to replace the mathematically estimated inbreeding coefficient by so-called identity-coefficient (IK), which would depend on the probe and the restriction endonuclease used (e.g. IKMZ 1.3/Hinf I). Using the MZ 1.3 probe, the DNA fingerprint technique allows a unique genetic identification of different strains of mice and offers, furthermore, the possibility to use a colour kit rather than the usual P-32 marker.

DNA fingerprinting of benign and malignant breast lesions

<p>The DNA fingerprinting techniques, 16S-restriction fragment length polymorphism (16S-RFLP), ribosomal intergenic spacer analysis (RISA) and repetitive extragenic palindrome PCR (Rep-PCR), were used for analyzing the bacterial communities of seven pulp and paper wastewater treatment systems. All three methods generate DNA fingerprints that can be compared using the computerassisted program, Gelcompar©. Community similarity coefficients were based on quantitative determinations of both the positions of the DNAbands and the band intensities in order to compare the relative differences in the populations. Unique 16S-RFLP DNA fingerprints were observed for each mill suggesting that individual mills contained phylogenetically different communities. However this method was not sensitive enough to detect differences within a mill treatment system from different locations or from different sampling times. The RISA method, which generated more complex fingerprints than 16S-RFLP, could, for some mills, discern differences between samples. The Rep-PCR technique, however, showed the highest degree of resolution and produced not only distinct patterns for each mill but also distinct fingerprints for the temporal and spatial samples from some of the treatment systems. The sensitivity of this method might potentially be used to monitor the stability of the bacterial community within a secondary treatment system.</p> <p><br></p>

Linearly arranged chemical structure in chromosome is known as DNA. It is a double helix made up of two strands of genetic material spiraled around each other. Each strand has a sequence of bases. There are four types of basis namely adenine, guanine, cytosine and thiamine which are very unique to each individual just like their actual fingerprint. The nitrogen base adenine always binds with thymine and cytosine also always binds with guanine. Thus the DNA profiling unique to each individual is collectively known as DNA fingerprinting. DNA determines individuality or uniqueness of the each human being except in uniovular twins. The chances of complete similarity are one in 30 billion to 300 billion i.e. half the population of world. The technique of DNA fingerprinting was first developed by Dr. Alec Jeffery’s from Britain in 1984. He discovered a minisatellite region close to the human myoglobin gene. He isolated this sequence and used it as a probe to investigate human DNA. He found that the minisatellite probe result was a complex band pattern for each individual. In India, initially it was done at CCMB, Hyderabad by Dr. Lalji Singh. Now there are various centers where DNA fingerprinting is carried out. In Maharashtra it is carried out at Sate Forensic Science Laboratory, Vidya Nagar, Kalina, Mumbai – 400 098 (Phone 022–26670755). Using this technique FBI formally concluded the participation of Mr. Bill Clinton in Monica Lewyninskey case. In India more than 79 cases have been solved by using this technique including important case of Dhanu and Shivarasan alleged assailant of Late Priminister Shr. Rajiv Gandhi, Tandori case, Madhumati murder case etc.

Seasonal and spatial variation in the genotypic richness of 3480 Escherichia coli isolates obtained from the Yeongsan River basin in South Korea was investigated by using the horizontal fluorophore-enhanced rep-PCR (HFERP) DNA fingerprinting technique. The relationship between 60 E. coli isolates from each of 58 freshwater samples was determined by using multidimensional scaling (MDS) analysis and self-organized maps (SOMs). The MDS analysis, done based on HFERP DNA fingerprints, showed that E. coli isolates obtained in October through December clustered tightly, while those obtained in other sampling periods were more genetically diverse. However, site-specific E. coli genotypes were not observed. SOMs analysis, done using the 10 most frequently isolated E. coli genotypes, showed the occurrence of season-specific E. coli genotypes and the main SOMs clusters were most influenced by temperature, strain diversity and biochemical oxygen demand. Diversity among E. coli genotypes tended to decrease as water temperature decreased, and the numbers of E. coli genotypes observed in urban area were greater, more diverse and less dependent on water temperature than those obtained from agricultural areas. Taken together, our findings indicate that that an ecological approach needs to be considered in order to obtain a better understanding of E. coli community dynamics in the environment and that SOMs analysis is useful to visualize the multidimensional dependent variables that are influencing the types and dynamics of specific E. coli genotypes in the environment.

Objective To study the epidemiology of clinical mastitis caused by Escherichia coli and Staphylococcus aureus by differentiating isolates with DNA fingerprinting techniques, using polymerase chain reaction. Design Milk samples were collected from cases of clinical mastitis in dairy cows. Escherichia coli and S aureus isolates from these cases were compared within and between cows and herds. Sample Population Seven dairy herds with an average bulk milk somatic cell count < 150,000/ml, and incidence of cows with clinical mastitis of > 25%/y. Procedure Chromosomal DNA was isolated from E coli and S aureus strains isolated from cases of clinical mastitis, and amplified by polymerase chain reaction, using enterobacterial repetitive intergenic consensus primers for E coli and a random amplified polymorphic DNA primer for S aureus. Escherichia coli and S aureus strains were identified and differentiated, using their DNA polymorphism pattern. Results Multiple E coli genotypes were found in each of the herds. Persistent infections with E coli were sporadic. Only a limited number of different S aureus genotypes was found in each of the herds studied. Recurrent cases of S aureus mastitis were found in 25% of quarters with clinical S aureus mastitis. Comparing S aureus isolates from different herds indicated that 1 S aureus genotype was most prevalent. Conclusions Because different quarters were infected with different genotypes, it was concluded that E coli is an environmental pathogen, and does not generally spread from quarter to quarter. The hypothesis that S aureus mastitis is a contagious disease, spreading from infected to uninfected quarters, could not be rejected. (Am J Vet Res 1996;57:39-42)

Samples of the stingless bee Trigona pagdeni Schwarz were collected from north, northeast, central and peninsular Thailand. Genetic variation and population structure were investigated using a DNA fingerprinting technique, TE-AFLP, and Analysis of Molecular Variance (AMOVA). We found high levels of genetic variation among individuals in all populations, but mean expected heterozygosity was highest in the Northeast. We used AMOVA to calculate φPT (analogous to Wright's φST for codominant markers) to compare genetic differentiation among populations. This revealed significant genetic differentiation among the four populations (φPT = 0.18, P = 0.01). We also detected differentiation (φPT = 0.13, P = 0.001) between samples collected north and south of the Kra ecotone, a biogeographical zone of transition between seasonal evergreen and mixed moist deciduous forests. However the greatest differentiation was detected between samples from the northeast and the other locations combined (φPT = 0.21, P = 0.001). This method can be applied to the study of population structure in T. pagdeni and other stingless bees, and may provide a useful tool for management and conservation of this species. Meliponini / Trigona / DNA fingerprinting / AFLP / geographic variation

DNA fingerprinting techniques for the analysis of genetic and epigenetic alterations in colorectal cancer

New Avenues in Drug Discovery and Bioactive Natural Products is the second volume of the Natural Medicine book series. It is devoted to current research in drug discovery from natural sources. The volume features 13 chapters that cover modern analytical and scientific approaches. The book starts with chapters on advanced analytical and research techniques, such as genomic mining, quality control of herbal drugs, DNA fingerprinting, high-throughput screening, molecular docking and extraction techniques. The contributors provide a summary of challenges for researchers and commercial applications where possible. The book also features chapters dedicated to specific medicinal agents that target a disease (glycosides, SARS-CoV2 spike protein inhibitors, and andrographolides. The collection of important research topics in natural product chemistry aims to help the scholars and researchers in the scientific community that are involved in the extraction and development of new medicines.

Genetic diversity of Gram-negative, proteolytic, psychrotrophic bacteria isolated from refrigerated raw milk

The Karasu River (Sinop, Turkey) was sampled for bacteriological analyses and investigation of antibiotic resistance at the bacterial isolates. In the study, the physical properties and numbers of total aerobic (TA), total coliform (TC), faecal coliform (FC), faecal Streptococci (FS) and Escherichia coli in water samples were investigated. E. coli strains were subjected to antibiotic susceptibility testing, antibiotic resistant genes, virulence genes and plasmid DNA analysis and fingerprinting techniques. The bacteriological analysis results showed that the difference between TA, TC, FC and FS numbers (p > 0.05) was not significant for a total of four stations. The Multiple Antibiotic Resistance Index (MARI) values of tested strains were found in range from 0.4 to 0.7. The antibiotic resistance genes of strains; tetA (33.9%), blacym‐2 (12.5%) and cat1 (16.7%) were screened by PCR method. Out of a total of 57 E. coli isolates, 11 E. coli strains were positive for the eae (19.6%) gene, representing atypical EPEC, whilst only one strain was positive for the eae and stx1 (1.7%) gene, representing STEC. Present study has shown that the aquatic environments can be reservoirs of pathogenic E. coli strains. The study also indicated that PCR‐based techniques might especially be helpful in the rapid identification of multidrug‐resistance analysis and virulence genes of strains. Karasu stream, which is important in Sinop province, Turkey, was investigated for the first time in this study in terms of a molecular microbiological perspective.

The molecular technique, enterobacterial repetitive intergenic consensus (ERIC)-polymerase chain reaction (PCR) produces genomic DNA fingerprint that discriminate bacterial species and strains. This technique was applied to the characterization of Listeria monocytogenes, an important food-borne pathogen implicated in numerous cases of listeriosis. The ERIC-PCR resulted in distinct DNA fingerprinting patterns of all L. monocytogene serotypes and Listeria species. Analysis of the genomic DNA fingerprints was accomplished using capillary electrophoresis (CE), an alternative technique to the conventional agarose gel method. The optimization of CE conditions (electrokinetic injection, applied voltage) resulted in the resolution of amplified DNA fragments up to 1000 bp. Comparisons of electropherograms provided genomic fingerprint templates which could be further used for supplementary information. The ERIC-PCR method coupled to CE provides a rapid technique in differentiating bacterial spp., and may contribute relevant information in food-borne outbreak studies.

Dormant cuttings from grapevine source areas in three Australian states were indexed for the presence of Agrobactenrium vitis, the causal agent of crown gall disease of grapevine. The bacterium was present at varying levels in both rootstock and scion material from New South Wales, Victoria, and Western Australia. A genomic DNA fingerprinting technique was used to compare these strains of A. vitis with strains previously isolated from South Australian propagation material and with strains from the United States of America, particularly strains from the University of California at Davis where a number of Australian grape cultivars originated. Nine distinct DNA fingerprint types were found, demonstrating that A, vitis is a genetically heterogeneous species. There were no DNA fingerprint types in common between Australian states, suggesting that little or no transfer of A. vitis between states has occurred. Two Californian strains were indistinguishable, when analysed by DNA fingerprinting, from strains isolated over a 14 year period in South Australia. This finding is strong circumstantial evidence that A. vitis was introduced to South Australia on grapevine cuttings imported from California. Given the fact that A. vitis is probably not indigenous to Australia, it is possible that the different strains found in each state were the result of independent introductions. Although grapevines from all Australian states tested did contain A. vitis, the existence of unique strains in each state means that the restrictions on movement of grapevine material between states should be maintained, and that quarantine authorities should avoid introducing further strains of the bacterium.

A collection of epidemiologically unrelated verocytotoxin (VT)-producing Escherichia coli (VTEC) strains of serogroup O111 isolated from human patients and cattle with diarrhoeal disease in five different countries were characterised by determination of their VT genotypes, the presence of other virulence factors such as the intimin-coding eae gene and the enterohaemorrhagic E. coli (EHEC) plasmid, and their antibiotic susceptibility patterns. The genetic relatedness among isolates was evaluated by genomic DNA fingerprinting techniques such as restriction fragment length polymorphism analysis of ribosomal RNA genes (ribotyping) and pulsed-field gel electrophoresis. The results indicated that the VTEC O111 examined belong to two distinct clonal lineages. The first group was constituted mainly of non-motile, eae-positive, EHEC plasmid-positive isolates from both man and cattle. The second lineage was represented by an O111:H2 epidemic strain, isolated during an outbreak of haemolytic uraemic syndrome in France and exhibiting an unusual combination of virulence factors: VT production and aggregative adhesion to HEp-2 cells associated with an enteroaggregative E. coli (EAEC) plasmid.

Forensic DNA analysis has revolutionized criminal investigations by providing valuable insights into identifying perpetrators, exonerating the innocent and unravelling mysteries of the past. Various molecular biology techniques, such as Short Tandem Repeat (STR) analysis, Single Nucleotide Polymorphism (SNP) profiling, mitochondrial DNA (mtDNA) sequencing, epigenetics and DNA methylation analysis, have played crucial roles in forensic investigations. While these techniques offer exceptional sensitivity and specificity, they also come with unique challenges. This review explores the current approaches of forensic DNA analysis, highlighting each technique. STR analysis, regarded as the gold standard in forensic DNA profiling, provides high discrimination power. However, it is limited in its ability to analyze degraded or mixed samples. On the other hand, SNP profiling offers advantages in analyzing highly degraded DNA samples but lacks the discriminatory power of STRs. Meanwhile, mtDNA analysis, particularly useful in cases involving compromised nuclear DNA, presents challenges due to its maternal inheritance pattern and lower discriminatory power. Furthermore, explore the exciting realm of epigenetics and the analysis of DNA methylation in forensic investigations. Epigenetic markers offer insights into gene expression patterns influenced by environmental factors, potentially aiding in the determination of tissue origin and chronological age estimation. DNA methylation analysis holds promise in forensic applications, providing additional layers of information for identity verification and tissue differentiation. Despite these advancements, several challenges persist in forensic DNA analysis, including the interpretation of complex DNA mixtures, standardization of methodologies, ethical considerations, and privacy concerns associated with the use of genetic information. Moreover, the integration of multi-omics data and machine learning approaches presents both opportunities and challenges in enhancing the accuracy and reliability of forensic DNA analysis. Looking ahead, future directions in molecular biology research for forensic DNA analysis involve the development of novel techniques with increased sensitivity, scalability, and robustness. By addressing these challenges and embracing emerging technologies, the field of forensic DNA analysis is poised for further advancements, offering enhanced capabilities in criminal justice and humanitarian efforts.