Random Amplified Polymorphic DNA-polymerase Chain Reaction Research Articles

Coexistence of armA and genes encoding aminoglycoside-modifying enzymes in Acinetobacter baumannii

A total number of 104 Acinetobacter baumannii isolates obtained from a teaching hospital were studied for the genetic basis of their resistance to aminoglycosides. The rate of aminoglycoside-resistant A. baumannii was 76.9% (80/104), and 66.25% (53/80) aminoglycoside-resistant isolates were high-level resistant to either aminoglycosides including amikacin, gentamicin and tobramycin (MIC≥512μg/ml). Genes encoding aminoglycoside-modifying enzymes including ant(2″)-Ia, aac(6′)-Ib, aph(3′)-Ia, aac(3)-Iaand aph(3′)-VI, and 16S ribosomal RNA (rRNA) methylase armA, rmtA, rmtB, rmtC, rmtD,rmtE and npmA were detected by polymerase chain reaction (PCR), armA gene waspresent in 50% (52/104) of A. baumannii isolates with high-level resistance to aminoglycosides (MICs≥512 μg/ml). No rmtA, rmtB, rmtC, rmtD, rmtE or npmA genes were found. The rates of ant(2″)-Ia, aac(6′)-Ib, aph(3′)-Ia, aac(3)-Ia and aph(3′)-VI were 68.25%,20%,70%,47.5% and 1.25%, respectively. armA, ant(2″)-Ia and aph(3')-Ia genes coexisted in 62.5%(50/80) of aminoglycoside-resistant isolates, armA, ant(2″)-Ia and aac(6′)-Ib were commonly positive in same isolates. The presence of aminoglycoside-modifying enzymes was responsible for the moderate level resistance to aminoglycosides in A. baumannii, whereas armA was responsible for the high-level resistance to aminoglycosides. Aminoglycoside-resistant A. baumannii isolates were classified into genotypes by Random Amplified Polymorphic DNA PCR (RAPD-PCR), pattern A was almost throughout wards of the hospital. In summary, most A. baumannii is resistant to aminoglycosides, in the hospital coexistence of armA and genes encoding aminoglycoside-modifying enzymes are the main reasons. Key words: Acinetobacter baumannii, aminoglycoside-resistant, 16S rRNA methylase, aminoglycoside-modifying enzymes, RAPD-PCR

Toxicity of Bacillus thuringiensis d-exotoxins and b-endotoxins to Drosophila melanogaster, Ephestia kuhniella and human erythrocytes

A total of 73 Bacillus thuringiensis (Bt) strains were screened for the presence of non-hemolytic insecticidal β -exotoxin-free δ -endotoxins. Out of them, 45 Bt strains produced δ -endotoxins with specific insecticidal activity against Drosophila melanogaster and/or Ephestia kuhniella larvae. The thermostable β -exotoxin was observed only in 15 Bt strains and appeared to exhibit dual non-specific insecticidal activity against both D. melanogaster and E. kuhniella larvae and showed in vitro hemolysis for human erythrocytes. It was found that β -exotoxin was produced by Bt strains belonging to five serovars ( israelensis , kenyae , kurstaki , pakistani , and tohokuensis ) and two non-serotypable strains. This result suggests that β-exotoxin production is a strain-specific property rather than a serovar-specific property. To our knowledge, this is the first study that demonstrates β -exotoxins production association with Bt strains belonging to serovars israelensis, pakistani, and tohokuensis. The plasmid DNA profiles of some β -exotoxin producing Bt strains shared large plasmid patterns which may have the common β -exotoxin regulatory gene(s). It was found that 16 local Bt strains, 15 of which belonged to five serovars ( aizawai , israelensis , kurstaki , morrisoni , and pakistani ) and one was autoagglutinated strain, produced non-hemolytic insecticidal β -exotoxin-free δ -endotoxins. Based on random amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR), the genotypic relatedness among these 16 Bt strains was investigated. The strains were grouped into two clusters. Bt strains within serovars israelensis were grouped in two subclusters, Bt strains within serovars aizawai were genomically homogeneous and clustered together, while the other serovars were grouped together in one subcluster. The autoagglutinated strain was clustered within serovar israelensis . Thus, these δ -endotoxins can be developed for the use in Bt-based insecticidal preparations. Keywords: Endotoxin, exotoxin, hemolytic, serovar, thuringiensis

Genetic Variability of the Tomato Leaf Miner (Tuta absoluta Meirick; Lepidoptera: Gelechiidae), in Tunisia, Inferred from RAPD-PCR

The tomato leaf miner Tuta absoluta Meyrick has invaded tomato ( Solanum lycopersicum L.) crop in Tunisia since 2008 and is representing today a major threat to the production of this crop. In this study, we used the Randomly Amplified Polymorphic DNA-Polymerase Chain Reaction (RAPD-PCR) technology to assess the genetic variability within and among seven populations of T. absoluta, collected on tomato from different regions in Tunisia. Using five RAPD-PCR primers and 108 individuals, 140 polymorphic fragments were recorded. From 335 different RAPD phenotypes generated, 71 were redundant and 264 unique to a specific population. The genetic structure of T. absoluta was investigated using analysis of molecular variance (AMOVA), genetic distances (Fst) and multidimensional scaling (MDS). We detected a high genetic diversity within and among populations in conjunction with a significant differentiation between populations, suggesting that different founder genotypes would have been responsible of the introduction of T. absoluta in Tunisia. The presence of overlapping phenotypes probably indicates migration events between populations, mainly through infested plant material carried by humans.

Characterization of Vibrio alginolyticus and Vibrio parahaemolyticus isolated from Penaeus monodon: Antimicrobial resistance, plasmid profiles and random amplification of polymorphic DNA analysis

Vibrio alginolyticus and Vibrio parahaemolyticus are important pathogens which cause serious episode to shrimp. We characterize the suitability of randomly amplified polymorphic DNA (RAPD), and plasmid profiling for rapid molecular characterization of pathogenic bacteria for epidemiological and antibiogram study to find out the relationship between plasmids and bacterial sensitivity to antibiotics. Nine primers, belonging to OPA and OPY series were selected for the RAPD study. Primer OPA3 and OPY2 amplified 3 to 7 bands ranging from 0.306 to 3.94 Kb in V. alginolyticus. In V. parahaemolyticus, primer OPA 3 and OPY 2 amplified 3 to 7 bands of molecular wt 0.306 to 4.11 Kb. V. alginolyticuspossess only one plasmid each with molecular weight 10. 23 Kb. V. parahaemolyticusstrains contained 1 to 3 plasmids with molecular weight 7.12 to 10.23 Kb. The strains were mostly resistant to Ampicillin and Cefuroxime. Among the strains 90 and 70% were resistant Bacitracin and Co-trimazole, respectively. All strains of V. alginolyticus and V. parahaemolyticus were sensitive to Amoxycillin, Chlorotetracycline and Ciprofloxacin. Unique distinct amplicons generated by RAPD polymerase chain reaction (PCR) and plasmid profiling will be used for several diagnostic markers for identification of these pathogenic microorganisms and in epidemiological study. Key words: Vibrio parahaemolyticus, Vibrio alginolyticus, RAPD, antibiogram, plasmid profiling.

Preliminary molecular analysis of the genetic diversity of some Atriplex species present in the Northeast of Algerian

In the arid and semi arid areas, salt bush ( Atriplex ) represents an important forage resource. The characterization of the genetic diversity of these species is useful for their classification, their conservation and their improvement. In this context, we used the random amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR) technique to test the genotypes of three different species of Atriplex: Atriplex halimus, Atriplex canescens and Atriplex nummularia . The Obtained results showed 319 amplified bands which were mostly polymorphic. The analysis of the results generated four different groups where the genotypes of two introduced species (A. canescens and A. nummularia) were clustered into two independent groups. However, genotypes of the autochthonous species ( A. halimus ) was subdivided into two different independent groups where we noted that individuals of one of these groups are genetically the nearest between them, by report, to the existing others within the same group. In general, results reveal inter and intra high level of genetic diversity in the Atriplex studied species. Key words: Atriplex, Chenopodiaceae, genetic diversity, molecular analysis, steppe, Algeria.

Selection of Dominant NSLAB from a Mature Traditional Cheese According to their Technological Properties and in vitro Intestinal Challenges

Isolates (47) of lactobacilli from 5 different productions of Melichloro cheese were examined for potential use as adjunct cultures. The sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) of whole-cell proteins classified 29 isolates as L. paraplantarum and 18 as L. paracasei subsp. paracasei. Randomly amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR) analysis differentiated the L. paraplantarum and L. paracasei subsp. paracasei isolates at strain level and both, RAPD analysis and whole-cell protein profiling provided useful information about the diversity of nonstarter lactic acid bacteria (NSLAB) in the different cheese productions. The isolates were slow acidifiers and about 70% of them degraded, preferentially α(s)-casein. The amounts of amino acids accumulated in the milk increased with the incubation time. A similar enzyme profile was exhibited by strains of both species, except for α-mannosidase and α-fucosidase, which were not detected in the L. paracasei subsp. paracasei strains. All strains grew in the presence of bile at 0.3% and the majority was able to withstand pH 2.5 and pancreatin at 0.1%. Moreover, all strains reduced cholesterol in vitro, with higher removal ability recorded for strains of L. paraplantarum. A narrow spectrum of antibacterial activity was recorded for 88% of the strains. Selected isolates with appropriate technological and interesting in vitro intestinal challenges could be used as adjuncts and deserve further studies.

Influence of inoculation time of an autochthonous selected malolactic bacterium on volatile and sensory profile of Tempranillo and Merlot wines

A study was carried out to determine the effect of the inoculation time of the lactic acid bacteria (LAB) on the kinetic of vinification and on chemical and sensory characteristics of Tempranillo and Merlot wines. Traditional vinifications, with LAB inoculated after completion of AF, were compared with vinifications where yeast and bacteria were co-inoculated. Two commercial yeast strains and an autochthonous Oenococcus oeni strain (C22L9) previously identified and selected at our laboratory were used. Monitoring of alcoholic and malolactic fermentations was carried out by yeast and lactic acid bacteria counts and by measuring contents of glucose+fructose, malic acid and lactic acid. The implantation rate of O. oeni C22L9 was calculated by typing isolates obtained from count plates using the RAPD-PCR (Randomly Amplified Polymorphic DNA-Polymerase Chain Reaction) technique. Wines were chemically characterised and analysed for biogenic amine and volatile compound contents. A sensory analysis, consisting in a descriptive and a triangular test was also carried out. Results from this study showed that for both grape varieties, the concurrent yeast/bacteria inoculation of musts produced a significant reduction in duration of the process, without a pronounced degradation of malic acid during AF, nor an excessive increase in volatile acidity. Biogenic amine content was also lower in wines produced by co-inoculation. Important differences in volatile compound contents were observed, although there was little impact on the sensorial profile of wines. These results suggest that co-inoculation using O. oeni C22L9 is a worthwhile alternative compared to traditional post AF inoculation for Tempranillo and Merlot winemaking.

Molecular Characterization of Leishmania Infection in Sand flies From Sistan Va Baluchistan Province, Southeastern Iran

: Background: Cutaneous leishmaniasis (CL) is a zoonotic disease that is caused by various species of the genus Leishmania. The disease is considered a major health problem in different areas of Iran and is an endemic disease in rural areas of Mirjaveh, Chabahar, and Konarak Counties, Sistan Va Baluchistan Province.Objectives: The aim of this study was to identify Leishmania species that was isolated from potential sand fly vectors by molecular analysis in Chabahar County.Materials and Methods: To collect Sand flies, sticky traps were placed at the entrance of rodents burrows in Dashtiyari division of Chabahar County, where CL is endemic. Freshly collected Sand flies were identified with regard to species, dissected in normal saline using binocular, and examined for leptomonads under a microscope. Leptomonads from the Sand flies were used to inoculate the base of Balb/c mice tails subcutaneously; after an incubation period and the development of lesions, the parasites were transferred to NNN + LIT medium culture. The harvested Leishmania parasites were subjected to DNA extraction and analyzed by random amplified polymorphic DNA polymerase chain reaction (RAPD-PCR).Results: DNA from Leishmania species from Phlebotomus papatasi and P. salehi Sand flies produced distinctive patterns of bands of L. major with all primers. However, the products at approximately 2100 bp and 800 bp that were amplified with primer 329 were stable and reproducible in all assays. This is the first report on the isolation and identification of L. major in P. salehi from Iran and P. papatasi from Sistan va Baluchistan.Conclusions: The study shows that P. papatasi and P. salehi Sand flies play a major role in the maintenance and transmission of disease to humans in this area. Implication for health policy/practice/research/medical education:Identification of P. papatasi and P. salehi Sand flies as the insect vectors as well as the Leishmania species they harbor would be the key elements in understanding the epidemiology of leishmaniasis and its control in Chabahar County. Please cite this paper as:Kassiri H, Naddaf SR, Mohebali M, Javadian EA. Molecular Characterization of Leishmania Infection in Sand flies From Sistan Va Baluchistan Province, Southeastern Iran. Jundishapur J Microbiol.2012;5(2):430-1. DOI: 10.5812/jjm.3523 Copyright © 2012 Kowsar Corp. All rights reserved.

Prevalence, molecular fingerprinting and drug resistance profile of enterovirulent Escherichia coli isolates from free-ranging yaks of Tawang district, Arunachal Pradesh, India

Of 273 samples (rectal swab) collected from free-ranging yaks of Tawang district, Arunachal Pradesh, 42 Shiga toxin-producing Escherichia coli (STEC), six enteropathogenic E. coli (EPEC) and 27 enterotoxigenic E. coli (ETEC) strains were isolated. All the STEC and EPEC strains were further investigated for respective stx variants (for STEC only) and additional putative virulence factors. The 27 ETEC strains were also screened for characteristic enterotoxin gene(s) and colonization factors. Occurrence of ETEC was significantly (p < 0.05) higher in the diarrheic yaks and yaks of less than 1 year of age. Majority of enterovirulent E. coli isolates were resistant to amikacin, azithromycin, chloramphenicol, colistin, doxycycline, furazolidone, nalidixic acid, nitrofurantoin, streptomycin and tetracycline. Dendrogram, constructed with molecular fingerprinting profiles obtained from RAPD (Randomly Amplified Polymorphic DNA) and ERIC (Enterobacterial Repetitive Intergenic Consensus) PCR, placed the isolates in different clusters irrespective of their serotypes, virulence gene and drug resistance pattern. Collectively, the study indicates that yaks, being a potential reservoir of multidrug resistant STEC and EPEC, may represent significant risk to public health in this region. Higher recovery of ETEC isolates from yaks with diarrhea points out that ETEC may be a major determinant for repeated occurrence of diarrhea in yaks.

Genetic Variability and Changes of Elemental Concentrations in Cells of Tetrix tenuicornis (Orthoptera: Tetrigidae) from Polluted and Unpolluted Areas

Genetic variability between populations of the orthopteran insect Tetrix tenuicornis, collected from six locations in Poland, was assayed by using the random amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR) method. The results show that insects in a population from metal polluted areas in Bolesław have reduced genetic variability in contrast to five other populations located in unpolluted areas. The insects from polluted sites also showed significant changes in elemental concentrations in nerve and muscle cells, measured by X-ray spectroscopy, when compared to insects of the same species collected from unpolluted sites.

Genetic diversity and identification of some Turkish cotton genotypes (Gossypium hirsutum L.) by RAPD-PCR analysis

Cotton (Gossypium hirsutum L.) is the leading fiber crop and the second most important oil seed crop in the world. This study aimed to analyze the genetic differences among 9 cotton genotypes of cultivars from Turkey using the random amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR) method. A total of 42 RAPD primers were used to determine polymorphism among the genotypes and 34 of the primers produced scorable results, with 319 RAPD-PCR bands obtained. The rate of polymorphism was calculated to be 18.1%, and genetic similarity between any 2 genotypes ranged from 90.2% to 96.5%. By evaluating the presence and absence of the RAPD-PCR bands, a dendrogram indicating the relative genetic similarity of the Turkish cotton genotypes was constructed and presented. The results also showed that the genotypes can be separated from each other at the molecular level by taking advantage of some of the RAPD markers. Specific markers were proposed for the identification of genotypes. To our knowledge, this is the first study to use RAPD markers to investigate the genetic relationships among the Gossypium genotypes grown in Turkey.

RAPD-PCR and Outer Membrane Protein Characterization of Vibrio alginolyticus and Vibrio parahaemolyticus Isolated from Diseased shrimp

Vibrio alginolyticus and V. parahaemolyticus were characterized by random amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR) and profiling of outer membrane proteins (OMP) to the genetic level to enable easier identification of these microorganisms from diseased shrimp (Penaeus monodon) and adopt control measures. Of 20 tested Operon primers (OPC), only OPC5 and OPC18 were used for PCR amlification of 70 isolates representing three strains of each species. OPC5 amplified 1-8 bands with sizes ranging 0.42-2.95 Kb in V. alginolyticus, 5-10 fragments with sizes ranging 0.449-3.55 Kb in V. parahaemolyticus, and one unique common band of 1.34 Kb in both species. When OPC18 was used, one dark band (2.4 Kb) was common to all V. alginolyticus strains and another dark band (1.77 Kb) was common to two strains and the V. alginolyticus ATCC reference strain. One protein band (37.63 kDa) was common to both V. alginolyticus and V. parahaemolyticus and could be used as a diagnostic OMP marker for both species. The simpler and more rapid dot-ELISA and indirect ELISA methods confirmed identification of these pathogenic bacteria.

Optimization of RAPD-PCR Protocol to Screen Jatropha Curcas and Gossypium Hirsutum Grown in Metal Contaminated Soil

The optimization of randomly amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR) protocol was performed to screen two plants ( Jatropha curcas & Gossypium hirsutum ) grown in metal contaminated soils. A CTAB method was employed for DNA extraction, but an overnight RNase treatment was carried out to eliminate RNA contaminants. The isolated DNA was used for RAPD-PCR amplification. The optimum condition for reliable amplification requires a higher concentration of MgCl 2 (3 mM), primer (2.5 μM), Taq DNA polymerase (1 unit) and 3μl of template DNA (sample) and an annealing temperature of 55°C. Reproducible amplified products were observed in these conditions for both the plant species.

Identification and determination of relatedness of lactobacilli using different DNA amplification methods

AbstractSeveral DNA amplification-based methods were used for identification and evaluation of the relation between lactobacilli isolated from breastfed full-term infant faeces (31 strains), dairy products (5 strains) and silage (1 strain). Twenty-seven strains isolated from infant faeces were identified as Lactobacillus rhamnosus (9), Lactobacillus gasseri (6), Lactobacillus paracasei (4), Lactobacillus fermentum (4), Lactobacillus salivarius (2), Lactobacillus plantarum (1), and Lactobacillus helveticus (1) using 10 species-specific polymerase chain reactions (PCRs), multiplex PCR for the Lactobacillus casei group, and sequencing of 16S rDNA. Four strains were not identified. Six strains isolated from dairy products and silage were identified as Lactobacillus rhamnosus. A repetitive extragenic palindromic polymerase chain reaction (rep-PCR) with primer (GTG)5 and a randomly amplified polymorphic DNA polymerase chain reaction (RAPD-PCR) with primer M13 were used for confirmation of species identification. Fingerprints were used for evaluation of the relatedness of lactobacilli. Differences between strains from infant faeces, dairy products, and silage were not detected.

Molecular analysis and dimorphism of azole-susceptible and resistant Candida albicans isolates

Candida albicans is responsible for superficial or systemic infections known as candidiasis, which may be found in infected tissue as unicellular budding yeasts, hyphae, or pseudohyphae. In this study, the effects of both fluconazole and itraconazole antifungal agents on the hyphal formation and genotypic characterization of C. albicans isolates classified as either susceptible or resistant were investigated. The hyphal production of five C. albicans isolates under the action of antifungal agents was investigated by culturing yeast on growth medium and on hyphal induction medium. The genotypic characterization was carried out for 13 isolates of C. albicans using the random amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR) method. The dimorphism analysis showed that the hyphal formation was higher in resistant than in the susceptible isolates to both azoles. The RAPD-PCR method identified the formation of two different groups. In group A, four resistant and two susceptible isolates were clustered, and in group B, one resistant and six susceptible isolates were clustered. Considering that hyphal formation was higher in resistant isolates in the presence of azole drugs, we confirmed that the hyphal production is closely related to susceptibility to azoles. These drugs may affect the morphogenesis of C. albicans depending on their susceptibility to these drugs. In relation to RAPD-PCR, most resistant isolates classified in group A and susceptible isolates in group B demonstrated that this method presented a similar standard between the two groups, suggesting that by this technique, a strong correlation between genotypes and fluconazole-resistant samples may be found.

Screening for glycosidase activities of lactic acid bacteria as a biotechnological tool in oenology

The aim of this study was to evaluate the ability from a number of lactic acid bacteria isolated from different sources to produce glycosidase enzymes. Representative isolates (225) from clusters obtained after genotyping, using randomly amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR) analysis, of 1,464 isolates, were screened for β-D-glucosidase activity. Thirty-five of them were selected for subsequent analysis. These strains were able to hydrolyze α-D-glucopyranoside, β-D-xylopyranoside and α-L-arabinofuranoside although β-D-glucosidase activity was the predominant activity for 22 of the selected strains. Only some of them did so with α-L-rhamnopyranoside. All of these were from wine samples and were identified as belonging to the Oenococcus oeni species using Amplification and Restriction Analysis of 16S-rRNA gene (16S-ARDRA). When the influence of pH, temperature and ethanol or sugars content on β-D-glucosidase activity was assayed, a strain-dependent response was observed. The β-D-glucosidase activity occurred in both whole and sonicated cells but not in the supernatants from cultures or obtained after cell sonication. Strains 10, 17, 21, and 23 retained the most β-D-glucosidase activity when they were assayed at the conditions of temperature, pH, ethanol and sugar content used in winemaking. These results suggest that these strains could be used as a source of glycosidase enzymes for use in winemaking.

Diversity and toxigenicity among members of the Bacillus cereus group

Members of the Bacillus cereus group were isolated from rice products by centrifugation-plating and conventional spread-plating methods. Random amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR) results showed broad diversity among the strains and revealed some associations among isolates from raw and cooked rice samples, at the genotypic level. A comparatively greater diversity among strains was observed in isolates from raw rice than those from cooked rice and, generally, the RAPD profiles of isolates from raw and cooked rice were different, with only a few of them common to both types of rice. The toxigenic potential of the isolates was also determined by molecular and immunoassay analyses. The results revealed that most isolates from the B. cereus group were potentially or actually toxigenic, and some isolates could produce both diarrhoeal and emetic toxins. Generally, isolates belonging to the B. cereus group with the same RAPD pattern were shown to have a similar profile of enterotoxigenicity.

Typing of Salmonella Typhi strains isolated from Egypt by RAPD PCR

PCR-based fingerprinting using random amplified polymorphic DNA (RAPD) has been used widely for genome identification. In this study, 13 Salmonella Typhi strains were isolated from typhoid patients from Aswan, Cairo, Fayoum, and Monofya Governorates of Egypt. The isolates, along with three reference strains, i.e., O901, H901, and Ty2 were subjected to whole genome typing by RAPD PCR. Three RAPD-PCR 10-mer primers generated a total of 85 RAPD bands (81 polymorphic bands), 12 distinct PCR profiles, and proved to be useful for discriminating the isolates and strains studied. Interestingly, the B1 and C1 PCR profile were found only in Cairo and Monofya, respectively; and some PCR types appeared only in certain Governorates of Egypt. By combining the profiles obtained with the primer trio used in this study, an excellent discrimination index (D) of 0.942 was reached. Pairwise comparisons of Jaccard’s similarity coefficients calculated among the 12 PCR types identified three major clusters; i.e., O901 branch and Ty2 and H901 sub-branches. Principal component analysis adequately resolved each of these three major clusters. Three principal components accounted for about 72% of the variation, with the first two components accounting for about 62% of the total variance among the genotypes studied. Biclustering improved the display of groups of RAPD amplicons (markers) that cluster similarly across the genomes and could delineate features pertaining to genome structure. In conclusion, RAPD PCR provided a fast method with high potentials in surveillance and epidemiological investigations of Salmonella Typhi infections.

Assessment of air pollution genotoxicity by RAPD in Evernia prunastri L. Ach. from around iron-steel factory in Karabük, Turkey

Lichens are widely used in biomonitoring studies of air pollution, either as bioindicators of air quality or as bioaccumulators of atmospheric deposition. Over the past decade, several molecular techniques have been developed to provide information on diversity, genotoxicology, genetic relationships, etc. The heavy metal contents of Evernia prunastri samples were determined by atomic absorption spectrometry. The Random Amplified Polymorphic DNA Polymerase Chain Reaction (RAPD-PCR) method was used to describe the pattern of DNA band variation in the samples influenced by the environmental pollution. The study was designed to describe the level of pollution in an area contaminated with smoke and waste from an iron-steel factory, and to reveal the level of potential genotoxic agents around this source of pollution. The study also examined the suitability of the lichen samples for the detection of genotoxicity.

Genetic diversity in 14 tomato (Lycopersicon esculentum Mill.) varieties in Nigerian markets by RAPD-PCR technique

A total of 226 tomato (Lycopersicon esculentum Mill.) fruits sold in markets within three states in Southwestern Nigeria were purchased and differentiated into 14 varietal groups based on morphological parameters. The genomic DNA from the young apical leaves of the seeded plants were extracted and analyzed by the randomly amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR) technique. Seventy-four (74) amplified products were scored with 62.2% of them revealing polymorphism. The OPB-18 primer recorded the highest polymorphism (83.3%) and OPU-14 the least (44.4%). The dendrogram revealed 3 clusters. The genotypic relationships between varieties within clusters ranged from 2 in clusters 1 and 3 to 10 in cluster 2. Cluster 2 revealed 4 subgroups with two pairs of varieties (var. c and i and l and m) showing 100% similarities. The local names, place of cultivation and phenotypic characters expressed by the varieties were not suitable for varietal differentiation since phenotypically different fruits showed genotypic relatedness. The impression that tomato varieties from a particular geographical zone are genetically similar was therefore nullified. Key words: Tomato, randomly amplified polymorphic DNA, polymerase chain reaction, molecular, genetic diversity, Lycopersicum.