Using Unweighted Pair Group Method With Arithmetic Mean Research Articles

SSRs MARKERS REVEAL HIGH GENETIC DIVERSITY AND LIMITED DIFFERENTIATION AMONG POPULATIONS OF NATIVE GUATEMALAN AVOCADO

Even though Guatemala is considered one of the three domestication centres of the avocado (Persea americana Mill), little is known about the genetic diversity of its native populations. This is the first study to explore avocado genetic diversity and population structure in native dispersal areas using microsatellite markers. We sampled 189 avocado trees grown from seeds in eight geographical populations, and 12 microsatellite loci were tested for diversity. There were a total of 289 alleles found throughout the 12 loci, with an average of 23.83 alleles per locus. The average observed and expected heterozygosity were 0.53 and 0.83, respectively. The analysis of molecular variance revealed that the eight populations comprised around 2% of the variation. Five of the 28 G'ST(Nei) pairwise comparisons indicated no genetic difference. We discovered a mixture of avocado trees from various populations using unweighted pair group method with arithmetic mean (UPGMA) hierarchical cluster analysis. The sampled individuals were classified into three main genetic clusters by the model-based STRUCTURE and discriminant analysis of principal components (DAPC). The weak genetic structure exhibited by mixing avocado trees from different populations could be mainly attributed to high human-mediated gene flow (Nm = 12.25) due to avocado's importance in food, culture, and Guatemala's religion. The high genetic diversity found in the avocado germplasm suggests that it might be a valuable source of variable alleles that could be used in breeding programs for this crop in Guatemala.

Genetic diversity analysis of Moroccan saffron (Crocus sativus L.) of different origins based on intermicrosatellite markers

Saffron (Crocus sativus L.) is an autumnal herbaceous triploid plant; it is the source of saffron spice, recognized as the most expensive spice in the world. In this study, genetic diversity among 14 saffron accessions collected from different ancestral geographic areas in Morocco, Greece and France, has been assessed using inter-simple sequence repeats (ISSRs) markers system. Ten ISSR primers were amplified, a total of 143 fragments of which 44.05 % are polymorphic with an average of 6.3 polymorphic fragments per each primer and average of polymorphic information content (PIC) of 0.236. ISSR markers proved to be a powerful tool for assessment of genetic diversity among saffron accessions. Cluster analysis using unweighted pair group method with arithmetic mean (UPGMA), based on Jaccard’s similarity coefficient and supported by the principal coordinate analysis (PCoA), divided the studied accessions into three major groups, and showed that genetic distance is independent of geographical distance. In this paper, we report for the first time the level of genetic diversity among Moroccan saffron; this information allows an unequivocal development of a suitable approach for the conservation of C. sativus germplasm and reduce its genetic erosion.

Genetic variation for important processing traits and identification of potential parents for developing hybrid clones suitable for processing in sweet potato (Ipomoea batatus L Lam)

Processing and value addition have gained tremendous focus in recent years and therefore breeding for improved quality traits and development of varieties suitable for various processing methods and products assumes significance. This study aimed at determining the genetic variation and identifying parents for developing hybrid clones suitable for processing in sweet potato based on five important processing traits namely starch content, sugar content, dry matter content, flour yield and peel loss. Significant variability for all the selected processing traits was found among the 18 genotypes studied which encompasses the scope of mining the germplasm further for processing and consumer traits. Nine genotypes recorded dry matter content of above 30%. Dry matter content had highly significant and positive correlation with starch and flour yield and significant positive correlation with sugar content. Flour yield exhibited the highest heritability followed by dry matter content, peel loss, sugar and starch contents. The Principal Component Analysis (PCA) analysis identified two principal components explaining 39.11% and 26.40% of total variation observed. Two major and six sub clusters were formed using unweighted pair group method with arithmetic mean (UPGMA) clustering approach. High and low starch varieties were grouped separately in two different clusters. The sensory evaluation of fried chips demonstrated that genotypes such as S1652, S1712, S27 and S1609 were found suitable for processing into fried chips. The score for willingness to buy was found to be high for fried chips of S1652, Kanhangad, S1609, Varsha and S27. Genotypes such as S1652, Bhu Sona, Bhu Krishna, Kamala Sundari and S1712 were found to be highly suitable for consumption after boiling. The genotypes identified in this study can be utilised for the development of varieties suitable for varied end uses. The findings of this study strongly suggest that varieties suitable for processing can be identified among the germplasm based on evaluation of processing traits.

EgyGene GelAnalyzer4: a powerful image analysis software for one-dimensional gel electrophoresis

BackgroundGel image analysis is a cornerstone in electrophoresis-based experiments. Nature of the experiment determines the target of the analysis. Implemented features within different software for gel images and their outputs are variable. EgyGene GelAnalyzer4 software aimed to make a powerful, easy, and almost all-in-one software for gel image analysis. ResultsEgyGene GelAnalyzer4 is a software for analyzing the gel images. The program consists of six main parts: gel image enhancement, image analysis, data merging, molecular markers detection, phylogenetic tree prediction, and population parameters estimation. Firstly, the gel image enhancement part enhances the gel image colors and clarity. Then, the image is analyzed in the second part using both automatic and manual detection of lanes and bands. The third part, data merging, is for merging several analyses for different samples/gels together to produce one table comprises the entire experiment. The fourth section, markers detection, collects several pre-analyzed data for the same samples to detect their molecular markers based on some user-defined criteria. The fifth part uses the outputs of the previous parts to draw the phylogenetic tree between samples. The program constructs a phylogenetic tree using unweighted pair group method with arithmetic mean (UPGMA) method. Fourteen different equations are used to calculate the similarity percentages: Anderberg, Czekanowski, Dice, Faith, Gower and Legendre, Jaccard, Nei and Li, Rogers and Tanimoto, Russel and Rao, simple matching, Sokal and Michener, Sokal and Sneath 1, Sokal and Sneath 2, and Sorensen. The last section calculates the population parameters for the pre-analyzed images before. EgyGene GelAnalyzer4 is designed and programed using Microsoft Visual Studio 2012 Express version, based on the Dot Net FrameWork 4.5 edition, whereas its icons are created using the freeware Greenfish Icon Editor Pro 3.2.5.1. ConclusionsEgyGene GelAnalyzer4 is a simple, fast, and user-friendly software with six novel features: changing the order of lanes within the gel image, splitting the detecting lane into two lanes, merging of several detected lanes into just one, splitting a single band into two bands, merging two or more bands to be a single band, and comparing data of several analyzed images with a big number of samples and merging it together in a single table and detection of molecular markers between samples. In addition to the basic features and outputs of other image analysis software such as calculation of relative mobility values, molecular weights/sizes estimation, DNA/RNA/protein quantity within bands, table of present/absent of bands, polymorphism percentage, similarity percentages and phylogeny, population parameters, enhancing the image colors and clarity, and merging data for several gels. In addition, writing names of samples and the detected markers on the image. EgyGene GelAnalyzer4 provides many several features, which may be not available together in any other alternative.

Identification of Genetic Distance of Exotic and Locally Developed Maize ( Zea mays L) Inbred Lines

Development of hybrid varieties is one of the strategies to increase the productivity and production of Maize (Zea mays L.) in Sri Lanka. Heterosis is an important aspect to measure the hybrid vigor. Genetic distance is a main factor, which affects on heterosis. Identification of suitable parents using molecular markers gives many advantages over morphological evaluation. This study was conducted at the Molecular Biology Laboratory at Plant Genetic Resources Centre, Gannoruwa to identify distant parents using Simple Sequence Repeats (SSR) primers. Genomic DNA was isolated from selected 23 inbred lines including exotic and locally developed lines using modified CTAB method. Molecular characterization was done using four SSR primers; umc1023, phi402893, phi 112 and phi 041. Amplified products were resolved in Polyacrylamide Gel Electrophoresis. Data were manually scored and analysis of genetic variation was carried out using the software POWEMARKER 3.1. Analysis revealed that polymorphism existed for all four SSR primers. Considerable genetic diversity was observed among the loci. Number of alleles per locus ranged from 2 to 3. Polymorphic information content (PIC) ranged from 0.24 (phi112) to 0.42 (phi402893). Gene diversity varied from 0.28 (phi112) to 0.46 (phi402893). Based on the values, it was revealed that there is a high genetic diversity present within the tested inbred lines. Genetic distance was calculated based on Nei’s genetic distance and phylogenetic tree was developed using Unweighted Pair Group Method with Arithmetic Mean (UPGMA). Tested 23 populations could be grouped into four clusters. Most of the exotic inbred lines grouped into one cluster revealed that there is less genetic distances present within the exotic lines. The genetic distance information obtained in this study will be useful to select better parents for development of maize hybrids with good performance.

Genetic characterization of iler (Plectranthus scutellarioides (L.) R. Br.) based on RAPD molecular marker

Iler (Plectranthus scutellarioides (L.) R. Br.) is an ornamental plant and also used as medicinal plant. The plant is one of medicinal plant for anti hemorrhoids Jamu Saintifik herbal formula in B2P2TOOT Indonesian Ministry of Health. This will lead to commercialization opportunities in herbal drug industry that recently has become increasingly popular. With the emergence of substitution and adulteration of herbal drug and the lack of information on coleus genetic diversity which may result in misidentification, this research was conducted to determine Iler genetic characteristics using RAPD markers. The genomic DNA was extracted from 15 accessions of iler and 3 accessions from genus Plectranthus as an outgroup. Six selected RAPD primers were used in amplification. Dice similarity index was used to calculate similarity index followed by cluster analysis and dendrogram contruction using Unweighted Pair Group Method with Arithmetic Mean (UPGMA). The results showed 52 DNA fragments were produce within the size range 293-2,667 bp and polymorphism level among accessions at 94.23%. Cluster results results in genetic diversity ranging from 40.45-97.87% and a dendogram that shows clustering trend of Iler accessions based on the similarity of morphological characters such as leaf shape and color.

Craniometric geographical variation in Hylaeamys megacephalus populations

ABSTRACT: Hylaeamys megacephalus (G. Fisher, 1814) presents great genetic diversity and wide geographical distribution, and occurs in both the Amazon and Cerrado biomes. Because of its generalist aspect, this species tolerates different eating habits and habitats. It occurs in flooded and dry areas and is predominantly terrestrial, which allows greater gene flow between populations even over long distances. Studies that seek a better understanding of morphological variations resulting from differences imposed by the environment throughout this species’ distribution are still lacking. This study aimed to analyze the differences between H. megacephalus populations based on craniometry, investigating whether the environment has an influence on morphology. We analyzed a total of 142 specimens from three scientific mammal collections: National Museum, “Universidade Federal do Rio de Janeiro” (MN-UFRJ); “Laboratório de Biologia e Parasitologia de Mamíferos Reservatórios Silvestres”, “Instituto Oswaldo Cruz”, “Fundação Oswaldo Cruz”(LBCE-Fiocruz); and “Laboratório de Biodiversidade”, “Universidade Federal de Goiás”, “Regional Jataí” (LZE-UFG), and took 20 craniometric measurements. Craniometry was explored using unweighted pair group method with arithmetic mean (UPGMA), canonical variate analysis, and principal component analysis (PCA). The results led us to conclude that there are three craniometric groups of H. megacephalus with a tendency to differentiate as a result of geographical influences.

Genetic Variability of Eggplant Germplasm Evaluated under Open Field and Glasshouse Cropping Conditions

Knowledge of agro-morphological genetic variation and cropping conditions on vegetative and yield-related traits plays a significant role in varietal improvement and production of eggplant (Solanum melongena L.). Following this premise, the current study was conducted to critically asses the genetic variation of 29 eggplant accessions by using agro-morphological characterization evaluated under two cropping conditions, namely, glasshouse and open field. The experiments were laid out in randomized complete block design (RCBD) with three replications. Data on vegetative and yield characteristics were collected and subjected to analysis of variance (ANOVA) using SAS 9.4, while variance components were estimated manually. The results obtained from the analysis of variance indicated a highly significant difference (p ≤ 0.01) for all characteristics studied in both cropping conditions. The evaluated accessions were grouped into six major clusters based on agro-morphological traits using Unweighted Pair Group Method with Arithmetic mean (UPGMA) dendrogram. Hence, crosses between group I with VI or V could be used to attain higher heterosis and vigor among the accessions. Also, this evaluation could be used as a selection criterion for important yield agronomic traits in eggplant. The methodology and the approaches used may provide a model for the enhancement of other vegetable crop diversity towards adaptability to the cropping condition decision. This result displayed importance for preserving eggplant germplasm for future varietal development and revealed that open field cropping condition is more suitable under Malaysia’s agroecology.

Screening of Blast Resistance Genes from South Tapanuli Rice Cultivars, North Sumatra, Indonesia

Rice blast disease caused by a phytopathogenic fungus, Magnaporthe oryzae, has been considered as a major threat to global rice productivity, including Indonesia. South Tapanuli is a regency in North Sumatra, known for its intensive and significant rice production activities. A collection of 13 rice cultivars originating from the region, is claimed for its resistances against the rice blast disease based on the practices by the local farmers. Molecular screening of blast resistance (R) genes was initiated to the 13 accessions using the Polymerase Chain Reaction (PCR) targeting five blast resistance (R) genes, Pup1, Pi-37, Pi-d2, Pi-ta2, Pib and Pi-kh. Out of the 13 accessions, the Pi-d2, Pi-ta2 and Pib were detected in all cultivars, showing 100% of R gene frequency. The second dominant, Pup1 and Pi-37 gene frequencies were 76.9 and 61.5%, respectively while the least dominant, Pi-kh gene frequency was 15.4%. The number of R genes detected among cultivars was ranged between 3 to 6 genes. The genetic associations among thirteen germplasms were determined using unweighted pair group method with arithmetic mean (UPGMA) analysis. Cluster analysis revealed that the least blast-resistant cultivars: Putri Kembar, Silatian and Siganteng, were grouped into cluster 1, showing polymorphism for Pi-d2, Pi-ta2 and Pi-b. The high-resistant cultivars, Martabe-Sicondong and Sayuti, Sitolas, Pulo Raja, Pulo Pandan, Siporang and Pulo Manggis, were grouped into cluster 2 and 3, respectively. The medium blast-resistant cultivars, IR 64 and Sitampan were grouped into cluster 4, showing polymorphism for Pup1, Pi-d2, Pi-ta2 and Pi-b. These results indicated that the utilization of blast-resistant cultivars in North Tapanuli was supported by the presence of R genes.

Genetic variation of Eruca sativa L. genotypes revealed by agro-morphological traits and ISSR molecular markers

Taramira (Eruca sativa L.) is an oilseed crop with extensive industrial and pharmaceutical applications. The present study was carried out to identify genetic diversity in 60 different genotypes of E. sativa L. using 19 inter simple sequence repeats (ISSR) molecular markers and 12 agro-morphological traits, collected from different geographical regions of the world. The results of analysis of variance (ANOVA) indicated that there was a significant difference among genotypes in terms of all agronomic traits (except for capitula length and 1000-seed weight). In this study, genotype 161 (with Iranian origin) had the highest grain yield per plant (519.57 gm−2). The number of 19 ISSR primers generated a total of 286 polymorphic bands which polymorphism information content (PIC) value varied from 0.17 (in UBC827 primer) to 0.31 (in UBC825 primer) with average of 0.25. Genetic diversity statistics showed that, the UBC840 primer had the most significant role in differentiating genotypes. Taramira genotypes were divided into three subgroups in cluster analysis of molecular traits using unweighted pair group method with arithmetic mean (UPGMA) method and the STRUCTURE methods. There was a considerable similarity between the geographic origin of genotypes and molecular classification pattern. Therefore, the selection of genotypes with a high genetic distance in terms of the molecular marker, along with desirable agronomic traits, can be effective in future breeding programs of taramira in order to produce new superior hybrids.

Molecular characterization of sesame germplasms of West Bengal, India using RAPD markers

The aim of this research was to assess the genetic diversity of sesame (Sesamum indicum L.) and also to reveal the genetic relationships using the Random Amplified Polymorphic DNA (RAPD) markers. Fifteen sesame germplasms were collected from seven districts or four zones of West Bengal, India. A high genetic diversity was revealed by ten RAPD primers within and among the fifteen germplasms. The value of Jaccard’s similarity coefficients among and within the fifteen germplasms ranged from 0.287 to 0.725 which indicated high degree of genetic variability. Cluster analysis using Unweighted Pair Group Method with Arithmetic Mean (UPGMA) grouped all the germplasms into three main clusters. Analysis of various genetic diversity indices strongly indicated high level of genetic diversity among the populations of four different regions. UPGMA analysis of four populations resulted into two groups and the results of Principal Coordinates Analysis (PCoA) depicted a clear distinction among the germplasms.

Use of Next Generation Sequencing to study two cowpox virus outbreaks.

BackgroundBetween 2008 and 2011 about 40 cases of human cowpox were reported from Germany and France. Infections had been acquired via close contact to infected, young pet rats. An identical and unique sequence of the hemagglutinin gene was found in various cowpox virus (CPXV) isolates pointing to a common source of infection. In a second CPXV outbreak in cats in a small animal clinic in Germany in 2015, four out of five hospitalized cats showed identical hemagglutinin sequences and thus, a hospital-acquired transmission had been assumed. Next-Generation Sequencing was performed in order to re-investigate the outbreaks, as epidemiological data could not confirm all cases.MethodsHomogenates of lesion material from rats, cats and humans were cultivated in cell culture. The genomes of four virus isolates, nine CPXVs from our strain collections and from DNA of three paraffin-embedded lesion materials were determined by Next Generation Sequencing (NGS). For phylogenetic analyses a MAFFT-alignment was generated. A distance matrix based on concatenated SNPs was calculated and plotted as dendrogram using Unweighted Pair Group Method with Arithmetic mean (UPGMA) for visualization.ResultsAligning of about 200.000 nucleotides of 8 virus isolates associated with the pet rat outbreak revealed complete identity of six genomes, the remainder two genomes differed in as little as 3 SNPs. When comparing this dataset with four already published CPXV genomes also associated with the pet rat outbreak, again a maximum difference of 3 SNPs was found. The outbreak which lasted from 2008 till 2011 was indeed caused by a single strain which has maintained an extremely high level of clonality over 4 years. Aligning genomic sequences from four cases of feline cowpox revealed 3 identical sequences and one sequence which differed in 65 nucleotides. Although identical hemagglutinin sequences had been obtained from four hospitalized cats, genomic sequencing proved that a hospital-acquired transmission had occurred in only three cats.ConclusionAnalyzing the rather short sequence of the hemagglutinin gene is not sufficient to conduct molecular trace back analyses. Instead, whole genome sequencing is the method of choice which can even be applied to paraffin-embedded specimens.

Genome-Wide Association Mapping of Major Root Length QTLs Under PEG Induced Water Stress in Wheat.

Roots are vital plant organs that determine adaptation to various soil conditions. The present study evaluated a core winter wheat collection for rooting depth under PEG induced early stage water stress and non-stress growing conditions. Analysis of phenotypic data indicated highly significant (p < 0.01) variation among genotypes. Broad sense heritability of 59 and 73% with corresponding genetic gains of 7.6 and 9.7 (5% selection intensity) were found under non-stress and stress conditions, respectively. The test genotypes were grouped in to three distinct clusters using unweighted pair group method with arithmetic mean (UPGMA) clustering based on maximum Euclidian distance. The first three principal components gave optimum mixed linear model for genome wide association study (GWAS). Linkage disequilibrium (LD) analysis showed significant LD (p < 0.05) amongst 15% of total marker pairs (25,125). Nearly 16% of the significant LDs were among inter chromosomal marker pairs. GWAS revealed five significant root length QTLs spread across four chromosomes. None of the identified QTLs were common between the two growing conditions. Stress specific QTLs, combined explaining 31% of phenotypic variation were located on chromosomes 2B (wPt6278) and 3B (wPt1159). Similarly, two of the three QTLs (wPt0021 and wPt8890) identified under the non-stress condition were found on chromosomes 3B and 5B, respectively. The B genome showed significant importance in controlling root growth both under stress and non-stress conditions. The identified markers can potentially be validated and used for marker assisted selection.

Genetic diversity and population structure of synthetic hexaploid-derived wheat (Triticum aestivum L.) accessions

A comprehensive understanding of the population structure and genetic diversity of potential germplasm is necessary for making breeding decisions and to fully interpret marker-trait associations. The purpose of this study was to examine the genetic diversity and population structure of a panel of 194 synthetic hexaploid-derived wheat (SHW; Triticum aestivum L.) accessions using 6904 polymorphic single nucleotide polymorphism (SNP) markers. Ancestry-based dissimilarity indices and marker-based genetic distances were positively correlated (r = 0.67). The variation in the primary synthetic parent in the pedigrees accounted for 4.52%, while the degree of the synthetic contribution accounted for only 1.06% of variation in the genetic distance. In addition, variation in the Aegilops tauschii Coss. (syn. Aegilops squarrosa auct. non L.) accession and T. turgidum accession used in the initial cross accounted for 3.48% and 2.75% of the variation in genetic distance, respectively. Using a model-based population structure approach, seven sub-populations were identified in the panel. Results of the model-based population structure analysis was for the most part in agreement with the distance-based clustering using unweighted pair group method with arithmetic mean (UPGMA) of the genetic distance or ancestry data and the principle component analysis of relatedness. We conclude that using a model-based approach provides a more statistically robust estimation of population structure. Results of this study, while highlighting the potential contribution of introgressed genome in the panel, provide the foundation for employing this panel in genome-wide association studies.

Numerical taxonomy contributes to delimitation of Iranian and Turkish Hesperis L. (Brassicaceae) species

Taxonomic descriptions of Iranian and Turkish Hesperis (Brassicaceae) species are generally insufficient and partly incomplete, which makes the species delimitation ambiguous. In order to clarify species circumscription, we scored 57 morphological descriptors (MDs) in 121 operational taxonomic units (OTUs) of Hesperis from Iran and Turkey and performed a multivariate analysis. The dendrogram was created from Gower’s distance matrix using Unweighted Pair Group Method with arithmetic mean (UPGMA) algorithm. The dendrogram clearly separates the 121 OTUs of Hesperis into five main phenons, which significantly deviate from the classical taxonomic treatment (sectional assignments) of the genus. Similar distinct delineation among the five phenons was revealed by a Principal Coordinate Analysis (PCoA), highlighting the resolving power of the multivariate analyses of quantitative and qualitative morphological characters. While there were significant variations among the OTUs for 57 MDs, the most distinctive morphological descriptors delimiting the phenons were estimated to be fruit, petal, stem, and leaf by a de-trended correspondence analysis (DCA). We also present a comparative discussion between the classical taxonomy and the delimitation of taxa revealed in our study.

Analysis of genetic diversity of passion fruit (Passiflora edulis Sims) genotypes grown in Kenya by sequence-related amplified polymorphism (SRAP) markers

Abstract Passion fruit (Passiflora edulis [Sims]) is currently ranked third among fruit exports from Kenya and has great potential since demand for both fresh fruit and processed juice is on a continuous increase. Although assessment of genetic variability of germplasm is indispensable for improvement and development of superior cultivars, little information is available on the genetic diversity of passion fruit cultivated in Kenya. The objective of this study was to determine the genetic diversity of passion fruit genotypes from major growing regions in Kenya using sequence-related amplified polymorphism (SRAP) markers. Twenty four SRAP primer combinations were screened using three passion fruit genotypes and only seven that displayed polymorphic and stable amplification profiles were used to analyze 22 genotypes. The seven primer combinations amplified a total of 931 clear bands with an average of 133 bands per primer pair, of which 610 (65.5%) bands were polymorphic. The similarity coefficients among the 22 passion fruit germplasms ranged from 0.51 to 1.0 with an average of 0.755. The 22 passion fruit genotypes were classified into two groups by cluster analysis using unweighted pair-group method with arithmetic mean (UPGMA) with 12% similarity. Shannon's diversity index was 0.0934 and Nei's gene diversity index was 0.1370 in the present study. The study findings demonstrate the existence of a considerable amount of genetic variability among passion fruit genotypes grown in different regions of Kenya. This indicates the potential application of these genotypes in breeding programs by exploiting the use of molecular markers for selection of specific agronomic traits.

Start codon targeted (SCoT) polymorphism-based genetic relationships and diversity among populations of Tecomella undulata (Sm.) Seem—an endangered timber tree of hot arid regions

Tecomella undulata (Sm.) Seem commonly called Rohida or Desert teak (family bignoniaceae) is an important agroforestry tree having an important pharmacological and therapeutic uses. Its distribution is restricted to hot arid regions of India and Pakistan having <150 to 500 mm annual rainfall. Genetic diversity status of this important endangered timber tree species designated state flower of Rajasthan remains unrevealed. Genetic diversity of 21 populations (108 accessions) encompassing yellow, red and orange coloured flower bearing morphotypes collected from all 12 districts of western Rajasthan, India, has been examined using start codon targeted (SCoT) polymorphism for the first time. Fingerprinting with 22 SCoT primers (out of 36 screened) generated 294 amplicons of 100 to 3000 bp size, of which 212 (71.6%) were polymorphic. Amplicon number varied from 4 (SCoT-9) to 24 (SCoT-15) with an average of 13.4 amplicons per primer. Average polymorphism information content (PIC), Nei’s diversity index (H) and Shannon index (I) were 0.54, 0.22 and 0.36, respectively. Dendrogram generated using unweighted pair-group method with arithmetic mean (UPGMA) delineated the 108 accessions into 5 clusters while 2 accessions out grouped. Principal coordinate analysis (PCoA) also revealed similar clustering. High level of genetic differentiation among accessions/populations was attributed to cross pollination and continuous evolution under harsh agro-climatic conditions.

Detection of neonatal unit clusters of Candida parapsilosis fungaemia by microsatellite genotyping: Results from laboratory-based sentinel surveillance, South Africa, 2009-2010.

Neonatal candidaemia is a common, deadly and costly hospital-associated disease. To determine the genetic diversity of Candida parapsilosis causing fungaemia in South African neonatal intensive care units (NICUs). From February 2009 through to August 2010, cases of candidaemia were reported through laboratory-based surveillance. C.parapsilosis isolates from neonatal cases were submitted for identification by internal transcribed spacer (ITS) region sequencing, antifungal susceptibility testing and microsatellite genotyping. Cluster analysis was performed using Unweighted Pair Group Method with Arithmetic Mean (UPGMA). Of 1671 cases with a viable Candida isolate, 393 (24%) occurred among neonates. Isolates from 143 neonatal cases were confirmed as C.parapsilosis sensu stricto. Many isolates were resistant to fluconazole (77/143; 54%) and voriconazole (20/143; 14%). Of 79 closely-related genotypes, 18 were represented by ≥2 isolates; 61 genotypes had a single isolate each. Seven clusters, comprised of 82 isolates, were identified at five hospitals in three provinces. Isolates belonging to certain clusters were significantly more likely to be fluconazole resistant: all cluster 7 isolates and the majority of cluster 4 (78%), 5 (89%) and 6 (67%) isolates (P<.001). Candida parapsilosis-associated candidaemia in public-sector NICUs was caused by closely related genotypes and there was molecular evidence of undetected outbreaks as well as intra-hospital transmission.

Assessment of genetic variability in Trigonella foenum-graecum L. germplasm by SDS-page analysis

The 96 genotypes of Trigonella foenum-graecum L. (Fenugreek) were estimated by using Sodium Dodecyl Sulphate Poly Acrylamide Gel Electrophoresis (SDS-PAGE). The accessions were maintained from diverse ecological areas of the world. Total seed storage proteins were electrophoresis on 12.5% polyacrylamide gels. A total of 17 protein bands were detected, of which seven were highly polymorphic and six were moderate polymorphic and four were low polymorphic with molecular weight extending from of ~15 to ~180 kDa. The dendrogram based on similarity matrix using Unweighted Pair Group Method with Arithmetic Mean (UPGMA) divided all the genotypes into four major clusters i.e., I, II, III and IV comprising 51, 24, 10 and 11 accessions, respectively. Although limited genetic diversity was detected amongst known germplasm but the presence/absence of polymorphic bands revealed a significant variances among different Trigonella genotypes. The differences exposed in this project work should be oppressed for the future breeding prospective of Trigonella genotypes by using other advanced molecular techniques. The SDS-PAGE per se seems inadequate tool for such kind of analysis, and may be integrated with other genetic and sequence based approaches for more precise estimation of the genetic variability within closely related accessions. Our results provide baseline for obtaining locally adapted and improved cultivars of fenugreek in Pakistan.

Cluster Analysis of Campylobacter jejuni Genotypes Isolated from Small and Medium-Sized Mammalian Wildlife and Bovine Livestock from Ontario Farms.

Using data collected from a cross-sectional study of 25 farms (eight beef, eight swine and nine dairy) in 2010, we assessed clustering of molecular subtypes of C.jejuni based on a Campylobacter-specific 40 gene comparative genomic fingerprinting assay (CGF40) subtypes, using unweighted pair-group method with arithmetic mean (UPGMA) analysis, and multiple correspondence analysis. Exact logistic regression was used to determine which genes differentiate wildlife and livestock subtypes in our study population. A total of 33 bovine livestock (17 beef and 16 dairy), 26 wildlife (20 raccoon (Procyon lotor), five skunk (Mephitis mephitis) and one mouse (Peromyscus spp.) C.jejuni isolates were subtyped using CGF40. Dendrogram analysis, based on UPGMA, showed distinct branches separating bovine livestock and mammalian wildlife isolates. Furthermore, two-dimensional multiple correspondence analysis was highly concordant with dendrogram analysis showing clear differentiation between livestock and wildlife CGF40 subtypes. Based on multilevel logistic regression models with a random intercept for farm of origin, we found that isolates in general, and raccoons more specifically, were significantly more likely to be part of the wildlife branch. Exact logistic regression conducted gene by gene revealed 15 genes that were predictive of whether an isolate was of wildlife or bovine livestock isolate origin. Both multiple correspondence analysis and exact logistic regression revealed that in most cases, the presence of a particular gene (13 of 15) was associated with an isolate being of livestock rather than wildlife origin. In conclusion, the evidence gained from dendrogram analysis, multiple correspondence analysis and exact logistic regression indicates that mammalian wildlife carry CGF40 subtypes of C.jejuni distinct from those carried by bovine livestock. Future studies focused on source attribution of C.jejuni in human infections will help determine whether wildlife transmit Campylobacter jejuni directly to humans.