Genetic Variation and Phylogeny of Wabisuke Camellias by Amplified Fragment Length Polymorphism (AFLP) Analysis

Amplified fragment length polymorphism (AFLP) analysis was conducted on the wabisuke camellia and its relative camellia species. Genetic polymorphism was identified among the ‘Uraku’ camellia, its offspring ‘Tosa-uraku’ and Camellia japonica, whereas the two accessions of the old ‘Uraku’ showed monomorphism in all the fragments. The results suggested that the two old ‘Uraku’ trees are asexually-propagated clonal strains. The genetic distance between wabisuke cultivars and Chinese camellias and between wabisuke camellias and C. sinensis was much further than that between wabisuke cultivars and Camellia japonica. It has also been suggested that wabisuke camellias can be classified into two subgroups, I-1 and I-2, and that Subgroup I-2 originated from C. japonica, while Subgroup I-1, including ‘Uraku’ (synonym: ‘Tarokaja’), was developed by the repeated hybridization of C. japonica to interspecific hybrids with the Chinese camellias, e.g., C. pitardii var. pitardii, or by the involvement of related species not investigated in this study.

Genetic Analyses of Saprolegnia Strains Isolated from Salmonid Fish of Different Geographic Origin Document the Connection Between Pathogenicity and Molecular Diversity

Saprolegnia parasitica is recognized as one of the most important oomycetes pests of salmon and trout species. The amplified fragment length polymorphism (AFLP) and method sequence data of the internal transcribed spacer (ITS) were used to study the genetic diversity and relationships of Saprolegnia spp. collected from Canada, Chile, Japan, Norway and Scotland. AFLP analysis of 37 Saprolegnia spp. isolates using six primer combinations gave a total of 163 clear polymorphic bands. Bayesian cluster analysis using genetic similarity divided the isolates into three main groups, suggesting that there are genetic relationships among the isolates. The unweighted pair group method with arithmetic mean (UPGMA) and principal coordinate analysis (PCO) confirmed the pattern of the cluster analyses. ITS analyses of 48 Saprolegnia sequences resulted in five well-defined clades. Analysis of molecular variance (AMOVA) revealed greater variation within countries (91.01%) than among countries (8.99%). We were able to distinguish the Saprolegnia isolates according to their species, ability to produce oogonia with and without long spines on the cysts and their ability to or not to cause mortality in salmonids. AFLP markers and ITS sequencing data obtained in the study, were found to be an efficient tool to characterize the genetic diversity and relationships of Saprolegnia spp. The comparison of AFLP analysis and ITS sequence data using the Mantel test showed a very high and significant correlation (r2 = 0.8317).

AFLP analysis of Genetic diversity and Phylogenetic relationships of Vigna radiata (L) Wilczek

Among the important pulse-yielding plants Vigna radiata is important. In the Indian subcontinent, it is an essential source of dietary protein particularly in the vegetarian population. It contains abundant nutrients with biological activities. Soaked seeds are eaten as a good source of protein and mature seeds are cooked or can be sprouted for human consumption. Mung beans and its sprouts contain chemical constituents like flavonoids, phenolic acids, organic acids, amino acids, carbohydrates, and lipids. It is related to different biological activities like antioxidant, antimicrobial, antitumor, anti-inflammatory, anti-diabetic, and antihypertensive effects, etc. The seeds are a traditional source of cures for paralysis, rheumatism, coughs, fevers, and liver ailments. As a green manure or cover crop, the plant can be grown, enriching the soil with the nitrogen formed on its roots. The powerful Amplified Fragment Length Polymorphism (AFLP) is a molecular marker used for a variety of applications like assessing genetic diversity within species or among closely related species, inferring population-level phylogenies, generating genetic maps, and determining the relatedness among cultivars. AFLP has become extremely beneficial in the study of taxa including bacteria, fungi, and plants, where much is still unknown about the genomic makeup of various organisms. A study was made to assess the genetic diversity and phylogenetic relationships of a set of five varieties of Vigna radiata (L) Wilczek using the AFLP technique. Five different varieties of Vigna radiata viz. B1, TARM 2, PDM 84, TM 99, and TM 98 were subjected to AFLP analysis. A total of 471 fragments were scored across all the 12 AFLP primer sets used and the results were used to plot a dendrogram. It was observed that these five varieties formed three distinct groups among which the aromatic B1 variety formed a distinct group.

Morpho-agronomic, biochemical and molecular analysis of genetic diversity in the Mesoamerican common bean panel

The common bean (Phaseolus vulgaris L.) is of great importance to the food and nutritional security of many populations, and exploitation of the crop’s genetic diversity is essential for the success of breeding programs. Thus, the aim of the present study was to evaluate the genetic diversity of 215 common bean accessions, which included cultivars, obsolete cultivars, improved lines, and landraces using morpho-agronomic and biochemical traits, and amplified fragment length polymorphism markers (AFLP). Genetic parameters, box plots, Pearson’s correlation analysis, and Ward’s hierarchical clustering were used to analyze the data. The Jaccard similarity coefficient and neighbour-joining clustering method were used for molecular analysis. A wide variability among the accessions was observed for morpho-agronomic and biochemical traits. Selective accuracy (Ac) and broad-sense heritability (h2) values were high to intermediate for all traits, except seed yield. Ward’s hierarchical clustering analysis generated six groups. AFLP analysis also revealed significant differences among the accessions. There was no correlation between the differences based on genetic markers and those based on morpho-agronomic and biochemical data, which indicates that both datasets are important for elucidating the differences among accessions. The results of the present study indicate great genetic diversity among the evaluated accessions.

Evaluation of genetic stability in olive callus-induced and meristem-induced shoots using flow cytometry and amplified fragment length polymorphism techniques

Background In vitro culture of olive, as an economically valuable tree, has fundamentally a genotype-dependant low micropropagation rate which needs to be improved in already established and newly released cultivars. Various plant tissue culture media, planting systems and growth factors were evaluated in two promissing Iranian olive cultivars ˈAminˈ and ˈMeshkatˈ and the commercial Spanish cultivar ˈArbequinaˈ. Results The results showed that cultivars have their specific optimal media, i.e. ˈAminˈ in the MS with 4 mg/L zeatin, ˈArbequinaˈ in the OM with 1 mg/L zeatin, and ˈMeshkatˈ in the OM and MS with 2 mg/L zeatin, which produced significantly a higher number of axillary shoots than other media. The results also indicated a significant improvement in the growth indices of ˈAminˈ (number of axillary shoots) when cultured using periodical mini bioreactor (PMB) in the VS medium. In comparison with VS, OM did not reveal any significant differences when both culturing systems (PMB and semi-solid media (SSM)) were used. Regarding the effect of carbon source and light intensity, mannitol and 2000 cd sr m−2 greatly enhanced ˈArbequinaˈ growth indices (main shoot length and growth quality). The results of genetic stability of callus induced shoots (CIS) and meristem induced shoots (MIS) revealed that 2C DNA value assessed by partec flow cytometery (FCM) had 0.01, 0.03 and 0.08 pg discrepencies in ˈAminˈ, ˈArbequinaˈ and ˈMeshkatˈ, repectively. The Amplified Fragment Length Polymorphism (AFLP) results also indicated that the cultivars were classified regardless of the micropropagation origin (CIS or MIS), except for ˈArbequinaˈ. The AFLP findings showed that ˈArbequinaˈ had the highest dispersal (7–38%) in CIS and MIS, while the Iranian cultivar of ˈMeshkatˈ (5–9%) had the highest stability. Conclusions This study indicated the importance of in vitro growth parameters for improving the micropropagation indices of olive cultivars. It showed that optimized protocols (OM, PMB, zeatin, mannitol and 2000 cd sr m−2) co-produced larger calli resulting in indirect organogenesis. Based on FCM and AFLP analysis, it can be concluded that true-to-typeness of micropropagated olive was cultivar-dependent.

Genetic and Epigenetic Changes Are Rapid Responses of the Genome to the Newly Synthesized Autotetraploid Carassius auratus

Whole genome duplication events have occurred frequently during the course of vertebrate evolution. To better understand the influence of polyploidization on the fish genome, we herein used the autotetraploid Carassius auratus (4n = 200, RRRR) (4nRR) resulting from the whole genome duplication of Carassius auratus (2n = 100, RR) (RCC) to explore the genomic and epigenetic alterations after polyploidization. We subsequently performed analyses of full-length transcriptome dataset, amplified fragment length polymorphism (AFLP) and methylation sensitive amplification polymorphism (MSAP) on 4nRR and RCC. By matching the results of 4nRR and RCC isoforms with reference genome in full-length transcriptome dataset, 649 and 1,971 novel genes were found in the RCC and 4nRR full-length geneset, respectively. Compared to Carassius auratus and Megalobrama amblycephala, 4nRR presented 3,661 unexpressed genes and 2,743 expressed genes. Furthermore, GO enrichment analysis of expressed genes in 4nRR revealed that they were enriched in meiosis I, whereas KEGG enrichment analysis displayed that they were mainly enriched in proteasome. Using AFLP analysis, we noted that 32.61% of RCC fragments had disappeared, while 32.79% of new bands were uncovered in 4nRR. Concerning DNA methylation, 4nRR exhibited a lower level of global DNA methylation than RCC. Additionally, 60.31% of methylation patterns in 4nRR were altered compared to RCC. These observations indicated that transcriptome alterations, genomic changes and regulation of DNA methylation levels and patterns had occurred in the newly established autotetraploid genomes, suggesting that genetic and epigenetic alterations were influenced by autotetraploidization. In summary, this study provides valuable novel insights into vertebrate genome evolution and generates relevant information for fish breeding.

Apple Autotetraploids with Enhanced Resistance to Apple Scab (Venturia inaequalis) Due to Genome Duplication-Phenotypic and Genetic Evaluation

Among the fungal diseases of apple trees, serious yield losses are due to an apple scab caused by Venturia inaequalis. Protection against this disease is based mainly on chemical treatments, which are currently very limited. Therefore, it is extremely important to introduce cultivars with reduced susceptibility to this pathogen. One of the important sources of variability for breeding is the process of polyploidization. Newly obtained polyploids may acquire new features, including increased resistance to diseases. In our earlier studies, numerous tetraploids have been obtained for several apple cultivars with ‘Free Redstar’ tetraploids manifesting enhanced resistance to apple scab. In the present study, tetraploids of ‘Free Redstar’ were assessed in terms of phenotype and genotype with particular emphasis on the genetic background of their increased resistance to apple scab. Compared to diploid plants, tetraploids (own-rooted plants) were characterized with poor growth, especially during first growing season. They had considerably shorter shoots, fewer branches, smaller stem diameter, and reshaped leaves. In contrast to own-rooted plants, in M9-grafted three-year old trees, no significant differences between diplo- and tetraploids were observed, either in morphological or physiological parameters, with the exceptions of the increased leaf thickness and chlorophyll content recorded in tetraploids. Significant differences between sibling tetraploid clones were recorded, particularly in leaf shape and some physiological parameters. The amplified fragment length polymorphism (AFLP) analysis confirmed genetic polymorphism of tetraploid clones. Methylation-sensitive amplification polymorphism (MSAP) analysis showed that the level of DNA methylation was twice as high in young tetraploid plants as in a diploid donor tree, which may explain the weaker vigour of neotetraploids in the early period of their growth in the juvenile phase. Molecular analysis showed that ‘Free Redstar’ cultivar and their tetraploids bear six Rvi genes (Rvi5, Rvi6, Rvi8, Rvi11, Rvi14 and Rvi17). Transcriptome analysis confirmed enhanced resistance to apple scab of ‘Free Redstar’ tetraploids since the expression levels of genes related to resistance were strongly enhanced in tetraploids compared to their diploid counterparts.

Virulence and Molecular Diversity in the Kabatiella zeae Population Causing Maize Eyespot in China

Maize eyespot, caused by Kabatiella zeae, has become a major yield-limiting factor in maize planting areas in northeast China. Limited information is available on pathotypes, virulence, and the genetic diversity of the K. zeae population. We analyzed virulence and genetic diversity of 103 K. zeae isolates collected from six provinces in China with differential hosts and the amplified fragment length polymorphism (AFLP) technique, respectively. To evaluate the virulence, 103 isolates were inoculated on nine differential hosts (maize inbred lines)—E28, Shen137, Qi319, B73, Danhuang34, Zi330, Mo17, Huangzaosi, and CN165—and grouped into 23 pathotypes and three virulence groups according to the coded triplet nomenclature system on differential hosts. AFLP analysis resolved the set of isolates into four genetic diversity clusters (DICE similarity values of 76%). Genetic variation of K. zeae among and between pathotypes revealed that the pathogen population had a high genotypic diversity. The correlation between pathotypes, virulence, and genetic diversity grouping was low. A correlation between AFLP groups and geographic locations was detected.

Vagococcus sp. a porcine pathogen: molecular and phenotypic characterization of strains isolated from diseased pigs in Brazil

Introduction: Vagococcus spp. is known for its importance as a systemic and zoonotic bacterial pathogen even though it is not often reported in pigs. This is related to the pathogen misidentification due to the lack of usage of more discriminatory diagnostic techniques. Here we present the first report of Vagococcus lutrae in swine and the characterization of Vagococcus fluvialis and Vagococcus lutrae isolated from diseased animals. Methodology: Between 2012 and 2017, 11 strains with morphological characteristics similar to Streptococcus spp. were isolated from pigs presenting different clinical signs. Bacterial identification was performed by matrix assisted laser desorption ionization time of flight (MALDI-TOF) mass spectrometry and confirmed by 16S rRNA sequencing and biochemical profile. Strains were further genotyped by single-enzyme amplified fragment length polymorphism (SE-AFLP). Broth microdilution was used to determine the minimal inhibitory concentration of the antimicrobials of veterinary interest. Results: Ten strains were identified as V. fluvialis and one was identified as V. lutrae. The SE-AFLP analysis enabled the species differentiation with specific clustering of all V. fluvialis separately from the V. lutrae strain. Most strains presented growth in the maximum antibiotic concentration values tested for eight of the 10 analyzed antimicrobial classes. Conclusions: The observed resistance pattern can represent a problem for veterinary and producers in the treatment of diseases associated Vagococcus spp. in swine production. Vagococcus species may also be a risk for pig industry workers. The data described here will be of great value in further understanding the behavior of this pathogen in animal production.