Development of microsatellite markers for the soft tick Ornithodoros phacochoerus.

BACKGROUND: Soft ticks of the genus Ornithodoros are responsible for the maintenance and transmission of the African swine fever (ASF) virus in the sylvatic and domestic viral cycles in Southern Africa. They are also the main vectors of the Borrelia species causing relapsing fevers. Currently, no genetic markers are available for Afrotropical Ornithodoros ticks. As ASF spreads globally, such markers are needed to assess the role of ticks in the emergence of new outbreaks. The aim of this study is to design microsatellite markers that could be used for ticks of the Ornithodoros moubata complex, particularly Ornithodoros phacochoerus, to assess population structure and tick movements in ASF endemic areas. METHODS: A total of 151 markers were designed using the O. moubata and O. porcinus genomes after elimination of repeated sequences in the genomes. All designed markers were tested on O. phacochoerus and O. porcinus DNA to select the best markers. RESULTS: A total of 24 microsatellite markers were genotyped on two populations of O. phacochoerus and on individuals from four other Ornithodoros species. Nineteen markers were selected to be as robust as possible for population genetic studies on O. phacochoerus. CONCLUSIONS: The microsatellite markers developed here represent the first genetic tool to study nidicolous populations of O. phacochoerus.

Population genetics assessment of two pocilloporid coral species from the northern red sea: Implications for urbanized reef sustainability.

Understanding the genetic makeup of key coral species is vital for effective coral reef management, as heightened genetic diversity directly influences long-term survival and resilience against environmental changes. This study focused on two widespread Indo-Pacific branching corals, Pocillopora damicornis (referred as Pocillopora cf. damicornis (as identified only morphologically) and Seriatopora hystrix, by genotyping 222 and 195 colonies, respectively, from 10 sites in the northern Gulf of Eilat, Red Sea, using six and five microsatellite markers, respectively. Both species exhibited low observed heterozygosity (0.47 for P. cf. damicornis, 0.32 for S. hystrix) and similar expected heterozygosity (0.576 for P. cf. damicornis, 0.578 for S. hystrix). Pocillopora cf. damicornis showed minimal deviations from Hardy-Weinberg equilibrium (HWE) and low but positive F values, indicating high gene flow, while S. hystrix exhibited higher diversion from HWE and positive F values, suggesting isolation by distance and possible non-random mating or genetic drift. As the Gulf of Eilat undergoes rapid urbanization, this study highlights the anthropogenic impacts on the population genetics of key ecosystem engineering species and emphasizes the importance of managing genetics of Marine Protected Areas while implementing active coral reef restoration. The differences in reproductive traits between the two species (S. hystrix being a brooder, while P. cf. damicornis a broadcast spawner), underscore the need for sustainable population genetics management of the coral reefs for the future and resilience of the coral reef ecosystem of the northern Red Sea region.

Pleistocene climate oscillations have shaped the expansion and contraction speciation model of the globose Eriosyce sect. Neoporteria cacti in Central Chile.

BACKGROUND AND AIMS: Pleistocene climatic oscillations, characterized by arid (interglacial) and pluvial (glacial) phases, have profoundly impacted the floras of Mediterranean climates. Our study investigates the hypothesis that these climatic extremes have promoted phases of range expansion and contraction in the Eriosyce sect. Neoporteria, resulting in pronounced genetic structuring and restricted gene flow. METHODS: Utilizing nuclear microsatellite markers, we genotyped 251 individuals across 18 populations, encompassing all 14 species and one subspecies within the Eriosyce sect. Neoporteria. Additionally, Species Distribution Models (SDMs) were employed to reconstruct past (Last Interglacial, Last Glacial Maximum, Mid-Holocene) and current potential distribution patterns, aiming to delineate the climatic influences on species' range dynamics. KEY RESULTS: The gene flow analysis disclosed disparate levels of genetic interchange among species, with marked restrictions observed between entities that are geographically or ecologically separated. Notably, E. subgibbosa from Hualpen emerged as genetically distinct, warranting its exclusion for clearer genetic clustering into north, central, and south clusters. The SDMs corroborated these findings, showing marked range expansions during warmer periods and contractions during colder times, indicating significant shifts in distribution patterns in response to climatic changes. CONCLUSIONS: Our findings emphasize the critical role of Pleistocene climatic fluctuations in driving the dynamic patterns of range expansions and contractions that have led to geographic isolation and speciation within the Eriosyce sect. Neoporteria. Even in the face of ongoing gene flow, these climate-driven processes have played a pivotal role in sculpting the species' genetic architecture and diversity. This study elucidates the complex interplay between climatic variability and evolutionary dynamics among Mediterranean cacti in central Chile, highlighting the necessity of considering historical climatic millenial oscillations in conservation and evolutionary biology studies.

Genetic Diversity and Population Structure of the Invasive Oriental Fruit Fly, Bactrocera dorsalis (Diptera: Tephritidae) in Burkina Faso.

Bactrocera dorsalis Hendel is a highly invasive horticultural pest that is of major economic importance worldwide. In Burkina Faso, it is one of the main insect pests that affects the production and exportation of mangos. Understanding the biology and the genetic dynamics of this insect pest provides crucial information for the development of effective control measures. The aim of this study was to understand the distribution, diversity, and genetic structure of B. dorsalis in Burkina Faso. Male flies were collected transversally in Burkina Faso and analyzed by PCR using 10 microsatellite markers. The results showed an abundance of B. dorsalis varying from 87 to 2986 flies per trap per day at the different sampling sites. The genetic diversity was high at all sites, with an average Shannon's Information Index (I) of 0.72 per site. The gene flow was high between study populations and ranged from 10.62 to 27.53 migrants. Bayesian admixture analysis showed no evidence of structure, while Discriminant Analysis of Principal Components identified three weakly separated clusters in the population of B. dorsalis in Burkina Faso. The results of this study could be used to optimize the effectiveness of current control interventions and to guide the implementation of new, innovative, and sustainable strategies.

Genetic Structure of Introduced Plasmodium vivax Malaria Isolates in Greece, 2015-2019.

Greece has been malaria-free since 1974, after an intense malaria control program. However, as Greece hosts migrant populations from P. vivax malaria-endemic countries, there is a risk of introducing the disease to specific vulnerable and receptive areas of the country. Knowledge of the genetic diversity of P. vivax populations is essential for understanding the dynamics of malaria disease transmission in a given region. We used nine highly polymorphic markers to genotype 124 P. vivax-infected archived DNA samples from human blood specimens referred to the NMRL from all over Greece throughout 2015-2019. The genotypic variability of the samples studied was noted, as they comprised several unique haplotypes, indicative of the importation of a large number of different P. vivax strains in the country. However, only a few events of local transmission were recorded. Genotyping revealed and confirmed the same clusters as those identified through epidemiological investigation. In only one introduction event was the index case found. No sustained/ongoing malaria transmissions in/between the studied regions or during consecutive years or additional foci of local transmission were observed. Genotyping is an important component in assisting malaria surveillance, as it provides information concerning the patterns of introduction and the effectiveness of implemented malaria control and elimination measures.

Molecular Diversity and Combining Ability in Newly Developed Maize Inbred Lines under Low-Nitrogen Conditions.

Nitrogen is an essential element for maize growth, but excessive application can lead to various environmental and ecological issues, including water pollution, air pollution, greenhouse gas emissions, and biodiversity loss. Hence, developing maize hybrids resilient to low-N conditions is vital for sustainable agriculture, particularly in nitrogen-deficient soils. Combining ability and genetic relationships among parental lines is crucial for breeding superior hybrids under diverse nitrogen levels. This study aimed to assess the genetic diversity of maize inbred lines using simple sequence repeat (SSR) markers and evaluate their combining ability to identify superior hybrids under low-N and recommended conditions. Local and exotic inbred lines were genotyped using SSR markers, revealing substantial genetic variation with high gene diversity (He = 0.60), moderate polymorphism information content (PIC = 0.54), and an average of 3.64 alleles per locus. Twenty-one F1 hybrids were generated through a diallel mating design using these diverse lines. These hybrids and a high yielding commercial check (SC-131) were field-tested under low-N and recommended N conditions. Significant variations (p < 0.01) were observed among nitrogen levels, hybrids, and their interaction for all recorded traits. Additive genetic variances predominated over non-additive genetic variances for grain yield and most traits. Inbred IL3 emerged as an effective combiner for developing early maturing genotypes with lower ear placement. Additionally, inbreds IL1, IL2, and IL3 showed promise as superior combiners for enhancing grain yield and related traits under both low-N and recommended conditions. Notably, hybrids IL1×IL4, IL2×IL5, IL2×IL6, and IL5×IL7 exhibited specific combining abilities for increasing grain yield and associated traits under low-N stress conditions. Furthermore, strong positive associations were identified between grain yield and specific traits like plant height, ear length, number of rows per ear, and number of kernels per row. Due to their straightforward measurability, these relationships underscore the potential of using these traits as proxies for indirect selection in early breeding generations, particularly under low-N stress. This research contributes to breeding nitrogen-efficient maize hybrids and advances our understanding of the genetic foundations for tolerance to nitrogen limitations.

Morphological and genetic characterization of jackfruit (Artocarpus heterophyllus) in the Kayunga and Luwero districts of Uganda.

BACKGROUND: Jackfruit (Artocarpus heterophyllus) is an economically valuable fruit tree in Uganda. However, the production of jackfruit in Uganda is low. Additionally, because of deforestation, genetic erosion of the resource is predicted before its exploitation for crop improvement and conservation. As a prerequisite for crop improvement and conservation, 100 A. heterophyllus tree isolates from the Kayunga and Luwero districts in Uganda were characterized using 16 morphological and 10 microsatellite markers. RESULTS: The results from the morphological analysis revealed variations in tree height, diameter at breast height (DBH), and crown diameter, with coefficient of variation (CV) values of 20%, 41%, and 33%, respectively. Apart from the pulp taste, variation was also observed in qualitative traits, including tree vigor, trunk surface, branching density, tree growth habit, crown shape, leaf blade shape, fruit shape, fruit surface, flake shape, flake color, flake flavor and pulp consistency/texture. Genotyping revealed that the number of alleles amplified per microsatellite locus ranged from 2 to 5, with an average of 2.90 and a total of 29. The mean observed (Ho) and expected (He) heterozygosity were 0.71 and 0.57, respectively. Analysis of molecular variance (AMOVA) indicated that 81% of the variation occurred within individual trees, 19% among trees within populations and 0% between the two populations. The gene flow (Nm) in the two populations was 88.72. The results from the 'partitioning around medoids' (PAM), principal coordinate analysis (PCoA) and genetic cluster analysis further revealed no differentiation of the jackfruit populations. The Mantel test revealed a negligible correlation between the morphological and genetic distances. CONCLUSIONS: Both morphological and genetic analyses revealed variation in jackfruit within a single interbreeding population. This diversity can be exploited to establish breeding and conservation strategies to increase the production of jackfruit and hence boost farmers' incomes. However, selecting germplasm based on morphology alone may be misleading.

Genetic variation and structure of endemic and endangered wild celery (Kelussia odoratissima Mozaff.) quantified using novel microsatellite markers developed by next-generation sequencing.

Kelussia odoratissima Mozaff. (Apiaceae) is a native plant that has been traditionally consumed in Iran's food and pharmaceutical industries. Overharvesting of the taxon, especially at the beginning of the growing season, due to its considerable medicinal and economic value, is believed to be the main reason for the extirpating of this plant. The consequences of the severe anthropogenic impacts on the genetic diversity of populations are poorly known. In order to investigate the level of genetic variation and patterns of the genetic structure of K. odoratissima, we developed novel microsatellite markers using the 454 Roche next-generation sequencing (NGS) platform for the first time. Out of 1,165 microsatellite markers bioinformatically confirmed, twenty-five were tested, of which 23 were used to screen genetic variation across 12 natural populations. Our results showed that the average number of alleles per locus and the polymorphic information content (PIC) were 10.87 (range 7 to 27), and 0.81 (range 0.67 to 0.94), respectively. The mean observed and expected heterozygosities (± SD) across all populations were 0.80 ± 0.31 and 0.72 ± 0.14, respectively. The average pairwise FST among the populations was 0.37 (range 0.04 to 0.81). Bayesian and distance-based clustering, and principal coordinate analyses revealed at least four major genetic clusters. Although high level of structure can be explained by landscape topography and geographic distance, presence of admixed populations can be associated to seed or pollen dispersal. Contrary to expectations, the high level of genetic variation and lack of inbreeding suggest that overexploitation has not yet significantly purged the allelic variability within the natural populations in protected areas.

Integrated Discriminant Evaluation of Molecular Genetic Markers and Genetic Diversity Parameters of Endangered Balearic Dog Breeds.

The genetic diversity analysis of six dog breeds, including Ca de Bestiar (CB), Ca de Bou (CBOU), Podenco Ibicenco (PI), Ca Rater (CR), Ca Mè (CM), and Ca de Conills (CC), reveals insightful findings. CB showcases the highest mean number of alleles (6.17) and heterozygosity values, with significant deviations from Hardy-Weinberg equilibrium (HWE) observed in five markers, indicating high intra-racial genetic diversity (average observed heterozygosity (Ho) = 0.754, expected heterozygosity (He) = 0.761). In contrast, CBOU presents the lowest mean number of alleles (5.05) and heterozygosity values, coupled with moderate polymorphic information content (PIC) values and a moderate level of intra-racial genetic diversity (average Ho = 0.313, He = 0.394). PI demonstrates moderate genetic diversity with an average of 5.75 alleles and highly informative PIC values, while CR displays robust genetic diversity with an average of 6.61 alleles and deviations from equilibrium, indicating potential risks of inbreeding (average Ho = 0.563, He = 0.658). CM exhibits moderate genetic diversity and deviations from equilibrium, similar to CBOU, with an average of 6.5 alleles and moderate PIC values (average Ho = 0.598, He = 0.676). Conversely, CC shows a wider range of allelic diversity and deviations from equilibrium (average Ho = 0.611, He = 0.706), suggesting a more diverse genetic background. Inter-racial analysis underscores distinct genetic differentiation between breeds, emphasizing the importance of informed breeding decisions and proactive genetic management strategies to preserve diversity, promote breed health, and ensure long-term sustainability across all breeds studied.

Genetic resources of common ash (Fraxinus excelsior L.) in Poland.

BACKGROUND: Knowledge of genetic structure and the factors that shape it has an impact on forest management practices. European ash (Fraxinus excelsior L.) has declined dramatically throughout its range as a result of a disease caused by the fungus Hymenoscyphus fraxineus. Despite the need for conservation and restoration of the species, genetic data required to guide these efforts at the country level are scarce. Thereofore, we studied the chloroplast and nuclear genetic diversity of 26 natural common ash populations (1269 trees) in Poland. RESULTS: Chloroplast polymorphisms grouped the populations into two geographically structured phylogenetic lineages ascribed to different glacial refugia (the Balkans and the Eastern Alps). However, the populations demonstrated high genetic diversity (mean AR = 12.35; mean Ho = 0.769; mean He = 0.542) but low differentiation based on nuclear microsatellites (FST = 0.045). Significant spatial genetic structure, consistent with models of isolation by distance, was detected in 14 out of 23 populations. Estimated effective population size was moderate-to-high, with a harmonic mean of 57.5 individuals per population. CONCLUSIONS: Genetic diversity was not homogeneously distributed among populations within phylogenetic gene pools, indicating that ash populations are not equal as potential sources of reproductive material. Genetic differences among populations could be related to their histories, including founder effects or gene flow between evolutionary lineages (admixture). Our results suggest that ash stands across Poland could be treated as two main management units (seed zones). Therefore, despite the homogenizing effect of pollen gene flow known for this species, the genetic structure should be taken into account in the management of the genetic resources of the common ash. Although ash dieback poses an additional challenge for the management of genetic resources, efforts should be directed towards protecting populations with high genetic diversity within defined phylogenetic units, as they may be an important source of adaptive variation for future stands.

Flight Dispersal in Supratidal Rockpool Beetles.

Flight dispersal is ecologically relevant for the survival of supratidal rockpool insects. Dispersal has important consequences for colonisation, gene flow, and evolutionary divergence. Here, we compared the flight dispersal capacity of two congeneric beetle species (Ochthebius quadricollis and Ochthebius lejolisii) that exclusively inhabit these temporary, fragmented, and extreme habitats. We estimated flight capacity and inferred dispersal in both species using different approaches: experimental flying assays, examination of wing morphology, and comparison of microsatellite markers between species. Our findings revealed that both species exhibited similar flight behaviour, with 60 to 80% of the individuals flying under water heating conditions. Notably, females of both species had larger body sizes and wing areas, along with lower wing loading, than males in O. quadricollis. These morphological traits are related to higher dispersal capacity and more energetically efficient flight, which could indicate a female-biassed dispersal pattern. The wing shapes of both species are characterised by relatively larger and narrower wings in relation to other species of the genus, suggesting high flight capacity at short distances. Molecular data revealed in both species low genetic divergences between neighbouring populations, non-significant differences between species, and no isolation by distance effect at the study scale (<100 km). These results point to passive dispersal assisted by wind.

Simultaneous determination of subspecies and geographic origins of 110 rice cultivars by microsatellite markers.

Rice varieties of different subspecies types (indica rice and japonica rice) across various geographical origins (Hunan, Jiangsu, and Northeast China) were monitored using microsatellite markers (simple sequence repeats, SSR). 110 representative rice cultivars were collected from the main crop areas. Multiple methods including clustering analysis (neighbor-joining (NJ) method, unweighted pair-group method with arithmetic mean (UPGMA) method), principal component analysis (PCA) and model-based grouping were applied. The study revealed that 25 pairs of SSR markers exhibited a broad range of polymorphism information content (PIC) values, ranging from 0.240 to 0.830. Furthermore, our study successfully achieved a higher overall mean correct rate of 99.09% in determining the geographical origin of rice. Simultaneously, it accurately classified indica rice and japonica rice. These findings are significant as they provide an SSR fingerprint of 110 high-quality rice cultivars, serving as a valuable scientific resource for the detection of rice adulteration and traceability of its origin.

Diversity assessment of a lesser known buffalo population from Central India and its comparative evaluation reveals presence of sufficient genetic variation and absence of selection.

Planned breeding and conservation strategies for a lesser-known population require an assessment of complete genetic diversity and population structure analysis in addition to its morphometric characteristics. In the present study, a comparative analysis of the genetic structure of a rare buffalo population, namely Chhattisgarhi, was extensively studied using a panel of FAO-recommended microsatellite markers along with well-established breeds namely Murrah, Nili-Ravi, Gojri, Kalahandi, and Nagpuri. Mode shift analysis indicated the absence of genetic bottleneck in the recent past. Assessment of genetic diversity indices across all loci indicated the presence of sufficient genetic variation within and between populations. Analysis of molecular variance between the six different buffalo populations attributed 19.05% of the variations to between-population differentiation. Cluster analyses using DAPC and Bayesian approach along with the phylogenetic tree based on UPGMA grouped six populations into three groups. The Chhattisgarhi population was revealed to be genetically closer to Nagpuri and Kalahandi populations. The study reveals the presence of sufficient genetic diversity within the Chhattisgarhi population and indicates the absence of a systematic selection program. We suggest improvement and conservation programs should be planned for this breed in the near future through short-term selection.

Wheat Stem Rust Surveillance Reveals Two New Races of Puccinia graminis f. sp. tritici in South Africa During 2016 to 2020.

Stem rust, caused by Puccinia graminis f. sp. tritici (Pgt), is an important disease of wheat in South Africa (SA) and is primarily controlled using resistant cultivars. Understanding virulence diversity of Pgt is essential for successful breeding of resistant cultivars. Samples of infected wheat stems were collected across the major wheat-growing regions of SA from 2016 to 2020 to determine the pathogenic variability of Pgt isolates. Seven races were identified from 517 isolates pathotyped. The most frequently found races were 2SA104 (BPGSC + Sr9h,27,Kw) (35% frequency) and 2SA88 (TTKSF + Sr8b) (33%). Race 2SA42 (PTKSK + Sr8b), which was found in 2017, and 2SA5 (BFGSF + Sr9h), identified in 2017, are new races. The Ug99 variant race 2SA42 is similar in its virulence to 2SA107 (PTKST + Sr8b) except for avirulence to Sr24 and virulence to Sr8155B1. Race 2SA5 is closely related in its virulence to existing races that commonly infect triticale. Certain races showed limited geographical distribution. Races 2SA5, 2SA105, and 2SA108 were found only in the Western Cape, whereas 2SA107 and 2SA42 were detected only in the Free State province. The new and existing races were compared using microsatellite (SSR) marker analysis and their virulence on commercial cultivars was also determined. Seedling response of 113 wheat entries against the new races, using 2SA88, 2SA88+9h, 2SA106, and 2SA107 as controls, revealed 2SA107 as the most virulent (67 entries susceptible), followed by 2SA42 (64), 2SA106 (60), 2SA88+9h (59), 2SA88 (25), and 2SA5 (17). Thus, 2SA5 may not pose a significant threat to local wheat production. SSR genotyping revealed that 2SA5 is genetically distinct from all other SA Pgt races.

Effect of Cucurbit Host, Production Region, and Season on the Population Structure of Pseudoperonospora cubensis in Florida.

Pseudoperonospora cubensis, the causal agent of Cucurbit downy mildew (CDM), is one of the most important diseases affecting cucurbit production in the United States. This disease is especially damaging to Florida production areas, as the state is a top producer of many cucurbit species. In addition, winter production in central and south Florida likely serves as a likely source of P. cubensis inoculum for spring and summer cucurbit production throughout the eastern United States, where CDM is unable to overwinter in the absence of a living host. Over 2 years (2017 and 2018) and four seasons (spring 2017, spring 2018, fall 2017, and fall 2018), 274 P. cubensis isolates were collected from cucurbit hosts at production sites in south, central, and north Florida. The isolates were analyzed with 10 simple sequence repeat (SSR) markers to establish population structure and genetic diversity and further assigned to a clade based on a qPCR assay. Results of population structure and genetic diversity analyses differentiated isolates based on cucurbit host and clade (1 or 2). Of the isolates assigned to clade by qPCR, butternut squash, watermelon, and zucchini were dominated by clade 1 isolates, whereas cucumber isolates were split 34 and 59% between clades 1 and 2, respectively. Clade assignments agreed with isolate clustering observed within discriminant analysis of principal components (DAPC) based on SSR markers, although watermelon isolates formed a group distinct from the other clade 1 isolates. For seasonal collections from cucumber at each location, isolates were typically skewed to one clade or the other and varied across locations and seasons within each year of the study. This variable population structure of cucumber isolates could have consequences for regional disease management. This is the first study to characterize P. cubensis populations in Florida and evaluate the effect of cucurbit host and clade-type on isolate diversity and population structure, with implications for CDM management in Florida and other United States cucurbit production areas.

Development and Application of Potentially Universal Microsatellite Markers for Pheasant Species.

Pheasants are widely distributed in the southwest of China, but many of them are endangered due to habitat fragmentation and environmental changes. Genetic diversity is crucial for species to maintain their evolutionary potential, and thus it is important to develop universal genetic markers for facilitating the assessment of genetic diversity and planning effective conservation actions in these endangered species. In this study, 471 microsatellite loci which are common among eight pheasant species were screened based on genome data, and 119 loci were selected to develop microsatellite markers. After PCR amplifications and reaction condition optimizations, and validation of microsatellite loci in 14 species of 11 genera within Phasianidae. Finally, 49 potentially universal microsatellite markers in pheasant species were obtained. These microsatellite markers were successfully applied to assess the genetic diversity of 3 pheasant species. The Sichuan hill partridge (Arborophila rufipectus), blood pheasant (Ithaginis cruentus), buff-throated partridge (Tetraophasis szechenyii) and Sichuan hill partridge had a relatively low genetic diversity level. These 49 microsatellite loci are potentially universal microsatellite loci for pheasants and are of great significance to establish a shared platform in population genetics study of pheasants.

Creation of a Biobank of the Sperm of the Honey Bee Drones of Different Subspecies of Apis mellifera L.

The cryopreservation of gametes and embryos is an important element of biodiversity conservation. One species in need of conservation is the honey bee Apis mellifera L. Changing environmental factors, especially the anthropogenic factor, have led to a reduction in the numbers of this insect species. In this study, we provide an example of the creation of a biobank of honey bee drone sperm. For sperm cryopreservation, drones of the most common subspecies of honey bees common in Russia were selected. These were the dark forest bee, Apis mellifera mellifera, from the Republic of Bashkortostan, with three subspecies (A. m. carnica, A. m. carpatica, and A. m. caucasica) from the southern regions of Russia, as well as two breeding stocks, the Far Eastern bee and Prioksky bee. For subspecies identification, morphometric and genetic methods were used. The subspecies of the studied samples were confirmed via the analysis of the tRNAleu-COII locus of mitochondrial DNA and nine microsatellite markers of nuclear DNA. It was shown that bees of the Prioksky breeding stock belong to the subspecies A. m. caucasica based on phylogenetic analysis, and the Far Eastern breeding stock is a stable hybrid, descending on the maternal line from the evolutionary lineage C or O. The results of the morphometric analysis are consistent with the results of the genetic analysis. For the cryopreservation of sperm, we used a cryoprotectant solution with honey. As a result, the viability of frozen-thawed sperm decreased by 20.3% compared to fresh sperm, and overall motility decreased 25-fold. The measurement of the sperm concentration in the spermatheca of artificially inseminated queens showed that it varied from 0.22 to 4.4 million/µL. Therefore, the use of honey in sperm cryopreservation has great potential.

Association Mapping for Evaluation of Population Structure, Genetic Diversity, and Physiochemical Traits in Drought-Stressed Maize Germplasm Using SSR Markers.

Globally, maize is one of the most consumed crops along with rice and wheat. However, maize is sensitive to different abiotic stress factors, such as drought, which have a significant impact on its production. The aims of this study were to investigate (1) genetic variation among 41 maize-inbred lines and the relationships among them and (2) significant marker-trait associations (SMTAs) between 7 selected physiochemical traits and 200 simple sequence repeat (SSR) markers to examine the genetics of these traits. A total of 1023 alleles were identified among the 41 maize-inbred lines using the 200 SSR loci, with a mean of 5.1 alleles per locus. The average major allele frequency, gene diversity, and polymorphism information content were 0.498, 0.627, and 0.579, respectively. The population structure analysis based on the 200 SSR loci divided the maize germplasm into two primary groups with an admixed group. Moreover, this study identified, respectively, 85 SMTAs and 31 SMTAs using a general linear model (Q GLM) and a mixed linear model (Q + K MLM) with statistically significant (p < 0.05 and <0.01) associations with the seven physiochemical traits (caffeic acid content, chlorogenic acid content, gallic acid content, ferulic acid content, 2,2-diphenyl-1-picrylhydrazyl free radical scavenging activity, leaf relative moisture content, total phenolic content). These SSR markers were highly correlated with one or more of the seven physiochemical traits. This study provides insights into the genetics of the 41 maize-inbred lines and their seven physiochemical traits and will be of assistance to breeders in the marker-assisted selection of maize for breeding programs.

Development of a novel molecular tool to study molecular ecology of Ornithomya (Hippoboscidae) avian louse flies.

Louse flies (Diptera: Hippoboscidae) are obligatory hematophagous ectoparasites of birds and mammals. These widely distributed parasitic flies may have a significant impact on wild and farm animals by feeding on their blood and transmitting bloodborne pathogens. However, despite their ecological importance, louse flies are clearly underrepresented in host-parasite research and implementation of genetic approaches in this group is generally hampered by lacking molecular tools. In addition, louse flies that parasitize long-distance migrants can travel long distances with their avian hosts, facilitating the large-scale spread of pathogens across landscapes and geographic regions. Given the wide diversity of louse flies that parasitize a variety of avian hosts, their direct negative impact on host survival, and their high potential to transmit bloodborne pathogens even along avian migration routes, it is surprising that our knowledge of louse fly ecology is rather modest and incomplete. Here, we aimed to develop a novel molecular tool for polyxenous avian louse flies from the genus Ornithomya, which are among the most common and widely distributed representatives of Hippoboscidae family, to improve research of their genetic population structure and molecular ecology. Using the Illumina Mi-seq sequencing, we conducted a genome-wide scan in Ornithomya avicularia to identify putative microsatellite markers. A panel of 26 markers was selected to develop amplification protocols and assess polymorphism in the Central European population of O. avicularia, as well as to test for cross-amplification in a congeneric species (O. chloropus). A genome-scan in O. avicularia identified over 12 thousand putative microsatellite markers. Among 26 markers selected for a population-wide screening; one did not amplify successfully and three were monomorphic. 22 markers were polymorphic with at least two alleles detected. Two markers showed presence of null alleles. A cross-amplification of microsatellite markers in O. chloropus revealed allelic polymorphism at 14 loci, with the mean allelic richness of 3.78 alleles per locus (range: 2-8). Our genome-wide scan in O. avicularia provides a novel and powerful tool for molecular research in Ornithomya louse flies. Our panel of polymorphic microsatellite loci should allow genotyping of louse flies from geographically distinct populations and from a wide spectrum of avian hosts, enhancing population genetic and phylogeographic research in Ornithomya.

Novel Microsatellite Loci, Cross-Species Validation of Multiplex Assays, and By-Catch Mitochondrial Genomes on Ochthebius Beetles from Supratidal Rockpools.

Here we focus on designing, for the first time, microsatellite markers for evolutionary and ecological research on aquatic beetles from the genus Ochthebius (Coleoptera, Hydraenidae). Some of these non-model species, with high cryptic diversity, exclusively inhabit supratidal rockpools, extreme and highly dynamic habitats with important anthropogenic threats. We analysed 15 individuals of four species (O. lejolisii, O. subinteger, O. celatus, and O. quadricollis) across 10 localities from the Mediterranean coasts of Spain and Malta. Using next-generation sequencing technology, two libraries were constructed to interpret the species of the two subgenera present consistently (Ochthebius s. str., O. quadricollis; and Cobalius, the rest of the species). Finally, 20 markers (10 for each subgenus) were obtained and successfully tested by cross-validation in the four species under study. As a by-catch, we could retrieve the complete mitochondrial genomes of O. lejolisii, O. quadricollis, and O. subinteger. Interestingly, the mitochondrial genome of O. quadricollis exhibited high genetic variability compared to already published data. The novel SSR panels and mitochondrial genomes for Ochthebius will be valuable in future research on species identification, diversity, genetic structure, and population connectivity in highly dynamic and threatened habitats such as supratidal coastal rockpools.