Respective Progeny Research Articles

The Role of a Soybean 14-3-3 Gene (Glyma05g29080) on White Mold Resistance and Nodulation Investigations Using CRISPR-Cas9 Editing and RNA Silencing.

The role of a soybean 14-3-3 gene (Glyma05g29080) in defense against white mold and in nodulation was investigated by loss-of-gene-function with CRISPR-Cas9 editing and silencing of RNA interference (RNAi). Particle bombardment was used to introduce the CRISPR expression cassette to target the soybean 14-3-3 gene and an RNAi construct to silence gene transcription. Transmission of the edited 14-3-3 gene and the RNAi construct was confirmed in their respective progeny. The recovered transgenic plants and their progeny were significantly more susceptible to Sclerotinia sclerotiorum infection and showed a significant reduction in nodulation, thus confirming the role of the 14-3-3 gene (Glyma05g29080) in both nodulation and defense.

Genetic variability of indigenous (Quercus robur L.) and late flushing oak (Quercus robur L. subsp. slavonica (Gáyer) Mátyás) in adult stands compared with their natural regeneration

Slavonian oak (Quercus robur subsp. slavonica (Gáyer) Mátyás) is currently gaining interest in forestry due to forest restructuring in Germany caused by climate change. Slavonian oaks originating from Croatia have been introduced into Germany mainly in the Münsterland region of North Rhine-Westphalia since the second half of the nineteenth century. They are characterized by their late bud burst, long clear bole, stem straightness and faster height and diameter growth compared to indigenous oaks in Germany. In this study, the genetic differentiation of adult trees and their respective progeny of two Slavonian and two indigenous stands in Hamm-Westtünnen, was evaluated. Genetic diversity and structure were estimated using 23 nuclear simple sequence repeat (SSRs) and 5 maternally inherited chloroplast microsatellite markers (cpSSRs). The mean expected heterozygosity of 0.545 and allelic richness of 6.23 indicate high genetic diversity in the studied populations. The group of progenies (AR = 8.40, Ho = 0.524, He = 0.559, FIS = 0.064) shows similar levels of genetic variation as the adult stands (AR = 8.37, Ho = 0.513, He = 0.554, FIS = 0.075). The genetic differentiation between adult stands and progeny was low (FST = 0.013). Genetic assignment of individuals using STRUCTURE revealed that the studied populations were divided into two clusters. There was no evidence of extensive hybridization or gene flow between Slavonian and native populations, possibly due to the different timing of bud burst of the two taxa.

Rate of Transfer of Infectious Anaemia Maternal Antibodies from Broiler Breeders To the Progeny: a Field Evaluation

Clinical manifestation of the disease caused by the chicken anaemia virus (CAV) occurs when chicken chicks are vertically contaminated or before the second week of life. CAV control is based on the vaccination of broiler breeders in order to promote progeny protection through maternal antibodies. This work aims to evaluate, under field conditions, the antibody title in commercial broiler breeders at 28, 48, and 68 weeks of age, the rate of transference to the progeny, as well as the duration of antibodies in the progeny up to 21 days of age. Thus, a total of 92 sera samples from 93,000 broiler breeders vaccinated with a live vaccine for CAV at 14 weeks of age and 366 sera samples from their respective progeny were analyzed using ELISA. Breeders’ antibody title for CAV ranged between 5051 and 8660, and these titles may provide sufficient protection for their progeny. On average, 63% of the maternal antibodies were transferred to the progeny and lasted up to the second week of chick’s life. It is possible to conclude that the vaccine and the vaccination procedure used by this company for breeders against CAV seems to be effective in inducing high antibody levels in the breeders and transfering protective maternal antibodies to the progeny.

Early warming stress on rainbow trout juveniles impairs male reproduction but contrastingly elicits intergenerational thermotolerance

The exposure of adult fish to warm or high temperatures is known to impair reproduction, yet the long-term reproductive impacts for treatments at early life are not well clarified. This study aimed to evaluate the effects of warm temperature (WT) during juvenile stage on gonad maturation, gamete quality, and offspring thermotolerance in rainbow trout. While the comparison of basic reproductive parameters in WT females did not reveal any kind of impairment, many WT males showed an atrophied, undeveloped gonad, or a smaller testis with lower milt volume; sperm quality parameters in WT males and deformity rates in the respective progeny were also highly affected. However, despite of such negative effects, many of the remaining progeny presented better rates of survival and growth when exposed to the same conditions as those of parental fish (WT), suggesting that thermal stress in parr stage males elicited intergenerational thermotolerance after a single generation. The present results support that prolonged warming stress during early life stages can adversely affect key reproductive aspects, but contrastingly increase offspring performance at upper thermal ranges. These findings have implications on the capacity of fish to adapt and to cope with global warming.

Disease-Relevant Single Cell Photonic Signatures Identify S100\u03b2 Stem Cells and their Myogenic Progeny in Vascular Lesions

A hallmark of subclinical atherosclerosis is the accumulation of vascular smooth muscle cell (SMC)-like cells leading to intimal thickening and lesion formation. While medial SMCs contribute to vascular lesions, the involvement of resident vascular stem cells (vSCs) remains unclear. We evaluated single cell photonics as a discriminator of cell phenotype in vitro before the presence of vSC within vascular lesions was assessed ex vivo using supervised machine learning and further validated using lineage tracing analysis. Using a novel lab-on-a-Disk(Load) platform, label-free single cell photonic emissions from normal and injured vessels ex vivo were interrogated and compared to freshly isolated aortic SMCs, cultured Movas SMCs, macrophages, B-cells, S100β+ mVSc, bone marrow derived mesenchymal stem cells (MSC) and their respective myogenic progeny across five broadband light wavelengths (λ465 - λ670 ± 20 nm). We found that profiles were of sufficient coverage, specificity, and quality to clearly distinguish medial SMCs from different vascular beds (carotid vs aorta), discriminate normal carotid medial SMCs from lesional SMC-like cells ex vivo following flow restriction, and identify SMC differentiation of a series of multipotent stem cells following treatment with transforming growth factor beta 1 (TGF- β1), the Notch ligand Jagged1, and Sonic Hedgehog using multivariate analysis, in part, due to photonic emissions from enhanced collagen III and elastin expression. Supervised machine learning supported genetic lineage tracing analysis of S100β+ vSCs and identified the presence of S100β+vSC-derived myogenic progeny within vascular lesions. We conclude disease-relevant photonic signatures may have predictive value for vascular disease.Graphical abstract

Radiation-induced toxicity in rectal epithelial stem cell contributes to acute radiation injury in rectum

BackgroundRadiation-induced rectal epithelial damage is a very common side effect of pelvic radiotherapy and often compromise the life quality and treatment outcome in patients with pelvic malignancies. Unlike small bowel and colon, effect of radiation in rectal stem cells has not been explored extensively. Here we demonstrate that Lgr5-positive rectal stem cells are radiosensitive and organoid-based transplantation of rectal stem cells mitigates radiation damage in rectum.MethodsC57Bl6 male mice (JAX) at 24 h were exposed to pelvic irradiation (PIR) to determine the radiation effect in pelvic epithelium. Effect of PIR on Lgr5-positive rectal stem cells (RSCs) was determined in Lgr5-EGFP-Cre-ERT2 mice exposed to PIR. Effect of PIR or clinically relevant fractionated PIR on regenerative response of Lgr5-positive RSCs was examined by lineage tracing assay using Lgr5-eGFP-IRES-CreERT2; Rosa26-CAG-tdTomato mice with tamoxifen administration to activate Cre recombinase and thereby marking the ISC and their respective progeny. Ex vivo three-dimensional organoid cultures were developed from Lgr5-EGFP-Cre-ERT2 mice. Organoid growth was determined by quantifying the budding crypt/total crypt ratio. Organoids from Lgr5-EGFP-ires-CreERT2-TdT mice were transplanted in C57Bl6 male mice exposed to PIR. Engraftment and repopulation of Lgr5-positive RSCs were determined after tamoxifen administration to activate Cre recombinase in recipient mice. Statistical analysis was performed using Log-rank (Mantel-Cox) test and paired two-tail t test.ResultExposure to pelvic irradiation significantly damaged rectal epithelium with the loss of Lgr5+ve rectal stem cells. Radiosensitivity of rectal epithelium was also observed with exposure to clinically relevant fractionated pelvic irradiation. Regenerative capacity of Lgr5+ve rectal stem cells was compromised in response to fractionated pelvic irradiation. Ex vivo organoid study demonstrated that Lgr5+ve rectal stem cells are sensitive to both single and fractionated radiation. Organoid-based transplantation of Lgr5+ve rectal stem cells promotes repair and regeneration of rectal epithelium.ConclusionLgr5-positive rectal stem cells are radiosensitive and contribute to radiation-induced rectal epithelial toxicity. Transplantation of Lgr5-positive rectal stem cells mitigates radiation-induced rectal injury and promotes repair and regeneration process in rectum.

Distinguishing mutations and null alleles from genotyping errors using mother progeny comparisons in Brazilian pine (Araucaria angustifolia)

The use of microsatellite markers provides a window into the evolutionary processes of a given species. As such, these markers are widely used in scientific and applied research and are praised for their practicality and ease of use; however, the unavoidable incidence of genotyping deviations has been broadly neglected in the literature. Therefore, the present study aimed to estimate the rate of null alleles, mutations, and genotyping errors in microsatellite loci, using Araucaria angustifolia, a threatened species, as a case study. We estimated the rates of the different types of genotyping deviations using mother-progeny genotype comparison from 50 seed-trees and their respective progeny (seeds). A total of 2336 A. angustifolia samples were genotyped, and we found that the rate of null alleles was 0.045. From the 1972 mother-progeny comparisons, the overall genotype deviation rate was 1.58%, consisting of 145 inconsistences (mutations), 339 null alleles, and 210 genotyping errors. In terms of seed numbers, 128 (6.5%) showed inconsistencies in at least one locus, 118 (6.0%) null alleles, and 321 (16.3%) genotyping errors. This is the first study to describe the inconsistences (mutations) between mother-progeny genotypes for A. angustifolia, and the outcome makes it clear that an understanding of these genotyping deviations must be considered in assessing the accuracy of inferences made based on population genetics analyses.

Temporal plasticity of apical progenitors in the developing mouse neocortex.

The diverse subtypes of excitatory neurons that populate the neocortex are born from apical progenitors located in the ventricular zone. During corticogenesis, apical progenitors sequentially generate deep-layer neurons followed by superficial-layer neurons directly or via the generation of intermediate progenitors. Whether neurogenic fate progression necessarily implies fate restriction in single progenitor types is unknown. Here we specifically isolated apical progenitors and intermediate progenitors, and fate-mapped their respective neuronal progeny following heterochronic transplantation into younger embryos. We find that apical progenitors are temporally plastic and can re-enter past molecular, electrophysiological and neurogenic states when exposed to an earlier-stage environment by sensing dynamic changes in extracellular Wnt. By contrast, intermediate progenitors are committed progenitors that lack such retrograde fate plasticity. These findings identify a diversity in the temporal plasticity of neocortical progenitors, revealing that some subtypes ofcells can be untethered from their normal temporal progression to re-enter past developmental states.

The Bos taurus maternal microbiome: Role in determining the progeny early-life upper respiratory tract microbiome and health.

Natural transference of maternal microbes to the neonate, especially at birth via the vaginal canal, has recently been recognized in humans and cows; however, its microbial influence on calf health has not yet been documented. We compared the bacterial communities in vaginal and fecal samples from 81 pregnant dairy cows versus those in nasopharyngeal and fecal samples collected at 3, 14 and 35 days of life from their respective progeny. The microbiota of the calf upper respiratory tract (URT), regardless of calf age, was found to be highly similar to the maternal vaginal microbiota. Calf fecal microbiota clustered closely to the maternal fecal microbiota, progressing toward an adult-like state over the first 35 days when relative abundances of taxa were considered. Sixty-four, 65 and 87% of the detected OTUs were shared between cow and calf fecal microbiota at days 3, 14 and 35 respectively, whereas 73, 76 and 87% were shared between maternal vaginal microbiome and calf URT microbiota at days 3, 14 and 35, respectively. Bacteroidetes, Ruminococcus, Clostridium, and Blautia were the top four genera identified in maternal and calf fecal samples. Mannheimia, Moraxella, Bacteroides, Streptococcus and Pseudomonas were the top five genera identified in maternal vaginal and calf URT samples. Mannheimia was relatively more abundant in the vaginal microbiota of cows whose progeny were diagnosed with respiratory and middle ear disease. Our results indicate that maternal vaginal microbiota potentially influences the initial bacterial colonization of the calf URT, and that might have an important impact on the health of the calf respiratory tract and middle ear.

BCN057 induces intestinal stem cell repair and mitigates radiation-induced intestinal injury

BackgroundRadiation-induced gastrointestinal syndrome (RIGS) results from the acute loss of intestinal stem cells (ISC), impaired epithelial regeneration, and subsequent loss of the mucosal barrier, resulting in electrolyte imbalance, diarrhea, weight loss, sepsis, and mortality. The high radiosensitivity of the intestinal epithelium limits effective radiotherapy against abdominal malignancies and limits the survival of victims of nuclear accidents or terrorism. Currently, there is no approved therapy to mitigate radiation toxicity in the intestine. Here we demonstrate that BCN057, an anti-neoplastic small molecular agent, induces ISC proliferation and promotes intestinal epithelial repair against radiation injury.MethodsBCN057 (90 mg/kg body weight, subcutaneously) was injected into C57Bl6 male mice (JAX) at 24 h following abdominal irradiation (AIR) and was continued for 8 days post-irradiation. BCN057-mediated rescue of Lgr5-positive ISC was validated in Lgr5-EGFP-Cre-ERT2 mice exposed to AIR. The regenerative response of Lgr5-positive ISC was examined by lineage tracing assay using Lgr5-EGFP-ires-CreERT2-TdT mice with tamoxifen administration to activate Cre recombinase and thereby marking the ISC and their respective progeny. Ex vivo three-dimensional organoid cultures were developed from surgical specimens of human colon or from mice jejunum and were used to examine the radio-mitigating role of BCN057 on ISC ex vivo. Organoid growth was determined by quantifying the budding crypt/total crypt ratio. Statistical analysis was performed using Log-rank (Mantel-Cox) test and paired two-tail t test.ResultsTreatment with BCN057 24 h after a lethal dose of AIR rescues ISC, promotes regeneration of the intestinal epithelium, and thereby mitigates RIGS. Irradiated mice without BCN057 treatment suffered from RIGS, resulting in 100% mortality within 15 days post-radiation. Intestinal organoids developed from mice jejunum or human colon demonstrated a regenerative response with BCN057 treatment and mitigated radiation toxicity. However, BCN057 did not deliver radio-protection to mouse or human colon tumor tissue.ConclusionBCN057 is a potential mitigator against RIGS and may be useful for improving the therapeutic ratio of abdominal radiotherapy. This is the first report demonstrating that a small molecular agent mitigates radiation-induced intestinal injury by inducing ISC self-renewal and proliferation.

The presence and dosimetry of radon and thoron in a historical, underground metalliferous mine

Abstract A combination of long term passive, and short term active radon-222, radon-220 and respective progeny measurements were conducted in both traverse and longitudinal axes of a historical metalliferous underground mine in North Queensland, Australia. While the passive monitor results provided average radon and thoron air concentrations over periods of 70–90 days, active measurements over a four day period provided significantly more detail into the dynamics of radon and progeny concentrations in the naturally ventilated mine environment. Passive monitor concentrations for radon and thoron ranged between 60 and 390 Bq m−3 (mean: 140 ±55 Bq m−3) and 140 and 2600 Bq m−3 (mean: 1070 ±510 Bq m−3) respectively, with passive thoron progeny monitors providing a mean concentration of 9 ±5 Bq m−3EEC. Active measurement mean concentrations for radon, thoron, radon progeny and thoron progeny in the centre of the mine drive were 130 ±90 Bq m−3, 300 ±100 Bq m−3, 20 ±20 Bq m−3EEC and 10 ±10 Bq m−3EEC respectively. It was identified that thoron passive detector placement is critical in establishing reliable monitoring data, and is the reason for the discrepancy between the active and passive thoron results in this study. Site specific progeny measurements are required for the accurate estimation of dose to persons entering the mine. Based on short term active measurements and passive thoron progeny monitor results, the dose contribution from thoron and progeny in the mine was observed to contribute up to 80% of the total radon/thoron inhalation dose, and therefore should not be underestimated in monitoring programs under similar conditions.

Inheritance of Fifteen Microsatellite Loci inCeratitis capitata(Diptera: Tephritidae)

Molecular methods that rely on microsatellite markers have been developed for population genetic studies and diagnostics of tephritid pest species such as the Mediterranean fruit fly, Ceratitis capitata (Wiedemann) (Diptera: Tephritidae). Whereas many of these markers are tested to see if they are within the Hardy—Weinberg equilibrium, very few markers developed for pest species are tested to ensure the selected alleles behave according to the laws of Mendelian inheritance. Fifteen previously developed microsatellite markers were examined for Mendelian inheritance. Nine parental groups consisting of a laboratory reared parent and a wild type parent and their respective progeny were examined. In total, 174 flies, consisting of 90 males and 84 females, were analyzed. Seventy-seven segregation ratio tests were performed to determine if any departures from expected Mendelian inheritance occurred. Representatives from each of the observed alleles were cloned and sequenced. Troubleshooting was performed on loci that did not conform to expected Mendelian inheritance ratios to confirm the cause and improve laboratory procedures. Issues observed included incomplete adenylation at the 5′ end in Ccmic3, the presence of artifactual bands leading to false calls in Ccmic25, and monomorphic alleles in Ccmic7. Only 1 locus, Ccmic25, deviated from Mendelian expectations after protocol optimization in the form of a detected transmission ratio distortion leading to excessive heterozygosity. Finally, 1 locus, Ccmic9, showed evidence of allelic homoplasy.

Live Monitoring of Blastemal Cell Contributions during Appendage Regeneration

The blastema is a mass of progenitor cells that enables regeneration of amputated salamander limbs or fish fins. Methodology to label and track blastemal cell progeny has been deficient, restricting our understanding of appendage regeneration. Here, we created a system for clonal analysis and quantitative imaging of hundreds of blastemal cells and their respective progeny in living adult zebrafish undergoing fin regeneration. Amputation stimulates resident cells within a limited recruitment zone to reset proximodistal (PD) positional information and assemble the blastema. Within the newly formed blastema, the spatial coordinates of connective tissue progenitors are predictive of their ultimate contributions to regenerated skeletal structures, indicating early development of an approximate PD pre-pattern. Calcineurin regulates size recovery by controlling the average number of progeny divisions without disrupting this pre-pattern. Our longitudinal clonal analyses of regenerating zebrafish fins provide evidence that connective tissue progenitors are rapidly organized into a scalable blueprint of lost structures.

DNA methylation changes and TE activity induced in tissue cultures of barley (Hordeum vulgare L.).

BackgroundIn vitro plant regeneration via androgenesis or somatic embryogenesis is capable of inducing (epi)mutations that may affect sexual progenies. While epimutations are associated with DNA methylation, mutations could be due to the movement of transposons. The common notion is that both processes are linked. It is being assumed that demethylation activates transposable elements (TEs). Analysis of methylation changes and their relation with TEs activation in tissue cultures requires uniquely derived donor plants (Ds), their regenerants (Rs) and respective progeny (Ps) that would allow discrimination of processes not related to changes introduced via in vitro cultures. Moreover, a set of methods (RP-HPLC, SSAP, and MSTD) is needed to study whether different TEs families are being activated during in vitro tissue culture plant regeneration and whether their activity could be linked to DNA methylation changes or alternative explanations should be considered.ResultsThe in vitro tissue culture plant regeneration in barley was responsible for the induction of DNA methylation in regenerants and conservation of the methylation level in the progeny as shown by the RP-HPLC approach. No difference between andro- and embryo-derived Rs and Ps was observed. The SSAP and MSTD approach revealed that Ds and Rs were more polymorphic than Ps. Moreover, Rs individuals exhibited more polymorphisms with the MSTD than SSAP approach. The differences between Ds, Rs and Ps were also evaluated via ANOVA and AMOVA.ConclusionsStressful conditions during plant regeneration via in vitro tissue cultures affect regenerants and their sexual progeny leading to an increase in global DNA methylation of Rs and Ps compared to Ds in barley. The increased methylation level noted among regenerants remains unchanged in the Ps as indicated via RP-HPLC data. Marker-based experiments suggest that TEs are activated via in vitro tissue cultures and that, independently of the increased methylation, their activity in Rs is greater than in Ps. Thus, the increased methylation level may not correspond to the stabilization of TEs movement at least at the level of regenerants. The presence of TEs variation among Ds that were genetically and epigenetically uniform may suggest that at least some mobile elements may be active, and they may mask variation related to tissue cultures. Thus, tissue cultures may activate some TEs whereas the others remain intact, or their level of movement is changed. Finally, we suggest that sexual reproduction may be responsible for the stabilization of TEs.

Epithelial–mesenchymal transition induces similar metabolic alterations in two independent breast cancer cell lines

Epithelial–mesenchymal transition (EMT) induces invasive properties in epithelial tumors and promotes metastasis. Although EMT-mediated cellular and molecular changes are well understood, very little is known about EMT-induced metabolic changes. HER2-positive BT-474 breast cancer cells were induced to undergo a stable EMT using mammosphere culture, as previously described by us for the ERα-positive MCF-7 breast cancer cells. Two epithelial breast cancer cell lines (BT-474 and MCF-7) were compared to their respective EMT-derived mesenchymal progeny (BT-474EMT and MCF-7EMT) for changes in metabolic pathways including glycolysis, glycogen metabolism, anabolic pathways and gluconeogenesis. Both EMT-derived cells displayed enhanced aerobic glycolysis along with the overexpression of specific glucose transporters, lactate dehydrogenase isoforms, monocarboxylate transporters and glycogen phosphorylase isoform. In contrast, both EMT-derived cells suppressed the expression of crucial enzymes in anabolic pathways and gluconeogenesis. STAT3, a transcription factor involved in tumor initiation and progression, plays a role in the EMT-related changes in the expression of specific enzymes and transporters. This study provides a broad overview of similar metabolic changes induced by EMT in two independent breast cancer cell lines. These metabolic changes may provide novel therapeutic targets for metastatic breast cancer.

Genetic variability of Scots pine (Pinus sylvestris L.) and Norway spruce (Picea abies L. Karst.) natural regeneration compared with their maternal stands

Abstract The genetic variability and biodiversity of tree populations ensure the stability and sustainability of forest ecosystems. New research tools based on molecular DNA markers enable precise characterisation of forest genetic resources, i.e. detection of different allele frequencies in mature trees and progeny populations. The paper describes the genetic structure of mature stands of Scots pine (Pinus sylvestris L.) in Oława Forest District and Norway spruce (Picea abies L. Karst.) in Jawor Forest District and stands of their respective progeny. In the Scots pine stand, there was a slight increase (0.6%) in heterozygosity level and a larger increase (4.9%) in the inbreeding coefficient of progeny trees. In the Norway spruce stand, despite a small reduction (0.9%) in heterozygosity, a similar increase (4.6%) in the inbreeding coefficient of progeny was revealed. In both stands, allele richness and the partition probability of basic clustering were high. Both pine and spruce adults and progeny trees were characterised by high levels of genetic similarity (96% and 79%, respectively). Gene flow between the mature and progeny populations was high (Nm > 1) for both Scots pine and Norway spruce. Conservation of the gene pool within forest tree stands requires an increase in the proportion of natural regeneration. To estimate the extent to which genes are transmitted between adult trees and their progeny, more studies are needed, especially taking into account the influence of silviculture measures, like selective tree cutting, on the genetic variability of the younger generation. These results confirm that the gene pool was conserved when transmitted between the stands studied, as well as highlight the usefulness of such a study for silvicultural purposes

Distribution of naturally occurring radionuclides activity concentration in East Malaysian marine sediment

Studies of naturally occurring radioactive materials (NORM) distribution of 226Ra, 228Ra and 40K in East Malaysia were carried out as part of a marine coastal environment project. The results of measurements will serve as baseline data and background reference level for Malaysia coastlines. Sediments from 21 coastal locations and 10 near shore locations were collected for analyses. The samples were dried, finely ground, sealed in a container and stored for a minimum of 30 days to establish secular equilibrium between 226Ra and 228Ra and their respective radioactive progenies. They were counted using a high-purity germanium (HPGe) spectrometer covering the respective progeny energy peak. For 40K, the presence of this was measured directly via its 1460 keV energy peak. The concentration of 226Ra, 228Ra and 40K in samples obtained from coastal Sarawak ranged between 23 and 41 (mean 30±2) Bq/kg, 27 and 45 (mean 39±4) Bq/kg and 142 and 680 (mean 462±59) Bq/kg, respectively. Meanwhile, the concentration of 226Ra, 228Ra and 40K for samples obtained from coastal Sabah ranged between 16 and 30 (mean 23±2) Bq/kg, 23 and 45 (mean 35±4) Bq/kg and 402 and 842 (mean 577±75) Bq/kg, respectively. For the Sarawak near shore stations, the concentration of 226Ra, 228Ra and 40K ranged between 11 and 36 (mean 22±2) Bq/kg, 21 and 65 (mean 39±5) Bq/kg and 149 and 517 (mean 309±41) Bq/kg, respectively. Meanwhile, the concentration of 226Ra, 228Ra and 40K for samples obtained from Sabah ranged between 9 and 31 (mean 14±2) Bq/kg, 10 and 48 (mean 21±3) Bq/kg and 140 and 580 (mean 269±36) Bq/kg, respectively. The calculated external hazard values of between 0.17 and 0.33 (less than unity) showed that there is little risk of external hazard to the workers handling the sediments.

Replication-Coupled Packaging Mechanism in Positive-Strand RNA Viruses: Synchronized Coexpression of Functional Multigenome RNA Components of an Animal and a Plant Virus in Nicotiana benthamiana Cells by Agroinfiltration

Flock house virus (FHV), a bipartite RNA virus of insects and a member of the Nodaviridae family, shares viral replication features with the tripartite brome mosaic virus (BMV), an RNA virus that infects plants and is a member of the Bromoviridae family. In BMV and FHV, genome packaging is coupled to replication, a widely conserved mechanism among positive-strand RNA viruses of diverse origin. To unravel the events that modulate the mechanism of replication-coupled packaging, in this study, we have extended the transfer DNA (T-DNA)-based agroinfiltration system to express functional genome components of FHV in plant cells (Nicotiana benthamiana). Replication, intracellular membrane localization, and packaging characteristics in agroinfiltrated plant cells revealed that T-DNA plasmids of FHV were biologically active and faithfully mimicked complete replication and packaging behavior similar to that observed for insect cells. Synchronized coexpression of wild-type BMV and FHV genome components in plant cells resulted in the assembly of virions packaging the respective viral progeny RNA. To further elucidate the link between replication and packaging, coat protein (CP) open reading frames were precisely exchanged between BMV RNA 3 (B3) and FHV RNA 2 (F2), creating chimeric RNAs expressing heterologous CP genes (B3/FCP and F2/BCP). Coinfiltration of each chimera with its corresponding genome counterpart to provide viral replicase (B1+B2+B3/FCP and F1+F2/BCP) resulted in the expected progeny profiles, but virions exhibited a nonspecific packaging phenotype. Complementation with homologous replicase (with respect to CP) failed to enhance packaging specificity. Taken together, we propose that the transcription of CP mRNA from homologous replication and its translation must be synchronized to confer packaging specificity.

Genetic variability of Scots pine maternal populations and their progenies

The genetic variability of Scots pine was investigated in six populations from Poland representing two maternal populations and their natural and artificial progenies. Thirteen enzyme systems controlled by 25 allozyme loci were analyzed using starch gel electrophoresis. Progeny populations maintained a high and similar level of genetic variation to that observed in the maternal populations. As expected, much closer genetic relationships were observed between maternal populations and their respective progeny than between the two maternal populations themselves. Progeny populations had the same major alleles, but differed in the number of rare alleles. Therefore, probably not all rare alleles were transferred from the maternal stands to the progenies. In addition, new rare alleles appeared in the progeny populations, possibly as a result of external pollen flow into the maternal populations.

Insecticidal effect of Beauveria bassiana (Balsamo) Vuillemin (Ascomycota: Hypocreaes) in combination with three diatomaceous earth formulations against Sitophilus granarius (L.) (Coleoptera: Curculionidae)

Abstract The insecticidal effect of the entomopathogenic fungus Beauveria bassiana (Balsamo) Vuillemin (Ascomycota: Hypocreaes) in combination with three diatomaceous earth (DE) formulations against adults of the granary weevil, Sitophilus granarius (L.) (Coleoptera: Curculionidae) was tested in the laboratory. The three DEs were Insecto™, SilicoSec ® and PyriSec ® . The fungus was applied at 400 ppm alone, or in combination with 200 ppm of each of the three DEs. Mortality was measured after 7 d of exposure. Bioassays were conducted at three temperatures 20, 25 and 30 °C and two relative humidities (rh) 55% and 75%. On wheat treated with B. bassiana alone, mortality was higher at 55% than at 75% rh. Also, the fungus alone was less effective at 20 °C than at the other two temperatures tested, but mortality did not exceed 52% for any of the conditions tested. Similar mortality levels were also noted on wheat treated with each of the three DEs alone. The simultaneous presence of B. bassiana and DE increased weevil mortality. In this combination, mortality was higher at high temperatures and low rh, and this effect was similar for all DEs tested. Progeny production on wheat treated with B. bassiana was higher that the respective progeny counts in the DE-treated wheat. The results indicate that a combination of B. bassiana and DEs is effective against S. granarius , under a broad range of temperature and rh levels in stored wheat.