Investigations on Biosuppression of Wilt Disease in Tomato Using Cell-Free Culture Filtrate of Phytopathogenic Fusarium oxysporum f.sp. lycopersici

Potency of cell-free culture filtrate of pathogenic Fusarium oxysporum f.sp. lycopersici as systemic defense inducer against wilt disease in tomato was tested by treating seedlings with the filtrate before pathogen challenge. Infective conidia and the cell-free culture filtrate were prepared from F. oxysporum (Sensu lato) previously isolated from wilt diseased affected plants. Growth relations of the isolates were characterized under ambient temperature (25 ± 2°C) and viability of the infective conidia of the two isolates were evaluated. Treatments were done as follows: (a) treatment of seedlings with infective conidia only (Treatment-A) (b) culture filtrate treatment only (Treatment-B) (c) treatment with culture filtrate followed by inoculation with infective conidia (Treatment-C) and (d) the control which consisted of plants sprayed with sterile distilled water only (Treatment-D). Effects of fungal conidia on tomato seed germination as well as some agronomic characters of the tomato plants and disease incidence under treatments A, B, C and D were evaluated. The growth and sporulation rates of the F. oxysporum F-isolate-1 were 12.1 mm day-1 and 6.5 x 104 conidia cm-2 colony area respectively and the values for the second isolate, F-Isolate 2 were 6.1 mm day-1 and 5.4 x 104 conidia cm-2 colony area. The germination rates of the infective conidia of F-isolate 1 and F-Isolate 2 after 24 hours incubation period at ambient temperature were 70% and 85% respectively. Treatment of tomato seeds with conidia suspension containing 1.0 x 106 had no statistically significant effects on seed germination [F(2,12)=0.148, P=0.64]; the mean percentage germination of the seeds treated with F-Isolate 1, F-Isolate 2 conidia and the control were 52%,46% and 52% respectively after five days. In the Treatment-A (Infective conidia only), F-Isolate 1 and F-isolate 2 caused 100% and 80% wilt of the plant populations at 7 weeks post-inoculation, when the tomato plants were 9 weeks old and no wilt was recorded in the control. There was no wilt recorded under Treatment-B and Treatment-C also, but there was reduced flowering and the mean percentage number of leaves showing chlorosis were significantly higher than the control [F (4, 192) =6.641, P=0.001]. When the plants were 10 weeks old, withered branches at the lower vegetal strata and the senescent leaves in the control were comparable with that recorded under Treatment-B and Treatment-C (F (4, 159) =3.563, P=0.08].

Retreating Shorelines as an Emerging Threat to Adélie Penguins on Inexpressible Island

Long-term observation of penguin abundance and distribution may warn of changes in the Antarctic marine ecosystem and provide support for penguin conservation. We conducted an unmanned aerial vehicle (UAV) survey of the Adélie penguin (Pygoscelis adeliae) colony on Inexpressible Island and obtained aerial images with a resolution of 0.07 m in 2018. We estimated penguin abundance and identified the spatial extent of the penguin colony. A total of 24,497 breeding pairs were found on Inexpressible Island within a colony area of 57,507 m2. Based on historical images, the colony area expanded by 30,613 m2 and abundance increased by 4063 pairs between 1983 and 2012. Between 2012 and 2018 penguin abundance further increased by 3314 pairs, although the colony area decreased by 1903 m2. In general, Adélie penguins bred on Inexpressible Island at an elevation <20 m, and >55% of penguins had territories within 150 m of the shoreline. This suggests that penguins prefer to breed in areas with a low elevation and close to the shoreline. We observed a retreat of the shoreline on Inexpressible Island between 1983 and 2018, especially along the northern coast, which may have played a key role in the expansion of the penguin colony on the northern coast. In sum, it appears that retreating shorelines reshaped penguin distribution on the island and may be an emerging risk factor for penguins. These results highlight the importance of remote sensing techniques for monitoring changes in the Antarctic marine ecosystem and providing reliable data for Antarctic penguin conservation.

Quantifying impacts of stony coral tissue loss disease on corals in Southeast Florida through surveys and 3D photogrammetry

Since 2014, stony coral tissue loss disease (SCTLD) has contributed to substantial declines of reef-building corals in Florida. The emergence of this disease, which impacts over 20 scleractinian coral species, has generated a need for widespread reef monitoring and the implementation of novel survey and disease mitigation strategies. This study paired SCTLD prevalence assessments with colony-level monitoring to help improve understanding of disease dynamics on both individual coral colonies and at reef-wide scales. Benthic surveys were conducted throughout the northern Florida Reef Tract to monitor the presence/absence of disease, disease prevalence, and coral species affected by SCTLD. Observed SCTLD prevalence was lower in Jupiter and Palm Beach than in Lauderdale-by-the-Sea or St. Lucie Reef, but there were no significant changes in prevalence over time. To assess colony-level impacts of the disease, we optimized a low-cost, rapid 3D photogrammetry technique to fate-track infected Montastraea cavernosa coral colonies over four time points spanning nearly four months. Total colony area and healthy tissue area on fate-tracked colonies decreased significantly over time. However disease lesion area did not decrease over time and was not correlated with total colony area. Taken together these results suggest that targeted intervention efforts on larger colonies may maximize preservation of coral cover. Traditional coral surveys combined with 3D photogrammetry can provide greater insights into the spatiotemporal dynamics and impacts of coral diseases on individual colonies and coral communities than surveys or visual estimates of disease progression alone.

Antagonism Test of Trichoderma atroviride and Gliocladium sp. Bali Local Isolates As a Disease Control of Blendok Disease (Botryodiplodia theobromae) in Grapefruit (Citrus grandis L. Osbeck)

This study aims to determine the ability of Trichoderma atroviride and Gliocladium sp. in inhibiting the fungus that causes Gumosis disease Botryodiplodia theobromae. This study used a completely randomized design with two single treatments and was repeated eight times. In the in-vitro antagonism test Trichoderma atroviride and Gliolcadium sp. can inhibit the fungus Botryodiplodia theobromae as well. The percentage of inhibition was 94.58% in the Trichoderma atroviride antagonism test and 81.67% in the Gliocladium sp. antagonism test. Whereas the Botryodiplodia theobromae Colony Area had no significant effect on both treatments. Each Botryodiplodia theobromae Colony area is 18.34 cm2 with T. atroviride treatments and 22.25 cm2 with Gliocladium sp. treatment. The growth rate of Botryodiplodia Theobroma fungi was superior to that of Trichoderma atroviride and Gliocladium sp.

How to characterize a strain? The neglected influence of clonal heterogeneity on the phenotypes of industrial Saccharomyces

Populations of microbes are constantly evolving heterogeneity that selection acts upon, yet heterogeneity is non-trivial to assess methodologically. The practice of isolating single cell colonies for establishing, transferring, and using a strain results in single-cell bottlenecks with a generally neglected effect on the characteristics of the strain. We used six industrial yeasts to assess the level of heterogeneity in clonal populations, especially in terms of stress tolerance. First, we uncovered the existence of genome structure variants in available sequenced genomes of clonal lineages of thes strains. Subsequent phenotyping of strains and their newly isolated subclones showed that single-cell bottlenecks during isolation can considerably influence the observable phenotype. Next, we decoupled fitness distributions on the level of individual cells from clonal interference by plating single cell colonies. We used the obtained data on colony area for statistical modeling of the heterogeneity in phenotypes. One strain was further used to show how individual subclonal lineages are remarkably different not just in phenotype, but also in the level of heterogeneity. Thereby we call attention to the fact that choosing an initial clonal lineage from an industrial yeast strain may vastly influence downstream performances and observations on geno- and phenotype, and also on heterogeneity.

Automatic detection and mapping of European ground squirrel burrows on UAV-based multi- and hyperspectral imagery with classification methods

&lt;p&gt;European ground squirrels (EGS) are members of the soil megafauna and part of the ecosystem engineers that shape physical, chemical, and biological characteristics of soil ecosystems in European grasslands. Thanks to their strict protection their abundance and distribution have been surveyed systematically and annually in Hungary. The results of their 20 year monitoring indicate that their population is declining, there are sudden extinctions of local populations, and a desynchronized variation of the abundance of local populations occur either spatially or temporally.&lt;/p&gt;&lt;p&gt;The monitoring protocol involves the estimation of their abundance in each colony by a strip-transect method and the habitat-colony area by visual observations or digital maps. Both approaches use animal burrows as proxies for either their presence (colony area) or density (colony size). These estimation methods, however, consist of systematical errors: first, they consider the animals&amp;#8217; density to be even over the entire habitat-area, second, they conjecture that animals occupy the habitable area completely, and third, evenly. If we were able to survey distribution and abundance of EGS more accurately, frequently, and efficiently, we could better intervene in time when local populations begin to decline or before they disappear. In addition, we could better estimate the effects (+ or -) of management strategies in real time.&lt;/p&gt;&lt;p&gt;The primary aims of our study were to develop a non-invasive, semi-automated method for (1) estimating abundance of EGS in the area of occupation of a colony, and (2) delineating their occupancy within the habitable area. We have defined burrow openings and mounds as quantitative proxies for the presence of animals. We have started to develop a monitoring technique to identify, locate, and count objects of interest in images automatically and to delineate the area of occupancy by identifying those objects of interest from the surroundings. To survey EGS colonies and habitats we have used a multirotor platform UAV equipped with either an RGB visible-range or a hyperspectral sensor.&lt;/p&gt;&lt;p&gt;To test our method several pilot areas with different vegetation and relief were surveyed. Acquired aerial images have been processed by photogrammetric software and resulting high spatial resolution orthomosaics are classified by machine-learning algorithms (randomforest, CART, C5.0) implemented in a custom R script. As detection of mounds and openings are visually restricted by vegetation height (e.g. grass, shrubs, weeds, herbs), we have studied the effect of grass height on detection success. Preliminary results suggest that successful classification can be performed either on RGB visible-range and hyperspectral images. However, the appropriate spatial resolution (below cm range) and the presence of high grass are more important key factors than number of spectral bands.&lt;/p&gt;&lt;p&gt;Detecting EGS burrow openings and mounds is based on surface characteristics of EGS burrow openings and mounds consequently the method is being developed for EGS specifically but can be modified to the characteristics of other burrowing mammals of this size (e.g. mole-rats, moles).&lt;/p&gt;

Quantification of the morphological characteristics of hESC colonies

The maintenance of the undifferentiated state in human embryonic stem cells (hESCs) is critical for further application in regenerative medicine, drug testing and studies of fundamental biology. Currently, the selection of the best quality cells and colonies for propagation is typically performed by eye, in terms of the displayed morphological features, such as prominent/abundant nucleoli and a colony with a tightly packed appearance and a well-defined edge. Using image analysis and computational tools, we precisely quantify these properties using phase-contrast images of hESC colonies of different sizes (0.1–1.1 $${{\bf{\text{mm}}}}^{{\bf{2}}}$$mm2) during days 2, 3 and 4 after plating. Our analyses reveal noticeable differences in their structure influenced directly by the colony area $${\boldsymbol{A}}$$A. Large colonies (A > 0.6 mm2) have cells with smaller nuclei and a short intercellular distance when compared with small colonies (A < 0.2 mm2). The gaps between the cells, which are present in small and medium sized colonies with A ≤ 0.6 mm2, disappear in large colonies (A > 0.6 mm2) due to the proliferation of the cells in the bulk. This increases the colony density and the number of nearest neighbours. We also detect the self-organisation of cells in the colonies where newly divided (smallest) cells cluster together in patches, separated from larger cells at the final stages of the cell cycle. This might influence directly cell-to-cell interactions and the community effects within the colonies since the segregation induced by size differences allows the interchange of neighbours as the cells proliferate and the colony grows. Our findings are relevant to efforts to determine the quality of hESC colonies and establish colony characteristics database.

Characterization of Growth and Virulence of Five Nigerian Isolates of Entomopathogenic Fungi Using Galleria mellonella Larvae for Pathogenicity Testing

Entomopathogenic fungi (EF) are naturally occurring insect population regulators, with several species that are exploited as biocontrol agents against insect pests. Five EF consisting of two strains of Isaria farinosa, (IF-I and IF-II) and one strain each of Metarhizium anisopliae, Beauveria bassiana and Entomophaga sp. (Sensu lato) were isolated from soil using Tribolium castaneum (Coleoptera: Tenebrionidae) (Herbst) larvae as bait. The isolates were cultured on standard Potato Dextrose Agar (PDA) (Sigma-Aldrich, UK) and identified based on phenotypic appearance and micro-morphology. Growth rates (mm day-1), number of conidia per cm2 colony area after incubation for 14 days at ambient temperature (25±2°C), viability of conidia (% germination), based on 24-hours incubation period and virulence of the infective conidia against Galleria mellonella were evaluated. The data on growth was subjected to analysis of Variance (ANOVA) procedure and means were separated using Tukeys Honestly Significant Difference (P=0.05). The number of conidia produced by Entomophaga sp was (7.0×105 conidia cm 2 per colony area), while the isolate of M. anisopliae produced (5.2×104 conidia cm2 per colony area). The number of conidia produced by the two isolates of I. farinosa, IF-I and IF-II and B. bassiana were 9.4×104, 7.2×104 and 2.1×105 conidia cm2 per colony area respectively. Eighty percent of Entomophaga sp conidia germinated after incubating for 24 hours at 25°C while 100% germination occurred in the other fungal isolates. There were statistically significant variabilities in the rates of growth of the EF isolates F(4,2.064) =12.97, P=0.001. The B. bassiana isolate had the fastest growth rate, with statistically significant value of 3.3 mm day-1. The rates of growth of the two I. farinosa isolates: IF-I, IF-II and M. anisopliae were comparable, being 1.53, 1.4 and 1.28 mm day-1 respectively, without statistically significant difference. The growth rate of Entomophaga sp was 2.0 mm day-1, which was significantly higher than the growth rates of I. farinosa and M. anisopliae. The mean percentage mortality values of, G. mellonella larvae treated with 1×108 conidia ml-1 of the infective conidia of I. farinosa, IF-I, IF-II, M. anisopliae, after five days were 70, 60, 60% respectively while Entomophaga sp and B. bassiana caused 50% mortality. The results suggest that the five isolates examined can potentially be developed into experimental formulations and tested against important horticultural pests in future studies.

Influence of mitochondrial DNA loss on the development of complex structured colonies of SK1 strains of yeast Saccharomyces cerevisiae

Aim. Determine the effect of mitochondrial DNA loss on the formation of complex yeast colonies of Saccharomyces cerevisiae. Methods. Development of giant colonies of the parent strain SK1 (rho+) and the "petit" strain SK1p, which lost mtDNA (rho0 mutation), was observed for 40 days,. To find out zones of cell death in colonies, both strains were cultured on solid YPD media containing dyes that accumulate in dead cells. The survival of the cells within colony was estimated by the ability to create microcolonies. Results. The loss of mitochondrial DNA in SK1p cells led to a decrease in colony area and to simplification of colony morphology on the YPD medium. When SK1p colonies were cultivated on media with addition of dyes, bright spot was formed in the center due to the dyes accumulation in dead cells. At the same time, parent strain developed a uniform insignificant coloration. Conclusions. rho0 mutation in SK1p yeast strain Saccharomyces cerevisiae led to a significant simplification of complex colony structure that formed on the nutrient medium YPD. The mitochondrial DNA loss in strain SK1p results in an accelerated cell death in the center of colony on YPD.Keywords: Saccharomyces cerevisiae, rho0, colonies, cell survival.