Second round of an interlaboratory comparison of SARS-CoV2 molecular detection assays used by 45 veterinary diagnostic laboratories in the United States

The COVID-19 pandemic presents a continued public health challenge. Veterinary diagnostic laboratories in the United States use RT-rtPCR for animal testing, and many laboratories are certified for testing human samples; hence, ensuring that laboratories have sensitive and specific SARS-CoV2 testing methods is a critical component of the pandemic response. In 2020, the FDA Veterinary Laboratory Investigation and Response Network (Vet-LIRN) led an interlaboratory comparison (ILC1) to help laboratories evaluate their existing RT-rtPCR methods for detecting SARS-CoV2. All participating laboratories were able to detect the viral RNA spiked in buffer and PrimeStore molecular transport medium (MTM). With ILC2, Vet-LIRN extended ILC1 by evaluating analytical sensitivity and specificity of the methods used by participating laboratories to detect 3 SARS-CoV2 variants (B.1; B.1.1.7 [Alpha]; B.1.351 [Beta]) at various copy levels. We analyzed 57 sets of results from 45 laboratories qualitatively and quantitatively according to the principles of ISO 16140-2:2016. More than 95% of analysts detected the SARS-CoV2 RNA in MTM at ≥500 copies for all 3 variants. In addition, for nucleocapsid markers N1 and N2, 81% and 92% of the analysts detected ≤20 copies in the assays, respectively. The analytical specificity of the evaluated methods was >99%. Participating laboratories were able to assess their current method performance, identify possible limitations, and recognize method strengths as part of a continuous learning environment to support the critical need for the reliable diagnosis of COVID-19 in potentially infected animals and humans.     

Use of Spinach, Radish, and Perennial Ryegrass to Assess the Availability of Metals in Waste Foundry Sands

Opencast coalmine effluent contains higher concentrations of Cd, Cr, Co, Ni and Pb. Biofiltration of these metals has been demonstrated successfully with the help of aquatic macrophytes i.e., E. crassipes, L. minor and A. pinnata. Experiments revealed E. crassipes reduced highest concentration of heavy metals followed by L. minor and A. pinnata on 20th days retention period. Plant tissue analysis revealed higher accumulation of metals in roots than leaves. Highly significant correlations have been noted between removal of heavy metals in effluent and their accumulation in roots and leaves of the experimental sets. Translocation factor also revealed lower transportation of metals from root to leaves. Reduction in chlorophyll and protein content was noted with the accumulation of heavy metals. N, P and K analysis in plant tissues indicated continuous decrease in their concentration with increasing metal concentration. Negative and significant correlations between metal accumulation and N, P and K concentrations in plant tissues showed adverse effects of heavy metals. Analysis of variance (Dunnett t-test) showed significant results (p?<?0.001) for all the metals in different durations.     

FUM13 encodes a short chain dehydrogenase/reductase required for C-3 carbonyl reduction during fumonisin biosynthesis in Gibberella moniliformis

Fumonisins are polyketide-derived mycotoxins produced by the filamentous fungus Gibberella moniliformis (anamorph Fusarium verticillioides). Wild-type strains of the fungus produce predominantly four B-series fumonisins, designated FB(1), FB(2), FB(3), and FB(4). Recently, a cluster of 15 putative fumonisin biosynthetic genes (FUM) was described in G. moniliformis. We have now conducted a functional analysis of FUM13, a gene in the cluster that is predicted by amino acid sequence similarity to encode a short chain dehydrogenase/reductase (SDR). Mass spectrometric analysis of metabolites from FUM13 deletion mutants revealed that they produce approximately 10% of wild-type levels of B-series fumonisins as well as two previously uncharacterized compounds. NMR analysis revealed that the new compounds are similar in structure to FB(3) and FB(4) but that they have a carbonyl function rather than a hydroxyl function at carbon atom 3 (C-3). These results indicate that the FUM13 protein catalyzes the reduction of the C-3 carbonyl to a hydroxyl group and are the first biochemical evidence directly linking a FUM gene to a specific reaction during fumonisin biosynthesis. The production of low levels of FB(1), FB(2), FB(3), and FB(4), which have a C-3 hydroxyl, by the FUM13 mutants suggests that G. moniliformis has an additional C-3 carbonyl reductase activity but that this enzyme functions less efficiently than the FUM13 protein.     

Modified Pollard transects for assessing tropical butterfly abundance and diversity

This paper introduces and discusses the consistency and effectiveness of an inexpensive modification of Pollard transects for assessing the diversity and abundance of tropical butterfly communities in two national parks in Rio de Janeiro, Brazil. To measure butterfly abundance, students walked simultaneous timed transects at the interface of forest and cleared areas. They either counted the number of individuals observed or tabulated the number of specimens collected with nets. After a short training period, the number of butterflies observed or collected on simultaneous transects was statistically indistinguishable among student groups, and there was a significant positive correlation between observation counts and collected number of individuals. As a measure of species richness, the number of butterfly species sampled on each simultaneous transect was tabulated and did not differ statistically. To measure diversity, alpha of the logseries model was calculated for each collected sample, and statistical fit to a logseries model was determined. Although virtually all daily samples and the year's accumulated sample at one park fit the logseries model, about 35% of the daily samples and the entire year's sample did not fit at the other park. Despite these differences between the two parks, values for alpha from daily samples at both parks varied similarly (from 15 to 50 in almost all cases), and values from the entire year's samples were statistically indistinguishable. The repeatability of results among novices, such as students, suggests that timed transects have great promise for furthering our understanding of butterfly community demographics.     

Lepidium latifolium: plant nutrient competition-soil interactions

Exotic weeds are invading rangelands of the western United States at unprecedented rates. Understanding plant-soil relationships and competitive interactions of invasive weeds is crucial in long-term control strategies. In a greenhouse experiment, we investigated the influence of soil nutrient depletion on plant growth and plant competition between the exotic invasive weeds, Lepidium latifolium (invading wetlands) and Bromus tectorum (invading a multitude of habitats). Plants were grown individually and in combination until L. latifolium flowered, then roots and aboveground mass were harvested. Soil in individual pots was homogenized, subsamples collected for nutrient analyses, and the soil was re-planted to the same species, grown, and harvested twice more for a total of three growth cycles. As nutrient supplying capacity of the soil declined through growth cycles, aboveground mass of L. latifolium decreased significantly (PА.05) and growth potential of B. tectorum surpassed that of L. latifolium. Only bicarbonate-extractable soil ortho-P positively correlated with plant mass of L. latifolium. A separate experiment demonstrated that L. latifolium has a narrow window of soil water potentials for optimal growth; greatest growth at -20 kPa with significantly declining growth at saturation and -400 kPa. Our data suggest that L. latifolium primarily invades wetlands because the high soil water content reduces tortuosity and allows efficient transport of nutrients to this sparsely rooted species. When soil moisture and/or the nutrient supplying capacity of the soil declines, plants with greater root density can out-compete L. latifolium. Monocultural stands of L. latifolium may be self-limiting in time as available nutrients, particularly P, are biocycled to drier upper soil layers.     

Use of adsorbent and supercritical carbon dioxide to concentrate flavor compounds from orange oil.

Orange oil is composed largely of terpene hydrocarbons but is a source of flavor and fragrance compounds (oxygenated) that are present in low concentrations. To increase the ratio of oxygenated compounds to terpene hydrocarbons, orange oil was partially fractionated by adsorption of the oxygenated compounds onto porous silica gel, with full utilization of its adsorbent capacity, and then further purified by desorption into supercritical carbon dioxide. The desorption of 24 compounds was monitored by GC and GC-MS. Adsorption alone removed three-fourths of the terpene hydrocarbons, and fractional extraction by supercritical carbon dioxide (SC-CO(2)) improved the separation further. Response surface methodology was used in the experimental design, and regression analysis was used to determine the effects of process variables. Extraction at low temperatures and flow rates improved separation by SC-CO(2). Decanal was concentrated to 20 times that of the feed oil by using SC-CO(2) at 13.1 MPa, 35 degrees C, and 2 kg/h. The systems were operating at close to equilibrium conditions because of the fine dispersal of the oils and the excellent mass transfer properties of supercritical carbon dioxide.     

Effects of soil pH and soil water content on prosulfuron dissipation

The sulfonylurea herbicide prosulfuron, 1-(4-methoxy-6-methyltriazin-2-yl)-3-[2-(3,3,3-trifluoropropyl)phenylsulfonyl]urea, is used for the selective control of broadleaf weeds in corn, sorghum, and cereal grains. To investigate its fate in soils, this study examined the effects of soil pH and water content on the rates of dissipation processes and the products formed under aerobic conditions. Radiometry and chromatography analyses were used to quantify the degradation products and bound residues formed in incubations of 10 different soils. The pH-dependent hydrolysis of the sulfonylurea bridge to form phenyl sulfonamide was the primary transformation process. Significant microbial degradation of prosulfuron occurred in 2 of the 10 soils, yielding (14)CO(2) and desmethyl prosulfuron among the major products. The time required for 50% dissipation of the herbicide (DT(50)) was determined for each soil and water content treatment. At equivalent water contents, prosulfuron DT(50) values were positively correlated with soil pH (P < 0.0001), varying from 6.5 days at pH 5.4 to 122.9 days at pH 7.9. Soil pH and water content strongly influence the fate of sulfonylurea herbicides in agricultural fields. Differences in the effect of soil water content on dissipation kinetics in a comparison of two soils were attributed to differences in soil pH, texture, and the ability of indigenous microorganisms to transform the herbicide.     

The empirical bias of estimates by restricted maximum likelihood, Bayesian method, and method R under selection for additive, maternal, and dominance models.

Bayesian analysis via Gibbs sampling, restricted maximum likelihood (REML), and Method R were used to estimate variance components for several models of simulated data. Four simulated data sets that included direct genetic effects and different combinations of maternal, permanent environmental, and dominance effects were used. Parents were selected randomly, on phenotype across or within contemporary groups, or on BLUP of genetic value. Estimates by Bayesian analysis and REML were always empirically unbiased in large data sets. Estimates by Method R were biased only with phenotypic selection across contemporary groups; estimates of the additive variance were biased upward, and all the other estimates were biased downward. No empirical bias was observed for Method R under selection within contemporary groups or in data without contemporary group effects. The bias of Method R estimates in small data sets was evaluated using a simple direct additive model. Method R gave biased estimates in small data sets in all types of selection except BLUP. In populations where the selection is based on BLUP of genetic value or where phenotypic selection is practiced mostly within contemporary groups, estimates by Method R are likely to be unbiased. In this case, Method R is an alternative to single-trait REML and Bayesian analysis for analyses of large data sets when the other methods are too expensive to apply.