Evaluation of a field deployable, high-throughput RT-LAMP device as an early warning system for COVID-19 through SARS-CoV-2 measurements in wastewater.

Quantification of SARS-CoV-2 RNA copies in wastewater can be used to estimate COVID-19 prevalence in communities. While such results are important for mitigating disease spread, SARS-CoV-2 measurements require sophisticated equipment and trained personnel, for which a centralized laboratory is necessary. This significantly impacts the time to result, defeating its purpose as an early warning detection tool. The objective of this study was to evaluate a field portable device (called MINI) for detecting SARS-CoV-2 viral loads in wastewater using real-time reverse transcriptase loop-mediated isothermal amplification (real-time RT-LAMP). The device was tested using wastewater samples collected from buildings (with 430 to 1430 inhabitants) that had known COVID-19-positive cases. Results show comparable performance of RT-LAMP against reverse transcriptase polymerase chain reaction (RT-qPCR) when detecting SARS-CoV-2 copies in wastewater. Both RT-LAMP and RT-qPCR detected SARS-CoV-2 in wastewater from buildings with at least three positive individuals within a 6-day time frame prior to diagnosis. The large 96-well throughput provided by MINI provided scalability to multi-building detection. The portability of the MINI device enabled decentralized on-site detection, significantly reducing the time to result. The overall findings support the use of RT-LAMP within the MINI configuration as an early detection system for COVID-19 infection using wastewater collected at the building scale.

Characterization of the public transit air microbiome and resistome reveals geographical specificity.

BACKGROUND: The public transit is a built environment with high occupant density across the globe, and identifying factors shaping public transit air microbiomes will help design strategies to minimize the transmission of pathogens. However, the majority of microbiome works dedicated to the public transit air are limited to amplicon sequencing, and our knowledge regarding the functional potentials and the repertoire of resistance genes (i.e. resistome) is limited. Furthermore, current air microbiome investigations on public transit systems are focused on single cities, and a multi-city assessment of the public transit air microbiome will allow a greater understanding of whether and how broad environmental, building, and anthropogenic factors shape the public transit air microbiome in an international scale. Therefore, in this study, the public transit air microbiomes and resistomes of six cities across three continents (Denver, Hong Kong, London, New York City, Oslo, Stockholm) were characterized. RESULTS: City was the sole factor associated with public transit air microbiome differences, with diverse taxa identified as drivers for geography-associated functional potentials, concomitant with geographical differences in species- and strain-level inferred growth profiles. Related bacterial strains differed among cities in genes encoding resistance, transposase, and other functions. Sourcetracking estimated that human skin, soil, and wastewater were major presumptive resistome sources of public transit air, and adjacent public transit surfaces may also be considered presumptive sources. Large proportions of detected resistance genes were co-located with mobile genetic elements including plasmids. Biosynthetic gene clusters and city-unique coding sequences were found in the metagenome-assembled genomes. CONCLUSIONS: Overall, geographical specificity transcends multiple aspects of the public transit air microbiome, and future efforts on a global scale are warranted to increase our understanding of factors shaping the microbiome of this unique built environment.

Dnmt3a regulates myeloproliferation and liver-specific expansion of hematopoietic stem and progenitor cells.

DNA methyltransferase 3A (DNMT3A) mutations are observed in myeloid malignancies, including myeloproliferative neoplasms (MPN), myelodysplastic syndromes (MDS) and acute myeloid leukemia (AML). Transplantation studies have elucidated an important role for Dnmt3a in stem cell self-renewal and in myeloid differentiation. Here, we investigated the impact of conditional hematopoietic Dnmt3a loss on disease phenotype in primary mice. Mx1-Cre-mediated Dnmt3a ablation led to the development of a lethal, fully penetrant MPN with myelodysplasia (MDS/MPN) characterized by peripheral cytopenias and by marked, progressive hepatomegaly. We detected expanded stem/progenitor populations in the liver of Dnmt3a-ablated mice. The MDS/MPN induced by Dnmt3a ablation was transplantable, including the marked hepatomegaly. Homing studies showed that Dnmt3a-deleted bone marrow cells preferentially migrated to the liver. Gene expression and DNA methylation analyses of progenitor cell populations identified differential regulation of hematopoietic regulatory pathways, including fetal liver hematopoiesis transcriptional programs. These data demonstrate that Dnmt3a ablation in the hematopoietic system leads to myeloid transformation in vivo, with cell-autonomous aberrant tissue tropism and marked extramedullary hematopoiesis (EMH) with liver involvement. Hence, in addition to the established role of Dnmt3a in regulating self-renewal, Dnmt3a regulates tissue tropism and limits myeloid progenitor expansion in vivo.

Dispersal behavior of neonate European corn borer (Lepidoptera: Crambidae) on Bt corn.

European corn borer, Ostrinia nubilalis (Hübner) (Lepidoptera: Crambidae), has historically been a significant economically important insect pest of corn (Zea mays L.) in the United States and Canada. The development in the 1990s of genetically modified corn expressing genes derived from Bacillus thuringiensis (Bt) that encodes insecticidal crystalline (Cry) proteins has proven to be effective in controlling this insect as well as other corn pests. The purpose of this study was to assess the movement and dispersal behavior of neonate European corn borer on Bt corn. We examined differences in neonate European corn borer dispersal behavior for the first 4 h after eclosion in the field among a stacked pyramid (Cry1F X Cry1Ab X Cry34/35Ab1) Bt corn, a Cry1F Bt corn, and a non-Bt sweet corn; and in the laboratory among a Bt corn hybrid containing Cry1F, a hybrid containing Cry1Ab, a pyramid combining these two hybrids (Cry1F X Cry1Ab), and a non-Bt near isoline corn. In field experiments, we found that dispersal was significantly higher on Bt corn compared with sweet corn. In laboratory experiments, dispersal was significantly higher on Cry1Ab Bt corn and Cry1F X Cry1Ab Bt corn than on non-Bt near isoline corn. Results indicated that neonate dispersal may be significantly greater in Bt cornfields compared with non-Bt cornfields. The findings on dispersal behavior in this study will be useful in evaluating the efficacy of a blended seed refuge system for managing European corn borer resistance in Bt corn.

Feeding behavior of neonate Ostrinia nubilalis (Lepidoptera: Crambidae) on Cry1Ab Bt corn: implications for resistance management.

The European corn borer, Ostrinia nubilalis (Hübner) (Lepidoptera: Crambidae), is an economically important insect pest of corn, Zea mays L., in the United States and Canada. The development of genetically modified corn expressing genes derived from Bacillus thuringiensis (Bt) that encodes insecticidal crystalline (Cry) proteins has proven to be effective in controlling this insect. To assess the feeding behavior of neonate O. nubilalis on Bt corn, we examined differences in feeding behavior, based on presence of plant material in the gut, between Cry1Ab Bt corn and non-Bt near isoline corn for four intervals over a 48-h period. Feeding experiments revealed that there was significantly less feeding on Bt corn compared with non-Bt near isoline corn. The behavior of neonates on the plant corresponded with the differences in feeding on the two corn lines. The findings also showed that > 50% of the larvae initially left the plant before there was evidence in the gut of feeding regardless of whether the source was Bt or non-Bt corn. A higher quantity of plant material was found in the gut of larvae recovered from leaves of non-Bt compared with Bt corn. At the end of 48 h among the larvae that had left the plant, a greater proportion from Bt corn had plant material in the gut than did those from non-Bt corn.

Genetic similarity among pheromone and voltinism races of Ostrinia nubilalis (Hübner) (Lepidoptera: Crambidae).

The genetic variability of seven European corn borer populations, Ostrinia nubilalis, from North America and Europe was assessed by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis and DNA sequencing. The nuclear ribosomal internal transcribed spacer 1 (ITS-1) region (approximately 500 base pair [bp]) and four mitochondrial (mtDNA) regions (1550 bp total) were examined. The smartweed borer, Ostrinia obumbratalis, and south-Western corn borer, Diatraea grandiosella, were used for comparisons. Of 106 restriction sites identified (80 in mtDNA and 26 in ITS-1), none differentiated geographical populations, pheromone races, or voltine ecotypes of the European corn borer. The lack of variation in the ITS-1 of European corn borer was confirmed by DNA sequence analysis. The genetic similarity of European corn borer populations, despite their wide geographical range and physiological differences, may be explained by a relatively recent origin for the voltinism and pheromone races, gene flow among races, and/or expansion from genetic bottlenecks.

Histochemical demonstration of skeletal muscle fibre types and capillaries on the same transverse section.

Serial transverse sections of m. vastus lateralis biopsies from six healthy men were reacted: (1) for myofibrillar adenosine triphosphatase (mATPase) to identify fibre types; or, (2) with alpha-amylase, periodic acid-Schiff (alpha PAS) to visualize capillaries. Sections were also processed with a new histochemical method for identification of fibre types and capillaries on the same skeletal muscle slice (mATPase/alpha PAS). Fibre type composition using either method was 41% type I, 37% type IIA and 22% type IIB. Types I, IIA and IIB least diameter areas (mean +/- SE, micron2) were similar in sections processed for mATPase/alpha PAS or mATPase (3976 +/- 338, 5187 +/- 373 and 4389 +/- 514 vs. 4092 +/- 345, 5100 +/- 360 and 4289 +/- 474, respectively). The number of capillaries per mm2 and per fibre did not differ in sections processed using the alpha PAS (315 +/- 29 and 1.48 +/- 0.12) or mATPase/alpha PAS (317 +/- 25 and 1.43 +/- 0.10) method. The number of capillaries was greater (P less than 0.05) around types I or IIA than type IIB fibres whether a section was processed for mATPase/alpha PAS (4.5 +/- 0.2 or 4.6 +/- 0.2 vs. 3.4 +/- 0.3) or when fibre profiles of serial sections reacted for mATPase or alpha PAS were 'matched' (4.5 +/- 0.2 or 4.4 +/- 0.2 vs. 3.4 +/- 0.3). The results indicate that histochemical and morphometric measures can be made on the same transverse section using the new method with substantial savings of time, cost and energy.