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.

Effects of microgravity or simulated launch on testicular function in rats.

Testes from flight rats on COSMOS 2044 and simulated-launch, vivarium, or caudal-elevation control rats (5/group) were analyzed by subjective and quantitative methods. On the basis of observations of fixed tissue, it was evident that some rats had testicular abnormalities unassociated with treatment and probably existing when they were assigned randomly to the four treatment groups. Considering rats without preexisting abnormalities, diameter of seminiferous tubules and numbers of germ cells per tubule cross section were lower (P less than 0.05) in flight than in simulated-launch or vivarium rats. However, ratios of germ cells to each other or to Sertoli cells and number of homogenization-resistant spermatids did not differ from values for simulated-launch or vivarium controls. Expression of testis-specific gene products was not greatly altered by flight. Furthermore, there was no evidence for production of stress-inducible transcripts of the hsp70 or hsp90 genes. Concentration of receptors for rat luteinizing hormone in testicular tissue and surface density of smooth endoplasmic reticulum in Leydig cells were similar in flight and simulated-launch rats. However, concentrations of testosterone in testicular tissue or peripheral blood plasma were reduced (P less than 0.05) in flight rats to less than 20% of values for simulated-launch or vivarium controls. Thus spermatogenesis was essentially normal in flight rats, but production of testosterone was severely depressed. Exposure to microgravity for greater than 2 wk might result in additional changes. Sequelae of reduced androgen production associated with microgravity on turnover of muscle and bone should be considered.

Effects of clinostat rotation on mouse meiotic maturation in vitro.

The effects of microgravity on meiosis, fertilization, and early embryonic development in mammals are being examined by using a clinostat to reorient the cells with respect to the gravity vector. A clinostat capable of supporting mammalian cells in tissue culture has been developed. Initial studies have focused on examining the effects of clinostat rotation on meiotic maturation in mouse oocytes. Oocytes recovered from ovarian follicles were subjected to clinostat rotation on a horizontal or vertical axis or to static conditions for a 16 hr period. No gross morphological changes and no effects on germinal vesicle breakdown were observed under any rotation conditions (1/4, 1, 10, 30, 100 RPM). Success of meiotic progression to Metaphase II was comparable among experimental and control groups except at 100 RPM, where a slight inhibition was observed.

Evidence for the involvement of the proto-oncogene c-mos in mammalian meiotic maturation and possibly very early embryogenesis.

The c-mos proto-oncogene exists as a maternal mRNA in mammalian oocytes, in that it has been shown to accumulate in mouse oocytes during the growth phase and to be present at high levels in fully grown oocytes. The function of c-mos during the subsequent development of the oocytes and embryos was examined by determining the fate of the oocyte c-mos mRNAs by in situ hybridization and Northern blot hybridization analysis. A substantial decrease in the levels of c-mos transcripts was observed in oocytes undergoing meiotic maturation. By the two-cell stage, levels of c-mos transcripts dropped to below the limits of detection using in situ hybridization. c-mos transcripts remained undectable through the blastocyst stage of embryogenesis. Analysis of meiotic maturation in vitro permitted finer temporal resolution of the initial drop in c-mos levels. Between approximately 7 and 17 h of culture, the amount of c-mos mRNA fell to 18-43% of the levels found in the fully grown oocyte. This interval corresponds to the progression of meiotic maturation from metaphase I to metaphase II. Our in vivo studies showed that ovulation per se is not the stimulus for the drop in c-mos transcript levels, since preovulatory metaphase II oocytes exhibited this decline to a degree comparable to that of ovulated metaphase II oocytes. The development specificity of c-mos transcript levels suggests a role of this putative serine kinase in the meiotic maturation of mammalian germ cells.