The lack of a permissive system for the propagation of viral stocks containing abundant human papillomavirus (HPV) particles has hindered the study of infectivity and the early stages of HPV replication. The organotypic (raft) culture system has permitted the study of a number of the differentiation-specific aspects of HPV, including amplification of viral DNA, expression of late genes, and viral morphogenesis. However, these investigations have been limited to a single virus type, namely, HPV type 31 (HPV31). We have artificially introduced linearized HPV18 genomic DNA into primary keratinocytes by electroporation, followed by clonal expansion and induction of epithelial stratification and differentiation in organotypic culture. We report the synthesis of infectious HPV18 virions. Virus particles approximately 50 nm in diameter were observed by electron microscopy. HPV18 virions purified by isopycnic gradient were capable of infecting keratinocytes in vitro, as shown by the expression of multiple HPV18-specific, spliced transcripts.

Satellite remote sensing plays an important role in the monitoring of surface water for historical analysis and near real-time applications. Due to its cloud penetrating capability, many studies have focused on providing efficient and high quality methods for surface water mapping using Synthetic Aperture Radar (SAR). However, few studies have explored the effects of SAR pre-processing steps used and the subsequent results as inputs into surface water mapping algorithms. This study leverages the Google Earth Engine to compare two unsupervised histogram-based thresholding surface water mapping algorithms utilizing two distinct pre-processed Sentinel-1 SAR datasets, specifically one with and one without terrain correction. The resulting surface water maps from the four different collections were validated with user-interpreted samples from high-resolution Planet Scope data. It was found that the overall accuracy from the four collections ranged from 92% to 95% with Cohen’s Kappa coefficients ranging from 0.7999 to 0.8427. The thresholding algorithm that samples a histogram based on water edge information performed best with a maximum accuracy of 95%. While the accuracies varied between methods it was found that there is no statistical significant difference between the errors of the different collections. Furthermore, the surface water maps generated from the terrain corrected data resulted in a intersection over union metrics of 95.8%–96.4%, showing greater spatial agreement, as compared to 92.3%–93.1% intersection over union using the non-terrain corrected data. Overall, it was found that algorithms using terrain correction yield higher overall accuracy and yielded a greater spatial agreement between methods. However, differences between the approaches presented in this paper were not found to be significant suggesting both methods are valid for generating accurate surface water maps. High accuracy surface water maps are critical to disaster planning and response efforts, thus results from this study can help inform SAR data users on the pre-processing steps needed and its effects as inputs on algorithms for surface water mapping applications.

Human papillomaviruses (HPVs) are epitheliotropic viruses, and their life cycle is intimately linked to the stratification and differentiation state of the host epithelial tissues. Defining a role for the E5 gene product in the differentiation-dependent viral life cycle has been difficult due to the lack of a suitable culture system. We used the organotypic (raft) culture system to investigate the spatial and temporal expression pattern of the E5 protein during the differentiation-dependent life cycle of HPV-31b. We report the generation of antisera specific to the HPV-31b E5a protein. The HPV-31b E5a protein was detected throughout the viral life cycle in raft cultures as determined by immunostaining analyses, and the protein was localized predominantly to the basal and granular layers. Expression of epidermal growth factor receptor or platelet-derived growth factor receptors, two proteins with which E5 has been shown to interact in cell culture, did not specifically colocalize with E5a expression. However, HPV-31b E5a expression did colocalize with the epithelial differentiation-specific marker filaggrin. The kinetics of E5a protein expression during the complete viral life cycle was analyzed by immunoblotting, and the highest level was found to be coincidental with the onset of virion morphogenesis.