Features of SARS-CoV-2 Replication in Various Types of Reptilian and Fish Cell Cultures.

BACKGROUND: SARS-CoV-2 can enter the environment from the feces of COVID-19 patients and virus carriers through untreated sewage. The virus has shown the ability to adapt to a wide range of hosts, so the question of the possible involvement of aquafauna and animals of coastal ecosystems in maintaining its circulation remains open. METHODS: the aim of this work was to study the tropism of SARS-CoV-2 for cells of freshwater fish and reptiles, including those associated with aquatic and coastal ecosystems, and the effect of ambient temperature on this process. In a continuous cell culture FHM (fathead minnow) and diploid fibroblasts CGIB (silver carp), SARS-CoV-2 replication was not maintained at either 25 °C or 29 °C. At 29 °C, the continuous cell culture TH-1 (eastern box turtle) showed high susceptibility to SARS-CoV-2, comparable to Vero E6 (development of virus-induced cytopathic effect (CPE) and an infectious titer of 7.5 ± 0.17 log10 TCID50/mL on day 3 after infection), and primary fibroblasts CNI (Nile crocodile embryo) showed moderate susceptibility (no CPE, infectious titer 4.52 ± 0.14 log10 TCID50/mL on day 5 after infection). At 25 °C, SARS-CoV-2 infection did not develop in TH-1 and CNI. CONCLUSIONS: our results show the ability of SARS-CoV-2 to effectively replicate without adaptation in the cells of certain reptile species when the ambient temperature rises.

Maps of Constitutive-Heterochromatin Distribution for Four Martes Species (Mustelidae, Carnivora, Mammalia) Show the Formative Role of Macrosatellite Repeats in Interspecific Variation of Chromosome Structure.

Constitutive-heterochromatin placement in the genome affects chromosome structure by occupying centromeric areas and forming large blocks. To investigate the basis for heterochromatin variation in the genome, we chose a group of species with a conserved euchromatin part: the genus Martes [stone marten (M. foina, 2n = 38), sable (M. zibellina, 2n = 38), pine marten (M. martes, 2n = 38), and yellow-throated marten (M. flavigula, 2n = 40)]. We mined the stone marten genome for the most abundant tandem repeats and selected the top 11 macrosatellite repetitive sequences. Fluorescent in situ hybridization revealed distributions of the tandemly repeated sequences (macrosatellites, telomeric repeats, and ribosomal DNA). We next characterized the AT/GC content of constitutive heterochromatin by CDAG (Chromomycin A3-DAPI-after G-banding). The euchromatin conservatism was shown by comparative chromosome painting with stone marten probes in newly built maps of the sable and pine marten. Thus, for the four Martes species, we mapped three different types of tandemly repeated sequences critical for chromosome structure. Most macrosatellites are shared by the four species with individual patterns of amplification. Some macrosatellites are specific to a species, autosomes, or the X chromosome. The variation of core macrosatellites and their prevalence in a genome are responsible for the species-specific variation of the heterochromatic blocks.

Modification of a Tumor-Targeting Bacteriophage for Potential Diagnostic Applications.

BACKGROUND: Tumor-targeting bacteriophages can be used as a versatile new platform for the delivery of diagnostic imaging agents and therapeutic cargo. This became possible due to the development of viral capsid modification method. Earlier in our laboratory and using phage display technology, phages to malignant breast cancer cells MDA-MB 231 were obtained. The goal of this study was the optimization of phage modification and the assessment of the effect of the latter on the efficiency of phage particle penetration into MDA-MB 231 cells. METHODS: In this work, we used several methods, such as chemical phage modification using FAM-NHS ester, spectrophotometry, phage amplification, sequencing, phage titration, flow cytometry, and confocal microscopy. RESULTS: We performed chemical phage modification using different concentrations of FAM-NHS dye (0.5 mM, 1 mM, 2 mM, 4 mM, 8 mM). It was shown that with an increase of the modification degree, the phage titer decreases. The maximum modification coefficient of the phage envelope with the FAM-NHS dye was observed with 4 mM modifying agent and had approximately 804,2 FAM molecules per phage. Through the immunofluorescence staining and flow cytometry methods, it was shown that the modified bacteriophage retains the ability to internalize into MDA-MB-231 cells. The estimation of the number of phages that could have penetrated into one tumor cell was conducted. CONCLUSIONS: Optimizing the conditions for phage modification can be an effective strategy for producing tumor-targeting diagnostic and therapeutic agents, i.e., theranostic drugs.