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Cyanobacteria are adaptable and dominant organisms that exist in many harsh and extreme environments due to their great ecological tolerance. They produce various secondary metabolites, including cyanotoxins. While cyanobacteria are well studied in surface waters and some aerial habitats, numerous other habitats and niches remain underexplored. We collected 61 samples of: (i) biofilms from springs, (ii) aerial microbial mats from buildings and subaerial mats from caves, and (iii) water from borehole wells, caves, alkaline, saline, sulphidic, thermal, and iron springs, rivers, seas, and melted cave ice from five countries (Croatia, Georgia, Italy, Serbia, and Slovenia). We used (q)PCR to detect cyanobacteria (phycocyanin intergenic spacer-PC-IGS and cyanobacteria-specific 16S rRNA gene) and cyanotoxin genes (microcystins- mcyE , saxitoxins- sxtA , cylindrospermopsins- cyrJ ), as well as amplicon sequencing and morphological observations for taxonomic identification. Cyanobacteria were detected in samples from caves, a saline spring, and an alkaline spring. While mcyE or sxtA genes were not observed in any sample, cyrJ results showed the presence of a potential cylindrospermopsin producer in a biofilm from a sulphidic spring in Slovenia. This study contributes to our understanding of cyanobacteria occurrence in diverse habitats, including rare and extreme ones, and provides relevant methodological considerations for future research in such environments.

As a result of the COVID-19 pandemic, many new materials and masks came onto the market. To determine their suitability, several standards specify which properties to test, including bacterial filtration efficiency (BFE), while none describe how to determine viral filtration efficiency (VFE), a property that is particularly important in times of pandemic. Therefore, we focused our research on evaluating the suitability and efficiency of different systems for determining VFE. Here, we evaluated the VFE of 6 mask types (e.g., a surgical mask, a respirator, material for mask production, and cloth masks) with different filtration efficiencies in four experimental setups and compared the results with BFE results. The study included 17 BFE and 22 VFE experiments with 73 and 81 mask samples tested, respectively. We have shown that the masks tested had high VFE (>99% for surgical masks and respirators, ≥98% for material, and 87-97% for cloth masks) and that all experimental setups provided highly reproducible and reliable VFE results (coefficient of variation < 6%). Therefore, the VFE tests described in this study can be integrated into existing standards for mask testing.

The outbreak of the worrisome coronavirus disease in 2019 has caused great concern among the global public, especially regarding the need for personal protective equipment with applied antiviral agents to reduce the spread and transmission of the virus. Thus, in our research, chitosan-based bioactive polymers as potential antiviral agents were first evaluated as colloidal macromolecular solutions by elemental analysis and charge. Three different types of low and high molecular weight chitosan (LMW Ch, HMW Ch) and a LMW Ch derivative, i.e., quaternary chitosan (quart-LMW Ch), were used. To explore their antiviral activity for subsequent use in the form of coatings, the macromolecular Chs dispersions were incubated with the model virus phi6 (surrogate for SARS-CoV-2), and the success of virus inactivation was determined. Inactivation of phi6 with some chitosan-based compounds was very successful (>6 log), and the mechanisms behind this were explored. The changes in viral morphology after incubation were observed and the changes in infrared bands position were determined. In addition, dynamic and electrophoretic light scattering studies were performed to better understand the interaction between Chs and phi6. The results allowed us to better understand the antiviral mode of action of Chs agents as a function of their physicochemical properties.

Early diagnosis with rapid detection of the virus plays a key role in preventing the spread of infection and in treating patients effectively. In order to address the need for a straightforward detection of SARS-CoV-2 infection and assessment of viral spread, we developed rapid, sensitive, extraction-free one-step reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and reverse transcription loop-mediated isothermal amplification (RT-LAMP) tests for detecting SARS-CoV-2 in saliva. We analyzed over 700 matched pairs of saliva and nasopharyngeal swab (NSB) specimens from asymptomatic and symptomatic individuals. Saliva, as either an oral cavity swab or passive drool, was collected in an RNA stabilization buffer. The stabilized saliva specimens were heat-treated and directly analyzed without RNA extraction. The diagnostic sensitivity of saliva-based RT-qPCR was at least 95% in individuals with subclinical infection and outperformed RT-LAMP, which had at least 70% sensitivity when compared to NSBs analyzed with a clinical RT-qPCR test. The diagnostic sensitivity for passive drool saliva was higher than that of oral cavity swab specimens (95% and 87%, respectively). A rapid, sensitive one-step extraction-free RT-qPCR test for detecting SARS-CoV-2 in passive drool saliva is operationally simple and can be easily implemented using existing testing sites, thus allowing high-throughput, rapid, and repeated testing of large populations. Furthermore, saliva testing is adequate to detect individuals in an asymptomatic screening program and can help improve voluntary screening compliance for those individuals averse to various forms of nasal collections.

Due to increased frequency of cyanobacterial blooms and emerging evidence of cyanotoxicity in biofilm, reliable methods for early cyanotoxin threat detection are of major importance for protection of human, animal and environmental health. To complement the current methods of risk assessment, this study aimed to evaluate selected qPCR assays for detection of potentially toxic cyanobacteria in environmental samples. In the course of one year, 25 plankton and 23 biofilm samples were collected from 15 water bodies in Slovenia. Three different analyses were performed and compared to each other; qPCR targeting mcyE , cyrJ and sxtA genes involved in cyanotoxin production, LC-MS/MS quantifying microcystin, cylindrospermopsin and saxitoxin concentration, and microscopic analyses identifying potentially toxic cyanobacterial taxa. qPCR analyses detected potentially toxic Microcystis in 10 lake plankton samples, and potentially toxic Planktothrix cells in 12 lake plankton and one lake biofilm sample. A positive correlation was observed between numbers of mcyE gene copies and microcystin concentrations. Potential cylindrospermopsin- and saxitoxin-producers were detected in three and seven lake biofilm samples, respectively. The study demonstrated a potential for cyanotoxin production that was left undetected by traditional methods in both plankton and biofilm samples. Thus, the qPCR method could be useful in regular monitoring of water bodies to improve risk assessment and enable timely measures.