Natural Virus Research Articles

Development of Virus-Like Particles (VLPs) for Hepatitis C Virus genotype 4: a novel approach for vaccine development in Egypt

BackgroundEgypt has the highest global prevalence of Hepatitis C Virus (HCV) infection, particularly of genotype 4. The development of a prophylactic vaccine remains crucial for HCV eradication, yet no such vaccine currently exists due to the vaccine development challenges. The ability of Virus-Like Particles (VLPs) to mimic the native virus and incorporate neutralizing and conformational epitopes, while effectively engaging both humoral and cellular immune responses, makes them a promising approach to addressing the challenges in HCV vaccine development.MethodsLentiviral-based vectors were constructed and employed to integrate the full-length sequence of Core, E1, E2, and P7 genes of HCV genotype 4 into the genome of Human Embryonic Kidney cells (HEK293T). Upon the expression, HCV structural proteins can oligomerize and self-assemble into VLPs mimicking the structure of HCV native virus. VLPs were purified and characterized for the development of a potential VLPs-based vaccine.ResultsIn this study, mammalian cells were successfully engineered to stably express HCV structural proteins and generate non-infectious VLPs for HCV genotype 4. The expression of HCV-integrated genes resulted in a successful production of HCV structural proteins, which oligomerized and self-assembled into two layers enveloped VLPs. Electron microscopy analysis of purified VLPs revealed spherical particles with an average diameter of 60–65 nm, closely resembling mature HCV virions. These results highlighted the potential of these VLPs as a vaccine candidate for HCV genotype 4.ConclusionsHCV genotype 4 remains an underexplored target in vaccine development, despite its significant public health burden, especially in Egypt. The successful generation of VLPs for this genotype represents a promising avenue for further vaccine development. The established system provides a robust platform for the production and study of VLP-based vaccines targeting HCV genotype 4.

Whole Genome Sequence Analysis of Terbinafine Resistant and Susceptible Trichophyton Isolates from Human and Animal Origin.

Trichophyton indotineae, first identified in India, has increasingly been reported in Asia, the Middle East, Europe, and recently in the USA. The global spread of terbinafine-resistant T. indotineae underscores the urgency of the issue. With its ability for human-to-human transmission, it can be considered anthropophilic. However, its highly virulent nature suggests a possible link to zoophilic species, raising the potential for disease transmission from animals. In this study, we have performed whole genome sequencing (WGS) of terbinafine susceptible and resistant Trichophyton species from animal and human origin to understand transmission dynamics of this species. Thirteen isolates of Trichophyton spp. from human (n = 9) and canine (n = 4) origin, respectively from Chandigarh and Bareilly, India, were included in this study. Isolate identification based on ITS extracted from WGS data identified six T. indotineae (ITS genotype VIII) and seven T. interdigitale (ITS genotype II) isolates. WGS single nucleotide polymorhpism (SNP) analysis separated the isolates into two distinct groups, T. indotineae and T. interdigitale and showed the clonal nature of both species. For both species, low SNP differences between isolates from humans and dogs were observed as well as low differences between isolates from Chandigarh and Bareilly, cities >350km apart from each other. These findings suggest zoonotic transmission, next to fast spread across large distances. The T. indotineae terbinafine-resistant strains exhibitedthe SQLEF397L substitution while susceptible strains had the SQLES395P substitution or demonstrated a wild-type (WT) SQLE sequence. However, all T. interdigitale strains displayed a WT SQLE sequence despite terbinafine minimum inhibitory concentrations (MICs) ranging between 0.031 to64µg/mL.

“It Takes a Village”: Parental Experiences with Cooperation to Overcome Cyberbullying in Norwegian Schools

AbstractThis study investigated parental beliefs and experiences with cooperation among adults to overcome cyberbullying in Norwegian schools. Data comprised 17 semi-structured interviews conducted with parents of adolescents enrolled in primary or lower secondary schools in Norway. Reflexive thematic analysis revealed four themes: (1) individual and (2) organizational aspects influencing cooperation, (3) cyberbullying as a form of aggression, and (4) interactional dimensions of cooperation. These findings highlight the complexity of cyberbullying characteristics and mesosystemic cooperation, which require expertise in various fields. The viral and covert nature of cyberbullying requires adults to create spaces, online or offline, where children may safely report unwanted behaviors without the fear of retaliation. Any barriers to cooperation, such as language, lack of technological knowledge, and resource allocation, must be addressed. Immigrant parents contributed insights regarding barriers to cooperation due to cultural and linguistic differences. Attitudes, behaviors, and outcomes need to be addressed before cooperation based on trust, support, and realistic goals can be implemented.

Immunogenic recombinant Mayaro virus-like particles present natively assembled glycoprotein

AbstractVirus-like particles (VLPs) are an established vaccine platform and can be strong immunogens capable of eliciting both humoral and cellular immune responses against a range of pathogens. Here, we show by cryo-electron microscopy that VLPs of Mayaro virus, which contain envelope glycoproteins E1-E2 and capsid, exhibit an architecture that closely resembles native virus. In contrast to monomeric and soluble envelope 2 (E2) glycoprotein, both VLPs as well as the adenovirus and modified vaccinia virus Ankara (MVA) vaccine platforms expressing the equivalent envelope glycoproteins E1-E2, and capsid induced highly neutralising antibodies after immunisation. The levels of neutralising antibodies elicited by the viral-vectored vaccines of structural proteins and VLPs increased significantly upon boosting. Immunisation of Mayaro virus VLPs in mice with or without an adjuvant (poly:IC) yielded similar levels of neutralising antibodies suggesting that the VLPs may be used for immunisation without the need for an adjuvant. A single or two doses of non-adjuvanted 5 µg of MAYV VLP vaccination provided significant protection against viremia and MAYV-induced foot swelling in the C57BL/6 mouse challenge model. MAYV VLPs represent a non-infectious vaccine candidate, which may constitute a complementary option for future immunisation strategies against this important emerging alphavirus.

Effect of pandemic influenza A virus PB1 genes of avian origin on viral RNA polymerase activity and pathogenicity.

Zoonotic influenza A virus (IAV) infections pose a substantial threat to global health. The influenza RNA-dependent RNA polymerase (RdRp) comprises the PB2, PB1, and PA proteins. Of the last four pandemic IAVs, three featured avian-origin PB1 genes. Prior research linked these avian PB1 genes to increased viral fitness when reassorted with human IAV genes. This study evaluated chimeric RdRps with PB1 genes from the 1918, 1957, and 1968 pandemic IAVs in a low pathogenic avian influenza (LPAI) virus background to assess polymerase activity and pathogenicity. Substituting in the pandemic PB1 genes reduced polymerase activity, virulence, and altered lung pathology, while the native LPAI PB1 showed the highest pathogenicity and polymerase activity. The native LPAI PB1 virus caused severe pneumonia and high early viral RNA levels, correlating with elevated host cytokine signaling. Increased genetic distance from the LPAI PB1 sequence correlated with reduced polymerase activity, IFN-β expression, viral replication, and pathogenicity.

Rapid voltammetric determination of hemagglutinin in saliva using magnetic nanoparticles modified with triazolotriazine in a portable cell design

Express determination of the viral nature of the diseases is one of the most important tasks of modern medicine. Portable electrochemicalanalytical devices are used to complement standard laboratory analytical methods and implement analysis at the patient's bedside. A rapid approach of voltammetric determination of the viral marker protein hemagglutinin (НA) in saliva using portable electrochemical microcell based on the three-electrode planar system (TPS) and magnetite nanoparticles modified with 3-nitro-4‑hydroxy-1,4-dihydro-7-propargylthio-1,2,4-triazolo[5,1-c][1,2,4]triazine (MNPs-Aza) was developed. MNPs-Aza were synthesized by carbodiimide cross-linking and characterized by infrared spectroscopy and square wave voltammetry. Modified MNPs were used simultaneously as carriers of the HA receptor and as agents for HA-Aza complexes concentrating on the surface of the working electrode. The normalized difference between the peak current values of the first stage of electroreduction of Aza before and after selective interaction with the HA in the analyzed sample (I*) was used as the analytical signal. Under the selected operating conditions of the developed portable microcell, linear calibration I* = (12.81±0.26) lg сHA + (136.75±1.73) was obtained in the range of 10–3–10–9 M. The limit of detection (LOD) calculated by the 3σ criteria was 8.4·10–10 M. The developed approach was successfully tested on model solutions and real saliva samples.

Structural insights into Semiliki forest virus receptor binding modes indicate novel mechanism of virus endocytosis.

The Very Low-Density Lipoprotein Receptor (VLDLR) is an entry receptor for the prototypic alphavirus Semliki Forest Virus (SFV). However, the precise mechanisms underlying the entry of SFV into cells mediated by VLDLR remain unclear. In this study, we found that of the eight class A (LA) repeats of the VLDLR, only LA2, LA3, and LA5 specifically bind to the native SFV virion while synergistically promoting SFV cell attachment and entry. Furthermore, the multiple cryo-electron microscopy structures of VLDLR-SFV complexes and mutagenesis studies have demonstrated that under physiological conditions, VLDLR primarily binds to E1-DIII of site-1, site-2, and site-1' at the twofold symmetry axes of SFV virion through LA2, LA3, and LA5, respectively. These findings unveil a novel mechanism for viral entry mediated by receptors, suggesting that conformational transitions in VLDLR induced by multivalent binding of LAs facilitate cellular internalization of SFV, with significant implications for the design of antiviral therapeutics.

Phenotypic Characterization and Genome Analysis of New Broad-Spectrum Virulent Salmophage, Salmonella Phage KKP_3822, for Biocontrol of Multidrug-Resistant Salmonella enterica Strains.

Salmonella is one of the main foodborne pathogens. Irrational antibiotic management has led to an increase in the incidence of multidrug-resistant strains. Bacteriophages may be an alternative method of food biopreservation and contribute to reducing the number of food poisonings requiring pharmacotherapy. This study aimed to isolate a bacteriophage (phage) targeting indigenous multidrug-resistant (MDR) Salmonella strains, followed by their biological, morphological, and genomic characterization. In this study we isolated Salmonella phage KKP_3822, targeting MDR Salmonella Manchester strain KKP 1213. Salmonella phage KKP_3822 retained high activity in the temperature range from -20 °C to 40 °C and active acidity from pH 3 to 11. Temperatures of 70 °C and 80 °C and extreme pH values (2 and 12) significantly reduced the phage titer. Its activity decreased proportionally to the time of UV exposure. Genome analysis (linear dsDNA with a length of 114,843 bp) revealed the presence of 27 tRNA genes. Proteins encoded by the vB_Sen-IAFB3822 phage were divided into functional modules related to (i) phage structure/assembly, (ii) DNA replication/modification/regulation, (iii) phage lysis, and (iv) DNA packaging into the capsid. No genes associated with antibiotic resistance or integration into the host genome, markers of temperate bacteriophages, were annotated in the Salmonella phage KKP_3822 genome. Based on morphological features and whole-genome sequence analysis, the newly isolated Salmonella phage KKP_3822 shows the greatest similarity to representatives of tailed phages from the Caudoviricetes class, Demerecviridae family, and Epseptimavirus genus. Genome analysis confirmed the virulent nature of the Salmonella phage KKP_3822, making it a potential candidate for food biocontrol.

Binding of Infectious Hypodermal and Haematopoietic Necrosis Virus-Like Particles to Mannose Receptor Stimulates Antimicrobial Responses in Immune-Related Tissues of Peneaus vannamei.

Mannose receptor (MR) is a transmembrane protein and a type of pattern-recognition receptor (PRR) that plays a critical role in the immunity of mammals and fish. In this study, we examined the role of MR in binding with infectious hypodermal and haematopoietic necrosis virus-like particle (IHHN-VLP) and the downstream immune pathway that it triggers in the shrimp Peneaus vannamei. Upon IHHN-VLP challenge, transcripts of MR in P. vannamei (PvMR) increased significantly in all examined tissues, particularly those related to shrimp immunity, including hemocyte, hepatopancreas and gill tissues. Specifically, IHHN-VLP bound to the 34-kDa PvMR protein in shrimp-tissue extracts. Immunohistochemistry results of hemocytes showed that PvMR was initially localised on the plasma membrane but later internalised and dispersed throughout the cytoplasm after IHHN-VLP administration. Binding between IHHN-VLP and PvMR also induced significant upregulation of genes for the antimicrobial peptides (AMPs) penaeidin 3 and crustin, presumably to protect the shrimp against the viral infection. However, knocking down PvMR resulted in down-regulation of all immune-related genes examined. Overall, as an immune-related PRR, PvMR serves as a receptor for invading viruses, which then trigger the expression of AMPs. Strategic designs using PvMR could be developed to either block the interaction of native virus with the host cells or provoke its up-regulation to enhance shrimp immunity, which could open up opportunities to fight against IHHNV infection in shrimp.

Exploring challenges and policy considerations in point-of-care testing for hospital preparedness ahead of infectious disease emergencies: A qualitative study

BackgroundDespite the uncertainty raised by several studies regarding the practicality of Point-of-Care Testing (POCT) in hospital settings, the urgency prompted by the COVID-19 pandemic led many hospitals to invest in these rapid diagnostics. As the COVID-19 pandemic showcased the potential of POCT in emergency situations, an urgent call arises to leverage the successes and lessons learned for the long-term benefit of healthcare systems, particularly in preparation for the next pandemic of viral respiratory nature. MethodsWe conducted semi-structured interviews with nine emergency care and hospital laboratory stakeholders with expertise and responsibility over POCT operations within hospital systems during the COVID-19 pandemic in Maryland, USA. The grounded theory approach was utilized, and the transcripts were thematically analyzed through an iterative process. ResultsThree distinct themes were identified as barriers to POCT implementation: Regulatory barriers encompassed inconsistencies between the federal and state regulations and a regulatory structure that lags technological advancements. Staffing and operational barriers comprised a shortage of POCT workforce including nurses and Point of Care coordinators. Economic barriers were linked to the overall cost of integrating new POCT devices into the existing testing menu at emergency departments. ConclusionWe conclude with strategies, including policy considerations, to enable and sustain decentralized POCT programs within hospitals. Some of these strategies include expanding the workforce trained in the regulatory procedures required to support POCT; harmonizing state and federal regulations for diagnostic testing, especially for FDA-waived POCT; and sustained funding for POCT development and use for public health preparedness.

Analysis of the antigenic and immunogenic properties of the native rabies virus glycoprotein purified by Lens culinaris lectin affinity chromatography

Rabies virus glycoprotein (RABV-G) is responsible for the recognition of specific cell surface receptors and induces the production of neutralizing antibodies (VNA). Since RABV-G is a glycoprotein, this work aimed to evaluate Lens culinaris (LCA) chromatography as a simple and effective purification method. The purity and identification of the protein obtained were analyzed by SDS-PAGE, ELISA and lectin-binding assay. The antigenic properties of the purified RABV-G were evaluated by direct ELISA using human serum samples from individuals who had received rabies pre-exposure vaccination. For the immunogenicity study, Swiss Webster mice were immunized with purified RABV-G and the specific antibodies were measured by direct ELISA and RFFIT. As results, it was observed that the purified protein reveled a molecular mass of 55 kDa and the presence of carbohydrate; additionally, it was recognized by anti-rabies virus glycoprotein monoclonal antibody. Purified RABV-G induced high VNA titers (>50.0 IU/ml) in vivo, as detected by RFFIT, as well as RABV-G specific IgG1 (0.8 mean OD±SD) and IgG2a (0.3 mean OD±SD) antibodies, with a predominance of IgG1 (p< 0.001). In addition, it was observed that RABV-G was efficient in selectively detecting anti- RABV-G IgG in the sera of vaccinated individuals compared to the negative control. Therefore, LCA chromatography was efficient in preserving the native properties of RABV-G that are essential in inducing an adequate humoral immune response. In addition, the purified RABV-G presented analytical potential as an ELISA reagent.

Glass-like Relaxation Dynamics during the Disorder-to-Order Transition of Viral Nucleocapsids.

Nucleocapsid self-assembly is an essential yet elusive step in virus replication. Using time-resolved small-angle X-ray scattering on a model icosahedral ssRNA virus, we reveal a previously unreported kinetic pathway. Initially, RNA-bound capsid subunits rapidly accumulate beyond the stoichiometry of native virions. This is followed by a disorder-to-order transition characterized by glass-like relaxation dynamics and the release of excess subunits. Our molecular dynamics simulations, employing a coarse-grained elastic model, confirm the physical feasibility of self-ordering accompanied by subunit release. The relaxation can be modeled by an exponential integral decay on the mean squared radius of gyration, with relaxation times varying within the second range depending on RNA type and subunit concentration. A nanogel model suggests that the initially disordered nucleoprotein complexes quickly reach an equilibrium size, while their mass fractal dimension continues to evolve. Understanding virus self-assembly is not only crucial for combating viral infections, but also for designing synthetic virus-inspired nanocages for drug delivery applications.

Maximizing Non-Profit Engagement on TikTok Through the Effective Application of AIDA in Indonesia

General Background: Social media platforms like TikTok have emerged as powerful marketing tools, enabling organizations to engage with audiences more effectively. Specific Background: Yayasan Lembaga Manajemen Infaq Ukhuwah Islamiyah (LMI Zakat) leverages TikTok to promote its programs, including MSIB (Magang dan Studi Independen Bersertifikat), through its official account @lmizakat. The integration of marketing strategies such as the AIDA (Awareness, Interest, Desire, Action) model provides insight into consumer behavior and the effectiveness of digital content. Knowledge Gap: While studies on social media marketing are extensive, limited research has explored the application of AIDA principles on platforms like TikTok within the context of non-profit organizations. Aims: This research aims to examine the implementation of Integrated Marketing Communication (IMC) by LMI Zakat through TikTok, focusing on how the AIDA model enhances marketing communications and program engagement. Results: Findings indicate that LMI Zakat successfully applied the AIDA model through creative and educational content, which effectively raised brand awareness, generated interest, and increased engagement with the MSIB program. Specifically, the content utilized viral trends to raise awareness and generated desire through informative and persuasive narratives. Novelty: The study provides a unique contribution by highlighting how a zakat organization can use TikTok’s content and viral nature to apply marketing principles, which are typically associated with for-profit sectors. Implications: This research underscores the potential for zakat and non-profit organizations to enhance their marketing strategies through digital platforms like TikTok. Future studies should explore diverse content types and apply AIDA across other social media platforms, contributing to a broader understanding of digital marketing in non-profit contexts. Highlights: TikTok applies AIDA effectively for LMI Zakat's marketing communication. Viral content boosts awareness and engagement with MSIB program. Future studies should explore content variation and cross-platform strategies. Keywords: TikTok marketing, AIDA model, LMI Zakat, non-profit promotion, digital communication

Native BK Virus Nephropathy in a Patient with Multiple Myeloma

Native BK Virus Nephropathy in a Patient with Multiple Myeloma

A case of Atypical Keratitis by the rare fungus Pseudallescheria Boydii in a young male

We present a case of a 24-year-old male with fungal keratitis caused by a rare, atypical fungus Pseudallescheria boydii. Initially, aggressive medical therapy was given, but due to the virulent nature of fungus and its rapid spread, therapeutic keratoplasty had to be done to control the infection. Following healing and scarring after 4 months, Optical PK (penetrating keratoplasty) with cataractous lens exchange was done for visual rehabilitation.

Beta Spike-Presenting SARS-CoV-2 Virus-like Particle Vaccine Confers Broad Protection against Other VOCs in Mice.

Virus-like particles (VLPs) are non-infectious and serve as promising vaccine platforms because they mimic the membrane-embedded conformations of fusion glycoproteins on native viruses. Here, we employed SARS-CoV-2 VLPs (SMEN) presenting ancestral, Beta, or Omicron spikes to identify the variant spike that elicits potent and cross-protective immune responses in the highly sensitive K18-hACE2 challenge mouse model. A combined intranasal and intramuscular SMEN vaccine regimen generated the most effective immune responses to significantly reduce disease burden. Protection was primarily mediated by antibodies, with minor but distinct contributions from T cells in reducing virus spread and inflammation. Immunization with SMEN carrying ancestral spike resulted in 100, 75, or 0% protection against ancestral, Delta, or Beta variant-induced mortality, respectively. However, SMEN with an Omicron spike provided only limited protection against ancestral (50%), Delta (0%), and Beta (25%) challenges. By contrast, SMEN with Beta spikes offered 100% protection against the variants used in this study. Thus, the Beta variant not only overcame the immunity produced by other variants, but the Beta spike also elicited diverse and effective humoral immune responses. Our findings suggest that leveraging the Beta variant spike protein can enhance SARS-CoV-2 immunity, potentially leading to a more comprehensive vaccine against emerging variants.

Virulence Adaptation by Rice Planthoppers and Leafhoppers to Resistance Genes and Loci: A Review.

In recent decades, research on developing and deploying resistant rice has accelerated due to the availability of modern molecular tools and, in particular, advances in marker-assisted selection. However, progress in understanding virulence adaptation has been relatively slow. This review tracks patterns in virulence adaptation to resistance genes (particularly Bph1, bph2, Bph3, and bph4) and examines the nature of virulence based on selection experiments, responses by virulent populations to differential rice varieties (i.e., varieties with different resistance genes), and breeding experiments that interpret the genetic mechanisms underlying adaptation. The review proposes that varietal resistance is best regarded as a combination of minor and major resistance traits against which planthoppers develop partial or complete virulence through heritable improvements that are reversable or through evolutionary adaptation, respectively. Agronomic practices, deployment patterns, and herbivore population pressures determine the rates of adaptation, and there is growing evidence that pesticide detoxification mechanisms can accelerate virulence adaptation. Research to delay adaptation has mainly focused on gene pyramiding (i.e., including ≥ two major genes in a variety) and multilines (i.e., including ≥ two resistant varieties in a field or landscape); however, these strategies have not been adequately tested and, if not managed properly, could inadvertently accelerate adaptation compared to sequential deployment. Several research gaps remain and considerable improvements in research methods are required to better understand and manage virulence adaptation.

Genome-wide search and gene expression studies reveal candidate effectors with a role in pathogenicity and virulence in Fusarium graminearum

ABSTRACT Fusarium graminearum causes Fusarium head blight (FHB) disease in wheat worldwide. Although F. graminearum is reported to secrete several effectors, their role in virulence and pathogenicity is unknown. The study aimed at identifying candidate genes with a role in pathogenicity and virulence using two different host systems, Arabidopsis thaliana and wheat, challenged with F. graminearum TN01. Detached leaf assay and histological studies revealed the virulent nature of TN01. A genome-wide in silico search revealed several candidate genes, of which 23 genes were selected based on reproducibility. Gene expression studies by reverse transcriptase–polymerase chain reaction (RT-PCR) in leaf tissues of Arabidopsis and the two wheat genotypes, the susceptible (Sonalika) and the resistant (Nobeoka Bozu/Nobeoka), compared with mock-treated controls in a time-course study using fungal- and plant-specific genes as internal controls revealed that these genes were differentially regulated. Further, expression of these candidates in F. graminearum–inoculated Sonalika and Nobeoka spikes compared with mock-treated controls revealed their role in pathogenicity and virulence. Gene ontology studies revealed that some of these secretory proteins possessed a role in apoptosis and ceratoplatanin and KP4 killer toxin syntheses. A three-dimensional protein configuration was performed by homology modeling using trRosetta. Further, real-time quantitative PCR (RT-qPCR) studies in F. graminearum–inoculated Arabidopsis and wheat at early time points of inoculation revealed an increased expression of the majority of these genes in Sonalika, suggesting their possible role in pathogenicity, whereas low mRNA abundance was observed for 11 of these genes in the resistant genotype, Nobeoka, compared with Sonalika, indicating their role in virulence of F. graminearum.

Immune status of patients with community-acquired pneumonia associated with a new coronavirus infection and other viral and bacterial pathogens

Despite the fact that the state of the individual’s immune system plays an important role in developing community-acquired pneumonia, its clinical features are determined not only by immune response characteristics, but also by the nature of the infectious agent. The aim of the work was to assess immune status of patients with community-acquired pneumonia with lung damage ranging from 2 to 12%, in whom pathogens of a viral, bacterial and fungal nature were verified (n = 96, aged 46,3±20,5) who were admitted to the hospital 5–7 days after disease onset. Materials and methods. The selection of smears and sputum specimens was carried out on day 1 after admission. COVID-19 pathogen was analyzed in smears by PCR using the Vector-PCRR-2019-nCoV-RG kit (SSC Vector of Rospotrebnadzor, Russia). Biochemical activity, colony morphology, and pathogen concentration (in the amount of ≥ 105 CFU/ml) were also evaluated. Based on the results of preliminary studies, target patients groups were formed: with a new coronavirus infection caused by the SARS-CoV-2 gene variant B.1.1.529 (Omicron) (n = 33); with COVID-19 and bacterial pathogens (n = 17); with COVID-19 and combined bacterial and fungal infection (n = 16); patients with identified pathogens of bacterial infections (n = 13); patients with combined bacterial and fungal infection (n = 17). The relative and absolute level of CD45+CD3+ lymphocytes, CD45+CD3+CD4+ cells, CD45+CD3+CD8+ lymphocytes, CD45+CD3-CD16+CD56+ cells, CD45+CD3+CD16+CD56+ lymphocytes, CD45+CD45RO–CD45RA+ and CD45+CD45RO+CD45RA– lymphocytes, CD45+CD19+ cells, CD45+CD5+CD19–CD27– and CD45+CD19+CD5–CD27– lymphocytes, CD45+CD19+CD5–CD27+ cells was assessed in all volunteers. Results. The analysis of the immune status of patients with community-acquired pneumonia associated with Omicron gene variant new coronavirus infection revealed T-lymphocytopenia, a decreased number of CD8+ lymphocytes, an increased relative and absolute number of NK cells. In COVID-19 patients, who also had bacterial pathogens verified, there was a decrease in relative and absolute number of T-helper cells, an increased relative and absolute number of B and B2 lymphocytes. In COVID-19-negative volunteers with bacterial and bacterial-fungal community-acquired pneumonia, the relative and absolute level of B and B2 lymphocytes, as well as the general memory B cell population, was increased. In contrast to COVID-19-positive patients, in the absence of T-lymphocytopenia, a decreased number of CD4+ cells and rise in number of cytotoxic lymphocytes were recorded in patients of these groups. Conclusions. The data obtained indicate that in the immune status of patients with community-acquired pneumonia, depending on the nature of the infectious agent, changes in relative and absolute level of T-helper cells, cytotoxic lymphocytes, as well as in the qualitative and quantitative B-lymphocyte population composition are recorded.

Safeguarding reverse osmosis membrane integrity in drinking water production: A pilot study using 2 native bacterial communities and viruses in surface water as integrity indicators

Water treatment train including a reverse osmosis barrier is capable of producing high-quality water coupled with microbial safety. However, analytically proving this with log removal value (LRV) > 3 has remained a challenge. Conservative techniques that quantify salt or sulfate ion passage as virus surrogates are currently used to credit the membrane system for its microbial removal capacity. These methods are insensitive, and it is now understood that the transport mechanism correlation between these surrogates with the actual passage of viruses is weak. There is also a huge gap between the current log credit assigned (using spiking of surrogates in feed) and the actual log removal that can be proven in the drinking water plant. In this paper, we introduce two techniques that exploit the high concentration of microbial communities in fresh surface water by quantifying bacteria through flow cytometry (FC) and native viruses through qPCR assays. These two methods are compared to LRV of conductivity, TOC, and fluorescent markers. That high concentration of bacterial community allowed for LRV > 4 by flow cytometry in an intact module. After inflicting damage with a 1.5 mm defect, the FC method detected the damage, and a bacterial cell count >103 cell count/ml was measured in the permeate, while conductivity lacked sensitivity to monitor any integrity loss. Furthermore, after chlorine exposure to the membrane, both bacteria and virus detected no leakage, proving the membrane's intactness and also demonstrating the effect of chlorination on the passage of small solutes. The implementation of the two microbial quantifying methods in full-scale water treatment plants facilitates higher sensitivity in membrane integrity loss than currently used methods.