Quantification of oxidative stress markers in the blood sera following subacute administration of different oximes in rats.

Oxidative stress status, as a disruption of redox homeostasis, in the blood sera of Wistar rats caused by repeated application of selected acetylcholinesterase reactivators - asoxime, obidoxime, K027, K048, K074, and K075 were evaluated. Throughout this study, each oxime in a dose of 0.1 of LD50/kg im was given 2x/week for 4 weeks. Then, seven days after the last oximes' application, markers of lipid peroxidation (malondialdehyde, MDA), and protein oxidation (advanced oxidation protein products, AOPP), as well as the activity of antioxidant enzymes (catalase, CAT, superoxide dismutase, SOD, reduced glutathione, GSH, and oxidized glutathione, GSSG), were determined. Oxidative stress parameters, MDA and AOPP were significantly highest in the K048-, K074- and K075-treated groups (p < 0.001). The activity of CAT was significantly elevated in the obidoxime-treated group (p < 0.05), while treatment with K027, K048, and K074 induced high elevation in SOD levels (p < 0.01, p < 0.001). Interestingly, the activity of GSH in each oxime-treated group was significantly elevated. Unlike, treatment with obidoxime caused elevation in GSSG levels (p < 0.01). As a continuation of our previously published data, these results assure that applied oximes following subacute treatment ameliorated the oxidative status and further adverse systemic toxic effects in rats.

Alterations to sphingolipid metabolism from antipsychotic administration in healthy volunteers are restored following the use of cannabidiol.

Randomized clinical trials substantiate cannabidiol (CBD) as a next-generation antipsychotic, effective in alleviating positive and negative symptoms associated with psychosis, while minimising the adverse effects seen with established treatments. Although the mechanisms remain debated, CBD is known to induce drug-responsive changes in lipid-based retrograde neurotransmitters. Lipid aberrations are also frequently observed with antipsychotics, which may contribute to their efficacy or increase the risk of undesirables, including metabolic dysfunction, obesity and dyslipidaemia. Our study investigated CBD's impact following lipid responses triggered by interaction with second-generation antipsychotics (SGA) in a randomized phase I safety study. Untargeted mass spectrometry assessed the lipidomic profiles of human sera, collected from 38 healthy volunteers. Serum samples were obtained prior to commencement of any medication (t = 0), 3 days after consecutive administration of one of the five, placebo-controlled, treatment arms designed to achieve steady-state concentrations of each SGA (amisulpride, 150 mg/day; quetiapine, 300 mg/day; olanzapine 10 mg/day; risperidone, 3 mg/day), and after six successive days of SGA treatment combined with CBD (800 mg/day). Receiver operating characteristics (ROC) refined 3712 features to a putative list of 15 lipids significantly altered (AUC > 0.7), classified into sphingolipids (53 %), glycerolipids (27 %) and glycerophospholipids (20 %). Targeted mass spectrometry confirmed reduced sphingomyelin and ceramide levels with antipsychotics, which mapped along their catabolic pathway and were restored by CBD. These sphingolipids inversely correlated with body weight after olanzapine, quetiapine, and risperidone treatment, where CBD appears to have arrested or attenuated these effects. Herein, we propose CBD may alleviate aberrant sphingolipid metabolism and that further investigation into sphingolipids as markers for monitoring side effects of SGAs and efficacy of CBD is warranted.

In silico study to explore the mechanism of Toxoplasma-induced inflammation and target therapy based on sero and salivary Toxoplasma.

We aimed to assess salivary and seroprevalence of Toxoplasma immunoglobulins in risky populations and evaluate drug docking targeting TgERP. A cross-sectional study was conducted in Alexandria University hospitals' outpatient clinics. 192 participants were enrolled from September 2022 to November 2023. Anti-Toxoplasma IgG and IgM were determined in serum and saliva by ELISA. An in-Silico study examined TgERP's protein-protein interactions (PPIs) with pro-inflammatory cytokine receptors, anti-inflammatory cytokine, cell cycle progression regulatory proteins, a proliferation marker, and nuclear envelope integrity-related protein Lamin B1. Our findings revealed that anti-T. gondii IgG were detected in serum (66.1%) and saliva (54.7%), with 2.1% of both samples were positive for IgM. Salivary IgG had 75.59% sensitivity, 86.15% specificity, 91.40% PPV, 64.40% NPP, 79.17% accuracy and fair agreement with serum IgG. On the other hand, the sensitivity, specificity, PPV, NPV, and accuracy in detecting salivary IgM were 75.0%, 99.47%, 75.0%, 99.47%, and 98.96%. AUC 0.859 indicates good discriminatory power. Examined synthetic drugs and natural products can target specific amino acids residues of TgERP that lie at the same binding interface with LB1 and Ki67, subsequently, hindering their interaction. Hence, salivary samples can be a promising diagnostic approach. The studied drugs can counteract the pro-inflammatory action of TgERP.

Cross-Reactive Antibody Responses to Coronaviruses Elicited by SARS-CoV-2 Infection or Vaccination.

BACKGROUND: The newly emerged SARS-CoV-2 possesses shared antigenic epitopes with other human coronaviruses. We investigated if COVID-19 vaccination or SARS-CoV-2 infection may boost cross-reactive antibodies to other human coronaviruses. METHODS: Prevaccination and postvaccination sera from SARS-CoV-2 naïve healthy subjects who received three doses of the mRNA vaccine (BioNTech, BNT) or the inactivated vaccine (CoronaVac, CV) were used to monitor the level of cross-reactive antibodies raised against other human coronaviruses by enzyme-linked immunosorbent assay. In comparison, convalescent sera from COVID-19 patients with or without prior vaccination history were also tested. Pseudoparticle neutralization assay was performed to detect neutralization antibody against MERS-CoV. RESULTS: Among SARS-CoV-2 infection-naïve subjects, BNT or CV significantly increased the anti-S2 antibodies against Betacoronaviruses (OC43 and MERS-CoV) but not Alphacoronaviruses (229E). The prevaccination antibody response to the common cold human coronaviruses did not negatively impact the postvaccination antibody response to SARS-CoV-2. Cross-reactive antibodies that binds to the S2 protein of MERS-CoV were similarly detected from the convalescent sera of COVID-19 patients with or without vaccination history. However, these anti-S2 antibodies do not possess neutralizing activity in MERS-CoV pseudoparticle neutralization tests. CONCLUSIONS: Our results suggest that SARS-CoV-2 infection or vaccination may potentially modulate population immune landscape against previously exposed or novel human coronaviruses. The findings have implications for future sero-epidemiological studies on MERS-CoV.

SARS-CoV-2 omicron BA.2.87.1 exhibits higher susceptibility to serum neutralization than EG.5.1 and JN.1.

As SARS-CoV-2 continues to spread and mutate, tracking the viral evolutionary trajectory and understanding the functional consequences of its mutations remain crucial. Here, we characterized the antibody evasion, ACE2 receptor engagement, and viral infectivity of the highly mutated SARS-CoV-2 Omicron subvariant BA.2.87.1. Compared with other Omicron subvariants, including EG.5.1 and the current predominant JN.1, BA.2.87.1 exhibits less immune evasion, reduced viral receptor engagement, and comparable infectivity in Calu-3 lung cells. Intriguingly, two large deletions (Δ15-26 and Δ136-146) in the N-terminal domain (NTD) of the spike protein facilitate subtly increased antibody evasion but significantly diminish viral infectivity. Collectively, our data support the announcement by the USA CDC that the public health risk posed by BA.2.87.1 appears to be low.

Development of real-time recombinase polymerase amplification (RPA) and RPA combined with lateral flow dipstick (LFD) assays for the rapid and sensitive detection of cyprinid herpesvirus 3.

In this issue, we established rapid, cost-effective, and simple detection methods including recombines polymerase amplification with lateral flow dipstick (RPA-LFD) and real-time RPA for cyprinid herpesvirus 3(CyHV-3), and evaluated their sensitivity, specificity, and applicability, the real-time RPA method could achieve sensitive diagnosis of CyHV-3 within 1.3 copies per reaction, respectively. The real-time RPA method is 10-fold more sensitive than RPA-LFD method. The exact number of CyHV-3 can be calculated in each sample by real-time RPA. The sera from koi also can be tested in these methods. In addition, no cross-reaction was observed with other related pathogens, including carp oedema virus (CEV), spring viraemia of carp virus (SVCV), cyprinid herpesvirus 1(CyHV-1), cyprinid herpesvirus 2(CyHV-2), type I grass carp reovirus (GCRV-I), type II GCRV (GCRV-II), type III GCRV (GCRV-III), and Aeromonas hydrophila.

Cross-reactivity of rPvs48/45, a recombinant Plasmodium vivax protein, with sera from Plasmodium falciparum endemic areas of Africa.

Background: Ps48/45, a Plasmodium gametocyte surface protein, is a promising candidate for malaria transmission-blocking (TB) vaccine. Due to its relevance for a multispecies vaccine, we explored the cross-reactivity and TB activity of a recombinant P. vivax Ps48/45 protein (rPvs48/45) with sera from P. falciparum-exposed African donors. Methods: rPvs48/45 was produced in Chinese hamster ovary cell lines and tested by ELISA for its cross-reactivity with sera from Burkina Faso, Tanzania, Mali, and Nigeria - In addition, BALB/c mice were immunized with the rPvs48/45 protein formulated in Montanide ISA-51 and inoculated with a crude extract of P. falciparum NF-54 gametocytes to evaluate the parasite-boosting effect on rPvs48/45 antibody titers. Specific anti-rPvs48/45 IgG purified from African sera was used to evaluate the ex vivo TB activity on P. falciparum, using standard mosquito membrane feeding assays (SMFA). Results: rPvs48/45 protein showed cross-reactivity with sera of individuals from all four African countries, in proportions ranging from 94% (Tanzania) to 40% (Nigeria). Also, the level of cross-reactive antibodies varied significantly between countries (p<0.0001), with a higher antibody level in Mali and the lowest in Nigeria. In addition, antibody levels were higher in adults (≥ 17 years) than young children (≤ 5 years) in both Mali and Tanzania, with a higher proportion of responders in adults (90%) than in children (61%) (p<0.0001) in Mali, where male (75%) and female (80%) displayed similar antibody responses. Furthermore, immunization of mice with P. falciparum gametocytes boosted anti-Pvs48/45 antibody responses, recognizing P. falciparum gametocytes in indirect immunofluorescence antibody test. Notably, rPvs48/45 affinity-purified African IgG exhibited a TB activity of 61% against P. falciparum in SMFA. Conclusion: African sera (exposed only to P. falciparum) cross-recognized the rPvs48/45 protein. This, together with the functional activity of IgG, warrants further studies for the potential development of a P. vivax and P. falciparum cross-protective TB vaccine.

Cell-Based Assay to Detect the Autoantibody Serostatus in Patients with Neuromyelitis Optica Spectrum Disorder (NMOSD).

Cell-based assay (CBA) is an immunofluorescence assay that is extensively used for the confirmatory diagnosis of inflammatory demyelinating diseases of the central nervous system, like neuromyelitis optica spectrum disorder (NMOSD). Detecting the type of autoantibody present in the sera of the patients is the primary goal. CBA is the most sensitive and recommended detection method among all similar tools. Briefly, serum autoantibody is screened by transfecting specific cells seeded on cover glasses with full-length specific antigen fused with green fluorescent protein (GFP), followed by treating them with the patient serum used here as the source of primary antibody. The autoantibody-treated cells are further labeled with a rhodamine-conjugated secondary antibody. The co-localization of GFP and rhodamine is visualized by confocal microscopy, and the intensity of fluorescence is evaluated to determine the presence of autoantibody. A detailed protocol to screen antibodies against AQP4 and MOG in human sera using this method is described.

Evaluation of species-specific polyclonal antibodies to detect and differentiate between Neospora caninum and Toxoplasma gondii.

Neosporosis and toxoplasmosis are major causes of abortion in livestock worldwide, leading to substantial economic losses. Detection tools are fundamental to the diagnosis and management of those diseases. Current immunohistochemistry (IHC) tests, using sera raised against whole parasite lysates, have not been able to distinguish between Toxoplasma gondii and Neospora caninum. We used T. gondii and N. caninum recombinant proteins, expressed in Escherichia coli and purified using insoluble conditions, to produce specific polyclonal rabbit antisera. We aimed to develop species-specific sera that could be used in IHC on formalin-fixed, paraffin-embedded (FFPE) tissue sections to improve the diagnosis of ruminant abortions caused by protozoa. Two polyclonal rabbit sera, raised against recombinant proteins, anti-Neospora-rNcSRS2 and anti-Toxoplasma-rTgSRS2, had specificity for the parasite they were raised against. We tested the specificity for each polyclonal serum using FFPE tissue sections known to be infected with T. gondii and N. caninum. The anti-Neospora-rNcSRS2 serum labeled specifically only N. caninum-infected tissue blocks, and the anti-Toxoplasma-rTgSRS2 serum was specific to only T. gondii-infected tissues. Moreover, tissues from 52 cattle and 19 sheep previously diagnosed by lesion profiles were tested using IHC with our polyclonal sera and PCR. The overall agreement between IHC and PCR was 90.1% for both polyclonal anti-rNcSRS2 and anti-rTgSRS2 sera. The polyclonal antisera were specific and allowed visual confirmation of protozoan parasites by IHC, but they were not as sensitive as PCR testing.

Prevalence of A2 and A2B Subgroups among Blood Groups A and AB in healthy donors.

Objective: To determine the frequency of A2 and A2B subgroups among blood groups A and AB in healthy donors. Methods: It was a Cross-Sectional study, conducted at the Department of Hematology & Transfusion Medicine, UCHS, The Children's Hospital Lahore and Sundas foundation Lahore from June 2022 to December 2022 including 13,120 healthy blood donors of both genders, after taking informed consent. Venous blood samples of donors were collected in EDTA vials (3ml) and serum gel vial for routine blood grouping which was done by standard tube method. Further testing of donors positive for an antigen (blood Group-A and AB) was performed using anti-A1 lectin by standard tube method as per manufacturer's instruction. The data was analyzed using SPSS version 23. Results: Among 13120 blood donors, 12857 (97.9%) were male and 263 (2.0%) were female with mean age of 36.7 years ± 15.04 years. Majority of them (91.7%) were of Punjabi ethnicity. Donors having blood group phenotype A and AB were 3890 (29.6%). Among blood Group-A donors, A1 was found in 97.8% and A2 in 2.2% donors. While among Blood Group-AB, 96.7% donors belonged to A1B blood group and 3.2% belonged to A2B blood group. Conclusions: Blood group A2 and A2B do exist in blood donors of Punjabi ethnicity. The knowledge of presence of these blood groups' phenotypes in our population can provide a better base for transfusion staff to do troubleshooting in compatibility testing and to avoid any rare but hazardous transfusion outcome.

Precursor carboxy-silica for functionalization with interactive ligands. III. Carbodiimide assisted preparation of immobilized lectin stationary phases for high performance lectin affinity chromatography of sub-glycoproteomics from cancer and disease free human sera.

In this study, a precursor carboxy-silica support was demonstrated in the immobilization of two different lectins, namely concanavalin A (Con A) and wheat germ agglutinin (WGA) for use in high performance lectin affinity chromatography (LAC) for the selective capturing and enrichment of glycoproteins from healthy/disease free and cancer human sera. The lectin columns thus obtained (i.e., Con A- and WGA-columns) showed no nonspecific interactions toward some chosen standard glycoproteins and non-glycoproteins. Both columns were shown in sub-glycoproteomics enrichment from human sera including disease free and adenocarcinoma cancer sera. The collected fractions were subjected to LC-MS/MS for identification of the captured glycoproteins, whereby the total number of identified proteins using Con A column from disease-free and cancer sera were 164 and 188, respectively while 133 and 103 proteins were identified in the fractions captured by the WGA column from disease-free and cancer sera samples, respectively. Differentially expressed proteins (DEPs) between the disease free and cancer sera in both the Con A and WGA column fractions were identified via the plot of the abundance vs. the protein ratio whereby the binary logarithm of average intensities of cancer and disease free sera were plotted against the binary logarithm of cancer/disease free sera ratios. The proteins that exhibit log 2 (cancer/healthy) ratio values greater than +2 and less than -2 in both categories are considered as DEPs. Furthermore, for visualization of the data arrangement, Q-Q scatterplot were also used whereby the binary logarithm of cancer serum was plotted against the binary logarithm of disease-free serum for both Con A and WGA. For Con A column, 28 up-regulated and 10 down regulated proteins were identified with a total of 38 DEPs while only two being non-glycoproteins. Furthermore, the up-regulated, and down regulated proteins recorded for WGA column are 14 and 6, respectively, totaling 20 proteins including 3 non-glycoproteins. Some of the non-specific binding to lectin are most likely due to protein-protein interactions.

Epidemiological surveys revealed the risk of TAMV spill-over from ticks to hosts.

BACKGROUND: Tick-borne viral diseases have become an increasingly important public health concern. Tamdy virus (TAMV) is a tick-borne virus of the genus Orthonairovirus in the family Nairoviridae. While some studies have suggested that TAMV is a pathogen associated with human febrile illness, its epidemiology and the risk of TAMV spill-over remain poorly understood. METHODS: Ticks were collected in Xinjiang, China, and grouped into pools. RT-PCR assays were used to detect TAMV RNA in these pools. The seroprevalence of TAMV was investigated using Immunofluorescence assays, Western blotting, and Luciferase immunoprecipitation system (LIPS) assays. RESULTS: TAMV RNA was detected in 17 out of 363 tick pools, resulting in a minimum infection rate (MIR) of 4.7%. Hyalomma asiaticum and Dermacentor nuttalli were identified as major tick vectors of TAMV. Phylogenetic analysis demonstrated that TAMV strains from Xinjiang are closely related to strains from other countries. Seroprevalence studies showed that TAMV exposure has been occurring in Xinjiang since at least 2006. Antibody responses to TAMV were detected in 1.1% (26/2296) of animals, including domestic animals and wild rodents. The seropositivity rates were as follows: sheep (1.7%), dog (2.3%), Marmota monax (0.8%), Meriones meridianus (3.5%). CONCLUSIONS: The research findings reveal that TAMV can be transmitted by ticks to various animal species, posing a significant public health risk. The wide distribution of TAMV and its tick vectors emphasise the importance of early preparedness and control measures. This study highlights the necessity for maintaining vigilance in addressing emerging zoonotic diseases transmitted by ticks.

Neutralization of Different Variants of SARS-CoV-2 by a F(ab')2 Preparation from Sera of Horses Immunized with the Viral Receptor Binding Domain.

The Receptor Binding Domain (RBD) of SARS-CoV-2, the virus responsible for the COVID-19 pandemic, is the functional region of the viral Spike protein (S), which is involved in cell attachment to target cells. The virus has accumulated progressively mutations in its genome, particularly in the RBD region, many of them associated with immune evasion of the host neutralizing antibodies. Some of the viral lineages derived from this evolution have been classified as Variant of Interest (VOI) or Concern (VOC). The neutralizing capacity of a F(ab')2 preparation from sera of horses immunized with viral RBD was evaluated by lytic plaque reduction assay against different SARS-CoV-2 variants. A F(ab')2 preparation of a hyperimmune serum after nine immunizations with RBD exhibited a high titer of neutralizing antibodies against the ancestral-like strain (1/18,528). A reduction in the titer of the F(ab')2 preparation was observed against the different variants tested compared to the neutralizing activity against the ancestral-like strain. The highest reduction in the neutralization titer was observed for the Omicron VOC (4.7-fold), followed by the Mu VOI (2.6), Delta VOC (1.8-fold), and Gamma VOC (1.5). Even if a progressive reduction in the neutralizing antibodies titer against the different variants evaluated was observed, the serum still exhibited a neutralizing titer against the Mu VOI and the Omicron VOC (1/7113 and 1/3918, respectively), the evaluated strains most resistant to neutralization. Therefore, the preparation retained neutralizing activity against all the strains tested.

ASCA-related antibodies in the blood sera of healthy donors and patients with colorectal cancer: characterization with oligosaccharides related to Saccharomyces cerevisiae mannan.

Mannans are polysaccharide antigens expressed on the cell wall of different fungal species including Saccharomyces cerevisiae and Candida spp. These fungi are components of the normal intestinal microflora, and the presence of antibodies to fungal antigens is known to reflect the features of the patient's immune system. Thus, titers of IgG and IgA antibodies against Saccharomyces cerevisiae mannan (ASCA) are markers for clinical diagnostics of inflammatory bowel diseases. The complex organization and heterogeneity of cell-wall mannans may reduce the quality and reproducibility of ELISA results due to interference by different antigenic epitopes. In this research, we analyzed the levels of IgG antibodies in the sera of healthy donors and patients with colorectal cancer using an array of synthetic oligosaccharides related to distinct fragments of fungal mannan. This study aimed to establish the influence of oligosaccharide structure on their antigenicity. Variations in the structure of the previously established ASCA epitope (changing type of linkage, chain length, and the presence of branches) significantly modified the ability of ligands to bind to circulating antibodies in blood sera. The study showed that surface presentation density of the ligand critically affects the results of enzyme immunoassay. The transition from natural coating antigens to their corresponding synthetic mimetics with a defined structure opens new opportunities for improving existing ELISA test systems, as well as developing diagnostic kits with new properties.

Safety and immunogenicity of BK-SE36/CpG malaria vaccine in healthy Burkinabe adults and children: a phase 1b randomised, controlled, double-blinded, age de-escalation trial.

Background: BK-SE36/CpG is a recombinant blood-stage malaria vaccine candidate based on the N-terminal Plasmodium falciparum serine repeat antigen5 (SE36), adsorbed to aluminium hydroxide gel and reconstituted, prior to administration, with synthetic oligodeoxynucleotides bearing CpG motifs. In healthy Japanese adult males, BK-SE36/CpG was well tolerated. This study assessed its safety and immunogenicity in healthy malaria-exposed African adults and children. Methods: A double-blind, randomised, controlled, age de-escalating clinical trial was conducted in an urban area of Ouagadougou, Burkina Faso. Healthy participants (n=135) aged 21-45 years (Cohort 1), 5-10 years (Cohort 2) and 12-24 months (Cohort 3) were randomised to receive three vaccine doses (Day 0, 28 and 112) of BK-SE36/CpG or rabies vaccine by intramuscular injection. Results: One hundred thirty-four of 135 (99.2%) subjects received all three scheduled vaccine doses. Vaccinations were well tolerated with no related Grade 3 (severe) adverse events (AEs). Pain/limitation of limb movement, headache in adults and fever in younger children (all mild to moderate in intensity) were the most frequently observed local and systemic AEs. Eighty-three of BK-SE36/CpG (91%) recipients and 37 of control subjects (84%) had Grade 1/2 events within 28 days post vaccination. Events considered by the investigator to be vaccine related were experienced by 38% and 14% of subjects in BK-SE36/CpG and control arms, respectively. Throughout the trial, six Grade 3 events (in 4 subjects), not related to vaccination, were recorded in the BK-SE36/CpG arm: 5 events (in 3 subjects) within 28 days of vaccination. All serious adverse events (SAEs) (n=5) were due to severe malaria (52-226 days post vaccination) and not related to vaccination. In all cohorts, BK-SE36/CpG arm had higher antibody titres after Dose 3 than after Dose 2. Younger cohorts had stronger immune responses (12-24-month-old > 5-10 years-old > 21-45 years-old). Sera predominantly reacted to peptides that lie in intrinsically unstructured regions of SE36. In the control arm, there were no marked fold changes in antibody titres and participants' sera reacted poorly to all peptides spanning SE36. Conclusion: BK-SE36/CpG was well-tolerated and immunogenic. These results pave the way for further proof-of-concept studies to demonstrate vaccine efficacy. Clinical trial registration: https://pactr.samrc.ac.za/TrialDisplay.aspx?TrialID=1921, PACTR201701001921166.

Homotypic antibodies target novel E glycoprotein domains after natural DENV 3 infection/vaccination.

The envelope (E) glycoprotein is the primary target of type-specific (TS) neutralizing antibodies (nAbs) after infection with any of the four distinct dengue virus serotypes (DENV1-4). nAbs can be elicited to distinct structural E domains (EDs) I, II, or III. However, the relative contribution of these domain-specific antibodies is unclear. To identify the primary DENV3 nAb targets in sera after natural infection or vaccination, chimeric DENV1 recombinant encoding DENV3 EDI, EDII, or EDIII were generated. DENV3 EDII is the principal target of TS polyclonal nAb responses and encodes two or more neutralizing epitopes. In contrast, some were individuals vaccinated with a DENV3 monovalent vaccine-elicited serum TS nAbs targeting each ED in a subject-dependent fashion, with an emphasis on EDI and EDIII. Vaccine responses were also sensitive to DENV3 genotypic variation. This DENV1/3 panel allows the measurement of serum ED TS nAbs, revealing differences in TS nAb immunity after natural infection or vaccination.

Biodistribution of Fluorescent Albumin Nanoparticles among Organs of Laboratory Animals after Intranasal and Peroral Administration.

Natural, environmental and engineered nanoparticles (NP) penetrate into cells by endocytosis and induce innate immunity. The behaviour of the nanomaterials both in vitro and in vivo should be assessed. Our goal was to study protein NP stability in biological fluids and distribution in organs of animals after intranasal and oral administration. Bovine serum albumin (BSA) was labelled with the fluorescent dye RhoB and NP were fabricated by nanoprecipitation. The fluorescent protein NPwere administered intranasally and orally in laboratory-outbred mice ICR and rabbits. RhoB-BSA NP distribution in organs was detected using spectrofluorometry and fluorescent microscopy. Innate immunity was evaluated using reverse transcription with random hexanucleotide primer and subsequent real-time PCR with specific fluorescent hydrolysis probes. The labelled BSA NP were shown to remain stable in blood sera and nasopharyngeal swabs for 5 days at +37 °C. In vivo the maximal accumulation was found in the brain in 2 days posttreatment without prevalent accumulation in olfactory bulbs. For the intestine, heart and liver, the BSA NP accumulation was similar in 1 and 2 days, whereas for kidney samples even decreased after 1 day. Both intranasal and peroral administration of RhoB-BSA NP did not induce innate immunity. Thus, after intranasal or oral instillation RhoB-BSA NP were found mainly in the brain and intestine without interferon gene expression.

Water-soluble biopolymers from heat-treated and high pressure homogenized vegetable purées: investigating their emulsion forming and stabilizing capacities.

The emulsion forming and stabilizing capacities of water-soluble biopolymers originating from the aqueous (serum) phase of heat-treated and high pressure homogenized purées were investigated. The serum biopolymers were characterized and then utilized as emulsifier/stabilizer in simple oil-in-water emulsions. The resulting emulsions were stored at 4 °C and monitored for 2 weeks. Results revealed that carrot and tomato sera contained higher amounts of pectin and lower protein compared to broccoli. The serum pectic biopolymers exhibited distinct molecular structures, depending on the vegetable origin. Given these natural biopolymer composition and characteristics, emulsions with small droplet sizes were observed at pH 3.5. However, emulsions at pH 6.0 showed large mean droplet sizes, except for the emulsion formulated with carrot serum. Regardless of the pH, emulsions containing carrot serum biopolymers exhibited high capacity to form fine emulsions that were stable during the 2-week storage period at low temperature. This study clearly shows the capacity of natural water-soluble biopolymers isolated from the serum phase of vegetable purées to form fine emulsion droplets and maintain its stability during storage, especially in the case of carrot serum biopolymers.

An accurate DFT study within conformational survey of the D-form serine-alanine protected dipeptide.

The conformational analysis of N-formyl-D-serine-D-alanine-NH2 dipeptide was studied using density functional theory methods at B3LYP, B3LYP‒D3, and M06‒2X levels using 6‒311 + G (d,p) basis set in the gas and water phases. 87 conformers of 243 stable ones were located and the rest of them were migrated to the more stable geometries. Migration pattern suggests the more stable dipeptide model bears serine in ßL, γD, γL and the alanine in γL and Î³D configurations. The investigation of side‒chain‒backbone interactions revealed that the most stable conformer, γD-γL, is in the ß‒turn region of Ramachandran map; therefore, serine-alanine dipeptide model should be adopted with a ß‒turn conformation. Intramolecular hydrogen bonding in ß‒turns consideration by QTAIM disclosed γD-γL includes three hydrogen bonds. The computed UV‒Vis spectrum alongside of NBO calculation showed the five main electronic transition bands derived of n → n* of intra‒ligand alanine moiety of dipeptide structure.

A novel Galleria mellonella experimental model for zoonotic pathogen Brucella.

Brucellosis is a major threat to public health and animal husbandry. Several in vivo vertebrate models, such as mice, guinea pigs, and nonhuman primates, have been used to study Brucella pathogenesis, bacteria-host interactions, and vaccine efficacy. However, these models have limitations whereas the invertebrate Galleria mellonella model is a cost-effective and ethical alternative. The aim of the present study was to examine the invertebrate G. mellonella as an in vivo infection model for Brucella. Infection assays were employed to validate the fitness of the larval model for Brucella infection and virulence evaluation. The protective efficacy of immune sera was evaluated by pre-incubated with a lethal dose of bacteria before infection. The consistency between the mouse model and the larval model was confirmed by assessing the protective efficacy of two Brucella vaccine strains. The results show that G. mellonella could be infected by Brucella strains, in a dose- and temperature-dependent way. Moreover, this larval model can effectively evaluate the virulence of Brucella strains in a manner consistent with that of mammalian infection models. Importantly, this model can assess the protective efficacy of vaccine immune sera within a day. Further investigation implied that haemolymph played a crucial role in the protective efficacy of immune sera. In conclusion, G. mellonella could serve as a quick, efficient, and reliable model for evaluating the virulence of Brucella strains and efficacy of immune sera in an ethical manner.