Probiotic Potential, Antiproliferative, and Proapoptotic Activities of Lactobacillus fermentum Isolated from Breast Milk.

Background: Breast milk is always the best choice for infant's nutrition due to its useful compounds such as immune cells and molecules, oligosaccharides, as well as bacteria and their metabolites. We identified and characterized the isolated strain from human breast milk in this study. Materials and Methods: A total of 20 lactating mothers aged 25 to 34 years were enrolled in our study. We collected the breast milk samples in sterile microtubes. 100 µl of each sample was spread on de Man-Rogosa-Sharpe (MRS) agar plates and incubated aerobically at 37°C for 48 hr. After identifying the isolated strain, initial tests (hemolysis inactivity and L-arginine hydrolysis, catalase), the acid tolerance, bile tolerance, and antibiotics susceptibility of the isolated strain were estimated. Furthermore, the antiproliferative and proapoptotic activities of heat-killed cells) HKC) and cell-free supernatant (CFS) of the strain on the HT-29 cell line were evaluated using MTT assay and flow cytometry analysis, respectively. Results: The isolated strain was Gram-positive, bacilli in shape, catalase-negative, non-hemolytic, and negative for L-arginine hydrolysis. By 16S rRNA gene sequencing, the isolated strain was Lactobacillus fermentum. According to MTT assay and flow cytometry results, the HKC and CFS of the isolated strain reduced the viability of the HT-29 cells. The total apoptosis induced in HT-29 cells by HKC and CFS was 65.98% and 70.1%, respectively. Conclusion: Our findings suggest that this strain, despite the properties of probiotic bacteria, has potential antiproliferative and proapoptotic capabilities.

Molecular and Antigen Detection, Phylogenetics, and Immunoinformatics Study of the Zoonotic Coronavirus in Iranian Diarrheic Calves.

Background: Bovine coronaviruses (BCoVs) are zoonotic diseases that result in substantial economic losses due to mortality, impaired growth, and increased medication expenses in large animals. These viruses pose a risk to children who live beside infected animal, as they can cause diarrhea. This study was dedicated to molecular and antigen detections and phylogenetic and immunoinformatics analysis of zoonotic coronavirus (CoV) in Iran. Materials and Methods: A total of 77 diarrheic samples were collected from Holstein dairy herds in selected provinces of Iran. Samples were tested by capture antibody enzyme-linked immunosorbent assay (ELISA) to detect CoV and reverse transcriptase-polymerase chain reaction (RT-PCR) for verification of detection and also genotyping of spike glycoprotein in CoV-positive samples. After statistical analysis, nucleotide sequence alignment, and nucleotide and protein phylogenetic tree construction, the centralized sequence for vaccine strains was obtained using computationally optimized broadly reactive antigen (COBRA)'s center-of-the-tree (COT) method. Results: Twenty-two (28.5%) and eight (10.3%) of 77 samples were positive according to RT-PCR and ELISA, respectively. (Basic Local Alignment Search Tool) BLAST and phylogenetic analysis revealed that most similar sequences to the Iranian CoV sequence were for European countries. Furthermore, there were strong correlations to other CoVs in humans and wild and domesticated animals. As CoV has variable COT, the most recent strains and COBRA vaccine strains were obtained. Conclusion: Based on the high prevalence of this viral disease in calves and its economic impact on the breeding industry, as well as the potential transmission to humans and correlation with World Health Organization (WHO) One Health approach guidelines, the study emphasizes the importance of implementing preventive strategies such as animal vaccination.

Comparative Analysis and Identification of Spike Mutations in Iranian COVID-19 Samples from the First Three Waves of Disease.

Background: The spike surface glycoprotein of SARS-CoV-2 is the essential protein in virus attachment to the target cell and cell entrance. As this protein contains immunodominant epitopes and is the main target for immune recognition, it is the critical target for vaccine and therapeutics development. In the current research, we analyzed the variability and mutations of the spike glycoprotein isolated from 72 COVID-19-positive patients from Iran's first three waves of disease. Materials and Methods: The RNA was extracted from nasopharyngeal samples of confirmed COVID-19 cases and served as a template for cDNA synthesis and reverse transcriptase polymerase chain reaction. The reverse transcriptase polymerase chain reaction products of each sample were assembled and sequenced. Results: After analysis of 72 sequences, we obtained 46 single nucleotide polymorphisms, including 23 that produce amino acid changes. Our analysis showed that the most frequent mutation was the D614G (in the samples of the second and third waves). Conclusions: Our findings suggest that developing effective vaccines requires identifying the predominant variants of SARS-CoV-2 in each community.

The effect of mutation on neurotoxicity reduction of new chimeric reteplase, a computational study.

Background and purpose: Excitotoxicity in nerve cells is a type of neurotoxicity in which excessive stimulation of receptors (such as N-methyl-d-aspartate glutamate receptors (NMDAR)) leads to the influx of high-level calcium ions into cells and finally cell damage or death. This complication can occur after taking some of the plasminogen activators like tissue plasminogen activator and reteplase. The interaction of the kringle2 domain in such plasminogen activator with the amino-terminal domain (ATD) of the NR1 subunit of NMDAR finally leads to excitotoxicity. In this study, we assessed the interaction of two new chimeric reteplase, mutated in the kringle2 domain, with ATD and compared the interaction of wild-type reteplase with ATD, computationally. Experimental approach: Homology modeling, protein docking, molecular dynamic simulation, and molecular dynamics trajectory analysis were used for the assessment of this interaction. Findings/Results: The results of the free energy analysis between reteplase and ATD (wild reteplase: -2127.516 ± 0.0, M1-chr: -1761.510 ± 0.0, M2-chr: -521.908 ± 0.0) showed lower interaction of this chimeric reteplase with ATD compared to the wild type. Conclusion and implications: The decreased interaction between two chimeric reteplase and ATD of NR1 subunit in NMDAR which leads to lower neurotoxicity related to these drugs, can be the start of a way to conduct more tests and if the results confirm this feature, they can be considered potential drugs in acute ischemic stroke treatment.

A Process Evaluation of the Isfahan Antibiotic Awareness Campaign: Developing Engagement on Antimicrobial Resistance.

Background: One of the most prominent global health threats is antibiotic resistance, leading to infection treatment failure. The first Iranian antibiotic awareness week campaign was initiated to improve the prudent use and wise prescription of antibiotics. Materials and Methods: The Isfahan antibiotic awareness campaign was held from November 30 to December 6, 2019, among two targeted populations; the general population and health-care workers by Isfahan University of Medical Sciences. In this campaign held in the main squares, streets, and a city's referral hospital, various educational methods were used to aware and sensitize the general population and medical staff about antibiotics and microbial resistance. These methods include face-to-face training, brochures, advertisement posters and billboards around the city, educational videos, social media messages, retraining for medical doctors and medical specialists, and interviewing in the Islamic Republic of Iran Broadcast. Results: Two hundred and twenty general practitioners, medical specialists, and residents participated in two retraining educational conferences in Al-Zahra Hospital, Isfahan, Iran. The mean score satisfaction of the two conferences was three from four. Nearly 2000 of the general population were under face-to-face educational programs whom after that, 83.6% had the correct answer to the questions around antimicrobial awareness. Conclusions: This campaign was an excellent experience as a pilot study with appealing issues. Further, activities are required to improve engagement with the target population and determine the impact of this campaign on antibiotic consumption and prescription behavior among the public and health-care professionals.

Association of Clinical Features with Spike Glycoprotein Mutations in Iranian COVID-19 Patients.

BACKGROUND: Mutations in spike glycoprotein, a critical protein of SARS-CoV-2, could directly impact pathogenicity and virulence. The D614G mutation, a non-synonymous mutation at position 614 of the spike glycoprotein, is a predominant variant circulating worldwide. This study investigated the occurrence of mutations in the crucial zone of the spike gene and the association of clinical symptoms with spike mutations in isolated viruses from Iranian patients infected with SARS-CoV-2 during the second and third waves of the COVID-19 epidemic in Isfahan, the third-largest city in Iran. METHODS: The extracted RNA from 60 nasopharyngeal samples of COVID-19 patients were subjected to cDNA synthesis and RT-PCR (in three overlapping fragments). Each patient's reverse transcriptase polymerase chain reaction (RT-PCR) products were assembled and sequenced. Information and clinical features of all sixty patients were collected, summarized, and analyzed using the GENMOD procedure of SAS 9.4. RESULTS: Analysis of 60 assembled sequences identified nine nonsynonymous mutations. The D614G mutation has the highest frequency among the amino acid changes. In our study, in 31 patients (51.66%), D614G mutation was determined. For all the studied symptoms, no significant relationship was observed with the incidence of D614G mutation. CONCLUSIONS: D614G, a common mutation among several of the variants of SARS-CoV-2, had the highest frequency among the studied sequences and its frequency increased significantly in the samples of the third wave compared to the samples of the second wave of the disease.

Herbal Medicines as Potential Inhibitors of SARS-CoV-2 Infection.

Coronavirus disease 2019 (COVID-19) is the result of infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Understanding molecular pathogenesis is an essential factor for the allocation of effective preventive measures and the development of targeted therapeutics against COVID-19. The genome of SARS-CoV-2 encodes structural and nonstructural proteins, which can be targets for compounds with potential therapeutic ability. On the other hand, the virus life cycle has stages susceptible to targeting by drug compounds. Many natural antiviral compounds have been studied and evaluated at the cellular and molecular levels with antiviral potential. Meanwhile, many studies over the past few months have shown that plant polysaccharides have a good ability to target proteins and stages of the virus life cycle. In this regard, in this review study, the virus specifications and infectious process and structural and functional components of SARSCoV- 2 will be reviewed, and then the latest studies on the effect of plant compounds with more focus on polysaccharides on viral targets and their inhibitory potential on the infectious process of COVID-19 will be discussed.

Gene Editing-Based Technologies for Beta-hemoglobinopathies Treatment.

Beta (ß)-thalassemia is a group of human inherited abnormalities caused by various molecular defects, which involves a decrease or cessation in the balanced synthesis of the ß-globin chains in hemoglobin structure. Traditional treatment for ß-thalassemia major is allogeneic bone marrow transplantation (BMT) from a completely matched donor. The limited number of human leukocyte antigen (HLA)-matched donors, long-term use of immunosuppressive regimen and higher risk of immunological complications have limited the application of this therapeutic approach. Furthermore, despite improvements in transfusion practices and chelation treatment, many lingering challenges have encouraged researchers to develop newer therapeutic strategies such as nanomedicine and gene editing. One of the most powerful arms of genetic manipulation is gene editing tools, including transcription activator-like effector nucleases, zinc-finger nucleases, and clustered regularly interspaced short palindromic repeat-Cas-associated nucleases. These tools have concentrated on γ- or ß-globin addition, regulating the transcription factors involved in expression of endogenous γ-globin such as KLF1, silencing of γ-globin inhibitors including BCL11A, SOX6, and LRF/ZBTB7A, and gene repair strategies. In this review article, we present a systematic overview of the appliances of gene editing tools for ß-thalassemia treatment and paving the way for patients' therapy.

Vaccine design and delivery approaches for COVID-19.

COVID-19 is still a deadly disease that remains yet a major challenge for humans. In recent times, many large pharmaceutical and non-pharmaceutical companies have invested a lot of time and cost in fighting this disease. In this regard, today's scientific knowledge shows that designing and producing an effective vaccine is the best possible way to diminish the disease burden and dissemination or even eradicate the disease. Due to the urgent need, many vaccines are now available earlier than scheduled. New technologies have also helped to produce much more effective vaccines, although the potential side effects must be taken into account. Thus, in this review, the types of vaccines and vaccine designs made against COVID-19, the vaccination programs, as well as the delivery methods and molecules that have been used to deliver some vaccines that need a carrier will be described.

Acetate Kinase a Antisense Delivery by PAMAM Dendrimer for Decreasing Acetate Production and Increasing the Production of Recombinant Albumin in E. coli.

BACKGROUND: Acetate accumulation in the culture medium is known as an inhibitor in recombinant protein production in Escherichia coli. Various approaches have been proposed and evaluated to overcome this challenge and reduce the concentration of acetate. In this study, we examined the effect of acetate kinase A antisense on acetate production rate in E. coli We also used PAMAM dendrimers as a suitable delivery agent for antisense transformation into E. coli host cell. OBJECTIVE: This study aimed to decrease acetate production as a by-product using an antisense-dendrimer complex to increase mass cell and subsequently recombinant Albumin production in E. coli. MATERIALS AND METHODS: Here, to study the effect of this treatment on recombinant protein production, we used pET22b/HAS construct. The ackA gene expression was inhibited by designed antisense to reduce acetate concentration in culture medium. AckA antisense was transferred to E. coli by PAMAM dendrimer. Finally, ackA expression and recombinant Albumin production were evaluated Real-Time PCR and densitometry, respectively. RESULTS: Our data showed, designed antisense lead to reduction of acetate kinase gene expression and subsequently acetate concentration in the culture medium. Finally, acetate concentration reduction and cell mass increase result in enhanced recombinant Alb production in the treated group (1.25 mg.mL-1) compare to the control group ( 0.59 mg.mL-1). CONCLUSIONS: Reduction of acetate in E. coli fermentation process decreased the recombinant Alb production following cell growth and cell mass increase. In the current study, we showed that an antisense can be a useful tool for ackA gene expression reduction. Also, we noted that PAMAM dendrimer could be a proper delivery agent for oligonucleotide antisense transformation into bacterial cells.

Novel and emerging mutations of SARS-CoV-2: Biomedical implications.

Coronavirus disease-19 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The SARS-CoV-2 virus strains has geographical diversity associated with diverse severity, mortality rate, and response to treatment that were characterized using phylogenetic network analysis of SARS-CoV-2 genomes. Although, there is no explicit and integrative explanation for these variations, the genetic arrangement, and stability of SARS-CoV-2 are basic contributing factors to its virulence and pathogenesis. Hence, understanding these features can be used to predict the future transmission dynamics of SARS-CoV-2 infection, drug development, and vaccine. In this review, we discuss the most recent findings on the mutations in the SARS-CoV-2, which provide valuable information on the genetic diversity of SARS-CoV-2, especially for DNA-based diagnosis, antivirals, and vaccine development for COVID-19.

Recent developments in antibody derivatives against colorectal cancer; A review.

Colorectal cancer (CRC) is the fourth most common cause of cancer and mortality worldwide and is the third most common cancer in men and women. Surgery, radiotherapy, and chemotherapy are conventionally used for the treatment of colorectal cancer. However, these methods are associated with various side effects on normal cells. Thus, new studies are moving towards more effective and non-invasive methods for treatment of colorectal cancer. Targeted therapy of CRC is a promising new approach to enhance the efficiency and decrease the toxicity of the treatment. In targeted therapy of CRC, antibody fragments can directly inhibit tumor cell growth and proliferation. They also can act as an ideal carrier for targeted delivery of anticancer drugs. In the present study, the structure and function of different formats of antibody fragments, immune-targeted therapy of CRC using antibody fragments will be discussed.

Current status of COVID-19 pandemic; characteristics, diagnosis, prevention, and treatment.

Humans have always been encountered to big infectious diseases outbreak throughout the history. In December 2019, novel coronavirus (COVID-19) was first noticed as an agent causing insidious pneumonia in Wuhan, China. COVID-19 was spread rapidly from Wuhan to the rest of the world. Until late June 2020, it infected more than 10,000,000 people and caused more than 500,000 deaths in almost all of countries in the world, creating a global crisis worse than all previous epidemics and pandemics. In the current review, we gathered and summarized the results of various studies on characteristics, diagnosis, treatment, and prevention of this pandemic crisis.

Bioinformatics prediction and experimental validation of VH antibody fragment interacting with Neisseria meningitidis factor H binding protein.

OBJECTIVES: We previously conducted an in silico research on the interactions between the ribosome display-selected single chain variable fragment (scFv) and factor H binding protein (fHbp) of Neisseria meningitidis. We found that heavy chain variable (VH) fragment of this scFv had considerable affinity to fHbp. These results led us to evaluate the ability of this small antibody fragment in binding and detection of fHbp antigen. MATERIALS AND METHODS: In this study, at first, the three-dimensional structure of VH fragment was simulated by Kotai Antibody Builder web server. By using ClusPro 2.0 web server, the 3D structure of the soluble form of fHbp (PDB: 2KC0) was docked to the modeled VH fragment to extract the structure of the complex's binding. Molecular dynamics (MD) simulation was carried out using GROMACS 4.5.3 package for 65 ns. Secondly, coding sequence of VH fragment was cloned separately and expressed in Escherichia coli. After purification of the VH fragment, its binding activity to fHbp protein was analyzed by enzyme-linked immunosorbent assay (ELISA) and surface plasmon resonance (SPR) method. RESULTS: Important amino acids involved in antigen- antibody interaction were identified by analyzing the fHbp-VH complex. The ability of the VH antibody fragment to bind and detect fHbp antigen has been confirmed by the results of in silico analysis, ELISA and SPR methods. CONCLUSION: These results showed that this small fragment of antibody could be used for designing diagnostic kits.

An update on antiviral antibody-based biopharmaceuticals.

Due to the vastness of the science virology, it is no longer an offshoot solely of the microbiology. Viruses have become as the causative agents of major epidemics throughout history. Many therapeutic strategies have been used for these microorganisms, and in this way the recognizing of potential targets of viruses is of particular importance for success. For decades, antibodies and antibody fragments have occupied a significant body of the treatment approaches against infectious diseases. Because of their high affinity, they can be designed and engineered against a variety of purposes, mainly since antibody fragments such as scFv, nanobody, diabody, and bispecific antibody have emerged owing to their small size and interesting properties. In this review, we have discussed the antibody discovery and molecular and biological design of antibody fragments as inspiring therapeutic and diagnostic agents against viral targets.

Bicyclic peptides: types, synthesis and applications.

Bicyclic peptides form one of the most promising platforms for drug development owing to their biocompatibility, similarity and chemical diversity to proteins, and they are considered as a possible practical tool in various therapeutic and diagnostic applications. Bicyclic peptides are known to have the capability of being employed as an effective alternative to complex molecules, such as antibodies, or small molecules. This review provides a summary of the recent progress on the types, synthesis and applications of bicyclic peptides. More specifically, natural and synthetic bicyclic peptides are introduced with their different production methods and relevant applications, including drug targeting, imaging and diagnosis. Their uses as antimicrobial agents, as well as the therapeutic functions of different bicyclic peptides, are also discussed.

Antibody-drug therapeutic conjugates: Potential of antibody-siRNAs in cancer therapy.

Codelivery is a promising strategy of targeted delivery of cytotoxic drugs for eradicating tumor cells. This rapidly growing method of drug delivery uses a conjugate containing drug linked to a smart carrier. Both two parts usually have therapeutic properties on the tumor cells. Monoclonal antibodies and their derivatives, such as Fab, scFv, and bsAb due to targeting high potent have now been attractive candidates as drug targeting carrier systems. The success of some therapeutic agents like small interfering RNA (siRNA), a small noncoding RNAs, with having problems such as enzymatic degradation and rapid renal filtration need to an appropriate carrier. Therefore, the aim of this study is to review the recent enhancements in development of antibody drug conjugates (ADCs), especially antibody-siRNA conjugates (SRCs), its characterizations and mechanisms in innovative cancer therapy approaches.

Targeted cancer therapy through antibody fragments-decorated nanomedicines.

Active targeting in cancer nanomedicine, for improved delivery of agents and diagnose, has been reviewed as a successful way for facilitating active uptake of theranostic agents by the tumor cells. The application of a targeting moiety in the targeted carrier complexes can play an important role in differentiating between tumor and healthy tissues. The pharmaceutical carriers, as main part of complexes, can be polymeric nanoparticles, micelles, liposomes, nanogels and carbon nanotubes. The antibodies are among the natural ligands with highest affinity and specificity to target pharmaceutical nanoparticle conjugates. However, the limitations, such as size and long circulating half-lives, hinder reproducible manufacture in clinical studies. Therefore, novel approaches have moved towards minimizing and engineering conventional antibodies as fragments like scFv, Fab, nanobody, bispecific antibody, bifunctional antibody, diabody and minibody preserving their functional potential. Different formats of antibody fragments have been reviewed in this literature update, in terms of structure and function, as smart ligands in cancer diagnosis and therapy of tumor cells.