Functional attributes of bioactive peptides of bovine milk origin and application of in silico approaches for peptide prediction and functional annotations.

Bovine milk peptides are the protein fragments with diverse bioactive properties having antioxidant, anticarcinogenic, other therapeutic and nutraceutical potentials. These peptides are formed in milk by enzymatic hydrolysis, gastrointestinal digestion and fermentation processes. They have significant health impact with high potency and low toxicity making them a suitable natural alternative for preventing and managing diseases. Antibiotic resistance has increased the quest for better peptide candidates with antimicrobial effects. This article presents a comprehensive review on well documented antimicrobial, immunological, opioid, and anti-hypertensive activities of bovine milk peptides. It also covers the usage of computational biology tools and databases for prediction and analysis of the food-derived bioactive peptides. In silico analysis of amino acid sequences of Bos taurus milk proteins have been predicted to generate peptides with dipeptidyl peptidase IV inhibitory and ACE inhibitory properties, making them favorable candidates for developing blood sugar lowering drugs and anti-hypertensives. In addition to the prediction of new bioactive peptides, application of bioinformatics tools to predict novel functions of already known peptides is also discussed. Overall, this review focuses on the reported as well as predicted biologically active peptide of casein and whey proteins of bovine milk that can be utilized to develop therapeutic agents.

Systematically Optimized Imiquimod-Loaded Novel Hybrid Vesicles by Employing Design of Experiment (DoE) Approach with Improved Biocompatibility, Stability, and Dermatokinetic Profile.

The present research work explored the possibility of harnessing the benefits of vesicular carriers for overcoming imiquimod-associated complaints or side effects. Hybrid vesicles were prepared by the most common and easily scalable method, i.e., thin film hydration. The chaffing of myriad of factors, both process and material related, affecting the desirable attributes of conceived vesicles, was performed through Taguchi design. Based upon the analysis of Pareto chart and prior experiences, concentration of phospholipid and poloxamer 407 was selected for optimization by 2 levels, 13 run central composite design (CCD). The optimized hybrid vesicles contained 1% w/v phospholipid and 3% w/v poloxamer 407. The optimized hybrid vesicles were incorporated into the 3% w/v carbopol 940 gel and characterized for morphology, physicochemical properties, and rheological behavior. The release (%) and skin retention (% of total dose) across rat skin from gel at same amount of formulation was more than Imiquad®. The gel delivered the loaded cargo, preferably, in the viable region of skin and formed local depot in confocal microscopic studies. The gel followed one compartment open body dermatokinetic model in rat skin. There was not any harmful effect on the mice skin after repeated applications. The gel was stable at room conditions for 1 year.

Fmoc-phenylalanine displays antibacterial activity against Gram-positive bacteria in gel and solution phases.

In the quest for new antimicrobial materials, hydrogels of Fmoc-protected peptides and amino acids have gained momentum due to their ease of synthesis and cost effectiveness; however, their repertoire is currently limited, and the mechanistic details of their function are not well understood. Herein, we report the antibacterial activity of the hydrogel and solution phases of Fmoc-phenylalanine (Fmoc-F) against a variety of Gram-positive bacteria including methicillin-resistant Staphylococcus aureus (MRSA). Fmoc-F, a small molecule hydrogelator, reduces the bacterial load both in vitro and in the skin wound infections of mice. The antibacterial activity of Fmoc-F is predominantly due to its release from the hydrogel. Fmoc-F shows surfactant-like properties with critical micelle concentration nearly equivalent to its minimum bactericidal concentration. Similar to Fmoc-F, some Fmoc-conjugated amino acids (Fmoc-AA) have also shown antibacterial effects that are linearly correlated with their surfactant properties. At low concentrations, where Fmoc-F does not form micelles, it inhibits bacterial growth by entering the cell and reducing the glutathione levels. However, at higher concentrations, Fmoc-F triggers oxidative and osmotic stress and, alters the membrane permeabilization and integrity, which kills Gram-positive bacteria. Herein, we proposed the use of the Fmoc-F hydrogel and its solution for several biomedical applications. This study will open up new avenues to enhance the repertoire of Fmoc-AA to act as antimicrobial agents and improve their structure-activity relationship.

Functional testing of a PF02458 homologue of putative rice arabinoxylan feruloyl transferase genes in Brachypodium distachyon.

MAIN CONCLUSION: We show that changing the expression of a putative feruloyl transferase gene belonging to the BAHD acyl-transferase family alters the levels of cell wall esterified ferulates and diferulates in Brachypodium distachyon cell walls. While the potential of grass cell walls for biofuel production has been realized, the technology for lignocellulosic biomass conversion for the production of ethanol is still inefficient because of structural mechanisms that plants have evolved to make the cell wall recalcitrant to enzymatic attack. One of these mechanisms in grasses involves the esterification of arabinoxylans in the cell wall with ferulic acid via an ester linkage to arabinose side chains on xylans. These ferulates undergo oxidative coupling reactions to form ferulate dimers, thus crosslinking polysaccharides. Arabinoxylan feruloylation is an important factor that determines cell wall recalcitrance because it directly cross-links xylans and because ferulates act as nucleating sites for the formation of lignin and for the linkage of lignin to the xylan/cellulose network. Here we report on the effects of changing the expression of Bradi2g43520 (BdAT1), a homologue of the rice feruloyl transferase gene Os01g42880 belonging to the Pfam PF02458 family, in Brachypodium distachyon. Down regulation in several independent RNAi::BdAT1 lines, resulted in up to a 35 % reduction of ferulate levels in both leaves and stems compared to control plants, over 2-3 generations of selfing. In contrast, overexpression of putative BdAT1 resulted in an increase of up to 58 and 47 % of ferulate levels in leaves and stems, respectively, compared to control plants and analyzed over 2-3 generations of selfing. These findings suggest that Bradi2g43520 may be a good candidate for feruloylation of AX in Brachypodium.

The relationship between capsular type and OmpA of Pasteurella multocida is associated with the outcome of disease.

The genes encoding OmpA of Pasteurella multocida recovered from diseased and apparently healthy animals have been characterized. The nucleotide sequence revealed ORFs of 1047-1077 bp encoding proteins of 349-360 amino acids. Domain analysis of OmpA showed signal peptide, N-terminal ompA domain and C-terminal ligand binding domain. The transmembrane topology of OmpA showed short turns at the periplasmic end and longer irregular loops at the extracellular end. The phylogenetic analysis based on OmpA showed affiliation of isolates to 7 groups representing different alleles. The identical segments in OmpA also suggested assortative recombination within classes IV, V and VI of distinct lineages. Principal component analysis separated isolates into groups based on capsular type and PmompA alleles. The alleles belonging to class VI exclusively associated with capsular type A, whereas class I-IV were associated with capsular type B. PmompA alleles in class V were recorded in both serogroups. PmompA6.1, 6.4 were distributed among strains with capsular type A, and PmompA6.2 and 6.3 among capsular type B. Despite internal OmpA variabilty, restrictive and well defined distribution was seen amongst P. multocida. A definitive association of "OmpA-capsular type" was observed with clinical status of animals. A cohort of pasteurellae comprising of OmpA(I-IV)-capB was recovered from diseased animals and OmpA(VI)-capA from healthy subjects. This study concludes that P. multocida with serogroup A and B from healthy and diseased animals represent distinct clusters also differentiated based on their OmpA-types and OmpA-capsular type relationship possibly determine the virulence and disease outcome.

An Unusual Cause of Fever, Bilateral Ear Discharge, Lung Nodules, and Polyuria in a Young Male Post Coronavirus Disease 19.

We present a case of a young male with pyrexia of unknown origin, bilateral ear discharge, lung nodules and polyuria within four weeks of recovering from a moderate Coronavirus disease-19 infection. To the best of our knowledge, this is the first case of post- Coronavirus disease-19 Granulomatosis with Polyangiitis with overlapping large vessel vasculitis with a favorable outcome.

Expression of flavonoid 3'-hydroxylase is controlled by P1, the regulator of 3-deoxyflavonoid biosynthesis in maize.

BACKGROUND: The maize (Zea mays) red aleurone1 (pr1) encodes a CYP450-dependent flavonoid 3'-hydroxylase (ZmF3'H1) required for the biosynthesis of purple and red anthocyanin pigments. We previously showed that Zmf3'h1 is regulated by C1 (Colorless1) and R1 (Red1) transcription factors. The current study demonstrates that, in addition to its role in anthocyanin biosynthesis, the Zmf3'h1 gene also participates in the biosynthesis of 3-deoxyflavonoids and phlobaphenes that accumulate in maize pericarps, cob glumes, and silks. Biosynthesis of 3-deoxyflavonoids is regulated by P1 (Pericarp color1) and is independent from the action of C1 and R1 transcription factors. RESULTS: In maize, apiforol and luteoforol are the precursors of condensed phlobaphenes. Maize lines with functional alleles of pr1 and p1 (Pr1;P1) accumulate luteoforol, while null pr1 lines with a functional or non-functional p1 allele (pr1;P1 or pr1;p1) accumulate apiforol. Apiforol lacks a hydroxyl group at the 3'-position of the flavylium B-ring, while luteoforol has this hydroxyl group. Our biochemical analysis of accumulated compounds in different pr1 genotypes showed that the pr1 encoded ZmF3'H1 has a role in the conversion of mono-hydroxylated to bi-hydroxylated compounds in the B-ring. Steady state RNA analyses demonstrated that Zmf3'h1 mRNA accumulation requires a functional p1 allele. Using a combination of EMSA and ChIP experiments, we established that the Zmf3'h1 gene is a direct target of P1. Highlighting the significance of the Zmf3'h1 gene for resistance against biotic stress, we also show here that the p1 controlled 3-deoxyanthocyanidin and C-glycosyl flavone (maysin) defence compounds accumulate at significantly higher levels in Pr1 silks as compared to pr1 silks. By virtue of increased maysin synthesis in Pr1 plants, corn ear worm larvae fed on Pr1; P1 silks showed slower growth as compared to pr1; P1 silks. CONCLUSIONS: Our results show that the Zmf3'h1 gene participates in the biosynthesis of phlobaphenes and agronomically important 3-deoxyflavonoid compounds under the regulatory control of P1.

Detection of Mycobacterium bovis and Mycobacterium tuberculosis from Cattle: Possible Public Health Relevance.

Mycobacterium bovis and Mycobacterium tuberculosis infect both animals and humans. The disease epidemiology by these agents differs in developed and developing countries due to the differences in the implementation of the prevention and control strategies. The present study describes the detection of M. bovis and M. tuberculosis from specimens of lungs and pulmonary lymph nodes of four cattle died in an organized herd of 183 cattle in the state of Himachal Pradesh, India, with inconclusive skin test results. Identification and distinction of these closely related mycobacterial species was done by PCR-RFLP targeting hsp65 gene followed by spacer oligonucleotide typing. Mixed infection of M. bovis and M. tuberculosis was detected in one cattle.

Investigation of an outbreak of sheeppox among native sheep breeds in the Western Himalayas of India.

An outbreak of sheeppox was investigated in a cluster of villages situated in Western Himalayan ranges of a Northern Indian state. Non-migratory sheep (n = 80) of native breeds namely Gaddi and Rampur Bushair were infected and 15 have died. The outbreak started after a few animals contracted the disease during the summer grazing period at the highland pastures from migrating flocks of sheep. This initial outbreak resulted in a further spreading of the disease into the valley. Clinical examination revealed varying degree of cutaneous papular lesions and respiratory distress. Upon necropsy, visceral lesions in the lungs, trachea and kidneys were also found. Clinical and morbid samples were found positive for sheeppox virus using group specific P32 gene and I3L gene based multiplex PCR differentiating sheeppox and goatpox viruses. Histopathological, hematological and blood biochemical analysis also supported the pathology of an acute viral infection. The causative sheeppox virus strain was isolated using lamb testicular cell culture and phylogenetic analysis, based upon P32 and RPO30 genes, showed its clustering with other Indian strains reported from neighboring states. This study demonstrated the spread of sheeppox virus to new niches by migratory sheep flocks leading to establishment of endemic infections in many new pockets of higher Western Himalayas.

Pathologies Associated with Second Mandibular Molar Due to Various Types of Impacted Third Molar: A Comparative Clinical Study.

Introduction: Impacted third molars are associated with various degrees of damage to the second molars. The possible complications also include distal cervical caries, root resorption of second molar, periodontal problems, odontogenic cysts, etc. Whether a particular impacted third molar is going to affect second molar depends upon its position and orientation in the bone. Materials and Method: This study was carried out in 418 cases. Three examiners evaluated the patient clinically and radiographically and only those cases were included in this study where at least two observers agreed. A total of 341 cases (163 males and 178 female), age range (15-40 years) with impacted mandibular third molars, were included. Clinically and radiographically, the impacted mandibular third and second molars were evaluated; simultaneously, the prevalence of various pathologies associated with mandibular second molar (dental caries, periodontal pockets, root resorption) due to impacted third molar was also evaluated and compared among various types and positions of impactions. Results: Statistical analysis was carried out using Pearson Chi-square and Asymp. Sig. (two-sided) test. Prevalence of mesioangular impactions was maximum (50.1%). Mesioangular impaction and position B (Pell and Gregory classification) were significantly associated with dental caries (32.20% and 33.90%, respectively), and periodontal pockets were seen higher with position B impactions (26.8%) {horizontal (14.7%), disto-angular (12.10%), vertical (14.5%) mesioangular (16.4%%)} in adjacent mandibular second molar. Root resorption was seen maximally in horizontal impaction (17.30%) with position c type (12.30%). The order of pathologies associated with second molar due to impacted third molar was dental caries (19.9%) > periodontal pockets (15.2%) > root resorption (8.5%). Discussion: Evidence regarding pathologies are associated with second molar due to impacted third molar aids in decision making for surgical removal of third molars. Different types of impaction and the prevalence of pathologies related to them would aid in treatment planning of the impacted tooth as certain types have high probability of pathologies associated.

Penicilliosis presenting as fungating skin lesion.

A 35-year-old HIV seropositive male patient presented with fever, weight loss, papules, nodules and fungating masses all over the body. Histopathological and mycological study of the skin biopsy tissue confirmed the diagnosis of penicilliosis. Although penicilliosis is restricted to Southeast Asia, more cases are being recognized in nonendemic countries.

Effects of intraoperative placement of tetracycline, tetracycline + gelatin sponge, and placebo on postoperative pain after mandibular molar extraction: A comparative prospective study.

BACKGROUND: Painless postoperative period is a major requisite following routine dental extractions. Reduction in the postextraction complication is beneficial to both clinician and patients. Hence, emphasis should be given to the techniques and agents that help reduce the complications for better postoperative recovery. MATERIALS AND METHODS: Three groups of 30 patients each requiring mandibular molar extractions were randomly selected with intrasocket placement of tetracycline, tetracycline plus gelatin sponge, and placebo control after extraction. A small piece of collagen membrane was used on the superior surface of the socket after the placement of the medicament in Group A and Group B. The postoperative pain scores were evaluated at 24 h, 48 h, and 7 days postoperatively. RESULTS: Ninety patients requiring mandibular molar teeth extraction were enrolled as the study participants. Forty-two patients (55.26%) were male and 34 (44.73%) were female. Tetracycline alone and tetracycline + gelatin sponge group shows no statistical significant difference in reducing postoperative pain after 7 days, but the pain score values were less in Tetracycline + gelatin sponge group. The comparison between tetracycline alone and control group showed no significant difference observed between the groups at 24 h but showed statistically significant difference between the groups after 48 h and 7 days, whereas comparison between tetracycline + gelatin sponge and control group showed statistically significant difference between the groups after 24 h, 48 h, and 7 days (P = 0.009, 0.001, and 0.017, respectively). CONCLUSION: Tetracycline and gelatin sponge intrasocket placement provides a good substitute to the prolong use of analgesics and as a good adjuvant in reducing pain in the first few days after routine dental extraction.

Theileria orientalis outbreak in an organized cattle breeding farm.

Theileriosis is an important tick borne disease of cattle caused by a haemoprotozoan of genus Theileria. Clinical bovine theileriosis is mainly caused by T. annulata or T. parva but the clinical disease due to T. orientalis is rare. T. orientalis mainly infect RBCs and causes "Oriental theileriosis" or Theileria-associated bovine anaemia in cattle and other livestock species. Two genotypes of T. orientalis (Chitose and Ikeda) are reported to cause severe disease in some countries. In this report, a spontaneous outbreak of Oriental theileriosis was studied in an organized Holstein-Friesian cattle breeding farm situated in the south-eastern Himalayan ranges of Himachal Pradesh State of India. Animal blood and tick samples were tested using cytological and PCR techniques. The disease episode occurred in a protracted manner spanning over 10 to 12 months and association of T. orientalis was confirmed in 93.3% of the blood and 21.7% of Rhipicephalus microplus (tick) samples. No other tick borne pathogen was detected except Anaplasma marginale in two blood samples. Haematological profiling of infected cattle showed characteristic indicators of anaemia like haemoblobin, RBC count, haematocrit value and mean corpuscular volume at either lower than normal or near the lower normal range. The prevailing persistent anaemic changes led to more severe clinical manifestations like abortion and joint inflammation. The detected T. orientalis strains and ticks species were further confirmed by nucleotide sequence analysis of 18S rRNA and 16S rRNA genes. Phylogenetically, T. orientalis strains showed clustering with other reported strains of T. orientalis from the surrounding regions. This first report of clinical Oriental theileriosis from India emphasises the importance of T. orientalis as an emerging tick borne pathogen and role of widely prevalent ticks species in disease transmission and their impact on livestock production.

A review: Lumpy skin disease and its emergence in India.

Lumpy skin disease (LSD) is a viral disease caused by lumpy skin disease virus (LSDV), a member of Capripoxvirus genus of Poxviridae family. It is a transboundary disease of the economic importance affecting cattle and water buffaloes. The disease is transmitted by arthropod vectors and causes high morbidity and low mortality. LSD has recently been reported first time in India with 7.1% morbidity among cattle. Generally, fever, anorexia, and characteristic nodules on the skin mucous membrane of mouth, nostrils, udder, genital, rectum, drop in milk production, abortion, infertility and sometimes death are the clinical manifestations of the disease. The disease is endemic in African and Middle East countries but has started spreading to Asian and other countries. It has been recently reported from China and Bangladesh sharing borders with India. We have summarized occurrence of LSD outbreaks in last 10 years in Asian countries for the first time. In India, currently epidemiological status of the disease is unknown. Vaccination along with strict quarantine measures and vector control could be effective for preventing the spread of the disease. This review aims to summarise the latest developments in the epidemiology with the focus on transboundary spread, aetiology and transmission, clinical presentations, diagnostics and management of the disease.

Holistic development of coal tar lotion by embedding design of experiments (DoE) technique: preclinical investigations.

Background: The research work endeavors to develop a liquid dosage form of an efficacious antipsoriatic drug, i.e., coal tar, but having problems like variability and patient noncompliance.Methods: The emulsion was prepared by the wet gum method from standardized coal tar. The optimized lotion obtained after sequential experimental designs was characterized for various dosage form and/or coal tar-related properties including efficacy.Results: The formulation deposited more coal tar in the unit area of rat skin than marketed lotions. The efficacy of lotion in psoriasis animal models was more or equivalent to marketed lotions. The formulation showed one compartment body model dermatokinetics, nonirritancy after repeated applications, and stability at room conditions for a year.Conclusion: The formulation with desired attributes was successfully developed.

Stability Kinetics of Imiquimod: Development and Validation of an Analytical Method.

For a new immune modulator imiquimod, various liquid chromatography methods have been described in literature but all of them are deficient in one or other aspects of complete method development. The present work intends to develop and validate the stability indicating reverse phase high performance liquid chromatographic (HPLC) method. The isocratic flow of mobile phase comprising equal volume ratio of acetate buffer BP pH 3.7 and acetonitrile at the rate of 1.5 mL/min through the C-18 column at 25°C lead to elution of drug around 2.3 min when analyzed at 244 nm using UV-detector. The linear regression equation in calibration plot was y = 61632×-1224 with 0.9992 coefficient of determination (r2). The percent relative standard variation (% RSD) in peak area at low, mid and high region of linearity range was less than 5% in precision studies. The method was able to detect 0.039 µg/mL of drug but practical limit of quantitation (LOQ) was 1.5 µg/mL. The imiquimod molecule was stable in all except oxidizing conditions where it degraded into more polar molecule in hydrogen peroxide (H2O2) concentration dependent manner. Therefore, an analytical method capable of accurately and specifically estimating the drug in microgram range was successfully developed.

Actinic keratosis and imiquimod: a review of novel carriers and patents.

INTRODUCTION: Actinic keratosis is one of the most common disorder characterized by erythematic and generally attached scaly lesions which are present either alone or in clusters. World Health Organization defines actinic keratosis as a common intraepidermal neoplasm of sun-damaged skin, characterized by variable atypia of keratinocytes. AREAS COVERED: At the beginning of the 20th century, a new immunomodulator molecule, imiquimod, appears in the market for the treatment of actinic keratosis but suffers from the pitfalls of the conventional approach of dosage form preparation including high dose, poor stability and more side effects. The present article attempts to compile the scatter information related to actinic keratosis and imiquimod at one place. The special emphasis will be made on the information available in various research articles and patents with respect to the efforts made for overcoming shortcomings associated with imiquimod by novel drug delivery or other approaches. EXPERT OPINION: The conventional drug delivery systems are unsuccessful to improve the actinic keratosis. The patient acceptance and compliance with these treatments are generally poor due to associated side effects, poor cosmetic outcomes and high costs. Therefore, several available and reported novel therapeutic approaches are being developed in order to provide better action.

Effect of Metamorphed Keratolytic Agent on the Behavior of Imiquimod Loaded Hybrid Vesicles Containing Gel.

The cost, side effects, and patient compliance-related issues of topically effective imiquimod have prevented its widespread acceptance. The present work intends to evaluate the feasibility of overcoming the shortcomings of poorly soluble and skin-penetrating immunomodulator by using biocompatible keratolytic agent with drug-loaded hybrid vesicles. Salicylic acid was complexed with phospholipid through simple mixing and incorporated into carbopol 940 gel containing drug-loaded vesicles, prepared by thin-film hydration method. The morphology, physicochemical properties, rheological behavior, release profile, and dermatokinetics of developed gel were compared with control gel (developed gel without keratolytic agent). In ex vivo drug release studies across the rat skin, there was significant increase in the steady-state permeation flux (Jss) and skin retention of drug from developed gel in comparison with control. There was favorable change in almost every evaluated dermatokinetic parameter. The innocuous nature of control gel had not changed on addition of skin structure-altering agent. The developed gel was found to be stable at room temperature and humidity for 1 year.

Anesthetic Efficacy of Single Buccal Infiltration of 4% Articaine and 2% Lignocaine in Extraction of Maxillary 1st Molar.

BACKGROUND: The extraction of tooth being the most common procedure in oral surgery should be pain free with limited dosage and limited needlepricks. Articaine being unique among amide local anesthetics contains a thiophene group, which increases its liposolubility, and an ester group which helps biotransformation in plasma. Because of the high diffusion properties, it can be used as a single buccal infiltration to extract a maxillary tooth. AIM AND OBJECTIVE: Objective of the study was to compare the efficacy of single buccal infiltration of 4% articaine with that of 2% lignocaine for maxillary first molar extraction. METHODOLOGY: A triple blind randomized controlled study was carried on 100 patients of age group 18-60 years who required maxillary first molar extraction, visiting the Department of Oral and Maxillofacial surgery. They were included in the study after obtaining informed consent. Buccal infiltration of 1.8 ml of anesthetic solution was given randomly to 100 patients with appropriate blinding of the cartridges. Objective signs were checked. If any additional injection was given, it was noted as type and number of rescue injection given. Postoperatively VAS score and surgeon's quality of anesthesia was noted. Duration of anesthesia was measured every 5 minutes for 50 minutes from infiltration. RESULTS: Out of 50 patients in group A (Articaine), in 44 patients extraction was done without the need of additional injection whereas in group B(Lignocaine), 29 patients require additional infiltration on the palatal side. The VAS score values for group A were also significantly less in comparison with group B. The mean duration of anesthesia for Group A being (71.70 ± 17.82 min) in 44 patients who only received buccal infiltration. INTERPRETATION AND CONCLUSION: The efficacy of single buccal injection of articaine is comparable to buccal and palatal injection of lignocaine.

Isolation of Moraxella bovis from frozen bovine semen and determination of microbial load.

In the present study, isolation of Moraxella bovis was done from the microbiological examination of frozen semen straws from clinically healthy twelve Jersey and one Jersey-cross bull supplied by a semen laboratory. The organism was identified on the basis of colonial morphology and biochemical characteristics. Further, viable cell count of the bacterium was studied in detail in the semen sample.