Molecular Cloning and Characterization of P4 Nuclease from Leishmania infantum

Parasite of the genus Leishmania is reliant on the salvage pathway for recycling of ribonucleotides. A class I nuclease enzyme also known as P4 nuclease is involved in salvage of purines in cutaneous Leishmania species but the relevant enzymes have not been characterized in Leishmania infantum (L. infantum). The aim of this study was to clone and characterize the gene encoding class I nuclease in L. infantum. DNA extracted from L. infantum was used for amplification of P4 nuclease gene (Li-P4) by PCR. The product was cloned, sequenced, and expressed in E. coli for further characterization. Analysis of the sequence of Li-P4 revealed that the gene consists of an ORF of 951 bp. Sequence similarity analysis indicated that Li-P4 has a high homology to relevant enzymes of other kintoplastids with the highest homology (88%) to p1/s1 class I nuclease from L. donovani. Western blotting of antirecombinant Li-P4 with promastigote and amastigote stages of L. infantum showed that this nuclease is present in both stages of parasite with higher expression in amastigote stage. The highly conserved nature of this essential enzyme in Leishmania parasites suggests it as a promising drug target for leishmaniasis

Tongue carcinoma: case series and demographic analysis

BACKGROUND: Oral tongue squamous cell carcinoma (SCC) accounts for approximately half of oral cavity SCCcases. Smoking and drinking are two major risk factors for SCC worldwide. Recently, increasing incidence amongyoung white individuals who were never-smoker, never-drinker have been reported. METHODS: To determine the demographic status of tongue SCC in our region, this study was conducted amongpatients who undergone surgery in Tabriz, Iran, due to tongue SCC. RESULTS: Mean age of 27 patients was 61 years and 22% of them were under 45 years old. Mean duration ofbeginning the symptoms to biopsy was 113 days for patients under the age of 60 years and 253 days for elderly patients(P = 0.026). Early diagnosis, especially in patients more than 60 years old, was associated with good prognosis. CONCLUSIONS: The results of this study indicated that considering a lower threshold for obtaining biopsy fromelderly patients, even in the first visit, is useful for early diagnosis and better prognosis

Isolation and characterization of a novel human scFv inhibiting EGFR vIII expressing cancers

EGFRvIII, a mutant form of epidermal growth factor receptor is highly expressed in glioblastoma, carcinoma of the breast, ovary, and lung but not in normal cells. This tumor specific antigen has emerged as a promising candidate for antibody based therapy of several cancers. The aim of the present study was isolation and characterization of a human single chain antibody against EGFRvIII as a promising target for cancer therapy. For this, a synthetic peptide corresponding to EGFRvIII protein was used for screening the naive human scFv phage library. Selection was performed using a novel screening strategy for enrichment of rare specific clones. After five rounds of screening, six positive scFv clones against EGFRvIII were selected using monoclonal phage ELISA, among them, a clone with an amber mutation in VH CDR2 coding sequence showed higher reactivity. The mutation was corrected through site directed mutagenesis and then scFv fragment was expressed after subcloning into the bacterial expression vector. Expression in BL21 pLysS resulted in a highly soluble scFv appeared in soluble fraction of E. coli lysate. Bioinformatic in silico analysis between scFv and EGFRvIII sequences confirmed specific binding of desired scFv to EGFRvIII in CDR regions. The specific reactivity of the purified scFv with native EGFRvIII was confirmed by cell based ELISA and western blot. In conclusion, human anti-EGFRvIII scFv isolated from a scFv phage library displayed high reactivity with EGFRvIII. The scFv isolated in this study can be the groundwork for developing more effective diagnostic and therapeutic agents against EGFRvIII expressing cancers. (C) 2016 European Federation of Immunological Societies. Published by Elsevier B.V. All rights reserved

Global mortality associated with 33 bacterial pathogens in 2019: a systematic analysis for the Global Burden of Disease Study 2019

Summary Background Reducing the burden of death due to infection is an urgent global public health priority. Previous studies have estimated the number of deaths associated with drug-resistant infections and sepsis and found that infections remain a leading cause of death globally. Understanding the global burden of common bacterial pathogens (both susceptible and resistant to antimicrobials) is essential to identify the greatest threats to public health. To our knowledge, this is the first study to present global comprehensive estimates of deaths associated with 33 bacterial pathogens across 11 major infectious syndromes. Methods We estimated deaths associated with 33 bacterial genera or species across 11 infectious syndromes in 2019 using methods from the Global Burden of Diseases, Injuries, and Risk Factors Study (GBD) 2019, in addition to a subset of the input data described in the Global Burden of Antimicrobial Resistance 2019 study. This study included 343 million individual records or isolates covering 11 361 study-location-years. We used three modelling steps to estimate the number of deaths associated with each pathogen: deaths in which infection had a role, the fraction of deaths due to infection that are attributable to a given infectious syndrome, and the fraction of deaths due to an infectious syndrome that are attributable to a given pathogen. Estimates were produced for all ages and for males and females across 204 countries and territories in 2019. 95% uncertainty intervals (UIs) were calculated for final estimates of deaths and infections associated with the 33 bacterial pathogens following standard GBD methods by taking the 2·5th and 97·5th percentiles across 1000 posterior draws for each quantity of interest. Findings From an estimated 13·7 million (95% UI 10·9–17·1) infection-related deaths in 2019, there were 7·7 million deaths (5·7–10·2) associated with the 33 bacterial pathogens (both resistant and susceptible to antimicrobials) across the 11 infectious syndromes estimated in this study. We estimated deaths associated with the 33 bacterial pathogens to comprise 13·6% (10·2–18·1) of all global deaths and 56·2% (52·1–60·1) of all sepsis-related deaths in 2019. Five leading pathogens—Staphylococcus aureus, Escherichia coli, Streptococcus pneumoniae, Klebsiella pneumoniae, and Pseudomonas aeruginosa—were responsible for 54·9% (52·9–56·9) of deaths among the investigated bacteria. The deadliest infectious syndromes and pathogens varied by location and age. The age-standardised mortality rate associated with these bacterial pathogens was highest in the sub-Saharan Africa super-region, with 230 deaths (185–285) per 100 000 population, and lowest in the high-income super-region, with 52·2 deaths (37·4–71·5) per 100 000 population. S aureus was the leading bacterial cause of death in 135 countries and was also associated with the most deaths in individuals older than 15 years, globally. Among children younger than 5 years, S pneumoniae was the pathogen associated with the most deaths. In 2019, more than 6 million deaths occurred as a result of three bacterial infectious syndromes, with lower respiratory infections and bloodstream infections each causing more than 2 million deaths and peritoneal and intra-abdominal infections causing more than 1 million deaths. Interpretation The 33 bacterial pathogens that we investigated in this study are a substantial source of health loss globally, with considerable variation in their distribution across infectious syndromes and locations. Compared with GBD Level 3 underlying causes of death, deaths associated with these bacteria would rank as the second leading cause of death globally in 2019; hence, they should be considered an urgent priority for intervention within the global health community. Strategies to address the burden of bacterial infections include infection prevention, optimised use of antibiotics, improved capacity for microbiological analysis, vaccine development, and improved and more pervasive use of available vaccines. These estimates can be used to help set priorities for vaccine need, demand, and development