A review on anti‐tuberculosis peptides: Impact of peptide structure on anti‐tuberculosis activity

Antibiotic resistance is a major public health problem globally. Particularly concerning amongst drug‐resistant human pathogens is Mycobacterium tuberculosis that causes the deadly infectious tuberculosis (TB) disease. Significant issues associated with current treatment options for drug‐resistant TB and the high rate of mortality from the disease makes the development of novel treatment options against this pathogen an urgent need. Antimicrobial peptides are part of innate immunity in all forms of life and could provide a potential solution against drug‐resistant TB. This review is a critical analysis of antimicrobial peptides that are reported to be active against the M tuberculosis complex exclusively. However, activity on non‐TB strains such as Mycobacterium avium and Mycobacterium intracellulare, whenever available, have been included at appropriate sections for these anti‐TB peptides. Natural and synthetic antimicrobial peptides of diverse sequences, along with their chemical structures, are presented, discussed, and correlated to their observed antimycobacterial activities. Critical analyses of the structure allied to the anti‐mycobacterial activity have allowed us to draw important conclusions and ideas for research and development on these promising molecules to realise their full potential. Even though the review is focussed on peptides, we have briefly summarised the structures and potency of the various small molecule drugs that are available and under development, for TB treatment.     

In vitro antibiofilm and anti‐adhesion effects of magnesium oxide nanoparticles against antibiotic resistant bacteria

The aim of the current investigation was to determine the antibacterial and antibiofilm potential of MgO nanoparticles (NPs) against antibiotic‐resistant clinical strains of bacteria. MgO NPs were synthesized by a wet chemical method and further characterized by scanning electron microscopy and energy dispersive X‐ray. Antibacterial activity was determined by broth microdilution and agar diffusion methods. The Bradford method was used to assess cellular protein leakage as a result of loss of membrane integrity. Microtiter plate assay following crystal violet staining was employed to determine the effect of MgO NPs on biofilm formation and removal of established biofilms. MIC values ranged between 125 and 500 μg/mL. Moreover, treatment with MgO NPs accelerated rate of membrane disruption, measured as a function of leakage of cellular proteins. Leakage of cellular protein content was greater among gram‐negative bacteria. Cell adherence assay indicated 25.3–49.8% inhibition of bacterial attachment to plastic surfaces. According to a static biofilm method, MgO NPs reduced biofilm formation potential from 31% to 82.9% in a time‐dependent manner. Moreover, NPs also significantly reduced the biomass of 48, 72, 96 and 120 hr old biofilms (P < 0.05). Cytotoxicity experiments using a neutral red assay revealed that MgO NPs are non‐toxic to HeLa cells at concentrations of 15–120 μg/mL. These data provide in vitro scientific evidence that MgO NPs are effective and safe antibiofilm agents that inhibit adhesion, biofilm formation and removal of established biofilms of multidrug‐resistant bacteria.

     

Survival of Mycobacterium tuberculosis during experimental aerosolization and implications for aerosol challenge models

Aims: We undertook a series of experiments to investigate factors that contribute to variation in Mycobacterium tuberculosis viability and infectivity, during experimental aerosolization, with an aim to optimize a strategy to enable a more reproducible delivered dose within animal models of tuberculosis. Methods and results: The viability and infectivity of the challenge suspension was determined in relation to aerosolization time, concentration, method of preparation and M. tuberculosis strain. Challenge stocks generated from frozen aliquots of M. tuberculosis were shown to undergo a 1 log₁₀ CFU ml^?1 decrease in viability during the first 10 min of aerosolization. This correlated with a decrease in surface lung lesions developing in guinea pigs challenged during this time. The phenomenon of decreased viability in vitro was not observed when using freshly grown, nonfrozen cells of M. tuberculosis. The viability of aerosolized bacilli at the point of inhalation relative to the point of aerosolization always remained constant. Conclusion: Based on these findings, we have developed an improved strategy by which to reproducibly deliver aerosol infection doses to individually challenged animals and separately challenged groups of animals. Significance and Impact of the Study: Study of the aerobiological characteristics of micro‐organisms is a critical step in the validation of methodology for aerosol infection animal models, particularly where large numbers of animals and nonhuman primates are used.     

Cloning and characterization of a gene from Streptomyces griseus coding for a streptomycin-phosphorylating activity

Abstract Six different plasmids expressing streptomycin (SM) resistance and SM phosphotransferase were obtained by cloning genomic DNA from Streptomyces griseus into Streptomyces lividans . The phosphorylating enzymatic activity formed in S. lividans differed in several biochemical properties from the one in S. griseus , though the phosphorylated products were identical.     

Molecular identification and characterization of rifampicin-resistant Mycobacterium tuberculosis isolates by line probe assay: an approach for rapid diagnosis of multidrug-resistant tuberculosis

Aim:  Early identification and characterization of rifampicin-resistant (R^r) Mycobacterium tuberculosis isolates recovered from the samples of tuberculosis (TB) patients in the Aegean (West Anatolian) Region was intended.
Methods and Results:  Sixty isolates [47 (78·3%) multidrug-resistant (MDR)], which were identified as M. tuberculosis complex and phenotypically resistant to rifampicin by both BACTEC mycobacteria growth indicator tube (MGIT) 960 and 460 systems were analysed by a commercial line probe assay (INNO-LiPA Rif TB). The concordance of LiPA with the in vitro susceptibility test was found as 98·3%. Among the isolates, S531L (R5 pattern; 46·7%) and L511P/R, S512T, Q513L/K (ΔS1 pattern; 11·7%) were the most frequent mutation patterns. As compared with the BACTEC systems and conventional techniques for cultivation, identification and in vitro susceptibility testing, INNO-LiPA Rif TB after cultivation in BACTEC MGIT 960 system provided an average of 20 days early diagnosis of R^r M. tuberculosis isolates.
Conclusions:  Rapid molecular identification and characterization of R^r M. tuberculosis isolates after BACTEC MGIT 960 cultivation would be useful for faster diagnosis, infection control and planning of accurate treatment in MDR-TB patients.
Significance and Impact of the Study:  Patients with MDR-TB need a specified treatment and efficient follow-up strategies. Rapid and practical methodologies to diagnose and follow these patients should be applied in routine use.     

Bioinspired molecular adhesive for water‐resistant oxygen indicator films

Mussels can attach themselves to nearly all types of hard surfaces in wet environments. Such attractive adhesive ability of mussels is believed to rely on the amino acid composition of proteins found near the plaque–substrate interface. Dopamine (DA) is identified as a simplified mimic of mussel proteins, which are rich in 3,4‐dihydroxy‐l ‐phenylalanine and lysine, because it contains both catechol and amine functional groups. In this work, we have first applied this bioinspired adhesive to tackle a dye leaching problem of colorimetric oxygen indicator films, which are widely used to ensure the absence of oxygen inside the package of oxygen‐sensitive materials. Simple immersion of packaging films into a DA solution resulted in poly(DA) deposition, decreasing the water contact angle of the films from 105° to 65°. The poly(DA) coating could reduce the thionine leakage of the UV‐activated oxygen indicator film. The effects of poly(DA) coating were found to be dependent on the DA solution pH, the coating time, and the DA concentration. The film resistant to dye leaching lost its dye color by 5 min UVB irradiation and regained the color in the presence of oxygen, demonstrating that it functioned successfully as UV‐activated oxygen indicators. © 2013 American Institute of Chemical Engineers Biotechnol. Prog., 29: 513–519, 2013     

Use of an anti‐apoptotic CHO cell line for transient gene expression

Transient gene expression in mammalian cells allows for rapid production of recombinant proteins for research and preclinical studies. Here, we describe the development of a polyethylenimine (PEI) transient transfection system using an anti‐apoptotic host cell line. The host cell line, referred to as the Double Knockout (DKO), was generated by deleting two pro‐apoptotic factors, Bax and Bak, in a CHO‐K1 cell line using zinc finger nuclease mediated gene disruption. Optimized DNA and PEI volumes for DKO transfections were 50% and 30% lower than CHO‐K1, respectively. During transfection DKO cells produced relatively high levels of lactate, but this was mitigated by a temperature shift to 31°C which further enhanced productivity. DKO cells expressed ～3‐ to 4‐fold higher antibody titers than CHO‐K1 cells. As evidence of their anti‐apoptotic properties post‐transfection, DKO cells maintained higher viability and had reduced levels of active caspase‐3 compared to CHO‐K1 cells. Nuclear plasmid DNA copy numbers and message levels were significantly elevated in DKO cells. Although DNA uptake levels, as early as 40 min post‐transfection, were higher in DKO cells this was not due to differences in cell surface heparan sulfate (HS) or initial endocytosis mechanism as both cell types utilized caveolae‐ and clathrin‐mediated endocytosis to internalize DNA:PEI complexes. These results suggest that the increased transfection efficiency and titers from DKO cells are attributed to their resistance to transfection‐induced apoptosis and not differences in endocytosis mechanism. © 2013 American Institute of Chemical Engineers Biotechnol. Prog., 29:1050–1058, 2013     

上海某高校新校区中抗生素抗性基因污染状况分析

探究新型环境污染物—抗生素抗性基因（ARGs）在校园环境中的分布状况。通过聚合酶链式反应(PCR)对上海某高校使用5年新校区不同区域污水检查井污泥中8种四环素类、4种磺胺类、7种β-内酰胺类、4种链霉素类和5种氯霉素类ARGs进行定性研究,并利用变性梯度凝胶电泳(DGGE)技术分析污泥中细菌群落的多样性。结果显示,校园各区域中共检出19种ARGs,有8种ARGs的检出率大于50%,其中磺胺类抗性基因sulI、sulII的检出率最高,为100%。实验区及餐饮区的ARGs检出种类最多,均为14种,其次为宿舍区(12种),教学区的ARGs检出最少(8种)。通过DGGE分析细菌群落结构,证明该地区的ARGs分布与细菌多样性无明显关系。新校区使用5年但ARGs污染严重,可能是由于人类活动（尤其是科研活动）对ARGs的产生及扩散存在促进作用。此外,细菌群落多样性与ARGs种类的关系表明ARGs在环境中的迁移可能受到除细菌种类之外其他环境因素的影响。     

Research progress on distribution,migration, transformation of antibiotics and antibiotic resistance genes (ARGs) in aquatic environment

Antimicrobial and antibiotics resistance caused by misuse or overuse of antibiotics exposure is a growing and significant threat to global public health. The spread and horizontal transfer of antibiotic resistant bacteria (ARB) and antibiotic resistance genes (ARGs) by the selective pressure of antibiotics in an aquatic environment is a major public health issue. To develop a better understanding of potential ecological risks die to antibiotics and ARGs, this study mainly summarizes research progress about: (i) the occurrence, concentration, fate, and potential ecological effects of antibiotics and ARGs in various aquatic environments, (ii) the threat, spread, and horizontal gene transfer (HGT) of ARGs, and (iii) the relationship between antibiotics, ARGs, and ARB. Finally, this review also proposes future research direction on antibiotics and ARGs.     

Identification of castration‐resistant prostate cancer‐related hub genes using weighted gene co‐expression network analysis

Prostate cancer is the most common malignancy in urinary system and brings heavy burdens in men. We downloaded gene expression profile of mRNA and related clinical data of GSE70768 data set from public database. Weighted gene co‐expression network analysis (WGCNA) was used to identify the relationships between gene modules and clinical features, as well as the candidate genes. Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO) analyses were developed to investigate the potential functions of related hub genes. Importantly, basic experiments were performed to verify the relationship between hub genes and the phenotype previously identified. Lastly, copy number variation (CNV) analysis was conducted to explore the genetical alteration. WGCNA identified that black module was the most relevant module which was tightly related to castration‐resistant prostate cancer (CRPC) phenotype. KEGG and GO analysis results revealed genes in black module were mainly related to RNA splicing. Additionally, 9 genes were chosen as hub genes and heterogeneous nuclear ribonucleoprotein A2/B1 (HNRNPA2B1), golgin A8 family member B (GOLGA8B) and mitogen‐activated protein kinase 8 interacting protein 3 (MAPK8IP3) were identified to be associated with PCa progression and prognosis. Moreover, all above three genes were highly expressed in CRPC‐like cells and their suppression led to hindered cell proliferation in vitro. Finally, CNV analysis found that amplification was the main type of alteration of the 3 hub genes. Our study found that HNRNPA2B1, GOLGA8B and MAPK8IP3 were identified to be tightly associated with tumour progression and prognosis, and further researches are needed before clinical application.     

环境耐药组及其健康风险的宏基因组学研究策略和方法

抗生素耐药性在环境中的发展和传播对人体健康造成潜在风险。随着高通量测序技术和生物信息学方法的不断发展,宏基因组学技术被广泛应用于不同环境样本的抗生素耐药组研究。本文介绍了两种针对环境耐药组筛查的宏基因组学分析方法,总结了当前主流的生物信息学软件和数据库,并阐述了环境耐药组的风险评估框架和基于宏基因组学技术的相关实践,以期为环境耐药组的监测、风险评估和管控提供可行的路线图。     

Detection and quantification of methicillin‐resistant Staphylococcus aureus (MRSA) clones in retail meat products

Aims: The objective of the study was to determine the prevalence of methicillin‐resistant Staphylococcus aureus (MRSA) contamination of retail meat and to determine the level of contamination. Methods and Results: Pork (pork chops and ground pork), ground beef and chicken (legs, wings and thighs) were purchased at retail outlets in four Canadian provinces and tested for the presence of methicillin‐resistant Staph. aureus using qualitative and quantitative methods. MRSA was isolated from 9·6% of pork, 5·6% of beef and 1·2% of chicken samples (P = 0·0002). Low levels of MRSA were typically present, with 37% below the detection threshold for quantification and <100 CFU g^?1 present in most quantifiable samples. All isolates were classified as Canadian epidemic MRSA‐2 (CMRSA‐2) by pulsed field gel electrophoresis (PFGE), with two different PFGE subtypes, and were spa type 24/t242. Conclusions: MRSA contamination of retail meat is not uncommon. While CMRSA‐2, a human epidemic clone, has been found in pigs in Canada, the lack of isolation of livestock‐associated ST398 was surprising. Significance and Impact of the Study: The relevance of MRSA contamination of meat is unclear but investigation is required because of the potential for exposure from food handling. Sources of contamination require investigation because these results suggest that human or animal sources could be involved.