Bacteriophages Mediating Effective Elimination of Multidrug-Resistant Avian Pathogenic Escherichia coli.

Background: Avian pathogenic Escherichia coli (APEC) causes colibacillosis and septicemia; in certain cases, mortality leads to economic losses and elicits potential foodborne zoonotic risk. The study aimed to determine the prevalence of APEC pathotypes and serotypes in poultry, followed by characterization for virulence markers and antibiotic sensitivity and analysis of lytic efficacy of bacteriophages in the eradication of APEC. Methods: We successfully isolated and characterized 34 E. coli isolates from poultry farms. The lytic efficacy of seven bacteriophages, as well as a phage cocktail, was evaluated for biological control of multiple drug resistance (MDR) APEC. Results: A total of 67.65% of isolated E. coli were APEC. A total of 94.11% of the isolates were multidrug-resistant bacteria harboring virulence genes. The lytic ability of seven bacteriophages ranged from 0.98% to 36.76%, with a cocktail of EscoΦA-06 and ΦA-07 exhibiting lysis of 48.04% isolates. Conclusion: As serological variability in APEC limits the application and development of vaccines, the findings support the employment of bacteriophages against elimination of MDR APEC in poultry settings.

Calcium phosphate nanoclusters modify periodontium remodeling and minimize orthodontic relapse.

Orthodontic relapse is one of the most prevalent concerns of orthodontic therapy. Relapse results in patients' teeth reverting towards their pretreatment positions, which increases the susceptibility to functional problems, dental disease, and substantially increases the financial burden for retreatment. This phenomenon is thought to be induced by rapid remodeling of the periodontal ligament (PDL) in the early stages and poor bone quality in the later stages. Current therapies, including fixed or removable retainers and fiberotomies, have limitations with patient compliance and invasiveness. Approaches using biocompatible biomaterials, such as calcium phosphate polymer-induced liquid precursors (PILP), is an ideal translational approach for minimizing orthodontic relapse. Here, post-orthodontic relapse is reduced after a single injection of high concentration PILP (HC-PILP) nanoclusters by altering PDL remodeling in the early stage of relapse and improving trabecular bone quality in the later phase. HC-PILP nanoclusters are achieved by using high molecular weight poly aspartic acid (PASP, 14 kDa) and poly acrylic acid (PAA, 450 kDa), which resulted in a stable solution of high calcium and phosphate concentrations without premature precipitation. In vitro results show that HC-PILP nanoclusters prevented collagen type-I mineralization, which is essential for the tooth-periodontal ligament (PDL)-bone interphase. In vivo experiments show that the PILP nanoclusters minimize relapse and improve the trabecular bone quality in the late stages of relapse. Interestingly, PILP nanoclusters also altered the remodeling of the PDL collagen during the early stages of relapse. Further in vitro experiments showed that PILP nanoclusters alter the fibrillogenesis of collagen type-I by impacting the protein secondary structure. These findings propose a novel approach for treating orthodontic relapse and provide additional insight into the PILP nanocluster's structure and properties on collagenous structure repair.

Hydroxide-Source-Dependent Polymorphism and Phase Stability of Cobalt(II) Hydroxides in Diffusion-Driven Systems.

Hydroxides of cobalt(II) exist predominantly in two polymorphic forms, namely, the blue-green α-form [α-Co(OH)2] and reddish ß-form [ß-Co(OH)2]. These hydroxides have a layered structure with interlayer galleries of around 7 and 4 Å, respectively, for α- and ß-Co(OH)2. In most of the previous studies, both the polymorphs were synthesized separately, and a few of them showed that the α-form gets converted to a thermodynamically more stable ß-form via physical processes. In the present work, we have optimized the conditions for the simultaneous synthesis of both polymorphs under identical conditions in the same reactor using the 1D reaction-diffusion framework by employing different outer electrolytes. We found that the polymorph chemistry of Co(OH)2 depends on the source and concentration of OH- rather than other reaction conditions or later physical transformation. The products are characterized to confirm their morphology, structure, and chemical environment. We observed that the use of NaOH and NH4OH as the OH- precursor leads to α-Co(OH)2 only; however, with NaOH, a continuous precipitate is formed, and with NH4OH, periodic precipitation is formed. On the other hand, with hydrazine (HYZ) as the OH- source, Liesegang bands of α-Co(OH)2 and ß-Co(OH)2 as granules are formed throughout the diffusion reactor. Another intriguing observation on the HYZ system is that at its high concentration, the bands of α-Co(OH)2 get converted to ß-Co(OH)2. We articulate the reasons and mechanism of those observations.

Multicentre, randomised, double-blind, placebo-controlled, proof of concept study of LSALT peptide as prevention of acute respiratory distress syndrome and acute kidney injury in patients infected with SARS-CoV-2 (COVID-19).

OBJECTIVE: Dipeptidase-1 (DPEP-1) is a recently discovered leucocyte adhesion receptor for neutrophils and monocytes in the lungs and kidneys and serves as a potential therapeutic target to attenuate inflammation in moderate-to-severe COVID-19. We aimed to evaluate the safety and efficacy of the DPEP-1 inhibitor, LSALT peptide, to prevent specific organ dysfunction in patients hospitalised with COVID-19. DESIGN: Phase 2a randomised, placebo-controlled, double-blinded, trial. SETTING: Hospitals in Canada, Turkey and the USA. PARTICIPANTS: A total of 61 subjects with moderate-to-severe COVID-19. INTERVENTIONS: Randomisation to LSALT peptide 5 mg intravenously daily or placebo for up to 14 days. PRIMARY AND SECONDARY OUTCOME MEASURES: The primary endpoint was the proportion of subjects alive and free of respiratory failure and/or the need for renal replacement therapy (RRT). Numerous secondary and exploratory endpoints were assessed including ventilation-free days, and changes in kidney function or serum biomarkers. RESULTS: At 28 days, 27 (90.3%) and 28 (93.3%) of subjects in the placebo and LSALT groups were free of respiratory failure and the need for RRT (p=0.86). On days 14 and 28, the number of patients still requiring more intensive respiratory support (O2 ≥6 L/minute, non-invasive or invasive mechanical ventilation or extracorporeal membrane oxygenation) was 6 (19.4%) and 3 (9.7%) in the placebo group versus 2 (6.7%) and 2 (6.7%) in the LSALT group, respectively (p=0.14; p=0.67). Unadjusted analysis of ventilation-free days demonstrated 22.8 days for the LSALT group compared with 20.9 in the placebo group (p=0.4). LSALT-treated subjects had a significant reduction in the fold expression from baseline to end of treatment of serum CXCL10 compared with placebo (p=0.02). Treatment-emergent adverse events were similar between groups. CONCLUSION: In a Phase 2 study, LSALT peptide was demonstrated to be safe and tolerated in patients hospitalised with moderate-to-severe COVID-19. TRIAL REGISTRATION NUMBER: NCT04402957.

Assessment of tympanic membrane perforation size using ImageJ software by ENT clinicians of different grades.

OBJECTIVE: To compare visual estimation versus ImageJ calculation of tympanic membrane perforation size in the paediatric population between clinicians of different experience. METHODS: Five images of tympanic membrane perforations in children, captured using an otoendoscope, were selected. The gold standard was the ImageJ results by one consultant otologist. Consultants, registrars and Senior House Officers or equivalent were asked to visually estimate and calculate the perforation size using ImageJ software. RESULTS: The mean difference in variation from gold standard between visual estimation and ImageJ calculation was 12.16 per cent, 95 per cent CI (10.55, 13.78) p < 0.05, with ImageJ providing a more accurate estimation of perforation. Registrars were significantly more accurate at visual estimation than senior house officers. There was no statistically significant difference in ImageJ results between the different grades. CONCLUSION: Using ImageJ software is more accurate at estimating tympanic membrane perforation size than visual assessment for all ENT clinicians regardless of experience.

The Role of Airway Management on Feeding Difficulties in Children With Pfeiffer Syndrome.

BACKGROUND: Pfeiffer syndrome is characterized by craniosynostosis, mid-face hypoplasia, broad thumbs, and often multilevel airway obstruction. Airway management is often required, including the use of positive airway ventilation, nasopharyngeal airway (NPA), or tracheostomy. OBJECTIVE: The objective of this study was to assess the impact an airway adjunct can have on feeding difficulties in children with Pfeiffer syndrome. METHODS: Retrospective review of patients diagnosed with Pfeiffer syndrome from January 1998 to January 2020 at one of England's 4 supraregional Craniofacial Units, Alder Hey Children's Hospital. Speech & Language Therapy case notes and medical notes were used to gather data, as well as the Oral Feeding Score component of the UK Craniofacial Outcome Score. RESULTS: Eleven patients were included. Six patients had no airway adjunct (55%): 3 had tracheostomy (27%) and 2 patients had NPA (18%). All patients with airway adjuncts were percutaneous endoscopic gastrostomy/percutaneous endoscopic jejunostomy fed. Those who did not require an airway adjunct had an Oral Feeding Score of 4.60 (SD: 0.49). The children who went on to have an airway adjunct had a mean preintervention Oral Feeding Score of 2.4 (SD: 0.8). The mean feeding score (postairway adjunct) in the NPA group was 2.0, compared with the tracheostomy group scoring 3.0. CONCLUSIONS: Children with Pfeiffer syndrome who require airway intervention have more significant feeding problems requiring feeding intervention. Although there were small numbers included in this study, there is a suggestion that airway adjuncts can contribute to feeding difficulties, particularly NPAs.

Effect of respiratory tract co-colonizers on initial attachment of Neisseria meningitidis.

Respiratory tract is a complex system comprising of unique microbiota inhabitants. Neisseria meningitidis, Staphylococcus aureus, Streptococcus pyogenes, Pseudomonas aeruginosa and Klebsiella pneumoniae are few prevalent bacteria in the community composition during lung infections. Although, N. meningitidis resides asymptomatically in nasopharynx of the human host, it can cause fatal infections like meningitis. However, factors affecting transit from carriage to symptomatic infection are not well understood. Various host metabolites and environmental conditions affect the virulence of bacteria. Here, we report that presence of co-colonizers significantly reduces the initial attachment of N. meningitidis to A549 nasopharyngeal epithelial cells. Further, significant decrease in invasion to A549 nasopharyngeal epithelial cells was observed. Moreover, survival in J774A.1 murine macrophage also increases significantly when conditioned media (CM) from S. pyogenes and L. rhamnosus is used for culturing N. meningitidis. The increase in survival could be attributed to increased capsule synthesis. The gene expression studies revealed increased expression of siaC and ctrB in CM prepared from the growth S. pyogenes and L. rhamnosus. Overall, the results suggest change in the virulence of N. meningitidis is assisted by lung microbiota.

Acyl Homoserine Lactone Sensitised Streptococcus Pyogenes Differentially Regulates the Transcriptional Expression of Early Growth Response 1 (EGR1) in Epithelial and Macrophage Cells.

The host transcriptional activator Early growth response 1 (EGR1) plays a vital role in cell cycle and differentiation, cell proliferation, and regulation of cytokines and several growth factors. It is an immediate-early gene that is expressed as an initial response to various environmental stimuli. Bacterial infection is one such factor that can trigger the expression of EGR1 in host. Therefore, it is imperative to understand expression of EGR1 during early stages of host-pathogen interaction. Streptococcus pyogenes is an opportunistic bacteria causing skin and respiratory tract infections in humans. The quorum-sensing molecule, N-(3-oxododecanoyl)-l-homoserine lactone (Oxo-C12), not synthesised by S. pyogenes, can be sensed by S. pyogenes leading to molecular changes in the pathogen. In this study, we investigated the role of Oxo-C12 on EGR1 regulation in lung epithelial and murine macrophage cell line upon S. pyogenes infection. We report that Oxo-C12 sensitised S. pyogenes upregulates the transcriptional expression of EGR1 through ERK1/2 pathway. It was observed that EGR1 was not involved in the intial attachment of S. pyogenes to A549 cells. However, inhibition of EGR1 in macrophage cell line, J774A.1, through the ERK1/2 pathway resulted in decreased adhesion of S. pyogenes. The EGR1 upregulation by Oxo-C12 sensitised S. pyogenes plays a vital role in enhancing the survival of S. pyogenes in murine macrophages, leading to persistent infection. Thus, understanding the molecular modulation in the host during bacterial infection will further help develop therapeutics to target specific sites.

Polyamine as a microenvironment factor in resistance to antibiotics.

One of the main issues in modern medicine is the decrease in the efficacy of antibiotic therapy against resistant microorganisms. The advent of antimicrobial resistance has added significantly to the impact of infectious diseases, in number of infections, as well as added healthcare costs. The development of antibiotic tolerance and resistance is influenced by a variety of environmental variables, and it is important to identify these environmental factors as part of any strategy for combating antibiotic resistance. The review aims to emphasize that biogenic polyamines are one of such environmental cues that impacts the antibiotic resistance in bacteria. The biogenic polyamines can help bacteria acquire resistance to antibiotics either by regulating the level of number of porin channels in the outer membrane, by modifying the outer membrane liposaccharides or by protecting macromolecule from antibiotic stress. Thus, understanding the way polyamines function in bacteria can thus be beneficial while designing the drugs to combat diseases.

Convalescent Plasma Therapy in Late-State, Severe COVID-19 Infection.

OBJECTIVES: Current evidence favors plasma to be effective against coronavirus disease 2019 (COVID-19) in critically ill patients in the early stages of infection. We investigated the safety and efficacy of convalescent plasma in specifically late-stage (designated as after 2 weeks of hospital admission) severe COVID-19 infection. We also conducted a literature review on the late-stage use of plasma in COVID-19. METHODS: This case series examined eight COVID-19 patients admitted to the intensive care unit (ICU) who met criteria for severe or life-threatening complications. Each patient received one dose (200 mL) of plasma. Clinical information was gathered in intervals of 1 day pretransfusion and 1 hour, 3 days, and 7 days posttransfusion. The primary outcome was effectiveness of plasma transfusion, measured by clinical improvement, laboratory parameters, and all-cause mortality. RESULTS: Eight ICU patients received plasma late in the course of COVID-19 infection, on average at 16.13 days postadmission. On the day before transfusion, the averaged initial Sequential Organ Failure Assessment (SOFA) score, PaO2:FiO2 ratio, Glasgow Coma Scale (GCS), and lymphocyte count were 6.5, 228.03, 8.63, and 1.19, respectively. Three days after plasma treatment, the group averages for the SOFA score (4.86), PaO2:FiO2 ratio (302.73), GCS (9.29), and lymphocyte count (1.75) improved. Although the mean GCS improved to 10.14 by posttransfusion day 7, the other means marginally worsened with an SOFA score of 5.43, a PaO2:FiO2 ratio of 280.44, and a lymphocyte count of 1.71. Clinical improvement was noted in six patients who were discharged from the ICU. CONCLUSIONS: This case series provides evidence that convalescent plasma may be safe and effective in late-stage, severe COVID-19 infection. Results showed clinical improvement posttransfusion as well as decreased all-cause mortality in comparison to pretransfusion predicted mortality. Randomized controlled trials are needed to conclusively determine benefits, dosage, and timing of treatment.

Antibiotics modulates the virulence of Neisseria meningitidis by regulating capsule synthesis.

The opportunistic pathogens residing are frequently exposed to range of antimicrobials which affects virulence attributes. Neisseria meningitidis, is a host-restricted commensal of human upper respiratory tract which is subjected to a variety of stresses within the host, including antibiotic exposure. One of the most important virulence factors for pathogenesis is the meningococcal lipo-oligosaccharide capsule. Role of capsules in antimicrobial resistance and persistence is not yet established. In this study, different virulence factors of N. meningitidis were examined in presence of sub-MIC of four antibiotics: penicillin, ciprofloxacin, erythromycin and chloramphenicol. We observed increased production of the capsule by N. meningitidis when grown in the presence of penicillin, erythromycin, and chloramphenicol at sub-inhibitory concentration. Capsular production increase concurrently with increased resistance to inducing antibiotic which also confers increased survival in human serum. Finally, we show that increased capsule production in response to antibiotic exposure is aided by siaC, ctrB, lipA gene expression. These findings show that capsule synthesis, a major pathogenicity determinant, is regulated in response to antibiotic stress. Our findings support a model in which gene expression changes caused by ineffective antibiotic treatment cause N. meningitidis transition between states of low and high virulence potential, contributing to pathogen's opportunistic nature.

Study on impact of flexible endoscopy training course for surgeons in India.

Context: Competence in flexible endoscopy is essential for all surgeons during this era of minimal access surgery. However, fewer surgeons have expertise in endoscopy due to a lack of training and interest. The Indian Association of Gastrointestinal Endo Surgeons devised a short-structured training course in the art and science of endoscopy. Aims: This study aimed to find the impact of the endoscopy training course (Endoscopic Fellowship of Indian Association of Gastrointestinal Endo Surgeons [EFIAGES]) in improving the endoscopic skill of surgeons. Settings and Design: Twenty-two-part electronic survey forms were sent to all 375 candidates who took the course between 2016 and 2019 for this retrospective observational study. Subjects and Methods: The following outcome measures were noted, namely technical competence in endoscopy before the course, delegate feedback about the course modules, volume of endoscopies before and after the course and quality indicators such as reaching up to duodenum (D2) and caecum before and after the course. Statistical Analysis Used: Statistical analysis of the impact of the course was done using Chi-square test. Results: Responses from 262 out of a total of 375 candidates were received. Seventy-seven per cent of trainees were pleased with content and mode of conduct of the course. The quality indicator of gastroscopy with the ability to reach D2 in 90% of the caseload was achieved by only 28% of trainees before the EFIAGES. This increased to 72% of candidates after the course and similar results were seen with colonoscopy also. Most of the candidates noted a distinct improvement in their endoscopic navigation skills subsequent to the course. Conclusions: Endoscopy skill transfer was possible with a short-structured endoscopy course. The surgical fraternity should realise the importance of endoscopy skills in the current era of surgical practice.

Cu(II)-Metallized Three-Layered Cu-8HQ Complex with Hierarchical Crystallite Morphologies Synthesized via Reaction-Diffusion.

Self-organization of regular band patterns of the precipitate via a reaction-diffusion (RD) framework is called Liesegang phenomenon. This non-equilibrium system is emerging as an efficient method for synthesizing materials with unique morphologies that may have desired properties. The formation of continuous precipitation inside a band with poor control over the shape and size of sparingly soluble salts has been well documented. However, only a few reports on forming organic-inorganic bonds are available. In the present work, we demonstrate the formation of 2D frameworks of bis-(8-hydroxyquinoline) copper(II) in the agar gel via RD. The macroscopic particles were dumbbell-shaped, with aspect ratios ranging from 2.7 (inner bands) to 0.7 (outer bands). The particles were composed of ribbon-shaped crystallites at the microscopic level, each with three layers of parallelogram prismatic monoclinic sheets stacked over one another, which could easily be exfoliated. The powder X-ray diffraction patterns at low angles and the surface areas of the crystallites indicated the formation of metal-organic frameworks. It was observed that the sizes of the particles could be tuned by controlling the extent of diffusion using reactant concentrations. Since such heterostructures have energy storage capacity, the cyclic voltammograms of the unexfoliated and exfoliated materials showed that they fall in the pseudocapacitor category with potential application as the electrode material. The frameworks were further characterized by techniques such as optical and electron microscopy, X-ray diffraction, IR spectroscopy, and UV-visible spectrophotometry.

Calcium Phosphate Delivery Systems for Regeneration and Biomineralization of Mineralized Tissues of the Craniofacial Complex.

Calcium phosphate (CaP)-based materials have been extensively used for mineralized tissues in the craniofacial complex. Owing to their excellent biocompatibility, biodegradability, and inherent osteoconductive nature, their use as delivery systems for drugs and bioactive factors has several advantages. Of the three mineralized tissues in the craniofacial complex (bone, dentin, and enamel), only bone and dentin have some regenerative properties that can diminish due to disease and severe injuries. Therefore, targeting these regenerative tissues with CaP delivery systems carrying relevant drugs, morphogenic factors, and ions is imperative to improve tissue health in the mineralized tissue engineering field. In this review, the use of CaP-based microparticles, nanoparticles, and polymer-induced liquid precursor (PILPs) amorphous CaP nanodroplets for delivery to craniofacial bone and dentin are discussed. The use of these various form factors to obtain either a high local concentration of cargo at the macroscale and/or to deliver cargos precisely to nanoscale structures is also described. Finally, perspectives on the field using these CaP materials and next steps for the future delivery to the craniofacial complex are presented.

Circulation of a single hepatitis A virus genotype IIIA with two distinct clusters in different states of India.

With the changing hepatitis A epidemiology in India, focal viral outbreaks are being reported from different parts of the country. This study presents Hepatitis A Virus (HAV) strain characterization (period 2009-2020) from 18 states of India. For that, blood and stool samples (n â€‹= â€‹280) were screened for HAV RNA and sequences for 5'non-coding and VP3 regions were generated from positive samples (n â€‹= â€‹68). Presence of a single IIIA genotype in all samples indicated IIIA being the only HAV genotype currently circulating in India. Interestingly, it was evident that these strains form two distinct groups suggesting independent evolution of these two clusters.

CRISPR in Modulating Antibiotic Resistance of ESKAPE Pathogens.

The ESKAPE (Enterococcus spp., Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter spp.) isolates both from the clinical settings and food products are demonstrated to gain resistance to multiple antimicrobials. Therefore, the ESKAPE pathogens pose a serious threat to public health, which warrants specific attention to developing alternative novel therapeutics. The clustered regularly interspaced short palindromic repeats associated (CRISPR-Cas) system is one of the novel methods for managing antibiotic-resistant strains. Specific Cas nucleases can be programmed against bacterial genomic sequences to decrease bacterial resistance to antibiotics. Moreover, a few CRISPR-Cas nucleases have the ability to the sequence-specific killing of bacterial strains. However, some pathogens acquire antibiotic resistance due to the presence of the CRISPR-Cas system. In brief, there is a wide range of functional diversity of CRISPR-Cas systems in bacterial pathogens. Hence, to be an effective and safe infection treatment strategy, a comprehensive understanding of the role of CRISPR-Cas systems in modulating antibiotic resistance in ESKAPE pathogens is essential. The present review summarizes all the mechanisms by which CRISPR confers and prevents antibiotic resistance in ESKAPE. The review also emphasizes the relationship between CRISPR-Cas systems, biofilm formation, and antibiotic resistance in ESKAPE.

Lactic acid bacteria: prominent player in the fight against human pathogens.

INTRODUCTION: The human microbiome is a unique repository of diverse bacteria. Over 1000 microbial species reside in the human gut, which predominantly influences the host's internal environment and plays a significant role in host health. Lactic acid bacteria have long been employed for multiple purposes, ranging from food to medicines. Lactobacilli, which are often used in commercial food fermentation, have improved to the point that they might be helpful in medical applications. AREAS COVERED: This review summarises various clinical and experimental evidence on efficacy of lactobacilli in treating a wide range of infections. Both laboratory based and clinical studies have been discussed. EXPERT OPINION: Lactobacilli are widely accepted as safe biological treatments and host immune modulators (GRAS- Generally regarded as safe) by the US Food and Drug Administration and Qualified Presumption of Safety. Understanding the molecular mechanisms of lactobacilli in the treatment and pathogenicity of bacterial infections can help with the prediction and development of innovative therapeutics aimed at pathogens which have gained resistance to antimicrobials. To formulate effective lactobacilli based therapy significant research on the effectiveness of different lactobacilli strains and its association with demographic distribution is required. Also, the side effects of such therapy needs to be evaluated.

Spermidine promotes lysozyme tolerance and acid stress resistance in Streptococcus pyogenes M3.

Streptococcus pyogenes are Gram-positive opportunistic pathogens residing in the human nasopharynx and skin. Changes in environmental conditions, such as pH, temperature and availability of essential ions, can stimulate the expression of S. pyogenes virulence factors. One such factor could be the availability of an extracellular pool of polyamines. Polyamines are synthesized from amino acids, and are universally present in the environment. Polyamines have been implicated in the ecology of pathogenesis by modulating quorum sensing, host adaptation and virulence. Polyamines mediate pathogenesis and help the pathogen resist environmental stress. In this study, we investigated the ability of the polyamine, spermidine, to promote acid stress survival of S. pyogenes. S. pyogenes does not synthesize spermidine, but the extracellular pool of spermidine constituted by the host and microbiome could be utilized as a signalling molecule. We report that spermidine promotes acid stress resistance in S. pyogenes. Moreover, spermidine affects the morphology of S. pyogenes by decreasing the cell size and increasing the dltA gene expression. Along with dltA, spermidine upregulated the gene expression of cell wall-modifying genes such as mur, pgdA, pepO and srtA, which might help the bacteria to resist acidic stress.