Protein kinase PfPK2 mediated signalling is critical for host erythrocyte invasion by malaria parasite.

Signalling pathways in malaria parasite remain poorly defined and major reason for this is the lack of understanding of the function of majority of parasite protein kinases and phosphatases in parasite signalling and its biology. In the present study, we have elucidated the function of Protein Kinase 2 (PfPK2), which is known to be indispensable for the survival of human malaria parasite Plasmodium falciparum. We demonstrate that it is involved in the invasion of host erythrocytes, which is critical for establishing infection. In addition, PfPK2 may also be involved in the maturation of the parasite post-invasion. PfPK2 regulates the release of microneme proteins like Apical Membrane Antigen 1 (AMA1), which facilitates the formation of Tight Junction between the merozoite and host erythrocyte- a key step in the process of invasion. Comparative phosphoproteomics studies revealed that PfPK2 may be involved in regulation of several key proteins involved in invasion and signalling. Furthermore, PfPK2 regulates the generation of cGMP and the release of calcium in the parasite, which are key second messengers for the process of invasion. These and other studies have shed light on a novel signalling pathway in which PfPK2 acts as an upstream regulator of important cGMP-calcium signalling, which plays an important role in parasite invasion.

Tympanostomy Tubes or Medical Management for Recurrent Acute Otitis Media.

BACKGROUND: Official recommendations differ regarding tympanostomy-tube placement for children with recurrent acute otitis media. METHODS: We randomly assigned children 6 to 35 months of age who had had at least three episodes of acute otitis media within 6 months, or at least four episodes within 12 months with at least one episode within the preceding 6 months, to either undergo tympanostomy-tube placement or receive medical management involving episodic antimicrobial treatment. The primary outcome was the mean number of episodes of acute otitis media per child-year (rate) during a 2-year period. RESULTS: In our main, intention-to-treat analysis, the rate (±SE) of episodes of acute otitis media per child-year during a 2-year period was 1.48±0.08 in the tympanostomy-tube group and 1.56±0.08 in the medical-management group (P = 0.66). Because 10% of the children in the tympanostomy-tube group did not undergo tympanostomy-tube placement and 16% of the children in the medical-management group underwent tympanostomy-tube placement at parental request, we conducted a per-protocol analysis, which gave corresponding episode rates of 1.47±0.08 and 1.72±0.11, respectively. Among secondary outcomes in the main analysis, results were mixed. Favoring tympanostomy-tube placement were the time to a first episode of acute otitis media, various episode-related clinical findings, and the percentage of children meeting specified criteria for treatment failure. Favoring medical management was children's cumulative number of days with otorrhea. Outcomes that did not show substantial differences included the frequency distribution of episodes of acute otitis media, the percentage of episodes considered to be severe, and antimicrobial resistance among respiratory isolates. Trial-related adverse events were limited to those included among the secondary outcomes of the trial. CONCLUSIONS: Among children 6 to 35 months of age with recurrent acute otitis media, the rate of episodes of acute otitis media during a 2-year period was not significantly lower with tympanostomy-tube placement than with medical management. (Funded by the National Institute on Deafness and Other Communication Disorders and others; ClinicalTrials.gov number, NCT02567825.).

Identifying Children Likely to Benefit From Antibiotics for Acute Sinusitis: A Randomized Clinical Trial.

Importance: The large overlap between symptoms of acute sinusitis and viral upper respiratory tract infection suggests that certain subgroups of children being diagnosed with acute sinusitis, and subsequently treated with antibiotics, derive little benefit from antibiotic use. Objective: To assess if antibiotic therapy could be appropriately withheld in prespecified subgroups. Design, Setting, and Participants: Randomized clinical trial including 515 children aged 2 to 11 years diagnosed with acute sinusitis based on clinical criteria. The trial was conducted between February 2016 and April 2022 at primary care offices affiliated with 6 US institutions and was designed to evaluate whether symptom burden differed in subgroups defined by nasopharyngeal Streptococcus pneumoniae, Haemophilus influenzae, or Moraxella catarrhalis on bacterial culture and by the presence of colored nasal discharge. Interventions: Oral amoxicillin (90 mg/kg/d) and clavulanate (6.4 mg/kg/d) (n = 254) or placebo (n = 256) for 10 days. Main Outcomes and Measures: The primary outcome was symptom burden based on daily symptom scores on a validated scale (range, 0-40) during the 10 days after diagnosis. Secondary outcomes included treatment failure, adverse events including clinically significant diarrhea, and resource use by families. Results: Most of the 510 included children were aged 2 to 5 years (64%), male (54%), White (52%), and not Hispanic (89%). The mean symptom scores were significantly lower in children in the amoxicillin and clavulanate group (9.04 [95% CI, 8.71 to 9.37]) compared with those in the placebo group (10.60 [95% CI, 10.27 to 10.93]) (between-group difference, -1.69 [95% CI, -2.07 to -1.31]). The length of time to symptom resolution was significantly lower for children in the antibiotic group (7.0 days) than in the placebo group (9.0 days) (P = .003). Children without nasopharyngeal pathogens detected did not benefit from antibiotic treatment as much as those with pathogens detected; the between-group difference in mean symptom scores was -0.88 (95% CI, -1.63 to -0.12) in those without pathogens detected compared with -1.95 (95% CI, -2.40 to -1.51) in those with pathogens detected. Efficacy did not differ significantly according to whether colored nasal discharge was present (the between-group difference was -1.62 [95% CI, -2.09 to -1.16] for colored nasal discharge vs -1.70 [95% CI, -2.38 to -1.03] for clear nasal discharge; P = .52 for the interaction between treatment group and the presence of colored nasal discharge). Conclusions: In children with acute sinusitis, antibiotic treatment had minimal benefit for those without nasopharyngeal bacterial pathogens on presentation, and its effects did not depend on the color of nasal discharge. Testing for specific bacteria on presentation may represent a strategy to reduce antibiotic use in this condition. Trial Registration: ClinicalTrials.gov Identifier: NCT02554383.

ImitateDB: A database for domain and motif mimicry incorporating host and pathogen protein interactions.

Molecular mimicry of host proteins by pathogens constitutes a strategy to hijack the host pathways. At present, there is no dedicated resource for mimicked domains and motifs in the host-pathogen interactome. In this work, the experimental host-pathogen (HP) and host-host (HH) protein-protein interactions (PPIs) were collated. The domains and motifs of these proteins were annotated using CD Search and ScanProsite, respectively. Host and pathogen proteins with a shared host interactor and similar domain/motif constitute a mimicry pair exhibiting global structural similarity (domain mimicry pair; DMP) or local sequence motif similarity (motif mimicry pair; MMP). Mimicry pairs are likely to be co-expressed and co-localized. 1,97,607 DMPs and 32,67,568 MMPs were identified in 49,265 experimental HP-PPIs and organized in a web-based resource, ImitateDB ( http://imitatedb.sblab-nsit.net ) that can be easily queried. The results are externally integrated using hyperlinked domain PSSM ID, motif ID, protein ID and PubMed ID. Kinase, UL36, Smc and DEXDc were frequent DMP domains whereas protein kinase C phosphorylation, casein kinase 2 phosphorylation, glycosylation and myristoylation sites were frequent MMP motifs. Novel DMP domains SANT, Tudor, PhoX and MMP motif microbody C-terminal targeting signal, cornichon signature and lipocalin signature were proposed. ImitateDB is a novel resource for identifying mimicry in interacting host and pathogen proteins.

Network analysis of host-pathogen protein interactions in microbe induced cardiovascular diseases.

Large-scale visualization and analysis of HPIs involved in microbial CVDs can provide crucial insights into the mechanisms of pathogenicity. The comparison of CVD associated HPIs with the entire set of HPIs can identify the pathways specific to CVDs. Therefore, topological properties of HPI networks in CVDs and all pathogens was studied using Cytoscape3.5.1. Ontology and pathway analysis were done using KOBAS 3.0. HPIs of Papilloma, Herpes, Influenza A virus as well as Yersinia pestis and Bacillus anthracis among bacteria were predominant in the whole (wHPI) and the CVD specific (cHPI) network. The central viral and secretory bacterial proteins were predicted virulent. The central viral proteins had higher number of interactions with host proteins in comparison with bacteria. Major fraction of central and essential host proteins interacts with central viral proteins. Alpha-synuclein, Ubiquitin ribosomal proteins, TATA-box-binding protein, and Polyubiquitin-C &B proteins were the top interacting proteins specific to CVDs. Signaling by NGF, Fc epsilon receptor, EGFR and ubiquitin mediated proteolysis were among the top enriched CVD specific pathways. DEXDc and HELICc were enriched host mimicry domains that may help in hijacking of cellular machinery by pathogens. This study provides a system level understanding of cardiac damage in microbe induced CVDs.

Corticosteroids to prevent kidney scarring in children with a febrile urinary tract infection: a randomized trial.

BACKGROUND: To evaluate the efficacy of adjuvant systemic corticosteroids in reducing kidney scarring. A previous study suggested that use of adjuvant systemic corticosteroids reduces kidney scarring in children radiologically confirmed to have extensive pyelonephritis. Efficacy of corticosteroids for children with febrile urinary tract infection (UTI) has not been studied. METHODS: Children aged 2 months to 6 years with their first febrile UTI were randomized to corticosteroids or placebo for 3 days (both arms received antimicrobial therapy); kidney scarring was assessed using 99mTc-dimercaptosuccinic acid kidney scan 5-24 months after the initial UTI. RESULTS: We randomized 546 children of which 385 had a UTI and 254 had outcome kidney scans (instead of the 320 planned). Rates of kidney scarring were 9.8% (12/123) and 16.8% (22/131) in the corticosteroid and placebo groups, respectively (p = 0.16), corresponding to an absolute risk reduction of 5.9% (95% confidence interval: - 2.2, 14.1). CONCLUSION: While children randomized to adjuvant corticosteroids tended to develop fewer kidney scars than children who were randomized to receive placebo, a statistically significant difference was not achieved. However, the study was limited by not reaching its intended sample size. CLINICAL TRIAL REGISTRATION: Clinicaltrials.gov , NCT01391793, Registered 7/12/2011 Graphical abstract.

Shortened Antimicrobial Treatment for Acute Otitis Media in Young Children.

BACKGROUND: Limiting the duration of antimicrobial treatment constitutes a potential strategy to reduce the risk of antimicrobial resistance among children with acute otitis media. METHODS: We assigned 520 children, 6 to 23 months of age, with acute otitis media to receive amoxicillin-clavulanate either for a standard duration of 10 days or for a reduced duration of 5 days followed by placebo for 5 days. We measured rates of clinical response (in a systematic fashion, on the basis of signs and symptomatic response), recurrence, and nasopharyngeal colonization, and we analyzed episode outcomes using a noninferiority approach. Symptom scores ranged from 0 to 14, with higher numbers indicating more severe symptoms. RESULTS: Children who were treated with amoxicillin-clavulanate for 5 days were more likely than those who were treated for 10 days to have clinical failure (77 of 229 children [34%] vs. 39 of 238 [16%]; difference, 17 percentage points [based on unrounded data]; 95% confidence interval, 9 to 25). The mean symptom scores over the period from day 6 to day 14 were 1.61 in the 5-day group and 1.34 in the 10-day group (P=0.07); the mean scores at the day-12-to-14 assessment were 1.89 versus 1.20 (P=0.001). The percentage of children whose symptom scores decreased more than 50% (indicating less severe symptoms) from baseline to the end of treatment was lower in the 5-day group than in the 10-day group (181 of 227 children [80%] vs. 211 of 233 [91%], P=0.003). We found no significant between-group differences in rates of recurrence, adverse events, or nasopharyngeal colonization with penicillin-nonsusceptible pathogens. Clinical-failure rates were greater among children who had been exposed to three or more children for 10 or more hours per week than among those with less exposure (P=0.02) and were also greater among children with infection in both ears than among those with infection in one ear (P<0.001). CONCLUSIONS: Among children 6 to 23 months of age with acute otitis media, reduced-duration antimicrobial treatment resulted in less favorable outcomes than standard-duration treatment; in addition, neither the rate of adverse events nor the rate of emergence of antimicrobial resistance was lower with the shorter regimen. (Funded by the National Institute of Allergy and Infectious Diseases and the National Center for Research Resources; ClinicalTrials.gov number, NCT01511107 .).

Reduced-Concentration Clavulanate for Young Children with Acute Otitis Media.

Amoxicillin-clavulanate (A/C) is currently the most effective oral antimicrobial in treating children with acute otitis media (AOM), but the standard dosage of 90 mg amoxicillin/6.4 mg clavulanate/kg of body weight/day commonly causes diarrhea. We examined whether an A/C formulation containing lower concentrations of clavulanate would result in less diarrhea while maintaining plasma levels of amoxicillin and clavulanate adequate to eradicate middle-ear pathogens and to achieve clinical success. We conducted an open-label study in children with AOM who were 6 to 23 months of age. In phase 1, we treated 40 children with a reduced-clavulanate A/C formulation providing 90 mg amoxicillin/3.2 mg clavulanate/kg/day for 10 days. In phase 2, we treated 72 children with the same formulation at a dosage of 80 mg amoxicillin/2.85 mg clavulanate/kg/day for 10 days. We compared the rates of protocol-defined diarrhea (PDD), diaper dermatitis, and AOM clinical response in these children with rates we had reported in children who received the standard A/C regimen, and we obtained plasma levels of amoxicillin and clavulanate at various time points. Outcomes in phase 1 children and in children who had received the standard regimen did not differ significantly. Rates of PDD in children receiving phase 2 and standard regimens were 17% and 26%, respectively (P = 0.10). The corresponding rates of diaper dermatitis were 21% and 33% (P = 0.04) and of AOM treatment failure were 12% and 16% (P = 0.44). Symptomatic responses did not differ significantly between regimens; both gave clavulanate levels sufficient to inhibit ß-lactamase activity. In young children with AOM, clavulanate dosages lower than those currently used may be associated with fewer side effects without reducing clinical efficacy. (This study has been registered at ClinicalTrials.gov under registration no. NCT02630992.).

Structural basis of focal adhesion targeting domain-mediated signaling in cardiac hypertrophy.

The focal adhesion targeting (FAT) domain of focal adhesion kinase (FAK) exists in monomeric closed (c) or arm exchanged (ae) dimeric state. FAT interaction with Grb2 necessitates an intermediate open (o) state that interacts with Grb2 and activates signaling pathways leading to pathological cardiac hypertrophy. Targeted molecular dynamics (TMD) simulation was carried out in order to capture the structure of the intermediate formed by opening of Helix1 (H1) from monomeric cFAT leading to the formation of monomeric aeFAT. During TMD, H1 separated from the four helices bundle of cFAT, completely unfolded and performed a full turn before folding back to a helix inclined at an acute angle to the helical bundle in aeFAT. The entire transition can be described in six distinct intermediate structural stages. The most significant correlation of H1 motion was observed with Loop3 (L3) and is the likely reason for the complete disruption of the FAT interaction with paxillin during the transition. High-affinity analogs of the paxillin LD4 region can be a promising strategy to drive the equilibrium towards cFAT, thus antagonizing FAT-Grb2 association. During transition, the overall shift in orientation of all the four helices rejects paxillin binding and approves Grb2 association. Exposure and ß-turn conformation of the YENV motif (residues 925-928) in oFAT-facilitated phosphorylation and Grb2 binding. Docking, MD simulation and conservation analysis of oFAT-Grb2 complex provided insight into the structural determinants of binding and specificity. Our work provides a structural basis for pharmacological modulation of dynamic conformational changes and interactions of FAT.

Calcium-induced conformational changes of Thrombospondin-1 signature domain: implications for vascular disease.

CONTEXT: Thrombospondin1 (TSP1) participates in numerous signaling pathways critical for vascular physiology and disease. The conserved signature domain of thrombospondin 1 (TSP1-Sig1) comprises three epidermal growth factor (EGF), 13 calcium-binding type 3 thrombospondin (T3) repeats, and one lectin-like module arranged in a stalk-wire-globe topology. TSP1 is known to be present in both calcium-replete (Holo-) and calcium-depleted (Apo-) state, each with distinct downstream signaling effects. OBJECTIVE: To prepare a homology model of TSP1-Sig1 and investigate the effect of calcium on its dynamic structure and interactions. METHODS: A homology model of Holo-TSP1-Sig1 was prepared with TSP2 as template in Swissmodel workspace. The Apo-form of the model was obtained by omitting the bound calcium ions from the homology model. Molecular dynamics (MD) simulation studies (100 ns) were performed on the Holo- and Apo- forms of TSP1 using Gromacs4.6.5. RESULTS AND DISCUSSION: After simulation, Holo-TSP1-Sig1 showed significant reorientation at the interface of the EGF1-2 and EGF2-3 modules. The T3 wire is predicted to show the maximum mobility and deviation from the initial model. In Apo-TSP1-Sig1 model, the T3 repeats unfolded and formed coils with predicted increase in flexibility. Apo-TSP1-Sig1model also predicted the exposure of the binding sites for neutrophil elastase, integrin and fibroblast growth factor 2. We present a structural model and hypothesis for the role of TSP1-Sig1 interactions in the development of vascular disorders. CONCLUSION: The simulated model of the fully calcium-loaded and calcium-depleted TSP1-Sig1 may enable the development of its interactions as a novel therapeutic target for the treatment of vascular diseases.

Utility of sedation for young children undergoing dimercaptosuccinic acid renal scans.

BACKGROUND: No studies have examined whether use of sedation during a Tc-99 m dimercaptosuccinic acid (DMSA) renal scan reduces patient discomfort. OBJECTIVE: To compare discomfort level during a DMSA scan to the discomfort level during other frequently performed uroradiologic tests, and to determine whether use of sedation during a DMSA scan modifies the level of discomfort. MATERIALS AND METHODS: We examined the discomfort level in 798 children enrolled in the Randomized Intervention for children with Vesicoureteral Reflux (RIVUR) and Careful Urinary Tract Infection Evaluation (CUTIE) studies by asking parents to rate their child's discomfort level with each procedure on a scale from 0 to 10. We compared discomfort during the DMSA scan and the DMSA image quality between centers in which sedation was used >90% of the time (sedation centers), centers in which sedation was used <10% of the time (non-sedation centers), and centers in which sedation was used on a case-by-case basis (selective centers). RESULTS: Mean discomfort level was highest for voiding cystourethrogram (6.4), followed by DMSA (4.0), followed by ultrasound (2.4; P<0.0001). Mean discomfort level during the DMSA scan was significantly higher at non-sedation centers than at selective centers (P<0.001). No difference was apparent in discomfort level during the DMSA scan between sedation centers and selective centers (P=0.12), or between the sedation centers and non-sedation centers (P=0.80). There were no differences in the proportion with uninterpretable DMSA scans according to sedation use. CONCLUSION: Selective use of sedation in children 12-36 months of age can reduce the discomfort level experienced during a DMSA scan.

Integrative analysis of ocular complications in atherosclerosis unveils pathway convergence and crosstalk.

Atherosclerosis is a life-threatening disease and a major cause of mortalities worldwide. While many of the atherosclerotic sequelae are reflected as microvascular effects in the eye, the molecular mechanisms of their development is not yet known. In this study, we employed a systems biology approach to unveil the most significant events and key molecular mediators of ophthalmic sequelae caused by atherosclerosis. Literature mining was used to identify the proteins involved in both atherosclerosis and ophthalmic diseases. A protein-protein interaction (PPI) network was prepared using the literature-mined seed nodes. Network topological analysis was carried out using Cytoscape, while network nodes were annotated using database for annotation, visualization and integrated discovery in order to identify the most enriched pathways and processes. Network analysis revealed that mitogen-activated protein kinase 1 (MAPK1) and protein kinase C occur with highest betweenness centrality, degree and closeness centrality, thus reflecting their functional importance to the network. Our analysis shows that atherosclerosis-associated ophthalmic complications are caused by the convergence of neurotrophin signaling pathways, multiple immune response pathways and focal adhesion pathway on the MAPK signaling pathway. The PPI network shares features with vasoregression, a process underlying multiple vascular eye diseases. Our study presents a first clear and composite picture of the components and crosstalk of the main pathways of atherosclerosis-induced ocular diseases. The hub bottleneck nodes highlight the presence of molecules important for mediating the ophthalmic complications of atherosclerosis and contain five established drug targets for future therapeutic modulation efforts.

Treatment of acute otitis media in children under 2 years of age.

BACKGROUND: Recommendations vary regarding immediate antimicrobial treatment versus watchful waiting for children younger than 2 years of age with acute otitis media. METHODS: We randomly assigned 291 children 6 to 23 months of age, with acute otitis media diagnosed with the use of stringent criteria, to receive amoxicillin-clavulanate or placebo for 10 days. We measured symptomatic response and rates of clinical failure. RESULTS: Among the children who received amoxicillin-clavulanate, 35% had initial resolution of symptoms by day 2, 61% by day 4, and 80% by day 7; among children who received placebo, 28% had initial resolution of symptoms by day 2, 54% by day 4, and 74% by day 7 (P=0.14 for the overall comparison). For sustained resolution of symptoms, the corresponding values were 20%, 41%, and 67% with amoxicillin-clavulanate, as compared with 14%, 36%, and 53% with placebo (P=0.04 for the overall comparison). Mean symptom scores over the first 7 days were lower for the children treated with amoxicillin-clavulanate than for those who received placebo (P=0.02). The rate of clinical failure--defined as the persistence of signs of acute infection on otoscopic examination--was also lower among the children treated with amoxicillin-clavulanate than among those who received placebo: 4% versus 23% at or before the visit on day 4 or 5 (P<0.001) and 16% versus 51% at or before the visit on day 10 to 12 (P<0.001). Mastoiditis developed in one child who received placebo. Diarrhea and diaper-area dermatitis were more common among children who received amoxicillin-clavulanate. There were no significant changes in either group in the rates of nasopharyngeal colonization with nonsusceptible Streptococcus pneumoniae. CONCLUSIONS: Among children 6 to 23 months of age with acute otitis media, treatment with amoxicillin-clavulanate for 10 days tended to reduce the time to resolution of symptoms and reduced the overall symptom burden and the rate of persistent signs of acute infection on otoscopic examination. (Funded by the National Institute of Allergy and Infectious Diseases; ClinicalTrials.gov number, NCT00377260.).

Structural effect of the H992D/H418D mutation of angiotensin-converting enzyme in the Indian population: implications for health and disease.

The Non synonymous SNPs (nsSNPs) of the renin-angiotensin-system (RAS) pathway, unique to the Indian population were investigated in view of its importance as an endocrine system. nsSNPs of the RAS pathway genes were mined from the IndiGenome database. Damaging nsSNPs were predicted using SIFT, PredictSNP, SNP and GO, Snap2 and Protein Variation Effect Analyzer. Loss of function was predicted based on protein stability change using I mutant, PremPS and CONSURF. The structural impact of the nsSNPs was predicted using HOPE and Missense3d followed by modeling, refinement, and energy minimization. Molecular Dynamics studies were carried out using Gromacsv2021.1. 23 Indian nsSNPs of the RAS pathway genes were selected for structural analysis and 8 were predicted to be damaging. Further sequence analysis showed that HEMGH zinc binding motif changes to HEMGD in somatic ACE-C domain (sACE-C) H992D and Testis ACE (tACE) H418D resulted in loss of zinc coordination, which is essential for enzymatic activity in this metalloprotease. There was a loss of internal interactions around the zinc coordination residues in the protein structural network. This was also confirmed by Principal Component Analysis, Free Energy Landscape and residue contact maps. Both mutations lead to broadening of the AngI binding cavity. The H992D mutation in sACE-C is likely to be favorable for cardiovascular health, but may lead to renal abnormalities with secondary impact on the heart. H418D in tACE is potentially associated with male infertility.Communicated by Ramaswamy H. Sarma.

Analysis of SARS-CoV-2 genome evolutionary patterns.

The spread of SARS-CoV-2 virus accompanied by public availability of abundant sequence data provides a window for the determination of viral evolutionary patterns. In this study, SARS-CoV-2 genome sequences were collected from seven countries in the period January 2020-December 2022. The sequences were classified into three phases, namely, pre-vaccination, post-vaccination, and recent period. Comparison was performed between these phases based on parameters like mutation rates, selection pressure (dN/dS ratio), and transition to transversion ratios (Ti/Tv). Similar comparisons were performed among SARS-CoV-2 variants. Statistical significance was tested using Graphpad unpaired t-test. The analysis showed an increase in the percent genomic mutation rates post-vaccination and in recent periods across all countries from the pre-vaccination sequences. Mutation rates were highest in NSP3, S, N, and NSP12b before and increased further after vaccination. NSP4 showed the largest change in mutation rates after vaccination. The dN/dS ratios showed purifying selection that shifted toward neutral selection after vaccination. N, ORF8, ORF3a, and ORF10 were under highest positive selection before vaccination. Shift toward neutral selection was driven by E, NSP3, and ORF7a in the after vaccination set. In recent sequences, the largest dN/dS change was observed in E, NSP1, and NSP13. The Ti/Tv ratios decreased with time. C→U and G→U were the most frequent transitions and transversions. However, U→G was the most frequent transversion in recent period. The Omicron variant had the highest genomic mutation rates, while Delta showed the highest dN/dS ratio. Protein-wise dN/dS ratio was also seen to vary across the different variants.IMPORTANCETo the best of our knowledge, there exists no other large-scale study of the genomic and protein-wise mutation patterns during the time course of evolution in different countries. Analyzing the SARS-CoV-2 evolutionary patterns in view of the varying spatial, temporal, and biological signals is important for diagnostics, therapeutics, and pharmacovigilance of SARS-CoV-2.

Positive selection and evolution of dengue type-3 virus in the Indian subcontinent.

BACKGROUND: Dengue virus infection has recently taken endemic proportions in India with dengu type-3 (DEN-3) as a predominant serotype. In this study, we carried out the selection pressure analysis of three critical immunogenic regions of DEN-3. Phylogenetic analysis was then carried out on the positively selected genomic region in the DEN-3 virus strains isolated in the Indian subcontinent over a time span of 25 yr (1984-2008). Bayesian Markov chain Monte Carlo (MCMC) calculation of the substitution rate was carried out for the DEN-3 genotype-III sequences. METHODS: Sequences corresponding to the C-prM, E-NS1 and NS1 sequence regions of DEN-3 strains were taken for the positive selection analysis. The C-prM junction sequences were then used to construct a maximum likelihood (ML) phylogenetic tree. Substitution rates were also calculated under various models of population growth. RESULTS: It was found that codon 86, corresponding to a conserved arginine residue in a crucial T-cell epitope of the C-protein was under significant positive selection. The K86R substitution was found to exist in almost all the Indian strains isolated after 2004. The ML tree constructed from the C-prM junction sequences indicated that strains from the 2006 dengue incidences in Delhi, namely: 04/03/del2006, 05/03/del2006, and 06/03/del2006 were the most rapidly evolving. Substitution rates of a DEN-3 genotype-III sequences from the Indian subcontinent were found to be ~3.0 times higher than those reported from other parts of the world. CONCLUSION: Positive selection in the codon corresponding to R86 of the highly conserved surface C-protein is important in view of its occurrence in a T-cell epitope as well as its strict conservation in all the DEN strains. Phylogenetic analysis of the C-prM junction sequences showed that three strains of 2006 are rapidly evolving. These results were also supported by calculations of the substitution rates. Their significance in the expansion of viral epidemics requires to be investigated.

Role of molecular mimicry in the SARS-CoV-2-human interactome for pathogenesis of cardiovascular diseases: An update to ImitateDB.

Mimicry of host proteins is a strategy employed by pathogens to hijack host functions. Domain and motif mimicry was explored in the experimental and predicted SARS-CoV-2-human interactome. The host first interactor proteins were also added to capture the continuum of the interactions. The domains and motifs of the proteins were annotated using NCBI CD Search and ScanProsite, respectively. Host and pathogen proteins with a common host interactor and similar domain/motif constitute a mimicry pair indicating global structural similarity (domain mimicry pair; DMP) or local sequence similarity (motif mimicry pair; MMP). 593 DMPs and 7,02,472 MMPs were determined. AAA, DEXDc and Macro domains were frequent among DMPs whereas glycosylation, myristoylation and RGD motifs were abundant among MMP. The proteins involved in mimicry were visualised as a SARS-CoV-2 mimicry interaction network. The host proteins were enriched in multiple CVD pathways indicating the role of mimicry in COVID-19 associated CVDs. Bridging nodes were identified as potential drug targets. Approved antihypertensive and anti-inflammatory drugs are proposed for repurposing against COVID-19 associated CVDs. The SARS-CoV-2 mimicry data has been updated in ImitateDB (http://imitatedb.sblab-nsit.net/SARSCoV2Mimicry). Determination of key mechanisms, proteins, pathways, drug targets and repurposing candidates is critical for developing therapeutics for SARS CoV-2 associated CVDs.

3D-QSAR and ADMET studies of morpholino-pyrimidine inhibitors of DprE1 from Mycobacterium tuberculosis.

DprE1 is involved in the synthesis of Mycobacterium tuberculosis cell wall and is a potent drug target for Tuberculosis (TB) treatment. The structure and dynamics of the loops L-I and L-II flanking the inhibitor binding site was studied using molecular dynamics (MD) simulation and MMPBSA in Amber v18. Docking and three-dimensional quantitative structure-activity relationship (3D-QSAR) of 55 Morpholino-pyrimidine (MP) inhibitors was carried out using Autodock v1.2.0 and Forge v10. ADMET analysis was done using SwissADME and pkCSM. All MP inhibitors docked in the DprE1 binding pocket, making contacts with L-II residues. MD studies showed that L-I and L-II unfold in the absence of the inhibitor but fold stably structure with reduced protein motions in the presence of MP-38, the highest affinity inhibitor. This was confirmed by k-means clustering and secondary structure analysis. L-II residues, L317, F320 and R325 contributed most towards the MMPBSA binding free energy of MP-38. A robust field-based 3D-QSAR model showed values of r2train = 0.982, r2test = 0.702 and q2 = 0.516. The MP inhibitor field points were broadly divided into negative electrostatics near the A, B rings and hydrophobic electrostatics near the D, E rings. Addition of negative groups at methanone position and ring B as well as addition of hydrophobic and bulky groups at ring E will improve activity. Highly active compounds 47, 49 and 50 of MP series exhibited highly favourable drug-like properties. SAR and ADMET insights attained from this model will help in the development of active DprE1 inhibitors in future.Communicated by Ramaswamy H. Sarma.