The acidic C-terminal tail of DNA Gyrase of Salmonella enterica serovar Typhi controls DNA relaxation in an acidic environment.

The intracellular bacteria, Salmonella Typhi adapts to acidic conditions in the host cell by resetting the chromosomal DNA topology majorly controlled by DNA Gyrase, a Type II topoisomerase. DNA Gyrase forms a heterodimer A2B2 complex, which manages the DNA supercoiling and relaxation in the cell. DNA relaxation forms a part of the regulatory mechanism to activate the transcription of genes required to survive under hostile conditions. Acid-induced stress attenuates the supercoiling activity of the DNA Gyrase, resulting in DNA relaxation. Salmonella DNA becomes relaxed as the bacteria adapt to the acidified intracellular environment. Despite comprehensive studies on DNA Gyrase, the mechanism to control supercoiling activity needs to be better understood. A loss in supercoiling activity in E. coli was observed upon deletion of the non-conserved acidic C-tail of Gyrase A subunit. Salmonella Gyrase also contains an acidic tail at the C-terminus of Gyrase A, where its deletion resulted in reduced supercoiling activity compared to wild-type Gyrase. Interestingly, we also found that wild-type Gyrase compromises supercoiling activity at acidic pH 2-3, thereby causing DNA relaxation. The absence of a C-tail displayed DNA supercoiling to some extent between pH 2-9. Hence, the C-tail of Gyrase A might be one of the controlling factors that cause DNA relaxation in Salmonella at acidic pH conditions. We propose that the presence of the C-tail of GyraseA causes acid-mediated inhibition of the negative supercoiling activity of Gyrase, resulting in relaxed DNA that attracts DNA-binding proteins for controlling the transcriptional response.

Guidelines for endobronchial ultrasound-transbronchial needle aspiration (EBUS-TBNA): Joint Indian Chest Society (ICS)/Indian Association for Bronchology (IAB) recommendations.

Over the past decade, endobronchial ultrasound-guided transbronchial needle aspiration (EBUS-TBNA) has become an indispensable tool in the diagnostic armamentarium of the pulmonologist. As the expertise with EBUS-TBNA has evolved and several innovations have occurred, the indications for its use have expanded. However, several aspects of EBUS-TBNA are still not standardized. Hence, evidence-based guidelines are needed to optimize the diagnostic yield and safety of EBUS-TBNA. For this purpose, a working group of experts from India was constituted. A detailed and systematic search was performed to extract relevant literature pertaining to various aspects of EBUS-TBNA. The modified GRADE system was used for evaluating the level of evidence and assigning the strength of recommendations. The final recommendations were framed with the consensus of the working group after several rounds of online discussions and a two-day in-person meeting. These guidelines provide evidence-based recommendations encompassing indications of EBUS-TBNA, pre-procedure evaluation, sedation and anesthesia, technical and procedural aspects, sample processing, EBUS-TBNA in special situations, and training for EBUS-TBNA.

Managing nitrate-nitrogen in the intensively drained upper Mississippi River Basin, USA under uncertainty: a perennial path forward.

The upper Mississippi River basin has been identified as the most significant contributor of excessive nutrients to the hypoxic zone in the Gulf of Mexico. The land-use changes from an internally drained prairie-wetland complex to an intensively managed corn-soybean production system drained by subsurface tile drainage system in the north-central Iowa and south-central Minnesota are the primary cause of nutrient loads into the Mississippi River and many other environmental stresses. The present study summarizes the water-quality degradation from land-use change and offers a fuzzy logic-based decision support for assessing degree of suitability of the four recommended perennial plant options for managing water and nitrate-nitrogen export. These options are designed based on landscape position that currently fails to produce high yielding row crops and scale: (1) marginal upland depressions for water storage by planting deep-rooted perennial grasses and fast-growing woody poplar, willow, and alder in poorly drained swales; (2) saturated buffers and/or subtle changes in landscape slope for draining high nitrate-nitrogen subsurface (through multi-species phytoremediation treatment buffers or strips of perennial vegetation); (3) two-stage ditches with linear floodplains planted with perennial grasses; and (4) riparian and in-channel ecologically engineered trees, shrubs, and grasses to better connect meander belt width to frequent peak stream flows at larger scales. When applied throughout a typical (Des Moines Lobe Till) DMLT watershed, each option can have positive cumulative environmental effects. Fuzzy logic enhanced the precision in watershed decision-making by incorporating the uncertainty associated with factors like cost effectiveness, nitrate reduction potential, water quality improvement, and level of acceptance.

Conformational flexibility enables catalysis of phthalate cis-4,5-dihydrodiol dehydrogenase.

Phthalate cis-4,5-dihydrodiol dehydrogenase (PhtC), the second enzyme of the phthalate catabolic pathway, catalyzes the dehydrogenation of cis-4,5-dihydrodiol phthalate (DDP). Here, we report the structural and biochemical characterization of PhtC from Comamonas testosteroni KF1 (PhtCKF1). With biochemical experiments, we have determined the enzyme's catalytic efficiency (kcat/Km) with DDP as 2.6 ± 0.5 M-1s-1, over 10-fold higher than with cis-3,4-dihydrodiol phthalate (CDP). To understand the structural basis of these reactions, the crystal structures of PhtCKF1 in apo-form, the binary complex with NAD+, and the ternary complex with NAD+ and 3-hydroxybenzoate (3HB) were determined. These crystal structures reveal that the binding of 3HB induces a conformational change in the substrate-binding loop. This conformational change causes the opening of the NAD + binding site while trapping the 3HB. The PhtCKF1 crystal structures show that the catalytic domain of PhtCKF1 is larger than that of other structurally characterized homologs and does not align with other cis-diol dehydrogenases. Structural and mutational analysis of the substrate-binding loop residues, Arg164 and Glu167 establish that conformational flexibility of this loop is necessary for positioning the substrate in a catalytically competent pose, as substitution of either of these residues to Ala did not yield the dehydrogenation activity. Further, based on the crystal structures of PhtCKF1 and related structural homologs, a reaction mechanism is proposed. Finally, with the biochemical analysis of a variant M251LPhtCKF1, the broader substrate specificity of this enzyme is explained.

Role of Triple Injection Platelet-Rich Plasma for Osteoarthritis Knees: A 2 Years Follow-Up Study.

PURPOSE: To assess the clinical outcomes of intra-articular Platelet-rich plasma (PRP) injection in knee osteoarthritis (OA) at 2 year follow-up. METHODS: This was a prospective interventional study. 68 cases (105 knees) with Kellgren-Lawrence (KL) grades I, II and III knee OA received 3 intra-articular injections of PRP 1 month apart. The cases were followed up for 2-years. Outcomes were measured using Western Ontario and McMaster Universities Arthritis Index (WOMAC) score and Visual analog scale (VAS) scores. RESULTS: The mean age was 51.7 years. 18 knees had KL grade I, 55 had grade II and 32 had grade III OA. The mean pre-treatment VAS score decreased significantly at 1 year (mean difference - 5.3, p = 0.003) and 2 year follow-up (mean difference - 6, p = 0.007). The mean pre-treatment WOMAC score decreased significantly at 1 year (mean difference - 45.9, p = 0.011) and at 2 year (mean difference - 52.4, p = 0.009). The WOMAC and VAS scores improved significantly from baseline to final follow-up across all KL grades (p = 0.001 and 0.001, 0.009 and 0.007, 0.021 and 0.017 for WOMAC and VAS across KL grade I, II and III, respectively). There was no significant differences in WOMAC and VAS scores between three KL grades at final follow-up (p = 0.17 and 0.09, respectively), although the baseline scores had significant difference (p = 0.001 for both VAS and WOMAC) with worse scores in higher KL grades. The variables like age, sex, BMI, KL grade, baseline VAS and baseline WOMAC did not predict the final VAS and WOMAC scores. CONCLUSION: Triple injection of intra-articular PRP given one month apart significantly relieves the symptoms of knee OA till 2 years in KL grade I, II and III. The improvement in symptoms at 2-year follow-up did not differ across different KL grades.

APOBEC3, TRIM5α, and BST2 polymorphisms in healthy individuals of various populations with special references to its impact on HIV transmission.

AIDS restriction genes (ARGs) like APOBEC3, TRIM5α, and BST2 can act as immunological detectors of the innate protective mechanism of the body. ARGs influence the course of viral pathogenesis and progression of the disease. The infection caused by different viruses including HIV activates the innate immune receptors leading to production of proinflammatory cytokines, interferons and signals that recruit and activate cells involved in the process of inflammation following induction of adaptive immunity. Differential expression of genes involved in viral infection decide the fate and subsequent susceptibility to infection and its clinical outcome. Nevertheless, comprehensive reports on the incidence of genetic polymorphism of APOBEC3s, TRIM5α, and BST-2 in the general population and its association with pathological conditions have not been described well. Therefore, the occurrence of APOBEC3, TRIM5α, and BST2 polymorphism in healthy individuals and its impact on HIV transmission was analyzed. We conducted an extensive search using the several databases including, EMBASE, PubMed (Medline), and Google Scholar. APOBEC3-D, -F, -G, and -H out of the seven human APOBEC3s, help in the control of viral infection. Amongst various restriction factors, TRIM5α and BST-2 also restrict the viral infection followed by the development of the disease. In the current review, a brief account of the polymorphism in the APOBEC3G, TRIM5α, and BST2 genes are explored among different populations along with the interaction of APOBEC3G with Vif protein. Furthermore, this review specifically focus on ARGs polymorphism (APOBEC3G, TRIM5α, and BST2) associated with HIV transmission.

Detecting SARS-CoV-2 RNA prone clusters in a municipal wastewater network using fuzzy-Bayesian optimization model to facilitate wastewater-based epidemiology.

The current pandemic disease coronavirus (COVID-19) has not only become a worldwide health emergency, but also devoured the global economy. Despite appreciable research, identification of targeted populations for testing and tracking the spread of COVID-19 at a larger scale is an intimidating challenge. There is a need to quickly identify the infected individual or community to check the spread. The diagnostic testing done at large-scale for individuals has limitations as it cannot provide information at a swift pace in large populations, which is pivotal to contain the spread at the early stage of its breakouts. Recently, scientists are exploring the presence of SARS-CoV-2 RNA in the faeces discharged in municipal wastewater. Wastewater sampling could be a potential tool to expedite the early identification of infected communities by detecting the biomarkers from the virus. However, it needs a targeted approach to choose optimized locations for wastewater sampling. The present study proposes a novel fuzzy based Bayesian model to identify targeted populations and optimized locations with a maximum probability of detecting SARS-CoV-2 RNA in wastewater networks. Consequently, real time monitoring of SARS-CoV-2 RNA in wastewater using autosamplers or biosensors could be deployed efficiently. Fourteen criteria such as population density, patients with comorbidity, quarantine and hospital facilities, etc. are analysed using the data of 14 lac individuals infected by COVID-19 in the USA. The uniqueness of the proposed model is its ability to deal with the uncertainty associated with the data and decision maker's opinions using fuzzy logic, which is fused with Bayesian approach. The evidence-based virus detection in wastewater not only facilitates focused testing, but also provides potential communities for vaccine distribution. Consequently, governments can reduce lockdown periods, thereby relieving human stress and boosting economic growth.