Publications by authors named "Bhupender Singh"

41 Publications

Base Composition and Host Adaptation of the SARS-CoV-2: Insight From the Codon Usage Perspective.

Front Microbiol 2021 6;12:548275. Epub 2021 Apr 6.

College of Veterinary Medicine, South China Agricultural University, Guangzhou, China.

The novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has been spreading rapidly all over the world and has raised grave concern globally. The present research aims to conduct a robust base compositional analysis of SARS-CoV-2 to reveal adaptive intricacies to the human host. Multivariate statistical analysis revealed a complex interplay of various factors including compositional constraint, natural selection, length of viral coding sequences, hydropathicity, and aromaticity of the viral gene products that are operational to codon usage patterns, with compositional bias being the most crucial determinant. UpG and CpA dinucleotides were found to be highly preferred whereas, CpG dinucleotide was mostly avoided in SARS-CoV-2, a pattern consistent with the human host. Strict avoidance of the CpG dinucleotide might be attributed to a strategy for evading a human immune response. A lower degree of adaptation of SARS-CoV-2 to the human host, compared to Middle East respiratory syndrome (MERS) coronavirus and SARS-CoV, might be indicative of its milder clinical severity and progression contrasted to SARS and MERS. Similar patterns of enhanced adaptation between viral isolates from intermediate and human hosts, contrasted with those isolated from the natural bat reservoir, signifies an indispensable role of the intermediate host in transmission dynamics and spillover events of the virus to human populations. The information regarding avoided codon pairs in SARS-CoV-2, as conferred by the present analysis, promises to be useful for the design of vaccines employing codon pair deoptimization based synthetic attenuated virus engineering.
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http://dx.doi.org/10.3389/fmicb.2021.548275DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8057303PMC
April 2021

Environmental survival of SARS-CoV-2 - A solid waste perspective.

Environ Res 2021 06 26;197:111015. Epub 2021 Mar 26.

Human Molecular Cytogenetics and Stem Cell Laboratory, Department of Human Genetics and Molecular Biology, Bharathiar University, Coimbatore, 641-046, India. Electronic address:

The advent of COVID-19 has kept the whole world on their toes. Countries are maximizing their efforts to combat the virus and to minimize the infection. Since infectious microorganisms may be transmitted by variety of routes, respiratory and facial protection is required for those that are usually transmitted via droplets/aerosols. Therefore this pandemic has caused a sudden increase in the demand for personal protective equipment (PPE) such as gloves, masks, and many other important items since, the evidence of individual-to-individual transmission (through respiratory droplets/coughing) and secondary infection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). But the disposal of these personal protective measures remains a huge question mark towards the environmental impact. Huge waste generation demands proper segregation according to waste types, collection, and recycling to minimize the risk of infection spread through aerosols and attempts to implement measures to monitor infections. Hence, this review focuses on the impact of environment due to improper disposal of these personal protective measures and to investigate the safe disposal methods for these protective measures by using the safe, secure and innovative biological methods such as the use of Artificial Intelligence (AI) and Ultraviolet (UV) lights for killing such deadly viruses.
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http://dx.doi.org/10.1016/j.envres.2021.111015DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7997151PMC
June 2021

Transcriptome Profiling of Associated Extracellular Vesicles Reveals Presence of Small RNA-Cargo.

Front Mol Biosci 2020 13;7:566207. Epub 2021 Jan 13.

Department of Medical Biology, Research Group for Host-Microbe Interactions, UiT The Arctic University of Norway, Tromsø, Norway.

Bacterial extracellular vesicles (EVs) have a vital role in bacterial pathogenesis. However, to date, the small RNA-cargo of EVs released by the opportunistic pathogen has not been characterized. Here, we shed light on the association of small RNAs with EVs secreted by MSSA476 cultured in iron-depleted bacteriologic media supplemented with a subinhibitory dosage of vancomycin to mimic infection condition. Confocal microscopy analysis on intact RNase-treated EVs indicated that RNA is associated with EV particles. Transcriptomic followed by bioinformatics analysis of EV-associated RNA revealed the presence of potential gene regulatory small RNAs and high levels of tRNAs. Among the EV-associated enriched small RNAs were SsrA, RsaC and RNAIII. Our finding invites new insights into the potential role of EV-associated RNA as a modulator of host-pathogen interaction.
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http://dx.doi.org/10.3389/fmolb.2020.566207DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7838569PMC
January 2021

Immuno-Informatics Quest against COVID-19/SARS-COV-2: Determining Putative T-Cell Epitopes for Vaccine Prediction.

Infect Disord Drug Targets 2020 Sep 21. Epub 2020 Sep 21.

School of Bioengineering and Biosciences, Lovely Professional University, Phagwara, Punjab. India.

Background: Since, December 2019 a novel coronavirus, SARS-CoV-2 has caused global public health issue after being reported for the first time in Wuhan province of China. So far, there have been approximately 14.8 million confirmed cases and 0.614 million deaths due to the SARS-CoV-2 infection globally, and still numbers are increasing. Although, the virus has caused a global public health concern, no effective treatment has been developed.

Objective: One of the strategies to combat the COVID-19 disease caused by SARS-CoV-2 is development of vaccines that can make humans immune to these infections. Considering this approach, in this study an attempt has been made to design epitope based vaccine for combatting COVID-19 disease by analyzing the complete proteome of the virus by using immuno-informatics tools.

Methods: The protein sequence of the SARS-CoV-2 was retrieved and the individual proteins were checked for their allergic potential. Then, from non-allergen proteins antigenic epitopes were identified that could bind with MHCII molecules. The epitopes were modeled and docked to predict the interaction with MHCII molecules. The stability of the epitopeMHCII complex was further analyzed by performing molecular dynamic simulation study. The selected vaccine candidates were also analyzed for their global population coverage and conservancy among SARS related coronavirus species.

Results: The study has predicted 5 peptide molecules that can act as potential candidate for epitope based vaccine development. Among the 5 selected epitopes, the peptide LRARSVSPK can be the most potent epitope because of its high geometric shape complementarity score, low ACE and very high response to it by the world population (81.81% global population coverage). Further, molecular dynamic simulation analysis indicated the formation of stable epitope-MHCII complex. The epitope LRARSVSPK was also found to be highly conserved among the SARS-CoV-2 isolated from different countries.

Conclusion: The study has predicted T-cell epitopes that can elicit robust immune response in global human population and act as potential vaccine candidates. However, the ability of these epitopes to act as vaccine candidate needs to be validated in wet lab studies.
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http://dx.doi.org/10.2174/1871526520666200921154149DOI Listing
September 2020

Evaluation of Normal Morphology of Mandibular Condyle: A Radiographic Survey.

J Clin Imaging Sci 2020 17;10:51. Epub 2020 Aug 17.

Department of Oral Medicine and Radiology, Government Dental College, Kozhikode (Calicut), Kerala, India.

Objectives: The temporomandibular joint (TMJ) is a complex, highly specialized joint. Along with the teeth, these joints are considered to be a "tri-joint complex." Mandibular condyle morphology is characterized by a rounded bone projection with an upper biconvex and oval surface in axial plane. Anatomical knowledge of the TMJ is one of the basic foundations of clinical practice, allowing the understanding of TMJ pathologies and fabrication of condylar prostheses. The cross-sectional descriptive study was undertaken to evaluate normal variation in the condylar morphology on radiographs in persons without TMJ symptomatology and its relation to age, gender, dentition status, chewing habits, parafunctional habits, history of orthodontic treatment, and denture wearing was assessed.

Material And Methods: A total of 350 subjects without TMJ symptomatology included in the study were further grouped by age, gender, dentition status, chewing habits, parafunctional habits, history of orthodontic treatment, and denture wearing history. Panoramic radiograph was taken for the assessment of condylar morphology.

Results: A significant association between dentition status and bilaterally similar condylar morphology was noticed. Bilaterally similar condyles were seen in 81.4% of subjects. Round-shaped condyles were seen in 176 (62%) persons. Loss of bilateral occlusion tends to alter the condylar morphology. Association between normal chewing habits and bilaterally similar condyle shapes was significant.

Conclusion: The study describes the normal morphology of mandibular condyles in a population attending the tertiary dental care center, Kozhikode. The dentition status and chewing habits of individuals had a significant role in determining condylar morphology.
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http://dx.doi.org/10.25259/JCIS_84_2020DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7451156PMC
August 2020

Drug-target binding quantitatively predicts optimal antibiotic dose levels in quinolones.

PLoS Comput Biol 2020 08 14;16(8):e1008106. Epub 2020 Aug 14.

Department of Pharmacy, Faculty of Health Sciences, UiT-The Arctic University of Norway, Tromsø, Norway.

Antibiotic resistance is rising and we urgently need to gain a better quantitative understanding of how antibiotics act, which in turn would also speed up the development of new antibiotics. Here, we describe a computational model (COMBAT-COmputational Model of Bacterial Antibiotic Target-binding) that can quantitatively predict antibiotic dose-response relationships. Our goal is dual: We address a fundamental biological question and investigate how drug-target binding shapes antibiotic action. We also create a tool that can predict antibiotic efficacy a priori. COMBAT requires measurable biochemical parameters of drug-target interaction and can be directly fitted to time-kill curves. As a proof-of-concept, we first investigate the utility of COMBAT with antibiotics belonging to the widely used quinolone class. COMBAT can predict antibiotic efficacy in clinical isolates for quinolones from drug affinity (R2>0.9). To further challenge our approach, we also do the reverse: estimate the magnitude of changes in drug-target binding based on antibiotic dose-response curves. We overexpress target molecules to infer changes in antibiotic-target binding from changes in antimicrobial efficacy of ciprofloxacin with 92-94% accuracy. To test the generality of our approach, we use the beta-lactam ampicillin to predict target molecule occupancy at MIC from antimicrobial action with 90% accuracy. Finally, we apply COMBAT to predict antibiotic concentrations that can select for resistance due to novel resistance mutations. Using ciprofloxacin and ampicillin as well defined test cases, our work demonstrates that drug-target binding is a major predictor of bacterial responses to antibiotics. This is surprising because antibiotic action involves many additional effects downstream of drug-target binding. In addition, COMBAT provides a framework to inform optimal antibiotic dose levels that maximize efficacy and minimize the rise of resistant mutants.
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http://dx.doi.org/10.1371/journal.pcbi.1008106DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7449454PMC
August 2020

Natural compounds from spp. as possible therapeutic candidates against SARS-CoV-2: An investigation.

J Biomol Struct Dyn 2020 Jun 19:1-12. Epub 2020 Jun 19.

Department of Biotechnology, Lovely Professional University, Phagwara, India.

The COVID-19 pandemic caused by the novel coronavirus SARS-CoV-2 has rattled global public health, with researchers struggling to find specific therapeutic solutions. In this context, the present study employed an approach to assess the inhibitory potential of the phytochemicals obtained from GC-MS analysis of twelve species against the imperative spike protein, main protease enzyme M and RNA-dependent RNA polymerase (RdRp) of SARS-CoV-2. An extensive molecular docking investigation of the phytocompounds at the active binding pockets of the viral proteins revealed promising inhibitory potential of the phytochemicals taraxerol, friedelin and stigmasterol. Decent physicochemical attributes of the compounds in accordance with Lipinski's rule of five and Veber's rule further established them as potential therapeutic candidates against SARS-CoV-2. Molecular mechanics-generalized Born surface area (MM-GBSA) binding free energy estimation revealed that taraxerol was the most promising candidate displaying the highest binding efficacy with all the concerned SARS-CoV-2 proteins included in the present analysis. Our observations were supported by robust molecular dynamics simulations of the complexes of the viral proteins with taraxerol for a timescale of 40 nanoseconds. It was striking to note that taraxerol exhibited better binding energy scores with the concerned viral proteins than the drugs that are specifically targeted against them. The present results promise to provide new avenues to further evaluate the potential of the phytocompound taraxerol and towards its successful deployment as a SARS-CoV-2 inhibitor and combat the catastrophic COVID-19.Communicated by Ramaswamy H. Sarma.
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http://dx.doi.org/10.1080/07391102.2020.1780947DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7309333PMC
June 2020

Cellular signalling pathways mediating the pathogenesis of chronic inflammatory respiratory diseases: an update.

Inflammopharmacology 2020 Aug 18;28(4):795-817. Epub 2020 Mar 18.

Discipline of Pharmacy, Graduate School of Health, University of Technology Sydney, Ultimo, NSW, 2007, Australia.

Respiratory disorders, especially non-communicable, chronic inflammatory diseases, are amongst the leading causes of mortality and morbidity worldwide. Respiratory diseases involve multiple pulmonary components, including airways and lungs that lead to their abnormal physiological functioning. Several signaling pathways have been reported to play an important role in the pathophysiology of respiratory diseases. These pathways, in addition, become the compounding factors contributing to the clinical outcomes in respiratory diseases. A range of signaling components such as Notch, Hedgehog, Wingless/Wnt, bone morphogenetic proteins, epidermal growth factor and fibroblast growth factor is primarily employed by these pathways in the eventual cascade of events. The different aberrations in such cell-signaling processes trigger the onset of respiratory diseases making the conventional therapeutic modalities ineffective. These challenges have prompted us to explore novel and effective approaches for the prevention and/or treatment of respiratory diseases. In this review, we have attempted to deliberate on the current literature describing the role of major cell signaling pathways in the pathogenesis of pulmonary diseases and discuss promising advances in the field of therapeutics that could lead to novel clinical therapies capable of preventing or reversing pulmonary vascular pathology in such patients.
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http://dx.doi.org/10.1007/s10787-020-00698-3DOI Listing
August 2020

Evaluation of Different Preosteotomy Determinants as Affecting the Success of Implant Therapy: A "CBCT"-based Clinical Study.

J Contemp Dent Pract 2019 Oct 1;20(10):1212-1216. Epub 2019 Oct 1.

Department of Prosthodontics, JCD Dental College, Sirsa, Haryana, India.

Aim: The success of implant therapy is of greatest concern for clinicians because a minor negligence can lead to ultimate failure of treatment. However, comprehensive and precise treatment planning can ensure high success rate of implant therapy. Cone-beam computed tomography (CBCT) is an innovation that allows clinicians to explore all related factors in details. This study was conducted to evaluate different preosteotomy determinants as affecting the success of implant therapy in the maxillary anterior region using CBCT.

Materials And Methods: This study was conducted on 98 partially or complete denture patients willing for artificial replacement of their missing teeth by implant or implant over-denture. Demographic details of participating patients were collected. Furthermore, comprehensive local examination was also done to finalize the site of placement of implant. Cone-beam computed tomography was attempted in all patients for determining accurate implant location, status of bone, and other interrelated determinants of implant success. Cone-beam computed tomography was also prescribed for patients so as to have presurgical idea of implant dimensions as shown in virtual placement of implant. To rule out any interobserver bias, the interpretations of CBCT images were completed by two independent experienced observers.

Results: In the 98 studied patients, 61 were males and 37 were females. The study was restricted to the maxillary anterior region only. The studied preosteotomy determinants were available bone height and width in the edentulous region from ridge crest up to the maxillary sinus floor or the nasal fossa floor. A total of 107 implants were placed virtually (on CBCT) in the maxillary anterior region and compared quantitatively in postosteotomy phases. Implant placement sites were the maxillary central incisor region (39), the lateral incisor region (31), and canine (37). Authors also noticed that the relative length and width of virtual implant remained unaffected in 97% of the cases.

Conclusion: Cone-beam computed tomography showed accurate status of various presurgical determinants like trabeculae, peri-ridiculer pathology, and amount of horizontal and vertical bone losses. Hence, it was further concluded that all these presurgical determinants greatly affect the final success rate of implant therapy. It is therefore deemed necessary to judiciously consider and clinically manage such factors before attempting implant in the maxillary anterior region.

Clinical Significance: Presurgical evaluation of factors associated with implant dimensions significantly assists clinicians in deciding the finest treatment option. All additional information provided by CBCT genuinely led to a change in the treatment plan that provides enhanced clinical outcome with lesser postoperative complications. How to cite this article: Jain S, Kapoor K, Sethi K, Evaluation of Different Preosteotomy Determinants as Affecting the Success of Implant Therapy: A "CBCT"-based Clinical Study. J Contemp Dent Pract 2019;20(10):1212-1216.
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October 2019

Red complex: Polymicrobial conglomerate in oral flora: A review.

J Family Med Prim Care 2019 Nov 15;8(11):3480-3486. Epub 2019 Nov 15.

Department of Periodontics, BRS Dental College and Hospital, Haryana, India.

Oral diseases are the complex host responses composed of a broad array of inflammatory cells, and cytokines, chemokines, and mediators derived from the cells resident in the gingival tissues, as well as from the emigrating inflammatory cells. A chronic polymicrobial challenge to the local host tissues triggers this response, which under certain circumstances, and in a subset of the population, leads to the progressing soft and hard tissue destruction that characterizes periodontitis. The red complex has been proposed as a pathogenic consortium, consisting of , and . This review has attempted to examine the virulence potential and determinants of these commensal opportunists.
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http://dx.doi.org/10.4103/jfmpc.jfmpc_759_19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6881954PMC
November 2019

Inequalities in healthcare access: how a man with exstrophy in rural India coped.

BMJ Case Rep 2018 Oct 31;2018. Epub 2018 Oct 31.

Department of Urology, King George's Medical University, Lucknow, Uttar Pradesh, India.

We report a case of a 22-year-old man with adult exstrophy. The patient made a self-made urine collection device, which helped him to lead a normal life, carrying out his routine as well as occupational activities smoothly. This patient is a prime example of inequalities in healthcare distribution in low-income and middle-income countries. He was never taken to a proper medical centre to correct his condition nor was his mother ever given proper antenatal healthcare access. His background of being a poor person from rural India highlights the problems of inequalities in healthcare access.
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http://dx.doi.org/10.1136/bcr-2018-226857DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6254381PMC
October 2018

Flagella-mediated secretion of a novel cytotoxin affecting both vertebrate and invertebrate hosts.

Commun Biol 2018 7;1:59. Epub 2018 Jun 7.

Department of Molecular Biology and The Laboratory for Molecular Infection Medicine Sweden (MIMS), Umeå University, SE-90187, Umeå, Sweden.

Using as an infection host model for predator interactions, we discovered a bacterial cytotoxin, MakA, whose function as a virulence factor relies on secretion via the flagellum channel in a proton motive force-dependent manner. The MakA protein is expressed from the polycistronic (otility-ssociated lling factor) operon. Bacteria expressing induced dramatic changes in intestinal morphology leading to a defecation defect, starvation and death in . The Mak proteins also promoted colonization of the zebrafish gut causing lethal infection. A structural model of purified MakA at 1.9 Å resolution indicated similarities to members of a superfamily of bacterial toxins with unknown biological roles. Our findings reveal an unrecognized role for flagella in cytotoxin export that may contribute both to environmental spread of the bacteria by promoting survival and proliferation in encounters with predators, and to pathophysiological effects during infections.
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http://dx.doi.org/10.1038/s42003-018-0065-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6123715PMC
June 2018

Fluorescent Sensing Assay for Trinitrotoluene Using Fluorescein Isothiocyanate Conjugated Mesoporous MCM-41 Particles.

J Nanosci Nanotechnol 2018 10;18(10):6838-6849

In the present study, Trinitrotoluene (TNT) has been detected by the formation of Meisenheimer complex using Fluorescein isothiocyanate (FITC) dye loaded Mesoporous silica particles (MCM-41). FITC dye loaded mesoporous silica particles (MCM-41/FITC) have been synthesized using (3-Aminopropyl)trimethoxysilane, APTMS (λex = 490 nm and λem = 512 nm). TNT forms Meisenheimer complex with the amine group of APTMS present on MCM-41 particles. The loading of FITC in the pores of MCM-41 particles has been confirmed by different advanced characterization techniques. The average diameter of mesoporous MCM-41 particles was found about 130 nm. Pore volume is observed to decrease from 1.06 cm3/g to 0.49 cm3/g after FITC loading. The selective detection of TNT up to 0.1 ppb level makes MCM-41/FITC particles a potential sensing material for TNT detection.
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http://dx.doi.org/10.1166/jnn.2018.15456DOI Listing
October 2018

A drag force interpolation model for capsule-shaped cells in fluid flows near a surface.

Microbiology (Reading) 2018 04 6;164(4):483-494. Epub 2018 Mar 6.

Department of Physics, Umeå University, 901 87 Umeå, Sweden.

We report an interpolation model to calculate the hydrodynamic force on tethered capsule-shaped cells in micro-fluidic flows near a surface. Our model is based on numerical solutions of the full Navier-Stokes equations for capsule-shaped objects considering their geometry, aspect ratio and orientation with respect to fluid flow. The model reproduced the results from computational fluid dynamic simulations, with an average error of <0.15 % for objects with an aspect ratio up to 5, and the model exactly reproduced the Goldman approximation of spherical objects close to a surface. We estimated the hydrodynamic force imposed on tethered Escherichia coli cells using the interpolation model and approximate models found in the literature, for example, one that assumes that E. coli is ellipsoid shaped. We fitted the 2D-projected area of a capsule and ellipsoid to segmented E. coli cells. We found that even though an ellipsoidal shape is a reasonable approximation of the cell shape, the capsule gives 4.4 % better agreement, a small difference that corresponds to 15 % difference in hydrodynamic force. In addition, we showed that the new interpolation model provides a significantly better agreement compared to estimates from commonly used models and that it can be used as a fast and accurate substitute for complex and computationally heavy fluid dynamic simulations. This is useful when performing bacterial adhesion experiments in parallel-plate flow channels. We include a MATLAB script that can track cells in a video time-series and estimate the hydrodynamic force using our interpolation formula.
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http://dx.doi.org/10.1099/mic.0.000624DOI Listing
April 2018

Cervical Vagal Schwannoma: Anesthetic Concerns.

Anesth Essays Res 2017 Jan-Mar;11(1):254-256

Department of Anaesthesia, Vardhman Mahaveer Medical College and Safdarjung Hospital, New Delhi, India.

Schwannomas arising from cervical vagus nerve is an extremely rare benign neoplasm. The majority of patients present with insidiously growing lateral neck mass without neurological deficit. We report a case of symptomatic cervical vagal nerve schwannoma in a 32-year-old female. Complete surgical resection is the treatment of choice. However, anesthetic management of these tumors can be challenging. We describe here the complications experienced during the management of the patient.
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http://dx.doi.org/10.4103/0259-1162.183163DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5341630PMC
March 2017

Antibodies Damage the Resilience of Fimbriae, Causing Them To Be Stiff and Tangled.

J Bacteriol 2017 01 13;199(1). Epub 2016 Dec 13.

Department of Physics, Umeå University, Umeå, Sweden

As adhesion fimbriae are a major virulence factor for many pathogenic Gram-negative bacteria, they are also potential targets for antibodies. Fimbriae are commonly required for initiating the colonization that leads to disease, and their success as adhesion organelles lies in their ability to both initiate and sustain bacterial attachment to epithelial cells. The ability of fimbriae to unwind and rewind their helical filaments presumably reduces their detachment from tissue surfaces with the shear forces that accompany significant fluid flow. Therefore, the disruption of functional fimbriae by inhibiting this resilience should have high potential for use as a vaccine to prevent disease. In this study, we show that two characteristic biomechanical features of fimbrial resilience, namely, the extension force and the extension length, are significantly altered by the binding of antibodies to fimbriae. The fimbriae that were studied are normally expressed on enterotoxigenic Escherichia coli, which are a major cause of diarrheal disease. This alteration in biomechanical properties was observed with bivalent polyclonal antifimbrial antibodies that recognize major pilin subunits but not with the Fab fragments of these antibodies. Thus, we propose that the mechanism by which bound antibodies disrupt the uncoiling of natural fimbria under force is by clamping together layers of the helical filament, thereby increasing their stiffness and reducing their resilience during fluid flow. In addition, we propose that antibodies tangle fimbriae via bivalent binding, i.e., by binding to two individual fimbriae and linking them together. Use of antibodies to disrupt physical properties of fimbriae may be generally applicable to the large number of Gram-negative bacteria that rely on these surface-adhesion molecules as an essential virulence factor.

Importance: Our study shows that the resiliency of colonization factor antigen I (CFA/I) and coli surface antigen 2 (CS2) fimbriae, which are current targets for vaccine development, can be compromised significantly in the presence of antifimbrial antibodies. It is unclear how the humoral immune system specifically interrupts infection after the attachment of enterotoxigenic Escherichia coli (ETEC) to the epithelial surface. Our study indicates that immunoglobulins, in addition to their well-documented role in adaptive immunity, can mechanically damage the resilience of fimbriae of surface-attached ETEC, thereby revealing a new mode of action. Our data suggest a mechanism whereby antibodies coat adherent and free-floating bacteria to impede fimbrial resilience. Further elucidation of this possible mechanism is likely to inform the development and refinement of preventive vaccines against ETEC diarrhea.
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http://dx.doi.org/10.1128/JB.00665-16DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5165098PMC
January 2017

Detecting Bacterial Surface Organelles on Single Cells Using Optical Tweezers.

Langmuir 2016 05 28;32(18):4521-9. Epub 2016 Apr 28.

Department of Physics, and ‡Department of Chemistry, Umeå University , 901 87 Umeå, Sweden.

Bacterial cells display a diverse array of surface organelles that are important for a range of processes such as intercellular communication, motility and adhesion leading to biofilm formation, infections, and bacterial spread. More specifically, attachment to host cells by Gram-negative bacteria are mediated by adhesion pili, which are nanometers wide and micrometers long fibrous organelles. Since these pili are significantly thinner than the wavelength of visible light, they cannot be detected using standard light microscopy techniques. At present, there is no fast and simple method available to investigate if a single cell expresses pili while keeping the cell alive for further studies. In this study, we present a method to determine the presence of pili on a single bacterium. The protocol involves imaging the bacterium to measure its size, followed by predicting the fluid drag based on its size using an analytical model, and thereafter oscillating the sample while a single bacterium is trapped by an optical tweezer to measure its effective fluid drag. Comparison between the predicted and the measured fluid drag thereby indicate the presence of pili. Herein, we verify the method using polymer coated silica microspheres and Escherichia coli bacteria expressing adhesion pili. Our protocol can in real time and within seconds assist single cell studies by distinguishing between piliated and nonpiliated bacteria.
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http://dx.doi.org/10.1021/acs.langmuir.5b03845DOI Listing
May 2016

Antibody-mediated disruption of the mechanics of CS20 fimbriae of enterotoxigenic Escherichia coli.

Sci Rep 2015 Sep 28;5:13678. Epub 2015 Sep 28.

Department of Physics, Umeå University, SE-901 87 Umeå, Sweden.

Preventive vaccines against enterotoxigenic Escherichia coli (ETEC) are being developed, many of which target common fimbrial colonization factors as the major constituent, based on empirical evidence that these function as protective antigens. Particularly, passive oral administration of ETEC anti-fimbrial antibodies prevent ETEC diarrhea. Little is, however, known regarding the specific mechanisms by which intestinal antibodies against ETEC fimbriae function to prevent disease. Using coli surface antigen 20 (CS20) fimbriae as a model ETEC colonization factor, we show using force spectroscopy that anti-fimbrial antibodies diminish fimbrial elasticity by inhibiting their natural capacity to unwind and rewind. In the presence of anti-CS20 antibodies the force required to unwind a single fimbria was increased several-fold and the extension length was shortened several-fold. Similar measurements in the presence of anti-CS20 Fab fragments did not show any effect, indicating that bivalent antibody binding is required to reduce fimbrial elasticity. Based on these findings, we propose a model for an in-vivo mechanism whereby antibody-mediated disruption of the biomechanical properties of CS20 fimbriae impedes sustained adhesion of ETEC to the intestinal mucosal surface. Further elucidation of the role played by intestinal antibodies in mechanical disruption of fimbrial function may provide insights relevant to ETEC vaccine development.
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http://dx.doi.org/10.1038/srep13678DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4585931PMC
September 2015

Biomechanical and structural features of CS2 fimbriae of enterotoxigenic Escherichia coli.

Biophys J 2015 Jul;109(1):49-56

Department of Physics, Umeå University, Umeå, Sweden; Umeå Centre for Microbial Research (UCMR), Umeå University, Umeå, Sweden. Electronic address:

Enterotoxigenic Escherichia coli (ETEC) are a major cause of diarrhea worldwide, and infection of children in under-developed countries often leads to high mortality rates. Isolated ETEC expresses a plethora of colonization factors (fimbriae/pili), of which CFA/I and CFA/II, which are assembled via the alternate chaperone pathway (ACP), are among the most common. Fimbriae are filamentous structures whose shafts are primarily composed of helically arranged single pilin-protein subunits, with a unique biomechanical ability to unwind and rewind. A sustained ETEC infection, under adverse conditions of dynamic shear forces, is primarily attributed to this biomechanical feature of ETEC fimbriae. Recent understanding about the role of fimbriae as virulence factors points to an evolutionary adaptation of their structural and biomechanical features. In this work, we investigated the biophysical properties of CS2 fimbriae from the CFA/II group. Homology modeling of its major structural subunit, CotA, reveals structural clues related to the niche in which they are expressed. Using optical-tweezers force spectroscopy, we found that CS2 fimbriae unwind at a constant force of 10 pN and have a corner velocity (i.e., the velocity at which the force required for unwinding rises exponentially with increased speed) of 1300 nm/s. The biophysical properties of CS2 fimbriae assessed in this work classify them into a low-force unwinding group of fimbriae together with the CFA/I and CS20 fimbriae expressed by ETEC strains. The three fimbriae are expressed by ETEC, colonize in similar gut environments, and exhibit similar biophysical features, but differ in their biogenesis. Our observation suggests that the environment has a strong impact on the biophysical characteristics of fimbriae expressed by ETEC.
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http://dx.doi.org/10.1016/j.bpj.2015.05.022DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4572467PMC
July 2015

Structure and function of enterotoxigenic Escherichia coli fimbriae from differing assembly pathways.

Mol Microbiol 2015 Jan 27;95(1):116-26. Epub 2014 Nov 27.

Department of Physics, Umeå University, SE-901 87, Umeå, Sweden.

Pathogenic enterotoxigenic Escherichia coli (ETEC) are the major bacterial cause of diarrhea in young children in developing countries and in travelers, causing significant mortality in children. Adhesive fimbriae are a prime virulence factor for ETEC, initiating colonization of the small intestinal epithelium. Similar to other Gram-negative bacteria, ETEC express one or more diverse fimbriae, some assembled by the chaperone-usher pathway and others by the alternate chaperone pathway. Here, we elucidate structural and biophysical aspects and adaptations of each fimbrial type to its respective host niche. CS20 fimbriae are compared with colonization factor antigen I (CFA/I) fimbriae, which are two ETEC fimbriae assembled via different pathways, and with P-fimbriae from uropathogenic E. coli. Many fimbriae unwind from their native helical filament to an extended linear conformation under force, thereby sustaining adhesion by reducing load at the point of contact between the bacterium and the target cell. CFA/I fimbriae require the least force to unwind, followed by CS20 fimbriae and then P-fimbriae, which require the highest unwinding force. We conclude from our electron microscopy reconstructions, modeling and force spectroscopy data that the target niche plays a central role in the biophysical properties of fimbriae that are critical for bacterial pathophysiology.
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http://dx.doi.org/10.1111/mmi.12847DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4275653PMC
January 2015

Surgical clips in the common bile duct suspected on endoscopic ultrasound and confirmed on endoscopic retrograde cholangiopancreatography.

Endosc Ultrasound 2013 Jul;2(3):157-8

Jaswant Rai Speciality Hospital, Uttar Pradesh, India.

We report a 56-year-old lady presented with cholangitis due to post-surgical clip migration associated choledocholithiasis. She under-went laparoscopic cholecystectomy 2 years ago. Endoscopic ultrasound revealed linear nature of hyper-echoic lesion casting acoustic shadow in the distal common bile duct suggestive of metallic surgical clip, which was later confirmed by endoscopic cholangiography.
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http://dx.doi.org/10.7178/eus.06.008DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4062261PMC
July 2013

Cefuroxime axetil loaded solid lipid nanoparticles for enhanced activity against S. aureus biofilm.

Colloids Surf B Biointerfaces 2014 Sep 18;121:92-8. Epub 2014 Apr 18.

Department of Pharmaceutics, Indian Institute of Technology (Banaras Hindu University), Varanasi 221 005, India. Electronic address:

The present research work is focused on the development of solid lipid nanoparticles of cefuroxime axetil (CA-SLN) for its enhanced inhibitory activity against Staphylococcus aureus produced biofilm. CA-SLN was prepared by solvent emulsification/evaporation method using single lipid (stearic acid (SA)) and binary lipids (SA and tristearin (TS)). Process variables such as volume of dispersion medium, concentration of surfactant, homogenization speed and time were optimized. The prepared SLN were characterized for encapsulation efficiency, drug polymer interaction studies (DSC and FT-IR), shape and surface morphology (SEM and AFM), in vitro drug release, stability studies and in vitro anti biofilm activity against S. aureus biofilm. Among the process variables, increased volume of dispersion medium, homogenization speed and time led to increase in particle size whereas increase in surfactant concentration decreased the particle size. SLN prepared using binary lipids exhibited higher entrapment efficiency than the single lipid. DSC and FT-IR studies showed no incompatible interaction between drug and excipients. CA-SLN showed two folds higher anti-biofilm activity in vitro than pristine CA against S. aureus biofilm.
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http://dx.doi.org/10.1016/j.colsurfb.2014.03.046DOI Listing
September 2014

Organization of ribosomes and nucleoids in Escherichia coli cells during growth and in quiescence.

J Biol Chem 2014 Apr 5;289(16):11342-11352. Epub 2014 Mar 5.

Department of Cell and Molecular Biology, Uppsala University, Box-596, BMC, 75124, Uppsala, Sweden. Electronic address:

We have examined the distribution of ribosomes and nucleoids in live Escherichia coli cells under conditions of growth, division, and in quiescence. In exponentially growing cells translating ribosomes are interspersed among and around the nucleoid lobes, appearing as alternative bands under a fluorescence microscope. In contrast, inactive ribosomes either in stationary phase or after treatment with translation inhibitors such as chloramphenicol, tetracycline, and streptomycin gather predominantly at the cell poles and boundaries with concomitant compaction of the nucleoid. However, under all conditions, spatial segregation of the ribosomes and the nucleoids is well maintained. In dividing cells, ribosomes accumulate on both sides of the FtsZ ring at the mid cell. However, the distribution of the ribosomes among the new daughter cells is often unequal. Both the shape of the nucleoid and the pattern of ribosome distribution are also modified when the cells are exposed to rifampicin (transcription inhibitor), nalidixic acid (gyrase inhibitor), or A22 (MreB-cytoskeleton disruptor). Thus we conclude that the intracellular organization of the ribosomes and the nucleoids in bacteria are dynamic and critically dependent on cellular growth processes (replication, transcription, and translation) as well as on the integrity of the MreB cytoskeleton.
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http://dx.doi.org/10.1074/jbc.M114.557348DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4036271PMC
April 2014

P-fimbriae in the presence of anti-PapA antibodies: new insight of antibodies action against pathogens.

Sci Rep 2013 Dec 2;3:3393. Epub 2013 Dec 2.

1] Department of Physics, Umeå University, SE-901 87 Umeå, Sweden [2].

Uropathogenic strains of Escherichia coli establish urinary tract infections by attaching to host epithelial cells using adhesive organelles called fimbriae. Fimbriae are helix-like structures with a remarkable adaptability, offering safeguarding for bacteria exposed to changing fluid forces in the urinary tract. We challenged this property of P-fimbriae by cross-linking their subunits with shaft-specific antibodies and measuring the corresponding force response at a single organelle level. Our data show compromised extension and rewinding of P-fimbriae in the presence of antibodies and reduced fimbrial elasticity, which are important properties of fimbriae contributing to the ability of bacteria to cause urinary tract infections. The reduced elasticity found by cross-linking fimbrial subunits could thus be another assignment for antibodies; in addition to marking bacteria as foreign, antibodies physically compromise fimbrial function. We suggest that our assay and results will be a starting point for further investigations aimed at inhibiting sustained bacterial adhesion by antibodies.
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http://dx.doi.org/10.1038/srep03393DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3848023PMC
December 2013

Optoelectronic properties of dual emitting RNA-mediated colloidal PbSe nanostructures.

Dalton Trans 2013 Aug 4;42(32):11455-64. Epub 2013 Jul 4.

Department of Chemistry, Indian Institute of Technology Roorkee, Roorkee 247667, India.

This paper presents the optical and photophysical properties of RNA templated PbSe nanostructures in the presence of excess Pb(2+). These particles exhibit an onset of absorption in the NIR range at about 1208 nm (0.97 eV) and display weak excitonic bands at 320 nm, 405 nm and 670 nm. The excitation of these particles by 670 nm light causes dual fluorescence in the red and NIR region peaking (range): at 770 nm (690-850 nm) and 1000 nm (850-1150 nm), respectively. The quantum efficiency of fluorescence in the red and NIR region were estimated to be 0.018 and 0.27, respectively. Fluorescence due to freshly prepared colloidal PbSe in the visible and NIR range is observed to have a fairly long lifetime of 320 ns and 31.8 ns, respectively unlike those of organic fluorophores emitting in this range. Aging of these particles leads to a change in morphology from QDs to nanotubes, which is associated with a decrease in absorption coefficient in the visible range along with an increase in absorption in the NIR region. These nanostructures become less emissive and depict a shorter emission lifetime. The poor absorption in the NIR range and a fairly intense fluorescence in the wavelength range of 850-1150 nm make these particles highly suitable for bioimaging of tissues.
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http://dx.doi.org/10.1039/c3dt50497bDOI Listing
August 2013

Congenital anterior urethral valve with or without diverticulum: a single-centre experience.

J Pediatr Urol 2013 Dec 10;9(6 Pt B):1183-7. Epub 2013 Jun 10.

Department of Urology, King George's Medical University, (Formerly, Chhatrapati Shahuji Maharaj Medical University), Lucknow, India.

Introduction: Congenital anterior urethral valves (AUV) are rare and can occur as an isolated entity or in association with proximal diverticula. Diagnosis may be overlooked and ideal treatment is not standardized when both the valve and diverticulum are simultaneously present. We present our experience of congenital AUV.

Material & Methods: From January 2007 to June 2012 a retrospective review of the medical records of 7 cases of AUV was performed.

Results: Three patients were diagnosed as isolated AUV while four presented with associated diverticula. The age of presentation ranged from 10 months to 6 years. Weak voiding stream and dribbling were the most common symptoms. Renal function was found to be deranged in two patients (28%). Hydro-ureteronephrosis was present in three boys (42%) and reflux was present in one patient. Post-void residual volume was >20 ml (mean 55 ml) in all children. Transurethral holmium laser fulguration was carried out on isolated AUV or AUV with small diverticula. Open resection and reconstruction or plication was performed in patients with AUV and proximal large (>3 cm) diverticula. Surgical outcome was successful in all patients except for occurrence of urethrocutaneous fistula in one patient.

Conclusions: In isolated AUV or valve with associated small diverticulum, transurethral holmium:YAG laser ablation is the treatment of choice. Primary excision and repair or plication are preferred if a large diverticula has formed. Eventual outcomes of AUV are good if irreversible changes have not been established.
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http://dx.doi.org/10.1016/j.jpurol.2013.05.006DOI Listing
December 2013

Use of holmium:YAG laser in posterior urethral valves: another method of fulguration.

J Pediatr Urol 2013 Dec 6;9(6 Pt B):1093-7. Epub 2013 May 6.

Department of Urology, King George's Medical University, (Formerly, Chhatrapati Shahuji Maharaj Medical University), Lucknow, India.

Objective: To evaluate the use of holmium:YAG laser for posterior urethral valve (PUV) fulguration and compare with electro-fulguration.

Methods: Forty boys underwent primary fulguration of PUV using 20-25 W holmium laser (Group 1) from January 2009 to December 2011. Data of last 40 boys (retrospective cohort: January 2005 to December 2008) who underwent electro-fulguration (Group 2) for PUV were compared with group 1. Ultrasonography was done at 2 weeks and 3 months and voiding cystourethrography at 3 and 6 months respectively after fulguration and as needed thereafter. DTPA scan and urodynamic study were performed during follow-up in select patients, as required.

Results: Pre-operative parameters were comparable between groups. Improvement in voiding occurred in 38 and 34 boys in group 1 and 2, respectively. Hydroureteronephrosis and vesico-ureteric reflux resolved in 53% and 60% in group 1 and 51% and 53% in group 2, respectively. Boys in group 1 had statistically significant greater success in voiding after catheter removal (40 vs 32), shorter period of catheterization (1 vs 1.8 days), lower mean operative time (15 vs 20 min), needed re-fulguration less commonly (2 vs 6), and were less likely to develop urethral stricture (0 vs 2) and urinary incontinence (0 vs 1).

Conclusion: PUV fulguration using holmium:YAG laser is a feasible, safe and effective alternative for endoscopic transurethral ablation with similar success, and appears to have fewer complications than electro-fulguration.
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http://dx.doi.org/10.1016/j.jpurol.2013.03.015DOI Listing
December 2013

Prospective evaluation of complications using the modified Clavien grading system, and of success rates of percutaneous nephrolithotomy using Guy's Stone Score: A single-center experience.

Indian J Urol 2012 Oct;28(4):392-8

Department of Urology, CSM Medical University (Upgraded King George's Medical College), Lucknow, Uttar Pradesh, India.

Introduction And Objectives: To prospectively document the perioperative complications of percutaneous nephrolithotomy (PCNL) using the modified Clavien grading system. Evaluation of complications and clearance rates according to stone complexity using the validated Guy's Stone Score (GSS) was also done.

Materials And Methods: A total of 221 renal units underwent 278 PCNL procedures at a urology resident training center between September 2010 and September 2011 and data were recorded prospectively in our registry. Patients with co-morbidities like diabetes, renal failure, hypertension and cardiopulmonary diseases were excluded. Stone complexity was classified according to the GSS while peri-operative complications were recorded using the modified Clavien grading system.

Results: Two hundred and forty-five complications were encountered in 278 PCNL procedures involving 116 renal units (41.72%). Complications of Grades 1, 2, 3a, 3b, 4a, 4b and 5 were seen in 52 (18.7%), 122 (43.8%), 42 (15.1%), 18 (6.4%), 6 (2.1%), 4 (1.4%) and 1 (0.3%) renal units respectively. There were 68, 98, 50 and 5 renal units in GSS I, II, III and IV groups, respectively. All grades of complications were more common in GSS III and IV (P<0.05). For GSS I, II, III and IV 100%, 74%, 56% and 0% of renal units, respectively, were stone-free after one session and 0%, 24%, 44% and 60% respectively needed two sessions to be stone-free.

Conclusion: Although the complication rates were higher most were of low grade and self-limiting. Complications were significantly more common with higher GSS and the GSS effectively predicted stone-free rates.
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http://dx.doi.org/10.4103/0970-1591.105749DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3579117PMC
October 2012

Grading complications after transurethral resection of prostate using modified Clavien classification system and predicting complications using the Charlson comorbidity index.

Int Urol Nephrol 2013 Apr 16;45(2):347-54. Epub 2013 Feb 16.

Department of Urology, Chhatrapati Shahuji Maharaj Medical University, (Formerly, King George Medical College), Lucknow, India.

Purpose: To retrospectively report and grade the peri-operative complications of transurethral resection of the prostate (TURP) using the modified Clavien classification system (MCCS) and validate whether Charlson comorbidity index (CCI) predicts the complications after TURP.

Materials And Methods: Between September 2006 and 2012, data of 722 patients who underwent TURP were analyzed after excluding patients with incomplete data (n = 40). Data recorded included the age, prostate volume, operative time, mean prostatic tissue removed and duration of hospitalization while complications were recorded using the MCCS. Preexisting comorbidities were evaluated using the CCI, and patients were classified into 3 CCI score categories (0, 1, ≥ 2).

Results: Two hundred and forty-four complications were seen in 145 (20 %) patients. CCI score was "0" for 480 patients (66.5 %), "1" for 184 patients (25.5 %) and "≥ 2" for 58 patients (8 %). Significant difference was observed between patient groups with CCI score 0, 1 and ≥ 2 for mean age, prostatic weight, operative time, weight of prostatic chips and duration of hospitalization. Similarly, a significant difference in occurrence of various MCCS grades of complication among patients groups with different CCI score was observed.

Conclusion: Grades I, II and III complications constituted the main bulk (90 %) while grade IV were less common (<8 %) and grade V was rare (1 %) after TURP. Men with higher CCI score had a significantly higher rate of morbidity than those with a lower score. The present study is the first to validate that CCI can predict complications of TURP recorded according to the MCCS.
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http://dx.doi.org/10.1007/s11255-013-0399-xDOI Listing
April 2013

Asymmetric growth and division in Mycobacterium spp.: compensatory mechanisms for non-medial septa.

Mol Microbiol 2013 Apr 6;88(1):64-76. Epub 2013 Mar 6.

Department of Cell and Molecular Biology, Uppsala University Biomedical Center, Box 596, 751 24 Uppsala, Sweden.

Mycobacterium spp., rod-shaped cells belonging to the phylum Actinomycetes, lack the Min- and Noc/Slm systems responsible for preventing the placement of division sites at the poles or over the nucleoids to ensure septal assembly at mid-cell. We show that the position for establishment of the FtsZ-ring in exponentially growing Mycobacterium marinum and Mycobacterium smegmatis cells is nearly random, and that the cells often divide non-medially, producing two unequal but viable daughters. Septal sites and cellular growth disclosed by staining with the membrane-specific dye FM4-64 and fluorescent antibiotic vancomycin (FL-Vanco), respectively, showed that many division sites were off-centre, often over the nucleoids, and that apical cell growth was frequently unequal at the two poles. DNA transfer through the division septum was detected, and translocation activity was supported by the presence of a putative mycobacterial DNA translocase (MSMEG2690) at the majority of the division sites. Time-lapse imaging of single live cells through several generations confirmed both acentric division site placement and unequal polar growth in mycobacteria. Our evidence suggests that post-septal DNA transport and unequal polar growth may compensate for the non-medial division site placement in Mycobacterium spp.
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http://dx.doi.org/10.1111/mmi.12169DOI Listing
April 2013