ECTFE (HALAR) as a New Material for Primary Sample Containment of Astromaterials

Fluoropolymers, such as Teflon (PTFE, PFA, FEP) and Viton (FKM), have been used for over 40 years in curating astromaterials at NASA JSC. In general, fluoropolymers have low outgassing and particle shedding properties that reduce cross-contamination to curated samples. Ethylene - Chlorotrifluoroethylene (ECTFE), commonly called Halar (trademark of Solvay Solexis), is a partially fluorinated semi-crystalline copolymer in the same class of fluoropolymers with superior abrasion resistance and extremely low permeability to liquids, gases, and vapors than any other fluoropolymer (fig. 1). ECTFE coatings are becoming more popular in the nuclear, semiconductor, and biomedical industry for lining isolation containment gloveboxes and critical piping as well as other clean room applications. A study was conducted at NASA JSC to evaluate the potential use of Halar on future sample return missions as a material for primary sample containment

Mathematical modeling supports substantial mouse neural progenitor cell death

<p>Abstract</p> <p>Background</p> <p>Existing quantitative models of mouse cerebral cortical development are not fully constrained by experimental data.</p> <p>Results</p> <p>Here, we use simple difference equations to model neural progenitor cell fate decisions, incorporating intermediate progenitor cells and initially low rates of neural progenitor cell death. Also, we conduct a sensitivity analysis to investigate possible uncertainty in the fraction of cells that divide, differentiate, and die at each cell cycle.</p> <p>Conclusion</p> <p>We demonstrate that uniformly low-level neural progenitor cell death, as concluded in previous models, is incompatible with normal mouse cortical development. Levels of neural progenitor cell death up to and exceeding 50% are compatible with normal cortical development and may operate to prevent forebrain overgrowth as observed following cell death attenuation, as occurs in caspase 3-null mutant mice.</p

Effect of Subinhibitory Concentrations of Antibiotics and Disinfectants on ISAba-Mediated Inactivation of Lipooligosaccharide Biosynthesis Genes in Acinetobacter baumannii

Inactivation of the lipooligosaccharide (LOS) biosynthesis genes lpxA, lpxC and lpxD by ISAba insertion elements results in high-level resistance to colistin in A. baumannii. In the present study, we quantify the rate of spontaneous insertional inactivation of LOS biosynthesis genes by ISAba elements in the ATCC 19606-type strain and two multidrug clinical isolates. Using insertional inactivation of lpxC by ISAba11 in the ATCC 19606 strain as a model, we determine the effect of several subinhibitory concentrations of the antibiotics, namely tetracycline, ciprofloxacin, meropenem, kanamycin and rifampicin, as well as the disinfectants ethanol and chlorhexidine on ISAba11 insertion frequencies. Notably, subinhibitory concentrations of tetracycline significantly increased ISAba11 insertion, and rifampicin completely inhibited the emergence of colistin resistance due to ISAba11 inactivation of lpxC. Sequencing of ISAba11 insertion sites within the lpxC gene demonstrated that insertions clustered between nucleotides 382 and 618 (58.3% of unique insertions detected), indicating that this may be a hotspot for ISAba11 insertion. The alignment of insertion sites revealed a semi-conserved AT-rich consensus sequence upstream of the ISAba11 insertion site, suggesting that ISAba11 insertion sites may be sequence-dependent. This study explores previously uncharacterized aspects regarding the acquisition of colistin resistance through insertional activation in LOS biosynthesis genes in A. baumannii.This research was supported by grants MPY 380/18 from the Instituto de Salud Carlos III (ISCIII) awarded to M.J.M. A.C.L. is supported by the Atracción de Talento Program of the Community of Madrid.S

Inhibition of LpxC Increases the Activity of Iron Chelators and Gallium Nitrate in Multidrug-Resistant Acinetobacter baumannii

Infections caused by multidrug-resistant Acinetobacter baumannii would benefit from the development of novel treatment approaches. Compounds that interfere with bacterial iron metabolism, such as iron chelators and gallium nitrate, have previously been shown to have antimicrobial activity against A. baumannii. In this study, we characterize the effect of LpxC inhibitors on the antimicrobial activity of previously characterized iron chelators, 2,2'-bipyridyl (BIP) and deferiprone (DFP), and gallium nitrate (Ga(NO3)3) against A. baumannii reference strains and multidrug-resistant clinical isolates. The LpxC inhibitor LpxC-2 was synergistic with BIP for 30% of strains tested (FICI values: 0.38-1.02), whereas inhibition with LpxC-4 was synergistic with BIP for 60% of strains tested (FICI values: 0.09-0.75). In time-kill assays, combinations of BIP with both LpxC inhibitors demonstrated synergistic activity, with a more than 3 log10 reduction in bacterial counts compared to BIP alone. LpxC-2 was synergistic with Ga(NO3)3 for 50% of strains tested (FICI values: 0.27-1.0), whereas LpxC-4 was synergistic with Ga(NO3)3 for all strains tested (FICI values: 0.08-≤0.50). In time-kill assays, combinations of Ga(NO3)3 with LpxC-2 and LpxC-4 decreased the growth of both strains compared to each compound separately; however, only the combination with LpxC-4 met the defined criteria for synergy. These results identify a novel synergy between two antimicrobial classes against A. baumannii strains.This research was supported by grants MPY 380/18 from the Instituto de Salud Carlos III (ISCIII) awarded to M.J.M. V.V. is supported by the Río Hortega Program from the Instituto de Salud Carlos III. The APC was funded by MPY 380/18 from the Instituto de Salud Carlos III (ISCIII) awarded to M.J.M.S

Identification of Promoter Region Markers Associated With Altered Expression of Resistance-Nodulation-Division Antibiotic Efflux Pumps in Acinetobacter baumannii

Genetic alterations leading to the constitutive upregulation of specific efflux pumps contribute to antibacterial resistance in multidrug resistant bacteria. The identification of such resistance markers remains one of the most challenging tasks of genome-level resistance predictors. In this study, 487 non-redundant genetic events were identified in upstream zones of three operons coding for resistance-nodulation-division (RND) efflux pumps of 4,130 Acinetobacter baumannii isolates. These events included insertion sequences, small indels, and single nucleotide polymorphisms. In some cases, alterations explicitly modified the expression motifs described for these operons, such as the promoter boxes, operators, and Shine-Dalgarno sequences. In addition, changes in DNA curvature and mRNA secondary structures, which are structural elements that regulate expression, were also calculated. According to their influence on RND upregulation, the catalog of upstream modifications were associated with "experimentally verified," "presumed," and "probably irrelevant" degrees of certainty. For experimental verification, DNA of upstream sequences independently carrying selected markers, three for each RND operon, were fused to a luciferase reporter plasmid system. Five out of the nine selected markers tested showed significant increases in expression with respect to the wild-type sequence control. In particular, a 25-fold expression increase was observed with the ISAba1 insertion sequence upstream the adeABC pump. Next, overexpression of each of the three multi-specific RND pumps was linked to their respective antibacterial substrates by a deep A. baumannii literature screen. Consequently, a data flow framework was then developed to link genomic upregulatory RND determinants to potential antibiotic resistance. Assignment of potential increases in minimal inhibitory concentrations at the "experimentally verified" level was permitted for 42 isolates to 7-8 unrelated antibacterial agents including tigecycline, which is overlooked by conventional resistome predictors. Thus, our protocol may represent a time-saving filter step prior to laborious confirmation experiments for efflux-driven resistance. Altogether, a computational-experimental pipeline containing all components required for identifying the upstream regulatory resistome is proposed. This schema may provide the foundational stone for the elaboration of tools approaching antibiotic efflux that complement routine resistome predictors for preventing antimicrobial therapy failure against difficult-to-threat bacteria.This research was supported by grants MPY 380/18 and MPY 509/19 from the Instituto de Salud Carlos III (ISCIII). ML-S is the recipient of a Sara Borrell contract by the ISCIII. AM-G is the recipient of a Miguel Servet contract by the ISCIII.S

System Integration and Intelligence Improvements for WPI’s UGV - Prometheus

This project focuses on realizing a series of operational improvements for WPI\u27s unmanned ground vehicle Prometheus with the end goal of a prize winning entry to the Intelligent Ground Vehicle Challenge. Operational improvements include a practical implementation of stereo vision on an NVIDIA GPU, a more reliable implementation of line detection, a better approach to mapping and path planning, and a modified system architecture realized by an easier to work with GPIO implementation. The end result of these improvements is better autonomy, accessibility, robustness, reliability, and usability for Prometheus

Subinhibitory Concentrations of Clinically-Relevant Antimicrobials Affect Resistance-Nodulation-Division Family Promoter Activity in Acinetobacter baumannii

Efflux pumps contribute to multidrug resistance in Acinetobacter baumannii due to their ability to expel a wide variety of structurally unrelated compounds. This study aimed to characterize the effect of subinhibitory concentrations of clinically-relevant antibiotics and disinfectants on the promoter activity of members of the Resistance-Nodulation-Division (RND) family in A. baumannii. The promoter regions from three RND efflux pumps (AdeABC, AdeFGH and AdeIJK) and the AdeRS regulatory system from three different A. baumannii strains (ATCC 17961, ATCC 17978, and ATCC 19606) were cloned into a luciferase reporter system (pLPV1Z). Promoter activity was quantitatively assessed in both exponential and stationary phase cultures after exposure to subinhibitory concentrations of four antibiotics from different classes (rifampicin, meropenem, tigecycline and colistin) and two disinfectants (ethanol and chlorhexidine). Subinhibitory concentrations of the compounds tested had variable effects on promoter activity that were highly dependent on the A. baumannii strain, the compound tested and the growth phase. Fold changes in AdeABC promoter activity ranged from 1.97 to 113.7, in AdeFGH from -5.6 to 1.13, in AdeIJK from -2.5 to 2, and in AdeRS from -36.2 to -1.32. Taken together, these results indicate that subinhibitory concentrations of clinically-relevant antibiotics and disinfectants affect the promoter activity of RND family members in A. baumannii in a strain and growth phase dependent manner. These results may have important implications for the treatment of infections caused by A. baumannii.AT is supported by the Garantía Juvenil Program of the Comunidad Autonoma de Madrid and ML-S is supported by the Sara Borrell Program of the Instituto de Salud Carlos III. MJM is supported by grants from the Instituto de Salud Carlos III (MP 516/19 and MPY 380/18).S

Role of peptidoglycan recycling enzymes AmpD and AnmK in Acinetobacter baumannii virulence features

Acinetobacter baumannii is an important causative agent of hospital acquired infections. In addition to acquired resistance to many currently-available antibiotics, it is intrinsically resistant to fosfomycin. It has previously been shown that AmpD and AnmK contribute to intrinsic fosfomycin resistance in A. baumannii due to their involvement in the peptidoglycan recycling pathway. However, the role that these two enzymes play in the fitness and virulence of A. baumannii has not been studied. The aim of this study was to characterize several virulence-related phenotypic traits in A. baumannii mutants lacking AmpD and AnmK. Specifically, cell morphology, peptidoglycan thickness, membrane permeability, growth under iron-limiting conditions, fitness, resistance to disinfectants and antimicrobial agents, twitching motility and biofilm formation of the mutant strains A. baumannii ATCC 17978 ΔampD::Kan and ΔanmK::Kan were compared to the wild type strain. Our results demonstrate that bacterial growth and fitness of both mutants were compromised, especially in the ΔampD::Kan mutant. In addition, biofilm formation was decreased by up to 69%, whereas twitching movement was reduced by about 80% in both mutants. These results demonstrate that, in addition to increased susceptibility to fosfomycin, alteration of the peptidoglycan recycling pathway affects multiple aspects related to virulence. Inhibition of these enzymes could be explored as a strategy to develop novel treatments for A. baumannii in the future. Furthermore, this study establishes a link between intrinsic fosfomycin resistance mechanisms and bacterial fitness and virulence traits.ML-S was supported by the Sara Borrell Program of the Instituto de Salud Carlos III (CD17CIII/00017), and AT was supported by the Garantıa Juvenil Program of the Comunidad ́Autónoma de Madrid (PEJ2018-004820-A -MPY 387/19), is currently supported by a FPU grant (FPU20/03261) and PhD student in Biomedical Sciences and Public Health, Universidad Nacional de Educación a Distancia (UNED), Madrid, Spain ([email protected]). MM is supported by grants from the Instituto de Salud Carlos III (MP 516/19 and MPY 380/18).S

Predicted Epitope Abundance Supports Vaccine-Induced Cytotoxic Protection Against SARS-CoV-2 Variants of Concern

The effect of emerging SARS-CoV-2 variants on vaccine efficacy is of critical importance. In this study, the potential impact of mutations that facilitate escape from the cytotoxic cellular immune response in these new virus variants for the 551 most abundant HLA class I alleles was analyzed. Computational prediction showed that most of these alleles, that cover >90% of the population, contain enough epitopes without escape mutations in the principal SARS-CoV-2 variants. These data suggest that the cytotoxic cellular immune protection elicited by vaccination is not greatly affected by emerging SARS-CoV-2 variants.This research was supported by grants from COV20_00679 (MPY 222-20), to MM, MPY 509/19 to AM-G, and MPY 388/18 to DL of “Acción Estratégica en Salud” from the ISCIII.S

Predicted impact of the viral mutational landscape on the cytotoxic response against SARS-CoV-2

The massive assessment of immune evasion due to viral mutations that increase COVID-19 susceptibility can be computationally facilitated. The adaptive cytotoxic T response is critical during primary infection and the generation of long-term protection. Here, potential HLA class I epitopes in the SARS-CoV-2 proteome were predicted for 2,915 human alleles of 71 families using the netMHCIpan EL algorithm. Allele families showed extreme epitopic differences, underscoring genetic variability of protective capacity between humans. Up to 1,222 epitopes were associated with any of the twelve supertypes, that is, allele clusters covering 90% population. Next, from all mutations identified in ~118,000 viral NCBI isolates, those causing significant epitope score reduction were considered epitope escape mutations. These mutations mainly involved non-conservative substitutions at the second and C-terminal position of the ligand core, or total ligand removal by large recurrent deletions. Escape mutations affected 47% of supertype epitopes, which in 21% of cases concerned isolates from two or more sub-continental areas. Some of these changes were coupled, but never surpassed 15% of evaded epitopes for the same supertype in the same isolate, except for B27. In contrast to most supertypes, eight allele families mostly contained alleles with few SARS-CoV-2 ligands. Isolates harboring cytotoxic escape mutations for these families co-existed geographically within sub-Saharan and Asian populations enriched in these alleles according to the Allele Frequency Net Database. Collectively, our findings indicate that escape mutation events have already occurred for half of HLA class I supertype epitopes. However, it is presently unlikely that, overall, it poses a threat to the global population. In contrast, single and double mutations for susceptible alleles may be associated with viral selective pressure and alarming local outbreaks. The integration of genomic, geographical and immunoinformatic information eases the surveillance of variants potentially affecting the global population, as well as minority subpopulations.This research was supported by Acción Estratégica en Salud from the ISCIII (https://www.isciii.es), grants MPY 380/18 (to MJM), 388/18 (to DL) and 509/19 (to AJM-G). AJM-G is the recipient of a Miguel Servet contract by the ISCIII. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.S