Apollo Next Generation Sample Analysis (ANGSA): an Apollo Participating Scientist Program to Prepare the Lunar Sample Community for Artemis.

As a first step in preparing for the return of samples from the Moon by the Artemis Program, NASA initiated the Apollo Next Generation Sample Analysis Program (ANGSA). ANGSA was designed to function as a low-cost sample return mission and involved the curation and analysis of samples previously returned by the Apollo 17 mission that remained unopened or stored under unique conditions for 50 years. These samples include the lower portion of a double drive tube previously sealed on the lunar surface, the upper portion of that drive tube that had remained unopened, and a variety of Apollo 17 samples that had remained stored at -27 °C for approximately 50 years. ANGSA constitutes the first preliminary examination phase of a lunar "sample return mission" in over 50 years. It also mimics that same phase of an Artemis surface exploration mission, its design included placing samples within the context of local and regional geology through new orbital observations collected since Apollo and additional new "boots-on-the-ground" observations, data synthesis, and interpretations provided by Apollo 17 astronaut Harrison Schmitt. ANGSA used new curation techniques to prepare, document, and allocate these new lunar samples, developed new tools to open and extract gases from their containers, and applied new analytical instrumentation previously unavailable during the Apollo Program to reveal new information about these samples. Most of the 90 scientists, engineers, and curators involved in this mission were not alive during the Apollo Program, and it had been 30 years since the last Apollo core sample was processed in the Apollo curation facility at NASA JSC. There are many firsts associated with ANGSA that have direct relevance to Artemis. ANGSA is the first to open a core sample previously sealed on the surface of the Moon, the first to extract and analyze lunar gases collected in situ, the first to examine a core that penetrated a lunar landslide deposit, and the first to process pristine Apollo samples in a glovebox at -20 °C. All the ANGSA activities have helped to prepare the Artemis generation for what is to come. The timing of this program, the composition of the team, and the preservation of unopened Apollo samples facilitated this generational handoff from Apollo to Artemis that sets up Artemis and the lunar sample science community for additional successes.

Predicting Antimicrobial Resistance Using Partial Genome Alignments.

Antimicrobial resistance (AMR) is an important global health threat that impacts millions of people worldwide each year. Developing methods that can detect and predict AMR phenotypes can help to mitigate the spread of AMR by informing clinical decision making and appropriate mitigation strategies. Many bioinformatic methods have been developed for predicting AMR phenotypes from whole-genome sequences and AMR genes, but recent studies have indicated that predictions can be made from incomplete genome sequence data. In order to more systematically understand this, we built random forest-based machine learning classifiers for predicting susceptible and resistant phenotypes for Klebsiella pneumoniae (1,640 strains), Mycobacterium tuberculosis (2,497 strains), and Salmonella enterica (1,981 strains). We started by building models from alignments that were based on a reference chromosome for each species. We then subsampled each chromosomal alignment and built models for the resulting subalignments, finding that very small regions, representing approximately 0.1 to 0.2% of the chromosome, are predictive. In K. pneumoniae, M. tuberculosis, and S. enterica, the subalignments are able to predict multiple AMR phenotypes with at least 70% accuracy, even though most do not encode an AMR-related function. We used these models to identify regions of the chromosome with high and low predictive signals. Finally, subalignments that retain high accuracy across larger phylogenetic distances were examined in greater detail, revealing genes and intergenic regions with potential links to AMR, virulence, transport, and survival under stress conditions. IMPORTANCE Antimicrobial resistance causes thousands of deaths annually worldwide. Understanding the regions of the genome that are involved in antimicrobial resistance is important for developing mitigation strategies and preventing transmission. Machine learning models are capable of predicting antimicrobial resistance phenotypes from bacterial genome sequence data by identifying resistance genes, mutations, and other correlated features. They are also capable of implicating regions of the genome that have not been previously characterized as being involved in resistance. In this study, we generated global chromosomal alignments for Klebsiella pneumoniae, Mycobacterium tuberculosis, and Salmonella enterica and systematically searched them for small conserved regions of the genome that enable the prediction of antimicrobial resistance phenotypes. In addition to known antimicrobial resistance genes, this analysis identified genes involved in virulence and transport functions, as well as many genes with no previous implication in antimicrobial resistance.

Oncolysis and suppression of tumor growth by a GFP-expressing oncolytic adenovirus controlled by an hTERT and CMV hybrid promoter.

One of the challenges of oncolytic virotherapy is the inability to easily track or monitor virus activity during treatment. Here we describe the construction and functional characterization of Ad/hTC-GFP-E1, an oncolytic virus whose transgenes GFP and E1A are both under the control of a synthetic promoter (hTC). This promoter consists of sequences from the human telomorase reverse transcriptase promoter and a minimal cytomegalovirus (CMV) early promoter. The tumor-specific expression of E1A and GFP was demonstrated by Western blot and fluorescent microscope analyses, and the tumor-specific cytotoxicity by crystal-violet staining and cell viability assays. Viral replication and tumor cell lysis occurred at multiplicities of infection (MOI) as low as 100 viral particles per cell in sensitive cell lines. No overt cytotoxic effect was observed in normal human fibroblasts, even at MOIs over 2000 vp. The presence of oncolytic vector was easily visualized and quantitated in vitro and in vivo, in correlation with viral replication. Intralesional administration of the virus into subcutaneous H1299 (NSCLC) tumor xenografts significantly suppressed tumor growth and provided a survival benefit. Together, these results demonstrate that an hTERT-specific oncolytic adenovirus expressing an hTERT-specific transgene is applicable for cancer therapy.

Monitoring interfacial bioelectrochemistry using a FRET switch.

Generation of functionally active biomolecular monolayers is important in both analytical science and biophysical analyses. Our ability to monitor the redox-active state of immobilized proteins or enzymes at a molecular level, from which stochastic and surface-induced variations would be apparent, is impeded by comparatively slow electron-transfer kinetics and associated signal:noise difficulties. We demonstrate herein that by covalently tethering an appropriate dye to the copper protein azurin a highly oxidation-state-sensitive FRET process can be established which enables redox switching to be optically monitored at protein levels down to the zeptomolar limit. The surface-potential-induced cycling of emission enables the redox potential of clusters of a few hundred molecules to be determined.

Longitudinal pulmonary function changes in pigeon breeders.

We studied 15 subjects (14 men and one woman; seven symptomatic and eight asymptomatic) at three- to four-year intervals from 1967 through 1985 (18 years) to determine if continued pigeon antigen exposure in these groups was associated with a decline in pulmonary function greater than expected in healthy individuals. We collected immunologic studies at the initial visit, performed sequential pulmonary function studies and obtained chest radiographs. After compensating for the normal changes expected with increasing age, we found the declines in FVC, FEV1, and Dsb in the symptomatic group were nearly four times greater than expected. There was no significant difference in the decline of pulmonary function between asymptomatic subjects and the normal predicted declines with increasing age. We conclude that, if an individual has episodes of acute hypersensitivity pneumonitis, long-term exposure to pigeon antigens will result in an accelerated decline in pulmonary function.

A scanning tunnelling microscopy study of Clostridium pasteurianum rubredoxin.

Scanning tunnelling microscopy (STM), which can provide 'direct' and 'non-averaged' information on molecular structure in three dimensions, has been used to achieve sub-molecular resolution in a 'single molecule' of rubredoxin, an important iron-sulphur protein, at the gold (111)/water interface. The metal-ligand site [Fe(III)-Cys4] appears distinct because of an enhancement of the tunnelling current over this region compared to the surrounding protein structure.

Modification of the silver impregnation technique of Bielschowsky for use in glycol methacrylate-embedded brain tissue.

The modified Bielschowsky stain has become the standard silver impregnation technique for use in paraffin-embedded tissue sections for identifying abnormalities of neuritic processes in a variety of neurodegenerative disorders such as Alzheimer's disease. Recently, glycol methacrylate embedding has been used increasingly as a substitute for paraffin embedding of tissue, because it results in less tissue distortion during processing and provides greater clarity and resolution of cellular architecture and structural details. We have been able to modify the Bielschowsky stain for use in glycol methacrylate-embedded brain sections. The use of this technique will permit more accurate quantitation of the histopathologic changes in neuritic processes in neurodegenerative diseases.

Paint as a source of recontamination of houses in urban environments and its role in maintaining elevated blood leads in children.

A detailed lead isotopic and scanning electron microscope investigation of particulates from three houses in urban Sydney, previously decontaminated by their owners, has shown that they have been recontaminated over varying periods, as short as 6 months. The source of recontamination is lead paint from adjoining dwellings whose paint is thoroughly deteriorated, as well as from unknown sources. In one house, the external to internal lead loading was > 10:1. The pathway for the lead paint contaminants is both airborne and mechanical transport into the houses. Recontamination of houses provides an explanation for the maintenance of elevated blood lead levels in the children residing in these houses. Recontamination can be a major urban problem applicable in any community which used leaded paints on dwellings in the past. It is a matter of concern for families with young children and couples, especially women who are, or intend to become, pregnant.

Effects of bovine somatotropin and ruminally undegraded protein on feed intake, live weight gain, and mohair production by yearling Angora wethers.

Yearling Angora wethers (n = 24; 24+/-1.0 kg BW) were used in an experiment with a 2 x 2 factorial arrangement of treatments to investigate effects of bovine somatotropin (bST) treatment and dietary level of ruminally undegraded protein on DMI, ADG, and mohair production. Untreated casein (UC) or casein treated with formaldehyde (TC) was included at 7% DM of a diet containing 11% CP and 46% concentrate. A slow-release bST form was administered weekly to deliver 0 (Control) or 100 microg/ (kg BW.d) of bST. Plasma concentrations of bST and IGF-I were increased (P < .05) during the 7-d period following bST injection. Ruminal fluid ammonia N concentration was lower (P < .01) for TC than for UC before feeding (6.6 vs 7.5 mg/dL) and 4 h later (8.2 vs 12.2 mg/dL), and total VFA concentration was lower (P < .01) for TC than for UC. Treatment with bST decreased (P = .08) DMI with UC (1.15 vs .91 kg/d) and increased (P = .08) DMI with TC (.95 vs 1.06 kg/d). Formaldehyde treatment of casein increased ADG (65, 74, 55, and 91 g/d; P = .03) and clean fleece production (P < .01; 14.1, 17.3, 15.0, and 18.4 g/d for UC-Control, TC-Control, UC-bST, and TC-bST, respectively), with no effect of bST during the 8-wk period of treatment or for the 8 wk thereafter (P > .10). In conclusion, with yearling Angora wethers, bST does not seem useful to enhance mohair production and may not alter effects of dietary level of ruminally undegradable protein on mohair production.

Technical note: tissue residues of mimosine and 2,3-dihydroxypyridine after intravenous infusion in goats.

Sixteen growing Alpine wethers (average BW 35 +/- 2 kg) were assigned to one of four treatments to evaluate tissue retention of the leucaena toxins mimosine (MIM) and 2,3-dihydroxypyridine (2,3-DHP). Treatments were infused i.v. for 2 d and were 1) saline control, 2) MIM (200 mg.kg BW-.75.d-1), 3) 2,3-DHP (200 mg.kg BW-.75.d-1), or 4) MIM (100 mg.kg BW-.75.d-1) + 2,3-DHP (100 mg.kg BW-.75.d-1). Immediately after the infusion, the goats were slaughtered and tissue concentrations of MIM and 2,3-DHP were determined via HPLC. No detectable levels of either toxin were found in spleen, heart, lung, or muscle; however, appreciable amounts of MIM and 2,3-DHP were found in plasma, kidney, and liver samples. Kidney MIM content was greater (P < .01) than that of liver, although liver tended to retain slightly more 2,3-DHP (P > .05). Infusion of MIM resulted in a plasma MIM content of 39 to 54 mumol/L and reduced (P < .01) plasma PHE and LEU. Infusion of 2,3-DHP resulted in a plasma 2,3-DHP content of 9.4 mumol/L and increased plasma THR, ARG, VAL, PHE, ILE, LEU, and LYS concentrations (P < .10). Humans consuming offals from ruminants consuming large amounts of the leguminous forage leucaena may be exposed to appreciable quantities of MIM and 2,3-DHP.

The effect of bovine somatotropin treatment on production of lactating angora does with kids.

Fourteen Angora does (35+/-2 kg), each with a single kid and in the first month of lactation, were used to determine ongoing (Period 1) and residual (Period 2) effects of chronic bovine somatotropin (bST) treatment. Specifically, we sought to determine whether chronic bST treatment was capable of improving milk yield, and thus kid growth, and mohair production of nursing does. The experiment consisted of a 2-wk pretreatment period, 5 wk of weekly subcutaneous treatment of slow-release bST (n = 7; Period 1), and a 4-wk posttreatment period (Period 2). The weekly dose of bST was calculated to release 100 microg/(kg BW.d(-1)). To estimate milk production, kids were separated from the does daily for 5 h, and their BW was recorded before and after suckling. The difference in BW was taken as milk production for 5 h. Fiber growth was measured by shearing does at the start of the experiment and at the end of Periods 1 and 2. Dry matter intake and BW of does were not affected by bST (P>.05). Average daily gain of kids that were suckling bST-treated does was higher (P<.05) than for kids of untreated does during Period 1 (184 vs. 139 g/d) but not during Period 2 (140 vs. 136 g/d; P>.10). Treatment with bST did not affect (P>.10) milk composition or clean fleece production in either period. Injection of bST did not affect (P>.10) plasma concentrations of glucose (mean = 49.5 mg/dL), urea N (mean = 19 mg/dL), total protein (mean = 72.5 g/d), or NEFA (mean = 122 microEq/L). During the period of bST treatment, plasma concentrations of somatotropin and IGF-I were increased (P<.05), concentrations of thyroxine and cortisol were decreased (P<.10), and plasma insulin levels were unchanged (P>.10) by bST. In conclusion, treatment of Angora dams with bST did not change DMI or mohair growth, but it improved growth of their kids.

Achilles tendon ruptures stratified by age, race, and cause of injury among active duty U.S. Military members.

A total of 865 members of the U.S. military underwent repair of Achilles tendon ruptures at U.S. military hospitals during calendar years 1994, 1995, and 1996. The discharge summaries of these patients were analyzed for patient demographic information, including age, race, and causative activity. Patients were then stratified by age, race, and cause of injury. Blacks were at increased risk for undergoing repair of the Achilles tendon compared with nonblacks (overall relative risk = 4.15, 95% confidence interval [CI] = 3.63, 4.74; summary odds ratio controlling for age = 3.69, CI = 3.25, 4.19). Participation in the game of basketball accounted for 64.9% of all injuries in black patients and 34.0% of all injuries in nonblack patients. Among those injured, blacks had a significantly increased risk for injury related to playing basketball than nonblacks (relative risk = 1.82, CI = 1.58, 2.10). This finding suggests that there may be other predisposing factor(s) that result in a higher risk of Achilles tendon ruptures in black individuals.

Lead bioavailability in the environment of children: blood lead levels in children can be elevated in a mining community.

Lower blood lead averages in mining communities, compared with other child exposure settings, e.g., innercity areas of the United States and smelter communities, have been attributed to lower bioavailability of lead to children in the mining areas. Direct supporting evidence of the lower bioavailability has, however, generally been lacking. Elevated blood lead levels for approximately 85% of children with > 10 micrograms/dl have been reported from the Broken Hill mining community in Australia. Lead isotope, optical, and scanning electron microscope analyses on the lead species from soils and dusts show them to be derived mainly from weathered ore body material. Solubility tests using 0.1M HCl on the -53 + 38 microns fraction of soil and dust show the lead species to have a high degree of bioavailability. Ingestion of soil and dust, either directly or via mouthing activity, is the main source and pathway for elevated blood lead in children from this community.

A blueprint for green marketing.

Companies have rushed to market environmentally acceptable products. But according to the author, many have ignored the planning considerations that should have preceded the development and promotion of these "green" products.

Isolated limb perfusion: a novel delivery system for wild-type p53 and fiber-modified oncolytic adenoviruses to extremity sarcoma.

Isolated limb perfusion (ILP) is a limb salvage surgical modality used to deliver chemotherapy and biologic agents to locally advanced and recurrent extremity soft tissue sarcoma (STS), and may be readily tailored for delivery of gene therapy. We set out to test the feasibility of delivering AdFLAGp53 (replication incompetent adenovirus bearing FLAG-tagged wild-type p53) and Ad.hTC.GFP/E1a.RGD (a fiber-modified, replication selective oncolytic adenovirus) into human leiomyosarcoma xenografts by ILP. Nude rats bearing SKLMS-1 tumors in their hind limbs underwent ILP with escalating doses of AdLacZ or AdFLAGp53 (study 1), or with Ad.CMV.GFP.RGD or Ad.hTC.GFP/E1a.RGD (study 2) following in vitro confirmation of therapeutic potential in STS cell lines and strains. Seventy-two hours after delivery, reverse transcription-polymerase chain reaction confirmed FLAGp53 expression, and immunohistochemistry confirmed diffuse upregulation of p21CIP1/WAF1 in ILP-treated tumors. Ad.hTC.GFP/E1a.RGD perfused tumors demonstrated robust macroscopic transgene expression throughout their substance, but not in perfused normal tissues, 21 days after delivery. Intra-tumoral viral replication was confirmed by immunohistochemical staining for early (E1a) and late (hexon) viral protein expression. Terminal deoxynucleotidyl transferase-mediated-digoxigenin nick end-labeling staining identified foci of cell death within regions of viral replication. In conclusion, therapeutic adenoviral gene therapy against limb borne human STS can be successfully delivered by ILP and warrants further investigation.

Oncolytic virotherapy for cancer treatment: challenges and solutions.

Advances in gene modification and viral therapy have led to the development of a variety of vectors in several viral families that are capable of replication specifically in tumor cells. Because of the nature of viral delivery, infection, and replication, this technology, oncolytic virotherapy, may prove valuable for treating cancer patients, especially those with inoperable tumors. Current limitations exist, however, for oncolytic virotherapy. They include the body's B and T cell responses, innate inflammatory reactions, host range, safety risks involved in using modified viruses as treatments, and the requirement that most currently available oncolytic viruses require local administration. Another important constraint is that genetically enhanced vectors may or may not adhere to their replication restrictions in long-term applications. Several solutions and strategies already exist, however, to minimize or circumvent many of these limitations, supporting viral oncolytic therapy as a viable option and powerful tool in the fight against cancer.

JNK1-dependent antimitotic activity of thiazolidin compounds in human non-small-cell lung and colon cancer cells.

We recently identified two thiazolidin compounds, 5-[(4-methylphenyl)methylene]-2-(phenylamino)-4(5H)-thiazolone (MMPT) and 5-(2,4-dihydroxybenzylidene)-2-(phenylimino)-1,3-thiazolidin (DBPT), that inhibit the growth of human non-small-cell lung and colon cancer cells independent of P-glycoprotein and p53 status. Here we further investigated the mechanism by which these thiazolidin compounds mediate their anticancer effects. Treatment of cancer cells with MMPT and DBPT led to a time-dependent accumulation of cells arrested in the G2/M phase with modulation of the expression of proteins such as cyclin B1, cdc25C, and phosphorylated histone H3. Moreover, treatment with MMPT and DBPT increased M-phase arrest with abnormal spindle formation. DBPT-mediated G2/M phase arrest and phosphorylation of cdc25C and histone H3 were abrogated when JNK activation was blocked either with SP600125, a specific JNK inhibitor, or a dominant-negative JNK1 gene. Moreover, DBPT-mediated microtubule disruption was also blocked by SP600125 treatment. Our results demonstrate that thiazolidin compounds can effectively induce G2/M arrest in cancer cells and that this G2/M arrest requires JNK activation.