Persistent Biofluid Small-Molecule Alterations Induced by Trypanosoma cruzi Infection Are Not Restored by Parasite Elimination.

Chagas disease (CD), caused by Trypanosoma cruzi (T. cruzi) protozoa, is a complicated parasitic illness with inadequate medical measures for diagnosing infection and monitoring treatment success. To address this gap, we analyzed changes in the metabolome of T. cruzi-infected mice via liquid chromatography tandem mass spectrometry of clinically accessible biofluids: saliva, urine, and plasma. Urine was the most indicative of infection status across mouse and parasite genotypes. Metabolites perturbed by infection in urine include kynurenate, acylcarnitines, and threonylcarbamoyladenosine. Based on these results, we sought to implement urine as a tool for the assessment of CD treatment success. Strikingly, it was found that mice with parasite clearance following benznidazole antiparasitic treatment had an overall urine metabolome comparable to that of mice that failed to clear parasites. These results provide a complementary hypothesis to explain clinical trial data in which benznidazole treatment did not improve patient outcomes in late-stage disease, even in patients with successful parasite clearance. Overall, this study provides insights into new small-molecule-based CD diagnostic methods and a new approach to assess functional responses to treatment.

Localized cardiac small molecule trajectories and persistent chemical sequelae in experimental Chagas disease.

Post-infectious conditions present major health burdens but remain poorly understood. In Chagas disease (CD), caused by Trypanosoma cruzi parasites, antiparasitic agents that successfully clear T. cruzi do not always improve clinical outcomes. In this study, we reveal differential small molecule trajectories between cardiac regions during chronic T. cruzi infection, matching with characteristic CD apical aneurysm sites. Incomplete, region-specific, cardiac small molecule restoration is observed in animals treated with the antiparasitic benznidazole. In contrast, superior restoration of the cardiac small molecule profile is observed for a combination treatment of reduced-dose benznidazole plus an immunotherapy, even with less parasite burden reduction. Overall, these results reveal molecular mechanisms of CD treatment based on simultaneous effects on the pathogen and on host small molecule responses, and expand our understanding of clinical treatment failure in CD. This link between infection and subsequent persistent small molecule perturbation broadens our understanding of infectious disease sequelae.

Persistent biofluid small molecule alterations induced by Trypanosoma cruzi infection are not restored by antiparasitic treatment.

Chagas Disease (CD), caused by Trypanosoma cruzi (T. cruzi) protozoa, is a complicated parasitic illness with inadequate medical measures for diagnosing infection and monitoring treatment success. To address this gap, we analyzed changes in the metabolome of T. cruzi-infected mice via liquid chromatography tandem mass spectrometry analysis of clinically-accessible biofluids: saliva, urine, and plasma. Urine was the most indicative of infection status, across mouse and parasite genotypes. Metabolites perturbed by infection in the urine include kynurenate, acylcarnitines, and threonylcarbamoyladenosine. Based on these results, we sought to implement urine as a tool for assessment of CD treatment success. Strikingly, it was found that mice with parasite clearance following benznidazole antiparasitic treatment had comparable overall urine metabolome to mice that failed to clear parasites. These results match with clinical trial data in which benznidazole treatment did not improve patient outcomes in late-stage disease. Overall, this study provides insights into new small molecule-based CD diagnostic methods and a new approach to assess functional treatment response.

Metabolomic Analysis of Polymicrobial Wound Infections and an Associated Adhesive Bandage.

Concerns about ion suppression, spectral contamination, or interference have led to avoidance of polymers in mass spectrometry (MS)-based metabolomics. This avoidance, however, has left many biochemical fields underexplored, including wounds, which are often treated with adhesive bandages. Here, we found that despite previous concerns, the addition of an adhesive bandage can still result in biologically informative MS data. Initially, a test LC-MS analysis was performed on a mixture of known chemical standards and a polymer bandage extract. Results demonstrated successful removal of many polymer-associated features through a data processing step. Furthermore, the bandage presence did not interfere with metabolite annotation. This method was then implemented in the context of murine surgical wound infections covered with an adhesive bandage and inoculated with Staphylococcus aureus, Pseudomonas aeruginosa, or a 1:1 mix of these pathogens. Metabolites were extracted and analyzed by LC-MS. On the bandage side, we observed a greater impact of infection on the metabolome. Distance analysis showed significant differences between all conditions and demonstrated that coinfected samples were more similar to S. aureus-infected samples compared to P. aeruginosa-infected samples. We also found that coinfection was not merely a summative effect of each monoinfection. Overall, these results represent an expansion of LC-MS-based metabolomics to a novel, previously under-investigated class of samples, leading to actionable biological information.

Untargeted Fecal Metabolomic Analyses across an Industrialization Gradient Reveal Shared Metabolites and Impact of Industrialization on Fecal Microbiome-Metabolome Interactions.

The metabolome is a central determinant of human phenotypes and includes the plethora of small molecules produced by host and microbiome or taken up from exogenous sources. However, studies of the metabolome have so far focused predominantly on urban, industrialized populations. Through an untargeted metabolomic analysis of 90 fecal samples from human individuals from Africa and the Americas-the birthplace and the last continental expansion of our species, respectively-we characterized a shared human fecal metabolome. The majority of detected metabolite features were ubiquitous across populations, despite any geographic, dietary, or behavioral differences. Such shared metabolite features included hyocholic acid and cholesterol. However, any characterization of the shared human fecal metabolome is insufficient without exploring the influence of industrialization. Here, we show chemical differences along an industrialization gradient, where the degree of industrialization correlates with metabolomic changes. We identified differential metabolite features such as amino acid-conjugated bile acids and urobilin as major metabolic correlates of these behavioral shifts. Additionally, coanalyses with over 5,000 publicly available human fecal samples and cooccurrence probability analyses with the gut microbiome highlight connections between the human fecal metabolome and gut microbiome. Our results indicate that industrialization significantly influences the human fecal metabolome, but diverse human lifestyles and behavior still maintain a shared human fecal metabolome. This study represents the first characterization of the shared human fecal metabolome through untargeted analyses of populations along an industrialization gradient. IMPORTANCE As the world becomes increasingly industrialized, understanding the biological consequences of these lifestyle shifts and what it means for past, present, and future human health is critical. Indeed, industrialization is associated with rises in allergic and autoimmune health conditions and reduced microbial diversity. Exploring these health effects on a chemical level requires consideration of human lifestyle diversity, but understanding the significance of any differences also requires knowledge of what molecular components are shared between human groups. Our study reveals the key chemistry of the human gut as defined by varied industrialization-based differences and ubiquitous shared features. Ultimately, these novel findings extend our knowledge of human molecular biology, especially as it is influenced by lifestyle and behavior, and provide steps toward understanding how human biology has changed over our species' history.

Spatial Metabolomics Reveals Localized Impact of Influenza Virus Infection on the Lung Tissue Metabolome.

The influenza virus (IAV) is a major cause of respiratory disease, with significant infection increases in pandemic years. Vaccines are a mainstay of IAV prevention but are complicated by IAV's vast strain diversity and manufacturing and vaccine uptake limitations. While antivirals may be used for treatment of IAV, they are most effective in early stages of the infection, and several virus strains have become drug resistant. Therefore, there is a need for advances in IAV treatment, especially host-directed therapeutics. Given the spatial dynamics of IAV infection and the relationship between viral spatial distribution and disease severity, a spatial approach is necessary to expand our understanding of IAV pathogenesis. We used spatial metabolomics to address this issue. Spatial metabolomics combines liquid chromatography-tandem mass spectrometry of metabolites extracted from systematic organ sections, 3D models, and computational techniques to develop spatial models of metabolite location and their role in organ function and disease pathogenesis. In this project, we analyzed serum and systematically sectioned lung tissue samples from uninfected or infected mice. Spatial mapping of sites of metabolic perturbations revealed significantly lower metabolic perturbation in the trachea compared to other lung tissue sites. Using random forest machine learning, we identified metabolites that responded differently in each lung position based on infection, including specific amino acids, lipids and lipid-like molecules, and nucleosides. These results support the implementation of spatial metabolomics to understand metabolic changes upon respiratory virus infection. IMPORTANCE The influenza virus is a major health concern. Over 1 billion people become infected annually despite the wide distribution of vaccines, and antiviral agents are insufficient to address current clinical needs. In this study, we used spatial metabolomics to understand changes in the lung and serum metabolome of mice infected with influenza A virus compared to uninfected controls. We determined metabolites altered by infection in specific lung tissue sites and distinguished metabolites perturbed by infection between lung tissue and serum samples. Our findings highlight the utility of a spatial approach to understanding the intersection between the lung metabolome, viral infection, and disease severity. Ultimately, this approach will expand our understanding of respiratory disease pathogenesis.

Enabling Quantitative Analysis of Surface Small Molecules for Exposomics and Behavioral Studies.

Workplace chemical exposures are a major source of occupational injury. Although over half of these are skin exposures, exposomics research often focuses on chemical levels in the air or in worker biofluids such as blood and urine. Until now, one limitation has been the lack of methods to quantitatively measure surface chemical transfer. Outside the realm of harmful chemicals, the small molecules we leave behind on surfaces can also reveal important aspects of human behavior. In this study, we developed a swab-based quantitative approach to determine small molecule concentrations across common surfaces. We demonstrate its utility using one drug, cyclobenzaprine, on metal surfaces, and two human-derived metabolites, carnitine and phenylacetylglutamine, on four common surfaces: linoleum flooring, plastified laboratory workbench, metal, and Plexiglas. We observed peak areas proportional to surface analyte concentrations at 45 min and 1 week after deposition, enabling quantification of molecule abundance on workplace built environment surfaces. In contrast, this method was unsuitable for analysis of oleanolic acid, for which we did not observe a strong linear proportional relationship following swab-based recovery from surfaces. Overall, this method paves the way for future quantitative exposomics studies in analyte-specific and surface-specific frameworks.

Alterations to the Cardiac Metabolome Induced by Chronic T. cruzi Infection Relate to the Degree of Cardiac Pathology.

Chronic Chagasic cardiomyopathy (CCC) is a Neglected Tropical Disease caused by the parasite Trypanosoma cruzi. The pathognomonic findings in symptomatic CCC patients and animal models includes diffuse cardiac fibrosis and inflammation with persistent parasite presence in the heart. This study investigated chemical alterations in different regions of the heart in relation to cardiac pathology indicators to better understand the long-term pathogenesis of this neglected disease. We used data from echocardiography, fibrosis biomarkers, and histopathological analysis to fully evaluate cardiac pathology. Metabolites isolated from the pericardial and endocardial sides of the right ventricular myocardium were analyzed by liquid chromatography tandem mass spectrometry. The endocardial sections contained significantly less cardiac inflammation and fibrosis than the pericardial sections. Cardiac levels of acylcarnitines, phosphocholines, and other metabolites were significantly disrupted in accordance with cardiac fibrosis, inflammation, and serum fibrosis biomarker levels. These findings have potential implications in treatment and monitoring for CCC patients.

Mass spectrometry studies of the fragmentation patterns and mechanisms of protonated peptoids.

Peptoids belong to a class of sequence-controlled polymers comprising of N-alkylglycine. This study focuses on using tandem mass spectrometry techniques to characterize the fragmentation patterns of a set of singly and doubly protonated peptoids consisting of one basic residue placed at different positions. The singly protonated peptoids fragment by producing predominately high-abundant C-terminal ions called Y-ions and low-abundant N-terminal ions called B-ions. Computational studies suggest that the proton affinity (PA) of the C-terminal fragments is generally higher than that of the N-terminal fragments, and the PA of the former increases as the fragments are elongated. The B-ions are likely formed upon dissociating the proton-activated amide bonds via an oxazolone structure, and the Y-ions are produced subsequently by abstracting a proton from the newly formed B-ions, which is energetically favored. The doubly protonated peptoids prefer to fragment closest to either the N- or the C-terminus and produce corresponding B/Y-ion pairs. The basic residue seems to dictate the preferred fragmentation site, which may be the result of minimizing the repulsion between the two charges. Water and terminal neutral losses are a facile process accompanying the peptoid fragmentation in both charge states. The patterns appear to be highly influenced by the location of the basic residue.

Local Phenomena Shape Backyard Soil Metabolite Composition.

Soil covers most of Earth's continental surface and is fundamental to life-sustaining processes such as agriculture. Given its rich biodiversity, soil is also a major source for natural product drug discovery from soil microorganisms. However, the study of the soil small molecule profile has been challenging due to the complexity and heterogeneity of this matrix. In this study, we implemented high-resolution liquid chromatography-tandem mass spectrometry and large-scale data analysis tools such as molecular networking to characterize the relative contributions of city, state and regional processes on backyard soil metabolite composition, in 188 soil samples collected from 14 USA States, representing five USA climate regions. We observed that region, state and city of collection all influence the overall soil metabolite profile. However, many metabolites were only detected in unique sites, indicating that uniquely local phenomena also influence the backyard soil environment, with both human-derived and naturally-produced (plant-derived, microbially-derived) metabolites identified. Overall, these findings are helping to define the processes that shape the backyard soil metabolite composition, while also highlighting the need for expanded metabolomic studies of this complex environment.

Epigenetic Regulation by Agonist-Specific Aryl Hydrocarbon Receptor Recruitment of Metastasis-Associated Protein 2 Selectively Induces Stanniocalcin 2 Expression.

The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor that regulates a plethora of target genes. Historically, the AhR has been studied as a regulator of xenobiotic metabolizing enzyme genes, notably cytochrome P4501A1 encoded by CYP1A1, in response to the exogenous prototypical ligand 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). AhR activity depends on its binding to the xenobiotic response element (XRE) in partnership with the AhR nuclear translocator (Arnt). Recent studies identified stanniocalcin 2 (Stc2) as a novel AhR target gene responsive to the endogenous AhR agonist cinnabarinic acid (CA). CA-dependent AhR-XRE-mediated Stc2 upregulation is responsible for cytoprotection against ectoplasmic reticulum/oxidative stress-induced apoptosis both in vitro and in vivo. Significantly, CA but not TCDD induces expression of Stc2 in hepatocytes. In contrast to TCDD, CA is unable to induce the CYP1A1 gene, thus revealing an AhR agonist-specific mutually exclusive dichotomous transcriptional response. Studies reported here provide a mechanistic explanation for this differential response by identifying an interaction between the AhR and the metastasis-associated protein 2 (MTA2). Moreover, the AhR-MTA2 interaction is CA-dependent and results in MTA2 recruitment to the Stc2 promoter, concomitant with agonist-specific epigenetic modifications targeting histone H4 lysine acetylation. The results demonstrate that histone H4 acetylation is absolutely dependent on CA-induced AhR and MTA2 recruitment to the Stc2 regulatory region and induced Stc2 gene expression, which in turn confers cytoprotection to liver cells exposed to chemical insults.

Photoelectron Spectroscopy Study of Quinonimides.

Structures and energetics of o-, m-, and p-quinonimide anions (OC6H4N-) and quinoniminyl radicals have been investigated by using negative ion photoelectron spectroscopy. Modeling of the photoelectron spectrum of the ortho isomer shows that the ground state of the anion is a triplet, while the quinoniminyl radical has a doublet ground state with a doublet-quartet splitting of 35.5 kcal/mol. The para radical has doublet ground state, but a band for a quartet state is missing from the photoelectron spectrum indicating that the anion has a singlet ground state, in contrast to previously reported calculations. The theoretical modeling is revisited here, and it is shown that accurate predictions for the electronic structure of the para-quinonimide anion require both an accurate account of electron correlation and a sufficiently diffuse basis set. Electron affinities of o- and p-quinoniminyl radicals are measured to be 1.715 ± 0.010 and 1.675 ± 0.010 eV, respectively. The photoelectron spectrum of the m-quinonimide anion shows that the ion undergoes several different rearrangements, including a rearrangement to the energetically favorable para isomer. Such rearrangements preclude a meaningful analysis of the experimental spectrum.

Fragmentation Patterns and Mechanisms of Singly and Doubly Protonated Peptoids Studied by Collision Induced Dissociation.

Peptoids are peptide-mimicking oligomers consisting of N-alkylated glycine units. The fragmentation patterns for six singly and doubly protonated model peptoids were studied via collision-induced dissociation tandem mass spectrometry. The experiments were carried out on a triple quadrupole mass spectrometer with an electrospray ionization source. Both singly and doubly protonated peptoids were found to fragment mainly at the backbone amide bonds to produce peptoid B-type N-terminal fragment ions and Y-type C-terminal fragment ions. However, the relative abundances of B- versus Y-ions were significantly different. The singly protonated peptoids fragmented by producing highly abundant Y-ions and lesser abundant B-ions. The Y-ion formation mechanism was studied through calculating the energetics of truncated peptoid fragment ions using density functional theory and by controlled experiments. The results indicated that Y-ions were likely formed by transferring a proton from the C-H bond of the N-terminal fragments to the secondary amine of the C-terminal fragments. This proton transfer is energetically favored, and is in accord with the observation of abundant Y-ions. The calculations also indicated that doubly protonated peptoids would fragment at an amide bond close to the N-terminus to yield a high abundance of low-mass B-ions and high-mass Y-ions. The results of this study provide further understanding of the mechanisms of peptoid fragmentation and, therefore, are a valuable guide for de novo sequencing of peptoid libraries synthesized via combinatorial chemistry. Graphical Abstract ᅟ.

Electronic structures of AlMoO(y)(-) (y = 1-4) determined by photoelectron spectroscopy and density functional theory calculations.

Vibrationally-resolved photoelectron spectra of AlMoO(y)(-) (y = 1-4) are presented and analyzed in conjunction with density functional theory computational results. The structures determined for the AlMoO(y) anion and neutral clusters suggest ionic bonding between Al(+) and a MoO(y)(-) or MoO(y)(-2) moiety, and point to the relative stability of Mo=O versus Al=O bonds. The highest occupied and partially occupied orbitals in the anions and neutrals can be described as Mo atomic-like orbitals, so while the Mo is in a higher oxidation state than Al, the most energetically accessible electrons are localized on the molybdenum center.

Electromagnetic interference-aware transmission scheduling and power control for dynamic wireless access in hospital environments.

We study the multiple access problem for e-Health applications (referred to as secondary users) coexisting with medical devices (referred to as primary or protected users) in a hospital environment. In particular, we focus on transmission scheduling and power control of secondary users in multiple spatial reuse time-division multiple access (STDMA) networks. The objective is to maximize the spectrum utilization of secondary users and minimize their power consumption subject to the electromagnetic interference (EMI) constraints for active and passive medical devices and minimum throughput guarantee for secondary users. The multiple access problem is formulated as a dual objective optimization problem which is shown to be NP-complete. We propose a joint scheduling and power control algorithm based on a greedy approach to solve the problem with much lower computational complexity. To this end, an enhanced greedy algorithm is proposed to improve the performance of the greedy algorithm by finding the optimal sequence of secondary users for scheduling. Using extensive simulations, the tradeoff in performance in terms of spectrum utilization, energy consumption, and computational complexity is evaluated for both the algorithms.

IEEE 802.15.4 MAC with GTS transmission for heterogeneous devices with application to wheelchair body-area sensor networks.

In wireless personal area networks, such as wireless body-area sensor networks, stations or devices have different bandwidth requirements and, thus, create heterogeneous traffics. For such networks, the IEEE 802.15.4 medium access control (MAC) can be used in the beacon-enabled mode, which supports guaranteed time slot (GTS) allocation for time-critical data transmissions. This paper presents a general discrete-time Markov chain model for the IEEE 802.15.4-based networks taking into account the slotted carrier sense multiple access with collision avoidance and GTS transmission phenomena together in the heterogeneous traffic scenario and under nonsaturated condition. For this purpose, the standard GTS allocation scheme is modified. For each non-identical device, the Markov model is solved and the average service time and the service utilization factor are analyzed in the non-saturated mode. The analysis is validated by simulations using network simulator version 2.33. Also, the model is enhanced with a wireless propagation model and the performance of the MAC is evaluated in a wheelchair body-area sensor network scenario.

CO2 reduction by group 6 transition metal suboxide cluster anions.

Reactions between small group 6 transition metal suboxide clusters, M(x)O(y)(-) (M = (98)Mo or (186)W; x = 1-4; y < or = 3x) and both CO(2) and CO were studied in gas phase using mass spectrometric analysis of high-pressure, fast flow reaction products. Both Mo(2)O(y)(-) and W(2)O(y)(-) show evidence of sequential oxidation by CO(2) of the form, M(2)O(y)(-)+CO(2)-->M(2)O(y+1)(-)+CO for the more reduced species. Similar evidence is observed for the trimetallic clusters, although Mo(3)O(6)(-) appears uniquely unreactive. Lower mass resolution in the M(4)O(y)(-) range precludes definitive product mass assignments, but intensity patterns suggest the continued trend of sequential oxidation of the more reduced end of the M(4)O(y)(-) oxide series. Based on thermodynamic arguments, cluster oxidation by CO(2) is possible if D(0)(O-Mo(x)O(y)(-)) > 5.45 eV. Although simple bond energy analysis suggests that tungsten oxides may be more reactive toward CO(2) compared to molybdenum oxides, this is not born out experimentally, suggesting that the activation barrier for the reduction of CO(2) by tungsten suboxide clusters is very high compared to analogous molybdenum suboxide clusters. In reactions with CO, suboxides of both metal-based oxides show CO addition, with the product distribution being more diverse for Mo(x)O(y)(-) than for W(x)O(y)(-). No evidence of cluster reduction by CO is observed.

An EMI-aware prioritized wireless access scheme for e-Health applications in hospital environments.

Wireless communications technologies can support efficient healthcare services in medical and patient-care environments. However, using wireless communications in a healthcare environment raises two crucial issues. First, the RF transmission can cause electromagnetic interference (EMI) to biomedical devices, which could critically malfunction. Second, the different types of electronic health (e-Health) applications require different quality of service (QoS). In this paper, we introduce an innovative wireless access scheme, called EMI-aware prioritized wireless access, to address these issues. First, the system architecture for the proposed scheme is introduced. Then, an EMI-aware handshaking protocol is proposed for e-Health applications in a hospital environment. This protocol provides safety to the biomedical devices from harmful interference by adapting transmit power of wireless devices based on the EMI constraints. A prioritized wireless access scheme is proposed for channel access by two different types of applications with different priorities. A Markov chain model is presented to study the queuing behavior of the proposed system. Then, this queuing model is used to optimize the performance of the system given the QoS requirements. Finally, the performance of the proposed wireless access scheme is evaluated through extensive simulations.

Disparate product distributions observed in Mo((3-x))W(x)O(y) (-) (x=0-3; y=3-9) reactions with D(2)O and CO(2).

Results of gas phase reactivity studies on group six transition metal suboxide clusters, Mo(3)O(y) (-), Mo(2)WO(y) (-), MoW(2)O(y) (-), and W(3)O(y) (-) (Mo((3-x))W(x)O(y) (-), x=0-3; y=ca. 3-9) with both D(2)O and CO(2) are reported. Sequential oxidation for the more reduced species, Mo((3-x))W(x)O(y) (-)+D(2)O/CO(2)-->Mo((3-x))W(x)O(y+1) (-)+D(2)/CO, and dissociative addition for certain species, Mo((3-x))W(x)O(y) (-)+D(2)O/CO(2)-->Mo((3-x))W(x)O(y+1)D(2) (-)/Mo((3-x))W(x)O(y+1)CO(-), is evident in the product distributions observed in mass spectrometric measurements. Reactions with D(2)O proceed at a rate that is on the order of 10(2) higher than for CO(2). The pattern of reaction products reveals composition-dependent chemical properties of these group six unary and binary clusters. At the core of this variation is the difference in Mo-O and W-O bond energies, the latter of which is significantly higher. This results in a larger thermodynamic drive to higher oxidation states in clusters with more tungsten atoms. However, addition products for more oxidized W-rich clusters are not observed, while they are observed for the more Mo-rich clusters. This is attributed to the following: In the higher oxides (e.g., y=8), addition reactions require distortion of local metal-oxygen bonding, and will necessarily have higher activation barriers for W-O bonds, since the vibrational potentials will be narrower. The binary (x=1,2) clusters generally show sequential oxidation to higher values of y. This again is attributed to higher W-O bond energy, the result being that stable binary structures have W atoms in higher oxidation states, and Mo centers both in more reduced states and sterically unhindered. The reduced Mo center provides a locus of higher reactivity. An unusual result that is not readily explained is the chemically inert behavior of Mo(3)O(6) (-).