Biology subjects

Gilman, R. H.

Publications and source records attributed to Gilman, R. H..

14 recordsLinked to original sources

bioRxiv2026

Pericystic brain transcriptomics reveals molecular signatures of immune activation and neurovascular remodelling in viable and post-treatment porcine neurocysticercosis

Neurocysticercosis (NCC), the infection of the central nervous system by Taenia solium larvae, is a leading cause of acquired epilepsy in endemic regions. While viable cysticerci can persist asymptomatically for extended periods, their spontaneous or drug-induced degradation triggers marked perilesional inflammation and severe neurological symptoms. Despite well-documented histopathological characterisation of these lesion states, the host transcriptional programmes associated with viable parasite persistence and early post-treatment lesion disruption remain poorly understood. To address this gap, we performed the first bulk RNA sequencing of pericystic brain tissue using a physiologically relevant porcine model of NCC. Comparing uninfected controls (n = 3), infected untreated pigs with intact viable cysts (n = 6), and antiparasitic-treated pigs with disrupted cysts (n = 3), we identified distinct transcriptional signatures associated with each disease state. Viable infection was associated with broad transcriptional changes (461 upregulated and 175 downregulated genes), characterised by local immune activation alongside suppression of blood-brain barrier (BBB) remodelling, vascular, and neuronal signalling molecular signatures. The post-treatment state with confirmed BBB disruption was associated with a smaller but directionally distinct response (160 upregulated and 57 downregulated genes), marked by inflammatory signalling and increased expression of genes associated with endothelial activation, vascular regulation, and BBB-associated remodelling. Together, these findings suggest that, while immune engagement is a feature shared across both lesion states, the BBB-associated transcriptional axis shifts substantially following treatment. These results provide an exploratory transcriptomic framework for understanding parasite persistence, treatment-induced neuroinflammation, and neurovascular remodelling in NCC, and highlight candidate pathways and genes for future mechanistic investigation. Author SummaryNeurocysticercosis is a major cause of epilepsy in regions where Taenia solium is endemic. Brain cysts can remain viable for long periods with limited symptoms, but parasite degeneration, whether spontaneous or drug-induced, can trigger damaging neuroinflammation. In this study, we used RNA sequencing in a pig model that closely resembles human disease to characterise how brain tissue responds to viable cysts and to early treatment-induced cyst disruption. We found that viable infection was associated with local immune activation alongside reduced expression of genes involved in blood-brain barrier function. Following antiparasitic treatment, disrupted lesions showed an increased expression of genes linked to vascular and barrier remodelling. These findings suggest that the host transcriptional environment changes substantially after parasite disruption, and highlight molecular pathways that may contribute to neuroinflammation, blood-brain barrier changes, and neurological disease in NCC. As an exploratory first transcriptomic survey in this model, these results provide a candidate framework for future studies aimed at identifying biomarkers and adjunctive therapeutic targets in NCC.

genomics↗

bioRxiv2023

Immunologic changes are detectable in the peripheral blood transcriptome of clinically asymptomatic Chagas cardiomyopathy patients

Chagas disease, caused by the protozoan parasite Trypanosoma cruzi, is a neglected parasitic disease that affects approximately 6 million individuals worldwide. Of those infected, 20-30% will go on to develop chronic Chagas cardiomyopathy (CCC), and ultimately many of these individuals will progress to advanced heart failure. The mechanism by which this progression occurs is poorly understood, as few studies have focused on early CCC. In this study, we sought to understand the physiologic changes associated with T. cruzi infection and the development of CCC. We analyzed gene expression in the peripheral blood of asymptomatic Chagas patients with early structural heart disease, Chagas patients without any signs or symptoms of disease, and Chagas-negative patients with and without early structural heart disease. Our analysis shows that early CCC was associated with a downregulation of various peripheral immune response genes, with gene expression changes suggestive of reduced antigen presentation and T cell activation. Notably, these genes and processes were distinct from those of early cardiomyopathy in Chagas-negative patients, suggesting that the processes mediating CCC may be unique from those mediating progression to other cardiomyopathies. This work highlights the importance of the immune response in early CCC, providing insight into the early pathogenesis of this disease. The changes we have identified may serve as biomarkers of progression and could inform strategies for the treatment of CCC in its early stages, before significant cardiac damage has occurred.

immunology↗

bioRxiv2023

Whole genome assembly of a hybrid Trypanosoma cruzi strain assembled with nanopore sequencing alone

Trypanosoma cruzi is the causative agent of Chagas disease, which causes 10,000 deaths per year. Despite the high mortality caused by the pathogen, relatively few parasite genomes have been assembled to date; even some commonly used laboratory strains do not have publicly available genome assemblies. This is at least partially due to T. cruzis highly complex and highly repetitive genome: while describing the variation in genome content and structure is critical to better understanding T. cruzi biology and the mechanisms that underlie Chagas disease, the complexity of the genome defies investigation using traditional short read sequencing methods. Here, we have generated a high-quality whole genome assembly of the hybrid Tulahuen strain, a commercially available Type VI strain, using long read Nanopore sequencing without short read scaffolding. Using automated tools and manual curation for annotation, we report a genome with 25% repeat regions, 17% variable multigene family members, and 27% transposable elements. Notably, we find that regions with transposable elements are significantly enriched for surface proteins, and that on average surface proteins are closer to transposable elements compared to other coding regions. This finding supports a possible mechanism for diversification of surface proteins in which mobile genetic elements such as transposons facilitate recombination within the gene family. This work demonstrates the feasibility of nanopore sequencing to resolve complex regions of T. cruzi genomes, and with these resolved regions, provides support for a possible mechanism for genomic diversification.

genomics↗

bioRxiv2023

Identification of differentially recognized T cell epitopes in the spectrum of Mtb infection

Tuberculosis caused by Mycobacterium tuberculosis is one of the leading causes of death from a single infectious agent. Identifying dominant epitopes and comparing their reactivity in different tuberculosis (TB) infection states can help design diagnostics and vaccines. We performed a proteome-wide screen of 20,610 Mtb derived peptides in 21 Active TB (ATB) patients 3-4 months post-diagnosis of pulmonary TB (mid-treatment) using an IFN{gamma} and IL-17 Fluorospot assay. Responses were mediated exclusively by IFN{gamma} and identified a total of 137 unique epitopes, with each patient recognizing, on average, 8 individual epitopes and 22 epitopes (16%) recognized by 2 or more participants. Responses were predominantly directed against antigens part of the cell wall and cell processes category. Testing 517 peptides spanning TB vaccine candidates and ESAT- 6 and CFP10 antigens also revealed differential recognition between ATB participants mid-treatment and healthy IGRA+ participants of several vaccine antigens. An ATB-specific peptide pool consisting of epitopes exclusively recognized by participants mid-treatment, allowed distinguishing participants with active pulmonary TB from healthy interferon-gamma release assay (IGRA)+/- participants from diverse geographical locations. Analysis of longitudinal samples indicated decreased reactivity during treatment for pulmonary TB. Together, these results show that a proteome-wide screen of T cell reactivity identifies epitopes and antigens that are differentially recognized depending on the Mtb infection stage. These have potential use in developing diagnostics and vaccine candidates and measuring correlates of protection.

immunology↗

bioRxiv2022

Amplicon sequencing reveals complex infection in infants congenitally infected with Trypanosoma cruzi and informs the dynamics of parasite transmission

Congenital transmission of Trypanosoma cruzi, the causative agent of Chagas disease, is an important source of new infections worldwide. The mechanisms of congenital transmission remain poorly understood, but there is evidence that parasite factors could play a role. Investigating changes in parasite strain diversity during transmission could provide insight into the parasite factors that influence the process. Here we use deep amplicon sequencing of a single copy gene in the T. cruzi genome to evaluate the diversity of infection in a collection of clinical blood samples from Chagas positive mothers and their infected infants. We found several infants and mothers infected with more than two parasite haplotypes, indicating infection with multiple parasite strains. Two haplotypes were detected exclusively in infant samples, while one haplotype was never found in infants, suggesting a relationship between the probability of transmission and parasite genotype. Finally, we found an increase in parasite population diversity in children after birth compared to their mothers, suggesting that there is no transmission bottleneck during congenital infection and that multiple parasites breach the placenta in the course of congenital transmission.

microbiology↗

bioRxiv2022

Peripheral blood T-cell deficiency and hyperinflammatory monocyte responses associate with MAC lung disease

RationaleAlthough nontuberculous mycobacterial (NTM) disease is a growing problem, available treatments are suboptimal and diagnostic tools are inadequate. Immunological mechanisms of susceptibility to NTM disease are poorly understood. ObjectiveTo understand NTM pathogenesis, we evaluated innate and antigen-specific adaptive immune responses to Mycobacterium avium complex (MAC) in individuals with MAC lung disease (MACDZ). MethodsWe synthesized 15mer MAC-, NTM-, or MAC/Mtb-specific peptides and stimulated peripheral blood mononuclear cells (PBMC) with pools of these peptides. We measured T-cell responses by cytokine production, expression of surface markers, and analysis of global gene expression in 27 MACDZ individuals and 32 healthy controls. We also analyzed global gene expression in Mav-infected and uninfected peripheral blood monocytes from 17 MACDZ and 17 healthy controls. Measurements and Main ResultsWe were unable to detect T-cell responses against the peptide libraries or Mav lysate that has increased reactivity in MACDZ subjects compared to controls. T-cell responses to non-mycobacteria derived antigens were preserved. MACDZ individuals had a lower frequency of Th1 and Th1* T-cell populations. By contrast, global gene expression analysis demonstrated upregulation of proinflammatory pathways in uninfected and Mav-infected monocytes derived from MACDZ subjects compared to controls. ConclusionsPeripheral blood T-cell responses to Mycobacterial antigens and the frequency of Th1 and Th1* cell populations are diminished in individuals with MAC disease. In contrast, MACDZ subjects had hyperinflammatory monocyte responses. Together, these data suggest a novel immunologic defect which underlies MAC pathogenesis and includes concurrent innate and adaptive dysregulation.

immunology↗

bioRxiv2022

Assembly and phylogeographic analysis of novel Taenia solium mitochondrial genomes reveal further differentiation between and within Asian and African-American genotypes

BackgroundTaenia solium is a parasite that hampers human health, causing taeniasis and cysticercosis. The genetic variability in its mitochondrial genome is related to the geographical origin of the specimen. Two main genotypes have been identified: The Asian and the African-American. The geographic genetic variability is expected to cause different clinical manifestations. Thus, characterizing differences between and within genotypes is crucial for completing the epidemiology of T. solium diseases. Methods/Principal FindingsHere, two Peruvian (one complete and one partial; 7,811X and 42X of coverage, respectively) and one Mexican (complete, 3,395X) T. solium mitochondrial genomes were assembled using the Chinese reference. Variant calling with respect to the reference was performed. Thirteen SNPs that involved a change in the amino acid physicochemical nature were identified. Those were present in all the assembled genomes and might be linked to differences in aerobic respiration efficiency between Latin American (African-American) and Asian genotypes. Then, phylogeographic studies were conducted using Cytochrome C oxidase subunit I and cytochrome B from these genomes and other isolates. The analysis showed that Indonesian samples are the most ancient and related to the modern T. solium ancestor of the Asian genotype. Finally, a consistent subdivision of the African-American genotype into two subgroups was found. One subgroup relates to East African countries, while the other is West Africa. The East African linage suggests a previously unnoticed influence of the Indian Ocean trade in the genetic structure of Latin America T. solium. Conclusions/SignificanceOverall, this study reports novel mitochondrial genomes valuable for further studies. New Latin American SNPs were identified and suggest metabolic differences between parasites of the Asian and African-American genotypes. Moreover, the phylogeographic analysis revealed differences within each genotype that shed light on T. soliums historical spread. Overall, the results represent an important step in completing T. solium genetic epidemiology. Author SummaryTaenia solium is a human parasite that causes taeniasis and cysticercosis. Eradicated from developed countries, they are still a public health problem in developing nations. T. solium differences in the mitochondrial genetic material depend on its geographical origin. This is expected to cause different clinical manifestations. Despite the importance of genetics to the epidemiology of T. solium diseases, few efforts have been made to assemble and compare their genomes. We aimed to help fill this knowledge gap by assembling three mitochondrial genomes from Latin America and comparing them to the Chinese reference. Additionally, two genes from the Latin American genomes and from other isolates were employed to assess T. solium genetic distribution. We found thirteen mutations with respect to the Chinese genome present in all Latin American samples, which involved a change in the amino acid physicochemical nature. Those might be causing metabolic differences between Asian and Latin American parasites that could change their affinity to specific human tissues. Moreover, we determined that Indonesian samples are the most ancient and related to the modern T. solium ancestor. Finally, we identified a previously unnoticed influence of East African countries in T. solium phylogeny, with which our assembled genomes are closely related.

genetics↗

bioRxiv2021

Silencing of an efflux pump coding gene decreases the efflux rate of pyrazinoic acid in Mycobacterium smegmatis

BackgroundTuberculosis (TB) is an infectious disease caused by Mycobacterium tuberculosis (MTB). The recommended treatment for TB is based on the use of first-line drugs, including pyrazinamide (PZA). PZA is also a drug used in the treatment of multidrug-resistant TB (MDR-TB) because of its main effect against the latent stage. The main cause of resistance to PZA is mutations in the pncA gene, which compromise the activity of the encoded enzyme pyrazinamidase (PZAse), which hydrolyzes PZA into POA, the active antituberculosis molecule. The mechanism of action of PZA requires that POA is expelled from the bacterium by an efflux mechanism. After that, if the extracellular medium is sufficiently acidic, POA is protonated and returns to the cytosol, releasing the proton and repeating the cycle, resulting lethal to the bacteria. The efflux pump responsible for extruding the POA to the extracellular environment is not yet known. Mycobacterium smegmatis is naturally resistant to PZA and has a 900-fold faster POA efflux rate than MTB, and has the advantage to be a faster growing mycobacterium. MethodsIn the present study we have silenced the transcription of several genes encoding efflux pumps in M. smegmatis by CRISPRi (CRISPR interference). These genes (MSMEG_0250, MSMEG_3815, MSMEG_0241, MSMEG_5046 and MSMEG_0410) were homologous to efflux pump genes in MTB. POA efflux rate was measured, and a quantitative Waynes test was performed after silencing each gene. ResultsSilencing of MSMEG_0250, resulted in approximately 5-fold decrease in the POA efflux rate in M. smegmatis (P<0.0001). None of the other silenced genes showed a notable decrease in POA efflux rate.

molecular biology↗

bioRxiv2021

NAToRA, a relatedness-pruning method to minimize the loss of dataset size in genetic and omics analyses

Genetic and omics analyses frequently require independent observations, which is not guaranteed in real datasets. When relatedness cannot be accounted for, solutions involve removing related individuals (or observations) and, consequently, a reduction of available data. We developed a network-based relatedness-pruning method that minimizes dataset reduction while removing unwanted relationships in a dataset. It uses node degree centrality metric to identify highly connected nodes (or individuals) and implements heuristics that approximate the minimal reduction of a dataset to allow its application to large datasets. NAToRA outperformed two popular methodologies (implemented in software PLINK and KING) by showing the best combination of effective relatedness-pruning, removing all relatives while keeping the largest possible number of individuals in all datasets tested and also, with similar or lesser reduction in genetic diversity. NAToRA is freely available, both as a standalone tool that can be easily incorporated as part of a pipeline, and as a graphical web tool that allows visualization of the relatedness networks. NAToRA also accepts a variety of relationship metrics as input, which facilitates its use. We also present a genealogies simulator software used for different tests performed in the manuscript.

bioinformatics↗

bioRxiv2020

EXPRESSION OF Mycobacterium tuberculosis RpsA IN Mycobacterium smegmatis INCREASES SUSCEPTIBILITY TO PYRAZINAMIDE

Pyrazinamide (PZA) is one of the most important drugs used in combined antituberculous therapy. After the drug enters Mycobacterium tuberculosis it is hydrolyzed by pyrazinamidase (PZAse) to the bactericidal molecule pyrazinoic acid (POA). Ribosomal protein S1 (RpsA) was recently identified as a possible target of PZA based on its binding activity to POA and capacity to inhibit trans-translation. However, its role is not completely understood. It has been proposed that Mycobacterium smegmatis RpsA is not capable of binding POA, unlike M. tuberculosis RpsA. This may be due to the different amino acid sequence in the carboxy-terminal region of the two molecules: in M. smegmatis RpsA it is much closer to the sites that may interact with POA than in M. tuberculosis RpsA. These differences could be contributing, along with the presence of highly active POA efflux, to the natural resistance to PZA in M. smegmatis. To further understand the mechanisms of action of PZA and the role of RpsA in PZA susceptibility, we evaluated the effect of complementing M. tuberculosis RpsA expression in M. smegmatis using pNIT mycobacterial non-integrative expression vector and then performed a PZA susceptibility test determining the minimum inhibitory concentration (MIC) of PZA. It was expected that chimeric ribosomes comprising M. tuberculosis RpsA may be present and may affect PZA susceptibility. Our results showed a reduction in PZA MIC in M. smegmatis complemented with overexpressed M. tuberculosis RpsA compared to non-overexpressed M. smegmatis (468 {micro}g/mL and >7500 {micro}g/mL respectively).

microbiology↗

bioRxiv2020

The genetic structure and adaptation of Andean highlanders and Amazonian dwellers is influenced by the interplay between geography and culture

Western South America was one of the worldwide cradles of civilization. The well known Inca Empire was the tip of the iceberg of a cultural and biological evolutionary process that started 14-11 thousand years ago. Genetic data from 18 Peruvian populations reveal that: (1) The between-population homogenization of the central-southern Andes and its differentiation with respect to Amazonian populations of similar latitudes do not extend northward. Instead, longitudinal gene flow between the northern coast of Peru, Andes and Amazonia accompanied cultural and socioeconomic interactions revealed by archeological studies. This pattern recapitulates the environmental and cultural differentiation between the fertile north, where altitudes are lower; and the arid south, where the Andes are higher, acting as a genetic barrier between the sharply different environments of the Andes and Amazonia (2). The genetic homogenization between the populations of the arid Andes is not only due to migration during the Inca Empire or the subsequent colonial period. It started at least during the earlier expansion of the pre-Inca Wari Empire (600-1000 YBP) (3) This demographic history allowed for cases of positive natural selection in the high and arid Andes vs. the low Amazon tropical forest: in the Andes, HAND2-AS1 (heart and neural crest derivatives expressed 2 antisense RNA1, related with cardiovascular function) and DUOX2 (dual oxidase 2, related to thyroid function and innate immunity) genes; in the Amazon, the gene encoding for the CD45 protein, essential for antigen recognition by T/B lymphocytes in viral-host interaction, consistent with the host-virus arms race hypothesis.

evolutionary biology↗

bioRxiv2019

Admixture/fine-mapping in Brazilians reveals a West African associated potential regulatory variant (rs114066381) with a strong female-specific effect on body mass- and fat mass-indexes

Admixed populations are a resource to study the global genetic architecture of complex phenotypes, which is critical, considering that non-European populations are severely under-represented in genomic studies. Leveraging admixture in Brazilians, whose chromosomes are mosaics of fragments of Native American, European and African origins, we used genome-wide data to perform admixture mapping/fine-mapping of Body Mass Index (BMI) in three population-based cohorts from Northeast (Salvador), Southeast (Bambui) and South (Pelotas) of the country. We found significant associations with African-associated alleles in children from Salvador (PALD1 and ZMIZ1 genes), and in young adults from Pelotas (NOD2 and MTUS2 genes). More importantly, in Pelotas, rs114066381, mapped in a potential regulatory region, is significantly associated only in females (p= 2.76 e-06). This variant is very rare in Europeans but with frequencies of ~3% in West Africa, and has a strong female-specific effect (95%CI: 2.32-5.65 kg/m2 per each A allele). We confirmed this sex-specific association and replicated its strong effect for an adjusted fat-mass index in the same Pelotas cohort, and for BMI in another Brazilian cohort from Sao Paulo (Southeast Brazil). A meta-analysis confirmed the significant association. Remarkably, we observed that while the frequency of rs114066381-A allele ranges from 0.8 to 2.1% in the studied populations, it attains ~9% among morbidly obese women from Pelotas, Sao Paulo, and Bambui. The effect size of rs114066381 is at least five-times the effect size of the FTO SNPs rs9939609 and rs1558902, already emblematic for their high effects, and for which we replicated associations in Pelotas. We demonstrate how, after a decade of GWAS mostly performed in European-ancestry populations, non-European and admixed populations are a source of new relevant phenotype-associated genetic variants.

genetics↗

bioRxiv2019

Origins, admixture dynamics and homogenization of the African gene pool in the Americas

The Transatlantic Slave Trade transported more than 9 million Africans to the Americas between the early 16th and the mid-19th centuries. We performed genome-wide analysis of 6,267 individuals from 22 populations and observed an enrichment in West-African ancestry in northern latitudes of the Americas, whereas South/East African ancestry is more prevalent in southern South-America. This pattern results from distinct geographic and geopolitical factors leading to population differentiation. However, we observed a decrease of 68% in the African gene pool between-population diversity within the Americas when compared to the regions of origin from Africa, underscoring the importance of historical factors favoring admixture between individuals with different African origins in the New World. This is consistent with the excess of West-Central Africa ancestry (the most prevalent in the Americas) in the US and Southeast-Brazil, respect to historical-demography expectations. Also, in most of the Americas, admixture intensification occurred between 1,750 and 1,850, which correlates strongly with the peak of arrivals from Africa. This study contributes with a population genetics perspective to the ongoing social, cultural and political debate regarding ancestry, race, and admixture in the Americas.\n\nSignificance StatementDifferently from most genetic studies, that have estimated the overall African ancestry in the Americas, we perform a finer geographic analysis and infer how different African groups contributed to North-, South-American and Caribbean populations, in the context of geographic and geopolitical factors. We also perform a formal comparison of information from demographic history records of the Transatlantic Slave Trade with inferences based on genomic diversity of current populations. Our approach reveals the distinct regional African ancestry roots of different populations from North-, South-America and the Caribe and other important aspects of the historical process of mestizaje and its dynamics in the American continent.

genetics↗

bioRxiv2018

No cell is an island: circulating T cell:monocyte complexes are markers of immune perturbations

Our results highlight for the first time that a significant proportion of cell doublets in flow cytometry, previously believed to be the result of technical artefacts and thus ignored in data acquisition and analysis, are the result of true biological interaction between immune cells. In particular, we show that cell:cell doublets pairing a T cell and a monocyte can be directly isolated from human blood, and high resolution microscopy shows polarized distribution of LFA1/ICAM1 in many doublets, suggesting in vivo formation. Intriguingly, T cell:monocyte complex frequency and phenotype fluctuate with the onset of immune perturbations such as infection or immunization, reflecting expected polarization of immune responses. Overall these data suggest that cell doublets reflecting T cell-monocyte in vivo immune interactions can be detected in human blood and that the common approach in flow cytometry to avoid studying cell:cell complexes should be revisited.

immunology↗