Microbiology
Abstract
Macrophages deploy numerous strategies to combat invasion by microbes. One tactic is to restrict acquisition of diverse nutrients including trace metals, a process termed nutritional immunity. Intracellular pathogens adapt to a resource poor environment by marshalling mechanisms to harvest nutrients. Carbon acquisition is crucial for pathogen survival; compounds that reduce availability are a potential strategy to control intracellular replication. Treatment of macrophages with the glucose analog, 2-deoxy-D-glucose (2-DG), armed phagocytes to eliminate the intracellular fungal pathogen Histoplasma capsulatum in vitro and in vivo. Killing did not rely on altering access to carbon-containing molecules, or changes in ATP, ER stress, or autophagy. Unexpectedly, 2-DG undermined import of exogenous zinc into macrophages decreasing the quantity of cytosolic and phagosomal zinc. The fungus perished as a result of zinc starvation. This change in metal ingress was not ascribed to a defect in a single importer; rather, there was a collective impairment in transporter activity. This undescribed effect promotes the antifungal machinery of macrophages and expands the complexity of 2-DG activities far beyond manipulating glycolysis. Mechanistic metabolic studies employing 2-DG will have to consider its effect on zinc transport. Our preclinical data support consideration of this agent as a possible adjunctive therapy for histoplasmosis.
Authors
Diego C.P. Rossi, Julio A. Landero Figueroa, William R. Buesing, Kathleen Candor, Logan T. Blancett, Heather M. Evans, Rena Lenchitz, Bradford L. Crowther, Waleed Elsegeiny, Peter R. Williamson, Jan Rupp, George S. Deepe Jr.
Abstract
Group B Streptococcus (GBS) is the major cause of human neonatal infections. A single clone, designated CC17-GBS, accounts for more than 80% of meningitis cases, the most severe form of the infection. However, the events allowing blood-borne GBS to penetrate the brain remain largely elusive. In this study, we identified the host transmembrane receptors α5β1 and αvβ3 integrins as the ligands of Srr2, a major CC17-GBS specific adhesin. Two motifs located in the binding region of Srr2 were responsible for the interaction between CC17-GBS and these integrins. We demonstrated, in a blood-brain barrier cellular model, that both integrins contributed to the adhesion and internalization of CC17-GBS. Strikingly, both integrins were overexpressed during the post-natal period in the brain vessels of the blood-brain and blood-cerebrospinal fluid barriers and contributed to the juvenile susceptibility to CC17-meningitis. Finally, blocking these integrins decreased CC17-GBS crossing into the juvenile mice central nervous system in an in vivo model of meningitis.Our study demonstrates that CC17-GBS exploits integrins for crossing the brain vessels leading to meningitis. Importantly, it provides host molecular insights into neonate’s susceptibility to CC17-GBS meningitis, thereby opening new perspectives for therapeutic and prevention strategies of GBS-elicited meningitis.
Authors
Romain Deshayes de Cambronne, Agnès Fouet, Amandine Picart, Anne-Sophie Bourrel, Cyril Anjou, Guillaume Bouvier, Cristina Candeias, Abdelouhab Bouaboud, Lionel Costa, Anne-Cécile Boulay, Martine Cohen-Salmon, Isabelle Plu, Caroline Rambaud, Eva Faurobert, Corinne Albiges-Rizo, Asmaa Tazi, Claire Poyart, Julie Guignot
Abstract
Unlike pathogens, which attack the host, commensal bacteria create a state of friendly coexistence. Here, we identified a mechanism of bacterial adaptation to the host niche, where they reside. Asymptomatic carrier strains were shown to inhibit RNA Polymerase II (Pol II) in host cells by targeting Ser2 phosphorylation; a step required for productive mRNA elongation. Assisted by a rare, spontaneous loss-of-function mutant from a human carrier, the bacterial NlpD protein was identified as a Pol II inhibitor. After internalization by host cells, NlpD was shown to target constituents of the Pol II phosphorylation complex (RPB1 and PAF1C), attenuating host gene expression. Therapeutic efficacy of the rNlpD protein was demonstrated in a urinary tract infection model, by reduced tissue pathology, accelerated bacterial clearance and attenuated Pol II-dependent gene expression. The findings suggest an intriguing, evolutionarily conserved mechanism for bacterial modulation of host gene expression, with a remarkable therapeutic potential.
Authors
Inès Ambite, Nina A. Filenko, Elisabed Zaldastanishvili, Daniel S.C. Butler, Thi Hien Tran, Arunima Chaudhuri, Parisa Esmaeili, Shahram Ahmadi, Sanchari Paul, Björn Wullt, Johannes Putze, Swaine L. Chen, Ulrich Dobrindt, Catharina Svanborg
Abstract
Group A Streptococcus (GAS), a Gram-positive human-specific pathogen yields 517,000 deaths annually worldwide, including 163,000 due to invasive infections and among them puerperal fever. Before efficient prophylactic measures were introduced, the mortality rate for mothers during childbirth was about 10%; puerperal fever still accounts for over 75,000 maternal deaths annually. Yet little is known regarding the factors and mechanisms of GAS invasion and establishment in postpartum infection. We characterized the early steps of infection in an ex vivo infection model of the human decidua, the puerperal fever portal of entry. Coordinate analysis of GAS behavior and the immune response led us to demonstrate that (i) GAS growth was stimulated by tissue products; (ii) GAS invaded tissue and killed ~50% of host cells within two hours; these processes required SpeB protease and Streptolysin O activities, respectively; (iii) GAS impaired the tissue immune response. Immune impairment occurred both at the RNA level, with only partial induction of the innate immune response, and protein level, in an SLO- and SpeB-dependent manner. Our study indicates that efficient GAS invasion of decidua and the restricted host immune response favored its propensity to develop rapid invasive infections in a gynecological-obstetrical context.
Authors
Antonin Weckel, Thomas Guilbert, Clara Lambert, Céline Plainvert, Francois Goffinet, Claire Poyart, Céline Méhats, Agnès Fouet
Abstract
Staphylococcus aureus remains a leading cause of human infection. These infections frequently recur when the skin is a primary site of infection, especially in infants and children. In contrast, invasive staphylococcal disease is less commonly associated with reinfection, suggesting that tissue-specific mechanisms govern the development of immunity. Knowledge of how S. aureus manipulates protective immunity has been hampered by a lack of antigen-specific models to interrogate the T cell response. Utilizing a chicken egg ovalbumin (OVA)-expressing S. aureus strain to analyze OVA-specific T cell responses, we demonstrated that primary skin infection is associated with impaired development of T cell memory. Conversely, invasive infection induced antigen-specific memory and protected against reinfection. This defect in adaptive immunity following skin infection was associated with a loss of dendritic cells, attributable to S. aureus α-toxin (Hla) expression. Genetic and immunization-based approaches to protect against Hla during skin infection restored the T cell response. Within the human population, exposure to α-toxin through skin infection may modulate the establishment of T cell-mediated immunity, adversely impacting long-term protection. These studies prompt consideration that vaccination targeting S. aureus may be most effective if delivered prior to initial contact with the organism.
Authors
Brandon Lee, Reuben Olaniyi, Jakub Kwiecinski, Juliane Bubeck Wardenburg
Abstract
Chimeric antigen receptor–engineered T cells targeting CD19 (CART19) provide an effective treatment for pediatric acute lymphoblastic leukemia but are less effective for chronic lymphocytic leukemia (CLL), focusing attention on improving efficacy. CART19 harbor an engineered receptor, which is delivered through lentiviral vector integration, thereby marking cell lineages and modifying the cellular genome by insertional mutagenesis. We recently reported that vector integration within the host TET2 gene was associated with CLL remission. Here, we investigated clonal population structure and therapeutic outcomes in another 39 patients by high-throughput sequencing of vector-integration sites. Genes at integration sites enriched in responders were commonly found in cell-signaling and chromatin modification pathways, suggesting that insertional mutagenesis in these genes promoted therapeutic T cell proliferation. We also developed a multivariate model based on integration-site distributions and found that data from preinfusion products forecasted response in CLL successfully in discovery and validation cohorts and, in day 28 samples, reported responders to CLL therapy with high accuracy. These data clarify how insertional mutagenesis can modulate cell proliferation in CART19 therapy and how data on integration-site distributions can be linked to treatment outcomes.
Authors
Christopher L. Nobles, Scott Sherrill-Mix, John K. Everett, Shantan Reddy, Joseph A. Fraietta, David L. Porter, Noelle Frey, Saar I. Gill, Stephan A. Grupp, Shannon L. Maude, Donald L. Siegel, Bruce L. Levine, Carl H. June, Simon F. Lacey, J. Joseph Melenhorst, Frederic D. Bushman
Abstract
An excess of fecal bile acids (BAs) is thought to be one of the mechanisms for diarrhea-predominant irritable bowel syndrome (IBS-D). However, the factors causing excessive BA excretion remain incompletely studied. Given the importance of gut microbiota in BA metabolism, we hypothesized that gut dysbiosis might contribute to excessive BA excretion in IBS-D. By performing BA-related metabolic and metagenomic analyses in 290 IBS-D patients and 89 healthy volunteers, we found that 24.5% of IBS-D patients exhibited excessive excretion of total BAs and alteration of BA-transforming bacteria in feces. Notably, the increase in Clostridia bacteria (e.g., C. scindens) was positively associated with the levels of fecal BAs and serum 7α-hydroxy-4-cholesten-3-one (C4), but negatively correlated with serum fibroblast growth factor 19 (FGF19) concentration. Furthermore, colonization with Clostridia-rich IBS-D fecal microbiota or C. scindens individually enhanced serum C4 and hepatic conjugated BAs but reduced ileal FGF19 expression in mice. Inhibition of Clostridium species with vancomycin yielded opposite results. Clostridia-derived BAs suppressed the intestinal FGF19 expression in vitro and in vivo. In conclusion, this study demonstrates that the Clostridia-rich microbiota contributes to excessive BA excretion in IBS-D patients, which provides a mechanistic hypothesis with testable clinical implications.
Authors
Ling Zhao, Wei Yang, Yang Chen, Fengjie Huang, Lin Lu, Chengyuan Lin, Tao Huang, Ziwan Ning, Lixiang Zhai, Linda L.D. Zhong, Waiching Lam, Zhen Yang, Xuan Zhang, Chungwah Cheng, Lijuan Han, Qinwei Qiu, Xiaoxiao Shang, Runyue Huang, Haitao Xiao, Zhenxing Ren, Dongfeng Chen, Silong Sun, Hani El-Nezami, Zongwei Cai, Aiping Lu, Xiaodong Fang, Wei Jia, Zhaoxiang Bian
Abstract
Successful infection by mucosal pathogens requires overcoming the mucus barrier. To better understand this key step, we performed a survey of the interactions between human respiratory mucus and the human pathogen S. pneumoniae. Pneumococcal adherence to adult human nasal fluid was seen only by isolates expressing pilus-1. Robust binding was independent of pilus-1 adhesive properties but required Fab-dependent recognition of RrgB, the pilus shaft protein, by naturally-acquired secretory immunoglobulin A (sIgA). Pilus-1 binding by specific sIgA led to bacterial agglutination, but adherence required interaction of agglutinated pneumococci and entrapment in mucus particles. To test the effect of these interactions in vivo, pneumococci were preincubated with human sIgA prior to intranasal challenge in a mouse model of colonization. sIgA-treatment resulted in rapid immune exclusion of pilus-expressing pneumococci. Our findings predict that immune exclusion would select for non-piliated isolates in individuals who acquired RrgB-specific sIgA from prior episodes of colonization with piliated strains. Accordingly, genomic data comparing isolates carried by mothers and their children showed that mothers are less likely to be colonized with pilus-expressing strains. Our study provides a specific example of immune exclusion involving naturally-acquired antibody in the human host, a major factor driving pneumococcal adaptation.
Authors
Ulrike Binsker, John A. Lees, Alexandria J. Hammond, Jeffrey N. Weiser
Abstract
Mucus-invasive bacterial biofilms are identified on the colon mucosa of approximately 50% of colorectal cancer (CRC) patients and approximately 13% of healthy subjects. Here, we test the hypothesis that human colon biofilms comprise microbial communities that are carcinogenic in CRC mouse models. Homogenates of human biofilm-positive colon mucosa were prepared from tumor patients (tumor and paired normal tissues from surgical resections) or biofilm-positive biopsies from healthy individuals undergoing screening colonoscopy; homogenates of biofilm-negative colon biopsies from healthy individuals undergoing screening colonoscopy served as controls. After 12 weeks, biofilm-positive, but not biofilm-negative, human colon mucosal homogenates induced colon tumor formation in 3 mouse colon tumor models (germ-free ApcMinΔ850/+;Il10–/– or ApcMinΔ850/+ and specific pathogen–free ApcMinΔ716/+ mice). Remarkably, biofilm-positive communities from healthy colonoscopy biopsies induced colon inflammation and tumors similarly to biofilm-positive tumor tissues. By 1 week, biofilm-positive human tumor homogenates, but not healthy biopsies, displayed consistent bacterial mucus invasion and biofilm formation in mouse colons. 16S rRNA gene sequencing and RNA-Seq analyses identified compositional and functional microbiota differences between mice colonized with biofilm-positive and biofilm-negative communities. These results suggest human colon mucosal biofilms, whether from tumor hosts or healthy individuals undergoing screening colonoscopy, are carcinogenic in murine models of CRC.
Authors
Sarah Tomkovich, Christine M. Dejea, Kathryn Winglee, Julia L. Drewes, Liam Chung, Franck Housseau, Jillian L. Pope, Josee Gauthier, Xiaolun Sun, Marcus Mühlbauer, Xiuli Liu, Payam Fathi, Robert A. Anders, Sepideh Besharati, Ernesto Perez-Chanona, Ye Yang, Hua Ding, Xinqun Wu, Shaoguang Wu, James R. White, Raad Z. Gharaibeh, Anthony A. Fodor, Hao Wang, Drew M. Pardoll, Christian Jobin, Cynthia L. Sears
Abstract
Necrotizing fasciitis and myositis are devastating infections characterized by high mortality. Group A streptococcus (GAS) is a common cause of these infections, but the molecular pathogenesis is poorly understood. We report a genome-wide analysis using serotype M1 and M28 strains that identified GAS genes contributing to necrotizing myositis in nonhuman primates (NHP), a clinically relevant model. Using transposon-directed insertion-site sequencing (TraDIS), we identified 126 and 116 GAS genes required for infection by serotype M1 and M28 organisms, respectively. For both M1 and M28 strains, more than 25% of the GAS genes required for necrotizing myositis encode known or putative transporters. Thirteen GAS transporters contributed to both M1 and M28 strain fitness in NHP myositis, including putative importers for amino acids, carbohydrates, and vitamins and exporters for toxins, quorum-sensing peptides, and uncharacterized molecules. Targeted deletion of genes encoding 5 transporters confirmed that each isogenic mutant strain was significantly (P < 0.05) impaired in causing necrotizing myositis in NHPs. Quantitative reverse-transcriptase PCR (qRT-PCR) analysis showed that these 5 genes are expressed in infected NHP and human skeletal muscle. Certain substrate-binding lipoproteins of these transporters, such as Spy0271 and Spy1728, were previously documented to be surface exposed, suggesting that our findings have translational research implications.
Authors
Luchang Zhu, Randall J. Olsen, Stephen B. Beres, Jesus M. Eraso, Matthew Ojeda Saavedra, Samantha L. Kubiak, Concepcion C. Cantu, Leslie Jenkins, Amelia R. L. Charbonneau, Andrew S. Waller, James M. Musser
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Copyright © 2021 American Society for Clinical Investigation
ISSN: 0021-9738 (print), 1558-8238 (online)