Central aspects in the pathogenesis of scrub typhus, an infection caused by Orientia (O.) tsutsugamushi, have remained obscure. Its organ and cellular tropism are poorly understood.The purpose of this study was to analyze the kinetics of bacterial dissemination and associated inflammatory responses in infected tissues in an experimental scrub typhus mouse model, following infection with the human pathogenic strain Karp. We provide a thorough analysis of O. tsutsugamushi infection in inbred Balb/c mice using footpad inoculation, which is close to the natural way of infection. By a novel, highly sensitive qPCR targeting the multi copy traD genes, we quantitatively monitored the spread of O. tsutsugamushi Karp from the skin inoculation site via the regional lymph node to the internal target organs. The highest bacterial loads were measured in the lung. Using confocal imaging, we also detected O. tsutsugamushi at the single cell level in the lung and found a predominant macrophage rather than endothelial localization. Immunohistochemical analysis of infiltrates in lung and brain revealed differently composed lesions with specific localizations: iNOS-expressing macrophages were frequent in infiltrative parenchymal noduli, but uncommon in perivascular lesions within these organs. Quantitative analysis of the macrophage response by immunohistochemistry in liver, heart, lung and brain demonstrated an early onset of macrophage activation in the liver. Serum levels of interferon (IFN)-γ were increased during the acute infection, and we showed that IFN-γ contributed to iNOS-dependent bacterial growth control.Our data show that upon inoculation to the skin, O. tsutsugamushi spreads systemically to a large number of organs and gives rise to organ-specific inflammation patterns. The findings suggest an essential role for the lung in the pathogenesis of scrub typhus. The model will allow detailed studies on host-pathogen interaction and provide further insight into the pathogenesis of O. tsutsugamushi infection.
Colistin, also known as polymyxin E, is an antimicrobial agent that is effective against a variety of Gram-negative bacilli, especially the Enterobacteriaceae family. Recently, the wide dissemination of colistin-resistance has brought strong attention to the scientific society because of its importance as the last resort for the treatment of carbapenem-resistant Enterobacteriaceae infections and its possible horizontal transmission. The mobilized colistin resistance (mcr) gene was identified as the gene responsible for unique colistin resistance. Indeed, despite many studies that have revealed a pan variation in the existence of this gene, not only for the mcr genes main group but also for its many subgroups, the problem is growing and worsening day after day. In this regard, this review paper is set to review the updated data that has been published up to the end of 2019 third quarter, especially when related to colistin resistance by the mcr genes. It will include the present status of colistin resistance worldwide, the mcr gene dissemination in different sectors, the discovery of the mcr variants, and the global plan to deal with the threat of antimicrobial resistance. In line with global awareness, and to stop antibiotic misuse and overuse, especially in agricultural animals, the study will further discuss in detail the latest alternatives to colistin use in animals, which may contribute to the elimination of inappropriate antibiotic use and to the help in preventing infections. This review will advance our understanding of colistin resistance, while supporting the efforts toward better stewardship, for the proper usage of antimicrobial drugs in humans, animals, and in the environment.
T cells are known to contribute to immune protection against scrub typhus, a potentially fatal infection caused by the obligate intracellular bacterium Orientia (O.) tsutsugamushi. However, the contribution of CD8+ T cells to protection and pathogenesis during O. tsutsugamushi infection is still unknown. Using our recently developed BALB/c mouse model that is based on footpad inoculation of the human-pathogenic Karp strain, we show that activated CD8+ T cells infiltrate spleen and lung during the third week of infection. Depletion of CD8+ T cells with monoclonal antibodies resulted in uncontrolled pathogen growth and mortality. Adoptive transfer of CD8+ T cells from infected animals protected naïve BALB/c mice from lethal outcome of intraperitoneal challenge. In C57Bl/6 mice, the pulmonary lymphocyte compartment showed an increased percentage of CD8+ T cells for at least 135 days post O. tsutsugamushi infection. Depletion of CD8+ T cells at 84 days post infection caused reactivation of bacterial growth. In CD8+ T cell-deficient beta 2-microglobulin knockout mice, bacterial replication was uncontrolled, and all mice succumbed to the infection, despite higher serum IFN-γ levels and stronger macrophage responses in liver and lung. Moreover, we show that CD8+ T cells but not NKT cells were required for hepatocyte injury: elevated concentrations of serum alanine aminotransferase and infection-induced subcapsular necrotic liver lesions surrounded by macrophages were found in C57Bl/6 and CD1d-deficient mice, but not in beta 2-microglobulin knockout mice. In the lungs, peribronchial macrophage infiltrations also depended on CD8+ T cells. In summary, our results demonstrate that CD8+ T cells restrict growth of O. tsutsugamushi during acute and persistent infection, and are required to protect from lethal infections in BALB/c and C57BL/6 mice. However, they also elicit specific pathologic tissue lesions in liver and lung.
Scrub typhus is a potentially lethal infection that is caused by the obligate intracellular bacterium Orientia tsutsugamushi. The roles of Toll-like receptor 2 (TLR2) and TLR4 in innate recognition of O. tsutsugamushi have not been elucidated. By overexpression of TLR2 or TLR4 in HEK293 cells, we demonstrated that TLR2, but not TLR4, recognizes heat-stable compounds of O. tsutsugamushi that were sensitive to treatment with sodium hydroxide, hydrogen peroxide, and proteinase K. TLR2 was required for the secretion of tumor necrosis factor alpha (TNF-␣) and interleukin-6 (IL-6) by dendritic cells. In an intradermal mouse infection model, TLR2-deficient mice did not show impaired control of bacterial growth or reduced survival. Moreover, after intraperitoneal infection, TLR2-deficient mice were even more resistant to lethal infection than C57BL/6 wild-type mice, which showed stronger symptoms and lower survival rates during the convalescent phase. Compared to the time of reduction of bacterial loads in TLR2-deficient mice, the reduction of bacterial loads in infected organs was accelerated in wild-type mice. The higher mortality of wild-type mice was associated with increased concentrations of serum alkaline phosphatase but not aspartate aminotransferase. The transcription of mRNA for TNF-␣ and IL-6 decreased more rapidly in peritoneum samples from wildtype mice than in those from TLR2-deficient mice and was therefore not a correlate of increased susceptibility. Thus, although TLR2 is an important mediator of the early inflammatory response, it is dispensable for protective immunity against O. tsutsugamushi. Increased susceptibility to O. tsutsugamushi infection in TLR2-competent mice rather suggests a TLR2-related immunopathologic effect. Scrub typhus is a neglected chigger-borne zoonosis caused by the obligate intracellular bacterium Orientia tsutsugamushi. Although scrub typhus has lost much of its former threat thanks to the advent of antibiotic therapy, fatal infections continue to be reported, yet it remains largely unknown how bacterial virulence factors and host immunity mutually contribute to the pathogenesis of scrub typhus (1).In mammals, the Toll-like receptor (TLR) system has evolved as a germ line-encoded, ancient repertoire of pattern recognition receptors. Conserved structures on the surface of microorganisms consisting of lipoproteins and lipopeptides (2, 3) and of lipopolysaccharide (LPS) in Gram-negative bacteria (4) are recognized by TLR2 and TLR4, respectively, and trigger immediate antimicrobial responses. As part of the first line of immune defense, these responses help to limit infection success and shape the development of protective adaptive immunity. TLR ligation, mediated by MyD88-or TRIF-dependent pathways, results in activation of transcription factors, such as nuclear factor (NF)-B, activated protein-1 (AP-1), or interferon regulatory transcription factors (IRFs) (5).Many studies have supported a protective effect for TLR2 and TLR4 during bacterial and fungal infections in mouse models...
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