C-C Motif Ligand 7 (CCL7) and C-C Motif Chemokine Receptor 3 (CCR3) dysregulation in patients with scrub typhus and association with mortality.

BACKGROUND: Scrub typhus, caused by Orientia tsutsugamushi involves infiltration of a mixture of perivascular lymphocytes and macrophages into affected organs. We investigated if this is characterized by chemokine dysregulation. METHODS: mRNA expression of chemokines and receptors were screened in whole blood by cDNA microarray in a subgroup of patients and controls. Regulated transcripts were analyzed in plasma by enzyme immunoassays (chemokines) and in whole blood by qPCR (receptors) from scrub typhus patients (n=129), patients with similar febrile illness without Orentia tsutsugamushi infection (n=31) and healthy controls (n=31). RESULTS: (i) cDNA microarray identified dysregulation of the chemokines CCL18 and CCL23 and CCR3 receptor, in severe scrub typhus. (ii) Plasma CCL7, a ligand for CCR3, CCL18 and CCL23 were higher in scrub typhus patients, with a decline during follow-up. (iii) Conversely, mRNA levels of CCR3 and CCR8, the receptor for CCL18, were decreased in whole blood at hospital admission followed by an increase during follow-up. (iv) CCL7 was independently associated with disease severity. (v) Admission CCL7 levels were associated with short-time mortality. CONCLUSION: Our findings suggest CCL7 could represent a hitherto unknown pathogenic mediator in Orentia tsutsugamushi infection contributing to local and systemic inflammation.

Orientia tsutsugamushi infection reduces host gluconeogenic but not glycolytic substrates.

Orientia tsutsugamushi a causal agent of scrub typhus, is an obligate intracellular bacterium that, akin to other rickettsiae, is dependent on host cell-derived nutrients for survival and thus pathogenesis. Based on limited experimental evidence and genome-based in silico predictions, O. tsutsugamushi is hypothesized to parasitize host central carbon metabolism (CCM). Here, we (re-)evaluated O. tsutsugamushi dependency on host cell CCM as initiated by glucose and glutamine. Orientia infection had no effect on host glucose and glutamine consumption or lactate accumulation, indicating no change in overall flux through CCM. However, host cell mitochondrial activity and ATP levels were reduced during infection and correspond with lower intracellular glutamine and glutamate pools. To further probe the essentiality of host CCM in O. tsutsugamushi proliferation, we developed a minimal medium for host cell cultivation and paired it with chemical inhibitors to restrict the intermediates and processes related to glucose and glutamine metabolism. These conditions failed to negatively impact O. tsutsugamushi intracellular growth, suggesting the bacterium is adept at scavenging from host CCM. Accordingly, untargeted metabolomics was utilized to evaluate minor changes in host CCM metabolic intermediates across O. tsutsugamushi infection and revealed that pathogen proliferation corresponds with reductions in critical CCM building blocks, including amino acids and TCA cycle intermediates, as well as increases in lipid catabolism. This study directly correlates O. tsutsugamushi proliferation to alterations in host CCM and identifies metabolic intermediates that are likely critical for pathogen fitness.IMPORTANCEObligate intracellular bacterial pathogens have evolved strategies to reside and proliferate within the eukaryotic intracellular environment. At the crux of this parasitism is the balance between host and pathogen metabolic requirements. The physiological basis driving O. tsutsugamushi dependency on its mammalian host remains undefined. By evaluating alterations in host metabolism during O. tsutsugamushi proliferation, we discovered that bacterial growth is independent of the host's nutritional environment but appears dependent on host gluconeogenic substrates, including amino acids. Given that O. tsutsugamushi replication is essential for its virulence, this study provides experimental evidence for the first time in the post-genomic era of metabolic intermediates potentially parasitized by a scrub typhus agent.

Canonical notch activation in patients with scrub typhus: association with organ dysfunction and poor outcome.

PURPOSE: The mechanisms that control inflammation in scrub typhus are not fully elucidated. The Notch pathways are important regulators of inflammation and infection, but have not been investigated in scrub typhus. METHODS: Plasma levels of the canonical Notch ligand Delta-like protein 1 (DLL1) were measured by enzyme immunoassay and RNA expression of the Notch receptors (NOTCH1, NOTCH2 and NOTCH4) in whole blood was analyzed by real-time PCR in patients with scrub typhus (n = 129), in patients with similar febrile illness without O. tsutsugamushi infection (n = 31) and in healthy controls (n = 31); all from the same area of South India. RESULTS: Our main results were: (i) plasma DLL1 was markedly increased in scrub typhus patients at hospital admission with a significant decrease during recovery. (ii) RNA expression of NOTCH4 was decreased at admission in whole blood. (iii) A similar pattern for DLL1 and NOTCH4 was seen in febrile disease controls. (iv) Admission DLL1 in plasma was associated with disease severity and short-term survival. (vi) Regulation of Notch pathways in O. tsutsugamushi-infected monocytes as evaluated by public repository data revealed enhanced canonical Notch activation with upregulation of DLL1 and downregulation of NOTCH4. CONCLUSION: Our findings suggest that scrub typhus patients are characterized by enhanced canonical Notch activation. Elevated plasma levels of DLL1 were associated with organ dysfunction and poor outcomes in these patients.

Discovery of a novel spotted fever group Rickettsia, "Candidatus Rickettsia kedanie," in unfed larval chigger mites, Leptotrombidium scutellare.

Spotted fever group (SFG) rickettsia, the causative agent of SFG rickettsiosis, is predominantly carried by ticks, whereas Orientia tsutusgamushi, the causative agent of scrub typhus, is primarily transmitted by chigger mites in Japan. In this study, we attempted to isolate intracellular eubacteria from Leptotrombidium scutellare, a major vector of O. tsutsugamushi; moreover, we isolated an SFG rickettsia using a mosquito-derived cell line. Draft genome sequences of this unique isolate, by applying criteria for species delimitation, classified this isolate as a novel strain, proposed as "Rickettsia kedanie." Further genetic analysis identified conserved virulence factors, and the isolate successfully propagated in mammalian cells, suggesting its ability to cause diseases in humans. The presence of SFG rickettsia in unfed larvae implies potential dual-pathogen carriage and reflects a symbiotic relationship similar to that between the mites and O. tsutsugamushi, indicating possibility of its transovarial transmission from female adults. Furthermore, conserved genomic similarity of the novel isolate to known SFG rickettsia suggests potential multiple hosts, including chiggers and ticks. In the natural environment, ticks, chigger mites, and wild animals may carry new isolates, complicating the infection cycle and increasing the transmission risks to humans. This discovery challenges the conventional association of SFG rickettsia with ticks, emphasizing its implications for research and disease control. However, this study was confined to a particular species of chigger mites and geographic area, underscoring the necessity for additional studies to comprehend the ecological dynamics, host interactions, and health implications linked to this newly identified SFG rickettsia.

Gaps and inconsistencies in the current knowledge and implementation of biosafety and biosecurity practices for rickettsial pathogens.

INTRODUCTION: Rickettsia spp. and Orientia spp. are the causes of neglected infections that can lead to severe febrile and systemic illnesses in humans. Implementing proper biosafety practices when handling these pathogens is crucial to ensure a safe and sustainable work environment. It is essential to assess the current knowledge and identify any potential gaps to develop effective measures that minimise the risk of exposure to these pathogens. By doing so, we can establish a comprehensive framework that promotes safety, mitigates hazards, and safeguards the well-being of personnel and the surrounding community. METHODS AND RESULTS: This review aimed to synthesise and determine the evidence base for biosafety precautions for Rickettsia spp. and Orientia spp. pathogens. Enhancing our understanding of the relative infectious risk associated with different strains of Rickettsia and Orientia spp. requires identifying the infectious dose of these pathogens that can cause human disease. The application of risk groups for Rickettsia and Orientia spp. is inconsistent across jurisdictions. There is also incomplete evidence regarding decontamination methods for these pathogens. With regards to Orientia spp. most of the available information is derived from experiments conducted with Rickettsia spp. CONCLUSIONS: Rickettsia and Orientia spp. are neglected diseases, as demonstrated by the lack of evidence-based and specific biosafety information about these pathogens. In the case of Orientia spp., most of the available information is derived from Rickettsia spp., which may not be appropriate and overstate the risks of working with this pathogen. The advent of effective antibiotic therapy and a better understanding of the true hazards and risks associated with pathogen manipulation should inform decisions, allowing a sustainable and safe work environment.

Changes in seasonality and sex ratio of scrub typhus: a case study of South Korea from 2003 to 2019 based on wavelet transform analysis.

BACKGROUND: Scrub typhus (ST, also known as tsutsugamushi disease) is a common febrile vector-borne disease in South Korea and commonly known as autumn- and female-dominant disease. Although understanding changes in seasonality and sex differences in ST is essential for preparing health interventions, previous studies have not dealt with variations in periodicity and demographic characteristics in detail. Therefore, we aimed to quantify the temporal dynamics of seasonal patterns and sex differences in the incidence of ST in South Korea. METHODS: We extracted epidemiological week (epi-week)-based ST cases from 2003 to 2019 Korean National Health Insurance Service data (ICD-10-CM code: A75.3). To determine changes in seasonality and sex differences, year-, sex-, and age-group-stratified male-to-female ratios and wavelet transform analyses were conducted. RESULTS: Between 2003 and 2019, 213,976 ST cases were identified. The incidence per 100,000 population increased by 408.8% from 9.1 in 2003 to 37.2 in 2012, and subsequently decreased by 59.7% from 2012 to 15.0 in 2019. According to the continuous wavelet transform results, ST exhibited a dual seasonal pattern with dominant seasonality in autumn and smaller seasonality in spring from 2005 to 2019. Overall, the periodicity of seasonality decreased, whereas its strength decreased in autumn and increased in spring. With an overall male-to-female ratio being 0.68:1, the ratio has increased from 0.67:1 in 2003 to 0.78:1 in 2019 (Kendall's τ = 0.706, p < 0.001). However, interestingly, the ratio varied significantly across different age groups. CONCLUSIONS: Our findings quantitatively demonstrated changes in seasonality with dual seasonal pattern and shortened overall periodicity and a decrease in sex differences of ST in South Korea. Our study suggests the need for continuous surveillance on populations of vector and host to address ST dynamics to preemptively prepare against global warming.

Clinical and laboratory findings in scrub typhus associated Guillain-Barré syndrome in South Korea.

BACKGROUND AND AIMS: Scrub typhus is an endemic disease in the fall season that occurs in a limited number of places known as the Tsutsugamushi Triangle. Peripheral neuropathy is a common complication of scrub typhus. Herein, we encountered several patients with ascending paralysis after scrub typhus infection, who were diagnosed with Guillain-Barré syndrome (GBS). We aimed to investigate the clinical and laboratory characteristics of patients who developed GBS after scrub typhus. METHODS: Patients were retrospectively recruited from six nationwide tertiary centers in South Korea from January 2017 to December 2021. Patients who had been clinically diagnosed with GBS and confirmed to have scrub typhus via laboratory examination and/or the presence of an eschar before the onset of acute limb paralysis were included. The GBS-associated clinical and electrophysiological characteristics, outcomes, and scrub typhus-associated features were collected. RESULTS: Of the seven enrolled patients, six were female and one was male. The median time from scrub typhus infection to the onset of limb weakness was 6 (range: 2-14) days. All patients had eschar on their bodies. Four patients (57.1%) were admitted to the intensive care unit and received artificial ventilation for respiratory distress. At 6 months, the median GBS disability score was 2 (range, 1-4) points. INTERPRETATION: Patients with scrub typhus-associated GBS have a severe clinical presentation and require intensive treatment with additional immunotherapies. Therefore, GBS should be included in the differential diagnosis when peripheral neuropathies develop during scrub typhus treatment. Notably, scrub typhus is associated to GBS.

Unveiling the burden of scrub typhus in acute febrile illness cases across India: A systematic review & meta-analysis.

Background & objectives Scrub typhus is an emerging mite-borne zoonotic infection that has been overlooked, despite being one of the most widespread severe vector-borne diseases. With an estimated one billion people at risk worldwide and one million annual cases, it poses a significant public health concern. While various studies have investigated the prevalence of scrub typhus in different regions of India, a comprehensive regional systematic review and meta-analysis on the seropositivity of scrub typhus among acute febrile cases has been lacking. To address this gap, we conducted a systematic review and meta-analysis to compile information on the current seroprevalence of scrub typhus in acute febrile illness cases in India. Methods A literature search of multiple databases on prevalence of scrub typhus in acute febrile illness in India, 60 eligible studies out of 573 studies. The prevalence of individual studies was double arcsine transformed, and the pooled prevalence was calculated using inverse variance method. Results In total, these studies encompassed 34,492 febrile cases. The overall seroprevalence of scrub typhus among acute febrile illness cases in India was found to be 26.41 per cent [95% confidence interval (CI): 22.03-31.03]. Additionally, the pooled case fatality rate (based on data from six studies) among scrub typhus-positive cases yielded a case fatality rate of 7.69 per cent (95% CI: 4.37-11.72). Interpretation & conclusions This meta-analysis shows that scrub typhus is a significant health threat in India. Preventive measures to control scrub typhus need to be given priority.

Scrub Typhus- An Underestimated Infectious Disease Attributable to Community Acquired Acute Kidney Injury.

Acute Kidney Injury (AKI) associated with Scrub typhus is an emerging health problem which is more common in the tropics including India. This study intended to find out the occurrence of Scrub typhus among the Community Acquired Acute Kidney Injury patients in a tertiary care hospital in Assam, North East India. AKI patients with acute febrile illness admitted to Gauhati Medical College and Hospital, Guwahati, Assam were included in the study and demographic characteristics along with clinical features were recorded. The detection of Scrub typhus was done by IgM Enzyme Linked Immunosorbent Assay (ELISA) test (Optical Density > 0.5) and polymerase chain reaction (PCR) analysis. Routine haematological and biochemical tests were performed. Molecular characterization of Orientia tsutsugamushi was done followed by phylogenetic analysis. The Graph Pad Prism software 9 was used for statistical analysis. Out of 221 AKI patients admitted to hospital, 45 patients (20.4%) were confirmed to be Scrub typhus positive and among them, 4 cases were co-infected with leptospirosis. Majority of Scrub typhus positive AKI patients were in Stage I (82.2%) under KDIGO guideline. "Karp" was the predominant circulating serotype. The study showed cases of Scrub typhus associated Acute Kidney Injury was high and mortality was 11.1%. Hence, in this region, further studies need to be done with large number of population and more emphasis need to be given on differential diagnosis. Supplementary Information: The online version contains supplementary material available at 10.1007/s12088-023-01137-x.

Detection of Orientia spp. Bacteria in Field-Collected Free-Living Eutrombicula Chigger Mites, United States.

Scrub typhus, a rickettsial disease caused by Orientia spp., is transmitted by infected larval trombiculid mites (chiggers). We report the molecular detection of Orientia species in free-living Eutrombicula chiggers collected in an area in North Carolina, USA, to which spotted fever group rickettsiae infections are endemic.

Association of Scrub Typhus in Children with Acute Encephalitis Syndrome and Meningoencephalitis, Southern India.

Scrub typhus is an established cause of acute encephalitis syndrome (AES) in northern states of India. We systematically investigated 376 children with AES in southern India, using a stepwise diagnostic strategy for the causative agent of scrub typhus, Orientia tsutsugamushi, including IgM and PCR testing of blood and cerebrospinal fluid (CSF) to grade its association with AES. We diagnosed scrub typhus in 87 (23%) children; of those, association with AES was confirmed in 16 (18%) cases, probable in 55 (63%), and possible in 16 (18%). IgM detection in CSF had a sensitivity of 93% and specificity of 82% compared with PCR. Our findings suggest scrub typhus as an emerging common treatable cause of AES in children in southern India and highlight the importance of routine testing for scrub typhus in diagnostic algorithms. Our results also suggest the potential promise of IgM screening of CSF for diagnosis of AES resulting from scrub typhus.

Nonnegligible Seroprevalence and Predictors of Murine Typhus, Japan.

To elucidate the epidemiology of murine typhus, which is infrequently reported in Japan, we conducted a cross-sectional study involving 2,382 residents of rickettsiosis-endemic areas in Honshu Island during August-November 2020. Rickettsia typhi seroprevalence rate was higher than that of Orientia tsutsugamushi, indicating that murine typhus is a neglected disease.

Environmental, Occupational, and Demographic Risk Factors for Clinical Scrub Typhus, Bhutan.

Underdiagnosis and underreporting of scrub typhus has increasingly affected public health in Bhutan since its initial detection in 2008. Identifying scrub typhus risk factors would support early diagnosis and treatment for this nonspecific febrile disease, reducing the incidence of potentially fatal complications. We conducted a hospital-based, case‒control study during October‒December 2015 in 11 scrub typhus‒prone districts. We identified harvesting cardamom as the major risk factor (odds ratio 1,519; p<0.001); other factors were traditional housing, largely caused by an outside toilet location, as well as owning a goat and frequently sitting on grass. Harvesting vegetables, herding cattle in the forest, and female sex were protective. Age had a nonlinear effect; children and the elderly were more likely to seek treatment for clinical scrub typhus. This study has informed public health policies and awareness programs for healthcare workers through development of National Guidelines for Prevention, Treatment and Control of Scrub Typhus in Bhutan.

Phylogenetic and immunological investigations of complete TSA56 ORF of Orientia tsutsugamushi present in acute encephalitis syndrome cases from eastern Uttar Pradesh, India.

Scrub typhus (ST) caused by Orientia tsutsugamushi (OT), has long been known to cause acute encephalitis syndrome (AES) and acute febrile illness (AFI). The immunodominant 56 kDa protein of OT, which is encoded by the 56 kDa gene (1600 bp encoding 516-541 amino acids) is a commonly studied antigen for genotype and serotype assignment. Previous studies from India have utilized partial type specific antigen (TSA) 56 kDa sequences for OT strain characterisation. On the other hand, understanding the antigenic diversity of current OT strains, is critical for developing specific diagnostic tests and vaccines against ST. As a result, the current study analyses antigenic variants using the entire TSA56 ORF of OT from AES cases. Phylogenetic investigation using complete TSA56 ORF sequences revealed Karp and Gilliam were the circulating predominant strains of OT. Furthermore, Immuno-informatical analysis demonstrated that the majority of high-binding affinity CD4 TCEs against the most prevalent Indian human leukocyte antigen alleles were present in the S-VDIII/IV and S-VDIV spacer regions of TSA56 ORF. TSA56 conserved spacer is crucial for OT immunological response investigations. Further, the pathophysiological effects of spacer domains in ST require further investigation. Furthermore, the characterization of the TSA56 spacer region of the OT from different parts of India is critical for developing region-specific ST diagnostic assays and vaccines.

Doxycycline vs azithromycin in patients with scrub typhus: a systematic review of literature and meta-analysis.

INTRODUCTION: Scrub typhus is a bacterial mite-borne disease associated with poor clinical outcomes if not treated adequately. The study aimed to compare the time to defervescence, clinical failure, mortality and treatment-related adverse effects of two common drugs (doxycycline and azithromycin) used for its treatment. METHODOLOGY: This was a systematic review and meta-analysis. All studies up to 20.03.2023 were screened for eligibility in Pubmed and Embase using a search string containing terms related to scrub typhus, doxycycline and azithromycin. After two phases of screening, all comparative studies where doxycycline and azithromycin were used to treat scrub typhus were included. The studies were critically appraised using standardised tools, and a meta-analysis was performed for time to defervescence (primary outcome), clinical failure, mortality and treatment-related adverse effects. RESULTS: Of 744 articles from two databases, ten were included in the meta-analysis. All but two studies had a high risk of bias. The meta-analysis for time to defervescence had a high heterogeneity and did not show any significant difference between doxycycline and azithromycin arms [Mean difference of -3.37 hours (95%CI: -10.31 to 3.57), p=0.34]. When the analysis was restricted to studies that included only severe scrub typhus, doxycycline was found to have a shorter time to defervescence [mean difference of -10.15 (95%CI: -19.83 to -0.46) hours, p=0.04]. Additionally, there was no difference between the two arms concerning clinical failure, mortality and treatment-related adverse effects. CONCLUSION: The current data from studies with a high risk of bias did not find statistically significant differences in clinical outcomes between doxycycline and azithromycin for scrub typhus.

Epidemiological and clinical characteristics of scrub typhus in northern Fujian, China, from 2015 to 2019.

BACKGROUND: This study aimed to analyze the epidemiological and clinical characteristics of scrub typhus in northern Fujian Province on the southeast coast of China. METHODS: A retrospective analysis was performed on 303 patients with scrub typhus admitted to the First Hospital of Nanping City, Fujian Province, from January 2015 to December 2019. The epidemic characteristics were analyzed, such as the annual number of cases, age distribution, sex distribution, and seasonal distribution in each region. The patient's clinical manifestations, signs, complications, auxiliary examinations, and prognosis were analyzed. RESULTS: From 2015 to 2019, the age distribution of scrub typhus cases was mainly concentrated in 40-49 y (17.16%), 50-59 y (24.09%), and 60-69 y (26.73%). There were no sex differences among the patients. 68.98% of the cases were concentrated in rural areas, with farmers having the highest proportion. However, this study compared prognostic factors in the cured and uncured groups, and found significant differences in non-farmer occupation and diagnosis time ≥ 8 days. Scrub typhus showed two peaks north of Fujian; the prominent peak was from June to July, and the other slight rise was from October to November. The SDE plot showed that the cases were mainly concentrated in Yanping, Shunchang, Zhenghe, and Songxi counties. The number of cases in hilly and mountainous areas was higher than in plain areas. The main diagnostic methods in this area are based on specific eschar and epidemiology, while the positive rate of the Weil-Felix test is low. CONCLUSIONS: The results of this study can guide primary care institutions to improve the level of diagnosis and treatment of scrub typhus and take effective public health intervention measures in endemic areas.

A case of pulmonary tuberculosis patient complicated with hemorrhagic fever with renal syndrome and scrub typhus in Yunnan, China: a case report.

BACKGROUND: Hemorrhagic fever with renal syndrome (HFRS) caused by Orthohantavirus (OHV) and scrub typhus (ST) caused by Orientia tsutsugamushi (OT) are two infectious diseases prevalent in southwest China. Rodents are the natural host and the main source of the two diseases. OT infection to humans is usually resulted from bite of an infective chigger mite on rodents, and OHV is transmitted through contact or inhalation of aerosols and secretions from infected rodent. The use of antibiotics and hormones is crucial for infectious diseases, although the clinical manifestations are not obvious and a definitive diagnosis becomes more difficult in the presence of these drugs. Clinically, fever is the first symptom of these two diseases, and most of them are accompanied by common symptoms such as chills and headaches. The clinical symptoms of these two diseases are very similar and therefore it is not easy to make a differential diagnosis. CASE PRESENTATION: In this case, a 44-year-old male famer with pulmonary tuberculosis and a history of working in coal transportation was admitted to the hospital because of respiratory symptoms accompanied by fever, headache, and skin rashes on his body. Biochemical and urinalysis revealed the hepatic and renal injury. The subsequent molecular testing confirmed he suffered from HFRS and scrub typhus simultaneously that the serological and clinical diagnosis could not identify the cause of infection before. Such case has not been reported in Yunnan Province before. CONCLUSION: The clinical diagnosis should be combined with serological and nucleic acid testing approaches for differential diagnosis in areas where HFRS and ST are endemic.

Determination of a cut-off value for the serological diagnosis of scrub typhus by detecting anti-Orientia tsutsugamushi immunoglobulin M.

Background & objectives: The diagnosis of scrub typhus (ST) is usually done using enzyme-linked immunosorbent assay (ELISA) due to its ease of performance and reading objectivity. The cut-off value for ELISA needs to be calculated for each geographical location as it depends on zonal endemicity of the disease. This study was, therefore, undertaken to calculate the pan-India cut-off for anti-Orientia tsutsugamushi (OT) immunoglobulin M (IgM) by ELISA. Methods: Samples from cases (cases of ST) and controls (voluntary, consenting, healthy adults) were collected by a network of 29 laboratories across India and tested for anti-OT IgM by immunofluorescence assay (IFA), the considered gold standard test. These samples were retested by ELISA for anti-OT IgM and their optical densities (ODs) were used for cut-off estimation by receiver operating characteristic (ROC) curve. Results: Anti-OT IgM ELISA ODs from 273 controls and 136 cases were used for the cut-off estimation. The ODs of the anti-OT IgM ELISA on healthy individuals and those of confirmed ST cases ranged from 0.1 to 0.75 and 0.5 to 4.718, respectively. ROC curve-based cut-off for ELISA was calculated as 0.554 at a sensitivity of 95.2 per cent and specificity of 95.1 per cent. A value of >1 was noted to have a specificity of 100 per cent in diagnosing ST. Interpretation & conclusions: The cut-off calculated for India was similar to the previous cut-off that was used until now.

Crystal structure of a guanine nucleotide exchange factor encoded by the scrub typhus pathogen Orientia tsutsugamushi.

Rho family GTPases regulate an array of cellular processes and are often modulated by pathogens to promote infection. Here, we identify a cryptic guanine nucleotide exchange factor (GEF) domain in the OtDUB protein encoded by the pathogenic bacterium Orientia tsutsugamushi A proteomics-based OtDUB interaction screen identified numerous potential host interactors, including the Rho GTPases Rac1 and Cdc42. We discovered a domain in OtDUB with Rac1/Cdc42 GEF activity (OtDUBGEF), with higher activity toward Rac1 in vitro. While this GEF bears no obvious sequence similarity to known GEFs, crystal structures of OtDUBGEF alone (3.0 Å) and complexed with Rac1 (1.7 Å) reveal striking convergent evolution, with a unique topology, on a V-shaped bacterial GEF fold shared with other bacterial GEF domains. Structure-guided mutational analyses identified residues critical for activity and a mechanism for nucleotide displacement. Ectopic expression of OtDUB activates Rac1 preferentially in cells, and expression of the OtDUBGEF alone alters cell morphology. Cumulatively, this work reveals a bacterial GEF within the multifunctional OtDUB that co-opts host Rac1 signaling to induce changes in cytoskeletal structure.

Detection of co-infection with Orientia tsutsugamushand and hemorrhagic fever with renal syndrome by next-generation sequencing.

Purpose: The co-infection with Orientia tsutsugamushand and hemorrhagic fever with renal syndrome is rare. There are many similarities in early clinical practice between the two diseases, and sometimes it is easy to misdiagnose, especially when co-infection occurs.Methods: We describe a patient who presented with fever and headache after bitten by an insect and whose physical examination showed conjunctival hyperemia, eschar and petechiae in tongue and the soft palate. To lead to a diagnosis, the serum antibody of Hantaan virus, Weil-Felix test and  next-generation sequencing of cerebrospinal fluid was performed.Results: The Weil-Felix test was negative on the 15th day after the onset of the disease and a repeated Weil-Felix test on the 21st day showed a titer of 1:160 and the IgM against Hantaan virus was positive. The number of sequence reads identified corresponding to O. tsutsugamushi was 239 with a genomic coverage of 0.9178%. This patient was diagnosed with intracranial infection with Orientia tsutsugamushi and co-infection with epidemic hemorrhagic fever. The symptoms in our patient quickly decreased after the administration of tetracycline.Conclusion: Next-generation sequencing is helpful for the early diagnosis of scrub typhus, especially when the Weil-Felix test is negative. Clinicians need to be reminded to screen for common pathogens that may be co-infected, such as epidemic hemorrhagic fever.