Detection of Severe Murine Typhus by Nanopore Targeted Sequencing, China.

We report a case of murine typhus in China caused by Rickettsia typhi and diagnosed by nanopore targeted sequencing of a bronchoalveolar lavage fluid sample. This case highlights that nanopore targeted sequencing can effectively detect clinically unexplained infections and be especially useful for detecting infections in patients without typical signs and symptoms.

Increased Seroprevalence of Typhus Group Rickettsiosis, Galveston County, Texas, USA.

Whether increases in typhus group rickettsiosis in Galveston County, Texas, USA, are caused by increased recognition or true reemergence is unclear. We conducted a serosurvey that demonstrated Rickettsia typhi antibodies increased from 1.2% in 2013 to 7.8% in 2021 (p<0.001). These findings support pathogen reemergence rather than enhanced recognition alone.

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.

Severe Rickettsia typhi Infections, Costa Rica.

Murine typhus is a febrile, fleaborne disease caused by infection with Rickettsia typhi bacteria. Cases can range from mild and nonspecific to fatal. We report 2 cases of murine typhus in Costa Rica, confirming the presence and circulation of R. typhi causing severe disease in the country.

Fleaborne Typhus-Associated Deaths - Los Angeles County, California, 2022.

Fleaborne typhus (also known as murine typhus), a widely distributed vectorborne zoonosis caused by Rickettsia typhi, is a moderately severe, but infrequently fatal illness; among patients who receive doxycycline, the case-fatality rate is <1%. Fleaborne typhus is a mandated reportable condition in California. Reported fleaborne typhus cases in Los Angeles County have been increasing since 2010, with the highest number (171) reported during 2022. During June-October 2022, Los Angeles County Department of Public Health learned of three fleaborne typhus-associated deaths. This report describes the clinical presentation, illness course, and methods used to diagnose fleaborne typhus in these three cases. Severe fleaborne typhus manifestations among these cases included hemophagocytic lymphohistiocytosis, a rare immune hyperactivation syndrome that can occur in the infection setting; myocarditis; and septic shock with disseminated intravascular coagulation. Increased health care provider and public health awareness of the prevalence and severity of fleaborne typhus and of the importance of early doxycycline therapy is essential for prevention and treatment efforts.

Zoonotic Pathogens in Wildlife Traded in Markets for Human Consumption, Laos.

We tested animals from wildlife trade sites in Laos for the presence of zoonotic pathogens. Leptospira spp. were the most frequently detected infectious agents, found in 20.1% of animals. Rickettsia typhi and R. felis were also detected. These findings suggest a substantial risk for exposure through handling and consumption of wild animal meat.

Murine Typhus in Canary Islands, Spain, 1999-2015.

To document the epidemiology, clinical features, and outcomes of murine typhus patients in the Canary Islands (Spain), we analyzed data that were retrospectively collected for 16 years for 221 patients. Murine typhus in the Canary Islands is characterized by a high rate of complications (31.6%), mainly liver, lung, kidney or central nervous system involvement.

Rise in Murine Typhus in Galveston County, Texas, USA, 2018.

Murine typhus, an undifferentiated febrile illness caused by Rickettsia typhi, is increasing in prevalence and distribution throughout Texas. In 2018, a total of 40 cases of murine typhus were reported in Galveston County. This increase, unprecedented since the 1940s, highlights the importance of awareness by physicians and public health officials.

The Isolation of Orientia tsutsugamushi and Rickettsia typhi from Human Blood through Mammalian Cell Culture: a Descriptive Series of 3,227 Samples and Outcomes in the Lao People's Democratic Republic.

In the Lao People's Democratic Republic (Laos), rickettsial infections, including scrub and murine typhus, account for a significant burden of fevers. The Mahosot Hospital Microbiology Laboratory in Vientiane, Laos, routinely performs rickettsial isolation from hospitalized patients with suspected rickettsioses using mammalian cell culture systems. We review the clinical and laboratory factors associated with successful Orientia tsutsugamushi and Rickettsia typhi isolations from this laboratory over a period of 6 years between 2008 and 2014. The overall isolation success was 7.9% for all samples submitted and 17.3% for samples for which the patient had a positive O. tsutsugamushi or R. typhi rapid diagnostic test (RDT), serology, or PCR. The frequency of successful isolation was highest for samples submitted in November, at the end of the wet season (28.3%). A longer median duration of reported illness, a positive result for a concurrent Orientia or Rickettsia spp. quantitative PCR, and the use of antibiotics by the patient in the week before admission were significantly associated with isolation success (P < 0.05). Buffy coat inoculation and a shorter interval between sample collection and inoculation in the laboratory were associated with a higher frequency of isolation (both P < 0.05). This frequency was highest if cell culture inoculation occurred on the same day as blood sample collection. Factors related to the initial rickettsial bacterial concentration are likely the main contributors to isolation success. However, modifiable factors do contribute to the rickettsial isolation success, especially delays in inoculating patient samples into culture.

Serum C-reactive protein and procalcitonin values in acute Q fever, scrub typhus, and murine typhus.

BACKGROUND: Although C-reactive protein (CRP) and procalcitonin (PCT) are widely used inflammatory markers for infectious diseases, their role and potential application for rickettsioses were rarely studied. METHODS: A retrospective chart review and serological study were conducted in patients with rickettsioses. The clinical presentations, characteristics, laboratory data, and treatment responses were recorded and their associations with CRP and PCT values were analyzed. RESULTS: A total of 189 cases of rickettsioses, including 115 cases of acute Q fever (60.8%), 55 cases of scrub typhus (29.1%), and 19 cases of murine typhus (10.1%) were investigated. Both CRP and PCT values increased in the acute phase and declined in the convalescent phase. In the acute phase, mean CRP and PCT values were 78.2 ± 63.7 mg/L and 1.05 ± 1.40 ng/mL, respectively. Percentages of patients falling under different cut-off values of CRP and PCT were calculated systematically. Only 10.8% of CRP was > 150 mg/L and 14.2% of PCT was > 2.0 ng/mL. Patients with delayed responses to doxycycline treatment (> 3 days from treatment to defervescence) had significantly higher CRP values (102.7 ± 77.1 vs. 72.2 ± 58.2 mg/L, p = 0.041) and more PCT > 1.0 ng/ml (48.4% vs. 26.0%, p = 0.019) in the acute phase; higher CRP values (19.1 ± 37.4 vs. 3.6 ± 13.1 mg/L, p = 0.049) and more PCT > 0.5 ng/ml (19.2% vs. 1.4%, p = 0.005) in the convalescent phase. Correlation analysis was conducted for patients with acute Q fever. CRP and PCT values were positively correlated to each other, and both markers also had a positive correlation with serum aspartate transaminase values. Both CRP and PCT values and white blood cell counts were positively correlated to the days needed from doxycycline treatment to defervescence. CONCLUSION: CRP and PCT values might be useful in clinical investigations for patients with suspected rickettsioses and in predicting the response to doxycycline treatment for rickettsioses.

Underdiagnoses of Rickettsia in patients hospitalized with acute fever in Indonesia: observational study results.

BACKGROUND: Reports of human rickettsial infection in Indonesia are limited. This study sought to characterize the epidemiology of human rickettsioses amongst patients hospitalized with fever at 8 tertiary hospitals in Indonesia. METHODS: Acute and convalescent blood from 975 hospitalized non-dengue patients was tested for Rickettsia IgM and IgG by ELISA. Specimens from cases with seroconversion or increasing IgM and/or IgG titers were tested for Rickettsia IgM and IgG by IFA and Rickettsia genomes using primers for Rickettsia (R.) sp, R. typhi, and Orientia tsutsugamushi. Testing was performed retrospectively on stored specimens; results did not inform patient management. RESULTS: R. typhi, R. rickettsii, and O. tsutsugamushi IgG antibodies were identified in 269/872 (30.8%), 36/634 (5.7%), and 19/504 (3.8%) of samples, respectively. For the 103/975 (10.6%) non-dengue patients diagnosed with acute rickettsial infection, presenting symptoms included nausea (72%), headache (69%), vomiting (43%), lethargy (33%), anorexia (32%), arthralgia (30%), myalgia (28%), chills (28%), epigastric pain (28%), and rash (17%). No acute rickettsioses cases were suspected during hospitalization. Discharge diagnoses included typhoid fever (44), dengue fever (20), respiratory infections (7), leptospirosis (6), unknown fever (6), sepsis (5), hepatobiliary infections (3), UTI (3), and others (9). Fatalities occurred in 7 (6.8%) patients, mostly with co-morbidities. CONCLUSIONS: Rickettsial infections are consistently misdiagnosed, often as leptospirosis, dengue, or Salmonella typhi infection. Clinicians should include rickettsioses in their differential diagnosis of fever to guide empiric management; laboratories should support evaluation for rickettsial etiologies; and public policy should be implemented to reduce burden of disease.

Assessing human exposure to spotted fever and typhus group rickettsiae in Ontario, Canada (2013-2018): a retrospective, cross-sectional study.

BACKGROUND: Assessing the burden of rickettsial infections in Ontario, Canada, is challenging since rickettsial infections are not reportable to public health. In the absence of reportable disease data, we assessed the burden of rickettsial infections by examining patient serological data and clinical information. METHODS: Our retrospective, cross-sectional study included patients who had Rickettsia serological testing ordered by their physician, in Ontario, from 2013 to 2018. We tested sera from 2755 non-travel patients for antibodies against spotted fever group rickettsiae (SFGR) and typhus group rickettsiae (TGR) using an indirect immunofluorescence assay (IFA) (positive IgG titers ≥1:64). We classified cases using a sensitive surveillance case definition: confirmed (4-fold increase in IgG titers between acute and convalescent sera with clinical evidence of infection), possible (single positive sera with clinical evidence) and previous rickettsial infection (single positive sera without clinical evidence). We classified cases seropositive for both SFGR and TGR as unspecified Rickettsia infections (URIs). RESULTS: Less than 5% of all patients had paired acute and convalescent sera tested, and of these, we found a single, laboratory-confirmed SFGR case, with a 4-fold increase in IgG titers and evidence of fever, maculopapular rash and headache. There were 45 possible (19 SFGR, 7 TGR, 19 URI) and 580 previous rickettsial infection (183 SFGR, 89 TGR, 308 URI) cases. The rate of positive tests for SFGR, TGR and URI combined (all case classifications) were 4.4 per 100,000 population. For confirmed and possible cases, the most common signs and symptoms were fever, headache, gastrointestinal complaints and maculopapular rash. The odds of having seropositive patients increased annually by 30% (odds ratio = 1.3, 95% confidence interval: 1.23-1.39). CONCLUSIONS: The rates of rickettsial infections in Ontario are difficult to determine. Based on confirmed and possible cases, rates are low, but inclusion of previous rickettsial infection cases would indicate higher rates. We highlight the need for education regarding the importance of testing acute and convalescent sera and consistent completion of the laboratory requisition in confirming rickettsial disease. We suggest further research in Ontario to investigate rickettsial agents in potential vectors and clinical studies employing PCR testing of clinical samples.

[Detection of Rickettsia typhi in Rhipicephalus sanguineus s.l. and Amblyomma mixtum in South of Mexico]. / Detección de Rickettsia typhi en Rhipicephalus sanguineus s.l. y Amblyomma mixtum en el sur de México.

OBJECTIVE: To determine the presence of Rickettsia typhi in Rhipicephalus sanguineus s.l. and Amblyomma mixtum in southern Mexico. MATERIALS AND METHODS: Ticks were collected in humans and domestic animals. The presence of Rickettsia was determined by PCR and sequencing. RESULTS: 10/39 work vials amplified fragments of the gltA, htrA and ompB genes. On 7/10 from Rh. sanguineus s.l. collected from dogs and in 3/10 of A. mixtum collected from horse and human. Sequencing indicated R. typhi in Rh. sanguineus and A. mixtum with 100% homology (LS992663.1) for a region of the htrA gene and 99% (LS992663.1) with the regions of the gltA and OmpB genes. The minimum infection rate (TMI) for R. typhi was 3.88. CONCLUSIONS: Rhipicephalus sanguineus s.l. and Amblyomma mixtum are naturally infected with R. typhi in Southern Mexico.

OBJETIVO: Determinar la presencia de Rickettsia typhi en Rhipicephalus sanguineus s.l. y Amblyomma mixtum, en el sur de México. MATERIAL Y MÉTODOS: Las garrapatas fueron colectadas en humanos y animales domésticos. Se determinó la presencia de Rickettsia por reacción en cadena de la polimerasa (PCR, por sus siglas en inglés) y secuenciación. RESULTADOS: 10/39 viales de trabajo amplificaron fragmentos de los genes gltA, htrA y ompB, en 7/10 proveniente de Rh. sanguineus s.l. colectadas de perros y en 3/10 de A. mixtum colectadas de caballo y humano. La secuenciación indicó R. typhi en Rh. sanguineus y A. mixtum con homología de 100% (LS992663.1), para una región del gen de htrA, y de 99% (LS992663.1), con las regiones de los genes de gltA y OmpB. La tasa mínima de infección (TMI) para R. typhi fue de 3.88. CONCLUSIONES: Las garrapatas Rhipicephalus sanguineus s.l. y Amblyomma mixtum están infectadas naturalmente con R. typhi en el sur de México.

Laboratory-acquired Scrub Typhus and Murine Typhus Infections: The Argument for a Risk-based Approach to Biosafety Requirements for Orientia tsutsugamushi and Rickettsia typhi Laboratory Activities.

This study examined the literature on laboratory-acquired infections (LAIs) associated with scrub typhus (Orientia tsutsugamushi) and murine typhus (Rickettsia typhi) research to provide an evidence base for biosafety and biocontainment. Scrub typhus LAIs were documented in 25 individuals, from 1931 to 2000 with 8 (32%) deaths during the preantibiotic era. There were 35 murine typhus LAI reports and no deaths. Results indicated that the highest-risk activities were working with infectious laboratory animals involving significant aerosol exposures, accidental self-inoculation, or bite-related infections. A risk-based biosafety approach for in vitro and in vivo culture of O. tsutsugamushi and R. typhi would require that only high-risk activities (animal work or large culture volumes) be performed in high-containment biosafety level (BSL) 3 laboratories. We argue that relatively low-risk activities including inoculation of cell cultures or the early stages of in vitro growth using low volumes/low concentrations of infectious materials can be performed safely in BSL-2 laboratories within a biological safety cabinet.

Rickettsia Lipid A Biosynthesis Utilizes the Late Acyltransferase LpxJ for Secondary Fatty Acid Addition.

Members of the Rickettsia genus are obligate intracellular, Gram-negative coccobacilli that infect mammalian and arthropod hosts. Several rickettsial species are human pathogens and are transmitted by blood-feeding arthropods. In Gram-negative parasites, the outer membrane (OM) sits at the nexus of the host-pathogen interaction and is rich in lipopolysaccharide (LPS). The lipid A component of LPS anchors the molecule to the bacterial surface and is an endotoxic agonist of Toll-like receptor 4 (TLR4). Despite the apparent importance of lipid A in maintaining OM integrity, as well as its inflammatory potential during infection, this molecule is poorly characterized in Rickettsia pathogens. In this work, we have identified and characterized new members of the recently discovered LpxJ family of lipid A acyltransferases in both Rickettsia typhi and Rickettsia rickettsii, the etiological agents of murine typhus and Rocky Mountain spotted fever, respectively. Our results demonstrate that these enzymes catalyze the addition of a secondary acyl chain (C14/C16) to the 3'-linked primary acyl chain of the lipid A moiety in the final steps of the Raetz pathway of lipid A biosynthesis. Since lipid A architecture is fundamental to bacterial OM integrity, we believe that rickettsial LpxJ may be important in maintaining membrane dynamics to facilitate molecular interactions at the host-pathogen interface that are required for adhesion and invasion of mammalian cells. This work contributes to our understanding of rickettsial outer membrane physiology and sets a foundation for further exploration of the envelope and its role in pathogenesis.IMPORTANCE Lipopolysaccharide (LPS) triggers an inflammatory response through the TLR4-MD2 receptor complex and inflammatory caspases, a process mediated by the lipid A moiety of LPS. Species of Rickettsia directly engage both extracellular and intracellular immunosurveillance, yet little is known about rickettsial lipid A. Here, we demonstrate that the alternative lipid A acyltransferase, LpxJ, from Rickettsia typhi and R. rickettsii catalyzes the addition of C16 fatty acid chains into the lipid A 3'-linked primary acyl chain, accounting for major structural differences relative to the highly inflammatory lipid A of Escherichia coli.

The Cat Flea (Ctenocephalides felis) Immune Deficiency Signaling Pathway Regulates Rickettsia typhi Infection.

Rickettsia species are obligate intracellular bacteria with both conserved and lineage-specific strategies for invading and surviving within eukaryotic cells. One variable component of Rickettsia biology involves arthropod vectors: for instance, typhus group rickettsiae are principally vectored by insects (i.e., lice and fleas), whereas spotted fever group rickettsiae are exclusively vectored by ticks. For flea-borne Rickettsia typhi, the etiological agent of murine typhus, research on vertebrate host biology is facilitated using cell lines and animal models. However, due to the lack of any stable flea cell line or a published flea genome sequence, little is known regarding R. typhi biology in flea vectors that, importantly, do not suffer lethality due to R. typhi infection. To address if fleas combat rickettsial infection, we characterized the cat flea (Ctenocephalides felis) innate immune response to R. typhi Initially, we determined that R. typhi infects Drosophila cells and increases antimicrobial peptide (AMP) gene expression, indicating immune pathway activation. While bioinformatics analysis of the C. felis transcriptome identified homologs to all of the Drosophila immune deficiency (IMD) and Toll pathway components, an AMP gene expression profile in Drosophila cells indicated IMD pathway activation upon rickettsial infection. Accordingly, we assessed R. typhi-mediated flea IMD pathway activation in vivo using small interfering RNA (siRNA)-mediated knockdown. Knockdown of Relish and Imd increased R. typhi infection levels, implicating the IMD pathway as a critical regulator of R. typhi burden in C. felis These data suggest that targeting the IMD pathway could minimize the spread of R. typhi, and potentially other human pathogens, vectored by fleas.

Typhus Group Rickettsiosis, Germany, 2010-20171.

Typhus group rickettsiosis is caused by the vectorborne bacteria Rickettsia typhi and R. prowazekii. R. typhi, which causes murine typhus, the less severe endemic form of typhus, is transmitted by fleas; R. prowazekii, which causes the severe epidemic form of typhus, is transmitted by body lice. To examine the immunology of human infection with typhus group rickettsiae, we retrospectively reviewed clinical signs and symptoms, laboratory changes, and travel destinations of 28 patients who had typhus group rickettsiosis diagnosed by the German Reference Center for Tropical Pathogens, Hamburg, Germany, during 2010-2017. Immunofluorescence assays of follow-up serum samples indicated simultaneous seroconversion of IgM, IgA, and IgG or concurrence in the first serum sample. Cytokine levels peaked during the second week of infection, coinciding with organ dysfunction and seroconversion. For 3 patients, R. typhi was detected by species-specific nested quantitative PCR. For all 28 patients, R. typhi was the most likely causative pathogen.

Rickettsia typhi as Cause of Fatal Encephalitic Typhus in Hospitalized Patients, Hamburg, Germany, 1940-1944.

We evaluated formalin-fixed paraffin-embedded tissue specimens from 7 patients who died with encephalitic typhus in Hamburg, Germany, during World War II. The archived specimens included only central nervous system tissues >70 years old that had been stored at room temperature. We demonstrated successful detection of Rickettsia typhi DNA by a nested qPCR specific to prsA in 2 patients. These results indicate that R. typhi infections contributed to typhus outbreaks during World War II. Immunohistochemical analyses of brain tissue specimens of R. typhi DNA-positive and -negative specimens showed perivascular B-cell accumulation. Around blood vessels, nodular cell accumulations consisted of CD4-positive and CD8-positive T cells and CD68-positive microglia and macrophages; neutrophils were found rarely. These findings are similar to those of previously reported R. prowazekii tissue specimen testing. Because R. typhi and R. prowazekii infections can be clinically and histopathologically similar, molecular analyses should be performed to distinguish the 2 pathogens.

Orientia tsutsugamushi Is Highly Susceptible to the RNA Polymerase Switch Region Inhibitor Corallopyronin A In Vitro and In Vivo.

Scrub typhus is a potentially lethal infection caused by the obligate intracellular bacterium Orientia tsutsugamushi Reports on the emergence of doxycycline-resistant strains highlight the urgent need to develop novel antiinfectives against scrub typhus. Corallopyronin A (CorA) is a novel α-pyrone compound synthesized by the myxobacterium Corallococcus coralloides that was characterized as a noncompetitive inhibitor of the switch region of the bacterial RNA polymerase (RNAP). We investigated the antimicrobial action of CorA against the human-pathogenic Karp strain of O. tsutsugamushiin vitro and in vivo The MIC of CorA against O. tsutsugamushi was remarkably low (0.0078 µg/ml), 16-fold lower than that against Rickettsia typhi In the lethal intraperitoneal O. tsutsugamushi mouse infection model, a minimum daily dose of 100 µg CorA protected 100% of infected mice. Two days of treatment were sufficient to confer protection. In contrast to BALB/c mice, SCID mice succumbed to the infection despite treatment with CorA or tetracycline, suggesting that antimicrobial treatment required synergistic action of the adaptive immune response. Similar to tetracycline, CorA did not prevent latent infection of O. tsutsugamushiin vivo However, latency was not caused by acquisition of antimicrobial resistance, since O. tsutsugamushi reisolated from latently infected BALB/c mice remained fully susceptible to CorA. No mutations were found in the CorA-binding regions of the ß and ß' RNAP subunit genes rpoB and rpoC Inhibition of the RNAP switch region of O. tsutsugamushi by CorA is therefore a novel and highly potent target for antimicrobial therapy for scrub typhus.

Structural Insights into Substrate Recognition and Catalysis in Outer Membrane Protein B (OmpB) by Protein-lysine Methyltransferases from Rickettsia.

Rickettsia belong to a family of Gram-negative obligate intracellular infectious bacteria that are the causative agents of typhus and spotted fever. Outer membrane protein B (OmpB) occurs in all rickettsial species, serves as a protective envelope, mediates host cell adhesion and invasion, and is a major immunodominant antigen. OmpBs from virulent strains contain multiple trimethylated lysine residues, whereas the avirulent strain contains mainly monomethyllysine. Two protein-lysine methyltransferases (PKMTs) that catalyze methylation of recombinant OmpB at multiple sites with varying sequences have been identified and overexpressed. PKMT1 catalyzes predominantly monomethylation, whereas PKMT2 catalyzes mainly trimethylation. Rickettsial PKMT1 and PKMT2 are unusual in that their primary substrate appears to be limited to OmpB, and both are capable of methylating multiple lysyl residues with broad sequence specificity. Here we report the crystal structures of PKMT1 from Rickettsia prowazekii and PKMT2 from Rickettsia typhi, both the apo form and in complex with its cofactor S-adenosylmethionine or S-adenosylhomocysteine. The structure of PKMT1 in complex with S-adenosylhomocysteine is solved to a resolution of 1.9 Å. Both enzymes are dimeric with each monomer containing an S-adenosylmethionine binding domain with a core Rossmann fold, a dimerization domain, a middle domain, a C-terminal domain, and a centrally located open cavity. Based on the crystal structures, residues involved in catalysis, cofactor binding, and substrate interactions were examined using site-directed mutagenesis followed by steady state kinetic analysis to ascertain their catalytic functions in solution. Together, our data reveal new structural and mechanistic insights into how rickettsial methyltransferases catalyze OmpB methylation.