Epidemiology of tsutsugamushi disease and its relationship with meteorological factors in Xiamen city, China.

Tsutsugamushi disease (TD) is an acute infectious disease caused by Orientia tsutsugamushi. This study aimed to analyze the epidemiological features of TD, investigate chigger mites and their hosts, and investigate the meteorological factors affecting TD incidence and the host of O. tsutsugamushi in Xiamen city, China. Data on reported TD cases were collected from 2006 to 2018. Spearman's correlation test were used for identifying the relationship between meteorological factors and TD incidence and whether meteorological factors affect the host of O. tsutsugamushi. The incidence of reported TD increased gradually from 2006, reached a peak of 4.59 per 100,000 persons in 2014, and then decreased gradually. The TD incidence was seasonal, with epidemic periods occurred mainly in summer and autumn. Patients aged 40-60 years had the highest proportion of cases, accounting for 44.44% of the total cases. Farmers had the largest number of cases among all occupational groups. Rattus Norvegicus was the most common host, accounting for the largest proportion of rats (73.00%), and the highest rat density was observed in March and October every year. There were significant positive correlations between the number of reported cases and average temperature, sunshine duration, and rainfall as well as between rat density and average temperature. On phylogenetic analysis, 7 sequences of hosts and human TD cases obtained from health records demonstrated the highest similarities to the Kato, Karp, and Gilliam strains. No correlations were observed between rat density, and sunshine duration and rainfall. The transmission of TD in Xiamen city, China, was seasonal, and its incidence was affected by several meteorological factors including average temperature, sunshine duration, and rainfall. However, the host of O. tsutsugamushi was only affected by average temperature.

Scrub typhus ecology: a systematic review of Orientia in vectors and hosts.

Scrub typhus, caused by Orientia tsutsugamushi, is an important and neglected vector-borne zoonotic disease with an expanding known distribution. The ecology of the disease is complex and poorly understood, impairing discussion of public health interventions. To highlight what we know and the themes of our ignorance, we conducted a systematic review of all studies investigating the pathogen in vectors and non-human hosts. A total of 276 articles in 7 languages were included, with 793 study sites across 30 countries. There was no time restriction for article inclusion, with the oldest published in 1924. Seventy-six potential vector species and 234 vertebrate host species were tested, accounting for over one million trombiculid mites ('chiggers') and 83,000 vertebrates. The proportion of O. tsutsugamushi positivity was recorded for different categories of laboratory test and host species. Vector and host collection sites were geocoded and mapped. Ecological data associated with these sites were summarised. A further 145 articles encompassing general themes of scrub typhus ecology were reviewed. These topics range from the life-cycle to transmission, habitats, seasonality and human risks. Important gaps in our understanding are highlighted together with possible tools to begin to unravel these. Many of the data reported are highly variable and inconsistent and minimum data reporting standards are proposed. With more recent reports of human Orientia sp. infection in the Middle East and South America and enormous advances in research technology over recent decades, this comprehensive review provides a detailed summary of work investigating this pathogen in vectors and non-human hosts and updates current understanding of the complex ecology of scrub typhus. A better understanding of scrub typhus ecology has important relevance to ongoing research into improving diagnostics, developing vaccines and identifying useful public health interventions to reduce the burden of the disease.

Analysis of the 56-kDa type specific antigen gene of Orientia tsutsugamushi from northern Vietnam.

Scrub typhus has been documented since 1932 in Vietnam, however, the disease burden of scrub typhus remains poorly understood in the country. We conducted this study to describe the phylogenetic analysis of the 56-kDa type-specific antigen (TSA) gene of Orientia tsutsugamushi associated with PCR positive cases of scrub typhus. Of 116 positive samples, 65 type-specific antigen gene sequences were obtained and classified into 3 genogroups: Karp, Kato and Gilliam. The Karp genogroup was the most frequently detected phylogenetic cluster in the study with 30 samples (46%), followed by Kato and Gilliam with 20 (31%) and 15 (23%), respectively. All sequences showed 94-100% nucleotide similarity to reference sequences collected in the central part of Vietnam in 2017. Patients infected with Karp genogroup were more likely to have significant thrombocytopenia than the other genogroups. These results suggest that any scrub typhus vaccine considered for use in Vietnam should provide protection against each of these 3 genogroups.

Immunization with a recombinant antigen composed of conserved blocks from TSA56 provides broad genotype protection against scrub typhus.

Scrub typhus is an acute febrile disease caused by Orientia tsutsugamushi infection. Despite the wide range of approaches explored during the last seventy years, an effective prophylactic vaccine is not yet available. Here, we developed a novel recombinant antigen derived from conserved regions of 56 kDa type-specific antigen (TSA56), a major outer membrane protein responsible for genetic heterogeneity and antigenicity, and evaluated it as a protective vaccine antigen. Our findings demonstrate that immunization with conserved blocks of TSA56 (cTSA56) not only provides protective immunity against lethal challenges with the homologous genotype, but also confers significantly better protection against heterologous genotypes than TSA56. Adoptive transfer of CD4+ or CD8+ T cells from immunized mice provided significantly enhanced protection against lethal challenge, whereas immune B cells failed to do so, indicating that cellular immunity against the conserved epitopes plays a protective role. Moreover, immunization with a 10-mer peptide mixture, screened from CD8+ T cell epitopes within the conserved region of TSA56, provided enhanced protection against lethal challenge with O. tsutsugamushi. Therefore, this novel recombinant antigen is a promising candidate for scrub typhus vaccine against a wide range of O. tsutsugamushi genotypes.

Genetic diversity of Orientia tsutsugamushi strains from patients in north India.

BACKGROUND: Scrub typhus has emerged as a major cause of acute febrile illness in India in recent years. The causative agent, Orientia tsutsugamushi has more than 20 prototype strains due to a variable 56-kDa outer membrane protein. It is crucial to know the prevailing types in India for the success of diagnostic immunoassays and prospective vaccine candidates. In north India, the principal types circulating are largely unknown. Our tertiary care hospital caters to a large area of north India (around 7 states and one union territory). Therefore, the current study was planned to identify the genotypes of O. tsutsugamushi strains circulating in this wide area of north India. MATERIALS AND METHODS: Adults and children presenting with suspected scrub typhus between July 2013 and December 2013 were included in this study. DNA was extracted from whole blood and a nested PCR was used to amplify a 483-bp region of the 56-kDa antigen gene of O. tsutsugamushi. The PCR products were purified and DNA sequencing was performed and aligned using the CLUSTAL_X2 program. A phylogenetic tree was constructed using neighbour- joining algorithms and analysed using the sequences obtained in this study and those obtained from the GenBank database. RESULTS: A total of 34 samples were positive for PCR. The amplicons were sequenced and analyzed. Karp-like strains predominated in all states studied (64.7%) followed by Gilliam-like (26.47 %) and 8.82% similar to Hualien 1 or S072. We did not find any Kato or Kawasaki-like strain. Karp like strains showed >99% similarity to TH2033, TH2191, TH2208, Xinjiang & Neimeng strains and Gilliam-like strains showed >99% similarity to Clone ISS -11. CONCLUSION: Orientia tsutsugamushi shows a great diversity in its strains over a large geographical area of north India. This has implications in the production of both diagnostic assays and vaccine for scrub typhus.

Phylogenetic diversity of Orientia tsutsugamushi isolates in patients with scrub typhus in Bengaluru, India.

Scrub typhus has re-emerged as an important cause of acute febrile illness in India. There is a dearth of information on strain diversity of Orientia tsutsugamushi from Karnataka, India, hence the present study sought to address this issue. One hundred clinically suspected cases of scrub typhus/rickettsiosis (as per the DHR-ICMR guidelines) were included. Nested-polymerase chain reaction (PCR) for 56-kDa gene and phylogenetic analysis was performed. PCR was positive in 22 cases and phylogenetic analysis showed the presence of different strains, with predominance of clustering (57%) with Gilliam-type for the first time in Karnataka. Knowledge of genetic diversity has implications in development of diagnostics and vaccine.

Use of eschar for the molecular diagnosis and genotypic characterisation of Orientia tsutsugamushi causing scrub typhus.

Scrub typhus caused by Orientia tsutsugamushi presents as an acute febrile illness with a varied presentation from mild illness to fatal disease in the absence of appropriate antibiotic treatment. Performing polymerase chain reaction (PCR) on eschar sample acts a rapid diagnostic tool in the early stage of scrub typhus when blood is negative. A total of eight patients from whom both whole blood and eschar samples were collected and tested by nested PCR targeting 56 kDa trichostatin A (TSA) gene to detect O. tsutsugamushi DNA. All (100%) eschar samples and three whole blood samples tested positive. Genetic analysis of the 56 kDa TSA gene sequences showed that the majority were related to Karp reference strains, while one clustered with Kawasaki strain. When present, eschar should be favoured as a diagnostic sample over whole blood in the early phase of infection.

Identification and Characterization of Orientia chuto in Trombiculid Chigger Mites Collected from Wild Rodents in Kenya.

We present data that concurs with the reported geographical expansion of scrub typhus outside the "Tsutsugamushi Triangle" and addition of Orientia chuto as a second species in the Orientia genus. Wild rodents were caught in Marigat, Baringo County, Kenya, and ectoparasites, including chiggers, were recovered. Rodent and chigger species were identified by taxonomic features. DNA was extracted from the chiggers and used to amplify and/or sequence the 47-kDa high temperature transmembrane protein (TSA47), the 56-kDa type-specific antigen (TSA56), and the 16S rRNA (rrs) Orientia genes. The main rodent hosts identified were Acomys wilsoni, Crocidura sp., and Mastomys natalensis, which accounted for 59.2% of the total collection. Of these, A. wilsoni and M. natalensis harbored most of the chiggers that belonged to the Neotrombicula and Microtrombicula genera. A pool of chiggers from one of M. natalensis was positive for Orientia by TSA47 PCR, but Orientia did not amplify with the TSA56 primers. On sequencing the 850 bp of the TSA47 gene, the closest phylogenetic relative was O. chuto, with 97.65% sequence homology compared to 84.63 to 84.76% for O. tsutsugamushi 16S rRNA deep sequencing also revealed O. chuto as the closest phylogenetic relative, with 99.75% sequence homology. These results and the existing immunological and molecular reports are strongly suggestive of the existence of Orientia species in Kenya.

Detection and distribution of Sca autotransporter protein antigens in diverse isolates of Orientia tsutsugamushi.

Orientia tsutsugamushi (Ots) frequently causes severe scrub typhus infections in the Asia-Pacific region. Korean investigators have demonstrated that Ots encodes five different autotransporter domain (ATD) proteins (ScaA-ScaE). ScaA functions as an adhesin and confers protective immunity in a lethal mouse model of Ots infection. Specific antibodies are detected against ScaA and ScaC in Korean scrub typhus patients. However, there is limited data on the distribution of the Sca protein genes in diverse isolates of Ots. By BLAST analysis with the conserved beta barrel autotransporter domain (ATD) regions of the sca proteins, we discovered a sixth gene scaF among 3 of 10 new partial Ots genome sequences available at NCBI GenBank (Sido, Karp, AFSC7). We designed two to seven specific TaqMan assays to detect the ATD for each of the six sca genes. The TaqMan assays among those for each sca gene which gave the greatest sensitivity and linearity with DNA log dilutions were then used for screening DNAs from Ots isolates grown in L929 mouse cells for sca genes. The sca prevalence survey was performed for all six sca genes with 178 DNAs from isolates from 12 countries. The survey results were confirmed by conventional PCR with primers from conserved regions of the passenger domains (PD) and ATD of the sca proteins. The ATD was highly conserved between the DNAs of different genotypes compared to the sca PD but each TaqMan assay was sca specific. The percentage positivity for 56 kDa and scaA genes in the 178 DNAs using Ha primers was 59.6% and 62.4%, respectively. Our scaA conventional ATD PCR assay was positive in 98.3% but scaA was present in all 178 DNAs (100%) by ATD TaqMan. scaB, scaC, scaD, scaE and scaF were detected in 33.7%, 97.8%, 93.8%, 97.2% and 43.3% isolates by ATD TaqMan, respectively. The ATDs of Ots sca genes are thus sufficiently conserved between different genotypes for molecular assay design. Four sca genes are widely distributed among diverse Ots isolates from diverse geographical areas. scaB and scaF were detected in fewer Ots isolates and absent from some available genome sequences. Whether the utility of the ScaA, ScaC, ScaD, and ScaE antigenic passenger protein domains exceeds that of the mixed 56 kDa type surface antigens of Ots now used in combination diagnostic assays needs to be determined before they can be considered as suitable alternative serological antigens for diagnosis of scrub typhus.

Heterogeneity of Orientia tsutsugamushi genotypes in field-collected trombiculid mites from wild-caught small mammals in Thailand.

Trombiculid mites are the vectors of scrub typhus, with infected larval mites (chiggers) transmitting the causative agent, Orientia tsutsugamushi, during feeding. Co-existence of multiple O. tsutsugamushi strains within infected mites has previously been reported in naturally infected, laboratory-reared mite lines using molecular methods to characterize the 56-kDa type-specific antigen (TSA) gene. In the current study, more advanced next-generation sequencing technology was used to reveal the heterogeneity of O. tsutsugamushi genotypes in field-collected trombiculid mites from rodents and small mammals in scrub typhus-endemic areas of Thailand. Twenty-eight trombiculid mites collected from 10 small mammals were positive for O. tsutsugamushi, corresponding to a prevalence rate of 0.7% within the mite population. Twenty-four of the infected mites were Leptotrombidium spp., indicating that this genus is the main vector for O. tsutsugamushi transmission in Thailand. In addition, O. tsutsugamushi was detected in the mite genera Ascoschoengastia, Blankaartia, Gahrliepia, and Lorillatum. Of the 10 infested small animal hosts, six had 2-10 infected mites feeding at the time of collection. Deep sequencing was used to characterize mixed infections (two to three O. tsutsugamushi genotypes within an individual mite), and 5 of the 28 infected mites (17.9%) contained mixed infections. Additionally, 56-kDa TSA gene sequence analysis revealed identical bacterial genotypes among co-feeding mites with single or mixed infections. These results suggest that co-feeding transmission may occur during the feeding process, and could explain the occurrence of mixed infections in individual mites, as well as the recovery of multiple infected mites from the same host. This study also revealed highly diverse within-host O. tsutsugamushi genotypes. The occurrence of multiple O. tsutsugamushi genotypes within individual mites has important implications, and could provide a mechanism for pathogen evolution/diversification in the mite vector.

Dual Genotype Orientia tsutsugamushi Infection in Patient with Rash and Eschar, Vietnam, 2016.

We report a dual genotype Orientia tsutsugamushi infection in Vietnam in 2016. The patient had fever, rash, and an eschar. The Kawasaki genotype was identified in the eschar specimen and Karp genotype in the whole blood specimen. The genotype co-infection rate for scrub typhus is unknown and should be further evaluated.

Comparative pan-genomic analyses of Orientia tsutsugamushi reveal an exceptional model of bacterial evolution driving genomic diversity.

Orientia tsutsugamushi, formerly Rickettsia tsutsugamushi, is an obligate intracellular pathogen that causes scrub typhus, an underdiagnosed acute febrile disease with high morbidity. Scrub typhus is transmitted by the larval stage (chigger) of Leptotrombidium mites and is irregularly distributed across endemic regions of Asia, Australia and islands of the western Pacific Ocean. Previous work to understand population genetics in O. tsutsugamushi has been based on sub-genomic sampling methods and whole-genome characterization of two genomes. In this study, we compared 40 genomes from geographically dispersed areas and confirmed patterns of extensive homologous recombination likely driven by transposons, conjugative elements and repetitive sequences. High rates of lateral gene transfer (LGT) among O. tsutsugamushi genomes appear to have effectively eliminated a detectable clonal frame, but not our ability to infer evolutionary relationships and phylogeographical clustering. Pan-genomic comparisons using 31 082 high-quality bacterial genomes from 253 species suggests that genomic duplication in O. tsutsugamushi is almost unparalleled. Unlike other highly recombinant species where the uptake of exogenous DNA largely drives genomic diversity, the pan-genome of O. tsutsugamushi is driven by duplication and divergence. Extensive gene innovation by duplication is most commonly attributed to plants and animals and, in contrast with LGT, is thought to be only a minor evolutionary mechanism for bacteria. The near unprecedented evolutionary characteristics of O. tsutsugamushi, coupled with extensive intra-specific LGT, expand our present understanding of rapid bacterial evolutionary adaptive mechanisms.

Geographical distribution of Orientia tsutsugamushi strains in chiggers from three provinces in Korea.

Chiggers were collected from the central and southern parts of South Korea between April and November, 2009 with the aim of investigating the seasonal and geographical distribution of Or. A total of 1136 chiggers were identified. They included eight species belonging to four genera, as follows: Leptotrombidium scutellare (27.2%, n = 309), L. pallidum (54.6%, n = 621), L. orientale (6.25%, n = 71), L. palpale (1.59%, n = 18), L. zetum (2.0%, n = 23), Euschoengastia koreaensis (1.5%, n = 17), Cheladonta ikaoensis (0.08%, n = 1) and Neotrombicula japonica (1.05%, n = 12). The density of L. pallidum was high from April to May, whereas L. scutallare was not found in spring, being observed from October. Serotype-specific nested PCR targeting the 56 kDa protein gene and sequencing analysis identified that the strains of 1136 O. tsutsugamushi in the chiggers as Boryong (6.8%), Kanda (0.4%), Oishi (0.3%), Jecheon (0.1%), Youngworl (0.1%) and Wonju (0.1%). Our findings indicate that L. pallidum and L. scutellare are dominant species in Korea and have geographical and seasonal variations.

Long-read whole genome sequencing and comparative analysis of six strains of the human pathogen Orientia tsutsugamushi.

BACKGROUND: Orientia tsutsugamushi is a clinically important but neglected obligate intracellular bacterial pathogen of the Rickettsiaceae family that causes the potentially life-threatening human disease scrub typhus. In contrast to the genome reduction seen in many obligate intracellular bacteria, early genetic studies of Orientia have revealed one of the most repetitive bacterial genomes sequenced to date. The dramatic expansion of mobile elements has hampered efforts to generate complete genome sequences using short read sequencing methodologies, and consequently there have been few studies of the comparative genomics of this neglected species. RESULTS: We report new high-quality genomes of O. tsutsugamushi, generated using PacBio single molecule long read sequencing, for six strains: Karp, Kato, Gilliam, TA686, UT76 and UT176. In comparative genomics analyses of these strains together with existing reference genomes from Ikeda and Boryong strains, we identify a relatively small core genome of 657 genes, grouped into core gene islands and separated by repeat regions, and use the core genes to infer the first whole-genome phylogeny of Orientia. CONCLUSIONS: Complete assemblies of multiple Orientia genomes verify initial suggestions that these are remarkable organisms. They have larger genomes compared with most other Rickettsiaceae, with widespread amplification of repeat elements and massive chromosomal rearrangements between strains. At the gene level, Orientia has a relatively small set of universally conserved genes, similar to other obligate intracellular bacteria, and the relative expansion in genome size can be accounted for by gene duplication and repeat amplification. Our study demonstrates the utility of long read sequencing to investigate complex bacterial genomes and characterise genomic variation.

Peptidoglycan in obligate intracellular bacteria.

Peptidoglycan is the predominant stress-bearing structure in the cell envelope of most bacteria, and also a potent stimulator of the eukaryotic immune system. Obligate intracellular bacteria replicate exclusively within the interior of living cells, an osmotically protected niche. Under these conditions peptidoglycan is not necessarily needed to maintain the integrity of the bacterial cell. Moreover, the presence of peptidoglycan puts bacteria at risk of detection and destruction by host peptidoglycan recognition factors and downstream effectors. This has resulted in a selective pressure and opportunity to reduce the levels of peptidoglycan. In this review we have analysed the occurrence of genes involved in peptidoglycan metabolism across the major obligate intracellular bacterial species. From this comparative analysis, we have identified a group of predicted 'peptidoglycan-intermediate' organisms that includes the Chlamydiae, Orientia tsutsugamushi, Wolbachia and Anaplasma marginale. This grouping is likely to reflect biological differences in their infection cycle compared with peptidoglycan-negative obligate intracellular bacteria such as Ehrlichia and Anaplasma phagocytophilum, as well as obligate intracellular bacteria with classical peptidoglycan such as Coxiella, Buchnera and members of the Rickettsia genus. The signature gene set of the peptidoglycan-intermediate group reveals insights into minimal enzymatic requirements for building a peptidoglycan-like sacculus and/or division septum.

Orientia tsutsugamushi infection in rodents in Anhui Province of China.

We conducted an investigation of Orientia tsutsugamushi infection among rodents in non-endemic areas in Anhui Province. Fifty-six (56) rodents including 44 Apodemus agrarius and 12 Niviventer niviventer were trapped and captured in autumn in the fields of three counties in Anhui Province. DNA samples were amplified and sequenced for the 56kDa protein gene of Orientia tsutsugamushi. The overall infection rate in the rodents was 23.2%(13/56). The rate of detection of O. tsutsugamushi in Apodemus agrarius and Niviventer niviventer were 27.3% and 8.3% respectively. Moreover, we identified two genotypes (Karp and Gilliam strains) of Orientia tsutsugamushi in rodents. Our study demonstrated that Apodemus agrarius is the main host for O. tsutsugamushi pathogen and this is the first report of Karp and Gilliam strains in these non-endemic areas in Anhui Province.

Seroprevalence of Scrub Typhus, Typhus, and Spotted Fever Among Rural and Urban Populations of Northern Vietnam.

AbstractRickettsial infections are recognized as important causes of fever throughout southeast Asia. Herein, we determined the seroprevalence to rickettsioses within rural and urban populations of northern Vietnam. Prevalence of individuals with evidence of prior rickettsial infections (IgG positive) was surprisingly low, with 9.14% (83/908) testing positive to the three major rickettsial serogroups thought to circulate in the region. Prevalence of typhus group rickettsiae (TG)-specific antibodies (6.5%, 58/908) was significantly greater than scrub typhus group orientiae (STG)- or spotted fever group rickettsiae (SFG)-specific antibodies (P < 0.05). The majority of TG seropositives were observed among urban rather than rural residents (P < 0.05). In contrast, overall antibody prevalence to STG and SFG were both very low (1.1%, 10/908 for STG; 1.7%, 15/908 for SFG), with no significant differences between rural and urban residents. These results provide data on baseline population characteristics that may help inform development of Rickettsia serological testing criteria in future clinical studies.

Diversification of Orientia tsutsugamushi genotypes by intragenic recombination and their potential expansion in endemic areas.

BACKGROUND: Scrub typhus is a mite-borne febrile disease caused by O. tsutsugamushi infection. Recently, emergence of scrub typhus has attracted considerable attention in several endemic countries in Asia and the western Pacific. In addition, the antigenic diversity of the intracellular pathogen has been a serious obstacle for developing effective diagnostics and vaccine. METHODOLOGY/PRINCIPAL FINDINGS: To understand the evolutionary pathway of genotypic diversification of O. tsutsugamushi and the environmental factors associated with the epidemiological features of scrub typhus, we analyzed sequence data, including spatiotemporal information, of the tsa56 gene encoding a major outer membrane protein responsible for antigenic variation. A total of 324 tsa56 sequences covering more than 85% of its open reading frame were analyzed and classified into 17 genotypes based on phylogenetic relationship. Extensive sequence analysis of tsa56 genes using diverse informatics tools revealed multiple intragenic recombination events, as well as a substantially higher mutation rate than other house-keeping genes. This suggests that genetic diversification occurred via frequent point mutations and subsequent genetic recombination. Interestingly, more diverse bacterial genotypes and dominant vector species prevail in Taiwan compared to other endemic regions. Furthermore, the co-presence of identical and sub-identical clones of tsa56 gene in geographically distant areas implies potential spread of O. tsutsugamushi genotypes. CONCLUSIONS/SIGNIFICANCE: Fluctuation and diversification of vector species harboring O. tsutsugamushi in local endemic areas may facilitate genetic recombination among diverse genotypes. Therefore, careful monitoring of dominant vector species, as well as the prevalence of O. tsutsugamushi genotypes may be advisable to enable proper anticipation of epidemiological changes of scrub typhus.

Use of eschar swabbing for the molecular diagnosis and genotyping of Orientia tsutsugamushi causing scrub typhus in Quang Nam province, Vietnam.

BACKGROUND: Scrub typhus is a rickettsiosis which is caused by Orientia tsutsugamushi and occurs throughout the Asia-Pacific region. Molecular diagnosis of rickettsioses using eschar swabs has recently emerged, and may be very useful for the diagnosis of these diseases in tropical settings. METHODOLOGY/PRINCIPAL FINDINGS: Quantitative polymerase chain reaction (qPCR) was used to detect O. tsutsugamushi DNA in whole blood and eschar swab specimens of 67 patients who were clinically suspected of scrub typhus in Quang Nam province, Vietnam. Among the 20 patients for whom both eschar and whole blood were obtained, 17 (85%) of the eschar specimens and 5 (25%) of the whole blood specimens tested positive for O. tsutsugamushi. Genetic analysis of the 56-kDa TSA gene sequences demonstrated that the 14 sequences obtained in this study, including 12 eschar swabs and 2 whole blood specimens, were related to 4 groups: Karp, Kawasaki, Gilliam (JG-v and TG-v) and TA716. The majority (9/14; 64.4%) of contemporary O. tsutsugamushi genotypes in Quang Nam province were related to the Karp group. CONCLUSIONS: These results suggest that polyclonal antigen pools used for serological testing in the future should contain at least Karp, Kawasaki, Gilliam and TA716 antigens for Vietnamese patients, as well as patients who have traveled to Vietnam. qPCR after eschar swabbing should be considered for molecular diagnosis of scrub typhus in endemic patients as well as in travelers, since it is easy to perform and appears very useful for the rapid detection of Orientia tsutsugamushi in the early phase of infection.

Scrub Typhus Leading to Acute Encephalitis Syndrome, Assam, India.

To determine the contribution of Orientia tsutsugamushi, the agent of scrub typhus, as a cause of acute encephalitis syndrome (AES) in Assam, India, we conducted a retrospective study of hospital patients with symptoms of AES during 2013-2015. Our findings suggest that O. tsutsugamushi infection leads to AES and the resulting illness and death.