[Show abstract][Hide abstract] ABSTRACT: Spotted Fever Group (SFG) Rickettsiae can cause febrile diseases with or without rash in humans worldwide. In Germany only limited data are available about their medical significance. Serological screening tests for antibodies against rickettsiae usually only distinguish between SFG and Typhus Group (TG) Rickettsiae due to the strong cross reactivities within the groups. Seroprevalence rates in dogs, as possible sentinels for tick-borne diseases, could serve as an indicator for the distribution of different Rickettsia species.
In this study, a micro-immunofluorescence assay (micro-IFA) was established for detection and differentiation of antibodies against five Rickettsia species in dogs (R. helvetica, R. raoultii, R. slovaca, R. monacensis and R. felis). Dogs that never left Germany (n = 605) previously investigated with an SFG-ELISA were included in this study and screened at a 1:128 dilution. Endpoint titres of fifty randomly selected seropositive samples of each of the five investigated regions in Germany were determined in order to allow a differentiation of the causative Rickettsia species. Sensitivity and specificity of the micro-IFA were compared with ELISA results of the previous study.
A total of 93.9% of the dogs were positive for antibodies of the SFG Rickettsiae at the screening titer of 1:128. Differentiation of SFG Rickettsiae with the micro-IFA was possible in 70.4%, but in 29.6% of the cases the detected antibodies were not differentiable. Considering a clear differentiation by a twofold titre difference between observed reactions, the seroprevalence rates were 66.0% for R. helvetica, 2.8% for R. raoultii, 1.6% for R. slovaca, but no serological reaction could be clearly attributed to R. monacensis or R. felis. No statistically significant regional differences were found for R. helvetica, R. slovaca and R. raoultii comparing the five regions of Germany. Comparison of micro-IFA with ELISA revealed a sensitivity of 82.0% and a specificity of 83.8% for the Rickettsia SFG ELISA.
The micro-IFA is a useful serological tool to differentiate antibodies against different Rickettsia species in dogs. Seroprevalence rates in dogs correspond to the prevalence rates and distribution of Rickettsia-carrying tick species.
[Show abstract][Hide abstract] ABSTRACT: Rickettsioses are endemic in sub-Sahara Africa. Burden of disease, risk factors and transmission are hitherto sparsely described.
From the EMINI (Evaluating and Monitoring the Impact of New Interventions) population cohort, we randomly selected 1,228 persons above the age of 5 years from the nine participating communities in Mbeya region, Southwestern Tanzania, stratified by age, altitude of residence and ownership of domestic mammals, to conduct a cross-sectional seroprevalence study in. The aim was to estimate the seroprevalence of IgG antibodies against Spotted Fever Group (SFG) rickettsiae and to assess socioeconomic and environmental risk factors. Serology (indirect immunofluorescence) was performed at a dilution of 1:64.
SFG-seropositivity in the cohort was found to be 67.9% (range among nine sites: 42.8-91.4%). Multivariable analysis revealed an association with age (prevalence ratio, PR per 10 years: 1.08; 95% CI 1.06-1.10), warmer temperatures (PR per °C: 1.38; 1.11-1.71), male gender (PR 1.08; 1.00-1.16), and low population density (PR per 1.000 persons/km²increase 0.96; 0.94-0.99). At higher elevations, higher cattle density was associated with higher seroprevalence.
SFG rickettsial infection seems to be common in the more rural population of Mbeya Region. Spread seems to be further limited by temperature and higher elevation. Examination of the contribution of SFG to febrile illnesses seems warranted in a prospective study to estimate the disease burden in the population. This will also allow determination of the causative pathogens.
[Show abstract][Hide abstract] ABSTRACT: Kemerovo group viruses are tick-transmitted members of Orbivirus genus of the Reoviridae family that can cause infections of the central nervous system of humans. In this work, Kemerovo virus (KEMV) RNA was detected for the first time in Novosibirsk region of Western Siberia, Russia, in Ixodes pavlovskyi and Ixodes persulcatus ticks.
[Show abstract][Hide abstract] ABSTRACT: Background
To date, Alphavirus infections and their most prominent member, chikungunya fever, a viral disease which first became apparent in Tanzania in 1953, have been very little investigated in regions without epidemic occurrence. Few data exist on burden of disease and socio-economic and environmental covariates disposing to infection.
A cross-sectional seroprevalence study was undertaken in 1,215 persons from Mbeya region, South-Western Tanzania, to determine the seroprevalence of anti-Alphavirus IgG antibodies, and to investigate associated risk factors.
18% of 1,215 samples were positive for Alphavirus IgG. Seropositivity was associated with participant age, low to intermediate elevation, flat terrain and with IgG positivity for Rift Valley fever, Flaviviridae, and rickettsiae of the spotted fever group. When comparing the geographical distribution of Alphavirus seropositivity to that of Rift Valley fever, it was obvious that Alphaviruses had spread more widely throughout the study area, while Rift Valley fever was concentrated along the shore of Lake Malawi.
Alphavirus infections may contribute significantly to the febrile disease burden in the study area, and are associated with several arthropod-borne infections. Their spread seems only limited by factors affecting mosquitoes, and seems less restricted than that of Rift Valley fever.
[Show abstract][Hide abstract] ABSTRACT: The 16th Meeting of the International Scientific Working Group on Tick-Borne Encephalitis (ISW-TBE) - a group of neurologists, general practitioners, clinicians, travel physicians, virologists, paediatricians, and epidemiologists - was held under the title "Tick-borne Encephalitis - a Notifiable Disease, a Review after One Year". With the inclusion of TBE in the list of notifiable diseases in 2012, an important measure was established to continue improving the level of evidence on TBE in Europe to better help guide policies and methods to lower the burden of this disease. The conference agenda was divided into six parts concerning Travel Medicine, Epidemiology & Risk Areas, Poster Session with an Epidemiological Update in Europe, Interactive Debate, Cases, and Social Communication and Recommendations. Important topics regarding current epidemiological investigations, risk areas, mobility, cases, TBE in children, treatment options, vaccination rates, and latest news on vaccination were presented and extensively discussed.
[Show abstract][Hide abstract] ABSTRACT: Abstract In Croatia, several rodent- and vector-borne agents are endemic and of medical importance. In this study, we investigated hantaviruses and, for the first time, tick-borne encephalitis virus (TBEV) and Rickettsia spp. in small wild rodents from two different sites (mountainous and lowland region) in Croatia. In total, 194 transudate and tissue samples from 170 rodents (A. flavicollis, n=115; A. agrarius, n=2; Myodes glareolus, n=53) were tested for antibodies by indirect immunoflourescence assays (IIFT) and for nucleic acids by conventional (hantaviruses) and real-time RT-/PCRs (TBEV and Rickettsia spp.). A total of 25.5% (24/94) of the rodents from the mountainous area revealed specific antibodies against hantaviruses. In all, 21.3% (20/94) of the samples from the mountainous area and 29.0% (9/31) from the lowland area yielded positive results for either Puumala virus (PUUV) or Dobrava-Belgrade virus (DOBV) using a conventional RT-PCR. All processed samples (n=194) were negative for TBEV by IIFT or real-time RT-PCR. Serological evidence of rickettsial infection was detected in 4.3% (4/94) rodents from the mountainous region. Another 3.2% (3/94) rodents were positive for Rickettsia spp. by real-time PCR. None of the rodents (n=76) from the lowland area were positive for Rickettsia spp. by real-time PCR. Dual infection of PUUV and Rickettsia spp. was found in one M. glareolus from the mountainous area by RT-PCR and real-time PCR, respectively. To our knowledge, this is the first detection of Rickettsia spp. in small rodents from Croatia. Phylogenetic analyses of S- and M-segment sequences obtained from the two study sites revealed well-supported subgroups in Croatian PUUV and DOBV. Although somewhat limited, our data showed occurrence and prevalence of PUUV, DOBV, and rickettsiae in Croatia. Further studies are warranted to confirm these data and to determine the Rickettsia species present in rodents in these areas.
[Show abstract][Hide abstract] ABSTRACT: Tick-transmitted diseases are of great importance for the general health of the German population. Several viruses, such as tick-borne encephalitis virus (TBEV), Uukuniemi virus, Tribec virus, Eyach virus or bacteria, such as Borrelia, Rickettsiae, Francisella tularensis, Anaplasma phagocytophilum, Candidatus Neoehrlichia mikurensis (CNM) and Coxiella burnetii were detected in the most prominent tick in Germany, the hard tick Ixodes ricinus. While infections, such as TBE and Lyme disease are well known, other infections are hardly known even among experts. Although there have been a few descriptions of isolated cases in Germany, a systematic investigation regarding the distribution and the pathogenic potential of these pathogens is still lacking. In particular elderly people and people with underlying diseases seem to be mostly affected. The importance of new infectious disease agents, such as Candidatus Neoehrlichia mikurensis but also of long known pathogens, such as Rickettsiae still remains unclear, while some of them could be detected in 20 % of investigated ticks. Whether climate change contributes to the further distribution of these infectious agents remains unclear and requires further investigation. The increasing initiatives to create natural environments and the trend towards spending more time in nature for recreational activities will increase the danger of coming into contact with ticks and the respective infectious agents. Considering these circumstances an increase of diseases caused by these pathogens is to be expected.
[Show abstract][Hide abstract] ABSTRACT: Viperin is an interferon-induced protein with a broad antiviral activity. This evolutionary conserved protein contains a radical S-adenosyl-L-methionine (SAM) domain which has been shown in vitro to hold a [4Fe-4S] cluster. We identified tick-borne encephalitis virus (TBEV) as a novel target for which human viperin inhibits production of the viral genome RNA. Wt viperin was found to require ER localization for full antiviral activity and to interact with the cytosolic Fe/S protein assembly factor CIAO1. Radiolabelling in vivo revealed incorporation of (55) Fe, indicative for the presence of an Fe-S cluster. Mutation of the cysteine residues ligating the Fe-S cluster in the central radical SAM domain entirely abolished both antiviral activity and incorporation of (55) Fe. Mutants lacking the extreme C-terminal W361 did not interact with CIAO1, were not matured, and were antivirally inactive. Moreover, intracellular removal of SAM by ectopic expression of the bacteriophage T3 SAMase abolished antiviral activity. Collectively, our data suggest that viperin requires CIAO1 for [4Fe-4S] cluster assembly, and acts through an enzymatic, Fe-S cluster and SAM-dependent mechanism to inhibit viral RNA synthesis.
[Show abstract][Hide abstract] ABSTRACT: The full genome sequences of three tick-borne encephalitis virus strains, two isolated from Ixodes ricinus ticks and one from the brain of a bank vole, Myodes glareolus, originating from the Slovak Republic were determined. Nucleotide sequences were found to be very similar (>99.5 % nt-identity) with only one distinct amino acid (aa) difference to each other. They all shared 30 aa-changes when compared to type strain Neudoerfl, isolated in neighboring Austria. An internal poly(A)-stretch similar to that of strain Neudoerfl was found only in TBEV strain 114 from a tick. Despite this heterogeneity in the 3'-NCR, the high level of sequence identity was striking, indicating a low rate of nucleotide substitutions of TBEV strains in Slovakia and no obvious relation to the host species.
[Show abstract][Hide abstract] ABSTRACT: A reliable and complete inactivation is an indispensable premise for any concentration of rickettsiae or for the development of diagnostic strategies based on their antigens. This study deals with the testing of methods to inactivate rickettsiae. Rickettsia honei was used as a model organism. The inactivating potency of formalin, Qiagen® antiviral lysozyme (AVL) buffer, heating to 56 °C, and β-propiolactone was analyzed in cell culture. The inactivation limits for rickettsiae were 0.1% formalin about 10 min, Qiagen AVL buffer about 5 min, 56 °C about 5 min, 0.125% β-propiolactone about 1 h, and 0.0125% β-propiolactone overnight. The interpretation was limited by cytotoxic effects of the inactivation procedures and by the culturally achievable rickettsial density in the cell culture supernatants that were used for the inactivation experiments. Reliable modes of inactivation were identified, allowing for the secure handling of rickettsial antigens for diagnostic purposes.
European Journal of Microbiology and Immunology 09/2013; 3(3):188-193. DOI:10.1556/EuJMI.3.2013.3.6
[Show abstract][Hide abstract] ABSTRACT: Rickettsiae tend to have a rapid decrease of viability outside living cells. Therefore, the transport of samples containing viable rickettsiae for culturing in cell culture for diagnostic purposes is challenging. The viability of rickettsiae in different transport media (commercially available transport medium COPAN "UTM-RT transport medium for viruses, chlamydia, mycoplasma, and ureaplasma," minimal essential medium (MEM) with and without 10% foetal calf serum) at various time points at 4 °C and at ambient temperature (22 °C) was compared. Rickettsia honei was used as model organism. After 2 weeks of storage at room temperature, no viable rickettsiae were detectable any more while storage at 4 °C kept rickettsiae viable for up to 4 weeks. The commercially available COPAN medium showed similarly good or slightly better stabilizing effects on rickettsiae compared with MEM + 10% foetal calf serum, pure MEM demonstrated the poorest results. It is important to transport and store media with potentially rickettsiae-containing samples at 4 °C to prevent inactivation. MEM + 10% foetal calf serum can be used if no commercial medium is available with similarly good results.
European Journal of Microbiology and Immunology 09/2013; 3(3):194-197. DOI:10.1556/EuJMI.3.2013.3.7
[Show abstract][Hide abstract] ABSTRACT: Rickettsiae are able to spread within infected cell mono-layers by modifying intra-cellular actin formations. The study analyzes whether a visualization of actin modifications in addition to specific immuno-fluorescence staining of rickettsiae might facilitate the proof of rickettsial growth in cell culture. Cell mono-layers of Vero E6 und BGM cells were infected with Rickettsia honei. Intra-cellular actin was fluorescence stained with TRITC-(tetra-methyl-5,6-isothiocyanate)-labeled phalloidin in addition to specific immuno-fluorescence staining of rickettsiae with FITC-(fluorescein-isothiocyanate)-labeled antibodies. DNA of bacteria and cells was counter-stained with DAPI (4´,6-diamino-2-phenyl-indole). Cell cultures infected with Vaccinia virus were used as positive controls, cell cultures infected with Coxiella burnetii as negative controls. High concentrations of R. honei are necessary to demonstrate characteristic modifications of the intra-cellular actin. This effect is more pronounced in Vero E6 cells than in BGM cells. Actin staining with phalloidin is not suited for an early proof of rickettsial growth in cell culture but may confirm unclear findings in specific immuno-fluorescence staining in case of sufficient bacterial density.
European Journal of Microbiology and Immunology 09/2013; 3(3):198-203. DOI:10.1556/EuJMI.3.2013.3.8
[Show abstract][Hide abstract] ABSTRACT: Arthropod-borne viral encephalitis of diverse origins shows similar clinical symptoms, histopathology and magnetic resonance imaging, indicating that the patho mechanisms may be similar. There is no specific therapy to date. However, vaccination remains the best prophylaxis against a selected few. Regardless of these shortcomings, there are an increasing number of case reports that successfully treat arboviral encephalitis with high doses of intravenous immunoglobulins.
To our knowledge, high dose intravenous immunoglobulin has not been tested systematically for treating severe cases of tick-borne encephalitis. Antibody-dependent enhancement has been suspected, but not proven, in several juvenile cases of tick-borne encephalitis. Although antibody-dependent enhancement during secondary infection with dengue virus has been documented, no adverse effects were noticed in a controlled study of high dose intravenous immunoglobulin therapy for dengue-associated thrombocytopenia. The inflammation-dampening therapeutic effects of generic high dose intravenous immunoglobulins may override the antibody-dependent enhancement effects that are potentially induced by cross-reactive antibodies or by virus-specific antibodies at sub-neutralizing levels.
Analogous to the increasing number of case reports on the successful treatment of other arboviral encephalitides with high dose intravenous immunoglobulins, we postulate whether it may be possible to also treat severe cases of tick-borne encephalitis with high dose intravenous immunoglobulins as early in the course of the disease as possible.
[Show abstract][Hide abstract] ABSTRACT: Tick-borne encephalitis virus (TBEV) strain A104 was isolated from the brain of a yellow-necked mouse in Austria in 1990. The complete genome sequence was 11,097 nucleotides long. Comparison with TBEV prototype strain Neudoerfl showed 32 amino acid exchanges and the absence of an internal poly(A) stretch within the 3' noncoding region.
[Show abstract][Hide abstract] ABSTRACT: In order to obtain a better understanding of tick-borne encephalitis virus (TBEV) strain movements in central Europe the E gene sequences of 102 TBEV strains collected from 1953-2011 at 38 sites in the Czech Republic, Slovakia, Austria and Germany were determined. Bayesian analysis suggests a 350-year history of evolution and spread in Central Europe of two main lineages A and B. In contrast to the east to west spread at the Eurasian continent level local Central European spreading patterns suggest historic west to east spread followed by more recent east to west spread. The phylogenetic and network analyses indicate TBEV ingressions from the Czech Republic and Slovakia into Germany via landscape features (Danube river system), biogenic factors (birds, red deer) and anthropogenic factors. The identification of endemic foci showing local genetic diversity is of paramount importance to the field as these will be a prerequisite for in depth analysis of focal TBEV maintenance and long distance TBEV spread.
Journal of General Virology 06/2013; 94(Pt_9). DOI:10.1099/vir.0.054478-0 · 3.18 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Rickettsioses caused by typhus group rickettsiae have been reported in various African regions. We conducted a cross-sectional survey of 1,227 participants from 9 different sites in the Mbeya region, Tanzania; overall seroprevalence of typhus group rickettsiae was 9.3%. Risk factors identified in multivariable analysis included low vegetation density and highway proximity.
[Show abstract][Hide abstract] ABSTRACT: The role of wild mammals in the dissemination and maintenance of Rickettsia in nature is still under investigation. European hedgehogs (Erinaceus europaeus) are often heavily infested by tick and flea species that are known to harbor and transmit different Rickettsia spp. We investigated ixodid ticks sampled from European hedgehogs for the presence of Rickettsia. A total of 471 Ixodes ricinus and 755 I. hexagonus were collected from 26 German and 7 British European hedgehogs. These were tested by a genus-specific real-time PCR assay targeting the citrate synthase gene (gltA). The rickettsia minimum infection rate was 11.7% with an increase detected with each parasitic tick stage. No significant difference in Rickettsia prevalence in the 2 Ixodes species was detected. Using sequencing of partial ompB, Rickettsia helvetica was the only species identified. More than half of the hedgehogs carried Rickettsia-positive ticks. In addition, tissue samples from 2/5 hedgehogs (where tissue DNA was available) were PCR-positive. These results show that European hedgehogs are exposed to R. helvetica via infected ticks and might be involved in the natural transmission cycle of this Rickettsia species.