Equine sentinels and one health: a comprehensive serological survey of Crimean-Congo hemorrhagic fever virus in southeastern and Central Europe.

To the Editor: Crimean Congo hemorrhagic fever, a tick-borne illness caused by Crimean Congo hemorrhagic fever virus (CCHFV), is endemic to Africa, the Balkans, the Middle East, and parts of Asia. The hard ticks (Ixodidae), especially those of the genus Hyalomma, serve as reservoirs and vectors for CCHFV, and a variety of animals, such as cattle, sheep, and camels, are considered amplifying hosts for the virus. Although CCHFV may cause little or no disease in zoonotic hosts, the virus can cause severe disease in humans who may be exposed by tick bites or by contact with blood or tissues from infected patients or animals (1). Surveillance for CCHFV in animal and vector populations provides an opportunity to monitor a disease of potentially severe impact. In North Africa and the Middle East, trade in live animals, meat, and meat products poses noticeable risk to human and animal health (2,3) and can serve as a mobile pool of diseases with potentially large economic and health effects. Animals often originate in distant areas of a country or its neighbors, where they may be exposed to zoonotic pathogens not endemic to their final location, and may collect vectors that carry additional pathogens. As part of a broader study examining occupational risk of exposure to vector-borne and zoonotic pathogens in a high-risk abattoir worker population, we collected ectoparasites from freshly-slaughtered livestock and examined them for CCHFV. This study and was conducted in compliance with the Animal Welfare Act and in accordance with the principles set forth in the Guide for the Care and Use of Laboratory Animals (4). Sample collection took place over a 2-week period in July 2009 at the Muneeb abattoir in the Giza Governorate of Egypt. Ectoparasites were removed from 43 freshly slaughtered animals by using blunt forceps and were placed in glass vials. In total, 342 ectoparasites were collected: 70 (20.5%) from 14 cattle, 52 (15.2%) from 17 buffalo, 6 (1.8%) from 2 sheep, and 214 (62.6%) from 10 camels. Cattle, buffalo, and sheep originated in Egypt; camels were imported from Sudan and Somalia. Ectoparasites were transported to US Naval Medical Research Unit No. 3 in Cairo, Egypt, for taxonomic identification and pathogen detection. Ninety-seven percent (334) of the ectoparasites were ticks from the family Ixodidae. The genus Hyalomma accounted for 254 (76.0%) of these ticks, including nearly all of those collected from sheep (100%) and camels (99.5%) but only 60.4% and 9.1% of those collected from buffalo and cattle, respectively. The remaining ticks of the family Ixodidae belonged to genus Boophilus. After identification, ticks were grouped into pools by species, sex, and animal source. RNA was then extracted by using the QIAamp Viral RNA Kit (QIAGEN, Valencia, CA, USA). CCHFV small fragment RNA was detected with SuperScript III Platinum SYBR Green One-Step qRT-PCR (Invitrogen, Carlsbad, CA, USA) and published primers as described (5). Of 138 pools tested (258 ticks of the genera Hyalomma and Boophilus), 6 pools were positive for CCHFV. These 6 pools contained ticks collected from 1 camel imported from Somalia and 4 from Sudan. One positive pool comprised female H. excavatum (Koch, 1844), and 5 comprised female H. dromedarii (Koch, 1844), both proven vectors of CCHFV. Sequence analysis showed that each CCHFV-positive pool contained an identical yet previously unrecorded 229-bp RNA fragment (GenBank accession no. {type:entrez-nucleotide,attrs:{text:JF706233,term_id:371444381}}JF706233; Figure). This fragment had 92% identity with many published CCHFV isolates (BLAST analysis [www.ncbi.nlm.nih.gov/BLAST], GenBank nonredundant database); however, most base changes were synonymous substitutions. Translated protein queries (tblastx) identified an amino acid (serine at codon 26) in this new variant that differed from all but 1 ({type:entrez-nucleotide,attrs:{text:AJ010648.1,term_id:10638944}}AJ010648.1) of the 100 most similar published sequences. The functional consequence of a S26N substitution is unknown but may be minimal because of the similar physicochemical properties of asparagine and serine. Figure Phylogenetic relationship of 110 Crimean-Congo hemorrhagic fever virus sequences obtained from this study (6 tick pools), the positive control, and 99 published isolates from GenBank. Sequences were aligned by using ClustalX (www.clustal.org), and a phylogenetic ... Despite the low number of camels sampled in this study, 5 of the 10 camels were found to harbor ticks carrying RNA from an undocumented variant of CCHF. Although none of the domestic animals harbored infected ticks, it is not possible to conclude if these data reflect importation of CCHFV or infection acquired within Egypt because details about conditions under which animals were kept before slaughter are unavailable. Previous serologic studies in Egypt have shown antibodies to CCHFV were prevalent among imported camels at a quarantine station in Aswan governorate (6) and among domestic cattle, buffalo, sheep, and goats in Sharkia Governorate (7). We plan to further investigate the presence of CCHFV within camel and ectoparasite populations in Egypt by expanding protocol activities to a camel market in the same area as the abattoir.

To the Editor: Crimean-Congo hemorrhagic fever (CCHF) virus, genus Nairovirus, family Bunyviridae, is transmitted to mammals and birds by ticks. Hyalomma ticks, the primary vectors in CCHF transmission, are widespread throughout Europe, Asia, the Middle East, and Africa; evidence of CCHF virus has been found in all these regions. CCHF in humans is an acute viral disease that is transmitted by the bite of infected ticks, direct contact with blood or infected tissues from viremic animals, and direct contact with the blood or secretions of an infected person (1). On January 26, 2003, a 22-year-old shepherd was treated at a health post in the Popenguine District, 60 km south of Dakar, Senegal; he reported fever, epistaxis, arthralgia, myalgia of the lower limbs, and dark urine for the past 2 days. Without biologic confirmation of the infection, he was treated for malaria with two intravenous injections of quinine, followed by oral administration of chloroquine. On January 31, the patient had a temperature of 39°C, conjunctival jaundice, bleeding gums, and was vomiting blood. He was seen again at the health post and was given antimicrobial drugs, intravenous quinine, and vitamin K; the next day, the bleeding stopped and the fever subsided. A serum sample was sent to the World Health Organization Collaborative Centre for Arboviruses and Viral Hemorrhagic Fevers at the Institut Pasteur, Dakar. Tests for anti-CCHF specific immunoglobulin (Ig) M antibody by enzyme-linked immunosorbent assay (ELISA) were positive, and CCHF virus by isolation on cell cultures (AP61 and Vero cells) and reverse transcriptase-polymerase chain reaction (RT-PCR) were negative. From January 31 to February 10, the IgM titer increased from 1/3,200 to >1/12,800 and IgG titer increased from 1/200 to 1/6,400. Examination of the patient on February 10 showed he had recovered without sequelae, and no trace of tick bites was found. The patient stated that he had not traveled, noticed any tick bites, slaughtered any animals, or been in contact with people with fever for several weeks before his illness. He lived in close proximity to goats and cattle, but no blood samples were taken from these animals. Although no ticks were found on nearby goats, 10 Amblyomma and Hyalomma ticks were collected from three cattle. Ticks were negative for CCHF virus isolation on suckling mice and RT-PCR amplification. No other case of fever accompanied by hemorrhage was reported in the area, and none of the patient's 14 close contacts became ill. Of the four close contacts from whom blood samples were taken, analyses for IgM and IgG antibodies against CCHF virus were negative by ELISA. While no clinical case of CCHF has ever been reported in Senegal, studies dating from 1969 indicate that CCHF virus had been found in various locations in the country (2,3). In the village of Bandia, in the same district where the reported case was observed, a study conducted from 1986 to 1988 showed a prevalence of anti-CCHF IgG of 3.2% in the human population (4). Another study, conducted in the same area from 1989 to 1992, showed seroconversions for several ruminants and isolated the virus from ticks (5). During CCHF outbreaks, an average of 30% of people who had the disease died (case-fatality ratio). It is often discovered during nosocomial outbreaks, as was the case in Mauritania, a country on Senegal's northern border, in 2003 (P. Nabeth, unpub. data). To prevent outbreaks of CCHF, public awareness campaigns aimed at the populations most at risk—livestock farmers, butchers, and health personnel—must be conducted, and the epidemiologic alert systems must be strengthened. In addition, conditions that enhance maintenance of the virus in nature and its transmission to humans must be better understood so adequate control measures can be developed.

Unique strain of Crimean-Congo hemorrhagic fever virus, Mali.

Crimean-Congo hemorrhagic fever (CCHF) is a viral disease transmitted to humans mainly by bite of Ixodid ticks, mainly those of the Hyalomma genus. CCHFV belongs to the genus Nairovirus in the family Bunyaviridae. CCHF virus is a segmented, single stranded, negative sense and RNA viruses. The onset of the disease is very sudden, with symptoms such as fever, rigors, intense headache, chills, and backache or leg pains, myalgia, nausea, and vomiting. CCHF originally identified in the former Soviet Union and the Congo, has rapidly spread across large sections of Europe, Asia, and Africa, and has been reported in more than 30 countries. The climatic changes may affect the life cycle of ticks and the routes of migratory birds, leading to tick abundance and virus distribution in CCHF-free areas. Extended use of land for agriculture and farming and changes in hunting activities play also a role in CCHF incidence, while livestock trade and movement may influence host-tick-virus dynamics resulting in transfer of CCHFV-infected ticks in non-endemic areas. Recent years, the epidemiology of CCHF is changing in Balkans and Turkey. Balkan Peninsula is a known endemic CCHF area, and sporadic cases and even outbreaks are being reported every year. The annual number of human CCHF cases is increasing in Balkans and Turkey. While Bulgaria, Kosovo and Albania were ZET Krm-Kongo Kanamal Atei (KKKA), zellikle Ixodid cinsi kene sr (esas olarak Hyalomma cinsi)

Tick-borne diseases pose a serious threat to human health in South-Eastern Europe, including Kosovo. While Crimean–Congo hemorrhagic fever (CCHF) is a well-known emerging infection in this area, there are no accurate data on Lyme borreliosis and tick-borne encephalitis (TBE). Therefore, we sampled and tested 795 ticks. Ixodes ricinus (n = 218), Dermacentor marginatus (n = 98), and Haemaphysalis spp. (n = 24) were collected from the environment by flagging (all from Kosovo), while Hyalomma marginatum (n = 199 from Kosovo, all from Kosovo) and Rhipicephalus bursa (n = 130, 126 from Albania) could be collected only by removal from animal pasture and domestic ruminants. Ticks were collected in the years 2014/2015 and tested for viral RNA of CCHF and TBE viruses, as well as for DNA of Borrelia burgdorferi sensu lato by real-time PCR. In Kosovo, nine ticks were positive for RNA of Crimean–Congo hemorrhagic fever virus and seven for DNA of B. burgdorferi s. l. None of the ticks tested positive for TBEV. CCHF virus was detected in one H. marginatum male specimen collected while feeding on grazing cattle from the Prizren region and in eight R. bursa specimens (five females and three males collected while feeding on grazing sheep and cattle) from the Prishtina region (Kosovo). B. burgdorferi s. l. was detected in seven questing ticks (four male and one female D. marginatus, two I. ricinus one female and one male) from the Mitrovica region (Kosovo). Our study confirmed that CCHF virus is circulating in Kosovo mainly in H. marginatum and R. bursa in the central areas of the country. B. burgdorferi s. l. was found in its major European host tick, I. ricinus, but also in D. marginatus, in the north of the Kosovo. In order to prevent the spread of these diseases and better control of the tick-borne infections, an improved vector surveillance and testing of ticks for the presence of pathogens needs to be established.

BackgroundCrimean-Congo hemorrhagic fever (CCHF) is an emerging infectious disease caused by a Nairovirus. CCHF is a tick-borne disease that is predominantly associated with Hyalomma ticks and have a widespread distribution in Africa, Asia and Europe. CCHF usually presents as a subclinical disease, but in some cases, it may present as a hemorrhagic fever with a high mortality rate. This systematic review of the literature was performed to identify the available evidence on the prevalence of CCHF in the European Region of the World Health Organization, based on seroprevalence (IgG antibodies).MethodologyA systematic review was performed following the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) statement protocol. PubMed, Embase, and the Web of Science were used for the search (up to January 31, 2019), combining the following MeSH terms: [“Crimean-Congo haemorrhagic fever” OR “Crimean-Congo hemorrhagic fever virus” OR “Congo-Crimea” OR “Crimea-Congo”] AND [“Europe”] AND [“epidemiology” OR “seroprevalence”]. The abstracts were screened. Subsequently, full-text articles were selected and reviewed based on the PICOS (Population-Intervention-Comparison-Outcomes-Study type) criteria by two independent reviewers for inclusion in the final analysis. The data were qualitatively synthesized without quantitative pooling due to the heterogeneity in the study populations and methodologies.Principal findingsThirty articles (9 from western Europe, 18 from central Europe and 3 from eastern Europe) were included in the analysis. All articles were cross-sectional studies (descriptive studies).ConclusionsThe highest seroprevalence of CCHF is found in central and eastern European countries. Southern and western Europe countries, such as Greece and Spain, have low levels of endemicity, but the spread of the infection, which is associated with climate change, is a possibility that we should keep in mind. Further studies, especially larger seroprevalence studies in humans and animals, are needed to establish the current status of the CCHF epidemiology and to generate standardized guidelines for action in the region.

Background: Crimean-Congo hemorrhagic fever (CCHF) is a tick-borne viral disease that is endemic in Africa, southeastern Europe, and Asia. Ticks are both the reservoir and the vector of the CCHF virus (CCHFV). Determining the virus infection rate of the tick population in different geographical regions is necessary todesign public health policies to prevent CCHF outbreaks. Objectives: To determine the prevalence, seasonal activity, and the frequency of CCHFV infection of the tick population in Hamadan province of west Iran. Methods: This cross-sectional study was conducted in 3 counties of Hamadan Province from June 2013 to May 2014. The study areas included both lowland (plains) and highland (mountains) and covered 5% of the villages where 10 herds per village of sheep and goats were randomly selected for hard tick collection. Results: We examined 983 sheep and goats, and 881 ticks were collected and identified before being preserved for molecular tests. The collected ticks belonged to 3 genera including, Rhipicephalus (95.6%, n = 842), Hyalomma (4.1%, n = 36) and Haemaphysalis (0.4%, n = 4). After species identification, 100 randomly selected ticks were analyzed using reverse transcriptase-polymerase chain reaction (RT-PCR) to detect viral infection. CCHFV infection was observed in 7 collected ticks, of which 4 belonged to R. sanguineus, 2 belonged to R. bursa, and one Hy. asiaticum. Conclusions: Hyalomma and Rhipicephalus ticks are the main vectors of CCHFV in Hamadan province where CCHF is focal and endemic. Keywords: Crimean-Congo hemorrhagic fever, Hamadan, Iran, molecular epidemiology, tick-borne diseases, zoonotic viral disease

Crimean-Congo haemorrhagic fever (CCHF) is a zoonotic disease caused by the Crimean-Congo hemorrhagic fever virus (CCHFV). Ticks of the genus Hyalomma are the main vectors and represent a reservoir for the virus. CCHF is maintained in nature in an endemic vertebrate-tick-vertebrate cycle. The disease is prevalent in wide geographical areas including Asia, Africa, South-Eastern Europe and the Middle East. It is of great importance for the public health given its occasionally high case/fatality ratio of CCHFV in humans. Climate change and the detection of possible CCHFV vectors in Central Europe suggest that the establishment of the transmission in Central Europe may be possible in future. We have developed a compartment-based nonlinear Ordinary Differential Equation (ODE) system to model the disease transmission cycle including blood sucking ticks, livestock and human. Sensitivity analysis of the basic reproduction number R0 shows that decreasing the tick survival time is an efficient method to control the disease. The model supports us in understanding the influence of different model parameters on the spread of CCHFV. Tick-to-tick transmission through co-feeding and the CCHFV circulation through transstadial and transovarial transmission are important factors to sustain the disease cycle. The proposed model dynamics are calibrated through an empirical multi-country analysis and multidimensional plot reveals that the disease-parameter sets of different countries burdened with CCHF are different. This information may help decision makers to select efficient control strategies.

Crimean-Congo hemorrhagic fever virus (CCHFV) is a widespread tick-borne zoonotic virus that causes Crimean-Congo hemorrhagic fever (CCHF). CCHF is asymptomatic in infected animals but can develop into severe illness in humans, with high case-fatality rates. Due to complex environmental and socio-economic factors, the distribution of CCHFV vectors is changing, leading to disease occurrence in previously unaffected countries. Neither an effective treatment nor a vaccine has been developed against CCHFV; thus, surveillance programs are essential to limit and control the spread of the virus. Furthermore, the WHO highlighted the need of assays that can cover a range of CCHFV antigenic targets, DIVA (differentiating infected from vaccinated animals) assays, or assays for future vaccine evaluation. Here, we developed a multiplex assay, based on a suspension microarray, able to detect specific antibodies in ruminants to three recombinantly produced CCHFV proteins: the nucleocapsid (N) protein and two glycoproteins, GN ectodomain (GNe), and GP38. This triplex assay was used to assess the antibody response in naturally infected animals. Out of the 29 positive field sera to the N protein, 40% showed antibodies against GNe or GP38, with 11 out of these 12 samples being positive to both glycoproteins. To determine the diagnostic specificity of the test, a total of 147 sera from Spanish farms free of CCHFV were included in the study. This multiplex assay could be useful to detect antibodies to different proteins of CCHFV as vaccine target candidates and to study the immune response to CCHFV in infected animals and for surveillance programs to prevent the further spread of the virus. IMPORTANCE Crimean-Congo hemorrhagic fever virus (CCHFV) causes Crimean-Congo hemorrhagic fever, which is one of the most important tick-borne viral diseases of humans and has recently been found in previously unaffected countries such as Spain. The disease is asymptomatic in infected animals but can develop into severe illness in humans. As neither an effective treatment nor a vaccine has been developed against CCHFV, surveillance programs are essential to limit and control the spread of the virus. In this study, a multiplex assay detecting antibodies against different CCHFV antigens in a single sample and independent of the ruminant species has been developed. This assay could be very useful in surveillance studies, to control the spread of CCHFV and prevent future outbreaks, and to better understand the immune response induced by CCHFV.

Sir, Crimean-Congo haemorrhagic fever (CCHF) is a tick-borne viral disease with average mortality rate of 30-50 per cent1. In India, presence of CCHF was first time confirmed in Gujarat State during a nosocomial outbreak in 20112,3. Since then, numerous outbreaks and sporadic cases of this disease have been reported from different districts of Gujarat State4,5,6. Studies conducted at the National Institute of Virology (NIV), Pune, had reported the presence of anti-CCHF IgG antibodies in domestic animals from Sirohi district, Rajasthan State. However, in the last four years none of the referred human samples were found positive3,4,5,6. On March 18, 2014, blood sample of a suspected CCHF case was referred to NIV, Pune, for aetiology confirmation. This suspected CCHF case was a 45 yr old male, shepherd by profession, residing at Veravilapur village, Sirohi district, Rajasthan. He was presented on March 14, 2014, with complains of abdominal discomfort since last six days and vomiting since one day along with history of intermittent fever since 15 days. He had moderate fever without chills and was associated with arthralgia, generalized body ache, constipation, decreased urine output, bleeding from nose, haematuria and bleeding per rectum. He was a known case of HBsAg reactivity, and was transferred from Rajasthan State to Civil hospital, Ahmadabad, Gujarat. On admission, patient had thrombocytopenia (20,000/μl). Serum creatinine was 1.2 mg/dl, prothrombin time 15.3 sec, International Normalized Ratio (INR) 1.15, and activated partial thromboplastin time (APTT) was 50.0 sec (Table). There was impairment of liver function test in the form of markedly elevated liver enzymes of serum glutamic-pyruvic transaminase (SGPT: 2620 U/l) with normal serum total bilirubin (0.40 mg/dl) and normal renal function. Parameters of complete blood count were in the normal range (Table). Table Day-wise clinical laboratory findings of Crimean-Congo haemorrhagic fever (CCHF) case in 2014 The differential diagnosis of CCHF at the pre-haemorrhagic stage is more difficult. As the disease progresses, clinical features become clearer and diagnosis becomes easier. The sample was differentially tested for some aetiological agents (hepatitis viruses, Leptospira and dengue viruses) which are endemic in the region and mimic the clinical illness of CCHF. Apart from HBsAg, patient was negative for viral markers (i.e. anti-HEV IgM, anti-HAV IgM, anti-HCV IgM). Anti-HIV antibodies, anti-Leptospira IgM and IgG antibodies, dengue IgM antibody, blood and urine culture were negative. The patient did not have any recent travel history to Gujarat State; but had close contact with livestock. Whole blood of the patient was collected on March 15, 2014 (2nd day of admission) and March 19, 2014 (6th day of admission). On day 19, urine sample was also collected. The patient's serum and urine samples were processed for CCHF virus specific real-time RT-PCR5,7. Anti-CCHF IgM antibodies were tested in serum samples using commercial CCHF IgM ELISA Kit (Vector-Best, Novosibirsk, Russia). Real-time RT-PCR results were found positive for both serum samples collected on day 15th [threshold cycle (Ct) =27] and on 19th day (Ct=34)]. On 19th day urine sample showed Ct=38. Both serum samples (of 15th and 19th days) were positive for IgM antibodies against CCHF virus. Real-time reverse-transcription RT-PCR data showed high CCHF viral copy number. On 19th post illness day, the urine sample also showed low level of CCHF viral RNA. As soon as the sample was laboratory confirmed as CCHF case, the patient was put in strict isolation. For treatment, oral ribavirin was administered on day 3 after admission on clinical suspicion at the dosage recommended by the World Health Organization8,9,10, along with the supportive and replacement therapy with blood products. The patient responded to the treatment and recovered completely, and was discharged on request on March 27, 2014 (14th day of admission). Livestock trade and movements of domestic animals infested with infected ticks might be the reason in distribution of infected ticks to newer areas, and eventually spread of CCHFV. As CCHF mimics a wide range of common illnesses caused by different aetiological agents which are endemic in India, differential diagnosis should be done based on clinical biochemical, haematological, bacteriological and virological findings. These include Kyasanur forest disease, hepatitis, Neisseria meningitidis infection, leptospirosis, borreliosis, typhoid, rickettsiosis, dengue and malaria. However, malaria diagnosis can be excluded in cases of suspected viral haemorrhagic fever (VHF)11. In conclusion, there is a need to initiate active serosurvey of CCHF among human population, and domestic animals in Rajasthan. This will be helpful in understanding the prevalence of this disease in Rajasthan State which eventually will alert the State health authorities.

Aspects of the spatial and temporal patterns of transmission of Crimean-Congo hemorrhagic fever (CCHF) virus were studied in Senegal, West Africa. A country-wide serological survey of domestic animals indicated that transmission was most intense in the northern dry sahelian zone and least in the southern, more humid guinean zone. Human IgG prevalence, ranging from nearly 20% to < 1% among 8 sites throughout the region, also was greatest in the north. A fatal human case of CCHF from Rosso, Mauritania in 1988 was studied and an accompanying serosurvey of human contacts and domestic animals indicated epidemic transmission during that period. Systematic samples of adult ixodid ticks on domestic animals allowed us to analyze the distribution and relative abundance of potential CCHF virus vectors, demonstrating that Hyalomma spp. predominated in those biotopes where transmission was most intense. A prospective study of CCHF virus infection and tick infestation in sheep exposed a period of epizootic transmission in 1988 that corresponded temporally with increased abundance of adult H. truncatum and H. impeltatum. Four strains of CCHF virus were isolated from pools of these ticks and of Rhipicephalus guilhoni. Our results suggest that CCHF virus is focally endemic throughout the region, although highly variable in time and space, and that the relative abundance of Hyalomma ticks may be the primary determinant of epidemic transmission.

To the Editor: Crimean-Congo hemorrhagic fever virus (CCHFV) (Nairovirus, Bunyaviridae), the causative agent of Crimean-Congo hemorrhagic fever, has been detected in sub-Saharan Africa, southeastern Europe, the Middle East, and central Asia. The virus has been detected in >31 species of ticks and is transmitted to humans by bite of infected ticks (mainly of the genus Hyalomma) or by contact with body fluids or tissue of viremic patients or livestock. The disease is characterized by fever, myalgia, headache, vomiting, and sometimes hemorrhage; reported mortality rate is 10%–50% (1). CCHFV strains currently constitute 7 evolutionary lineages, 1 of which (Europe 2) contains the prototype strain AP92, which was isolated in 1975 from Rhipicephalus bursa ticks collected from goats in Greece (2). This strain seems to have low or no pathogenicity for humans; only a few mild cases have been reported (3). This observation is supported by the relatively high (14.4%) seroprevalence but no clinical cases in humans in northwestern Greece (4). The documented tick carriers of this strain are R. bursa and Hyalomma marginatum (5). Hyalomma aegyptium ticks are highly host specific; adults feed almost entirely on tortoises of the genus Testudo (6) and occassionally on hedgehogs and hares. Unlike adult ticks, the larvae and nymphs are less host specific and feed on a wide spectrum of hosts (e.g., other reptiles, birds, and mammals [including humans]) (7). This trait elevates the epidemiologic role of the tick as a possible bridge vector connecting wildlife, domestic animals, and humans. To determine the biological and epidemiological role of H. aegyptium ticks, during 2009–2010, we collected 56 adult ticks from 12 Testudo graeca tortoises at a locality near the city of Aflou in Laghouat Province, Algeria. We tested the ticks for probable CCHFV infection by using nested reverse transcription PCR (8), which amplifies a partial fragment of the CCHFV small RNA segment. We slightly modified the assay: reverse transcription time was 60 minutes and annealing temperature was 52°C (9). In total, 16 (28.6%) ticks were positive for CCHFV. The PCR products of 15 (26.8%) positive samples were sequenced. BLAST (http://blast.ncbi.nlm.nih.gov//Blast.cgi) analysis identified all 15 sequences as CCHFV with 98%–100% identity to the AP92 strain (GenBank accession no. {type:entrez-nucleotide,attrs:{text:DQ211638,term_id:78191750,term_text:DQ211638}}DQ211638). Two variants of AP92 were detected and differed by 0.6%. A phylogenetic tree was constructed by Bayesian inference, using MrBayes version 3.1.2 (http://mrbayes.sourceforge.net/index/php) under a general time-reversible plus gamma distribution plus invariable site model with 107 generations setup (Figure). Sequences are available in GenBank (accession nos. KT99097 and KT99098). Figure Phylogenetic analysis of Crimean-Congo hemorrhagic fever virus small RNA segment sequences, performed by using Bayesian inference in MrBayes version 3.1.2. (http://mrbayes.csit.fsu.edu/) under a general time-reversible plus gamma distribution plus invariable ... Our findings demonstrate the presence of CCHFV in Algeria, either recently introduced or overlooked. The nearest location where CCHFV has been reported is the Zouala region in Morocco, where the virus was detected in H. marginatum tick larvae and nymphs collected from migratory birds (10). It also confirms association of AP92-like sequences with H. aegyptium ticks. This study shows that the Europe 2 lineage is not restricted to the Balkan region and Turkey. The role of H. aegyptium ticks as CCHFV vectors should be further tested. Further investigation of the distribution of CCHFV in ticks in Algeria is also needed. To date, CCHFV strains of lineage Europe 2 have not been associated with severe disease in humans. However, physicians in Algeria should be aware of potential Crimean-Congo hemorrhagic fever cases.

The impact of climate change and emerging infectious diseases on the blood supply

Ongoing and emerging arbovirus threats in Europe

Crimean-Congo hemorrhagic fever (CCHF) is a medically relevant tick-borne viral disease caused by the Bunyavirus, Crimean-Congo hemorrhagic fever virus (CCHFV). CCHFV is endemic to Asia, the Middle East, South-eastern Europe, and Africa and is transmitted in enzootic cycles among ticks, mammals, and birds. Human infections are mostly subclinical or limited to mild febrile illness. Severe disease may develop, resulting in multi-organ failure, hemorrhagic manifestations, and case-fatality rates up to 30%. Despite the widespread distribution and life-threatening potential, no treatments have been approved for CCHF. Antiviral inhibitory peptides, which antagonize viral entry, are licensed for clinical use in certain viral infections and have been experimentally designed against human pathogenic bunyaviruses, with in vitro and in vivo efficacies. We designed inhibitory peptides against CCHFV with and without conjugation to various polyethylene glycol and sterol groups. These additions have been shown to enhance both cellular uptake and antiviral activity. Peptides were evaluated against pseudotyped and wild-type CCHFV via neutralization tests, Nairovirus fusion assays, and cytotoxicity profiling. Four peptides neutralized CCHFV with two of these peptides shown to inhibit viral fusion. This work represents the development of experimental countermeasures for CCHF, describes a nairovirus immunofluorescence fusion assay, and illustrates the utility of pseudotyped CCHFV for the screening of entry antagonists at low containment settings for CCHF.