Field survey and molecular characterization of apicomplexan parasites in small mammals from military camps in Afghanistan.

Small mammals are an important reservoir for causative agents of numerous infectious diseases, including zoonotic and vector-borne diseases. The occurrence of these pathogens represents a regional but permanent threat for humans and animals in general and might especially weaken military personnel and companion animals in abroad missions. In our study, small mammals collected in military camps in Afghanistan (Feyzabad, Mazar-e Sharif, and Kunduz) were investigated for the presence of apicomplexans using histopathology and molecular methods. For this purpose, well-established and newly developed real-time PCR assays were applied. A high prevalence was detected not only in house mice (Mus musculus), but also in shrews (Crocidura cf. suaveolens) and grey dwarf hamsters (Cricetulus migratorius). The molecular characterization based on the 18S rRNA gene revealed a close relationship to a cluster of Hepatozoon sp. detected in voles of the genus Microtus. Hepatozoon canis DNA was detected in one house mouse as well as in two Rhipicephalus ticks from a dog puppy. In addition, around 5% of the house mice were found to be infected with far related adeleorinids showing the highest sequence identity of 91.5% to Klossiella equi, the only published Klossiella sequence at present. For their better phylogenetic characterization, we conducted metagenomics by sequencing of two selected samples. The resulting 18S rRNA gene sequences have a length of about 2400 base pairs including an insertion of about 500 base pairs and are 100% identical to each other. Histopathology together with organ tropism and detection rates verified this sequence as of Klossiella muris. In conclusion, we documented naturally occurring protozoan stages and the additional taxonomic characterization of a well-known commensal in mice by applying a combination of different approaches. The study is of medical, social, and biological importance for ensuring human and animal health in military camps and also stresses the required awareness for the potential risk of zoonoses.

Five years following first detection of Anopheles stephensi (Diptera: Culicidae) in Djibouti, Horn of Africa: populations established-malaria emerging.

The Asian malaria mosquito, Anopheles stephensi, is a well-known and important vector of Plasmodium falciparum and P. vivax. Until 2013, its geographical distribution was confined to central and southern Asia including the Arabian Peninsula. In the Horn of Africa (HoA) Region, An. stephensi was first recorded from Djibouti in 2012, when it was linked geographically and temporally with an unusual outbreak of urban P. falciparum malaria. In 2016, An. stephensi was detected in the neighbouring Somali Region of Ethiopia. In order to determine whether An. stephensi populations have become established in Djibouti and contributed to the unusual rise in local malaria cases there, we carried out continuous vector surveillance from January 2013 to December 2017, investigated seasonal changes in An. stephensi population densities and bionomics, analysed available literature describing malaria in Djibouti since 2013, and investigated whether An. stephensi may have contributed to local malaria transmission by detecting circumsporozoite antigen of P. falciparum and P. vivax in female anophelines. From 2013 to 2016, seasonal activity of An. stephensi in urban Djibouti City primarily occurred during the colder, wetter season between September and May, with either no or rare trap catches from June to August. Unlike past years, this species was detected year-round, including the extremely hot summer months of June to August 2017. This change in seasonal occurrence may indicate that An. stephensi populations are adapting to their new environment in sub-Saharan Africa, facilitating their spread within Djibouti City. Among the 96 female An. stephensi investigated for malaria infectivity, three (3.1%) were positive for P. falciparum circumsporozoite antigen, including one P. falciparum/P. vivax VK 210 double infection. Subsequent to the unusual resurgence of local malaria in 2013, with 1684 confirmed cased reported for that year, malaria case numbers increased continuously, peaking at 14,810 in 2017. Prior to 2016, only P. falciparum malaria cases had been reported, but in 2016, autochthonously acquired P. vivax malaria cases occurred for the first time at a rate of 16.7% among all malaria cases recorded that year. This number increased to 36.7% in 2017. Our data indicate that the dynamics of malaria species in Djibouti is currently changing rapidly, and that An. stephensi can be involved in the transmission of both P. falciparum and P. vivax, simultaneously. Considering the extremely high potential impact of urban malaria on public health, the timely deployment of optimal multinational vector surveillance and control programs against An. stephensi is strongly recommended, not only for the HoA Region, but for the entire African continent.

Long-lasting permethrin-impregnated clothing: protective efficacy against malaria in hyperendemic foci, and laundering, wearing, and weathering effects on residual bioactivity after worst-case use in the rain forests of French Guiana.

Personal protective measures against hematophagous vectors constitute the first line of defense against arthropod-borne diseases. However, guidelines for the standardized testing and licensing of insecticide-treated clothing are still lacking. The aim of this study was to analyze the preventive effect of long-lasting polymer-coated permethrin-impregnated clothing (PTBDU) against malaria after exposure to high-level disease transmission sites as well as the corresponding loss of permethrin and bioactivity during worst-case field use. Between August 2011 and June 2012, 25 personnel wearing PTBDUs and exposed for 9.5 person-months in hyperendemic malaria foci in the rain forest of French Guiana contracted no cases of malaria, whereas 125 persons wearing untreated uniforms only, exposed for 30.5 person-months, contracted 11 cases of malaria, indicating that PTBDU use significantly (p = 0.0139) protected against malaria infection. In the field, PTBDUs were laundered between 1 and 218 times (mean 25.2 ± 44.8). After field use, the mean remaining permethrin concentration in PTBDU fabric was 732.1 ± 321.1 min varying between 130 and 1270 mg/m2 (mean 743.9 ± 304.2 mg/m2) in blouses, and between 95 and 1290 mg/m2 (mean 720.2 ± 336.9 mg/m2) in trousers. Corresponding bioactivity, measured according to internal licensing conditions as KD99 times against Aedes aegypti mosquitoes, varied between 27.5 and 142.5 min (mean 47.7 ± 22.1 min) for blouses, and between 25.0 and 360 min (mean 60.2 ± 66.1 min) for trousers. We strongly recommend the use of long-lasting permethrin-impregnated clothing for the prevention of mosquito-borne diseases, including chikungunya, dengue, and zika fevers, which are currently resurging globally.

Bioactivity and laundering resistance of five commercially available, factory-treated permethrin-impregnated fabrics for the prevention of mosquito-borne diseases: the need for a standardized testing and licensing procedure.

Personal protective measures against hematophagous vectors constitute the first line of defense against arthropod-borne diseases. In this regard, a major advance has been the development of residual insecticides that can be impregnated into clothing. Currently, however, information on specific treatment procedures, initial insecticide concentrations, arthropod toxicity, residual activity, and laundering resistance is either fragmentary or non-existent, and no World Health Organization Pesticides Evaluation Scheme or other guidelines exist for the standardized testing and licensing of insecticide-treated clothing. The aim of this study was to analyze the insecticide content, contact toxicity, laundering resistance, and residual activity of five commercially available and commonly used permethrin-treated fabrics-Insect Shield, ExOfficio, Sol's Monarch T-shirts, battle dress uniforms (BDUs), and Labonal socks-against vector-competent Aedes aegypti, Anopheles stephensi, and Culex pipiens mosquitoes under laboratory conditions. Prior to laundering, permethrin concentrations ranged from 4300 to 870 mg/m(2) whereas, after 100 defined machine launderings, the remaining permethrin content fell to between 1800 and 20 mg/m(2), a percentage permethrin loss of 58.1 to 98.5 %. The highest 99 % knockdown (KD99) efficacy of permethrin was detected in Ae. aegypti, followed by An. stephensi and Cx. pipiens demonstrating that Ae. aegypti is the most sensitive species and Cx. pipiens the least sensitive. After 100 launderings, the remaining biocidal efficacy differed markedly among the five brands, with KD99 times varying from 38.8 ± 2.9 to >360 min for Ae. aegypti, from 44 ± 3.5 to >360 min for An. stephensi, and from 98 ± 10.6 to >360 min for Cx. pipiens. Overall, the ranking of the residual biocidal efficacies within the five brands tested was as follows: BDU ≈ Labonal > Sol's Monarch > ExOfficio > Insect Shield. When applying German Armed Forces licensing conditions, none of the four products available in the civilian market would completely meet all the necessary efficacy and safety requirements fulfilled by BDUs. Therefore, we strongly recommend standardized testing and licensing procedures for insecticide-treated clothing, with defined cutoff values for initial maximum and post-laundering minimum concentrations of permethrin as well as figures for permethrin migration rates, arthropod toxicity, homogeneity on fabrics, residual activity, and laundering resistance.

Pilot study assessing the effectiveness of factory-treated, long-lasting permethrin-impregnated clothing for the prevention of tick bites during occupational tick exposure in highly infested military training areas, Germany.

The protective effectiveness of factory-based permethrin-impregnated polymer-coated battle dress uniforms (PTBDUs) against tick bites was evaluated at four military training areas in southwestern and central Germany where tick bite incidence is known to be high. Data were analyzed by comparing tick bite incidence using non-permethrin-treated BDUs (NTBDUs) during 2009 versus PTBDUs during 2010 and 2011, the first two years after their formal introduction for in-country use in the German Bundeswehr. During 2009, 262 individual tick bites were reported at the four training sites, resulting in a tick bite incidence of 8.8 % per exposed person when wearing NTBDUs only. In 2010 and 2011, one tick bite case occurred under field conditions each year that PTBDUs were worn, corresponding to a protective effectiveness of 99.6 and 98.6 %. These data imply an annual tick bite incidence of 0.035 and 0.078 % per exposed person, respectively. Between 2010 and 2011, a 0.8 % decline in the protective effectiveness of PTBDUs was observed. Five tick bite incidents occurred while wearing non-impregnated parkas over correctly worn PTBDUs. Ixodes ricinus ticks were collected by standard tick drags from 2009 to 2011, with high mean annual densities ranging from 28.9 to 106.5 ticks per 100 m(2), while single drags revealed tick densities between zero and 381 ticks per 100 m(2). Overall, 4596 I. ricinus ticks (54 ♂, 82 ♀, 1776 nymphs, and 2684 larvae) were collected, of which 128 (2.8 %; mean annual range, 0-10.1 %) were Borrelia burgdorferi s.l. positive. The Borrelia genospecies distribution was as follows: 112 (87.5 %) Borrelia afzelii, 10 (7.8 %) B. burgdorferi s.s., and 6 (4.7 %) Borrelia garinii. Neither the tick density means from 2009 to 2011 nor associated B. burgdorferi s.l. prevalences differed significantly among the military locations investigated. The documented tick bite reductions clearly demonstrate the powerful protective effectiveness of properly worn PTBDUs against tick bites. Nevertheless, all apparel worn over PTBDUs should also be impregnated with permethrin in order to prevent tick infestation and subsequent bites.

Synergistic insecticidal and repellent effects of combined pyrethroid and repellent-impregnated bed nets using a novel long-lasting polymer-coating multi-layer technique.

New and improved strategies for malaria control and prevention are urgently needed. As a contribution to an optimized personal protection strategy, a novel long-lasting insecticide and repellent-treated net (LLIRN) has been designed by binding combinations of permethrin plus N,N-diethyl-m-toluamide (DEET), or insect repellent 3535 (IR3535), and etofenprox plus DEET, onto fibres of bed net fabric employing a new multi-layer polymer-coating technique. Protective repellent efficacy, toxicological effectiveness and residual activity of 12 LLIRN types have been evaluated by laboratory testing against adult Aedes aegypti. The novel multi-layer LLIRN design allowed simultaneous embedding at concentrations up to 5,930 mg/m(2) for DEET, 3,408 mg/m(2) for IR3535, 2,296 mg/m(2) for permethrin and 2,349 mg/m(2) for etofenprox, respectively. IR3535 layers prevented co-binding of additional pyrethroid-containing polymer layers, thus making pyrethroids plus DEET LLIRNs an ideal combination. All LLIRNs revealed synergistic insecticidal effects which, when measured against concentration controls of the isolated compounds, were significant in all LLIRN types designed. DEET in DEET plus permethrin LLIRNs significantly (p < 0.0001) reduced the concentration-dependent permethrin 100 % knockdown (KD) time from 55 to 75 %, the corresponding 100 % kill time (p < 0.0001) from 55 to 64 %. DEET in DEET plus etofenprox LLIRNs reduced the dose-specific 100 % knockdown (KD) time of etofenprox from 42 to 50 % (p = 0.004), the 100 % kill time from 25 to 38 % (p < 0.0001). Permethrin or etofenprox did not influence spatial repellency of DEET or IR3535 on LLIRNs. Vice versa, DEET and IR3535 increased spatial and excitatory repellency and reduced landing and probing frequency on LLIRNs resulting in strongly enhanced biting protection, even at low concentrations. One hundred percent biting and probing protection of stored LLIRNs was preserved for 83 weeks with the 5,930 mg/m(2) DEET and 2,139 mg/m(2) etofenprox LLIRN, for 72 weeks with the 5,002 mg/m(2) DEET and 2,349 mg/m(2) etofenprox LLIRN, for 63 weeks with the 3,590 mg/m(2) DEET and 1,208 mg/m(2) permethrin LLRN, and for 61 weeks with the 4,711 mg/m(2) DEET and 702 mg/m(2) etofenprox LLIRN. Because 100 % bite protection with up to 75 % quicker contact toxicity of pyrethroids were documented, synergistic toxicological and repellent effects of multi-layer polymer-coating LLIRNs may overcome LLIN-triggered selection pressure for development of new kdr- and metabolic pyrethroid resistances while simultaneously increasing protective efficacy also against kdr- and metabolic pyrethroid-resistant mosquitoes substantially due to the repellent-induced effects of LLIRNs thus indicating that this approach is a promising new candidate for future bed net, curtain, and window screen impregnation aiming at optimized prevention from mosquito-borne diseases.

Novel long-lasting impregnation technique transferred from clothing to bednets: extended efficacy and residual activity of different pyrethroids against Aedes aegypti as shown by EN ISO 6330-standardized machine laundering.

A novel long-lasting insecticide-treated net (LLIN) has been designed by embedding the pyrethroids deltamethrin, cyfluthrin, permethrin, or etofenprox, onto the fibres of bed net cotton fabric by transferring a new polymer-coating technique from clothing to netting material. EN ISO 6330, the more stringent European Norm for domestic washing and drying procedures for textile testing, has been newly employed to monitor and validate more precise wash durability and residual bioactivity of LLINs. Wash durability, residual insecticidal activity and mosquito landing/biting protection has been investigated and compared with four commercially available LLINs. The pyrethroid-embedding impregnation technique allows binding of high concentrations exceeding 8,000 mg permethrin/m(2) within one single-polymer layer. Recovery rates of 95.7 ± 5.8%, 92.4 ± 14.0%, 70.2 ± 9.1%, and 64.2 ± 6.3% for cyfluthrin; 32.4 ± 11.4%, 32.4 ± 12%, 35.1 ± 16.2%, and 35.8 ± 15.7% for deltamethrin; 75.3 ± 12.9%, 57.1 ± 15.8%, 48.5 ± 4.0%, and 35.6 ± 4.7% for etofenprox; and 95.7 ± 5.8%, 80.2 ± 8.6%, 39.1 ± 7.9%, and 34.1 ± 8.8% were measured after 1, 5, 10, and 20 launderings. Laundering resistance was highest with cyfluthrin > deltamethrin ≈ etofenprox ≈ permethrin. After one and five launderings, commercial LLINs revealed percentage pyrethroid recovery rates of 26.3 ± 11.8% and 0.9 ± 1.2% for the Care Plus(®) net; 31.8% and 28.9% for the Permanet(®) 2.0; 61.4% and 45.6% for the Net Protect(®); and 80.4% and 68.3% for the Conmanet(®). Recovery rates reveal that the polymer-coating method resulted in extended wash durability. Dose-dependent 100% knockdown rates were most effective with deltamethrin > cyfluthrin > permethrin ≈ etofenprox. LLINs tested at concentrations up to 8,000 mg permethrin/m(2) did not protect from mosquito bites. Results on long-lasting efficacy and bioactivity of the polymer-coating technique show that this new LLIN technique is a highly promising potential candidate for future malaria control strategies. Standardized machine laundering and drying according to EN ISO 6330 is highly recommended to monitor and validate wash durability and residual activity of LLINs.

Small Mammals as Reservoir for Zoonotic Agents in Afghanistan.

INTRODUCTION: Rodents and other small mammals can serve as reservoirs for a large number of zoonotic pathogens. A higher risk of infection with rodent-borne pathogens exists for humans with direct contact to rodents and/or their excretions, e.g., soldiers in operation areas. To date, little is known about endemic human pathogenic disease agents that are naturally associated with small mammals in Afghanistan. The aim of this study was to screen abundant rodents and insectivores collected from 2009 to 2012 in four field camps of the German Federal Armed Forces (Bundeswehr) in Northern Afghanistan for the presence of different pathogens. MATERIALS AND METHODS: Isolated nucleic acids from ear pinna were screened by real-time PCR for spotted fever group (SFG) rickettsiae and from liver samples for Francisella spp., Coxiella burnetii, Brucella spp., Yersinia pestis, and poxvirus. Chest cavity lavage (CCL) samples were tested for antibodies against SFG and typhus group (TG) rickettsiae, as well as against flaviviruses using an indirect immunofluorescence assay. RESULTS: Rickettsial DNA was detected in 7/750 (1%) ear pinna samples with one being identified as Rickettsia conorii. Antibodies against SFG rickettsiae were detected in 15.3% (n = 67/439) of the small mammals; positive samples were only from house mice (Mus musculus). Antibodies against TG rickettsiae were found in 8.2% (n = 36/439) of the samples, with 35 from house mice and one from gray dwarf hamster (Cricetulus migratorius). Flavivirus-reactive antibodies were detected in 2.3% (n = 10/439) of the investigated CCL samples; again positive samples were exclusively identified in house mice. All 199 investigated liver-derived DNA preparations were negative in the Francisella spp., C. burnetii, Brucella spp., Y. pestis, and poxvirus-specific PCRs. CONCLUSIONS: Further investigations will have to prove the potential value of rodents in army camps as sentinel animals.

[Diagnosis and therapy of cutaneous and mucocutaneous Leishmaniasis in Germany]. / S1-Leitlinie--Revision November 2010 Diagnostik und Therapie der kutanen und mukokutanen Leishmaniasis in Deutschland.

The incidence of cutaneous and mucocutaneous Leishmaniasis (CL/MCL) is increasing globally, also in Germany, although the cases are imported and still low in number. The current evidence for the different therapies has many limitations due to lack of sufficient studies on the different Leishmania species with differing virulence. So far there is no international gold standard for the optimal management. The aim of the German joint working group on Leishmaniasis, formed by the societies of Tropical Medicine (DTG), Chemotherapy (PEG) and Dermatology (DDG), was to establish a guideline for the diagnosis and treatment of CL and MCL in Germany, based on evidence (Medline search yielded 400 articles) and, where lacking, on consensus of the experts. As the clinical features do not necessarily reflect the involved Leishmania species and, as different parasite species and even geographically distinct strains of the same species may require different treatments or varying dosages or durations of therapy, the guidelines suggest for Germany to identify the underlying parasite prior to treatment. Because of relevant differences in prognosis and ensuing therapy species should be identified in i) New World CL/MCL (NWCL/ MCL) to distinguish between L. mexicana-complex and subgenus Viannia, ii) in suspected infections with L. mexicana-complex to distinguish from L. amazonensis, and iii) in Old World CL (OWCL) to distinguish between L. infantum and L. major, L. tropica, or L. aethiopica. A state-of-the-art diagnostic algorithm is presented. For recommendations on localized and systemic drug treatment and physical procedures, data from the accessible literature were adjusted according to the involved parasite species and a clinical differentiation into uncomplicated or complex lesions. Systemic therapy was strictly recommended for i) complex lesions (e. g. > 3 infected lesions, infections in functionally or cosmetically critical areas such as face or hands, presence of lymphangitis), ii) lesions refractory to therapy, iii) NWCL by the subgenus Viannia or by L. amazonensis, iv) in MCL and v) in recalcitrant, or disseminating or diffuse cutaneous courses. In e. g. infection with L. major it encompasses miltefosine, fluconazole and ketoconazole, while antimony or allopurinol were here considered second choice. Local therapy was considered appropriate for i) uncomplicated lesions of OWCL, ii) L. mexicana-complex and iii) pregnant women. In e. g. infection with L. major it encompasses perilesional antimony, combined with cryotherapy, paromomycin 15 %/in methylbenzethoniumchlorid 12 % and thermotherapy. The group also stated that there is an urgent need for improving the design and the way of publishing of clinical trials in leishmaniasis.

Insecticidal, acaricidal and repellent effects of DEET- and IR3535-impregnated bed nets using a novel long-lasting polymer-coating technique.

A novel long-lasting repellent-treated net (LLRTN) has been designed by binding the skin repellents N,N-diethyl-m-toluamide (DEET), or IR3535, onto the fibres of bed net fabric using a new polymer-coating technique. The repellent toxicological effectiveness and residual activity of a factory-based repellent-impregnated fabric has been evaluated by laboratory testing against adult Aedes aegypti mosquitoes and nymphal Ixodes ricinus ticks. By using this repellent-embedding impregnation technique, concentrations exceeding 10 g/m(2) could be achieved with one single polymer layer. Both DEET- and IR3535-impregnated fabrics revealed a dose-dependent insecticidal as well as acaricidal activity. One hundred percent knockdown times of DEET-treated bed nets ranged from 187.5 +/- 31.8 to 27.5 +/- 3.5 min against A. aegypti, and between 214 +/- 47 and 22.6 +/- 5 min against nymphal I. ricinus, linked to a DEET concentration of 1.08 and 10.58 g/m(2), respectively. With IR3535, A. aegypti produced dose-dependent 100% knockdown times varying from 87.5 +/- 10.6 to 57.5 +/- 3.5 min and between 131.4 +/- 6.5 and 33.8 +/- 5 min against nymphal I. ricinus, respectively, linked to concentrations between 1.59 and 10.02 g/m(2). One hundred percent repellency measured by complete landing and biting protection of impregnated fabric by using the arm-in-cage test could be achieved at DEET concentrations exceeding 3.7 to 3.9 g/m(2), and for IR3535 concentrations over 10 g/m(2). One hundred percent landing and biting protection could be preserved with DEET-treated fabrics for 29 weeks at an initial concentration of 4.66 g/m(2), 54 weeks at 8.8 g/m(2), 58 weeks at 9.96 g/m(2) and 61 weeks at 10.48 g/m(2) for DEET, and 23 weeks for IR3535-treated fabric at a concentration of 10.02 g/m(2). Unlike repellent-treated fabric, a brand of a commercially available long-lasting insecticide-treated net tested containing 500 mg permethrin/m(2) did not protect from mosquito bites. First results on bioactivity and long-lasting efficacy show that the new LLRTN technique is highly promising as a potential candidate for future malaria control strategies, especially in areas where pyrethroid resistance occurs.

Use of vector diagnostics during military deployments: recent experience in Iraq and Afghanistan.

Vector-borne diseases such as malaria, dengue, and leishmaniasis are a threat to military forces deployed outside of the United States. The availability of specific information on the vector-borne disease threat (e.g., presence or absence of a specific disease agent, temporal and geographic distribution of competent vectors, and vector infection rates) allows for effective implementation of appropriate measures to protect our deployed military forces. Vector diagnostics can provide critical, real-time information crucial to establishing effective vector prevention/control programs. In this article we provide an overview of current vector diagnostic capabilities, evaluate the use of vector diagnostics in Operation Enduring Freedom and Operation Iraqi Freedom, and discuss the concept of operations under which vector diagnostics are employed.

Zoonotic cutaneous leishmaniasis outbreak in Mazar-e Sharif, northern Afghanistan: an epidemiological evaluation.

In 2005, 200 cases of zoonotic cutaneous leishmaniasis (ZCL) were recorded among International Security Assistance Force (ISAF) troops stationed in the Mazar-e Sharif airport area. Within the local population, investigations revealed 3782 cases of ZCL, 174 cases of anthroponotic cutaneous leishmaniasis (ACL), and 2 cases of visceral leishmaniasis (VL) in the period from March 21, 2004 to March 20, 2005, and 4045 cases of ZCL, 198 cases of ACL, and no cases of VL from March 21, 2005 to March 20, 2006. The previously unknown transmission dynamics of ZCL, and differing seasonal distribution of ZCL and ACL, are here defined, thus permitting quantification and prediction of infection rates in deployed troops for the first time. At Mazar-e Sharif, Phlebotomus papatasi and Rhombomys opimus occurred in the highest densities yet observed, together with record-high Leishmania major infection rates. Data indicate the existence of high-density, anthropogenically induced ZCL in Afghanistan.

Differences in transmission seasons as an epidemiological tool for characterization of anthroponotic and zoonotic cutaneous leishmaniasis in northern Afghanistan.

Regional epidemiological data, when available from Afghan or international health authorities, usually include cutaneous leishmaniasis cases without further elaboration. Scientific reports from Afghanistan mainly focus on the current status of war and refugee-related anthroponotic cutaneous leishmaniasis (ACL), but little is known about zoonotic cutaneous leishmaniasis (ZCL), its regional and seasonal distribution, or other disease characteristics. Multiple field investigations revealed that both ACL and ZCL are widespread in Afghanistan and may show sharp differences in specific epidemiology and incubation periods. The previously unknown transmission dynamics and differing seasonality of ZCL, with maximum clinical cases in September and October, as opposed to the ACL peak in March and April, are here described, thus permitting for the first time prediction of the causative Leishmania species in undiscriminated CL reports. Results show that epidemiological differences may serve as a convenient tool for discriminating between ACL and ZCL, at least in northern and central Afghanistan, which can be important because specific treatment and control measures may be different.

Risk assessment of Bundeswehr (German Federal Armed Forces) permethrin-impregnated battle dress uniforms (BDU).

In an age when vector-borne diseases are emerging worldwide, personal protective measures are essential for shielding soldiers and other exposed persons from arthropod attack. The development of permethrin-impregnated clothing has been one recent advance in protecting persons at-risk. However, to date risk assessment has not been performed related to wearing permethrin-impregnated clothing over longer time periods. Therefore, this paper describes relevant toxicological aspects of permethrin and estimates the extent of dermal permethrin uptake by soldiers wearing impregnated uniforms by determining urine metabolites of permethrin. The exposure monitoring conducted in wearers of untreated uniforms did not show any signs of increased permethrin uptake and was similar to that of the general population in Germany. By contrast, studies involving the soldiers wearing permethrin-impregnated uniforms identified far higher internal exposure, the amounts of urine metabolites clearly above the reference value for the background exposure of the German population at large. Comparing the median excretion values, an approximately 200 times higher exposure can be assumed. The excretion levels of the subject with the maximum amount of metabolites correspond to an internal exposure of around 5-6microg/kg body weight and day thereby considering that biomonitoring could not take all urine metabolites and other elimination routes into account. Based on an oral absorption rate of 50%, the internal dose of 5-6microg/kg body weight and day would correspond to an oral uptake of permethrin which is around 20% of the ADI value of 50microg/kg body weight and day. In addition, based on these data and using a dermal absorption rate of 2% the permethrin dose reaching the skin was estimated to be 250microg/kg body weight and day. Considering a standard body weight and the area covered by the uniform, an exposure level of about 1.25microg permethrin/cm(2) skin and day can be calculated. Clinical subjective symptoms were recorded by means of a self-reporting questionnaire which has been developed and used for this specific purpose in environmental outpatient departments in both groups (wearers of impregnated versus non-impregnated uniforms). Only minor sensory impairments were identified in one of the studies (Kabul/Afghanistan) which may represent skin paraesthesiae. Based on these results, it can be assumed that the normal use of permethrin-treated uniforms does not affect human health to a clinical relevant extent. We recommend that the release rate of permethrin from the textile material should be strictly monitored by means of a quality assurance method. It should comply with standards to which the results of this study may contribute.

Factory-based permethrin impregnation of uniforms: residual activity against Aedes aegypti and Ixodes ricinus in battle dress uniforms worn under field conditions, and cross-contamination during the laundering and storage process.

The factory-based permethrin coating technique has only recently been developed. Consequently, no data are available on residual activity, laundering, and weathering resistance in impregnated battle dress uniforms (BDUs) worn under military deployment conditions, or on the cross-contamination potential of such uniforms. Herein, factory-impregnated BDUs worn-out during military deployment to Afghanistan were investigated for residual permethrin concentration, residual efficacy against arthropod vectors, and cross-contamination during laundering and storage. When compared with BDUs subjected to 50 defined washings using the U.S. Insect/Arthropod Repellent Fabric Treatment method, no significant differences in efficacy were observed against Aedes mosquitoes, but remaining knockdown activity in Ixodes ticks was significantly better in polymer-coated BDUs. BDUs impregnated by the polymer-coating method were found to be effective for the life of the uniform, ensuring protection of soldiers in the field from arthropod vectors, while causing less cross-contamination than those treated by the Insect/Arthropod Repellent Fabric Treatment method.

Genetic evidence for tula virus in Microtus arvalis and Microtus agrestis populations in Croatia.

To determine the threat of hantavirus infection to U.S. Forces, small mammals were sampled from training areas within Croatia. Of the 152 samples, 20 were positive for Tula virus (TUL), 12 common voles (Microtus arvalis) and eight field voles (Microtus agrestis). Sequences from M. agrestis were found in five and sequences from M. arvalis were found in six of seven sequence groups. The high percentage of the same TUL sequences in M. agrestis and M. arvalis suggests the co-occurrence of this virus in both Microtus species is not an accident. If M. agrestis field voles were accidentally infected with TUL, the percentage of polymerase chain reaction-positive animals should be lower than that of M. arvalis. Because the infection rate in M. arvalis (11.8%) was less than half of that found in M. agrestis (27.6%), it is unlikely that the predominance of positive M. agrestis could be due to accidental exposure. It is much more likely that the Croatian virus is circulating between both rodent species.

Contact toxicity and residual activity of different permethrin-based fabric impregnation methods for Aedes aegypti (Diptera: Culicidae), Ixodes ricinus (Acari: Ixodidae), and Lepisma saccharina (Thysanura: Lepismatidae).

The effectiveness and residual activities of permethrin-impregnated military battle dress uniforms were evaluated by comparing a new company-manufactured ready-to-use polymer-coating method with two "dipping methods" that are currently used to treat uniforms. Residual permethrin amounts and remaining contact toxicities on treated fabrics before and after up to 100 launderings were tested against Aedes aegypti (L.), Ixodes ricinus (L.), and Lepisma saccharina (L.). The residual amount of permethrin was considerably higher with the polymer-coating method: 280 mg a.i./m2 after 100 launderings, compared with 16 and 11 mg a.i./m2, respectively, obtained when using the two dipping methods. Hard ticks were most susceptible to the new polymer-coating method, resulting in prelaundering 100% knockdown times of 7.0 +/- 0.9 min, whereas equivalent times for the dipping methods were 7.9 +/- 0.35 min and 8.0 +/- 0.54 min, respectively. After 100 launderings, 100% knockdown of I. ricinus nymphs was reached at 15.2 +/- 1.04 min using the polymer-coating method, compared with 178.8 +/- 24.7 min and 231 +/- 53.6 min, respectively, using the dipping methods. Similar results were obtained for Ae. aegypti and L. saccharina, indicating that the polymer-coating method is more effective and efficient when compared with the dipping methods.

Hantavirus host/virus interactions within Southeast Europe.

Viral studies have historically approached their phylogenetic analysis without consideration of the impact of the role the host plays in evolution. Our study examines host/viral interactions through analysis of the phylogenetic relationship between hantavirus genetic sequences and host cytochrome B sequences. Phylogenetic analysis of known Hantavirus genetic sequences were performed using PAUP 3.1.1 (vers. 4.0.0d64). Only sequences available through GENBANK were analyzed. Phylogenetic analysis of hantavirus sequences revealed distinct patterns based upon geographic area. These patterns coincided with the known ranges of reservoir hosts. Multiple hosts for individual viruses and multiple viruses in a single host species for hantaviruses have been described. This may be due to accidental exposure, host-switching, co-speciation, or broad co-accommodation. Since the host is the actual environment that the virus survives in, changes in the host over time could potentially directly influence changes in the virus. Multiple viruses and hosts collide in Southeastern Europe increasing the prospect of finding distinct viral/host relationships. Rodent Cytochrome B is very well conserved and can be used to tract host lineage. By tracking the relationship of infected hosts, we theorize that patterns in host DNA will emerge that will mirror patterns in viral sequences. This analysis of the host DNA could provide an understanding into the causes of variation in hantaviral sequences, pathogenicity, transmissibility, infectivity, viral range and expand our knowledge of viral/host interactions. Surveillance for viruses in the field should include analysis of the host DNA in combination with the viral analysis.