Genetic detection of Dobrava-Belgrade hantavirus in the edible dormouse (Glis glis) in central Serbia.

Hantaviruses are endemic in the Balkans, particularly in Serbia, where sporadic cases and/or outbreaks of hantaviral human disease have been reported repeatedly, and evidenced serologically. Here, we present genetic detection of Dobrava-Belgrade virus (DOBV) hantaviral sequences in wild rodents trapped in central Serbia. All the animals were pre-screened serologically by indirect immunofluorescence (IF) test and only those with a positive finding of hantaviral antigens were further tested by polymerase chain reaction. Of the total of 104 trapped animals, 20 were found to be IF positive and of those three were positive for hantaviral RNA: one Microtus arvalis for Tula virus, and one each of Apodemus agrarius and Glis glis for DOBV. Phylogenetic analysis of the obtained sequences implies putative DOBV spillover infection of A. agrarius and G. glis from Apodemus flavicollis. However, future investigations should help to identify the most common natural host and geographical distribution of DOBV in its reservoir hosts in Serbia.

Assessment of Efficiency of Repellent Formulations with N,N-diethyl-m-toluamide in Laboratory Conditions.

The affinity of N,N-diethyl-m-toluamide) (DEET) for various polymers that are potential inert carriers (PEG, PVP, NaCMC, PAA) was studied. Mixtures of DEET and the polymers were prepared and analysed for DEET vapour release using head space analysis. Polyvinylpyrrolidone (PVP) showed the highest affinity for DEET. Several insect repellent formulations in the form of a hydrogel, an o/w cream and a stick were prepared with 5% w/w DEET and either 1% w/w or no PVP. The release of DEET from the formulations was investigated by Head Space gas-mass spectrometry at 32 degrees C. The degree of absorption of DEET through the skin was evaluated by applying these topical preparations to shaved rabbit skin and measuring DEET content in the blood samples by gas-mass spectrometry at various time intervals after application. Based on these results the pharmacokinetic parameters were determined. The minimum penetration of DEET through rabbit skin was obtained with a hydrogel containing PVP as a carrier. The biological efficiency of repellent formulations containing 5% DEET (w/w) was evaluated for repellency using the mosquito Aedes aegypti. The results demonstrated that the hydrogel formulation based on polyacrylic acid containing 5% DEET (w/w) supplemented with 1% PVP (w/w), could serve as a suitable vehicle for repellent preparations containing DEET.

[Use of repellents for protection against vectors of Lyme borreliosis]. / Primena repelenata u zastiti od vektora lajm borelioze.

Preparations with repellent effects in the form of lotion and ointment are formulated. The active substance is Deet (N,N-diethyl-toluamide) and the base constitutes of isopropanil and polyethylene glycol 300 and 1500. The physico-chemical examinations (Deet identification, examination of density, refraction index and Deet contents) have been previously performed. By these investigations the Deet structure was conformed, relative density was determined (1,0014) and also the refraction index (1,5211) (BP 80; USA XXI). The nitrogen (N) content in the resulting substance was 6.44%, in lotion 2.27% and in ointment 0.49%, which correspond to the contents of Deet in the formulated preparations. The evolution of biological efficacy was performed on Ixodes ricinus ticks. The preparation has shown protective effects to the presence of ticks over 60 min under field conditions and up to 146 min under laboratory conditions.

[Lyme borreliosis in Belgrade]. / Lyme borreliosis u Beogradu.

In the period 1990-1992 in Belgrade area a diagnosis of Lyme disease was registered in 487 patients. There were 195 men and 292 women. The disease was registered in all ages groups, median age 38 y. The tick bite has been confirmed in 379 (77.8%) patients. Erythema migrans was the first sign of illness in 387 (84.6%) patients. The disease resoted in neurologic symptoms in 1.8%, joint manifestation in 4.8% and cardiac involvement in 0.9%. Antibodies to Borrelia burgdorferi have been detected by indirect immunofluorescence microscopy in 23.4%. Lyme disease has been registered in all districts of Belgrade. Acarological investigations have detected an infection by Borrelia burgdorferi in 29.0% of investigated ticks.

[Modern immunologic tests in the diagnosis of Lyme borreliosis]. / Savremeni imunoloski testovi u dijagnostici lajm borelioze.

In this work we described the results that were obtained using various immunodiagnostic assays for the detection of lyme borreliosis. Sera of the patients that were in acute or chronical phase of the disease were analysed in indirect immunofluorescent, immunoenzyme and immunoblot assays which were prepared and carried out in our laboratory. As a control for the validity of these investigations, we used sera of the healthy people, as well as of the patients suffering of lues or rheumatoid illnesses. Results that we obtained pointed out the factors responsible for the nonspecific reactions in indirect immunofluorescent and immunoenzyme test. The advantage of the immunoblot analysis in detecting lyme borreliosis is described in this work.

[Lyme disease in human medicine]. / Problem i znacaj lajmske bolesti u humanoj medicini.

Six years of investigations of Lyme disease in our country has offered an opportunity for better understanding of this "new" illness, which is both a medical and a health problem. The number of patients with Lyme disease is found to be similar to the number of patients in other countries. Human infections occur in different areas, rural and urban settings. A high level of collected ticks with Borrelia burgdorferi, suggests a considerable risk of human infection after a tick bite. Difficulties in diagnosing of Lyme disease are connected with different clinical symptoms and signs of the illness a poor health service personnel knowledge og the disease, and the lack of reliable and more convenient laboratory tests for confirmation of the diagnosis. A team work and a multidisciplinary aspect in the investigations and the routine work provide a solid base for a better understanding and gathering much more experience in this field. The individual protection remains a basic control measure, and each person can do the most for self protection. An activity in the health education is also of a very great importance.

[Ecology of Borrelia burgdorferi]. / Ekologija Borelije burgdorferi.

Much still remains to be understood about ecology of Borrelia burgdorferi which is transmitted by a number of ticks belonging primarily to the Ixodes complex, known to feed on different animal hosts and humans. This article pretends to be a review of previous knowledge of Borrelia burgdorferi ecology. Special emphasis is placed on the preliminary results of an investigation conducted in Yugoslavia on the risk of developing lyme borreliosis after a tick bite (Ixodes ricinus). Lyme borreliosis was studied prospectively in 147 persons with tick fastened on them. Dark-field microscopic examination of tick mitgut revealed that 11 (7.9%) od 147 Ixodes ricinus were infected with Borrelia burgdorferi. Lyme borreliosis was demonstrated in only one (0.6%) of all persons bitten by ticks infected with Borrelia burgdorferi. In conclusion, these preliminary data indicate that in Yugoslavia lyme borreliosis infrequently occurred after a tick bite.

[Vectors of Borrelia burgdorferi]. / Vektori Borrelia burgdorferi.

During the period 1990-1992-5,915 ticks were collected and identified in the area of Belgrade. Identification has shown that 99.8% were Ixodes ricinus and 0.2% Dermacentor marginatus. Infection with Borrelia burgdorferi was found in 27.0% to 31.7%. Females were more often infected than males and nymphs more often than larvae. Tick infection was at lowest degree in parks, little bit higher in woods and highest in mixed park-wood areas. Out of 28 locations, six can not be defined as foci of Lyme borreliosis, two are defined as potential foci and 20 as active foci of Lyme borreliosis. In a three year period we have isolated Borrelia burgdorferi in five occasions; in one occasion from ticks collected in Barajevo and in four occasions from ticks collected in Kosutnjak.

[A theoretical model for the study of Lyme borreliosis]. / Teorijski model izucavanja lajm borelioze.

This article is a short review of: a) development of theory on natural foci of communicable diseases in man, b) methodological approach to the investigation of natural foci, e.g. theoretical model in their investigation, and c) the use of theoretical model in the investigation of Lyme borreliosis. A great number of multidisciplinary studies on natural foci zooanthroponoses conducted through many years, in which ecological approach and numerous methods were applied, commencing from studies of the processes at the level of molecules, genes, cells, tissues, organismus, populations and species, contributed to the knowledge that epizootical process of natural foci of zooanthroponoses is almost an ideal natural model. Its investigation allows the establishment of the rules of epidemiological process which, for many reasons (ethical above all), is not accessible for experimental studies. On these principles a general block model has been worked out, which serves for the estimation of epidemiological manifestations of natural foci zooanthroponoses, and which, applied to Lyme borreliosis, enables the determination of active and potential foci and the estimation of the degree of risk of man infection in determined foci. Data obtained in such a way allow planning and application of adequate preventive measures.

[Exposure to Ixodes ricinus bites among soldiers]. / Izlozenost vojnika ubodima Ixodes ricinusa.

Results of an investigation on the exposure to tick bites among soldiers in 6 permanent army structures on the territory of Belgrade are shown. Four army structures were found to be active foci of Lyme borreliosis. Infection rates among Ixodes ticks were from 10% in urban structures to 42.1% in rural structures. Exposure of soldiers to tick bites was the largest in rural army structures--3.40%, while in urban army structures it was 0.14%. Among 38 tick bitten army members, 1 developed clinical manifestation as Lyme borreliosis (2.6%).

[The first isolation of Borrelia burgdorferi in Apodemus flavicollis in Yugoslavia]. / Prva izolacija Borrelia burgdorferi is Apodemus flaviccolis-a u Yugoslaviji.

Small rodent species, as well as their ectoparasitic fauna, and the presence of Borrelia burgdorferi in their blood and internal organs (liver and spleen) was investigated, at two locations (Kosutnjak, Lipovicka suma) in Belgrade. Animals were caught between September 15-17, 1992. On location Kosutnjak two Apodemus flavicollis were trapped and Borrelia burgdorferi was isolated from the spleen of one of them. On location Lipovicka suma 13 rodents were trapped as follows: 10 Apodemus flavicollis, 2 Apodemus agrarius and one Mus musculus. In these the presence of Borrelia burgdorferi was not determined. Ectoparasits Ixodes ricinus, Nesopyllus fasciatus and Haemogamasus spp. have been found. The location Kosutnjak can be considered as an active focus of lyme borreliosis.

[Ixodes ricinus proven as a vector of Lyme borreliosis in Yugoslavia]. / Ixodes ricinus dokazan vektor lajm borelioze i u Jugoslaviji.

263 fasting female Ixodes ricinus were examined for Borrelia burgorferi, the vector of Lyme borreliosis. Female ticks were collected by flagella in the biotopes (Belgrade and Osijek) in which ticks bite patients with Lyme borreliosis. Borrelia burgorferi was proved in 58 (22%) of 263 female ticks in the native preparations of the intestinal contents by darkfield microscopy. In macerates of two groups of 5 female ticks each, Borrelia burgorferi was isolated by cultivation in the modified Kelly nutrient media for borrelia. The supposition that Ixodes ricinus is the vector of Lyme borreliosis in Yugoslavia is confirmed.

[Lyme disease in Yugoslavia]. / Lajmska bolest u Jugoslaviji.

Presented are results of the last five-year period study of the lyme disease in Yugoslavia. On the basis of published papers and professional communications over 2500 cases of lyme disease.were detected in our country within this period. Presented are common epidemiologic and clinical features of the disease and particularly cases diagnosed in Belgrade. Since 1987 a group of various specialists has been working on the research project in collaboration with many centres in the country. Indirect immunofluorescence test as the diagnostic laboratory technique has been introduced in the Military Medical Academy in Belgrade in 1987. First species of Borrelia burgdorferi from ticks Ixodes ricinus have been isolated and cultivated in May and June 1990. Current problems concerning research work, medical and epidemiological aspects of the lyme disease are presented.