Gene-specific features of tick-borne borreliosis in Russia

Aim. To study the gene-specific composition of Borrelia spp. in ixodid ticks inhabiting the foci of ixodid tick-borne borreliosis (ITBB) in the south of Western Siberia.Materials and Methods. The gene-specific composition of Borrelia spp. in ixodid ticks was determined by inoculation on a BSK-H nutrient medium, real-time polymerase chain reaction, and sequencing (1148 specimens of ixodid ticks collected from vegetation and 2183 specimens withdrawn from humans).Results. Infection of ticks with borrelia ranged from 22.4% in the Altai Republic to 56.9% in the Novosibirsk Region. There were no significant differences in the levels of Borrelia infection between two major ixodides, I. persulcatus and I. pavlovskiy (average infection levels 40.0% and 38.8%, respectively). At least five gene species of pathogenic Borrelia (B. garinii, B. afzelii, B. bavariensis, B. miyamotoi, and B. spielmanii) have been identified. The GenBank database contains 45 nucleotide sequences of the intergenic spacer rrf (5S)-rrl (23S). Prevalence of gene species B. garinii and B. afzelii in different ticks (I. persulcatus and I. pavlovskiy) did not differ significantly, yet B. garinii was found to be more frequent than B. afzelii. The frequency of infection of I. persulcatus ticks with B. miyamotoi was significantly (3.5-fold) as compared to those of B. garinii and B. afzelii. In D. reticulatus ticks, the DNA of B. spielmanii and B. miyamotoi was detected.Conclusion. It is necessary to continue studies to assess the role of the meadow ticks D. reticulatus in the circulation of different Borrelia spp. in various natural foci within the Russian Federation.

О находках Rhipicephalus (Boophilus) annulatus (Say, 1821) (Acari, Ixodidae) на западе Атырауской области

Blood-sucking ticks are vectors of protozoal, bacterial, viral and rickettsial diseases. The study of ixodes ticks: fauna, ecology, biology is of great importance not only for epidemiology but is also of scientific interest. Changes in natural and climatic conditions, hydrological regime, rapid development of human economic activities lead to the changes in the fauna of invertebrates including ixodid ticks. As they invade new territories ticks may expand their habitats for these dangerous for animals and humans infections. That is why the study of tick fauna, changes in species diversity of ticks and monitoring of their abundance is so important for monitoring of natural foci. The emergence of a new species of ixodid tick Rhipicephalus annulatus, in western Kazakhstan, which began to be registered from 2019 in Kurmangazy district of Atyrau region, bordering the Astrakhan region of the Russian Federation and its re-finding in 2020 is a very important fact to assess the epidemiological danger for humans and animals in this territory.

Molecular and microscopic detection of Babesia caballi and Theileria equi among working horses and donkeys in Cairo and Giza provinces of Egypt

Equine piroplasmosis (EP) is an ixodid tick-borne disease caused by Theileria equi and/or Babesia caballi that can lead to severe health issues and economic losses among equine population. This study aimed to determine the prevalence of T. equi and B. caballi among Egyptian equines based on microscopy and conventional PCR. Also, to determine the effect of season, age, and sex of on their prevalence and determining the difference in sensitivity between microscopy and conventional PCR in the diagnosis of EP. This study was carried out on 432 blood samples randomly collected from 146 horses and 286 donkeys during a period from April 2016 to March 2018. Microscopic examination revealed that among horses, 13 (8.9%) and 4 (2.7%) were infected by T. equi and B. caballi respectively. While among donkeys, 22 (7.7%), 16 (5.6%) respectively. While mixed infections were detected in 4 (1.4%) donkeys. There was a statistically nonsignificant relation between prevalence of infection and season and sex of equines but the highest prevalence was recorded in age group less than 5 years old. By conventional PCR, among 64 horses, 15 (23.4%) and 8 (12.5%) were infected by T. equi and B. caballi, respectively. While among 76 donkeys, 36 (47.4%), 16 (21.1%), and 5 (6.6%) were infected by T. equi, B. caballi, and mixed infection, respectively. Our finding proved the existence of T. equi and B. caballi among equines.

THE ABUNDANCE OF BORRELIA INIXODID TICKS OF BAIKAL REGION

Ixodid ticks (n=3714) from natural foci of Baikal Region (Irkutsk Region and Republic of Buryatia) have been examined for pathogenic Borrelia DNA during 2013-2010. On average 40.9 % of the samples were positive for Borrelia markers during the survey period; the range of variantion was – 32–55 %. The increasing of infection rate in ticks is traced in multi-year trends. During the ticks’ activity season, maximal infection rates have been noted at the end of the season (60 %), minimal – at the very beginning of the season (28,6 %) and on the peak of vectors’ abundance (36-39 %). The significant geographical, species and age differences have been detected in Borrelia infection rate of Ixodid ticks. Borrelia DNA have been detected considerably more often in taiga tick (the main vector of Ixodid tick borrelioses in Siberia) and in its nymphs more often, than in adult ticks. There was no difference in the infection rate of male and female ticks, and also between ticks, collected from the vegetation and from human and animals. The ratio og genotyped Borrelia species had been as follows: 64.2 % – B. garinii, 21,7 % – B. afzelii, 14,2 % – B. miyamotoi. Key words: Ixodid ticks, pathogenic Borrelia, PCR, genotyping, Baikal Region.

Acaricidal activity and biochemical analysis of Citrus limetta seed oil for controlling Ixodid Tick Rhipicephalus microplus infesting cattle

The present study was performed with an objective to determine acaricidal activity of Citrus limetta seed oil (CLO) against cattle tick Rhipicephalus microplus. The mode of action of CLO was determined by evaluating biochemical enzymatic levels in treated ticks. CLO was obtained by extracting with n-hexane as solvent in soxhlet apparatus. The active compounds in CLO were analyzed through Gas chromatography. Acaricidal activity of CLO was evaluated using larval packet test (LPT) and adult immersion test (AIT). Results of chromatographic analysis showed that palmitic acid, stearic acid, α-Tocopherol and bergamol were present as major active constituents along with other compounds. The lethal concentration (LC50) of CLO was found to be 2.87% and 3.96% for larvae and adult of R. microplus ticks, respectively. In vitro acaricidal bioassay revealed 100% mortality of ticks in both LPT and AIT at 12.5% dose. CLO significantly (p < 0.001) reduced oviposition rate, hatching ability of eggs and reproduction efficiency of ticks. Biochemical analysis of ticks homogenate showed different enzymatic levels. The results revealed that CLO disturbed protective enzymes functions by reducing level of SOD, GST, MAO and AChE but increasing NOS level in ticks as compared with control group. Thus, CLO may be used as an effective acaricidal agent in controlling R. microplus ticks.

Surveillance of Borrelia miyamotoi-carrying ticks and genomic analysis of isolates in Inner Mongolia, China

Background Borrelia miyamotoi is a newly described relapsing fever spirochete transmitted by ixodid tick species. Little is known about the prevalence of B. miyamotoi infections in humans and ticks in Inner Mongolia, China. Therefore, we investigated the prevalence of B. miyamotoi in Ixodes persulcatus ticks, and we aimed to isolateB. miyamotoi from I. persulcatus from four regions of Greater Khingan, Inner Mongolia, China. Methods From May to June each year during the period 2016–2019, host-seeking adult I. persulcatus ticks were collected from vegetation. Genomic DNA was prepared from half of each tick body for PCR template, and the remaining half was used to cultivate B. miyamotoi in BSK-M medium. We employed quantitative real-time PCR (qPCR) to detect Borrelia DNA in the ticks and to calculate the prevalence of B. miyamotoi and infections with other borreliae. For characterization of the isolated B. miyamotoi, we performed draft genome sequencing and multilocus sequencing analysis (MLSA). Results A total of 2656 adult I. persulcatus ticks were collected. The overall prevalence of relapsing fever (RF) borreliae in ticks was 5.0% (134/2656) and that of Lyme disease (LD) borreliae was 43.8% (1164/2656). Co-infection with RF and LD borreliae was observed in 63 ticks (2.4%). Ticks that were positive for RF borreliae by qPCR were subjected to glycerophosphodiester diester phosphodiesterase gene (glpQ) PCR amplification and sequencing, through which we identified the RF borrelia specimens as B. miyamotoi. Furthermore, the B. miyamotoi strain Hetao-1 was isolated from I. persulcatus, and a draft genome sequence was obtained from the isolate. Sequencing determined the strain Hetao-1 genome to be approximately 906.1 kbp in length (28.9% average GC content), and MLSA identified the strain as ST633, which has previously been reported in Japan and Mongolia. Conclusion We detected B. miyamotoi from I. persulcatus ticks collected in Inner Mongolia, and successfully isolated a B. miyamotoi strain. To our knowledge, this is the first study to culture a B. miyamotoi isolate from China. The data on the prevalence of B. miyamotoi and other borreliae in I. persulcatus ticks will be fundamental for future epidemiological studies of B. miyamotoi disease in Inner Mongolia.

Development and application of the technique for identification of Borrelia miyamotoi surface antigens

Introduction. Borrelia miyamotoi is a pathogen causing erythema-free ixodid tick-borne borreliosis (ITBB), a disease widespread in Russia. The genome of B. miyamotoi contains genes of multiple variable major proteins (Vmps). Vmps fall into two families — Vsps and Vlps (with subfamilies δ, γ, α and β). At a particular time, a single B. miyamotoi expresses only one variant of Vmp gene.The purpose of the work is to develop a technique for identification of the Vmp present at the expression site.Materials and methods. The technique is designed in the format of a real-time multiplex PCR. It was tested by using B. miyamotoi DNA samples extracted from blood collected from 172 ITBB patients and 109 ticks. The samples were collected in 14 regions of Russia.Results. The new technique made it possible to identify the expressed Vmp in 82% of the examined samples, thus having demonstrated its efficiency. Negative results were much less often observed with samples from patients than with samples from ticks. At the same time, the percentage of samples with one type of Vmp is identical for clinical samples and ticks, while the percentage of samples containing concurrently two types of Vmps is significantly higher among samples from patients with the most frequent occurrence of the Vlp-δ and Vsp combination.Discussion. The frequent occurrence of the combination of two Vmp types in the blood samples can indicate the concurrent presence of several subpopulations of B. miyamotoi in ITBB patients. A new antigenic Vmp variant is synthesized after protective antibodies have been produced for the major protein of the strain transmitted by a tick. This phenomenon known as immune evasion allows the pathogen to persist within a host.Conclusion. The developed technique of real-time multiplex PCR allows to simultaneous detect of several antigenic variants of the variable basic surface proteins of B. miyamotoi. The study of the antigenic spectrum of B. miyamotoi strains in comparison with the characteristics of conserved regions of the genome by the method of multilocus sequencing will clarify the stages of evolution and distribution of B. miyamotoi sensu lato.

Ticks on the turf: investigating the presence of ixodid ticks on and around football fields in Germany

Ixodes ricinus is the most abundant tick species and an important vector of pathogens in Germany and in large parts of Europe. A few other ixodid tick species, e.g., Dermacentor reticulatus, may also be of eco-epidemiological relevance. As ticks are not only found in natural but also in suburban areas (parks, gardens), the present study investigated whether ticks occur on and near football grounds thus posing a potential risk to players and visitors. Thirty-two football grounds from all 16 German federal states were selected, mainly situated adjacent to a green area (forest, park). Ticks were collected by the conventional flagging method in spring 2018, and nymphs and adults were counted and morphologically determined. Altogether 807 nymphal and adult ticks were collected from 29 football grounds: 714 I. ricinus, 64 Ixodes inopinatus, 2 Ixodes frontalis, 24 Ixodes sp. ticks, and 3 D. reticulatus. Ixodes inopinatus was found in 13 out of 16 German states. Three ticks were even found on the turf of two football fields. It can be concluded that ticks occur quite frequently and sometimes in high abundance near football grounds situated close or adjacent to a forest or a park.