FAUNA, MORPHOLOGY AND SYSTEMATICS OF PARASITES
Objective of research. A study of species composition of nematodes parasitizing the digestive system of domestic ruminants in Uzbekistan, analysis of seasonal dynamics of infestation and features of the nematode fauna in plains and foothill-mountain areas.
Material and methods. In the period of 2014–2015, the digestive tracts of 206 head of cattle and 878 head of small cattle from the farms of Tashkent, Syr-Darya, Dzhizak, Namangan, Kashkadarya, Surkhandarya, Navoiy and Bukhara regions as well as Karakalpakstan autonomous republic were investigated by the method of full helminthological dissection.
Results and discussion. 30 nematode species were found in digestive tracts of domestic ruminants of Uzbekistan; 28 nematode species in sheep, 26 species in goats, and 22 species in cattle. The highest rates of extensity of infection (EI) and intensity of infection (II) were detected in Ostertagia ostertagi, Marshallagia marshalli, Teladorsagia circumcincta and Parabronema skrjabini. An increase of EI and II in summer and autumn period was typical for the most of the discovered nematode species.
However, EI in O. ostertagi, T. circumcincta, Marshallagia spp. and P. skrjabini was relatively stable in all seasons, but II increased to autumn. 26 nematode species were registered in plains; only 3 of them with high EI and II.
28 nematode species were registered in foothill-mountain areas, 8 of them showed high rates of EI and II. 24 species of nematodes are common in both areas.
The obtained results allow us to conclude that it is necessary to improve helminth control of ruminants in Uzbekistan. Alternation of pastures and regulation of grazing, as well as preventive dehelmintization should not be ignored; observe the optimal requirements for feeding and management of animals.
Objective of research: The purpose of research is to study the fauna and ecology of helminths in cattle from Central Uzbekistan.
Material and methods. The cattle was investigated by the method of complete and incomplete helminthological autopsies. Cestodes, trematodes and nematodes were examined using the common methods. Larvae of trematodes and nematodes from dominant parasite species found in intermediate hosts (molluscs, insects, ants and dipterous) are also described in this paper.
Results and discussion: 32 helminth species were identified in cattle from the central region of Uzbekistan: 5 species belong to the class Cestoda, 6 species — to the class Trematoda and 21 species — to the class Nematoda. Based on the nature of helminth localization in the host body, three groups of parasite communities were detected.
Objective of research. The purpose of this research is to carry out morphological and molecular genetic identification and to determine phylogenetic relationship among Protostrongylidae species.
Materials and methods. Helminthological material was collected from domestic hollowhorned ruminants and land mollusks in different areas of Uzbekistan. The morphology of protostrongylids was studied using the methods of Boev (1975) and Anderson (1978). The firststage larvae were investigated by examination of fecal samples from animals taking into account remarkable morphological features of larvae without dorsal cuticular thorn at the tail point (for Protostrongylinae) and with thorn (for Muelleriinae, Varestrongylinae et al.] as well as length, tail form and body size of larvae. To study the morphology of the third-stage protostrongylid larvae, the feet of infected mollusks X. candaсharica were separated and placed into the artificial gastric juice. Nucleotide sequences ITS-2 regions of species Protostrongylus rufescens, P. shiozawai, Ortostrongylus macrotis, Cystocaulus ocreatus and Umingmakstrongylus pallikuukensis were used to study phylogenetic relations between protostrongylids. The phylogenetic analysis was conducted using the software Clustal X 2.0.
Results and discussion. Based on morphological and molecular examinations, five species of protostrongylid nematodes: Protostrongylus rufescens, P. hobmaieri, Protostrongylus sp., Spiculocaulus leuckarti and Cystocaulus ocreatus were found in hollow-horned ruminants. The morphological and molecular-genetic analysis of detected nematodes enables precise identification of species and spatial distribution of endemic protostrongylids.
ECOLOGY AND BIOLOGY OF PARASITES
Objective of research: To study the role of phoresy of the louse Mallophaga on the population of the louse-fly Hippoboscidae.
Material and methods: In 1997–2015 more than 10000 birds were caught; from them about 2000 louse-flies Hippoboscidae were collected and studied.
Results and discussion: 12 cases of phoresy of the louse Mallophaga on the louse-fly Hippoboscidae were recorded: 7 cases of phoresy of Columbicola columbae on pigeon louse flies Pseudolynchia canariensis collected from pigeons (Columba livia); one case — phoresy of the louse Ricinus rubeculae on the louse fly Orniyhomyia fringillina from the robin (Erithacus rubecula); one case — phoresy of the louse on the louse fly O. avicularia from the rook (Corvus frugilegus); one — phoresy of the louse on louse fly O. avicularia from the cuckoo (Cuculus canorus); one — phoresy of the louse on the louse fly Ornithoica turdi from the nightingale (Luscinia luscinia); one case — phoresy of the louse on the louse fly O. chloropus from Luscinia svecica.
EPIZOOTOLOGY, EPIDEMIOLOGY AND MONITORING OF PARASITIC DISEASES
Objective of research: The target of the paper is to develop a model of GIS-based risk monitoring of zoonotic cestodiasis in human.
Materials and methods: The use of geographic information systems (GIS) as an epizootiological and epidemiological method for the risk-based monitoring of human cestodiasis enables the development of a multi-level platform for solution of a wide range of tasks related to the control of this disease. The modern GIS tools use the methods of geoinformatics applying powerful software and hardware: open access geographic web servers, tools for multidimensional complex analysis, creating most accurate electronic and paper maps. Full-featured GIS contain a full set for processing geospatial data including acquisition of data, its integration and storage, automatic data processing, editing, creation and maintenance of topology, spatial analysis, access to the database management system (DBMS), visualization and creation of hard copies of any cartographic data.
Results and discussion: The use of GIS enables to study more closely the regularities of epizootic process, geography of human cestodiasis and to improve the methodology both for short-term and long-term retrospective epizootiological analyses.
Objective of research: To study the possibility of using temperature-based models for prevention of dirofilariasis.
Materials and methods. For mathematical modeling of dirofilariasis we use the HDUstemperature model based on the impact of the average daily temperature on the rate of development of Dirofilaria larvae in mosquitoes.
The amount of 130 DDU (Dirofilaria development units) accumulated in the period no more than 30 days at average daily temperature more than 14◦C is required for the development of Dirofilaria up to the infective stage.
Daily data on average air temperature in Rostov-on-Don (1996 — 2012), Veliky Novgorod (2008 — 2012), Anapa (2008 -2012) and Astrakhan (2008 -2012) were used for the calculation.
Results and discussion. The results of temperature simulation revealed that the DDU model is a low-priority forecasting model for canine dirofilariasis (Pearson’s correlation coefficient minus 0.45). The model considers only the average daily temperature and does not consider other factors affecting the incidence rates.
The epidemic season of dirofilariasis in Rostov-on-Don in 1999 — 2012 differed depending on average daily temperatures. Therefore, the earliest date of the transmission of dirofilariasis in that period fell on the 12th of May, 2012; the latest date — on 29th of June, 2001. The optimal time for prevention of canine dirofilariasis has been defined (in Rostov-on-Don, Anapa and Astrakhan microfilaricides should be given to dogs from May 15 to November 15; in Veliky Novgorod — from June 15 to August 31).
Thus, DDU-models (Dirofilaria Development Units) can be used to set time limits for epidemiological season of dirofilariasis and preventive treatment of dogs against dirofilaria.
Objective of research: To study the epizootiology of trematodoses of cattle from dairy cattle farms in the Vologda region; to estimate the anthelmintic efficacy of Fascocide, Helmicide, Oxiclozanide, Albendazole, Fezole, Alben.
Materials and methods: A preliminary analysis of veterinary reports of the Department of Veterinary Medicine, the regional Stations on Fight against Diseases in Animals, meat-processing plants and slaughter units, Laboratories of Veterinary and Sanitary Expertise was carried out for the period 2005–2009 according to the Forms «1-Vet» and «5-Vet». Coproovoscopic examinations of cattle were conducted monthly to determine seasonal and age dynamics of infection. Biological features of intermediate hosts were studied by standard methods. Fascocide at the dose of 10 mg a.i./kg, Helmicide — 7,5 g/100 kg, Oxiclozanide — 5,25 mg/kg, Albendazole — 15 mg a.i./kg and Alben — 10 mg a.i./kg were used against fasciolosis and paramphistomosis in cattle under production conditions in dairy farms and complexes of Vologda region.
Results and discussion: The rates of trematode infection in various climatic and geographic zones of the region are different. The highest level of infection was registered in the North Eastern and South Eastern zones, and the lowest — in the South Western.
Adult cattle are infected with Fasciola hepatica, Paramhistomum cervi and P. ichikawai all year round; the maximum peak of infection is registered in winter and spring seasons. First signs of trematode infection in calves of the first grazing season are observed at the beginning of the pasture season, which is the evidence for the overwintering of helminth larvae in intermediate hosts. Trematode infections manifest commonly as mixed infections.
First individuals of Lymnaea truncatula and Planorbis planorbis appear in biotopes in the second decade of May, reaching the maximum number in August. Their infection with trematode parthenites is observed in the period from the first half of June to October; the maximum peak of infection — in August. The most effective preparations for dehelmintization against Fasciola spp. and Paramhistomum spp. are Fascocide, Helmicide and Fezole.
Objective of research. To study the species composition of the parasite fauna and the infestation rate of the spiny loach Cobitis taenia Linnaeus, 1758 from Saratov Reservoir.
Materials and methods. The material was collected in the Mordovo-Kol’tsovsky area of the Reservoir (the middle part of the reservoir) in 2009 and 2014–2015. Fishing was carried out by a hydrobiological net. Totally 47 individuals of spiny loach were investigated by the method of full parasitological autopsy. Collection, fixation and office studies were carried out using the standard technique; diagnostics of species composition of parasites — by identification guides. To estimate the rate of fish invasion, we used the common parasitological indices: extensity of invasion, intensity of invasion and the index of abundance of parasites.
Results and discussion. 7 species of parasites belonging to 5 classes: Monogenea — 1, Cestoda — 1, Trematoda — 3, Nematoda — 1, Bivalvia — 1 were found in the spiny loach Cobitis taenia Linnaeus from Saratov Reservoir.
The species composition of parasites and the level of host infestation directly depend on the lifestyle and nutrition of fishes. Eating the infected benthic and weed bed invertebrates (oligochaetes, larvae and adult insects), the spiny loach becomes infected with 3 species of helminths.
4 species of parasites infect the host using the active infestation way. Most of the parasites found in fish from different families and groups are wide specific. The parasite fauna includes 1 species, narrow specific for the spiny loach (monogenea Gyrodactylus latus Bychowsky, 1933) and 1 species specific for loaches (fam. Cobitidae) — metacercariae of Holostephanus cobitis Opravilova, 1968. For 4 species of parasites, the spiny loach is the final (definitive) host, for 3 — intermediate. The dominant species in the composition of the parasite fauna C. taenia is the trematode H. cobitidis (mtc.); the extensity of infection with it reaches 97,9%, the abundance index is 10,8 ind.
Fluke larvae infect fishes (using the active way) penetrating through the skin. The most pathogenic for spiny loach parasites are the monogenea G. latus, metacercariae of trematodes H. cobitidis and Paracoenogonimus ovatus Katsurada, 1914, the larva of the nematode Raphidascaris acus Bloch, 1779.
Objective of research: To study the infection rate with B. sulcata in various age groups of cattle in summer at different management technologies in the Kurgan region.
Materials and methods. Samples were collected from animals kept under different management technologies (private sector, stabling&pasture, stabling) in the Kurgan region in July 2016. Samples weighing 10 g were taken from animals of different age groups from the rectum or by usual fecal gathering in animal premises. The collected material was preserved with 2% potassium dichromate. At the Laboratory for Protozoology and Sanitary Parasitology of the All-Russian Scientific Research Institute of Fundamental and Applied Parasitology of Animals and Plants named after K.I. Skryabin, 68 samples were examined under the microscope by methods of successive washings and formalinether sedimentation; cyst counting was performed in a counting chamber Mc Master.
Results and discussion. The results of random coproscopic examination of cattle of different age groups in farms of the Kurgan region in summer period revealed their infection with Buxtonella sulcata. Extensity of infection in the private sector ranged from 33% to 50%, in farms with stable management — from 10 to 62,5%, stable and pasture management — from 0 to 85,7%.
PATHOGENEZIS, PATHOLOGY AND ECONOMIC DAMAGE
Intensive development of the parasitic diseases immunology in the past decade allows making a definite conclusion about the features of the immune response formation in helminthiasis and its key problems, such as short duration, low-efficiency and the ability to cause the development of immunopathological processes.
Objective of research: The aim of this review was to identify most significant stages of helminthiasis immunogenesis wich can cause its low protective ability, and for which further comprehensive study of modern molecular biologists, geneticists, biochemists, immunologists and parasitologists should be held to clarify the immunopathology induction mechanisms and improve methods of prophylaxis, diagnosis and treatment of these diseases.
Results and discussion. Studies have revealed that the main condition for relationship building in the system “parasite-host” is the presence of protection mechanisms in helminths against exposure to the host’s immune system and immunomodulation mechanisms up to the complete immunosuppression in host.
Products of helminths vital activity, so-called secretory-excretory products (SEPs), as well as changed in the process of pathogenesis host proteins and cells become a powerful immune stimulus and activate mechanisms for general and local immunity.
The following defense mechanism in helminthiasis is the most effective: participation of IgE antibodies class and IgG2 subclass which play a major role in the activation of cell adhesive activity; influence of cytotoxic T cells (T killer cells, CD8+ -cells); involvement of macrophages, activated with T-cells; work of induced effector cells (eosinophils, neutrophils, mast cells, platelets, etc.), and the results of natural killer (NK) and regulating Treg- lymphocyte population (CD4 + CD25 + -cells) activity.
It was shown that helminthiasis is accompanied by oxidative stress, which is characterized by decreased activity of catalase, superoxide dismutase, and an increase of lipid peroxidation products, which may cause primary DNA damage underlying in gene and chromosomal mutations that is shown in prior studies.
Parasites metabolites have a cytotoxic effect on somatic, generative and immune cells of host, causing the increase of apoptotic cells among them.
Theoretical significance of the data on identification problems of helminthiases immunology is undoubted. Its practical implementation offers the significant increase in the effectiveness of helminthiasis prevention and control.
BIOCHEMISTRY, BIOTECHNOLOGY AND DIAGNOSTICS
Objective of research: To study the activities and properties of nucleoside- diphosphatase (NDPase) in cestode Bothriocephalus scorpii.
Materials and methods: Cestodes were homogenized with 10 vol. of extraction medium. NDPase was detected in mitochondria and microsomes with substrates (IDP, GDP, UDP). Inorganic phosphorus was determined by the method of Kochetov (1980). The effects of 10 anthelmintic drugs on the activity of NDPase were studied.
Results and discussion: It was found that the mitochondrial and microsomal fractions of cestodes B. scorpii have nucleoside diphosphatase activity.
The activity of nucleoside diphosphatase depends on substrates and Mg2+ ions. The impact of various effectors and ions (Ca2+, Mn2+, Zn2+) on enzyme activity was determined.
Effects of 10 anthelmintic drugs on activity of nucleoside diphosphatase were studied. The anthelmintics Bitionol and Trichlorophen have been proved effective.
TREATMENT AND PREVENTION
Objective of research: Determination of the efficiency of 5% concentration Diminazene aceturate at the dose of 0.25 mg/kg body in combination with polyethylene glycol solution applied against piroplasmidosis in cattle.
Materials and methods: Trial of the new method of prolonged chemoprevention was carried out in one of the farms of Dagestan, which is unfavorable for piroplasmidosis. Aceturate contained in this drug rapidly inhibits DNA in cells of blood parasites, which leads to their death within several hours. Diminazene aceturate proved to be effective against piroplasmidosis, babesiosis, taleriosis, hemosporidosis of animals. The experiment was conducted on 30 head of cattle divided into 3 groups 10 head in each. Animals of two groups received the preparation and prolongator at the dose of 0,25 mg/kg in the 15% aqueous solutions of polyethylene glycols (PEG), and at the dose of 0,25 mg/kg in the 20% aqueous solutions; the third group served as a control (received the preparation without prolongator). Efficacy of the method was estimated by periodic examination of blood smears from experimental animals for plasmodia in erythrocytes and synchronous measurement of body temperature of animals.
Results and discussion: It was found that the method of prolonged chemoprophylaxis applied against piroplasmidosis in animals (at the dose of 5 ml per 100 kg body weight, 6 times every 25 days) enables disease prevention during the whole disease season. Such method allows reducing the number of treatments from 24 to 12-13 times in a season. In addition, data on the distribution of hemosporidia infections among the cattle in the Republic of Dagestan were clarified.
Purpose of the research — to conduct analysis of the status and perspective of application of prolonged forms of anthelmintic drugs in veterinary.
Materials and methods. On the topic of development and application of prolonged forms of anthelmintic in veterinary was conducted the analysis of 27 literature sources including 23 foreign authors. Were analyzed the data about kinetics of avermectins and praziquantel in the animals body and comparative results about dependence and correlation of pharmacokinetic parameters with their anthelmintics efficiency.
The results and discussion. Prolonged forms of avermectins which prevent the infections for long period are developing to prevent infection of animals during grazing season. Attempts to create prolonged dosage forms of praziquantel to prevent the Schistosoma japonicum infections are currently in progress. Was shown the possibility of obtaining polymeric microspheres, which contains ivermectins in the form of implants. As solvents are used N-methylpyrrolidone, triacetin, benzyl benzoate etc. Mixes of composite based on sucrose, acetate isobutyrate and polylactic acid is offered for prolongation of the action of ivermectin, which is provides release of ivermectin within 80 days. Were described prolonged forms of praziquantel as the implants on a base of polycaprolactone and it’s mix with polyethylene glycol, which is obtained by extrusion of a melt of a mixture of praziquantel and polymers.
PHARMACOLOGY, TOXICOLOGY
Objective of research: To evaluate the tolerability of the drug, “Inspector Collar” based on Fipronil, Pyriproxyfen and Ivermectin in dogs and cats applied at a therapeutic dose and double increased therapeutic doses.
Materials and methods: Research was conducted at the shelter for dogs and cats «Bano Eco» (Moscow). Effect of drug “Inspector Collar” was studied on 15 clinically healthy outbreed dogs between 2 and 6 years with the body mass of 10–14 kg and 15 clinically healthy outbreed cats between 2 and 4 years with the body mass of 2–3,5 kg, receiving complete and balanced nutrition. Animals were divided into six groups (three groups of dogs and three of cats) five animals in each group. Dogs and cats from the first experimental group had put on one collar (equivalent of single therapeutic dose) for 45 days. The second experimental group had put two collars (double therapeutic dose) for 45 days. Animals from the third (control) group did not carry collars. During the experiment, daily supervision of animals was conducted; their general health status, behavior, appetite were observed, body mass and temperature estimated. Before and 15 and 30 days after the beginning of drug application, blood and urine tests were performed to study morphological and biochemical parameters. Statistical processing of data were carried out using Student’s t-test.
Results and discussion: The research results showed that the drug “Inspector Collar” is well tolerated even at double therapeutic dose. All observed indicators remained within the physiological norm.
Objective of research: Development of methods for the determination of fenbendazole and its metabolites in milk by liquid chromatography coupled with tandem mass spectrometry.
Materials and methods: Fenbendazole was administered orally to five cows. Samples of milk were taken on 1, 3, 5 and 10 days of drug application. The research method includes a description of reagents, plates and equipment; mass-spectrometric conditions for analysis of fenbendazole and its metabolites; preparation of the equipment to operation; preparation of eluent solution; preparation of the chromatograph to analysis; determination of chromatographic parameters of standard drug samples; preparation of milk samples to analysis; establishment of parameters of extracts’ chromatography; procedure of calibration of fenbendazole and its metabolites in eluent.
Results and discussion: When studying the pharmacokinetics of fenbendazole and its metabolites (sulfone and sulfoxide) in milk, it was found that the maximal concentrations were determined 24 h after drug administration and were 22,6 ng/ml for fenbendazole sulfone, 34,0 ng/ml for fenbendazole sulfoxide and 19,7 ng/ml for fenbendazole. 10 days after treatment, the concentrations of fenbendazole and its metabolites in milk did not exceed permitted values.
Objective of research: To estimate the effect of new combination drugs Aversect Forte and Aversect Combi on rats.
Materials and methods: The present experiment for the evaluation of eventual embyotoxic and teratogenic properties of drugs Aversect Forte and Aversect Combi was conducted on 32 female and 8 male rats based on the current methodological guidelines.
Pregnant female rats were divided into 2 experimental and 2 control groups.
Drugs were applied between the 7th and the 14th day of pregnancy taking into account the highest sensitivity of embryos towards different types of influence within this time period.
Aversect Forte and Aversect Combi were injected to pregnant rats subcutaneously at a dose of 0,3 mg a.i./kg of individual body weight. Rats from the control group received a forming mixture in comparable volume. Within the whole period of pregnancy, we observed the general clinical condition of female rats.
Results and discussion: Effects of new domestic preparations Aversect Forte and Aversect Combi on the antenatal development of the rat via subcutaneous injection of preparations to female rats at a dose of 0,3 mg a.i./kg between the 7th and the 14th day of pregnancy were estimated. The test preparations did not cause any external and internal developmental anomalies; indicators of embryo death, mass and dimensions of embryos were at the actual level of control and physiological parameters for that type of animals.
PARASITES OF PLANTS
Objective of research. To elaborate a framework for integrated protection of agricultural crops from oat, sugar beet, Golden potato, alfalfa and clover nematodes.
Materials and methods. The effectiveness of different anti-nematode measures was studied in 1991-2016 in Vinnyza, Kiev, Sumy, Chernigov and other regions of Ukraine. As research material served samples of soil, plants; eggs, larvae, imago; beet, oat, Golden potato, alfalfa and clover cyst nematodes. Preparation of temporary and permanent slides, determination of the species composition of cyst nematodes were carried out by standard methods [3, 6].
Results and discussion. The framework of integrated protection of agricultural crops from oat, sugar beet, Golden potato, alfalfa and clover nematodes has been elaborated. To prevent mass reproduction of cyst-forming nematodes, the saturation of crop rotations with host plants must not exceed 40% — with cereal grains; 20% — beet and oilseed Brassica (canola, radish, and mustard); 15% — potatoes; 20% — perennial legumes. Involvement of crops and green manure in the by-products cycle has a positive effect on the level of soil fertility and activation of life activity of natural antagonists of Heteroderidae.
Objective of research: to study the mechanisms of induced tomato plant resistance to rootknot nematode Meloidogyne incognita and potato to cyst nematode Globodera rostochiensis.
Materials and methods: The biogenic elicitors — chitosan and signal molecules — SA, JA for the modulation of immune plant responses were used. In experiment 1, tubers of potato cultivars Istrinskii (PCN-susceptibility) and Krinitsa (PCN-resistant), were treated with aqueous solutions of the immunomodulators at the specified concentrations. A low molecular weight soluble chitosan and acetylation degree of 15% and signal molecule — SA was used as an elicitor. In experiment 2, system tomato M. incognia was studied. Water solutions of chitosan, signal molecules: SA, JA were used for treatment of tomato seeds for 2 h and then the seeds were planted in sterile soil. The control plants were treated with water. Cultivation of plants was carried out by the standard technique. Plants were maintained in a greenhouse long enough for the nematodes to complete their life cycle. Development of nematodes in the processed plants estimated on morphophysiological and population characteristics. Biochemical indicators of roots and leaves of tomatoes estimated for 14 days after infection of plants. Previously identified major biochemical indicators of the plants in the genome that contain genes that determine the resistance of plants. The effects of biogenic elicitors on plant resistance were also evaluated by some metabolic changes related to natural plant resistance to tomato and potato to plant nematodes. These indicators were studied in clarifying mechanisms of induced resistance.
Results and discussion: Biogenic elicitors induce systemic resistance of plants to plant parasitic nematodes — Meloidogyne incognita and Globodera rostochiensis (decrease in the parasitic invasion of the roots; an inhibition of the vital activity of the parasite; a decrease in fertility and the amount of agents sources (larvae and eggs) capable of infecting the plants). The addition of signal molecules (salicylic and jasmonic acid) to elicitors increased their activity as immunomodulators. In present investigation, the mechanisms of induced plant resistance nematode were studied. The data obtained suggest that the mechanisms natural and induced by biogenic elicitors tomato resistance to the nematode have the same origin. These features meet all requirements of the new generation of methods of plant protection and the use of biogenic elicitors to raise plant resistance to parasitic nematodes may be promising.
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