Nghiên cứu nhiễm giun tròn đường tiêu hóa, bệnh giun lươn (strongyloidosis) trên lợn tại tỉnh bắc giang và biện pháp phòng trị tt tiếng anh

MINISTRY OF EDUCATION AND TRAINING THAI NGUYEN UNIVERSITY PhD CANDIDATE NGUYEN THI HUONG GIANG The thesis title STUDY ON THE PREVALENCE OF GASTROINTESTINAL NEMATODE INFECTION AND STRONGYLOIDIASIS ON PIGS IN BAC GIANG PROVINCE, THE PREVENTIVE MEASURES AND TREATMENT OF THE DISEASE Speciality: Veterinary parasitology and microbiology Code: 9.64.01.04 DISSERTATION IN VETERINARY MEDICINE Thai Nguyen - 2020 The dissertation has been completed at: AT COLLEGE OF AGRICULTURE AND FORESTRY THAI NGUYEN UNIVERSITY Supervisers: Professor Doctor Nguyen Thi Kim Lan Doctor Nguyen Van Quang Reviewer1: Reviewer2 Reviewer3: The dissertation will be defended at the Dissertation committee in National level COLLEGE OF AGRICULTURE AND FORESTRY - TNU Time date month year 2020 The dissertation can be found at - Viet Nam national library: - Learning Resource Centre in Thai Nguyen university - Library of Thai Nguyen university of Agriculture and Forestry 1 INTRODUCTION 1.1 The urgency of the project Bac Giang is a province with a quite development of pig husbandry For many years, pig production has made an important contribution to hunger eradication, poverty reduction and enrichment for farmers in the province According to the Department of Animal Science and Veterinary Medicine of Bac Giang province, there were 1,043,749 pigs in 2017 in the whole province, in 2018 there were 1,105,291 pigs Along with the rapid increase in the number of pigs, farmers have applied scientific and technical advances in pig farrming practice, thus bringing significant economic benefits to them However, pig farming in Bac Giang province has faced with many difficulties, including disease problems In addition to common infectious diseases, gastrointestinal nematode infections in pigs ofen occur chronically Although nematode infections not cause high mortality of pigs as infectious diseases, nematode infections usually occur in chronic form, making pigs stunted, growing slowly, reducing resistance of their bodies, and susceptible to other diseases Phan Dich Lan et al (2005), Pham Sy Lang et al (2011) reported that diseases caused by gastrointestinal nematodes were very common diseases and was one of reasons resulting in stunted pigs, reducing weight gain by 15-20%, and were susceptible to infectious diseases such as rotavirus infection, paratyphoid leading to more severe diarrhea syndrome Although there is quite development of pig farming in Bac Giang province, there have been no studies on the situation of nematode infections and Strongyloidosis in pigs, so there are no measures for effective prevention and treatment of these diseases From the urgent requirements of animal farming practice and disease prevention for pigs, we conducted the entitled: "Study on the prevalence of gastrointestinal nematode infection and Strongyloidiasis on pigs in Bac Giang province, the preventive measures and treatment of the disease" Objective of the project - Determining prevalence and intensity of infection of gastrointestinal nematodes in pigs; - Identification of some epidemiological, pathological and clinical characteristics, the prevention and control measures of Strongyloidosis in pigs in Bac Giang province Scientific and practical significance of the topic 3.1 Scientific significance The project provides scientific information about the situation of gastrointestinal nematode infection in pigs in Bac Giang province; Epidemiological, pathological and clinical characteristics of Strongyloidosis, on the basis of which, set up a process of prevention and treatment of strongyloidosis for pigs with high efficiency 3.2 Practical significance The results from the project are the basis for recommending the pig producers applying preventive measures and treatment of nematode infection in general, and Strongyloidosis in particular in order to limit the prevalence of nematode infection in pigs and limit damage caused by Strongyloidosis, making contribution to sustainable development of pig farming in Bac Giang province New contributions of the project - The project is the first systematic study on the situation of gastrointestinal nematode infection in pigs in Bac Giang province including ; Epidemiological, pathological and clinical characteristics of pig threadworm infection ( strongyloidosis) - Effective procedures for the prevention and treatment of Strongyloidosis in pigs have been designed, recommended and applied widely to farms and pig farming households in Bac Giang province and the neighboring provinces The dissertation structure The dissertation consists of 129 pages (excluding references list): introduction 03 pages; overview of literatures 31 pages; objects, materials, contents and methods of the study 22 pages; study results and discussion 72 pages; Conclusions and recommendations 02 pages In the disertation, there are 36 tables, 13 charts and graphs, 120 colour pictures depicting the results of the project PhD student has referred to 151 references (including 68 published in recent years ( past five years) Chapter DOCUMENT OVERVIEW Nguyen Thi Le et al (1996) indicated that nematodes in pigs are distributed all over the world In particular, in the tropic and subtropic regions there is hot and humid climate, which is a very favourable condition for eggs and nematode larvae to develop in the outside, making the gastrointestinal nematode infection in pigs occur through out the years According to Pham Sy Lang et al (2011), Gomathi M et al (2016), pigs infected with threadworms though gastrointestinal tract and though skin, piglets can be infected with S ransomi very early from the colostrum so it is possible to see adult worms from days old piglets Pigs infected with strongyloides are characterized by diarrhea, loose stools with blood due to hemorrhagic intestine, anemia, emaciation and sudden death In addition, when the larvae enter the alveoli, they can cause pneumonia with fever at temperature of 40 - 41.5oC, coughing a lot, difficulty breathing According to Nguyen Thi Kim Lan (2011), In order to prevent Strongyloidosis in pigs effectively, it is necessary to implement integrated prevention measures, including: hygiene of pig pens, cleaning livestock instruments and equipments; Pig manure and waste must be collected daily and properly composted at the stipulated places; implement disinfection of pig pens regularly, not keep pigs at different age together, using a reasonable diet for pigs According to Bui Thi Tho et al (2015), Nguyen Tai Nang et al (2016), there are many new anthelmintic drugs that have a good activities for dewornming nematodes, including drugs of ivermectin , benzimidazole, imidazothiazole groups Chapter OBJECTS, MATERIALS, CONTENTS AND METHODS OF STUDY 2.1 Object, time period and places of study 2.1.1 objects of study - Pigs reared in various locations of Bac Giang province - Gastrointestinal nematode species of pigs - Pigs affected by Strongyloidosis (caused by Strongyloides spp ) 2.1.2 Time period and place of study * Study period: from 2016 - 2019 * Study places - The project was implemented in districts of Bac Giang province including: Viet Yen, Hiep Hoa, Lang Giang, Yen Dung and Son Dong - Sample testing places: Laboratory of Faculty of Animal Science, and Veterinary Medicine - Bac Giang Agriculture and Forestry University; Bac Giang General Hospital and Preventive Medicine Center of Province; Institute of Ecology and Biological Resources; National Institute of Hygiene and Epidemiology 2.2 Study materials: *Experimental animals: pigs raised in districts of Bac Giang province * Types of samples for studying: fresh feces samples from pigs of different age; samples from sediment of pig house floor, wastewater samplesfrom pig pens, samples from surface soil in garden for growing feed crops; samples from nematodes collected from infected pigs; blood samples from pigs infected with Strongyloides spp and healthy pigs; samples collected from small intestines, heart, lungs, liver, kidneys of pigs affected by Strongyloidosis * Instruments, equipments and chemicals: light microscope, olympus CX221 microscope, scanning electron microscope; stool test kits; blood microsampling device; Hematology Analyzer.Erma PCE 210 and TC - Matrix; Microtome Equipment, automatic DNA sequencing machine; ABI Prism 3130 Genetic Analyzer; centrifuge machine, micropipetes, electrophoresis machine,UV Transilluminator, PCR machine, DNeasy Tissue Kit (QIAgen) Chemicals used for preparing smear slide from specimens, saturated brine solution; Barbagallo solution; solution for drawing water from nematode bodies; QIAquick PCR purification kit Disinfectants: P.V.D iodin 10%; Chloramin B; Good Farm L anthelmintic drugs for deworming Strongyloides in pigs: including fenbendazole, ivermectin and thiabendazole 2.3 Contents of study 2.3.1 Study on prevalence and intensity of gastrointestinal nematode infection on pigs in Bac Giang province - Investigation of situation of gastrointestinal nematode prevention for pigs in Bac Giang - Identification of gastrointestinal nematode species and their distribution in Bac Giang - Dissection of pigs to determine: + The prevalence and intensity of gastrointestinal nematode infection in locations +The prevalence and intensity of infection of various gastrointestinal nematode species in pigs - Stool examination to determine: + The prevalence of gastrointestinal nematode species infection in various locations + The prevalence and intensity of gastrointestinal nematode species infection in pigs through faecal examination + The prevalence of gastrointestinal nematode infection depending pig age, Farming methods and seasons 2.3.2 Study on Strongyloidiasis in pigs (Swine Strongyloidosis) * Identify the species of Strongyloides in pigs - Results from detection and collection of Strongyloides in pigs in Bac Giang through dissection - Results from designation of Strongyloides by using morphological technique, and by molecular biology engineering * Studying Strongyloidiasis infection through stool analysis - The prevalence and intensity of Strongyloides infection depending on locations - The prevalence and intensity of Strongyloides infection depending on age of pig, season, and farrming method * Study on contamination of strongyloides eggs and larvae in the environment * Study on pathological and clinical characteristics of Strongyloidosis in experimental infected pigs and naturally infected pigs * Study on preventive measures for strongyloidosis in pigs -Testing and determination of anthelmintic drugs fo deworming Strongyloides with high efficacy and safe for pigs - Study and recommendation of procesdures for prevention and treatment of Strongyloidosis in pigs 2.4 Methods of study 2.4.1 Study on prevalence and intensity of gastrointestinal nematodes infection on pigs in Bac Giang 2.4.1.1 Investigation of actual situation of prevention and control of gastrointestinal nematode infection in pigs in Bac Giang province: Prevention and control of gastrointestinal nematode infection in pigs in Bac Giang province was carried out by direct observation, using questionaire for pig farminghouseholds and investigation results were filling out the paper form 2.4.1.2.Method of study on prevalence and intensity of gastrointestinal nematodes infection in pigs - Method of descriptive epidemiology as discribed by (Nguyễn Nhu Thanh et all, 2001) was applied; - Minimum sample size was calculated using Win episcope 2.0 software; - Samples were collected using method of.Stratified sampling; - Pigs were dissected using methods of incomplete helminthological dissection for gastrointestinal organs as described by Skrjabin (1928); Samples of parasitic nematodes in the gastrointestinal tract of pigs were collected then, the nematode smear slide were fixed using method as discribed by De Grisse A T (1969) Species designation was made according to species identification key of Phan The Viet et al (1977), De Ley P and Blaxter M (2004); - Nematode eggs in pigs were detected by using lleborn's flotation technique The intensity of nematode infection was determined by using Mc Master egg counting technique for counting eggs per gram of faeces in Mc master counting chamber (According to Hansen J and Perry P., 1994) 2.4.2 Method of study on Strongloidosis in pigs 2.4.2.1 Species identification made by using morphological method Faecal smears of pigs infected with Strongyloides were fixed by using method of De Grisse A T (1969), the morphology of Strongyloides was observed under the light microscope, along with plotting to identify the species according to the identification key of Phan The Viet et al (1977), De Ley P and Blaxter M (2004) At the same time, the superstructural morphology of Strongyloides spp was observed based on the method of Sato H et al (2008) 2.4.2.2.Molecular technique for identification of Strongyloides - PCR technique was used for testing samples of Strongyloides collected from studied districts of Bac Giang province The obtained DNA sequences compared with the DNA sequences on the gene bank using MEGA software version 6.0 (Tamura K et al., 2013), the phylogenetic tree analysis and drawing were performed by using the Maximum Likelihood (ML) method with the most appropriate model The group confidence interval was assessed by bootstrap values with 1000 replicate samples 2.4.2.3.Methods for assessing the environment contaminated with Strongyloides eggs and larvae Soil samples for Strongyloides eggs were examined using techique discribed by Romanenko N A (1968) Strongyloides larvae in soil samples were determined by using Baerman nmethod for larvae isolation * Methods of identifying duration of Strongyloides eggs and larvae development and survival in faeces of pigs in laboratory experimental groups were designed in seasons: Spring, Summer, Autumn and Winter Faeces were collected from pigs severe infected with Strongyloides A plastic pot in 15- 20 cm diameter was used for each experiment , the start of the experiment was labeled and dated The samples were placed at normal temperature and humidity On the first day, every hours, samples were taken for testing by using Fulleborn flotation technique and Bearmann method for larvae isolation to identify the duration time when Strongyloides eggs hatch to larvae Then, about 3-5 grams of faeces were taken daily for testing using Bearmann method for larvae isolation to identify the time duration for larvae to develop into infective larvae Infective larvae were determined by observing the morphology of larvae under light microscope to recognize infective larvae based on morphology as described by Viney M E and Lok J B (2015) 2.4.2.4 Establishing experimental infection of pigs for studying Strongyloidosis experimental groups were designed by using healthy crossbred pigs (♂ Yorkshire x ♀ Mong Cai) at - months of age Pigs were divided into phases: Phase1: experimental infection was caused in order to study the pathological characteristics of the disease; experimental infection was used for testing efficacy of specific antheminthic drugs for treatment of Strongyloidosis: In phase 1: 15 pigs were used and divided into groups, group1, experimentally gastrointestinal infection was caused ; Group 2: skin infection was caused ;and group for control also set up The infecting dose was at 10,000 larvae phase was designed for experimental Strongyloides infection of 20 pigs which were divided into groups pigs for each Each pig was ingested 10,000 infective larvae 2.4.2.5 Study on pathological and clinical characteristics of Strongyloidosis in experimentally infected pigs and naturally infected pigs in the field * Study on infected pigs - Method of assessing clinical symptoms: the symptoms of infected pigs were observed, compared to pigs in the control group Hematological indices were determined using Erma PCE - 210 and TC Matrix hematology analyzers Macropic lesions score were Identified by dissection of pigs experimentally infected with Strongyloides, observing with the naked eye and magnifying glass lesions in liver, lungs and gastrointestinal tract Study on microscopic lesions score using preparation of slides from paraffin embedded tissues stained with Haematoxilin – Eosin, The slides were observed under a microscope to identify microscopic lesions * Methods of study on Strongyloidosis in pigs in the locations Methods of identifying clinical symptoms, signs and macropic lesions score in pigs infected with Strongyloides in the field were performed by using the same methods as conducted in experimentally infected pigs 2.4.2.6 Experimental design for testing efficacy of some selected anthelminthic drug against Strongyloides in pigs Efficacy and safety of 03 anthelminthic drugs including fenbendazole at dosage of mg/kg B.W, thiabendazole at dosage of 6.5 mg/kg B.W, ivermectin, at dosage of 0.3 mg/kg B.W and experimental study on preventive measures for the diseases were conducted by using - group experimental design, comparing them by testing for differences between them 2.4.5.Method for analysis of Experimental data Experimental data were analyzed by using method of biological statistics (according to documents of Do Duc Luc et al 2017), on Microsorf Office software, Excel 2010 program and minitab software version 16.0 Chapter RESULTS AND DISCUSSION 3.1 Study on the prevalene and intensity of gastrointestinal nematode infection in pigs in Bac Giang 3.1.1 Situation of prevention of gastrointestinal nematodiasis for pigs in Bac Giang province The findings are shown in table 3.1 : Many of 950 investigated households in districts of Bac Giang province had applied measures for prevention and control of nematodiasis 12 The results are shown in table 3.7, pigs of all ages were infected with gastrointestinal nematodes Pigs under months of age were infected with the highest prevalence (78.84%), followed by pigs from - months old (69.71%), pigs from months old (55.82%), the lowest prevalence of infection was in pigs over months of age (27.23%) Thus, the infection prevalence of gastrointestinal nematodes in pigs decreased gradually with the pig's age 3.1.3.6.The prevalence of gastrointestinal nematode infection in pigs depending on farming methods The results are shown in Table 3.8: Pigs raised by traditional farming methods, Infection prevalence of nematodes was the highest (85.59%); followed by prevalence of infection in semi industrial pig farming (72.35%); In industrial pig farming prevalence of infection was the lowest (28.64%) (P