Single and Low-level Oocyst Infections of Drug-resistant Field Strains of Eimeria tenella in Medicated Birds

Five out of ten birds infected with a single oocyst of strain Gt2 of Eimeria tenella and medicated with the recommended level of robenidine were found positive in the first experiment and four in the second in comparison with seven and six respectively in nonmedicated birds. Six birds out of ten were found positive in the two groups of similarly medicated birds infected with two or four oocysts each. Although single oocyst infections of strain Lilly 155 were unsuccessful, six out of ten birds were found positive in birds infected with ten oocysts each.All single and low-level oocyst infections were accomplished with oocysts previously treated with beta-glucuronidase and broken into sporocysts prior to infections.The overall results suggested that when a coccidium became resistant to an anticoccidial drug, only one or a few occysts were needed to start an infection if the drug was continued. The results also showed that, perhaps, successful single or low-level oocyst infections can also be used as a criterion for demonstrating drug resistance in coccidia.     

Isolation, propagation, identification and comparative pathogenicity of five Egyptian field strains of Eimeria tenella from broiler chickens in five different provinces in Egypt

A battery trial was conducted to evaluate the comparative pathogenicity of five field strains of Eimeria tenella from Behera, Khafr El-Sheikh, Alexandria, Gharbia and Matrouh provinces in Egypt. Two-week-old broiler chickens were infected with 25×10(3) sporulated oocysts of each strain of E. tenella. The comparative pathogenicity of the strains was assessed by calculating body weight gain, feed conversion ratio, lesion scores, dropping nature scores, cecal scrapings, mortality percentage and oocysts count. Hematological parameters including hemoglobin (Hb) content, packed cell volume (PCV%) and total erythrocytic count, were also evaluated. There were different degrees of pathogenicity between the strains.     

The extraintestinal stages of Eimeria tenella and E. maxima in the chicken

Donor chickens given feed medicated with one or two levels of decoquinate or given non-medicated feed were infected with oocysts of Eimeria tenella or E. maxima per os. Twelve hours after inoculation with oocysts liver, mid-intestine or ceca homogenates were fed to previously uninfected recipient chickens. The results showed that continuous medication with decoquinate was effective in preventing the transfer of sporozoites from the intestine to the liver. Oocysts were detected in the feces of all recipients of tissue from non-medicated donors, showing that some sporozoites of E. maxima and E. tenella are normally transferred to liver. Young broiler chickens were immunized by oral inoculation of E. maxima oocysts. The immune status of similar chickens inoculated with sporozoites of the same species directly into the liver or spleen were assessed. During the experimental period half of the chicks were provided with non-medicated food and the remainder were given feed supplemented with decoquinate; decoquinate was effective in arresting the development of the sporozoites. Two weeks after initial infection the birds were challenged with oocysts of E. maxima per os. Injection of sporozoites into the spleen did not protect against challenge. Birds inoculated with sporozoites into the liver were unable to develop a significant level of immunity. When the drug pressure was removed from these birds, parasitism of the intestine occurred and immunity developed.     

Assessment of dietary supplementation with probiotics on performance, intestinal morphology and microflora of chickens infected with Eimeria tenella

We evaluated the effects of dietary supplementation with different preparations of probiotics on the performance of broiler chickens experimentally infected with 2 × 10(4) sporulated oocysts of Eimeria tenella at 14 days of age. Three hundred, day-old, Cobb-500 chicks, as hatched, were separated into 10 equal groups with three replicates. Two of the groups, one challenged with E. tenella oocysts and the other not, were given a basal diet and served as controls without medication. The other challenged groups were given the anticoccidial lasalocid (60 mg/kg) or Enterococcus faecium (5 × 10(8) or 5 × 10(9)cfu/kg feed), Bifidobacterium animalis (5 × 10(8)cfu/kg feed), Lactobacillus reuteri (5 × 10(8)cfu/kg feed), Bacillus subtilis (5 × 10(8)cfu/kg feed), or a multi-species probiotic mix at 5 × 10(8) or 5 × 10(9)cfu/kg feed, respectively. The trial lasted 6 weeks. Individual body weight, feed intake per pen and feed conversion ratio values were recorded weekly, along with the extent of bloody diarrhea, excreta oocyst numbers and bird mortality. Caecal lesions were assessed and intestinal samples were taken for histopathological and bacteriological evaluation from ileum and caecum. Overall growth performance of chickens fed the multi-species probiotic mix at both levels was higher (P<0.05) compared to the infected control. Overall oocyst shedding was lowest (P<0.05) in the lasalocid supplemented group. Villous height was higher (P<0.05) in Bacillus supplemented groups compared to infected controls. The Lactobacillus supplemented group had the highest (P<0.05) numbers of both Lactobacillus and Bifidobacterium in ileum and caecum. In conclusion, dietary probiotics are promising for further investigation on improving intestinal health and growth performance of broiler chickens experimentally challenged with E. tenella.     

Efficacy of decoquinate against drug sensitive laboratory strains of Eimeria tenella and field isolates of Eimeria spp. in broiler chickens in China

The efficacy of decoquinate against Eimeria infections in broiler chickens was evaluated using two drug sensitive laboratory strains of Eimeria tenella and 20 field isolates of Eimeria spp. collected from farms in China where various anticoccidials (including maduramicin) had been used. Decoquinate (20-40 ppm in feed) and maduramicin (5 ppm) were efficacious against E. tenella laboratory strains, but decoquinate more so than maduramicin. Body weight gains of E. tenella infected chickens were significantly improved, and caecal lesions were prevented, by feeding either decoquinate or maduramicin. Decoquinate also prevented oocyst production, but maduramicin did not. Most (18/20) Eimeria field isolates were resistant to maduramicin, judged by oocyst production; decoquinate at > or =20 ppm completely controlled all 20 field isolates. Decoquinate has potential value as a broiler anticoccidial in China and other countries where it has not been previously used.     

Chemotherapy of caecal coccidiosis: efficacy of toltrazuril, sulphaquinoxaline/pyrimethamine and amprolium/ethopabate, given in drinking water, against field isolates of Eimeria tenella

Treatment with toltrazuril, sulphaquinoxaline/pyrimethamine and amprolium/ethopabate prevented mortality in chickens infected with field isolates of Eimeria tenella. Amprolium/ethopabate was the most effective drug in reducing lesions caused by the parasites. Few oocysts of E tenella were produced in birds medicated with sulphaquinoxaline/pyrimethamine or amprolium/ethopabate and none in those medicated with toltrazuril.     

Effect of dietary supplementation with oregano essential oil on performance of broilers after experimental infection with Eimeria tenella.

A study was carried out to examine the effect of dietary supplementation of oregano essential oil on performance of broiler chickens experimentally infected with Eimeria tenella at 14 days of age. A total of 120 day-old Cobb-500 chicks separated into 4 equal groups with three replicates each, were used in this study. Two groups, one infected with 5 x 10(4) sporulated oocysts of E. tenella and the other not, were given a basal diet and served as controls. The other two groups also infected with E. tenella were administered diets supplemented with oregano essential oil at a level of 300 mg/kg, or with the anticoccidial lasalocid at 75 mg/kg. Following this infection, survival rate, bloody diarrhoea and oocysts excretion as well as lesion score were determined. Throughout the experimental period of 42 days, body weight gain and feed intake were recorded weekly, and feed conversion ratios were calculated. Two weeks after the infection with E. tenella supplementation with dietary oregano oil resulted in body weight gains and feed conversion ratios not differing from the non-infected group, but higher than those of the infected control group and lower than those of the lasalocid group. These parameters correspond with the extent of bloody diarrhoea, survival rate, lesion score and oocyst numbers and indicated that oregano essential oil exerted an anticoccidial effect against E. tenella, which was, however, lower than that exhibited by lasalocid.     

柔嫩艾美耳球虫感染对肉仔鸡粪便水分的影响

采用单因子试验设计法,测定肉仔鸡感染柔嫩艾美耳球虫(E.tenella)后对粪便中水分变化的影响。试验表明:感染柔嫩艾美耳球虫后,鸡的生产性能受到显著的影响(P<0.05)。粪便中水分含量高于未感染组,感染后4d粪便中水分含量呈上升趋势,研究认为,粪便中水分含量的增加是球虫感染后的鸡的病理性变化的一种生理反映。在临床上,可将鸡粪便中水分含量的变化作为评判鸡感染球虫的参考指标之一。     

鸡胚接种柔嫩艾美耳球虫和堆形艾美耳球虫活卵囊的免疫保护试验

给18日龄鸡胚接种一定剂量的柔嫩艾美耳球虫(Eim eria tenella)和/或堆形艾美耳球虫(E.acervulina)孢子化卵囊,出雏后在无球虫环境中笼养,1~10日龄每天收集各组粪便样本,计数克粪便卵囊数(OPG),并于14日龄时以大剂量同源孢子化卵囊攻虫,以相对增重率(RWG)、饲料转化率(FCR)、相对卵囊产量(ROP)评价免疫保护效果。结果显示,以E.tenella或E.acervulina卵囊免疫18日龄鸡胚,其卵囊排出的潜隐期及达到峰值的时间与1日龄雏鸡接种组相一致,有相似的排卵囊曲线,提示其诱导免疫的建立是在出雏后开始建立的。攻虫后各免疫组的RWG由攻虫对照组的31.9%~51.7%提高到了76.5%~83.6%,RCR由攻虫对照组的4.11~4.89改善为2.72~2.96,ROP降至4.7%~23.5%。结果表明以一定剂量E.tenella和E.acervulina卵囊单独或混合经羊膜腔免疫18日龄鸡胚都可以建立起针对出雏后14日龄同源攻虫的良好免疫保护力。比较混合免疫E.tenella和E.acervulina卵囊组与单一接种E.tenella或E.acervulina卵囊组的免疫效果发现,混合免疫组的各项指标均稍优于后者。     

Isolation and selection of ionophore-tolerant Eimeria precocious lines: E. tenella, E. maxima and E. acervulina

Eimeria parasites were isolated from Nanhai Guangdong province (southern China) and studied in chickens in wire cages to evaluate their drug resistance against commonly used ionophores: monensin (100 mg/kg of feed), lasolacid (90 mg/kg), salinomycin (60 mg/kg), maduramicin (5 mg/kg) and semduramicin (25 mg/kg). Chinese Yellow Broiler Chickens were infected with 40,000 crude sporulated Eimeria oocysts at 15 days of age and prophylactic medication commenced a day prior to infection. Drug resistance was assessed for each ionophore drug by calculating the anticoccidial index (ACI) and percentage optimum anticoccidial activity (POAA) based on relative weight gain, rate of oocyst production and lesion values. Results revealed that Nanhai Eimeria oocysts comprising of E. tenella, E. maxima and E. acervulina, were resistant to monensin, sensitive to both salinomycin and lasolacid and partially sensitive to maduramicin and semduramicin. By selection for early development of oocysts during passage through chickens, the prepatent time of E. tenella, E. maxima and E. acervulina were reduced by 49, 36 and 22 h, respectively. The precocious lines are less pathogenic than the parent strains from which they were selected and conferred a satisfactory protection for chickens against coccidiosis. These ionophore-tolerant precocious lines could have wider applications in the development of anticoccidial vaccines for sustainable control of coccidiosis.     

Blackhead disease (Histomonas meleagridis) aggravated in broiler chickens by concurrent infection with cecal coccidiosis (Eimeria tenella).

The effect of concurrent cecal coccidiosis infections on severity of Histomonas meleagridis (blackhead disease) in chickens was investigated in a series of experiments. Cecal lesions from H. meleagridis were severe in all inoculated control groups and did not appear to be affected by the introduction of Eimeria tenella infection. However, the severity of liver lesions and number of birds positive for liver lesions of H. meleagridis increased significantly with the presence of E. tenella. The increase was similar when 10(3) or 10(4) oocysts of E. tenella were given and was the same when oocysts were given at the same time as H. meleagridis or 4 days prior. The liver lesions increased directly as doses of H. meleagridis increased from 7.5 x 10(3) cells to 30, 100, or 300 x 10(3) when E. tenella was given along with H. melelagridis but not when H. meleagridis was given alone. Administration of a live coccidiosis vaccine containing very low levels of E. tenella also gave a significant boost to liver lesions but at a much lower level than that observed with larger doses of E. tenella. The positive relationship between infections of cecal coccidiosis and H. meleagridis in chickens suggests that such dual exposure may contribute to increased clinical outbreaks of blackhead disease in chickens under field conditions.     

Comparison of disease susceptibility and subclass-specific antibody response in SC and FP chickens experimentally inoculated with Eimeria tenella, E. acervulina, or E. maxima

Susceptibility to disease and the subclass-specific antibody response to Eimeria tenella, E. acervulina, and E. maxima were compared in two inbred strains of chickens, FP (B15B21) and SC (B2B2). FP strain was more susceptible to coccidiosis than SC chickens based on oocyst production, lesion score, and clinical signs. FP chickens infected with E. tenella had more severe cecal lesions and a significantly lower hematocrit level than SC chickens. FP chickens infected with E. acervulina excreted five times as many oocysts at 6 days postinfection as SC and showed a 71% reduction in plasma carotenoid level compared with controls (56% reduction in SC chickens). Body-weight change did not correlate with other signs of disease. Both SC and FP chickens produced high levels of serum IgM and IgG and biliary IgA. Although SC chickens had a slightly higher antibody response than FP chickens at 7 days postinoculation, both strains maintained high levels of IgM, IgG, and IgA for a prolonged period post primary inoculation. Although SC and FP chickens show different disease susceptibility to coccidiosis, they demonstrate similar antibody response.     

Protective effects of sugar cane extracts (SCE) on Eimeria tenella infection in chickens

The effects of oral administration of sugar cane extracts (SCE) on Eimeria tenella oocysts infection in chickens were studied with 2 different experiments. In Experiment 1, 3-week-old inbred chickens (MHC; H.B15) were inoculated into the crop with SCE (500 mg/kg/day) for 1 day or 3 consecutive days, and then challenged with E. tenella sporulated oocysts (2 x 10(4) cells/chicken). In Experiment 2, 1-week-old chickens were orally administered SCE at the same dose for 3 consecutive days, and then initially infected with E. tenella sporulated oocysts (2 x 10(3) cells/chicken). At 2 and 3 weeks of age, these chickens were immunized intravenously with the mixed antigens of sheep red blood cells (SRBC) and Brucella abortus (BA). At 4 weeks of age, chickens were challenged with E. tenella sporulated oocysts (1 x 10(5)/chicken). Challenged chickens with E. tenella oocysts showed markedly decreased body weight gain/day, severe hemorrhage and great number of shedding oocysts in feces and high lesion scores. Oral administration of SCE and initial infection with oocysts (2 x 10 (3)/chicken) resulted in a remarkable improvement in body weight gain/day, hemorrhage, the number of shedding oocysts and lesion score, compare to other infected groups. In addition, SCE-inoculated chickens with the initial infection showed a significant increase in antibody responses against SRBC and BA and also improvement in decreased relative proportions of Bu-1a(+) and CD4( )cells in cecal tonsil lymphocytes of E. tenella-challenged chickens. Cecal tissues of chickens administered SCE and initially infected with E. tenella oocysts showed lower numbers of schizonts, gametocytes and oocysts than those of infected control chickens. These results suggest that SCE have immunostimulating and protective effects against E. tenella infection in chickens.     

Cecal coccidiosis in poultry as affected by prior exposure to aflatoxin B1.

Young New Hampshire and broiler chickens were given feed containing 2 concentrations of aflatoxin B1 (0.2 or 2.0 ppm). The larger concentration caused severe toxicosis and death in the New Hampshire chicks, but did not cause gross signs in the broiler chicks. However, temporary stunting was seen on both New Hampshire and broiler chicks during periods of aflatoxin feeding, along with persisting decreased weight gains in the broiler chicks; the latter apparently recovered during the next 21 days of feeding the starter ration. New Hampshire and broiler chicks which were given feed containing aflatoxin B1 at concentrations of 0.2 and 2.0 ppm for 28 days followed by a 21-day "recovery period" and which were not given a coccidiostat were more susceptible to severe cecal coccidiosis and had more persisting hepatic and cecal lesions than did chicks not given aflatoxin. The coccidiostat was protective against both cecal damage and losses in checks challenge exposed at 49 days of age to 100,000 infective Eimeria tenella oocysts.     

An enzyme-linked immunosorbent assay for coccidiosis in chickens: correlation of antibody levels with prior exposure to coccidia in the laboratory and in the field

An enzyme-linked immunosorbent assay was developed for detecting antibody to coccidia to facilitate the survey of laboratory and field infections. Serum antibody levels in chickens were measured against soluble Eimeria tenella oocyst antigen. Sera from breeders aged 10, 23, 37, and 43 weeks were positive with uniformly high antibody titers. Broiler chick sera showed high maternal antibody titer at hatch, decreasing to an almost negligible response at 3 weeks of age. Two-week-old broiler chicks had variable responses to a single infection of E. tenella: titers were elevated at 8 to 10 days postinfection and generally increased through day 24. Weekly reinfection of 2-week-old broiler chickens produced an antibody titer in proportion to the number of oocysts per dose and stimulated protection against challenge with 2 x 10(5) E. tenella. Inbred birds raised in a pathogen-free environment for 6 weeks had no detectable antibody titers.     

Studies on the stage of action of lasalocid against Eimeria tenella and Eimeria acervulina in the chicken

Broiler chickens in battery pens were either fed a diet containing 100 ppm lasalocid or no drug for 24 h prior to inoculation with sporulated oocysts of Eimeria tenella or Eimeria acervulina. Different groups of birds remained on medicated feed for 24, 48, 72, 96, 120 or 144 h after inoculation. Conversely, other groups started on an unmedicated diet, were given medicated feed at different times after oocyst inoculation. Starting lasalocid medication 24 h (E. tenella) or 48 h (E. acervulina) after inoculation reduced the lesions and improved the weight gain. There was no significant difference in performance of birds after withdrawal of the drug at 48 h (E. tenella) or 72 h (E. acervulina) and thereafter. Starting lasalocid medication at 96 or 120 h did not suppress but rather reduced oocyst production.     

柔嫩艾美耳球虫对盐酸氟乙基硫胺素耐药性的诱导

为评估盐酸氟乙基硫胺素产生抗药性的倾向,采用药物浓度递增法,在实验条件下进行柔嫩艾美耳球虫耐药性的诱导研究。每组动物用10只快大黄肉鸡,每只鸡接种1×105个孢子化卵囊,以50 mg/kg盐酸氨丙啉和盐酸氟乙基硫胺素为起始诱导浓度连续传代,以病变记分、存活率和卵囊数三项指标综合判定耐药性。经过10次传代,盐酸氨丙啉对敏感虫株的ACI为192.6,而对耐药虫株的ACI为109.1;经过12次传代,盐酸氟乙基硫胺素对敏感虫株的ACI为197.5,而对耐药虫株的ACI为110.6,对盐酸氨丙啉耐药虫株的ACI为163.2;结果表明成功地诱导出对盐酸氨丙啉、盐酸氟乙基硫胺素完全耐药的虫株。盐酸氟乙基硫胺素产生耐药速度慢于盐酸氨丙啉,与盐酸氨丙啉有部分交叉耐药性。     

球虫抗药性分子生物学检测技术的建立

选择在敏感虫株中沉默、抗药虫株中表达的特异序列AGD5设计检测引物，采用RT—PCR方法。以总RNA反转录的cDNA为模板，RT—PCR方法能够鉴别马杜霉素抗药虫株和3株马杜霉素敏感虫株（2株对所有药物敏感的不同地理株，1株地克珠利抗药性虫株对马杜霉素敏感），此鉴定结果与鸡体试验的结果一致，说明敏感虫株和抗药虫株在该序列上的差异发生在转录水平。而以卵囊总DNA为模板的PCR方法不能鉴别敏感和马杜霉素抗药性虫株。     

Evaluation of interspecific hybrids of the chicken, guinea fowl, and Japanese quail for innate resistance to coccidia

Experimental chicken/guinea fowl hybrids, guinea fowl, and chickens were orally inoculated with Eimeria acervulina or E. tenella, which are specific for chickens, or with E. grenieri, which is specific for guinea fowl. No intact oocysts were found in feces within 24 hr of inoculation, suggesting that excystation occurred in the normal and abnormal hosts. No oocysts were found in the feces of hybrids during a 9-day postinoculation period. The guinea fowl passed oocysts of guinea fowl coccidia (E. grenieri) but not those of chicken coccidia, and the chickens passed oocysts of chicken coccidia (E. acervulina and E. tenella) but not those of guinea fowl coccidia. Some asexual development (schizogony) occurred in hybrids inoculated with E. tenella, but sexual development (gametogony) did not. In contrast, quail/chicken hybrids became infected with oocysts of chicken coccidia (E. acervulina, E. tenella, and E. maxima) and quail coccidia (E. bateri) and passed a few oocysts during the normal patent period; control chickens and quails became heavily infected with oocysts of chicken and quail coccidia, respectively.