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.     

堆型艾美耳球虫早熟株对常用抗球虫药的敏感性测定

The study was aimed to explore the sensitivity of Eimeria acervulina （E.acervulina） precocious strain to some common anticoccidials (diclazuril, toltrazuril, sulfamonomethoxine, dinitolmide, decoquinate, nicarbazin, maduramicin, salinomycin and oxytetracycline). The sensitivity of E.acervulina precocious strain to 9 kinds anticoccidials was assessed by using 4 indices: anticoccidial activity percentage (POAA), reduction of lesion scores (RLS), relative oocyst production (ROP) and anticoccidial index (ACI). The results of these sensitivity trials indicated that the four indices POAA, RLS, ROP and ACI of groups toltrazuril, sulfamonomethoxine, dinitolmide, decoquinate, nicarbazin, salinomycin and oxytetracycline were all negative; the ROP index of diclazuril group was positive, the other three indices were negative. The ROP and ACI indices of maduramicin group were positive, the other two indices were negative. The results showed that the Eimeria acervulina precocious strain was sensitive to diclazuril, toltrazuril, sulfamonomethoxine, dinitolmide, decoquinate, nicarbazin, salinomycin and oxytetracycline, it was partly sensitive to maduramicin.     

巨型艾美耳球虫和柔嫩艾美耳球虫早熟株对8种抗球虫药的敏感试验

为检测巨型艾美耳球虫和柔嫩艾美耳球虫早熟株对常用抗球虫药的敏感性,选用地克珠利、氯苯胍、二硝托胺、癸氧喹酯、尼卡巴嗪、马杜米星、盐霉素和拉沙洛西等8种抗球虫药进行鸡体试验,通过抗球虫活性百分比（percent of optimum anticoccidial aetivity,POAA）、病变记分减少率（reduction of lesion scores,RLS）、相对卵囊产量（relative oocyst production,ROP）和抗球虫指数（anticoccidial index,ACI）4项指标进行综合评价。结果表明,巨型艾美耳球虫早熟株对8种药物敏感;柔嫩艾美耳球虫早熟株对盐霉素轻度敏感,对地克珠利、氯苯胍、二硝托胺、癸氧喹酯、尼卡巴嗪、马杜米星和拉沙洛西敏感。本研究结果可为对球虫早熟株疫苗的开发与应用提供实验依据。     

堆形艾美耳球虫早熟株选育及相关生物学特性研究

为了选育堆形艾美耳球虫（Eimeria acervulina）早熟株及观测其生物学特性,运用Jeffers创立的艾美耳属球虫早熟株选育方法,对实验室保存的堆形艾美耳球虫进行连续传代早熟选育,并对获得的早熟株与母株孢子化卵囊大小、繁殖能力、致病性以及免疫保护力等指标进行了测定,比较堆形艾美耳球虫早熟株与母株之间的基本生物学特性差异。经过18代早熟选育后,堆形艾美耳球虫的潜在期由母株的99h缩短至84h。与母株相比,早熟株的孢子化卵囊大小明显减小（P〈0．05）,卵囊繁殖能力降低了17％～42%。致病性试验中,早熟株感染组的平均增重均高于母株,而病变记分相当。免疫保护试验中,早熟株与母株的免疫保护力相当,其卵囊抑制率略低于母株,差异不明显（P〉0．05）。结果表明,本研究成功选育出堆形艾关耳球虫早熟株,对母株攻击具有保护力,为鸡球虫疫苗的研制奠定了基础。     

The efficacy and economic benefits of Supercox, a live anticoccidial vaccine in a commercial trial in broiler chickens in China

The efficacy and economic benefits of Supercox, a live anticoccidial vaccine were examined and compared with an anticoccidial drug in a trial in broiler chickens under modern commercial conditions in China. In total, 40,660 chickens were used in the present study, half of which were vaccinated with the Supercox vaccine comprising a precocious line of Eimeria tenella and non-attenuated lines of Eimeria maxima and Eimeria acervulina, and the other half were medicated with Diclazuril delivered as feed additive at the dosage of 1mg/kg of feed. The vaccine was administered orally to 7-day-old chickens. No clinical diseases were diagnosed in any of the vaccinated birds. However, clinical coccidiosis occurred in a large proportion of medicated control birds and these chickens had to be treated with anticoccidial drugs (Diclazuril and Toltrazuril). Comparison of production performance between vaccinated birds and medicated control birds revealed that the vaccine Supercox performed better than anticoccidial drugs in terms of mortalities, costs and overall economic benefits (profits). These findings demonstrated that the use of the Supercox vaccine could control clinical coccidiosis in broilers and achieve production performance superior to that using anticoccidial drugs, particularly where drug resistance might result in failure to control clinical diseases.     

Does Plasmodium gallinaceum induce relapse of Eimeria acervulina infection in chickens?

An experiment was carried out to test the hypothesis that chickens previously infected with Eimeria acervulina, but having ceased producing oocysts, recommence E acervulina oocyst production when infected with Plasmodium gallinaceum. No relapse of coccidiosis was caused by the malarial infection. Subsequent treatment with the immunosuppressant betamethasone of the control chicks infected with E acervulina only did not reveal any occult coccidial infection. The results are critically compared with previously published results which apparently supported the hypothesis.     

Selection and characterisation of two attenuated vaccine lines of Eimeria tenella in Australia

OBJECTIVE: To attenuate two strains of Eimeria tenella by selecting for precocious development and evaluate the strains in characterisation trials and by field evaluation, to choose one precocious line for incorporation into an Australian live coccidiosis vaccine for poultry. DESIGN: Two strains from non-commercial flocks were passaged through chickens while selecting for precocious development. Each strain was characterised for drug sensitivity, pathogenicity, protection against homologous and heterologous challenge, and oocyst output in replicated experiments in which the experimental unit was a cage of three birds. Oocyst output and/or body weight gain data collected over a 10 to 12 day period following final inoculation were measured. Feed conversion ratios were also calculated where possible. RESULTS: Fifteen passages resulted in prepatent periods reduced by 24 h for the Redlands strain (from 144 h to 120 h) and 23 h for the Darryl strain (from 139 h to 116 h). Characterisation trials demonstrated that each precocious line was significantly less pathogenic than its parent strain and each effectively induced immunity that protected chickens against challenge with both the parent strain and other virulent field strains. Both lines had oocyst outputs that, although significantly reduced relative to the parent strains, remained sufficiently high for commercial vaccine production, and both showed susceptibility to coccidiostats. CONCLUSION: Two attenuated lines have been produced that exhibit the appropriate characteristics for use in an Australian live coccidiosis vaccine.     

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.     

Effect of Eimeria acervulina infection on nutrient retention with special reference to fat malabsorption in chickens.

Nutritional balance studies were conducted to determine the effect of Eimeria acervulina infection on the retention of nutrients and the efficiency of utilization of protein and energy in the diet during the course of intestinal coccidiosis in chickens. Both light and electronmicroscopic studies were conducted to elucidate the pathogenesis of fat malabsorption in the infected chickens. The infection with E. acervulina during the acute phase of the disease (day 4 to day 8 postinfection) caused a reduction (P smaller than 0.01) in the retention of percent protein, percent ether extract and percent gross energy of the diet with a concomitant increase (P smaller than 0.01) in the concentration of ether extract and gross energy of the excreta. The metabolizable energy value of the diet was also depressed during this phase. During the recovery phase of the disease (day 9 to day 13 postinfection) infected chickens voided excreta containing a slightly greater concentration of ether extract when compared to the controls but differences in the retention of percent protein and percent gross energy were no longer observed. An increased retention of percent phosphorus also occured during the recovery phase. Studies involving the carcass composition revealed that the efficiency of utilization of protein and metabolizable energy of the diet for tissue deposition was reduced (P smaller than 0.05) during day 0 to day 8 postinfection but only the efficiency of metabolizable energy utilization was found to be depressed (P smaller than 0.05) when data for day 0 to day 14 postinfection were analyzed. Both light and electronmicroscopic studies of the duodenal villi of infected chickens during day 5 to day 6 postinfection revealed accumulation of large globules of fat in the villus epithelial cells parasitized by the gammonts of E. acervulina indicating a possible blackade of "fat exit" from these cells.     

柔嫩艾美球虫常州株对5种抗球虫药的抗药性分析

将柔嫩艾美耳球虫(E.tenella)常州分离株感染14日龄苏禽黄羽肉鸡,设立5个用药组、1个感染不用药对照组和1个不感染不用药对照组,以相对卵囊产量、病变记分减少率、最适抗球虫百分数和抗球虫指数为指标对该虫株对球虫宁、地克珠利、马杜霉素、磺胺氯吡嗪、癸氧喹酯5种常用抗球虫药的抗药性进行了综合评定,分析了该地区球虫的抗药性及用药对策。结果表明,该虫株对癸氧喹酯敏感,对球虫宁、地克珠利、马杜霉素、磺胺氯吡嗪具有完全抗药性,提示目前常州地区球虫抗药性非常严重,癸氧喹酯抗球虫疗效最好,其他药物应合理使用、应慎用、减少或暂停使用。     

Relative value of oocyst counts in evaluating anticoccidial activity.

Birds medicated with roxarsone and in another experiment with zoalene in the feed produced higher oocysts counts than unmedicated control birds receiving the same oocyst dose of Eimeria tenella or a mixture of six species (E. tenella, E. necatrix, E. brunetti, E. maxima, E. acervulina, and E. mitvati). These experiments confirm the conclusion that oocyst counts constitute an unsatisfactory and unreliable parameter for judging effectiveness of an anticoccidial even though such increases are a relatively rare occurrence in anticoccidial evaluation experiments.     

柔嫩艾美耳球虫盐霉素耐药株的实验室诱导

为了研究耐药性柔嫩艾美耳球虫产生耐药性的机理,本研究在实验室条件下,采用药物浓度递增的方法,以15 mg/kg为起始诱导浓度,对柔嫩艾美耳球虫盐霉素耐药性进行诱导.经过10次传代,获得了对60 mg/kg盐霉素耐药的柔嫩艾美耳球虫虫株.以抗球虫指数、最适抗球虫活性百分率、病变记分减少率和相对卵囊产量四项指标综合判定柔嫩...     

Higher incidence of Eimeria spp. field isolates sensitive for diclazuril and monensin associated with the use of live coccidiosis vaccination with paracox-5 in broiler farms

Twenty European Eimeria spp. field isolates were subjected to an anticoccidial sensitivity test (AST). The anticoccidial drugs tested were diclazuril (Clinacox) and monensin (Elancoban). The assay was performed in a battery cage trial. Infected medicated birds were compared with an unmedicated control group. Coccidial lesion scores and oocyst shedding were used as parameters. The results of the AST show that resistance is common amongst coccidiosis field isolates, especially Eimeria acervulina (68% and 53% resistance for diclazuril and monensin, respectively). Resistance is less frequent amongst Eimeria maxima (38% and 50% resistance for diclazuril and monensin, respectively) and Eimeria tenella isolates (23% and 38% resistance for diclazuril and monensin, respectively). A highly significant influence of the coccidiosis prevention program (live coccidiosis vaccination with Paracox-5 vs. anticoccidial drugs in feed) on the sensitivity patterns of Eimeria spp. field isolates for both diclazuril (P= 0.000) and monensin (P= 0.001) was found. Further, when looking at the single species and each anticoccidial drug level, significantly more sensitivity of E. acervulina for monensin (P= 0.018), E. maxima for diclazuril (P = 0.009), and E. tenella for diclazuril (P = 0.007) was found in isolates originating from vaccinated flocks. Moreover, for E. acervulina and diclazuril, E. maxima and monensin, and E. tenella and monensin a trend toward higher sensitivity of isolates for these products was found when live coccidiosis vaccination was applied. The present study shows that sensitivity for the anticoccidial drugs diclazuril and monensin is more frequent in Eimeria spp. field isolates originating from broiler farms where a coccidiosis vaccination policy is followed.     

柔嫩艾美耳球虫早熟耐药株对几种抗球虫药的敏感性

为了探讨鸡柔嫩艾美耳球虫早熟耐药株对常用抗球虫药的敏感性,采用尼卡巴嗪、氯苯胍、球痢灵、地克珠利和常山酮5种抗球虫药,对由晋球1号球虫疫苗虫株复制的病鸡进行防治试验,通过相对卵囊产量(ROP)、相对增重率、抗球虫指数(ACI)、病变记分减少率(RLS)和最适抗球虫活性百分率(POAA)5项指标进行综合评定.结果表明,柔嫩艾美耳球虫早熟耐药株对盐酸氯苯胍无抗药性,对地克珠利有轻度抗药性,对常山酮、球痢灵、尼卡巴嗪有部分抗药性,它们的抗球虫指数为187.46～246.75,达到了抗球虫效果.     

柔嫩艾美耳球虫莆田株抗药性研究

为掌握鸡场球虫抗药程度,以便制定合理用药方案,采集福建省莆田市某鸡场盲肠球虫,进行柔嫩艾美耳球虫单卵囊分离和纯化;采用鸡体试验法,设立5个用药组、1个感染不用药对照组和1个不感染不用药对照组,以病变记分减少率(Reduction of lesion sores,RLS)、抗球虫指数(Anticoccidial index,ACI)、相对盲肠卵囊产量(Relative at oocyst production,ROP)和最适抗球虫活性百分率(Percent of optimum anticoccidial activity,POAA)4项为指标,检测该球虫对癸氧喹酯、尼卡巴嗪、地克珠利、二硝托胺和氯苯胍等5种常用抗球虫药的抗药性。结果表明:该虫株对癸氧喹酯敏感,对地克珠利、二硝托胺和氯苯胍具有完全抗药性,对尼卡巴嗪为中度抗药性,说明目前莆田市球虫抗药性非常严重,癸氧喹酯可用于抗球虫病,其他药物应慎用或暂停使用。     

柔嫩艾美耳球虫扬州分离株对8种抗球虫药的抗药性

将柔嫩艾美耳球虫(Eimeria tenella)扬州分离株感染14日龄苏禽黄羽肉鸡,设立8个用药组、1个感染不用药对照组和1个不感染不用药对照组,以相对卵囊产量、病变记分减少率、最适抗球虫百分数和抗球虫指数为指标对该虫株对盐霉素、马杜拉霉素、球痢灵、氯羟吡啶、氨丙啉、尼卡巴嗪、甲基三嗪酮、地克珠利等8种常用抗球虫药的抗药性进行了综合评定,分析了该地区球虫的抗药性及对策。结果表明,该虫株对甲基三嗪酮敏感,对氨丙啉具有中度抗药性,对盐霉素、马杜拉霉素、球痢灵、氯羟吡啶、尼卡巴嗪、地克珠利等其他6种药物具有完全抗药性。提示目前在扬州地区甲基三嗪酮抗球虫疗效最好,氨丙琳效果稍差,应合理使用,盐霉素等其他6种药物应慎用、减少或暂停使用。     

Coccidiosis: a radiological study of sulphaquinoxaline distribution in infected and uninfected chickens

Using scintillation counting and autoradiographical techniques, the whole-body distribution in week-old uninfected chickens of the anticoccidial agent sulphaquinoxaline (SQ) labelled with ³⁵S was established at various time intervals after a single oral dose either alone or following continuous in-feed medication with unlabelled SQ, and after a single intravenous dose. The distribution was also established in chickens infected with the coccidia Eimeria acervulina or E. tenella , after a single oral dose of radiolabelled SQ administered either alone or following continuous in-feed medication with unlabelled SQ, as for uninfected chicks. In all uninfected chicks, SQ was rapidly absorbed from the gut and was distributed to all tissues. It appeared at high concentrations in the bile and kidneys 0.5 h after dosing. In chickens that had previously received unlabelled SQ in the diet, a radiolabelled dose maintained steadier tissue concentrations than the sharp rise and fall detected after a single oral dose. Intravenous dosing of uninfected chicks showed that SQ was secreted by the crop, gizzard and caecal epithelia into their lumina. Infection with E. acervulina or E. tenella coincided with an overall 3.5-fold sustained increase of SQ concentration in chick tissues. An updated hypothesis including these new observations for the anticoccidial mode of action of SQ in chickens is expounded.     

Drug resistance in coccidia: a robenidine-resistant strain of eimeria tenella.

Tests for resistance to a recently introduced anticoccidial drug, robenidine, were performed on a strain of Eimeria tenella recovered from broiler chickens infected with acute cecal coccidiosis. The strain was identified previously and again confirmed by the timing of mortality of the infected chickens (five to six days postinfection), the appearance of lesions and parasites in ceca only, and the measurements (21.6 +/- 1.9 micron X 19.8 +/- 2.4 micron) of the oocysts. The tests showed that the strain of E. tenella could establish infections in chickens medicated with fourfold the recommended level of robenidine in feed but no oocysts could be recovered from the feces of infected chickens when the level of drug was increased to eightfold. The decrease in mortality, in the severity of lesions and oocyst output and the increase in the average weight gain of infected chickens followed closely the increase in the level of robenidine in feed. In contrast, no infections were found in chickens infected with a sensitive strain of E. tenella and maintained on feed mixed with the recommended level of robenidine. This and other findings discussed here show that resistance to robenidine is developing in coccidia commonly found in broiler houses.     

Response of 21 Canadian field isolates of chicken coccidia to commercial anticoccidial drugs

Coccidia were isolated from 30 broiler farms in major poultry-producing provinces of Canada; 21 mixed-species isolates were tested for sensitivity to anticoccidial drugs. Eimeria acervulina was identified in 20 samples, E. maxima in 15, and E. tenella in 10 of the mixed-species isolates. On the basis of weight gain after infection, 3 isolates were judged sensitive to amprolium, 5 to monensin, 9 to salinomycin, 14 to lasalocid, 19 to halofuginone, and 20 to nicarbazin. Similar trends in sensitivity/tolerance were identified from the percent reduction of intestinal lesion scores of medicated birds compared with scores of unmedicated, infected controls. These results were consistent with the previous pattern of use of anticoccidial drugs in commercial poultry in Canada.     

毒害艾美耳球虫早熟株与亲本株繁殖力的观察

为选育毒害艾美耳球虫(E.necatri)早熟株,了解其繁殖力,为早熟弱毒活苗的研制奠定基础,本试验运用球虫的单卵囊分离技术得到一株E.necatrix(P0),并对其进行了早熟选育,得到了E.necatrix早熟株P8。P0和P8分别以0.05×104、0.1×104、0.5×104和1×104个/只的剂量接种11日龄雏鸡,接种后第7～13天每天检测粪便中的卵囊产量。结果表明,P0与P8均为接种0.5×104个/只的卵囊产量最大;P8的繁殖力是P0的55%;P8的卵囊高峰期较P0有提前趋势。