鸡隐孢子虫病有效药物筛选

为寻求治疗鸡隐孢子虫病的有效药物。本试验选取140只1日龄健康无感染鸡,随机分为7组,每组20只,除Ⅰ组外,Ⅱ-Ⅶ组均接种火鸡隐孢子虫卵囊1×106个/只。试验组鸡粪便检查呈阳性后,分别用硝唑尼特、阿奇霉素、巴龙霉素、常山酮和大蒜素等对Ⅲ-Ⅶ组进行治疗试验,以各组雏鸡日采食量、增重量、卵囊排出情况作为疗效指标。结果显示,经大蒜素治疗后的感染鸡,日采食量58.6g、日增重74.9g、每克粪便卵囊排出量减少1 739个,均优于其他药物治疗组,治疗效果显著。大蒜素可以提高病鸡的采食量、机体的免疫力并有效抑制卵囊的排出,可以作为治疗火鸡隐孢子虫病的备选药物。     

保存在自来水中的微小隐孢子虫卵囊活性及感染性研究

微小隐孢子虫卵囊(CPO)保存在4℃自来水中1~30个月,通过体外脱囊技术检测CPO的脱囊率评价其活性,通过检测CPO感染免疫抑制BALB/c小鼠的潜伏期、排卵囊数量和末端回肠绒毛中的隐孢子虫数量来评价其感染性。结果表明,保存在自来水中1~13个月的CPO出现脱囊;小鼠在感染保存1~13个月的CPO后3~8 d开始排出大量的CPO,在末端回肠绒毛中寄生有大量的隐孢子虫;CPO的保存时间与潜伏期之间存在强烈的相关性(r2=0.98)。因此,CPO在自来水中能保持活性和感染性至少13个月,水是保存CPO的良好介质,水中活性CPO的长期存在对饮用水工业是一个严重的挑战。     

微小隐孢子虫感染绵羊模型的建立

为建立稳定、可靠的微小隐孢子虫(Cryptosporidium parvum)感染绵羊模型,将4只2日龄绵羊分成2组,分别经口接种1×106个(低剂量组)和1×107个(高剂量组)的C.parvum卵囊,观察绵羊每日精神状况,并采集粪便样品,提取DNA,用基于TaqMan探针的实时荧光定量PCR方法进行检测,计算每日排卵囊数量。结果表明,两组羔羊均从第4天开始出现精神沉郁、食欲下降及严重的腹泻症状,镜检发现有大量的C.parvum卵囊排出,其中高剂量组的2只羔羊在感染后的第10天死亡。TaqMan实时荧光定量PCR检测结果显示,两组羔羊均从第1天开始有少量的卵囊排出,从第5天开始增多。其中低剂量组的1#羔羊第9天达到高峰,2#羔羊第7天达到高峰,随后排卵囊量开始下降,2只羔羊直到实验结束的第36天仍有卵囊排出,并且首次发现羔羊感染C.parvum后出现3个排卵囊高峰期。高剂量组的2只羔羊均在第6天达到高峰,其中3#羔羊第7天仍然有大量卵囊排出,4#羔羊第7天和第8天仍有大量卵囊排出,随后开始下降;高剂量组羔羊高峰期排卵囊数量明显高于低剂量组羔羊高峰期排卵囊数量。对死亡羔羊进行的肠道组织学观察,发现虫体寄生部位肠黏膜层出现大面积实质细胞脱落,局部可见肠腺数量减少,被增生的结缔组织取代。提取死亡羔羊肠道内容物和粪便DNA,进行套式PCR扩增,经测序鉴定,证实为C.parvum感染,与羔羊接种的虫种一致。上述结果表明,接种1×106个C.parvum卵囊,绵羊出现明显的临床症状,并伴有大量的卵囊排出,可以作为C.parvum的感染模型;而接种1×107个C.parvum卵囊,绵羊会发生死亡,但是能够排出更多的卵囊,适合进行隐孢子虫的繁殖试验。为进一步利用该模型研究C.parvum的致病机理、疫苗和药物筛选奠定了良好的基础。     

隐孢子虫卵囊形态学观察及动物交叉感染试验

作者观察了从湖南６种动物分离出的隐孢子虫卵囊的光镜下形态，并以鸡源、牛源和猪源隐孢子虫卵囊人工感染实验动物作交叉感染性研究。结果鉴定出隐孢子虫３个种，即贝氏隐孢子虫（Ｃ．ｂａｉｌｅｙｉ），寄生于鸡、鸭；微小隐孢子虫（Ｃ．Ｐａｒｖｕｍ），寄生于牛、山羊、猪、家兔和小鼠；鼠隐孢子虫（Ｃ．ｍｕｒｉｓ），寄生于牛。交叉感染结果表明，来自鸡的隐孢子虫可以感染雏鸡和雏鸭，而不能感染小鼠和家兔；来自牛的隐孢子虫可以感染小鼠和家兔而不能感染雏鸡，来自猪的隐孢子虫可以感染小鼠而对雏鸡无感染性。作者认为，哺乳类和鸟类的隐孢子虫可能在宿主纲的水平上具有宿主持异性。     

贝氏隐孢子虫对樱桃谷鸭的致病性试验

隐孢子虫病是一种全球性的人畜共患原虫病；禽类隐孢子虫发现于鸡、火鸡、鹌鹑、鹅、孔雀等，有关鸭隐孢子虫病的报道较少，特别是不同剂量感染的致病性差异未见报道，本试验采用鸭源贝氏隐孢子虫卵囊，雏鸡传代后，经口感染2日龄樱桃谷鸭，通过临床症状观察和病理变化观察，旨在探讨河南省鸭源贝氏隐孢子虫的致病性及不同剂量的卵囊接种后所引起的临床症状和病理变化的差异。     

乳酸常山酮治疗犊牛实验性小隐孢子虫感染的效果

检测了乳酸常山酮预防犊牛实验性隐孢子虫病的效果。２０头２日龄犊牛分成四组，口服接种１×１０６个小隐孢子虫卵囊。感染对照组不投药，而其它３组从接种后（０天为接种天）第２天到第８天连续７天每天分别按３０、６０和１２０μ／ｋｇ给药。感染后每周称重两次，并根据粪便的粘稠度、卵囊的排出和死亡率、０～３０天，每天评价疾病的发展和药物的功效。没有投药的对照组，实验性小隐孢子虫感染引起水泻、卵囊排出量大和死亡率高（５头中死亡３头）的严重疾病。结果清楚地证实了乳酸常山酮可以降低临床隐孢子虫病的严重程度，其功效具有剂量依赖性，最低剂量（每日３０μｇ／ｋｇ）不能够预防本病和降低死亡率（５头中死亡３头），每日６０和１２０μｇ／ｋｇ剂量组未观察到临床症状，但停药后，犊牛则排出卵囊，给予较高剂量的药物，排出卵囊时间明显推迟，但排卵囊时间的推迟对犊牛的生长没有影响。     

硝唑尼特的多重作用机制及应用研究进展

硝唑尼特(NTZ)是上世纪七十年代开发的一个抗寄生虫药物,已被美国食品药品监督管理局(FDA)批准用于治疗隐孢子虫或蓝氏贾第鞭毛虫引起的腹泻,被中国农业农村部批准用于治疗犬绦虫病。近年来的研究发现,硝唑尼特不仅能抗多种寄生虫,而且对多种危害严重的人或动物病原菌、病毒也具有很好的活性,但是研究发现硝唑尼特对各种病原体的活性作用机制却各不相同。本文主要综述了硝唑尼特对多种病原体的作用及其可能的机制,从而为硝唑尼特的利用和相关研究提供资料。     

鸡隐孢子虫病治疗试验

作者以高免血清和隐孢子虫卵囊裂解物等10种药物,对113例人工感染鸡病例分4批次共12组进行了治疗试验,以临床症状、剖检变化、排出卵囊的数量及增重额度等指标判定疗效,其结果表明,高免血清及卵囊裂解物对本病具有良好的治疗效果,尤其高免血清的疗效更佳,为本病的治疗创出了新路。     

隐孢子虫某些生物学特性研究

(1)对长春地区兔、小鼠、牛以及婴幼儿腹泻的隐孢子虫感染情况作了调查,其感染率分别为36%、90%、9/10和1/22;(2)对兔隐孢子虫卵囊排出规律的观察结果表明,每隔6～8d出现一个高峰期,在两个高峰期之间有时检不到卵囊;(3)交叉感染试验,从兔粪便中分离的卵囊经口感染BALB/c乳鼠和雏鸡后,在鼠粪便中检到卵囊,而在鸡粪便中未发现卵囊;(4)建立了兔和鼠的隐孢子虫病动物模型;(5)成功地进行了隐孢子虫体外培养试验;(6)鼠隐孢子虫病的治疗药物筛选结果,10~(-3)mol/L SM520和10~(-6)mol/LSMW在体外对隐孢子虫有杀灭作用;动物试验发现,中药配方I和SMW对鼠隐孢子虫有一定抑制作用.     

合肥市犬隐孢子虫感染情况初步调查

目的了解合肥市犬隐孢子虫的感染情况。方法在合肥市随机采集不同年龄、性别与饲养条件的犬粪样69份,采用金胺-酚改良抗酸染色法检测粪样隐孢子虫感染情况,并根据卵囊大小和形态,进行虫种鉴定。结果犬隐孢子虫感染率为28.99%。对68个隐孢子虫卵囊进行观察和测量,椭圆形卵囊大小平均为6.20μm×4.34μm,卵囊形状指数平均为1.44,初步鉴定为鼠隐孢子虫（Cryptosporidium muris）;近圆形卵囊,大小平均为4.69μm×4.58μm,卵囊形状指数平均为1.125,初步鉴定为微小隐孢子虫（Cryptosporidium parvum）。结论犬隐孢子虫的感染率存在年龄差异,年龄越小,感染率越高;饲养管理条件差的犬隐孢子虫感染率高;而隐孢子虫感染率与性别没有关系。     

Infectivity of Cryptosporidium sp isolated from wild mice for calves and mice

A study was conducted to determine the incidence of cryptosporidiosis in wild mice (Mus musculus) and the infectivity of oocysts from their feces for susceptible calves. The presence of oocysts and the duration of shedding of oocysts in the feces were evaluated in 115 wild mice. Approximately 30% of the mice shed Cryptosporidium sp oocysts, without evidence of clinical infection; recurrence of oocyst shedding was found in about 50% of the mice. Oocysts from the feces of naturally infected mice were infective for calves and mice. Calves began shedding oocysts at 7 days and shed oocysts for about 10 days. Nonfatal, clinical cryptosporidiosis developed in 7 infected calves. The mice began shedding oocysts at 6 days and shed oocysts for 12 days. Fatalities or clinical infection did not develop in 5 infected mice. The results indicated that Cryptosporidium-infected wild mice may be a source of cryptosporidiosis in susceptible calves.     

Concurrent response to challenge infection with Cryptosporidium parvum in immunosuppressed C57BL/6N mice

We investigated the response to challenge infection with Cryptosporidium parvum oocysts in immunosuppressed C57BL/6N mice. In the primary infection, fecal oocyst shedding and parasite colonization were greater in immunosuppressed mice than in nonimmunosuppressed mice. Compared with primary infection, challenge infection with C. parvum didn''t show any oocyst shedding and parasite colonization. Especially, oocyst shedding and parasite colonization from the mice infected with heat-killed oocysts were not detected. After challenge infection with C. parvum oocysts, however, these mice were shedding small numbers of oocysts and parasite colonization. Except normal control and uninfected groups, the antibody titers of other groups appear similar. Based on the fecal oocyst shedding, parasite colonization of ilea, and antibody titers in the mice, these results suggest that the resistance to challenge infection with C. parvum in immunosuppressed C57BL/6N mice has increased.     

Effect of halofuginone lactate on treatment and prevention of lamb cryptosporidiosis: an extensive field trial

This study aimed to investigate the effects of halofuginone lactate (100 mug/kg BW) on treatment and prevention of cryptosporidiosis in lambs. It consisted of three field trials. The first trial was designed to assess the efficacy of halofuginone in treating lamb diarrhoea caused by cryptosporidiosis, and in preventing the disease, using two schemes; halofuginone given for seven and for three consecutive days respectively. Halofuginone was effective in the treatment of diarrhoea caused by cryptosporidiosis (P < 0.01). In addition, halofuginone administered as a 7-day treatment was significantly (P < 0.05) more effective than a 3-day treatment in preventing diarrhoea in the infected flocks. The second trial was designed to evaluate the preventive effect of halofuginone, which was administered for 7 days in lambs infected with cryptosporidiosis, on diarrhoea incidence, oocyst shedding and body weight gain. Halofuginone significantly (P < 0.01) reduced the diarrhoea incidence, the time of oocyst shedding and the mean intensity of shedding, but did not affect body weight gain. The third trial was designed to examine the ability of halofuginone to reduce the death rate in flocks with cryptosporidiosis. Halofuginone treatment was effective in preventing and in reducing the death rate of cryptosporidiosis in these flocks.     

Infectivity of Cryptosporidium parvum isolated from asymptomatic adult goats to mice and goat kids.

An experimental study was carried out in neonatal goat kids to examine the infectivity of Cryptosporidium oocysts, pattern of oocyst shedding and morphological changes in the intestine during the infection. Cryptosporidium oocysts isolated from adult asymptomatic goats, and identified as C. parvum by polymerase chain reaction (PCR) were used in this study. Of three 4-day-old goat kids, two were orally infected with C. parvum oocysts (10(5) oocysts in 10 ml PBS/kid). One goat kid given 10 ml PBS only by the oral route served as a control. Cryptosporidium oocysts were detected in the faeces of one infected kid on day 3 post-inoculation (pi) whereas in the other 6 days pi. The faecal oocyst counts gradually increased and the peak counts in the two kids were 2 x 10(6)g(-1) (on day 12 pi) and 3.2 x 10(6)g(-1) (on day 14 pi). The increase in faecal oocyst output coincided with diarrhoea in an infected kid from days 10-17 pi. Although the oocyst excretion declined gradually after the peak, both infected kids excreted oocysts until euthanized on days 20 and 22 pi. Light and scanning electron microscopic investigations of the ileum revealed the endogenous stages on the brush border of the enterocytes, infiltration of neutrophils and mononuclear cells into the lamina propria, atrophy, stunting and fusion of villi. For purposes of comparison, goat Cryptosporidium oocysts were inoculated orally (10(3) oocysts/mouse) to eight, 1-week-old mice. All experimental mice excreted oocysts from day 3 pi, and four infected mice continued to excrete oocysts up to day 42 pi. The experimental infection described in goat kids resembled the natural disease in terms of oocyst excretion, clinical signs and intestinal pathology. The ability of oocysts excreted by asymptomatic goats, to infect goat kids and mice is likely to have a major impact on the epidemiology of cryptosporidiosis in livestock and man.     

Efficacy of azithromycin dihydrate in treatment of cryptosporidiosis in naturally infected dairy calves

The objective of this study was to evaluate the therapeutic efficacy of azithromycin treatment of cryptosporidiosis in naturally infected calves under field conditions. Fifty Holstein calves with cryptosporidiosis infection were divided into 5 groups: 1 group (10 calves) was unmedicated and served as the control group and was given distilled water only, whereas the other groups (10 animals per group) were medicated orally with azithromycin at the doses of 500 (group 1), 1,000 (group 2), 1,500 (group 3), and 2,000 mg (group 4) PO once daily for 7 days. The animals were examined clinically and fecal samples were collected on the 1st (inclusion day), 7th, 14th, and 21st days of the study. Drug efficacy was assessed by evaluating diarrhea, oocyst shedding, and weight gains from days 1 to 21 (4 assessments). Significant differences were observed in reductions of oocyst shedding (P < .05) and the fecal diarrhea incidence (P < .05) in groups 3 and 4 when compared with groups 1 and 2 and the control group. Weight gain of medicated calves was significantly higher than that of the unmedicated calves throughout the study (P < .05). The drug significantly suppressed oocyst shedding and resulted in significant improvements in clinical signs. Therefore, this suppression may have significant effect on the reduction of environmental contamination by cryptosporidial oocysts. From the economic point view, authors suggest that the most effective dose of azithromycin for the treatment of cryptosporidiosis in calves should be at 1,500 mg/d for 7 days.     

Analysis of the kinetics, isotype and specificity of serum and coproantibody in lambs infected with Cryptosporidium parvum

Enteric cryptosporidiosis was studied in colostrum-deprived lambs each infected at five days old with 10(6) oocysts. The prepatent period was three to five days and faecal oocyst concentration fell below detectable levels by day 16 after infection. Specific IgA, the only isotype detected by immunofluorescent assay in faecal extracts from infected lambs, was first evident on day 10 and titres continued to rise until day 16 of infection in association with declining oocyst output. Specific IgM and IgG antibodies were first detected in serum seven days after infection. No specific antibody was detected in uninfected control lambs. Immunoblotting methods showed that serum antibody and faecal IgA had similar profiles of antigen recognition. Antigens with approximate molecular weights of 180,000, 23,000 and 15,000 were consistent features on immunoblots performed with convalescent sera and faecal extracts. The results suggest that specific IgA in intestinal secretions has an important role in immunity to cryptosporidiosis.     

Field trial on the therapeutic efficacy of paromomycin on natural Cryptosporidium parvum infections in lambs

The objective of this study was to evaluate the therapeutic efficacy of paromomycin against cryptosporidiosis in naturally infected lambs under field conditions. The 36 cross-bred neonatal lambs, 3-10 days old, were used. On the first day that lambs showed diarrhea (Day 1) they were randomly divided into three groups. The infected control group (14 lambs) remained unmedicated whereas the two other groups were orally medicated with paromomycin solution (Humatin((R)), Parke Davis, France): 12 lambs (Group A) at 100mg/kg per day for three consecutive days (Days 1-3) and 10 lambs (Group B) at 200mg/kg per day for two days (Days 1 and 2). Drug efficacy was assessed by evaluating the presence of diarrhea, oocyst shedding and weight gains from Days 1 to 23. The results show the efficacy of paromomycin in reducing both cryptosporidial oocyst output and severity of clinical signs. On Day 4, all unmedicated lambs remained infected and excreted large numbers of cryptosporidial oocysts (mean score: 2.5) whereas oocyst output had stopped in most medicated lambs (>60%) and low numbers of oocysts were excreted in the remaining lambs (mean score: 0.45 in Group A and 1 in Group B). Mean oocyst excretion was significantly reduced in medicated lambs from Days 2 to 5 (P<0.05). Treatment also reduced, but not completely prevented, clinical symptoms although diarrhea stopped in most medicated lambs just after drug withdrawal. The mean weight gains of Group A lambs were higher than that of unmedicated lambs throughout the study and statistically significant differences were found from Days 1 to 11 (1.99+/-0.81 versus 1.47+/-0.53) (P<0.05). By contrast, the growth rate of Group B lambs from Days 11 to 23 was impaired when compared with the two other groups (P<0.05) although no significant differences were found at the end of the study (Days 1-23).     

Methylprednisolone acetate immune suppression produces differing effects on Cryptosporidium muris oocyst production depending on when administered

At different times after inoculation with Cryptosporidium muris, infected CF-1 female mice were immunosuppressed with a single subcutaneous dose of methylprednisolone acetate (MPA; 600 mg/kg). MPA immunosuppression decreases circulating CD3, CD4 and CD8 T-lymphocytes and B-lymphocytes by greater than 90% for approximately 14 days with numbers not returning to pre-suppression levels until after 41 days post-suppression. Immunosuppression was initiated at selected times before, during, and after oocyst production. Immunosuppression initiated prior to oocyst production delayed the start of production by 4-5 days and extended oocyst shedding by 16 days. Initiation of immunosuppression during oocyst production both extended oocyst shedding and greatly increased the number of oocysts shed per day over most of the extended shedding period. Immunosuppression during the decline of oocyst production resulted in only a moderate extension of shedding and a moderate increase in oocyst numbers. Immunosuppression initiated soon after oocyst shedding had ceased resulted in the re-initiation of limited oocyst production for only a few days. Suppression initiated on days 40 and 46 post-infection, 11 and 17 days after oocysts could no longer be detected in the feces, did not result in a resumption of oocyst production. In all cases, where oocyst production was extended or reinitiated, the shedding of oocysts halted between days 45 and 53 post-oocyst inoculation. These studies demonstrate that the effect of MPA immunosuppression depends on the immunologic conditions existing in the host at the time immunosuppression was initiated. Immunosuppression initiated during oocyst production allows an overwhelming parasitism to exist, implying that T- and B-lymphocytes play an important role in moving the host immune process along during this period of the infection. Conversely, severe suppression of T- and B-lymphocytes initiated as oocyst production is decreasing does not result in a complete relapse of the disease suggesting that T- and B-lymphocytes are not critical to the continuation of the immune process after this point. These studies also show that the C. muris infection persists beyond the end of the detection of oocysts in the feces.     

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.     

Performance and oocyst shedding in broiler chickens orally infected with Cryptosporidium baileyi and Cryptosporidium meleagridis

To study effects of experimental cryptosporidiosis, broiler chickens were infected per os with 5 x 10(5) oocysts of Cryptosporidium baileyi and Cryptosporidium meleagridis. In the first experiment, chickens were infected with oocysts of C. baileyi at the age of 7, 14, and 21 days. In the second experiment, chickens were infected with oocysts of C. baileyi, C. meleagridis, or both cryptosporidial species at the age of 7 days. Although clinical signs of infection were apparent, neither final live weight nor mortality was significanty influenced in chickens infected with a single Cryptosporidium species. In chickens infected with C. meleagridis, the growth retardation was observed in the 2-wk period after infection. The compensatory growth, however, started when the oocyst shedding had ceased. The number of oocysts in excreta specimens of chickens infected with C. meleagridis was two to three times lower than in excreta of chickens infected with C. baileyi. Chickens infected with both C. baileyi and C. meleagridis (5 x 10(5) oocysts of each) had significantly lower final live weight and worse feed efficiency than chickens of other groups. Concurrent infection did not influence individual C. baileyi or C. meleagridis oocyst shedding.