Seroprevalence of Toxoplasma gondii infection in sheep and alpacas (Llama pacos) in Chile

Serum samples from 408 sheep from different regions of Chile and 447 alpacas (Llama pacos) from the north of the country were tested for Toxoplasma gondii antibodies. The indirect haemagglutination test (IHAT) was used in both species and the indirect immunofluorescence test (IIFT) was also used on the sheep samples in order to compare the performance of the tests in that species. In both tests, titers ≥1:16 were considered diagnostically significant. Sera from 49 sheep (12%) were positive to T. gondii antibodies by the IHAT. When using the IIFT, 114 sheep sera (28%) were positive. The different results obtained in sheep sera between the tests were significant (p<0.0001). No differences were observed between geographical locations or sex of the sampled sheep regarding serological detection of T. gondii antibodies in sheep. As expected, adult sheep showed higher T. gondii reactivity than young sheep (p=0.0008). The corrected prevalence of toxoplasmosis in alpaca was 16.3% (32 positive out of 447). The rather low prevalence in alpacas may be associated with their extensive management as well as the extreme climatic conditions of The Andes which apparently would not be favorable for the transmission of the parasite.     

Protection against Toxoplasma gondii brain cyst formation in mice immunized with Toxoplasma gondii cytoskeleton proteins and Lactobacillus casei as adjuvant

The aim of the study was to evaluate the protection generated in mice against Toxoplasma gondii brain cyst burden by vaccination with T. gondii cytoskeleton proteins using Lactobacillus casei as adjuvant. One hundred and sixty-eight NIH mice were randomly allocated into eight groups of 21 mice each. Animals were immunized as follows: in group 1 with Toxoplasma lysate antigen (TLA) in Freund's modified adjuvant, containing L. casei (FMA), in group 2 with Toxoplasma cytoskeleton proteins (TCPs) in FMA, in group 3 with FMA, in group 4 with phosphate buffered saline (PBS), in group 5 with L. casei dead by heath (Lc), in group 6 with Freund's complete adjuvant (FCA), in group 7 with TLA in FCA, and in group 8 with TCP in FCA. Mean brain cyst burden (±S.E.M.) was assessed in mice 8 weeks after challenge with T. gondii Me49 strain (20 cysts per mouse). The percentages of reduction in cyst burden per brain (P < 0.01) as compared with the group 4 (control: mean 3181 ± 97.5) were 77.25% (724 ± 98) in group 1, 88.02% (381 ± 97.5) in group 2, 38.92% (1943 ± 130.3) in group 3, 44.31% (1771.4 ± 102) in group 5, 59.28% (1295.2 ± 99.1) in group 7 and 55.69% (1409.5 ± 89.9) in group 8. In order of importance, the best protection was obtained in groups 2, 1, 7, 8, 5 and 3. Noticeably the mice inoculated with L. casei alone showed a significant reduction in T. gondii brain cysts (P < 0.01), while those animals treated with FCA alone did not. Additionally, IgM anti-T. gondii antibody levels, as determined by ELISA 2 weeks after challenge, were highest in group 2 (P < 0.01) than in the other seven groups. Results suggest that T. gondii cytoskeleton proteins with L. casei as adjuvant constitute a good anti-toxoplasmosis vaccine candidate.     

Prevalence and molecular characterization of bovine Cryptosporidium in Qazvin province, Iran

The aim of this study was to determine the prevalence, variability with host age, and the genotypes of species of Cryptosporidium in cattle from 15 dairy farms in Qazvin province, Iran. Fecal samples, collected from 272 cattle during May 2006 to December 2007, were characterized microscopically. Oocysts from 51 positive samples were analyzed using PCR assay of 18S SSU rRNA, restriction fragment length polymorphism (RFLP) and sequencing. We identified 72.6% of the positive samples as Cryptosporidium parvum, 17.7% as Cryptosporidium andersoni, 7.8% as Cryptosporidium bovis and 1.9% as a novel genotype of C. parvum possessing a single mutation on MboII restriction. An infection rate of 19.5% of C. parvum among 174 pre-weaned calves was significantly higher than the 3.1% among 98 post-weaned calves (P < 0.0006). This is the first report of C. bovis and the new subgenotype of C. parvum in Iranian cattle.     

Seroprevalence of Toxoplasma gondii and Neospora caninum in wild and domestic ruminants sharing pastures in Galicia (Northwest Spain)

The prevalence of antibodies to the protozoan parasites Toxoplasma gondii and Neospora caninum were investigated by the direct agglutination test (DAT) and cELISA, respectively, in 160 roe deer (Capreolus capreolus), 177 sheep and 178 cattle sharing pastures in Galicia (Northwest Spain). The seroprevalence for T. gondii was 13.7% in roe deer, 57% in sheep and 7.3% in cattle. The seroprevalence for N. canimum was 6.8%, 10.1% and 24.1% in roe deer, sheep and cattle, respectively. Statistically significant differences were observed between sheep and the other species for T. gondii and between cattle and the other ruminants for N. caninum. Only 19/515 animals were positive for both, T. gondii and N. caninum. Statistically significant differences were observed among different geographical areas for T. gondii but not for Neospora, seroprevalence being higher in the coastal area lower than in other areas. This study reveals a widespread exposure to T. gondii in Galician ruminants, and therefore, those species, particularly sheep, should be regarded as a potential source of infection for humans.     

Prevalence of anti-Toxoplasma gondii and anti-Neospora caninum antibodies in sheep from Mossoró, Rio Grande do Norte, Brazil

Toxoplasma gondii is an obligate intracellular parasite with a variety of hosts, responsible for reproductive problems and economic losses in sheep flocks. Neospora caninum was recently identified and its clinical presentation in sheep is similar to that of toxoplasmosis, which can cause repeated abortions, though less frequently in this species. In order to confirm the prevalence of these agents in the city of Mossoró, Rio Grande do Norte, Brazil, 409 serum samples from adult sheep (364 females and 45 males) were tested by the indirect immunofluorescence antibody test, using cut-off point at a dilution of 1:64 and 1:50 for T. gondii and N. caninum, respectively. From the 35 properties examined, 23 (65.7%) had at least one seropositive animal for T. gondii and six (17.1%) for N. caninum. The prevalence of seropositive animals for T. gondii was 20.7% and for N. caninum 1.8%. There was no association between the presence of the agent’s antibody and gender, reports of reproductive problems and presence of dogs and/or cats in the properties. T. gondii is well distributed and N. caninum has low prevalence in sheep and in the properties of the studied region.     

Toxoplasma gondii infection in domestic ducks, free-range and caged chickens in southern China

Toxoplasma gondii is widely distributed in humans and other animals including domestic poultry throughout the world, but little is known of the prevalence of T. gondii in chickens and ducks in People's Republic of China. In the present study, antibodies to T. gondii were investigated in 349 domestic ducks (Anas spp.), 361 free-range, and 244 caged chickens (Gallus domesticus) raised in commercial flocks in Southern China's Guangdong Province using the modified agglutination test (MAT). Antibodies to T. gondii (MAT titer of 1:5 or higher) were found in 56 (16%) of 349 ducks, 41 (11.4%) of 361 free-range, and 10 (4.1%) of 244 caged chickens. The results indicate soil contamination due to T. gondii oocysts because free-range chickens feed from the ground, and suggest that the meat from the domestic poultry may be an important source for human infection by T. gondii in People's Republic of China.     

Associations of Neospora caninum seropositivity with gestation number and pregnancy outcome in Danish dairy herds

The prevalence and distribution of seropositivity towards the protozoan parasite Neospora caninum were studied in single blood samples from 1561 cows from 31 Danish dairy herds. Blood samples were analysed by an indirect enzyme-linked immunoassay and an indirect fluorescent-antibody test. Seroprevalence in 15 herds with previous abortions assigned to neosporosis ranged from 1% to 58%, with a mean frequency of 22%. In eight out of 16 herds without a history of N. caninum related abortions, no seroreactors were found. In the remaining eight herds, the seroprevalence ranged from 6% to 59%. The prevalence and distribution of seropositivity, gestation number prior to sampling, and breed were related to abortions and perinatal deaths using a random-effects logistic-regression model. Abortion risk was significantly increased in seropositive animals (OR=3) and in 2nd-gestation cows (OR=3). Perinatal death was significantly influenced by gestation number and breed, but not by serostatus. Reproductive performance and culling risk of cows were not affected by serostatus. Seropositivity increased with “age” (i.e. gestation number) (P=0.02). In open cows, seropositivity tended to decrease with distance from calving (P=0.05). The proportion of seropositive pregnant cows increased with trimester (P=0.02).     

Effects of crossbreed pregnancies on the abortion risk of Neospora caninum-infected dairy cows

Previous studies have shown that the use of beef bull semen significantly reduces the rate of abortions due to Neospora caninum in artificially inseminated (AI) seropositive dairy cows. In addition, certain beef breeds could be more resistant to N. caninum infection and abortion than others. The aim of the present study was to determine whether different crossbreed pregnancies, those derived from Limousin, Charolais, Piedmontese or Belgian Blue semen, carry different risks of abortion in Neospora-infected dairy cows. The effects of possible interactions between maternal levels of N. caninum antibodies and the different breed crosses were also evaluated. The study was performed on five commercial Holstein–Friesian dairy herds in Northeast Spain with previously confirmed diagnoses of N. caninum infection in aborted foetuses. The study population was comprised of 1115 pregnancies: 633 pregnancies recorded after AI using Holstein–Friesian semen from 18 bulls and 482 after AI using beef semen from 27 bulls (304 inseminations using semen from Limousin bulls, 191 from Belgian Blue bulls, 89 from Piedmontese bulls and 49 from Charolais bulls). Abortion rates were 32.2% (155/482) and 15.2% (96/633) for seropositive cows inseminated with Holstein–Friesian and beef breed semen, respectively. Logistic regression analysis revealed the herd and the interaction between maternal N. caninum antibody titre and the different crossbreeds as significant factors affecting the abortion rate. Lowest abortion rates, similar to that shown by seronegative animals in the analysed herds (3.2%, 239/7432), were observed in dams AI using Limousin semen that had low (<30 relative index (RI) units) N. caninum antibody titres (2.1% abortion, 3/145) and these cows were used as reference. Compared to the cows used as reference, cows with low N. caninum antibody titres (<30 RI units) showed a similar risk of abortion when inseminated with Piedmontese or Charolais bull semen, but higher risk of abortion when inseminated with Holstein (17.9 times) or Belgian Blue (7.2 times) bull semen. All cows with high N. caninum antibody titres (≥30 RI units) had a higher risk of abortion, ranging from 8.9 times (cows inseminated with Limousine semen) to 37.8 times (cows inseminated with Piedmontese semen), compared to the cows used as reference. In conclusion, different crossbreed pregnancies carried different abortion risks in Neospora-infected dairy cows. The use of beef bull semen dramatically reduced the risk of abortion in dairy cows, especially if Limousin breed semen was used. Moreover, this reduction was found to be dependent on the N. caninum antibody titre such that the lowest incidence of abortions was recorded in Limousin semen inseminated cows with low antibody titres. Insemination of Neospora-seropositive cows with beef bull semen could both reduce the risk of abortion and avoid breeding replacements for infected cattle.     

Peripheral white blood cell counts throughout pregnancy in non-aborting Neospora caninum-seronegative and seropositive high-producing dairy cows in a Holstein Friesian herd

Pregnancy is characterized by transient changes in the maternal immune system, also evident at peripheral level. The present study analyzes the kinetics and possible factors affecting peripheral white blood cell populations throughout pregnancy in a herd of high-producing dairy cows chronically infected or not with Neospora caninum. We examined 54 pregnant parous cows: 29 Neospora-seronegative and 25 Neospora-seropositive cows. Blood samples were collected on Days 90, 120, 150, 180 and 210 of gestation. General Linear Model (GLM) repeated measures analysis of variance showed that the interaction Neospora-seropositivity × parity significantly affected total leukocyte, neutrophil and monocyte counts with lower levels of total leukocytes, lower neutrophil and higher monocyte counts recorded in primiparous Neospora-seropositive cows. In addition, N. caninum-seropositive cows had significantly increased monocyte counts on Day 180 of gestation compared to seronegative ones. Other factors significantly associated with changes in total and/or differential leukocyte profiles were period of pregnancy, season, twin pregnancy and milk production. In conclusion, a parity-associated effect of chronic N. caninum infection was observed on peripheral blood cell profiles in dairy cattle during gestation.     

Prevalence and associated management factors of Cryptosporidium shedding in 50 Swedish dairy herds

Cryptosporidium parvum is a protozoan parasite causing diarrhoea in young calves. This cross-sectional study was performed to estimate the prevalence of Cryptosporidium infected herds in a sample of Swedish dairy herds and to identify potential risk factors associated with shedding of oocysts. Fifty dairy herds, selected by stratified random sampling, were included. The herds were visited once during the indoor seasons of 2005–2006 and 2006–2007. Faecal samples were collected from 10 calves, 10 young stock and 5 cows in each herd. Clinical observations of sampled animals and environmental status were recorded, and farmers were interviewed about management procedures. Faecal samples were cleaned by sodium chloride flotation and detection of oocysts was made by epifluorescence microscopy. Cryptosporidium parvum-like oocysts were found in 96% of the herds. Prevalence was 52% in calves, 29% in young stock and 5.6% in cows. Three two-day-old calves shed oocysts. Cryptosporidium andersoni was found in seven animals from four different herds. Factors associated with prevalence of shedders among sampled animals in a herd were age at weaning, cleaning of single calf pens, placing of young stock, system for moving young stock, and year of sampling. Factors associated with shedding in calves were age, placing of young stock, routines for moving young stock and time calf stays with the cow. The only significant factor in young stock was age. In cows, number of calves in the herd and type of farming (organic vs. conventional) affected shedding.     

Preventive and therapeutic efficacy of halofuginone-lactate against Cryptosporidium parvum in spontaneously infected calves: a centralised, randomised, double-blind, placebo-controlled study

The preventive and therapeutic efficacy of halofuginone-lactate (HFL) against Cryptosporidium parvum was evaluated in a study conducted from November 2004 to March 2005 on a dairy farm in Central Bohemia, Czech Republic, using 260 spontaneously infected calves. HFL (0.1 mg/kg/day) was administered orally for 7 days to 1-day-old and 8-day-old calves, respectively. In both treated groups the drug significantly, and in almost the same manner, decreased the intensity of diarrhoea (P<0.001) and faecal oocyst count (P<0.001) when compared to corresponding placebo groups. The only difference between both treated groups was the time of onset of symptoms of the infection. Over time, the clinical pattern of cryptosporidiosis in the animals treated at 8-14 days of age was similar to that seen in the groups receiving the placebo. In contrast, infection in the preventively treated group peaked about 10 days later but with the same intensity. The results of this study confirm the anticryptosporidial activity of HFL in calves, but show that the outcome of infection following preventive treatment is comparable to that observed in calves treated after the onset of symptoms.     

新疆和静县黑头羊犬新孢子虫的检测及分子鉴定

犬新孢子虫是一种细胞内原生动物寄生虫,可导致包括绵羊和山羊在内的多种哺乳动物流产和新生胎儿死亡。本研究为了预估巴音郭楞蒙古自治州和静县黑头羊中犬新孢子虫的流行情况,采集了8 个乡镇的165 份血样,运用聚合酶链式反应（PCR）扩增犬新孢子虫Nc5基因片段,并对部分阳性产物进行基因测序。结果显示,在165 只黑头羊中有13 只（7.9%）检测到犬新孢子虫阳性;年龄、流产史、饲养犬只与感染率有统计学差异（P< 0.05）;2 岁以上的母羊感染率最高,有流产史的母羊,新孢子虫感染率较高。饲养场地周边犬的存在对感染率也有所影响,有饲养犬的羊群感染率11.4%,高于未饲养犬的羊群。将Nc5 基因的部分序列与GenBank 上传的基因序列进行比对,结果显示同源性为92%~100%。本研究是首次对和静县黑头羊犬新孢子虫进行系统进化分析,为犬新孢子虫病防控方案的制订提供参考,对减少畜牧业的经济损失具有实际意义。     

Assessment of enzyme linked immunosorbent assay based diagnostic kits (Toxokit-G and Toxokit-M) for the detection of IgG and IgM antibodies to Toxoplasma gondii in human serum

An enzyme linked immunosorbent assay (ELISA) using penicillinase was developed in the form of diagnostic kits (Toxokit-G and Toxokit-M) for the detection of IgG and IgM antibodies to Toxoplasma gondii. The performance of both the kits was compared with commercially available diagnostic kits, i.e. Enzygnost-Toxoplasmosis/IgG (Behring Co., Germany), TOXOTEK-G (Flow Lab., U.K.) and Toxoplasma IgM Microassay (Diamedix Corp., U.S.A.) by testing toxoplasma-suspected human serum samples. The results indicate a good reliability between these diagnostic kits. Toxokit-G has 86.66 and 96.05% sensitivity and specificity respectively. The main advantage of Toxokit-G is that the end result can be assessed visually without using sophisticated instruments. Toxokit-M has 100% sensitivity and specificity and test results were not affected by the presence of antitoxoplasma IgG antibodies, rheumatoid factor or antinuclear antibodies.     

Seroprevalence of antibodies to Encephalitozoon cuniculi and Encephalitozoon intestinalis in humans and animals

The presence of antibodies against Encephalitozoon cuniculi (E. cuniculi) and Encephalitozoon intestinalis (E. intestinalis) was examined in 215 samples from humans and in 488 samples from five different species of domestic and companion animals in Slovakia. The 215 human samples and samples from 90 swine, 123 non-infected cattle (cattle), 24 cattle infected with bovine leukosis virus (BLV-positive cattle), 140 sheep and 111 dogs were examined by the enzyme-linked immunosorbent assay (ELISA). Samples with serum titres 1:200 or higher were considered as positive. Specific anti-E. cuniculi antibodies were found in humans (0.9%), swine (52%), cattle (2%), sheep (9%) and dogs (15%) except for the BLV-positive cattle at the titre of 1:200. The titre of 1:400 was detected only in humans (0.5%). The presence of specific anti-E. intestinalis antibodies at the titre of 1:200 was confirmed in humans (6%), swine (51%), cattle (11%), BLV-positive cattle (13%) and dogs (6%) but not in sheep. The anti-E. intestinalis antibodies reached the 1:400 in humans (1%), swine (4%) and BLV-positive cattle (17%). The presence of specific anti-E. intestinalis antibodies at the titre of 1:600 was observed only in one swine (1%). Significant differences were observed in animals at titres 1:200 and 1:400 (chi-squared test: p < 0.0001) for both pathogens and in humans only for E. cuniculi at the titre of 1:400 (chi-squared test: p < 0.0075).     

Prevalence and age-related infection of Cryptosporidium suis, C. muris and Cryptosporidium pig genotype II in pigs on a farm complex in the Czech Republic

A total of 413 pig faecal samples were collected from pre-weaners (119), starters (131), pre-growers (123) and sows (40) from a farm with a closed breeding system segmented into two breeding complexes and a growing complex in the region of Vysočina, Czech Republic and screened for the presence of Cryptosporidium using staining methods and genotyping (SSU rRNA). Cryptosporidium oocysts were detected by microscopy in the faeces of 21.1% of the samples (87/413). Sequence analyses and RFLP identified C. suis in 44, Cryptosporidium pig genotype II in 23 and C. muris in 2 samples. No mixed infections were found.Pigs under 7 weeks of age were infected with C. suis only. Cryptosporidium pig genotype II was found in animals from 7 weeks of age. No relationship was found between diarrhoea and any Cryptosporidium infection in any of the different age groups (P < 0.05). The pre-weaned pigs shed significantly more Cryptosporidium oocysts than older pigs and it was associated with C. suis infection.     

Prevalence of vancomycin-resistant Enterococcus faecium (VREF) in pig faeces from slaughterhouses in Spain

A study to estimate the prevalence of vancomycin-resistant Enterococcus faecium in faecal samples from pigs at slaughterhouses in Spain was carried out between November 1998 and January 1999 with 900 samples taken from four abattoirs representing 9.7% of all pig slaughtered in 1998. Using a selective enrichment broth with vancomycin (8 μg/ml), 64 samples (7.1%; 95% CI: 5.5, 9.0%) had E. faecium vancomycin-resistant strains that showed minimal inhibitory concentrations of 256 μg/ml (62 strains) and 512 μg/ml (two strains). Results by farm showed that 43 of the 240 pig farms represented in the sampling had at least one faecal sample with vancomycin-resistant E. faecium.     

Seroprevalence and risk factors associated with Toxoplasma gondii infection in pig farms from Catalonia, north-eastern Spain

Seroprevalence and associated risk factors for Toxoplasma gondii infection in pigs were analyzed in 1202 sera samples, including sows and pigs of three, seven, 11, 15 and 20 weeks of age, from 23 farms in Catalonia, north-eastern Spain. Antibodies were tested by the modified agglutination test (MAT) at titers ?1:25. Antibodies to T. gondii were found in 228 samples (19.0%; 95% CI: 16.8-21.2). The individual prevalence in animals higher than 7 weeks of age was 22.8% (174/762; 95% CI: 16.6-29.0) and the within-farm prevalence ranged from 7.1% to 36.4%. Statistically significant differences were found among age classes. The risk factors significantly associated with T. gondii seroprevalence were the presence of cats, percentage of mortality at weaning and the presence of outdoor facilities in the farms. The seroprevalence observed in the present study indicates widespread exposure to T. gondii among domestic pigs in Catalonia, which may have important implications for public health.     

弓形虫细胞核因子3基因的克隆及序列分析

为预测弓形虫细胞核因子3(TgNF3)基因作为抗弓形虫疫苗候选抗原的可能性,本试验以弓形虫RH株的总RNA为模板,应用逆转录-聚合酶链式反应(RT-PCR)扩增TgNF3基因并连接至pMD18-T载体。筛选阳性克隆后,测序鉴定。将测序结果正确的核苷酸序列翻译成氨基酸序列,运用生物信息学软件分析TgNF3蛋白的结构,预测抗原表位。结果显示,TgNF3基因长约950bp。TgNF3蛋白中包含1个跨膜结构域,9个α-螺旋区,4个β-折叠区,8个亲水区域和10个柔性区域,并预测含有9个线性B细胞抗原表位。结果表明,TgNF3蛋白可能具有良好的免疫原性,为研制以TgNF3为抗原的弓形虫亚单位疫苗及表位疫苗奠定了基础。     

The relationship of soil province to molecular characterization and phylogenetic analysis of Cryptosporidium parvum isolated from calves in Georgia

Although Cryptosporidium spp. are found throughout the world and in multiple environmental conditions, few data are available that explore the possibility of an association between specific environmental parameters and the species or strain of Cryptosporidium. This study examines the potential association between a particular Cryptosporidium species/strain found in calves and soil provinces in Georgia, USA. Necropsy cases spanning the years 1996-2002 were tested. No significant differences (P=0.962, chi(2) test of homogeneity) between numbers of positive cases were noted among soil provinces. Phylogenetic analysis of the sequences for the PCR products revealed sequence similarity of the products with Cryptosporidium parvum strain C1. Although, clinical Cryptosporidiosis in calves was not found to be affected by soil province and may be caused by a single genotype, other genotypes may be responsible for subclinical infection and warrant further investigation.     

The first report of Gyrodactylus salaris Malmberg, 1957 (Platyhelminthes, Monogenea) on Italian cultured stocks of rainbow trout (Oncorhynchus mykiss Walbaum)

The monogenean Gyrodactylus salaris Malmberg, 1957 is considered one of the most important parasites of wild salmonids in the European Community due to the heavy ecological and economical damage it has inflicted on Atlantic salmon (Salmo salar) parr populations. Rainbow trout (Oncorhynchus mykiss) is susceptible to G. salaris and can act as a suitable carrier host and, consequently, its trade in EU territory is restricted in relation to the status of “recognized free” zones. Despite the economic importance of rainbow trout farming in Italy, information on the Italian gyrodactylid fauna is lacking and prior to this study, G. salaris had not been officially reported. During a routine health examination of farmed rainbow trout stock throughout Central and Northern Italy in 2004–2005, five fish farms were found to be infected with G. salaris alongside three other gyrodactylids. Morphological and molecular characterisation confirmed the presence of G. salaris, Gyrodactylus teuchis Lautraite, Blanc, Thiery, Daniel et Vigneulle, 1999 and Gyrodactylus derjavinoides Malmberg, Collins, Cunningham et Jalali, 2007, while Gyrodactylus truttae Gläser, 1974 was identified by morphological analysis only. The findings from this study extend the distribution of G. salaris within Europe and highlight the importance of the rainbow trout trade in its dissemination.