Prevalence of Giardia duodenalis genotypes in pre-weaned dairy calves

To determine the prevalence of Giardia genotypes in pre-weaned dairy calves, fecal samples were collected from a minimum of 18, 1-7-week-old dairy calves per farm on two farms each in the states of Vermont, New York, Pennsylvania, Maryland, Virginia, North Carolina, and Florida. Samples cleaned of fecal debris and concentrated using CsCl density gradient centrifugation were stained and examined by immunofluorescence microscopy and also subjected to PCR and gene sequence analysis. Prevalence by PCR ranged from 9% on a farm in Pennsylvania to 93% on a farm in Vermont, with an average prevalence for 407 calves on 14 farms of 40%. Gene sequence analysis of the TPI, beta-giardin and 16S rRNA genes revealed 85% of the positive samples to be Assemblage E, while 15% were Assemblage A, although the percentages of these genotypes varied greatly from farm to farm. Some farms had no Assemblage A Giardia. Thus, while a majority of the calves were infected with a genotype that is not known to be infectious for humans, calves on 7 of 14 farms did harbor Assemblage A Giardia. Calves should be considered as a potential source of human infectious cysts in the environment.     

Prevalence and genotypes of Giardia duodenalis in post-weaned dairy calves

To determine the prevalence of Giardia genotypes in post-weaned dairy calves, fecal specimens were collected from 3 to 11-month-old dairy calves per farm on two farms in Vermont, New York, Pennsylvania, Maryland, Virginia, North Carolina, and Florida. Specimens cleaned of fecal debris and concentrated using CsCl density gradient centrifugation were stained and examined by immunofluorescence microscopy and also subjected to PCR and DNA sequence analysis. Overall, PCR provided more sensitive detection than IFA. Prevalence of Giardia infection, as detected by PCR ranged from 20% on NC-2 to 81% on VT-2, with an overall prevalence of 52% (237 positive samples out of 456 total). DNA sequence analysis of the 16S rRNA gene revealed 87% of the 237 Giardia isolates were Assemblage E, and 13% were Assemblage A although the prevalence of these genotypes varied greatly from farm to farm, with five farms having no Assemblage A Giardia. Therefore, Assemblage E was present in 45% of all animals tested and Assemblage A was present in 7% of the animals. Thus, while many of the calves were infected with a genotype that is not known to be infectious for humans, post-weaned calves on nine of 14 farms did harbor Assemblage A Giardia. Therefore calves should be considered as a potential source of human infectious cysts in the environment, with some farms representing a much higher risk than others.     

Prevalence and genotyping of Giardia duodenalis from beef calves in Alberta,Canada

Giardia infections in domestic cattle has come under increasing scrutiny owing to the potential contamination of surface and ground waters through manure distribution on fields and pasture runoff. The objective of the study was to determine the prevalence and genotypes of Giardia duodenalis in beef calves in major beef cow calf farms in Alberta, Canada. Fecal samples were collected from beef calves aged 2-10 weeks at nine farms in Alberta. Samples were examined for the presence of G. duodenalis cysts by immunofluorescent staining. Giardia cysts were found in 168 of the 495 fecal samples examined, with prevalence ranging from 7 to 60% among farms. Genotypic analysis of positive isolates utilizing PCR and sequencing of a 292 bp fragment of the 16S-rRNA locus, revealed the hoofed livestock genotype in 41 of the 42 isolates. One isolate was identical to the Assemblage A genotype. The results of this study demonstrate that beef calves in this area are primarily infected with the livestock genotype which is thought to be specific to artiodactyl hosts and non-infective to humans. This suggests that the Giardia carried by beef cattle may be a minimal zoonotic threat.     

Severe weight loss in lambs infected with Giardia duodenalis assemblage B

An outbreak of giardiasis was observed in a sheep farm in Central Italy. Infected lambs (30-90 days of age) showed a malabsorption syndrome, decreased weight gain and impairment in feed efficiency. The most relevant clinical sign was the excretion of malodorous and poorly formed faeces, whereas diarrhoea was rarely observed in the flock. Laboratory investigations revealed the presence of Giardia in affected animals, while no other significant viral, bacterial or parasitic pathogens were identified in faeces or tissue samples. A mild to severe infiltrative enteritis with eosinophils, lymphocytes and plasma cells was detected in histological sections of the gut. Giardia parasites collected from duodenal aspirates were typed as Giardia duodenalis Assemblage B, by PCR amplification and sequencing of the TPI gene. Treatment with fenbendazole at a dose of 10mg/kg for 3 consecutive days, successfully cleared the infection. These results show that G. duodenalis can cause significant economic losses in sheep farming.     

Mixed Giardia duodenalis assemblage A, B, C and E infections in pet chinchillas (Chinchilla lanigera) in Flanders (Belgium)

Fourteen isolates of Toxoplasma gondii were isolated from cats from 4 different geographic provinces (Anhui, Hubei, Shanxi and Guangdong) in China and their genetic diversity with 8 nuclear loci SAG2, SAG3, BTUB, GRA6, L358, PK1, c22-8, c29-2, and an apicoplast locus Apico, was analysed by restriction fragment length polymorphisms (RFLPs). Two genotypes from these 14 isolates were identified but none of them belongs to the typical genetic types (types I, II and III). It is unexpected that such high similarity was observed in these 14 isolates although their original regions are significantly distant. Our results strongly indicate that the three traditional clonal lineages of types I, II and III of this parasite may not be preponderant in China. In addition, our results show that the genotypes of T. gondii in China may be highly clonal with atypical genotypes and higher virulence.     

牛十二指肠贾第虫的分子流行病学研究进展

贾第虫是一类在全球范围内广泛分布的寄生虫,包括7个虫种。其中的十二指肠贾第虫（Giardia duodenalis）是一种重要的肠道寄生虫,能感染人和大多数哺乳动物,可引起腹泻、营养不良和体重减轻等症状。分子分型工具的发展促进了贾第虫检测、基因分型和溯源的发展,大大改变了人们对贾第虫人兽共患潜力的理解。利用分子分型工具可将十二指肠贾第虫分为8种集聚体（A~H）,8种集聚体的宿主范围都存在差异,其中,集聚体A和B是人兽共患型。牛是十二指肠贾第虫的重要宿主,但目前对牛十二指肠贾第虫病的分子流行病学认识不够,其公共卫生意义也一直被忽视。本文汇总了国内牛十二指肠贾第虫的分子流行病学调查结果。结果发现,我国牛十二指肠贾第虫的感染是普遍存在的,其中,集聚体E为优势集聚体,集聚体A和B呈散发流行。近年来,一些国家出现了集聚体E感染人的报道,同时,集聚体A在牛中的感染率有上升的迹象,牛十二指肠贾第虫的人兽共患潜力正在逐步得到认识。     

Molecular characterization of potentially zoonotic isolates of Giardia duodenalis in horses

Giardia isolates from eight horses from New York State (NY), USA and two horses from Western Australia (WA) were genetically characterized at the SSU-rDNA and triose-phosphate isomerase (TPI) genes. Phylogenetic analysis of the TPI gene provided strong support for the placement of both isolates of Giardia from horses in WA and a single isolate from a horse in NY within the assemblage AI genotype of G. duodenalis. Another two isolates from horses in NY placed within the assemblage AII genotype of G. duodenalis. Phylogenetic analysis of the TPI gene also provided strong bootstrap support for the placement of four G. duodenalis isolates from horses in NY into a potentially host-specific sub-assemblage of assemblage BIV. The results of this study are consistent with previous studies showing that assemblages AI and AII of G. duodenalis provide the greatest potential zoonotic risk to humans. Horses may therefore constitute a potential source for human infection of Giardia either directly or via watersheds.     

长春狐狸、貉和家兔养殖场十二指肠贾第虫基因型分析

为了解长春地区狐狸、貉和家兔养殖场的十二指肠贾第虫感染情况,基于十二指肠贾第虫GDH基因位点分别设计2对引物,对养殖场的120份狐狸粪便样品、60份貉粪便样品和148份家兔粪便样品十二指肠贾第虫进行检测和基因型分析。结果表明,在来自狐狸的粪便中检测出阳性样品44份,阳性率为36.7%,基因型均为基因亚型AI;6份貉阳性样品,阳性率为10.0%,基因型均为集聚体D;41份家兔阳性样品,阳性率为27.7%,其中4份是集聚体B,其他是集聚体A中的基因亚型AI。结果发现在长春地区貉感染贾第虫集聚体D,长春地区的狐狸感染人兽共患型的集聚体A,家兔感染人兽共患型的集聚体A和B。研究结果为该地区十二指肠贾第虫病的防控提供了参考。     

广东省猪十二指肠贾第虫感染现状和分子特征分析

【目的】 了解广东省不同地区猪源十二指肠贾第虫(Giardia duodenalis,简称贾第虫)的流行现状和分子特征,并评估其人兽共患风险。【方法】 从广东省10个地区采集新鲜猪粪样品,采用显微镜镜检,并应用巢式PCR对贾第虫谷氨酸脱氢酶(glutamate dehydrogenase,gdh)基因进行扩增,根据扩增结果计算不同地区和生长阶段猪贾第虫的阳性率。对所有的PCR阳性产物进行测序,将获得的贾第虫gdh基因序列进行BLAST比对,确定贾第虫的基因型;运用Mega 7.0软件的最大似然法构建进化树。【结果】 贾第虫包囊呈椭圆形,囊壁较厚,大小为(10~14)μm ×(7~10)μm,具有明显纵向轴柱,细胞核分布于轴柱两侧。521份受检样品中共检测出94份PCR阳性样品,阳性率为18.04%(94/521),其中茂名和清远的阳性率较高,分别达40.00%(10/25)和35.06%(27/77);肇庆和韶关的阳性率较低,分别为3.90%(3/77)和8.57%(3/35);佛山、江门、阳江、河源、广州和惠州的阳性率分别为11.11%(2/18)、15.31%(15/98)、11.43%(4/35)、13.33%(4/30)、9.38%(3/32)和24.47%(23/94)。不同饲养阶段的猪群的阳性率调查发现,母猪的贾第虫阳性率(24.55%)显著高于断奶仔猪(13.30%)(P<0.05),与育肥猪的贾第虫阳性率(19.23%)差异不显著(P>0.05)。基于gdh基因序列的基因分型和系统进化分析共发现5种贾第虫亚集聚体,即集聚体AⅠ和集聚体E的4种不同亚型,包括集聚体E1、E2、E10和E13,其中61.70%(58/94)测序阳性样品属于集聚体AⅠ,38.30%(36/94)属于集聚体E的不同亚型。【结论】 广东猪源贾第虫的感染较普遍,不同地区和不同生长阶段的猪感染率存在一定差异。其中,人兽共患性集聚体AⅠ为优势亚集聚体,表明广东省猪源贾第虫存在人兽跨物种传播的风险,应重视该病可能引起的公共卫生问题。     

Passive immunity and serological immune response in dairy calves associated with natural Giardia duodenalis infections

In a previous study, Giardia infection patterns were studied in newborn dairy calves over a 4-month period. Chronic Giardia infections were observed in all calves with initial cyst excretion occurring at approximately 1 month of age. In the work presented here, the passive immunity and serological immune response associated with these Giardia infections were examined. Colostrum and milk samples were collected from the dams of these calves, and monthly serum samples were collected from each calf. The colostrum, milk and sera samples were analyzed by ELISA and Western blot for the presence of anti-Giardia IgG antibodies. In addition, the in vitro anti-Giardia activity of milk and colostrum was examined using a miniculture adherence assay. When examined by ELISA, mean anti-Giardia antibody titres were found to be significantly higher in colostrum compared to milk. The monthly mean serum antibody titres in the calves were not found to differ significantly at any time point during the study. Western blot analysis revealed that colostrum from the dams reacted strongly with many different Giardia antigens between 205 and 7.5kDa, while milk reacted with few antigens in the same size range. Sera collected from the calves when 30 and 60 days of age reacted with few Giardia antigens, but as the calves aged, IgG antibodies in their sera began to react with antigens of 21, 50, 65, 73 and 79kDa. The miniculture adherence assay demonstrated that colostrum had significantly more anti-Giardia activity in vitro compared to milk. These results suggest that the calves in this dairy did not mount a significant humoral immune response against Giardia following infection. However, colostrum contained a high level of anti-Giardia antibodies and exhibited anti-Giardia activity in vitro. Therefore, colostrum may have the potential to provide initial protection against Giardia infections in calves, but the lack of a strong, specific humoral immune response by these calves could account for the high prevalence and chronic duration of the infections.     

Molecular evidence for zoonotic transmission of Giardia duodenalis among dairy farm workers in West Bengal, India

No study in the past has examined the genetic diversity and zoonotic potential of Giardia duodenalis in dairy cattle in India. To assess the importance of these animals as a source of human G. duodenalis infections and determine the epidemiology of bovine giardiasis in India, fecal samples from 180 calves, heifers and adults and 51 dairy farm workers on two dairy farms in West Bengal, India were genotyped by PCR-RFLP analysis of the β-giardin gene of G. duodenalis followed by DNA sequencing of the nested PCR products. The overall prevalence of G. duodenalis in cattle was 12.2% (22/180), the infection being more prevalent in younger calves than in adult cattle. Zoonotic G. duodenalis Assemblage A1 was identified in both calves and workers although the most prevalent genotype detected in cattle was a novel Assemblage E subgenotype. These findings clearly suggest that there is a potential risk of zoonotic transmission of G. duodenalis infections between cattle and humans on dairy farms in India.     

Genotypic analysis of Giardia duodenalis in domestic cats

BACKGROUND: Giardia duodenalis is an intestinal flagellated protozoan that affects many mammalian species often causing severe diarrheal disease. Several different genotypes have been identified (Assemblages A-G). Most isolates recovered from domestic cats have been assigned to either Assemblage A, the zoonotic form of the parasite, or Assemblage F, identified thus far only in cats. Genotypic variation within G. duodenalis may influence clinical presentation and course of disease. Therefore, host-adapted genotypes may not be responsible for diarrheal disease (eg, Assemblage F in cats). HYPOTHESIS: Multiple Giardia genotypes will be present in domestic cats, including Assemblage F, which will not be correlated with clinical signs. ANIMALS: 250 domestic cats from eastern Mississippi and northwestern Alabama. METHODS: Prevalence survey. Fecal samples evaluated for cysts using a centrifugation concentration technique and a commercially available direct immunoflourescent antibody kit. Giardia isolates were characterized by PCR amplification and sequencing of the glutamate dehydrogenase gene. RESULTS: Both Assemblage A-I (6/17) and Assemblage F (11/17) were identified. Although Assemblage was significantly associated with age and housing, no association was detected between Assemblage and a variety of other factors including the presence of gastrointestinal signs (acute vomiting, diarrhea, and constipation). CONCLUSIONS AND CLINICAL IMPORTANCE: The presence of diarrhea in domestic cats with Giardia cannot be used as a predictor of the presence of zoonotic genotypes in animals within the study area. Although Assemblage A was associated with age and housing, veterinarians should consider any isolation of Giardia from domestic cats as potentially zoonotic.     

Laboratory diagnosis of Giardia duodenalis infection in dogs

Results of trichrome staining of fecal samples and intestinal contents preserved in polyvinyl alcohol fixative, fecal flotation utilizing unpreserved feces, and enzyme-linked immunosorbent assay of serum specimens were compared for the diagnosis of Giardia duodenalis infections in dogs. Trichrome staining of preserved fecal samples resulted in the identification of 44 (92%) of the 48 infected dogs from a group of 200 dogs. Trichrome staining of preserved intestinal contents resulted in the identification of 26 (54%) of the infected dogs, and fecal flotation resulted in the identification of 23 (48%) of the infected dogs. Giardia duodenalis antibodies were not detected consistently in the sera of infected dogs.     

Prevalence of Giardia duodenalis genotypes in adult dairy cows

The prevalence of Giardia duodenalis genotypes was determined in adult dairy cows. Fecal specimens were collected from two farms each in Vermont, New York, Pennsylvania, Maryland, Virginia, North Carolina, and Florida. Specimens, cleaned of fecal debris and concentrated using CsCl density gradient centrifugation, were subjected to PCR and DNA sequence analysis. The prevalence of G. duodenalis infection, ranged from 3% to 64%, with an average prevalence of 27% (144 positive cows out of 541 examined). DNA sequence analysis of the 16S rRNA gene revealed the presence of both Assemblage A and Assemblage E, G. duodenalis. Overall, Assemblage E was present in 25% of all animals tested and Assemblage A was present in 2% of the animals. As a percentage of G. duodenalis isolates, Assemblage E represented 94% and Assemblage A represented 6%. Although, most of the cows were infected with a genotype that is not known to be infectious for humans, adult cows on five farms did harbor varying levels of zoonotic Assemblage A, G. duodenalis. Therefore, although adult cows do not appear to be a significant source of human infectious cysts in the environment, the risk from this age group should not entirely be discounted.     

Prevalence and molecular characterization of Giardia duodenalis in dogs in Israel

Three hundred and two stool samples were collected from municipal shelters and owned dogs in different geographical locations in Israel from December 2016 to September 2017 and examined for Giardia and assemblage type by PCR targeting the 18S rRNA and β-giardin genes. Overall Giardia prevalence was 24.5 % (74/30). Giardia prevalence was 1.9-fold higher in dogs ≤ 6 months old compared to > 6 ≤ 12 months old and older dogs [25/61 (41 %), 18/73 (24.6 %) and 31/166 (18.7 %), respectively, (p = 0.001)], 2.3-fold higher in winter [32/90 (35.5 %)] compared to its prevalence during autumn [15/60 (25 %)], spring [10/62 (16.1 %)] and summer [17/89 (19.1 %), p = 0.003)], and 2.7-fold more frequent among diarrheic dogs [23/43 (53.4 %)] compared to those with formed stools [51/253 (20.1 %)], (p = 0.001)]. The Giardia sp. assemblages detected were C and D. Higher infection rates in young, diarrheic dogs, sampled during winter, and housed in municipal shelters, indicates the need for targeted preventive measures.     

Molecular identification of Cryptosporidium parvum and Giardia duodenalis in the Italian water buffalo (Bubalus bubalis)

Livestock are commonly infected with protozoan parasites of the genera Cryptosporidium and Giardia, and some of the species and genotypes found in these animals have zoonotic significance. We characterized isolates of both parasites recovered from the Italian water buffalo (Bubalus bubalis), an economically important species whose milk is used for the production of "buffalo mozzarella" fresh cheese. Molecular analysis of the Cryptosporidium small subunit ribosomal DNA gene and of the Giardia beta-giardin gene shows the presence of both zoonotic parasites (Cryptosporidium parvum and Giardia duodenalis assemblage A) and host-specific parasites (G. duodenalis assemblage E), suggesting that water buffaloes can contribute to environmental contamination with oocysts and cysts potentially infectious to humans if their faeces are improperly disposed of. On the other hand, mozzarella cheese is probably a safe product, given that its production involves the treatment of cheese curd at 85-95 degrees C, which is likely to kill or inactivate the parasites.     

Prevalence and risk factors of Giardia duodenalis in dogs from Romania

The protozoan Giardia duodenalis is a mammalian-infecting parasite that produces diarrhoea and malabsorption in its hosts. A survey to investigate canine infections with G. duodenalis in Romania was undertaken between June 2008 and December 2009. The objectives of the study were to (i) estimate the prevalence of infection in different dog populations (kennels, shelters, shepherd, household) using microscopy and a commercially available enzyme-linked immunosorbent assay (ELISA) test kit; (ii) to establish the level of agreement and characteristics of the tests; and (iii) to identify risk factors for infection by multivariate logistic regression models. Faecal samples were collected from 614 dogs aged from 1 month to 16 years (mean ± SD=2.88 ± 2.86 years). Each sample was tested for the presence of cysts using a flotation method with saturated sodium chloride solution and 416 out of 614 stool samples were further examined for the presence of G. duodenalis specific antigens using Giardia Microwell ELISA (SafePath? Laboratories). Giardia cysts were identified in 8.5% of total dogs (52/614) and statistical significantly more frequently in dogs living in communities. The cysts prevalence according with dog populations was as follows: 7.2%(9/125) in kennel dogs; 16.5%(27/164 in shelter dogs; 4.3%(2/46) in shepherd dogs; 4.8%(4/84) in household dogs from urban areas; and 5.1%(10/195) in household dogs from rural areas. The overall prevalence of Giardia infection by ELISA was 34.6% (144/416). The prevalence was significantly higher in kennel dogs (50%; 13/26), shelter dogs (47.7%; 74/155) and shepherd dogs (40.5%; 17/42) than in household dogs from urban areas (34.1%; 15/44) and household dogs from rural areas (16.8%; 25/149). It was noticed poor agreement between microscopy and ELISA (k=0.19). The microscopy performed best, with an Youden Index of 0.74, a Se of 73.68% and a Sp of 100%. ELISA had 100% Sp, but only 19.44% Se. Young dogs (up to 12 months age) and living in communities were identified as risk factors for infection by multivariate logistic regression analysis. 71.2% (37/52) Giardia cysts positive dogs presented co-infections with other intestinal parasites: Toxocara canis (14/52; 26.9%), Isospora ohioensis (12/52; 23.1%), Ancylostoma caninum (9/52; 17.3%), Uncinaria stenocephala (7/52; 13.5%), Trichocephalus vulpis (6/52; 11.5%), Hammondia heydorni/Neospora caninum (5/52; 9.6%), Sarcocystis spp. (5/52; 9.6%), Isospora canis (4/52; 7.7%), Capillaria aerophila (3/52; 5.8%), Strongyloides stercoralis (2/52; 93.8%), Dipylidium caninum (1/52; 1.9%) and Toxascaris leonina (1/52; 1.9%).     

Effects of supplementation on intake and growth of nursing calves grazing native range in southeastern North Dakota

A 2-yr study was conducted to determine the first limiting nutrient for gain in nursing calves grazing native range in southeastern North Dakota. Thirty-two calves (20 steers, 12 heifers) in Trial 1 (169 +/- 5 kg initial BW) and 31 (16 steers, 15 heifers) in Trial 2 (214 +/- 5 kg initial BW) grazed common pastures. Calves were blocked by sex and stratified by weight. Calves were stratified by age of dam in Trial 1 and by pretrial milk intake (MI) in Trial 2. Treatments were nonsupplemented control (CON); energy supplement (ENERGY; 100% soyhulls); degradable intake protein supplement (DIP; 68% soyhulls, 32% SBM); and degradable with undegradable intake protein supplement (DIP+UIP; 80% sulfite-liquor treated SBM, 16% feather meal, 4% blood meal). In Trial 2, 5% molasses was added to all supplements with the ratios of other ingredients held constant. Supplements were formulated to be similar in NE. The DIP and DIP+UIP supplements supplied equal amounts of degradable protein. Supplemented calves were fed individually, with similar supplement DMI. Weight and MI were measured in July, August, and September. Forage intake (FI) was measured in July, August, and September of Trial 1 and July and August of Trial 2. Gain data were analyzed as a randomized complete block and MI and FI as a split-plot in time. Orthogonal contrasts were used to separate means and included CON vs supplemented, ENERGY vs protein, and DIP vs DIP+UIP. No trial effect or trial x treatment interactions (minimum P-value = 0.30) were detected for ADG. Supplemented calves gained faster than CON (P = 0.06). No other contrast differences were observed (minimum P-value = 0.50). Treatment did not affect FI (P > or = 0.55). Forage intake was lower (P < 0.001) in Trial 1 than in Trial 2. A linear increase (P = 0.0001) in FI (kg OM/d and percentage BW) occurred over time. Calves in Trial 2 consumed more (P = 0.004) fluid milk than calves in Trial 1, though no difference (P = 0.28) was observed relative to BW. No treatment or period differences were detected for fluid MI (minimum P-value = 0.23). Relative to BW, MI declined linearly (P = 0.0001) with successive periods. Energy may be limiting weight gain of nursing calves grazing native range in southeastern North Dakota.