Rapid visual detection of Giardia duodenalis in faecal samples using an RPA-CRISPR/Cas12a system.

Giardiasis is a common intestinal infection caused by Giardia duodenalis, which is a major economic and health burden for humans and livestock. Currently, a convenient and effective detection method is urgently needed. CRISPR/Cas12a-based diagnostic methods have been widely used for nucleic acid-based detection of pathogens due to their high efficiency and sensitivity. In this study, a technique combining CRISPR/Cas12a and RPA was established that allows the detection of G. duodenalis in faecal samples by the naked eye with high sensitivity (10-1 copies/µL) and specificity (no cross-reactivity with nine common pathogens). In clinical evaluations, the RPA-CRISPR/Cas12a-based detection assay detected Giardia positivity in 2% (1/50) of human faecal samples and 47% (33/70) of cattle faecal samples, respectively, which was consistent with the results of nested PCR. Our study demonstrated that the RPA-CRISPR/Cas12a technique for G. duodenalis is stable, efficient, sensitive, specific and has low equipment requirements. This technique offers new opportunities for on-site detection in remote and poor areas.

Molecular characterization and zoonotic potential of Cryptosporidium spp. and Giardia duodenalis in humans and domestic animals in Heilongjiang Province, China.

BACKGROUND: Cryptosporidiosis and giardiasis are significant parasitic diseases shared between humans and domestic animals. Due to the close contact between humans and domestic animals in rural areas, it is important to consider the potential transmission of zoonotic parasites from infected domestic animals to humans. This investigation aimed to determine the prevalence and molecular characteristics of Cryptosporidium spp. and Giardia duodenalis in domestic animals and villagers. METHODS: A total of 116 fecal samples from villagers and 686 fecal samples from domestic animals in Heilongjiang Province, China, were analyzed for two parasites using nested polymerase chain reaction (PCR) targeting various genetic loci and DNA sequence analysis of the PCR products. RESULTS: By sequence analysis of the SSU rRNA gene, the prevalence of Cryptosporidium in humans was 0.9% (1/116), with one species of C. parvum (n = 1) detected; among domestic animals, the prevalence was 2.6% (18/686), with five species identified: C. suis (n = 7) and C. scrofarum (n = 7) in pigs, C. meleagridis (n = 1) in chickens, C. andersoni (n = 1) in cattle, and C. canis (n = 2) in foxes. C. parvum and C. canis were further subtyped as IIdA19G1 and XXa4 on the basis of gp60 gene. Regarding G. duodenalis, based on the SSU rRNA, bg, gdh, and tpi genes, the prevalence in domestic animals was 5.1% (31/608), with three assemblages identified: A (n = 1) in pigs, D (n = 1) in foxes, and E (n = 27) in geese, cattle, pigs, ducks, and sheep, along with mixed infection of A + E (n = 1) in one pig and B + E (n = 1) in one sheep. No G. duodenalis was detected in humans (0/116). CONCLUSIONS: The present results show that no overlap of subtypes between animals and villagers was found in Cryptosporidium spp. and G. duodenalis, indicating a minor role of domestic animals in infecting humans in this population. However, the presence of zoonotic protozoa in domestic animals highlights the need for special attention to high-risk individuals during close contact with domestic animals.

Minimal zoonotic risk of cryptosporidiosis and giardiasis from frogs and reptiles.

The zoonotic potential of the protist parasites Cryptosporidium spp. and Giardia duodenalis in amphibians and reptiles raises public health concerns due to their growing popularity as pets. This review examines the prevalence and diversity of these parasites in wild and captive amphibians and reptiles to better understand the zoonotic risk. Research on Giardia in both groups is limited, and zoonotic forms of Cryptosporidium or Giardia have not been reported in amphibians. Host-adapted Cryptosporidium species dominate in reptiles, albeit some reptiles have been found to carry zoonotic (C. hominis and C. parvum) and rodent-associated (C. tyzzeri, C. muris and C. andersoni) species, primarily through mechanical carriage. Similarly, the limited reports of Giardia duodenalis (assemblages A, B and E) in reptiles may also be due to mechanical carriage. Thus, the available evidence indicates minimal zoonotic risk associated with these organisms in wild and captive frogs and reptiles. The exact transmission routes for these infections within reptile populations remain poorly understood, particularly regarding the importance of mechanical carriage. Although the risk appears minimal, continued research and surveillance efforts are necessary to gain a more comprehensive understanding of the transmission dynamics and ultimately improve our ability to safeguard human and animal health.

First report of Giardia duodenalis in dairy cattle and beef cattle in Shanxi, China.

BACKGROUND: Giardia duodenalis is an important intestinal parasitic protozoan that infects several vertebrates, including humans. Cattle are considered the major source of giardiasis outbreak in humans. This study aimed to investigate the prevalence and multilocus genotype (MLG) of G. duodenalis in Shanxi, and lay the foundation for the prevention and control of Giardiosis. METHODS AND RESULTS: DNA extraction, nested polymerase chain reaction, sequence analysis, MLG analysis, and statistical analysis were performed using 858 bovine fecal samples from Shanxi based on three gene loci: ß-giardin (bg), glutamate dehydrogenase (gdh), and triosephosphate isomerase (tpi). The overall prevalence of G. duodenalis was 28.3%, while its prevalence in Yingxian and Lingqiu was 28.1% and 28.5%, respectively. The overall prevalence of G. duodenalis in dairy cattle and beef cattle was 28.0% and 28.5%, respectively. G. duodenalis infection was detected in all age groups evaluated in this study. The overall prevalence of G. duodenalis in diarrhea and nondiarrhea samples was 32.4% and 27.5%, respectively, whereas that in intensively farmed and free-range cattle was 35.0% and 19.9%, respectively. We obtained 83, 53, and 59 sequences of bg, gdh, and tpi in G. duodenalis, respectively. Moreover, assemblage A (n = 2) and assemblage E (n = 81) by bg, assemblage A (n = 1) and assemblage E (n = 52) by gdh, and assemblage A (n = 2) and assemblage E (n = 57) by tpi were identified. Multilocus genotyping yielded 29 assemblage E MLGs, which formed 10 subgroups. CONCLUSIONS: To the best of our knowledge, this is the first study to report cattle infected with G. duodenalis in Shanxi, China. Livestock-specific G. duodenalis assemblage E was the dominant assemblage genotype, and zoonotic sub-assemblage AI was also detected in this region.

Occurrence and genotypic identification of Blastocystis sp., Enterocytozoon bieneusi, and Giardia duodenalis in dairy cattle in Heilongjiang Province, China.

Blastocystis sp., Enterocytozoon bieneusi, and Giardia duodenalis are three common zoonotic intestinal parasites, and cattle are important hosts of these three intestinal protozoa. In this study, 1632 fecal samples were collected from dairy farms in Heilongjiang Province, China, and screened for Blastocystis sp., E. bieneusi, and G. duodenalis using polymerase chain reaction. Of these, 149 (9.13%) were positive for three zoonotic pathogens, including 104 (6.40%), 22 (1.35%), and 23 (1.41%) for Blastocystis sp., E. bieneusi, and G. duodenalis, respectively. Based on partial SSU rRNA gene sequencing analysis, 104 positive samples of Blastocystis sp. were found, and a total of nine known subtypes were identified, including ST10 (61), ST3 (18), ST14 (6), ST26 (7), ST24 (3), ST25 (2), ST1 (2), ST5 (2), and ST21 (1). Among these, three subtypes (ST1, ST3, and ST5) were recognized as zoonotic subtypes, and two subtypes (ST10 and ST14) were specific to animals. All 23 Giardia duodenalis-positive samples belonged to assemblage E (n = 23) based on sequenced beta-giardin (bg) and triosephosphate isomerase (tpi) genes. Three known genotypes of E. bieneusi, namely J (n = 9), I (n = 6), and BEB4 (n = 7), were identified by sequence analysis of the internal transcriptional spacer region gene. Our study provides basic data for prevention and control in Heilongjiang Province; however, further research is required to better understand the prevalence and public health significance of these pathogens in the Heilongjiang region.

Host-specific Cryptosporidium, Giardia and Enterocytozoon bieneusi in shelter dogs from central Europe.

Cryptosporidium spp., Giardia intestinalis and microsporidia are unicellular opportunistic pathogens that can cause gastrointestinal infections in both animals and humans. Since companion animals may serve as a source of infection, the aim of the present screening study was to analyse the prevalence of these intestinal protists in fecal samples collected from dogs living in 10 animal shelters in central Europe (101 dogs from Poland and 86 from the Czech Republic), combined with molecular subtyping of the detected organisms in order to assess their genetic diversity. Genus-specific polymerase chain reactions were performed to detect DNA of the tested species and to conduct molecular subtyping in collected samples, followed by statistical evaluation of the data obtained (using χ2 or Fisher's tests). The observed prevalence was 15.5, 10.2, 1 and 1% for G. intestinalis, Enterocytozoon bieneusi, Cryptosporidium spp. and Encephalitozoon cuniculi, respectively. Molecular evaluation has revealed the predominance of dog-specific genotypes (Cryptosporidium canis XXe1 subtype; G. intestinalis assemblages C and D; E. cuniculi genotype II; E. bieneusi genotypes D and PtEbIX), suggesting that shelter dogs do not pose a high risk of human transmission. Interestingly, the percentage distribution of the detected pathogens differed between both countries and individual shelters, suggesting that the risk of infection may be associated with conditions typical of a given location.

Rabbits as reservoirs: An updated perspective of the zoonotic risk from Cryptosporidium and Giardia.

Rabbits are highly abundant in many countries and can serve as reservoirs of diseases for a diversity of pathogens including the enteric protozoan parasites, Cryptosporidium and Giardia. Both parasites shed environmentally robust environmental stages (oo/cysts) and have been responsible for numerous waterborne outbreaks of diseases. Cryptosporidium hominis and C. parvum are responsible for most infections in humans, while Giardia duodenalis assemblages A and B, cause most human cases of giardiasis. Cryptosporidium cuniculus, the dominant species infecting rabbits, is the only spceies other than C. hominis and C. parvum to have caused a waterborne outbreak of gastritis, which occurred in the United Kingdom in 2008. This review examines the prevalence of Cryptosporidium and Giardia species in rabbits to better understand the public health risks of contamination of water sources with Cryptosporidium and Giardia oo/cysts from rabbits. Despite the abundance of C. cuniculus in rabbits, reports in humans are relatively rare, with the exception of the United Kingdom and New Zealand, and reports of C. cuniculus in humans from the United Kingdom have declined substantially since the 2008 outbreak. Subtyping of C. cuniculus has supported the potential for zoonotic transmission. Relatively few studies have been conducted on Giardia, but assemblage B dominates. However, improved typing methods are required to better understand the transmission dynamics of Giardia assemblages in rabbits. Similarly, it is not well understood if pet rabbits or contaminated water are the main source of C. cuniculus infections in humans. Well-planned studies using high-resolution typing tools are required to understand the transmission dynamics better and quantify the public health risk of Cryptosporidium and Giardia from rabbits.

Comparison of Giardia duodenalis point-of-care antigen faecal tests to reference laboratory assays in non-symptomatic dogs.

Giardia duodenalis is one of the most prevalent enteric parasites of dogs. Point-of-care antigen tests (POC) are rapid and do not require additional equipment, or a specialised diagnostic laboratory. The aim of this study was to compare diagnostic tests available in veterinary practices and in a diagnostic laboratory for the detection of G. duodenalis on a cohort of group-housed dogs from New South Wales, Australia. Two different POC tests were used for the detection of G. duodenalis. Laboratory tests used were the multiplexed-tandem PCR panel (MT-PCR) that includes detection of G. duodenalis DNA, and two reference tests (an in-house TaqMan real-time PCR and a direct immunofluorescence assay, DFA). Canine faecal samples (n = 40) were tested simultaneously for the detection of G. duodenalis. Using either DFA or TaqMan real-time PCR as reference tests, 77.5% (31/40) and 82.5% (33/40) of dogs tested positive, respectively. Agreement (Kappa) between the DFA and TaqMan real-time PCR was 0.84 (95% CI 0.64 to 1.00). There was substantial G. duodenalis test outcome agreement between the two POC tests, Kappa = 0.75. Combining the two POC tests yielded 77% sensitivity and 100% specificity with DFA as reference, and for TaqMan real-time PCR it was 73% sensitivity and 100% specificity. The MT-PCR was in excellent agreement with each reference test, DFA or TaqMan real-time PCR. Due to the high specificity of both POC tests, they can be confidently used as rule-in diagnostics. Confirmatory testing that detects different biological parameters such as DNA, e.g. PCR (inc. MT-PCR), should be implemented before concluding that a dog is negative for the presence of G. duodenalis.

Molecular detection and genetic characteristics of Giardia duodenalis in dairy cattle from large-scale breeding farms in Xinjiang, China.

Giardia duodenalis is an intestinal protozoan that can infect both humans and animals, leading to public health issues and economic losses in the livestock industry. G. duodenalis has been reported to infect dairy cattle, but there is limited information available on large-scale dairy farms in Xinjiang, China. The study collected 749 fresh faecal samples from five large-scale cattle farms in Xinjiang, China. The study used a nested PCR assay of the small subunit ribosomal RNA (SSU rRNA*) gene to determine the presence of G. duodenalis. The results showed that 24.0% (180/749) of dairy cattle were positive for G. duodenalis, with the highest infection rate observed in pre-weaned calves (45.1%, 69/153). Among the 180 G. duodenalis positive samples, three assemblages were identified: assemblage E (n = 176), assemblage A (n = 3) and assemblage B (n = 1). Sixty-nine, 67 and 49 sequences were obtained for the beta-giardin (bg*) gene, the glutamate dehydrogenase (gdh*) gene and the triose phosphate isomerase (tpi*) gene, respectively. Thirteen novel sequences of assemblage E were identified, including five sequences from the bg* gene, four sequences from the gdh* gene and four sequences from the tpi* gene. This study found that 32 G. duodenalis assemblage E isolates formed 26 MLGs, indicating genetic variation and geographic isolation-based differentiation in bovine-derived G. duodenalis assemblage E. These findings provide fundamental insights into the genetic diversity of G. duodenalis in dairy cattle and can aid in the prevention and control of its occurrence in large-scale dairy cattle farms.

Zoonotic Cryptosporidium and Giardia in marsupials-an update.

Marsupials, inhabiting diverse ecosystems, including urban and peri-urban regions in Australasia and the Americas, intersect with human activities, leading to zoonotic spill-over and anthroponotic spill-back of pathogens, including Cryptosporidium and Giardia. This review assesses the current knowledge on the diversity of Cryptosporidium and Giardia species in marsupials, focusing on the potential zoonotic risks. Cryptosporidium fayeri and C. macropodum are the dominant species in marsupials, while in possums, the host-specific possum genotype dominates. Of these three species/genotypes, only C. fayeri has been identified in two humans and the zoonotic risk is considered low. Generally, oocyst shedding in marsupials is low, further supporting a low transmission risk. However, there is some evidence of spill-back of C. hominis into kangaroo populations, which requires continued monitoring. Although C. hominis does not appear to be established in small marsupials like possums, comprehensive screening and analysis are essential for a better understanding of the prevalence and potential establishment of zoonotic Cryptosporidium species in small marsupials. Both host-specific and zoonotic Giardia species have been identified in marsupials. The dominance of zoonotic G. duodenalis assemblages A and B in marsupials may result from spill-back from livestock and humans and it is not yet understood if these are transient or established infections. Future studies using multilocus typing tools and whole-genome sequencing are required for a better understanding of the zoonotic risk from Giardia infections in marsupials. Moreover, much more extensive screening of a wider range of marsupial species, particularly in peri-urban areas, is required to provide a clearer understanding of the zoonotic risk of Cryptosporidium and Giardia in marsupials.

Giardiasis and diarrhea in dogs: Does the microbiome matter?

BACKGROUND: Giardia duodenalis (Gd) causes intestinal parasitosis. The involvement of the intestinal microbiome in determining the infection's clinical phenotype is unknown. OBJECTIVE: Investigate the fecal microbiome features in dogs with giardiasis. ANIMALS AND METHODS: Cross-sectional study, including fecal samples of kenneled dogs with Gd diagnosed by fecal Giardia antigen dot ELISA. The fecal microbial compositional characteristics and dysbiosis index (DI) were compared between diarrheic and nondiarrheic dogs. RESULTS: Fecal samples of 38 Gd-infected dogs (diarrheic, 21; nondiarrheic, 17) were included. No differences were found in Faith's phylogenic diversity and beta diversity (weighted UniFrac distances) and in specific taxa abundances at the phylum, genus, and species levels, as well as in alpha and beta diversities between diarrheic and nondiarrheic dogs, and also when divided by sex or age. Among diarrheic dogs, alpha diversity was higher in males than in females (pairwise Kruskal-Wallis, q = 0.01). Among males, fecal abundances of the genus Clostridium (W = 19) and Clostridium spiroforme species (W = 33) were higher in diarrheic compared to nondiarrheic dogs. In diarrheic dog fecal samples, Proteobacteria were more prevalent (W = 1), whereas Verrucomicrobia were less prevalent in dogs <1 year of age than in older dogs. The fecal sample DI of 19 diarrheic and 19 nondiarrheic dogs was similar (median, -0.2; range, -4.3 to 4.5 and median, -1.0; range, -4.3 to 5.8, respectively). CONCLUSIONS: The fecal microbial composition of symptomatic and asymptomatic dogs with giardiasis is similar. Based on fecal DI, giardiasis is not characterized by prominent dysbiosis. Other host and parasite characteristics might determine the severity of giardiasis in dogs.

The risk of wild birds contaminating source water with zoonotic Cryptosporidium and Giardia is probably overestimated.

Cryptosporidium and Giardia are important waterborne protozoan parasites that are resistant to disinfectants commonly used for drinking water. Wild birds, especially wild migratory birds, are often implicated in the contamination of source and wastewater with zoonotic diseases, due to their abundance near water and in urban areas and their ability to spread enteric pathogens over long distances. This review summarises the diversity of Cryptosporidium and Giardia in birds, with a focus on zoonotic species, particularly in wild and migratory birds, which is critical for understanding zoonotic risks. The analysis revealed that both avian-adapted and zoonotic Cryptosporidium species have been identified in birds but that avian-adapted Cryptosporidium species dominate in wild migratory birds. Few studies have examined Giardia species and assemblages in birds, but the non-zoonotic Giardia psittaci and Giardia ardeae are the most commonly reported species. The identification of zoonotic Cryptosporidium and Giardia in birds, particularly C. parvum and G. duodenalis assemblages A and B in wild migratory birds, is likely due to mechanical carriage or spillback from birds co-grazing pastures contaminated with C. parvum from livestock. Therefore, the role of wild migratory birds in the transmission of zoonotic Cryptosporidium and Giardia to source water is likely overestimated. To address knowledge gaps, it is important to conduct more extensive studies on the prevalence of Cryptosporidium and Giardia in a broader range of migratory wild birds. There is also a need to investigate the extent to which zoonotic infections with C. hominis/C. parvum and G. duodenalis assemblages A and B are mechanical and/or transient, and to assess the load and viability of zoonotic oo/cysts shed in avian faeces. Understanding the contribution of birds to zoonoses is essential for effective disease surveillance, prevention, and control.

[Feline and canine giardiosis: An Update]. / Ein Update zur felinen und caninen Giardiose.

Giardia duodenalis is a facultative pathogenic intestinal parasite. Giardiosis in dogs and cats may appear with or without clinical signs. Typical signs include diarrhea with or without vomiting. The prevalence in young animals is high and may amount to up to 50%. There are 8 different genotypes (A - H), which are called assemblages. Assemblages C and D are most common in dogs and assemblage F most frequent in cats. However, animals may also be infected with the zoonotically effective assemblages A and B or exhibit mixed infections. The immunofluorescence test (IFA), the enzyme-linked immunosorbent assay (ELISA) and fecal centrifugation using zinc sulphate solution are currently recommended as diagnostic methods. Polymerase chain reaction (PCR) may be used to determine the corresponding assemblage. Approved treatments for giardiosis include fenbendazole and metronidazole. In addition, undertaking specific hygiene measures is warranted. Only animals showing clinical signs or those living in the same household with high-risk patients (e. g. immunosuppressed humans) are recommended to receive medication. The aim of treatment is clinical improvement of the diseased dogs and cats. Frequently, complete elimination of Giardia is not attained.

First identification and coinfection detection of Enterocytozoon bieneusi, Encephalitozoon spp., Cryptosporidium spp. and Giardia duodenalis in diarrheic pigs in Southwest China.

BACKGROUND: Enterocytozoon bieneusi, Encephalitozoon spp., Cryptosporidium spp., and Giardia duodenalis (G. intestinalis) are enteric pathogens that cause diarrhea in pigs. This study aimed to determine the prevalence of these enteric parasites and their coinfection with E. bieneusi in diarrheic pigs in Southwest China (Chongqing and Sichuan) using nested polymerase chain reaction (nPCR) based methods. RESULTS: A total of 514 fecal samples were collected from diarrheic pigs from 14 pig farms in Chongqing (five farms) and Sichuan (nine farms) Provinces. The prevalence of Encephalitozoon spp., Cryptosporidium spp. and G. duodenalis was 16.14% (83/514), 0% (0/514), and 8.95% (46/514), respectively. Nested PCR revealed 305 mono-infections of E. bieneusi, six of E. cuniculi, two of E. hellem, and nine of G. duodenalis and 106 concurrent infections of E. bieneusi with the other enteric pathogens. No infections of E. intestinalis and Cryptosporidium species were detected. The highest coinfection was detected between E. bieneusi and E. cuniculi (10.5%, 54/514), followed by E. bieneusi and G. duodenalis (5.8%, 30/514) and E. bieneusi and E. hellem (2.9%, 15/514). E. bieneusi was the most frequently detected enteric pathogen, followed by E. cuniculi, G. duodenalis and E. hellem. There was a significant age-related difference in the prevalence of E. cuniculi in fattening pigs (χ2 = 15.266, df = 3, P = 0.002) and G. duodenalis in suckling pigs (χ2 = 11.92, df = 3, P = 0.008) compared with the other age groups. Sequence analysis of the ITS region of Encephalitozoon species showed two genotypes (II and III) for E. cuniculi and one (TURK1B) for E. hellem. Only G. duodenalis assemblage A was identified in all nested PCR-positive samples. E. bieneusi was found more often than other enteric pathogens. CONCLUSIONS: This study showed that E. bieneusi, Encephalitozoon spp. [E. cuniculi and E. hellem] and G. duodenalis were common enteric parasites in diarrheic pigs in Chongqing and Sichuan Provinces. In case of both mono-infection and coinfection, E. bieneusi was the most common enteric pathogen in diarrheic pigs. Thus, it may be a significant cause of diarrhea in pigs. Precautions should be taken to prevent the spread of these enteric parasites.

The detection and phylogenetic characterization of Cryptosporidium, Cystoisospora, and Giardia duodenalis of cats in South Korea.

Introduction: Cryptosporidium, Cystoisospora, and Giardia duodenalis are gastrointestinal protozoa parasites that cause diarrhea in various animals. However, information regarding the detection and phylogenetic characterization of gastrointestinal protozoa parasites in cats is limited throughout South Korea. Therefore, this study aimed to determine the detection and identify subspecies of gastrointestinal protozoa parasites in cats from South Korea. Methods: A total of 290 fecal samples were collected from stray, companion, and shelter cats in six provinces. Cryptosporidium, Cystoisospora, and G. duodenalis were identified by PCR. All positive samples were subtyped by PCR and sequencing of gp60, ITS-1, tpi, bg, and gdh. Results: The overall detection of gastrointestinal protozoan parasitic infection was 17.93%. G. duodenalis was the most prevalent, with 7.93%, followed by Cystoisospora spp. (7.24%) and Cryptosporidium spp. (4.48%). In addition, C. felis (n=10), C. parvum (n=2), C. ryanae (n=1), Cystoisospora felis (n=14), Cystoisospora suis (n=5), Cystoisospora ohioensis (n=1), Cystoisospora spp. were identified in subspecies analysis of positive samples. C. felis showed a significant association with diarrhea (7.81%) and living condition (6.04%), and Cystoisospora felis in diarreha (9.38%) according to detection. Through phylogenetic analysis of the tpi, bg, and gdh genes from 23 G. duodenalispositive samples, it was confirmed that the samples of present study belonged to assemblage A, B, C, and D. Discussion: South Korean cats have a high rate of gastrointestinal protozoan parasites infection with cat-specific Cryptosporidium and Cystoisospora, which are associated with living conditions and diarrhea symptoms. Moreover, zoonotic and other animal-specific subtype of protozoan parasites have been detected in cat feces.

Occurrence and genetic characteristics of Giardia duodenalis in donkeys in Xinjiang, China.

Giardia duodenalis is a common enteric parasite in humans and animals. To examine the occurrence and genetic characteristics of Giardia in donkeys in Xinjiang, China, 758 fecal samples from donkeys were collected, and Giardia was screened via PCR at the SSU rRNA gene. A total of 17.0% (129/758) of samples tested positive for Giardia, with the infection rate in large-scale farm and domestic donkeys being 21.4% (124/580) and 2.8% (5/178), respectively; the infection rates in <1-year-old and ≥1-year-old donkeys were 19.3% (72/374) and 12.7% (41/323), respectively. Three Giardia assemblages were identified, with assemblage B (n = 102) as the prevalent assemblage, followed by assemblage A (n = 23) and assemblage E (n = 4). Of the 129 Giardia-positive isolates, 40, 34 and 59 sequences were obtained at the bg, gdh and tpi genes, respectively. Twenty-one isolates successfully allowed multilocus genotyping (MLG), with four novel assemblage A MLGs, named MLG-AI-1 (n = 1), MLG-AI-2 (n = 1), MLG-AI-3 (n = 1), and MLG-AI-4 (n = 1) and three novel assemblage B MLGs, named MLG-B1 (n = 1), MLG-B2 (n = 14), and MLG-B3 (n = 1). Moreover, two isolates formed two MLG-mixed sequences. The results suggest that donkeys are commonly infected with Giardia in Xinjiang, and there is genetic diversity and host adaptability among the isolates.

Title: Présence et caractéristiques génétiques de Giardia duodenalis chez les ânes du Xinjiang, Chine. Abstract: Giardia duodenalis est un parasite entérique courant chez les humains et les animaux. Pour étudier la présence et les caractéristiques génétiques de Giardia chez les ânes du Xinjiang, en Chine, 758 échantillons fécaux d'ânes ont été collectés et Giardia a été criblé par PCR du gène de l'ARNr SSU. Au total, 17,0 % (129/758) des échantillons ont été testés positifs pour Giardia. Le taux d'infection, respectivement chez les ânes des élevages à grande échelle et domestiques, étaient de 21,4 % (124/580) et 2,8 % (5/178). Les taux chez les ânes de < 1 an et ≥ 1 an étaient respectivement de 19,3 % (72/374) et 12,7 % (41/323). Trois assemblages de Giardia ont été identifiés, l'assemblage B (n = 102) étant l'assemblage prédominant, suivi de l'assemblage A (n = 23) et de l'assemblage E (n = 4). Sur les 129 isolats positifs pour Giardia, 40, 34 et 59 séquences ont été obtenues respectivement au niveau des gènes bg, gdh et tpi. Vingt et un isolats ont permis du génotypage multilocus (MLG), avec quatre nouveaux MLG de l'assemblage A, nommés MLG-AI-1 (n = 1), MLG-AI-2 (n = 1), MLG-AI-3 (n = 1) et MLG-AI-4 (n = 1) et trois nouveaux MLG de l'assemblage B, nommés MLG-B1 (n = 1), MLG-B2 (n = 14) et MLG-B3 (n = 1). De plus, deux isolats formaient deux séquences MLG mélangés. Les résultats suggèrent que les ânes sont couramment infectés par Giardia au Xinjiang, et qu'il existe une diversité génétique et une adaptabilité à l'hôte parmi les isolats.

First report of prevalence and assemblage analysis of Giardia duodenalis in pigs from Guangxi Zhuang Autonomous Region, southern China.

Giardia duodenalis is a common intestinal protozoan that can cause diarrhea and intestinal disease in animals and in humans. However, the prevalence and assemblages of G. duodenalis in pigs from Guangxi Zhuang Autonomous Region have not been reported. In this study, a total of 724 fecal samples (201 from nursery pigs, 183 from piglets, 175 from breeding pigs, and 165 from fattening pigs) were obtained in four areas of the region (Nanning, Yulin, Hezhou, and Guigang). The gene of the small subunit ribosomal RNA (SSU rRNA) of G. duodenalis was amplified by nested PCR. The results show that the prevalence of G. duodenalis in pigs was 3.59% (26/724), of which 14 samples belonged to assemblage A (53.85%) and 12 samples belonged to assemblage E (46.15%). The infection rates of G. duodenalis in Hezhou, Yulin, Nanning, and Guigang were 0%, 0.7%, 10.8% and 1.1%, respectively (χ2 = 45.616, p < 0.01); whereas 5.1% of breeding pigs, 6.0% of piglets, 2.4% of fattening pigs, and 1.0% of nursery pigs were infected with G. duodenalis (χ2 = 8.874, p < 0.05). The SSU rRNA-positive samples were amplified by PCR based on the ß-giardin (bg), glutamate dehydrogenase (gdh), and triphosphate isomerase (tpi) genes. Ten, eight and seven positive samples were detected, respectively. Based on phylogenetic analysis of the three genetic loci sequences, a multilocus genotyping A1 was found. The findings of this study provide basic data for the development of prevention and control of G. duodenalis infections in pigs and humans in the Guangxi Zhuang Autonomous Region.

Title: Premier rapport sur la prévalence et l'analyse des assemblages de Giardia duodenalis chez les porcs de la région autonome Zhuang du Guangxi, dans le sud de la Chine. Abstract: Giardia duodenalis est un protozoaire intestinal commun qui peut provoquer des diarrhées et des maladies intestinales chez les animaux et les humains. Cependant, la prévalence et les assemblages de G. duodenalis chez les porcs de la région autonome Zhuang du Guangxi n'ont pas été rapportés. Dans cette étude, un total de 724 échantillons fécaux (201 provenant de jeunes porcelets, 183 de porcelets, 175 de porcs reproducteurs et 165 de porcs à l'engrais) ont été obtenus dans quatre zones de la région (Nanning, Yulin, Hezhou et Guigang). Le gène de la petite sous-unité de l'ARN ribosomal (ARNr SSU) de G. duodenalis a été amplifié par PCR nichée. Les résultats ont montré que la prévalence de G. duodenalis chez les porcs était de 3,59 % (26/724), dont 14 échantillons appartenaient à l'assemblages A (53,85 %) et 12 échantillons à l'assemblage E (46,15 %). Les taux d'infection par G. duodenalis à Hezhou, Yulin, Nanning et Guigang étaient respectivement de 0, 0,7 %, 10,8 % et 1,1 % (χ2 = 45,616, p < 0,01), alors que 5,1 % des porcs reproducteurs, 6,0 % des porcelets, 2,4 % de porcs à l'engrais et 1,0 % des jeunes porcelets étaient infectés par G. duodenalis (χ2 = 8,874, p < 0,05). Les échantillons positifs pour l'ARNr SSU ont été amplifiés par PCR basée sur les gènes de la ß-giardine (bg), de la glutamate déshydrogénase (gdh) et de la triphosphate isomérase (tpi), et dix, huit et sept échantillons positifs ont été détectés, respectivement. Sur la base de l'analyse phylogénétique des trois séquences de loci génétiques, un génotypage multilocus A1 a été trouvé. Les résultats de cette étude fournissent des données de base pour le développement de la prévention et du contrôle des infections à G. duodenalis chez les porcs et les humains dans la région autonome Zhuang du Guangxi.

Zoonotic Giardia duodenalis assemblage A in northern sloth from Brazilian Amazon.

BACKGROUND: The parasite Giardia duodenalis infects a wide range of vertebrate hosts, including domestic and wild animals as well as humans. Giardia is genotyped into eight assemblages (A-H). Zoonotic assemblages A and B have already been identified in humans and wild and domestic animals (non-human primates and cats) from Brazilian Amazon and in the world. Due to its zoonotic/zooanthroponotic nature, surveillance initiatives and the definition of Giardia assemblages are important in order to characterise the epidemiological scenario and to implement further control measures. OBJECTIVES: Determine assemblages of G. duodenalis in sloths from the Brazilian Amazon Region. METHODS: Faecal parasitological examination of sloths from Amazonas State. Polymerase chain reaction (PCR) targeting the beta giardin (BG), and genes from multilocus sequence typing (MLST) scheme, amplicon sequencing and phylogenetic analysis. FINDINGS: Here, we identified, by microscopy, Giardia in two northern sloths (Bradypus tridactylus). These two samples were submitted to molecular assays and it was revealed that both were infected by G. duodenalis assemblage A. Phylogenetic analysis showed that they belong to assemblage A within sequences from humans and wild and domestic animals. CONCLUSION: Therefore, besides showing, by the first time, the current presence of this parasite in sloths, our findings reveals that this wild animal species would be part of the zoonotic/zooanthroponotic scenario of this parasite in the Brazilian Amazon.

Prevalence and public health relevance of enteric parasites in domestic dogs and cats in the region of Madrid (Spain) with an emphasis on Giardia duodenalis and Cryptosporidium sp.

BACKGROUND: Pet dogs and cats exert an unquestionable beneficial effect in the well-being of their owners, but can also act as a source of zoonotic infections if improperly cared. OBJECTIVES: We investigated the occurrence, risk factors, genetic variability and zoonotic potential of intestinal parasites in dogs and cats attended in a clinical veterinary setting in Spain. METHODS: Canine (n = 252) and feline (n = 35) faecal samples were collected during 2017-2019 and analysed by coproparasitological methods. A rapid lateral immunochromatographic test (ICT) was used for detecting Giardia duodenalis and Cryptosporidium sp. Samples positive at microscopy examination and/or ICT were reassessed by molecular methods. RESULTS: Overall, 48.8% (123/252) of dogs and 48.6% (17/35) of cats were infected by enteric parasites. In dogs, G. duodenalis was the most prevalent species (40.9%), followed by Cystoisospora sp. (7.1%), and Toxocara canis (5.2%). In cats, Joyeuxiella sp. and Toxocara cati were the dominant species (20.0% each), followed by G. duodenalis (14.3%), D. caninum (5.7%) and Cystoisospora felis and Toxascaris leonina (2.9% each). Pups and kittens were more likely to harbour intestinal parasites and develop clinical signs. Sequence analyses of dog isolates revealed the presence of assemblages A (n = 1), C (n = 4), D (n = 4) and C+D (n = 1) within G. duodenalis; C. parvum (n = 1) and C. canis (n = 4) within Cryptosporidium and PtEb IX (n = 1) in Enterocytozoon bieneusi. A novel C. canis subtype family, named XXi, is reported. CONCLUSIONS: Our results highlight that (i) well-cared dogs carry zoonotic enteric protozoan parasites of public health relevance, (ii) proper hygiene practices and routine veterinary treatment are essential to prevent zoonotic infections, (iii) vulnerable populations should avoid contact with pups/kittens with diarrhoea and (iv) infected dogs might be major contributors to the environmental contamination with soil-transmitted helminths (STHs) eggs.

Thieno[3,2-b]pyrrole 5-carboxamides as potent and selective inhibitors of Giardia duodenalis.

Giardia duodenalis is the causative agent of the neglected diarrhoeal disease giardiasis. While often self-limiting, giardiasis is ubiquitous and impacts hundreds of millions of people annually. It is also a common gastro-intestinal disease of domestic pets, wildlife, and livestock animals. However, despite this impact, there is no vaccine for Giardia currently available. In addition, treatment relies on chemotherapies that are associated with increasing failure rates. To identify new treatment options for giardiasis we recently screened the Compounds Australia Scaffold Library for new chemotypes with selective anti-Giardia activity, identifying three compounds with sub-µM activity and promising selectivity. Here we extended these studies by examining the anti-Giardia activity of series CL9569 compounds. This compound series was of interest given the promising activity (IC50 1.2 µM) and selectivity demonstrated by representative compound, SN00798525 (1). Data from this work has identified an additional three thieno [3,2-b]pyrrole 5-carboxamides with anti-Giardia activity, including 2 which displayed potent cytocidal (IC50 ≤ 10 nM) and selective activity against multiple Giardia strains, including representatives from both human-infecting assemblages and metronidazole resistant parasites. Preclinical studies in mice also demonstrated that 2 is well-tolerated, does not impact the normal gut microbiota and can reduce Giardia parasite burden in these animals.