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Background: Giardia duodenalis is a common enteric protozoan parasite that is categorized into eight assemblages (A-H). In particular, assemblages A and B are zoonotic, capable of infecting both humans and animals worldwide, resulting in significant economic losses and public health challenges in epidemic regions. Thus, the development of rapid, accurate and non-laboratory-based diagnostic methods for infected animals is crucial for the effective prevention and control of giardiasis. Recent advancements in clustered, regularly interspaced, short palindromic repeats (CRISPR) and CRISPR-associated (Cas) protein (Cas12a) systems allow promising avenues for nucleic acid detection, characterized by their high flexibility, sensitivity and specificity., Methods: Combined recombinase polymerase amplification and CRISPR/Cas12a systems were combined and used as end-point diagnostic methods (termed REPORT) to detect G. duodenalis assemblage A and B. The diagnostic results can be observed by fluorescence readouts with the naked eye under blue light or colorimetric signals using a lateral flow strip (LFS)., Results: The limit of detection (LOD) of the REPORT‑based G. duodenalis assemblage A detection was 2.04 CFU/ml and 10 trophozoites per gram (TPG), and the LOD of assemblage B was 1.1 CFU/ml and 10 cysts per gram (CPG). The REPORT‑based G. duodenalis assemblage A and assemblage B detection methods have strong specificity and no cross-reactivity with other assemblages of G. duodenalis or common enteric parasitic protozoa and have excellent performance in clinical sample detection., Conclusions: This study presents a novel strategy for the direct identification of G. duodenalis assemblages A and B, requiring neither highly trained personnel nor costly specialized equipment., Competing Interests: Declarations Ethics approval and consent to participate All the research procedures used in this study were approved by the Institutional Review Board of Henan Agricultural University (approval no. IRB-HENAU-20190820-02). The use of positive human samples in this study complied with the 1975 Declaration of Helsinki, as revised in 2013. Consent for publication Not applicable. Competing interests The authors declare no competing interests., (© 2024. The Author(s).)

Intestinal protozoa Cryptosporidium spp., Giardia duodenalis , and Enterocytozoon bieneusi have been implicated in serious waterborne outbreaks worldwide. Wastewater-based epidemiology (WBE) is a promising approach for evaluating the disease prevalence in a catchment population in that it monitors the contamination level of the intestinal pathogens in wastewater. We collected 48 urban wastewater samples (24 from influents and 24 from effluents) from the Yangpu Wastewater Treatment Plant (YPWTP) in Shanghai, China. We identified Cryptosporidium spp., G. duodenalis , and E. bieneusi by nested polymerase chain reaction (PCR) amplification. Cryptosporidium hominis and subtype IdA14 were identified in two samples by analyzing the sequences of small subunit ribosomal RNA (SSU rRNA) and 60-kDa glycoprotein ( gp60 ) genes, respectively. The G. duodenalis sub-assemblage AII ( n = 8) and assemblage C ( n = 4) in 12 samples were determined by analyzing triosephosphate isomerase ( tpi ) gene sequences. The E. bieneusi genotype A was identified in one sample by analyzing the sequence of the internal transcribed spacer (ITS) region of the rRNA gene. These findings suggest that improving wastewater treatment and monitoring the virility of pathogens in effluents is critical. We observed similar prevalence and genotypes/subtypes of the three intestinal protozoa in our wastewater samples as those reported in previous studies, providing evidence that WBE can be used as an effective epidemic management tool.IMPORTANCE Cryptosporidium spp., Giardia duodenalis, and Enterocytozoon bieneusi are common intestinal protozoa causing diarrhea. The infective oocysts, cysts, and spores released in feces can survive in different environments, including multiple types of water bodies. Humans can acquire these intestinal protozoan infections via the fecal-oral route as in waterborne transmission. Wastewater-based epidemiology can rapidly and reliably detect and monitor the emergence and spread of waterborne diseases. We detected Cryptosporidium spp., G. duodenalis, and E. bieneusi in a wastewater treatment plant in Shanghai, China, reflecting the occurrence and genetic characterizations of the three intestinal pathogens from community members served by the wastewater treatment plant., Competing Interests: The authors declare no conflict of interest.

Giardia intestinalis is a major diarrhea-causing parasite that colonizes the proximal small intestine of humans and various other mammalian species, including pets and livestock. Despite its global occurrence, there is limited information about the epidemiology of Giardia in reptiles, particularly snakes. The aim of this study was to amplify the beta-giardin (bg) gene of Giardia by polymerase chain reaction (PCR), followed by a comparative evolutionary tree analysis to determine the occurrence and molecular characterization of Giardia in snakes. We collected 603 asymptomatic samples from 26 provinces in China, representing species such as Pantherophis guttatus, Pantherophis obsoletus, Pituophis melanoleucus, Thamnophis sirtalis, Lampropeltis getula, and Heterodon nasicus. Ultimately, a occurrence of Giardia infection of 4.15 % was detected in these snakes, with corn snakes (Pantherophis guttatus) having a significantly higher occurrence than other species. Molecular analysis identified assemblage F as the predominant genotype, and also identified an assemblage B that can infect humans and a variety of mammals. The results of this study illustrate the potential risk of transmission of Giardia from snakes to humans, especially in environments where close contact occurs. The present epidemiological study examines epidemiological investigations of Giardia in reptiles, provides data to understand the zoonotic risk of Giardia, and emphasizes the need for targeted surveillance, stringent hygiene measures, and public awareness campaigns to reduce these risks., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

This study aimed to carry out a molecular screening for the presence of Giardia, Cryptosporidium, and/or Entamoeba in the feces of pet and stray/feral cats in Jordan. G. duodenalis was found in 27.9% (95% CI, 23.2-32.9) of the 348 sampled cats overall; E. histolytica was found in only 0.6% (95% CI, 0.1-2.1) of the cats, while none of the sampled cats had Cryptosporidium infections. The infection rate of G. duodenalis among indoor cats (32.3%) did not differ significantly from that among outdoor cats (24.1%). There were significantly more infections (p = 0.0004) geographically in the cold semiarid areas (67%) than in the cold desert areas (24%). Multilocus sequence typing analysis of amplicons based on the bg, tpi, and gdh genes revealed that the majority of G. duodenalis infections were zoonotic assemblage B (65.9%; 64 of 97 positive samples); followed by feline-specific assemblage F (18.5%, 18/97); cattle-specific assemblage E (5.2%, 5/97); and then assemblage C that was shared with canids (1.0%; 1/97). Within Giardia isolates, a substitution mutation (A/G) was found at position 297 of the complete protein coding sequence (cds) of tpi-assemblage B, which may represent a new spreading mutation within this gene among the cat population in Jordan. The results of the present study suggest that close human-cat interactions could play a role in zoonotic transmission of Giardia, but further research is needed to determine the possible contribution of cats to the transmission of other protozoa to humans., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Cryptosporidium and Giardia duodenalis are common gastrointestinal protozoan parasites that cause diarrhea in humans and animals. Although dogs in animal shelters in South Korea can be reintroduced into human society as companions, no continuous investigations have been conducted on the prevalence and genetic characteristics of the protozoan parasites. In the present study, 345 fecal samples from dogs were obtained from animal shelters in six provinces between January and December 2022. The prevalence of Cryptosporidium and G. duodenalis were molecularly identified at the 18S rRNA gene. Cryptosporidium canis isolates were subtyped at the gp60 locus, and G. duodenalis assemblages were identified at the tpi, bg and gdh loci. Overall, 6.67% of the fecal samples tested positive for Cryptosporidium spp. and two species were identified: C. canis (5.51%) and Cryptosporidium parvum (1.16%). Significant regional differences in prevalence were identified for C. canis. Subtyping analysis of C. canis isolates revealed a predominance of the subtype families XXa and XXe over XXb, and XXc. G. duodenalis was detected in 17.68% of the samples, and significant regional differences were identified; its prevalence was also significantly highest in diarrhoeic animals. Molecular characterization of G. duodenalis reveal that most isolates belonged to the canine-specific assemblages C and D. The present results can contribute to further insights into the prevalence and genotypes of Cryptosporidium and G. duodenalis in shelter dogs in South Korea., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Giardia duodenalis is an intestinal pathogen that is found globally. Children are more susceptible and often suffer severe consequences after infection. Despite this, the health effects of this pathogen continue to be poorly understood and neglected. In Wenzhou, Zhejiang province, China, stool samples were obtained from 1032 children who were admitted to Yuying Children's Hospital. Out of these, 684 presented with diarrhea, while 348 were asymptomatic. The stool samples were screened for G. duodenali by targeting the small subunit of the ribosomal RNA (SSU rRNA) gene. Subtypes of G. duodenalis were identified via amplification of the glutamate dehydrogenase (gdh), beta-giardin (bg), and triosephosphate isomerase (tpi) genes in samples positive for the G. duodenalis. The findings indicated the presence of G. duodenalis in 0.9 % (9/1032) of the samples, with 9/684 (1.3 %) of the samples originating from children with diarrhea and none from the asymptomatic samples. All 9 samples that tested positive for G. duodenalis were determined to be of assemblage A. Of these, 6 samples were effectively genotyped at all 3 loci, resulting in the identification of 3 distinct MLGs: MLG-AII1 (n = 1), MLG-AII2 (n = 4), and MLG-AII2 (n = 1), all belonging to G. duodenalis assemblage AII. This was the first study that confirmed G. duodenalis infections in children residing in southern Zhejiang, China, with comparatively low rates of infection. The detection of G. duodenalis assemblage AII indicates a possibility of transfer from one human to another. The parasite's effect on the health of young children requires special attention and consideration., Competing Interests: Declaration of competing interest The authors declare that they have no competing interests., (Copyright © 2024. Published by Elsevier B.V.)

The domestic chinchilla (Chinchilla lanigera) is kept as a pet and previous studies suggest that it may play an important role as a source of zoonotic parasites, including Giardia intestinalis, Cryptosporidium spp. and microsporidia. In this study, we examined the occurrence and genetic diversity of above mentioned parasites in pet chinchillas in the Czech Republic by PCR/sequencing of the 18S rRNA, TPI, and ITS genes. Of 149 chinchillas from 24 breeders, 91.3 % were positive for G. intestinalis, 8.1 % for Cryptosporidium spp., 2.0 % for Encephalitozoon spp., and 5.4 % for E. bieneusi. Molecular analyses revealed presence of G. intestinalis assemblage B, C. ubiquitum (XIIa family), E. bieneusi genotypes D, SCF2, and, CHN-F1, and E. intestinalis. The infection intensity of G. intestinalis determined by qRT-PCR reached up to 53,978 CPG, C. ubiquitum up to 1409 OPG, E. intestinalis up to 1124 SPG, and E. bieneusi up to 1373 SPG. Only two chinchillas with C. ubiquitum and five with G. intestinalis had diarrhoea at the time of the screening. Three chinchillas in the long-term study were consistently positive for G. intestinalis, with intermittent excretion of C. ubiquitum, E. intestinalis, and E. bieneusi over 25 weeks. The findings indicate that chinchillas are frequently infected with zoonotic parasitic protists, but that these infections rarely show clinical signs. The lack of visible signs could reduce the vigilance of pet owners when handling their chinchillas, increasing the risk of transmission within breeding groups and possibly to humans., Competing Interests: Declaration of Competing Interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Martin Kváč reports financial support was provided by Czech Science Foundation. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier B.V.)

Giardia duodenalis is a common cause of diarrheal illness in regions with limited resources. The demand for rapid and cost-effective detection and genotyping methods in large-scale epidemiological studies and clinical diagnostics is imperative. Hence, we developed a multiplex PCR-RFLP technique targeting the tpi gene of G. duodenalis. The assay successfully screened G. duodenalis positive clinical samples (6.33 %; 36/565). It was also able to categorize the isolates into assemblages A (41.66 %; 13/36) and B (58.33 %; 23/36), as well as into subassemblages: AI (13.8 %; 5/36), AII (27.77 %; 10/36), BIII (36.11 %; 15/36) and BIV (22.22 %; 8/36). High diagnostic sensitivity (94.2 %), specificity (100 %) and accuracy (97.1 %) of the PCR assay were obtained, indicating its reliability for diagnosing giardiasis. Notably, the assay demonstrated close concordance with microscopy (κ=0.85) and reference PCR (κ=0.98) results. The optimized method offers a cost-effective and rapid approach for G. duodenalis detection and genotyping, convenient for epidemiological studies and clinical diagnostics., Competing Interests: Declaration of competing interest The authors affirm that they do not have any competing interests related to the publication of the article., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Background: The parasitic protozoan Giardia duodenalis is an important cause of diarrheal disease in humans and animals that can be spread by fecal-oral transmission through water and the environment, posing a challenge to public health and animal husbandry. Little is known about its impact on large-scale sheep farms in China. In this study we investigated G. duodenalis infection of sheep and contamination of the environment in large-scale sheep farms in two regions of China, Henan and Ningxia., Methods: A total of 528 fecal samples, 402 environmental samples and 30 water samples were collected from seven large-scale sheep farms, and 88 fecal samples and 13 environmental samples were collected from 12 backyard farms. The presence of G. duodenalis was detected by targeting the β-giardin (bg) gene, and the assemblage and multilocus genotype of G. duodenalis were investigated by analyzing three genes: bg, glutamate dehydrogenase (gdh) and triphosphate isomerase (tpi)., Results: The overall G. duodenalis detection rate was 7.8%, 1.4% and 23.3% in fecal, environmental and water samples, respectively. On the large-scale sheep farms tested, the infection rate of sheep in Henan (13.8%) was found to be significantly higher than that of sheep in Ningxia (4.2%) (P < 0.05). However, the difference between the rates of environmental pollution in Henan (1.9%) and Ningxia (1.0%) was not significant (P > 0.05). Investigations of sheep at different physiological stages revealed that late pregnancy ewes showed the lowest infection rate (1.7%) and that young lambs exhibited the highest (18.8%). Genetic analysis identified G. duodenalis belonging to two assemblages, A and E, with assemblage E being dominant. A total of 27 multilocus genotypes were identified for members of assemblage E., Conclusions: The results suggest that G. duodenalis is prevalent on large-scale sheep farms in Henan and Ningxia, China, and that there is a risk of environmental contamination. This study is the first comprehensive examination of the presence of G. duodenalis on large-scale sheep farms in China. Challenges posed by G. duodenalis to sheep farms need to be addressed proactively to ensure public health safety., (© 2024. The Author(s).)

Non-human primates (NHPs) are the group that most share infectious agents with humans due to their close taxonomic relationship. The southern brown howler monkeys (Alouatta guariba clamitans) are endemic primates from Brazil and Argentina's Atlantic Forest. This study aimed to investigate the presence of intestinal parasites in free-living (FL) and captive (CA) southern brown howler monkeys. Thirty-nine stool samples were collected in two areas in southern Brazil, 15 FL and 24 CA. Stool sediments obtained by centrifugal sedimentation technique were used for microscopic analysis and direct immunofluorescence assay and evaluated by molecular analysis through amplification and sequencing of TPI fragments. Intestinal parasites Giardia duodenalis, Cryptosporidium spp., and Trypanoxyuris minutus were detected at coproparasitological analysis. This is the first report of the presence of Cryptosporidium spp. in free-living howlers. The molecular characterization of G. duodenalis isolates indicated assemblage B for the first time found in free-living A. guariba clamitans. The high prevalence of G. duodenalis transmission in CA howler monkeys can be explained by direct contact with humans and frequent soil contact. The presence of a potentially zoonotic assemblage in these animals indicates that the process of fragmentation and cohabitation with humans and livestock affects the wildlife, thus indicating a need for eco-health measures., Competing Interests: Declaration of competing interest All authors declare no conflicts of interest., (Copyright © 2024. Published by Elsevier B.V.)

Giardiasis is a small intestinal disease caused by the zoonotic parasite, Giardia duodenalis. This study presents the molecular findings of G. duodenalis infection in companion dogs, domestic livestock and wildlife in the Northern Jordan Basin, Israel. Identification of G. duodenalis was accomplished by nested PCR (nPCR) targeting the 18S rRNA gene. Samples were collected from water (five samples from four sources of which one was recycled water), as well as feces from wolves (Canis lupus) (n = 34), jackals (Canis aureus) (n = 24), wild boars (Sus scrofa) (n = 40), cattle (Bos taurus) (n = 40), dogs (Canis lupus familiaris) (n = 37) and nutria (Mayocastor coypus) (n = 100). All positive samples were sequenced and a phylogenetic tree was drawn using the Bayesian Inference (BI) algorithm. Differences in G. duodenalis prevalence between the different hosts were analyzed by Pearson's chi-square (p < 0.05). Of the total 275 fecal samples, 36 were positive for G. duodenalis (13%). Frequency rates among different animal species was highest in wolves (32.3%), whilst rates in wild boars (22.5%), dogs (16.2%), cattle (12.5%) and jackals (4.2%), were observed to be significantly lower (p < 0.001). Three out of 5 recycled water (RW) samples were G. duodenalis positive. Three clusters with high posterior probabilities (PP) were found in the BI: Cluster 1: samples from wolves, wild boars, water and cattle together with database sequences of assemblages A, B and F, Cluster 2: samples from dogs, nutria and a jackal with sequences from assemblage D and Cluster 3: samples from cattle, wild boars, wolves and dogs with sequences from assemblage C and D. We suggest that wolves serve as reservoirs of G. duodenalis in this region. The finding of Giardia in RW suggests that this vehicle may further contaminate crops intended for human consumption as this water source is used for agricultural irrigation., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Eran Dvir reports financial support and administrative support were provided by Tel-Hai College. Eran Dvir reports a relationship with Tel-Hai College that includes: employment, (Copyright © 2024 Elsevier B.V. All rights reserved.)

Cryptosporidium spp. and G. duodenalis often infect humans, cats, and other mammals, causing diarrhea and being responsible for numerous outbreaks of waterborne and foodborne infections worldwide. The rapid increase in the number of pet cats poses a substantial public health risk. However, there were few reports about the infection of Cryptosporidium spp. and G. duodenalis infections in pet cats in Henan Province, central China. Thus, to understand the prevalence and genetic distribution of Cryptosporidium spp. and G. duodenalis in pet cats, and to evaluate the zoonotic potential, possible transmission routes and public health implications of isolates, fecal samples (n = 898) were randomly collected from pet cats in 11 cities in Henan Province, central China. Nested PCR based on the SSU rRNA gene and bg gene was used to the prevalence of Cryptosporidium spp. and G. duodenalis, respectively. The prevalence was 0.8 % (7/898) and 2.0 % (18/898) for Cryptosporidium spp. and G. duodenalis respectively. Additionally, the Cryptosporidium spp. positive isolates were identified as C. parvum subtype IIdA19G1 by gp60 gene. In the present study, the IIdA19G1 subtype was discovered in pet cats for the first time in China, enriching the information on the host type and geographical distribution of Cryptosporidium spp. in China. For G. duodenalis, a total of 18 G. duodenalis positive samples were identified, belonging to four assemblages: a zoonotic assemblage A1 (4/898), three host-specific assemblages C (8/898), D (5/898), and F (1/898). Interestingly, we found that pet cats infected with Cryptosporidium spp. and G. duodenalis are more likely to experience emaciation symptoms compared to the negative group. More importantly, the prevalence of Cryptosporidium spp. and G. duodenalis detected in the present study were low, but the subtype IIdA19G1 of Cryptosporidium spp. and the assemblages A1, C, D, and F of G. duodenalis have the potential for zoonotic transmission. Thus, we should focus on preventing and controlling the risk of cross-species transmission that may occur in pet cats in Henan Province., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier B.V.)

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 χ 2 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.

Diarrhea caused by zoonotic pathogens is one of the most common diseases in dairy calves, threatening the health of young animals. Humans are also at risk, in particular children. To explore the pathogens causing diarrhea in dairy calves, the present study applied PCR-based sequencing tools to investigate the occurrence and molecular characteristics of three parasites (Cryptosporidium spp., Giardia duodenalis, and Enterocytozoon bieneusi) and three bacterial pathogens (Escherichia coli, Clostridium perfringens, and Salmonella spp.) in 343 fecal samples of diarrheic dairy calves from five farms in Lingwu County, Ningxia Hui Autonomous Region, China. The total positive rate of these pathogens in diarrheic dairy calves was 91.0% (312/343; 95% CI, 87.9-94.0), with C. perfringens (61.5%, 211/343; 95% CI, 56.3-66.7) being the dominant one. Co-infection with two to five pathogens was found in 67.3% (231/343; 95% CI, 62.4-72.3) of investigated samples. There were significant differences (p < 0.05) in the positive rates of Cryptosporidium spp. and diarrheagenic E. coli among farms, age groups, and seasons. Two Cryptosporidium species (C. parvum and C. bovis) and five gp60 subtypes of C. parvum (IIdA15G1, IIdA20G1, IIdA19G1, IIdA14G1, and a novel IIdA13G1) were identified. Two assemblages (assemblage E and zoonotic assemblage A) of G. duodenalis and six ITS genotypes of E. bieneusi (J, Henan-IV, EbpC, I, EbpA, and ESH-01) were observed. Four virulence genes (eaeA, stx1, stx2, and st) of diarrheagenic E. coli and one toxin type (type A) of C. perfringens were detected. Our study enriches our knowledge on the characteristics and zoonotic potential of diarrhea-related pathogens in dairy calves., (© J.-Q. Zhao et al., published by EDP Sciences, 2024.)

Captive and free-living wildlife serve as significant hosts for Giardia duodenalis. Asiatic black bears, valued for their economic and medicinal importance, are extensively farmed in China and also prevalent in zoos. However, studies on G. duodenalis in these animals in China are limited. Here, 218 feces samples of Asiatic black bears were collected: 36 from a zoo in Heilongjiang Province, and 182 from a farm in Fujian Province. Nested PCR of the SSU rRNA gene, followed by sequencing, was employed to determine the frequency and assemblage distribution of G. duodenalis. Positive samples underwent further analysis through multilocus genotyping (MLG) by amplifying the genes for glutamate dehydrogenase (gdh), β-giardin (bg), and triosephosphate isomerase (tpi). Of the 218 samples, G. duodenalis was detected in 22 cases at the SSU rRNA gene locus, including three from Heilongjiang and 19 from Fujian. Three assemblages were identified: A (n = 1), B (n = 16), and E (n = 2) in Fujian; and B (n = 3) in Heilongjiang. Out of the 22 positive samples, 20, 19, and 9 were effectively amplified and sequenced across the tpi, gdh, and bg loci, respectively. Seven samples were genotyped successfully at all three loci, identifying MLG-B1 (n = 1), MLG-B2 (n = 1), and MLG-B3 (n = 1), MLG-B4 (n = 1), MLG-B5 (n = 2), and MLG-B6 (n = 1) as the six assemblage B MLGs. This study marks the first documentation of G. duodenalis in Asiatic black bears in captivity in Fujian and Heilongjiang. The identification of zoonotic assemblages A and B, along with E, underscores potential public health concerns., (© J. Chen et al., published by EDP Sciences, 2024.)

Almost all human giardiasis infections are caused by Giardia duodenalis assemblages A and B. Differentiation between human infections with these assemblages, as well as between single-assemblage (A or B) and mixed-assemblage (A and B) infections, is therefore needed to better understand the pathological impact of infection with either, or both, assemblages. We assessed the prevalence of G. duodenalis assemblages A and B using 305 fecal samples provided by school-age children situated along the southern shoreline of Lake Malawi. Concurrently, intestinal pathology data were also collected to test for association(s) between assemblage infection status and intestinal health. Prevalence of G. duodenalis infection was 39.3% by real-time polymerase chain reaction. Of all identified infections, 32% were single G. duodenalis assemblage A and 32% were single G. duodenalis assemblage B, whereas 33% were mixed-assemblage infections. Fifteen unique G. duodenalis assemblage A and 13 unique G. duodenalis assemblage B β-giardin haplotypes were identified. There was a positive association between single infection with G. duodenalis assemblage B and both self-reporting of abdominal pain (odds ratio [OR]: 3.05, P = 0.004) and self-reporting of diarrhea (OR: 3.1, P = 0.003). No association between single infection with assemblage A and any form of intestinal pathology was found. Additionally, there was a positive association between mixed-assemblage infections and self-reporting of abdominal pain (OR: 3.1, P = 0.002). Our study highlights the importance G. duodenalis assemblage typing and reaffirms the need for improved access to water, sanitation and hygiene infrastructure in rural areas of low- and middle-income countries.

Giardia duodenalis genotypes A and B have been reported in Colombia. The population consisted of 235 schoolchildren whose ages ranged from 2 to 10 yr of age from the municipalities of Soledad and Galapa in the department of Atlántico, Colombia. Fecal samples were obtained and then analyzed in triplicate using the sedimentation in formalin-ether (Ritchie's method) and direct examination techniques. Of the 235 fecal samples, 35 samples were positive for G. duodenalis; positive samples were concentrated in a sucrose gradient and sonicated for 3 cycles of 20 sec. DNA extraction was performed, and the parasites were genotyped by conventional PCR amplifying a region of the β-giardin gene. A general prevalence of G. duodenalis of 13.2% was found, and of these genotyped samples, 13 (56.7%) and 7 (20%) corresponded to genotype A, 1 (4.3%), and 3 (25%) corresponded to genotype B, and 9 (39.1%) and 2 (16.7%) were not defined, in the municipalities Soledad and Galapa, respectively. Additionally, 23 children were diagnosed with symptomatologic giardiasis, and 12 were asymptomatic; the most relevant symptoms were abdominal pain (7, 20%) and diarrhea (13, 56.7%). The nutritional status of children with Giardia genotypes A and B were as follows: 3 in a state of malnutrition (10%), 10 normal (33.3%), and 6 overweight and obese (20%) with genotype A, and 1 in a state of malnutrition (3.3%) and 3 normal (10%) with genotype B. The genotypes found in G. duodenalis did not show an association with nutritional status or with the clinical manifestations evaluated in schoolchildren., (© American Society of Parasitologists 2022.)

Giardia duodenalis is a common zoonotic intestinal pathogen. It has been increasingly reported in humans and animals; however, genotyping information for G. duodenalis in captive animals is still limited. This study was conducted to assess the prevalence and multilocus genotyping of G. duodenalis in captive animals in zoological gardens in Shanghai, China. A total of 678 fresh fecal samples were randomly collected from captive animals including non-human primates (NHPs) (n = 190), herbivores (n = 190), carnivores (n = 151), birds (n = 138) and reptiles (n = 9) in a zoo and were examined for the presence of G. duodenalis using nested polymerase chain reaction (nested PCR). All G. duodenalis positive samples were assayed with PCR followed by sequencing at β-giardin (bg), glutamate dehydrogenase (gdh) and triose phosphate isomerase (tpi) genes. In this study, 42 specimens (6.2%) were tested G. duodenalis-positive of the 678 fecal samples examined based on a single locus. A total of 30 (4.4%), 30 (4.4%) and 22 (3.2%) specimens were successfully amplified and sequenced at gdh, tpi and bg loci, respectively. Assemblages A and B were identified with assemblage B dominating in NHPs. Sequence analysis demonstrated that one, two and five new isolates were identified at bg, gdh and tpi loci. DNA sequences and new assemblage-subtypes of zoonotic G. duodenalis assemblages A and B were identified in the current study. Our data indicate the occurrence and molecular diversity of G. duodenalis and the potential zoonotic transmission in captive animals in China.

Background: Giardia duodenalis is a zoonotic protozoan parasite causing diarrhea through waterborne or fecal-oral infection. The cysts can live in the drinking water and cause pandemic diseases. In Taiwan, very little information is available regarding the epidemiology of G. duodenalis in domestic animals., Methods: Fecal samples were collected from cattle (n = 156) and pigs (n = 141) in Hualien country, eastern Taiwan. Detection and genotyping were done by microscopy examination of fecal samples and amplification of the β-giardin gene using nested PCR., Results: The prevalence of G. duodenalis infection was 19.87% for cattle (31/156) and 4.26% for pigs (6/141). Using nested PCR, 30 infected samples found in cattle belonged to Assemblage E, and one sample belonged to Assemblage D. For pigs, four samples belonged to Assemblage E, one belonged to Assemblage D, and another one belonged to Assemblage A. In addition, these results showed that G. duodenalis Assemblage A was detected in pigs and may cause zoonotic transmission., Conclusion: This is the first epidemiological investigation of G. duodenalis infection in animals in Hualien, Taiwan. These results could provide epidemiological information for disease control and public health protection., Competing Interests: Declaration of Competing Interest All authors declare that they have no conflicts of interest related to this article., (Copyright © 2020. Published by Elsevier B.V.)

This epidemiological study assesses the occurrence of enteric parasites in 4303 patients attended at two public hospitals in Ankara (Turkey) during 2018-2019. Microscopy was used as a screening test. Giardia duodenalis was also identified using a commercial ELISA for the detection of parasite-specific coproantigens. Giardia-positive samples by microscopy/ELISA were confirmed by real-time PCR and characterized using a multilocus genotyping scheme. Blastocystis sp. was genotyped in a sample subset. Blastocystis sp. (11.1%, 95% CI 11.4‒14.8%) and G. duodenalis (1.56%, 95% CI 1.22‒1.96) were the most prevalent pathogens found. Cryptosporidium spp., Entamoeba histolytica and intestinal helminths were only sporadically (<0.5%) found. For G. duodenalis, sequence (n = 30) analyses revealed the presence of sub-assemblages AII (23.3%), discordant AII/AIII (23.3%) and mixed AII + AIII (6.7%) within assemblage A, and BIII (10.0%), BIV (3.3%) and discordant BIII/BIV (23.3%) within assemblage B. Two additional sequences (6.7%) were assigned to the latter assemblage but sub-assemblage information was unknown. No associations between G. duodenalis assemblages/sub-assemblages and sociodemographic and clinical variables could be demonstrated. For Blastocystis sp., sequence (n = 6) analyses identified subtypes ST1, ST2 and ST3 at equal proportions. This is the first molecular characterization of G. duodenalis based on MLG conducted in Turkey to date.

Background: Giardia duodenalis is a leading cause of gastroenteritis worldwide. Humans are mainly infected by two different subtypes, i.e., assemblage A and B. Genotyping is hampered by allelic sequence heterozygosity (ASH) mainly in assemblage B, and by occurrence of mixed infections. Here we assessed the suitability of current genotyping protocols of G. duodenalis for epidemiological applications such as molecular tracing of transmission chains., Methodology/principal Findings: Two G. duodenalis isolate collections, from an outpatient tropical medicine clinic and from several primary care laboratories, were characterized by assemblage-specific qPCR (TIF, CATH gene loci) and a common multi locus sequence typing (MLST; TPI, BG, GDH gene loci). Assemblage A isolates were further typed at additional loci (HCMP22547, CID1, RHP26, HCMP6372, DIS3, NEK15411). Of 175/202 (86.6%) patients the G. duodenalis assemblage could be identified: Assemblages A 25/175 (14.3%), B 115/175 (65.7%) and A+B mixed 35/175 (20.0%). By incorporating allelic sequence heterozygosity in the analysis, the three marker MLST correctly identified 6/9 (66,7%) and 4/5 (80.0%) consecutive samples from chronic assemblage B infections in the two collections, respectively, and identified a cluster of five independent patients carrying assemblage B parasites of identical MLST type. Extended MLST for assemblage A altogether identified 5/6 (83,3%) consecutive samples from chronic assemblage A infections and 15 novel genotypes. Based on the observed A+B mixed infections it is estimated that only 75% and 50% of assemblage A or B only cases represent single strain infections, respectively. We demonstrate that typing results are consistent with this prediction., Conclusions/significance: Typing of assemblage A and B isolates with resolution for epidemiological applications is possible but requires separate genotyping protocols. The high frequency of multiple infections and their impact on typing results are findings with immediate consequences for result interpretation in this field., Competing Interests: The authors have declared that no competing interests exist.

The aim of this study was to detect and characterize Giardia lamblia in animals in the UAE. Eighty-seven fecal samples were tested for G. lamblia using the conserved fragment of small subunit (SSU)-rRNA by nested PCR. Giardia-positive isolates were genotyped for assemblages A and B using assemblage specific primers of the triosephosphate isomerase (tpi) gene. Thirty samples (34.5%) were positive for G. lamblia. Conversely, neither genotype A nor B were detected using tpi genotyping on the studied samples. Further investigations are required using higher number of samples including both human and animals in the country taking into consideration the analysis of other genotypes to provide more detailed understanding about the zoonotic transmission of this parasite.

Giardia duodenalis (syn. Giardia lamblia , Giardia intestinalis ) is the causative agent of giardiasis, one of the most common diarrheal infections in humans. Evolutionary relationships among G. duodenalis genotypes (or subtypes) of assemblage B, one of two genetic assemblages causing the majority of human infections, remain unclear due to poor phylogenetic resolution of current typing methods. In this study, we devised a methodology to identify new markers for a streamlined multilocus sequence typing (MLST) scheme based on comparisons of all core genes against the phylogeny of whole-genome sequences (WGS). Our analysis identified three markers with resolution comparable to that of WGS data. Using newly designed PCR primers for our novel MLST loci, we typed an additional 68 strains of assemblage B. Analyses of these strains and previously determined genome sequences showed that genomes of this assemblage can be assigned to 16 clonal complexes, each with unique gene content that is apparently tuned to differential virulence and ecology. Obtaining new genomes of Giardia spp. and other eukaryotic microbial pathogens remains challenging due to difficulties in culturing the parasites in the laboratory. Hence, the methods described here are expected to be widely applicable to other pathogens of interest and advance our understanding of their ecology and evolution. IMPORTANCE Giardia duodenalis assemblage B is a major waterborne pathogen and the most commonly identified genotype causing human giardiasis worldwide. The lack of morphological characters for classification requires the use of molecular techniques for strain differentiation; however, the absence of scalable and affordable next-generation sequencing (NGS)-based typing methods has prevented meaningful advancements in high-resolution molecular typing for further understanding of the evolution and epidemiology of assemblage B. Prior studies have reported high sequence diversity but low phylogenetic resolution at standard loci in assemblage B, highlighting the necessity of identifying new markers for accurate and robust molecular typing. Data from comparative analyses of available genomes in this study identified three loci that together form a novel high-resolution typing scheme with high concordance to whole-genome-based phylogenomics and which should aid in future public health endeavors related to this parasite. In addition, data from newly characterized strains suggest evidence of biogeographic and ecologic endemism.

Background: The waterborne pathogens Cryptosporidium spp., Giardia duodenalis, Enterocytozoon bieneusi and Cyclospora cayetanensis can cause intestinal diseases in humans. An understanding of their occurrence and transport in the environment is essential for accurate quantitative microbial risk assessment., Methods: A total of 238 influent samples were collected from four wastewater treatment plants (WWTPs) and 88 samples from eight sewer locations in Guangzhou, China. PCR-based tools were used to detect and genetically characterize Cryptosporidium spp., G. duodenalis and E. bieneusi. Eimeria spp. and Cyclospora spp. were also analyzed to assess the sources of Cryptosporidium spp., G. duodenalis and E. bieneusi in wastewater., Results: The overall occurrence rates in the WWTP and sewer samples were 14.3% (34/238) and 13.6% (12/88) for Cryptosporidium spp., 55.5% (132/238) and 33.0% (29/88) for G. duodenalis, 56.3% (134/238) and 26.1% (23/88) for E. bieneusi and 45.4% (108/238) and 47.7% (42/88) for Eimeria spp., respectively. Altogether, 11 Cryptosporidium species and genotypes, six G. duodenalis genotypes, 11 E. bieneusi genotypes and four C. cayetanensis were found, together with the presence of nine Eimeria species. The common occurrence of Cryptosporidium rat genotype IV, C. muris and Eimeria papillata and E. nieschulzi suggested that rodents were significant sources of the enteric pathogens detected in the wastewater samples., Conclusions: While the dominant Cryptosporidium spp. detected in the raw wastewater sampled in this study are not pathogenic to humans, the widely detected G. duodenalis assemblage A and E. bieneusi genotypes D and Type IV are well-known zoonotic pathogens. Further studies are needed to monitor the occurrence of these waterborne pathogens in WWTPs to better understand their transmission and environmental transport in China.

According to a few parasitological and epidemiological studies, Giardia is the most prevalent parasitic infection among pet dogs in the city of Medellín, the second-largest city in Colombia. This study determined the assemblages of Giardia in the fecal samples of dogs obtained from 18 veterinary centers of Medellín. One hundred fecal samples of dogs diagnosed with Giardia using microscopy were analyzed via nested polymerase chain reaction (PCR) using three genes (gdh, bg, and tpi). The PCR products were purified and sequenced, and phylogenetic analyses were conducted using the maximum likelihood algorithm of the three loci. From the 100 samples analyzed, 47 were Giardia-positive via PCR. Genotypes C and D were detected in six samples, neither of which were associated with human infection. However, the zoonotic potential of Giardia cannot be ruled out because of the small number of samples that could be sequenced for assemblage assignation., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Introduction: Giardia duodenalis, a unicellular, eukaryotic, and flagellated protozoan, presents two evolutionary forms in its life cycle, namely, trophozoites and cysts. It causes diarrhea in humans, dogs, cats, rodents, and ungulates. Despite being morphologically similar, the isolates of G. duodenalis are genetically diverse, affecting the stability and unanimity of taxonomic classification. Since different Giardia assemblages may occur within one isolate, multilocus genotyping is recommended for the genetic identification., Methodology: To determine the frequency of G. duodenalis infections in domiciled dogs in Cuiabá Municipality (State of Mato Grosso, Midwestern Brazil) and characterize its genetic variability, fecal samples were collected from 147 dogs., Results: Overall, 6.8% (10/147) of the samples presented cysts of G. duodenalis, which sequencing and genotypic characterization using tpi and gluD revealed assemblages C and A, genetic grouping of G. duodenalis. Only three samples amplified by tpi and one sample amplified by gluD., Conclusions: The risk factors age, gender, breed, diet and the presence of other dogs in the same house were not correlationated with giardiasis. The host-specific and zoonotic genotype warns of the risk of inter and intraspecies transmission and it provides, for the first time, information about genetic characterization of G. duodenalis isolates in dogs in Cuiabá, Midwest region of Brazil., Competing Interests: No Conflict of Interest is declared, (Copyright (c) 2020 Yolanda Paim Arruda Trevisan, Arleana do Bom Parto Ferreira de Almeida, Luciano Nakazato, Thabata dos Anjos Pacheco, Jessica Iglesias de Souza, Darlan Henrique Canei, Mariana Elisa Pereira, Maerle Oliveira Maia, Richard Campos Pacheco, Valeria Regia Franco Sousa.)

Background: In dogs, infections with Giardia duodenalis are mainly caused by assemblages C and D, but also by the potentially zoonotic assemblages A and B. The aims of this study were to assess differences in assemblages (i) between dogs living mainly in close proximity to humans (synanthropic dogs) versus dogs living mainly among other dogs, (ii) between samples of dogs with or without loose stool, and (iii) related to the amount of cysts shedding., Methods: One hundred eighty-nine qPCR Giardia positive fecal samples of dogs originating from four groups (household, sheltered, hunting, and dogs for which a veterinarian sent a fecal sample to a diagnostic laboratory) were used for genotyping. For this, multi-locus genotyping of beta-giardin, triose phosphate isomerase, and glutamate dehydrogenase and genotyping of SSU rDNA gene fragments were performed. Fecal consistency was scored (loose or non-loose stool), and cysts per gram of feces were determined with qPCR., Results: Assemblage D was the most prevalent in all groups, followed by the other canid assemblage C. Also, mixed C/D was common. In two (synanthropic) household dogs, the potentially zoonotic assemblage AI was present. Although occurrence of assemblage AI in household dogs was not significantly different from dogs living among other dogs (sheltered and hunting dogs), it was significantly higher compared to dogs for which a sample was sent to a diagnostic laboratory. Dogs with assemblage D shed significantly more cysts than dogs with other assemblages (except for mixed C/D results) or dogs in which no assemblage could be determined. None of the assemblages was significantly associated with loose stool., Conclusion: Not only do dogs mainly shed the canid Giardia duodenalis assemblages D and/or C, the numbers of cysts per gram for the canid assemblage D were also higher than for the potential zoonotic assemblage AI. Based on the assemblages shed by dogs, the risk to public health posed by dogs is estimated to be low, even though the dogs that shed AI were synanthropic household dogs. Loose stool in infected dogs was not associated with any particular Giardia assemblage.

Giardia lamblia is a common enteric pathogen associated with diarrheal diseases. There are some reports of G. lamblia infection among different breeds of cattle in recent years worldwide. However, it is yet to know whether cattle in Jiangxi province, southeastern China is infected with G. lamblia. The objectives of the present study were to investigate the prevalence and examine the multilocus genotypes of G. lamblia in cattle in Jiangxi province. A total of 556 fecal samples were collected from 3 cattle breeds (dairy cattle, beef cattle, and buffalo) in Jiangxi province, and the prevalence and genotypes of G. lamblia were determined by the nested PCR amplification of the beta-giardin (bg) gene. A total of 52 samples (9.2%) were positive for G. lamblia. The highest prevalence of G. lamblia was detected in dairy cattle (20.0%), followed by that in beef cattle (6.4%), and meat buffalo (0.9%). Multilocus sequence typing of G. lamblia was performed based on sequences of the bg, triose phosphate isomerase and glutamate dehydrogenase loci, and 22, 42, and 52 samples were amplifiable, respectively, forming 15 MLGs. Moreover, one mixed G. lamblia infection (assemblages A and E) was found in the present study. Altogether, 6 novel assemblage E subtypes (E41*-E46*) were identified for the first time. These results not only provided baseline data for the control of G. lamblia infection in cattle in this southeastern province of China, but also enriched the molecular epidemiological data and genetic diversity of G. lamblia in cattle.

Giardia duodenalis (syn. G. intestinalis, G. lamblia) is an important zoonotic parasite infecting livestock (including pigs) through ingesting cysts in contaminated food or water. This parasite has been classified into eight different genetic assemblages, A to H. Here, we examined the individual-level prevalence of G. duodenalis in domestic pig farms and confirmed host specificity by genotype comparisons. Samples were collected from southern and central Korea, between May 2017 and January 2019. DNA directly extracted from 745 pig fecal specimens were tested by PCR for G. duodenalis small subunit ribosomal RNA (ssu rRNA), glutamate dehydrogenase (gdh), and β-giardin gene sequences. Based on ssu rRNA PCR, 110 (14.8%) were positive for G. duodenalis. Infection risk was the highest in the fattener group (31/139, 22.3%) and during the autumn season (52/245, 21.2%: p < .001). No statistically significant differences in risk for infection were observed between fecal types (normal versus diarrheal). Fifty ssu rRNA samples, three gdh samples, and five β-giardin samples were successfully sequenced and genotyped. Ssu rRNA assemblage sequence analysis identified E (40.0%, 20/50), D (34.0%, 17/50), C (24.0%, 12/50), and A (2.0%, 1/50). The gdh locus identified three samples as assemblage E, and the β-giardin locus identified four samples as assemblage E and one as assemblage C. Assemblage A sequences obtained (ssu rRNA; MK430919) had 100% identity with Giardia sequences isolated from a Korean individual (AJ293301), indicating the potential of zoonotic transmission. Continuous management and monitoring for prevention of transmission and protection of animal and human health are essential., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Giardia duodenalis is a common zoonotic protozoan parasite with a broad host distribution. The main objectives of the present study were to determine the prevalence of giardiasis and to reveal the genetic and haplotype diversity of G. duodenalis in symptomatic cats in Turkey. Fecal samples were collected from cats (n = 102) with diarrhea that were admitted to different pet clinics in the Central Anatolia region of Turkey. All samples were analyzed by microscopic examination (ME), rapid immunochromatographic test (ICT), and PCR targeting the β-giardin (bg) loci of the parasite. Phylogenetic, haplotype, and network analyses of G. duodenalis based on the bg gene were carried out. Overall, G. duodenalis was detected in 70/102 (68.6%) of the cats with diarrhea by ME (38/102, 37.3%), ICT (51/102, 50%), and PCR (30/102, 29.4%). According to sequence analyses of the bg gene region, all isolates were identified as G. duodenalis assemblage B. Haplotype analyses revealed 2 known and 8 novel haplotypes for G. duodenalis assemblage B. This study provides first prevalence and genetic and haplotype diversity data on G. duodenalis assemblage B from cats in Turkey., (© American Society of Parasitologists 2020.)

We assessed the potential contribution of hospitals to contaminations of wastewater by enteric protists, including Cryptosporidium spp., Giardia duodenalis, and Enterocytozoon bieneusi in raw wastewater. Wastewater samples were collected from storage tanks in two hospitals and one associated wastewater treatment plant in Shanghai, China, from March to November 2009. Enteric pathogens were detected and identified using PCR and DNA sequencing techniques. Among a total of 164 samples analyzed, 31 (18.9%), 45 (27.4%), and 122 (74.4%) were positive for Cryptosporidium spp., G. duodenalis, and E. bieneusi, respectively. Altogether, three Cryptosporidium species, four G. duodenalis assemblages, and 12 E. bieneusi genotypes were detected. Cryptosporidium hominis, G. duodenalis sub-assemblage AII, and E. bieneusi genotype D were the dominant ones in wastewater from both hospitals and the wastewater treatment plant. A similar distribution in genotypes of enteric pathogens was seen between samples from hospitals and the wastewater treatment plant, suggesting that humans are one of the major sources for these pathogens and hospitals are important contributors of enteric parasites in urban wastewater. Data from this study might be useful in the formulation of preventive measures against environmental contamination of waterborne pathogens.

Little is known of the prevalence and genetic identity of Giardia duodenalis in sheep in Algeria. The present study aimed at characterizing G. duodenalis in lambs up to 6 months of age in Djelfa, Algeria. A total of 346 fecal specimens were collected from 28 farms and screened for G. duodenalis cysts by zinc sulfate flotation microscopy, and positive specimens were confirmed using a direct immunofluorescence assay. Microscopy-positive specimens were analyzed by PCR and sequence analysis of the triosephosphate isomerase and glutamate dehydrogenase genes to determine G. duodenalis assemblages. Coprological examination indicated that the overall infection rate was 7.0% (24/346). Lambs under 3 months of age had higher infection rate (18/197, 9.0%) than older (6/149, 4.0%) animals, and animals with diarrhea (7/44, 16.0%) had higher infection rate than animals without diarrhea (17/302, 5.6%). PCR sequence analyses of the 15 G. duodenalis isolates revealed the presence of assemblages A in 6 isolates, assemblage E in 7 isolates, and both in 2 isolates. Assemblage A was only found in pre-weaned lambs with diarrhea, while assemblage E was mostly found in post-weaned lambs without diarrhea. The assemblage E isolates from sheep were genetically related to those from cattle in Algeria, while assemblage A isolates were from a well-known subtype prevalent in humans. Data generated from the study improve our understanding of the transmission of G. duodenalis in Algeria.

Giardia duodenalis is a common intestinal protozoa, which can cause the occurrence of diarrhea, weight loss, and even death in animals or human, this threatens the husbandry industry and public health. It can infect virtually humans and all domestic animals including sheep. Tan sheep is one of the most important sheep breeds, which is short-tailed indigenous sheep breed used for production of high quality meat and pelts in China. However, there are no report regarding the occurrence and multilocus genotyping of G. duodenalis in Tan sheep in northwestern China. Thus, the objective of the present study was to investigate the prevalence and multilocus genotypes of G. duodenalis in Tan sheep. 1014 fecal samples were collected from Tan sheep from Ningxia Hui Autonomous Region, and three loci (β-giardin (bg), glutamate dehydrogenase (gdh) and triosephosphate isomerase (tpi) genes) were amplified by nested PCR. The prevalence of G. duodenalis in Tan sheep was 14.5% (147/1014), two assemblages (assemblage A, n = 43; and E, n = 90) were detected, including one novel assemblage A at bg locus, one novel assemblage A at tpi locus, and 10 and 11 novel subtypes of assemblage E were detected at the bg and gdh loci, respectively. One MLGs was formed based on sequence variation among the three loci. Moreover, 9 Tan sheep were infected with two assemblages (A and E) based on the three loci. These findings expand the host range of G. duodenalis and revealed genetic diversity of G. duodenalis assemblages in Tan sheep., Competing Interests: Declaration of Competing Interest The authors declare that they have no competing interests., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Giardia intestinalis is an intestinal protozoan most commonly found in humans. It has been grouped into 8 assemblages (A-H). Markers such as the glutamate dehydrogenase gene, triose phosphate isomerase and beta-giardin (β-giardin) have been widely used for genotyping. In addition, different genetic targets have been proposed as a valuable alternative to assess diversity and genetics of this microorganism. Thus, our objective was to evaluate new markers for the study of the diversity and intra-taxa genetic structure of G. intestinalis in silico and in DNA obtained from stool samples. We analysed nine constitutive genes in 80 complete genome sequences and in a group of 24 stool samples from Colombia. Allelic diversity was evaluated by locus and for the concatenated sequence of nine loci that could discriminate up to 53 alleles. Phylogenetic reconstructions allowed us to identify AI, AII and B assemblages. We found evidence of intra- and inter-assemblage recombination events. Population structure analysis showed genetic differentiation among the assemblages analysed.

Background: Giardia intestinalis is a common cause of gastrointestinal illness especially in children of developing countries. Giardia assemblages A and B are the major human infective genotypes., Objective: The present study aimed to investigate the role of water supply in the epidemiology of giardiasis via genotyping G. intestinalis detected in diarrheic children and in water samples in Egyptian rural areas., Methods: Stool samples of 100 diarrheic children, 40 drinking water samples and 10 raw water samples of canals were examined microscopically for Giardia. DNA was extracted from microscopically positive faecal samples and from all of the collected water samples. Amplification of Giardia tpi gene was performed by a nested PCR using assemblage A- and assemblage B-specific primers. Giardia gdh gene was amplified by a heminested PCR. Giardia genotypes were determined by restriction fragment polymorphism (RFLP) analysis of the amplified products. Sequencing of the amplified products was performed in two faecal and two water samples RESULTS: Giardia intestinalis was detected in 24 children, in none of the drinking water samples and in all canal water samples. Giardia sub-assemblage AII was identified in all stool and raw water samples. The RFLP pattern was confirmed in sequenced samples., Conclusion: The presence of the same Giardia sub-assemblage in diarrheic children and in raw water samples shows by molecular evidence the potential for waterborne dissemination of Giardia in Egypt. Further studies are needed to monitor cyst levels and infectivity of the genotype detected in water for risk assessment and management.

Background: Approximately 30% of children worldwide are infected with gastrointestinal parasites. Depending on the species, parasites can disrupt intestinal bacterial microbiota affecting essential vitamin biosynthesis., Methods: Stool samples were collected from 37 asymptomatic children from a previous cross-sectional Argentinian study. A multi-parallel real-time quantitative PCR was implemented for Ascaris lumbricoides, Ancylostoma duodenale, Necator americanus, Strongyloides stercoralis, Trichuris trichiura, Cryptosporidium spp., Entamoeba histolytica and Giardia duodenalis. In addition, whole-genome sequencing analysis was conducted for bacterial microbiota on all samples and analyzed using Livermore Metagenomic Analysis Toolkit and DIAMOND software. Separate analyses were carried out for uninfected, Giardia-only, Giardia + helminth co-infections, and helminth-only groups., Results: For Giardia-only infected children compared to uninfected children, DNA sequencing data showed a decrease in microbiota biodiversity that correlated with increasing Giardia burden and was statistically significant using Shannon's alpha diversity (Giardia-only > 1 fg/µl 2.346; non-infected group 3.253, P = 0.0317). An increase in diversity was observed for helminth-only infections with a decrease in diversity for Giardia + helminth co-infections (P = 0.00178). In Giardia-only infections, microbiome taxonomy changed from Firmicutes towards increasing proportions of Prevotella, with the degree of change related to the intensity of infection compared to uninfected (P = 0.0317). The abundance of Prevotella bacteria was decreased in the helminths-only group but increased for Giardia + helminth co-infections (P = 0.0262). Metagenomic analysis determined cobalamin synthesis was decreased in the Giardia > 1 fg/µl group compared to both the Giardia < 1 fg/µl and the uninfected group (P = 0.0369). Giardia + helminth group also had a decrease in cobalamin CbiM genes from helminth-only infections (P = 0.000754)., Conclusion: The study results may provide evidence for an effect of parasitic infections enabling the permissive growth of anaerobic bacteria such as Prevotella, suggesting an altered capacity of vitamin B12 (cobalamin) biosynthesis and potential impact on growth and development in children .

Limited data are available on infection rates and genetic identity of Enterocytozoon bieneusi and Giardia duodenalis in horses and donkeys. In this study, 865 fecal specimens were collected from donkeys (n = 540) and horses (n = 325) in three provinces and autonomous regions in northern China during 2015-2019. Enterocytozoon bieneusi was detected and genotyped by PCR and sequence analyses of the ribosomal internal transcribed spacer (ITS) and G. duodenalis was detected and genotyped by PCR and sequence analyses of the β-giardin, glutamate dehydrogenase, and triosephosphate isomerase genes. The overall infection rates of E. bieneusi and G. duodenalis were 21.9% (118/540) and 11.5% (62/540) in donkeys, and 7.4% (24/325) and 2.8% (9/325) in horses, respectively. These differences in infection rates of E. bieneusi and G. duodenalis between donkeys and horses were significant (χ 2  = 30.9, df = 1, P < 0.0001; χ 2  = 20.4, df = 1, P < 0.0001, respectively). By age, the 28.9% infection rate of E. bieneusi in donkeys under 6 months was significantly higher than that in animals over 6 months (6.0%; χ 2  = 35.2, df = 1, P < 0.0001). In contrast, donkeys of 6-12 months had higher infection rate (35.9%) of G. duodenalis than donkeys under 6 months (9.9%; χ 2  = 22.1, df = 1, P < 0.0001) and over 12 months (8.7%; χ 2  = 17.3, df = 1, P < 0.0001). In horses, animals of > 12 months had significantly higher infection rate (31.1%) of E. bieneusi than horses under 6 months (3.4%; χ 2  = 29.4, df = 1, P < 0.0001) and 6-12 months (3.8%; χ 2  = 26.1, df = 1, P < 0.0001). Twenty genotypes of E. bieneusi were detected, including six known ones and 14 new genotypes. Among them, nine genotypes in 45% E. bieneusi-positive specimens belonged to the zoonotic group 1. Similarly, three G. duodenalis assemblages were detected, including A (in 2 horses and 30 donkeys), B (in 6 horses and 29 donkeys), and E (in 1 horse); three donkeys had coinfections of assemblages A and B. The assemblage A isolates identified all belong to the sub-assemblage AI. These results indicate that unlike in other farm animals, there is a common occurrence of zoonotic E. bieneusi and G. duodenalis genotypes in horses and donkeys.

Giardia duodenalis is a zoonotic enteric parasite that can infect humans and a number of animal species including rabbits with a worldwide distribution. Infection with G. duodenalis can cause serious public health problems and significant economic losses to animal husbandry. So accurate understanding of the prevalence and genotype distribution of G. duodenalis in rabbits is necessary. In the present study, a total of 616 fecal samples were collected from rabbits in Shandong province, eastern China, and examined in G. duodenalis prevalence and genotypes by nested PCR amplification of β -giardin (bg), glutamate dehydrogenase (gdh), and triosephosphate isomerase (tpi) gene loci of G. duodenalis . Sixty-nine (11.2%) of the examined rabbit fecal samples were G. duodenalis- positive. Of them, the prevalence of G. duodenalis is 8.4% (41/490) in Rizhao city and 22.2% (28/126) in Weihai city. Breeds, region, and feeding modes were highly correlated with G. duodenalis infection in rabbits. Moreover, three genotypes (assemblages A, B, and E) were identified in rabbits at three gene loci, and the assemblage E was the dominant genotype, while the assemblage A was reported in rabbits in China for the first time. It is noticeable that two rabbits were found to be infected with two different G. duodenalis assemblages (assemblages A and E, assemblages B and E, respectively). These findings enrich the genotype distribution of G. duodenalis in rabbits and provide baseline data for preventing and controlling G. duodenalis infection in rabbits in eastern China., Competing Interests: The authors declare that they have no conflicts of interest., (Copyright © 2020 Tao-Shan Li et al.)

Giardia duodenalis is one of the main enteric pathogens associated with diarrheal disease. In developing countries, giardiasis is a major public health concern, particularly in children under five years of age. This study aimed to evaluate the occurrence and genetic diversity of G. duodenalis causing human infections in Shushtar County, Southwestern Iran. Individual faecal specimens were collected from 1,163 individuals (male/female ratio: 0.9; age range 2-75 years) with (n = 258) and without (n = 905) gastrointestinal symptoms living in rural and urban settings during the period 2017-2018. Conventional (sucrose flotation and microscopy) methods were used for the initial detection of G. duodenalis cysts in faecal specimens. Microscopy-positive samples were confirmed by PCR amplification and sequencing of the small subunit rRNA (ssu rRNA) gene of the parasite. A multilocus genotyping (MLG) scheme targeting the triose phosphate isomerase (tpi), the glutamate dehydrogenase (gdh), and the beta-giardin (bg) genes was used for genotyping purposes. Giardia duodenalis cysts were detected in 7.7% (90/1,163) of samples by microscopy, of which 82 were confirmed by ssu-PCR. Successful amplification and sequencing results were obtained for 23.2% (19/82), 9.8% (8/82), and 8.5% (7/82) of the confirmed samples at the tpi, gdh, and bg loci, respectively. MLG data for the three loci were available for two samples only. Out of the 24 samples genotyped at any loci, 50% (12/24) were identified as assemblage A and the remaining half as assemblage B. Overall, AII was the most prevalent sub-assemblage detected (41.7%, 10/24), followed by BIII (25.0%, 6/24), discordant BIII/BIV (5/24) or AII/AIII (2/24) sequences, and BIV (1/24). No significant correlation was demonstrated between a given assemblage/sub-assemblage and the occurrence of clinical symptoms. No genotypes adapted to animal hosts other than humans (e.g. assemblages C-F) were found circulating in the investigated human population, suggesting that transmission of human giardiasis in this Iranian region is primarily of anthroponotic nature. Further molecular-based studies are needed to confirm and expand these results, and to ascertain the presence and public health relevance of the parasite in environmental (e.g. drinking water) samples., Competing Interests: The authors have declared that no competing interests exist.

Background: Giardiasis is one of the most common gastrointestinal infections of humans and animals attributable to complex of eight morphologically identical genetic assemblages, further divided into sub-assemblages. Disease is common for a wide range of hosts and genetic characterization is needed for better understanding of multifaceted epidemiology for this protozoan parasite. The aim of this study was to identify genetic heterogeneity in assemblages and sub-assemblages of Giardiaduodenalis circulating among the children population living in deprived socioeconomic conditions., Methods: A total of 333 stool samples from children in eastern Slovakia were collected during the period of 2015-2016 and analysed by molecular methods. Molecular characterization of G. duodenalis was performed by sequence analysis of triose phosphate isomerase gene (tpi) and glutamate dehydrogenase gene (gdh)., Results: G. duodenalis DNA was detected in 21 samples (6.3%), out of which 14 isolates (66.7%) belonged to assemblage B, 4 isolates (19.0%) to sub-assemblage AII and 3 isolates (14.3%) corresponded to assemblage F. As regards the determination of sub-assemblages of assemblage B, 4 isolates were characterized as sub-assemblage BIII and 6 isolates as sub-assemblage BIV., Conclusion: This study is the first finding of cat specific assemblage F in man not only in Slovakia, but also in Europe. The absence of molecular data about G. duodenalis in companion animals in Slovakia establishes a strong need for further investigation for potential sources of giardiasis and understanding the epidemiology will help to improve the preventive strategies in eradication of infection in this population., (Copyright © 2018. Published by Elsevier B.V.)

Giardia duodenalis is one of the most important intestinal parasites globally, especially in children, and in Cuba is the leading cause of chronic paediatric diarrhoea in this population. G. duodenalis is composed of eight genetic groups (or assemblages), two of which (A and B) are apparently zoonotic, occurring in both humans and other animals. However, consensus on the most appropriate genotyping scheme for optimal characterization of G. duodenalis isolates is lacking. In this article we present the results of three descriptive observational studies conducted in Havana, Cuba between 2010 and 2013, with the aim of comparing the results from molecular (PCR) approaches targeting different genes in order to assign with confidence 224 isolates of G. duodenalis to the correct assemblages. In each sub-study, following DNA isolation by the phenol/chloroform/isoamyl alcohol extraction method, PCR targeting the triose phosphate isomerase (tpi) gene was used for molecular characterization, as well as one additional PCR-method targeting another gene or pair of genes. DNA amplification was obtained in 87%, 83%, and 80% in the three sub-studies. Although excellent agreement (kappa index = 1) was recorded between results from some pairs of genes, for other combinations only moderate or substantial agreement was achieved. These results highlight the importance of interpretation of genotyping data, especially when single genetic markers are used. From the results of our studies, PCR targeting a combination of the tpi gene and the intergenic spacer region of rDNA may be a useful approach for the molecular characterization of G. duodenalis isolates., (Copyright © 2019 The Authors. Published by Elsevier Inc. All rights reserved.)

Giardia lamblia has proved to be the most common intestinal protozoan parasite in humans that causes giardiasis. Given the high mutations in the genome of this parasite, the present study was conducted to determine the prevalence of Giardia lamblia subtypes and their relationship with clinical symptoms in patients who appear to have giardiasis. In 69 stool specimens with an appropriate number of giardia cysts, the DNA was first purified, and the genotype was then determined based on the glutamate dehydrogenase (gdh) gene sequence using PCR-RFLP. Data were collected on the clinical symptoms of the patients through a questionnaire, and their relationship with molecular results was studied. Four samples (5.8%) were found to be of subtype AI, 51 (73.9%) of subtype AII and 14 (20.3 %) of subtype BIII. No BIV subtype was found in the present study. A correlation was observed between Giardia lamblia genotypes (AI, AII and BIII) and abdominal pain. All of the people (100%) who had the AI genotype (i.e. the zoonosis subtype) had been losing weight. There was a significant correlation between weight loss and the AI subtype. All of the people (100%) infected with the BIII genotype experienced diarrhea, and this genotype was found to be associated with diarrhea. The present study found abdominal pain to be the most common symptom of giardiasis in Ilam province. Moreover, humans were found to be the main reservoir for giardia lamblia, although zoonosis subtypes such as AI and BIII still exist in the region and pose a risk for a giardiasis epidemic., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

In the present study, a total of 615 fecal samples from veterinary clinics were screened by microscopy for the presence of Cryptosporidium and Giardia oocysts. Molecular genotyping of Cryptosporidium and Giardia were carried out using PCR and sequence analysis. Overall, Cryptosporidium and Giardia oocysts were detected in the 0.6% (2/315) and 1.9% (6/315) of dogs and in the 0.7% (2/300) and 1.3% (4/300) of cats, respectively. Sequencing revealed the presence of C. canis (n = 2) in dogs and C. felis (n = 2) in cats. Moreover, G. intestinalisassemblage D (n = 2), C (n = 3) and A, sub-assemblage AII (n = 1) were identified in dogs; G. intestinalis assemblage F (n = 3) and assemblage A, sub-assemblage AI (n = 1) were identified in cats. The highest prevalence of Giardia was observed in dogs younger than one year (6/315), and in those with diarrhea (p < 0.05). Data of the study suggest that dogs and cats play a minor role in the zoonotic transmission of cryptosporidiosis and giardiasis in Southwestern Iran.

Background: Cryptosporidium spp. and Giardia duodenalis are major intestinal pathogens that can cause diarrheal diseases in humans, especially children. Enterocytozoon bieneusi is another parasite which can cause gastrointestinal tract disorders, with diarrhea being the main clinical symptom. However, few genetic studies of these parasites in pediatric inpatients in China have been published., Methods: To assess the genetic characteristics and epidemiological status of these parasites, a total of 2284 fecal samples were collected from children in the pediatric departments of three hospitals in Zhengzhou, central China, and screened for these protozoans with PCR, based on the small subunit ribosomal RNA (SSU rRNA) genes of Cryptosporidium spp. and G. duodenalis and the internal transcribed spacer (ITS) of E. bieneusi., Results: Six (0.26%), 14 (0.61%), and 27 (1.18%) of the samples were positive for Cryptosporidium spp., G. duodenalis and E. bieneusi, respectively. Of the 12 successfully sequenced G. duodenalis isolates, four were identified as assemblage A and eight as assemblage B. In subtype and multilocus genotype (MLG) analyses, C. parvum IIdA19G1 (n = 4) and two novel G. duodenalis MLGs belonging to subassemblage AII (n = 3) and BIV (n = 5) were successfully identified. The E. bieneusi isolates included genotypes D (n = 17), J (n = 2), PigEBITS7 (n = 1), BEB6 (n = 1), and CM8 (n = 1). This is the first report of C. parvum subtype IIdA19G1 in HIV-negative children and E. bieneusi genotype CM8 in humans., Conclusions: The dominance of zoonotic C. parvum subtype IIdA19G1 indicates that this parasite is turning into zoonotic origin from human-to-human transmission. The phylogenetic analysis also revealed the zoonotic origins and anthroponotic transmission potential of G. duodenalis and E. bieneusi, suggesting more efforts must be made to minimize the threat these pathogens pose to public health.

Enterocytozoon bieneusi and Giardia duodenalis are important opportunistic enteric zoonotic pathogens that cause diarrhoea and intestinal diseases in animals and humans. China is the largest producer of pigs, but whether Tibetan pigs, a unique pig breed in Tibet, are infected with E. bieneusi and G. duodenalis is unknown. Therefore, we conducted a molecular epidemiological survey to determine the prevalence of E. bieneusi and G. duodenalis in Tibetan pigs in Tibet, China, and identified the genotypes of these causative agents. A total of 345 faecal specimens were collected from Tibetan pigs from three Tibet counties (Milin, Cuona and Gongbujiangda), examined by nested PCR and sequenced utilizing genetic markers in the ribosomal internal transcribed spacer (ITS) region of the rRNA and glutamate dehydrogenase (gdh) gene for E. bieneusi and G. duodenalis, respectively. Moreover, using multilocus sequence typing, the subtypes of E. bieneusi were identified based on four loci (MS1, MS3, MS4 and MS7). A total of 41 (11.88%) faecal samples from Tibetan pigs were E. bieneusi-positive, and 2 (0.58%) were G. duodenalis-positive. The multivariate logistic regression analysis showed that age was considered a risk factor for Tibetan pig infection of E. bieneusi. Two novel (GB11, GB31) and four known E. bieneusi genotypes (EbpC, EbpD, PigEBITS5 and CHS12) were identified and were all classified as zoonotic group 1 according to the phylogenetic analysis. Two MLGs (MLGI and MLGII) were further identified in the E. bieneusi EbpC genotype by multilocus sequence typing analysis. In addition, two G. duodenalis assemblages (D and E) were found in the present study. To our knowledge, the current study is the first to detect the prevalence and perform genetic characterization of G. duodenalis in Tibetan pigs in Tibet, China. The results could provide essential data for controlling E. bieneusi and G. duodenalis infections in Tibetan pigs that are in contact with other animals and humans, as Tibetan pigs could be a potential source for human infection by these pathogens., (Copyright © 2019. Published by Elsevier B.V.)

Giardia duodenalis (syn. G. intestinalis, G. lamblia), a protozoan parasite that infects most mammals (including humans), has recently been classified into eight different groups/assemblages (A to H) based on host range and genetic characteristics. Assemblages A and B have been reported in humans and in many other animals, including dogs. This study analyzed Giardia infections, implementing multilocus genotyping, including the 18S rRNA, gdh, and β-giardin genes, in sheltered, companion, and special-purpose dogs throughout Korea. Of 640 dog stool samples, 99 (15.5%) were G. duodenalis-positive; this prevalence was similar to that previously reported in Korean calves. In addition, the prevalence was significantly higher in the southern region, and lower in non-sheltered dogs, older dogs, and in the autumn season. Phylogenetic analysis revealed that all G. duodenalis-positive samples belonged to assemblages C and D. Although no G. duodenalis assemblages A and B (mainly found in humans) were identified in this study, assemblage C was detected in the main types of dogs in contact with humans. Assemblage C, thought to be dog-specific, was recently identified in humans. As Giardia (including zoonotic assemblages) is distributed throughout Korea, dog living environments and the potential for dog-human transmission should be monitored., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Giardia duodenalis is one of the most common intestinal parasites in humans as well as livestock and wildlife. It is of both public and veterinary health importance in developing nations. A molecular survey of Giardia duodenalis assemblages in ruminants from Yazd Province, Iran was conducted on 484 animal faecal samples collected per rectum from slaughtered ruminants including 192 cattle, 192 sheep and 100 goats from June to November 2017. Species-specific and assemblage-specific PCRs for assemblages A, B and E at the triose phosphate isomerase (tpi) gene were performed, and samples positive for Giardia were confirmed by sequencing. In total, 25 (5.16%) of examined faecal samples including eight cattle (4.2%), twelve sheep (6.2%) and five goats (5%) were infected with G. duodenalis. Assemblage-specific PCR detected G. duodenalis assemblage E in seven faecal samples (six in sheep and one in a goat). Assemblages A and B were not detected. This study provides the first insight into Giardia infection in slaughtered livestock in Iran. Although the prevalence of infection with Giardia in this hot-arid area of Iran was low, educating people about direct contact with livestock such as farmers and abattoirs workers about this zoonotic infection is important., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Cryptosporidium parvum and Giardia duodenalis are the main diarrhea-causing parasitic pathogens; however, their prevalence in Korea is unknown. Here, we conducted a survey to determine the prevalence and genotype distribution of these 2 pathogens causing acute diarrhea in 8,571 patients hospitalized in 17 Regional Institute of Health Environment sites in Korea, during 2013-2016. C. parvum and G. duodenalis were detected and genotyped by nested PCR, and the isolate were molecularly characterized by sequencing the glycoprotein 60 (Gp60) and β-giardin genes, respectively. The overall prevalence of C. parvum and G. duodenalis was 0.37% (n=32) and 0.55% (n=47), respectively, and both pathogens were more prevalent in children under 9 years old. Molecular epidemiological analysis showed that the C. parvum isolates belonged to the IIa family and were subtyped as IIaA13G2R1, IIaA14G2R1, IIaA15G2R1, and IIaA18G3R1. Analysis of the β-giardin gene fragment from G. duodenalis showed that all positive strains belong to assemblage A. This is the first report on the molecular epidemiology and subtyping of C. parvum and G. duodenalis in such a large number of diarrheal patients in Korea. These results highlight the need for continuous monitoring of these zoonotic pathogens and provide a basis for implementing control and prevention strategies. Further, the results might be useful for epidemiological investigation of the source of outbreak.

Background: Although the distribution of Giardia duodenalis genotypes in humans has been increasingly reported in recent years, data on possible differences in pathogen transmission between age groups and virulence between genotypes are scarce. The purpose of this study is to investigate the genetic diversity of G. duodenalis in humans in Spain and compare the distribution of G. duodenalis assemblages A and B between children and adults and clinical presentations between the two genotypes., Methods: In the present study, 125 microscopy-positive fecal samples were collected from humans in Spain over a 7-year period. PCR and sequence analyses of the triosephosphate isomerase, β-giardin and glutamate dehydrogenase genes were used to identify the multilocus genotypes of G. duodenalis., Results: Sequence analysis of three genetic loci identified both G. duodenalis assemblages A (29) and B (66), with co-infections of the two in two patients. Among the sequences obtained in this study, four multilocus genotypes (MLGs) of the sub-assemblage AII were observed within assemblage A. In contrast, 19 MLGs were detected within assemblage B due to the high sequence diversity at each locus. One MLG, however, was found in 51.9% (27/52) of assemblage B samples. Children were more commonly infected by assemblage B (44/53 or 83%) than adults (22/42 or 52.4%; χ 2  = 10.371, df = 1, P = 0.001). Asymptomatic infection was more common in patients with assemblage A (4/29 or 13.8%) than in those with assemblage B (1/66 or 1.5%; χ 2  = 6.091, df = 1, P = 0.029), and the frequency of abdominal pain occurrence was higher in assemblage B patients (65/66 or 98.5%) than assemblage A patients (25/29 or 86.2%; χ 2  = 6.091, df = 1, P = 0.029)., Conclusions: These results illustrate the existence of differences in genotype distribution between children and adults and clinical presentations between G. duodenalis genotypes. They are useful in understanding the transmission of G. duodenalis in humans in Spain.

Giardia duodenalis is a common intestinal parasite in humans and other mammals, and it causes major public and veterinary health problems worldwide. China is a major pig-raising country, and studies on Giardia in pigs have important public health significance. The present study was conducted to investigate the prevalence of Giardia and assess its genetic characterization. A total of 93 samples were collected from two farms in Shanghai. The presence of Giardia was determined using PCR and sequence analysis of glutamate dehydrogenase, beta-giardin and triose phosphate isomerase genes. The average prevalence of G. duodenalis infection was 26.88% (25/93) in the pigs, with 28.13% (18/64) in farm 1 vs 24.14% (7/29) in farm 2. All the PCR-positive products were successfully sequenced, and assemblage E was more prevalent. Zoonotic assemblages A and B and canine-specific assemblage C were identified in farm 1, whereas, only assemblage E was detected in farm 2. Interestingly, two pig isolates showed 100% homology with human-derived isolates from Australia and China at the bg and tpi loci respectively. Pigs infected with Giardia infect humans by polluting the environment; whether pigs are a potential environmental source of the human pathogen in China requires more epidemiological data.