The zoonotic transmission of Giardia and Cryptosporidium

The molecular characterisation of Giardia and Cryptosporidium has given rise to a more epidemiological meaningful and robust taxonomy. Importantly, molecular tools are now available for 'typing' isolates of the parasites directly from clinical and environmental samples. As a consequence, information on zoonotic potential has been obtained although the frequency of zoonotic transmission is still poorly understood. Analysis of outbreaks and case-control studies, especially when coupled with genotyping data, is slowly providing information on the public health significance of zoonotic transmission. Such studies support the hypothesis that Cryptosporidium hominis is spread only between humans but that the major reservoir for Cryptosporidium parvum is domestic livestock, predominantly cattle, and that direct contact with infected cattle is a major transmission pathway along with indirect transmission through drinking water. The situation is less clearcut for Giardia duodenalis but the evidence does not, in general, support zoonotic transmission as a major risk for human infections. However, for both parasites there is a need for molecular epidemiological studies to be undertaken in well-defined foci of transmission in order to fully determine the frequency and importance of zoonotic transmission.     

Giardiasis as a re-emerging infectious disease and its zoonotic potential

The reasons for considering giardiasis as a re-emerging infectious disease are presented, with emphasis on Giardia infections in child care centres, livestock and pets, and the role of zoonotic transmission. However, the aetiology and control of giardiasis is complicated by the genetic and phenotypic variability of Giardia species infective to mammals. Of particular significance has been the uncertainty about host specificity and the question of zoonotic transmission. The recent application of molecular characterisation procedures based on PCR has made an enormous contribution to an understanding of the genetic structure of Giardia populations, and this is reviewed in the context of the zoonotic transmission and molecular epidemiology of Giardia infections.     

Molecular characterizations of Cryptosporidium, Giardia, and Enterocytozoon in humans in Kaduna State, Nigeria

The use of molecular diagnostic tools in epidemiological investigations of Cryptosporidium, Giardia, and Enterocytozoon has provided new insights into their diversity and transmission pathways. In this study, 157 stool specimens from 2-month to 70-year-old patients were collected, a polymerase chain reaction (PCR)-restriction fragment length polymorphism (RFLP) analysis of the small subunit (SSU) rRNA gene was used to detect and differentiate Cryptosporidium species, and DNA sequence analysis of the 60 kDa glycoprotein (gp60) gene was used to subtype Cryptosporidium hominis and Cryptosporidium parvum. Giardia duodenalis, and Enterocytozoon bieneusi in the specimens were detected using PCR and sequence analysis of the triosephosphate isomerase (tpi) gene and internal transcribed spacer (ITS), respectively. C. hominis and C. parvum were found in two (1.3%) and one (0.6%) specimen respectively, comprising of Ia and IIe (with 8 nucleotide substitutions) subtype families. The G. duodenalis A2 subtype was detected in five (3.2%) specimens, while four genotypes of E. bieneusi, namely A, type IV, D and WL7 were found in 10 (6.4%) specimens. Children aged two years or younger had the highest occurrence of Cryptosporidium (4.4%) and Enterocytozoon (13.0%) while children of 6 to 17 years had the highest Giardia infection rate (40.0%). No Cryptosporidium, Giardia, and Enterocytozoon were detected in patients older than 60 years. Enterocytozoon had high infection rates in both HIV-positive (3.3%) and HIV-negative (8.3%) patients. Results of the study suggest that anthroponotic transmission may be important in the transmission of Cryptosporidium spp. and G. duodenalis while zoonotic transmissions may also play a role in the transmission of E. bieneusi in humans in Kaduna State, Nigeria.     

Cryptosporidium and Giardia in marine-foraging river otters (Lontra canadensis) from the Puget Sound Georgia Basin ecosystem

Species of Cryptosporidium and Giardia can infect humans and wildlife and have the potential to be transmitted between these 2 groups; yet, very little is known about these protozoans in marine wildlife. Feces of river otters (Lontra canadensis), a common marine wildlife species in the Puget Sound Georgia Basin, were examined for species of Cryptosporidium and Giardia to determine their role in the epidemiology of these pathogens. Using ZnSO4 flotation and immunomagnetic separation, followed by direct immunofluorescent antibody detection (IMS/DFA), we identified Cryptosporidium sp. oocysts in 9 fecal samples from 6 locations and Giardia sp. cysts in 11 fecal samples from 7 locations. The putative risk factors of proximate human population and degree of anthropogenic shoreline modification were not associated with the detection of Cryptosporidium or Giardia spp. in river otter feces. Amplification of DNA from the IMS/DFA slide scrapings was successful for 1 sample containing > 500 Cryptosporidium sp. oocysts. Sequences from the Cryptosporidium 18S rRNA and the COWP loci were most similar to the ferret Cryptosporidium sp. genotype. River otters could serve as reservoirs for Cryptosporidium and Giardia species in marine ecosystems. More work is needed to better understand the zoonotic potential of the genotypes they carry as well as their implications for river otter health.     

Unravelling Cryptosporidium and Giardia epidemiology

Molecular biology has provided insights into the taxonomy and epidemiology of Cryptosporidium and Giardia, which are major causes of protozoal diarrhoea in humans worldwide. For both genera, previously unrecognized differences in disease, symptomatology, zoonotic potential, risk factors and environmental contamination have been identified using molecular tools that are appropriate for species, genotype and subtype analysis. In this article, to improve understanding of the epidemiology of cryptosporidiosis and giardiasis, we consider specific requirements for the development of more-effective molecular identification and genotyping systems that should be applicable to both clinical and environmental samples.     

Prevalence of Cryptosporidium and Giardia species in animals in irrigation catchments in the southwest of Australia

Screening of 445 animal faecal samples in irrigation catchments in Western Australia (WA) was conducted to identify the prevalence of Cryptosporidium and Giardia species. Of the samples positive for Giardia duodenalis, 30.7% (12/36) were the zoonotic Assemblage A, while approximately 13% (4/30) of Cryptosporidium positives were zoonotic. This is the first finding of Giardia Assemblage A in native marsupials and birds and indicates that marsupials and possibly birds may potentially be a reservoir of zoonotic Giardia.     

Zoonotic cryptosporidiosis

The widespread usages of molecular epidemiological tools have improved the understanding of cryptosporidiosis transmission. Much attention on zoonotic cryptosporidiosis is centered on Cryptosporidium parvum. Results of genotype surveys indicate that calves are the only major reservoir for C. parvum infections in humans. The widespread presence of human-adapted C. parvum, especially in developing countries, is revealed by recent subtyping and multilocus typing studies, which have also demonstrated the anthroponotic transmission of C. parvum subtypes shared by humans and cattle. Developing and industrialized countries differ significantly in disease burdens caused by zoonotic species and in the source of these parasites, with the former having far fewer human infections caused by C. parvum and little zoonotic transmission of this species. Exclusive anthroponotic transmission of seemingly zoonotic C. parvum subtypes was seen in Mid-Eastern countries. Other zoonotic Cryptosporidium spp. are also responsible for substantial numbers of human infections in developing countries, many of which are probably transmitted by anthroponotic pathways. The lower pathogenicity of some zoonotic species in some populations supports the occurrence of different clinical spectra of Cryptosporidium spp. in humans. The use of a new generation of molecular diagnostic tools is likely to produce a more complete picture of zoonotic cryptosporidiosis.     

Evaluation of PCR, nested PCR, and fluorescent antibodies for detection of Giardia and Cryptosporidium species in wastewater.

Giardiasis and cryptosporidiosis are diseases caused by the protozoan parasites Giardia lamblia and Cryptosporidium parvum. Waterborne transmission of these organisms has become more prevalent in recent years, and regulatory agencies are urging that source and finished water be screened for these organisms. A major problem associated with testing for these organisms is the lack of reliable methodologies and baseline information on the prevalence of these parasites in various water sources. Our study addressed both of these issues. We evaluated the presence and reduction of Giardia cysts and Cryptosporidium oocysts in sewage effluent by a combination of indirect fluorescent antibody (IFA) staining and PCR. Our results indicated a 3-log reduction of Giardia cysts and a 2-log reduction of Cryptosporidium oocysts through the sewage treatment process as determined by IFA. We developed a nested PCR to detect Cryptosporidium oocysts and used a double PCR to detect Giardia cysts. A 100% correlation was noted between IFA and PCR detection of Giardia cysts while correlation for Cryptosporidium oocysts was slightly less. On the basis of these results, PCR may be a useful tool in the environmental analysis of water samples for Giardia and Cryptosporidium organisms.     

Canine gastrointestinal parasitic zoonoses in India

Although well recognized and studied in developed countries, canine parasitic zoonoses pose a lowly prioritized public health problem in developing countries such as India, where conditions are conducive for transmission. A study of the most recent parasite survey determining prevalence and epidemiology of canine parasitic zoonoses among tea-growing communities of northeast India demonstrated the endemicity of the problem. This particular study serves as a model using conventional, as well as molecular parasitological, tools to provide novel insights into the role of dogs as mechanical transmitters of human parasites such as Ascaris and Trichuris, and discusses the risks dogs pose with regards to zoonotic transmission of hookworms and Giardia.     

Giardia and Cryptosporidium in inflowing water and harvested shellfish in a lagoon in Southern Italy

Giardia and Cryptosporidium spp. are important enteric protozoan pathogens for humans and animals, and have been found to contaminate water as well as edible shellfish all over the world. This is the first study to simultaneously investigate the presence of Giardia and Cryptosporidium in inflowing water and harvested shellfish in a geographically closed environment (Varano Lagoon, Southern Italy). Samples of treated wastewater were collected each month - at the outlet from the treatment plant, and downstream at the inlet into the lagoon - from the channels flowing into the Lagoon, together with specimens of Ruditapes decussatus and Mytilus galloprovincialis from shellfish-farms on the same lagoon. Giardia cysts were found by immunofluorescence (IF) microscopy in 16 out of 21 samples of treated wastewater and in 7 out of 21 samples from downstream water channels, and viable cysts were also detected by a beta-giardin RT-PCR. G. duodenalis Assemblages A and B were identified by small ribosomal subunit (18S-rDNA) and triosephosphate isomerase (tpi)-PCR, followed by sequencing. Cryptosporidium oocysts were found by IF in 5 out of 21 wastewater samples, and in 8 out of 21 samples from water channels. Molecular analysis identified the zoonotic species Cryptosporidium parvum by oocyst wall protein (COWP)-PCR and sequencing. Higher concentrations of Giardia cysts than Cryptosporidium oocysts were registered in almost all wastewater and water samples. IF and molecular testing of shellfish gave negative results for both protozoa. Wastewaters carrying Giardia and Cryptosporidium (oo)cysts are discharged into the Lagoon; however, the shellfish harvested in the same environment were found to be unaffected, thus suggesting that physical, ecological and climatic conditions may prevent contamination of harvested shellfish.     

Detection and characterisation of parasites causing emerging zoonoses

Understanding the epidemiology of zoonotic parasitic infections is dependent upon the availability of accurate and sensitive diagnostic techniques. The development of molecular diagnostic methods, particularly those utilising PCR for the detection of zoonoses will contribute greatly to the identification and control of these pathogens, by increasing the speed of diagnosis, specificity and sensitivity, reproducibility and ease of interpretation. Molecular characterisation studies allow us to distinguish between closely related infectious agents and to document the patterns of transmission of 'strains' and species within populations. This will allow precise determinations to be made about the aetiological agent, its characteristics and the source of infection. This review focuses on recent detection and characterisation techniques for both emerging and re-emerging parasite zoonoses.     

Cryptosporidium and cryptosporidiosis in Denmark--current status

The genus Cryptosporidium comprises a group of protozoan parasites that infect a broad variety of vertebrates causing severe diarrhoeal illness in immunocompromised as well as immunocompetent hosts. Although molecular heterogeneity of the genus is being increasingly recognised, traditional diagnostic methods do not discriminate all species/subtypes, and population genetic studies of these parasites, using discriminatory molecular markers, have only been published recently. In Denmark, Cryptosporidium research has focussed mainly on detection methods, pathogenicity and veterinary aspects. The present paper gives an overview of recent and ongoing Cryptosporidium research in Denmark with an emphasis on molecular approaches to study epidemiology and transmission.     

Giardia and Cryptosporidium in mammalian wildlife--current status and future needs

Environmental pollution with human and domestic-animal fecal material is recognized as a potential pathogen pathway for wildlife infections with zooanthropomorphic protozoan parasites such as Giardia and Cryptosporidium. In this article, we review current knowledge about the diversity of free-living and captive terrestrial and marine mammalian wildlife species infected with Giardia and Cryptosporidium. The combination of prevalence studies with modern molecular-genotyping techniques is providing valuable insights into the host specificity and possible transmission routes of these two important parasites.     

Zoonotic protozoa--food for thought

Outbreaks of water- and foodborne diseases caused by Cryptosporidium, Giardia and Toxoplasma are well documented. Three features of these zoonotic protozoa ensure a high level of environmental contamination and enhance the likelihood of waterborne transmission. Firstly, they are responsible for disease in a broad range of hosts including man, have a low infectious dose enhancing the possibility of zoonotic transmission, secondly, their transmissive stages are small in size and environmentally robust and thirdly are insensitive to the disinfectants commonly used in the water industry. In addition, there is growing evidence for the role that water and food can play in the transmission of the microsporidia, Balantidium and Blastocystis to humans.     

Giardia sp. and Cryptosporidium sp. infections in primates in fragmented and undisturbed forest in western Uganda

In June 2005, we collected 115 fecal samples from wild primates in western Uganda and examined them for Cryptosporidium sp. and Giardia sp. with the use of immunofluorescent antibody (IFA) detection. We sampled primates from an undisturbed forest in Kibale National Park and from 3 highly disturbed forest fragments outside the park. Of disturbed forest samples, red colobus (Pilocolobus tephrosceles) and red-tailed guenons (Cercopithecus ascanius) harbored species of Cryptosporidium or Giardia, but black-and-white colobus (Colobus guereza) did not. All primate samples from undisturbed forest were negative for both parasites. Seven of 35 (20%) red colobus and 1 of 20 red-tailed guenons (5%) from forest fragments were infected with either Cryptosporidium sp. or Giardia sp. The presence of Cryptosporidium and Giardia species in primates living in forest fragments, but not in primates in undisturbed forest, suggests that habitat disturbance may play a role in transmission or persistence of these pathogens.     

Zoonotic potential of Giardia

Giardia duodenalis (syn. Giardia lamblia and Giardia intestinalis) is a common intestinal parasite of humans and mammals worldwide. Assessing the zoonotic transmission of the infection requires molecular characterization as there is considerable genetic variation within G. duodenalis. To date eight major genetic groups (assemblages) have been identified, two of which (A and B) are found in both humans and animals, whereas the remaining six (C to H) are host-specific and do not infect humans. Sequence-based surveys of single loci have identified a number of genetic variants (genotypes) within assemblages A and B in animal species, some of which may have zoonotic potential. Multi-locus typing data, however, has shown that in most cases, animals do not share identical multi-locus types with humans. Furthermore, interpretation of genotyping data is complicated by the presence of multiple alleles that generate “double peaks” in sequencing files from PCR products, and by the potential exchange of genetic material among isolates, which may account for the non-concordance in the assignment of isolates to specific assemblages. Therefore, a better understanding of the genetics of this parasite is required to allow the design of more sensitive and variable subtyping tools, that in turn may help unravel the complex epidemiology of this infection.     

Variation in Cryptosporidium: towards a taxonomic revision of the genus

Cryptosporidium is an important cause of enteric disease in humans and other animals. Limitations associated with conventional diagnostic methods for cryptosporidiosis based on morphological features, coupled with the difficulty of characterising parasites isolated in the laboratory, have restricted our ability to clearly identify species. The application of sensitive molecular approaches has obviated the necessity for laboratory amplification. Such studies have found considerable evidence of genetic heterogeneity among isolates of Cryptosporidium from different species of vertebrate, and there is now mounting evidence suggesting that a series of host-adapted genotypes/strains/species of the parasite exist. In this article, studies on the molecular characterisation of Cryptosporidium during the last 5 years are reviewed and put into perspective with the past and present taxonomy of the genus. The predictive value of achieving a sound taxonomy for the genus Cryptosporidium with respect to understanding its epidemiology and transmission and controlling outbreaks of the disease is also discussed.     

Evidence supporting zoonotic transmission of Cryptosporidium in rural New South Wales

Cryptosporidium hominis, which has an anthroponotic transmission cycle and Cryptosporidium parvum, which is zoonotic, are the primary species of Cryptosporidium that infect humans. The present study identified the species/genotypes and subgenotypes of Cryptosporidium in 7 human and 15 cattle cases of sporadic cryptosporidiosis in rural western NSW during the period from November 2005 to January 2006. The species/genotype of isolates was determined by PCR sequence analysis of the 18S rRNA and C. parvum and C. hominis isolates were subgenotyped by sequence analysis of the GP60 gene. Fourteen of 15 cattle-derived isolates were identified as C. parvum and 1 as a C. bovis/C. parvum mixture. Of the human isolates, 4 were C. parvum and 3 were C. hominis. Two different subgenotypes were identified with the human C. hominis isolates and six different subgenotypes were identified within the C. parvum species from humans and cattle. All four of the C. parvum subtypes found in humans were also found in the cattle, indicating that zoonotic transmission may be an important contributor to sporadic human cases cryptosporidiosis in rural NSW.