Epidemiology of Giardia duodenalis infection in ruminant livestock and children in the Ismailia province of Egypt: insights by genetic characterization

Abstract

Background: Giardia duodenalis is a common flagellated protozoan parasite that infects the small intestine of a wide range of vertebrate hosts. This study aimed to determine whether tracing of G. duodenalis isolates by current genetic typing tools is possible using an exemplary set of samples from infected cattle, buffalo and children from the Ismailia province, Egypt.

Method: A total of 804 fecal samples from ruminant animals was collected from 191 herds and 165 samples from diarrheal children below the age of 10 years. Parasites were detected in these samples using the copro-antigen RIDA®QUICK test and by real-time PCR. Samples were then genetically characterized based on the triosephosphate isomerase, glutamate dehydrogenase and β-giardin genes.

Results: The prevalence of G. duodenalis was 53% in ruminants and 21% in symptomatic children and infection was not positively correlated with diarrheal symptoms. Sequence typing analysis confirmed predominance of B-type sequences (>67%) in humans and E-type sequences (>81%) in ruminants over A-type sequences. For 39 samples the complete sequence information of the three marker gene fragments could be derived. Integration of the concatenated sequence information of the three marker gene fragments with the spatial data of the respective sample revealed that identical or near identical (only up to 1 out of 1358 bp different) concatenated sequencing types were spatially related in 4 out of 5 cases.

Conclusion: The risk of zoonotic infection emanating from ruminants even in high prevalence areas is negligible. Genetic characterization indicated a predominant anthropogenic cycle of infection within the pediatric population studied. Integration of sequence typing data with information on geographic origins of samples allows parasite sub-population tracing using current typing tools.

Figure 1

Relatedness of assemblage B and E sequencing types using distance matrix analysis. The sequencing fragments of samples with the complete information at all three gene loci were concatenated in the order of the tpi-bg-gdh sequences. The resulting sequencing fragments (1358 bp) were subsequently aligned using ClustalW. Shown are the deduced distance matrices for assemblage B (A) and assemblage E (B) sequencing types. Numbers and heatmap indicate nucleotide residues not identical between two sequences. We revealed only two complete type A sequences and these were excluded from the analysis. (A) Eight unique type B sequences could be analyzed. (B) 25 unique sequences (of a total of 29 type E sequences) were retrieved and analyzed. Samples 31b/C7 and 33c/B7 were identical as well as samples 40b/C7, 41a/C1, 43a/B1 and 43a/C2.

Figure 2

Spatial distribution of samples containing assemblage B and E type parasites. Clusters of concatenated B and E type sequences were defined based on sequence identity, i.e. sequences that were identical or that differed in maximally one base pair were combined in one cluster. The spatial distribution of sequence types was visualized on a map of the study region. (A) Analysis of the 8 B type sequences revealed one cluster with 2 sequences that differed in one base pair (samples H66 and H90). (B) Analysis of 29 E type sequences revealed 4 clusters. Cluster 1 comprising of the identical sequences of samples 31b/C7 and 33c/B7. Cluster 2 comprising of the 4 identical sequences of samples 40b/C7, 41a/C1, 43a/B1 and 43a/C25, and of sequence of sample 32a/C2 that differed in one base pair. Cluster 3 comprising of sequences of samples 5a/C1 and 7a/C9 that differ in one base pair. Cluster 4 comprising of sequences of samples 35a/C2 and 35c/C8 that differ in one base pair. Note that sample 40c/C10 (shaded grey area) differed only in three base pairs from sample 40b/C7 of cluster 2.