Efficacy and safety of prazequantel for the treatment of Schistosoma mansoni infection across different transmission settings in Amhara Regional State, northwest Ethiopia.

BACKGROUND: Schistosoma mansoni and S. haematobium infections have been public health problems in Ethiopia, S. mansoni being more prevalent. To reduce the burden of schistosomiasis, a national school-based prazequantel (PZQ) mass drug administration (MDA) program has been implemented since November 2015. Nevertheless, S. mansoni infection is still a major public health problem throughout the country. Reduced efficacy of PZQ is reported by a few studies in Ethiopia, but adequate data in different geographical settings is lacking. Hence, this study aimed to assess the efficacy and safety of PZQ for the treatment of S. mansoni infection across different transmission settings in Amhara Regional State, northwest Ethiopia. METHODS: A school-based single-arm prospective cohort study was conducted from February to June, 2023 among 130 S. mansoni-infected school-aged children (SAC). Forty-two, 37, and 51 S. mansoni-infected SAC were recruited from purposely selected schools located in low, moderate, and high transmission districts, respectively. School-aged children who were tested positive both by Kato Katz (KK) using stool samples and by the point of care circulating cathodic antigen (POC-CCA) test using urine samples at baseline were treated with a standard dose of PZQ and followed for 21 days for the occurrence of adverse events. After three weeks post-treatment, stool and urine samples were re-tested using KK and POC-CCA. Then the cure rate (CR), egg reduction rate (ERR), and treatment-associated adverse events were determined. The data were analyzed using SPSS version 21. RESULTS: Out of the total 130 study participants, 110 completed the follow-up. The CR and ERR of PZQ treatment were 88.2% (95%CI: 82.7-93.6) and 93.5% (95%CI: 85.4-98.5), respectively, by KK. The CR of PZQ based on the POC-CCA test was 70.9% (95%CI: 62.7-79.1) and 75.5% (95%CI: 67.3-83.6) depending on whether the interpretation of 'trace' results was made as positive or negative, respectively. After treatment on the 21st day, 78 and 83 participants tested negative both by KK and POC-CCA, with respective interpretations of 'trace' POC-CCA test results as positive or negative. The CR in low, moderate and high transmission settings was 91.7%, 91.2% and 82.5%, respectively (p = 0.377) when evaluated by KK. The CR among SAC with a light infection at baseline (95.7%) by KK was higher than that of moderate (81.5%) and heavy (64.3%) infections (χ2 = 12.53, p = 0.002). Twenty-six (23.6%) participants manifested at least one adverse event. Eleven (10.0%), eight (7.3%), six (5.5%), and three (2.7%) participants complained about abdominal pain, nausea, headache, and anorexia, respectively. All adverse events were mild, needing no intervention. Occurrence of adverse events was slightly higher in high endemic areas (32.5%) than moderate (23.5%) and low endemic areas (p = 0.279). CONCLUSIONS: A single dose of 40 mg/kg PZQ was efficacious and safe for the treatment of S. mansoni infection when it was evaluated by the KK test, but a lower efficacy was recorded when it was evaluated by the POC-CCA test. However, the POC-CCA test's specificity, clearance time of CCA from urine after treatment, and interpretation of weakly reactive (trace) test results need further research.

Knowledge, attitude and practice towards intestinal schistosomiasis among school-aged children and adults in Amhara Regional State, northwest Ethiopia. A cross-sectional study.

BACKGROUND: Schistosoma mansoni causes intestinal schistosomiasis (SCH) in all regions of Ethiopia. Despite many years of mass treatment, the prevalence has not dropped significantly. The reduction of SCH transmission demands the integration of deworming with safe water, sanitation, and hygiene (WASH) activities. Adequate knowledge and a positive attitude towards SCH are critical to practicing those interventions. However, data on the knowledge, attitude, and practice (KAP) level in school and community settings is limited in Ethiopia. METHODS: School and community-based cross-sectional studies were conducted from February to June 2023 among 634 school-aged children (SAC) and 558 adults. A pre-tested questionnaire was used to collect socio-demographic and KAP data. Records were entered and analyzed using SPSS software version 21. Correct responses for each of the KAP questions were scored as one, while incorrect or 'I don't know' responses were scored as zero. Good knowledge, a positive attitude and good practice were declared if percentage scores were ≥ 80%, ≥ 90% and ≥ 75%, respectively. RESULTS: Only 229 (19.2%) respondents, comprising 91 (14.4%) SAC and 138 (24.7%) adults, had ever heard of SCH. Adults, males, and urban residents had higher awareness level compared to their respective counterparts (p < 0.05). Only 28.4% of aware respondents knew that swimming or bathing in freshwater is a risk factor for schistosoma infection; 10.9% knew the etiologic agent; and 14.4% mentioned at least one sign and symptom associated with SCH. The majority (97.8%) of the respondents were willing to take therapeutic drugs, but only 37.6% believed that SCH is a serious disease. Regarding risky practices, 89.5% practiced swimming or bathing in freshwater, and 25.3% had no access to piped water. Among the aware respondents, only 18 (7.9%) had good knowledge, while 30 (13.1%) had a positive attitude towards SCH. Ninety-nine (43.2%) respondents had good Schistosoma infection prevention practices. CONCLUSIONS: The knowledge, attitude, and preventive practice level towards schistosomiasis are low in the study area. Therefore, strengthening school and community-based health education, along with mass drug administration (MDA), WASH, and a vector control program, is recommended for preventing SCH.

Deletion patterns, genetic variability and protein structure of pfhrp2 and pfhrp3: implications for malaria rapid diagnostic test in Amhara region, Ethiopia.

BACKGROUND: Although rapid diagnostic tests (RDTs) play a key role in malaria-control strategies, their efficacy has been threatened by deletion and genetic variability of the genes pfhrp2/3. This study aims to characterize the deletion, genetic patterns and diversity of these genes and their implication for malaria RDT effectiveness, as well as their genetic evolution in the Amhara region of Ethiopia. METHODS: The study included 354 isolates from symptomatic patients from the Amhara region of Ethiopia who tested positive by microscopy. Exon 1-2 and exon 2 of genes pfhrp2 and -3 were amplified, and exon 2 was sequenced to analyse the genetic diversity, phylogenetic relationship and epitope availability. RESULTS: The deletion frequency in exon 1-2 and exon 2 was 22 and 4.6% for pfhrp2, and 68 and 18% for pfhrp3, respectively. Double deletion frequency for pfhrp2 and pfhrp3 was 1.4%. High genetic diversity, lack of clustering by phylogenetic analysis and evidence of positive selection suggested a diversifying selection for both genes. The amino-acid sequences, classified into different haplotypes, varied widely in terms of frequency of repeats, with novel amino-acid changes. Aminoacidic repetition type 2 and type 7 were the most frequent in all the sequences. The most frequent epitopes among protein sequences were those recognized by MAbs 3A4 and C1-13. CONCLUSION: Deletions and high amino acidic variation in pfhrp2 and pfhrp3 suggest their possible impact on RDT use in the Amhara region, and the high genetic diversity of these genes could be associated with a diversifying selection in Ethiopia. Surveillance of these genes is, therefore, essential to ensure the effectiveness of public health interventions in this region.

Prevalence of Strongyloides stercoralis infection and associated clinical symptoms among schoolchildren living in different altitudes of Amhara National Regional State, northwest Ethiopia.

BACKGROUND: Strongyloides stercoralis is a parasite that causes strongyloidiasis in humans. It is prevalent in the tropics and sub-tropics where poor sanitation is a common problem. The true prevalence of S. stercoralis in Ethiopia is underestimated due to the lack of a "Gold" standard diagnostic method. Moreover, its prevalence across altitudinal gradient in Amhara Region has not been studied. METHODS: A cross-sectional study was conducted among 844 schoolchildren in Amhara Region from April to December 2019. A stool sample was collected from each study participant and processed using formol ether concentration technique (FECT), spontaneous tube sedimentation technique (STST), Baermann concentration technique (BCT), agar plate culture (APC) and real-time polymerase chain reaction (RT-PCR). Data were entered using EpiData and analyzed by SPSS version 23 statistical software. Prevalence of S. stercoralis infection was determined using a single diagnostic technique and combination of techniques. Association of clinical variables with S. stercoralis infection was assessed by logistic regression and independent variables with p<0.05 were considered statistically significant. RESULTS: Prevalence of soil-transmitted helminths (STHs) and S. mansoni infections was 38.0% and 20.4%, respectively. Among STHs, the prevalence of hookworm infection was 32.8%. Prevalence of S. stercoralis infection was 39.0%, 28.8%, 10.9%, 10.3%, 4.0% and 2.0% by the respective, combinations of the five methods, RT-PCR, APC, BCT, STST and FECT. The highest prevalence rates, 48.2%, 45.0% and 41.1% of S. stercoralis were recorded in the age group of 12-14 years, males and rural dwellers, respectively. Prevalence rates of S. stercoralis infection in highland, semi-highland and lowland areas were 40.4%, 41.8% and 25.9%, respectively. Having abdominal pain (AOR = 2.48; 95% CI:1.65-3.72), cough (AOR = 1.63;95%CI:1.09-2.42), urticaria (AOR = 2.49;95%CI:1.50-4.01) and being malnourished (AOR = 1.44;95%:1.10-2.01) were significantly associated with strongyloidiasis. CONCLUSION: Prevalence of S. stercoralis infection was high and varied across different altitudes in Amhara Region. Some clinical syndromes were found to be significantly associated with S. stercoralis infection. Therefore, proper diagnosis and preventive strategies against S. stercoralis infection are highly recommended to be devised and implemented in Amhara Region.

Evaluation of five diagnostic methods for Strongyloides stercoralis infection in Amhara National Regional State, northwest Ethiopia.

BACKGROUND: Strongyloides stercoralis is an intestinal parasite that can cause chronic infection, hyperinfection and/or a dissemination syndrome in humans. The use of techniques targeting ova fails to detect S. stercoralis, as only larvae of the parasite are excreted in faeces. Due to the absence of "Gold" standard diagnostic method for S. stercoralis, there is a paucity of reported data worldwide. OBJECTIVE: This study aimed to evaluate the performance of diagnostic methods of S. stercoralis infection by taking the composite reference as a "Gold" standard. METHODS: A cross-sectional study was conducted among 844 schoolchildren in Amhara Region, Ethiopia, from April to December 2019. Stool samples were collected and processed with formol-ether concentration technique (FECT), spontaneous tube sedimentation technique (STST), Baermann concentration technique (BCT), agar plate culture (APC) and real-time polymerase chain reaction (RT-PCR). Sensitivity, specificity, positive predictive value, and negative predictive value of each diagnostic method were computed against the composite reference. The agreements of diagnostic methods were evaluated by Kappa value at 95% CI. RESULTS: The composite detection rate of S. stercoralis by the five diagnostic methods was 39.0% (329/844). The detection rate of the parasite from stool samples by FECT, STST, BCT, APC and RT-PCR was 2.0% (17/844), 4.0% (34/844), 10.2% (86/844), 10.9% (92/844) and 28.8% (243/844), respectively. The highest detection rate (37.8%; 319/844) of S. stercoralis was recorded by a combination of BCT, APC, and RT-PCR followed by a combination of STST, BCT, APC and RT-PCR (37.3%; 315/844). The sensitivity of FECT, STST, BCT, APC and RT-PCR against the composite reference was 5.2%, 10.3%, 26.4%, 28.0% and 73.9%, respectively. The diagnostic agreements of RT-PCR, APC, BCT, STST and FECT with the composite reference in detection of S. stercoralis were substantial (0.775), fair (0.321), fair (0.305), slight (0.123), and slight (0.062), respectively. CONCLUSION: RT-PCR detected the highest number of S. stercoralis infections. A combination of RT-PCR with APC and/or BCT better detected S. stercoralis from stool samples compared to other combinations or single diagnostic methods. Therefore, RT-PCR and combination of RT-PCR with APC and/or BCT diagnostic methods should be advocated for detection of S. stercoralis infection.

Feature importance: Opening a soil-transmitted helminth machine learning model via SHAP.

In the field of landscape epidemiology, the contribution of machine learning (ML) to modeling of epidemiological risk scenarios presents itself as a good alternative. This study aims to break with the "black box" paradigm that underlies the application of automatic learning techniques by using SHAP to determine the contribution of each variable in ML models applied to geospatial health, using the prevalence of hookworms, intestinal parasites, in Ethiopia, where they are widely distributed; the country bears the third-highest burden of hookworm in Sub-Saharan Africa. XGBoost software was used, a very popular ML model, to fit and analyze the data. The Python SHAP library was used to understand the importance in the trained model, of the variables for predictions. The description of the contribution of these variables on a particular prediction was obtained, using different types of plot methods. The results show that the ML models are superior to the classical statistical models; not only demonstrating similar results but also explaining, by using the SHAP package, the influence and interactions between the variables in the generated models. This analysis provides information to help understand the epidemiological problem presented and provides a tool for similar studies.

Determinant factors of chronic Strongyloides stercoralis infection among schoolchildren in Amhara National Regional State, northwest Ethiopia.

BACKGROUND: Strongyloides stercoralis is one of the most neglected parasites that is commonly found in tropical and sub-tropical countries. The majority of chronic S. stercoralis infections are characterized by minor clinical signs and symptoms. The extent to which socio-demographic, personal and environmental risk factors associated with S. stercoralis infection is not well known in Amhara Regional, Ethiopia. OBJECTIVE: This study aimed to assess socio-demographic, personal and environmental risk factors associated with S. stercoralis infection among schoolchildren in Amhara Region. METHODS: A total of 844 schoolchildren were randomly selected and participated in the study. Stool samples were collected and checked for S. stercoralis infection using five diagnostic methods. A structured questionnaire was also used to obtain socio-demographic, personal and environmental data from parents of the children. The degree of association between socio-demographic, personal and environmental factors with S. stercoralis infection was first assessed by univariate logistic regression and then analyzed by multivariate logistic regression. Variables with p<0.05 were considered as statistically significant. RESULTS: Among 844 participants, 329 (39.0%) were infected with S. stercoralis. Prevalences of S. stercoralis among 12-14 years old, males, and rural dwellers were 48.2% (109/226), 45.0% (196/436) and 41.1% (306/745), respectively. Being in the age group of 10-11years (AOR=1.56;95%CI:1.10-2.24), and 12-14 years (AOR=1.96;95%CI:1.32-2.90), male (AOR=1.55;95%CI:1.16-2.06), and rural dweller (AOR=1.86;95%CI:1.12-3.10); having family income of < 1000 Birr per month (AOR=2.03;95%CI:1.48-2.77), and family size of 5-9 members (AOR=2.36; 95%CI:1.57-3.54) were significantly associated with S. stercoralis infection. The habit of not wearing shoes (AOR=1.41; 95%CI:1.05-1.90), close contacting with pets (AOR=1.49;95%:1.09-2.04), participating in irrigation (AOR=1.49; 95%CI:1.10-2.00) and playing with soil (AOR=1.62;95%CI:1.20-2.18) were risk factors of S. stercoralis infection. Besides, presence of wet environment (AOR=1.62;95%CI:1.19-2.22), living around sandy soil (AOR=2.75; 95%CI: 1.81-4.19) and red clay soil (AOR=2.24;95%CI:1.58-3.17), living in semi-highland folds (AOR=2.47;95%CI:1.48-4.14) and highland areas (AOR=1.65;95%CI:1.06-2.57) were determinant factors for S. stercoralis. CONCLUSION: S. stercoralis infection among schoolchildren was highly prevalent in Amhara Region. Several determinant factors were also identified. Therefore, proper implementation of sanitation and hygiene programs and health care of pets should be advocated to break the transmission cycle of S. stercoralis.

Environmental characteristics around the household and their association with hookworm infection in rural communities from Bahir Dar, Amhara Region, Ethiopia.

Soil-Transmitted Helminths (STH) are highly prevalent Neglected Tropical Disease in Ethiopia, an estimated 26 million are infected. Geographic Information Systems and Remote Sensing (RS) technologies assist data mapping and analysis, and the prediction of the spatial distribution of infection in relation to environmental variables. The influence of socioeconomic, environmental and soil characteristics on hookworm infection at the individual and household level is explored in order to identify spatial patterns of infection in rural villages from Zenzelema (Amhara region). Inhabitants greater than 5 years old were recruited in order to assess the presence of STH. Socioeconomic and hookworm infection variables at the household level and environmental variables and soil characteristics using RS were obtained. The dominant STH found was hookworm. Individuals which practiced open defecation and those without electricity had a significant higher number of hookworm eggs in their stool. Additionally, adults showed statistically higher hookworm egg counts than children. Nonetheless, the probability of hookworm infection was not determined by socioeconomic conditions but by environmental characteristics surrounding the households, including a combination of vigorous vegetation and bare soil, high temperatures, and compacted soils (high bulk density) with more acidic pH, given a pH of 6.0 is optimal for hatching of hookworm eggs. The identification of high-risk environmental areas provides a useful tool for planning, targeting and monitoring of control measures, including not only children but also adults when hookworm is concerned.

Agar Plate Culture: An Alternative Sensitive Routine Laboratory Detection Method for Strongyloides stercoralis and Hookworm Parasites.

BACKGROUND: Human infection with Strongyloides stercoralis and hookworm parasites is usually under reported due to less sensitive diagnostic methods. Agar plate culture (APC) is the most sensitive technique for parasites having larval stage. However, using APC in routine diagnosis is uncommon. This study aimed to determine the detection rate and sensitivity of APC in comparison with formal ether concentration technique (FECT) and spontaneous tube sedimentation techniques (STSTs) for S. stercoralis and hookworm larvae. METHODS: Stool samples collected from 844 schoolchildren in Amhara Regional State, northwestern Ethiopia in 2019, transported to nearby health institutions and processed by APC, FECT and STSTs. The prevalence of S. stercoralis and hookworm was computed by descriptive statistics and Chi-square. The diagnostic agreement among the three techniques was evaluated using Kappa value. RESULTS: The overall prevalence of S. stercoralis and hookworm infections by combining the three methods was 13.2% (111/844) and 33.8% (277/844), respectively. Using APC alone, the prevalence of S. stercoralis and hookworm were found to be 10.9% (92/844) and 24.5% (207/844), respectively. Agar plate culture was 5.4 and 2.7 times respectively more sensitive than FECT and STST, with slight and fair agreement in the detection of S. stercoralis. Hookworm diagnostic agreement was moderate between APC and FECT, and APC and STST. The Kappa value between STST and FECT diagnostic methods was substantial. CONCLUSION: APC has a better detection rate of S stercoralis and hookworm larvae. Therefore, APC can be used as an alternative routine diagnostic method to S. stercoralis and hookworm co-endemic countries.

Efficacy of Single Dose Ivermectin Against Strongyloides stercoralis Infection Among Primary School Children in Amhara National Regional State.

BACKGROUND: Ivermectin has been proven to be highly effective against Strongyloides stercoralis in some countries. However, its single dose (200 µg/kg) efficacy has not been proven up until now in Ethiopia. OBJECTIVE: This study aimed to evaluate the efficacy of single dose ivermectin against S. stercoralis infection among school children. METHODS: Stool sample was collected from April 2019 to December 2019 among 844 school children and screened by formol ether concentration, spontaneous tube sedimentation, Baermann concentration, and agar plate techniques. Single oral dose (200 µg/kg) ivermectin was given to 101 S. stercoralis-infected student and posttreatment diagnosis was done for 92 students after 2 weeks. RESULTS: Of the total 92 S. stercoralis-infected students who took ivermectin treatment, 87 were negative with cure rate of 94.6%. No side effect of ivermectin was observed. CONCLUSION: Single dose ivermectin is an effective dose in uncomplicated chronic strongyloidiasis.

Epidemiology of intestinal helminthiases in a rural community of Ethiopia: Is it time to expand control programs to include Strongyloides stercoralis and the entire community?

BACKGROUND: Soil transmitted helminths are highly prevalent worldwide. Globally, approximately 1.5 billion people are infected with Ascaris lumbricoides, Trichuris trichiura or hookworm. Endemic countries carry out periodic mass treatment of at-risk populations with albendazole or mebendazole as a control measure. Most prevalence studies have focused on school aged children and therefore control programs are implemented at school level, not at community level. In this study, the prevalence of intestinal helminths, including Strongyloides stercoralis, was examined using a comprehensive laboratory approach in a community in north-western Ethiopia. METHODS: A cross-sectional survey was conducted on 792 individuals ≥5 years old in randomly selected houses in a rural district. Stools were examined using three techniques: a formol-ether concentration, the Baermann technique and a real time polymerase chain reaction test (these last two specific for S. stercoralis). Statistical analyses were performed between two large age groups, children (≤14 years old) and adults (≥15 years old). RESULTS: The prevalence of helminths was 91.3%; (95% CI: 89.3-93.3%). Hookworm was the most prevalent, 78.7% (95% CI 75.6-81.4%), followed by S. stercoralis 55.7% (95% CI 52.2-59.1%). Co-infection with both was detected in 45.4% (95% CI 42.0-49.0%) of the participants. The mean age of hookworm-infected individuals was significantly higher than non-infected ones (p = 0.003). Also, S. stercoralis infection was significantly associated with age, being more prevalent in adults (p = 0.002). CONCLUSIONS: This is the highest prevalence of S. stercoralis detected in Ethiopia so far. Our results highlight the need of searching specifically for infection by this parasite since it usually goes unnoticed if helminth studies rely only on conventional diagnostic techniques, i.e. Kato-Katz. Moreover, the focus of these programs on children undermines the actual prevalence of hookworm. The adult population acts as a reservoir for both hookworm and S. stercoralis and this fact may negatively impact the current control programs in Ethiopia which only target treatment of school aged children. This reservoir, together with a lack of adequate water, sanitation and hygiene, increases the probability of re-infection in children. Finally, the high prevalence of S. stercoralis found calls for a comprehensive diagnostic approach in endemic areas in addition to a revision of control measures that is, adding ivermectin to current albendazole/mebendazole, since it is the drug of choice for S. stercoralis.

High prevalence of Strongyloides stercoralis in school-aged children in a rural highland of north-western Ethiopia: the role of intensive diagnostic work-up.

BACKGROUND: Soil-transmitted helminthiases (hookworms, Ascaris lumbricoides and Trichuris trichiura) are extremely prevalent in school-aged children living in poor sanitary conditions. Recent epidemiological data suggest that Strongyloides stercoralis is highly unreported. However, accurate data are essential for conducting interventions aimed at introducing control and elimination programmes. METHODS: We conducted a cross-sectional survey of 396 randomly selected school-aged children in Amhara region in rural area in north-western Ethiopia, to assess the prevalence of S. stercoralis and other intestinal helminths. We examined stools using three techniques: conventional stool concentration; and two S. stercoralis-specific methods, i.e. the Baermann technique and polymerase chain reaction. The diagnostic accuracy of these three methods was then compared. RESULTS: There was an overall prevalence of helminths of 77.5%, with distribution differing according to school setting. Soil-transmitted helminths were recorded in 69.2%. Prevalence of S. stercoralis and hookworm infection was 20.7 and 54.5%, respectively, and co-infection was detected in 16.3% of cases. Schistosoma mansoni had a prevalence of 15.7%. Prevalence of S. stercoralis was shown 3.5% by the conventional method, 12.1% by the Baermann method, and 13.4% by PCR, which thus proved to be the most sensitive. CONCLUSIONS: Our results suggest that S. stercoralis could be overlooked and neglected in Ethiopia, if studies of soil-transmitted helminths rely on conventional diagnostic techniques alone. A combination of molecular and stool microscopy techniques yields a significantly higher prevalence. In view of the fact that current control policies for triggering drug administration are based on parasite prevalence levels, a comprehensive diagnostic approach should instead be applied to ensure comprehensive control of helminth infections.

Prevalence and Genetic Diversity of Giardia duodenalis and Cryptosporidium spp. among School Children in a Rural Area of the Amhara Region, North-West Ethiopia.

BACKGROUD: Giardia duodenalis and Cryptosporidium spp. are enteric protozoan causing gastrointestinal illness in humans and animals. Giardiasis and cryptosporidiosis are not formally considered as neglected tropical diseases, but belong to the group of poverty-related infectious diseases that impair the development and socio-economic potential of infected individuals in developing countries. METHODS: We report here the prevalence and genetic diversity of G. duodenalis and Cryptosporidium spp. in children attending rural primary schools in the Bahir Dar district of the Amhara Region, Ethiopia. Stool samples were collected from 393 children and analysed by molecular methods. G. duodenalis was detected by real-time PCR, and the assemblages and sub-assemblages were determined by multilocus sequence-based genotyping of the glutamate dehydrogenase and ß-giardin genes of the parasite. Detection and identification of Cryptosporidium species was carried out by sequencing of a partial fragment of the small-subunit ribosomal RNA gene. PRINCIPAL FINDINGS: The PCR-based prevalences of G. duodenalis and Cryptosporidium spp. were 55.0% (216/393) and 4.6% (18/393), respectively. A total of 78 G. duodenalis isolates were successfully characterized, revealing the presence of sub-assemblages AII (10.3%), BIII (28.2%), and BIV (32.0%). Discordant typing results AII/AIII and BIII/BIV were identified in 7.7% and 15.4% of the isolates, respectively. An additional five (6.4%) isolates were assigned to assemblage B. No mixed infections of assemblages A+B were found. Extensive genetic variation at the nucleotide level was observed within assemblage B (but no within assemblage A), resulting in the identification of a large number of sub-types. Cryptosporidium diversity was demonstrated by the occurrence of C. hominis, C. parvum, and C. viatorum in the population under study. CONCLUSIONS: Our data suggest an epidemiological scenario with an elevated transmission intensity of a wide range of G. duodenalis genetic variants. Importantly, the elevated degree of genetic diversity observed within assemblage B is consistent with the occurrence of intra-assemblage recombination in G. duodenalis.