Since very little is known on the epidemiology of intestinal protozoa in sub-Saharan Africa, we analyzed formalin-fixed stool samples obtained from 550 school-aged children who participated in a randomized controlled trial on Pemba Island to assess the efficacy and safety of nitazoxanide, albendazole and a combination of both drugs against T. trichiura and other soil-transmitted helminths. This trial found low CRs against soil-transmitted helminth species. Importantly, the study provided an opportunity to shed new light on the extent of intestinal protozoa infections in a child cohort and to determine whether the different study treatments had an effect on intestinal protozoa species.
Our results confirm that intestinal protozoa are a public health issue on Pemba Island. Indeed, almost half of the children were infected with at least one of the three (potentially) pathogenic intestinal protozoa. When considering the results from the baseline and the 3-week post-treatment follow-up, assuming that a child who was diagnosed positive at follow-up was diagnosed as a false-negative case at baseline, the prevalence of intestinal protozoa infection was even higher (Figure 1). This issue is most likely explained by the lack of sensitivity when examining only a single stool sample with the ether-concentration technique, an important limitation of our study. Hence, in future studies, multiple stool samples should be examined and subjected to the ether-concentration method or more sensitive molecular approaches employed to improve diagnostic accuracy[26, 27].
A recent study reported only moderate sensitivity for the ether-concentration technique compared to the FLOTAC technique when examining the same formalin-fixed stool samples. However, a study among five European reference laboratories showed that the agreement of diagnostic results was only moderate for pathogenic intestinal protozoa although the participating centres adhered to the same standard operating procedures for the ether-concentration technique. In particular, E. histolytica is frequently misdiagnosed, even by experienced laboratory personnel.
We found that girls were generally at higher odds of an infection with any of the intestinal protozoa encountered than boys. Similar results were found by Mohammed Mahady and colleagues in Malaysia. On the other hand, Traoré et al., in a study carried out in school-aged children in Côte d’Ivoire, reported a considerably higher prevalence of intestinal protozoa among boys than girls, corroborating findings by Cifuentes et al. from Mexico, where boys were at higher odds of a G. intestinalis infection that girls. Other studies found no gender difference at all. These findings indicate that intestinal protozoa infections may be related to gender-specific behaviour within a community. In addition, we observed statistically significant differences for some of the intestinal protozoa between the two schools. Hence, even though the schools are located only a few kilometers apart and the two settings were quite similar, at least in terms of socioeconomic status, availability of safe water supply and sanitation infrastructure, different infection profiles were observed. This highlights the possibility of ‘micro-geographic’ variability in endemicity of intestinal protozoa infection. One explanation could be that Al-Sadik school is located close to an orphanage, where transmission of intestinal protozoa might be enhanced. Other behavioural, infrastructure or environmental factors that may account for our observation should be investigated in future studies.
Intestinal protozoa often co-occur with intestinal nematodes and it is therefore important to determine whether anthelminthic and other antiparasitic drugs have an effect on concomitant intestinal protozoa infections. Albendazole, for example, was found in a recent meta-analysis to be as effective as metronidazole against G. intestinalis. The observed CRs against all intestinal protozoa were moderate to high in the three treatment groups. However – and contrary to what we expected – there was also a moderate treatment efficacy in the placebo group, which is difficult to explain other than a diagnostic dilemma. Our findings therefore have to be interpreted with caution. The high ‘cure rate’ observed within the placebo group underscores that analysis of a single stool sample with the formalin-ether concentration technique is unreliable, and hence, the CRs for the three treatment schemes investigated here are likely overestimated. Still, our results show that none of the drugs administered as single dose resulted in cure of all infected children. Only moderate CRs were observed against G. intestinalis, E. histolytica/E. dispar and the potentially pathogenic B. hominis. A further limitation is the relatively small number of positive children for specific intestinal protozoa. Infection intensity did not significantly decrease after treatment with the exception of E. coli in the nitazoxanide group. Note that intestinal protozoa multiply within the host, which can also influence infection intensity results. Further, the incubation time for different intestinal protozoa is between 7 and 28 days, meaning that it is possible that within the 3-week period before and after treatment, re-infection had occured[36–40]. The clinical relevance of the moderate CRs suggests that single-dose albendazole or nitazoxanide or a combination of the two drugs do not have sufficient efficacy against pathogenic intestinal protozoa. However, multiple stool samples should be examined to strengthen the diagnostic accuracy for these infections.