Environmental surveillance of zoonotic protozoa and fecal indicators in surface waters of a subtropical transition zone in Southern Brazil

Waterborne protozoa pose a significant challenge to public health, particularly in regions with deficient sanitation. This study aimed to evaluate the occurrence of Giardia spp., Cryptosporidium spp., and Cystoisospora spp. in surface waters of the Pelotas microregion, Southern Brazil, and their association with fecal indicator bacteria ( Escherichia coli ) and rainfall events. From March to August 2025, eight sampling points (streams, urban canals, and recreational beaches) were monitored across four campaigns ( n = 32 samples). Water samples were analyzed for physico-chemical parameters, E. coli quantification, and molecular detection (PCR) of protozoa. Giardia spp. and Cystoisospora spp. DNA were frequently and consistently detected across campaigns (detection rates of 37.5–50%), irrespective of the marked rainfall gradient (Spearman’s ρ = -0.08 to -0.11, p > 0.5). In contrast, E. coli counts declined significantly with increasing rainfall and decreasing water temperature (ρ = -0.47 and 0.73, respectively; p < 0.05), with a pronounced reduction during the wettest, coolest campaign (180.4 mm; <14 °C). Cryptosporidium spp. was not detected in any sample. These findings indicate that E. coli counts are not a reliable proxy for the molecular detection of Giardia and Cystoisospora in this watershed, underscoring the value of complementing bacterial indicators with protozoan surveillance in subtropical transition zones, while the host origin (zoonotic versus anthroponotic) and viability of the detected protozoa remain to be established.