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In Nigeria and tropical Africa, the bias towards the protection of natural sites on account of terrestrial biodiversity or their resultant ecosystem services has led to a considerably low number of freshwater conservation studies in the region. To close this gap, six lotic freshwater systems in three different natural sites (Obudu Mountains, Agbokim Waterfalls, and Cross River National Park) along the Cameroon border of Nigeria were evaluated for the biodiversity of macroinvertebrates and some aspects of water quality in December 2021 and March 2022. Among other macroinvertebrate-related indices, the community conservation index (CCI), which considers the presence of threatened, rare, and notable species, was used for the conservation evaluation of the freshwater systems. The Ephemeroptera-Plecoptera-Trichoptera (EPT) species were notably rich at the sites, with over 12 species each at the Afundu Stream, Kwa River, Cataract Stream, and Agbokim River. All of the sites had a very high conservation value, according to the CCI, which was greater than 20. Some Obudu sites (Grotto and Becheve streams), despite having fewer EPT taxa, had higher CCI than the Kwa or Agbokim River. This was due to the presence of the endangered damselflies (Africocypha centripunctata and Allocnemis vicki) at the Obudu sites and a rare mayfly species in Nigeria (Tricorythus tinctus), which was reported at Agbokim and Kwa Rivers. The presence of a notable damselfly genus (Pentaphlebia) at the Afundu stream of the Obudu Mountains also raises some hope that the critically endangered damselfly (Pentaphlebia gamblesi), whose adult was last seen in 1973, could still be alive at the site. Canonical Correspondence Analysis revealed that the proportion of scrapers had a strong association with high BOD and EC in Agbokim River, while the Becheve and Grotto streams which had high vegetal cover was positively associated with shredders. Also, increasing DO in Afundu Stream, Kwa River and Cataract Stream was positively associated with EPT richness. This study revealed that more freshwater ecological studies need to be conducted at isolated natural sites and protected areas in Africa. Findings from such studies have implications for preserving freshwater ecosystems of high conservation value from the various threats that characterize the Anthropocene., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)