Free-living amoebae (FLA) can be found both in natural aquatic environments and in artificial, man-made aquatic environments. For a long time, (naked) FLA were considered to be harmless protozoa of soil and water. However, research since the 1960s has demonstrated that FLA can be pathogenic to humans and animals with nearly 100% morbidity from some strains. As etiological agents of the so-called Acanthamoebiasis, the Acanthamoebae can trigger several specific diseases (or symptoms) in humans. The amoebiasis of the central nervous system is called granulomatous amebic encephalitis (GAE), when Acanthamoebae are the etiological agents, showing subacute to chronic progress. GAE differs clinically from the primary amoebic meningoencephalitis (PAM(E)), which is caused by Naegleria fowleri. The Acanthamoeba keratitis is not necessarily associated with an immune suppression, but rather with a trauma, exposure to contaminated water or, particularly, the improper handling of contact lenses, which promotes infection. The clinical picture of GAE by Balamuthia mandrillaris is characterized by headache and neck stiffness. The infection is chronic and the time between infection and appearance of neurological symptoms may range from one month to about two years. The prevalence of FLA in water networks is associated with biofilms, where the amoebae live within a biocoenosis sympatric with other microorganisms. These biofilms serve as feeding grounds for the FLA and provide protection to a certain degree while the FLA adhere to the surfaces. In such a biocoenosis there are multiple interactions between FLA and other microorganisms: In addition to their role as pathogens, FLA are known to serve as host of and vehicles for diverse intracellular organisms (bacteria, viruses, eucaryonts), some of them being natural human pathogens. They act as reservoir or vehicle for various microorganisms such as various Legionella sp., Listeria monocytogenes, Pseudomonas aeruginosa and some Mycobacterium sp. In the cyst-stage of the FLA these intracellular organisms (endocytobionts) are protected to a high degree against any adverse environment (FLA as “Trojan horse”). This host-endocytobiont relationship may further lead to health risks in terms of the development of pathogenicity/virulence and antibiotic resistance (FLA as “Trainings ground”).

Acanthamoeba is a ubiquitous genus of amoebae that can act as opportunistic parasites in both humans and animals, causing a variety of ocular, nervous and dermal pathologies. Despite advances in Acanthamoeba therapy, the management of patients with Acanthamoeba infections remains a challenge for health services. Therefore, there is a need to search for new active substances against Acanthamoebae. In the present study, we evaluated the amoebicidal activity of nitroxoline against the trophozoite and cyst stages of six different strains of Acanthamoeba. The strain A. griffini showed the lowest IC₅₀ value in the trophozoite stage (0.69 ± 0.01 µM), while the strain A. castellanii L-10 showed the lowest IC₅₀ value in the cyst stage (0.11 ± 0.03 µM). In addition, nitroxoline induced in treated trophozoites of A. culbertsoni features compatibles with apoptosis and autophagy pathways, including chromatin condensation, mitochondrial malfunction, oxidative stress, changes in cell permeability and the formation of autophagic vacuoles. Furthermore, proteomic analysis of the effect of nitroxoline on trophozoites revealed that this antibiotic induced the overexpression and the downregulation of proteins involved in the apoptotic process and in metabolic and biosynthesis pathways.

The diversity of nematodes infecting amphibians is understudied in tropical Africa and unknown in Rwanda. Diversity assessment is hampered by the fact that species descriptions refer mostly to morphological features that are unlinked to DNA sequences of marker genes available in public databases. In this paper, we explore the abundance and diversity of parasitic nematodes in reed frogs Hyperolius kivuensis (n = 115), H. parallelus (n = 45) and H. viridiflavus (n = 100) collected in Rwanda. Five nematode species were identified morphologically as Orneoascaris chrysanthemoides, O. schoutedeni, Gendria leberrei, Aplectana chamaeleonis and Rhabdias collaris. Corresponding DNA sequences of 18S and COI genes were determined and subsequently deposited in GenBank. Aplectana chamaeleonis showed the highest prevalence (8.7%), but O. chrysanthemoides the highest mean intensity of infection (6.0) and largest number (24) of individuals in H. kivuensis. To the best of our knowledge, all amphibian hosts are new records for these nematode species, which are known to infect a wide range of amphibian and reptile species. Our findings suggest that nematode diversity is probably lower than previously assumed due to low host specificity. As morphological species identification is often challenging, our data facilitate molecular identification of adult and specifically larval nematodes found in amphibians of Sub-Saharan Africa.

Clinostomum metacercariae are common endoparasites of fish and frogs. In this study, we examine taxonomic identity and developmental strategy of Clinostomum metacercariae infesting reed frogs Hyperolius kivuensis and H. viridiflavus in Rwanda. Moreover, we evaluate the impact of infestation on demographic and morphological life-history traits of the hosts. Morphological and molecular features, particularly genital morphology and COX1 sequences, provided evidence that the metacercariae belong to C. chabaudi Vercammen-Grandjean, 1960. Depending on the host’s defensive behavior and the availability of resources, metacercariae develop either as sedentary “yellow grubs” encysted in the lymphatic sacs or mouth of the host or as initially encysted, but later free-ranging individuals invading the host´s body cavity. Nutrition on lymphatic fluid within the cyst leads to yellow-colored gut content, feeding on blood or host tissue, to brownish green gut content in free-ranging individuals. Almost all metacercariae opted for the first developmental strategy in H. kivuensis, whereas the second strategy dominated in metacercariae infesting H. viridiflavus. Hyperolius kivuensis suffered significant morphological modifications, when infested with encysted metacercariae. Both developmental modes permitted a coexistence with the host of less than one year. We hypothesize that the presence of alternative development modes is an adaptation of C. chabaudi to cope with resource limitation within host-produced cysts.

Most of our knowledge on the processes structuring parasite communities in amphibians originate from temperate-zone taxa, whereas Afrotropical communities have been neglected so far. We found evidence that ecological fitting of the hosts and, probably, differential immune response may influence the variation in parasite species richness, prevalence, and infestation intensity of East African frogs Hyperolius kivuensis and H. viridiflavus. The most closely related host species share the same macrohabitat (that implies the same pool of potential parasites), but differ in microhabitat preference, so that a comparative analyses of syntopic and allopatric populations is expedient to reveal ecological fitting. We detected 11 parasite species (one annelid, four nematodes, five trematodes, one cestode) and two endocommensal species (protozoans). The component parasite communities included 4–5 helminth species in H. kivuensis and 6–8 in the more aquatic H. viridiflavus, supporting the hypothesis that trematode diversity increases with the amount of time spent in water. Five parasite species (Orneoascaris chrysanthemoides, Clinostomum chabaudi, an undetermined echinostomatid) and two protozoans (Nyctotheroides sp., and Protoopalina sp.) are shared among the syntopic amphibian populations. This finding indicates a similar susceptibility of these amphibians to infestation from the local parasite pool. Yet, the low prevalence of single- and multi-species infestations in H. kivuensis indicates that parasite clearing by its immune response is probably more effective and prominent than in H. viridiflavus. Therefore, H. viridiflavus suffered from significantly reduced short-term survival due to the infection. Thus, we conclude that the processes structuring component parasite communities in amphibians do not differ generally between temperate-zone and Afrotropical host species, but they do in the magnitude of ecological fitting.