Effects of Lactiplantibacillus plantarum and Lacticaseibacillus paracasei supplementation on the single-cell fecal parasitome in children with celiac disease autoimmunity: a randomized, double-blind placebo-controlled clinical trial.

BACKGROUND: Lactiplantibacillus plantarum HEAL9 and Lacticaseibacillus paracasei 8700:2 positively affect the fecal bacteriome in children with celiac disease autoimmunity after 6 months of supplementation. The aim of the present investigation was to study the effects of Lactiplantibacillus plantarum HEAL9 and Lacticaseibacillus paracasei 8700:2 on the single-cell parasitome, with a primary focus on Blastocystis. METHODS: Stool samples were collected from 78 Swedish children with celiac disease autoimmunity participating in a randomized, double-blind, placebo-controlled clinical trial to either receive a mixture of supplementation with L. plantarum HEAL9 and L. paracasei 8700:2 (n = 38) or placebo (n = 40). A total of 227 stool samples collected at baseline and after 3 and 6 months of intervention, respectively, were retrospectively analyzed for Blastocystis by quantitative real-time PCR and subtyped by massively parallel amplicon sequencing. Other single-cell parasites were detected by untargeted 18S rDNA amplicon sequencing and verified by real-time PCR. The relation between the parasites and the bacteriome community was characterized by using 16S rDNA profiling of the V3-V4 region. RESULTS: Three different single-cell protists were identified, of which the highest prevalence was found for Dientamoeba fragilis (23.1%, 18/78 children), followed by Blastocystis (15.4%, 12/78) and Entamoeba spp. (2.6%, 2/78). The quantity of the protists was stable over time and not affected by probiotic intervention (P = 0.14 for Blastocystis, P = 0.10 for D. fragilis). The positivity of the protists was associated with increased bacteriome diversity (measured by multiple indices, P < 0.03). Bacterial composition was influenced by the presence of the protists: positivity of Blastocystis was inversely associated with Akkermansia (at the levels of the genus as well as its family, order, class and phylum); P < 0.002), Faecalibacterium (P = 0.003) and Romboutsia (P = 0.029); positivity of D. fragilis was inversely associated with families Enterobacteriaceae (P = 0.016) and Coriobacteriaceae (P = 0.022) and genera Flavonifractor (P < 0.001), Faecalibacterium (P = 0.009), Lachnoclostridium (P = 0.029), Ruminococcus (P < 0.001) and Granulicatella (P = 0.018). CONCLUSIONS: The prevalence of single-cell protists is low in children with celiac disease autoimmunity. The colonization was stable regardless of the probiotic intervention and associated with increased diversity of the fecal bacteriome but inversely associated with some beneficial bacteria.

The opportunistic protist, Giardia intestinalis, occurs in gut-healthy humans in a high-income country.

Giardia intestinalis, a cosmopolitan gastrointestinal protist, is detected mainly in patients with clinical giardiasis in high-income countries. In contrast, there is very little information on the presence of Giardia in asymptomatic individuals. Therefore, the aim of this study was to determine the presence and prevalence of Giardia in gut-healthy volunteers in the Czech Republic and to perform a comparative evaluation of different diagnostic methods, since Giardia diagnostics is complicated. Our results confirmed that the qPCR method is the most sensitive method for detecting Giardia and revealed a prevalence of 7% (22/296) in asymptomatic individuals. In most cases, the colonization intensity ranged from 10-1-101. A conventional PCR protocol targeting the TPI gene was used to identify the assemblages. However, this protocol had limited sensitivity for Giardia amplification, effectively detecting colonization above an intensity of 104. In addition, Giardia was detected in 19% of the animals, which were closely associated with the study participants. However, due to methodological limitations, zoonotic transmission could not be clearly confirmed. Notably, contact with animals proved to be the only factor that had a significant impact on the incidence of Giardia in gut-healthy humans.

A Cross-Sectional Study on the Occurrence of the Intestinal Protist, Dientamoeba fragilis, in the Gut-Healthy Volunteers and Their Animals.

Dientamoeba fragilis is a cosmopolitan intestinal protist colonizing the human gut with varying prevalence depending on the cohort studied and the diagnostic methods used. Its role in human health remains unclear mainly due to the very sporadic number of cross-sectional studies in gut-healthy populations. The main objective of this study was to expand knowledge of the epidemiology of D. fragilis in gut-healthy humans and their animals. A total of 296 stool samples from humans and 135 samples from 18 animal species were analyzed. Using qPCR, a prevalence of 24% was found in humans in contrast to conventional PCR (7%). In humans, several factors were found to influence the prevalence of D. fragilis. A more frequent occurrence of D. fragilis was associated with living in a village, traveling outside Europe and contact with farm animals. In addition, co-infection with Blastocystis spp. was observed in nearly half of the colonized humans. In animals, D. fragilis was detected in 13% of samples from eight species using qPCR. Our molecular phylogenies demonstrate a more frequent occurrence of Genotype 1 in gut-healthy humans and also revealed a likely a new protist species/lineage in rabbits related to D. fragilis and other related organisms.

Comparison of molecular diagnostic approaches for the detection and differentiation of the intestinal protist Blastocystis sp. in humans.

Blastocystis is the most commonly found intestinal protist in the world. Accurate detection and differentiation of Blastocystis including its subtypes (arguably species) are essential to understand its epidemiology and role in human health. We compared (i) the sensitivity of conventional PCR (cPCR) and qPCR in a set of 288 DNA samples obtained from stool samples of gut-healthy individuals, and (ii) subtype diversity as detected by next-generation sequencing (NGS) versus Sanger sequencing. Real-time PCR resulted in more positive samples than cPCR, revealing high fecal load of Blastocystis based on the quantification curve in most samples. In subtype detection, NGS was largely in agreement with Sanger sequencing but showed higher sensitivity for mixed subtype colonization within one host. This fact together with use of the combination of qPCR and NGS and obtaining information on the fecal protist load will be beneficial for epidemiological and surveillance studies.

Title: Comparaison des approches de diagnostic moléculaire pour la détection et la différenciation du protiste intestinal Blastocystis sp. chez l'homme. Abstract: Blastocystis est le protiste intestinal le plus répandu dans le monde. La détection et la différenciation précises de Blastocystis, y compris ses sous-types (sans doute des espèces), sont essentielles pour comprendre son épidémiologie et son rôle dans la santé humaine. Nous avons comparé (i) la sensibilité de la PCR conventionnelle (cPCR) et de la qPCR dans un ensemble de 288 échantillons d'ADN obtenus à partir d'échantillons de selles d'individus en bonne santé intestinale et (ii) la diversité des sous-types détectée par le séquençage de nouvelle génération (NGS) par rapport au séquençage Sanger. La PCR en temps réel a donné plus d'échantillons positifs que la cPCR, révélant une charge fécale élevée de Blastocystis sur la base de la courbe de quantification dans la plupart des échantillons. Dans la détection des sous-types, le NGS était largement en accord avec le séquençage de Sanger mais a montré une sensibilité plus élevée pour la colonisation de sous-types mixtes au sein d'un hôte. Ce fait, associé à l'utilisation de la combinaison de qPCR et de NGS et à l'obtention d'informations sur la charge fécale de protistes, sera bénéfique pour les études épidémiologiques et de surveillance.

Altered gut ecosystems plus the microbiota's potential for rapid evolution: A recipe for inevitable change with unknown consequences.

In a single human gut, which is estimated to produce 1000-times more bacteria in a single day than the entire human population on Earth as of 2020, the potential for evolution is vast. In addition to the sheer volume of reproductive events, prokaryotes can transfer most genes horizontally, greatly accelerating their potential to evolve. In the face of this evolutionary potential, Westernization has led to profound changes in the ecosystem of the gut, including increased chronic inflammation in many individuals and dramatically reduced fiber consumption and decreased seasonal variation in the diet of most individuals. Experimental work using a variety of model systems has shown that bacteria will evolve within days to weeks when faced with substantial environmental changes. However, studies evaluating the effects of inflammation of the gut on the microbiota are still in their infancy and generally confounded by the effects of the microbiota on the immune system. At the same time, experimental data indicate that complete loss of fiber from the diet constitutes an extinction-level event for the gut microbiota. However, these studies evaluating diet may not apply to Westernized humans who typically have reduced but not absent levels of fiber in their diet. Thus, while it is expected that the microbiota will evolve rapidly in the face of Westernization, experimental studies that address the magnitude of that evolution are generally lacking, and it remains unknown to what extent this evolutionary process affects disease and the ability to treat the disease state.

How monoxenous trypanosomatids revealed hidden feeding habits of their tsetse fly hosts.

Tsetse flies are well-known vectors of trypanosomes pathogenic for humans and livestock. For these strictly blood-feeding viviparous flies, the host blood should be the only source of nutrients and liquids, as well as any exogenous microorganisms colonising their intestine. Here we describe the unexpected finding of several monoxenous trypanosomatids in their gut. In a total of 564 individually examined Glossina (Austenia) tabaniformis (Westwood) (436 specimens) and Glossina (Nemorhina) fuscipes fuscipes (Newstead) (128 specimens) captured in the Dzanga-Sangha Protected Areas, Central African Republic, 24 (4.3%) individuals were infected with monoxenous trypanosomatids belonging to the genera Crithidia Léger, 1902; Kentomonas Votýpka, Yurchenko, Kostygov et Lukes, 2014; Novymonas Kostygov et Yurchenko, 2020; Obscuromonas Votýpka et Lukes, 2021; and Wallacemonas Kostygov et Yurchenko, 2014. Moreover, additional 20 (3.5%) inspected tsetse flies harboured free-living bodonids affiliated with the genera Dimastigella Sandon, 1928; Neobodo Vickerman, 2004; Parabodo Skuja, 1939; and Rhynchomonas Klebs, 1892. In the context of the recently described feeding behaviour of these dipterans, we propose that they become infected while taking sugar meals and water, providing indirect evidence that blood is not their only source of food and liquids.

Blastocystis Colonization Alters the Gut Microbiome and, in Some Cases, Promotes Faster Recovery From Induced Colitis.

Protists are a normal component of mammalian intestinal ecosystems that live alongside, and interact with, bacterial microbiota. Blastocystis, one of the most common intestinal eukaryotes, is reported as a pathogen that causes inflammation and disease, though health consequences likely vary depending on host health, the gut ecosystem, and genetic diversity. Accumulating evidence suggests that Blastocystis is by and large commensal. Blastocystis is more common in healthy individuals than those with immune mediated diseases such as Inflammatory Bowel Diseases (IBD). Blastocystis presence is also associated with altered composition and higher richness of the bacterial gut microbiota. It is not clear whether Blastocystis directly promotes a healthy gut and microbiome or is more likely to colonize and persist in a healthy gut environment. We test this hypothesis by measuring the effect of Blastocystis ST3 colonization on the health and microbiota in a rat experimental model of intestinal inflammation using the haptenizing agent dinitrobenzene sulfonic acid (DNBS). We experimentally colonized rats with Blastocystis ST3 obtained from a healthy, asymptomatic human donor and then induced colitis after 3 weeks (short term exposure experiment) or after 13 weeks (long term exposure experiment) and compared these colonized rats to a colitis-only control group. Across experiments Blastocystis ST3 colonization alters microbiome composition, but not richness, and induces only mild gut inflammation but no clinical symptoms. Our results showed no effect of short-term exposure to Blastocystis ST3 on gut inflammation following colitis induction. In contrast, long-term Blastocystis exposure appears to promote a faster recovery from colitis. There was a significant reduction in inflammatory markers, pathology 2 days after colitis induction in the colonized group, and clinical scores also improved in this group. Blastocystis colonization resulted in a significant reduction in tumor necrosis factor alpha (TNFα) and IL-1ß relative gene expression, while expression of IFNγ and IL17re/17C were elevated. We obtained similar results in a previous pilot study. We further found that bacterial richness rebounded in rats colonized by Blastocystis ST3. These results suggest that Blastocystis sp. may alter the gut ecosystem in a protective manner and promote faster recovery from disturbance.

Helminth Interactions with Bacteria in the Host Gut Are Essential for Its Immunomodulatory Effect.

Colonization by the benign tapeworm, Hymenolepis diminuta, has been associated with a reduction in intestinal inflammation and changes in bacterial microbiota. However, the role of microbiota in the tapeworm anti-inflammatory effect is not yet clear, and the aim of this study was to determine whether disruption of the microflora during worm colonization can affect the course of intestinal inflammation. We added a phase for disrupting the intestinal microbiota using antibiotics to the experimental design for which we previously demonstrated the protective effect of H. diminuta. We monitored the immunological markers, clinical parameters, bacterial microbiota, and histological changes in the colon of rats. After a combination of colonization, antibiotics, and colitis induction, we had four differently affected experimental groups. We observed a different course of the immune response in each group, but no protective effect was found. Rats treated with colonization and antibiotics showed a strong induction of the Th2 response as well as a significant change in microbial diversity. The microbial results also revealed differences in the richness and abundance of some bacterial taxa, influenced by various factors. Our data suggest that interactions between the tapeworm and bacteria may have a major impact on its protective effect.

A Study on the Prevalence and Subtype Diversity of the Intestinal Protist Blastocystis sp. in a Gut-Healthy Human Population in the Czech Republic.

Blastocystis sp. is a common intestinal protist colonizing the human intestine the prevalence of which varies across non-industrialized and industrialized countries. Its role in the human gut ecosystem remains unclear due to persisting gaps in knowledge of epidemiology and factors affecting gut colonization. Here, we aimed to expand the knowledge of the epidemiology of Blastocystis sp. in the gut-healthy humans in one of the industrialized European countries, including the distribution of its subtypes, the correlation between its occurrence and several factors such as lifestyle, contact with animals, age, and sex. A total of 288 stool samples were obtained from asymptomatic individuals over the entire age-range and 136 samples from animals with which the volunteers were in frequent contact. All samples were examined in parallel by PCR and xenic in vitro culture. Blastocystis sp. was detected in samples from both human and non-human hosts. In humans, the overall prevalence was 24% and eight subtypes were found; in animals, the prevalence was 10%, and only five subtypes were detected. A higher incidence of Blastocystis sp. was observed in individuals (i) traveling outside Europe, (ii) in frequent contact with livestock, and (iii) over 50 years of age. We found no effect on gender on Blastocystis sp. colonization. Summary: This study provides data on the prevalence and diversity of the gut protist Blastocystis sp. and its subtypes in a gut-healthy human population with emphasis on several factors such as contact with animals, lifestyle, age, and gender.

Canine thelaziosis in the Czech Republic: the northernmost autochthonous occurrence of the eye nematode Thelazia callipaeda Railliet et Henry, 1910 in Europe.

The eye nematode Thelazia callipaeda Railliet et Henry, 1910 (Spirurida: Thelaziidae) is a vector-borne zoonotic nematode infecting a range of wild and domestic carnivores as well as humans. It is considered to be a causative agent of emerging and neglected disease and currently invades central part of Europe. Nematodes were collected from the eye of a dog living in Prague, which never travelled outside the Czech Republic. The nematodes were identified based on their morphology and partial sequence of the cox1 gene as T. callipaeda haplotype 1. This finding represents the northernmost record of autochthonous canine thelaziosis in Europe. The insufficient control of imported animals as well as free movement of dogs and wild carnivores within Europe probably facilitates spreading of T. callipaeda throughout the continent. To better understand the spreading of T. callipaeda and to prevent its zoonotic transmissions, information about the risk of this infection in newly invaded countries should be disseminated not only among veterinarians and physicians, but also within the community of pet owners and hunters.

Genetic diversity of the potentially therapeutic tapeworm Hymenolepis diminuta (Cestoda: Cyclophyllidea).

The cestode Hymenolepis diminuta is highly prevalent in wild rat populations and has also been observed rarely in humans, generally causing no apparent harm. The organism has been studied for decades in the laboratory, and its colonization of laboratory rats has recently been shown as protective against some inflammation-associated disorders. Recently, H. diminuta has become a leading candidate for helminth therapy, an emerging method of "biota enrichment" used to treat or prevent inflammatory diseases of humans in Western society. While most of the experimental isolates of H. diminuta are identified based on typical morphological features, hymenolepidid tapeworms may represent complexes of cryptic species as detected by molecular sequence data. In the present study, we explored the diversity of laboratory-kept strains using partial sequences of two genes (lsrDNA and cox1) and determined that H. diminuta isolates currently considered for therapeutic purposes in the US and Europe belong to a single, genetically nearly uniform lineage, showing only little genetic deviation from wild-caught isolates.

Evaluating rodent experimental models for studies of Blastocystis ST1.

Blastocystis is a common inhabitant of the human gut, colonizing at least one billion people at a prevalence ranging from <10% to 100% in healthy human populations globally. The majority of carriers remain asymptomatic, suggesting that Blastocystis is largely a commensal, though Blastocystis has also been implicated in disease in some people. However, there are no in vivo model systems in which to experimentally test the impact of Blastocystis on mammalian hosts and the gut ecosystem and determine which factors underlie these variable clinical outcomes. We evaluated a rat model for sustaining of a human-derived Blastocystis ST1 and assess colonization success and longevity. Because of the broad host range of Blastocystis, we compared the rat with three other rodent species to establish the reproducibility of our method. Blastocystis was introduced by esophageal gavage and colonization success evaluated by Blastocystis culture. Culture was also used to determine that all animals were negative prior to colonization and negative controls remain Blastocystis-free. In this study, Blastocystis ST1 established in 100% of the outbred rats (Rattus norvegicus) and gerbils (Meriones unguiculatus) challenged. Rats were colonized asymptomatically for more than one year, but Blastocystis ST1 was not transmitted between rats. Mus musculus strain CD1 and Mastomys coucha were not susceptible to Blastocystis ST1. Thus, rats appear to be a suitable in vivo model for studies of Blastocystis ST1, as do gerbils though testing was less extensive. This work lays the foundation for experimental work on the role of Blastocystis in health and disease.

The benign helminth Hymenolepis diminuta ameliorates chemically induced colitis in a rat model system.

The tapeworm Hymenolepis diminuta is a model for the impact of helminth colonization on the mammalian immune system and a candidate therapeutic agent for immune mediated inflammatory diseases (IMIDs). In mice, H. diminuta protects against models of inflammatory colitis by inducing a strong type 2 immune response that is activated to expel the immature worm. Rats are the definitive host of H. diminuta, and are colonized stably and over long time periods without harming the host. Rats mount a mild type 2 immune response to H. diminuta colonization, but this response does not generally ameliorate colitis. Here we investigate the ability of different life cycle stages of H. diminuta to protect rats against a model of colitis induced through application of the haptenizing agent dinitrobenzene sulphonic acid (DNBS) directly to the colon, and monitor rat clinical health, systemic inflammation measured by TNFα and IL-1ß, and the gut microbiota. We show that immature H. diminuta induces a type 2 response as measured by increased IL-4, IL-13 and IL-10 expression, but does not protect against colitis. In contrast, rats colonized with mature H. diminuta and challenged with severe colitis (two applications of DNBS) have lower inflammation and less severe clinical symptoms. This effect is not related the initial type 2 immune response. The gut microbiota is disrupted during colitis and does not appear to play an overt role in H. diminuta-mediated protection.

A benign helminth alters the host immune system and the gut microbiota in a rat model system.

Helminths and bacteria are major players in the mammalian gut ecosystem and each influences the host immune system and health. Declines in helminth prevalence and bacterial diversity appear to play a role in the dramatic rise of immune mediated inflammatory diseases (IMIDs) in western populations. Helminths are potent modulators of immune system and their reintroduction is a promising therapeutic avenue for IMIDs. However, the introduction of helminths represents a disturbance for the host and it is important to understand the impact of helminth reintroduction on the host, including the immune system and gut microbiome. We tested the impact of a benign tapeworm, Hymenolepis diminuta, in a rat model system. We find that H. diminuta infection results in increased interleukin 10 gene expression in the beginning of the prepatent period, consistent with induction of a type 2 immune response. We also find induction of humoral immunity during the patent period, shown here by increased IgA in feces. Further, we see an immuno-modulatory effect in the small intestine and spleen in patent period, as measured by reductions in tissue immune cells. We observed shifts in microbiota community composition during the patent period (beta-diversity) in response to H. diminuta infection. However, these compositional changes appear to be minor; they occur within families and genera common to both treatment groups. There was no change in alpha diversity. Hymenolepis diminuta is a promising model for helminth therapy because it establishes long-term, stable colonization in rats and modulates the immune system without causing bacterial dysbiosis. These results suggest that the goal of engineering a therapeutic helminth that can safely manipulate the mammalian immune system without disrupting the rest of the gut ecosystem is in reach.

Molecular identification of Entamoeba species in savanna woodland chimpanzees (Pan troglodytes schweinfurthii).

To address the molecular diversity and occurrence of pathogenic species of the genus Entamoeba spp. in wild non-human primates (NHP) we conducted molecular-phylogenetic analyses on Entamoeba from wild chimpanzees living in the Issa Valley, Tanzania. We compared the sensitivity of molecular [using a genus-specific polymerase chain reaction (PCR)] and coproscopic detection (merthiolate-iodine-formaldehyde concentration) of Entamoeba spp. We identified Entamoeba spp. in 72 chimpanzee fecal samples (79%) subjected to species-specific PCRs for six Entamoeba species/groups (Entamoeba histolytica, Entamoeba nuttalli, Entamoeba dispar, Entamoeba moshkovskii, Entamoeba coli and Entamoeba polecki ST2). We recorded three Entamoeba species: E. coli (47%), E. dispar (16%), Entamoeba hartmanni (51%). Coproscopically, we could only distinguish the cysts of complex E. histolytica/dispar/moshkovskii/nuttalli and E. coli. Molecular prevalence of entamoebas was higher than the prevalence based on the coproscopic examination. Our molecular phylogenies showed that sequences of E. dispar and E. coli from Issa chimpanzees are closely related to sequences from humans and other NHP from GenBank. The results showed that wild chimpanzees harbour Entamoeba species similar to those occurring in humans; however, no pathogenic species were detected. Molecular-phylogenetic methods are critical to improve diagnostics of entamoebas in wild NHP and for determining an accurate prevalence of Entamoeba species.

A tsetse and tabanid fly survey of African great apes habitats reveals the presence of a novel trypanosome lineage but the absence of Trypanosoma brucei.

Tsetse and tabanid flies transmit several Trypanosoma species, some of which are human and livestock pathogens of major medical and socioeconomic impact in Africa. Recent advances in molecular techniques and phylogenetic analyses have revealed a growing diversity of previously unidentified tsetse-transmitted trypanosomes potentially pathogenic to livestock and/or other domestic animals as well as wildlife, including African great apes. To map the distribution, prevalence and co-occurrence of known and novel trypanosome species, we analyzed tsetse and tabanid flies collected in the primary forested part of the Dzanga-Sangha Protected Areas, Central African Republic, which hosts a broad spectrum of wildlife including primates and is virtually devoid of domestic animals. Altogether, 564 tsetse flies and 81 tabanid flies were individually screened for the presence of trypanosomes using 18S rRNA-specific nested PCR. Herein, we demonstrate that wildlife animals are parasitized by a surprisingly wide range of trypanosome species that in some cases may circulate via these insect vectors. While one-third of the examined tsetse flies harbored trypanosomes either from the Trypanosoma theileri, Trypanosoma congolense or Trypanosoma simiae complex, or one of the three new members of the genus Trypanosoma (strains 'Bai', 'Ngbanda' and 'Didon'), more than half of the tabanid flies exclusively carried T. theileri. To establish the putative vertebrate hosts of the novel trypanosome species, we further analyzed the provenance of blood meals of tsetse flies. DNA individually isolated from 1033 specimens of Glossina spp. and subjected to high-throughput library-based screening proved that most of the examined tsetse flies engorged on wild ruminants (buffalo, sitatunga, bongo), humans and suids. Moreover, they also fed (albeit more rarely) on other vertebrates, thus providing indirect but convincing evidence that trypanosomes can be transmitted via these vectors among a wide range of warm- and cold-blooded hosts.

Molecular phylogeny of anoplocephalid tapeworms (Cestoda: Anoplocephalidae) infecting humans and non-human primates.

Anoplocephalid tapeworms of the genus Bertiella Stiles and Hassall, 1902 and Anoplocephala Blanchard, 1848, found in the Asian, African and American non-human primates are presumed to sporadic ape-to-man transmissions. Variable nuclear (5.8S-ITS2; 28S rRNA) and mitochondrial genes (cox1; nad1) of isolates of anoplocephalids originating from different primates (Callicebus oenanthe, Gorilla beringei, Gorilla gorilla, Pan troglodytes and Pongo abelii) and humans from various regions (South America, Africa, South-East Asia) were sequenced. In most analyses, Bertiella formed a monophyletic group within the subfamily Anoplocephalinae, however, the 28S rRNA sequence-based analysis indicated paraphyletic relationship between Bertiella from primates and Australian marsupials and rodents, which should thus be regarded as different taxa. Moreover, isolate determined as Anoplocephala cf. gorillae from mountain gorilla clustered within the Bertiella clade from primates. This either indicates that A. gorillae deserves to be included into the genus Bertiella, or, that an unknown Bertiella species infects also mountain gorillas. The analyses allowed the genetic differentiation of the isolates, albeit with no obvious geographical or host-related patterns. The unexpected genetic diversity of the isolates studied suggests the existence of several Bertiella species in primates and human and calls for revision of the whole group, based both on molecular and morphological data.

Wild chimpanzees are infected by Trypanosoma brucei.

Although wild chimpanzees and other African great apes live in regions endemic for African sleeping sickness, very little is known about their trypanosome infections, mainly due to major difficulties in obtaining their blood samples. In present work, we established a diagnostic ITS1-based PCR assay that allows detection of the DNA of all four Trypanosoma brucei subspecies (Trypanosoma brucei brucei, Trypanosoma brucei rhodesiense, Trypanosoma brucei gambiense, and Trypanosoma brucei evansi) in feces of experimentally infected mice. Next, using this assay we revealed the presence of trypanosomes in the fecal samples of wild chimpanzees and this finding was further supported by results obtained using a set of primate tissue samples. Phylogenetic analysis of the ITS1 region showed that the majority of obtained sequences fell into the robust T. brucei group, providing strong evidence that these infections were caused by T. b. rhodesiense and/or T. b. gambiense. The optimized technique of trypanosome detection in feces will improve our knowledge about the epidemiology of trypanosomes in primates and possibly also other endangered mammals, from which blood and tissue samples cannot be obtained. Finally, we demonstrated that the mandrill serum was able to efficiently lyse T. b. brucei and T. b. rhodesiense, and to some extent T. b. gambiense, while the chimpanzee serum failed to lyse any of these subspecies.

Ecology of malaria infections in western lowland gorillas inhabiting Dzanga Sangha Protected Areas, Central African Republic.

African great apes are susceptible to infections with several species of Plasmodium, including the predecessor of Plasmodium falciparum. Little is known about the ecology of these pathogens in gorillas. A total of 131 gorilla fecal samples were collected from Dzanga-Sangha Protected Areas to study the diversity and prevalence of Plasmodium species. The effects of sex and age as factors influencing levels of infection with Plasmodium in habituated gorilla groups were assessed. Ninety-five human blood samples from the same locality were also analysed to test for cross-transmission between humans and gorillas. According to a cytB PCR assay 32% of gorilla's fecal samples and 43·1% human individuals were infected with Plasmodium spp. All Laverania species, Plasmodium vivax, and for the first time Plasmodium ovale were identified from gorilla samples. Plasmodium praefalciparum was present only from habituated individuals and P. falciparum was detected from human samples. Although few P. vivax and P. ovale sequences were obtained from gorillas, the evidence for cross-species transmission between humans and gorillas requires more in depth analysis. No association was found between malaria infection and sex, however, younger individuals aged ≤6 years were more susceptible. Switching between two different Plasmodium spp. was observed in three individuals. Prolonged monitoring of Plasmodium infection during various seasons and recording behavioural data is necessary to draw a precise picture about the infection dynamics.