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Human health and the human microbiome are inevitably intertwined, increasing their relevance in clinical research. However, the collection, transportation and storage of faecal samples may introduce bias due to methodological differences, especially since postal shipping is a common practise in large-scale clinical cohort studies. Using four different Omics layer, we determined the structural (16S rRNA sequencing, cytometric microbiota profiling) and functional integrity (SCFAs, global metabolome) of the microbiota in relation to different easy-to-handle conditions. These conditions were storage at −20 °C, −20 °C as glycerol stock, 4 °C and room temperature with and without oxygen exposure for a maximum of one week. Storage time affected the microbiota on all Omics levels. However, the magnitude was donor-dependent, highlighting the need for purpose-optimized sample collection in clinical multi-donor studies. The effects of oxygen exposure were negligible for all analyses. At ambient temperature, SCFA and compositional profiles were stable for 24 h and 48 h, respectively, while at 4 °C, SCFA profiles were maintained for 48 h. The global metabolome was highly susceptible, already changing at 24 h in non-frozen conditions. Thus, faecal microbiota was best preserved on all levels when transported as a native sample frozen within 24 h, leading to the least biased outcomes in the analysis. We conclude that the immediate freezing of native stool samples for transportation to the lab is best suited for planned multi-Omics analyses that include metabolomics to extend standard sequencing approaches.

Human health and the human microbiome are inevitably intertwined, increasing their relevance in clinical research. However, the collection, transportation and storage of faecal samples may introduce bias due to methodological differences, especially since postal shipping is a common practise in large-scale clinical cohort studies. Using four different Omics layer, we determined the structural (16S rRNA sequencing, cytometric microbiota profiling) and functional integrity (SCFAs, global metabolome) of the microbiota in relation to different easy-to-handle conditions. These conditions were storage at -20 °C, -20 °C as glycerol stock, 4 °C and room temperature with and without oxygen exposure for a maximum of one week. Storage time affected the microbiota on all Omics levels. However, the magnitude was donor-dependent, highlighting the need for purpose-optimized sample collection in clinical multi-donor studies. The effects of oxygen exposure were negligible for all analyses. At ambient temperature, SCFA and compositional profiles were stable for 24 h and 48 h, respectively, while at 4 °C, SCFA profiles were maintained for 48 h. The global metabolome was highly susceptible, already changing at 24 h in non-frozen conditions. Thus, faecal microbiota was best preserved on all levels when transported as a native sample frozen within 24 h, leading to the least biased outcomes in the analysis. We conclude that the immediate freezing of native stool samples for transportation to the lab is best suited for planned multi-Omics analyses that include metabolomics to extend standard sequencing approaches., Competing Interests: The authors declare no conflict of interest.

Trypanosomes are a group of pathogens distributed in the continents of Africa, America, Asia and Europe, and they affect all vertebrates including the neotropical primate group. Information about the trypanosome’s diversity, phylogeny, ecology and pathology in non-human primates (NHPs) from the neotropical region is scarce. The objective of the study was to identify Trypanosoma and Babesia molecularly in NHPs under the phylogenetic species concept. We extracted DNA from a total of 76 faecal samples collected between 2019 and 2021, from a total of 11 non-human primate species of which 46 are from captive NHPs and 30 are free-living NHPs in the Western Amazon region of Ecuador. We did not detect DNA of Babesia sp. by polymerase chain reaction test in any of the faecal samples. However, the nested-PCR-based method revealed Trypanosoma parasites by ITS gene amplification in two faecal samples; one for the species Leontocebus lagonotus (from the captive population) and a second one for Cebus albifrons (from the free-ranging population). Maximum parsimony and likelihood methods with the Kimura2+G+I model inferred the evolutionary history of the two records, which showed an evolutionary relationship with the genus Trypanosoma. Two sequences are monophyletic with Trypanosoma. However, the number of sequences available in GenBank for their species identification is limited. The two samples present different molecular identifications and evolutionary origins in the tree topology. We are most likely referring to two different species, and two different localities of infection. We suggest that health management protocols should be implemented to prevent the transmission of blood-borne pathogens such as Trypanosoma sp. among captive populations. In addition, these protocols also protect the personnel of wildlife rehabilitation centers working in close proximity to NHPs and vice versa.

This research described the co-infection prevalence of endoparasites in Tremartus ornatus and domestic animals in the rural high mountains of Colombia by copro-parasitological examination. Some parasites have a zoonotic potential in wild endangered species and domestic animals in Colombian regions. T. ornatus had a notable infection with Eimeria spp., Ascaris spp., Ancylostoma spp., and Baylisascaris spp. Cryptosporidium spp., Balantidium coli, Anoplocephala spp., and Acanthamoeba spp. In B. taurus, Eimeria spp. is coinfecting with Cryptosporidium spp. (6.6%) and represents 18% of the total parasitism. In E. caballus and B. taurus. Eimeria spp. coinfecting (34.7%), with the Strongylus spp. (21.9–25%). In T. ornatus, Eimeria spp. is coinfecting with Ancylostoma spp. (36.2%), Cryptosporidium spp., Ascaris spp., Baylisascaris spp., and B. coli.

Accurate diagnosis of colorectal cancer (CRC) still relies on invasive colonoscopy. Noninvasive methods are less sensitive in detecting the disease, particularly in the early stage. In the current work, a metabolomics analysis of fecal samples was carried out by ultra-high-performance liquid chromatography–tandem mass spectroscopy (UPLC-MS/MS). A total of 1380 metabolites were analyzed in a cohort of 120 fecal samples from patients with normal colonoscopy, advanced adenoma (AA) and CRC. Multivariate analysis revealed that metabolic profiles of CRC and AA patients were similar and could be clearly separated from control individuals. Among the 25 significant metabolites, sphingomyelins (SM), lactosylceramides (LacCer), secondary bile acids, polypeptides, formiminoglutamate, heme and cytidine-containing pyrimidines were found to be dysregulated in CRC patients. Supervised random forest (RF) and logistic regression algorithms were employed to build a CRC accurate predicted model consisting of the combination of hemoglobin (Hgb) and bilirubin E,E, lactosyl-N-palmitoyl-sphingosine, glycocholenate sulfate and STLVT with an accuracy, sensitivity and specificity of 91.67% (95% Confidence Interval (CI) 0.7753–0.9825), 0.7 and 1, respectively.

Background: Malaria is a disease caused by hemoparasites of the Plasmodium genus. Non-human primates (NHP) are hosts of Plasmodium sp. around the world. Several studies have demonstrated that Plasmodium sp. emerged from Africa. However, little information is currently available about Plasmodium falciparum in the neotropical NHP and even less in Ecuador. Indeed, the objective of our study was to identify by molecular phylogenetic analyses the Plasmodium species associated with NHP from the Western Amazon region of Ecuador, and to design a molecular taxonomy protocol to use in the NHP disease ecology. Methods: We extracted DNA from faecal samples (n = 26) from nine species of captive (n = 19) and free-ranging (n = 7) NHP, collected from 2011 to 2019 in the Western Amazon region of Ecuador. Results: Using a pan-Plasmodium PCR, we obtained one positive sample from an adult female Leontocebus lagonotus. A maximum likelihood phylogenetic analysis showed that this sequence unequivocally clustered with Plasmodium falciparum. Conclusions: The identification of Plasmodium sp. in NHP of the Ecuadorian Amazon would be essential to identify their role as potential zoonotic reservoirs, and it is also important to identify their origin in wildlife and their transmission in captive NHP.

This study aimed to characterise the gut microbiome composition of European hares (Lepus europaeus) and its potential changes after a short-term diet modification. The high sensitivity of European hare to habitat changes makes this species a good model to analyse possible alterations in gut microbiome after the introduction of additional nourishment into the diet. In total, 20 pairs were chosen for the experiments; 10 pairs formed the control group and were fed with standard fodder. The other 10 pairs represented the experimental group, whose diet was integrated with apples and carrots. The DNA from fresh faecal pellets collected after 4 days from the start of the experiment was extracted and the V3-V4 hypervariable regions were amplified and sequenced using the Illumina MiSeq® platform. The obtained amplicon sequence variants were classified into 735 bacterial genera belonging to 285 families and 36 phyla. The control and the experimental groups appeared to have a homogenous dispersion for the two taxonomic levels analysed with the most abundant phyla represented by Bacteroidetes and Firmicutes. No difference between control and experimental samples was detected, suggesting that the short-term variation in food availability did not alter the hares’ gut microbiome. Further research is needed to estimate significant time threshold.

Trypanosomes are a group of pathogens distributed in the continents of Africa, America, Asia and Europe, and they affect all vertebrates including the neotropical primate group. Information about the trypanosome's diversity, phylogeny, ecology and pathology in non-human primates (NHPs) from the neotropical region is scarce. The objective of the study was to identify Trypanosoma and Babesia molecularly in NHPs under the phylogenetic species concept. We extracted DNA from a total of 76 faecal samples collected between 2019 and 2021, from a total of 11 non-human primate species of which 46 are from captive NHPs and 30 are free-living NHPs in the Western Amazon region of Ecuador. We did not detect DNA of Babesia sp. by polymerase chain reaction test in any of the faecal samples. However, the nested-PCR-based method revealed Trypanosoma parasites by ITS gene amplification in two faecal samples; one for the species Leontocebus lagonotus (from the captive population) and a second one for Cebus albifrons (from the free-ranging population). Maximum parsimony and likelihood methods with the Kimura2+G+I model inferred the evolutionary history of the two records, which showed an evolutionary relationship with the genus Trypanosoma . Two sequences are monophyletic with Trypanosoma. However, the number of sequences available in GenBank for their species identification is limited. The two samples present different molecular identifications and evolutionary origins in the tree topology. We are most likely referring to two different species, and two different localities of infection. We suggest that health management protocols should be implemented to prevent the transmission of blood-borne pathogens such as Trypanosoma sp. among captive populations. In addition, these protocols also protect the personnel of wildlife rehabilitation centers working in close proximity to NHPs and vice versa.

Background: Malaria is a disease caused by hemoparasites of the Plasmodium genus. Non-human primates (NHP) are hosts of Plasmodium sp. around the world. Several studies have demonstrated that Plasmodium sp. emerged from Africa. However, little information is currently available about Plasmodium falciparum in the neotropical NHP and even less in Ecuador. Indeed, the objective of our study was to identify by molecular phylogenetic analyses the Plasmodium species associated with NHP from the Western Amazon region of Ecuador, and to design a molecular taxonomy protocol to use in the NHP disease ecology. Methods: We extracted DNA from faecal samples (n = 26) from nine species of captive (n = 19) and free-ranging (n = 7) NHP, collected from 2011 to 2019 in the Western Amazon region of Ecuador. Results: Using a pan-Plasmodium PCR, we obtained one positive sample from an adult female Leontocebus lagonotus . A maximum likelihood phylogenetic analysis showed that this sequence unequivocally clustered with Plasmodium falciparum . Conclusions: The identification of Plasmodium sp. in NHP of the Ecuadorian Amazon would be essential to identify their role as potential zoonotic reservoirs, and it is also important to identify their origin in wildlife and their transmission in captive NHP.