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Tritrichomonas foetus is a flagellated and anaerobic parasite able to infect cattle and felines. Despite its prevalence, there is no effective standardized or legal treatment for T. foetus-infected cattle; the vaccination still has limited success in mitigating infections and reducing abortion risk; and nowadays, the diagnosis of T. foetus presents important limitations in terms of sensitivity and specificity in bovines. Here, we characterize the plasma membrane proteome of T. foetus and identify proteins that are represented in different isolates of this protozoan. Additionally, we performed a bioinformatic analysis that revealed the antigenicity potential of some of those proteins. This analysis is the first study to identify common proteins at the plasma membrane of different T. foetus isolates that could be targets for alternative diagnostic or vaccine techniques in the future., (© 2024. The Author(s).)

Bovine trichomonosis (BT), a disease of the bovine urogenital tract, is caused by the protozoan Tritrichomonas foetus (Tf). Tf causes endometritis, infertility, and premature death of the embryo, which generates considerable economic losses. The proteins released can mediate fundamental interactions between the pathogen and the host, triggering factors associated with the symptomatology, immune evasion and pathogenesis characteristic of the species. However, little is known about the profile of the proteins released by Tf. In order to contribute to their knowledge, we performed an isolation protocol and a proteomic profiling of the supernatant (SN) content of six Tf isolates. A total of 662 proteins present in the SN of Tf were detected, out of which 121 were shared by the six isolates, while the remaining 541 were found in at least one of the isolates studied. The comparative analyses using the databases of Tf strain genome K revealed 32.9% of uncharacterized proteins. The bioinformatic analyses showed that the main molecular functions predicted were binding (47.9%) and catalytic activity (38.2%). Additionally, we performed immunodetection assays to evidence the antigenic potential of SN proteins. Interestingly, we observed great ability to detect SN proteins from all six isolates using serum from immunized mice and infected bulls. A complementary mass spectrometry assay allowed us to determine that the proteins that showed the strongest signal intensity in the immunoassays were Grp78 (A0A1J4IZS3) and Ap65 (A0A1J4JSR1). This work represents the first proteomic characterization of Tf SN proteins and their antigenic potential, which might be interesting for the future design of new diagnosis and treatment methods for BT., 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 © 2023 Elsevier B.V. All rights reserved.)

Psychostimulant use is a major comorbidity in people living with HIV, which was initially explained by them adopting risky behaviors that facilitate HIV transmission. However, the effects of drug use on the immune system might also influence this phenomenon. Psychostimulants act on peripheral immune cells even before they reach the central nervous system (CNS) and their effects on immunity are likely to influence HIV infection. Beyond their canonical activities, classic neurotransmitters and neuromodulators are expressed by peripheral immune cells (e.g., dopamine and enkephalins), which display immunomodulatory properties and could be influenced by psychostimulants. Immune receptors, like Toll-like receptors (TLRs) on microglia, are modulated by cocaine and amphetamine exposure. Since peripheral immunocytes also express TLRs, they may be similarly affected by psychostimulants. In this review, we will summarize how psychostimulants are currently thought to influence peripheral immunity, mainly focusing on catecholamines, enkephalins and TLR4, and shed light on how these drugs might affect HIV infection. We will try to shift from the classic CNS perspective and adopt a more holistic view, addressing the potential impact of psychostimulants on the peripheral immune system and how their systemic effects could influence HIV infection.

Objective: The aim of the present study was to determine the effect of supplementing pasture-finished steers with corn silage on the expression level of the calpain system proteins and beef tenderization., Methods: Thirty Braford steers grazing on summer pasture were used for the study. For 120 days fifteen animals were supplemented with corn silage at 1% of body weight per head per day (Suppl) whereas the remaining 15 steers only received pasture (Contr). Carcass and meat traits were evaluated and compared between groups. Gene expression and activities of proteases (calpain 1 and calpain 2) and inhibitor (calpastatin) were measured using real-time polymerase chain reaction and casein zymography., Results: Carcass and meat traits were significantly different between feeding systems. Supplemented steers showed higher hot carcass weight (p<0.01), fat content (p = 0.02), and Warner-Bratzler shear force (p = 0.03). Furthermore, the control group showed higher protease:inhibitor ratios, at mRNA (p = 0.01) and protein levels (p<0.10). Warner-Bratzler shear force and mRNA calpains:calpastatin ratio were associated in both feeding systems (p<0.01)., Conclusion: Based on the results obtained in the study, beef tenderness differences among finishing strategies could be modulated through differential expression of the calpain system proteins.

Background: Tritrichomonas foetus is the etiologic agent of the sexually transmitted disease Bovine Trichomonosis (BT). In Argentina, BT is endemic and represents a relevant health problem that causes reproductive inefficiency in cattle and large economic losses. Metronidazole is the drug of choice in the treatment of BT. Treatment has been associated with a temporary resolution of the clinical signs but is not able to control the disease. In recent years, the apparition of in vivo and in vitro aerobic and anaerobic resistance leading to ineffective treatments has been reported., Aims: Thus, the aim of the present study was to explore the susceptibility of six different isolates of T. foetus under aerobic (AC) and anaerobic (ANC) conditions., Results and Discussion: Six isolates of T. foetus were obtained from samples of preputial smegma of bovine origin. Values of minimum lethal concentration and minimum inhibitory concentration were higher than those observed in other works and represent current data in Argentina and provide information to establish new treatment protocols.

Bovine Trichomonosis, an endemic sexually transmitted disease in countries with extensive livestock and natural service, represents one of the most common causes of reproductive failure. 5-nitroimidazoles and their derivatives are used for its treatment, mainly metronidazole (Mz). The emergence of resistance mechanisms adopted by the parasites against the drug and failure of the treatments suggest the need to investigate susceptibility and obtain current values. The available information of in vitro susceptibility of these drugs comes from the use of a diversity of methodologies and criteria, especially observation of the mobility of the parasite under the optical microscope to evaluate its viability. These techniques are arduous and time consuming and lead to a subjective assessment dependent on the operator, the methodology used, and the morphology adopted by the protozoan. In this sense, flow cytometry has proven to be a fast and efficient method to evaluate viability in other protozoa. The aim of this study was to evaluate the in vitro susceptibility of six bovine isolates of Tritrichomonas foetus to Mz in aerobic (AC) and anaerobic (ANC) conditions by means of the calculation of the 50% inhibitory concentration (IC 50 ), by flow cytometry, and also to analyze minimum lethal concentration (MLC) by means of recovery tests post-treatment in vitro. IC 50 values ranged from 1.06 to 1.25 μM in ANC and from 1.44 to 3.03 μM in AC, these being the only ones reported at 48 h for these protozoa. With respect to MLC at 48 h, the results were from 3.67 to 7.35 μM in ANC, and from 7.35 to 14.7 μM for AC, where two isolates (Tf0 and Tf2) for AC and one (Tf2) for ANC showed higher values than those described in the literature. Flow cytometry has proven to be an effective, rapid and objective methodology and very useful in susceptibility tests. The data obtained through these tests allow us to describe the susceptibility exhibited by these protozoa, this being valuable information when establishing dosages in Mz treatments., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Intestinal and free-living protozoa, such as Giardia lamblia, express a dense coat of variant-specific surface proteins (VSPs) on trophozoites that protects the parasite inside the host's intestine. Here we show that VSPs not only are resistant to proteolytic digestion and extreme pH and temperatures but also stimulate host innate immune responses in a TLR-4 dependent manner. We show that these properties can be exploited to both protect and adjuvant vaccine antigens for oral administration. Chimeric Virus-like Particles (VLPs) decorated with VSPs and expressing model surface antigens, such as influenza virus hemagglutinin (HA) and neuraminidase (NA), are protected from degradation and activate antigen presenting cells in vitro. Orally administered VSP-pseudotyped VLPs, but not plain VLPs, generate robust immune responses that protect mice from influenza infection and HA-expressing tumors. This versatile vaccine platform has the attributes to meet the ultimate challenge of generating safe, stable and efficient oral vaccines.

During evolution, parasitic microorganisms have faced the challenges of adapting to different environments to colonize a variety of hosts. Giardia lamblia, a common cause of intestinal disease, has developed fascinating strategies to adapt both outside and inside its host's intestine, such as trophozoite differentiation into cyst and the switching of its major surface antigens. How gene expression is regulated during these adaptive processes remains undefined. Giardia lacks some typical eukaryotic features, like canonical transcription factors, linker histone H1, and complex promoter regions; suggesting that post-transcriptional and translational control of gene expression is essential for parasite survival. However, epigenetic factors may also play critical roles at the transcriptional level. Here, we describe the most common post-translational histone modifications; characterize enzymes involved in these reactions, and analyze their association with the Giardia's differentiation processes. We present evidence that NAD + -dependent and NAD + -independent histone deacetylases regulate encystation; however, a unique NAD + -independent histone deacetylase modulate antigenic switching. The rates of acetylation of H4K8 and H4K16 are critical for encystation, whereas a decrease in acetylation of H4K8 and methylation of H3K9 occur preferentially during antigenic variation. These results show the complexity of the mechanisms regulating gene expression in this minimalistic protozoan parasite., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Methionine is an amino acid susceptible to being oxidized to methionine sulfoxide (MetSO). The reduction of MetSO to methionine is catalyzed by methionine sulfoxide reductase (MSR), an enzyme present in almost all organisms. In trypanosomatids, the study of antioxidant systems has been mainly focused on the involvement of trypanothione, a specific redox component in these organisms. However, no information is available concerning their mechanisms for repairing oxidized proteins, which would be relevant for the survival of these pathogens in the various stages of their life cycle. We report the molecular cloning of three genes encoding a putative A-type MSR in trypanosomatids. The genes were expressed in Escherichia coli, and the corresponding recombinant proteins were purified and functionally characterized. The enzymes were specific for L-Met(S)SO reduction, using Trypanosoma cruzi tryparedoxin I as the reducing substrate. Each enzyme migrated in electrophoresis with a particular profile reflecting the differences they exhibit in superficial charge. The in vivo presence of the enzymes was evidenced by immunological detection in replicative stages of T. cruzi and Trypanosoma brucei. The results support the occurrence of a metabolic pathway in Trypanosoma spp. involved in the critical function of repairing oxidized macromolecules., (Copyright © 2010 Elsevier Inc. All rights reserved.)

Giardia lamblia is a human intestinal pathogen. Like many protozoan microorganisms, Giardia undergoes antigenic variation, a mechanism assumed to allow parasites to evade the host's immune response, producing chronic and/or recurrent infections. Recently, we found that the mechanism controlling variant-specific surface protein (VSP) switching in Giardia involves components of the RNA interference machinery and that disruption of this pathway generates trophozoites simultaneously expressing many VSPs. Here we use these altered trophozoites to determine the role of antigenic variation in a gerbil model of giardiasis. Our results show that either primary infection with trophozoites simultaneously expressing many VSPs or immunization with purified VSPs from the transgenic cells protects gerbils from subsequent Giardia infections. These results constitute, to our knowledge, the first experimental evidence that antigenic variation is essential for parasite survival within hosts and that artificial disruption of this mechanism might be useful in generating vaccines against major pathogens that show similar behavior.

Giardia lamblia is one of the most common causes of intestinal disease in humans. To adapt to environments both inside and outside of the host's small intestine, this protozoan parasite undergoes significant developmental changes during its life cycle. In this review, we analyze and discuss the most recent findings regarding the process of Giardia trophozoites differentiation into infective cysts as well as the mechanism of antigenic variation, which allows the parasite to cause chronic and recurrent infections in susceptible individuals.

Giardia lamblia is a medically important protozoan parasite with a basal position in the eukaryotic lineage and is an interesting model to explain the evolution of biochemical events in eukaryotic cells. G. lamblia trophozoites undergo significant changes in order to survive outside the intestine of their host by differentiating into infective cysts. In the present study, we characterize the previously identified Orf-C4 (G. lamblia open reading frame C4) gene, which is considered to be specific to G. lamblia. It encodes a 22 kDa protein that assembles into high-molecular-mass complexes during the entire life cycle of the parasite. ORF-C4 localizes to the cytoplasm of trophozoites and cysts, and forms large spherical aggregates when overexpressed. ORF-C4 overexpression and down-regulation do not affect trophozoite viability; however, differentiation into cysts is slightly delayed when the expression of ORF-C4 is down-regulated. In addition, ORF-C4 protein expression is modified under specific stress-inducing conditions. Neither orthologous proteins nor conserved domains are found in databases by conventional sequence analysis of the predicted protein. However, ORF-C4 contains a region which is similar structurally to the alpha-crystallin domain of sHsps (small heat-shock proteins). In the present study, we show the potential role of ORF-C4 as a small chaperone which is involved in the response to stress (including encystation) in G. lamblia.

Giardia lamblia (also called Giardia intestinalis) is one of the most common intestinal parasites of humans. To evade the host's immune response, Giardia undergoes antigenic variation-a process that allows the parasite to develop chronic and recurrent infections. From a repertoire of approximately 190 variant-specific surface protein (VSP)-coding genes, Giardia expresses only one VSP on the surface of each parasite at a particular time, but spontaneously switches to a different VSP by unknown mechanisms. Here we show that regulation of VSP expression involves a system comprising RNA-dependent RNA polymerase, Dicer and Argonaute, known components of the RNA interference machinery. Clones expressing a single surface antigen efficiently transcribe several VSP genes but only accumulate transcripts encoding the VSP to be expressed. Detection of antisense RNAs corresponding to the silenced VSP genes and small RNAs from the silenced but not for the expressed vsp implicate the RNA interference pathway in antigenic variation. Remarkably, silencing of Dicer and RNA-dependent RNA polymerase leads to a change from single to multiple VSP expression in individual parasites.

The components of the redox metabolism in Entamoeba histolytica have been recently revisited by Arias et al. (Free Radic. Biol. Med. 42:1496-1505; 2007), after the identification and characterization of a thioredoxin-linked system. The present work deals with studies performed for a better understanding of the localization and identification of different components of the redox machinery present in the parasite. The gene encoding for amoebic thioredoxin 8 was cloned and the recombinant protein typified as having properties similar to those of thioredoxin 41. The ability of these thioredoxins and the specific reductase to assemble a system utilizing NADPH to metabolize hydroperoxides in association with a peroxiredoxin has been kinetically characterized. The peroxiredoxin behaved as a typical 2 cysteine enzyme, exhibiting a ping-pong mechanism with hyperbolic saturation kinetics for thioredoxin 8 (K(m)=3.8 microM), thioredoxin 41 (K(m)=3.1 microM), and tert-butyl hydroperoxide (K(m) about 35 microM). Moreover, the tandem system involving thioredoxin reductase and either thioredoxin proved to be operative for reducing low molecular weight disulfides, including putative physiological substrates as cystine and oxidized trypanothione. Thioredoxin reductase and thioredoxin 41 (by association also the functional redox system) have been immunolocalized underlying the plasma membrane in Entamoeba histolytica cells. These findings suggest an important role for the metabolic pathway involving thioredoxin as a redox interchanger, which could be critical for the maintenance and virulence of the parasite when exposed to highly toxic reactive oxygen species.

Giardia lamblia undergoes antigenic variation, a process that might allow the parasite to evade the host's immune response and adapt to different environments. Here we show that Giardia muris, a related species that naturally infects rodents, possesses multiple variant-specific surface proteins (VSPs) and expresses VSPs on its surface, suggesting that it undergoes antigenic variation similar to that of G. lamblia.

A novel variant-specific surface protein (VSP) from Giardia was identified using the monoclonal antibody 9B10, raised against purified cyst walls. VSP9B10B is preferentially induced during encystation and expressed simultaneously with other VSPs on the surface of encysting trophozoites. These results support the hypothesis that encystation and antigenic variation are processes that are mechanistically related.