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Violence is the second most common cause of death in Panama; however, there is a surprising paucity of research addressing the dynamics of this social phenomenon, particularly from a neuropsychological perspective. Evidence suggests that intelligence and executive function (EF) are strongly associated with delinquency and criminal behavior (Moffitt, 1993; Morgan & Lilienfeld, 2000). In the present study, three groups of men convicted for criminal offenses such as intimate partner femicide (FPI; n=27), non-relational homicide (HNR; n=28) and non-violent offenses (DNV; n=29) were assessed. The following instruments were administered: verbal (WAIS-III) and non-verbal (TONI-2) intelligence tests, as well as various executive function tests (TMT, Stroop, and COWAT). The FPI group exhibited significant performance deficits in the Stroop test,, which suggests impairments in the speed of information processing in this group. Additionally, a profound deficit in verbal functioning across all groups emerged as the defining element in the cognitive profile of men serving sentences for criminal offenses. Verbal CI was significantly correlated with years of study, non-verbal CI, and most of the EF measures. These results provide preliminary evidence that supports the development of early intervention programs as a means to prevent violent behavior

The occurrence of esophagitis and ingluvitis caused by Salmonella Typhimurium in passerines seized from illegal wildlife trafficking is described. This illegal activity causes stress and leads to lowered immunity in the birds. Additionally, inadequate hygiene conditions predispose the birds to diseases such as salmonellosis. Few studies report the occurrence of Salmonella -induced lesions in the esophagus and crop of passerines; therefore, this study aimed to describe the disease in birds of the genus Sporophila , as well as to investigate the presence of the bacterium in the environment and determine the antimicrobial resistance profile of the isolated bacteria. Three birds of the genus Sporophila were necropsied. In the esophagus and crop, yellowish plaques corresponding to necrosis with bacterial aggregates were observed. Salmonella spp. was isolated from these lesions, with genus confirmation via MALDI-TOF mass spectrometry. Environmental samples were collected from the enclosures and cages where the animals were quarantined, and the same bacterium was isolated. In two birds, the serotype S. Typhimurium was identified. Antibiograms performed on the strains from the birds and the environment revealed resistance to antibiotics in the classes of penicillins, sulfonamides, aminoglycosides, monobactams, tetracyclines, and first and third-generation cephalosporins. To the authors' knowledge, this is the first report of this agent causing death in Sporophila due to esophagitis and ingluvitis. It is also the first report of salmonellosis in three species of passerines in Brazil. The study underscores the importance of understanding the pathogens circulating in wild animals, especially within the context of One Health.

Introduction: Ethion is an organophosphate used as an acaricide and insecticide, that is restricted worldwide. In Colombia, pesticide poisoning is the third most common cause of chemical intoxication. On 9 October 2022, an outbreak of ethion poisoning occurred in Pereira. The aim of this study was to describe the clinical and epidemiological characteristics of the outbreak., Methods: This is a descriptive study of an outbreak of organophosphate poisoning. The onset of symptoms occurred on 9 October 2022, following the consumption of empanadas. Information was collected on sociodemographic characteristics and clinical manifestations, as well as from paraclinical examinations. Data were obtained from clinical histories, field epidemiological investigations, and inspection visits. Food samples were collected for analysis by gas chromatography-mass spectrometry. Attack rates, proportions, and measures of central tendency, dispersion, and position were calculated., Results: The case definition was met by 37 individuals with a median age of 30 years; all presented with muscarinic symptoms, 29 patients presented with nicotinic symptoms, and 20 patients presented with neurological symptoms. Males were the most affected (57%), and the most common time of symptom onset was 10:00 am. Twenty-three patients (62%) required intensive care unit admission, of whom 14 (38%) required mechanical ventilation. No deaths were reported. Erythrocyte acetylcholinesterase activity was reduced in all patients. Ethion was detected in mass-prepared maize and empanadas at concentrations greater than 0.1 mg/kg. The consumption of empanadas was identified as the common source., Discussion: In Colombia, pesticide poisonings are the third most common type of poisoning caused by chemical substances reported to the National Health Institute through the National Public Health Surveillance System. In the present outbreak, ethion was in empanadas, likely due to contamination of cooking oil., Conclusions: We describe a large ethion-contaminated food poisoning outbreak reported in Colombia. The main symptoms were muscarinic, and the main treatment measures employed were atropine and respiratory support. Increased awareness of pesticide poisoning and training for food handlers are needed.

Background: The periparturient period in dairy cows is marked by immunosuppression which increases the likelihood of infectious disorders, particularly also mastitis. An in-depth understanding of peripartum leukocyte biology is vital for the implementation of highly successful post-partum disease prevention measures. Immune checkpoint molecules, such as programmed death 1 (PD-1) and cytotoxic T-lymphocyte antigen 4 (CTLA-4), are critical inhibitory receptors expressed on immune cells, particularly T cells, that drive immunosuppressive signaling pathways. However, the potential role of immune checkpoint molecules expression in T-cells on udder health has never been explored. Thus, the association between the occurrence of new postpartum intramammary infections (IMIs) and the expression of programmed cell death protein 1 (PD-1) and cytotoxic T-lymphocyte-associated antigen-4 (CTLA-4) on blood T-cells during the peripartum period was investigated., Results: In this study, the incidence of IMIs by any pathogen in early lactation was not associated with a higher expression of PD-1 and CTLA-4 in the periparturient period. However, the incidence of IMIs by major pathogens throughout the first month of lactation was significantly associated with higher expression of PD-1 at 14 days before calving (P = 0.03) and CTLA-4 at parturition (P = 0.03) by blood T-cells. Also, the expression of CTLA-4 at D0 (P = 0.012) by T-cells was associated with the occurrence of persistent IMIs during the first month of lactation., Conclusions: To our knowledge, this is the first report to investigate the expression of PD-1 and CTLA-4 by blood T-lymphocytes during the periparturient period in dairy cows and to explore their relationship with the incidence of new IMIs in the postpartum period. Thus, a comprehensive understanding of leukocyte biology during peripartum would appear to be a prerequisite for the identification of resilient dairy cows or targets innovative (immunological) non-antibiotic approaches in the transition period., (© 2024. The Author(s).)

Significance: The integrity of the intestinal barrier is gaining recognition as a significant contributor to various pathophysiological conditions, including inflammatory bowel disease, celiac disease, environmental enteric dysfunction (EED), and malnutrition. EED, for example, manifests as complex structural and functional changes in the small intestine leading to increased intestinal permeability, inflammation, and reduced absorption of nutrients. Despite the importance of gut function, current techniques to assess intestinal permeability (such as endoscopic biopsies or dual sugar assays) are either highly invasive, unreliable, and/or difficult to perform in certain patient populations (e.g., infants)., Aim: We present a portable, optical sensor based on transcutaneous fluorescence spectroscopy to assess gut function (in particular, intestinal permeability) in a fast and noninvasive manner., Approach: Participants receive an oral dose of a fluorescent contrast agent, and a wearable fiber-optic probe detects the permeation of the contrast agent from the gut into the blood stream by measuring the fluorescence intensity noninvasively at the fingertip. We characterized the performance of our compact optical sensor by comparing it against an existing benchtop spectroscopic system. In addition, we report results from a human study in healthy volunteers investigating the impact of skin tone and contrast agent dose on transcutaneous fluorescence signals., Results: The first study with eight healthy participants showed good correlation between our compact sensor and the existing benchtop spectroscopic system [correlation coefficient ( r ) > 0.919 , p < 0.001 ]. Further experiments in 14 healthy participants revealed an approximately linear relationship between the ingested contrast agent dose and the collected signal intensity. Finally, a parallel study on the impact of different skin tones showed no significant differences in signal levels between participants with different skin tones ( p > 0.05 )., Conclusions: In this paper, we demonstrate the potential of our compact transcutaneous fluorescence sensor for noninvasive monitoring of intestinal health., (© 2024 The Authors.)

Guinea pigs have historically been used as a food source and are also an important model for studying the human intestines. Fasting is the act of temporarily stopping the intake of food. This process can alter the microbiota of various animals. This study is the first to investigate the impact of fasting on the cecum microbiome of three guinea pig breeds. We investigated the impact of fasting on the microbiome population structure in the cecum of three guinea pig breeds. This was done by sequencing and analyzing the V4 hypervariable region of the 16S rRNA gene in bacterial communities found in cecum mucosa samples. To achieve this, we established two treatment groups (fasting and fed), for each of the three guinea pig breeds: Andina, Inti, and Peru. The study involved twenty-eight guinea pigs, which were divided into the following groups: Andina-fed (five), Andina-fasting (five), Inti-fed (four), Inti-fasting (five), Peru-fed (five), and Peru-fasting (four). The results indicated a significant difference in beta diversity between the treatment groups for the Peru breed ( P -value = 0.049), but not for the treatment groups of the Andina and Inti breeds. The dominant phyla across all groups were Firmicutes and Bacteroidetes. We observed variations in the abundance of different taxa in the cecum microbiota when comparing the treatment groups for each breed. Additionally, there was a higher number of unique taxa observed in the fasting groups compared to the fed groups. We discovered that the genus Victivallis was the only one present in all fasting groups across all breeds. Despite the findings, the resilience of the gut microbiome was not challenged in all three breeds, which can lead to disruptive changes that may affect the overall maintenance of the cecum microbiome. Based on the observed differences in the treatment groups of the Peru breed, it can be suggested that fasting has a greater impact on this particular breed., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Frias, Murga Valderrama, Flores Durand, Cornejo, Romani, Bardales, Segura, Polveiro, Vieira, Ramos Sanchez, Lopez Lapa and Maicelo Quintana.)

Bovine leukemia virus (BLV) is caused by a deltaretrovirus and has been associated with immunosuppression as well as comorbidities such as bovine mastitis, the costliest disease in the dairy sector. However, no previous study has explored at the synergistic immunosuppressive effect of the peripartum period with an immunosuppressive viral disease such as BLV. Thus, our study explored the effect of BLV infection in the periparturient period on the expression of PD-1 and CTLA-4 in blood T lymphocytes, and the impact of BLV infection on the rate of new intramammary infections during the early lactation. Here, we found that BLV-infected dairy cows always had a statistically significant higher expression of CTLA-4 and PD-1 in blood T cells. Furthermore, our findings indicated that BLV infection prolongs immunosuppression in dairy cows during the periparturient period by sustaining higher expression of immunological checkpoints in T cells. In addition, BLV-infected dairy cows have a higher rate of new intramammary infections during early lactation. Thus, our study provides new insights of the immunosuppressive effect of BLV on the most critical period of the cows' life with marked detrimental effect on protective T-cell immunity and comorbidities, such as bovine mastitis., 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.)

The discovery and tracking of antimicrobial resistance genes are essential for understanding the evolution of bacterial resistance and restraining its dispersion. Mammaliicoccus sciuri (formerly Staphylococcus sciuri) is the most probable evolutionary repository of the mecA gene, that later disseminated to S. aureus. In this study, we describe the first double mecA/mecC homologue-positive non-aureus staphylococci and mammaliicocci (NASM) from the American continent, also representing the first report of mecC-positive NASM in Brazil. Two clonally related methicillin-resistant M. sciuri strains co-carrying mecA and mecC genes were isolated from the teat skin swab and milk sample collected from an ewe's left udder half. Both M. sciuri strains belonged to the sequence type (ST) 71. Besides mecA and mecC genes, the M. sciuri strains carried broad resistomes for clinically important antimicrobial agents, including β-lactams, tetracyclines, lincosamide, streptogramin, streptomycin, and aminoglycosides. Virulome analysis showed the presence of the clumping factor B (clfB), ATP-dependent protease ClpP (ClpP) and serine-aspartate repeat proteins (sdrC and sdrE) virulence-associated genes. Phylogenomic analysis revealed that these M. sciuri strains are part of a globally disseminated branch, associated with farm and companion animals and even with food. Our findings suggest that M. sciuri is likely to emerge as a pathogen of global interest, carrying a broad repertoire of antimicrobial resistance genes with a remarkable co-presence of mecA and mecC genes. Finally, we strongly encourage to monitor M. sciuri under the One Health umbrella since this bacterial species is spreading at the human-animal-environment interface., 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.)

We examined whether distinct staphylococcal and mammaliicoccal species and strains trigger B- and T-lymphocyte proliferation and interleukin (IL)-17A and interferon (IFN)-γ production by peripheral blood mononuclear cells in nulliparous, primiparous, and multiparous dairy cows. Flow cytometry was used to measure lymphocyte proliferation with the Ki67 antibody, and specific monoclonal antibodies were used to identify CD3, CD4, and CD8 T lymphocyte and CD21 B lymphocyte populations. The supernatant of the peripheral blood mononuclear cell culture was used to measure IL-17A and IFN-γ production. Two distinct, inactivated strains of bovine-associated Staphylococcus aureus [one causing a persistent intramammary infection (IMI) and the other from the nose], 2 inactivated Staphylococcus chromogenes strains [one causing an IMI and the other from a teat apex), as well as an inactivated Mammaliicoccus fleurettii strain originating from sawdust from a dairy farm, and the mitogens concanavalin A and phytohemagglutinin M-form (both specifically to measure lymphocyte proliferation) were studied. In contrast to the "commensal" Staph. aureus strain originating from the nose, the Staph. aureus strain causing a persistent IMI triggered proliferation of CD4 + and CD8 + subpopulations of T lymphocytes. The M. fleurettii strain and the 2 Staph. chromogenes strains had no effect on T- or B-cell proliferation. Furthermore, both Staph. aureus and Staph. chromogenes strains causing persistent IMI significantly increased IL-17A and IFN-γ production by peripheral blood mononuclear cells. Overall, multiparous cows tended to have a higher B-lymphocyte and a lower T-lymphocyte proliferative response than primiparous and nulliparous cows. Peripheral blood mononuclear cells of multiparous cows also produced significantly more IL-17A and IFN-γ. In contrast to concanavalin A, phytohemagglutinin M-form selectively stimulated T-cell proliferation., (The Authors. Published by Elsevier Inc. and Fass Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

Perovskite solar cells (PSCs) are an evolving photovoltaic field with the potential to disrupt the established silicon solar cell market. However, the presence of many transport barriers and defect trap states at the interfaces and grain boundaries has negative effects on PSCs; it decreases their efficiency and stability. The purpose of this work was to investigate the effects on efficiency and stability achieved by quaternary theophylline additives in MAPbI 3 PSCs with the structure FTO/TiO 2 /perovskite/spiro-OMeTAD/Ag. The X-ray photoelectron spectroscopy (XPS) and theoretical calculation strategies were applied to study the additive's interaction in the layer. The tetrafluoroborinated additive results in an increase in device current density ( J SC ) (23.99 mA cm -1 ), fill factor (FF) (65.7%), and open-circuit voltage ( V OC ) (0.95 V), leading to significant improvement of the power conversion efficiency (PCE) to 15.04% compared to control devices (13.6%). Notably, films exposed to controlled humidity of 30% using the tetrafluoroborinated additive maintained their stability for more than 600 hours (h), while the control films were stable for less than 240 hours (h)., Competing Interests: There are no conflicts to declare., (This journal is © The Royal Society of Chemistry.)

Background: Staphylococcus aureus is one of the most frequently major mastitis pathogens that cause clinical and subclinical mastitis worldwide. Current antimicrobial treatments are usually ineffective, and the commercially available vaccines lack proven effectiveness. The immunological response elicited by the recombinant S. aureus -cure-associated proteins phosphoglycerate kinase (PGK), enolase (ENO), and elongation factor-G (EF-G) in combination with the granulocyte-macrophage colony-stimulating factor (GM-CSF) DNA vaccination was studied in this work., Methods: Here, twenty-three C57BL/6 mice were divided into four groups and vaccinated with: G1: none (control); G2: GM-CSF DNA plasmid DNA vaccine; G3: the combination of EF-G+ENO+PGK; and G4: the combinations of EF-G+ENO+PGK proteins plus GM-CSF plasmid DNA vaccine. After 44 days, spleen cells were collected for immunophenotyping and lymphocyte proliferation evaluation by flow cytometry upon S. aureus stimulus., Results: Immunization with the three S. aureus recombinant proteins alone resulted in a higher percentage of IL-17A + cells among CD8 + T central memory cells, as well as the highest intensity of IL-17A production by overall lymphocytes indicating that the contribution of the combined lymphocyte populations is crucial to sustaining a type 3 cell immunity environment., Conclusion: The immunization with three S. aureus- cure-associated recombinant proteins triggered type 3 immunity, which is a highly interesting path to pursue an effective bovine S. aureus mastitis vaccine.

Staphylococcus aureus mastitis constitutes a serious threat to dairy cows. The reasons why available vaccines are not fully effective remain poorly understood; thus, in the present study, we investigated CD4 + and CD8 + T lymphocyte proliferation in dairy cows vaccinated with a polyvalent mastitis vaccine that had distinct precedent Staphylococcus aureus mastitis. We studied 17 S. aureus-infected dairy cows (11 vaccinated and six unvaccinated) and eight vaccinated healthy dairy cows with no previous S. aureus mastitis infections. Flow cytometry was used to assess lymphocyte proliferation using an anti-Ki67 antibody, and monoclonal antibodies were used to identify T cell subsets. S. aureus-infected cows exhibited reduced overall lymphocyte proliferation, including CD4 + T lymphocyte proliferation, and memory lymphocyte proliferation in response to S. aureus isolate stimulus. Immunization did not influence the expansion of blood lymphocyte populations. Furthermore, CD8 + T cells, memory CD8 + T lymphocytes, and effector memory CD8 + T lymphocytes displayed reduced proliferation 21 days after the third vaccine dose compared with before vaccination at time zero. The present data demonstrates an overall negative regulation of the T-cell response suggesting its detrimental impact leading to the persistence of S. aureus intramammary infections. Furthermore, the lack of vaccination effect on T-cell mediated immunity (e.g., proliferation) may be related to poor vaccine efficacy., Competing Interests: Declaration of interest None., (Copyright © 2022 Elsevier B.V. All rights reserved.)

The present study sought to evaluate the expression of PD-1 and CTLA-4 in blood T lymphocytes during the periparturient period and their relationship with uterine health in dairy cows, as determined by endometrial cytology and serum concentrations of β-hydroxybutyrate (BHB) and non-esterified fatty acids (NEFAs), which are indicators of a negative energy balance. The second objective of this study was to investigate whether the expression of PD-1 and CTLA-4 in T lymphocytes is associated with the serum acute phase-protein haptoglobin concentration during the periparturient period. To address these objectives, 26 clinically healthy dairy cows were used. Peripheral blood was collected 14 days prepartum (T-14), at calving (T0), and 30 days postpartum (T30) to measure the expression of PD-1 and CTLA-4 in blood T lymphocytes by flow cytometry. In addition, we collected blood at T0, 10 days after parturition (T10), and T30 to obtain serum and determine the serum concentrations of NEFA, BHB, and Hp. Endometrial cytology was performed at T10, 20 days after parturition (T20), and T30. In the present study, we observed higher expression of CTLA-4 and PD-1 in T lymphocytes at parturition and in the prepartum period, which could indicate a relationship between these immune checkpoints and immunological tolerance during gestation in dairy cattle. In addition, a negative association between the expression of these immune checkpoints prepartum or at parturition and endometrial cytology at T20 and T30 was observed, indicating the negative implications of these immune response regulators in susceptibility to infections. This finding was further corroborated by the relationship between the serum concentration of haptoglobin and the expression of CTLA-4 and PD-1 by T lymphocytes. However, we did not observe a relationship between the indicators of negative energy balance, evaluated by the serum concentrations of BHB and NEFA, and the expression of the immune checkpoint markers studied. Thus, our findings represent an initial step that paves the way for the development of new therapeutic alternatives directed by the host with the objective of increasing the resistance of dairy cattle to infections in this critical period of life., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Souza, Lima, Ordonho, Oliveira, Rodrigues, Moura, Lima, Blagitz, Ramos Sanchez, Medeiros, Souza and Fernandes.)

Visceral leishmaniasis caused by Leishmania (Leishmania) infantum in Latin America progress with hepatosplenomegaly, pancytopenia, hypergammaglobulinemia, and weight loss and maybe lethal mainly in untreated cases. miRNAs are important regulators of immune and inflammatory gene expression, but their mechanisms of action and their relationship to pathogenesis in leishmaniasis are not well understood. In the present study, we sought to quantify changes in miRNAs associated with immune and inflammatory pathways using the L. (L.) infantum promastigote infected- human monocytic THP-1 cell model and plasma from patients with visceral leishmaniasis. We identified differentially expressed miRNAs in infected THP-1 cells compared with non-infected cells using qPCR arrays. These miRNAs were submitted to in silico analysis, revealing targets within functional pathways associated with TGF-β, chemokines, glucose metabolism, inflammation, apoptosis, and cell signaling. In parallel, we identified differentially expressed miRNAs in active visceral leishmaniasis patient plasma compared with endemic healthy controls. In silico analysis of these data indicated different predicted targets within the TGF-β, TLR4, IGF-I, chemokine, and HIF1α pathways. Only a small number of miRNAs were commonly identified in these two datasets, notably with miR-548d-3p being up-regulated in both conditions. To evaluate the potential biological role of miR-548d-3p, we transiently transfected a miR-548d-3p inhibitor into L. (L.) infantum infected-THP-1 cells, finding that inhibition of miR-548d-3p enhanced parasite growth, likely mediated through reduced levels of MCP-1/CCL2 and nitric oxide production. Further work will be required to determine how miR-548d-3p plays a role in vivo and whether it serves as a potential biomarker of progressive leishmaniasis., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Ramos-Sanchez, Reis, Souza, Muxel, Santos, Lagos, Pereira, Brito, Kaye, Floeter-Winter and Goto.)

Staphylococcus aureus mastitis remains a major challenge for dairy farming. Here, 24 mice were immunized and divided into four groups: G1: control; G2: Granulocyte Macrophage Colony-Stimulating Factor (GM-CSF) DNA vaccine; G3: F0F1 ATP synthase subunit α (SAS), succinyl-diaminopimelate (SDD), and cysteinyl-tRNA synthetase (CTS) recombinant proteins; and G4: SAS+SDD+CTS plus GM-CSF DNA vaccine. The lymphocyte subpopulations, and the intracellular interleukin-17A (IL-17A) and interferon-γ production in the draining lymph node cells were immunophenotyped by flow cytometry. The immunophenotyping and lymphocyte proliferation was determined in spleen cells cultured with and without S. aureus stimulus. Immunization with S. aureus recombinant proteins generated memory cells in draining lymph nodes. Immunization with the three recombinant proteins plus GM-CSF DNA led to an increase in the percentage of IL-17A + cells among overall CD44 + (memory), T CD4 + , CD4 + T CD44 + CD27 - , γδ TCR, γδ TCR + CD44 + CD27 + , and TCRVγ4 + cells. Vaccination with S. aureus recombinant proteins associated with GM-CSF DNA vaccine downregulated T H 2 immunity. Immunization with the three recombinant proteins plus the GM-CSF DNA led to a proliferation of overall memory T, CD4 + , and CD4 + TEM cells upon S. aureus stimulus. This approach fostered type 3 immunity, suggesting the development of a protective immune response against S. aureus .

The implications of bovine leukemia virus (BLV) on innate and adaptive immune responses have been widely investigated; however, the effects of BLV on mammary gland immunity require further investigation. The present study investigated the viability, phagocytic capacity, and intracellular production of reactive oxygen and nitrogen species (RONS) by macrophages in milk samples from dairy cows naturally infected with BLV with or without persistent lymphocytosis (PL). No effect of BLV infection in the overall number of macrophages per milliliter and in the percentage of viable macrophages among overall milk viable cells was found. Furthermore, BLV-infected dairy cows had a higher frequency of viable milk macrophages, while healthy animals had a tendency toward a higher percentage of apoptotic milk macrophages. The percentage of milk macrophages that phagocytosed Staphylococcus aureus in seronegative animals was higher than that in BLV-infected dairy cows. No effect of BLV infection on the intracellular RONS production and the intensity of phagocytosis by milk macrophages was observed. Thus, this study provides new insights into the implications of BLV infections in the bovine mammary gland., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Lima, Blagitz, Batista, Alves, Fernandes, Ramos Sanchez, Frias Torres, Diniz, Silva, Della Libera and de Souza.)

American Tegumentary Leishmaniasis (ATL) is an endemic disease in Latin America, mainly caused in Brazil by Leishmania (Viannia) braziliensis . Clinical manifestations vary from mild, localized cutaneous leishmaniasis (CL) to aggressive mucosal disease. The host immune response strongly determines the outcome of infection and pattern of disease. However, the pathogenesis of ATL is not well understood, and host microRNAs (miRNAs) may have a role in this context. In the present study, miRNAs were quantified using qPCR arrays in human monocytic THP-1 cells infected in vitro with L. (V.) braziliensis promastigotes and in plasma from patients with ATL, focusing on inflammatory response-specific miRNAs. Patients with active or self-healed cutaneous leishmaniasis patients, with confirmed parasitological or immunological diagnosis, were compared with healthy controls. Computational target prediction of significantly-altered miRNAs from in vitro L. (V.) braziliensis -infected THP-1 cells revealed predicted targets involved in diverse pathways, including chemokine signaling, inflammatory, cellular proliferation, and tissue repair processes. In plasma, we observed distinct miRNA expression in patients with self-healed and active lesions compared with healthy controls. Some miRNAs dysregulated during THP-1 in vitro infection were also found in plasma from self-healed patients, including miR-548d-3p, which was upregulated in infected THP-1 cells and in plasma from self-healed patients. As miR-548d-3p was predicted to target the chemokine pathway and inflammation is a central to the pathogenesis of ATL, we evaluated the effect of transient transfection of a miR-548d-3p inhibitor on L. (V.) braziliensis infected-THP-1 cells. Inhibition of miR-548d-3p reduced parasite growth early after infection and increased production of MCP1/CCL2, RANTES/CCL5, and IP10/CXCL10. In plasma of self-healed patients, MCP1/CCL2, RANTES/CCL5, and IL-8/CXCL8 concentrations were significantly decreased and MIG/CXCL9 and IP-10/CXCL10 increased compared to patients with active disease. These data suggest that by modulating miRNAs, L. (V.) braziliensis may interfere with chemokine production and hence the inflammatory processes underpinning lesion resolution. Our data suggest miR-548d-3p could be further evaluated as a prognostic marker for ATL and/or as a host-directed therapeutic target., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Souza, Ramos-Sanchez, Muxel, Lagos, Reis, Pereira, Brito, Zampieri, Kaye, Floeter-Winter and Goto.)

Leishmaniases are diseases caused by several Leishmania species, and many factors contribute to the development of the infection. Because the adaptive immune response does not fully explain the outcome of Leishmania infection and considering that the initial events are crucial in the establishment of the infection, we investigated one of the growth factors, the insulin-like growth factor-I (IGF-I), found in circulation and produced by different cells including macrophages and present in the skin where the parasite is inoculated. Here, we review the role of IGF-I in leishmaniasis experimental models and human patients. IGF-I induces the growth of different Leishmania species in vitro and alters the disease outcome increasing the parasite load and lesion size, especially in L. major- and L. amazonensis- infected mouse leishmaniasis. IGF-I affects the parasite interacting with the IGF-I receptor present on Leishmania . During Leishmania -macrophage interaction, IGF-I acts on the arginine metabolic pathway, resulting in polyamine production both in macrophages and Leishmania . IGF-I and cytokines interact with reciprocal influences on their expression. IL-4 is a hallmark of susceptibility to L. major in murine leishmaniasis, but we observed that IGF-I operates astoundingly as an effector element of the IL-4. Approaching human leishmaniasis, patients with mucosal, disseminated, and visceral diseases presented surprisingly low IGF-I serum levels, suggesting diverse effects than parasite growth. We observed that low IGF-I levels might contribute to the inflammatory response persistence and delayed lesion healing in human cutaneous leishmaniasis and the anemia development in visceral leishmaniasis. We must highlight the complexity of infection revealed depending on the Leishmania species and the parasite's developmental stages. Because IGF-I exerts pleiotropic effects on the biology of interaction and disease pathogenesis, IGF-I turns up as an attractive tool to explore biological and pathogenic processes underlying infection development. IGF-I pleiotropic effects open further the possibility of approaching IGF-I as a therapeutical target., Competing Interests: The authors declare no conflicts of interest., (Copyright © 2021 Luiza C. Reis et al.)

Enucleated cells or cytoplasts (cells whose nucleus is removed in vitro ) represent an unexplored biological model for intracellular infection studies due to the abrupt interruption of nuclear processing and new RNA synthesis by the host cell in response to pathogen entry. Using enucleated fibroblasts hosting the protozoan parasite Leishmania amazonensis , we demonstrate that parasite multiplication and biogenesis of large parasitophorous vacuoles in which parasites multiply are independent of the host cell nucleus. Dual RNA sequencing of both host cytoplast and intracellular parasite transcripts identified host transcripts that are more preserved or degraded upon interaction with parasites and also parasite genes that are differentially expressed when hosted by nucleated or enucleated cells. Cytoplasts are suitable host cells, which persist in culture for more than 72 h and display functional enrichment of transcripts related to mitochondrial functions and mRNA translation. Crosstalk between nucleated host de novo gene expression in response to intracellular parasitism and the parasite gene expression to counteract or benefit from these host responses induces a parasite transcriptional profile favoring parasite multiplication and aerobic respiration, and a host-parasite transcriptional landscape enriched in host cell metabolic functions related to NAD, fatty acid, and glycolytic metabolism. Conversely, interruption of host nucleus-parasite cross talk by infection of enucleated cells generates a host-parasite transcriptional landscape in which cytoplast transcripts are enriched in phagolysosome-related pathway, prosurvival, and SerpinB-mediated immunomodulation. In addition, predictive in silico analyses indicated that parasite transcript products secreted within cytoplasts interact with host transcript products conserving the host V-ATPase proton translocation function and glutamine/proline metabolism. The collective evidence indicates parasite-mediated control of host cell transcripts half-life that is beneficial to parasite intracellular multiplication and escape from host immune responses. These findings will contribute to improved drug targeting and serve as database for L. amazonensis -host cell interactions., (Copyright © 2020 American Society for Microbiology.)

Prediction parameters of possible outcomes of canine leishmaniasis (CanL) therapy might help with therapeutic decisions and animal health care. Here, we aimed to develop a diagnostic method with predictive value by analyzing two groups of dogs with CanL, those that exhibited a decrease in parasite load upon antiparasitic treatment (group: responders) and those that maintained high parasite load despite the treatment (group: non-responders). The parameters analyzed were parasitic load determined by q-PCR, hemogram, serum biochemistry and immune system-related gene expression signature. A mathematical model was applied to the analysis of these parameters to predict how efficient their response to therapy would be. Responder dogs restored hematological and biochemical parameters to the reference values and exhibited a Th1 cell activation profile with a linear tendency to reach mild clinical alteration stages. Differently, non-responders developed a mixed Th1/Th2 response and exhibited markers of liver and kidney injury. Erythrocyte counts and serum phosphorus were identified as predictive markers of therapeutic response at an early period of assessment of CanL. The results presented in this study are highly encouraging and may represent a new paradigm for future assistance to clinicians to interfere precociously in the therapeutic approach, with a more precise definition in the patient's prognosis., Competing Interests: The authors declare no conflict of interest. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript, or in the decision to publish the results.

Staphylococcus aureus is one of the pathogens most frequently isolated from cases of mastitis worldwide. To decrease the effect of S. aureus mastitis in dairy farming, alternative strategies for controlling mastitis are needed that depend on a better knowledge of cow-to-cow variations in S. aureus antibody production. The present study sought to explore the diversity of S. aureus antibodies produced by dairy cows with a distinct mastitis history and vaccinated with a polyvalent mastitis vaccine. We obtained protein extracts from S. aureus isolates derived from persistent subclinical mastitis. Proteins were fractionated using 2-dimensional gel electrophoresis and Western blotting. Then, Western blotting membranes were exposed to sera from 24 dairy cows that had been divided into the following groups: vaccinated dairy cows that were infected with S. aureus, further subdivided according to whether they (a) remained infected by S. aureus or (b) recovered from the intramammary infection; unvaccinated dairy cows infected with S. aureus; and vaccinated healthy dairy cows with no history of S. aureus mastitis. Proteins found to be reactive by Western blot were identified by mass spectrometry (MALDI/TOF-TOF). Our most important finding was that F 0 F 1 ATP synthase subunit α, succinyl-diaminopimelate desuccinylase, and cysteinyl-tRNA synthetase were potential candidate proteins for the prevention of S. aureus mastitis. This study strengthens the notion that variations among animals should not be ignored and shows that the heterogeneity of antibody production against anti-staphylococcal antigens in animals may enable the identification of new immunotherapy targets., (Copyright © 2020 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.)

V-ATPases are part of the membrane components of pathogen-containing vacuoles, although their function in intracellular infection remains elusive. In addition to organelle acidification, V-ATPases are alternatively implicated in membrane fusion and anti-inflammatory functions controlled by ATP6V0d2, the d subunit variant of the V-ATPase complex. Therefore, we evaluated the role of ATP6V0d2 in the biogenesis of pathogen-containing vacuoles using ATP6V0d2 knock-down macrophages infected with the protozoan parasite Leishmania amazonensis. These parasites survive within IFNγ/LPS-activated inflammatory macrophages, multiplying in large/fusogenic parasitophorous vacuoles (PVs) and inducing ATP6V0d2 upregulation. ATP6V0d2 knock-down decreased macrophage cholesterol levels and inhibited PV enlargement without interfering with parasite multiplication. However, parasites required ATP6V0d2 to resist the influx of oxidized low-density lipoprotein (ox-LDL)-derived cholesterol, which restored PV enlargement in ATP6V0d2 knock-down macrophages by replenishing macrophage cholesterol pools. Thus, we reveal parasite-mediated subversion of host V-ATPase function toward cholesterol retention, which is required for establishing an inflammation-resistant intracellular parasite niche., Competing Interests: The authors have declared that no competing interests exist.

Certain cytokines modulate the expression of insulin-like growth factor- (IGF-) I. Since IL-4 and IGF-I promote growth of the protozoan Leishmania major , we here addressed their interaction in downregulating the expression of Igf-I mRNA using small interfering RNA (siRNA) in Leishmania major -infected macrophages. Parasitism was decreased in the siRNA-treated cells compared with the nontreated cells, reversed by the addition of recombinant IGF-I (rIGF-I). In IL-4-stimulated macrophages, parasitism and the Igf-I mRNA amount were increased, and the effects were nullified upon siRNA transfection. IGF-I downregulation inhibited both parasite and macrophage arginase activation even in IL-4-stimulated cells. Searching for intracellular signaling components shared by IL-4 and IGF-I, upon siRNA transfection, phosphorylated p44, p38, and Akt proteins were decreased, affecting the phosphatidylinositol-3-kinase (PI3K)/Akt pathway. In L. major -infected C57BL6-resistant mice, the preincubation of the parasite with rIGF-I changed the infection profile to be similar to that of susceptible mice. We conclude that IGF-I constitutes an effector element of IL-4 involving the PI3K/Akt pathway during L. major infection.

The present study aimed to validate the use of R-phycoerythrin (R-PE)-labeled Mannheimia haemolytica to simultaneously stimulate phagocytosis and intracellular production of reactive oxygen species (ROS) by blood phagocytes in bronchoalveolar lavage (BAL) fluid. Initially, R-PE-labeled M. haemolytica was inactivated using a water bath at 60 °C for 60 min. Afterwards, R-PE labelling of bacteria was confirmed by flow cytometry. The geometric mean fluorescence intensity of R-PE-labeled bacteria (FL2 detector, 585 ± 42 nm) was analyzed by flow cytometry and was 41.5-fold higher than the respective unlabeled controls, confirming the success of bacterial conjugation to R-PE. Phagocytosis and intracellular production of ROS by blood neutrophils and monocytes, and by BAL CD14 + macrophages, in 12 healthy 6-month-old male calves were then performed using R-PE-labeled bacteria and 2',7'-dichlorofluorescein diacetate (DCFH-DA) as probes. Confocal microscopy was used to confirm phagocytosis of R-PE-labeled M. haemolytica by phagocytes and to exclude erroneous measurements of bacteria adhering to the leukocyte membrane. The present study showed that there is no difference in the ROS production without stimulus and in the presence of M. haemolytica by peripheral blood neutrophils and monocytes, in contrast to the increased ROS production by local alveolar macrophages upon stimulation by M. haemolytica. This emphasizes the importance of alveolar macrophages in the maintenance of homeostasis and health of the respiratory system, which can be supported during the inflammatory process by the rapid recruitment of neutrophils with high microbicidal and phagocytic capacity. The method described here provides an easy and feasible tool to measure phagocytosis and intracellular ROS production by phagocytes, especially when commonly used probes for intracellular ROS production were used, such as DCFH-DA and dihydrorhodamine 123., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Background: Transfusion-transmitted malaria due to asymptomatic Plasmodium infections is a challenge for blood banks. There is a lack of data on the prevalence of asymptomatic infected blood donors and the incidence of transfusion-transmitted malaria in low endemicity areas worldwide. We estimated the frequency of blood donors harbouring Plasmodium in an area in which asymptomatic infections have been reported., Material and Methods: To estimate the frequency of blood donors harbouring Plasmodium we used microscopy and molecular tools. Serological tests were applied to measure the exposure of candidates to Plasmodium antigens. Venous blood was collected from 91 candidates attending the "Pró-Sangue" Blood Centre Foundation in São Paulo, who lived in the municipality of Juquitiba, São Paulo, Brazil, where sporadic autochthonous cases of malaria have been described. Blood samples were used for parasitological, molecular and serological studies., Results: Among the 91 samples examined, rare Plasmodium forms were observed in two donors. Genus real-time polymerase chain reaction analysis demonstrated Plasmodium amplification in three candidates and species-specific nested polymerase chain reaction identified P. malariae in two. ELISA-IgG was reactive in 42.9% of samples for P. vivax (Pv-MSP1 19 ) and in 6.6% for P. falciparum (Pf-Zw). ELISA-IgM was reactive in 2.2% of samples for P. vivax and in 4.4% for P. falciparum. An indirect immunofluorescence assay was reactive for P. malariae in 15.4% of cases., Discussion: Reservoirs of Plasmodium represent a challenge for blood banks, since studies have shown that high levels of submicroscopic infections can occur in low transmission areas. The risk of transfusion-transmitted malaria presented here points to the need to conduct molecular investigations of candidate donors with any positive malarial antibody test.

Background: Tubercidin (TUB) is a toxic adenosine analog with potential antiparasitic activity against Leishmania, with mechanism of action and resistance that are not completely understood. For understanding the mechanisms of action and identifying the potential metabolic pathways affected by this drug, we employed in this study an overexpression/selection approach using TUB for the identification of potential targets, as well as, drug resistance genes in L. major. Although, TUB is toxic to the mammalian host, these findings can provide evidences for a rational drug design based on purine pathway against leishmaniasis., Methodology/principal Findings: After transfection of a cosmid genomic library into L. major Friedlin (LmjF) parasites and application of the overexpression/selection method, we identified two cosmids (cosTUB1 and cosTU2) containing two different loci capable of conferring significant levels of TUB resistance. In the cosTUB1 contained a gene encoding NUPM1-like protein, which has been previously described as associated with TUB resistance in L. amazonensis. In the cosTUB2 we identified and characterized a gene encoding a 63 kDa protein that we denoted as tubercidin-resistance protein (TRP). Functional analysis revealed that the transfectants were less susceptible to TUB than LmjF parasites or those transfected with the control vector. In addition, the trp mRNA and protein levels in cosTUB2 transfectants were higher than LmjF. TRP immunolocalization revealed that it was co-localized to the endoplasmic reticulum (ER), a cellular compartment with many functions. In silico predictions indicated that TRP contains only a hypothetical transmembrane domain. Thus, it is likely that TRP is a lumen protein involved in multidrug efflux transport that may be involved in the purine metabolic pathway., Conclusions/significance: This study demonstrated for the first time that TRP is associated with TUB resistance in Leishmania. The next challenge is to determine how TRP mediates TUB resistance and whether purine metabolism is affected by this protein in the parasite. Finally, these findings may be helpful for the development of alternative anti-leishmanial drugs that target purine pathway., Competing Interests: The authors have declared that no competing interests exist.