Membrane fusion is essential for entry of the biomedically-important paramyxoviruses into their host cells (viral-cell fusion), and for syncytia formation (cell-cell fusion), often induced by paramyxoviral infections [e.g. those of the deadly Nipah virus (NiV)]. For most paramyxoviruses, membrane fusion requires two viral glycoproteins. Upon receptor binding, the attachment glycoprotein (HN/H/G) triggers the fusion glycoprotein (F) to undergo conformational changes that merge viral and/or cell membranes. However, a significant knowledge gap remains on how HN/H/G couples cell receptor binding to F-triggering. Via interdisciplinary approaches we report the first comprehensive mechanism of NiV membrane fusion triggering, involving three spatiotemporally sequential cell receptor-induced conformational steps in NiV-G: two in the head and one in the stalk. Interestingly, a headless NiV-G mutant was able to trigger NiV-F, and the two head conformational steps were required for the exposure of the stalk domain. Moreover, the headless NiV-G prematurely triggered NiV-F on virions, indicating that the NiV-G head prevents premature triggering of NiV-F on virions by concealing a F-triggering stalk domain until the correct time and place: receptor-binding. Based on these and recent paramyxovirus findings, we present a comprehensive and fundamentally conserved mechanistic model of paramyxovirus membrane fusion triggering and cell entry. The medically-important Paramyxovirus family includes the deadly Nipah virus (NiV). After paramyxoviruses attach to a receptor at a cell surface, fusion between viral and cellular membranes must occur before the virus genetic material can enter the cell and replication of the virus inside the cell can begin. For most paramyxoviruses, viral/cell membrane fusion requires the concerted actions of two viral glycoproteins. After binding to a cell surface receptor, the viral attachment glycoprotein triggers the viral fusion glycoprotein to execute viral/cell membrane fusion so the genetic material of the virus can enter the cell. However, the mechanism of this receptor-induced triggering of membrane fusion is not well understood. We identified several sequential receptor-induced structural changes in the attachment glycoprotein of NiV that are part of the viral/cell membrane fusion-triggering cascade. Importantly, we propose a mechanism of cell receptor-induced paramyxovirus entry into cells, based on the findings described here, similarities between NiV and other paramyxoviruses, and other recent advances.

Plague is a flea-borne rodent-associated zoonotic disease caused by Yersinia pestis . The disease is characterized by epizootics with high rodent mortalities, punctuated by interepizootic periods when the bacterium persists in an unknown reservoir. This study investigates the interaction between Y. pestis and the ubiquitous soil free-living amoeba (FLA) Acanthamoeba castellanii to assess if the bacterium can survive within soil amoebae and whether intracellular mechanisms are conserved between infection of mammalian macrophages and soil amoebae. The results demonstrate that during coculture with amoebae, representative Y. pestis strains of epidemic biovars Medievalis, Orientalis, and Antiqua are phagocytized and able to survive within amoebae for at least 5 days. Key Y. pestis determinants of the intracellular interaction of Y. pestis and phagocytic macrophages, PhoP and the type three secretion system (T3SS), were then tested for their roles in the Y. pestis -amoeba interaction. Consistent with a requirement for the PhoP transcriptional activator in the intracellular survival of Y. pestis in macrophages, a PhoP mutant is unable to survive when cocultured with amoebae. Additionally, induction of the T3SS blocks phagocytic uptake of Y. pestis by amoebae, similar to that which occurs during macrophage infection. Electron microscopy revealed that in A. castellanii , Y. pestis resides intact within spacious vacuoles which were characterized using lysosomal trackers as being separated from the lysosomal compartment. This evidence for prolonged survival and subversion of intracellular digestion of Y. pestis within FLA suggests that protozoa may serve as a protective soil reservoir for Y. pestis . IMPORTANCE Yersinia pestis is a reemerging flea-borne zoonotic disease. Sylvatic plague cycles are characterized by an epizootic period during which the disease spreads rapidly, causing high rodent mortality, and an interepizootic period when the bacterium quiescently persists in an unknown reservoir. An understanding of the ecology of Y. pestis in the context of its persistence in the environment and its reactivation to initiate a new epizootic cycle is key to implementing novel surveillance strategies to more effectively predict and prevent new disease outbreaks. Here, we demonstrate prolonged survival and subversion of intracellular digestion of Y. pestis within a soil free-living amoeba. This suggests the potential role for protozoa as a protective soil reservoir for Y. pestis , which may help explain the recrudescence of plague epizootics.

Conformational changes in the glycoproteins of enveloped viruses are critical for membrane fusion, which enables viral entry into cells and the pathological cell-cell fusion (syncytia) associated with some viral infections. However, technological capabilities for identifying viral glycoproteins and their conformational changes on actual enveloped virus surfaces are generally scarce, challenging, and time-consuming. Our model, Nipah virus (NiV), is a syncytium-forming biosafety level 4 pathogen with a high mortality rate (40 to 75%) in humans. Once the NiV attachment glycoprotein (G) (NiV-G) binds the cell receptor ephrinB2 or -B3, G triggers conformational changes in the fusion glycoprotein (F) that result in membrane fusion and viral entry. We demonstrate that confocal micro-Raman spectroscopy can, within minutes, simultaneously identify specific G and F glycoprotein signals and receptor-induced conformational changes in NiV-F on NiV virus-like particles (VLPs). First, we identified reproducible G- and F-specific Raman spectral features on NiV VLPs containing M (assembly matrix protein), G, and/or F or on NiV/vesicular stomatitis virus (VSV) pseudotyped virions via second-derivative transformations and principal component analysis (PCA). Statistical analyses validated our PCA models. Dynamic temperature-induced conformational changes in F and G or receptor-induced target membrane-dependent conformational changes in F were monitored in NiV pseudovirions in situ in real time by confocal micro-Raman spectroscopy. Advantageously, Raman spectroscopy can identify specific protein signals in relatively impure samples. Thus, this proof-of-principle technological development has implications for the rapid identification and biostability characterization of viruses in medical, veterinary, and food samples and for the analysis of virion glycoprotein conformational changes in situ during viral entry.

The aim of this study was to measure the prevalence of gastrointestinal parasites (GI) in domestic animals and children in high mountain populations in the districts of Combia and Toche, Valle del Cauca–Colombia. These communities have been affected by the armed conflict in Colombia and are susceptible to health risk factors related to the Colombian post-conflict. Prevalence and risk factors were measured using Bayesian methods on 45 structured interviews applied to 29 families in Combia and 16 in Toche. This inquire aimed to analyze the socio-economic and demographic factors associated with the presence of parasites. This interview was conducted with 50 children: 40 (80%) from Rita Sabogal school district of Toche, and 10 (20%) from Tablones—Atanasio Girardot schools. 23 faecal samples from asymtomatic children from these schools were collected. Subsequently, 308 animals were characterized through the analysis of 64 faecal samples from asymptomatic individuals (20,8%); 18/41 from dogs (43,9%), 18/175 from poultry (10,3%), 7/13 from cats 56,84%, 6/20 from equines (30%) and from 15/59 cattle (25,43%). The prevalence of intestinal parasites among children under six years was 60% [95% CI = 41%-78%]; Endolimax nana, 24% [95% CI = 9,8%-42%]; Iodamoeba buetschlii, 16% [95% CI = 4,7%-32%]; Entamoeba coli, 35% [95% CI = 18%-55%]; Giardia lamblia, 12% [95% CI = 2,7%-27%]. In Equids the presence of Strongylus spp was 37% [95% CI = 10%-71%]; Parascaris equorum, 37% [95% CI = 10%-71%]; in dogs, Dipyllidium caninum was 20% [95% CI = 6%-39%]; Trichuris trichiura, 9% [95% CI = 1,3%-26%]; Toxocara canis, 25% [95% CI = 9%-46%]; in cats, Toxocara cati had a prevalence of 44% [95% CI = 16%-75%]; cyst of Eimeria spp, 15% [95% CI = 3,4%-33%]; in poultry and Eimeria zuernii in cattle, 50% [95% CI = 23%-77%]. There was no association with exposure of humans to animal parasites. However, we conclude that female and children under 6 years of age are more likely OR (6,72–2,3) to get parasites. The Colombian armed conflict affected vulnerable communities with already precarious governmental and health care assistance. Epidemiological surveillance applied to public health is fundamental to prevent zoonotic diseases in vulnerable Colombian populations. Systematic analysis are needed in order to diagnose the problem, to design, to implement public health practices and to evaluate the interventions associated with the identification of parasitic risk factors. In livestock there are several risks linked to human behaviors that have an impact on the occurrence, dissemination, perpetuity of parasitic agents and the quality of life of people. Therefore, it is required to study the risk of parasitic diseases for the farmer production at the high mountain rural communities of Palmira in the post-conflict period. Overall, this study represents an important approach to understand endemic parasites in high elevations of the Andes region of Colombia, to prevent zoonotic diseases and to recognize the relevance of the One Health approach considering ecosystems, domestic animals, wild animals and human health as a unit.

Yersinia pseudotuberculosis is a foodborne enteric pathogen that causes a mild self-limiting diarrhea in humans. Yersinia pseudotuberculosis is able to persist in soil and water and in association with fresh produce, but the mechanism by which it persists is unknown. It has been shown that Y. pseudotuberculosis co-occurs with protozoans in these environments; therefore, this study investigates if bacterivorous free-living amoeba (FLA) are able to support persistence of Y. pseudotuberculosis. Coculture studies of Y. pseudotuberculosis and the prototype FLA, Acanthamoeba castellanii revealed that bacteria had an enhanced capacity to survive in association with amoeba and in the absence of any cytotoxic effects. Yersinia pseudotuberculosis is able to survive and replicate in trophozoites specifically localized within vacuoles, and persists within cysts over a period of at least a week. These data present the first evidence that Y. pseudotuberculosis is able to resist the bacterivorous nature of FLA and instead exhibits an enhanced ability to replicate and persist in coculture with amoeba. This study sheds light on the potential role of FLA in the ecology of Y. pseudotuberculosis which may have implications for food safety.

Objetivos: Reconocer las especies de Ixodidae en el Valle del Cauca permite establecer medidas preventivas para el manejo de las enfermedades transmitidas por estos ectoparásitos y el riesgo de propagación de estos vectores. Alcance: Inventario de garrapatas duras del Valle del Cauca y fortalecimiento de la Colección Entomológica de la Universidad Nacional de Colombia, Palmira (CEUNP). Metodología: Se recolectaron 1.897 garrapatas en todos los estadios de desarrollo, sobre animales domésticos y silvestres en 11 localidades del departamento y se identificaron mediante claves taxonómicas. Principales resultados: 11 especies de los géneros Rhipicephalus (Koch, 1844), Anocentor (Neumann, 1897), Amblyomma (Koch, 1844) e Ixodes (Latreille, 1795). El 53% de los individuos colectados correspondió a R. (B.) microplus, siempre asociada a bovinos, seguido por Anocentor nitens asociada a equinos en un 33%. Las otras especies encontradas se presentaron en menor abundancia, así: R. sanguineus sobre caninos, A. nodosum sobre Tamandua tetradactyla, Amblyomma cajennense sobre equinos y bovinos, Ixodes spp. sobre equinos, Didelphis marsupialis, Amblyomma rotundatum sobre Rhinella marina, Amblyomma spp. sobre bovinos, Amblyomma maculatum sobre equinos, Rhipicephalus spp. Sobre T. Tetradactyla; y Amblyomma dissimile sobre Bothrops asper. Conclusiones: Este es el primer reporte de la presencia de Amblyomma nodosum en Tamandua tetradactyla en la Reserva Forestal Bosque de Yotoco y se confi rmó la presencia de ejemplares previamente reportados para el Valle del Cauca.