Immune protection induced by E2 recombinant glycoprotein of bovine viral diarrhea virus in a murine model.

Bovine viral diarrhea virus (BVDV) is considered the most important viral pathogen in ruminants worldwide due to the broad range of clinical manifestations displayed by infected animals. Therefore, infection with BVDV leads to severe economic losses in several countries' beef and dairy industries. Vaccination prevents reproductive failure and gastrointestinal and respiratory disorders caused by BVDV infection. However, considering their limitations, conventional vaccines such as live, attenuated, and killed viruses have been applied. Hence, different studies have described subunit vaccines as an effective and safe alternative for BVDV protection. Therefore, in this study, the ectodomain of E2 (E2e) glycoprotein from NADL BVDV strain was expressed in mammalian cells and used in two vaccine formulations to evaluate immunogenicity and protection against BVDV conferred in a murine model. Formulations consisted of solo E2e glycoprotein and E2e glycoprotein emulsified in adjuvant ISA 61 VG. Five groups of 6 mice of 6-to-8-week-old were immunized thrice on days 1, 15, and 30 by intraperitoneal injection with the mentioned formulations and controls. To evaluate the conferred protection against BVDV, mice were challenged six weeks after the third immunization. In addition, the humoral immune response was evaluated after vaccination and challenge. Mice groups inoculated with solo E2e and the E2e + ISA 61 VG displayed neutralizing titers; however, the E2 antibody titers in the E2e + ISA 61 VG group were significantly higher than the mice group immunized with the solo E2e glycoprotein. In addition, immunization using E2e + ISA 61 VG prevents animals from developing severe lesions in surveyed tissues. Moreover, this group acquired protection against the BVDV challenge, evidenced by a significant reduction of positive staining for BVDV antigen in the lungs, liver, and brain between the experimental groups. Our findings demonstrated that using E2e + ISA 61 VG induces greater BVDV protection by an early humoral response and reduced histopathological lesions and BVDV antigen detection in affected organs, indicating that E2e + ISA 61 VG subunit formulation can be considered as a putative vaccine candidate against BVDV. The efficacy and safety of this vaccine candidate in cattle requires further investigation.

Bovine Viral Diarrhea Virus in Cattle From Mexico: Current Status.

Bovine viral diarrhea (BVD) is an infectious disease, globally-distributed, caused by bovine Pestiviruses, endemic of cattle and other ruminant populations. BVD leads to significant economic losses to the cattle industry due to the wide range of clinical manifestations, including respiratory and gastrointestinal diseases and reproductive disorders. Within the Pestivirus genus of the family Flaviviridae three viral species are associated with BVD; Pestivirus A (Bovine viral diarrhea virus 1, BVDV-1), Pestivirus B (Bovine viral diarrhea virus 2, BVDV-2), and Pestivirus H (HoBi-like pestivirus, atypical ruminant pestivirus). These species are subdivided into subgenotypes based on phylogenetic analysis. The extensive genetic diversity of BVDV has been reported for several countries, where the incidence and genetic variation are more developed in Europe than in the Americas. The first report of BVDV in Mexico was in 1975; this study revealed seropositivity of 75% in cows with a clinical history of infertility, abortions, and respiratory disease. Other studies have demonstrated the presence of antibodies against BVDV with a seroprevalence ranging from 7.4 to 100%. Recently, endemic BVDV strains affecting cattle populations started to be analyzed, providing evidence of the BVDV diversity in several states of the country, revealing that at least four subgenotypes (BVDV-1a, 1b, 1c, and 2a) are circulating in animal populations in Mexico. Little information regarding BVD epidemiological current status in Mexico is available. This review summarizes available information regarding the prevalence and genetic diversity viruses associated with BVD in cattle from Mexico.

Detection and genotyping of bovine viral diarrhea virus found contaminating commercial veterinary vaccines, cell lines, and fetal bovine serum lots originating in Mexico.

In this communication, we report the presence of RNA of bovine viral diarrhea virus (BVDV) as a contaminant of different biological products used in Mexico for veterinary vaccine production. For this purpose, six batches of monovalent vaccines, eight cell line batches used for vaccine production, and 10 fetal bovine serum lots (FBS) commercially available in Mexico from different suppliers were tested by reverse transcription polymerase chain reaction (RT-PCR). Viral RNA was detected in 62.5% of the samples analyzed. Phylogenetic analysis revealed the presence of the subgenotypes BVDV-1a, 1b, and BVDV-2a in the tested samples. Collectively, these findings indicate that contamination by BVDV RNA occurs in commercial vaccines and reagents used in research and production of biological products. The ramifications of this contamination are discussed.

Canine distemper in neotropical procyonids: Molecular evidence, humoral immune response and epidemiology.

Canine Distemper Virus (CDV) can produce a fatal multisystem disease in carnivores and other mammals and is an important threat for wildlife conservation. However, integrative and comparative studies in wild carnivores are scarce and some areas of the world lack of genetic studies. We explore the dynamic of host-CDV in a procyonid community during an outbreak. This study reports for the first time an index case occurred in a common raccoon (Procyon lotor) and for which a complete CDV diagnosis was performed. The long-term epidemiological analysis in two sympatric populations of common raccoons and white-nosed coatis (Nasua narica) was achieved through seroneutralization, RT-PCR and direct immunofluorescence assays. Additionally, hematologic analyses were performed and phylogenetic reconstruction of CDV was done using molecular data from this study. Overall prevalence for white-nosed coatis was 19.6 % and for common raccoons was 25.3 % by seroneutralization, and 13.3 % and 17.3 % by RT-PCR. Antibodies titer average for white-nosed coatis was 1:512 and 1:156 for common raccoons. Significant difference in prevalence between white-nosed coatis and common raccoons was detected during one season (summer 2013). White-nosed coatis showed differences in erythrocytes and monocytes counts between positives and negative animals. A 100 % similarity was found between CDV of white-nosed coati and CDV of common raccoon and is a new CDV sequence not previously described; this sequence is close to Asian and European lineage. An endemic state of distemper in both species was observed but showed different dynamics over time per host species. Differences in cellular and humoral responses were also detected between procyonids. The evidence found here may have serious implications for CDV understanding in wild carnivores, it reveals clear differences in the response over time to the same CDV strain, in two close related carnivore species.

Complete Genome Sequences of Two Vesicular Stomatitis New Jersey Viruses Representing the 2012 U.S. Epidemic Strain and Its Closest Relative Endemic Strain from Southern Mexico.

We report here the complete genome sequences of two vesicular stomatitis New Jersey virus (VSNJV) field strains isolated from epithelial lesions from naturally infected animals in Mexico and the United States. The close phylogenetic relationship of these isolates makes them an ideal model for assessing potential genetic factors linked with the emergence of VSNJV in the United States.

Genetic diversity of bovine viral diarrhea virus in cattle from Mexico.

Bovine viral diarrhea virus (BVDV) infects cattle populations worldwide, causing significant economic losses though its impact on animal health. Previous studies have reported the prevalence of BVDV species and subgenotypes in cattle from the United States and Canada. We investigated the genetic diversity of BVDV strains detected in bovine serum samples from 6 different Mexican regions. Sixty-two BVDV isolates from Mexico were genetically typed based on comparison of sequences from the 5' untranslated region (5'-UTR) of the viral genome. Phylogenetic reconstruction indicated that 60 of the samples belonged to the BVDV-1 genotype and 2 to the BVDV-2 genotype. Comparison of partial 5'-UTR sequences clustered 49 samples within BVDV-1c, 8 samples within BVDV-1a, 3 samples within BVDV-1b, and 2 samples clustered with the BVDV-2a subgenotypes. Our study, combined with information previously published on BVDV field strain diversity in the United States and Canada, benefits the development of effective detection assays, vaccines, and control programs for North America.

Omp31 plays an important role on outer membrane properties and intracellular survival of Brucella melitensis in murine macrophages and HeLa cells.

Brucellosis is an infectious disease that affects practically all species of mammals, including human, and is a major zoonosis worldwide. Brucella spp. are facultative intracellular pathogens that have the ability to survive and multiply in phagocytic and nonphagocytic cells such as trophoblast and epithelial cells. Among the six recognized species of the genus Brucella, Brucella melitensis is the main etiological agent involved in goat brucellosis and is also the most pathogenic for human. It causes significant losses in livestock production as a result of abortions, metritis, infertility, and birth of weak animals. Outer membrane proteins (OMPs) are exposed on the bacterial surface and are in contact with cells and effectors of the host immune response, whereby they could be important virulence factors of Brucella species. To evaluate this hypothesis, the gene encoding for the major outer membrane protein Omp31 was amplified, cloned into pUC18 plasmid, and inactivated by inserting a kanamycin cassette, rendering pLVM31 plasmid which was transformed into B. melitensis wild-type strain to obtain LVM31 mutant strain. The Outer membrane (OM) properties of the mutant strain were compared with B. melitensis Bm133 wild-type and B. melitensis Rev1 vaccine strains, in assessing its susceptibility to polymyxin B, sodium deoxycholate, and nonimmune serum. The mutant strain was assessed in vitro with survival assays in murine macrophages J774.A1 and HeLa cells. Our results demonstrate that LVM31 mutant is more susceptible to polymyxin B, sodium deoxycholate, and nonimmune serum than control strains; moreover, Omp31 mutation caused a decrease in the internalization and a significant decrease in the intracellular survival compared with the reference strains in both cell lines. These results allow us to conclude that Omp31 is important for maintaining OM integrity, but also it is necessary for bacterial internalization, establishment and development of an optimal replication niche, and essential for survival and intracellular multiplication.

Phylogeographic characteristics of vesicular stomatitis New Jersey viruses circulating in Mexico from 2005 to 2011 and their relationship to epidemics in the United States.

We analyzed the phylogenetic and time-space relationships (phylodynamics) of 181 isolates of vesicular stomatitis New Jersey virus (VSNJV) causing disease in Mexico and the United States (US) from 2005 through 2012. We detail the emergence of a genetic lineage in southern Mexico causing outbreaks in central Mexico spreading into northern Mexico and eventually into the US. That emerging lineage showed higher nucleotide sequence identity (99.5%) than that observed for multiple lineages circulating concurrently in southern Mexico (96.8%). Additionally, we identified 58 isolates from Mexico that, unlike previous isolates from Mexico, grouped with northern Central America clade II viruses. This study provides the first direct evidence for the emergence and northward migration of a specific VSNJV genetic lineage from endemic areas in Mexico causing VS outbreaks in the US. In addition we document the emergence of a Central American VSNJV genetic lineage moving northward and causing outbreaks in central Mexico.

Rotavirus infections in Galapagos sea lions.

Group A rotaviruses infect and cause diarrhea in the young of a broad range of terrestrial mammals, but it is unknown, to our knowledge, whether they infect marine mammals. During February and March of 2002 and 2003, we collected 125 serum samples and 18 rectal swab samples from Galapagos sea lion pups (GSL, Zalophus wollebaeki), and 22 serum samples from Galapagos fur seal pups (GFS, Arctocephalus galapagoensis) from nine islands of the Galapagos archipelago, Ecuador. Sera were tested for antibodies (immunoglobulin G [IgG]) to rotavirus by an enzyme immunoassay using rhesus rotavirus as the capture antigen. In addition, rectal swabs were analyzed for the presence of rotavirus genomic double-stranded RNA by silver-stained polyacrylamide gel electrophoresis. Antibodies to rotavirus were detected in 27 GSL pups (22%) and five GFS pups (23%), and rotavirus RNA was detected in the fecal sample from one GSL pup (6%). These results provide the first evidence that rotavirus infections are prevalent at an early age in Galapagos sea lions and Galapagos fur seals.

The BMEI0216 gene of Brucella melitensis is required for internalization in HeLa cells.

Brucella is an intracellular facultative bacterium able to survive and multiply in professional and non-professional phagocytes. However, its adhesion and invasion mechanisms have not been elucidated yet. In this work, we assess the interruption of a BMEI0216 gene of Brucella melitensis, by using HeLa epithelial cells and murine macrophages for invasion and replication assays. The mutation did not affect survival or multiplication within macrophages. Likewise, invasion assays with HeLa cells revealed no differences at 30 and 45 min, whereas, at 1 and 2h, the infection ability of the mutant was drastically reduced. These results suggest that the BMEI0216 gene is required for B. melitensis internalization.

[Molecular mechanism for pathogenicity of Salmonella sp]. / Mecanismos moleculares de patogenicidad de Salmonella sp.

Salmonella is a Gram negative bacillus that behaves like a facultative intracellular pathogen. Its environment is the human and animal gastrointestinal tracts, it is never found like a normal microbiota. It is associated with gastrointestinal problems, septicaemic disease and abortion, due to its cellular invasion capacity and its intraphagocytic survival. Nowadays, it is known that Salmonella contains five pathogenicity islands. Several genes involved in the cellular invasion of nonphagocytic cells such as epithelial cells, apoptosis of macrophages, activation of routes of MAP kinases and transcription factors are located in centisome 63, constituting the pathogenicity island 1 (SPI-1). The SPI-2 and SPI-3 islands control the intracellular survival and replication. The SPI-4 island encodes a putative type I secretion system and its believed that it participates in the intracellular survival. Finally, the SPI-5 island encodes for factors involved in the fluid secretion and inflammatory reaction in the intestinal mucosa. Due to a coordinated and precise regulation of the Salmonella genes, it allows for adaptation to environmental changes that occur during an inflammatory process.