scholarly journals Equine Arteritis Virus Elicits a Mucosal Antibody Response in the Reproductive Tract of Persistently Infected Stallions

2017 ◽  
Vol 24 (10) ◽  
Author(s):  
Mariano Carossino ◽  
Bettina Wagner ◽  
Alan T. Loynachan ◽  
R. Frank Cook ◽  
Igor F. Canisso ◽  
...  
Keyword(s):  
Antibody Response ◽  
Seminal Plasma ◽  
Persistent Infection ◽  
Reproductive Tract ◽  
Equine Arteritis Virus ◽  
Serological Response ◽  
Local Inflammatory Response ◽  
Accessory Sex Glands ◽  
Immunoglobulin Isotypes ◽  
Mucosal Antibody

ABSTRACT Equine arteritis virus (EAV) has the ability to establish persistent infection in the reproductive tract of the stallion (carrier) and is continuously shed in its semen. We have recently demonstrated that EAV persists within stromal cells and a subset of lymphocytes in the stallion accessory sex glands in the presence of a significant local inflammatory response. In the present study, we demonstrated that EAV elicits a mucosal antibody response in the reproductive tract during persistent infection with homing of plasma cells into accessory sex glands. The EAV-specific immunoglobulin isotypes in seminal plasma included IgA, IgG1, IgG3/5, and IgG4/7. Interestingly, seminal plasma IgG1 and IgG4/7 possessed virus-neutralizing activity, while seminal plasma IgA and IgG3/5 did not. However, virus-neutralizing IgG1 and IgG4/7 in seminal plasma were not effective in preventing viral infectivity. In addition, the serological response was primarily mediated by virus-specific IgM and IgG1, while virus-specific serum IgA, IgG3/5, IgG4/7, and IgG6 isotype responses were not detected. This is the first report characterizing the immunoglobulin isotypes in equine serum and seminal plasma in response to EAV infection. The findings presented herein suggest that while a broader immunoglobulin isotype diversity is elicited in seminal plasma, EAV has the ability to persist in the reproductive tract, in spite of local mucosal antibody and inflammatory responses. This study provides further evidence that EAV employs complex immune evasion mechanisms during persistence in the reproductive tract that warrant further investigation.

scholarly journals Downregulation of MicroRNA eca-mir-128 in Seminal Exosomes and Enhanced Expression of CXCL16 in the Stallion Reproductive Tract Are Associated with Long-Term Persistence of Equine Arteritis Virus

2018 ◽  
Vol 92 (9) ◽  
Author(s):  
Mariano Carossino ◽  
Pouya Dini ◽  
Theodore S. Kalbfleisch ◽  
Alan T. Loynachan ◽  
Igor F. Canisso ◽  
...  
Keyword(s):  
Persistent Infection ◽  
Molecular Mechanisms ◽  
Reproductive Tract ◽  
Antibody Responses ◽  
Equine Arteritis Virus ◽  
Putative Target ◽  
Mucosal Antibody ◽  
Enhanced Expression ◽  
Specific Allele

ABSTRACTEquine arteritis virus (EAV) can establish long-term persistent infection in the reproductive tract of stallions and is shed in the semen. Previous studies showed that long-term persistence is associated with a specific allele of theCXCL16gene (CXCL16S) and that persistent infection is maintained despite the presence of a local inflammatory and humoral and mucosal antibody responses. In this study, we demonstrated that equine seminal exosomes (SEs) are enriched in a small subset of microRNAs (miRNAs). Most importantly, we demonstrated that long-term EAV persistence is associated with the downregulation of an SE-associated miRNA (eca-mir-128) and with an enhanced expression of CXCL16 in the reproductive tract, a putative target of eca-mir-128. The findings presented here suggest that SE eca-mir-128 is implicated in the regulation of the CXCL16/CXCR6 axis in the reproductive tract of persistently infected stallions, a chemokine axis strongly implicated in EAV persistence. This is a novel finding and warrants further investigation to identify its specific mechanism in modulating the CXCL16/CXCR6 axis in the reproductive tract of the EAV long-term carrier stallion.IMPORTANCEEquine arteritis virus (EAV) has the ability to establish long-term persistent infection in the stallion reproductive tract and to be shed in semen, which jeopardizes its worldwide control. Currently, the molecular mechanisms of viral persistence are being unraveled, and these are essential for the development of effective therapeutics to eliminate persistent infection. Recently, it has been determined that long-term persistence is associated with a specific allele of theCXCL16gene (CXCL16S) and is maintained despite induction of local inflammatory, humoral, and mucosal antibody responses. This study demonstrated that long-term persistence is associated with the downregulation of seminal exosome miRNA eca-mir-128 and enhanced expression of its putative target, CXCL16, in the reproductive tract. For the first time, this study suggests complex interactions between eca-mir-128 and cellular elements at the site of EAV persistence and implicates this miRNA in the regulation of the CXCL16/CXCR6 axis in the reproductive tract during long-term persistence.

scholarly journals Intrahost Selection Pressure Drives Equine Arteritis Virus Evolution during Persistent Infection in the Stallion Reproductive Tract

2019 ◽  
Vol 93 (12) ◽  
Author(s):  
Bora Nam ◽  
Zelalem Mekuria ◽  
Mariano Carossino ◽  
Ganwu Li ◽  
Ying Zheng ◽  
...  
Keyword(s):  
Persistent Infection ◽  
Viral Genome ◽  
Reproductive Tract ◽  
Purifying Selection ◽  
Buffy Coat ◽  
Equine Arteritis Virus ◽  
Nucleotide Substitutions ◽  
Genome Wide ◽  
Nasal Secretions

ABSTRACTEquine arteritis virus (EAV) is the causative agent of equine viral arteritis (EVA), a reproductive and respiratory disease of horses. Following natural infection, 10 to 70% of infected stallions can become carriers of EAV and continue to shed virus in the semen. In this study, sequential viruses isolated from nasal secretions, buffy coat cells, and semen of seven experimentally infected and two naturally infected EAV carrier stallions were deep sequenced to elucidate the intrahost microevolutionary process after a single transmission event. Analysis of variants from nasal secretions and buffy coat cells lacked extensive positive selection; however, characteristics of the mutant spectra were different in the two sample types. In contrast, the initial semen virus populations during acute infection have undergone a selective bottleneck, as reflected by the reduction in population size and diversifying selection at multiple sites in the viral genome. Furthermore, during persistent infection, extensive genome-wide purifying selection shaped variant diversity in the stallion reproductive tract. Overall, the nonstochastic nature of EAV evolution during persistent infection was driven by active intrahost selection pressure. Among the open reading frames within the viral genome, ORF3, ORF5, and the nsp2-coding region of ORF1a accumulated the majority of nucleotide substitutions during persistence, with ORF3 and ORF5 having the highest intrahost evolutionary rates. The findings presented here provide a novel insight into the evolutionary mechanisms of EAV and identified critical regions of the viral genome likely associated with the establishment and maintenance of persistent infection in the stallion reproductive tract.IMPORTANCEEAV can persist in the reproductive tract of infected stallions, and consequently, long-term carrier stallions constitute its sole natural reservoir. Previous studies demonstrated that the ampullae of the vas deferens are the primary site of viral persistence in the stallion reproductive tract and the persistence is associated with a significant inflammatory response that is unable to clear the infection. This is the first study that describes EAV full-length genomic evolution during acute and long-term persistent infection in the stallion reproductive tract using next-generation sequencing and contemporary sequence analysis techniques. The data provide novel insight into the intrahost evolution of EAV during acute and persistent infection and demonstrate that persistent infection is characterized by extensive genome-wide purifying selection and a nonstochastic evolutionary pattern mediated by intrahost selective pressure, with important nucleotide substitutions occurring in ORF1a (region encoding nsp2), ORF3, and ORF5.

scholarly journals Distribution of hyaluronan in bull reproductive organs.

1994 ◽  
Vol 42 (11) ◽  
pp. 1479-1486 ◽  
Author(s):  
R Tammi ◽  
S Rönkkö ◽  
U M Agren ◽  
M Tammi
Keyword(s):  
Seminal Vesicle ◽  
Seminal Plasma ◽  
Reproductive Tract ◽  
Reproductive Organs ◽  
Accessory Gland ◽  
Seminiferous Tubules ◽  
Tissue Sections ◽  
Accessory Sex Glands ◽  
Cartilage Proteoglycan

To study the expression of hyaluronan in male reproductive organs and the origin of seminal plasma hyaluronan, we stained various parts of the bull reproductive tract for hyaluronan using a biotinylated probe derived from cartilage proteoglycan (bHABC). The potential loss of hyaluronan during tissue processing was checked with a novel technique by blotting frozen tissue sections on nitrocellulose and staining the blots with bHABC. In the same tissues the CD44 receptor was visualized by Hermes 1 antibody. The testes showed only traces of hyaluronan, whereas both the epithelium and the connective tissue of seminal vesicle, prostate, Cowper's gland, and epididymis were positive in bHABC staining. Hyaluronan was localized on the basolateral surfaces of these epithelial cells. The secretions inside the seminal vesicle and in the ducts of prostate and Cowper's gland were HA-positive, whereas the luminal contents of seminiferous tubules and epididymis were unstained both in paraffin sections and in the in situ blocks. The data indicate that hyaluronan in seminal plasma originates from the accessory sex glands. The co-localization of CD44 with hyaluronan in the basolateral surfaces of the accessory gland epithelia and its absence from other epithelia with little or no hyaluronan supports its role as a hyaluronan receptor.

scholarly journals Equine Arteritis Virus Has Specific Tropism for Stromal Cells and CD8+ T and CD21+ B Lymphocytes but Not for Glandular Epithelium at the Primary Site of Persistent Infection in the Stallion Reproductive Tract

2017 ◽  
Vol 91 (13) ◽  
Author(s):  
Mariano Carossino ◽  
Alan T. Loynachan ◽  
Igor F. Canisso ◽  
R. Frank Cook ◽  
Juliana R. Campos ◽  
...  
Keyword(s):  
Viral Replication ◽  
B Lymphocytes ◽  
Immune Evasion ◽  
Stromal Cells ◽  
Persistent Infection ◽  
Reproductive Tract ◽  
Inflammatory Responses ◽  
Viral Persistence ◽  
Equine Arteritis Virus ◽  
Glandular Epithelium

ABSTRACT Equine arteritis virus (EAV) has a global impact on the equine industry as the causative agent of equine viral arteritis (EVA), a respiratory, systemic, and reproductive disease of equids. A distinctive feature of EAV infection is that it establishes long-term persistent infection in 10 to 70% of infected stallions (carriers). In these stallions, EAV is detectable only in the reproductive tract, and viral persistence occurs despite the presence of high serum neutralizing antibody titers. Carrier stallions constitute the natural reservoir of the virus as they continuously shed EAV in their semen. Although the accessory sex glands have been implicated as the primary sites of EAV persistence, the viral host cell tropism and whether viral replication in carrier stallions occurs in the presence or absence of host inflammatory responses remain unknown. In this study, dual immunohistochemical and immunofluorescence techniques were employed to unequivocally demonstrate that the ampulla is the main EAV tissue reservoir rather than immunologically privileged tissues (i.e., testes). Furthermore, we demonstrate that EAV has specific tropism for stromal cells (fibrocytes and possibly tissue macrophages) and CD8+ T and CD21+ B lymphocytes but not glandular epithelium. Persistent EAV infection is associated with moderate, multifocal lymphoplasmacytic ampullitis comprising clusters of B (CD21+) lymphocytes and significant infiltration of T (CD3+, CD4+, CD8+, and CD25+) lymphocytes, tissue macrophages, and dendritic cells (Iba-1+ and CD83+), with a small number of tissue macrophages expressing CD163 and CD204 scavenger receptors. This study suggests that EAV employs complex immune evasion mechanisms that warrant further investigation. IMPORTANCE The major challenge for the worldwide control of EAV is that this virus has the distinctive ability to establish persistent infection in the stallion's reproductive tract as a mechanism to ensure its maintenance in equid populations. Therefore, the precise identification of tissue and cellular tropism of EAV is critical for understanding the molecular basis of viral persistence and for development of improved prophylactic or treatment strategies. This study significantly enhances our understanding of the EAV carrier state in stallions by unequivocally identifying the ampullae as the primary sites of viral persistence, combined with the fact that persistence involves continuous viral replication in fibrocytes (possibly including tissue macrophages) and T and B lymphocytes in the presence of detectable inflammatory responses, suggesting the involvement of complex viral mechanisms of immune evasion. Therefore, EAV persistence provides a powerful new natural animal model to study RNA virus persistence in the male reproductive tract.

scholarly journals Genetic characterization of equine arteritis virus during persistent infection of stallions

2004 ◽  
Vol 85 (2) ◽  
pp. 379-390 ◽  
Author(s):  
Udeni B. R. Balasuriya ◽  
Jodi F. Hedges ◽  
Victoria L. Smalley ◽  
Andrea Navarrette ◽  
William H. McCollum ◽  
...  
Keyword(s):  
Persistent Infection ◽  
Reproductive Tract ◽  
Genetic Characterization ◽  
Neutralization Assay ◽  
High Titre ◽  
Equine Arteritis Virus ◽  
Replicase Gene ◽  
Defective Interfering Particles ◽  
Genomic Deletions

Equine arteritis virus (EAV) causes a persistent infection of the reproductive tract of carrier stallions. The authors determined the complete genome sequences of viruses (CW96 and CW01) that were present 5 years apart in the semen of a carrier stallion (CW). The CW96 and CW01 viruses respectively had only 85·6 % and 85·7 % nucleotide identity to the published sequence of EAV (EAV030). The CW96 and CW01 viruses had two 1 nt insertions and a single 1 nt deletion in the leader sequence, and a 3 nt coding insertion in ORF1a; thus their genomes included 12 708 nt as compared to the 12 704 nt in EAV030. Variation between viruses present in the semen of stallion CW and EAV030 was especially marked in the replicase gene (ORF1a and 1b), and the greatest variation occurred in the portion of ORF1a encoding the nsp2 protein. The ORFs 3 and 5, which respectively encode the GP3 and GP5 envelope proteins, showed greatest variation amongst ORFs encoding structural EAV proteins. Comparative sequence analyses of CW96 and CW01 indicated that ORFs 1a, 1b and 7 were highly conserved during persistent infection, whereas there was substantial variation in ORFs 3 and 5. Although the variation that occurs in ORF5 results in the emergence of novel phenotypic viral variants as determined by neutralization assay, all variants were neutralized by high-titre polyclonal equine antisera, suggesting that immune evasion is unlikely to be responsible for the establishment of persistent EAV infection of carrier stallions. Northern blot analyses of RNA extracted from cell culture propagated viruses isolated from 10 different persistently infected stallions failed to demonstrate any large genomic deletions, suggesting that defective interfering particles are also unlikely to be important in either the maintenance or clearance of persistent EAV infection of the reproductive tract of carrier stallions.

scholarly journals Genetic Divergence with Emergence of Novel Phenotypic Variants of Equine Arteritis Virus during Persistent Infection of Stallions

1999 ◽  
Vol 73 (5) ◽  
pp. 3672-3681 ◽  
Author(s):  
Jodi F. Hedges ◽  
Udeni B. R. Balasuriya ◽  
Peter J. Timoney ◽  
William H. McCollum ◽  
N. James MacLachlan
Keyword(s):  
Persistent Infection ◽  
Reproductive Tract ◽  
Variable Region ◽  
Open Reading Frames ◽  
Equine Arteritis Virus ◽  
Phenotypic Properties ◽  
Equine Viral Arteritis ◽  
Venereal Infection ◽  
Reading Frames

ABSTRACT The persistently infected carrier stallion is the critical natural reservoir of equine arteritis virus (EAV), as venereal infection of mares frequently occurs after breeding to such stallions. Two Thoroughbred stallions that were infected during the 1984 outbreak of equine viral arteritis in central Kentucky subsequently became long-term EAV carriers. EAV genomes amplified from the semen of these two stallions were compared by sequence analysis of the six 3′ open reading frames (ORFs 2 through 7), which encode the four known structural proteins and two uncharacterized glycoproteins. The major variants of the EAV population that sequentially arose within the reproductive tract of each carrier stallion varied by approximately 1% per year, and the heterogeneity of the viral quasispecies increased during the course of long-term persistent infection. The various ORFs of the dominant EAV variants evolved independently, and there was apparently strong selective pressure on the uncharacterized GP3 protein during persistent infection. Amino acid changes also occurred in the V1 variable region of the GL protein. This region has been previously identified as a crucial neutralization domain, and selective pressures exerted on the V1 region during persistent EAV infection led to the emergence of virus variants with distinct neutralization properties. Thus, evolution of the EAV quasispecies that occurs during persistent infection of the stallion clearly can influence viral phenotypic properties such as neutralization and perhaps virulence.

scholarly journals Proteins of the canine seminal plasma

2016 ◽  
Vol 46 (5) ◽  
pp. 901-908 ◽  
Author(s):  
Annice Aquino-Cortez ◽  
Lúcia Daniel Machado da Silva ◽  
Airton Alencar de Araújo ◽  
Erika da Silva Bezerra de Menezes ◽  
Arlindo de Alencar Araripe Noronha Moura
Keyword(s):  
Alkaline Phosphatase ◽  
Seminal Plasma ◽  
Binding Proteins ◽  
Reproductive Tract ◽  
Sperm Quality ◽  
Zinc Binding ◽  
Heparin Binding ◽  
Reproductive Disorders ◽  
The Matrix ◽  
Peroxidase Enzymes

ABSTRACT: Studies have been performed to identify the proteins present in canine seminal plasma (SP) and relate them to sperm quality as well as to discover molecular markers of reproductive tract diseases. There is evidence that heparin-binding proteins, zinc-binding proteins, and lactoferrin as well as the matrix metalloproteinase, superoxide dismutase, catalase, and glutathione peroxidase enzymes are associated with canine sperm quality. Other studies indicate that prolactin and enzymes like arginine esterase, acid phosphatase, and alkaline phosphatase could be successfully used as biomarkers of reproductive disorders. Thus, the present literature review aims to address aspects related to proteins of the canine SP, their influence on fertility, and their importance as biomarkers of reproductive disorders.

TESTOSTERONE AND DIHYDROTESTOSTERONE CONCENTRATIONS IN THE FLUID MILIEU OF SPERMATOZOA IN THE REPRODUCTIVE TRACT OF THE BULL

1973 ◽  
Vol 74 (1) ◽  
pp. 186-200 ◽  
Author(s):  
Venkataseshu K. Ganjam ◽  
Rupert P. Amann
Keyword(s):  
Mass Spectrometry ◽  
Seminal Plasma ◽  
Reproductive Tract ◽  
Binding Assay ◽  
Rete Testis ◽  
Protein Binding Assay ◽  
Highly Sensitive ◽  
Competitive Protein Binding Assay ◽  
Competitive Protein Binding ◽  
Isolated Compound

ABSTRACT Total 17β-hydroxyandrogen concentrations were determined using a competitive protein binding assay, for bovine reproductive fluids. Rete testis fluid and cauda epididymal plasma were separated from the spermcontaining fluids obtained through cannulae from conscious bulls. Al-through the concentration of total 17β-hydroxyandrogens in rete testis fluid was similar (P > 0.05) to that in cauda epididymal plasma (25 and 19 ng/ml), both fluids contained higher (P < 0.01) androgen concentrations than seminal plasma, accessory sex gland fluid or serum from peripheral blood (3–5 ng/ml). However, since the amount of cauda epididymal plasma recovered was much less than for rete testis fluid (0.25 vs 35 ml/day), cauda epididymal plasma contained less than 1 % of the total 17β-hydroxyandrogens which entered the epididymis in rete testis fluid (5 vs 883 ng/day). Testosterone and/or dihydrotestosterone were isolated from the reproductive fluids by Sephadex LH-20 chromatography and quantified by a simple, specific and highly sensitive microassay. Dihydrotestosterone was found only in cauda epididymal plasma (14 ng/ml); identification of the isolated compound was confirmed by mass spectrometry. Dihydrotestosterone accounted for 52% of the 17β-hydroxyandrogens in cauda epididymal plasma while 23 % was testosterone. Testosterone represented 70 % of the 17β-hydroxyandrogens in rete testis fluid and 91 % of those in blood serum. Physiological implications of this shift in androgen balance are discussed.

scholarly journals Spermatozoa Obtained From Alpaca vas deferens. Effects of Seminal Plasma Added at Post-thawing

2021 ◽  
Vol 8 ◽  
Author(s):  
Eduardo G. Aisen ◽  
Wilfredo Huanca López ◽  
Manuel G. Pérez Durand ◽  
Edita Torres Mamani ◽  
Juan C. Villanueva Mori ◽  
...  
Keyword(s):  
Seminal Plasma ◽  
Artificial Insemination ◽  
Reproductive Tract ◽  
Vas Deferens ◽  
Female Reproductive Tract ◽  
Low Fertility ◽  
Sperm Cells ◽  
South American Camelids ◽  
Thawing Process

The viscous seminal plasma (SP) is currently a major impediment to the handling of ejaculate and the development of some biotechnologies in South American camelids. The vas deferens-collected spermatozoa of alpacas is a useful technique to avoid this problem. On the other hand, SP contains a large protein component that has been implicated in the function of spermatozoa within the female reproductive tract. In this sense, the low fertility achieved using transcervical insemination with frozen-thawed spermatozoa in alpacas could be improved by adding SP. This study aimed to evaluate the effect of the whole SP on some in vitro parameters of alpaca spermatozoa after the freezing-thawing-process and the fertility after artificial insemination. It would contribute to a better understanding of the interaction between thawed sperm cells and SP. Spermatozoa were obtained by surgically diverted vas deferens. The samples were diluted with a Tris-based extender, packaged in straws, and frozen. At thawing, each straw was divided into two post-thawing conditions: with the addition of 10% of PBS (control) or with 10% SP (treatment). The sperm cells were evaluated using dynamic parameters, sperm cell morphology, and morphometry. Fertility was assessed by an artificial insemination trial. All in vitro parameters were analyzed by ANOVA. A heterogeneity test was scheduled for the fertility trial. After the freezing-thawing process, motility and plasma membrane functionality was improved when SP was added. No differences were found for post-thaw viability between the control and treatment samples. The percentage of normal cells was higher with SP at post-thawing, and a decrease of the presence of bent tailed spermatozoa with a droplet in the SP group was observed. The length of the head spermatozoa was 3.4% higher in the samples with PBS compared to those in which SP was added. Females pregnant at day 25 post-insemination were 0/12 (with SP inside the straw) and 1/10 (without SP inside the straw). In conclusion, the presence of 10% SP at post-thawing improves sperm cells' motility, functionality, and morphology, indicating that it would be beneficial to improve the frozen-thawed alpaca's physiology spermatozoa. More fertility trials must be developed to increase this knowledge.

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