Immunized Goats Research Articles

PSII-6 Changes in Galectin Gene Expression and Secretion in Response to Gum Arabica Drench in Goats

Abstract Acacia senegal is part of the natural diet of goats. Gum Arabica (GA) is a water-soluble complex polysaccharide antimicrobial, anti-inflammatory prebiotic, derived from Acacia Senegal. Its mechanism of action has not been defined. Galectins are secreted sugar-binding animal proteins involved in the regulation of innate and adaptive immunity. They function in microbe recognition and as regulators of inflammation. This study evaluated the effect of GA on Galectin transcription. Clinically healthy Boer X Spanish goats (n = 20) were randomly assigned to two groups of 10. Goats in the treatment group received 10 mL of GA in water daily for six weeks, controls received sterile water. Blood was collected weekly from the jugular vein using acid citrate dextrose as an anticoagulant. Subsequently, the blood was centrifuged to isolate the cells and plasma. Total RNA isolation was isolated from the cell pellet using Trizol (Sigma-Aldrich). The RNA concentration (ng/μL) and purity was assessed using a Nanodrop Spectrophotometer (Thermo Scientific Inc). Galectin-1, -3 and -9 transcription was conducted using goat specific primers and housekeeping genes for real-time PCR. Fold changes were calculated using the Livak method. The Log2FC values for Galectin 1, 3, and 9 expressions were 12.47, 0.29, and 2.22 respectively. Thus, GA had varying effects on structurally distinct galectins. Galectin 1 gene (Lgals1) is an important modulator of the immune response and was highly induced. Changes in Galectin transcription can impact microbial recognition and immune activation for goat health. Further analyses are needed to evaluate the functional effects of GA for application as a prebiotic and immune modulator lain goat health.

PSV-25 Effect of Garlic on Galectin 3 in Goats

Abstract The objective of this study was to determine the effect of garlic extracts on expression of Galectin 3 in goats. Inflammatory diseases such as mastitis and gastrointestinal parasitic diseases cause significant problems for goat production. Galectins (Gal) are a family of highly conserved mammalian carbohydrate-binding proteins that can regulate inflammation. Galectin-3 is the only member of the lectin family with a chimeric structure. It is a potential therapeutic target to regulate the activity of inflammatory cells. Garlic extracts are natural anti-inflammatory agents that are being evaluated for use in goat management. Twelve adult female clinically healthy Boer X Spanish goats were divided into 2 groups. The treatment group (n = 6) was drenched with a 10 mL garlic (Garlic barrier) and the control group (n = 6) was given sterile water for 30 days. Blood was collected weekly from the jugular vein using acid citrate dextrose as an anticoagulant. Blood was centrifuged to separate cells and plasma. Galectin 3 secretion in plasma was determined using goat specific commercial ELISA kits (My-BioSource). Total RNA was isolated from the cell pellet using Trizol and it was then converted into cDNA. Primers specific for goat Gal 3 specific and the housekeeping genes RPLP0 and UCHL5 (Eurofins Genomics) were used in Real-time PCR to evaluate the gene expression. Fold change was calculated using the Livak method. All variables were assessed using SAS Statistical analysis software version 9.4 (SAS Institute) at P < 0.05. Treatment had an effect over time and a significant increase (P < 0.0141) was observed in the GAL- 3 concentrations. Gal-3 was expressed in greater amounts in treatment group animals compared with the control group with a fold change of 1.15. Garlic may be used to modulate Gal 3 expression and secretion in goat blood. It is important to consider the impact of garlic on Gal 3 modulation. Studies needed to define possible impacts on immune function and goat health.

Immunization With Recombinant Haemonchus contortus Y75B8A.8 Partially Protects Local Crossbred Female Goats From Haemonchus contortus Infection.

Haemonchus contortus Y75B8A.8 (Hc8) derived from H. contortus excretory–secretory (ES) products was identified as a functional inhibitor of goat interleukin 2 (IL-2). It may act as a vaccine candidate for the development of therapeutic strategies against H. contortus infection. In this research, recombinant Hc8 (rHc8) and goat anti-rHc8 polyclonal antibodies were employed to evaluate the protective capacities of Hc8 antigen against H. contortus infections via active and passive immunization trials, respectively. In both trials, local crossbred female goats aged 9–12 months old were randomly divided into three groups, five in each group, respectively. Parasitological examinations, including fecal egg counts (FEC), cumulative FEC (cFEC), and worm burdens, were performed. In addition, antibody levels in mucosal homogenate (MH) samples and hematological and immunological parameters were detected. In the passive trial, goats were intravenously immunized with 5 mg total IgG containing anti-rHc8 goat polyclonal antibodies. After twice immunization, compared with the challenged control group, cFEC was reduced by 39%. In addition, there was a 46% reduction of worm burdens compared with the challenged controls. In the active immunization trials, 500 μg of recombinant Hc8 protein was given subcutaneously twice to 9–12-month-old local crossbred female goats with a 2-week interval, resulting in the generation of high levels of antigen-specific circulating antibodies. Besides, cFEC and abomasal worm burden were reduced by 70 and 55%, respectively, compared with the challenged control group. In addition, immunized goats had higher mucosal homogenate IgA and hemoglobin levels than the challenged controls in both passive and active immunization trials. These preliminary results demonstrated the immunoprophylactic effects of Hc8 antigen and will inform new studies on ES proteins in developing subunit recombinant vaccines against H. contortus.

Evaluation of mucoadhesive property and the effect of Boswellia carteri gum on intranasal vaccination against small ruminant morbillivirus infection (PPR)

ABSTRACTA study was conducted to evaluate mucoadhesive property and immunomodulatory effect of phytogenic gums from Boswellia frereana, Boswellia carteri andCommiphora myrrha on intranasal Peste des petits ruminants (PPR) vaccination in goats and sheep in an ex-vivo and in-vivo situations. Plant gums were purified, dried and compressed into 500gm tablets. Modified shear stress measurement technique was used on freshly excised trachea and intestine tissues of goat to measure peak adhesion time. Forty eight animals (24 goats and 24 sheep) were divided into eight groups (of 3 goats and 3 sheep) and immunized intranasally with gum-vaccine combinations in two ratios (1:1, 1:2). Antibody against PPR virus was measured on day 14, 28, 42 and 56 post vaccination using H-based PPR bELISA. The peak adhesion time of the different gums was transient. PPR virus antibodies were detected in all immunized goats and sheep but not in unvaccinated control. The best percentage inhibition was recorded for Boswellia carteri-vaccine combination group at a ratio of 1:1. Administration of Boswellia carteri-PPR vaccine combination through intranasal or subcutaneous route, elicited similar antibody titre, implying that the intranasal route may be used as a non-invasive alternative delivery in PPR vaccination of small ruminants.

Haemonchus contortus transthyretin domain - containing protein (HcTTR): A promising vaccine candidate against Haemonchus contortus infection

Haemonchus contortus transthyretin domain-containing protein (HcTTR) with 136 amino acids belongs to a transthyretin-like (TTL) family member. In our previous study, it was reported that HcTTR was a novel antagonist of the goat cytokine Interleukin 4 (IL-4), and was involved in the regulation of host immune responses, implying that it might be applied for therapeutic strategies or vaccine development against the infection of H. contortus. Thus, the protective capacities of HcTTR against H. contortus infections via active and passive immunization trials were examined. For the passive protection trials, H. contortus-infected goats were intravenously immunized twice with 5 mg of total IgG containing anti-rHcTTR goat polyclonal antibodies. The results showed that the significant rates of reduction in egg shedding and worm burden was 58.12% and 64.61%, respectively, as compared with the positive control group. For the active protection trials, local goats were vaccinated twice with 500 μg of recombinant HcTTR to generate antigen-specific circulating antibodies, resulting in 63.7% reduction in eggs shedding and 66.4% reduction in worm burdens after H. contortus challenge. In both passive and active trials, the immunized goats displayed higher mucosal IgA levels and less anaemic compared to the challenged positive controls. Pen trials indicated that HcTTR generated partial immune protective effects against H. contortus challenge and it could be a promising vaccine candidate for development of effective strategy to control H. contortus.

Immunization of Goats with Recombinant Protein 14-3-3 Isoform 2(rHcftt-2) Induced Moderate Protection against Haemonchus contortus Challenge.

A previous study identified that isoform 2 (Hcftt-2) of the 14-3-3 protein of Haemonchus contortus (H. contortus) could suppress immune functions of goat peripheral blood mononuclear cells (PBMCs) and might be a potential vaccine target, as neutralization of the protein function may enhance anti-parasite immunity. In this research, the recombinant Hcftt-2 was evaluated for its immunoprotective efficacy against H. contortus infection in goats. Five experimental goats were immunized twice with rHcftt-2 along with Freund’s adjuvant. The five immunized goats and five nonimmunized goats (adjuvant only) were challenged with 5000 L3-stage H. contortus larvae after 14 days of second immunization. Five nonimmunized and uninfected goats (adjuvant only) were set as the uninfected group. A significant increase in the serum immunoglobin G(IgG) and serum IgA levels were identified in the rHcftt-2 immunized animals. The mean eggs per gram in feces (EPG) and the worm burdens of rHcftt-2 immunized group were reduced by 26.46% (p < 0.05) and 32.33%, respectively. In brief, immunization of goats with rHcftt-2 induced moderate protection against H. contortus challenge.

Recombinant fusion protein of Heparin-Binding Hemagglutinin Adhesin and Fibronectin Attachment Protein (rHBHA-FAP) of Mycobacterium avium subsp. paratuberculosis elicits a strong gamma interferon response in peripheral blood mononuclear cell culture

BackgroundMycobacterium avium subsp. paratuberculosis (MAP) is a causative agent of Johne’s disease in all ruminants worldwide. Economic problems in dairy cattle and sheep industries, public health concern, persistence of MAP in the environment and lack of effective vaccines mentioned necessity of research about various antigens to introduce as vaccine candidates. Based on MAP pathogenesis, it seems that research about the production of new recombinant proteins to stimulate cell-mediated immunity is helpful. This study describes successful expression and purification of a chimeric fusion protein which consists of Heparin-Binding Hemagglutinin Adhesin (HBHA) and high antigenic region of Fibronectin Attachment Protein (FAP-P). Triggered antigen-specific IFN-γ response of isolated PBMCs from immunized goats to rHBHA-FAP and all crude proteins of MAP (PPD), was measured by ELISA.ResultsSignificant increases were observed in the IFN-γ production level of peripheral blood mononuclear cells (PBMCs) stimulated by constructed chimeric protein from rHBHA-FAP and PPD vaccinated goats. Antigen-specific gamma interferon (IFN-γ) secretion in positive group (immunized by PPD) against rHBHA-FAP and test group (immunized by rHBHA-FAP) against PPD, also statistically insignificant rises between stimulation with rHBHA-FAP and PPD, suggested the potential and specificity of our chimeric protein to stimulate cell mediated immunity against MAP.ConclusionsCollectively, these results demonstrate that rHBHA-FAP elicits a strong IFN-γ production in PBMC culture. Therefore, further studies of the present product as a candidate vaccine in naturally infected animals should be conducted, to analyze its potential.

Biomarkers for Early Stages of Johne's Disease Infection and Immunization in Goats.

Background: Mycobacterium avium subsp. paratuberculosis (M. paratuberculosis) is the causative agent of Johne’s disease, a chronic enteric infection of ruminants. Infection occurs within the first few months of life but remains subclinical for an average of 2–5 years. Current diagnostics to detect early subclinical infections lack diagnostic sensitivity, which hinders disease control resulting in significant economic losses to the dairy industry worldwide. The pathophysiology of early infection with M. paratuberculosis is still not well understood and represents a key hurdle toward the development of better diagnostics.Methods: The present study employed a large-scale RNA-Sequencing technology to better understand early stages of M. paratuberculosis infection and immunization. Specifically, gene expression profiles of peripheral blood mononuclear cells (PBMCs) from infected or vaccinated goats were compared to controls.Results: When compared to the naïve control goats, we identified a large number of transcripts (N = 226, 1018, 1714) that were differentially expressed in the M. paratuberculosis-infected goats, goats vaccinated with live attenuated or inactivated vaccines. There were also 1133 differentially expressed (DE) transcripts between vaccinated goats and infected ones. Bioinformatics evaluation of the DE genes indicated the regulation of a large number of genes with immunity and inflammatory functions including IL-18BP, IFN-γ, IL-17A, NOS2, LIPG, and IL-22. Interestingly, a large number of goat genes (N = 667) were regulated whether live or inactivated vaccine were used. Some of the regulated genes (e.g., IL-17A, IFN-γ) continued its unique transcriptional profile up to 12 months post-challenge.Conclusion: Overall, transcriptome analysis of infected and/or immunized goats identified potential targets for developing early diagnostics for Johne’s disease and a potential approach to differentiate infected from vaccinated animals. A similar approach could be used to analyze later stages of Johne’s disease or other chronic infections.

Vaccination of goats with a combination Salmonella vector expressing four Brucella antigens (BLS, PrpA, Omp19, and SOD) confers protection against Brucella abortus infection.

Salmonella is an intracellular pathogen with a cellular infection mechanism similar to that of Brucella, making it a suitable choice for use in an anti-Brucella immune boost system. This study explores the efficacy of a Salmonella Typhimurium delivery-based combination vaccine for four heterologous Brucella antigens (Brucella lumazine synthase, proline racemase subunit A, outer-membrane protein 19, and Cu/Zn superoxide dismutase) targeting brucellosis in goats. We inoculated the attenuated Salmonella delivery-based vaccine combination subcutaneously at two different inoculation levels; 5 × 109 colony-forming unit (CFU)/mL (Group B) and 5 × 1010 CFU/mL (Group C) and challenged the inoculations with virulent Brucella abortus at 6 weeks post-immunization. Serum immunoglobulin G titers against individual antigens in Salmonella immunized goats (Group C) were significantly higher than those of the non-immunized goats (Group A) at 3 and 6 weeks after vaccination. Upon antigenic stimulation, interferon-γ from peripheral blood mononuclear cells was significantly elevated in Groups B and C compared to that in Group A. The immunized goats had a significantly higher level of protection as demonstrated by the low bacterial loads in most tissues from the goats challenged with B. abortus. Relative real-time polymerase chain reaction results revealed that the expression of Brucella antigens was lower in spleen, kidney, and lung of immunized goats than of non-immunized animals. Also, treatment with our combination vaccine ameliorated histopathological lesions induced by the Brucella infection. Overall, the Salmonella Typhimurium delivery-based combination vaccine was effective in delivering immunogenic Brucella proteins, making it potentially useful in protecting livestock from brucellosis.

Use of the mice passive protection test to evaluate the humoral response in goats vaccinated with Sterne 34F2 live spore vaccine

The Sterne live spore vaccine (34F2) is the most widely used veterinary vaccine against anthrax in animals. Antibody responses to several antigens of Bacillus anthracis have been described with a large focus on those against protective antigen (PA). The focus of this study was to evaluate the protective humoral immune response induced by the live spore anthrax vaccine in goats. Boer goats vaccinated twice (week 0 and week 12) with the Sterne live spore vaccine and naive goats were used to monitor the anti-PA and toxin neutralizing antibodies at week 4 and week 17 (after the second vaccine dose) post vaccination. A/J mice were passively immunized with different dilutions of sera from immune and naive goats and then challenged with spores of B. anthracis strain 34F2 to determine the protective capacity of the goat sera. The goat anti-PA ELISA titres indicated significant sero-conversion at week 17 after the second doses of vaccine (p = 0.009). Mice receiving undiluted sera from goats given two doses of vaccine (twice immunized) showed the highest protection (86%) with only 20% of mice receiving 1:1000 diluted sera surviving lethal challenge. The in vitro toxin neutralization assay (TNA) titres correlated to protection of passively immunized A/J mice against lethal infection with the vaccine strain Sterne 34F2 spores using immune goat sera up to a 1:10 dilution (rs ≥ 0.522, p = 0.046). This study suggests that the passive mouse protection model could be potentially used to evaluate the protective immune response in livestock animals vaccinated with the current live vaccine and new vaccines.

Effect of steroid hormones on the innate immune response induced by Staphylococcus aureus in the goat mammary gland.

The objective of this study was to compare the dynamics of innate immune components after intramammary infusion of Staphylococcus aureus (SA) under conditions of high oestrogen and high progesterone in goats. In one group ("E-group"), controlled internal drug release (CIDR) devices were inserted intravaginally from days -11 to -4. Prostaglandin F2α was administered immediately after removal of the CIDR device at day -3, and then oestradiol benzoate (E) was injected intramuscularly once a day from days -2 to 3. Heat-inactivated SA was then administered via intramammary infusion to the left udder at day 0, whilst only saline was infused to the right udder as a control. In a second group ("P-group"), CIDR devices were inserted intravaginally from days -3 to 7 and SA was infused at day 0 in the same way as in the E-group. The milk yield and the concentration of innate immune components (somatic cell count (SCC), lactoferrin (LF), S100A7 and goat ß-defensin 1 (GBD-1)) in the milk were measured. Milk yield decreased drastically in both SA and control udders in the E-group, whereas the P-group exhibited increased milk yield in both SA and control udders. SCC increased after SA infusion in both E- and P-groups, although it was higher in the E-group than in the P-group. There was no significant change in LF concentration in the E-group, but a decrease was observed in the P-group. Concentrations of S100A and GBD-1 were significantly increased after SA infusion in the E-group but not in the P-group. These results suggest that E enhances the innate immune response induced by SA in the goat mammary gland. This effect may be due to the reduction in milk yield and upregulation of innate immune components.

BoHV-4-based vector delivering Ebola virus surface glycoprotein.

BackgroundEbola virus (EBOV) is a Category A pathogen that is a member of Filoviridae family that causes hemorrhagic fever in humans and non-human primates. Unpredictable and devastating outbreaks of disease have recently occurred in Africa and current immunoprophylaxis and therapies are limited. The main limitation of working with pathogens like EBOV is the need for costly containment. To potentiate further and wider opportunity for EBOV prophylactics and therapies development, innovative approaches are necessary.MethodsIn the present study, an antigen delivery platform based on a recombinant bovine herpesvirus 4 (BoHV-4), delivering a synthetic EBOV glycoprotein (GP) gene sequence, BoHV-4-syEBOVgD106ΔTK, was generated.ResultsEBOV GP was abundantly expressed by BoHV-4-syEBOVgD106ΔTK transduced cells without decreasing viral replication. BoHV-4-syEBOVgD106ΔTK immunized goats produced high titers of anti-EBOV GP antibodies and conferred a long lasting (up to 6 months), detectable antibody response. Furthermore, no evidence of BoHV-4-syEBOVgD106ΔTK viremia and secondary localization was detected in any of the immunized animals.ConclusionsThe BoHV-4-based vector approach described here, represents: an alternative antigen delivery system for vaccination and a proof of principle study for anti-EBOV antibodies generation in goats for potential immunotherapy applications.Electronic supplementary materialThe online version of this article (doi:10.1186/s12967-016-1084-5) contains supplementary material, which is available to authorized users.

A study on prevalence of peste des petits ruminant (PPR) in goat at Bagmara upazilla at Rajshahi district in Bangladesh

A study was conducted at Bagmara upazilla under the district of Rajshahi in Bangladesh during the period of 2 month from 20 September 2015 to 14 November 2015 to determine the prevalence of PPR in goat. A total of 72 diseases cases were recorded randomly irrespective of age, sex and breed over the study period. In the present study the overall prevalence of PPR in goat was 27.78%. In black Bengal goat, the prevalence of PPR in different age group i.e. up to 6 month, 7-12 month, 13-19 month, above 19 month were 18.18%, 42.30%, 22.22% and 12.50%, respectively. Over all, female black bengal goat (46.42%) was more infected than male (12.00%). The prevalence of PPR in different age group i.e. 7-12 month, 13-19 month were 42.85% and 25.00%, respectively; and males (16.66%) and female (23.07%) of Jamunapari goat were recorder. In general, black bengal goats have the higher prevalence (30.18%) than Jamunapari goat (21.05%). The nonimmunized goat showed higher prevalence (38.77%) than immunized (4.34%) goat. All data were analyzed by using ?2 tests with P value at (P&lt;0.05) level of significance.Res. Agric., Livest. Fish.3(2): 339-344, August 2016

Intraperitoneally administered IgG from patients with amyotrophic lateral sclerosis or from an immune-mediated goat model increase the levels of TNF-α, IL-6, and IL-10 in the spinal cord and serum of mice.

BackgroundAmyotrophic lateral sclerosis (ALS) is a neurodegenerative disease that involves the selective loss of the upper and lower motor neurons (MNs). Neuroinflammation has been implicated in the pathogenesis of the sporadic form of the disease. We earlier developed immune-mediated animal models of ALS and demonstrated humoral and cellular immune reactions in the nervous system and in the sera of patients and animals. The accumulation of immunoglobulin G (IgG), an elevated intracellular level of calcium, ultrastructural alterations in the MNs, and activation of the microglia were noted in the spinal cord of ALS patients. Similar alterations developed in mice inoculated intraperitoneally with IgG from ALS patients or from an immune-mediated goat model.MethodsWe have now examined whether the intraperitoneal injection of mice with IgG from sporadic ALS patients or from immunized goats with the homogenate of the anterior horn of the bovine spinal cord is associated with changes in the pro-inflammatory (TNF-α and IL-6) and anti-inflammatory (IL-10) cytokines in the spinal cord and serum of the mice. The levels of cytokines were measured by ELISA.ResultsIntraperitoneally administered IgG from the ALS patients induced subclinical signs of MN disease, while the injection of IgG from immunized goats resulted in a severe respiratory dysfunction and limb paralysis 24 h after the injections. Significantly increased levels of TNF-α and IL-10 were detected in the spinal cord of the mice injected with the human ALS IgG. The level of IL-6 increased primarily in the serum. The IgG from the immunized goats induced highly significant increases in the levels of all three cytokines in the serum and the spinal cord of mice.ConclusionsOur earlier experiments had proved that when ALS IgG or IgG from immune-mediated animal models was inoculated into mice, it was taken up in the MNs and had the ability to initiate damage in them. The pathological process was paralleled by microglia recruitment and activation in the spinal cord. The present experiment revealed that these forms of IgG cause significant increases in certain cytokine levels locally in the spinal cord and in the serum of the inoculated mice. These results suggest that IgG directed to the MNs may be an initial element in the damage to the MNs both in human ALS and in its immune-mediated animal models.

Hydrolysis with Cucurbita ficifolia serine protease reduces antigenic response to bovine whey protein concentrate and αs-casein.

In the present study the effect of hydrolysis with non-commercial Cucurbita ficifolia serine protease on a reduction of the IgE and IgG binding capacity of whey protein concentrate and αs-casein was investigated. The intensity of the protein degradation was analyzed by the degree of hydrolysis, the free amino groups content and RP-HPLC. The ability to bind the antibodies by native proteins and their hydrolysates was determined using a competitive ELISA test. Deep hydrolysis contributed to a significant reduction of immunoreactive epitopes present in WPC. In the case of IgE and IgG present in the serum pool of children with CMA, the lowest binding capacity was detected in the 24 h WPC hydrolysate, where the inhibition of the reaction with native WPC was ≤23 and ≤60 %, respectively. The analysis of the IgG reactivity in the antiserum of the immunized goat showed that the lowest antibody binding capacity was exhibited also by 24 h WPC hydrolysate at a concentration of 1000 μg/ml where the inhibition of the reaction with nWPC was ≤47 %. One-hour hydrolysis of α-casein was sufficient to significant reduction of the protein antigenicity, while the longer time (5 h) of hydrolysis probably lead to the appearance of new epitopes reactive with polyclonal.

Induction of neutralizing antibodies specific for the envelope proteins of the koala retrovirus by immunization with recombinant proteins or with DNA

BackgroundThe koala retrovirus (KoRV) is the result of a transspecies transmission of a gammaretrovirus with fatal consequences for the new host. Like many retroviruses, KoRV induces lymphoma, leukemia and an immunodeficiency that is associated with opportunistic infections in the virus-infected animals. We recently reported the induction of neutralizing antibodies by immunization with the recombinant ectodomain of the transmembrane envelope protein p15E of KoRV. Since the neutralization titers of the p15E-specific sera were only moderate, we investigated the use of the surface envelope protein gp70 to induce neutralizing antibodies.FindingsWe immunized rats and goats with the recombinant gp70 protein of the KoRV, an unglycosylated protein of 52kD (rgp70/p52) or with the corresponding DNA. In parallel we immunized with recombinant rp15E or with a combination of rp15E and rgp70/p52. In all cases binding and neutralizing antibodies were induced. The gp70-specific sera had titers of neutralizing antibodies that were 15-fold higher than the p15E-specific sera. Combining rp15E and rgp70/p52 did not significantly increase neutralizing titers compared to rgp70/p52 alone. High titers of neutralizing antibodies specific for gp70 were also induced by immunization with DNA. Since KoRV and PERV are closely related, we investigated cross-neutralization of the antisera. The antisera against p15E and gp70 of PERV and KoRV inhibited infection by both viruses.ConclusionThe envelope proteins of the KoRV may therefore form the basis of an effective preventive vaccine to protect uninfected koalas from infection and possibly an immunotherapeutic treatment for those already infected.Electronic supplementary materialThe online version of this article (doi:10.1186/s12985-015-0296-2) contains supplementary material, which is available to authorized users.

Immunization with envelope proteins of the KoRV as a basis for a preventive vaccine

The rapid spread of the koala retrovirus (KoRV) in Australia and in international zoos calls for effective counter measures. As is the case with the human immunodeficiency virus (HIV) epidemic, a preventive vaccine is urgently needed. Vaccines inducing neutralizing antibodies are a good way to prevent retrovirus infections. Although for HIV there is still no effective vaccine available, commercial vaccines protecting cats from disease caused by the feline leukemia virus (FeLV) already exist and have been proven effective. KoRV is a retrovirus more closely related to FeLV than to HIV. Immunizing different species (rats, goats, hamsters, guinea pigs, mice, cats) with the transmembrane (TM) and surface (SU) envelope proteins of FeLV, as well as of the porcine endogenous virus (PERV) we always obtained neutralizing antibodies. PERV is also closely related to the KoRV. Based on the immunization studies with the envelope proteins of FeLV and PERV, we cloned and expressed the corresponding envelope proteins of the KoRV and immunized goats and rats. In all cases we obtained antibodies neutralizing the KoRV. However this does not mean that neutralizing antibodies will be obtained when immunizing koalas (Phascolarctos cinereus) with the envelope proteins of the KoRV or immunizing pigs with the envelope proteins of PERV. Therefore, koalas should be immunized with KoRV envelope antigens to determine whether neutralizing antibodies are induced and if so, whether such antibodies are able to protect healthy animals from infection. Furthermore, whether immunization with these antigens has a therapeutic effect on animals already infected with KoRV should be investigated. If Chlamydia infection of koalas is an opportunistic infection made possible by KoRV-induced immunodeficiency, immunization against KoRV will also protect animals from Chlamydia infection. Denner, Joachim. 2014. Immunization with envelope proteins of the KoRV as a basis for a preventive vaccine. In The Koala and its Retroviruses: Implications for Sustainability and Survival, ed. Geoffrey W. Pye, Rebecca N. Johnson and Alex D. Greenwood. Technical Reports of the Australian Museum, Online 24: 71–77. Infection of koalas with the KoRV, and infection of humans with HIV-1 leading to AIDS. Retroviruses have long been known to be capable of infecting new host species by transspecies transmission. Interest in this subject has been boosted by the finding that the human immunodeficiency viruses (HIV-1 and HIV-2) are the product of such a transspecies transmission (Gao 1994,1999) and by recent concerns over the potential transmission of PERVs after xenotransplantation of pig organs into humans (Denner & Tonjes, 2012). The koala retrovirus (KoRV) is the result of such a transspecies transmission which is even associated with endogenization of the virus into the germ line of the animals (Hanger et al., 2000; Denner & Young, 2013). The KoRV is closely related to the gibbon ape leukemia virus (GaLV), which however remained exogenous in gibbons (Hanger et al., 2000). Both are related to endogenous retroviruses of South Eastern Asian mice, (Martin et al., 1999) and bats, (Cui et al., 2012a,b) however the origin and the transmission routes are still unknown. Retroviruses are known to induce tumors and immunodeficiencies and HIV is the most prominent retrovirus induc ing an acquired immunodeficiency syndrome. Although HIV, a lenti(retro)virus, and the KoRV, a gammaretrovirus, are not closely related, the clinical picture of the syndrome

Protective immune response of live attenuated thermo-adapted peste des petits ruminants vaccine in goats.

Virulent isolate of peste des petits ruminants virus (PPRV) of Indian origin (PPRV Jhansi 2003) initially adapted in Vero cells was further propagated in thermo-adapted (Ta) Vero cells grown at 40°C for attaining thermo-adaption and attenuation of virus for development of Ta vaccine against PPR in goats and sheep. The virus was attenuated up to 50 passages in Ta Vero cells, at which, the virus was found sterile, innocuous in mice and guinea pigs and safe in seronegative goats and sheep. The developed vaccine was tested for its immunogenicity in goats and sheep by subcutaneous inoculation of 100 TCID50 (0.1field dose), 10(3) TCID50 (one field dose) and 10(5) TCID50 (100 field doses) of the attenuated virus along with controls as per OIE described protocols for PPR vaccine testing and were assessed for PPRV-specific antibodies 7-28days post vaccination (dpv) by PPR competitive ELISA and serum neutralization tests. The PPRV antibodies were detected in all immunized goats and sheep and goats were protective when challenged with virulent PPRV at 28th dpv along with controls for potency testing of the vaccine. The attenuated vaccine did not induce any adverse reaction at high dose (10(5) TCID50) in goats and sheep and provided complete protection even at low dose (10(2) TCID50) in goats when challenged with virulent virus. There was no shedding and horizontal transmission of the attenuated virus to in-contact controls. The results indicate that the developed PPR Ta attenuated virus is innocuous, safe, immunogenic and potent or efficacious vaccine candidate alternative to the existing vaccines for the protection of goats and sheep against PPR in the tropical countries like India.

Combination therapy of anti-cancer bioactive peptide with Cisplatin decreases chemotherapy dosing and toxicity to improve the quality of life in xenograft nude mice bearing human gastric cancer

BackgroundA great challenge of cancer chemotherapy is to eliminate cancer cells and concurrently maintain the quality of life (QOL) for cancer patients. Previously, we identified a novel anti-cancer bioactive peptide (ACBP), a peptide induced in goat spleen or liver following immunization with human gastric cancer protein extract. ACBP alone exhibited anti-tumor activity without measurable side effects. Thus, we hypothesize that ACBP and combined chemotherapy could improve the efficacy of treatment and lead to a better QOL.ResultsIn this study, ACBP was isolated and purified from immunized goat liver, and designated as ACBP-L. The anti-tumor activity was investigated in a previously untested human gastric cancer MGC-803 cell line and tumor model. ACBP-L inhibited cell proliferation in vitro in a dose and time dependent manner, titrated by MTT assay. The effect of ACBP-L on cell morphology was observed through light and scanning electron microscopy. In vivo ACBP-L alone significantly inhibited MGC-803 tumor growth in a xenograft nude mouse model without measurable side effects. Treatment with the full dosage of Cisplatin alone (5 mg/kg every 5 days) strongly suppressed tumor growth. However, the QOL in these mice had been significantly affected when measured by food intakes and body weight. The combinatory regiment of ACBP-L with a fewer doses of Cisplatin (5 mg/kg every 10 days) resulted in a similar anti-tumor activity with improved QOL. 18F-FDG PET/CT scan was used to examine the biological activity in tumors of live animals and indicated the consistent treatment effects. The tumor tissues were harvested after treatment, and ACBP-L and Cisplatin treatment suppressed Bcl-2, and induced Bax, Caspase 3, and Caspase 8 molecules as detected by RT-PCR and immunohistochemistry. The combinatory regiment induced stronger Bax and Caspase 8 protein expression.ConclusionOur current finding in this gastric cancer xenograft animal model demonstrated that ACBP-L could lower Cisplatin dose to achieve a similar anti-tumor efficacy as the higher dose of Cisplatin alone, through enhanced modulation of apoptotic molecules. This newly developed combination regiment improved QOL in tumor bearing hosts, which could lead to clinical investigation for the new strategy of combination therapy.

Sodium dodecyl sulfate reduces bacterial contamination in goat colostrum without negative effects on immune passive transfer in goat kids

To investigate the use of sodium dodecyl sulfate (SDS) as a biocide on goat colostrum, 2 experiments were performed. In the first, 20 goat colostrum samples were divided into 3 aliquots. A different treatment was performed on each aliquot: pasteurization (56°C, 30min) or addition of SDS to a final concentration of either 0.1 or 1% (36°C, 10min). Immunoglobulin G and colony-forming units were evaluated before and after treatment. Both pasteurization and treatment with 1% SDS significantly reduced the colony-forming units in colostrum. Treatment with 0.1% SDS was not effective at reducing the colony-forming units in colostrum. The IgG concentration of pasteurized colostrum was significantly lower than that of untreated colostrum, whereas treatment with 1% SDS did not affect the colostrum IgG concentration. In the second experiment, the effects of SDS colostrum treatment on immune passive transfer were evaluated. Forty goat kids were fed either refrigerated colostrum or colostrum treated with 1% SDS twice daily for 2 d. Blood samples were obtained at birth and every day for 5 d. IgG, IgM, and IgA were measured in blood serum to monitor the passive immune transfer process. Creatinine, glucose, total cholesterol, blood urea nitrogen, bilirubin, and aspartate transaminase were also monitored to evaluate the health of kids. No differences in serum IgG, IgM, IgA, creatinine, glucose, total cholesterol, blood urea nitrogen, bilirubin, or aspartate transaminase levels were observed between groups. Our findings indicate that SDS is an efficient colostrum biocide that, unlike pasteurization, does not affect immune passive transfer or goat kid health.