A novel modified-indirect ELISA based on spherical body protein 4 for detecting antibody during acute and long-term infections with diverse Babesia bovis strains

Cattle sera positive by the RAP-1-based cELISA but negative by the SBP4-based MI-ELISA and IFA had negative results by Western blot analysis, suggesting possible false positive results in the cELISA. A. Molecular weight marker (48 to 180 Kd), B. K42-#21, C. W31-#Y-3, D. W31-#Y-11, E. W31-#0-3, F. W31-#Y-9, G. W31-#0-9, H. W31-#Y-10, I. W31-#Y-15, J. P21-#224, K. positive control serum with a band at 75kd representing B. bovis RAP-1 protein, J. negative control serum. Figure S2. Technical difference between the modified indirect ELISA and conventional indirect ELISA using rGST-SBP4 was illustrated in this figure. (DOCX 645 kb

Environmental performance assessment of european countries

The European Union(EU) has been promoting an integrated approach to climate protection and energy policy, through a set of key objectives for 2020, 2030 and 2050, linking Europe’s green agenda with its need for energy security and competitiveness.This paper aims to evaluate the environmental efficiency of European Countries from 2010 to 2015 towards the set targets, through a Data Envelopment Analysis (DEA) model.The DEA model assesses the ability of each country in minimizing current resources while maximizing the gross domestic product and minimizing undesirable outputs,such as GHG emissions.The DEA model is based on directional distance Function, imposing weak disposability for the undesirable output. Results obtained show that globally, in the period under analysis, the EU has increased its environmental efficiency which is consistent with the analysis of the indicators of the 2020 climate and energy package.info:eu-repo/semantics/publishedVersio

Recognition of Highly Diverse Type-1 and -2 Porcine Reproductive and Respiratory Syndrome Viruses (PRRSVs) by T-Lymphocytes Induced in Pigs after Experimental Infection with a Type-2 PRRSV Strain

<div><p>Background/Aim</p><p>Live attenuated vaccines confer partial protection in pigs before the appearance of neutralizing antibodies, suggesting the contribution of cell-mediated immunity (CMI). However, PRRSV-specific T-lymphocyte responses and protective mechanisms need to be further defined. To this end, the hypothesis was tested that PRRSV-specific T-lymphocytes induced by exposure to type-2 PRRSV can recognize diverse isolates.</p><p>Methods</p><p>An IFN-gamma ELISpot assay was used to enumerate PRRSV-specific T-lymphocytes from PRRSV_SD23983-infected gilts and piglets born after in utero infection against 12 serologically and genetically distinct type-1 and -2 PRRSV isolates. The IFN-gamma ELISpot assay using synthetic peptides spanning all open reading frames of PRRSV_SD23983 was utilized to localize epitopes recognized by T-lymphocytes. Virus neutralization tests were carried out using the challenge strain (type-2 PRRSV_SD23983) and another strain (type-2 PRRSV_VR2332) with high genetic similarity to evaluate cross-reactivity of neutralizing antibodies in gilts after PRRSV_SD23983 infection.</p><p>Results</p><p>At 72 days post infection, T-lymphocytes from one of three PRRSV_SD23983-infected gilts recognized all 12 diverse PRRSV isolates, while T-lymphocytes from the other two gilts recognized all but one isolate. Furthermore, five of nine 14-day-old piglets infected in utero with PRRSV_SD23983 had broadly reactive T-lymphocytes, including one piglet that recognized all 12 isolates. Overlapping peptides encompassing all open reading frames of PRRSV_SD23983 were used to identify ≥28 peptides with T-lymphocyte epitopes from 10 viral proteins. This included one peptide from the M protein that was recognized by T-lymphocytes from all three gilts representing two completely mismatched MHC haplotypes. In contrast to the broadly reactive T-lymphocytes, neutralizing antibody responses were specific to the infecting PRRSV_SD23983 isolate.</p><p>Conclusion</p><p>These results demonstrated that T-lymphocytes recognizing antigenically and genetically diverse isolates were induced by infection with a type 2 PRRSV strain (SD23983). If these reponses have cytotoxic or other protective functions, they may help overcome the suboptimal heterologous protection conferred by conventional vaccines.</p></div

T-lymphocyte responses from 14-day-old piglets infected in utero with PRRSV_SD23983 in an IFN-gamma ELISpot assay against phytohemagglutin (PHA) and various type-1 and type-2 PRRSV isolates.

<p>T-lymphocyte responses from 14-day-old piglets infected in utero with PRRSV_SD23983 in an IFN-gamma ELISpot assay against phytohemagglutin (PHA) and various type-1 and type-2 PRRSV isolates.</p

T-lymphocyte responses from one-year-old gilts infected with PRRSV_SD23983 at 44 and 72 dpi in an IFN-gamma ELISpot assay against various type-1 and type-2 PRRSV isolates.

<p>T-lymphocyte responses from one-year-old gilts infected with PRRSV_SD23983 at 44 and 72 dpi in an IFN-gamma ELISpot assay against various type-1 and type-2 PRRSV isolates.</p

Peptides containing epitopes recognized by T-lymphocytes from three gilts infected with PRRSV_SD23983.

<p>Peptides containing epitopes recognized by T-lymphocytes from three gilts infected with PRRSV_SD23983.</p

Mapping of T-lymphocyte epitopes on PRRSV proteins using overlapping peptide pools encompassing all open reading frames of PRRSV_SD23983.

<p>Mapping of T-lymphocyte epitopes on PRRSV proteins using overlapping peptide pools encompassing all open reading frames of PRRSV_SD23983.</p

Comprehensive Immunological Evaluation Reveals Surprisingly Few Differences between Elite Controller and Progressor Mamu-B*17-Positive Simian Immunodeficiency Virus-Infected Rhesus Macaques▿

The association between particular major histocompatibility complex class I (MHC-I) alleles and control of human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV) replication implies that certain CD8+ T-lymphocyte (CD8-TL) responses are better able than others to control viral replication in vivo. However, possession of favorable alleles does not guarantee improved prognosis or viral control. In rhesus macaques, the MHC-I allele Mamu-B*17 is correlated with reduced viremia and is overrepresented in macaques that control SIVmac239, termed elite controllers (ECs). However, there is so far no mechanistic explanation for this phenomenon. Here we show that the chronic-phase Mamu-B*17-restricted repertoire is focused primarily against just five epitopes—VifHW8, EnvFW9, NefIW9, NefMW9, and envARFcRW9—in both ECs and progressors. Interestingly, Mamu-B*17-restricted CD8-TL do not target epitopes in Gag. CD8-TL escape variation occurred in all targeted Mamu-B*17-restricted epitopes. However, recognition of escape variant peptides was commonly observed in both ECs and progressors. Wild-type sequences in the VifHW8 epitope tended to be conserved in ECs, but there was no evidence that this enhances viral control. In fact, no consistent differences were detected between ECs and progressors in any measured parameter. Our data suggest that the narrowly focused Mamu-B*17-restricted repertoire suppresses virus replication and drives viral evolution. It is, however, insufficient in the majority of individuals that express the “protective” Mamu-B*17 molecule. Most importantly, our data indicate that the important differences between Mamu-B*17-positive ECs and progressors are not readily discernible using standard assays to measure immune responses

Gag-Specific CD8 +

CD8(+) T cells are a key focus of vaccine development efforts for HIV. However, there is no clear consensus as to which of the nine HIV proteins should be used for vaccination. The early proteins Tat, Rev, and Nef may be better CD8(+) T cell targets than the late-expressed structural proteins Gag, Pol, and Env. In this study, we show that Gag-specific CD8(+) T cells recognize infected CD4(+) T lymphocytes as early as 2 h postinfection, before proviral DNA integration, viral protein synthesis, and Nef-mediated MHC class I down-regulation. Additionally, the number of Gag epitopes recognized by CD8(+) T cells was significantly associated with lower viremia (p = 0.0017) in SIV-infected rhesus macaques. These results suggest that HIV vaccines should focus CD8(+) T cell responses on Gag

The Major Histocompatibility Complex Class II Alleles Mamu-DRB1*1003 and -DRB1*0306 Are Enriched in a Cohort of Simian Immunodeficiency Virus-Infected Rhesus Macaque Elite Controllers▿

The role of CD4+ T cells in the control of human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV) replication is not well understood. Even though strong HIV- and SIV-specific CD4+ T-cell responses have been detected in individuals that control viral replication, major histocompatibility complex class II (MHC-II) molecules have not been definitively linked with slow disease progression. In a cohort of 196 SIVmac239-infected Indian rhesus macaques, a group of macaques controlled viral replication to less than 1,000 viral RNA copies/ml. These elite controllers (ECs) mounted a broad SIV-specific CD4+ T-cell response. Here, we describe five macaque MHC-II alleles (Mamu-DRB*w606, -DRB*w2104, -DRB1*0306, -DRB1*1003, and -DPB1*06) that restricted six SIV-specific CD4+ T-cell epitopes in ECs and report the first association between specific MHC-II alleles and elite control. Interestingly, the macaque MHC-II alleles, Mamu-DRB1*1003 and -DRB1*0306, were enriched in this EC group (P values of 0.02 and 0.05, respectively). Additionally, Mamu-B*17-positive SIV-infected rhesus macaques that also expressed these two MHC-II alleles had significantly lower viral loads than Mamu-B*17-positive animals that did not express Mamu-DRB1*1003 and -DRB1*0306 (P value of <0.0001). The study of MHC-II alleles in macaques that control viral replication could improve our understanding of the role of CD4+ T cells in suppressing HIV/SIV replication and further our understanding of HIV vaccine design