The highly pathogenic Avian Influenza subtype H5N1 can lead to 100 % mortality in chickens. The main issue in prevention of H5N1 is the development of efficient poultry vaccines. Influenza haemagglutinin (HA) derived recombinant polypeptides would not elicit an immune response against internal viral proteins. Thus HA polypeptide use facilitates differentiation between infected and vaccinated animals (DIVA). Serological tests using recombinant immune-dominant proteins devoid of non-specific moieties present in whole cell preparations might have higher sensitivity and specificity. In the present study, four non-overlapping sequences of different functional domains of influenza A virus subtype H5 virus (A / Thailand / 1 (Kan-1) / 2004) designated P1, P2, P5 and rHA1 were cloned and expressed in Pichia pastoris for vaccination and diagnosis purposes. The four polypeptides were expressed successfully in P. pastoris using peptone methanol (1 % (w/v) yeast extract, 2 % (w/v) peptone, 2 % (v/v) methanol). P1, P2 and rHA1 polypeptides were purified using nickel affinity chromatography, whereas, P5 was purified using lectin affinity chromatography. Correct expression was analysed by SDS-PAGE and western blot, glycosylation analysis and MALDI-TOF.The immune responses of P1, P2 and rHA1 polypeptides were assessed in BALB/C mice. To enhance antibody response, recombinant polypeptides were mixed with the Gerbu adjuvant and injected subcutaneously. Vaccination of mice induced high subtype specific antibody titres in mice as analysed by Elisa (using recombinant antigens or whole H5N1 antigen) and Immunofluorescence assay (IFA) performed on Vero cells infected with H5 (A / Thailand / 1 (Kan-1) / 2004). The immunogenicity of P1, P2, P5 and rHA1 polypeptides was determined in commercial layer chickens. Results showed that P1, P2 and rHA1 polypeptides induced high subtype specific antibody titres in chickens as analysed by Elisa (using recombinant antigens or whole H5N1 antigen), IFA (performed on Vero cells infected with H5N1 A / Thailand / 1 (Kan-1) / 2004) and microneutralization test (µNT). However, P5 polypeptide was not immunogenic in chickens. Neutralizing antibodies could be detected in chicken sera immunized with P1, P2 and rHA1 polypeptides as analyzed with microneutralization test. IgY was analysed in egg yolk of chickens immunized with recombinant polypeptides. The IgY of chicken immunized with P1 and rHA1, transferred to the egg yolk was proportional to maternal serum IgY. However, IgY could not be detected in egg yolk of chickens immunized with P2 and P5 recombinant polypeptides. The more immunogenic polypeptides P1 and rHA1 were used in an recombinant Elisa (rElisa) for detection of influenza A subtype H5 in chickens and duck sera.The optimal antigen for the concentrations of rHA1, P1 was 50 ng / well, 50 ng / well. Analysis of 25 positive sera and 25 negative sera to H5 antibodies revealed that, the sensitivity of Western blot, whole H5N1 Elisa, agar gel immunodiffusion test (AGID), P1-Elisa and rHA1-Elisa was 100 %, 100 %, 52 %, 80 % and 100 %, respectively, while the specificity was 100 %, 100 %, 100 %, 72 %, and 100 %, respectively. Moreover, duck sera, with haemagglutination inhibiting titer ranged from 4 - 8 log2, were tested positive by rHA1 Elisa compared with negative duck sera. Further analysis of 179 serum samples with rHA1-Elisa in comparison with haemagglutination inhibition (HI) and commercial Elisa proved to be highly sensitive and specific. The agreement ratio between rElisa and HI was 84.9 % and between commercial Elisa (Flock check) and HI was 76.5 %. In conclusion, P. pastoris may allow development of an effective recombinant influenza vaccine based on truncated sequences of HA that might provide broader protection against H5 influenza viruses. The possibilities to use rHA1, P1 and P5 recombinant polypeptides as a vaccine against H5 influenza should be further studied. Also our study demonstrates the potential utility of recombinant Elisa as a tool for improvement of serological diagnosis of influenza A subtype H5 in chickens and ducks. / Die hochpathogene aviäre Influenza des Subtyps H5N1 erreicht beim Ausbruch von Infektionen in Nutzgeflügelbeständen Mortalitätsraten von bis zu 100 %. Effektive und kostengünstige Impfstoffe werden benötigt, die möglichst auch eine Differenzierung zwischen geimpften Tieren und mit Wild-Virus infizierten Tieren zulassen. In diesem Zusammenhang könnten Peptid-Vakzine eine mögliche Alternative zu den herkömmlichen Impfstoffen darstellen, bei denen unter Verwendung des Vollvirus Antikörper gegen mehrere Virusproteine induziert werden. Außerdem, könnten rekombinante Antigene in serologischen Tests zur Diagnose von H5 Virus in Nutzgeflügel eingesetzt werden. Von dem Einsatz spezifischer rekombinanter Antigene ist eine Verbesserung der Serodiagnostik zu erwarten. In dieser Arbeit, wurden vier verkürzte Sequenzen des Hämagglutinins (P1, P2, P5 und rHA1) von Subtyp H5 (A / Thailand / 1 (Kan-1) / 2004) rekombinant in Pichia Pastoris exprimiert. Dazu erfolgten zunächst eine Klonierung in der Expressionsvektor pAOX und die Transformation von Pichia Pastoris. Die Expression wurde durch Methanol induziert. Der Nachweis der rekombinanten Fusionspeptiden mit C-terminalen Histidin-Tag erfolgte durch SDS-PAGE, Western Blot, Glycolysierungsanalyse, und MALDI-TOF. Der Histidin-Tag ermöglichte die Reinigung von P1, P2 und rHA1 mit Metall-Affinitätschromatographie. Polypeptid P5 hingegen wurde mittels Lectin-Affinitäts- chromatographie gereinigt. Balb/c Mäuse wurden mit Polypeptid P1, P2 bzw. rHA1, versetzt mit Gerbu Adjuvans, immunisiert. Zur Untersuchung der Immunantwort wurden die murinen Seren mittels Elisa (unter Verwendung rekombinanter Antigene oder Voll-H5N1 Antigen) sowie IFA (durchgeführt in Vero- Zellen infiziert mit A / Thailand / 1 (Kan-1) / 2004) analysiert. Dabei wurde die präferentielle Induktion von H5-spezifischen Antikörpern detektiert. Die Immunogenität der P1, P2, P5 und rHA1-Polypeptide wurde in kommerziellen Legehennen bestimmt. Seren wurden mit ELISA, IFA, und Mikroneutralizationstest (μNT) analysiert. Die ELISA-Ergebnisse zeigten, dass die Polypeptide P1, P2 und rHA1 hohe Subtyp-spezifische Antikörpertiter in Hühnern induzierten. Im µNT konnte nur ein niedriger neutralisierender Antikörpertiter nachgewiesen werden. Das P5- Polypeptid ist bei Hühnern nicht immunogen. Im Eigelb von Hühnern, die mit den rekombinanten Polypeptiden P1 und rHA1 immunisiert wurden, konnten H5-spezifische IgY Antikörper detektiert werden. Hühner, die mit P2 und P5 immunisiert wurden, zeigten keine IgY im Eigelb. Die rekombinanten Antigene P1 und rHA1 wurden im ELISA auf ihre potenzielle Eignung für die Serodiagnostik untersucht. Die optimale Antigenkonzentration war 50 ng / well. Die serologische Analyse von 25 positiven und 25 negativen Seren auf Antikörper gegen H5 zeigte, dass Sensitivität und Spezifität von Western Blot, Voll-H5N1 ELISA und rHA1-ELISA bei jeweils 100 % lagen. Bei Agargel- Immunodiffusiontest (AGID) lagen Sensitivität und Spezifität bei 52 % und 100 %, während im P1-Elisa lediglich eine Sensitivität von 80 % und eine Spezifität von 72 % erreicht wurden. Somit eignet sich rHA1 für die Anwendung in der Serodiagnostik. Bei der serologischen Untersuchung von 175 Hühnerseren wurde eine Überbestimmung zwischen rHA1-ELISA und Hämagglutinationshemmungstest (HAI) 84.9 % festgestellt, während diese zwischen dem kommerziellen ELISA (Flock Check) und HAI 76.5 % betrug. Die Ergebnisse zeigten, dass das Expressionssystem P. pastoris als Produktionssystem rekombinanter Antigene für die Serodiagnostik von H5 Influenza geeignet ist. Challenge-Versuche sind nötig, um die Eignung von rekombinanten Antigenen als möglichen Impfstoff gegen H5 Influenza zu untersuchen.

Zöliakie ist eine immunologisch vermittelte Erkrankung bei der die Dünndarm-schleimhaut durch das in zahlreichen Getreidesorten vorkommende Klebereiweiß Gliadin geschädigt wird. Dabei wird die typische Architektur der Mukosa zerstört und imponiert histologisch als Zottenatrophie. In Folge dessen zeigen Betroffene Mangelerscheinungen und Verdauungsbeschwerden sowie zahlreiche extra-intestinale, atypische Symptome. Bei Kindern können zusätzlich gravierende Wachstums- und Entwicklungsstörungen auftreten. Die Therapie besteht in einer lebenslangen glutenfreien Diät. Die Diagnostik der Erkrankung basiert auf vier Säulen: Neben der Beurteilung der klinischen Symptomatik werden zöliakie-typische Antikörper nachgewiesen, welche bei hoher Konzentration die Indikation zur Biopsie darstellen. Die bioptische Untersuchung mit anschließendem histolo-gischem Nachweis der Zottenatrophie stellt den Goldstandard der Diagnostik dar und wird durch die Besserung der klinischen Symptomatik unter glutenfreier Diät gestützt. Bei der serologischen Untersuchung haben Antikörper gegen natives Gliadin auf Grund niedriger diagnostischer Genauigkeit an Bedeutung verloren. Sie wurden durch die Bestimmung von Autoantikörpern gegen die Gewebstransglutaminase abgelöst, die eine höhere Sensitivität und Spezifität aufweisen. Im Jahr 2000 konn-te jedoch gezeigt werden, dass sich Antikörper von Zöliakiepatienten an Gliadin-peptide besser nach selektiver Deamidierung (Austausch der Aminosäure Glutamin durch Glutaminsäure) binden. Darauf aufbauend entwickelte die Firma INOVA Diagnostics im Jahr 2006 erst-mals einen ELISA zur Zöliakiediagnostik mit synthetisch hergestellten, deamidier-ten Gliadinpeptiden (DGP) als Antigen. Zu Beginn unserer Arbeit existierten keine Veröffentlichungen zur diagnostischen Genauigkeit dieser Tests bei Kindern und nur eine Veröffentlichung, die zwei der ELISA an Seren von erwachsenen Patienten untersuchte. Bei Kindern ist es jedoch besonders wichtig, durch die Antikörperbestimmung eine hohe Diagnosesicher-heit zu erlangen, weil für sie die Biopsie eine große Belastung darstellt und eine nicht erkannte Zöliakieerkrankung zu schwerwiegenden Entwicklungsstörungen führen kann. Aus diesem Grund soll die vorliegende Studie den Nutzen dieser neuen ELISA in der Diagnostik der Zöliakie im Kindesalter evaluieren. Es wurden dazu insgesamt 340 Seren von bioptisch bestätigten Zöliakiepatienten und Kontrollen, bei denen die Erkrankung histologisch ausgeschlossen wurde, gesammelt, verblindet und retrospektiv analysiert. Dabei wurden vier verschiede-ne ELISA der Firma INOVA Diagnostics eingesetzt: drei ELISA mit DGP als An-tigen sowie ein weiterer ELISA, in dem DGP mit Gewebstransglutaminase kombiniert war. Es wurden folgende Erkenntnisse gewonnen: 1. Alle vier ELISA eignen sich zur Diagnostik von Zöliakie bei Kindern und weisen eine hohe Trennschärfe auf. Die Fläche unter der Receiver Operating Characteristic (ROC)-Kurve ist für alle vier Tests größer als 0,96. Mit den Tests kann somit die Indikation zur bioptischen Untersuchung mit großer Sicherheit gestellt werden. 2. Die Bestimmung der Antikörper gegen DGP ist der Antikörperbestimmung gegen natives Gliadin überlegen. Die DGP-Tests weisen eine signifikant größe-re Fläche unter der ROC-Kurve als die Tests auf Antikörper gegen natives Gli-adin auf. Die Bestimmung der Antikörper gegen DGP ist der Bestimmung von Antikörpern gegen Gewebstransglutaminase nicht unterlegen. Die Flächen un-ter den ROC-Kurven unterscheiden sich nicht signifikant voneinander. 3. Überraschenderweise zeigt die IgG-Klasse der DGP eine signifikant höhere diagnostische Genauigkeit als die IgA-Klasse. Somit ist auch bei Patienten mit IgA-Mangel eine sichere Diagnosestellung gegeben und es kann auf die gene-relle Bestimmung des Gesamt-IgA verzichtet werden. 4. Der DGP-IgG-Test weist von den vier validierten Antikörpertests bei einer Sensitivität von 100 % die höchste Spezifität (90 %) und bei einer Spezifität von 100 % die höchste Sensitivität (64 %) auf. 5. Durch den kombinierten Test mit zwei Antigenen und den gleichzeitigen Nachweis von IgA und IgG in einem ELISA (tTG/DGP-Screen-Test) lassen sich Kosten und Zeit sparen. Dieser kombinierte Test weist die höchste diagnosti-sche Genauigkeit der untersuchten DGP-Tests auf. 6. Durch Angabe von Likelihood Ratios und mittels grafischer Darstellung der Posttest-Wahrscheinlichkeit in Abhängigkeit der Antikörperkonzentration und Prävalenz können den behandelnden Ärzten wertvolle Informationen zur Di-agnosesicherheit eines einzelnen Testergebnisses vermittelt werden. Mit den evaluierten DGP-Tests stehen neue und zuverlässige ELISA zur Diagnos-tik der Zöliakie im Kindesalter zur Verfügung. Sie weisen eine hohe diagnostische Sicherheit auf und unterscheiden sicher zwischen Gesunden und Zöliakiepatien-ten. Die Indikation für eine Dünndarmbiopsie kann mit Hilfe der DGP-Tests sehr sicher gestellt werden. Ob darüber hinaus auf der Basis sehr hoher Antikörper-konzentrationen ohne histologische Untersuchung die Diagnose Zöliakie ausrei-chend sicher gestellt werden kann oder bei sehr niedrigen Konzentrationen auf einen bioptischen Ausschluss verzichtet werden kann, soll in einer weiteren be-reits in Planung befindlichen prospektiven Studie geklärt werden. Zusätzlich sollte die Diagnosestellung durch DGP-Tests bei besonders jungen Kindern in prospektiven Studien untersucht werden. Die vorliegende Arbeit legt gemeinsam mit anderen Untersuchungen den Grundstein dafür.

Legeleistungseinbußen – vor allem mit verminderter Eischalenqualität – stellen in einem Legehennenbetrieb hohe wirtschaftliche Verluste dar. Impfungen gegen entsprechende Erreger, u.a. gegen das aviäre Metapneumovirus (aMPV), sind daher weit verbreitet. AMPV ist seit den 70er Jahren als Auslöser der Rhinotracheitis der Puten (Turkey Rhinotracheitis; TRT) und des sogenannten Swollen Head Syndroms (SHS) der Hühner bekannt. Jedoch liegen nur wenige epidemiologische Studien zu der Verbreitung des Virus und dessen Subtypen in Legehennenbetrieben vor. Ziel der vorliegenden Studie war es daher, die Verbreitung des aMPV, vor allem der Subtypen A und B, zu unterschiedlichen Zeiten der Legeperiode zu untersuchen, um ein besseres Verständnis über den Zeitpunkt der Erstinfektionen sowie evtl. Re- oder Neuinfektionen zu erhalten. Dafür wurden erstmals 18 Legehennenherden in Sachsen alle drei Monate über die gesamte Legeperiode auf das Vorkommen von aMPV-RNA und aMPV-spezifischer Antikörper untersucht. Verschiedene Haltungssysteme wurden berücksichtigt, um ein unterschiedliches seuchenhygienisches Risiko unter Praxisbedingungen bewerten zu können. Pro Herde wurden von je zehn Hühnern Trachealtupfer und Serumproben entnommen. Die Tupferproben wurden mittels duplex nested RT-PCR untersucht, die Serumproben mittels zweier kommerzieller ELISA-Tests. In jeder Herde gelang der aMPV-RNA-Nachweis mindestens einmal zu unterschiedlichen Zeitpunkten. Bereits bei der Einstallung konnten in 17 Herden aMPV-spezifische Antikörper und/oder aMPV-RNA nachgewiesen werden. Diese Ergebnisse zeigen die hohe Verbreitungsrate des aMPV in Legehennenbetrieben. Bereits in der Aufzucht fand in der Mehrzahl der Herden eine aMPV-Infektion statt; während der Legeperiode kam es zu häufigen Re- oder Neuinfektionen bzw. zu einer langen Persistenz des Virus. Subtyp A kam alleine (51%) mehr als doppelt so häufig vor wie ausschließlich Subtyp B (22%). Doppelinfektionen mit den Subtypen A und B (27%) wurden ungefähr so häufig gefunden wie eine Infektion ausschließlich mit Subtyp B. Ein Wechsel der Subtypen A und B während einer Legeperiode wurde am häufigsten beobachtet: zehn der 18 Herden (56%) zeigten diesen Verlauf. Ausschließlich Subtyp A in allen positiven Entnahmen pro Betrieb wurde in vier von 18 Herden gefunden, ausschließlich Subtyp B in drei von 18 Herden, Subtyp A gemeinsam mit Subtyp B in einer von 18 Herden. Dies verdeutlicht die Dominanz des Subtyps A in Legehennenbetrieben. Obwohl drei Herden während der Aufzucht mit einer Subtyp B-Vakzine geimpft wurden, gelang der aMPV-RNA Nachweis in bis zu vier Probenentnahmen. Der Subtyp A dominierte auch in den geimpften Herden. Neben dem Subtyp B Feldvirus wurde in einer Herde zum Zeitpunkt der Einstallung auch ein Subtyp B ähnlich dem Impfstamm nachgewiesen. Es ist daher davon auszugehen, dass trotz bekannter Kreuzimmunität eine Impfung nicht vor Infektionen schützt, aber die Persistenz von Subtyp B vermindert. Die Analyse der Serumproben mit zwei kommerziellen ELISA-Tests ergab zum Teil konträre Ergebnisse. Da die Diagnose einer aMPV-Infektion häufig nur über diese Methode gestellt wird, ist dies von praktischer Relevanz. Eine Evaluierung des ELISA-Tests mit der höchsten Spezifität und Sensitivität sollte daher folgen.

Introdução: A relevância clínica da presença de anticorpos anti-HLA após o transplante renal tem sido foco de recente atenção para os estudiosos da histocompatibilidade. Pacientes que possuem anticorpos anti-HLA no póstransplante apresentam maior incidência de rejeição aguda (RA) e de nefropatia crônica do enxerto (NCE). Como conseqüência, alguns perdem o órgão transplantado ou sofrem com as reações imunopatológicas correspondentes. Entretanto, existem algumas controvérsias sobre o grau de valorização da presença desses anticorpos na etiopatogenia da RA e da NCE, pois nem todos os pacientes com anticorpos evoluem mal. Objetivo: Avaliar a presença de anticorpos anti-HLA no primeiro ano do transplante renal e verificar sua associação com a ocorrência de RA e NCE. Pacientes e Método: Este estudo incluiu consecutivamente 88 pacientes submetidos a transplante renal no Serviço de Nefrologia do Hospital de Clínicas de Porto Alegre, entre outubro de 2002 a outubro de 2004. Amostras de sangue foram colhidas no 1º, 3º, 6º e 12º meses pós-transplante renal, visando à pesquisa de anticorpos IgG anti-HLA de classes I e II. Nos pacientes que consentiram, biópsias renais de protocolo foram realizadas entre o 2º e o 3º mês e no 12º mês póstransplante. A detecção dos anticorpos foi realizada através de ensaio ELISA (LATM e LAT-1240, One Lambda Inc., USA). Rejeição aguda e a NCE foram diagnosticadas por critérios clínicos, laboratoriais e histopatológicos. Resultados: Oitenta e oito pacientes foram avaliados, sendo 40 (45,5%) do sexo feminino e setenta e dois (81,8%) de etnia caucasóide. Setenta e um (80,6%) receberam rins de doador falecido. Foi detectada a presença de anticorpos anti-HLA em vinte pacientes (22,7%). Desses, somente 3 (4,4%) desenvolveram anticorpos anti-HLA (classe I) no período pós-transplante; os demais (17) já os apresentavam no período prétransplante. No seguimento até um ano, 23 pacientes (26,1%) apresentaram RA e 43 (51,2%) desenvolveram NCE. Nove (45%) pacientes com anticorpos no póstransplante desenvolveram RA contra 14 (20,6%) dos sem anticorpos (P=0,058). Entre os pacientes com anticorpos no pós-transplante, 11 (64,7%) desenvolveram NCE contra 32 (47,8%) dos sem anticorpos (P=0,329). Na análise histológica, os anticorpos anti-HLA foram associados à RA IIA (P=0,001) e à NCE grau II (P= 0,012). As variáveis preditoras para a RA e NCE foram, respectivamente, presença de anticorpos anti-HLA de classe I no 1º mês pós-transplante (OR= 4,30; IC 95%= 1,32-14,1; P= 0,016) e transplante com órgão de doador-limítrofe (OR= 4,81; IC 95%= 1,18-20,3; P= 0,028). Setenta por cento (70%) dos pacientes com RA desenvolveram NCE, contra 45,3% dos pacientes sem RA (P= 0,054). Conclusão: Os anticorpos anti-HLA presentes no primeiro ano do transplante renal foram associados a RA e NCE. A pesquisa de anticorpos anti-HLA no pós-transplante renal realizada por outros pesquisadores e aqui também avaliada, se adotada como rotina, possibilitaria a identificação de casos de mau prognóstico e a escolha de planos terapêuticos mais adequados. A correlação entre anticorpos anti-HLA e rejeição deverá se tornar mais evidente com o passar dos anos, sendo que nossos resultados fortalecem a convicção da necessidade de continuidade desses estudos. / Introduction: The clinical relevance of the presence of anti-HLA antibodies following kidney transplant has been the recent focus of attention of histocompatibility researchers. Patients who present anti-HLA antibodies in the post-transplant period have shown higher incidence of acute rejection (AR) and of chronic allograft nephropathy (CAN). As a result, some lose the transplanted organ or suffer from the corresponding immunopathological reactions. However, there has been some controversy as to the importance of the presence of these antibodies in the ethiopathology of AR and CAN, since not all patients who have these antibodies present the same outcome. Objective: To evaluate the presence of anti-HLA antibodies during the first year of kidney transplantation and to check its association with the occurrence of AR and CAN. Patients and Method: This research included consecutively 88 patients who had undergone kidney transplants in the Hospital de Clínicas de Porto Alegre Nephrology Service between October 2002 and October 2004. Blood samples were taken during the 1st, 3rd, 6th and 12th months post kidney transplant, aiming at researching for Class I and II IgG anti-HLA antibodies. In consenting patients, protocol kidney biopsies were carried out between the 2nd and 3rd months and in the 12th month after the transplant. Detection of antibodies was done through ELISA test (LAT-M and LAT-1240, One Lambda Inc., USA). Acute rejection and CAN were diagnosed through clinical, laboratorial and histopathological criteria. Results: Eighty-eight patients were evaluated, among which 40 (45.5%) were female and seventy-two (81.8%) were Caucasian. Seventy-one (80.6%) received kidneys from deceased donors. The presence of anti-HLA antibodies was found in 20 patients (22.7%). Among these, only 3 (4.4%) developed anti-HLA antibodies (class I) during the post-transplant period; the remaining (17) already presented these antibodies during the pre-transplant period. In the follow-up up to one year, 23 patients (26.1%) presented AR and 43 (51.2%) developed CAN. Nine patients (45%) with antibodies in the post-transplant period developed AR as opposed to 14 (20.6%) patients without antibodies (P=0.058). Among the patients with antibodies in the post-transplant period, 11 (64.7%) developed CAN as opposed to 32 (47.8%) of those without antibodies (P=0.329). In the histological analysis, the anti-HLA antibodies were associated to AR IIA (P=0.001) and to CAN degree II (P= 0.012). The predictive variables for AR and CAN were, respectively, the presence of Class I anti-HLA antibodies in the first month post-transplant (OR= 4.30; IC 95%= 1.32-14.1; P= 0.016) and transplant with expanded criteria donors (OR= 4.81; IC 95%= 1.18-20.3; P= 0.028). Seventy per cent of the patients presenting AR developed CAN, as opposed to 45.3% of the patients without AR (P= 0.054). Conclusion: The anti-HLA antibodies present in the first year of the kidney transplant were associated to AR and CAN. The research of anti-HLA antibodies in the kidney post-transplant period carried by other researchers, as well as in this study, if done routinely, would allow the identification of cases with a poor prognosis and the choice of more adequate treatments. The correlation of anti-HLA antibodies and rejection will become more evident with time, and our results reinforce the certainty that these studies must continue.

Made available in DSpace on 2015-03-26T13:47:05Z (GMT). No. of bitstreams: 1 texto completo.pdf: 2838276 bytes, checksum: ab260e36167467b459860c7472987dd6 (MD5) Previous issue date: 2012-05-29 / The bovine cysticercosis is a zoonotic disease that causes damage both in terms of health and in terms of economy. This research was developed in order to better understand the anatomical and microscopic characteristics related to the implantation of cysticerci and the immune evolution of the disease, and evaluate the performance of the immunoassay ELISA in the diagnosis of nine experimentally infected animals by oral administration of Taenia saginata eggs. Field work with the animals included the monthly collection of blood every 30 days, being that from 120 days post-infection an animal was slaugthered every 30 days to perform the anatomic-pathological examination and proceed with the collection of cysticerci. The samples from naturally infected animals were obtained by collecting in commercial slaughterhouse. Regarding muscular location of cysticercus in Group 1, the sites routinely inspected with predominance of occurrence were the heart (37.7%), head muscles (17.1%) and the tongue (2.3%). In relation to the assessed commercial cuts, 8.2% of cysticerci were found in the sparerib, 6.6% in the shoulder, 6.2% in the tenderloin and 5.8 in the rump. Other not-routinely inspected tissues that presented cysticerci were the diaphragm, the liver and the esophagus with 2.7%, 12.0% and 1.2% of the total cysticerci. Not viable cysticercus was the majority in head muscles (77.3%), followed by 76.3% in heart muscle and 71.0% in the liver and tongue (50%). Concerning viability, liver presented 87.5% of the not viable cysticerci, followed by the tongue 66.7%, and the heart with 63.2%, but the highest frequency of viable cysticerci was found in the head muscles (68.3%). Of the nine animals studied, five had the kinetics of antibody production against the cysticerci in a similar way when measured the production of antibodies of the IgM and IgG classes, once the peak of antibody production occurred in the period of 0-30 days post-infection followed by a natural decline and maintained detectable until the end of the study. However, the other two groups, each containing two animals, showed the kinetics of antibody production different from most animals, being that one group had a delayed response to the presence of cysticerci and the other group showed no immune response against antigens secreted by cysticerci. For the production of IgG1, it was possible to detect increased levels of antibodies from 30 days post-infection, having the production peak at 270 days, and IgG2 at 60 days. Regarding the cellular response observed against the cysticerci, it was possible to observe that in the case of viable cysticerci, inflammatory cells were present in the major part; and in the case of unviable cysticerci, the presence of repairing tissue cells, being also correlated the amount of calcareous corpuscles with the parasite's stage of death. With regard to the performance of the ELISA test against experimentally and naturally infected animals using a cut-off 1 and 2 added with 2 standard-deviation (SD), it was of 12% and 24.4%, respectively; and when using the cut-off 1 and 2 added with 3 SD the test sensitivity was at 1.99% and 14.4%, respectively. As for the samples from experimentally infected animals using the cut point 1 and 2 added with 2 SD the sensitivity was at 55.9% and 92.5%; and adding 3 SD the values found were 31.2% and 86%, respectively; the specificity of the test in all situations tested was at 100%. The combination of these results should be considered for diagnosis of bovine cysticercosis through the ELISA test, suggesting its use in combination with the anatomic-pathological test routinely performed in slaughterhouses. / A cisticercose bovina é uma doença de caráter zoonótico que acarreta prejuízos tanto de ordem sanitária quanto de ordem econômica. Essa pesquisa foi desenvolvida com o objetivo de conhecer melhor as características macroscópicas e microscópicas relacionadas à implantação dos cisticercos e a evolução imunológica da doença, bem como avaliar o desempenho do imunoensaio ELISA no diagnóstico de bovinos com infecção natural e experimental, através da administração oral de ovos de Taenia saginata. Os trabalhos de campo com os nove animais infectados experimentalmente envolveram coleta mensal de sangue nos intervalos de 30 dias, sendo que a partir de 120 dias pós-infecção um animal era abatido a cada 30 dias para realizar o exame anátomo-patológico e proceder com a coleta de cisticercos. As amostras dos animais naturalmente infectados foram obtidas através da coleta em matadouro-frigorífico comercial. Com relação à localização muscular dos cisticercos no grupo 1 (animais experimentalmente infectados), os sítios inspecionados rotineiramente predominantes de ocorrência foram o coração (37,7%), músculos mastigatórios (17,1%) e língua (2,3%). Em relação aos cortes comerciais estudados, foram encontrados os valores de 8,2% de cisticercos no acém, 6,6% na paleta, 6,2% no contra-filé e 5,8% na alcatra. Os outros tecidos não inspecionados rotineiramente que apresentaram cisticercos foram o diafragma, fígado e o esôfago, com 2,7%, 12,0% e 1,2% dos cisticercos totais. Houve predominância de cisticercos inviáveis nos músculos mastigatórios (77,3%), seguidos de 76,3% na musculatura do coração e 71,0% do fígado e na língua (50%). Os cisticercos viáveis predominaram na alcatra (80,0%), diafragma (71,4%) e esôfago (66,7%). Nos sítios de predileção dos animais naturalmente infectados, foram encontrados 61,78% dos cisticercos totais no coração, seguido dos músculos mastigatórios (38,21%) e fígado (10,19%). Em relação à viabilidade, o fígado apresentou 87,5% de cisticercos inviáveis, seguidos da língua com 66,7%, coração 63,2%, entretanto nos tecidos mastigatórios foi encontrada a maior frequência de cisticercos viáveis (68,3%). Dos nove animais estudados, cinco apresentaram a cinética de produção de anticorpos contra os cisticercos de forma semelhante quando mensurados a produção de anticorpos da classe IgM e IgG, constituindo-se o grupo 1. O pico de produção de anticorpos do grupo 1 ocorreu no período de 0-30 dias pós-infecção seguido de um declínio natural, mantendo-se detectável até o final do estudo, no entanto os outros dois grupos contendo dois animais cada apresentaram a cinética de produção de anticorpos diferente do grupo 1, sendo que, o grupo 2 não apresentou resposta imunológica contra os antígenos secretados pelos cisticercos e o grupo 3 apresentou resposta tardia à presença dos cisticercos. Em relação à produção média de IgG1 para todos os grupos, foi possível detectar elevação dos níveis de anticorpos a partir de 30 dias pós-infecção, tendo-se o pico de produção aos 270 dias e IgG2 aos 60 dias. Já em relação à resposta celular hospedeiro-parasito, foi possível observar que no caso de cisticercos viáveis estavam presentes em sua maioria células inflamatórias, e no caso de cisticercos inviáveis a presença de células reparadoras de tecido, sendo também correlacionada a quantidade de corpúsculos calcáreos com o estádio de morte do parasito. Com relação ao desempenho do teste ELISA frente a animais infectados naturalmente e experimentalmente utilizando o ponto de corte 1 (calculado a partir de amostras de animais negativos para a cisticercose durante inspeção de rotina em matadouro-frigorífico) e ponto de corte 2 (amostras de animais negativos para a cisticercose criados em isolamento) adicionados de 2 desvios-padrão (SD), a sensibilidade do teste foi de 12% e 24,4%. Ao utilizar o ponto de corte 1 e 2 adicionados de 3 SD a sensibilidade ficou em 1,99% e 14,4%, respectivamente. Já para as amostras de animais experimentalmente infectados utilizando o ponto de corte 1 e 2 adicionados de 2 SD a sensibilidade ficou em 55,9% e 92,5% e, adicionando 3 SD foram encontrados os valores de 31,2% e 86%. A especificidade do teste em todas as situações testadas ficou em 100%. A combinação desses resultados deve ser valorizada no diagnóstico da cisticercose bovina por meio do teste ELISA sugerindo sua utilização em associação com o teste anátomo-patológico realizado rotineiramente em matadouros-frigoríficos.

Made available in DSpace on 2016-12-08T16:24:07Z (GMT). No. of bitstreams: 1 PGCA12MA056.pdf: 526904 bytes, checksum: 7a4c82b6cb16c016c29f0182cfbbaa51 (MD5) Previous issue date: 2012-02-27 / The analysis methods for Salmonella spp. detection are of great importance in the system of animal products processing industries, because they reflect directly on the microbiological quality of food, impacting the storage time and therefore the release of the product for sale. The present study verified the efficiency of a new ELFA ELISA test using a recombinant protein from bacteriophage (VIDAS UP®) to detect the presence of Salmonella spp. in products of animal origin. Two hundred fifteen samples (15 samples artificially contaminated and 200 samples from agribusiness routine), from different products of animal origin were analyzed. The samples were tested by five methods of diagnosis: ELFA ELISA, conventional methodology in accordance with ISO 6579, polymerase chain reaction - PCR, semi-solid Rappaport-Vassiliadis Modified - MSRV and MSRV medium + supplement. Eleven out of 215 (5.12%) samples were positive in at least one test, which four samples were from agribusiness routine and seven were from artificially contaminated samples. The sensitivity and specificity of the ELFA ELISA when compared with the conventional method was 90.0% and 99.51% respectively. The ELFA ELISA was equivalent to the other methodologies, showing to be effective and rapid to detect Salmonella spp. in different products of animal origin / Os métodos de análise para detecção de Salmonella spp. são de grande importância no processo da agroindústria de produtos de origem animal, pois refletem diretamente na qualidade microbiológica dos alimentos, impactando no tempo de estocagem e por conseguinte na liberação do produto para venda. O presente trabalho verificou a eficiência de um novo teste ELFA ELISA utilizando proteína recombinante oriunda de bacteriófago (VIDAS UP®) para detectar a presença de Salmonella spp. em produtos de origem animal. Foram analisadas 215 amostras (15 amostras artificilamente contaminadas e 200 amostras de rotina da agroindústria), de diferentes produtos de origem animal. As amostras foram submetidas a cinco métodos de diagnóstico: ELFA ELISA, metodologia convencional de acordo com ISO 6579, reação em cadeia da polimerase - PCR, meio semi-sólido Rappaport-Vassiliadis Modificado - MSRV e meio MSRV + suplemento. Onze amostras das 215 (5,12%) foram positivas em pelo menos um dos testes, sendo quatro amostras de rotina da agroindústria e sete amostras artificialmente contaminadas. A sensibilidade e especificidade do teste ELFA ELISA com a metodologia convencional foi de 90,0% e 99,51%, respectivamente. O teste ELFA ELISA apresentou equivalência com as demais metodologias, mostrando ser eficaz e rápido na detecção de Salmonella spp. em diferentes produtos de origem animal

Made available in DSpace on 2016-12-08T16:24:11Z (GMT). No. of bitstreams: 1 PGCA10MA077.pdf: 251261 bytes, checksum: 659afb14e9609405d069749d1029eb68 (MD5) Previous issue date: 2010-08-09 / This study evaluated the prevalence of antibodies against Toxoplasma gondii in sheeps in the municipality of Lages, in the state of Santa Catarina, Brazil, and identified possible risk factors for infection. Blood samples of 360 animals from 13 properties were collected by puncturing of the jugular vein, stored in sterile tubes and carried to the Laboratory of Parasitology and Parasitic Diseases CAV / UDESC, for later processing. Each creator answered a questionnaire with data of the propertie and each individual animal for identification of risk factors for infection. Indirect Immunofluorescence Assay (IFA) where serum samples were considered positive at dilutions ≥1:64 and Enzyme Linked Immunosorbent Assay (ELISA) were used to detect IgG anti Toxoplasma gondii antibodies. 100% of properties were found positive animals. By IFA, 205 (56.94%) sheeps had antibodies against Toxoplasma gondii and by ELISA, 153 (42.50%) sheeps were positive. Considering the serological techniques and statistical analysis, were risk factors by ELISA: the age, the water source and the animal category; and by IFA, the racial type. It has been found sensitivity of 61%, specificity of 82% and Kappa of 0.7 between the ELISA and the IFA (1:64), considered good, allowing to indicate the ELISA as technique adjusted for the diagnosis of Toxoplasma gondii in the ovina species / Com os objetivos de estimar a prevalência de ovinos portadores de Toxoplasma gondii no município de Lages, Santa Catarina, Brasil, e de identificar possíveis fatores de risco para a infecção, foi coletado sangue por punção da veia jugular externa de 360 animais, de 13 propriedades, armazenado em tubos estéreis e transportado ao Laboratório de Parasitologia e Doenças Parasitárias CAV/UDESC, para posterior processamento. Cada criador respondeu a um questionário com dados da propriedade e individuais de cada animal para permitir a identificação dos fatores de risco da infecção. A pesquisa de anticorpos foi realizada por meio da Reação de Imunofluorescência Indireta (RIFI) utilizando como ponto de corte a titulação 1:64 e do Ensaio Imunoenzimático Indireto (ELISA). Em 100% das propriedades foram encontrados animais positivos. Pela RIFI, 205 (56,94%) ovinos apresentaram anticorpos contra Toxoplasma gondii e pelo ELISA, 153 (42,50%). Considerando-se as técnicas sorológicas e a análise estatística, foram fatores de risco pelo ELISA: a idade, a fonte de água e a categoria animal; e pela RIFI, o tipo racial. Foi constatada sensibilidade de 61%, especificidade de 82% e concordância Kappa de 0,7 entre o ELISA e a RIFI (1:64), considerada boa, permitindo indicar o ELISA como técnica adequada para o diagnóstico de Toxoplasma gondii na espécie ovina

Made available in DSpace on 2014-12-17T14:10:22Z (GMT). No. of bitstreams: 1 AndreaDM_DISSERT.pdf: 1136289 bytes, checksum: 4971fd9b2aab3f08935061c736892689 (MD5) Previous issue date: 2010-03-31 / Toxoplasmosis is one zoonosis caused by Toxoplasma gondii protozoan. Goats, amongst the production animals, are one of the species most susceptible to this parasite, being one them main involved agents in ovine and goat abortions, determining great economic losses and implications for public health, since the presence it parasite in the products of goat origin, consist in one of the main sources of infection for the man. In this study 244 blood samples in 8 farms situated in 4 cities from the Sert?o do Cabugi region, Rio Grande do Norte State, northeast of Brazil and, tested by ELISA assay. The results had shown a prevalence of 47.13% for anti- T. gondii antibodies and a significant association between positivity and variable evaluated as age, locality and property. The IgG avidity assay evaluated in 115 positive samples was carried to discriminate acute and chronic infection. Twelve samples (10.4%) had presented antibodies of low avidity while 103 (89.6%) presented high avidity antibodies; indicating that most of the animals was precocious exposure to the parasite. Significant difference was verified only for the variable sex. We also evaluate the capacity of recombinant adenoviruses codifying SAG1, SAG2, SAG3 and CMV in inducing activation of specific immune response in goat. These 109 animals received 109 pfu of the AdSAG1, AdSAG2, AdSAG3, AdCMV or PBS in vaccine protocol with 3 immunizations. Serum samples of the each animal, before and after mmunization, had been submitted to the ELISA. The results demonstrate that the immunizations had induced the production of IgG antibodies specific against T. gondii proteins / A toxoplasmose ? uma zoonose causada pelo protozo?rio Toxoplasma gondii. Os caprinos, dentre os animais de produ??o, s?o uma das esp?cies mais suscet?veis a este parasito, sendo um dos principais agentes envolvidos no abortamento de ovinos e caprinos, determinando grandes perdas econ?micas e tendo implica??es para sa?de p?blica, j? que a presen?a do parasita nos produtos de origem caprina, constituem-se em uma das principais fontes de infec??o para o homem. Neste estudo foram coletadas 244 amostras de sangue em 8 fazendas situadas em 4 munic?pios da regi?o do Sert?o do Cabugi, Estado do Rio Grande do Norte, regi?o Nordeste do Brasil e testadas atrav?s do Ensaio Imunoenzim?tico (ELISA). Os resultados obtidos mostraram uma preval?ncia de 47,13% para anticorpos anti-T. gondii e uma associa??o significativa entre positividade e as vari?veis avaliadas tais como idade, localidade e propriedade. Foi realizado o ensaio IgG de avidez em 115 amostras positivas com o intuito de discriminar infec??o aguda e cr?nica. Doze amostras (10,4%) apresentaram anticorpos de baixa avidez enquanto 103 (89,6%) anticorpos de alta avidez; indicando que a maior parte dos animais foi exposto ao parasita precocemente. Foi verificada diferen?a estat?stica significativa apenas para a vari?vel sexo. Avaliamos tamb?m a capacidade de adenov?rus recombinantes codificando SAG1, SAG2, SAG3 e CMV em induzir ativa??o de resposta imune em caprinos. Estes animais receberam 109 pfu de AdSAG1, AdSAG2, AdSAG3, AdCMV ou PBS em protocolo vacinal com 3 imuniza??es. Amostras de soro obtidas dos animais, antes e ap?s cada imuniza??o, foram submetidas ao ELISA. Os resultados demonstram que as imuniza??es induziram a produ??o anticorpos IgG espec?ficos contra as prote?nas de T. gondii

Made available in DSpace on 2014-12-17T15:55:05Z (GMT). No. of bitstreams: 1 JessicaRF_DISSERT.pdf: 1324969 bytes, checksum: b7d1df5d394f9f4ec1bafce2c8803e4c (MD5) Previous issue date: 2014-02-21 / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior / Blooms negatively compromise the aquatic environment, plants, animals and human health.Some species are toxin-producing, such as the neurotoxin saxitoxin and the hepatotoxin microcystin, which may contaminate water reservoirs, as those existing in the semiarid region of Rio Grande do Norte (Brazil) which are used to supply the population, fishing, aquiculture and recreational activities, thereby providing the risk of human exposure through water intake, dermal contact and respiratory tract. Thus, it is recommended a constant monitoring of the density of cyanobacteria with the quantification of cyanotoxins. One goal with this work was the monitoring of water in four reservoirs in semiarid of RN through the identification and enumeration of cyanobacteria and through the identification and quantification of cyanotoxins by ELISA. Furthermore, we intended to assess the environmental perception of farmers and artisanal fishers in reservoirs of semiarid of RN through semi-structured interviews with questions mostly related to water and eutrophication. Through these objectives the aim was the development of management strategies for aquaculture and prevention of risks to public health. The results showed that the highest values of microcystins were found in the rainy season. Standards for drinking water, according to the guidelines of Ministry of Health 2914/2011 and CONAMA 357/05, setting the maximum values for raw water density of cyanobacteria: 50,000 cel.mL-1; microcystin: 1 μg. L-1 and saxitoxin: 3 μg. L-1. The values found for microcystin ranged between 0.00227 μg. L-1 and 24.1954 μg. L-1. From 128 samples analyzed, 27% were above the limit. There was no clear seasonal pattern for saxitoxins and their values ranged between 0.003 μg. L-1 and 0.766 μg. L-1 with none of the values above the limit. Furthermore, 76% of the densities of cyanobacteria values were above the limit. About environmental perception, 52 interviews were conducted and the results show that the respondents recognize the main uses of water of the rervoirs, recognize the importance and have a positive view about the reservoirs. They also realize that the water has a poor quality and can cause health problems. The results provide data showing the persistence of cyanobacteria and cyanotoxins, many times over the limit, reinforcing the importance of constant monitoring. The assessment of environmental perception gives foundation for later proposed environmental education linked to public health management into the context of this particular population, making it more effective / As flora??es de cianobact?rias comprometem negativamente o meio ambiente aqu?tico, plantas, animais e podem gerar problemas de sa?de p?blica. Algumas esp?cies s?o produtoras de toxinas, como a neurotoxina saxitoxinas e a hepatotoxina microcistina, as quais podem contaminar reservat?rios de ?gua, como os a?udes do semi?rido do Rio Grande do Norte, os quais s?o utilizados para abastecimento da popula??o, pesca, atividades aquicultoras e de lazer, propiciando assim o risco de exposi??o humana pelas vias oral, d?rmica e respirat?ria. Assim, ? recomendado o monitoramento permanente da densidade de cianobact?rias juntamente com a quantifica??o de cianotoxinas. Um dos objetivos com esse trabalho foi realizar o monitoramento da ?gua de quatro a?udes do semi?rido norteriograndense atrav?s da identifica??o e contagem de cianobact?rias e atrav?s da identifica??o e quantifica??o das cianotoxinas pelo do m?todo ELISA. Al?m disso, pretendeuse avaliar a percep??o ambiental de aquicultores e pescadores em a?udes do semi?rido do RN atrav?s de entrevistas semiestruturadas com quest?es majoritariamente relacionadas ? ?gua e eutrofiza??o. Atrav?s desses objetivos visou-se o desenvolvimento de estrat?gias de manejo para aquicultura e preven??o de riscos ? sa?de p?blica. Os resultados demonstraram que os maiores valores de microcistinas foram encontrados no per?odo chuvoso. Os padr?es de potabilidade da ?gua, de acordo com a portaria do Minist?rio da Sa?de 2914/2011 e CONAMA 357/05, estabelecem os valores m?ximos para ?gua bruta de densidade de cianobact?rias: 50 mil cel.mL-1; microcistina: 1 μg.L-1 e saxitoxina: 3 μg.L-1. Os valores encontrados de microcistinas variaram entre 0,00227 μg.L-1 e 24,1954 μg.L-1. Das 128 amostras analisadas, 27% estavam acima do permitido. N?o foi encontrado um padr?o sazonal para saxitoxinas e seus valores variaram entre 0,003 μg.L-1 e 0,766 μg.L-1 com nenhum dos valores acima do permitido. Al?m disso, 76% dos valores de densidade de cianobact?rias estavam acima do permitido. Em rela??o ? percep??o ambiental, foram realizadas 52 entrevistas e os resultados mostram que os entrevistados reconhecem os principais usos da ?gua dos a?udes, reconhecem a import?ncia e enxergam os mesmos de forma positiva. Eles tamb?m percebem que a ?gua est? com a qualidade ruim e que pode trazer problemas de sa?de. Os resultados fornecem dados que demonstram a perman?ncia de cianobact?rias e cianotoxinas, muitas vezes acima do limite, refor?ando a import?ncia do monitoramento constante. A avalia??o da percep??o ambiental d? embasamento a uma posterior proposta de educa??o ambiental ligada ? gest?o p?blica da sa?de inserida no contexto dessa determinada popula??o, tornando-a mais efetiva

Introdução: A relevância clínica da presença de anticorpos anti-HLA após o transplante renal tem sido foco de recente atenção para os estudiosos da histocompatibilidade. Pacientes que possuem anticorpos anti-HLA no póstransplante apresentam maior incidência de rejeição aguda (RA) e de nefropatia crônica do enxerto (NCE). Como conseqüência, alguns perdem o órgão transplantado ou sofrem com as reações imunopatológicas correspondentes. Entretanto, existem algumas controvérsias sobre o grau de valorização da presença desses anticorpos na etiopatogenia da RA e da NCE, pois nem todos os pacientes com anticorpos evoluem mal. Objetivo: Avaliar a presença de anticorpos anti-HLA no primeiro ano do transplante renal e verificar sua associação com a ocorrência de RA e NCE. Pacientes e Método: Este estudo incluiu consecutivamente 88 pacientes submetidos a transplante renal no Serviço de Nefrologia do Hospital de Clínicas de Porto Alegre, entre outubro de 2002 a outubro de 2004. Amostras de sangue foram colhidas no 1º, 3º, 6º e 12º meses pós-transplante renal, visando à pesquisa de anticorpos IgG anti-HLA de classes I e II. Nos pacientes que consentiram, biópsias renais de protocolo foram realizadas entre o 2º e o 3º mês e no 12º mês póstransplante. A detecção dos anticorpos foi realizada através de ensaio ELISA (LATM e LAT-1240, One Lambda Inc., USA). Rejeição aguda e a NCE foram diagnosticadas por critérios clínicos, laboratoriais e histopatológicos. Resultados: Oitenta e oito pacientes foram avaliados, sendo 40 (45,5%) do sexo feminino e setenta e dois (81,8%) de etnia caucasóide. Setenta e um (80,6%) receberam rins de doador falecido. Foi detectada a presença de anticorpos anti-HLA em vinte pacientes (22,7%). Desses, somente 3 (4,4%) desenvolveram anticorpos anti-HLA (classe I) no período pós-transplante; os demais (17) já os apresentavam no período prétransplante. No seguimento até um ano, 23 pacientes (26,1%) apresentaram RA e 43 (51,2%) desenvolveram NCE. Nove (45%) pacientes com anticorpos no póstransplante desenvolveram RA contra 14 (20,6%) dos sem anticorpos (P=0,058). Entre os pacientes com anticorpos no pós-transplante, 11 (64,7%) desenvolveram NCE contra 32 (47,8%) dos sem anticorpos (P=0,329). Na análise histológica, os anticorpos anti-HLA foram associados à RA IIA (P=0,001) e à NCE grau II (P= 0,012). As variáveis preditoras para a RA e NCE foram, respectivamente, presença de anticorpos anti-HLA de classe I no 1º mês pós-transplante (OR= 4,30; IC 95%= 1,32-14,1; P= 0,016) e transplante com órgão de doador-limítrofe (OR= 4,81; IC 95%= 1,18-20,3; P= 0,028). Setenta por cento (70%) dos pacientes com RA desenvolveram NCE, contra 45,3% dos pacientes sem RA (P= 0,054). Conclusão: Os anticorpos anti-HLA presentes no primeiro ano do transplante renal foram associados a RA e NCE. A pesquisa de anticorpos anti-HLA no pós-transplante renal realizada por outros pesquisadores e aqui também avaliada, se adotada como rotina, possibilitaria a identificação de casos de mau prognóstico e a escolha de planos terapêuticos mais adequados. A correlação entre anticorpos anti-HLA e rejeição deverá se tornar mais evidente com o passar dos anos, sendo que nossos resultados fortalecem a convicção da necessidade de continuidade desses estudos. / Introduction: The clinical relevance of the presence of anti-HLA antibodies following kidney transplant has been the recent focus of attention of histocompatibility researchers. Patients who present anti-HLA antibodies in the post-transplant period have shown higher incidence of acute rejection (AR) and of chronic allograft nephropathy (CAN). As a result, some lose the transplanted organ or suffer from the corresponding immunopathological reactions. However, there has been some controversy as to the importance of the presence of these antibodies in the ethiopathology of AR and CAN, since not all patients who have these antibodies present the same outcome. Objective: To evaluate the presence of anti-HLA antibodies during the first year of kidney transplantation and to check its association with the occurrence of AR and CAN. Patients and Method: This research included consecutively 88 patients who had undergone kidney transplants in the Hospital de Clínicas de Porto Alegre Nephrology Service between October 2002 and October 2004. Blood samples were taken during the 1st, 3rd, 6th and 12th months post kidney transplant, aiming at researching for Class I and II IgG anti-HLA antibodies. In consenting patients, protocol kidney biopsies were carried out between the 2nd and 3rd months and in the 12th month after the transplant. Detection of antibodies was done through ELISA test (LAT-M and LAT-1240, One Lambda Inc., USA). Acute rejection and CAN were diagnosed through clinical, laboratorial and histopathological criteria. Results: Eighty-eight patients were evaluated, among which 40 (45.5%) were female and seventy-two (81.8%) were Caucasian. Seventy-one (80.6%) received kidneys from deceased donors. The presence of anti-HLA antibodies was found in 20 patients (22.7%). Among these, only 3 (4.4%) developed anti-HLA antibodies (class I) during the post-transplant period; the remaining (17) already presented these antibodies during the pre-transplant period. In the follow-up up to one year, 23 patients (26.1%) presented AR and 43 (51.2%) developed CAN. Nine patients (45%) with antibodies in the post-transplant period developed AR as opposed to 14 (20.6%) patients without antibodies (P=0.058). Among the patients with antibodies in the post-transplant period, 11 (64.7%) developed CAN as opposed to 32 (47.8%) of those without antibodies (P=0.329). In the histological analysis, the anti-HLA antibodies were associated to AR IIA (P=0.001) and to CAN degree II (P= 0.012). The predictive variables for AR and CAN were, respectively, the presence of Class I anti-HLA antibodies in the first month post-transplant (OR= 4.30; IC 95%= 1.32-14.1; P= 0.016) and transplant with expanded criteria donors (OR= 4.81; IC 95%= 1.18-20.3; P= 0.028). Seventy per cent of the patients presenting AR developed CAN, as opposed to 45.3% of the patients without AR (P= 0.054). Conclusion: The anti-HLA antibodies present in the first year of the kidney transplant were associated to AR and CAN. The research of anti-HLA antibodies in the kidney post-transplant period carried by other researchers, as well as in this study, if done routinely, would allow the identification of cases with a poor prognosis and the choice of more adequate treatments. The correlation of anti-HLA antibodies and rejection will become more evident with time, and our results reinforce the certainty that these studies must continue.