COX24 codes for a mitochondrial protein required for processing of the COX1 transcript

In most strains of Saccharomyces cerevisiae the mitochondrial gene COX1, for subunit 1 of cytochrome oxidase, contains multiple exons and introns. Processing of COX1 primary transcript requires accessory proteins factors, some of which are encoded by nuclear genes and others by reading frames residing in some of the introns of the COX1 and COB genes. Here we show that the low molecular weight protein product of open reading frame YLR204W, for which we propose the name COX24, is also involved in processing of COX1 RNA intermediates. The growth defect of cox24 mutants is partially rescued in strains harboring mitochondrial DNA lacking introns. Northern blot analyses of mitochondrial transcripts indicate cox24 null mutants to be blocked in processing of introns aI2 and aI3. The dependence of intron aI3 excision on Cox24p is also supported by the growth properties of the cox24 mutant harboring mitochondrial DNA with different intron compositions. The intermediate phenotype of the cox24 mutant in the background of intronless mitochondrial DNA, however, suggests that in addition to its role in splicing of the COX1 pre-mRNA, Cox24p still has another function. Based on the analysis of a cox14-cox24 double mutant, we propose that the other function of Cox24p is related to translation of the COX1 mRNA. © 2006 by The American Society for Biochemistry and Molecular Biology, Inc.

Plesiomonas shigelloides: Um enteropatógeno emergente?

Plesiomonas shigelloides is a Gram-negative rod-shaped bacterium, of the family Enterobacteriaceae, which has been isolated from freshwater and salt water, freshwater fish, shellfish and many species of animals. Most human P. shigelloides infections are suspected to be waterborne. The organism can be found in untreated water used as drinking water, in recreational water, or in water used to rinse food that is consumed without cooking or heating. The ingestion of P. shigelloides does not always cause illness in the host animal, and the organism may be present temporarily as a transient, noninfectious member of the intestinal flora. It has been isolated from the stools of patients with diarrhea,but it is also sometimes isolated from healthy individuals. P. shigelloides has been implicated in gastroenteritis, usually a self-limiting disease characterized by fever, chills, abdominal pain, nausea, diarrhea or vomiting; in severe cases the diarrhea may be yellowish-green, foamy and tinged with blood. The bacteria may also cause extra-intestinal infection. Furthermore, it can produce toxins and may be invasive. The evidence in favor of considering P. shigelloides as an enteropathogen is not totally convincing. Although it has been isolated from patients with diarrhea and incriminated in some outbreaks involving contaminated water and food, it was not possible, in many P. shigelloides samples associated with gastrointestinal infections, to identify a definite mechanism of virulence.

Fatores antinutricionais e coeficientes de digestibilidade aparente da proteína de produtos de soja para o pacu (Piaractus mesopotamicus)

This study aimed to determine the activities of trypsin inhibitors, hemagglutinant and tannin levels in soybean meal and in raw and processed soy, as well evaluate the protein apparent digestibility coefficient for pacu juveniles. The apparent coefficients of raw, extruded, toasted and milled soy were determined using chromium oxide (0.5%) as marker. A reference diet was created with 26% crude protein and 4,352 kcal kg-1, with each feed containing 30% of the test diet. Feces were collected by abdominal pressure. All analyzed products presented anti-nutritional factors, but the lowest trypsin inhibitor activity was observed in soybean meal. Soy that received thermal treatment presented better digestibility coefficients and lower hemagglutinating activity values than raw soy. No effects of trypsin and tannin inhibitor were observed on the protein digestibility coefficient, but a negative relationship was observed between hemagglutinin levels and protein digestibility coefficient. The use of soybean meal and extruded or toasted soy is recommended for pacu feeding.

Presence of antibodies against H5, H7 and H9 influenza a virus in wild birds in the state of São Paulo, Brazil

Although the natural reservoirs of the avian influenza (AI) virus have been extensively studied in many countries, there is a clear lack of information on this subject in South America, particularly in Brazil. The objective of this study was to conduct a serological survey for H5, H7 and H9 antibodies to AI-subtype viruses in wild birds in the state of São Paulo, Brazil. Serum samples were tested using the hemagglutination-inhibition assay. Out of the 31 wild birds sampled between January and December of 2006, seven (22.58%), were seropositive for H5, H7 and H9; four (12.90%) were seropositive for H5 and H7; 13 (41.94%), were seropositive only for H7; three (9.7%), were seropositive only for H9; and four (12.90%) were negative for all three hemagglutinin subtypes. These results indicate that AI viruses belonging to H5, H7 and H9 subtypes circulate among wild birds in the state of São Paulo in the form of either concurrent or consecutive infections. This study contributes to the knowledge of AI epidemiology in Brazil, and stresses the need of further detailed and long-term epidemiological and ecological investigation to determine the current status of this virus.

Análise citogenética clássica e molecular em Chelonoidis carbonaria e Phrynops geoffroanus (Testudines)

Pós-graduação em Genética - IBILCE

Análise da expressão gênica global da bactéria Xylella fastidiosa em laranja doce por microarranjos de DNA

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Caracterização dos genes codificadores da hemaglutinina e PB2 do vírus Influenza A (H1N1) pandêmico isolado na mesorregião metropolitana de Belém

A recente pandemia de gripe de 2009/2010 causada pelo vírus A (H1N1) pandêmico mostrou um perfil de gravidade diferente da gripe sazonal, pois um percentual considerável de casos graves e fatais ocorreu em indivíduos adultos jovens, sem comorbidade. A virulência dos vírus Influenza A (H1N1) pandêmico resulta de interações protéicas complexas e depende essencialmente de alguns genes virais. O objetivo deste estudo foi caracterizar os genes codificadores da hemaglutinina (H1) e polimerase básica 2 (PB2) do vírus Influenza A (H1N1) pandêmico mediante a obtenção de cepas provenientes de pacientes com gripe procedente da mesorregião metropolitana de Belém-PA. O tamanho amostral foi constituído de 87 amostras aleatórias de ambos os sexos de 0 a 96 anos, com síndrome respiratória aguda grave (SRAG) sem nenhuma comorbidade relatada, no período de maio de 2009 a agosto de 2010. As amostras foram isoladas em cultura de célula MDCK e analisadas por técnicas de biologia molecular que compreenderam três etapas principais: a) extração do RNA viral (RNAv) a partir do sobrenadante celular; b) amplificação do RNAv pela técnica de Reação em Cadeia mediada pela Polimerase precedida de Transcrição Reversa (RT-PCR); c) sequenciamento completo dos genes codificadores da H1 e PB2. Das 87 cepas amplificadas pelo RT-PCR, em 82 tornou-se possível a obtenção e análise de sequências para o gene HA, enquanto que de 81 amostras virais obteve-se sequências para o gene PB2. A análise comparativa das sequências obtidas com a sequência da cepa vacinal (A/California/07/2009(H1N1)) revelou substituições aminoacídicas na HA (P83S; D97N; S203T; D222G; Q293H e I321V) e na PB2 (K340N; K526R e M631L), no entanto sem associação a hospitalização. Ao nível de substituição na HA, a D97N isolada ou associada com a S203T, foi detectada com mais frequência na primeira onda. Já ao nível da PB2 a substituição K526R foi mais encontrada em cepas que circularam na primeira onda, enquanto que, a M631L foi mais evidenciada na segunda. A substituição D222G na HA só foi encontrada em casos de óbitos. Por fim, observou-se uma tendência de alterações nos sítios antigênicos da HA. Sendo assim, a contínua vigilância genética e antigênica do vírus Influenza A (H1N1) pdm em circulação, bem como o compartilhamento de informações é de extrema importância para a melhor recomendação possível para os vírus que entram na composição vacinal evitando assim maior risco de epidemias severas no futuro.

Polymorphic toxin systems: Comprehensive characterization of trafficking modes, processing, mechanisms of action, immunity and ecology using comparative genomics

Background: Proteinaceous toxins are observed across all levels of inter-organismal and intra-genomic conflicts. These include recently discovered prokaryotic polymorphic toxin systems implicated in intra-specific conflicts. They are characterized by a remarkable diversity of C-terminal toxin domains generated by recombination with standalone toxin-coding cassettes. Prior analysis revealed a striking diversity of nuclease and deaminase domains among the toxin modules. We systematically investigated polymorphic toxin systems using comparative genomics, sequence and structure analysis. Results: Polymorphic toxin systems are distributed across all major bacterial lineages and are delivered by at least eight distinct secretory systems. In addition to type-II, these include type-V, VI, VII (ESX), and the poorly characterized "Photorhabdus virulence cassettes (PVC)", PrsW-dependent and MuF phage-capsid-like systems. We present evidence that trafficking of these toxins is often accompanied by autoproteolytic processing catalyzed by HINT, ZU5, PrsW, caspase-like, papain-like, and a novel metallopeptidase associated with the PVC system. We identified over 150 distinct toxin domains in these systems. These span an extraordinary catalytic spectrum to include 23 distinct clades of peptidases, numerous previously unrecognized versions of nucleases and deaminases, ADP-ribosyltransferases, ADP ribosyl cyclases, RelA/SpoT-like nucleotidyltransferases, glycosyltranferases and other enzymes predicted to modify lipids and carbohydrates, and a pore-forming toxin domain. Several of these toxin domains are shared with host-directed effectors of pathogenic bacteria. Over 90 families of immunity proteins might neutralize anywhere between a single to at least 27 distinct types of toxin domains. In some organisms multiple tandem immunity genes or immunity protein domains are organized into polyimmunity loci or polyimmunity proteins. Gene-neighborhood-analysis of polymorphic toxin systems predicts the presence of novel trafficking-related components, and also the organizational logic that allows toxin diversification through recombination. Domain architecture and protein-length analysis revealed that these toxins might be deployed as secreted factors, through directed injection, or via inter-cellular contact facilitated by filamentous structures formed by RHS/YD, filamentous hemagglutinin and other repeats. Phyletic pattern and life-style analysis indicate that polymorphic toxins and polyimmunity loci participate in cooperative behavior and facultative 'cheating' in several ecosystems such as the human oral cavity and soil. Multiple domains from these systems have also been repeatedly transferred to eukaryotes and their viruses, such as the nucleo-cytoplasmic large DNA viruses. Conclusions: Along with a comprehensive inventory of toxins and immunity proteins, we present several testable predictions regarding active sites and catalytic mechanisms of toxins, their processing and trafficking and their role in intra-specific and inter-specific interactions between bacteria. These systems provide insights regarding the emergence of key systems at different points in eukaryotic evolution, such as ADP ribosylation, interaction of myosin VI with cargo proteins, mediation of apoptosis, hyphal heteroincompatibility, hedgehog signaling, arthropod toxins, cell-cell interaction molecules like teneurins and different signaling messengers.

A phase I randomized, double-blind, controlled trial of 2009 influenza A (H1N1) inactivated monovalent vaccines with different adjuvant systems

Methods We conducted a phase I, multicenter, randomized, double-blind, placebo-controlled, multi-arm (10) parallel study involving healthy adults to evaluate the safety and immunogenicity of influenza A (H1N1) 2009 non-adjuvanted and adjuvanted candidate vaccines. Subjects received two intramuscular injections of one of the candidate vaccines administered 21 days apart. Antibody responses were measured by means of hemagglutination-inhibition assay before and 21 days after each vaccination. The three co-primary immunogenicity end points were the proportion of seroprotection >70%, seroconversion >40%, and the factor increase in the geometric mean titer >2.5. Results A total of 266 participants were enrolled into the study. No deaths or serious adverse events were reported. The most commonly solicited local and systemic adverse events were injection-site pain and headache, respectively. Only three subjects (1.1%) reported severe injection-site pain. Four 2009 influenza A (H1N1) inactivated monovalent candidate vaccines that met the three requirements to evaluate influenza protection, after a single dose, were identified: 15 μg of hemagglutinin antigen without adjuvant; 7.5 μg of hemagglutinin antigen with aluminum hydroxide, MPL and squalene; 3.75 μg of hemagglutinin antigen with aluminum hydroxide and MPL; and 3.75 μg of hemagglutinin antigen with aluminum hydroxide and squalene. Conclusions Adjuvant systems can be safely used in influenza vaccines, including the adjuvant monophosphoryl lipid A (MPL) derived from Bordetella pertussis with squalene and aluminum hydroxide, MPL with aluminum hydroxide, and squalene and aluminum hydroxide.

Induktion und Analyse spezifischer Immunantworten nach intranodaler Immunisierung mit RNA im murinen Modell

Die Wirksamkeit einer Vakzine ist von vielen Parametern abhängig. Dazu gehören unter anderen: das ausgewählte Antigen, die Formulation in der das Antigen benutzt wird sowie die Applikationsroute. Antigen-kodierende Ribonukleinsäuren (RNA) gilt heutzutage als eine sichere und effiziente Alternative zu traditionellen Impfstoff-Formulierungen, wie Peptiden, rekombinanten Proteinen, viralen Systemen oder DNA basierten Impfstoffen. Bezüglich des Applikationsortes repräsentiert der Lymphknoten ein optimales Milieu für die Interaktion zwischen antigenpräsentierenden Zellen und T-Zellen. Vor diesem Hintergrund war die Zielsetzung dieser Arbeit, ein auf direktem in vivo Transfer von Antigen-kodierender in vitro transkribierter RNA (IVT-RNA) basierendes Impfverfahren zu entwickeln, zu charakterisieren und auf seine anti-tumorale Wirksamkeit zu testen. In der vorliegenden Arbeit konnte gezeigt werden, dass dendritische Zellen (DCs) in vitro hocheffizient mit IVT-RNA transfiziert werden können und eine hohe stimulatorische Kapazität besitzen. Durch Sequenzmodifikation der IVT-RNA konnten wir die Transkriptstabilität und Translationseffizienz erhöhen was zu einer Steigerung der stimulatorischen Kapazität in vivo führte. Darüber hinaus untersuchten wir die Auswirkung der Insertion eines Signalpeptides 5’ sowie einer C-terminalen transmembran- und zytosolischen-Domäne eines MHC-Klasse-I-Moleküls am 3’ der Antigen-kodierenden Sequenz auf die Effizienz der MHC-Klasse-I und -II Präsentation. Wir konnten in vitro und in vivo nachweisen, dass diese Modifikation zu einer gesteigerten, simultanen Stimulation von antigenspezifischen CD4+ und CD8+ T-Zellen führt. Auf der Basis der optimierten Vektorkassetten etablierten wir die intranodale (i.n.) Transfektion von antigenpräsentierenden Zellen in der Maus. Dazu nutzten wir verschiedene Reportersysteme (eGFP-RNA, fluoreszensmarkierte RNA) und konnten zeigen, dass die intranodale Applikation von IVT-RNA zu selektiven Transfektion und Maturation lymphknotenresidenter DCs führt. Zur Untersuchung der immunologischen Effekte wurden in erster Linie auf Influenza-Hemagglutinin-A und Ovalbumin basierende Modellantigensysteme verwendet. Beide Antigene wurden als Antigen-MHC-Fusionskonstrukte genutzt. Als Responderzellen wurden TCR-transgene Lymphozyten verwendet, die MHC-Klasse-I oder -Klasse-II restringierte Epitope des Influenza-Hemagglutinin-A bzw. des Ovalbumin-Proteins erkennen. Wir konnten in vivo zeigen, dass die intranodale Immunisierung mit IVT-RNA zu einer effizienten Stimulation und Expansion von antigenspezifischen CD4+ und CD8+ T-Zellen in einer dosisabhängigen Weise führt. Funktionell konnte gezeigt werden, dass diese T-Zellen Zytokine sezernieren und zur Zytolyse befähigt sind. Wir waren in der Lage durch repetitive i.n. RNA Immunisierung ein ‚Priming’ CD8+ T-Zellen in naiven Mäusen sowohl gegen virale als auch gegen Tumor assoziierte Antigene zu erreichen. Die geprimten T-Zellen waren befähigt eine zytolytische Aktivität gegen mit spezifischem Peptid beladene Targetzellen zu generieren. Darüber hinaus waren wir in der Lage Gedächtnisszellen expandieren zu können. Abschließend konnten wir in Tumormodellen sowohl in prophylaktischen als auch in therapeutischen Experimenten zeigen dass die i.n. RNA Vakzination die Potenz zur Induktion einer anti-tumoralen Immunität besitzt.

Caratterizzazione genomica di virus influenzali suini in Italia mediante next generation sequencing

I sottotipi H1N1, H1N2 e H3N2 di influenza A virus sono largamente diffusi nella popolazione suina di tutto il mondo. Nel presente lavoro è stato sviluppato un protocollo di sequenziamento di c.d. nuova generazione, su piattaforma Ion Torrent PGM, idoneo per l’analisi di tutti i virus influenzali suini (SIV). Per valutare l’evoluzione molecolare dei SIV italiani, sono stati sequenziati ed analizzati mediante analisi genomica e filogenetica un totale di sessantadue ceppi di SIV appartenenti ai sottotipi H1N1, H1N2 e H3N2, isolati in Italia dal 1998 al 2014. Sono stati evidenziati in sei campioni due fenomeni di riassortimento: tutti i SIV H1N2 esaminati presentavano una neuraminidasi di derivazione umana, diversa da quella dei SIV H1N2 circolanti in Europa, inoltre l’emoagglutinina (HA) di due isolati H1N2 era originata dal riassortimento con un SIV H1N1 avian-like. L’analisi molecolare dell’HA ha permesso di rivelare un’inserzione di due amminoacidi in quattro SIV H1N1 pandemici e una delezione di due aminoacidi in quattro SIV H1N2, entrambe a livello del sito di legame con il recettore cellulare. E’ stata inoltre evidenziata un’elevata omologia di un SIV H1N1 con ceppi europei isolati negli anni ’80, suggerendo la possibile origine vaccinale di questo virus. E’ stato possibile, in aggiunta, applicare il nuovo protocollo sviluppato per sequenziare un virus influenzale aviare altamente patogeno trasmesso all’uomo, direttamente da campione biologico. La diversità genetica nei SIV esaminati in questo studio conferma l’importanza di un continuo monitoraggio della costellazione genomica dei virus influenzali nella popolazione suina.

Improving adoptive T cell therapies of cancer by ectopic expression of miR181a to repress inhibitory phosphatases

Adoptive T cell therapy using antigen-specific T lymphocytes is a powerful immunotherapeutic approach against cancer. Nevertheless, many T cells against tumor-antigens exhibit only weak anti-tumoral response. To overcome this barrier it is necessary to improve the potency and anti-tumoral efficacy of these T cells. Activation and activity of T cells are tightly controlled to inhibit unwanted T cell responses and to reduce the risk of autoimmunity. Both are regulated by extrinsic signals and intrinsic mechanisms which suppress T cell activation. The intrinsic mechanisms include the expression of phosphatases that counteract the activation-inducing kinases. Modifying the expression of these phosphatases allows the targeted modulation of T cell reactivity. MicroRNAs (miRNAs) are regulatory small noncoding RNA molecules that control gene expression by targeting messenger RNAs in a sequence specific manner. Gene-specific silencing plays a key role in diverse biological processes, such as development, differentiation, and functionality. miR181a has been shown to be highly expressed in immature T cells that recognize low-affinity antigens.rnThe present study successfully shows that ectopic expression of miR181a is able to enhance the sensitivity of both murine and human T cells. In CD4+ T helper cells as well as in CD8+ cytotoxic T cells the overexpression of miR181a leads to downregulation of multiple phosphatases involved in the T cell receptor signaling pathway. Overexpression of miR181a in human T cells achieves a co-stimulatory independent activation and has an anti-apoptotic effect on CD4+ T helper cells. Additionally, increasing the amount of miR181a enhances the cytolytic activity of murine CD8+ TCRtg T cells in an antigen-specific manner.rnTo test miR181a overexpressing T cells in vivo, a mouse tumor model using a B cell lymphoma cell line (A20-HA) expressing the Influenza hemagglutinin (Infl.-HA) antigen was established. The expression of model antigens in tumor cell lines enables targeted elimination of tumors using TCRtg T cells. The transfer of miR181a overexpressing Infl.-HA TCRtg CD8+ T cells alone has no positive effect neither on tumor control nor on survival of A20-HA tumor-bearing mice. In contrast, the co-transfer of miR181a overexpressing Infl.-HA TCRtg CD8+ and CD4+ T cells leads to improved tumor control and prolongs survival of A20-HA tumor-bearing mice. This effect is characterized by higher amounts of effector T cells and the expansion of Infl.-HA TCRtg CD8+ T cells.rnAll effects were achieved by changes in expression of several genes including molecules involved in T cell differentiation, activation, and regulation, cytotoxic effector molecules, and receptors important for the homing process of T cells in miR181a overexpressing T cells. The present study demonstrates that miR181a is able to enhance the anti-tumoral response of antigen-specific T cells and is a promising candidate for improving adoptive cell therapy.

Adsorption of Components of the Plasma Kinin-forming System on the Surface of Porphyromonas gingivalis Involves Gingipains as the Major Docking Platforms

Enhanced production of proinflammatory bradykinin-related peptides, the kinins, has been suggested to contribute to the pathogenesis of periodontitis, a common inflammatory disease of human gingival tissues. In this report, we describe a plausible mechanism of activation of the kinin-generating system, also known as the contact system or kininogen-kallikrein-kinin system, by the adsorption of its plasma-derived components such as high-molecular-mass kininogen (HK), prekallikrein (PK), and Hageman factor (FXII) to the cell surface of periodontal pathogen Porphyromonas gingivalis. The adsorption characteristics of mutant strains deficient in selected proteins of the cell envelope suggested that the surface-associated cysteine proteinases, gingipains, bearing hemagglutinin/adhesin domains (RgpA and Kgp) serve as the major platforms for HK and FXII adhesion. These interactions were confirmed by direct binding tests using microplate-immobilized gingipains and biotinylated contact factors. Other bacterial cell surface components such as fimbriae and lipopolysaccharide were also found to contribute to the binding of contact factors, particularly PK. Analysis of kinin release in plasma upon contact with P. gingivalis showed that the bacterial surface-dependent mechanism is complementary to the previously described kinin generation system dependent on HK and PK proteolytic activation by the gingipains. We also found that several P. gingivalis clinical isolates differed in the relative significance of these two mechanisms of kinin production. Taken together, these data show the importance of this specific type of bacterial surface-host homeostatic system interaction in periodontal infections.

Physiologic cold shock of Moraxella catarrhalis affects the expression of genes involved in the iron acquisition, serum resistance and immune evasion

Background Moraxella catarrhalis, a major nasopharyngeal pathogen of the human respiratory tract, is exposed to rapid downshifts of environmental temperature when humans breathe cold air. It was previously shown that the prevalence of pharyngeal colonization and respiratory tract infections caused by M. catarrhalis are greatest in winter. The aim of this study was to investigate how M. catarrhalis uses the physiologic exposure to cold air to upregulate pivotal survival systems in the pharynx that may contribute to M. catarrhalis virulence. Results A 26°C cold shock induces the expression of genes involved in transferrin and lactoferrin acquisition, and enhances binding of these proteins on the surface of M. catarrhalis. Exposure of M. catarrhalis to 26°C upregulates the expression of UspA2, a major outer membrane protein involved in serum resistance, leading to improved binding of vitronectin which neutralizes the lethal effect of human complement. In contrast, cold shock decreases the expression of Hemagglutinin, a major adhesin, which mediates B cell response, and reduces immunoglobulin D-binding on the surface of M. catarrhalis. Conclusion Cold shock of M. catarrhalis induces the expression of genes involved in iron acquisition, serum resistance and immune evasion. Thus, cold shock at a physiologically relevant temperature of 26°C induces in M. catarrhalis a complex of adaptive mechanisms that enables the bacterium to target their host cellular receptors or soluble effectors and may contribute to enhanced growth, colonization and virulence.

Immune response to influenza vaccination in children treated with methotrexate or/and tumor necrosis factor-alpha inhibitors

In children treated with immunosuppressive medication such as methotrexate and tumor necrosis factor-alpha (TNF-α) inhibitors, additional immunizations are recommended because of increased susceptibility to infections. However, it is unclear if adequate antibody response to vaccinations can be established in children receiving methotrexate and/or TNF-α inhibitors. In a prospective open label study, we assessed seroprotection and seroconversion following influenza vaccination during 2 seasons (6 strains) in 36 children with autoimmune disease treated either with methotrexate (n=18), TNF-α inhibitors (n=10) or both (n=8) and a control group of 16 immunocompetent children. Influenza antibody titers were determined by hemagglutinin inhibition assay, before and 4-8 weeks after vaccination. Post-vaccination seroprotection (defined as a titer ≥1:40) did not significantly differ between immunosuppressed and immunocompetent subjects. Seroconversion, defined as the change from a nonprotective (< 1:40) to a protective titer (≥1:40) with at least a 4-fold titer increase, was less likely to occur in immunosuppressed patients, although no significant difference from the control group was established. Safety evaluation of vaccination showed no serious adverse events. Children receiving methotrexate and/or TNF-α inhibitors can be safely and effectively immunized against influenza, with a seroprotection after vaccination comparable to immunocompetent children.