Ornithobacterium rhinotracheale: An Update Review about An Emerging Poultry Pathogen.

Respiratory diseases in birds generate sanitary and economic impacts and may be related to the environment and climate. Mycoplasma gallisepticum, Ornithobacterium rhinotracheale (ORT), Pasteurella multocida, Avibacterium paragallinarum, Escherichia coli, Riemerella anatipestifer, and Bordetella avium are among the most important avian respiratory pathogens. ORT is responsible for causing ornitobacteriosis, a disease characterized by clinical signs ranging from mild to severe respiratory conditions, with high mortality rates, mainly affecting turkeys and chickens. The first report of ornitobacteriosis was in 1981 in Germany. Despite its importance, few studies on ORT have been published. In addition, the presence of this pathogen has been neglected in poultry farms, mainly due to the lack of appropriate diagnostic protocols. The lack of correct isolation and diagnostic protocols along with inappropriate use of antimicrobial agents have been contributing to treatment failure. Due to its economic importance to the poultry industry, ornitobacteriosis should be monitored and included in national programs for the prevention and control of avian respiratory diseases. This review aimed to update and discuss important issues related to ORT since this pathogen has great economic and sanitary implications for the chicken production chain.

Characterization of epidemic clones of Listeria monocytogenes serotype 4b isolated from humans and meat products in Brazil.

INTRODUCTION: Listeria monocytogenes is an important foodborne pathogen and the 4b serotype is responsible for many cases of human listeriosis reported in Brazil. Several listeriosis outbreaks worldwide have involved a small number of well-defined clonal groups, designated as epidemic clones (ECs). METHODOLOGY: We studied 71 strains of serotype 4b, including 25 isolates from human cases of listeriosis and 46 from meat-based foods, collected in Brazil between 1977 and 2010. The presence of ECs (I and II) markers and virulence genes (inlA, inlB, ilnC, inlJ and actA) were evaluated by PCR assay. The genetic relationship of ECs-positive strains was assessed by pulsed field gel electrophoresis. RESULTS: ECI and ECII markers were found both in human and food strains, with 19.7% positive for the ECI marker and 40.8% for ECII. Most strains (97.2%) were positive for the virulence genes that were studied. Nevertheless, the actA gene amplicons showed two distinct sizes, with all ECI positive strains exhibiting a 105bp deletion. Pulsed field gel electrophoresis (PFGE) analysis allowed the recognition of highly related strains, particularly from two outbreaks of neonatal listeriosis in São Paulo State occurred in 1992 and 1997, both ECII-positive; and two ECI strains from a human case (1982) and from bovine meat (2009). CONCLUSIONS: The presence of ECs among clinical samples and beef isolates of serotype 4b from some regions of Brazil highlights the need for rigorous control of production procedures. Furthermore, the association of ECII with two nosocomial outbreaks suggests its ability to spread in these settings.

Genetic diversity of Ehrlichia canis strains from naturally infected dogs in Rio de Janeiro, Brazil.

The aim of this study was to characterize Ehrlichia canis strains from naturally infected dogs in Rio de Janeiro, Brazil. In addition, all the clinical and hematological findings observed in these dogs were reported. PCR targeting the 16S rRNA gene was used for diagnostic purposes, and the TRP19 and TRP36 genes were sequenced to evaluate the genetic diversity. Fifteen samples were positive for E. canis. The polymerase chain reaction for the TRP19 gene resulted in 11 amplicons (11/15), which were cloned into the pGEM-T easy vector for sequencing. The complete sequence of TRP19 gene was compared to those in the GenBank, revealing high identicalness. Phylogenetic analysis on the TRP36 gene sequences demonstrated two distinct strains from two dogs, named 56C and 70C. The 56C strain was grouped with the strain Cuiaba 16, which is a hybrid strain formed by Brazilian and US genogroups; and the 70C strain was grouped with other strains of the US genogroup, thus suggesting that there are at least two genogroups of E. canis in Rio de Janeiro (US and Brazilian). Those animals, in which the 70C and 56C strains were isolated, showed distinct clinical and hematological manifestations of the disease. The appearance of different genotypes may express new phenotypes, thus resulting in different forms of presentation of the disease and making its diagnosis more complex.

Genetic diversity of Ehrlichia canis strains from naturally infected dogs in Rio de Janeiro, Brazil / Diversidade genética de cepas de Ehrlichia canis encontradas em cães naturalmente infectados no Rio de Janeiro, Brasil

The aim of this study was to characterize Ehrlichia canis strains from naturally infected dogs in Rio de Janeiro, Brazil. In addition, all the clinical and hematological findings observed in these dogs were reported. PCR targeting the 16S rRNA gene was used for diagnostic purposes, and the TRP19 and TRP36 genes were sequenced to evaluate the genetic diversity. Fifteen samples were positive for E. canis. The polymerase chain reaction for the TRP19 gene resulted in 11 amplicons (11/15), which were cloned into the pGEM-T easy vector for sequencing. The complete sequence of TRP19 gene was compared to those in the GenBank, revealing high identicalness. Phylogenetic analysis on the TRP36 gene sequences demonstrated two distinct strains from two dogs, named 56C and 70C. The 56C strain was grouped with the strain Cuiaba 16, which is a hybrid strain formed by Brazilian and US genogroups; and the 70C strain was grouped with other strains of the US genogroup, thus suggesting that there are at least two genogroups of E. canis in Rio de Janeiro (US and Brazilian). Those animals, in which the 70C and 56C strains were isolated, showed distinct clinical and hematological manifestations of 1the disease. The appearance of different genotypes may express new phenotypes, thus resulting in different forms of presentation of the disease and making its diagnosis more complex.

O objetivo deste estudo foi caracterizar as cepas de Ehrlichia canis em cães naturalmente infectados no Rio de Janeiro, Brasil. Além disso, os achados clínicos e hematológicos observados nos cães foram relatados. O gene 16S rRNA foi utilizado como alvo da PCR para fins diagnósticos, e os genes TRP19 e TRP36 para avaliar a diversidade genética. Quinze amostras foram positivas para E. canis. PCR para o gene TRP19 produziu 11 amplicons (11/15) que foram clonados no pGEM-T easy vector para sequenciamento. A comparação das sequências completas do gene TRP19 com outras sequências depositadas no GenBank revelou uma alta identidade. Duas amostras (56C e 70C) após o ensaio da PCR, tendo como alvo o gene TRP36, geraram sequências, e a análise filogenética mostrou que a cepa 56C foi agrupada com a cepa Cuiabá 16, que é uma cepa híbrida, formada pelo genogrupo Brasileiro e o genogrupo US; e a cepa 70C agrupou com as outras cepas do genogrupo US, sugerindo a existência de pelo menos dois genogrupos de E. canis no Rio de Janeiro (US e Brasileiro). Esses animais apresentaram manifestações clínicas e hematológicas distintas, e diferentes genótipos podem expressar novos fenótipos, resultando em diferentes formas de apresentação da doença e fazendo com que o diagnóstico seja mais complexo.

Cytauxzoon felis and 'Candidatus Mycoplasma haemominutum' coinfection in a Brazilian domestic cat (Felis catus).

This article describes the first detection of Cytauxzoon felis, using molecular techniques, in a naturally infected domestic cat from Brazil, South America. Coinfection with 'Candidatus Mycoplasma haemominutum' was also found. The molecular identification of the piroplasmid species was performed by Polymerase Chain Reaction (PCR) and sequencing analysis. A 284 pb fragment of the gene encoding the 18S ribosomal RNA region was amplified and showed 99% identity with other C. felis strains from North America. In addition, PCR-RFLP (restriction fragment length polymorphism) analysis, which amplifies a 595 bp fragment of the gene encoding 16S ribosomal RNA of some bacterial species, identified the co-infecting species as 'Candidatus M. haemominutum'.

Cytauxzoon felis and 'Candidatus Mycoplasma haemominutum' coinfection in a brazilian domestic cat (Felis catus) / Co-infecção por Cytauxzoon felis e 'Candidatus Mycoplasma haemominutum' em um gato doméstico (Felis catus) no Brasil

This article describes the first detection of Cytauxzoon felis, using molecular techniques, in a naturally infected domestic cat from Brazil, South America. Coinfection with 'Candidatus Mycoplasma haemominutum' was also found. The molecular identification of the piroplasmid species was performed by Polymerase Chain Reaction (PCR) and sequencing analysis. A 284 pb fragment of the gene encoding the 18S ribosomal RNA region was amplified and showed 99% identity with other C. felis strains from North America. In addition, PCR-RFLP (restriction fragment length polymorphism) analysis, which amplifies a 595 bp fragment of the gene encoding 16S ribosomal RNA of some bacterial species, identified the co-infecting species as 'Candidatus M. haemominutum'.

Este artigo descreve a primeira detecção de Cytauxzoon felis em um gato doméstico naturalmente infectado no Brasil, América do Sul, através de técnicas moleculares. Também foi encontrada co-infecção com 'Candidatus Mycoplasma haemominutum'. A detecção molecular da espécie do piroplasmídeo foi realizada através da reação em cadeia pela polimerase (PCR) e sequenciamento. Um fragmento de 284 pb do gene codificador da região 18S do RNA ribossomal do parasito foi sequenciada e mostrou 99% de identidade com outros isolados de C. felis da América do Norte. Ademais, através da análise por meio de PCR-RFLP (Polimorfismo no comprimento de fragmentos de restrição), que amplifica um fragmento de 595 pb do gene codificador da porção 16 do RNA ribossomal de algumas espécies de bactérias, concluiu-se que a espécie com-infectante era 'Candidatus M. haemominutum'.

Detection and molecular characterization of piroplasms species from naturally infected dogs in southeast Brazil.

Rangelia vitalii is a protozoon described from dogs in the south and southeast regions of Brazil. It is phylogenetically related to Babesia spp. that infects dogs, but data on this enigmatic parasite is still limited. The aim of this work was to detect piroplasm species in dogs in the state of Rio de Janeiro, Brazil, by 18S rRNA gene-based PCR assay, restriction fragment length polymorphism (RFLP) and sequence analyses. Of 103 dogs examined, seven (6.8%) were positive for Babesia spp. by PCR. The amplified products were digested by restriction enzymes to differentiate the Babesia species, and one sample was identified as Babesia vogeli. The pattern observed for the other six amplification products did not match with pattern described for large Babesia infecting dogs. Sequencing analysis confirmed these six samples as R. vitalii, with high homologies (99-100%) with a sequence from south Brazil. This study confirms the presence of Babesia vogeli and Rangelia vitalii circulate in domestic dogs in Teresópolis, Rio de Janeiro, Brazil.

Detection and molecular characterization of piroplasms species from naturally infected dogs in southeast Brazil / Detecção e caracterização molecular de piroplasmas em cães naturalmente infectados no Sudeste do Brasil

Rangelia vitalii is a protozoon described from dogs in the south and southeast regions of Brazil. It is phylogenetically related to Babesia spp. that infects dogs, but data on this enigmatic parasite is still limited. The aim of this work was to detect piroplasm species in dogs in the state of Rio de Janeiro, Brazil, by 18S rRNA gene-based PCR assay, restriction fragment length polymorphism (RFLP) and sequence analyses. Of 103 dogs examined, seven (6.8%) were positive for Babesia spp. by PCR. The amplified products were digested by restriction enzymes to differentiate the Babesia species, and one sample was identified as Babesia vogeli. The pattern observed for the other six amplification products did not match with pattern described for large Babesia infecting dogs. Sequencing analysis confirmed these six samples as R. vitalii, with high homologies (99-100%) with a sequence from south Brazil. This study confirms the presence of Babesia vogeli and Rangelia vitalii circulate in domestic dogs in Teresópolis, Rio de Janeiro, Brazil.

Rangelia vitalii é um protozoário que infecta cães e foi descrito nas regiões Sul e Sudeste do Brasil. R. vitalii é filogeneticamente próxima à Babesia spp., mas dados deste misterioso parasito ainda são escassos. O objetivo deste trabalho foi detectar a presença de piroplasmas em cães naturalmente infectados no estado do Rio de Janeiro, através da amplificação do gene 18S rRNA pela PCR, clivagem com enzimas de restrição (RFLP) e caracterização genética através do sequenciamento. De 103 cães, sete (6,8%) foram positivos para Babesia spp. pela PCR. Os produtos amplificados foram digeridos por enzimas de restrição para a diferenciação das espécies de Babesia e uma amostra foi identificada como Babesia vogeli. O padrão de amplificação observado nas outras seis amostras não correspondeu ao padrão descrito para babesias que infectam cães. O sequenciamento das seis amostras confirmou ser uma espécie geneticamente idêntica a R. vitalii apresentando grande homologia (99-100%) com a sequência do sul do Brasil. Este estudo confirma a presença de Babesia vogeli e Rangelia vitalii infectando cães em Teresópolis, Rio de Janeiro, Brasil.

Characterization of atypical Enteropathogenic Escherichia coli (aEPEC) isolated from dogs.

Enteropathogenic Escherichia coli (EPEC), an important human pathogen has the ability to form attaching and effacing lesions on the intestinal epithelium and has been isolated from a wide range of species. Two EPEC subgroups are recognized: typical (tEPEC) and atypical (aEPEC) strains, differing by the presence of EAF plasmid and bundle-forming pilus (BFP) in typical strains and their absence in atypical strains. This study searched the presence of EPEC strains in 101 fecal samples of diarrheic (n=65) and non-diarrheic (n=36) dogs from two cities in Rio de Janeiro State, Brazil. The isolates were evaluated for the presence of eae, tir, espA, espB and bfpA genes, EAF plasmid, and for the insertion site of the LEE locus. Cell-adherence assays, fluorescent actin staining (FAS) test, hemolysin production and serotyping were also performed. Twenty eight aEPEC isolates were recovered from 48 eae-positive fecal samples, 24 from diarrheic animals and 4 from non-diarrheic ones. PCR showed that most isolates was positive for ß or γ intimin, and for ß or α subtypes of tir, espA and espB. Six isolates showed a selC insertion of locus LEE. Only two isolates from the same diarrheic animal harbored the bfpA gene, and none presented the EAF plasmid. Most isolates was FAS-positive and showed a localized adherence-like (LAL) in a 6h HeLa cell-adherence assay. A wide diversity of serotypes was detected including O4:H16 and O51:H40, previously described in human disease. The phenotypic and genotypic markers of aEPEC isolated from diarrheic and non-diarrheic dogs were similar to those found in isolates recovered from human disease.

Bartonelose: análise molecular e sorológica em gatos do Rio de Janeiro Brasil / Bartonellosis: molecular and serological analysis in cats from Rio de Janeiro – Brazil

Bartonella henselae e mais recentemente B. quintana têm sido apontados como agentes causais de diversas moléstias emhumanos, entre as quais a doença da arranhadura do gato, endocardite, meningoencefalite e neuroretinite, podendo levarao óbito, principalmente os imunocomprometidos. O gato doméstico é considerado o principal animal envolvido na transmissãodestes patógenos. Constituiu-se objetivo deste estudo a avaliação da frequência de Bartonella spp. em gatos domésticosdomiciliados do município de Vassouras (RJ) comparando-se os achados na reação em cadeia pela polimerase (PCR) e nasorologia por imunofluorescência indireta (IFA). Amostras sanguíneas de 37 (100%) gatos de um abrigo da cidade deVassouras (RJ) foram analisadas, sendo 36 (97,3%) positivas na PCR para Bartonella spp. Das amostras PCR positivas,nove (25%) e 27 (75%) apresentaram, respectivamente, reatividade e ausência de reatividade ao IFA. Apenas uma (2,7%)amostra de sangue foi concomitantemente negativa na PCR e IFA para Bartonella spp. Este é o primeiro registro de infecçãopor Bartonella spp. em felinos domésticos no estado do Rio de Janeiro (Brasil) identificada por análise molecular e sorológica,o que nos permite concluir que este agente zoonótico está presente em alta frequência em gatos domésticos do municípiode Vassouras (RJ).

Bartonella henselae and B. quintana have been pointed as causal agents of many diseases in humans, and can lead to death, mainly immunodefficient people. Domestic cat is considered the unique animal in transmission of these pathogens. The purpose of this study was to evaluate the frequency of Bartonella spp. in domestic cats from Vassouras city (RJ) by polimerase chain reaction (PCR) and indirect immunofluorescence test assay (IFA) and compare the results. Blood samples from 37 (100%) domestic cats from a shelter of Vassouras city (RJ) were analyzed and 36 (97.3%) were considered positive by polymerase chain reaction (PCR). The indirect immunofluorescence test assay (IFA) revealed 9 (25.0%) and 27 (75.0%) of that PCR positive samples showed, respectively, reaction and absence of reaction to IFA. Only one sample (2.7%) was negative in PCR and IFA. This is the first communication of Bartonella spp. infection in domestic cats in Rio de Janeiro State (Brazil) identified by molecular and serological assays, thus it can be concluded that this zoonotic agent is present in high frequencies in domestic cats from Vassouras city (RJ).

Tratamento isoterápico de dermatofilose em eqüino: relato de caso / Isotherapic treatment of dermatophytosis in a horse: case report

Relata-se um caso de dermatofilose em eqüino puro-sangue inglês de 11 anos, macho, castrado, de 540 kg, com histórico de dermatopatia recorrente há dois anos, previamente tratado com penicilina e banhos de solução à base de iodo povidona.. Foram realizados exames físico e parasitológico de raspado de pele, assim como cultura e antibiograma dos exsudatos presentes nas lesões. À inspeção, foram detectadas marcante emaciação, anidrose na garupa, lesões cutâneas exsudativas, áreas de alopecia localizadas na garupa, dorso e boleto, como também a presença de descamação furfurácea e pêlo fosco. Os resultados da cultura e do antibiograma permitiram detectar a presença de Dermatophilus congolensis, com sensibilidade ao Ceftiofur sódico. Apesar do resultado do antibiograma, optou-se por tratamento isoterápico à base de autonosódio,observando-se melhora clínica do paciente. O resultado obtido com a utilização do isoterápico nos chama atenção para as possibilidades de utilização de terapias não convencionais, em situações clínicas especiais, como a verificada neste caso.

This is a report on a dermatophytosis case affecting an Thoroughbred horse. The animal was a gelded, 540-kg male of 11 years old showing a recurrent dermatopathy for two years, which has been previously treated using penicillin and baths with a povidone-iodine solution. The animal was submitted to physical examination, culture and antibiogram of lesion exudates and parasitological examination of skin scraping. Inspection of the animal showed it to be emaciated; it also showed anhydrosis onthe croup, exsudative cutaneous lesions, localized areas of alopecia on the croup, back and fetlock, as well as furfuraceous scaling and dry hair. Culture and antibiogram showed the presence of Dermatophilus congolensis sensitive to Ceftiofur sodium.In spite of the antibiogram results, an isotherapic treatment was determined based on nosode, with clear clinical improvement. Results obtained with the use of an isotherapic compound shows the possibility of following non-conventional therapies inspecial clinical situations, such as the one reported here.

Shigatoxigenic Escherichia coli serogroups O157, O111 and O113 in feces, water and milk samples from dairy farms

Este trabalho teve como objetivo determinar a prevalência de Escherichia coli Shigatoxigênicas dos sorogrupos O157, O111 e O113 em bovinos leiteiros, água e leite de propriedades rurais do Município de Jaboticabal/SP. Para isso, foram colhidas 454 amostras de fezes, 54 amostras de água e 30 amostras de leite e pesquisada a presenca de seqüências stx1, stx2 e eae pela reacão em cadeia da polimerase (PCR). Todas as amostras stx e/ eae positivas foram submetidas a uma nova reacão de PCR para deteccão das seqüências rfb O157, rfb O111 e rfb O113. Uma alta ocorrência (59,9%) de stx foi detectada nas fezes dos bovinos. Os coeficientes de prevalência das seqüências rfb O157, O111 e O113 nas fezes dos bovinos foram, respectivamente, 18,9%, 3,3% e 30,4%. Todas as seqüências foram encontradas com maior freqüência em bezerros e novilhas. As seqüências eae e stx2 foram as mais detectadas entre as amostras de fezes. Foram detectados 1,9% e 3,3% de seqüências stx na água e no leite, respectivamente. Nas amostras de água observou-se uma baixa prevalência de O113 e não foram detectadas O157 e O111. Nenhum dos sorogrupos pesquisados foi encontrado nas amostras de leite. STEC foram identificadas em todos os rebanhos (100%) e os sorogrupos O157, O111 e O113 foram observados em 40,0%, 50,0% e 90,0% dos rebanhos. Conclui-se que a prevalência de STEC em fezes de bovino, no Município de Jaboticabal/SP é alta, caracterizando-a como importante via de contaminacão ambiental para esses microrganismos.

Prevalence of Ehrlichia canis infection in thrombocytopenic dogs from Rio de Janeiro, Brazil.

BACKGROUND: Infection with Ehrlichia canis causes a highly variable, multisystemic disease in dogs. Nevertheless, many clinicians in Rio de Janeiro, Brazil, use the presence of only thrombocytopenia to make a presumptive diagnosis of E canis infection. OBJECTIVE: The objective of this study was to determine the prevalence of E canis in thrombocytopenic dogs from Rio de Janeiro, Brazil, using polymerase chain reaction (PCR). METHODS: Following DNA extraction of whole blood samples from 226 dogs, PCR assays were done using primers for rickettsial DNA (including Ehrlichia spp, Anaplasma platys and A phagocytophilum) and using E canis-specific primers (16S rRNA gene). Dogs were grouped as thrombocytopenic and nonthrombocytopenic based on platelet counts. The null hypothesis that there was no difference in the prevalence of E canis in these groups was rejected at P<.05. RESULTS: Thirty-six (32.1%) of the thrombocytopenic dogs and 4 (3.5%) of the nonthrombocytopenic dogs were positive for rickettsial gene sequences (P<.0001). Further, 30 (26.8%) of thrombocytopenic dogs and 4 (3.5%) nonthrombocytopenic dogs were positive for E canis-specific gene sequences (P<.0001). CONCLUSIONS: Although the prevalence of E canis infection was higher in thrombocytopenic dogs, less than one third of these dogs had demonstrable E canis infection. Thus, thrombocytopenia is not specific for the detection of E canis infection and should not be used solely to establish a diagnosis of canine ehrlichiosis, even in a geographic area with relatively high disease prevalence.

Escherichia coli produtora de toxina Shiga: prevalência no reservatório animal, marcadores de virulência e análise da estrutura clonal / Shiga toxin-producing Escherichia coli (STEC): prevalence in animal reservoir, virulence markers and analysis of clonal structure

Dentre as diversas categorias diarreiagênicas de Escherichia coli, as amostras produtoras de toxina Shiga (STEC) destacam-se pela potencial letalidade, principalmente em crianças, decorrente de graves seqüelas extra-intestinais e pelo importante papel de animais, especialmente bovinos, como reservatórios destas bactérias para o ser humano. Apesar da relativa baixa ocorrência de STEC como agente de diarréia humana em nosso meio, dados prévios revelaram a predominância desta categoria em produtos cárneos comercializados no Rio de Janeiro. O presente estudo buscou avaliar em relação a STEC a sua ocorrência em bovinos no Estado do Rio de Janeiro, caracterizar a presença de mercadores de virulência em amostras previamente isoladas de alimentos e analisar a possível estrutura clonal de amostras isoladas de diferentes origens em nosso meio. Uma elevada ocorrência de STEC foi caracterizada pela amplificação por PCR de seqüências genéticas de stx em 71 por cento das 197 amostras fecais de bovinos analisadas, sendo maior em animais de regime leiteiro (82 por cento) quando comparada a animais de corte (53 por cento). Constatou-se ainda, após ensaios de hibridização com sondas genéticas, a predominância da presença simultânea das toxinas Stx1 e Stx2 (58 por cento) nas amostras amplificadas. Através da utilização de meio seletivo (TC-SMAC), foi possível o isolamento de 3 amostras STEC pertencentes ao sorotipo 0157:H7 provenientes de 3 animais distintos, sendo este o primeiro relato do isolamento de amostras deste sorotipo no Brasil. Doze amostras STEC adicionais foram obtidas a partir de amostras fecais positivas ufilizando-se um protocolo de isolamento baseado em PCR. A predominância do genótipo stxl/stx2 foi confinada entre as amostras isoladas. A expressão da citotoxina foi caracterizada em todas as amostras à exceção de 2 pertencentes ao sorotipo 0157:H7. A caracterização de mercadores de virulência foi analisada em 21 amostras STEC de origem alimentar acrescidas de 8 amostras de origem bovina (5 isoladas de animais diarréicos e as 3 amostras 0157:H7 isoladas neste estudo) e 4 amostras isoladas de crianças diarréicas. De modo geral as amostras de origem humana e animal apresentaram um perfil de virulência compatível com aquele descrito para o subgrupo mais virulento das STEC, constituído pelas amostras enterohemorrágicas (EHEC), principalmente pela presença do gene eae, da produção de enterohemolisina e por uma interação eficiente e característica ...(au)