Diarrhea outbreaks in suckling piglets due to rotavirus group C single and mixed (rotavirus groups A and B) infections

Porcine group A rotavirus (PoRVA) is a major cause of neonatal diarrhea in suckling and recently weaned piglets worldwide. The involvement of non-group A rotavirus in cases of neonatal diarrhea in piglets are sporadic. In Brazil there are no reports of the porcine rotavirus group C (PoRVC) as etiologic agent of the diarrhea outbreaks in piglets. The aim of this study was to describe the identification of rotavirus group C in single and in mixed infection with rotavirus groups A and B in three neonatal diarrhea outbreaks in suckling (<21-day-old) piglets, with 70% to 80% and 20% to 25% of morbidity and lethality rates, respectively, in three pig herds located in the state of Santa Catarina, Brazil. The diagnosis of PoRV in the diarrheic fecal samples was performed using polyacrylamide gel electrophoresis (PAGE) to identify the presence of porcine rotavirus groups A, B (PoRVB), and C, and by RT-PCR (PoRVA and PoRVC) and semi-nested (SN)-PCR (PoRVB) to partially amplify the VP4 (VP8*)-VP7, NSP2, and VP6 genes of PoRVA, PoRVB, and PoRVC, respectively. One RT-PCR (PoRVA and PoRVC) and SN-PCR (PoRVB) product of each group of rotavirus of each diarrhea outbreak was submitted to nucleotide (nt) sequence analysis. Based on the PAGE technique, 4 (25%) and 1 (6.25%) of the 16 diarrheic fecal samples evaluated in the first outbreak presented PoRVA and PoRVC electropherotype, respectively, and 11 (68.75%) were negative. In the second outbreak, 3 (42.85%) of the 7 fecal samples evaluated presented PoRVA electropherotype, and in 3 (42.85%) and in 1 (14.3%) fecal samples were detected inconclusive and negative results, respectively. Three (30%) of the 10 fecal samples of the third outbreak presented PoRVC electropherotype; 5 (50%) and 2 (20%) samples showed negative and inconclusive results, respectively. Based on the RT-PCR and SN-PCR assays in the first neonatal diarrhea outbreak, PoRVC was detected in 13 (81.2%) of the 16 diarrheic fecal samples evaluated. PoRVC single infection was identified in 4 (25%) of these samples and mixed infections with PoRVA and PoRVB in 9 (56.2%) fecal samples. All of the seven diarrheic fecal samples evaluated from the second neonatal diarrhea outbreak were positive for PoRVC, whereas its mixed infection with other PoRV groups was detected in 4 (57.2%) samples. In the third outbreak, PoRVC in single infection was detected in all of the 10 diarrheic fecal samples analyzed. In the nt sequence analysis, the PoRVA strains of the first and second outbreaks demonstrated higher nt identity with G4P[6] and G9P[23] genotypes, respectively. The PoRVB strains (first and second outbreaks) and the PoRVC strains (first, second, and third outbreaks) showed higher nt identity and clustered in the phylogenetic tree with PoRVB and PoRVC strains that belong to the N4 and I1 genotypes, respectively. This is the first description in Brazil of the involvement of PoRVC in the etiology of diarrhea outbreaks in suckling piglets. The results of this study demonstrated that PoRVC, in both single and mixed infections, is an important enteropathogen involved in neonatal diarrhea outbreaks in piglets and that the use of more sensitive diagnostic techniques allows the identification of mixed infections involving two or even three groups of PoRV, which may be more common than previously reported.

Severe diarrhea outbreak in beef calves (Bos indicus) caused by G6P[11], an emergent genotype of bovine rotavirus group A

The episodes of diarrhea caused by neonatal bovine rotavirus group A (BoRVA) constitute one of the major health problems in the calf rearing worldwide. The main G (VP7) and P (VP4) genotypes of BoRVA strains involved in the etiology of diarrhea in calves are G6P[1], G10P[11], G6P[5], and G8P[1]. However, less frequently, other G and P genotypes have been described in BoRVA strains identified in diarrheic fecal samples of calves. This study describes the identification and molecular characterization of an emerging genotype (G6P[11]) in BoRVA strains involved in the etiology of a diarrhea outbreak in beef calves in a cattle herd of high production in extensive management system. The diarrhea outbreak, which showed high morbidity (60%) and lethality (7%) rates, occurred in calves (n= 384) Nelore (Bos indicus) up to 30-day-old from the State of Mato Grosso do Sul, Brazil. BoRVA was identified in 80% (16/20) of the fecal samples analyzed by polyacrylamide gel electrophoresis (PAGE) technique. In all PAGE-positive fecal samples were amplified products with 1,062-bp and 876-bp in the RT-PCR assays for VP7 (G type) and VP4 (VP8*) (P type) of BoRVA, respectively. The nucleotide sequence analysis of VP7 and VP4 genes of four wild-type BoRVA strains showed G6-III P[11]-III genotype/lineage. The G6P[11] genotype has been described in RVA strains of human and animal hosts, however, in calves this genotype was only identified in some cross-sectional studies and not as a single cause of diarrhea outbreaks in calves with high morbidity and lethality rates as described in this study. The monitoring of the G and P genotypes of BoRVA strains involved in diarrhea outbreaks in calves is important for both animal and public health by allowing the identification of the most frequent genotypes, the characterization of novel genotypes and to identify reassortments with genotypes described in animal and human hosts. The results of this study show the importance of the monitoring of the genotypes of BoRVA strains involved in episodes of bovine neonatal diarrhea as for characterization of frequency of occurrence and pathogenic potential of uncommon genotypes as for monitoring of the emergency of different BoRVA genotypes not included in commercial vaccines.

Identification of canine papillomavirus type 1 (CPV1) DNA in dogs with cutaneous papillomatosis

Canine oral papillomavirus (COPV), also known as Canine Papillomavirus type 1 (CPV1), induces papillomas at the mucous membranes of the oral cavity and at the haired skin of dogs. The classification of Papillomavirus (PV) types is based on the L1 capsid protein and nucleotide sequence; so far, 14 CPV types have been described in several countries, but the molecular characterization of CPV in Brazil is lacking. This study investigated the presence of the PV in seven papillomas from four mixed breed dogs from Londrina/PR, Southern Brazil, by partial sequencing of the L1 gene. Seven exophytic cutaneous lesions were surgically removed and processed for histopathological and molecular characterization. Histopathology confirmed the lesions as viral papillomas due to typical histological features. Polymerase Chain Reaction (PCR) assay using the FAP59 and FAP64 primers targeted the L1 gene followed by sequence analysis of the amplicons identified CPV1 in all evaluated papilloma samples. This study represents the first description of CPV1 DNA associated with canine papillomatosis in Brazil.

Histophilus somni-induced thrombotic meningoencephalitis in cattle from northern Paraná, Brazil

Thrombotic meningoencephalitis (TME) is a fatal neurological disease of cattle, predominantly from North America, that is caused by Histophilus somniwith sporadic descriptions from other countries. This manuscript describes the occurrence of spontaneous TME in cattle from northern Paraná, Brazil. Most cattle had acute neurological manifestations characteristic of brain dysfunction. Hematological and cerebrospinal fluid analyses were not suggestive of bacterial infections of the brain. Histopathology revealed meningoencephalitis with vasculitis and thrombosis of small vessels that contained discrete neutrophilic and/or lymphocytic infiltrates admixed with fibrin at the brainstem, cerebral cortex, and trigeminal nerve ganglion of all animals. All tissues from the central nervous system used during this study were previously characterized as negative for rabies virus by the direct immunofluorescence assay. PCR and RT-PCR assays investigated the participation of infectious agents associated with bovine neurological disease by targeting specific genes of H. somni, Listeria monocytogenes, bovine herpesvirus -1 and -5, bovine viral diarrhea virus, and ovine herpesvirus-2. PCR and subsequent sequencing resulted in partial fragments of the 16S rRNA gene of H. somni from brain sections of all animals with histopathological diagnosis of TME; all other PCR/RT-PCR assays were negative. These findings confirmed the participation of H. somni in the neuropathological disease observed in these animals, extend the geographical distribution of this disease, and support previous findings of H. somni from Brazil.

Histopathological, immunohistochemical, and molecular study of BHV-5 infection in the central nervous system of experimentally infected calves

Bovine meningoencephalitis caused by BHV-5, a double-stranded DNA enveloped virus that belongs to the family Herpesviridae and subfamily Alphaherpesvirinae, is an important differential diagnosis of central nervous diseases. The aim of this study was to describe the histological changes in the central nervous system of calves experimentally infected with BHV-5 and compare these changes with the PCR and IHC results. Formalin-fixed paraffin-embedded central nervous system samples from calves previously inoculated with BHV-5 were microscopically evaluated and tested using IHC and PCR. All the animals presented with nonsuppurative meningoencephalitis. From 18 evaluated areas of each calf, 32.41% and 35.19% were positive by IHC and PCR, respectively. The telencephalon presented more accentuated lesions and positive areas in the PCR than other encephalic areas and was the best sampling area for diagnostic purposes. Positive areas in the IHC and PCR were more injured than IHC and PCR negative areas. The animal with neurological signs showed more PCR- and IHC-positive areas than the other animals.

Molecular survey of porcine teschovirus, porcine sapelovirus, and enterovirus G in captive wild boars (Sus scrofa scrofa) of Paraná state, Brazil

AbstractPorcine teschovirus (PTV), porcine sapelovirus (PSV), and enterovirus G (EV-G) are infectious agents specific to pig host species that are endemically spread worldwide. This study aimed to investigate the natural infection by these porcine enteric picornaviruses in wild boars (Sus scrofa scrofa) of Paraná state, Brazil, and to evaluate peccaries (Pecari tajacu and Tayassu pecari) as alternative host species for these viruses. Fecal samples (n=36) from asymptomatic wild boars (n=22) with ages ranging from 2 to 7 months old (young, n=14) and 2 to 4 years old (adult, n=8) and from peccaries (6 to 8 months old, n=14) were collected from a farm and a zoo, respectively, both located in Paraná state. Reverse transcription-polymerase chain reaction (RT-PCR) and nested-PCR (n-PCR) assays targeting the 5'non-translated region of the virus genome were used for screening the viruses. Porcine enteric picornaviruses were detected in 12 out of the 22 wild boar fecal samples. According to each of the viruses, EV-G was most frequently (11/22, 50%) detected, followed by PTV (10/22, 45.5%) and PSV (4/22, 18.2%). Regarding the age groups, young wild boars were more frequently (9/14, 64.3%) infected with PTV, PSV, and EV-G than adult animals (3/8, 37.4%). One n-PCR amplified product for each of the viruses was submitted to sequencing analysis and the nucleotide sequences were compared with the related viruses, which showed similarities varying from 97.7% to 100% for PTV, 92.4% to 96.2% for PSV, and 87.1% to 100% for EV-G. Peccaries tested negative for the viruses and in this study they did not represent infection reservoirs. This study is the first to report the molecular detection of PTV, PSV, and EV-G from captive wild boars in a South American country and the first to screen peccaries as alternative host species for porcine enteric picornavirus.

Caracterização molecular de DNA de Delta papillomavirus bovino (BPV1, 2 e 13) em sarcoides equinos

Resumo:Sarcoides são tumores fibroblásticos, considerados os tumores de pele mais comuns em pele de equinos e que raramente apresentam regressão espontânea. Papilomavírus bovino (BPV) tipos 1 e 2 são relacionados com a patogenia do sarcoide e, provavelmente, o BPV tipo 13 (BPV13), recentemente descrito, também pode estar associado com a formação dessa lesão. Neste estudo, 20 amostras de lesões cutâneas, sendo 12 constituídas por tecidos frescos e 8 amostras de tecido fixado em formalina e embebido em parafina, provenientes de 15 cavalos foram utilizadas para a identificação do DNA de BPV. A análise histopatológica (HE) confirmou todas as lesões como sarcoide. Para a amplificação do DNA de papilomavírus (PV) foram realizadas três reações de PCR. Como triagem, os primers IFNR2/IDNT2 foram utilizados para amplificar um fragmento da ORF L1 do PV. O segundo par de primersutilizado é complementar a sequência dos genes E5 e L2 de BPVs 1, 2 e 13. O terceiro par de primers(FAP59/FAP64) utilizado tem o gene L1 como alvo. A primeira e a segunda PCRs permitiram amplificar produtos em todas as amostras avaliadas. Entretanto, na terceira reação, na qual foram utilizados os primers FAP, foi possível amplificar produtos com tamanho molecular esperado somente nas amostras constituídas por tecidos frescos. O sequenciamento de nucleotídeos e as análises filogenéticas realizadas nos fragmentos E5L2 resultaram na identificação de BPV1, 2 e 13 em 14 (70%), 2 (10%) e em 4 (20%) amostras de sarcoides, respectivamente. As amostras de sarcoides de um dos animais continha somente o DNA de BPV1. Entretanto, nas amostras provenientes do segundo cavalo foi possível identificar o DNA de três tipos de Deltapapillomavirus bovino (BPV1, 2 e 13) em lesões distintas. Este estudo ratifica a presença do DNA de BPV1, 2 e 13 em lesões de sarcoides em equinos, além de identificar três tipos de BPVs em um mesmo animal e descrever pela primeira vez no Brasil a presença de BPV1 e 2 nesse tipo de lesão.

Detection of cytotoxic activity on Vero cells in clinical isolates of Serratia marcescens

Cytotoxin production was studied in 60 Serratia marcescens strains isolated from hospitalized patients. Association of cytotoxic activity with serotype, source of isolation and presence of plasmids was also evaluated. Thirteen of the 60 S. marcescens strains produced a cytotoxic effect on Vero cells. These strains were isolated from distinct clinical sources and classified into seven different serotypes (O1:H7; O4:NM; O10:NT; O19:NM; O6,14:H4; O6,14:NM and O6,14:H1). No relationship was observed between cytotoxic activity and clinical source or serotypes of the strains. Plasmids from five cytotoxin-producing S. marcescens strains were transferred to E. coli K12/711. The transconjugants did not exhibit cytotoxicity, indicating that the cytotoxic effect is not plasmid-mediated among these strains. Although a cytotoxic activity was demonstrated in filtrates of some S. marcescens strains, further studies should be performed to assess the role of this toxin in pathogenesis

Human group C rotavirus in children with diarrhea in the Federal District, Brazil

Group C rotaviruses are fastidious in their in vitro cell culture requirements. Recent serosurveys indicate that antibody to group C rotavirus is present in 3-45% of the human population in certain geographic locations, suggesting that rotavirus group C infection is more prevalent than previously believed and that the low rate of detection of these agents is probably due to the lack of sensitive diagnostic assays. From March to December 1994, 406 fecal specimens were collected from children under five years of age who were outpatients at the emergency services of nine public hospitals in Brasília, Federal District, Brazil. In addition to the samples from children, one public outpatient unit requested virological investigation of a stool sample from an HIV-seropositive adult male with diarrhea of sudden onset. All samples were analyzed by enzyme immunoassay for group A rotavirus and adenovirus (EIARA) and by polyacrylamide gel electrophoresis (PAGE). One hundred and seven (26%) were positive for group A rotavirus. Four samples from children and the sample from the HIV-seropositive patient, although negative by EIARA, showed a group C rotavirus profile by PAGE and were positive for rotavirus by electron microscopy. Using specific VP6 and VP7 primers for group C rotavirus, a reverse transcriptase-polymerase chain reaction (RT-PCR) was performed and products were detected by agarose gel electrophoresis and ethidium bromide staining. These products were confirmed to be specific for group C rotavirus by using digoxigenin-oligonucleotide probes, Southern hybridization and chemiluminescent detection. The five positive group C rotavirus samples were detected in August (3 samples) and September (2 samples). To the best of our knowledge, this is the first report of group C rotavirus detected in the Federal District, Brazil and in an HIV-seropositive patient with acute gastroenteritis.

Preservation of graft function in low-risk living kidney transplant recipients treated with a combination of sirolimus and cyclosporine

The use of sirolimus (SRL) in combination with full doses of cyclosporin A (CsA) results in reduced one-year kidney allograft function, which is associated with shorter long-term allograft survival. We determined the effect of reduced CsA exposure on graft function in patients receiving SRL and prednisone. Ninety recipients of living kidney transplants receiving SRL (2 mg/day, po) were compared to 35 recipients receiving azathioprine (AZA, 2 mg kg-1 day-1, po). All patients also received CsA (8-10 mg kg-1 day-1, po) and prednisone (0.5 mg kg-1 day-1). Efficacy end-point was a composite of biopsy-confirmed acute rejection, graft loss, or death at one year. Graft function was measured by creatinine, creatinine clearance, and graft function deterioration between 3 and 12 months (delta1/Cr). CsA concentrations in patients receiving SRL were 26% lower. No differences in one-year composite efficacy end-point were observed comparing SRL and AZA groups (18 vs 20%) or in the incidence of biopsy-proven acute rejection (14.4 and 14.3%). There were no differences in mean ± SD creatinine (1.65 ± 0.46 vs 1.60 ± 0.43 mg/dl, P = 0.48) or calculated creatinine clearances (61 ± 15 vs 62 ± 13 ml/min, P = 0.58) at one year. Mean ± SD delta1/Cr (-11 ± 17 vs -14 ± 15%, P = 0.7) or the percentage of patients with >20% (26 vs 31%, P = 0.6) or >30% delta1/Cr (19 vs 17%, P = 1) did not differ between the two groups. The use of 2-mg fixed oral doses of SRL and reduced CsA exposure was effective in preventing acute rejection and preserving allograft function.

Molecular analysis of the bovine coronavirus S1 gene by direct sequencing of diarrheic fecal specimens

Bovine coronavirus (BCoV) causes severe diarrhea in newborn calves, is associated with winter dysentery in adult cattle and respiratory infections in calves and feedlot cattle. The BCoV S protein plays a fundamental role in viral attachment and entry into the host cell, and is cleaved into two subunits termed S1 (amino terminal) and S2 (carboxy terminal). The present study describes a strategy for the sequencing of the BCoV S1 gene directly from fecal diarrheic specimens that were previously identified as BCoV positive by RT-PCR assay for N gene detection. A consensus sequence of 2681 nucleotides was obtained through direct sequencing of seven overlapping PCR fragments of the S gene. The samples did not undergo cell culture passage prior to PCR amplification and sequencing. The structural analysis was based on the genomic differences between Brazilian strains and other known BCoV from different geographical regions. The phylogenetic analysis of the entire S1 gene showed that the BCoV Brazilian strains were more distant from the Mebus strain (97.8% identity for nucleotides and 96.8% identity for amino acids) and more similar to the BCoV-ENT strain (98.7% for nucleotides and 98.7% for amino acids). Based on the phylogenetic analysis of the hypervariable region of the S1 subunit, these strains clustered with the American (BCoV-ENT, 182NS) and Canadian (BCQ20, BCQ2070, BCQ9, BCQ571, BCQ1523) calf diarrhea and the Canadian winter dysentery (BCQ7373, BCQ2590) strains, but clustered on a separate branch of the Korean and respiratory BCoV strains. The BCoV strains of the present study were not clustered in the same branch of previously published Brazilian strains (AY606193, AY606194). These data agree with the genealogical construction and suggest that at least two different BCoV strains are circulating in Brazil.

Genetic analysis of the porcine group B rotavirus NSP2 gene from wild-type Brazilian strains

Group B rotaviruses (RV-B) were first identified in piglet feces, being later associated with diarrhea in humans, cattle, lambs, and rats. In human beings, the virus was only described in China, India, and Bangladesh, especially infecting adults. Only a few studies concerning molecular analysis of the RV-B NSP2 gene have been conducted, and porcine RV-B has not been characterized. In the present study, three porcine wild-type RV-B strains from piglet stool samples collected from Brazilian pig herds were used for analysis. PAGE results were inconclusive for those samples, but specific amplicons of the RV-B NSP2 gene (segment 8) were obtained in a semi-nested PCR assay. The three porcine RV-B strains showed the highest nucleotide identity with the human WH1 strain and the alignments with other published sequences resulted in three groups of strains divided according to host species. The group of human strains showed 92.4 to 99.7% nucleotide identity while the porcine strains of the Brazilian RV-B group showed 90.4 to 91.8% identity to each other. The identity of the Brazilian porcine RV-B strains with outer sequences consisting of group A and C rotaviruses was only 35.3 to 38.8%. A dendrogram was also constructed to group the strains into clusters according to host species: human, rat, and a distinct third cluster consisting exclusively of the Brazilian porcine RV-B strains. This is the first study of the porcine RV-B NSP2 gene that contributes to the partial characterization of this virus and demonstrates the relationship among RV-B strains from different host species.

A sintaxe da violência entre Hegel e Marx

Trata-se de investigar a tese marxiana acerca do papel da violência na história, tal como enunciada em O Capital, analisando sua sintaxe de matriz hegeliana e o modo como Engels articula tal tese, para então defender uma interpretação não-teleológica da violência, segundo a qual esta apresenta uma pluralidade de formas, um caráter totalmente difuso e uma pesada materialidade.

Escatologia à la cantonade: Althusser para além de Derrida

O autor propõe um confronto entre Althusser e Derrida acerca da questão da temporalidade, focalizando a leitura que os dois autores fazem de Marx sub specie theatri. A partir desse confronto, emerge uma teoria da temporalidade althusseriana, a qual se encontra para além, quer da teleologia da tradição hegeliano-marxista, quer do messianismo sem Messias proposto por Derrida, em sua releitura benjaminiana de Marx. Nesse sentido, a escatologia em Althusser é pensável somente à la cantonade.