RESUMO
A wild peninsula ribbon snake (Thamnophis sauritus sackenii) in Florida was found to have hypochromic erythrocytes containing two different types of inclusions: purple granular inclusions, and pale orange or pink crystalloid inclusions that were round, oval, rectangular, or hexagonal in shape. Transmission electron microscopy revealed hexagonal or pleomorphic, homogenous inclusions and enveloped particles morphologically consistent with a member of the Iridoviridae. Histopathology of the animal revealed necrotizing hepatitis consistent with sepsis. Consensus PCR was used to amplify a 628-bp region of iridoviral DNA-dependent DNA polymerase. Bayesian and maximum likelihood phylogenetic analysis found that this virus was distinct from other known iridoviral genera and species, and may represent a novel genus and species.
Assuntos
Colubridae/virologia , Infecções por Vírus de DNA/veterinária , Eritrócitos/virologia , Iridoviridae/classificação , Iridoviridae/isolamento & purificação , Filogenia , Sequência de Aminoácidos , Animais , Teorema de Bayes , Colubridae/sangue , Infecções por Vírus de DNA/patologia , Infecções por Vírus de DNA/virologia , DNA Viral/química , DNA Viral/genética , Eritrócitos/ultraestrutura , Amplificação de Genes , Iridoviridae/ultraestrutura , Funções Verossimilhança , Microscopia Eletrônica de Transmissão/veterinária , Dados de Sequência Molecular , Reação em Cadeia da Polimerase/veterinária , Alinhamento de SequênciaRESUMO
BACKGROUND: Anaplasma phagocytophilum (formerly known as the human granulocytic ehrlichia, Ehrlichia equi and Ehrlichia phagocytophila) is an obligate intracellular organism causing clinical disease in humans and various species of domestic animals. OBJECTIVES: The objectives of this investigation were to sequence and clone the major surface protein 5 (MSP5) of A phagocytophilum and to evaluate the suitability of this antigen in the serologic diagnosis of anaplasmosis in humans and dogs. METHODS: The msp5 gene of A phagocytophilum was sequenced, cloned, and expressed in Escherichia coli. The predicted amino acid sequence homology of the various MSP5/major antigenic protein 2 orthologs was compared among various Anaplasma and Ehrlichia species. Recombinant MSP5 of A phagocytophilum was used in an ELISA to detect antibodies in serum samples from humans and dogs infected with the organism. RESULTS: Serum samples from 104 individuals previously diagnosed with A phagocytophilum infection, as well as samples from clinically healthy humans, were tested. In addition, multiple samples from 4 dogs experimentally infected with 2 different geographic isolates of A phagocytophilum and 5 dogs naturally infected with a Swiss isolate were tested using ELISA. Using this group of immunofluorescent antibody test-positive and immunofluorescent antibody test-negative samples, we found the overall agreement between assays to be >90%. CONCLUSIONS: These results indicate that recombinant MSP5 has potential for use as a diagnostic test antigen to detect infection with A phagocytophilum in both dogs and humans. However, sequence similarities among orthologs of MSP5 in related species of anaplasma and ehrlichia suggest that cross-reactivity among these pathogens is likely if the entire peptide is used as a test antigen.
Assuntos
Anaplasma phagocytophilum/genética , Anaplasma phagocytophilum/metabolismo , Proteínas da Membrana Bacteriana Externa/genética , Proteínas da Membrana Bacteriana Externa/metabolismo , Doenças do Cão/diagnóstico , Doenças do Cão/microbiologia , Ehrlichiose/veterinária , Sequência de Aminoácidos , Anaplasma phagocytophilum/química , Animais , Clonagem Molecular , Cães , Ehrlichiose/diagnóstico , Ehrlichiose/microbiologia , Humanos , Dados de Sequência Molecular , Especificidade da EspécieRESUMO
Three 3-month-old guinea pigs (Cavia porcellus) were evaluated for purulent ocular discharge. Conjunctival swabs were obtained for cytologic evaluation of Wright's-Giemsa-stained preparations. The specimen from the most severely affected guinea pig consisted primarily of karyolytic neutrophils and small lymphocytes. Epithelial cells occasionally were observed that contained intracytoplasmic coccoid basophilic organisms, 0.5-1.5 microm in diameter. The intraepithelial inclusions were most consistent with Chlamydia sp elementary and reticulate bodies. Specimens from the other 2 guinea pigs had a similar inflammatory response, but organisms were not observed. Polymerase chain reaction (PCR) analysis of a conjunctival swab from the most severely affected guinea pig was positive for C psittaci, which also is referred to as Chlamydophila caviae, immunotype 8, formerly known as the guinea pig inclusion conjunctivitis strain of C psittaci. Chlamydial conjunctivitis is a common problem in guinea pig populations, with C caviae being specific for this species. Cytologic identification of elementary or reticulate bodies within epithelial cells is diagnostic for the organism in Giemsa-stained preparations. However, PCR is an important complementary tool when organisms are not observed and for accurate classification of the Chlamydia species.