Ana M. S. Guimaraes

Ehrlichiosis in Brazil.

Erliquiose e uma doenca causada por rickettsias pertencentes ao genero Ehrlichia. No Brasil, estudos sorologicos e moleculares tem avaliado a ocorrencia de especies de Ehrlichia em caes, gatos, animais selvagens e seres humanos. Ehrlichia canis e a principal especie em caes no Brasil, embora a infeccao por E. ewingii tenha, recentemente, despertado suspeita em cinco caes. O DNA de E. chaffeensis foi detectado e caracterizado em cervo-do-pantanal, enquanto que E. muris e E. ruminantium ainda nao foram identificadas no Brasil. A erliquiose monocitica canina causada pela E. canis parece ser altamente endemica em muitas regioes do Brasil, embora dados de prevalencia nao estejam disponiveis em muitas delas. O DNA de E. canis tambem foi detectado e caracterizado em tres gatos domesticos, enquanto anticorpos contra E. canis foram detectados em felideos neotropicais de vida livre. Evidencias sorologicas sugerem a ocorrencia de erliquiose humana no Brasil, entretanto, o agente etiologico ainda nao foi identificado. A melhoria do diagnostico molecular promovera a identificacao e caracterizacao de especies associadas a erliquiose humana no Brasil.Ehrlichiosis is a disease caused by rickettsial organisms belonging to the genus Ehrlichia. In Brazil, molecular and serological studies have evaluated the occurrence of Ehrlichia species in dogs, cats, wild animals and humans. Ehrlichia canis is the main species found in dogs in Brazil, although E. ewingii infection has been recently suspected in five dogs. Ehrlichia chaffeensis DNA has been detected and characterized in mash deer, whereas E. muris and E. ruminantium have not yet been identified in Brazil. Canine monocytic ehrlichiosis caused by E. canis appears to be highly endemic in several regions of Brazil, however prevalence data are not available for several regions. Ehrlichia canis DNA also has been detected and molecularly characterized in three domestic cats, and antibodies against E. canis were detected in free-ranging Neotropical felids. There is serological evidence suggesting the occurrence of human ehrlichiosis in Brazil but its etiologic agent has not yet been established. Improved molecular diagnostic resources for laboratory testing will allow better identification and characterization of ehrlichial organisms associated with human ehrlichiosis in Brazil.

Exploratory study of Mycoplasma suis (Eperythrozoon suis) on four commercial pig farms in southern Brazil

Mycoplasma suis (Eperythrozoon suis) was detected by pcr and Southern blot in 186 pigs (121 sows, 61 piglets and four boars) on four farms in southern Brazil. dna was extracted from blood samples and a 16S rrna gene fragment of M suis was amplified by pcr; Southern blot analysis was then performed on all the samples. Twenty-two of the sows (18·2 per cent) were positive by pcr, and 40 (33·1 per cent) were positive by Southern blot; only one piglet and one boar were positive. The packed cell volume and total plasma protein of the pigs and their pcr and Southern blot results were not significantly different on the four farms, but higher proportions of the pigs were positive by Southern blot than by pcr (P<0·05). The packed cell volume and total plasma protein concentrations of the M suis positive and negative sows were not significantly different.

Mycoplasma ovis in captive cervids: prevalence, molecular characterization and phylogeny.

Hemotrophic mycoplasmas (hemoplasmas) are bacteria that attach to red blood cells of mammals, leading to acute and/or subclinical disease in infected animals. It has been suggested that Mycoplasma ovis, a hemoplasma that infects sheep and goats worldwide, may also infect deer. The aim of this study was to evaluate whether South American deer are infected with M. ovis. EDTA-anticoagulated blood samples from a herd of 32 captive South American deer were collected. DNA extraction of blood samples was performed followed by PCR amplification of the 16S and 23S rRNA genes, and sequencing of products. Using M. ovis PCR, 27/31 (87%) were positive, including 21/22 Mazama nana; 2/3 Mazama americana and 4/6 Blastocerus dichotomus. Sequencing of the nearly entire 16S rRNA gene of 26/27 positive samples showed 98.2-98.8% identity to M. ovis of sheep (GenBank, AF338268) and 98.6-99.4% identity to M. ovis-like of a fawn (FJ824847); the 23S rRNA gene from one of these isolates and the fawns had 97.6% identity. The remaining isolate had just 94.9% identity to the 16S rRNA gene of M. ovis and only 89.4% identity to the 23S rRNA gene of the fawns M. ovis. This is the first report of M. ovis in captive South American deer, revealing a high prevalence of hemoplasma infection in these animals.

Complete Genome Sequences of Two Hemotropic Mycoplasmas, Mycoplasma haemofelis Strain Ohio2 and Mycoplasma suis Strain Illinois

We report the complete and fully assembled genomes of Mycoplasma haemofelis strain Ohio2 and Mycoplasma suis strain Illinois, which are the first available genomes of these uncultivatable hemoplasma species. The single circular chromosomes of 1,152,484 bp and 742,431 bp for M. haemofelis and M. suis, respectively, are typical of mycoplasma species, having reduced size and low G+C content (38.8% for M. haemofelis and 31.1% for M. suis). Their metabolic pathways are reduced, with evidence of adaption to the blood environment.

Co-infection with Mycoplasma haemofelis and ‘Candidatus Mycoplasma haemominutum’ in three cats from Brazil☆

The two most common haemotropic Mycoplasma of cats, Mycoplasma haemofelis and ‘Candidatus Mycoplasma haemominutum’ have been identified using molecular techniques in all continents, except Antarctica. We report the first molecular characterization in South America of a dual infection with M haemofelis and ‘Candidatus Mycoplasma haemominutum’ in three domestic cats. The 16S ribosomal RNA gene was amplified in three anaemic cats in which haemoplasma organisms were seen attached to the erythrocytes in the peripheral blood smear. Bands of the expected size for M haemofelis and ‘Candidatus Mycoplasma haemominutum’ were observed in all three cats. The 393 bp segment of one of the amplicons had a similarity value of 100% to M haemofelis, whereas the other amplicon, a 192 bp segment, was 100% similar to ‘Candidatus Mycoplasma haemominutum’. After diagnosis, two cats received blood transfusion and they were all treated with doxycycline. All three cats recovered uneventfully.

Survey of Owned Feline and Canine Populations in Apartments from a Neighbourhood in Curitiba, Brazil

Animal population estimates are essential for public health services to ensure the success of zoonoses control programmes. Canine and feline populations vary among different regions mainly because of local human income status and type of human residence. Accordingly, the present study estimated the pet population size living in apartments of a vertical neighbourhood in Curitiba, Brazil. We chose a neighbourhood with a predominance of apartment buildings. All apartment buildings were visited, and questionnaires were completed by doormen or residents. Data were obtained from 120 of 173 apartment buildings. Survey questions included the number of apartments, residents, dogs and cats. Two thousand nine hundred and sixty six apartments with a total of 7429 residents were surveyed. The number of dogs and cats was 569 and 86 respectively. Thus, the human : dog and human : cat ratios were 13.05 : 1 and 86.38 : 1. These ratios were higher than those observed in other neighbourhoods in Curitiba. The present study indicates that the number of pets from apartments may be different from houses, and different among distinct areas within the same city.

Identification of a Mycoplasma ovis-like organism in a herd of farmed white-tailed deer (Odocoileus virginianus) in rural Indiana

Mycoplasma ovis is a hemoplasma parasite of sheep, goats, and reindeer; however, natural hemoplasma infection in white-tailed deer has not previously been reported. Subsequent to finding many coccoid, bacillary, and ring-shaped organisms, consistent with hemotropic mycoplasmas, on RBCs from a 72-day-old female white-tailed fawn, we sought to (1) identify the putative hemoplasma observed in blood from the fawn, (2) evaluate others in the herd for hemoplasma infection, and (3) identify clinicopathologic characteristics of hemoplasma-infected white-tailed deer. EDTA-anticoagulated whole blood was collected from the fawn and 8 apparently healthy does in the same herd. CBCs were performed on 7 nonclotted samples from the fawn and 6 does. DNA was extracted from all samples, followed by PCR amplification of bacterial (16S rDNA) and protozoal (18S rDNA) genes. The nearly complete 16S rDNA product from the fawns sample was directly sequenced and compared with known sequences in the GenBank database. Samples from the fawn and 7 of 8 does were PCR-positive using hemoplasma-specific and M ovis-specific protocols. The fawn was PCR-negative for Anaplasma spp., Babesia spp., and Theileria spp. The 16S rDNA sequence from the fawn (GenBank accession number, FJ824847) was most closely related to M ovis (AF338268), having 98.5% sequence identity. The fawn had a mild nonregenerative anemia, a neutrophilic left-shift with toxic change, aspiration bronchopneumonia, and gastrointestinal disease. Hematologic values, including blood film evaluation, in infected does were unremarkable. The M ovis-like organism may have acted as either an opportunistic or primary pathogen in the fawn. The high occurrence of subclinical infections in the does suggests that white-tailed deer may act as wildlife reservoirs for M ovis.

A quantitative TaqMan PCR assay for the detection of Mycoplasma suis

Aim:  To develop a TaqMan probe‐based, highly sensitive and specific quantitative PCR (qPCR) assay for the detection and quantification of Mycoplasma suis in the blood of pigs.

Invasion of Ureaplasma diversum in Hep-2 cells

BackgroundUnderstanding mollicutes is challenging due to their variety and relationship with host cells. Invasion has explained issues related to their opportunistic role. Few studies have been done on the Ureaplasma diversum mollicute, which is detected in healthy or diseased bovine. The invasion in Hep-2 cells of four clinical isolates and two reference strains of their ureaplasma was studied by Confocal Laser Scanning Microscopy and gentamicin invasion assay.ResultsThe isolates and strains used were detected inside the cells after infection of one minute without difference in the arrangement for adhesion and invasion. The adhesion was scattered throughout the cells, and after three hours, the invasion of the ureaplasmas surrounded the nuclear region but were not observed inside the nuclei. The gentamicin invasion assay detected that 1% of the ATCC strains were inside the infected Hep-2 cells in contrast to 10% to the clinical isolates. A high level of phospholipase C activity was also detected in all studied ureaplasma.ConclusionsThe results presented herein will help better understand U. diversum infections, aswell as cellular attachment and virulence.

Serological survey of Toxoplasma gondii in captive Neotropical felids from Southern Brazil

Toxoplasma gondii is the causative intracellular protozoan of toxoplasmosis in human being and animals. Members of the Felidae family are considered the single definitive host for the infection; both wild and domestic cats are able to excrete oocysts in the environment. Wild cats maintained in captivity may serve as source of infection for other clinically susceptible animals in the same environment. The aim of this study was to determine the frequency of T. gondii IgG antibodies in 57 neotropical felids (1 Leopardus geoffroyi; 3 Puma yagouaroundi; 17 Leopardus wiedii; 22 Leopardus tigrinus; and 14 Leopardus pardalis) kept at the Bela Vista Biological Sanctuary, Itaipu Binacional, Southern Brazil, by the modified agglutination test (MAT) using titer 16 as cut-off point. Seropositivity was observed in 38/57 (66.67%; 95% CI 53.66-77.51%) samples, with higher frequency in ocelots (71.43%). Wild-caught felids were three times more likely to be infected when compared to zoo-born animals (P<or=0.05) and age of wild-caught animals (P=0.6892; 95% CI=0.7528-1.66) was not significant as a risk factor for the infection, the same occurring with zoo-born animals (P=0.05; 95% CI=0.6267-24.052). These results suggest that, despite efforts to control T. gondii infection in zoo facilities, such as individual pens, hygiene monitoring, veterinary care and pre-frozen meat offered as food, non-domestic felids kept in captivity, particularly the wild-caught specimens, may be invariably exposed to infection due to other environmental sources.