Jane Megid

Clinical manifestations of brucellosis in domestic animals and humans.

Brucellosis in domestic animals is a chronic disease that is characterized mainly by reproductive signs in cattle, buffaloes, pigs, sheep, goats and dogs. In females the disease is characterized by abortion, placenta retention, vaginal secretions, low fertility rate and also embryonic and neonatal death. In males, regular findings include epididymitis, orchitis, uni- or bilateral testicular atrophy, sperm abnormalities and infertility. Lymphadenopathy, hepatopathy, splenomegaly, uveitis and discospondylitis may also be observed in dogs. In horses, the typical clinical sign is characterized by a granulomatous supraspinous or supra-atlantal bursa lesion. Infected animals can also be asymptomatic. Infected symptomatic or asymptomatic animals represent an important source of infection to other animals and humans. Brucellosis in humans can cause undulant fever, malaise, insomnia, anorexia, headache, arthralgia, constipation, sexual impotence, nervousness and depression. For all species the presentation of clinical signs are only suggestive of disease infection and thus must be differentiated from other diseases.

Comparison of agar gel immunodiffusion test, rapid slide agglutination test, microbiological culture and PCR for the diagnosis of canine brucellosis

The performance of the rapid slide agglutination test, with and without 2-mercaptoethanol (RSAT and 2ME-RSAT) and agar gel immunodiffusion test (AGID) was evaluated for the diagnosis of brucellosis in naturally infected dogs. The microbiological culture, PCR and clinical parameters were used as reference. A total of 167 dogs were clinically examined and tested by blood culture, culture of semen/vaginal swab and PCR in blood and semen/vaginal swab. According to the results observed the 167 dogs were divided into three groups: Brucella canis infected dogs (Group 1), B. canis non-infected dogs (Group 2) and dogs with suspected brucellosis (Group 3). The dogs were then tested by RSAT, 2ME-RSAT and AGID. Groups 1 and 2 were used to calculate the diagnostic sensitivity and specificity of the serological tests and the results observed in Group 3 were also discussed. The diagnostic sensitivity of RSAT, 2ME-RSAT and AGID was respectively 70.58%, 31.76%, and 52.94%. The diagnostic specificity of RSAT, 2ME-RSAT and AGID was respectively 83.34%, 100%, and 100%. In dogs with suspected brucellosis 15% were RSAT positive, none was 2ME-RSAT positive and 5% were AGID positive. Although the serological tests are the most commonly used methods for brucellosis diagnosis, a significant proportion of false-negative results were observed highlighting the importance of the direct methods of diagnosis, like blood culture and PCR to improve the diagnosis of canine brucellosis.

Diagnosis of canine brucellosis: Comparison between serological and microbiological tests and a PCR based on primers to 16S-23S rDNA interspacer

A pair of primers directed to 16S-23S rDNA interspacer (ITS) was designed directed to Brucella genetic sequences in order to develop a polymerase chain reaction (PCR) putatively capable of amplifying DNA from any Brucella species. Nucleic acid extracts from whole-blood from naive dogs were spiked with decreasing amounts of Brucella canis RM6/66 DNA and the resulting solutions were tested by PCR. In addition, the ability of PCR to amplify Brucella spp. genetic sequences from naturally infected dogs was evaluated using 210 whole-blood samples of dogs from 19 kennels. The whole-blood samples collected were subjected to blood culture and PCR. Serodiagnosis was performed using the rapid slide agglutination test with and without 2-mercaptoethanol. The DNA from whole blood was extracted using proteinase-K, sodium dodecyl sulphate and cetyl trimethyl ammonium bromide followed by phenol–chloroform purification. The PCR was capable of detecting as little as 3.8 fg of Brucella DNA mixed with 450 ng of host DNA. Theoretically, 3.8 fg of Brucella DNA represents the total genomic mass of fewer than two bacterial cells. The PCR diagnostic sensitivity and specificity were 100%. From the results observed in the present study, we conclude that PCR could be used as confirmatory test for diagnosis of B. canis infection.

Vaccinia Virus Zoonotic Infection, São Paulo State, Brazil

To the Editor: Since 1999, vaccinia virus (VACV) has been isolated frequently from dairy cattle and humans (1–3). During bovine vaccinia outbreaks, VACV can be transmitted to farmers and those who milk cows; it frequently causes lesions on the hands and forearms. Bovine vaccinia causes economic losses and affects public health services in Brazil (1–4). One of the first VACV viruses isolated during Brazilian bovine vaccinia outbreaks was Aracatuba virus (ARAV), which was collected in Sao Paulo State, and since that time, other VACVs have been isolated in this state (2,5,6).

Epidemiological assessment of canine brucellosis

Observou-se brucelose canina em quatro canis diferentes. Os canis apresentaram animais com historia de aborto, mortalidade em neonatos e nascimentos prematuros. A porcentagem de animais soropositivos para brucelose canina, pela prova de imunodifusao em agar gel, variou de 4,6 a 57,1%. Observou-se correlacao positiva entre porcentagem de animais positivos e aspectos reprodutivos e condicoes de aglomeracao.

Serological study of vaccinia virus reservoirs in areas with and without official reports of outbreaks in cattle and humans in São Paulo, Brazil

Vaccinia virus (VACV), the etiological agent of an exanthematic disease, has been associated with several bovine outbreaks in Brazil since the end of the global vaccination campaign against smallpox. It was previously believed that the vaccine virus used for the WHO global campaign had adapted to an unknown wild reservoir and was sporadically re-emerging in outbreaks in cattle and milkers. At present, it is known that Brazilian VACV is phylogenetically different from the vaccinia virus vaccinal strain, but its origin remains unknown. This study assessed the seroprevalence of orthopoxviruses in domestic and wild animals and farmers from 47 farms in three cities in the southwest region of the state of São Paulo with or without official reports of outbreaks in cattle or humans. Our data indicate a low seroprevalence of antibodies in wild animals and raise interesting questions about the real potential of wild rodents and marsupials as VACV reservoirs, suggesting other routes through which VACV can be spread.

Heminested reverse-transcriptase polymerase chain reaction (hnRT-PCR) as a tool for rabies virus detection in stored and decomposed samples

BackgroundThe use of methods, both sensitive and specific, for rabies diagnosis are important tools for the control and prophylaxis of the disease. Reverse-Transcriptase Polymerase Chain Reaction (RT-PCR) has been used in rabies diagnosis with good results, even in decomposed materials. Additionally, molecular techniques have been used for epidemiological studies and to gain a better knowledge of viral epidemiology.FindingsThe aim of this work was to evaluate the RT-PCR and hnRT-PCR for rabies virus detection in original tissues stored at -20°C for different periods considering their use for rabies virus detection in stored and decomposed samples. RT-PCR and hnRT-PCR were evaluated in 151 brain samples from different animal species, thawed and left at room temperature for 72 hours for decomposition.The RT-PCR and hnRT-PCR results were compared with previous results from Direct Fluorescent Antibody Test and Mouse Inoculation Test. From the 50 positive fresh samples, 26 (52%) were positive for RT-PCR and 45 (90%) for hnRT-PCR. From the 48 positive decomposed samples, 17 (34, 3%) were positive for RT-PCR and 36 (75%) for hnRT-PCR. No false-positives results were found in the negatives samples evaluated to the molecular techniques.ConclusionThese results show that the hnRT-PCR was more sensitive than RT-PCR, and both techniques presented lower sensibility in decomposed samples. The hnRT-PCR demonstrated efficacy in rabies virus detection in stored and decomposed materials suggesting its application for rabies virus retrospective epidemiological studies.

First Identification of Canine Distemper Virus in Hoary Fox (Lycalopex vetulus): Pathologic Aspects and Virus Phylogeny

Canine distemper virus (CDV) has been reported in several wild animal species, but there have been no reports of CDV in hoary fox (Lycalopex vetulus). This paper characterizes the first case of natural CDV infection in hoary fox, including the clinical and pathologic aspects of the disease as well as the viral strain phylogeny.

Canine Distemper Virus in a Crab-eating Fox (Cerdocyon thous) in Brazil: Case Report and Phylogenetic Analyses

Although canine distemper is en-zootic worldwide and has a wide host range, there are no reports of canine distemper virus in crab-eating foxes (Cerdocyon thous) that provide information on virus phylogeny and histopathologic lesions. The objective of this study is report and describe canine distemper in a crab-eating fox (C. thous), with a focus on the phylogeny of the virus strain and the histopathologic lesions in the animal.

Rabies virus distribution in tissues and molecular characterization of strains from naturally infected non-hematophagous bats

Bats are main reservoirs for Lyssavirus worldwide, which is an important public health issue because it constitutes one of the big challenges in rabies control. Yet, little is known about how the virus is maintained among bats, and the epidemiological relationships remain poorly understood. The aim of the present study was to investigate the distribution of the rabies virus (RABV) in bat tissues and organs and to genetically characterize virus isolates from naturally infected non-hematophagous bats. The heminested reverse transcriptase polymerase chain reaction (hnRT-PCR) and sequencing using primers to the nucleoprotein coding gene were performed. The results showed a dissemination of the RABV in different tissues and organs, particularly in the salivary glands, tongue, lungs, kidneys, bladder, intestine and feces, suggesting other possible forms of RABV elimination and the possibility of transmission among these animals. The phylogenetic analysis confirmed that different variants of RABV are maintained by non-hematophagous bats in nature and have similar tissue distribution irrespective of bat species and phylogenetic characterization.