Serological and Molecular Survey of Brucella Species in Owners and Their Dogs Living on Island and Mainland Seashore Areas of Brazil.

Background: Although Brucella abortus, Brucella suis, and Brucella canis may infect humans and dogs worldwide, no study to date has assessed and compared owners and their dogs between island and mainland seashore areas. Materials and Methods: Accordingly, the study herein has applied serological tests, including Microplate Agglutination Test with 2-Mercaptoethanol, immunochromatographic assay, and Rose Bengal Test, and a Brucella genus-specific PCR assay to 195 owners and their 148 dogs living on 1 mainland seashore area and three nearby oceanic islands of southern Brazil. Results: No seropositivity to B. abortus and B. suis was detected in owner or dog sera. Anti-B. canis seropositivity was observed in 3/148 (2.0%) dogs, but no owner sample was seropositive to B. canis. In addition, all blood samples from both owners and dogs were negative on Brucella genus-specific PCR assay. Conclusions: The seropositive dogs were not related and lived on the seashore mainland area of Guaraqueçaba city. The absence of seropositivity on the islands and the low seropositivity on the seashore mainland could be attributed to geographic isolation, and suggest the low impact of the disease in the region. Despite being a zoonotic disease, brucellosis by B. canis is not included in the National Program for Control and Eradication of Brucellosis, and its diagnosis and notification are not mandatory. The presence of seropositive dogs highlights the risk to human health and the importance of epidemiological surveillance actions in the region, as well as the need for the implantation of preventive measures to avoid the transmission of the pathogen.

Combination of the electronic nose with microbiology as a tool for rapid detection of Salmonella.

Salmonella is one of the most important foodborne pathogens and its analysis in raw and processed products is mandatory in the food industry. Although microbiological analysis is the standard practice for Salmonella determination, these assays are commonly laborious and time-consuming, thus, alternative techniques based on easy operation, few manipulation steps, low cost, and reduced time are desirable. In this paper, we demonstrate the use of an e-nose based on ionogel composites (ionic liquid + gelatine + Fe3O4 particles) as a complementary tool for the conventional microbiological detection of Salmonella. We used the proposed methodology for differentiating Salmonella from Escherichia coli, Pseudomonas fluorescens, Pseudomonas aeruginosa, and Staphylococcus aureus in nonselective medium: pre-enrichment in brain heart infusion (BHI) (incubation at 35 °C, 24 h) and enrichment in tryptone soy agar (TSA) (incubation at 35 °C, 24 h), whereas Salmonella differentiation from E. coli and P. fluorescens was also evaluated in selective media, bismuth sulfite agar (BSA), xylose lysine deoxycholate agar (XLD), and brilliant green agar (BGA) (incubation at 35 °C, 24 h). The obtained data were compared by principal component analysis (PCA) and different machine learning algorithms: multilayer perceptron (MLP), linear discriminant analysis (LDA), instance-based (IBk), and Logistic Model Trees (LMT). For the nonselective media, under optimized conditions, taking merged data of BHI + TSA (total incubation time of 48 h), an accuracy of 85% was obtained with MLP, LDA, and LMT, while five separated clusters were presented in PCA, each cluster corresponding to a bacterium. In addition, for evaluation of the e-nose for discrimination of Salmonella using selective media, considering the combination of BSA + XLD and total incubation of 72 h, the PCA showed three separated and well-defined clusters corresponding to Salmonella, E. coli, and P. fluorescens, and an accuracy of 100% was obtained for all classifiers.

Molecular detection of Coxiella burnetii in aborted bovine fetuses in Brazil.

In the past decade, cases of Q fever have been reported in Brazil. Although the previous report of Coxiella burnetii in humans and animals, the knowledge about the occurrence of this pathogen in livestock in Brazil is scarce. This study aimed to search C. burnetii and possible coinfections in tissues of aborted bovine fetuses from Brazil. Tissue samples from seventy-six aborted bovine fetuses sent to the laboratory of molecular diagnosis of infectious diseases from 2013 to 2019 were evaluated by real-time PCR for C. burnetii. Overall, 9.2% (7/76) of the samples were positive for C. burnetii. Moreover, the molecular diagnostic history of our lab revealed the coinfection with Neospora spp. in three fetuses and the presence of histopathological features suggestive with fetal neosporosis in another one. The previous report of C. burnetii in humans and animals in the country, with the detection of C. burnetii from tissues of aborted bovine fetuses reported here, reinforces the neglected state of the disease in Brazil and raises the question of the role of the pathogen in reproductive disorders in national livestock.

Coxiella burnetii in slaughterhouses in Brazil: A public health concern.

Q fever is an important zoonosis, yet it is often neglected and can present large outbreaks, as observed in the Netherlands. In the past few years, cases of Q fever have been described in Brazil; however, the epidemiological situation of Q fever in ruminants, the main reservoir of the pathogen, is unknown in this country. Our study aimed to estimate the prevalence of C. burnetii in cattle sent to slaughterhouses using an immunofluorescence assay (IFA) and quantitative real-time PCR (qPCR). From 1515 cattle serum samples collected from nine slaughterhouses, 23.8% (360/1515) were serologically positive by IFA (cutoff titer>1:64), indicating past or recent exposure to C. burnetii infection. Among the 54 cities sampled during the study, 83.3% (45/54) had at least one seropositive animal. Subsequently, all seropositive samples were submitted to qPCR for C. burnetii DNA, and 12.2% (44/360) of the sera were qPCR positive, which indicates bacteremia and suggests active or recent infection. The results highlight the risk for abattoir workers that results from exposure to contaminated aerosols produced during slaughter procedures. Moreover, the heat maps that were construction from the positive samples demonstrate the widespread distribution of C. burnetii in the State of São Paulo, Brazil and denotes the need for surveillance and preventive measures to reduce the prevalence in cattle.

Real-time quantitative PCR-based detection of Coxiella burnetii in unpasteurized cow's milk sold for human consumption.

Coxiella burnetii is a zoonotic pathogen with a worldwide distribution that is responsible for Q fever in humans. It is a highly infectious bacterium that can be transmitted from cattle to humans through the consumption of unpasteurized milk. We report the molecular identification of C. burnetii in raw cow's milk being sold directly for human consumption in Brazil without official inspection or pasteurization. One hundred and twelve samples of raw milk were analysed by real-time quantitative PCR (qPCR), and C. burnetii was detected in 3.57% (4/112) of the samples at a concentration ranging from 125 to 404 bacteria per millilitre. The identification of this zoonotic pathogen in raw milk sold directly for human consumption is a public health concern since C. burnetii can be transmitted through the oral route. This result indicates that health education and other preventive measures should be officially implemented in Brazil to prevent the spread of infection. To our knowledge, this is the first qPCR-based detection of C. burnetii in raw milk samples from cows sold in Brazil that do not undergo official inspection or pasteurization.

Determination of neutral diols and carboxylic acids formed during glycerol electrooxidation by capillary electrophoresis with dual C4D.

Methods for determination of glycerol and its electrooxidation products (neutral diols and carboxylates) by capillary electrophoresis (CE) with dual capacitively coupled contactless conductivity detectors (C4D) are presented. Glycerol, dihydroxyacetone and glyceraldehyde were detected as anionic borate complexes in less than 3min under counter Electroosmotic Flow (EOF) mode (resolution of the critical pair of 1.8). Limits of detection (LODs) of 15, 15 and 10µmolL-1 were obtained for glycerol, dihydroxyacetone and glyceraldehyde, respectively. Two methods of separation were used for the separation of carboxylates. The first one used the same Back Ground Electrolyte (BGE) containing borate, and the second used a BGE (pH 6.1) composed by 2-(N-morpholino)ethanesulfonic acid (MES), L-Histidine and a flow modifier. Better separation and LODs for carboxylates were obtained using Mes/Histidine as BGE. However, along with the non-applicability of this BGE to the determination of neutral diols, observation of the C4D signals at two different points of the capillary (10 and 50cm apart from the injection tip) revealed interaction of the flow modifier with some species (mesoxalate and glyoxylate). The electrooxidation of a glycerol sample in alkaline media on an 8cm2 gold working electrode was evaluated by the developed methods. After 16h of electrolysis, 87% of the glycerol had been oxidized and formate, glycolate, hydroxypyruvate and glycerate were detected as the main products.