Antibiofilm activity of the biosurfactant and organic acids against foodborne pathogens at different temperatures, times of contact, and concentrations.

Biofilm formation has been suggested to play a significant role in the survival of pathogens in food production. Interest in evaluating alternative products of natural origin for disinfectant use has increased. However, there is a lack of information regarding the effects of biosurfactants and organic acids on Salmonella enterica serotype Enteritidis, Escherichia coli, and Campylobacter jejuni biofilms, mainly considering temperatures found in environments of poultry processing, as well as simulating the contact times used for disinfection. The aim of this study was to evaluate the antibiofilm activity of rhamnolipid, malic acid, and citric acid on the adhesion of S. Enteritidis, E. coli, and C. jejuni on polystyrene surfaces at different temperatures (4, 12, and 25 °C), compound concentrations, and times of contact (5 and 10 min), and to analyze the potential use of these compounds to disrupt formed biofilms. All three compounds exhibited antibiofilm activity under all analyzed conditions, both in the prevention and removal of formed biofilms. Contact time was less important than temperature and concentration. The antibiofilm activity of the compounds also varied according to the pathogens involved. In the food industry, compound selection must consider the temperature found in each stage of product processing and the target pathogens to be controlled.

Bacterial community identification in poultry carcasses using high-throughput next generation sequencing.

Poultry products are susceptible to contamination by pathogenic and spoilage bacteria during the slaughtering process. Molecular techniques have been used to assist in the identification of microorganisms in various microbiomes. The aim of this study was to identify bacterial components of the microbiome in poultry carcasses during the slaughter process, using high-throughput next generation sequencing (HT-NGS). Samples were collected from three slaughterhouses (A, B, and C) located in southern Brazil and included those taken from three points (initial, middle, and end) in the chiller tanks and two carcass pools (at the entrance to the clean area and after the final carcass packaging) at each establishment. A total of 104 carcasses were collected from each slaughterhouse. For this study, HT-NGS allows for a precise, quantitative and culture-independent microbiome assessment in poultry products. Three phyla (Firmicutes, Bacteroidetes, and Proteobacteria) were found in all establishments, and one phylum (Verrucomicrobia) was found only in Establishment A. Common set of genera (Anaerotruncus, Bacteroides, Campylobacter, Erysipelatoclostridium, Faecalibacterium, Lachnoclostridium, and Subdoligranulum) was identified in processing establishments along with the groups unique to a particular site. Pathogenic and spoilage bacteria, as well as other microorganisms that were not expected in poultry products, were detected by HT-NGS technique. The Shannon diversity index was the highest in Establishment B (2.40), followed by establishments C (1.98) and A (1.43). As we progressed through sample analysis, from the entrance of the clean area to the final carcass packaging area, we found significant reductions (p < 0.05) in the quantities of sequences of all phyla in establishments A and B. Significant differences (p < 0.05) in the quantities of sequences of all phyla were found between different stages in the slaughtering process. More stringent control procedures in establishments A and B were associated with reduced contamination even though all establishments followed the official sanitary standards. Our findings provide new insight into the chicken meat microbiome, and can be used in future studies to help ensure food safety in slaughterhouses.

Antimicrobial susceptibility and detection of virulence-associated genes of Escherichia coli and Salmonella spp. isolated from domestic pigeons (Columba livia) in Brazil.

Overpopulation of domestic pigeons is considered to be one of the major problems of urban centers, as these birds are responsible for the dissemination of relevant pathogens to animal and human health. The aim of this study was to detect potentially pathogenic Escherichia coli and Salmonella spp. in domestic pigeons captured in areas near silos used for grain and feed storage, analyzing the antimicrobial sensitivity and the presence of virulence-associated genes. We evaluated 41 pigeons. From each bird, cecal contents and a pool of viscera (heart, spleen, and liver) were collected. Fifty strains of E. coli and three strains of S. Typhimurium were isolated. The antimicrobial susceptibility assay showed that 2% of the isolates of E. coli were resistant to chloramphenicol and the combination of sulfamethoxazole + trimethoprim and 4% to tetracycline, doxycycline, and sulfonamide. The three S. Typhimurium strains were sensitive to all antimicrobials tested. The pathogenicity profile demonstrated that no E. coli isolates showed a STEC compatible profile. Regarding the APEC pathotype, all genes were observed in 8% of E. coli, 6% had only the iss gene and 4% presented ompT, hlyF, and iutA genes. invA, hilA, avrA, and lpfA genes were detected in 100% of Salmonella isolates. The sitC and pefA genes were only present in one strain and the remaining genes were detected in two. In conclusion, it was found that pigeons living in the vicinity of silos are carriers of important pathogens, and control measures should be taken to minimize animal and human health risks.

Occupational Health and Safety in Aquaculture: Insights on Brazilian Public Policies.

Aquaculture has many occupational hazards, including those that are physical, chemical, biological, ergonomic, and mechanical. The risks in aquaculture are inherent, as this activity requires particular practices. The objective of the present study was to show the risks associated with the aquaculture sector and present a critical overview on the Brazilian public policies concerning aquaculture occupational health. Methods include online research involved web searches and electronic databases including Pubmed, Google Scholar, Scielo and government databases. We conducted a careful revision of Brazilian labor laws related to occupational health and safety, rural workers, and aquaculture. The results and conclusion support the idea that aquaculture requires specific and well-established industry programs and policies, especially in developing countries. Aquaculture still lacks scientific research, strategies, laws, and public policies to boost the sector with regard to occupational health and safety. The establishment of a safe workplace in aquaculture in developing countries remains a challenge for all involved in employer-employee relationships.

Aplicação da técnica de RT-PCR para metapneumovírus aviário (aMPV) / Application of RT-PCR technique for avian metapneumovirus (aMPV) / Aplicación de la técnica RT-PCR para metapneumovirus aviario (aMPV)

Os principais hospedeiros do Metapneumovírus aviário (aMPV) são os frangos de corte e perus. O vírus acomete o trato respiratório superior dos perus desencadeando a Rinotraqueíte Viral dos Perus (RVP). O principal objetivo deste trabalho foi padronizar uma técnica de RT-PCR para a detecção do aMPV, por meio do uso do kit AccessQuick™ RT-PCR system (Promega®). Foram utilizados amostras de suabes de traqueia e pulmão de 38 perus comerciais com sintomatologia respiratória e dois suabes oculares de faisão. O RNA viral foi extraído utilizando-se o kit RTP® DNA/RNA Virus Mini Kit (STRATEC Molecular). Em seguida as amostras foram submetidas à RT-PCR One Step, utilizando o kit AccessQuick™ RT-PCR system (Promega®). Todas as 40 amostras testadas por RT-PCR foram negativas, exceto a amostra vacinal que foi utilizada como controle positivo. O aMPV não causa latência em frangos de corte ou perus, logo a excreção viral é limitada. Dessa forma, a ausência da detecção de genoma viral neste estudo pode ser justificada devido à idade que as amostras foram coletadas em perus, com 140 dias no abatedouro, impossibilitando dessa maneira a amplificação do genoma do aMPV. Porém, esse estudo também mostra que a RT-PCR se mostrou eficaz para detectar o genoma viral do aMPV, podendo dessa forma ser utilizado como uma ferramenta de diagnóstico rápido para investigação e estudo de casos de aMPV em rebanho de perus.

The main hosts of Avian metapneumovirus (aMPV) are broilers and turkeys. This virus affects the upper respiratory tract of turkeys, triggering Turkey Rhinotracheitis (TRT). The aim of this study was to optimize a RT-PCR technique in order to detect aMPV using the AccessQuick™ RT-PCR system (Promega®) kit. Tracheal and lung swab samples from 38 commercial turkeys with respiratory symptoms and two ocular swabs from pheasants were analyzed. Viral RNA was extracted using RTP® DNA/RNA Virus Mini Kit (Molecular STRATEC) kit. All 40 samples tested were negative in the RT-PCR. The only positive sample was a vaccine strain, used as the positive control. The aMPV does not cause latency in broilers, chickens or turkeys, thus, the viral excretion is limited. However, the absence of viral genome detection in this study may be justified due to the age the samples were collected, since they were collected in turkeys with about 140 days in the slaughterhouse, thus preventing the amplification of the aMPV genome. This study shows that the RT-PCR is effective to detect aMPV viral genome and may be used as a rapid diagnostic tool for research and for the studying of aMPV cases in turkey flocks in Brazil.

Los principales anfitriones de Metapneumovirus aviario (aMPV) son los pollos de engorde y pavos. El virus afecta el tracto respiratorio superior de los pavos desencadenando la Rinotraqueitis Viral de los pavos (RVP). El principal objetivo de ese estudio fue estandarizar una técnica de RT-PCR para la detección del aMPV, a través del uso del kit AccessQuick™-PCRsystem (Promega®). Se utilizaron muestras de hisopos traqueales y pulmonares de 38 pavos comerciales con síntomas respiratorios y dos hisopos oculares de faisán. El RNA viral se extrajo utilizando el kit DNA RTP® DNA/RNA Virus Mini Kit (STRATEC Molecular). A continuación, las muestras se sometieron a la RT-PCR OneStep utilizando el kit AccessQuick™ RT-PCR (Promega®). Todas las 40 muestras analizadas por RT-PCR fueron negativas, excepto la muestra de vacuna que se utilizó como control positivo. El aMPV no causa latencia en pollos de engorde o pavos, por lo que la excreción viral es limitada. Así, la ausencia de la detección de genoma viral en este estudio puede ser justificada debido a la edad que se recogieron las muestras en los pavos, con 140 días en el matadero, imposibilitando de este modo la amplificación del genoma del aMPV. Sin embargo, ese estudio también muestra que la RT-PCR se ha demostrado eficaz para detectar el genoma viral del aMPV, pudiendo así ser utilizado como una herramienta de diagnóstico rápido para investigación y estudio de casos de aMPV en bandada de pavos.

Antimicrobial susceptibility and pathogenicity of Escherichia coli strains of environmental origin / Suscetibilidade antimicrobiana e patogenicidade de amostras de Escherichia coli de origem ambiental

The study aimed to evaluate the antimicrobial susceptibility of 109 samples of Escherichia coli (E. coli) of environmental origin and to characterize these isolates according to the degree of pathogenicity in vivo, verifying a possible relationship between this variable and susceptibility to the active principles tested. The isolates were subjected to disc diffusion test to 14 antibiotics. From 16.5% to 90% of the samples were sensitive; 1 - 28.5% showed intermediate degree of susceptibility and between 9 to 78% of E. coli analyzed were resistant. The highest resistance percentages were seen in the class of quinolones and tetracyclines (>75%), and for sensitivity in the class of amphenicols (68.8%). By inoculating 1- day - old chicks, the isolates were classified as highly pathogenic (2.7%), intermediate (10.1%), low (42.2%) and apathogenic (45%). It was observed a wide variation in the susceptibility profile of isolates in relation to antimicrobials. It was also found that most of the samples had pathogenic potential (55%), thus being considered as APEC (avian pathogenic E. coli). No relationship between pathogenicity and antimicrobial susceptibility (P≤0.05) was observed.

O estudo teve como objetivo avaliar a suscetibilidade antimicrobiana de 109 amostras de Escherichia coli (E. coli) de origem ambiental frente a antibióticos e caracterizar esses isolados quanto ao grau de patogenicidade in vivo, verificando-se uma possível relação entre esta variável e a suscetibilidade aos princípios ativos testados. Os isolados foram submetidos ao teste de disco-difusão para 14 antibióticos. Entre 16.5% a 90% das amostras foram sensíveis, 1-28.5% apresentaram grau de suscetibilidade intermediário e entre 9-78% das E. coli analisadas foram resistentes. Os maiores percentuais de resistência foram encontrados para a classe das quinolonas e das tetraciclinas (>75%), e de sensibilidade para a classe dos anfenicóis (68.8%). Por meio da inoculação em pintinhos de um dia de idade, os isolados foram classificados como sendo de patogenicidade alta (2.7%), intermediária (10.1%), baixa (42.2%) e apatogênicos (45%). Foi observada uma ampla variação no perfil de suscetibilidade das amostras frente aos antimicrobianos. Verificou-se também que a maioria apresentou potencial patogênico (55%), sendo, portanto, consideradas APEC (E. coli patogênica para aves). Não foi observada relação entre a patogenicidade e a suscetibilidade aos antimicrobianos (P≤0.05).

Laparoscopic-assisted treatment of pyometra associated with mammary fibroadenomatous hyperplasia in a cat / Tratamento de piometra associada à hiperplasia mamária fibroadenomatosa por videolaparoscopia assistida em um gato

This paper describes a case of laparoscopic-assisted ovariohysterectomy in a female cat presenting pyometra and mammary fibroadenomatous hyperplasia. Using four portals, mesovarium were ligated by titanium ligature clips whereas the uterine vessels were occluded by video-assisted conventional ligatures. There were no postoperative complications. Video-assisted technique can be an alternative method for treatment of pyometra and cystic endometrial hyperplasia in female cats.

Este artigo descreve um caso de ovariohisterectomia por laparoscopia assistida em uma gata com sinais de piometra e hyperplasia mamária fibroadenomatosa. Usando quatro portais, os mesovários foram ligados com grampos de ligadura de titânio enquanto que os vasos uterinos foram ocluídos por ligaduras convencionais vídeo assistidas. Não houve complicações pós-operatórias. Esta técnica pode ser uma alternativa para tratamento de piometra e hiperplasia cística endometrial em gatas.

The constitutive expression of the V gene of Parainfluenza virus 5 affects the growth properties of bovine herpesvirus 5

This study aimed to analyze the effect of the expression of Parainfluenza virus 5 (PIV5) V protein in bovine cells on the replication of Bovine herpesvirus 5 (BoHV-5). Growth properties of BoHV-5 were evaluated in parental and PIV5 transfected cells. In one-step growth experiments, the BoHV-5 reached higher titers at earlier time points in the transfected cells when compared to the parental cells. The mean plaque size produced by the BoHV-5 in transfected cells was larger than the parental cells. This indicated that the expression of the PIV5 V gene facilitated the release and cell-to-cell spread of BoHV-5 in bovine cells.

Detection of Alphacoronavirus in velvety free-tailed bats (Molossus molossus) and Brazilian free-tailed bats (Tadarida brasiliensis) from urban area of Southern Brazil.

A survey was carried out in search for bat coronaviruses in an urban maternity roost of about 500 specimens of two species of insectivorous bats, Molossus molossus and Tadarida brasiliensis, in Southern Brazil. Twenty-nine out of 150 pooled fecal samples tested positive by reverse transcription-PCR contained fragments of the RNA-dependent RNA polymerase gene of coronavirus-related viruses. The sequences clustered along with bat alphacoronaviruses, forming a subcluster within this group. Our findings point to the need for risk assessment and continued surveillance of coronavirus infections of bats in Brazil.

Bovine herpesviruses do not play a major role in the differential diagnosis of rabies in cattle in southern Brazil

Background: Rabies has long been recognized as the major cause of encephalitis in cattle in Latin American countries. It has been estimated that nearly 50.000 cattle heads per year are lost due to encephalitis in that subcontinent, with a significant economic impact on cattle productive chains. In Brazil only, 2.500 to 3.000 cattle heads are estimated to be lost every year due to rabies. However, it is believed that rabies incidence in cattle is much larger, since usually only a few samples from affected animals in disease outbreaks are submitted to diagnostic laboratories. Rabies encephalitis is promptly and accurately diagnosed; however, particularly when rabies is excluded as causa mortis, the agent responsible for neurological disease of infectious origin often remains undetermined. Two bovine herpesviruses (BoHVs), bovine herpesvirus type 1 (BoHV-1) and bovine herpesvirus type 5 (BoHV-5) are major pathogens of cattle which are widely disseminated in Brazil. As usual in herpesvirus' biology, these tend to infect a large number of hosts and establish lifelong latent infections which may occasionally be reactivated. Both viruses, particularly BoHV-5, are often recovered from cases of neurological disease in cattle. The participation of BoHVs in the differential diagnosis of rabies must be evaluated. Besides, there might be associations between the occurrence of rabies and BoHV infections that deserve investigation. The aim of this study was to investigate whether bovine herpesvirus 1 and 5 would play a significant role in cases of neurological disease where rabies was the presumptive clinical diagnosis. In addition, associations between the occurrence of rabies and BoHV infections were searched for. The approach adopted for conducting such investigations was based on the search for viral nucleic acids as well as classical virus isolation on tissues of cattle submitted to rabies diagnosis over a two-year period, including rabies-positive and rabies-negative specimens. Materials, Methods & Results: Brain tissue samples of 101 cattle originally submitted to rabies diagnosis were collected over a two year period (2009-2010) from various municipalities within the state of Rio Grande do Sul, Brazil. Thirty nine of these samples had the diagnosis of rabies confirmed by standard laboratory diagnostic methods. Aliquots of tissues were submitted to DNA extraction and examined in search for genomes of bovine herpesviruses (BoHV) types 1 (BoHV-1) and 5 (BoHV-5) by as well as for infectious virus. Bovine herpesvirus genomes were detected in 78/101 (77.2%) samples, in which BoHV-1 genomes were detected in 26/78 (25.7%), BoHV-5 genomes in 22/78 (21.8%) and mixed BoHV infections (BoHV-1 and BoHV-5 genomes) were detected in 30/101 (29.7%) samples. In the 39 samples with confirmed rabies diagnosis, BoHV-1 DNA was detected in 9/39 (23%), BoHV-5 DNA in 6/39 (15.4%) and mixed infections with both BoHV types in 16/39 (41%) samples. However, no infectious herpesvirus was recovered from any of the specimens examined. Discussion: The high prevalence of BoHV1 and BoHV-5 infections was evidenced in the sampled population, but the absence of infectious BoHVs indicate that these were not associated to the occurrence of the cases of encephalitis where rabies was the primary suspicion. In addition, no association was detected between occurrence of rabies and detection of BoHVs, since the frequency of detection of herpesvirus genomes did not significantly differ between rabies-positive and rabies-negative samples. The detection of BoHV DNA in scattered areas of the brain with no infectious virus suggests that latency may take place in different regions of the brain.