Fecal bacterial communities of wild black capuchin monkeys (Sapajus nigritus) from the Atlantic Forest biome in Southern Brazil are divergent from those of other non-human primates.

Gut microbiota are influenced by factors such as diet, habitat, and social contact, which directly affect the host's health. Studies related to gut microbiota in non-human primates are increasing worldwide. However, little remains known about the gut bacterial composition in wild Brazilian monkeys. Therefore, we studied the fecal microbiota composition of wild black capuchin monkey (Sapajus nigritus) (n=10) populations from two different Atlantic Forest biome fragments (five individuals per fragment) in south Brazil. The bacterial community was identified via the high-throughput sequencing and partial amplification of the 16S rRNA gene (V4 region) using an Ion Personal Genome Machine (PGMTM) System. In contrast to other studies involving monkey microbiota, which have generally reported the phyla Firmicutes and Bacteroidetes as predominant, black capuchin monkeys showed a high relative abundance of Proteobacteria ( χ ¯ = 80.54%), followed by Firmicutes ( χ ¯ = 12.14%), Actinobacteria ( χ ¯ = 4.60%), and Bacteriodetes ( χ ¯ = 1.31%). This observed particularity may have been influenced by anthropogenic actions related to the wild habitat and/or diet specific to the Brazilian biome's characteristics and/or monkey foraging behavior. Comparisons of species richness (Chao1) and diversity indices (Simpson and InvSimpson) showed no significant differences between the two groups of monkeys. Interestingly, PICRUSt2 analysis revealed that metabolic pathways present in the bacterial communities were associated with xenobiotic biodegradation and the biosynthesis of secondary metabolites, which may suggest positive effects on monkey health and conservation in this anthropogenic habitat. Infectious disease-associated microorganisms were also observed in the samples. The present study provides information about the bacterial population and metabolic functions present in fecal microbiota, which may contribute to a better understanding of the ecology and biology of black capuchin monkeys living in forest fragments within the Atlantic Forest biome in southern Brazil. Additionally, the present study demonstrates that the fecal bacterial communities of wild black capuchin monkeys in this area are divergent from those of other wild non-human primates.

Respiratory microbiota of healthy broilers can act as reservoirs for multidrug-resistant Escherichia coli.

This study aimed at evaluate the presence and to study characteristics of Escherichia coli in the respiratory system microbiota of healthy broilers. Trachea, air sacs, and lungs of 20 broilers were analyzed at 21 days of age, reared in experimental conditions, without receiving antimicrobials. E. coli strains were isolated and identified using conventional bacteriology through morphological and biochemical characterization. The production of bacteriocin-like substances, the presence of virulence-associated genes (VAGs) of APEC (Avian Pathogenic Escherichia coli) predictors, and the antimicrobial susceptibility were evaluated. E. coli was found in 85 % of the animals (17/20), in the trachea, air sacs or lungs; and it was not found in 15 % of the animals (3/20). A total of 34 isolates were recovered, 13 from the air sacs, 13 from the lungs, and 8 from the trachea, which showed no production of bacteriocin-like substances nor virulence genes associated with APEC. Most isolates, 59 % (20/34), showed resistance to at least one of the tested antimicrobials, and six multiresistant strains were identified. The results demonstrated that strains of E. coli were commensal of the respiratory microbiota, and that they did not present pathogenicity to the host, since there were no clinical signs of disease, macroscopic lesions in the organs of the evaluated broilers, production of bacteriocin-like substances, nor virulence-associated genes considered as predictors of APEC in bacteria. These strains of E. coli were mostly susceptible to antimicrobials. However, the occurrence of multidrug-resistant strains suggests that these animals can act as reservoirs of resistant to antimicrobials E. coli.

Detection of ESBL/AmpC-Producing and Fosfomycin-Resistant Escherichia coli From Different Sources in Poultry Production in Southern Brazil.

This study discussed the use of antimicrobials in the commercial chicken production system and the possible factors influencing the presence of Extended-spectrum ß-lactamase (ESBL)/AmpC producers strains in the broiler production chain. The aim of this study was to perform longitudinal monitoring of ESBL-producing and fosfomycin-resistant Escherichia coli from poultry farms in southern Brazil (Paraná and Rio Grande do Sul states) and determine the possible critical points that may be reservoirs for these strains. Samples of poultry litter, cloacal swabs, poultry feed, water, and beetles (Alphitobius sp.) were collected during three distinct samplings. Phenotypic and genotypic tests were performed for characterization of antimicrobial resistant strains. A total of 117 strains were isolated and 78 (66%) were positive for ESBL production. The poultry litter presented ESBL positive strains in all three sampled periods, whereas the cloacal swab presented positive strains only from the second period. The poultry litter represents a significant risk factor mainly at the beginning poultry production (odds ratio 6.43, 95% confidence interval 1-41.21, p < 0.05). All beetles presented ESBL positive strains. The predominant gene was bla CTX-M group 2, which occurred in approximately 55% of the ESBL-producing E. coli. The cit gene was found in approximately 13% of the ESBL-producing E. coli as AmpC type determinants. A total of 19 out of 26 fosfomycin-resistant strains showed the fosA3 gene, all of which produced ESBL. The correlation between fosA3 and bla CTX-M group 1 (bla CTX-M55 ) genes was significant among ESBL-producing E. coli isolated from Paraná (OR 3.66, 95% CI 1.9-9.68) and these genetic determinants can be transmitted by conjugation to broiler chicken microbiota strains. Our data revealed that poultry litter and beetles were critical points during poultry production and the presence of fosfomycin-resistant strains indicate the possibility of risks associated with the use of this antimicrobial during production. Furthermore, the genetic determinants encoding CTX-M and fosA3 enzymes can be transferred to E. coli strains from broiler chicken microbiota, thereby creating a risk to public health.

Antimicrobial Resistance Profiles in Enterococcus spp. Isolates From Fecal Samples of Wild and Captive Black Capuchin Monkeys (Sapajus nigritus) in South Brazil.

The environment, human, and animals play an important role in the spread of antibiotic-resistant bacteria. Enterococci are members of the gastrointestinal tracts of humans and animals and represent important reservoirs of antibiotic resistance genes. Until today, few studies have examined antibiotic susceptibility in enterococci isolated from primates. Therefore, the present study investigated species distribution, antibiotic susceptibility, and resistance genes in enterococci isolated from wild and captive black capuchins monkeys (Sapajus nigritus) in Rio Grande do Sul, South Brazil. A total of 24 swabs/fecal samples were collected, including 19 from wild monkeys living in two forest fragments [São Sebastião do Caí (SSC) and Santa Cruz do Sul (SCS)], and five in captive [Parque Zoológico da Fundação Zoobotânica (ZOO)], between August 2016 and November 2017. Fifteen colonies were randomly selected from each sample. Enterococci were identified by MALDI-TOF, tested for susceptibility to 12 antibiotics; and screened for tet(S), tet(M), tet(L), msrC, and erm(B) genes by PCR. Two-hundred ninety-six enterococci were isolated (SSC n = 137; SCS n = 86; ZOO n = 73) and differences in Enterococcus species distribution were detected on three monkey groups, with low abundance in SCS (1 - D = 0.2), followed by ZOO (1 - D = 0.68), and SSC (1 - D = 0.73). The enterococci frequently recovered include the following: Enterococcus faecalis (42.6%), E. hirae (29.1%), and E. faecium (15.9%). Antibiotic-nonsusceptible was observed in 202 (67.9%) strains. The rate of non-susceptibility to rifampicin, tetracycline, erythromycin, nitrofurantoin, chloramphenicol, and ampicillin was 46%, 26%, 22% and 19%, 13%, 0.3%, and 0.3%, respectively. All strains were susceptible to vancomycin, streptomycin, gentamycin, and linezolid. Forty-three (14.52%) isolates were identified as multidrug resistant (MDR), and the highest number of MDR enterococci were E. faecium recovered from wild monkeys living close to a hospital and water treatment plant. Elevated rates of antibiotic resistance genes msrC and tet(L) were isolates from ZOO. In conclusion, differences in the frequency of enterococci species, antibiotic-nonsusceptible and antibiotic resistance genes in all groups of monkeys were identified. These data suggest that anthropogenic activities could have an impact in the resistome of primate gut enterococci communities.

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.

Prevalence of ColV Plasmid-Linked Genes and In Vivo Pathogenicity of Avian Strains of Escherichia coli.

Avian pathogenic Escherichia coli (APEC) causes extraintestinal infections in birds, leading to an increase in the cost of poultry production. The ColV plasmid-linked genes iroN, ompT, hlyF, iss, and iutA have previously been suggested to be predictors of the virulence of APEC. In this research, we analyzed the frequencies of these genes in a Brazilian collection of E. coli isolated from birds with colibacillosis (APEC) and from apparently healthy birds (avian fecal [A(fecal)]), as well as from the litter of poultry houses of apparently healthy flocks (avian litter [A(litter)]). All the isolates that harbored ompT also harbored hlyF, so they were considered as one trait for statistical analysis. The relationship between in vivo virulence in 1-day-old chicks, expressed as a pathogenicity score, and the number of genes in each isolate showed that isolates with less than two of the four genes were rarely pathogenic, while most pathogenic isolates contained two or more genes. Nevertheless, about half of the nonpathogenic isolates also harbored two or more genes, in agreement with previous observations that commensal E. coli isolates from the birds' microbiota can serve as a reservoir of virulence genes. Thus, the pentaplex polymerase chain reaction can be used to indicate that a strain carrying none or only one gene would be nonpathogenic, but it cannot be used to indicate that a strain with two to four genes would be an APEC. Isolates allocated to phylogenetic group B2, which is frequently associated with extraintestinal infections, had the highest pathogenicity scores, while isolates allocated to group B1 had the lowest.

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).