Diagnostic and field investigations in Mycoplasma hyosynoviae and Mycoplasma hyorhinis

Swine mycoplasmas are acknowledged for their potential to contribute to several swine disease complexes. In addition, mycoplasma-associated arthritis has recently been recognized as a re-emergent issue in the swine industry. Reports from Veterinary Diagnostic Laboratories suggest an increasing incidence of cases occurring in the major regions of swine production in the U.S. Although previous studies reproduced polyarthritis by inoculating pigs with either Mycoplasma hyosynoviae or M. hyorhinis, there is a lack of information regarding the epidemiology of these microorganisms within swine populations. In addition, although new diagnostic tools have become available, information regarding the application and performance of these assays is lacking. Therefore, a major objective of the work described in this thesis was to review and to provide new information regarding the use of diagnostic methods in the detection of M. hyosynoviae and M. hyorhinis. In order to evaluate the performance of a new real-time polymerase chain reaction (RT-PCR), a comparison of various DNA extraction methods using different clinical specimens was conducted in the laboratory. Thereafter, a field study was carried using RT-PCR- and antibody-based assays to monitor the infection dynamics of these microorganisms in a wean-to-finish population

Systems-Based Approach for Optimization of Assembly-Free Bacterial MLST Mapping

Epidemiological surveillance of bacterial pathogens requires real-time data analysis with a fast turnaround, while aiming at generating two main outcomes: (1) species-level identification and (2) variant mapping at different levels of genotypic resolution for population-based tracking and surveillance, in addition to predicting traits such as antimicrobial resistance (AMR). Multilocus sequence typing (MLST) aids this process by identifying sequence types (ST) based on seven ubiquitous genome-scattered loci. In this paper, we selected one assembly-dependent and one assembly-free method for ST mapping and applied them with the default settings and ST schemes they are distributed with, and systematically assessed their accuracy and scalability across a wide array of phylogenetically divergent Public Health-relevant bacterial pathogens with available MLST databases. Our data show that the optimal k-mer length for stringMLST is species-specific and that genome-intrinsic and -extrinsic features can affect the performance and accuracy of the program. Although suitable parameters could be identified for most organisms, there were instances where this program may not be directly deployable in its current format. Next, we integrated stringMLST into our freely available and scalable hierarchical-based population genomics platform, ProkEvo, and further demonstrated how the implementation facilitates automated, reproducible bacterial population analysis

ProkEvo: an automated, reproducible, and scalable framework for high-throughput bacterial population genomics analyses

Whole Genome Sequence (WGS) data from bacterial species is used for a variety of applications ranging from basic microbiological research, diagnostics, and epidemiological surveillance. The availability of WGS data from hundreds of thousands of individual isolates of individual microbial species poses a tremendous opportunity for discovery and hypothesis-generating research into ecology and evolution of these microorganisms. Flexibility, scalability, and user-friendliness of existing pipelines for population-scale inquiry, however, limit applications of systematic, population-scale approaches. Here, we present ProkEvo, an automated, scalable, reproducible, and open-source framework for bacterial population genomics analyses using WGS data. ProkEvo was specifically developed to achieve the following goals: (1) Automation and scaling of complex combinations of computational analyses for many thousands of bacterial genomes from inputs of raw Illumina paired-end sequence reads; (2) Use of workflow management systems (WMS) such as Pegasus WMS to ensure reproducibility, scalability, modularity, fault-tolerance, and robust file management throughout the process; (3) Use of high-performance and high-throughput computational platforms; (4) Generation of hierarchical-based population structure analysis based on combinations of multi-locus and Bayesian statistical approaches for classification for ecological and epidemiological inquiries; (5) Association of antimicrobial resistance (AMR) genes, putative virulence factors, and plasmids from curated databases with the hierarchically-related genotypic classifications; and (6) Production of pan-genome annotations and data compilation that can be utilized for downstream analysis such as identification of population-specific genomic signatures. The scalability of ProkEvo was measured with two datasets comprising significantly different numbers of input genomes (one with ~2,400 genomes, and the second with ~23,000 genomes). Depending on the dataset and the computational platform used, the running time of ProkEvo varied from ~3-26 days. ProkEvo can be used with virtually any bacterial species, and the Pegasus WMS uniquely facilitates addition or removal of programs from the workflow or modification of options within them. To demonstrate versatility of the ProkEvo platform, we performed a hierarchical-based population structure analyses from available genomes of three distinct pathogenic bacterial species as individual case studies. The specific case studies illustrate how hierarchical analyses of population structures, genotype frequencies, and distribution of specific gene functions can be integrated into an analysis. Collectively, our study shows that ProkEvo presents a practical viable option for scalable, automated analyses of bacterial populations with direct applications for basic microbiology research, clinical microbiological diagnostics, and epidemiological surveillance

Quantitative real-time polymerase chain reaction for detecting Mycoplasma hyosynoviae and Mycoplasma hyorhinis in pen-based oral, tonsillar, and nasal fluids

Mycoplasma (M.) hyorhinis and M. hyosynoviae are pathogens known to cause disease in pigs post-weaning. Due to their fastidious nature, there is increased need for culture-independent diagnostic platforms to detect these microorganisms. Therefore, this study was performed to develop and optimize quantitative real-time PCR (qPCR) assays to rapidly detect M. hyorhinis and M. hyosynoviae in pen-based oral fluids as well as nasal and tonsillar fluids as proxies for samples used in swine herd surveillance. Two methods of genomic DNA extraction, automated versus manual, were used to compare diagnostic test performance. A wean-to-finish longitudinal study was also carried out to demonstrate the reproducibility of using pen-based oral fluids. Overall, pen-based oral and tonsillar fluids were more likely to be positive for both types of bacteria whereas only M. hyorhinis was detected in nasal fluids. DNA extraction protocols were shown to significantly influence test result. Although the initial detection time somewhat differed, both organisms were repeatedly detected in the longitudinal study. Overall, this study evaluated two qPCR methods for rapid and specific detection of either mycoplasma. Results from the present investigation can serve as a foundation for future studies to determine the prevalence of the two microorganisms, environmental load, and effectiveness of veterinary interventions for infection control

\u3ci\u3eSalmonella enterica\u3c/i\u3e induces biogeography-specific changes in the gut microbiome of pigs

Swine are a major reservoir of an array of zoonotic Salmonella enterica subsp. enterica lineage I serovars including Derby, Typhimurium, and 4,[5],12:i:- (a.k.a. Monophasic Typhimurium). In this study, we assessed the gastrointestinal (GI) microbiome composition of pigs in different intestinal compartments and the feces following infection with specific zoonotic serovars of S. enterica (S. Derby, S. Monophasic, and S. Typhimurium). 16S rRNA based microbiome analysis was performed to assess for GI microbiome changes in terms of diversity (alpha and beta), community structure and volatility, and specific taxa alterations across GI biogeography (small and large intestine, feces) and days post-infection (DPI) 2, 4, and 28; these results were compared to disease phenotypes measured as histopathological changes. As previously reported, only S. Monophasic and S. Typhimurium induced morphological alterations that marked an inflammatory milieu restricted to the large intestine in this experimental model. S. Typhimurium alone induced significant changes at the alpha- (Simpson’s and Shannon’s indexes) and beta-diversity levels, specifically at the peak of inflammation in the large intestine and feces. Increased community dispersion and volatility in colonic apex and fecal microbiomes were also noted for S. Typhimurium. All three Salmonella serovars altered community structure as measured by co-occurrence networks; this was most prominent at DPI 2 and 4 in colonic apex samples. At the genus taxonomic level, a diverse array of putative short-chain fatty acid (SCFA) producing bacteria were altered and often decreased during the peak of inflammation at DPI 2 and 4 within colonic apex and fecal samples. Among all putative SCFA producing bacteria, Prevotella showed a broad pattern of negative correlation with disease scores at the peak of inflammation. In addition, Prevotella 9 was found to be significantly reduced in all Salmonella infected groups compared to the control at DPI 4 in the colonic apex. In conclusion, this work further elucidates that distinct swine-related zoonotic serovars of S. enterica can induce both shared (high resilience) and unique (altered resistance) alterations in gut microbiome biogeography, which helps inform future investigations of dietary modifications aimed at increasing colonization resistance against Salmonella through GI microbiome alterations

Salmonella enterica induces biogeography-specific changes in the gut microbiome of pigs

Swine are a major reservoir of an array of zoonotic Salmonella enterica subsp. enterica lineage I serovars including Derby, Typhimurium, and 4,[5],12:i:- (a.k.a. Monophasic Typhimurium). In this study, we assessed the gastrointestinal (GI) microbiome composition of pigs in different intestinal compartments and the feces following infection with specific zoonotic serovars of S. enterica (S. Derby, S. Monophasic, and S. Typhimurium). 16S rRNA based microbiome analysis was performed to assess for GI microbiome changes in terms of diversity (alpha and beta), community structure and volatility, and specific taxa alterations across GI biogeography (small and large intestine, feces) and days post-infection (DPI) 2, 4, and 28; these results were compared to disease phenotypes measured as histopathological changes. As previously reported, only S. Monophasic and S. Typhimurium induced morphological alterations that marked an inflammatory milieu restricted to the large intestine in this experimental model. S. Typhimurium alone induced significant changes at the alpha- (Simpson’s and Shannon’s indexes) and beta-diversity levels, specifically at the peak of inflammation in the large intestine and feces. Increased community dispersion and volatility in colonic apex and fecal microbiomes were also noted for S. Typhimurium. All three Salmonella serovars altered community structure as measured by co-occurrence networks; this was most prominent at DPI 2 and 4 in colonic apex samples. At the genus taxonomic level, a diverse array of putative short-chain fatty acid (SCFA) producing bacteria were altered and often decreased during the peak of inflammation at DPI 2 and 4 within colonic apex and fecal samples. Among all putative SCFA producing bacteria, Prevotella showed a broad pattern of negative correlation with disease scores at the peak of inflammation. In addition, Prevotella 9 was found to be significantly reduced in all Salmonella infected groups compared to the control at DPI 4 in the colonic apex. In conclusion, this work further elucidates that distinct swine-related zoonotic serovars of S. enterica can induce both shared (high resilience) and unique (altered resistance) alterations in gut microbiome biogeography, which helps inform future investigations of dietary modifications aimed at increasing colonization resistance against Salmonella through GI microbiome alterations

Transversal study about acute acromioclavicular lesions

The objective of the present transversal study is to check how Brazilian orthopedists treat these injuries, their criteria for choosing the treatment, results and complications. METHODS: During the 6th Brazilian Congress of Shoulder and Elbow Surgery (CBOC) and the 38th Brazilian Congress of Orthopaedics and Traumatology (CBOT) 507 questionnaires were distributed (148 CBOC and 359 CBOT), with 478 being considered for analysis. RESULTS: Regarding type-I and -II ACIs, most of the respondents use traditional non-surgical methods. On the other hand, for type-IV, V and VI injuries, 475 (99.4%) of the respondents indicate surgical methods. Concerning type-III injuries, there is no consensus in the selection between traditional and surgical treatment for 386 (80.7%) respondents, with the most important factor for selecting a given treatment method being the patient's level of sports practice and age. CONCLUSION: There is no consensus regarding type-III ACIs, and the selection of the best treatment method is made according to patient's individual characteristics. However, current literature shows a trend towards non-operative methods.OBJETIVO: O objetivo do presente estudo transversal é verificar como o ortopedista brasileiro interpreta as lesões acromioclaviculares quanto aos critérios para a indicação do tratamento cirúrgico ou não, seus métodos preferidos, as complicações mais frequentes e os resultados obtidos. MÉTODOS: Durante o 6º Congresso Brasileiro de Cirurgia do Ombro e Cotovelo (CBOC) e o 38º Congresso Brasileiro de Ortopedia e Traumatologia (CBOT) foram distribuídos 507 questionários, sendo considerados 478 para análise. RESULTADOS: Em relação ao tratamento das LAC tipo I e II, a maioria dos entrevistados utilizam métodos não cirúrgicos. Em contraposição nas LAC IV, V e VI 475 (99,4%) dos entrevistados tratam essas lesões cirurgicamente. Nas LAC tipo III não existe uma definição na escolha do tratamento cirúrgico ou não cirúrgico para 386 (80,7%) entrevistados, sendo que o fator mais importante para tomada de decisão para a maioria dos entrevistados é a atividade esportiva do paciente e a idade. CONCLUSÃO: Nas LAC tipo III não há consenso, sendo o tratamento determinado conforme as características do paciente, contudo na literatura atual há uma tendência para o tratamento não cirúrgico destas lesões.Universidade Federal de São Paulo (UNIFESP) Departamento de Ortopedia e TraumatologiaUNIFESP, Depto. de Ortopedia e TraumatologiaSciEL

Heuristic and Hierarchical-Based Population Mining of Salmonella enterica Lineage I Pan-Genomes as a Platform to Enhance Food Safety

The recent incorporation of bacterial whole-genome sequencing (WGS) into Public Health laboratories has enhanced foodborne outbreak detection and source attribution. As a result, large volumes of publicly available datasets can be used to study the biology of foodborne pathogen populations at an unprecedented scale. To demonstrate the application of a heuristic and agnostic hierarchical population structure guided pan-genome enrichment analysis (PANGEA), we used populations of S. enterica lineage I to achieve two main objectives: (i) show how hierarchical population inquiry at different scales of resolution can enhance ecological and epidemiological inquiries; and (ii) identify population-specific inferable traits that could provide selective advantages in food production environments. Publicly available WGS data were obtained from NCBI database for three serovars of Salmonella enterica subsp. enterica lineage I (S. Typhimurium, S. Newport, and S. Infantis). Using the hierarchical genotypic classifications (Serovar, BAPS1, ST, cgMLST), datasets from each of the three serovars showed varying degrees of clonal structuring. When the accessory genome (PANGEA) was mapped onto these hierarchical structures, accessory loci could be linked with specific genotypes. A large heavy-metal resistance mobile element was found in the Monophasic ST34 lineage of S. Typhimurium, and laboratory testing showed that Monophasic isolates have on average a higher degree of copper resistance than the Biphasic ones. In S. Newport, an extra sugEgene copy was found among most isolates of the ST45 lineage, and laboratory testing of multiple isolates confirmed that isolates of S. Newport ST45 were on average less sensitive to the disinfectant cetylpyridimium chloride than non-ST45 isolates. Lastly, data-mining of the accessory genomic content of S. Infantis revealed two cryptic Ecotypes with distinct accessory genomic content and distinct ecological patterns. Poultry appears to be themajor reservoir for Ecotype 1, and temporal analysis further suggested a recent ecological succession, with Ecotype 2 apparently being displaced by Ecotype 1. Altogether, the use of a heuristic hierarchical-based population structure analysis that includes bacterial pan-genomes (core and accessory genomes) can (1) improve genomic resolution for mapping populations and accessing epidemiological patterns; and (2) define lineage-specific informative loci that may be associated with survival in the food chain

The tropical forest and fire emissions experiment: Trace gases emitted by smoldering logs and dung from deforestation and pasture fires in Brazil

Earlier work showed that Amazonian biomass burning produces both lofted and initially unlofted emissions in large amounts. A mobile, Fourier transform infrared spectrometer (FTIR) measured the unlofted emissions of 17 trace gases from residual smoldering combustion (RSC) of logs as part of the Tropical Forest and Fire Emissions Experiment (TROFFEE) during the 2004 Amazonian dry season. The RSC emissions were highly variable and the few earlier RSC measurements lay near the high end of combustion efficiency observed in this study. Fuel consumption by RSC was ∼5% of total for a planned deforestation fire. Much regional RSC probably occurs in the residual woody debris burned during pasture maintenance fires. RSC could increase estimated total fire emissions for the Amazon region by 20–50% for several important VOC. FTIR emissions measurements of burning dung (in a pasture) showed high emission ratios for acetic acid and ammonia to CO (6.6 ± 3.4% and 8.9 ± 2.1%). Large emissions of nitrogen containing trace gases from burning dung and crop waste could mean that biomass burning in India produces more particle mass than previously assumed. Measurements of late-stage kiln emissions suggested that VOC/CO may increase as carbonization is extended. A cook stove emitted many VOC and NH3 far outside the range observed for open wood cooking fires. Enclosed/vented cooking stoves may change the chemistry of the smoke that is emitted

Estimating cardiovascular risk in patients with type 2 diabetes: a national multicenter study in Brazil

According to Brazilian National Data Survey diabetes is the fifth cause for hospitalization and is one of the ten major causes of mortality in this country.Aimsto stratify the estimated cardiovascular risk (eCVR) in a population of type 2 diabetics (T2DM) according to the Framingham prediction equations as well as to determine the association between eCVR with metabolic and clinical control of the disease.MethodsFrom 2000 to 2001 a cross-sectional multicenter study was conducted in 13 public out-patients diabetes/endocrinology clinics from 8 Brazilian cities. the 10-year risk of developing coronary heart disease (CHD) was estimated by the prediction equations described by Wilson et al (Circulation 1998). LDL equations were preferably used; when patients missed LDL data we used total cholesterol equations instead.ResultsData from 1382 patients (59.0% female) were analyzed. Median and inter-quartile range (IQ) of age and duration of diabetes were 57.4 (51-65) and 8.8 (3-13) years, respectively without differences according to the gender. Forty-two percent of these patients were overweight and 35.4% were obese (the prevalence of higher BMI and obesity in this T2DM group was significantly higher in women than in men; p 20%) in 738 (53.4%), intermediate in 202 (14.6%) and low in 442 (32%) patients. Men [25.1(15.4-37.3)] showed a higher eCVR than women [18.8 (12.4-27.9) p < 0.001]. the most common risk factor was high LDL-cholesterol (80.8%), most frequently found in women than in men (p = 0.01). the median of risk factors present was three (2-4) without gender differences. Overall we observed that 60 (4.3%) of our patients had none, 154(11.1%) one, 310 (22.4%) two, 385 (27.9%) three, 300 (21.7%) four, 149 (10.5%) five and six, (2%) six risk factors. A higher eCVR was noted in overweight or obese patients (p = 0.01 for both groups). No association was found between eCVR with age or a specific type of diabetes treatment. A correlation was found between eCVR and duration of diabetes (p < 0.001), BMI (p < 0.001), creatinine (p < 0.001) and triglycerides levels (p < 0.001) but it was not found with HbA1c, fasting blood glucose and postprandial glucose. A higher eCVR was observed in patients with retinopathy (p < 0.001) and a tendency in patients with microalbuminuria (p = 0.06). Conclusion: our study showed that in this group of Brazilian T2DM the eCVR was correlated with the lipid profile and it was higher in patients with microvascular chronic complications. No correlation was found with glycemic control parameters. These data could explain the failure of intensive glycemic control programs aiming to reduce cardiovascular events observed in some studies.Univ Estado Rio de Janeiro, Diabet Unit, Rio de Janeiro, BrazilUniv São Paulo Scholl Med, Hosp Clin, Lab Clin & Expt Gatroenterol LIM07, São Paulo, BrazilUniv Fed Maranhao, Div Endocrinol, Sao Luis, BrazilUniv Estadual Campinas, Div Endocrinol, Campinas, BrazilCtr Estudos Diabet & Endocrinol Estado Bahia CEDE, Salvador, BA, BrazilUniv Fed Parana, Div Endocrinol, BR-80060000 Curitiba, Parana, BrazilHosp Agamenon Magalhaes, Div Endocrinol, Recife, PE, BrazilPosto Assistencia Med Jaguribe, Joao Pessoa, Paraiba, BrazilSanta Casa Porto Alegre, Div Endocrinol, Porto Alegre, RS, BrazilInst Assistencia & Previdencia Servidor Estado Ba, Div Endocrinol, Salvador, BA, BrazilSanta Casa Belo Horizonte, Diabet Unit, Belo Horizonte, MG, BrazilHosp Geral Goiania, Div Endocrinol, Goiania, Go, BrazilSecretaria Municipal Saude Brasilia, Brasilia, DF, BrazilUniversidade Federal de São Paulo, Div Endocrinol, São Paulo, BrazilUniversidade Federal de São Paulo, Div Endocrinol, São Paulo, BrazilWeb of Scienc