Non-alcoholic fatty liver disease (NAFLD) includes simple hepatic steatosis (SS) and non-alcoholic steatohepatitis (NASH). NAFLD is tightly linked to obesity and is thought to be secondary to various noxious signals, some of which may originate from the intestinal microbiota (IM). Despite a growing body of evidence supporting a link between obesity and altered IM, there are no studies assessing the IM of patients with NAFLD. In this cross-sectional study we aimed at comparing fecal levels of total bacteria, Bacteroidetes, C. coccoides, C. leptum, Bifidobacteria, E. coli, and Archaea between healthy controls (HC) and patients with SS or NASH. We found higher C. coccoides levels in NASH compared to SS and lower percentage Bacteroidetes in NASH compared to SS and HC. Controlling for body mass index and fat intake we found an association between presence of NASH and percentage Bacteroidetes. The latter inversely correlated with insulin resistance.

Orientador: Iveraldo dos Santos Dutra / Resumo: Periodontite e desgaste dentário excessivo são as duas enfermidades orais que mais comumente afetam a dentição de ovinos. De etiologias multifatoriais, são responsáveis por grandes perdas econômicas em diversas regiões do mundo, decorrentes das dificuldades na preensão e mastigação dos alimentos e na ruminação. A periodontite ovina é caracterizada por inflamação gengival e formação de bolsa periodontal, além de perda precoce da unidade dental. O desgaste dentário é a perda gradual e irreversível de estruturas dos dentes, pelo contato físico repetitivo ou por agente químico. O presente estudo teve como objetivos relatar a ocorrência de periodontite e desgaste excessivo da unidade dental, bem como avaliar a presença de biofilme supragengival, em ovelhas em fase reprodutiva. A ocorrência de lesão periodontal, avaliada pela recessão gengival, foi observada em 58% das 129 ovelhas examinadas, das quais 16% apresentaram lesão nos incisivos e 53% nos dentes mastigatórios. Todos os animais avaliados apresentaram algum grau de biofilme aderido à superfície dos dentes, representando um importante fator de risco para o desenvolvimento de periodontite. O desgaste da coroa dental foi observado em 100% dos dentes mastigatórios e 88% dos incisivos, em graus variados. Em estudo complementar, para a caracterização da microbiota envolvida na periodontite ovina, foi realizado o exame da cavidade oral de 63 animais, de diferentes regiões do Brasil, sem distinção de gênero, raça e idade, e nesse... (Resumo completo, clicar acesso eletrônico abaixo) / Doutor

Orientador: Wagner Bettiol / Coorientador: Rodrigo Mendes / Banca: Kátia de Lima Nechet / Banca: Flávia Rodrigues Alves Patrício / Banca: Adriana Zanin Kronka / Banca: Antonio Carlos Maringoni / Resumo: Os microrganismos endofíticos ocorrem naturalmente em todas as partes do cafeeiro e produzem metabólitos secundários que podem influenciar as características da expressão da bebida, bem como a resistência da planta às doenças e pragas. O aumento da concentração atmosférica de CO2 vem sendo observado nas últimas décadas e pode afetar positiva, negativamente ou ser neutro na planta de café, e também influenciar na comunidade de microrganismos endofíticos. A melhor forma de se realizar os estudos com o aumento da concentração de CO2 é por meio de experimento tipo FACE (Free Air Carbon Dioxide Enrichement), o qual possibilita obter uma resposta sem alterar o microclima em áreas extensas de amostragens. Diante disso, o presente trabalho teve por objetivo estudar a comunidade bacteriana endofítica em folhas, frutos e ramos de cafeeiro da variedade Catuaí Vermelho IAC 144, cultivados no experimento ClimapestFACE da Embrapa Meio Ambiente, bem como identificar as bactérias endofíticas presentes em frutos e ramos de café por meio de sequenciamento em larga escala. O experimento FACE é composto por 12 anéis, dos quais seis com a concentração atual de CO2 que é de aproximadamente 400 ppm e os outros seis enriquecidos com CO2 até uma concentração de 550 ppm. Em cada anel, foram coletados folhas, ramos e frutos nas safras de 2013 a 2015. Com os tecidos amostrados foi realizada análises por meio da técnica de T-RFLP (Restriction Fragment Length Polymorphism), onde na amplificação do gene 16... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Endophytic microorganisms occur naturally in coffee plant and produce secondary metabolites that can influence the beverage characteristics, as well as the resistance of plant diseases and pests. The increase in the atmospheric CO2 concentration has been observed in the last decades and can affect positively, negatively or be neutral to coffee plant, and also influence the endophytic microorganisms. The best way to carry out the studies with the increase of CO2 concentration is in FACE experiment (Free Air Carbon Dioxide Enrichement). The objective of this study was to study the endophytic bacterial community in leaves, fruits and branches of coffee grown in the ClimapestFACE experiment at Embrapa Environment, as well as to identify the endophytic bacteria in fruits and branches of through large-scale sequencing. The FACE experiment consists of 12 rings, of which six maintains the ambient CO2 concentration (400 pm) and other six enriched with CO2 at 550 ppm. In each ring were collected leaves, branches and fruit between 2013 and 2015. Samples were analyzed using T-RFLP technique, where specific universal bacteria primers were used in the amplification of 16S rDNA gene. For identification of endophytic community sequencing was performed through Ion Torrent technique. The data of T-RFLP technique were analyzed by ANOSIM test and differences were observed when comparing the groupings between leaves, fruits and branches, however no differences were observed between treatments with or without CO2 enrichment. The Principal Component Analysis confirmed the ANOSIM test and revealed that the structure of endophytic bacterial communities presents greater dissimilarity when comparing leaf, fruit and branch. However, in the comparison between treatments with and without CO2 enrichment there was a similarity and in some cases overlap, which shows that there was no distinction between communities... / Doutor

Inflammatory bowel disease (IBD) is associated with an inappropriate immune response to the gut microbiota and disruption of intestinal homeostasis. IBD patients and experimental animal models have consistently shown alterations in the gut microbiota composition. However, these studies have mainly focused on faecal microbiota samples taken after the onset of inflammation and IBD establishment. The colonic microbiota inhabits both the gut lumen and the mucus layer covering the intestinal epithelium. Thus, information about mucus-resident microbiota is not necessarily conveyed in the routine microbiota analyses of faecal samples. To address potential changes in microbial composition and function before the onset of IBD, we compared both mucus and faecal microbiota in the mdr1a-/- spontaneous model of colitis over times that we histologically defined as before onset of colitis, during and after colitis onset. We showed that alterations in microbiota composition preceded the onset of intestinal inflammation and that these changes were evident in the mucus, but not in faeces. This altered microbiota composition was coupled with a reduced inner mucus layer, indicating a compromised mucus barrier prior to colitis development. Upon emergence of inflammation, compositional differences were found in both mucus and faecal microbial communities. Spatial segregation of microbiota with intestinal mucosa was also disrupted on disease onset which we hypothesise contributes to a more severe intestinal pathology. Therefore, our data indicate that microbial changes start locally in the mucus and then proceed to the faecal matter concomitantly with colitis development. Next, we examined whether microbial gene functional potential and endogenous metabolite profiles followed alterations in gut microbiota taxonomic composition. Our findings showed that the microbial gene content was similar between mdr1a-/- mice and wild-type littermate controls, demonstrating stability of the gut microbiome at the face of ensuing gut inflammation. In further support of these findings, urinary metabolite analysis revealed that metabolite profiles were unaffected by intestinal inflammation. Metabolites previously reported to change in IBD were similar between mdr1a-/- and wild-type mice at stages preceding and during inflammation. We also found that changes in metabolite profiles did not correlate with colitis scores. However, metabolite changes could discriminate mdr1a-/- mice from wild-type controls, suggesting they could have value in predicting risk of IBD with a potential clinical use in at least a subset of individuals with MDR1A polymorphisms. To assess whether changes in antimicrobial proteins (AMPs) accounted for observed differences in mucus microbiota composition, we also investigated the expression of regenerating islet-derived protein 3 γ (Reg3γ), angiogenin 4 (Ang4), β-defensin 1 and resistin-like molecule beta (Relm-β) in the colon. We found similar levels of these AMPs as well as IgA-producing plasma cells between mdr1a-/- and wild-type mice, suggesting that other factors contribute to alterations in microbiota composition. Overall, our data indicate that the mdr1a-/- is a good model of colitis, as it enables us to look at pre-clinical changes in the gut microbiota. This work suggests the importance of mucus sampling for sensitive detection of microbiota changes. Furthermore, metabolite profiling may be a helpful way to discriminate genetic susceptibility to disease.

Objective: To determine the variations in the composition of the intestinal microbiota in asymptomatic children infected with Helicobacter pylori in comparison with children without the infection. Results: Children infected with H. pylori doubled their probability of presenting 3 of 9 genera of bacteria from the gut microbiota, including: Proteobacteria (p = 0.008), Clostridium (p = 0.040), Firmicutes (p = 0.001) and Prevotella (p = 0.006) in comparison to patients without the infection. We performed a nutritional assessment and found that growth stunting was statistically significantly higher in patients infected with H. pylori (p = 0.046). / Revisión por pares / Revisión por pares

Recent advances have increased the understanding that dysbiosis of the gut microbiome may be a significant contributor to the pathophysiology of ulcerative colitis. Because of this, the use of fecal microbiota transplants (FMT) has become more popular as a potential supplemental treatment option for patients suffering from this disease. Research has shown a possible benefit of FMT in conjunction with varying conventional therapies for patients with mild to moderate disease severity. However, there are scarce publications that have investigated the benefit of FMT in conjunction with a single conventional therapy for patients with moderate to severe disease, specifically. The proposed study is a multicenter, double blind, randomized controlled study of FMT, mercaptopurine (6-MP), and prednisone vs 6-MP and prednisone alone in patients with moderate to severe ulcerative colitis. The study subjects will have a baseline evaluation and the treatment trial will last 8 weeks with follow up throughout the study. Investigators will analyze the primary outcome of clinical remission and secondary outcomes of improvement of fecal calprotectin levels, Inflammatory Bowel Disease Questionnaire (IBDQ) score, C-reactive protein (CRP) and erythrocyte sedimentation rate (ESR) in the treatment vs control groups. The data from this study will help to identify if FMT would be an additional safe, efficacious treatment modality to the current medical management of ulcerative colitis.

Clostridium difficile infection (CDI) is a disease associated with the wide-spread use of antibiotics and causes 450,000 infections and almost 30,000 deaths in the United States annually. Recurrence is a major problem, with approximately 1/3rd of patients relapsing after antibiotic treatment for CDI. Fecal Microbiota Transplantation (FMT) has emerged as a novel therapy for recurrent CDI, but the majority of the literature to date is made up of uncontrolled case series, so FMT’s true efficacy compared with standard antibiotic regimens remains unknown. Only a few randomized control trials (RCTs) have been published, and these have studied small numbers of patients and exhibited marked methodological heterogeneity. As such, there is uncertainty about the appropriate indications for FMT with respect to recurrent CDI, as well as the best methodology for the procedure, which has been carried our using various fecal preparations and modes of delivery. In particular, questions remain about if FMT should be recommended for patients with a first CDI recurrence, and if minimally invasive methods of performing FMT such as administration of enteric coated capsules are more efficacious than standard antibiotic treatments. We propose a double blind, placebo controlled, RCT that will be run as two parallel RCTs, where Trial 1 will enroll patients experiencing a first CDI recurrence, and Trial 2 will enroll patients experiencing a second or later CDI recurrence. The treatment arms in each trial will receive FMT in the form of orally administered frozen capsules, while the control arms will receive standard antibiotic treatments based on the number of recurrences they have experienced. If shown to be efficacious in a large RCT, oral capsulized FMT alone as treatment for recurrent CDI has the potential to increase access to FMT, decrease unnecessary antibiotic use, and substantially reduce morbidity and mortality attributable to CDI.

Diabetes is a metabolic disease characterized by the loss of functional pancreatic beta cells. The incidence of diabetes has risen rapidly in recent decades, which has been attributed at least partially to alterations in host-associated microbial communities, or microbiota. It is hypothesized that the loss of important microbial functions from the microbiota of affected host populations plays a role in the mechanism of disease onset. Because the immune system also plays a causative role in diabetes progression, and it is well documented that immune cell development and function are regulated by the microbiota, most diabetes microbiota research has focused on the immune system. However, microbial regulation is also required for the development of many other important tissues, including stimulating differentiation and proliferation. We therefore explored the possibility that the microbiota plays a role in host beta cell development. Using the larval zebrafish as a model, we discovered that sterile or germ free (GF) larvae have a depleted beta cell mass compared to their siblings raised in the presence of bacteria and other microbes. This dissertation describes the discovery and characterization of a rare and novel bacterial gene, whose protein product is sufficient to rescue this beta cell developmental defect in the GF larvae. Importantly, these findings suggest a possible role for the microbiota in preventing or prolonging the eventual onset of diabetes through induction of robust beta cell development. Furthermore, the loss of rare bacterial products such as the one described herein could help to explain why low diversity microbial communities are correlated with diabetes.

Submitted by Jaqueline Jarusevicius (jaquejaru@ibb.unesp.br) on 2018-08-10T12:31:45Z No. of bitstreams: 1 Manuscrito Defesa - Jaqueline Jarusevicius.pdf: 1898282 bytes, checksum: d3ff76c36419bc9f757606d4e8af71c7 (MD5) / Approved for entry into archive by Sulamita Selma C Colnago null (sulamita@btu.unesp.br) on 2018-08-10T16:49:22Z (GMT) No. of bitstreams: 1 jarusevicius_j_dr_bot.pdf: 1898282 bytes, checksum: d3ff76c36419bc9f757606d4e8af71c7 (MD5) / Made available in DSpace on 2018-08-10T16:49:22Z (GMT). No. of bitstreams: 1 jarusevicius_j_dr_bot.pdf: 1898282 bytes, checksum: d3ff76c36419bc9f757606d4e8af71c7 (MD5) Previous issue date: 2018-07-20 / Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) / Aedes aegypti (Ae. aegypti) é o principal vetor de dengue e é também responsável por transmitir outras arboviroses de importância em saúde pública, como as febres zika e chikungunya. Devido a falhas no controle da transmissão destas arboviroses, que tem como base a eliminação do mosquito vetor, o Brasil é um país endêmico para a dengue e a cada ano nos deparamos com epidemias cada vez mais graves. Ao se alimentar de sangue humano infectado o primeiro local de interação do vírus com o organismo do mosquito é o intestino. Além das respostas imunológicas antivirais para conter a infecção, neste ambiente também está presente a microbiota intestinal do mosquito, um importante modulador na infecção de patógenos. Compreender como a microbiota intestinal de mosquitos é definida e se modifica em uma determinada população é de grande interesse uma vez que isso pode elucidar a relação entre mosquitos e seus organismos simbiontes, e consequentemente auxiliar em processos de paratransgênese. A principal forma de aquisição das bactérias intestinais é através do contato com o ambiente, mas outros mecanismos como transmissão transestadial e vertical também devem influenciar no estabelecimento da microbiota intestinal. Neste estudo, nós analisamos a composição da microbiota intestinal de mosquitos Ae. aegypti de uma população de campo, coletados na cidade de Botucatu, SP, através do sequenciamento em larga escala da região hipervariavel V4 do gene 16S rRNA, e acompanhamos como esta composição é alterada durante colonização em insetário. Nossos resultados revelaram uma transição da composição da microbiota intestinal, principalmente pela diminuição progressiva da diversidade de bactérias como efeito da colonização. Os gêneros mais abundantes encontrados, como Pantoea, Burkholderia e Enterobacter, permaneceram em proporções similares ao longo destas gerações. Estes dados sugerem que existe transmissão de bactérias entre as gerações e que o ambiente intestinal possui mecanismos para manter a composição original de sua microbiota. / 2016/16952-9

Submitted by Nathália Faria da Silva (nathaliafsilva.ufv@gmail.com) on 2017-07-28T12:20:01Z No. of bitstreams: 1 texto completo.PDF: 355690 bytes, checksum: 2bbfc33e03e4341d6021cdd4cefb4d35 (MD5) / Made available in DSpace on 2017-07-28T12:20:01Z (GMT). No. of bitstreams: 1 texto completo.PDF: 355690 bytes, checksum: 2bbfc33e03e4341d6021cdd4cefb4d35 (MD5) Previous issue date: 2002-02-25 / Conselho Nacional de Desenvolvimento Científico e Tecnológico / Foi avaliada a microbiota do ar dos ambientes de recepção, embalagem e pasteurização de leite e produção de queijos, iogurte, doce de leite e manteiga no Laticínios/Funarbe-UFV, pelas técnicas da impressão em ágar (IA) e da sedimentação (S). Avaliou-se, ainda, a eficiência da pulverização de soluções sanificantes de digluconato de clorhexidina (DC) a 1.000 e 2.000 mg/L (pH 5,3 e 5,2, respectivamente), de ácido peracético (AP) a 45 e 75 mg/L (pH 4,2 e 3,8, respectivamente) e de quaternário de amônia (QA) a 700 e 1.200 mg/L (pH 9,2 e 9,3, respectivamente), à temperatura ambiente (20 a 25 °C) no controle da microbiota. As contagens de microrganismos mesófilos aeróbios (MA) e de fungos filamentosos e leveduras (FL) pela técnica IA ultrapassaram 90 UFC· m -3 de ar, valor máximo recomendado pela APHA. Pela técnica S, as contagens do -2 ar de quatro ambientes também ultrapassaram -1 30 UFC· cm · semana , conforme recomendação da APHA. Os ambientes diferiram (p<0,05) apenas quanto aos números de Staphylococcus spp. (<1,0 a 4,3 UFC· m -3 ). As contagens microbianas por IA foram de 2 a 10 vezes maiores que as obtidas por S, evidenciando-se a maior capacidade da IA em determinar microrganismos do ar, inclusive patógenos. Quanto à distribuição da microbiota do ar, houve a predominância de FL pela técnica IA e de MA pela técnica S. A elevação da temperatura ambiente, ao contrário do aumento da umidade relativa do ar, não contribuiu para maiores contagens microbianas no ar. Avaliou-se o uso dos sanificantes nos diferentes tempos de análise do ar dos ambientes: T o antes e T 1 ,T 2 e T 3 , respectivamente, 0,5, 12 e 24 horas depois da pulverização. Em função da literatura disponível e dos resultados, considerou- se que houve ação antimicrobiana do sanificante quando a redução das contagens de T 1 foi de, pelo menos, 15%, em duas aplicações do agente químico. Considerou-se, ainda, que o sanificante apresentou efeito residual quando ocorreram reduções de pelo menos 10% nas contagens de T 0 , da primeira para a segunda e da segunda para a terceira aplicação. A ação antimicrobiana contra FL foi observada para DC/2.000 mg/L e para QA/700 mg/L. Contra MA, AP/45 mg/L apresentou ação antimicrobiana. Efeito residual foi constatado contra FL nas soluções de DC/1.000 mg/L e AP/75 mg/L. As contagens elevadas de 12 e 24 horas após as aplicações foram devidas à variabilidade das condições de análise, principalmente nesses tempos, como a atividade de pessoal. A pulverização do ar de ambientes pareceu uma técnica viável no controle da qualidade microbiológica do processamento de produtos lácteos. / In a dairy plant (Funarbe-UFV), the airborne microrganisms were determined by using setting culture plates (SCP) and an Andersen one-stage sieve sampler (ASS), in the following ambients: milk reception, packing and pasteurization; production of cheese, yogurt and “doce de leite” (concentrated condensed milk). Air sanitation, by the pulverization with clorhexidin digluconate/CD (1000 and 2000 mg/L), peracetic acid/PA (45 and 75 mg/L) and amonium quaternary AQ (700 and 1200 mg/L), was evaluated against mesophilic aerobics (MA) and yeasts and molds (YM). The numbers of MA and YM, determined by the ASS, exceeded 90 CFU· m -3 of air, like a APHA’s suggestion. By the SCP, the microrganisms numbers of four ambients also exceeded 30 CFU· cm 2 · week -1 , according APHA recomendation. The ambients differed (p< 0,05) only for Staphylococcus spp. numbers (<1,0 a 4,3 CFU· m -3 ). The ASS microrganisms numbers were about 2 to 10 times larger than SCP numbers, testifing the major hability of ASS on determining airborne microrganisms, including pathogens. There was a predominance at the enviromenthal dairy plant air of YM, by the ASS and of MA, by SCP determinations. In opposition to air moisture increase, the temperature increase didn’t contributed to the increase of airborne microrganisms numbers. An evaluation of the air pulverization was realized at these different air sampling times: T 0 before, and T 1 ,T 2 ,T 3 , respectively 0,5; 12; and 24 hours after the pulverization. Based on results and available literature, it was considered that the sanitizer antimicrobial activity occured when the T 1 reduction was about, at least, 15% at two pulverizations. When the reductions was about 10% at T 0 , between the first and the second pulverization, it was considered a residual effect. The antimicrobial activity against YM was observed for CD/2000 mg/L and to QA/700 mg/L. Against MA, the chemical solution that shows antimicrobian action was PA/45mg/L. A residual effect was observed against YM for CD/1000 mg/L and PA/75 mg/L. The higher numbers of 12 and 24 hours after pulverizations were due to the experimental conditions variability, mainly at these times, like personal activity, for instance. For the microbiologic quality control at the dairy products manufacturing process, the enviromenthal air pulverization shows to be a practicable technic.