scholarly journals Abundant DNase I-Sensitive Bacterial DNA in Healthy Porcine Lungs and Its Implications for the Lung Microbiome

2013 ◽  
Vol 79 (19) ◽  
pp. 5936-5941 ◽  
Author(s):  
Alejandro A. Pezzulo ◽  
Patrick H. Kelly ◽  
Boulos S. Nassar ◽  
Cedric J. Rutland ◽  
Nicholas D. Gansemer ◽  
...  
Keyword(s):  
Bacterial Diversity ◽  
Defense Mechanisms ◽  
16S Rrna Gene Sequencing ◽  
Dnase I ◽  
Rrna Gene ◽  
Bacterial Dna ◽  
Lung Microbiome ◽  
Rrna Gene Sequencing ◽  
Porcine Lungs

ABSTRACTHuman lungs are constantly exposed to bacteria in the environment, yet the prevailing dogma is that healthy lungs are sterile. DNA sequencing-based studies of pulmonary bacterial diversity challenge this notion. However, DNA-based microbial analysis currently fails to distinguish between DNA from live bacteria and that from bacteria that have been killed by lung immune mechanisms, potentially causing overestimation of bacterial abundance and diversity. We investigated whether bacterial DNA recovered from lungs represents live or dead bacteria in bronchoalveolar lavage (BAL) fluid and lung samples in young healthy pigs. Live bacterial DNA was DNase I resistant and became DNase I sensitive upon human antimicrobial-mediated killingin vitro. We determined live and total bacterial DNA loads in porcine BAL fluid and lung tissue by comparing DNase I-treated versus untreated samples. In contrast to the case for BAL fluid, we were unable to culture bacteria from most lung homogenates. Surprisingly, total bacterial DNA was abundant in both BAL fluid and lung homogenates. In BAL fluid, 63% was DNase I sensitive. In 6 out of 11 lung homogenates, all bacterial DNA was DNase I sensitive, suggesting a predominance of dead bacteria; in the remaining homogenates, 94% was DNase I sensitive, and bacterial diversity determined by 16S rRNA gene sequencing was similar in DNase I-treated and untreated samples. Healthy pig lungs are mostly sterile yet contain abundant DNase I-sensitive DNA from inhaled and aspirated bacteria killed by pulmonary host defense mechanisms. This approach and conceptual framework will improve analysis of the lung microbiome in disease.

scholarly journals The growth response of rice (Oryza sativa L. var. FARO 44) in vitro after inoculation with bacterial isolates from a typical ferruginous ultisol

2021 ◽  
Vol 45 (1) ◽  
Author(s):  
Musa Saheed Ibrahim ◽  
Beckley Ikhajiagbe
Keyword(s):  
16S Rrna ◽  
Bacillus Cereus ◽  
Proteus Mirabilis ◽  
Growth Response ◽  
Gene Sequencing ◽  
16S Rrna Gene Sequencing ◽  
Rrna Gene ◽  
Rice Seedlings ◽  
Rrna Gene Sequencing

Abstract Background Rice forms a significant portion of food consumed in most household worldwide. Rice production has been hampered by soil factors such as ferruginousity which has limited phosphorus availability; an important mineral component for the growth and yield of rice. The presence of phosphate-solubilizing bacteria (PSB) in soils has been reported to enhance phosphate availability. In view of this, the present study employed three bacteria species (BCAC2, EMBF2 and BCAF1) that were previously isolated and proved P solubilization capacities as inocula to investigate the growth response of rice germinants in an in vitro setup. The bacteria isolates were first identified using 16S rRNA gene sequencing and then applied as inoculum. The inolula were prepared in three concentrations (10, 7.5 and 5.0 ml) following McFarland standard. Viable rice (var. FARO 44) seeds were sown in petri dishes and then inoculated with the three inocula at the different concentrations. The setup was studied for 28 days. Results 16S rRNA gene sequencing identified the isolates as: isolate BCAC2= Bacillus cereus strain GGBSU-1, isolate BCAF1= Proteus mirabilis strain TL14-1 and isolate EMBF2= Klebsiella variicola strain AUH-KAM-9. Significant improvement in rice germination, morphology, physiology and biomass parameters in the bacteria-inoculated setups was observed compared to the control. Germination percentage after 4 days was 100 % in the inoculated rice germinants compared to 65% in the control (NiS). Similarly, inoculation with the test isolates enhanced water-use efficiency by over 40%. The rice seedlings inoculated with Bacillus cereus strain GGBSU-1 (BiS) showed no signs of chlorosis and necrosis throughout the study period as against those inoculated with Proteus mirabilis strain TL14-1 (PiS) and Klebsiella variicola strain AUH-KAM-9 (KiS). Significant increase in chlorophyll-a, chlorophyll-b and alpha amylase was observed in the rice seedlings inoculated with BiS as against the NiS. Conclusion Inoculating rice seeds with Bacillus cereus strain GGBSU-1, Proteus mirabilis strain TL14-1 and Klebsiella variicola strain AUH-KAM-9 in an in vitro media significantly improved growth parameters of the test plant. Bacillus cereus strain GGBSU-1 showed higher efficiency due to a more improved growth properties observed.

scholarly journals Live Bacillus subtilis natto Promotes Rumen Fermentation by Modulating Rumen Microbiota In Vitro

Animals ◽  
2021 ◽  
Vol 11 (6) ◽  
pp. 1519
Author(s):  
Meinan Chang ◽  
Fengtao Ma ◽  
Jingya Wei ◽  
Junhao Liu ◽  
Xuemei Nan ◽  
...  
Keyword(s):  
Bacillus Subtilis ◽  
16S Rrna ◽  
Rumen Fluid ◽  
Gene Sequencing ◽  
16S Rrna Gene Sequencing ◽  
Rumen Fermentation ◽  
Rrna Gene ◽  
Bacillus Subtilis Natto ◽  
Rrna Gene Sequencing

Previous studies have shown that Bacillus subtilis natto affects rumen fermentation and rumen microbial community structure, which are limited to detect a few microbial abundances using traditional methods. However, the regulation of B. subtilis natto on rumen microorganisms and the mechanisms of microbiota that affect rumen fermentation is still unclear. This study explored the effects of live and autoclaved B. subtilis natto on ruminal microbial composition and diversity in vitro using 16S rRNA gene sequencing and the underlying mechanisms. Rumen fluid was collected, allocated to thirty-six bottles, and divided into three treatments: CTR, blank control group without B. subtilis natto; LBS, CTR with 109 cfu of live B. subtilis natto; and ABS, CTR with 109 cfu of autoclaved B. subtilis natto. The rumen fluid was collected after 0, 6, 12, and 24 h of fermentation, and pH, ammonia nitrogen (NH3-N), microbial protein (MCP), and volatile fatty acids (VFAs) were determined. The diversity and composition of rumen microbiota were assessed by 16S rRNA gene sequencing. The results revealed LBS affected the concentrations of NH3-N, MCP, and VFAs (p < 0.05), especially after 12 h, which might be attributed to changes in 18 genera. Whereas ABS only enhanced pH and NH3-N concentration compared with the CTR group (p < 0.05), which might be associated with changes in six genera. Supplementation with live B. subtilis natto improved ruminal NH3-N and propionate concentrations, indicating that live bacteria were better than autoclaved ones. This study advances our understanding of B. subtilis natto in promoting ruminal fermentation, providing a new perspective for the precise utilization of B. subtilis natto in dairy rations.

scholarly journals Transition of Bacterial Diversity and Composition in Tongue Microbiota during the First Two Years of Life

mSphere ◽  
2019 ◽  
Vol 4 (3) ◽  
Author(s):  
Shinya Kageyama ◽  
Mikari Asakawa ◽  
Toru Takeshita ◽  
Yukari Ihara ◽  
Shunsuke Kanno ◽  
...  
Keyword(s):  
Oral Cavity ◽  
Bacterial Diversity ◽  
Bacterial Species ◽  
16S Rrna Gene Sequencing ◽  
Rrna Gene ◽  
Oral Microbiota ◽  
Bacterial Composition ◽  
Content Type ◽  
Operational Taxonomic Units ◽  
Rrna Gene Sequencing

ABSTRACTNewborns are constantly exposed to various microbes from birth; hence, diverse commensal bacteria colonize the oral cavity. However, how or when these bacteria construct a complex and stable ecosystem remains unclear. This prospective cohort study examined the temporal changes in bacterial diversity and composition in tongue microbiota during infancy. We longitudinally collected a total of 464 tongue swab samples from 8 infants (age of <6 months at baseline) for approximately 2 years. We also collected samples from 32 children (aged 0 to 2 years) and 73 adults (aged 20 to 29 years) cross-sectionally as control groups. Bacterial diversities and compositions were determined by 16S rRNA gene sequencing. The tongue bacterial diversity in infancy, measured as the number of observed operational taxonomic units (OTUs), rapidly increased and nearly reached the same level as that in adults by around 80 weeks. The overall tongue bacterial composition in the transitional phase, 80 to 120 weeks, was more similar to that of adults than to that of the early exponential phase (EEP), 10 to 29 weeks, according to analysis of similarities. Dominant OTUs in the EEP corresponding toStreptococcus perorisandStreptococcus lactariusexponentially decreased immediately after EEP, around 30 to 49 weeks, whereas several OTUs corresponding toGranulicatella adiacens,Actinomyces odontolyticus, andFusobacterium periodonticumreciprocally increased during the same period. These results suggest that a drastic compositional shift of tongue microbiota occurs before the age of 1 year, and then bacterial diversity and overall bacterial composition reach levels comparable to those in adults by the age of 2 years.IMPORTANCEEvaluating the development of oral microbiota during infancy is important for understanding the subsequent colonization of bacterial species and the process of formation of mature microbiota in the oral cavity. We examined tongue microbiota longitudinally collected from 8 infants and found that drastic compositional shifts in tongue microbiota occur before the age of 1 year, and then bacterial diversity and overall bacterial composition reach levels comparable to those in adults by the age of 2 years. These results may be helpful for preventing the development of various diseases associated with oral microbiota throughout life.

scholarly journals Loss of bacterial diversity in the sinuses is associated with lower smell discrimination scores

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Kristi Biswas ◽  
Brett Wagner Mackenzie ◽  
Charlotte Ballauf ◽  
Julia Draf ◽  
Richard G. Douglas ◽  
...  
Keyword(s):  
Bacterial Diversity ◽  
Bacterial Communities ◽  
Gene Sequencing ◽  
16S Rrna Gene Sequencing ◽  
Olfactory Dysfunction ◽  
Rrna Gene ◽  
Olfactory Loss ◽  
Olfactory Impairment ◽  
Rrna Gene Sequencing ◽  
Normal Sense

Abstract Olfactory impairment affects ~ 20% of the population and has been linked to various serious disorders. Microbes in the nasal cavity play a key role in priming the physiology of the olfactory epithelium and maintaining a normal sense of smell by the host. The aim of this study was to explore the link between olfactory dysfunction and nasal bacterial communities. A total of 162 subjects were recruited for this study from a specialized olfactory dysfunction clinic and placed into one of three groups: anosmia, hyposmia or normosmia. Swabs from the nasal middle meatus were collected from each subject then processed for bacterial 16S rRNA gene sequencing. No overall differences in bacterial diversity or composition were observed between the three cohorts in this study. However, the relative abundances of Corynebacterium spp. and Streptococcus spp. were significantly (p < 0.05) different in subjects with olfactory loss. Furthermore, subjects with deficiencies in discriminating between smells (based on discrimination scores) had a lower bacterial diversity (Simpson’s evenness p < 0.05). While these results are preliminary in nature, potential bacterial biomarkers for olfactory loss were identified. These findings need to be further validated and biologically tested in animal models.

scholarly journals Findings from the microbiota of tooth apical periodontitis and the search for pathogen forgranulomatous inflammation

2021 ◽  
Author(s):  
Yuanyuan Wang ◽  
Hao Xu ◽  
Minghui Wei ◽  
Yuhong Wang ◽  
Wenzhe Wang ◽  
...  
Keyword(s):  
Granulomatous Inflammation ◽  
16S Rrna Gene Sequencing ◽  
Apical Periodontitis ◽  
Rrna Gene ◽  
Normal Microbiota ◽  
Rrna Gene Sequencing ◽  
Neisseria Subflava ◽  
Foot Pad

Abstract BackgroundOrofacial granulomatosis (OFG) is a granulomatous inflammation (GI) disease in maxillofacial region, the underlying cause of it remains unknown. Our previous study demonstrated that tooth apical periodontitis (AP) plays a significant role in the pathogenesis of OFG, we aimed here to characterize the AP bacterial signatures of OFG patients, and identify bacteria that may be important pathogens capable of inducing OFG.ResultsThe composition of AP microbiota in OFG cases and common AP controls was compared using 16S rRNA gene sequencing, the results showed a specific AP microbiota signature in OFG patients, characterized by domination of phyla Firmicutes and Proteobacteria , notably members of Streptococcus, Lactobacillus and Neisseria. To assess the pathogenicity of the potential pathogens in OFG, we isolated and successfully in vitro cultured Streptococcus, Lactobacillus casei, Neisseria subflava, Veillonella parvula and Actinomyces from OFG patients, and injected the clinical isolates into mice respectively. Ultimately, foot pad injection with N. subflava elicited granulomatous inflammation, and the virulence of N. subflava was verified based on Koch’s postulates.ConclusionsOur findings confirmed the role of bacteria in OFG, and first suggested that the component of the host normal microbiota, N. subflava is likely a pathogen for GI.

scholarly journals Personalized Bioconversion of Panax Notoginseng Saponins Mediated by Gut Microbiota Between Chinese-diet and Western-diet Healthy Subjects

2021 ◽  
Author(s):  
Li Wang ◽  
Man-Yun Chen ◽  
Li Shao ◽  
Wei Zhang ◽  
Xiang-Ping Li ◽  
...  
Keyword(s):  
Gut Microbiota ◽  
Lactobacillus Rhamnosus ◽  
16S Rrna Gene Sequencing ◽  
Panax Notoginseng ◽  
Rrna Gene ◽  
Bifidobacterium Adolescentis ◽  
Correlative Analysis ◽  
Rrna Gene Sequencing ◽  
The Relationship

Abstract Background: Panax notoginseng saponins (PNS) as the main effective substances from P. notoginseng with low bioavailability could be bio-converted by human gut microbiota. In our previous study, PNS metabolic variations mediated by gut microbiota have been observed between high fat, high protein (HF-HP)-diet and low fat, plant fiber-rich (LF-PF)-diet subjects. In this study, we aimed to correspondingly characterize the relationship between distinct gut microbiota profiles and PNS metabolites. Methods: Gut microbiota were collected from HF-HP and LF-PF healthy adults, respectively and profiled by 16S rRNA gene sequencing. PNS were incubated with gut microbiota in vitro. A LC-MS/MS method was developed to quantify the five main metabolites yields including ginsenoside F1 (GF1), ginsenoside Rh2 (GRh2), ginsenoside compound K (GC-K), protopanaxatriol (PPT) and protopanaxadiol (PPD). The selected microbial species, Bifidobacterium adolescentis and Lactobacillus rhamnosus, were employed to metabolize PNS for the corresponding metabolites.Results: The five main metabolites were significantly different between the two diet groups. Compared with HF-HP group, the microbial genus Blautia, Bifidobacterium, Clostridium, Corynebacterium, Dorea, Enhydrobacter, Lactobacillus, Roseburia, Ruminococcus, SMB53, Streptococcus, Treponema and Weissella were enriched in LF-PF group, while Phascolarctobacterium and Oscillospira were relatively decreased. Furthermore, Spearman’s correlative analysis revealed gut microbiota enriched in LF-PF and HF-HP groups were positively and negatively associated with PNS metabolites yields, respectively. Conclusions: Our data showed gut microbiota diversity led to the personalized bioconversion of PNS.

scholarly journals The Viable Microbiome of Human Milk Differs from the Metataxonomic Profile

Nutrients ◽  
2021 ◽  
Vol 13 (12) ◽  
pp. 4445
Author(s):  
Lisa F. Stinson ◽  
Michelle L. Trevenen ◽  
Donna T. Geddes
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Human Milk ◽  
Dna Content ◽  
Gene Sequencing ◽  
16S Rrna Gene Sequencing ◽  
Rrna Gene ◽  
Bacterial Dna ◽  
Cutibacterium Acnes ◽  
Rrna Gene Sequencing

Bacteria in human milk contribute to the establishment of the infant gut microbiome. As such, numerous studies have characterized the human milk microbiome using DNA sequencing technologies, particularly 16S rRNA gene sequencing. However, such methods are not able to differentiate between DNA from viable and non-viable bacteria. The extent to which bacterial DNA detected in human milk represents living, biologically active cells is therefore unclear. Here, we characterized both the viable bacterial content and the total bacterial DNA content (derived from viable and non-viable cells) of fresh human milk (n = 10). In order to differentiate the living from the dead, a combination of propidium monoazide (PMA) and full-length 16S rRNA gene sequencing was used. Our results demonstrate that the majority of OTUs recovered from fresh human milk samples (67.3%) reflected DNA from non-viable organisms. PMA-treated samples differed significantly in their bacterial composition compared to untreated samples (PERMANOVA p < 0.0001). Additionally, an OTU mapping to Cutibacterium acnes had a significantly higher relative abundance in PMA-treated (viable) samples. These results demonstrate that the total bacterial DNA content of human milk is not representative of the viable human milk microbiome. Our findings raise questions about the validity of conclusions drawn from previous studies in which viability testing was not used, and have broad implications for the design of future work in this field.

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