scholarly journals Identification and Differentiation of Pseudomonas Species in Field Samples Using an rpoD Amplicon Sequencing Methodology

mSystems ◽  
2021 ◽  
Author(s):  
Jonas Greve Lauritsen ◽  
Morten Lindqvist Hansen ◽  
Pernille Kjersgaard Bech ◽  
Lars Jelsbak ◽  
Lone Gram ◽  
...  
Keyword(s):  
High Throughput Sequencing ◽  
16S Rrna Gene Sequencing ◽  
Amplicon Sequencing ◽  
Rrna Gene ◽  
Content Type ◽  
Field Samples ◽  
Plant Growth Promoting ◽  
Growth Promoting ◽  
Rrna Gene Sequencing ◽  
Specific Species

A high-throughput sequencing-based method for profiling of Pseudomonas species in soil microbiomes was developed and identified more species than 16S rRNA gene sequencing or cultivation. Pseudomonas species are used as biocontrol organisms and plant growth-promoting agents, and the method will allow tracing of specific species of Pseudomonas as well as enable screening of environmental samples for further isolation and exploitation.

Assessing the Plant Growth-promoting Traits and Host Specificity of Endophytic Bacteria of Pulse Crops

2021 ◽  
Author(s):  
R. Thamizh Vendan ◽  
D. Balachandar
Keyword(s):  
Plant Growth ◽  
Endophytic Bacteria ◽  
Gene Sequencing ◽  
16S Rrna Gene Sequencing ◽  
Growth And Yield ◽  
Rrna Gene ◽  
Plant Growth Promoting ◽  
Growth Promoting ◽  
Rrna Gene Sequencing ◽  
Plant Growth Promoting Traits

Background: Symbiotic associations between legumes and Rhizobia are ancient and fundamental. However, the plant growth-promoting endophytes other than Rhizobia are not yet fully explored for pulses productivity. The present study was aimed to isolate efficient endophytic bacteria from pulses, assess their diversity, screen their plant growth-promoting activities and to test their potential as bio inoculants for pulses.Methods: We have isolated several endophytic bacteria from pulse crops more specifically from blackgram (Vigna mungo) and greengram (Vigna radiata). After careful screening, 15 promising endophytic isolates were selected for this study. The identification of endophytic bacterial isolates was performed by 16S rRNA gene sequencing. The isolates were tested for their potential for the plant growth-promoting traits such as nitrogen fixation, phosphate solubilization, indole-3-acetic acid production, siderophore secretion and antifungal activity. Pot culture experiments were conducted with the screened potential endophytic cultures.Result: The 16S rRNA gene sequencing revealed that species of Enterobacter, Bacillus, Pantoea, Pseudomonas, Acromobacter, Ocrobacterium were found as endophytes in blackgram and greengram. The in vitro screening identified Bacillus pumilus (BG-E6), Pseudomonas fluorescens (BG-E5) and Bacillus licheniformis (BG-E3) from blackgram and Pseudomonas chlororaphis (GG-E2) and Bacillus thuringiensis (GG-E7) from greengram as potential plant growth-promoting endophytes. These strains showed antagonism against plant pathogenic fungi. Upon inoculation of these endophytic PGPR strains, the blackgram and greengram growth and yield got increased. Among the strains, BG-E6 recorded 14.7% increased yield in blackgram and GG-E2 accounted for a 19.5% yield increase in greengram compared to respective uninoculated control. The experimental results showed that there was a host specificity found among the endophytic bacterial cultures with pulses. The cross inoculation of endophytic strains did not perform well to enhance the growth and yield of their alternate hosts. 

scholarly journals Screening of Potent Arsenic Resistant and Plant Growth Promoting Bacillus species from the Soil of Terai Region of Nepal

2019 ◽  
Vol 6 ◽  
pp. 1-9
Author(s):  
Pramod Poudel ◽  
Ashish Nepal ◽  
Rashmi Roka Magar ◽  
Pratibha Rauniyar ◽  
Lil Buda Magar
Keyword(s):  
Plant Growth ◽  
Sodium Arsenite ◽  
16S Rrna Gene Sequencing ◽  
Bacillus Species ◽  
Rrna Gene ◽  
Sodium Arsenate ◽  
Bacillus Spp ◽  
Plant Growth Promoting ◽  
Growth Promoting ◽  
Rrna Gene Sequencing

Objectives: To isolate arsenic resistant Bacillus spp. and to determine plant growth promoting activities.  Methods: Eighteen soil samples were collected from the agricultural soil of Terai region of Nepal. Selective isolation of Bacillus species was done by heating the soil at 80 ºC for 15 minutes before the isolation. Nutrient agar was used as an isolation medium. Screening of arsenic resistant Bacillus species was done using nutrient agar supplemented with 100 ppm sodium arsenate and sodium arsenite. For plant growth promoting activity; IAA production was detected taking 0.1% tryptophane and measuring absorbance at 540 nm, NH3 production was tested by Nessler’s reagent and phosphate solubilization activity was detected by growing colonies on Pikovskaya’s agar. Sugar assimilation test was performed to identify the isolates. Most potent arsenic resistant isolate was identified by 16S rRNA gene sequencing. Results: Among 54 randomly selected isolates, 42 were found to be Gram-positive rod-shaped, spore-forming while 12 isolates were Gram-negative bacteria. The isolates IN12a, M12a and BG34a showed growth on 100 ppm sodium arsenite containing NA. Only isolate M12a tolerated up to 1000 ppm and 15000 ppm of sodium arsenite and sodium arsenate respectively, while other isolates could not grow above 400 ppm sodium arsenite. The isolates IN12a and M12a were able to produce IAA and solubilize phosphate while BG34a could not. Both the isolates IN12a and M12a were able to utilize the sugars glucose, fructose, lactose, sucrose, galactose, mannose, mannitol, maltose and xylose.  Based on the 16S rRNA gene sequencing, isolate M12a was identified to be Bacillus flexus with highest similarity of 99.2%. Conclusion: Arsenic resistant and plant growth promoting Bacillus spp. was isolated from the agricultural soil of Terai region of Nepal

scholarly journals Diversity of Culturable Endophytic bacteria from Wild and Cultivated Rice showed potential Plant Growth Promoting activities

2018 ◽  
Author(s):  
Madhusmita Borah ◽  
Saurav Das ◽  
Himangshu Baruah ◽  
Robin C. Boro ◽  
Madhumita Barooah
Keyword(s):  
Plant Growth ◽  
Gene Sequencing ◽  
16S Rrna Gene Sequencing ◽  
Rrna Gene ◽  
Plant Growth Promoting Activities ◽  
Plant Growth Promoting ◽  
Growth Promoting ◽  
Rrna Gene Sequencing ◽  
Solubilizing Activity ◽  
Pot Culture

AbstractIn this paper, we report the endophytic microbial diversity of cultivated and wild Oryza sativa plants including their functional traits related to multiple traits that promote plant growth and development. Around 255 bacteria were isolated out of which 70 isolates were selected for further studies based on their morphological differences. The isolates were characterized both at biochemical and at the molecular level by 16s rRNA gene sequencing. Based on 16S rRNA gene sequencing the isolates were categorized into three major phyla, viz, Firmicutes (57.1 %), Actinobacteria (20.0 %) and Proteobacteria (22.8 %). Firmicutes was the dominant group of bacteria of which the most abundant genus was Bacillus. The isolates were further screened in vitro for plant growth promoting activities which revealed a hitherto unreported endophytic bacterial isolate, Microbacteriaceae bacterium RS01 11 as the highest secretor of a phytohormone, IAA (28.39 ± 1.39 μg/ml) and GA (67.23 ± 1.83 μg/ml). Bacillus subtilis RHS 01 displayed highest phosphate solubilizing activity (81.70 ± 1.98 μg/ml) while, Microbacterium testaceum MK LS01, and Microbacterium trichothecenolyticum MI03 L05 exhibited highest potassium solubilizing activity (53.42±0.75μg/ml) and zinc solubilizing efficiency (157.50%) respectively. Bacillus barbaricus LP20 05 produced highest siderophore units (64.8 %). Potential plant growth promoting isolated were tested in vivo in pot culture under greenhouse conditions. A consortium consisting of Microbacteriaceae bacterium RS01 11, Bacillus testaceum MK LS01 and Bacillus subtilis RHS promoted plant growth and increased the yield 3.4 fold in rice when compared to control T0 when tested in pot culture and reduce application rates of chemical fertilizer to half the recommended dose. Our study confirms the potentiality of the rice endophytes isolated as good plant growth promoter and effective biofertilizer.

scholarly journals Much ado about nothing? Off-target amplification can lead to false-positive bacterial brain microbiome detection in healthy and Parkinson’s disease individuals

Microbiome ◽  
2021 ◽  
Vol 9 (1) ◽  
Author(s):  
Janis R. Bedarf ◽  
Naiara Beraza ◽  
Hassan Khazneh ◽  
Ezgi Özkurt ◽  
David Baker ◽  
...  
Keyword(s):  
Parkinson’S Disease ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
False Positive ◽  
Gene Sequencing ◽  
Bacterial Biomass ◽  
16S Rrna Gene Sequencing ◽  
Amplicon Sequencing ◽  
Rrna Gene ◽  
Rrna Gene Sequencing

Abstract Background Recent studies suggested the existence of (poly-)microbial infections in human brains. These have been described either as putative pathogens linked to the neuro-inflammatory changes seen in Parkinson’s disease (PD) and Alzheimer’s disease (AD) or as a “brain microbiome” in the context of healthy patients’ brain samples. Methods Using 16S rRNA gene sequencing, we tested the hypothesis that there is a bacterial brain microbiome. We evaluated brain samples from healthy human subjects and individuals suffering from PD (olfactory bulb and pre-frontal cortex), as well as murine brains. In line with state-of-the-art recommendations, we included several negative and positive controls in our analysis and estimated total bacterial biomass by 16S rRNA gene qPCR. Results Amplicon sequencing did detect bacterial signals in both human and murine samples, but estimated bacterial biomass was extremely low in all samples. Stringent reanalyses implied bacterial signals being explained by a combination of exogenous DNA contamination (54.8%) and false positive amplification of host DNA (34.2%, off-target amplicons). Several seemingly brain-enriched microbes in our dataset turned out to be false-positive signals upon closer examination. We identified off-target amplification as a major confounding factor in low-bacterial/high-host-DNA scenarios. These amplified human or mouse DNA sequences were clustered and falsely assigned to bacterial taxa in the majority of tested amplicon sequencing pipelines. Off-target amplicons seemed to be related to the tissue’s sterility and could also be found in independent brain 16S rRNA gene sequences. Conclusions Taxonomic signals obtained from (extremely) low biomass samples by 16S rRNA gene sequencing must be scrutinized closely to exclude the possibility of off-target amplifications, amplicons that can only appear enriched in biological samples, but are sometimes assigned to bacterial taxa. Sequences must be explicitly matched against any possible background genomes present in large quantities (i.e., the host genome). Using close scrutiny in our approach, we find no evidence supporting the hypothetical presence of either a brain microbiome or a bacterial infection in PD brains.

scholarly journals Citroniella saccharovorans gen. nov. sp. nov., a member of the family Peptoniphilaceae isolated from a human fecal sample from a coastal traditional community member

2019 ◽  
Vol 69 (4) ◽  
pp. 1142-1148 ◽  
Author(s):  
Nisha B. Patel ◽  
Alexandra J. Obregón-Tito ◽  
Raul Y. Tito ◽  
Omar Trujillo-Villaroel ◽  
Luis Marin-Reyes ◽  
...  
Keyword(s):  
Fecal Sample ◽  
16S Rrna Gene Sequencing ◽  
Rrna Gene ◽  
The Novel ◽  
Content Type ◽  
Link Type ◽  
Novel Bacterium ◽  
The Family ◽  
Diamino Acid ◽  
Rrna Gene Sequencing

A novel Gram-stain-positive, non-motile, non-spore-forming coccus-shaped obligately anaerobic bacterium was recovered from a fecal sample obtained from an individual from a traditional community located on the southern coast of Peru. The results of analysis based on 16S rRNA gene sequencing indicated the novel bacterium to be phylogenetically distinct from other genera of members of the Peptoniphilaceae family, sharing a loose affinity with the genera Ezakiella , Finegoldia , Gallicola and Parvimonas . The major cellular fatty acids of the novel isolate were determined to be C16:0, C17:1ω8c, and C18:1ω9c. The DNA G+C content was 29.9 mol%. End products of metabolism from peptone yeast glucose broth (PYG) were determined to be acetate and methyl succinate. The diagnostic diamino acid present in the cell wall was lysine. On the basis of the phenotypic, chemotaxonomic and phylogenetic results the organism is a member of a novel genus belonging to the family Peptoniphilaceae for which the name Citroniella saccharovorans gen nov. sp. nov., is proposed. The type strain is M6.X9T (DSM 29873T=CCUG 66799T).

scholarly journals Impact of Oral Fidaxomicin Administration on the Intestinal Microbiota and Susceptibility to Clostridium difficile Colonization in Mice

2018 ◽  
Vol 62 (5) ◽  
Author(s):  
N. J. Ajami ◽  
J. L. Cope ◽  
M. C. Wong ◽  
J. F. Petrosino ◽  
L. Chesnel
Keyword(s):  
Clostridium Difficile ◽  
Gut Microbiota ◽  
16S Rrna Gene Sequencing ◽  
Taxonomic Structure ◽  
Rrna Gene ◽  
Colonization Resistance ◽  
Content Type ◽  
Rrna Gene Sequencing ◽  
Hospital Acquired ◽  
Predetermined Time

ABSTRACT Clostridium difficile infection (CDI), a common cause of hospital-acquired infections, typically occurs after disruption of the normal gut microbiome by broad-spectrum antibiotics. Fidaxomicin is a narrow-spectrum antibiotic that demonstrates a reduced impact on the normal gut microbiota and is approved for the treatment of CDI. To further explore the benefits of this property, we used a murine model to examine the effects of fidaxomicin versus vancomycin on gut microbiota and susceptibility to C. difficile colonization while tracking microbiota recovery over time. Mice were exposed to fidaxomicin or vancomycin by oral gavage for 3 days and subsequently challenged with C. difficile spores at predetermined time points up to 21 days postexposure to antibiotics. Fecal samples were subsequently collected for analysis. Twenty-four hours postchallenge, mice were euthanized and the colon contents harvested. The microbiota was characterized using 16S rRNA gene sequencing. All fidaxomicin-exposed mice (except for one at day 8) were resistant to C. difficile colonization. However, 9 of 15 vancomycin-exposed mice were susceptible to C. difficile colonization until day 12. All vancomycin-exposed mice recovered colonization resistance by day 16. Bacterial diversity was similar prior to antibiotic exposure in both arms and decreased substantially after exposure. A shift in taxonomic structure and composition occurred after both exposures; however, the shift was greater in vancomycin-exposed than in fidaxomicin-exposed mice. In summary, compared with vancomycin, fidaxomicin exposure had less impact on microbiota composition, promoted faster microbial recovery, and had less impact on the loss of C. difficile colonization resistance.

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 Whole-Genome Sequencing ofAggregatibacterSpecies Isolated from Human Clinical Specimens and Description ofAggregatibacter kilianiisp. nov.

2018 ◽  
Vol 56 (7) ◽  
Author(s):  
May Murra ◽  
Lisbeth Lützen ◽  
Aynur Barut ◽  
Reinhard Zbinden ◽  
Marianne Lund ◽  
...  
Keyword(s):  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
16S Rrna Gene Sequencing ◽  
Rrna Gene ◽  
Whole Genome ◽  
Content Type ◽  
Clinical Specimens ◽  
Phylogenetic Cluster ◽  
Invasive Infections ◽  
Rrna Gene Sequencing

ABSTRACTAggregatibacterspecies are commensal bacteria of human mucosal surfaces that are sometimes involved in serious invasive infections. During the investigation of strains cultured from various clinical specimens, we encountered a coherent group of 10 isolates that could not be allocated to any validly named species by phenotype, mass spectrometry, or partial 16S rRNA gene sequencing. Whole-genome sequencing revealed a phylogenetic cluster related to but separate fromAggregatibacter aphrophilus. The meanin silicoDNA hybridization value for strains of the new cluster versusA. aphrophiluswas 56% (range, 53.7 to 58.0%), whereas the average nucleotide identity was 94.4% (range, 93.9 to 94.8%). The new cluster exhibited aggregative properties typical of the genusAggregatibacter. Key phenotypic tests for discrimination of the new cluster from validly namedAggregatibacterspecies are alanine-phenylalanine-proline arylamidase,N-acetylglucosamine, and β-galactosidase. The nameAggregatibacter kilianiiis proposed, with PN_528 (CCUG 70536Tor DSM 105094T) as the type strain.

scholarly journals Cloacibacillus sp., a Potential Human Pathogen Associated with Bacteremia in Quebec and New Brunswick

2015 ◽  
Vol 53 (10) ◽  
pp. 3380-3383 ◽  
Author(s):  
M.-C. Domingo ◽  
C. Yansouni ◽  
C. Gaudreau ◽  
F. Lamothe ◽  
S. Lévesque ◽  
...  
Keyword(s):  
16S Rrna ◽  
New Brunswick ◽  
Gene Sequencing ◽  
16S Rrna Gene Sequencing ◽  
Rrna Gene ◽  
Human Pathogen ◽  
Emerging Pathogens ◽  
Content Type ◽  
Rrna Gene Sequencing ◽  
Anaerobic Infections

Bacteremia due toCloacibacillusspecies is poorly described. We present three cases involving eitherCloacibacillusevryensisorCloacibacillusporcorum. The isolates were identified by 16S rRNA gene sequencing and were susceptible to antibiotics commonly used for anaerobic infections. The clinical significance of these organisms as potential emerging pathogens is discussed.

Emendation of the description of the species Corynebacterium propinquum to include strains which produce urease

2013 ◽  
Vol 63 (Pt_6) ◽  
pp. 2146-2154 ◽  
Author(s):  
Kathryn Bernard ◽  
Ana Luisa Pacheco ◽  
Ian Cunningham ◽  
Navdeep Gill ◽  
Tamara Burdz ◽  
...  
Keyword(s):  
Type Species ◽  
Sequence Similarity ◽  
Gene Sequencing ◽  
16S Rrna Gene Sequencing ◽  
Rrna Gene ◽  
Single Test ◽  
Content Type ◽  
Link Type ◽  
Corynebacterium Pseudodiphtheriticum ◽  
Rrna Gene Sequencing

Corynebacterium propinquum is a Gram-positive rod occasionally recovered from clinical infections which, according to 16S rRNA gene sequencing, is most closely related (>99 % sequence similarity) to Corynebacterium pseudodiphtheriticum . The two species are very similar biochemically, commonly differentiated by a single test, the detection of urease, where strains of C. propinquum are described as being urease-non-producing and strains of C. pseudodiphtheriticum are described as urease-producing. In this study, historical and contemporary strains of C. propinquum and C. pseudodiphtheriticum from this laboratory were definitively characterized, which included use of rpoB sequencing. Urease-producing strains of C. propinquum as well as typical urease-non-producing isolates were identified after rpoB sequencing, with six of these being originally identified as C. pseudodiphtheriticum . Based on these observations, we propose emendation of the description of C. propinquum to include strains which produce urease. MALDI-TOF analysis may be a useful tool to differentiate these taxa. Existing commercial databases should be updated to include urease-positive strains of C. propinquum .

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