scholarly journals Optimization of Preservation Methods Allows Deeper Insights into Changes of Raw Milk Microbiota

2020 ◽  
Vol 8 (3) ◽  
pp. 368 ◽  
Author(s):  
Alexandre J. Kennang Ouamba ◽  
Gisèle LaPointe ◽  
Simon Dufour ◽  
Denis Roy
Keyword(s):  
Community Structure ◽  
16S Rrna ◽  
Raw Milk ◽  
Cow Milk ◽  
Rrna Gene ◽  
Significant Shift ◽  
Propidium Monoazide ◽  
Suitable Alternative ◽  
Milk Samples ◽  
The 16S Rrna Gene

The temporal instability of raw milk microbiota drastically affects the reliability of microbiome studies. However, little is known about the microbial integrity in preserved samples. Raw cow milk samples were preserved with azidiol or bronopol and stored at 4 °C, or with dimethyl sulfoxide (DMSO) or a mixture of azidiol and DMSO and stored at −20 °C for up to 30 days. Aliquots of 5-, 10-, and 30-day post-storage were treated with propidium monoazide (PMA), then analyzed by sequencing the 16S rRNA gene V3-V4 and V6-V8 regions. The V6-V8 gave a higher richness and lower diversity than the V3-V4 region. After 5-day storage at 4 °C, the microbiota of unpreserved samples was characterized by a drastic decrease in diversity, and a significant shift in community structure. The treatment with azidiol and DMSO conferred the best community stabilization in preserved raw milk. Interestingly, the azidiol treatment performed as well for up to 10 days, thus appearing as a suitable alternative. However, neither azidiol nor bronopol could minimize fungal proliferation as revealed by PMA-qPCR assays. This study demonstrates the preservative ability of a mixture of azidiol and DMSO and provides deeper insights into the microbial changes occurring during the cold storage of preserved raw milk.

Risk from Staphylococcus aureus in informally marketed raw cow milk

2019 ◽  
Vol 2 (1) ◽  
pp. 74 ◽  
Author(s):  
Desislava Bangieva ◽  
Valentin Rusev
Keyword(s):  
Staphylococcus Aureus ◽  
16S Rrna ◽  
Pcr Amplification ◽  
Raw Milk ◽  
Cow Milk ◽  
Pcr Analysis ◽  
Milk Samples ◽  
Staphylococcus Spp ◽  
And Staphylococcus Aureus

This study aimed to investigate the presence of Staphylococcus spp. and Staphylococcus aureus in raw cow milk samples taken from peddlers of five regions in Bulgaria. The results showed that all 44 samples tested were positive for Staphylococcus spp. All isolates were tested for coagulase production and subjected to PCR analysis. PCR amplification of 16S rRNA and nuc genes found the presence of Staphylococcus aureus in 23 (52.3%) of a total of 44 raw milk samples. The number of Staphylococcus spp. ranged from 3x102 to 1.08x106 cfu/ml, and that of Staphylococcus aureus from 1.5x102 to 3.19x105 cfu/ml. It is concluded that control over the hygiene of handling and processing raw milk is essential for its safety.

scholarly journals Primer Design for an Accurate View of Picocyanobacterial Community Structure by Using High-Throughput Sequencing

2019 ◽  
Vol 85 (7) ◽  
Author(s):  
Paula Huber ◽  
Francisco M. Cornejo-Castillo ◽  
Isabel Ferrera ◽  
Pablo Sánchez ◽  
Ramiro Logares ◽  
...  
Keyword(s):  
Community Structure ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
High Throughput ◽  
High Throughput Sequencing ◽  
Rrna Gene ◽  
Content Type ◽  
Structure And Dynamics ◽  
Hypervariable Regions ◽  
The 16S Rrna Gene

ABSTRACTHigh-throughput sequencing (HTS) of the 16S rRNA gene has been used successfully to describe the structure and dynamics of microbial communities. Picocyanobacteria are important members of bacterioplankton communities, and, so far, they have predominantly been targeted using universal bacterial primers, providing a limited resolution of the picocyanobacterial community structure and dynamics. To increase such resolution, the study of a particular target group is best approached with the use of specific primers. Here, we aimed to design and evaluate specific primers for aquatic picocyanobacterial genera to be used with high-throughput sequencing. Since the various regions of the 16S rRNA gene have different degrees of conservation in different bacterial groups, we therefore first determined which hypervariable region of the 16S rRNA gene provides the highest taxonomic and phylogenetic resolution for the generaSynechococcus,Prochlorococcus, andCyanobium. Anin silicoanalysis showed that the V5, V6, and V7 hypervariable regions appear to be the most informative for this group. We then designed primers flanking these hypervariable regions and tested them in natural marine and freshwater communities. We successfully detected that most (97%) of the obtained reads could be assigned to picocyanobacterial genera. We defined operational taxonomic units as exact sequence variants (zero-radius operational taxonomic units [zOTUs]), which allowed us to detect higher genetic diversity and infer ecologically relevant information about picocyanobacterial community composition and dynamics in different aquatic systems. Our results open the door to future studies investigating picocyanobacterial diversity in aquatic systems.IMPORTANCEThe molecular diversity of the aquatic picocyanobacterial community cannot be accurately described using only the available universal 16S rRNA gene primers that target the whole bacterial and archaeal community. We show that the hypervariable regions V5, V6, and V7 of the 16S rRNA gene are better suited to study the diversity, community structure, and dynamics of picocyanobacterial communities at a fine scale using Illumina MiSeq sequencing. Due to its variability, it allows reconstructing phylogenies featuring topologies comparable to those generated when using the complete 16S rRNA gene sequence. Further, we successfully designed a new set of primers flanking the V5 to V7 region whose specificity for picocyanobacterial genera was testedin silicoand validated in several freshwater and marine aquatic communities. This work represents a step forward for understanding the diversity and ecology of aquatic picocyanobacteria and sets the path for future studies on picocyanobacterial diversity.

scholarly journals Phyllobacterium loti sp. nov. isolated from nodules of Lotus corniculatus

2014 ◽  
Vol 64 (Pt_3) ◽  
pp. 781-786 ◽  
Author(s):  
Maximo Sánchez ◽  
Martha-Helena Ramírez-Bahena ◽  
Alvaro Peix ◽  
María J. Lorite ◽  
Juan Sanjuán ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Sequence Similarity ◽  
Carbon Sources ◽  
Lotus Corniculatus ◽  
Culture Conditions ◽  
Rrna Gene ◽  
Content Type ◽  
Link Type ◽  
The 16S Rrna Gene

Strain S658T was isolated from a Lotus corniculatus nodule in a soil sample obtained in Uruguay. Phylogenetic analysis of the 16S rRNA gene and atpD gene showed that this strain clustered within the genus Phyllobacterium . The closest related species was, in both cases, Phyllobacterium trifolii PETP02T with 99.8 % sequence similarity in the 16S rRNA gene and 96.1 % in the atpD gene. The 16S rRNA gene contains an insert at the beginning of the sequence that has no similarities with other inserts present in the same gene in described rhizobial species. Ubiquinone Q-10 was the only quinone detected. Strain S658T differed from its closest relatives through its growth in diverse culture conditions and in the assimilation of several carbon sources. It was not able to reproduce nodules in Lotus corniculatus. The results of DNA–DNA hybridization, phenotypic tests and fatty acid analyses confirmed that this strain should be classified as a representative of a novel species of the genus Phyllobacterium , for which the name Phyllobacterium loti sp. nov. is proposed. The type strain is S658T( = LMG 27289T = CECT 8230T).

scholarly journals Diagnosis of Fulminant Pneumonia Caused by Legionella pneumophila Serogroup 8 with the Sequence Analysis of the 16S rRNA Gene

2011 ◽  
Vol 225 (1) ◽  
pp. 65-69 ◽  
Author(s):  
Toshinori Kawanami ◽  
Kazuhiro Yatera ◽  
Kazumasa Fukuda ◽  
Kei Yamasaki ◽  
Masamizu Kunimoto ◽  
...  
Keyword(s):  
Sequence Analysis ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
Legionella Pneumophila ◽  
Rrna Gene ◽  
The 16S Rrna Gene

scholarly journals The Western Progression of Lyme Disease: Infectious and NonclonalBorrelia burgdorferi Sensu LatoPopulations in Grand Forks County, North Dakota

2014 ◽  
Vol 81 (1) ◽  
pp. 48-58 ◽  
Author(s):  
Brandee L. Stone ◽  
Nathan M. Russart ◽  
Robert A. Gaultney ◽  
Angela M. Floden ◽  
Jefferson A. Vaughan ◽  
...  
Keyword(s):  
Lyme Disease ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
North Dakota ◽  
Intergenic Spacer ◽  
Rrna Gene ◽  
Content Type ◽  
Intergenic Spacer Region ◽  
Grand Forks ◽  
The 16S Rrna Gene

ABSTRACTScant attention has been paid to Lyme disease,Borrelia burgdorferi,Ixodes scapularis, or reservoirs in eastern North Dakota despite the fact that it borders high-risk counties in Minnesota. Recent reports ofB. burgdorferiandI. scapularisin North Dakota, however, prompted a more detailed examination. Spirochetes cultured from the hearts of five rodents trapped in Grand Forks County, ND, were identified asB. burgdorferi sensu latothrough sequence analyses of the 16S rRNA gene, the 16S rRNA gene-ileTintergenic spacer region,flaB,ospA,ospC, andp66. OspC typing revealed the presence of groups A, B, E, F, L, and I. Two rodents were concurrently carrying multiple OspC types. Multilocus sequence typing suggested the eastern North Dakota strains are most closely related to those found in neighboring regions of the upper Midwest and Canada. BALB/c mice were infected withB. burgdorferiisolate M3 (OspC group B) by needle inoculation or tick bite. Tibiotarsal joints and ear pinnae were culture positive, andB. burgdorferiM3 was detected by quantitative PCR (qPCR) in the tibiotarsal joints, hearts, and ear pinnae of infected mice. Uninfected larvalI. scapularisticks were able to acquireB. burgdorferiM3 from infected mice; M3 was maintained inI. scapularisduring the molt from larva to nymph; and further, M3 was transmitted from infectedI. scapularisnymphs to naive mice, as evidenced by cultures and qPCR analyses. These results demonstrate that isolate M3 is capable of disseminated infection by both artificial and natural routes of infection. This study confirms the presence of unique (nonclonal) and infectiousB. burgdorferipopulations in eastern North Dakota.

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