Studies on bacterial community composition are affected by the time and storage method of the rumen content
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Fecha
2017-04-28
Autores
Granja-Salcedo, Yury Tatiana
Ramirez-Uscategui, Ricardo Andrés
Machado, Elwi Guillermo
Duarte Messana, Juliana
Takeshi Kishi, Luciano
Lino Dias, Ana Veronica
Berchielli, Telma Teresinha
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Public Library of Science
Resumen
The objective of this study was to investigate three storage methods and four storage times
for rumen sampling in terms of quality and yield of extracted metagenomic DNA as well as
the composition of the rumen bacterial community. One Nellore steer fitted with a ruminal silicone-
type cannula was used as a donor of ruminal contents. The experiment comprised 11
experimental groups: pellet control (PC), lyophilized control (LC), P-20: pellet stored frozen
at -20ÊC for a period of 3, 6, and 12 months, P-80: pellet stored frozen at -80ÊC for a period
of 3, 6, and 12 months, and L-20: lyophilized sample stored frozen at -20ÊC for a period of 3,
6, and 12 months. Metagenomic DNA concentrations were measured spectrophotometrically
and fluorometrically and ion torrent sequencing was used to assess the bacterial community
composition. The L-20 method could not maintain the yield of DNA during storage. In
addition, the P-80 group showed a greater yield of metagenomic DNA than the other groups
after 6 months of storage. Rumen samples stored as pellets (P-20 and P-80) resulted in
lower richness Chao 1, ACE, and Shannon Wiener indices when compared to PC, while LC
and PC were only different in richness ACE. The storage method and storage time influenced
the proportions of 14 of 17 phyla identified by sequencing. In the P-20 group, the proportion
of Cyanobacteria, Elusimicrobia, Fibrobacteres, Lentisphaerae, Proteobacteria, and
Spirochaetes phyla identified was lower than 1%. In the P-80 group, there was an increase
in the proportion of the Bacteroidetes phylum (p = 0.010); however, the proportion of Actinobacteria,
Chloroflexi, SR1, Synergistetes, TM7, and WPS.2 phyla were unchanged compared
to the PC group (p > 0.05). The class Clostridium was the most abundant in all stored
groups and increased in its proportion, especially in the L-20 group. The rumen sample storage
time significantly reduced the yield of metagenomic DNA extracted. Therefore, the storage method can influence the abundance of phyla, classes, and bacterial families studied
in rumen samples and affect the richness and diversity index.
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Palabras clave
Microbioma ruminal, Acidosis ruminal subagudo