Analysis of the methanogenic and acetogenic community structure in young calves

Abstract: The newborn calf that has not been fed colostrum lacks immune defense and ruminal microorganisms needed for digestion of polysaccharides from plants and cellulose. Establishment of a gastrointestinal microbiota is vital. This study aimed to shed light on the establishment of the methanogenic and acetogenic community in young calves. Nine naturally born bull calves at the age of 2 or 7 days were used for collecting fecal samples and intestinal tissue and digesta from various segments of the gut. In addition, 31 heifer calves of various ages (2 days - 6 months) were used only for fecal sampling. DNA was isolated from the samples and polymerase chain reaction (PCR) used to amplify the methyl coenzyme-M reductase (mcrA) genes for detection of methanogens and the formyltetrahydrofolate synthetize gene (FHS) for the acetogens. Terminal-restriction fragment length polymorphism (T-RFLP) analysis was performed on PCR products generated from methanogens and acetogens. T-RFLP does not give any data about which species that are present in the microbial community. In order to get more information on species level, clone libraries were created from representative samples from both methanogens and acetogens. Fecal samples from one 6 months old and one 2 weeks old calf were chosen for generation of a library for the methanogens whereas for the acetogens, four libraries were created from a fecal sample from a 6 month old calf and feces, rumen- and cecum digesta from a 2 days old calf. Gel electrophoresis of the PCR confirmed presence of methanogens in 21 faecal samples from calves 14 days of age and older. Not all of the samples from 14 and 28 days old calves were positive for methanogens. However, from 6 weeks and older the presence of methanogens seemed to be established and all samples contained methanogens. Analysis of the acetogens gave positive results for both faecal, GI digesta and tissue samples. Interestingly the tissue that contained acetogens came mainly from rumen and abomasum samples from 7 days old calves. This may indicate that the acetogens primarily adhere to the tissue of those regions but pass through the rest of the GIT and get picked up in digesta samples from other sites. T-RFLP analysis on the methanogenic community composition revealed a shift according to age of the calves. The PCA analysis of the T-RFLP data revealed two clusters of samples from calves at 2-8-weeks age and a third cluster of samples from 6-months old calves. Among the acetogenic samples there was a large spread among 2-day calves whereas the 6-month old calves clustered more closely in the PCA plot. Which indicates that the within group similarity in the microbiota increases as the calves get older. The results from this study indicate that the methanogens are not present in young animals. Acetogens were found in intestinal and faeces samples from calves 2 days and older. The method could not confirm whether the acetogens were homoacetogenic or not.