Effects of early-life rumen inoculation on the long-term microbial community development and production outcomes in dairy cows | Maitokarjalle varhaiskasvun aikana annostellun pötsinesteen pitkäaikaisvaikutukset mikrobiston kehittymiseen ja tuotokseen

Modulating rumen microbiota during the pre-weaning period has been suggested as a means to enhance animal performance later in life. This thesis aimed to elucidate how a rumen liquid inoculum from an adult cow, provided to calves during the pre-weaning period, influences the establishment of rumen (I) and fecal (III) bacterial, archaeal, fungal, and ciliate protozoan communities in monozygotic twin calves, and whether the treatment induces long-term post-treatment effects on microbiota composition (II) and production performance (II). Six pairs of twin calves were divided into treatment (T-group) and control (C-group) groups immediately after birth. The T-group received fresh rumen liquid from an adult cow as an oral inoculum from 2 to 8 weeks of age, while the C-group was managed identically but not inoculated. Fecal samples were collected weekly, and rumen samples every other week (weeks 2, 4, 6, 8), between 1–8 weeks of age, to monitor the microbial community composition. After weaning at 8 weeks of age, the heifers were group-housed and were no longer treated. Their rumen microbial composition, rumen fermentation, and growth were monitored from the age of 3 months until they were 1 year old. Later during their first lactation, the effects of rumen liquid dosing on production parameters, such as weight gain, dry matter intake, residual energy intake, energy-corrected milk, milk composition, and methane production were measured. Rumen and fecal microbial community compositions were determined using the bacterial and archaeal 16S ribosomal RNA (rRNA) gene, the protozoal 18S rRNA gene, and fungal ITS1 region amplicon sequencing. In the first publication (I), the aim was to elucidate how dosing with fresh rumen liquid impacts the establishment of rumen microbial communities and whether it affects rumen function, feed intake, and growth during the treatment period. The treatment tended to increase concentrate intake and increased the weekly weight gain. The inoculum stimulated the earlier establishment of bacteria related to the mature rumen in the T-group. The archaeal communities differed significantly between groups until week 4. The inoculum also increased archaeal OTU diversity but did not affect archaeal quantity. The C-group remained protozoa-free until week 6, while in the T-group ciliate protozoa were detected by week 2. Likely due to different seeding sources, the ciliate protozoan community composition differed between the C- and T-groups by week 8. All calves were observed to have anaerobic fungi at week 2, but in some calves the fungi disappeared towards weaning. Anaerobic fungal diversity in the rumen decreased with age and was likely influenced by changes in diet and interactions with other rumen microorganisms. The results indicate that the rumen liquid inoculum from an adult cow enhanced the maturation of bacterial and archaeal rumen communities in pre-weaning calves, though its effect on eukaryotic communities requires further investigation. The second publication (II) aimed to assess whether the effects of rumen liquid dosing on microbiota and rumen function persisted post-treatment and whether the treatment impacted production performance. Although weight gain was unaffected, the butyrate proportion was higher in the T-group at month 3. The richness of bacteria and ciliate protozoa increased until month 7 and anaerobic fungi until month 11, but the archaea richness did not significantly increase with age. The community composition of archaea, ciliate protozoa, bacteria, and anaerobic fungi continued to develop until months 3, 6, 7, and 10, respectively. The treatment stimulated the earlier maturation of the prokaryote community in the T-group, with effects persisting up to month 4, and the earlier maturation of ciliate protozoa until month 2. However, no effect on the maturity of the anaerobic fungal community was observed. During the first lactation, the T-group exhibited lower variation in energy-corrected milk yield, a tendency for different residual energy intake patterns, and numerically lower somatic cell counts, but there were no significant differences in dry matter intake or methane emissions. The results demonstrated that the oral microbial inoculant induced transient changes in early rumen microbiome maturation and may influence later production performance, though the mechanisms require further exploration. The third publication (III) aimed to investigate how the dosing with rumen liquid inoculum affected hindgut microbiota composition during the pre-weaning treatment period, and after weaning when the treatment had ended. Secondly, the aim was to assess if the treatment affected gut health during the pre-weaning period. Additionally, the contribution of donor rumen liquid inoculum, colostrum, and the calf's own rumen as seeding sources for microbes was assessed. The fecal microbiota composition was monitored from one week old until the heifers were one year old. The rumen modulation had an insignificant effect on age-related fecal microbiota development. The fecal bacterial community gradually evolved in response to dietary changes, forming distinct pre-weaning and post-weaning communities. Bacterial richness increased with age and stabilized by month 9, while variation between samples decreased after weaning. The quantity of archaea in fecal samples increased after month 4, with archaeal richness stabilizing by month 9. Anaerobic fungi were detected in feces at month 4, reaching a stable community richness at month 7. Prior to month 6, the fungal community composition differed from that of mature communities. The calf's own rumen was the primary seeding source for fecal bacteria and fungi. The donor inoculum did not have an impact on gut health, as diarrhea rates were similar between the T-group and C-group. In conclusion, early-life microbiota modulation shows potential in improving rumen microbial community development. However, a more targeted approach with microbes adapted to the hindgut environment may be necessary for the effective modulation of the hindgut.

Eläinten varhaiskasvun aikana toteutettavaa suolistomikrobiston muokkaamista on tutkittu yhtenä keinoista parantaa tuotantotehokkuutta ja terveyttä, sekä vähentää eläintuotannon ilmastopäästöjä. Tämä väitöskirjatutkimus pyrki selvittämään, miten vasikalle annosteltu aikuisen lehmän pötsineste vaikuttaa vasikoiden pötsin ja ulosteen mikrobiyhteisöjen kehittymiseen ja onko käsittelyllä pitkäaikaisia vaikutuksia mikrobiston koostumukseen tai tuotantotehokkuuteen. Tutkimus toteutettiin seuraamalla identtisten kaksosvasikkaparien suolistomikrobiston kehittymistä, pötsin toimintaa ja vasikoiden kasvua viikon iästä vuoden ikään saakka. Koeryhmän (n=6) vasikoille annosteltiin tuoretta pötsinestettä (5–10 ml) kolmesti viikossa vieroituskauden ajan. Kontrolliryhmän (n=6) vasikat kasvatettiin samoissa olosuhteissa, mutta ne eivät saaneet pötsinestettä. Myöhemmin, ensimmäisen lypsykauden aikana, mitattiin käsittelyn vaikutuksia tuotantoparametreihin. Tulokset osoittivat, että pötsinesteannostelu vaikutti mikrobiryhmiin eri tavalla. Annostelu edesauttoi pötsin bakteeri- ja arkeoniyhteisöjen kehittymistä ja alkueläinten kolonisaatiota, kun taas vaikutukset anaerobisiin sieniin olivat kaksijakoisia. Vakaa sieniyhteisö muodostui kaikille vasikoille vasta vieroituksen jälkeen. Käsittelyn vaikutukset olivat nähtävissä bakteereissa ja alkueläimissä vielä noin kaksi kuukautta annostelun päättymisen jälkeen. Arkeoniyhteisössä tai anaerobisessa sieniyhteisössä ei kuitenkaan havaittu pitkäaikaisia vaikutuksia. Pötsinesteen annostelulla ei ollut merkittävää vaikutusta ulostemikrobiston kehitykseen tai vieroituskauden suolistoterveyteen. Ulosteen bakteeriyhteisö kehittyi vasikoilla aikuisen kaltaiseksi aiemmin kuin pötsin bakteeriyhteisö, mahdollisesti vakaamman elinympäristön vuoksi. Vaikka arkeoneja esiintyi vasikoiden ulosteissa vieroituskaudella, niiden määrä ja monimuotoisuus lisääntyivät vasta eläinten siirryttyä kiinteään rehuun. Anaerobisia sieniä esiintyi ulosteissa vasta vieroituksen jälkeen, ja niiden monimuotoisuus lisääntyi vuoden ikään saakka. Pötsinesteannostelu edesauttoi lievästi vasikoiden painonnousua ja paransi kiinteän rehun syöntiä vieroituskaudella, joskaan vaikutuksia ei voitu selittää pötsin fermentaatioparametreilla. Ensimmäisen lypsykauden aikana koeryhmän lehmillä oli hieman tasaisempi maitotuotos ja jäännösenergian saanti, sekä hieman pienempi maidon soluluku kuin kontrollilehmillä, mutta metaanintuotannossa ei havaittu eroja. Tarvitaan kuitenkin lisätutkimuksia suuremmalla eläinmäärällä, jotta tulokset sekä mekanismit, joiden kautta mikrobisto vaikuttaa lehmien tuotantotehokkuuteen, voidaan vahvistaa.

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