High throughput sequencing has emerged as one of the most important techniques for characterizing microbial dynamics and revealing bacteria and host interactions. However, data interpretation using this technique is mainly based on relative abundance and ignores total bacteria load. In certain cases, absolute abundance is more important than compositional relative data, and interpretation of microbiota data based solely on relative abundance can be misleading. The available approaches for absolute quantification are highly diverse and challenging, especially for quantification in differing biological situations, such as distinguishing between live and dead cells, quantification of specific taxa, enumeration of low biomass samples, large sample size feasibility, and the detection of various other cellular features. In this review, we first illustrate the importance of integrating absolute abundance into microbiome data interpretation. Second, we briefly discuss the most widely used cell-based and molecular-based bacterial load quantification methods, including fluorescence spectroscopy, flow cytometry, 16S qPCR, 16S qRT-PCR, ddPCR, and reference spike-in. Last, we present a specific decision-making scheme for absolute quantification methods based on different biological questions and some of the latest quantitative methods and procedure modifications.

Weaning is one of the most stressful events in the life of a pig. Unsuccessful weaning often leads to intestinal and immune system dysfunctions, resulting in poor growth performance as well as increased morbidity and mortality. The gut microbiota community is a complex ecosystem and is considered an “organ,” producing various metabolites with many beneficial functions. In this review, we briefly introduce weaning-associated gut microbiota dysbiosis. Then, we explain the importance of maintaining a balanced gut microbiota. Finally, we discuss dietary supplements and their abilities to restore intestinal balance and improve the growth performance of weaning pigs.

Archaea are an essential class of gut microorganisms in humans and animals. Despite the substantial progress in gut microbiome research in the last decade, most studies have focused on bacteria, and little is known about archaea in mammals. In this study, we investigated the composition, diversity, and functional potential of gut archaeal communities in pigs by re-analyzing a published metagenomic dataset including a total of 276 fecal samples from three countries: China (n = 76), Denmark (n = 100), and France (n = 100). For alpha diversity (Shannon Index) of the archaeal communities, Chinese pigs were less diverse than Danish and French pigs (p < 0.001). Consistently, Chinese pigs also possessed different archaeal community structures from the other two groups based on the Bray–Curtis distance matrix. Methanobrevibacter was the most dominant archaeal genus in Chinese pigs (44.94%) and French pigs (15.41%), while Candidatus methanomethylophilus was the most predominant in Danish pigs (15.71%). At the species level, the relative abundance of Candidatus methanomethylophilus alvus, Natrialbaceae archaeon XQ INN 246, and Methanobrevibacter gottschalkii were greatest in Danish, French, and Chinese pigs with a relative abundance of 14.32, 11.67, and 16.28%, respectively. In terms of metabolic potential, the top three pathways in the archaeal communities included the MetaCyc pathway related to the biosynthesis of L-valine, L-isoleucine, and isobutanol. Interestingly, the pathway related to hydrogen consumption (METHANOGENESIS-PWY) was only observed in archaeal reads, while the pathways participating in hydrogen production (FERMENTATION-PWY and PWY4LZ-257) were only detected in bacterial reads. Archaeal communities also possessed CAZyme gene families, with the top five being AA3, GH43, GT2, AA6, and CE9. In terms of antibiotic resistance genes (ARGs), the class of multidrug resistance was the most abundant ARG, accounting for 87.41% of archaeal ARG hits. Our study reveals the diverse composition and metabolic functions of archaea in pigs, suggesting that archaea might play important roles in swine nutrition and metabolism.

In humans and animals, probiotics are widely accepted as crucial for host health and growth. The investigation of the probiotic colonization and expression of probiotics in the host is beneficial for proper usage of probiotics and isolation of indigenous probiotics. In this study, we analyzed commonly used probiotic strains in the intestines/rumen of humans and animals by analyzing metagenomic and paired meta-transcriptomic data from the gut or rumen microbiome of humans (n = 13), pigs (n = 6), chickens (n = 6), cattle (n = 14), sheep (n = 10), and mice (n = 8). First, we generated an expression profile based on 192 selected representative probiotic strains from a published database. A total of 58 probiotic strains were not detected in any samples, while 3 strains were presented and expressed in all individuals. Overall, the probiotic expression of probiotics as detected by meta-transcriptome was significantly higher than the relative abundance of probiotic as detected by metagenomics in cattle, sheep, mice, and humans; however, this difference was not significant in pigs and chickens. In total, 17 (cattle), 21 (sheep), 22 (pig), 14 (chicken), 13 (mouse), and 3 (human) probiotic strains were identified as probiotic strains with significantly higher expression levels [Fold Change (FC) ≥ 2, False Discovery Rate (FDR) ≤ 0.05]. Among them, Clostridium butyricum TOA was found to be significantly expressed in the rumen or gut of all host species. In addition, network analysis based on the expression of probiotics as detected by meta-transcriptomics revealed that several probiotic strains were significantly negatively linked with Salmonella spp., Mycoplasma spp., and Escherichia coli. The results in this study provide a useful reference for developing indigenous probiotics.