Metabarcoding Analysis Research Articles

Salps in the NW Atlantic Slope Water: metabarcoding and compound-specific stable isotope analysis of diet diversity and trophic interactions

Salps in the NW Atlantic Slope Water: metabarcoding and compound-specific stable isotope analysis of diet diversity and trophic interactions

Comparison of commonly used software pipelines for analyzing fungal metabarcoding data.

Metabarcoding targeting the internal transcribed spacer (ITS) region is commonly used to characterize fungal communities of various environments. Given their size and complexity, raw ITS sequences are necessarily processed and quality-filtered with bioinformatic pipelines. However, such pipelines are not yet standardized, especially for fungal communities, and those available may produce contrasting results. While some pipelines cluster sequences based on a specified percentage of base pair similarity into operational taxonomic units (OTUs), others utilize denoising techniques to infer amplicon sequencing variants (ASVs). While ASVs are now considered a more accurate representation of taxonomic diversity for prokaryote communities based on 16S rRNA amplicon sequencing, the applicability of this method for fungal ITS sequences is still debated. Here we compared the performance of two commonly used pipelines DADA2 (inferring ASVs) and mothur (clustering OTUs) on fungal metabarcoding sequences originating from two different environmental sample types (fresh bovine feces and pasture soil). At a 99% OTU similarity threshold, mothur consistently identified a higher fungal richness compared to DADA2. In addition, mothur generated homogenous relative abundances across multiple technical replicates (n = 18), while DADA2 results for the same replicates were highly heterogeneous. Our study highlights a potential pipeline-associated bias in fungal metabarcoding data analysis of environmental samples. Based on the homogeneity of relative abundances across replicates and the capacity to detect OTUs/ASVs, we suggest using OTU clustering with a similarity of 97% as the most appropriate option for processing fungal metabarcoding data.

A baroduric immobilized composite material promoting remediation of oil-polluted sediment at typical deep-sea condition: The performances and potential mechanisms

Contriving immobilized bioreagent is of great significance to enhance bioremediation of marine oil pollution. However, there remains a notable scarcity of correlational study conducted at deep sea condition. Herein, we first developed a baroduric microsphere encasing biotic and chemical materials to remediate oil-contaminated sediments at deep-sea microcosm. Total oil degradation efficiency of microsphere-treated group reached 71% within a month, representing an approximate 35% increase compared to natural remediation. Absorption and biodegradation by microsphere provided a comparable contribution to oil elimination. Together with scanning electron microscope observation, the physical mechanism was that the reticulate structure of microsphere surface facilitating oil adsorption and bacteria attachment. Via metabarcoding analysis for meta and metabolically-active microbes, we demonstrated the primary working center was located at the microsphere. Proteobacteria, Firmicutes, Bacteroidota and Desulfobacterota were the key activated bacteria. More importantly, we revealed the ecological mechanisms were associated with the following aspects: 1) the addition of microsphere significantly improved the metabolic activity of bacteria (particularly including several oil-degrading taxa); 2) the microspheres enhanced ecological stability and microbial functional diversification during bioremediation; 3) expressing activity of pathways involving oil component degradation, biosurfactant production, biofilm architecture, biogeochemical and energy cycling all were observed to be up-regulated in microsphere-treated samples. Altogether, our results provide important theoretical guidance and data support on application of immobilization technology in removing in-situ oil pollution of deep-sea sediment.

Unsuspected pollinator potential? First DNA dietary analysis in Euleptes europaea (Gené, 1839) and confirmation of an omnivorous diet using video surveillance

Abstract DNA analysis of the faeces of wild animals is a major step towards our understanding of the dietary requirements of cryptic and sometimes threatened species. The presence of digested and liquefied matter undetectable to the naked eye can be identified and used to detect the presence of plant resources. Validating the consumption of plant material is tricky and often controversial, as it may have been taken unintentionally while hunting prey. Here, we present the very first dietary DNA metabarcoding analysis of the gecko Euleptes europaea. We identified one class and three orders of Arthropoda, and two classes of plants. To validate the results of plant presence, we set up a novel feeding selectivity protocol, followed by video surveillance in the wild. We offered four plant resources, two of which were close relatives to the found metabacoded taxa. While a food selectivity protocol combined with DNA analysis validated the consumption of plant resources, they showed that these geckos had a preference for plant pollen but refused other sweet resources. These results raise the possibility of a pollinating role of this gecko, which can sometimes be the only vertebrate to inhabiting the smallest islets in the Mediterranean Sea.

Can eDNA Present in Aquatic Environments of Rural Areas Help Identify Species Diversity in the Order Anura?

Paddy fields are classified as wetland environments, and they comprise freshwater ecosystems. They are ecologically important habitats and breeding grounds for many aquatic insects, amphibians, and reptiles. However, paddy field ecosystems are constantly threatened by climate change and the indiscriminate use of pesticides. The metabarcode analysis of eDNA (environmental DNA) method is highly effective at accumulating information on many organisms that inhabit paddy field ecosystems. It can indirectly identify the existence of taxa that are no longer found in the target ecosystem due to behavioral characteristics, such as those exhibited by amphibians. In the metabarcoding results of this study, genes of five species of frogs were found, but it was impossible to confirm all of the frogs’ taxa, morphological pictures, and croak sounds. On the other hand, some frog taxa were only found in the metabarcoding analysis. The eDNA of the frogs found only in the metabarcoding analysis is estimated to have been introduced from nearby areas inhabited by frogs rather than the target region. Due to the powerful analytical resolution of eDNA metabarcoding, this eDNA-based paddy field search is expected to help investigate the biodiversity in agricultural ecosystems.

Are passive collectors effective samplers of microbes in natural aquatic systems?

Biodiversity surveys of aquatic systems often include DNA metabarcoding analyses of environmental samples that are collected through filtration of large volumes of water. The standard practice of sterile collection and filtration in or near the field sites is challenging to implement in remote locations, and filtration of large volumes is a limiting step, especially for water from highly productive systems or with high suspended sediment loads. Recent trials have shown that passive samplers can be effective for aquatic metabarcoding to document metazoan diversity, but that this approach needs to be trialed under a wider variety of conditions and across more diverse taxa. Here we assess the utility of passive sampling for documenting the diversity of bacteria in six tropical aquatic environments (one lake, one reservoir, two mountain streams and two blackwater rivers). We find that passive collectors generally recover significantly higher diversity of Bacteria compared to filtered samples, despite capturing significantly less overall DNA than active water filtering. However, the communities captured by the two methods show significant differences within sites, with only 26% of the Bacteria ASVs recovered by both methods. These differences were largely driven by relative abundances of taxa within Actinobacteriota, Campilobacterota, Desulfobacterota, and Proteobacteria. Our results demonstrate that passive collectors can be a cost-effective solution for monitoring aquatic microbial diversity but that the two methods are not interchangeable. Additional work is necessary to understand the selectivity of both passive collectors and active water filtering for eDNA studies.

DNA metabarcoding analyses reveal fine-scale microbiome structures on Western Canadian bat wings.

Microbiomes play important roles in host health. White-nose syndrome (WNS), a fungal infection of bat wings and muzzles, has threatened bat populations across North America since 2006. Recent research suggest that the skin microbiome of bats may play a significant role in bat's susceptibility to WNS. However, relatively little is known about the skin microbiome composition and function in bats in Western Canada, a region with a high diversity of bats, but WNS has yet to be a major issue. Here, we revealed high bacterial and fungal diversities on the skin of three common bat species in Lillooet, British Columbia, including several highly prevalent microbial species that have been rarely reported in other regions. Our analyses showed fine-scale structures of bat wing microbiome based on local sites and bat species. The knowledge obtained from WNS-naïve bat populations in this study may help develop mitigation and management strategies against WNS.

Looking for Pathogens in Dust from North Africa Arriving in the French West Indies Using Metabarcoding and Cultivable Analysis.

Periodically, the French West Indies receive dust originating from North Africa (NA). Microorganisms associated with desert dust can be transported over long distances through the atmosphere and could represent a means for the remote colonization of new habitats by putatively pathogenic microorganisms. The aim of this study was to determine the diversity and frequency of microbial agents (bacteria, eukaryotes) in NA dusts and the potential threat toward human and/or animal health by comparing microbial air composition during dust events and in control samples. In 2017 and 2018, 16 samples were collected during seven NA dust episodes and there were 9 controls. The microbial composition of the samples was characterized using a cultivable approach and by metabarcoding analyses (16S and 18S). A greater bacterial load and greater diversity were observed during the dust events, and some genera were significantly associated with the events. Some, such as Geodermatophilus, can be considered signature species of NA dust. No pathogenic species were found with the cultivable approach, whereas the metabarcoding analyses highlighted the presence of several potentially pathogenic species or known human pathogens such as Naegleria fowleri.

Irradiance level and elevation shape the soil microbiome communities of Coffea arabica L.

BackgroundThe nexus plant-microbe-environment is essential to understand the ecosystem processes shaping plant health and fitness. Within this triangle, soils and associated microflora are among the key ecosystem’s drivers, underpinning plant productivity and evolution. In this study, we conducted a comprehensive analysis (physicochemical properties, enzyme activities, and taxonomic diversity) of soils under the canopy projection of Coffea arabica trees along a gradient of elevation (600, 800, and 900 m) and shade (0, 50, 100%).ResultsWhile shade had no influence on most parameters, altitude shaped the dynamics of microbial communities. Available phosphorus, soil organic carbon, and nitrate were significantly higher at 800 m, likely due to the higher activities of β-glucosidase and phosphatases at this altitude. Microbial biomass (carbon and nitrogen) and moisture were significantly higher at 600 and 900 m, which might be attributed to the abundance and richness of soil microorganisms. Indeed, metabarcoding analysis revealed a complex pattern of microbial consortia (bacteria, archaea, fungi) at the three altitudes, with the lowest index of richness recorded at 800 m. The highest number of Amplicon Sequence Variants was observed in bacteria, whose functional analysis revealed distinct metabolic adaptations across different altitudes. At 900 m, the main functional attributes favored the responses to environmental stimuli and microbial interactions; at 800 m, the predominant metabolic pathways were related to organic matter, fermentation, and bioremediation; and at the lower 600 m, the pathways shifted towards the breakdown of plant-derived compounds (e.g. geraniol, limonene, and pinene degradation).ConclusionOverall, the results indicate a higher effectiveness of the microbial consortium at 800 m, which might result in better nutrient cycling. The study highlights the importance of canopy shade species and elevation for the composition of microbial consortia in C. arabica, unveiling ecological functions beyond plant health, with implications for bio-based solutions and biotechnology.

Environmental RNA metabarcoding for ballast water microbial diversity: Minimizing false positives

While maritime transport boosts global trade by shipping bulk goods, it raises concerns about the spread of harmful bacteria via ballast water. Moreover, the dark and cold environments of ballast tanks often harbor extracellular DNA from dead organisms, leading to false positives in traditional environmental DNA (eDNA) metabarcoding analyses. Here, we alternatively employed environmental RNA (eRNA) metabarcoding to assess its potential for reducing false positive in ballast water monitoring. We collected eDNA and eRNA in parallel from ballast water before and after disinfection in three vessels. High-throughput sequencing of the 16S rRNA V4-V5 regions and cDNA counterparts was conducted to compare bacterial community composition. Our findings showed that over 80 % of the top 150 abundant amplicon sequence variants (ASVs) were detected by both eRNA and eDNA metabarcoding. Samples sequenced separately using DNA and RNA consistently clustered together, indicating similar community recovery efficacy. However, 42 % of ASVs were detected exclusively in DNA, resulting in significantly higher bacterial diversity compared to RNA, which suggests false positives in the DNA method. In treated samples with higher dead cell counts, the RNA method showed significantly lower bacterial diversity, indicating its effectiveness in detecting live bacteria. In summary, eRNA metabarcoding offers comparable recovery efficiency while maintaining a lower false-positive rate than eDNA metabarcoding.

Metabarcoding and morphological analysis of diets of mesopelagic fishes in the NW Atlantic Slope Water

Metabarcoding and morphological analysis of diets of mesopelagic fishes in the NW Atlantic Slope Water

Water and nutrient availability modulate the salinity stress response in Olea europaea cv. Arbequina

Salinity stress represents a key factor for global agriculture. Plants can respond to salinity stress by adapting their physiology in different ways with the aim of limiting reductions in growth and development. Importantly, moisture retention capacity, permeability and nutrient availability of substrates represent critical variables for plants as they may further influence the effect of osmotic stress. Here, a multidisciplinary approach was applied to evaluate the role of two different substrates, peat and perlite, on 2-year-old potted cuttings of Olea europaea (cultivar Arbequina) under different salinity stress conditions (0, 100 and 200 mM NaCl). Biometric and physiological data indicate that plants potted in perlite (AP) generally present lower growth and photosynthetic rates when compared with peat (AS) in combination with salinity stress. Ion measurements indicate a rise in Na+ accumulation with increasing stress severity, which alters the ion ratio in both substrates. In addition, differences occurred in polyphenol contents, with a general increase in quinic acid and rutin contents in AS and AP samples, respectively. Metabolomic and biometric data were also coupled with metabarcoding analysis, which indicates that the moderate salinity treatment (100 mM NaCl, T100) reshaped the endophytic community of plants grown on both substrates. Taken together, the data suggest that the strategy used by a glycophytic species such as the olive tree to cope with salinity stress seems to be highly related to availability of water and nutrients. The lack of both may be simulated by perlite, enhancing the effect of salinity stress response in woody plants. Lastly, applying the beneficial endophytic bacterial taxa identified here could represent a step forward in increasing plant defence and nutrient uptake and reducing inputs for modern and more sustainable agriculture.

Strong seasonality and unsuspected diversity of haptophytes explored by metabarcoding analysis in the Chinese seas

The haptophytes, are essential components of the marine pico- and nano-plankton but little is known about their diversity and abundance due to the small size. In this study, the taxonomic composition, geographical distribution, and seasonal variation of the haptophytes in the Bohai Sea, the Yellow Sea, and the East China Sea were investigated using DNA metabarcoding in April and October of 2021. A total of 623 and 3756 haptophyte amplicon sequence variants (ASVs) were obtained in spring and autumn, respectively. All currently described or detected haptophyte orders were retrieved, including several deep-branching novel environmental lineages with relative high abundance. The predominant groups were Chrysochromulina, Clade HAP 2-3-4-5, Phaeocystis, and Prymnesium in spring, and Chrysochromulina, Phaeocystis, and Emiliania/Gephyrocapsa Compelx in autumn. The richness and diversity showed seasonal variation, with higher alpha diversity occurred in autumn than that of spring. Different haptophyte taxa exhibited unique spatial distribution patterns and water temperature was significantly correlated with the observed community dissimilarities and was the most influential driving factor in both seasons. Our results highlight the high hidden diversity and seasonal variations of haptophytes in the Chinese seas.

Horse Meat Microbiota: Determination of Biofilm Formation and Antibiotic Resistance of Isolated Staphylococcus Spp.

Domestic horses could be bred for leisure activities and meat production, as is already the case in many countries. Horse meat is consumed in various countries, including Kazakhstan and Kyrgyzstan, and with the increase in this consumption, horses are registered as livestock by the Food and Agricultural Organization. In this study, horse meat microbiota of horse samples (n = 56; 32 samples from Kazakhstan and 24 samples from Kyrgyzstan) from two countries, Kazakhstan (n = 3) and Kyrgyzstan (n = 1), were investigated for the first time by next-generation sequencing and metabarcoding analysis. The results demonstrated that Firmicutes, Proteobacteria, and Actinobacteria were the dominant bacterial phyla in all samples. In addition, three (5.4%) Staphylococcus strains were isolated from the Uzynagash region, Kazakhstan. Staphylococcus strains were identified as Staphylococcus warneri, S. epidermidis, and S. pasteuri by partial 16S rRNA DNA gene Sanger sequencing. All three Staphylococcus isolates were nonbiofilm formers; only the S. pasteuri was detected as multidrug-resistant (resistant to penicillin, cefoxitin, and oxacillin). In addition, S. pasteuri was found to carry mecA, mecC, and tetK genes. This is the first study to detect potentially pathogenic Staphylococcus spp. in horse meat samples originating from Kazakhstan. In conclusion, it should be carefully considered that undercooked horse meat may pose a risk to consumers in terms of pathogens such as antibiotic-resistant Staphylococcus isolates.

Contribution of data acquired from spectroscopic, genomic and microbiological analyses to enhance mussels’ quality assessment

In this study, a large amount of heterogeneous data (i.e., microbiological, spectral and Next Generation Sequencing data) were obtained analyzing mussels of different species and origin, to acquire a comprehensive view about the quality and safety of these products. More specifically, spectral data were collected through Fourier transform Infrared (FTIR) spectroscopy, while the overall profile of microorganisms present in these samples, affecting quality and safety of mussels throughout storage, was determined through Next Generation Sequencing (NGS) using 16S rRNA metabarcoding analysis. In parallel, conventional microbiological analysis for the estimation of culturable spoilage microorganisms (total aerobes, Pseudomonas spp., B. thermosphacta, Shewanella spp. and Enterobacteriaceae) was applied. Different machine learning algorithms, namely Partial Least Square (PLS), Support Vector Machines (SVM), k-Nearest Neighbors (kNN), Random Forest (RF) Neural Networks (NN)) were applied accordingly, to assess the potential of FTIR and NGS data to provide useful information about mussels’ microbiological quality.Microbial counts ranged from 3.5 to 9.0 log CFU/g, while NGS revealed several bacterial genera such as Pseudoalteromonas, Psychrobacter, Acinetobacter, Pseudomonas, B. thermosphacta, Psychrobacter, Kistimonas, Psychrilyobacter to affect the quality of mussels, depending on the mussel species, batch and storage conditions. According to the performance metrics, the SVM algorithm in tandem with FTIR achieved the highest prediction accuracy for microbial counts in M. chilensis samples (Rsquared; 0.89, RMSE; 0,74), while in the case of predicting the abundance of microbial genera using spectroscopic data, the best performing algorithm varied by bacterial genus. Indicatively, in M. chilensis, RF, kNN and NN performed better in predicting Enterococcus, Enhydrobacterium and Pseudoalteromonas, respectively (Rsquared = 0.92, 0.93, 0.99). Associations between genomics data and specific spectral regions were further investigated, revealing certain spectral regions that are associated with mussels’ quality and safety.The application of “multi-omics” in seafood supply chain can provide insightful information about mussels’ quality and safety compared to the methodologies followed in current quality and safety management systems.

Comparison of the trophic sources and pathways of mesozooplankton and ichthyoplankton in the Kuroshio current and its neighboring waters

Commercially important fish spend their vulnerable early life stages in the Kuroshio Current, resulting in high fishery production even in the vicinity of poor prey availability under oligotrophic conditions. Nevertheless, little information is available on how ichthyoplankton are supported by trophodynamics in complicated food webs. Here, we have explored trophic sources and pathways toward ichthyoplankton in the Kuroshio and its neighboring waters based on metabarcoding analysis of gut DNA content for major taxonomic groups of mesozooplankton and ichthyoplankton. Calanoids were found to be the most predominant and frequently appearing prey, whereas non-crustaceans were the secondary prey for most mesozooplankton and ichthyoplankton groups. Trophic networks based on gut DNA content demonstrated that calanoids were the most important sector with multiple linkages among their prey and predators, and gelatinous and non-crustacean mesozooplankton were the secondary sectors. These findings suggest that calanoids are important hubs of trophic pathways toward ichthyoplankton, and that gelatinous and non-crustacean mesozooplankton groups strengthen trophic relationships with multiple components. Contrary to general thought, our metabarcoding analysis has revealed that trophodynamics toward ichthyoplankton are not strongly dependent on the grazing food chain, but are supported by multiple trophic pathways in the Kuroshio and its neighboring waters.

Impact of the use of an indigenous starter on microbial community structuring and aromatic profile of rocamadour protected designation of origin cheese

The use of indigenous strains as starters offers new opportunities that challenge the use of industrial starters, especially to strengthen the link between Protected Designation of Origin (PDO) cheeses and their terroir. This study investigated the effect of two starters, industrial and indigenous, on the microbial ecosystem and volatile organic compounds (VOCs) in Rocamadour PDO cheese. Metabarcoding analysis showed that regardless of the type of starter used, Lactococcus lactis and Geotrichum candidum largely dominated the microbial communities of Rocamadour (>96 %). The analysis of β-diversity based on the Bray-Curtis indices differentiated the starters according to their bacterial communities. VOC analysis showed that the starters were distinguished by three fatty acids characteristic of the goat cheese aroma (p-value <0.05): decanoic, hexanoic, and octanoic acids. Hexanoic acid was not present in cheeses produced with the indigenous starter. This fatty acid was positively correlated with the presence of Paucilactobacillus nenjiangensis, Lactiplantibacillus sp. and Enterobacteriaceae, these microorganisms that were also characteristic of cheeses made with the industrial starter. Octanoic and decanoic acid, on the other hand, were suggested to be correlated with the presence of Leuconostoc mesenteroides.Cheeses produced with the indigenous starter differed in bacterial communities and aromatic signatures.

From small water bodies to lakes: Exploring the diversity of freshwater bacteria in an Alpine Biosphere Reserve

Small water bodies, although supporting high biodiversity, are often understudied in the Alpine region. In this work, we characterized the planktic and benthic bacterial communities, as well as the water chemistry, of a wide physiographic range of 19 freshwater bodies within an Alpine Biosphere Reserve, including ponds, pasture ponds, peat bogs, shallow lakes, and lakes. We collected both water and surface sediment samples, followed by metabarcoding analysis based on the V3-V4 regions of the 16S rRNA gene. We investigated the changes in biodiversity and the distribution of unique and shared amplicon sequence variants (ASVs) between water (11,829 ASVs) and surface sediment (19,145 ASVs) habitats, as well as across different freshwater typologies. The majority of ASVs (78 %) were unique to a single sample, highlighting the variability and uniqueness of bacterial communities in such freshwater bodies. Most freshwater environments showed higher α-diversity in sediment samples (median, 1469 ASVs) compared to water (468 ASVs). We found that water and sediment habitats harboured unique bacterial communities with significant differences in their taxonomic compositions. Benthic bacteria were associated with several biogeochemical and degradative processes occurring in the sediments, with no notable differences among freshwater typologies and with phylogenetically and ecologically similar species. Conversely, planktic communities showed greater heterogeneity: small water bodies and peat bogs were characterized by higher relative abundances of Patescibacteria (up to 33 %), while lakes and shallow lakes were dominated by Actinobacteriota (up to 36 %). Cyanobacteria (426 ASVs) were generally distributed at low abundances in both water and sediment habitats. Overall, our results provided essential insights into the bacterial ecology of understudied environments such as ponds and pasture ponds and highlighted the importance of further exploring their rich pelagic and benthic bacterial biodiversity.

Molecular diversity and seasonal dynamics of Ostreococcus (Mamiellophyceae, Chlorophyta) in typical mariculture bays based on metabarcoding analysis

Ostreococcus (Mamiellophyceae, Chlorophyta) is a cosmopolitan genus of marine pico-phytoplankton and the smallest free-living photosynthetic eukaryotes with cell size of 1–2 μm. To understand the diversity and spatio-temporal distribution of Ostreococcus in the Rongcheng coastal regions in northern China, metabarcoding analysis based on the 18S rDNA V4 molecular marker was applied to study the molecular diversity and seasonal dynamics of Ostreococcus in three typical mariculture bays (Rongcheng Bay, Lidao Bay and Sanggou Bay). A total of 103 amplicon sequence variants (ASVs) annotated as Ostreococcus were detected in these three typical mariculture bays throughout the year. The top five ASVs in terms of abundance were ASV4, ASV9, ASV14, ASV28 and ASV109, totally occupying 99.1% of Ostreococcus reads. Phylogenetic analysis showed that these five dominant ASVs represented two Ostreococcus ecotypes (OI and OII) and were grouped into four Ostreococcus clades including Ostreococcus lucimarinus (ASV9) and Ostreococcus tauri (ASV28 and ASV109) in OI, and Ostreococcus sp. RC1 (ASV4) and Ostreococcus sp. RC2 (ASV14) in OII, which provided direct evidence to support the co-existence of two ecotypes in the Rongcheng coastal regions. Five dominant ASVs in OI and OII exhibited two distinct seasonal distribution patterns. Three dominant ASVs (ASV9, ASV28 and ASV109) in OI could be detected in all four seasons of the year, exhibiting native distribution properties, while two ASVs (ASV4 and ASV14) in OII decreased sharply in winter and could not be detected in spring, exhibiting characteristics of alien inputs. The composition, succession and association of Ostreococcus community were mainly driven by water temperature in these mariculture bays. This study helps us systematically understand the molecular diversity and distribution patterns of Ostreococcus in typical mariculture bays in northern China, laying the foundation for understanding and revealing the ecological functions of pico-phytoplankton.

Daily Intake of Household-Produced Milk Kefir on Salmonella Typhimurium Infection in C57BL/6 Mice: Mortality, Microbiota Modulation and Immunological Implications.

Salmonellosis, a major global cause of diarrheal diseases, significantly impacts the intestinal microbiome. Probiotic-rich beverages, such as kefir, are increasingly utilized as alternative health-promoting beverages associated with various microbiota benefits. This study investigated the repercussions of daily consumption of household-produced milk kefir on Salmonella enterica serovar Typhimurium infection in C57BL-6 mice. Kefir consumption pre infection reduced the presence of inflammatory cells in the colon and altered the cytokine profile by reducing IL-10 and increasing IFN-γ. Despite reducing intestinal inflammation, kefir intake did not yield a prompt response to an acute infection caused by the aggressive pathogen Salmonella. This contributed to increased mortality in the mice, evidenced by higher fecal Salmonella counts post-infection. Metabarcoding analysis demonstrated that the use of kefir before infection increases butyric acid by the higher abundance of Lachnospiraceae and Prevotellaceae families and genus in feces, coupled with an increase in Muribaculaceae family and Bacteroides genus among infected kefir-treated mice. While kefir hinted at microbiota alterations reducing enterobacteria (Helicobacter), decrease IL-10, and increased IFN-γ, butyric acid on pre-infection, the beverage potentially facilitated the systemic translocation of pathogens, intensifying the infection's severity by altering the immune response. The use of kefir in the dosage of 10% w/v (109 CFU), for acute infections with Salmonella Typhimurium, may not be enough to combat the infection and worsen the prognosis, leaving the intestine less inflamed, favoring the replication and translocation of the pathogen. These findings underscore the importance of prudently evaluating the widespread use of probiotics and probiotic-rich beverages, especially during acute infections, given their potential association with adverse effects during these diseases.