Raw milk cheeses harbor complex microbial communities. Some of these microorganisms are technologically essential, but undesirable microorganisms can also be present. While most of the microbial dynamics and cross-talking studies involving interaction between food-derived bacteria have been carried out on agar plates in laboratory-controlled conditions, the present study evaluated the modulation of the resident microbiota and the changes of metabolite production directly in ripening raw milk cheese inoculated with Listeria innocua strains. Using a proxy of the pathogenic Listeria monocytogenes, we aimed to establish the key microbiota players and chemical signals that characterize Latteria raw milk cheese over 60 days of ripening time. The microbiota of both the control and Listeria-inoculated cheeses was analyzed using 16S rRNA targeted amplicon sequencing, while direct analysis in real time mass spectrometry (DART-HRMS) was applied to investigate the differences in the metabolic profiles of the cheeses. The diversity analysis showed the same microbial diversity trend in both the control cheese and the inoculated cheese, while the taxonomic analysis highlighted the most representative genera of bacteria in both the control and inoculated cheese: Lactobacillus and Streptococcus. On the other hand, the metabolic fingerprints revealed that the complex interactions between resident microbiota and L. innocua were governed by continuously changing chemical signals. Changes in the amounts of small organic acids, hydroxyl fatty acids, and antimicrobial compounds, including pyroglutamic acid, hydroxy-isocaproic acid, malic acid, phenyllactic acid, and lactic acid, were observed over time in the L. innocua-inoculated cheese. In cheese that was inoculated with L. innocua, Streptococcus was significantly correlated with the volatile compounds carboxylbenzaldheyde and cyclohexanecarboxylic acid, while Lactobacillus was positively correlated with some volatile and flavor compounds (cyclohexanecarboxylic acid, pyroxidal acid, aminobenzoic acid, and vanillic acid). Therefore, we determined the metabolic markers that characterize a raw milk cheese inoculated with L. innocua, the changes in these markers with the ripening time, and the positive correlation of flavor and volatile compounds with the resident microbiota. This multi-omics approach could suggest innovative food safety strategies based on the enhanced management of undesirable microorganisms by means of strain selection in raw matrices and the addition of specific antimicrobial metabolites to prevent the growth of undesirable microorganisms.

Giardia duodenalis (Giardia) is a worldwide cause of acute diarrheal disease both in humans and animals. The primary aim of this study was to investigate possible variations in gut microbiota in a population of asymptomatic dogs (n = 31), naturally infected or not by Giardia. Gut microbiota and the hematological, biochemical, and fecal parameters related to intestinal function were investigated. Giardia infection was associated with a significant shift of beta diversity, showing a relevant reduction of Gammaproteobacteria and an increase of Fusobacteria in male-positive dogs if compared with negatives. A significant imbalance of different bacterial taxa, with particular reference to the Erysipelotrichales, Lactobacillales, Clostridiales, and Burkholderiales orders, was observed, with the first two being higher in Giardia-positive dogs. Giardia-positive males displayed significantly higher values of cCRP than negative males as well as positive females, supporting the presence of a pro-inflammatory state. Taken together, these results indicate that the presence of Giardia does not substantially modify the microbial ecology of the intestine nor the hematological markers of disease. Thus treatments against Giardia should be considered with caution in asymptomatic subjects.

The western honey bee (Apis mellifera) represents a key pollinator for both crops and wild plants, and its global decline raises serious concerns for ecosystem stability and agricultural productivity. Several biotic and abiotic factors are responsible for colony losses, including alterations in the bee microbiota, which is essential for host metabolism, development, and immune responses. In this study, we employed both molecular protocols and metagenomic approaches based on Next-Generation Sequencing (NGS) to characterize the microbial composition and identify commensal, symbiotic, and pathogenic microorganisms, both known and emerging, associated with A. mellifera colonies from 20 apiaries across the Italian territory. Molecular screening revealed Vairimorpha ceranae, Lotmaria passim, Crithidia mellificae and several viruses, including Sacbrood virus (SBV), Black Queen Cell virus (BQCV), Deformed Wing virus (DWV), Chronic Bee Paralysis virus (CBPV) and Acute Bee Paralysis virus (ABPV). 16S rRNA gene sequencing highlighted a bacterial community mainly composed of the Lactobacillus, Gilliamella, and Snodgrassella genera. Virome analysis detected members belonging to the families Dicistroviridae and Iflaviridae, as well as previously unreported viruses in Italy, such as Apis rhabdovirus (ARV-1, ARV-2), Bee Macula-like virus (BeeMLV), and Lake Sinai virus (LSV). This research expands current knowledge of the A. mellifera metagenome, offering valuable insights for epidemiological surveillance and diagnostic assay development.

Honey bees are valuable bioindicators of environmental contamination due to their wide foraging range and exposure to diverse biotic and abiotic stressors. This study evaluates the use of honey bees as bioindicators for detecting per- and polyfluoroalkyl substances (PFASs) in areas with varying levels of exposure risk (high-, intermediate-, and low-), as well as their potential effects on gut-microbial communities. Analytical chemistry to detect PFASs and pesticide residues were combined with molecular and metagenomic analyses to identify common honey bee pathogens and characterize gut-microbial communities. Short-chain PFASs were predominated over long-chain compounds, particularly in high-risk areas, while pesticide residues occurred in only a few apiaries. Molecular analyses revealed several widespread pathogens, while metagenomics showed a stable gut microbiome mainly composed of the genera Lactobacillus, Bartonella, Bombilactobacillus, Snodgrassella, Gilliamella, Bifidobacterium, and Gluconacetobacter. However, microbial-community composition differed significantly between apiaries with high- versus low-risk PFAS exposure. These results support honey bees as effective bioindicators for environmental PFAS monitoring and suggest that PFAS exposure may be associated with alterations in their gut-microbial communities. Further research is needed to elucidate the impact of PFASs on honey bee toxicity, gut-microbiota composition, and overall health.