Systematic interrogation of CRISPR antimicrobials in Klebsiella pneumoniae reveals nuclease-, guide- and strain-dependent features influencing antimicrobial activity.

CRISPR-Cas systems can be utilized as programmable-spectrum antimicrobials to combat bacterial infections. However, how CRISPR nucleases perform as antimicrobials across target sites and strains remains poorly explored. Here, we address this knowledge gap by systematically interrogating the use of CRISPR antimicrobials using multidrug-resistant and hypervirulent strains of Klebsiella pneumoniae as models. Comparing different Cas nucleases, DNA-targeting nucleases outperformed RNA-targeting nucleases based on the tested targets. Focusing on AsCas12a that exhibited robust targeting across different strains, we found that the elucidated modes of escape varied widely, restraining opportunities to enhance killing. We also encountered individual guide RNAs yielding different extents of clearance across strains, which were linked to an interplay between improper gRNA folding and strain-specific DNA repair and survival. To explore features that could improve targeting across strains, we performed a genome-wide screen in different K. pneumoniae strains that yielded guide design rules and trained an algorithm for predicting guide efficiency. Finally, we showed that Cas12a antimicrobials can be exploited to eliminate K. pneumoniae when encoded in phagemids delivered by T7-like phages. Altogether, our results highlight the importance of evaluating antimicrobial activity of CRISPR antimicrobials across relevant strains and define critical parameters for efficient CRISPR-based targeting.

Blood Flow Suppresses Vascular Anomalies in a Zebrafish Model of Cerebral Cavernous Malformations.

RATIONALE: Pathological biomechanical signaling induces vascular anomalies including cerebral cavernous malformations (CCM), which are caused by a clonal loss of CCM1/KRIT1 (Krev interaction trapped protein 1), CCM2/MGC4607, or CCM3/PDCD10. Why patients typically experience lesions only in lowly perfused venous capillaries of the cerebrovasculature is completely unknown. OBJECTIVE: In contrast, animal models with a complete loss of CCM proteins lack a functional heart and blood flow and exhibit vascular anomalies within major blood vessels as well. This finding raises the possibility that hemodynamics may play a role in the context of this vascular pathology. METHODS AND RESULTS: Here, we used a genetic approach to restore cardiac function and blood flow in a zebrafish model of CCM1. We find that blood flow prevents cardiovascular anomalies including a hyperplastic expansion within a large Ccm1-deficient vascular bed, the lateral dorsal aorta. CONCLUSIONS: This study identifies blood flow as an important physiological factor that is protective in the cause of this devastating vascular pathology.

Copper, iron, zinc and tannin concentrations throughout the digestive tract of tropical goats and sheep fed a high-fibre tannin-rich diet.

The dry season in tropical regions urges livestock to feed on nutritionally very poor diets. It has not been explored how tropical sheep-assumed grazers-and tropical goats-intermediate browsers-cope with a high-fibre tannin-rich diet. This study was designed to determine the effects of a high-fibre and tannin-rich diet on the flow of important microminerals iron (Fe), zinc (Zn) and copper (Cu) throughout the digestive tract of tropical sheep and goats. The feeding trial was set up with twelve adult male animals, six sheep with mean body weight (BW) of 30.3 ± 1.6 kg and six goats with mean BW of 26.4 ± 2.2 kg. The feed consisted of 36% leaves of Millettia ferruginea, 61% hay and 3% concentrate and was offered at 3% of BW (all on dry matter (DM) basis). The total faecal collection was carried out for 7 consecutive days. At the end of the experimental period, the animals were slaughtered to collect liver and digesta samples from the gastrointestinal tract. Feed, digesta and faecal samples underwent analysis of Fe, Zn, and Cu and total tannins (TT). Goats had significantly higher reticulum Cu concentrations expressed on DM as compared to sheep. Faecal Cu concentrations were higher for goats compared to sheep. Reticulum and colon digesta Zn levels were higher in goats than sheep. Abomasum and colon Fe levels were higher in sheep than goats when expressed on DM. These results suggest differences in feed intake, micromineral absorption, secretion and excretion between sheep and goats, pointing to a divergent mineral metabolism as an adaptation to the challenge of a dry season diet having very low nutritive value.

A Mutational Hotspot and Strong Selection Contribute to the Order of Mutations Selected for during Escherichia coli Adaptation to the Gut.

The relative role of drift versus selection underlying the evolution of bacterial species within the gut microbiota remains poorly understood. The large sizes of bacterial populations in this environment suggest that even adaptive mutations with weak effects, thought to be the most frequently occurring, could substantially contribute to a rapid pace of evolutionary change in the gut. We followed the emergence of intra-species diversity in a commensal Escherichia coli strain that previously acquired an adaptive mutation with strong effect during one week of colonization of the mouse gut. Following this first step, which consisted of inactivating a metabolic operon, one third of the subsequent adaptive mutations were found to have a selective effect as high as the first. Nevertheless, the order of the adaptive steps was strongly affected by a mutational hotspot with an exceptionally high mutation rate of 10-5. The pattern of polymorphism emerging in the populations evolving within different hosts was characterized by periodic selection, which reduced diversity, but also frequency-dependent selection, actively maintaining genetic diversity. Furthermore, the continuous emergence of similar phenotypes due to distinct mutations, known as clonal interference, was pervasive. Evolutionary change within the gut is therefore highly repeatable within and across hosts, with adaptive mutations of selection coefficients as strong as 12% accumulating without strong constraints on genetic background. In vivo competitive assays showed that one of the second steps (focA) exhibited positive epistasis with the first, while another (dcuB) exhibited negative epistasis. The data shows that strong effect adaptive mutations continuously recur in gut commensal bacterial species.

Recurrent Reverse Evolution Maintains Polymorphism after Strong Bottlenecks in Commensal Gut Bacteria.

The evolution of new strains within the gut ecosystem is poorly understood. We used a natural but controlled system to follow the emergence of intraspecies diversity of commensal Escherichia coli, during three rounds of adaptation to the mouse gut (∼1,300 generations). We previously showed that, in the first round, a strongly beneficial phenotype (loss-of-function for galactitol consumption; gat-negative) spread to >90% frequency in all colonized mice. Here, we show that this loss-of-function is repeatedly reversed when a gat-negative clone colonizes new mice. The regain of function occurs via compensatory mutation and reversion, the latter leaving no trace of past adaptation. We further show that loss-of-function adaptive mutants reevolve, after colonization with an evolved gat-positive clone. Thus, even under strong bottlenecks a regime of strong-mutation-strong-selection dominates adaptation. Coupling experiments and modeling, we establish that reverse evolution recurrently generates two coexisting phenotypes within the microbiota that can or not consume galactitol (gat-positive and gat-negative, respectively). Although the abundance of the dominant strain, the gat-negative, depends on the microbiota composition, gat-positive abundance is independent of the microbiota composition and can be precisely manipulated by supplementing the diet with galactitol. These results show that a specific diet is able to change the abundance of specific strains. Importantly, we find polymorphism for these phenotypes in indigenous Enterobacteria of mice and man. Our results demonstrate that natural selection can greatly overwhelm genetic drift at structuring the strain diversity of gut commensals and that competition for limiting resources may be a key mechanism for maintaining polymorphism in the gut.

The response of canine faecal microbiota to increased dietary protein is influenced by body condition.

BACKGROUND: High protein diets shift the faecal microbiota into a more unfavourable composition in obese humans. In lean dogs, higher protein consumption is accompanied with increased production of putrefactive fermentation products, whereas obese dogs have a different gut microbiota compared to lean dogs. Still, the impact of high dietary protein on gut microbiota in obese dogs remains unclear. The aim of this study was to investigate faecal microbial changes in lean and obese dogs in response to two different levels of dietary protein. Six healthy lean and six obese Beagles were fed a high protein diet (HP) and a low protein diet (LP) for 28 days each in a crossover design. Denaturing gradient gel electrophoresis and quantitative PCR were performed on faecal samples for microbial profiling. Plasma acylcarnitine and fermentation metabolites were measured. RESULTS: Dogs fed HP had higher concentrations of protein fermentation metabolites including faecal ammonia, isovalerate, isobutyrate, phenol, indole, serum indoxyl sulphate and plasma 3-OH isovalerylcarnitine compared to dogs fed LP, whereas no changes in faecal concentrations of acetate and butyrate were observed. The abundances of clostridial clusters IV and XIVa, covering the majority of butyrate-producing bacteria, and of the butyrate kinase gene, one of the terminal genes of the butyrate synthesis pathway were higher in dogs on HP compared to LP. Significant interactions between diet and body condition were found for the abundance of Firmicutes, Lactobacillus and clostridial cluster I. The similarity coefficient of faecal microbiota between the two diets was smaller in obese dogs than in lean dogs. CONCLUSIONS: High protein diet increased the abundance and activity of butyrate-producing bacteria in Beagles independent of the body condition. In addition, increasing dietary protein content had a greater overall impact on faecal microbiota in obese compared to lean dogs.

The first steps of adaptation of Escherichia coli to the gut are dominated by soft sweeps.

The accumulation of adaptive mutations is essential for survival in novel environments. However, in clonal populations with a high mutational supply, the power of natural selection is expected to be limited. This is due to clonal interference--the competition of clones carrying different beneficial mutations--which leads to the loss of many small effect mutations and fixation of large effect ones. If interference is abundant, then mechanisms for horizontal transfer of genes, which allow the immediate combination of beneficial alleles in a single background, are expected to evolve. However, the relevance of interference in natural complex environments, such as the gut, is poorly known. To address this issue, we have developed an experimental system which allows to uncover the nature of the adaptive process as Escherichia coli adapts to the mouse gut. This system shows the invasion of beneficial mutations in the bacterial populations and demonstrates the pervasiveness of clonal interference. The observed dynamics of change in frequency of beneficial mutations are consistent with soft sweeps, where different adaptive mutations with similar phenotypes, arise repeatedly on different haplotypes without reaching fixation. Despite the complexity of this ecosystem, the genetic basis of the adaptive mutations revealed a striking parallelism in independently evolving populations. This was mainly characterized by the insertion of transposable elements in both coding and regulatory regions of a few genes. Interestingly, in most populations we observed a complete phenotypic sweep without loss of genetic variation. The intense clonal interference during adaptation to the gut environment, here demonstrated, may be important for our understanding of the levels of strain diversity of E. coli inhabiting the human gut microbiota and of its recombination rate.

The metabolic response in fish to mildly elevated water temperature relates to species-dependent muscular concentrations of imidazole compounds and free amino acids.

Fish species show distinct differences in their muscular concentrations of imidazoles and free amino acids (FAA). This study was conducted to investigate whether metabolic response to mildly elevated water temperature (MEWT) relates to species-dependent muscular concentrations of imidazoles and FAA. Thirteen carp and 17 Nile tilapia, housed one per aquarium, were randomly assigned to either acclimation (25°C) or MEWT (30°C) for 14 days. Main muscular concentrations were histidine (HIS; P<0.001) in carp versus N-α-acetylhistidine (NAH; P<0.001) and taurine (TAU; P=0.001) in tilapia. Although the sum of imidazole (HIS+NAH) and TAU in muscle remained constant over species and temperatures (P>0.05), (NAH+HIS)/TAU ratio was markedly higher in carp versus tilapia, and decreased with MEWT only in carp (P<0.05). Many of the muscular FAA concentrations were higher in carp than in tilapia (P<0.05). Plasma acylcarnitine profile suggested a higher use of AA and fatty acids in carp metabolism (P<0.05). On the contrary, the concentration of 3-hydroxyisovalerylcarnitine, a sink of leucine catabolism, (P=0.009) pointed to avoidance of leucine use in tilapia metabolism. Despite a further increase of plasma longer-chain acylcarnitines in tilapia at MEWT (P=0.009), their corresponding beta-oxidation products (3-hydroxy-longer-chain acylcarnitines) remained constant. Together with higher plasma non-esterified fatty acids (NEFA) in carp (P=0.001), the latter shows that carp, being a fatter fish, more readily mobilises fat than tilapia at MEWT, which coincides with more intensive muscular mobilization of imidazoles. This study demonstrates that fish species differ in their metabolic response to MEWT, which is associated with species-dependent changes in muscle imidazole to taurine ratio.

Does canine inflammatory bowel disease influence gut microbial profile and host metabolism?

BACKGROUND: Inflammatory bowel disease (IBD) refers to a diverse group of chronic gastrointestinal diseases, and gut microbial dysbiosis has been proposed as a modulating factor in its pathogenesis. Several studies have investigated the gut microbial ecology of dogs with IBD but it is yet unclear if this microbial profile can alter the nutrient metabolism of the host. The aim of the present study was to characterize the faecal bacterial profile and functionality as well as to determine host metabolic changes in IBD dogs. Twenty-three dogs diagnosed with IBD and ten healthy control dogs were included. Dogs with IBD were given a clinical score using the canine chronic enteropathy clinical activity index (CCECAI). Faecal short-chain fatty acids (SCFA) and ammonia concentrations were measured and quantitative PCR was performed. The concentration of plasma amino acids, acylcarnitines, serum folate, cobalamin, and indoxyl sulfate was determined. RESULTS: No significant differences in the abundance of a selection of bacterial groups and fermentation metabolites were observed between the IBD and control groups. However, significant negative correlations were found between CCECAI and the faecal proportion of Lactobacillus as well as between CCECAI and total SCFA concentration. Serum folate and plasma citrulline were decreased and plasma valine was increased in IBD compared to control dogs. Increased plasma free carnitine and total acylcarnitines were observed in IBD compared with control dogs, whereas short-chain acylcarnitines (butyrylcarnitine + isobutyrylcarnitine and, methylmalonylcarnitine) to free carnitine ratios decreased. Dogs with IBD had a higher 3-hydroxyisovalerylcarnitine + isovalerylcarnitine to leucine ratio compared to control dogs. CONCLUSIONS: Canine IBD induced a wide range of changes in metabolic profile, especially for the plasma concentrations of short-chain acylcarnitines and amino acids, which could have evolved from tissue damage and alteration in host metabolism. In addition, dogs with more severe IBD were characterised by a decrease in faecal proportion of Lactobacillus.

Response to dietary tannin challenges in view of the browser/grazer dichotomy in an Ethiopian setting: Bonga sheep versus Kaffa goats.

It has been suggested that goats (typical browser) are better adapted to digest tannin-rich diets than sheep (typical grazer). To evaluate this, Bonga sheep and Kaffa goats were used in a 2 × 3 randomized crossover design with two species, three diets, and three periods (15-day adaptation + 7-day collection). The dietary treatments consisted of grass-based hay only (tannin-free diet = FT), a high-tannin diet (36% Albizia schimperiana (AS) + 9% Ficus elastica (FE) + 55% FT (HT)), and HT + polyethylene glycol 6000 (PEG). Animals were individually fed at 50 g dry matter (DM)/kg body weight (BW) and had free access to clean drinking water and mineralized salt licks. Nutrient intake, apparent nutrient digestibility, nutrient conversion ratios, and live weight changes were determined. Condensed tannin concentrations in AS and FE were 110 and 191 g/kg DM, respectively. Both sheep and goats ate 47% more of HT than FT, and dry matter intake further increased by 9% when PEG was added, with clear difference in effect size between goats and sheep (P < 0.001). The effects of the tannin-rich diet and PEG addition were similarly positive for DM digestibility between sheep and goats, but crude protein (CP) digestibility was higher in HT + PEG-fed goats than in sheep fed the same diet. However, PEG addition induced a larger improvement in growth performance and feed efficiency ratio in sheep than in goat (P < 0.001). The addition of PEG as a tannin binder improved digestion and performance in both species, but with the highest effect size in sheep.

Chestnut Wood Tannins in Broiler Diets: Pharmacokinetics, Serum Levels during Rearing, and Intestinal Absorption Pattern of Gallic Acid.

Studies on the bioavailability, serum levels, and absorption of hydrolyzable tannin compounds are lacking. In this study, we performed a pharmacokinetic trial, measured the serum levels of compounds in broilers that were reared with different feed added or not with tannins, and tested the digestibility of tannins throughout the intestinal tract. Only gallic acid and 4-O-methyl gallic acid were found in the serum. Moreover, gallic acid showed a 41.8% absolute oral bioavailability and a 72.3% relative bioavailability of gallic acid from chestnut extract compared to the standard. The rapid metabolization caused alternating serum levels during the day and night. These patterns were not affected by the feed type or the previous addition of tannins in the feed. The absorption and metabolization in the intestines occurred gradually throughout the intestinal tract. The latter was true for gallic acid as well as ellagic acid, which was not found in the serum. We can conclude that components from chestnut tannins are absorbed throughout all components of the intestinal tract and are eliminated quickly with little interaction from the feed and previous addition of tannins. Moreover, ellagic acid seems to be absorbed but would remain accumulated in the intestinal tissue or be metabolized by the microbiome.

Horizontal gene transfer among host-associated microbes.

Horizontal gene transfer is an important evolutionary force, facilitating bacterial diversity. It is thought to be pervasive in host-associated microbiomes, where bacterial densities are high and mobile elements are frequent. These genetic exchanges are also key for the rapid dissemination of antibiotic resistance. Here, we review recent studies that have greatly extended our knowledge of the mechanisms underlying horizontal gene transfer, the ecological complexities of a network of interactions involving bacteria and their mobile elements, and the effect of host physiology on the rates of genetic exchanges. Furthermore, we discuss other, fundamental challenges in detecting and quantifying genetic exchanges in vivo, and how studies have contributed to start overcoming these challenges. We highlight the importance of integrating novel computational approaches and theoretical models with experimental methods where multiple strains and transfer elements are studied, both in vivo and in controlled conditions that mimic the intricacies of host-associated environments.

Phages against Noncapsulated Klebsiella pneumoniae: Broader Host range, Slower Resistance.

Klebsiella pneumoniae (Kp), a human gut colonizer and opportunistic pathogen, is a major contributor to the global burden of antimicrobial resistance. Virulent bacteriophages represent promising agents for decolonization and therapy. However, the majority of anti-Kp phages that have been isolated thus far are highly specific to unique capsular types (anti-K phages), which is a major limitation to phage therapy prospects due to the highly polymorphic capsule of Kp. Here, we report on an original anti-Kp phage isolation strategy, using capsule-deficient Kp mutants as hosts (anti-Kd phages). We show that anti-Kd phages have a broad host range, as the majority are able to infect noncapsulated mutants of multiple genetic sublineages and O-types. Additionally, anti-Kd phages induce a lower rate of resistance emergence in vitro and provide increased killing efficiency when in combination with anti-K phages. In vivo, anti-Kd phages are able to replicate in mouse guts colonized with a capsulated Kp strain, suggesting the presence of noncapsulated Kp subpopulations. The original strategy proposed here represents a promising avenue that circumvents the Kp capsule host restriction barrier, offering promise for therapeutic development. IMPORTANCE Klebsiella pneumoniae (Kp) is an ecologically generalist bacterium as well as an opportunistic pathogen that is responsible for hospital-acquired infections and a major contributor to the global burden of antimicrobial resistance. In the last decades, limited advances have been made in the use of virulent phages as alternatives or complements to antibiotics that are used to treat Kp infections. This work demonstrates the potential value of an anti-Klebsiella phage isolation strategy that addresses the issue of the narrow host range of anti-K phages. Anti-Kd phages may be active in infection sites in which capsule expression is intermittent or repressed or in combination with anti-K phages, which often induce the loss of capsule in escape mutants.

Chestnut tannins in broiler diets: Affecting intestinal development in different feeding phases.

It is known that high doses of various tannins could impair broiler growth, and this seems to be linked to a lowered protein availability. However, effects on protein digestion under the influence of hydrolysable tannins were minimal in previous research and literature. Other possible proposed reasons to explain reduced growth are scarce. In this experiment we studied the effect of hydrolysable tannins on body allometry by using different feeding schemes throughout the rearing period. In total 112 individually reared male Ross 308 broilers received a 3-phase basal diet with chestnut wood extract (+: 2,000 mg/kg) or not (-: 0 mg/kg) (Tanno-SAN®, Sanluc International NV, Belgium). This resulted in 2 groups during the starter period (S+, S-), 4 groups in the grower period (G++, G+-, G-+, G-) and 8 groups in the finisher period (F+++, F++-, F+-+, F+--, F-++, F-+-, F--+, F--). Similar to previous studies, growth reduction was also observed in this study. Effects were the largest in broilers that were given the tannins during the grower phase. At the end of each phase 8 broilers per group were euthanized and sampled. Liver, pancreas, pectoralis muscle, intestinal weights and intestinal length were recorded. The largest effects were seen on the intestine. Broilers that received tannins during the grower phase, had longer intestines at the end of the finisher period. Furthermore, histological differences between treatment groups were observed at the end of the grower period. Addition of tannins in the grower phase (G-+, G++) resulted in longer villi, whereas addition of tannins in the starter (G+-, G++) caused deeper crypts at the end of the grower phase, with the group (G-+) having the highest villi-to-crypt ratio. These results tentatively prove that tannins influence intestinal growth, both macroscopically as well as histologically. We hypothesize that the observed growth reduction with tannins could be the result of a changed energy and nutrient partitioning, i.e., more nutrients are directed to intestinal growth than for muscle growth.

The gut environment regulates bacterial gene expression which modulates susceptibility to bacteriophage infection.

Abundance and diversity of bacteria and their viral predators, bacteriophages (phages), in the digestive tract are associated with human health. Particularly intriguing is the long-term coexistence of these two antagonistic populations. We performed genome-wide RNA sequencing on a human enteroaggregative Escherichia coli isolate to identify genes differentially expressed between in vitro conditions and in murine intestines. We experimentally demonstrated that four of these differentially expressed genes modified the interactions between E. coli and three virulent phages by either increasing or decreasing its susceptibility/resistance pattern and also by interfering with biofilm formation. Therefore, the regulation of bacterial genes expression during the colonization of the digestive tract influences the coexistence of phages and bacteria, highlighting the intricacy of tripartite relationships between phages, bacteria, and the animal host in intestinal homeostasis.

Chestnut tannins in broiler diets: performance, nutrient digestibility, and meat quality.

In 2 broiler trials, the effects of chestnut tannins on performance and meat quality (trial 1), and digestion (trial 2) were evaluated. In both trials, Ross 308 broilers received one of 2 basal diets: one basal diet contained corn and soy as main feed ingredients, while the challenge basal diet contained wheat, palm oil, and rapeseed meal. The composition of the basal diets was chosen to assess the interaction between chestnut tannins and diet composition. To both basal diets, chestnut tannins were added at 3 doses: 0 mg/kg (T-), 500 mg/kg (T+), or 2,000 mg/kg (T++), resulting in a total of 6 treatments. In trial 1, both basal diets containing 2,000 mg/kg chestnut tannins lowered broiler performance in grower and finisher phases. A tannin dose of 500 mg/kg had no effect on performance in either basal diet. Corn-based diets resulted in lower meat pH compared to wheat diets. Further, addition of chestnut tannins resulted in increased meat pH, and caused proportionally a lower meat drip loss and shear force for both basal diets. During the digestibility study (trial 2), blood was also collected. None of the treatments affected digestibility or blood parameters (glucose, non-esterified fatty acids, and triacylglycerols). Malondialdehyde (MDA) was measured in plasma to assess antioxidative properties of chestnut tannins. In wheat diets, chestnut tannins significantly lowered plasma MDA demonstrating its antioxidative nature. Regarding gut health, crypt depth decreased proportionally with the dosage of chestnut tannins in both basal diets with significantly shallower crypts for the wheat diets compared to the corn diets. Relative intestinal growth was stimulated in the wheat diets proportionally to the tannin dose based on the larger relative gut length. In conclusion, chestnut tannins did not influence digestive metabolism, yet they lowered performance at higher doses regardless of feed ingredients used in the diet. Tannins positively affected meat quality and when added to wheat diets, intestinal growth was stimulated and the antioxidative status of the broilers improved.

Prophylactic Administration of a Bacteriophage Cocktail Is Safe and Effective in Reducing Salmonella enterica Serovar Typhimurium Burden in Vivo.

Nontyphoidal Salmonella bacteria are the causative agent of salmonellosis, which accounts for the majority of foodborne illness of bacterial etiology in humans. Here, we demonstrate the safety and efficacy of the prophylactic administration of a bacteriophage preparation termed FOP (foodborne outbreak pill), which contains lytic phages targeting Salmonella (SalmoFresh phage cocktail), Shiga toxin-producing Escherichia coli (STEC), and Listeria monocytogenes, for lowering Salmonella burdens in OMM12 gnotobiotic mice. Prophylactic administration of FOP significantly reduced the levels of Salmonella in feces and in intestinal sections compared to the levels in controls. Moreover, the overall symptoms of the disease were also considerably lessened. Dose-dependent administration of FOP showed that phage amplification reached similarly high levels in less than 48 h independent of dose. In addition, 16S rRNA gene analysis showed that FOP did not alter the intestinal microbiota of healthy OMM12 mice and reduced microbiota perturbations induced by Salmonella. FOP maintained its full potency against Salmonella in comparison to that of SalmoFresh, its Salmonella-targeting component phages alone. Altogether, the data support that preventive administration of FOP may offer a safe and effective approach for reducing the risk of foodborne infections caused by Salmonella and, potentially, other foodborne bacteria (namely, STEC and L. monocytogenes) targeted by the FOP preparation. IMPORTANCE Foodborne bacterial infections cause worldwide economic loss. During an epidemic, the use of antibiotics to slow down the spread of the disease is not recommended because of their side effects on the resident microbiota and the selection of antibiotic-resistant bacteria. Here, we investigated the potential for the prophylactic administration of bacteriophages (viruses infecting bacteria) to reduce the burden of Salmonella in vivo using mice colonized by a synthetic microbiota. We found that the repeated administration of bacteriophages was safe and efficient in lowering the Salmonella burden. Perturbations of the microbiota by the Salmonella infection were also reduced when mice received bacteriophages. Altogether, these data support the use of bacteriophages as a prophylactic intervention to lower the spread of foodborne epidemics.

Forecasting COVID-19 Severity by Intelligent Optical Fingerprinting of Blood Samples.

Forecasting COVID-19 disease severity is key to supporting clinical decision making and assisting resource allocation, particularly in intensive care units (ICUs). Here, we investigated the utility of time- and frequency-related features of the backscattered signal of serum patient samples to predict COVID-19 disease severity immediately after diagnosis. ICU admission was the primary outcome used to define disease severity. We developed a stacking ensemble machine learning model including the backscattered signal features (optical fingerprint), patient comorbidities, and age (AUROC = 0.80), which significantly outperformed the predictive value of clinical and laboratory variables available at hospital admission (AUROC = 0.71). The information derived from patient optical fingerprints was not strongly correlated with any clinical/laboratory variable, suggesting that optical fingerprinting brings unique information for COVID-19 severity risk assessment. Optical fingerprinting is a label-free, real-time, and low-cost technology that can be easily integrated as a front-line tool to facilitate the triage and clinical management of COVID-19 patients.

The Spatial Heterogeneity of the Gut Limits Predation and Fosters Coexistence of Bacteria and Bacteriophages.

The ecological dynamics underlying the coexistence between antagonistic populations of bacteria and their viruses, bacteriophages (phages), in the mammalian gut microbiota remain poorly understood. We challenged a murine synthetic bacterial community with phages to study the factors allowing phages-bacteria coexistence. Coexistence was not dependent on the development of phage-resistant clones nor on the ability of phages to extend their host range. Instead, our data suggest that phage-inaccessible sites in the mucosa serve as a spatial refuge for bacteria. From there, bacteria disseminate in the gut lumen where they are predated by luminal phages fostering the presence of intestinal phage populations. The heterogeneous biogeography of microbes contributes to the long-term coexistence of phages with phage-susceptible bacteria. This observation could explain the persistence of intestinal phages in humans as well as the low efficiency of oral phage therapy against enteric pathogens in animal models and clinical trials.

"I will survive": A tale of bacteriophage-bacteria coevolution in the gut.

Viruses that infect bacteria, or bacteriophages, are among the most abundant entities in the gut microbiome. However, their role and the mechanisms by which they infect bacteria in the intestinal tract remain poorly understood. We recently reported that intestinal bacteria are an evolutionary force, driving the expansion of the bacteriophage host range by boosting the genetic variability of these viruses. Here, we expand these observations by studying antagonistic bacteriophage-bacteria coevolution dynamics and revealing that bacterial genetic variability is also increased under the pressure of bacteriophage predation. We propose a model showing how the expansion of bacteriophage-bacteria infection networks is relative to the opportunities for coevolution encountered in the intestinal tract. Our data suggest that predator-prey dynamics are perpetuated and differentiated in parallel, to generate and maintain intestinal microbial diversity and equilibrium.