Genetic Context of optrA and poxtA in Florfenicol-Resistant Enterococci Isolated from Flowing Surface Water in Switzerland.

Linezolid is an important last-resort antibiotic for the treatment of multidrug-resistant enterococci. The aim of this study was to further characterize the genetic context of optrA and poxtA in 10 florfenicol-resistant enterococci isolated from flowing surface water. In most genomes, optrA and poxtA were embedded in transposition units integrated into plasmids or into the chromosomal radC. For the first time, a chromosomally integrated optrA in an Enterococcus raffinosus isolate is described.

CXCR4 and CXCR7 Inhibition Ameliorates the Formation of Platelet-Neutrophil Complexes and Neutrophil Extracellular Traps through Adora2b Signaling.

Peritonitis and peritonitis-associated sepsis are characterized by an increased formation of platelet-neutrophil complexes (PNCs), which contribute to an excessive migration of polymorphonuclear neutrophils (PMN) into the inflamed tissue. An important neutrophilic mechanism to capture and kill invading pathogens is the formation of neutrophil extracellular traps (NETs). Formation of PNCs and NETs are essential to eliminate pathogens, but also lead to aggravated tissue damage. The chemokine receptors CXCR4 and CXCR7 on platelets and PMNs have been shown to play a pivotal role in inflammation. Thereby, CXCR4 and CXCR7 were linked with functional adenosine A2B receptor (Adora2b) signaling. We evaluated the effects of selective CXCR4 and CXCR7 inhibition on PNCs and NETs in zymosan- and fecal-induced sepsis. We determined the formation of PNCs in the blood and, in addition, their infiltration into various organs in wild-type and Adora2b-/- mice by flow cytometry and histological methods. Further, we evaluated NET formation in both mouse lines and the impact of Adora2b signaling on it. We hypothesized that the protective effects of CXCR4 and CXCR7 antagonism on PNC and NET formation are linked with Adora2b signaling. We observed an elevated CXCR4 and CXCR7 expression in circulating platelets and PMNs during acute inflammation. Specific CXCR4 and CXCR7 inhibition reduced PNC formation in the blood, respectively, in the peritoneal, lung, and liver tissue in wild-type mice, while no protective anti-inflammatory effects were observed in Adora2b-/- animals. In vitro, CXCR4 and CXCR7 antagonism dampened PNC and NET formation with human platelets and PMNs, confirming our in vivo data. In conclusion, our study reveals new protective aspects of the pharmacological modulation of CXCR4 and CXCR7 on PNC and NET formation during acute inflammation.

East and West African milk products are reservoirs for human and livestock-associated Staphylococcus aureus.

Staphylococcus aureus frequently isolated from milk products in sub-Saharan Africa (SSA) is a major pathogen responsible for food intoxication, human and animal diseases. SSA hospital-derived strains are well studied but data on the population structure of foodborne S. aureus required to identify possible staphylococcal food poisoning sources is lacking. Therefore, the aim was to assess the population genetic structure, virulence and antibiotic resistance genes associated with milk-derived S. aureus isolates from Côte d'Ivoire, Kenya and Somalia through spa-typing, MLST, and DNA microarray analysis. Seventy milk S. aureus isolates from the three countries were assigned to 27 spa (7 new) and 23 (12 new) MLST sequence types. Milk-associated S. aureus of the three countries is genetically diverse comprising human and livestock-associated clonal complexes (CCs) predominated by the CC5 (n = 10) and CC30 (n = 9) isolates. Panton-Valentine leukocidin, toxic shock syndrome toxin and enterotoxin encoding genes were predominantly observed among human-associated CCs. Penicillin, fosfomycin and tetracycline, but not methicillin resistance genes were frequently detected. Our findings indicate that milk-associated S. aureus in SSA originates from human and animal sources alike highlighting the need for an overarching One Health approach to reduce S. aureus disease burdens through improving production processes, animal care and hygienic measures.

Phylogenetic, epidemiological and functional analyses of the Streptococcus bovis/Streptococcus equinus complex through an overarching MLST scheme.

BACKGROUND: The Streptococcus bovis/Streptococcus equinus complex (SBSEC) comprises seven (sub)species classified as human and animal commensals, emerging opportunistic pathogens and food fermentative organisms. Changing taxonomy, shared habitats, natural competence and evidence for horizontal gene transfer pose difficulties for determining their phylogeny, epidemiology and virulence mechanisms. Thus, novel phylogenetic and functional classifications are required. An SBSEC overarching multi locus sequence type (MLST) scheme targeting 10 housekeeping genes was developed, validated and combined with host-related properties of adhesion to extracellular matrix proteins (ECM), activation of the immune responses via NF-KB and survival in simulated gastric juice (SGJ). RESULTS: Commensal and pathogenic SBSEC strains (n = 74) of human, animal and food origin from Europe, Asia, America and Africa were used in the MLST scheme yielding 66 sequence types and 10 clonal complexes differentiated into distinct habitat-associated and mixed lineages. Adhesion to ECMs collagen I and mucin type II was a common characteristic (23 % of strains) followed by adhesion to fibronectin and fibrinogen (19.7 %). High adhesion abilities were found for East African dairy and human blood isolate branches whereas commensal fecal SBSEC displayed low adhesion. NF-KB activation was observed for a limited number of dairy and blood isolates suggesting the potential of some pathogenic strains for reduced immune activation. Strains from dairy MLST clades displayed the highest relative survival to SGJ independently of dairy adaptation markers lacS/lacZ. CONCLUSION: Combining phylogenetic and functional analyses via SBSEC MLST enabled the clear delineation of strain clades to unravel the complexity of this bacterial group. High adhesion values shared between certain dairy and blood strains as well as the behavior of NF-KB activation are concerning for specific lineages. They highlighted the health risk among shared lineages and establish the basis to elucidate (zoonotic-) transmission, host specificity, virulence mechanisms and enhanced risk assessment as pathobionts in an overarching One Health approach.

Effect of Bifidobacterium thermophilum RBL67 and fructo-oligosaccharides on the gut microbiota in Göttingen minipigs.

Modulating the gut microbiota via dietary interventions is a common strategy to enhance the natural defence mechanisms of the host. Several in vitro studies have highlighted the probiotic potential of Bifidobacterium thermophilum RBL67 (RBL67) selected for its anti-Salmonella effects. The present study aimed to investigate the impact of RBL67 alone and combined with fructo-oligosaccharides (FOS) on the gut microbiota of Göttingen minipigs. Minipigs were fed a basal diet supplemented with 8 g/d probiotic powder (1×109 CFU/g in skim milk matrix) (probiotic diet (PRO)), 8 g/d probiotic powder plus 8 g/d FOS (synbiotic diet (SYN)) or 8 g/d skim milk powder (control), following a cross-sectional study design. Faecal and caecal microbiota compositions were analysed with pyrosequencing of 16S rRNA genes and quantitative PCR. Metabolic activity in the caecum and colon was measured by HPLC. 16S rRNA gene amplicon sequencing revealed that minipig faeces show close similarity to pig microbiota. During the treatments and at the time of killing of animals, RBL67 was consistently detected in faeces, caecum and colon at numbers of 105-106 16S rRNA copies/g content after feeding PRO and SYN diets. At the time of killing of animals, significantly higher Bifidobacterium numbers in the caecum and colon of SYN-fed minipigs were measured compared with PRO. Our data indicate that the Göttingen minipig may be a suitable model for gut microbiota research in pigs. Data from this first in vivo study of RBL67 colonisation suggest that the combination with FOS may represent a valuable symbiotic strategy to increase probiotic bacteria levels and survival in gastrointestinal tracts for feed and food applications.

Comparative genome analysis of Streptococcus infantarius subsp. infantarius CJ18, an African fermented camel milk isolate with adaptations to dairy environment.

BACKGROUND: Streptococcus infantarius subsp. infantarius (Sii) belongs to the Streptococcus bovis/Streptococcus equinus complex associated with several human and animal infections. Sii is a predominant bacterium in spontaneously fermented milk products in Africa. The genome sequence of Sii strain CJ18 was compared with that of other Streptococcus species to identify dairy adaptations including genome decay such as in Streptococcus thermophilus, traits for its competitiveness in spontaneous milk fermentation and to assess potential health risks for consumers. RESULTS: The genome of Sii CJ18 harbors several unique regions in comparison to Sii ATCC BAA-102T, among others an enlarged exo- and capsular polysaccharide operon; Streptococcus thermophilus-associated genes; a region containing metabolic and hypothetical genes mostly unique to CJ18 and the dairy isolate Streptococcus gallolyticus subsp. macedonicus; and a second oligopeptide transport operon. Dairy adaptations in CJ18 are reflected by a high percentage of pseudogenes (4.9%) representing genome decay which includes the inactivation of the lactose phosphotransferase system (lacIIABC) by multiple transposases integration. The presence of lacS and lacZ genes is the major dairy adaptation affecting lactose metabolism pathways also due to the disruption of lacIIABC.We constructed mutant strains of lacS, lacZ and lacIIABC and analyzed the resulting strains of CJ18 to confirm the redirection of lactose metabolism via LacS and LacZ.Natural competence genes are conserved in both Sii strains, but CJ18 contains a lower number of CRISPR spacers which indicates a reduced defense capability against alien DNA. No classical streptococcal virulence factors were detected in both Sii strains apart from those involved in adhesion which should be considered niche factors. Sii-specific virulence factors are not described. Several Sii-specific regions encoding uncharacterized proteins provide new leads for virulence analyses and investigation of the unclear association of dairy and clinical Sii with human diseases. CONCLUSIONS: The genome of the African dairy isolate Sii CJ18 clearly differs from the human isolate ATCC BAA-102T. CJ18 possesses a high natural competence predisposition likely explaining the enlarged genome. Metabolic adaptations to the dairy environment are evident and especially lactose uptake corresponds to S. thermophilus. Genome decay is not as advanced as in S. thermophilus (10-19%) possibly due to a shorter history in dairy fermentations.

Dissemination of ESBL-producing E. coli ST131 through wastewater and environmental water in Switzerland.

The E. coli lineage ST131 is a major cause of multidrug-resistant urinary tract and bloodstream infections worldwide. Recently emerged ST131 sublineages spread globally within few years, but their dissemination routes are incompletely understood. In this study, we investigate the potential role of wastewater and surface water in the spread of extended-spectrum ß-lactamase (ESBL)-producing ST131. Streams, lakes, and two wastewater treatment plants (WWTPs) in the canton of Zug, Switzerland, were consecutively sampled over 1.5 years. ST131 was detected in 38% of the samples taken downstream (1-5 km) of WWTP discharge sites, but usually absent in water bodies distant from urban areas or WWTP discharge. Specific strains were repeatedly isolated (≤5 pairwise cgSNP distance) from wastewater or river sites downstream of effluent discharge, indicating their repeated entry or persistence in WWTPs in large concentrations. Genetic characterization of the ESBL-producing water isolates revealed a predominance of clades A and C1 and an emerging ciprofloxacin-resistant sublineage with mutations in quinolone resistance determining regions (QRDR) within clade A. Multiple isolates belonged to internationally circulating sublineages, including C1-M27 and papGII + sublineages with chromosomally encoded ESBLs. This study demonstrates that the clinically relevant E. coli lineage ST131 pollutes river ecosystems, representing a significant challenge to public health and to technologies to minimize their entry into the water environment.

Complete genome sequence of the African dairy isolate Streptococcus infantarius subsp. infantarius strain CJ18.

Streptococcus infantarius subsp. infantarius, a member of the Streptococcus bovis/Streptococcus equinus complex, is highly prevalent in artisanal dairy fermentations in Africa. Here the complete genome sequence of the dairy-adapted S. infantarius subsp. infantarius CJ18 strain--a strain predominant in traditionally fermented camel milk (suusac) from Kenya--is presented.

Novel Streptococcus infantarius subsp. infantarius variants harboring lactose metabolism genes homologous to Streptococcus thermophilus.

Streptococcus infantarius subsp. infantarius belongs to the Streptococcus bovis/Streptococcus equinus complex (SBSEC) commonly associated with human and animal infections. We elucidated the lactose metabolism of S. infantarius subsp. infantarius predominant in African fermented milk products. S. infantarius subsp. infantarius isolates (n = 192) were identified in 88% of spontaneously fermented camel milk suusac samples (n = 24) from Kenya and Somalia at log10 8.2-8.5 CFU mL⁻¹. African S. infantarius isolates excreted stoichiometric amounts of galactose when grown on lactose, exhibiting a metabolism similar to Streptococcus thermophilus and distinct from their type strain. African S. infantarius subsp. infantarius CJ18 harbors a regular gal operon with 99.7-100% sequence identity to S. infantarius subsp. infantarius ATCC BAA-102(T) and a gal-lac operon with 91.7-97.6% sequence identity to S. thermophilus, absent in all sequenced SBSEC strains analyzed. The expression and functionality of lacZ was demonstrated in a ß-galactosidase assay. The gal-lac operon was identified in 100% of investigated S. infantarius isolates (n = 46) from suusac samples and confirmed in Malian fermented cow milk isolates. The African S. infantarius variant potentially evolved through horizontal gene transfer of an S. thermophilus-homologous lactose pathway. Safety assessments are needed to identify any putative health risks of this novel S. infantarius variant.

Comparative genomics of dairy-associated Staphylococcus aureus from selected sub-Saharan African regions reveals milk as reservoir for human-and animal-derived strains and identifies a putative animal-related clade with presumptive novel siderophore.

Staphylococcus aureus infection is considered to be a neglected tropical disease with huge impact on human and animal health alike. Dairy production in sub-Saharan Africa (SSA) relies heavily on various animals such as cows, goats, and camels, depending on the region. S. aureus causes mastitis and exhibits high prevalence in raw milk. The population structure including genotypic and phenotypic traits of dairy S. aureus in relation to animal and human isolates is, however, unknown for SSA. In this work, 20 S. aureus dairy isolates from East and West Africa were selected for comparative genomics and phenotypic analysis. Comparing their population structure revealed a large diversity of different origins suggesting milk to be a reservoir for human and animal strains alike. Furthermore, a novel putative siderophore was detected in multiple strains in a distinct animal-clade with strains of global origin. This putative siderophore shares a high genetic identity with that from Streptococcus equi suggesting possible horizontal gene transfer. These findings combined with the virulence genes harbored by these dairy-derived strains such as pvl, human evasion factor scn, various enterotoxin, leucocidin and antibiotic resistance genes, stresses the need for an integrative One Health approach to tackle the problem of S. aureus infections in animals and humans in sub-Saharan Africa.

Complete Genome Sequence of Hafnia paralvei Isolate AVS0177, Harboring mcr-9 on a Plasmid.

Here, we report the complete genome sequence of a Hafnia paralvei strain isolated from a lake in Switzerland in 2020. The genome consists of a 4.7-Mbp chromosome, a large plasmid (213 kb) harboring mcr-9, and a small plasmid (6 kb).

Complete Genome Sequence of Colistin-Resistant, mcr-10-Harboring, Enterobacter cloacae Isolate AVS0889, Recovered from River Water in Switzerland.

Here, we report the complete genome sequence of colistin-resistant Enterobacter cloacae sequence type 1 (ST1) isolate AVS0889, which was recovered from a river in Switzerland in 2021. The genome consists of a 4.95-Mbp chromosome and five plasmids, including a large plasmid (90.8 kb) harboring a disrupted mcr-10 gene.

Identification of Valerate as Carrying Capacity Modulator by Analyzing Lactiplantibacillus plantarum Colonization of Colonic Microbiota in vitro.

Humans ingest many microorganisms, which may colonize and interact with the resident gut microbiota. However, extensive knowledge about host-independent microbe-microbe interactions is lacking. Here, we investigated such colonization process using a derivative of the model probiotic Lactiplantibacillus plantarum WCFS1 into continuously cultivated gut microbiota in the intestinal PolyFermS fermentation model inoculated with five independently immobilized human adult fecal microbiota. L. plantarum successfully colonized and organized itself spatially in the planktonic, that is, the reactor effluent, and sessile, that is, reactor biofilm, fractions of distinct human adult microbiota. The microbiota carrying capacity for L. plantarum was independent of L. plantarum introduction dose and second supplementation. Adult microbiota (n = 3) dominated by Prevotella and Ruminoccocus exhibited a higher carrying capacity than microbiota (n = 2) dominated by Bacteroides with 105 and 103 CFU/ml of L. plantarum, respectively. Cultivation of human adult microbiota over 3 months resulted in decreased carrying capacity and correlated positively with richness and evenness, suggesting enhanced resistance toward colonizers. Our analyses ultimately allowed us to identify the fermentation metabolite valerate as a modulator to increase the carrying capacity in a microbiota-independent manner. In conclusion, by uncoupling microbe-microbe interactions from host factors, we showed that L. plantarum colonizes the in vitro colonic community in a microbiota-dependent manner. We were further able to demonstrate that L. plantarum colonization levels were not susceptible to the introduction parameters dose and repeated administration but to microbiota features. Such knowledge is relevant in gaining a deeper ecological understanding of colonizer-microbiota interactions and developing robust probiotic strategies.

In Vitro Gut Modeling as a Tool for Adaptive Evolutionary Engineering of Lactiplantibacillus plantarum.

Research and marketing of probiotics demand holistic strain improvement considering both the biotic and abiotic gut environment. Here, we aim to establish the continuous in vitro colonic fermentation model PolyFermS as a tool for adaptive evolutionary engineering. Immobilized fecal microbiota from adult donors were steadily cultivated up to 72 days in PolyFermS reactors, providing a long-term compositional and functional stable ecosystem akin to the donor's gut. Inoculation of the gut microbiota with immobilized or planktonic Lactiplantibacillus plantarum NZ3400, a derivative of the probiotic model strain WCFS1, led to successful colonization. Whole-genome sequencing of 45 recovered strains revealed mutations in 16 genes involved in signaling, metabolism, transport, and cell surface. Remarkably, mutations in LP_RS14990, LP_RS15205, and intergenic region LP_RS05100

Complete Genome Sequence of Escherichia coli Sequence Type 1193 Isolate AVS0096, Recovered from River Water in Switzerland.

Escherichia coli sequence type 1193 (ST1193) is an important cause of multidrug-resistant extraintestinal infections. Here, we report the complete genome sequence of strain AVS0096, isolated from river water in Switzerland in 2020. The genome consists of a chromosome (4.9 Mbp), a multidrug resistance plasmid (101 kb), and two small plasmids.

Inhibition of CXCR4 and CXCR7 Is Protective in Acute Peritoneal Inflammation.

Our previous studies revealed a pivotal role of the chemokine stromal cell-derived factor (SDF)-1 and its receptors CXCR4 and CXCR7 on migratory behavior of polymorphonuclear granulocytes (PMNs) in pulmonary inflammation. Thereby, the SDF-1-CXCR4/CXCR7-axis was linked with adenosine signaling. However, the role of the SDF-1 receptors CXCR4 and CXCR7 in acute inflammatory peritonitis and peritonitis-related sepsis still remained unknown. The presented study provides new insight on the mechanism of a selective inhibition of CXCR4 (AMD3100) and CXCR7 (CCX771) in two models of peritonitis and peritonitis-related sepsis by injection of zymosan and fecal solution. We observed an increased expression of SDF-1, CXCR4, and CXCR7 in peritoneal tissue and various organs during acute inflammatory peritonitis. Selective inhibition of CXCR4 and CXCR7 reduced PMN accumulation in the peritoneal fluid and infiltration of neutrophils in lung and liver tissue in both models. Both inhibitors had no anti-inflammatory effects in A2B knockout animals (A2B-/-). AMD3100 and CCX771 treatment reduced capillary leakage and increased formation of tight junctions as a marker for microvascular permeability in wild type animals. In contrast, both inhibitors failed to improve capillary leakage in A2B-/- animals, highlighting the impact of the A2B-receptor in SDF-1 mediated signaling. After inflammation, the CXCR4 and CXCR7 antagonist induced an enhanced expression of the protective A2B adenosine receptor and an increased activation of cAMP (cyclic adenosine mono phosphate) response element-binding protein (CREB), as downstream signaling pathway of A2B. The CXCR4- and CXCR7-inhibitor reduced the release of cytokines in wild type animals via decreased intracellular phosphorylation of ERK and NFκB p65. In vitro, CXCR4 and CXCR7 antagonism diminished the chemokine release of human cells and increased cellular integrity by enhancing the expression of tight junctions. These protective effects were linked with functional A2B-receptor signaling, confirming our in vivo data. In conclusion, our study revealed new protective aspects of the pharmacological modulation of the SDF-1-CXCR4/CXCR7-axis during acute peritoneal inflammation in terms of the two hallmarks PMN migration and barrier integrity. Both anti-inflammatory effects were linked with functional adenosine A2B-receptor signaling.

Traditional milk transformation schemes in Côte d'Ivoire and their impact on the prevalence of Streptococcus bovis complex bacteria in dairy products.

The Streptococcus bovis/Streptococcus equinus complex (SBSEC) and possibly Streptococcus infantarius subsp. infantarius (Sii) are associated with human and animal diseases. Sii predominate in spontaneously fermented milk products with unknown public health effects. Sii/SBSEC prevalence data from West Africa in correlation with milk transformation practices are limited. Northern Côte d'Ivoire served as study area due to its importance in milk production and consumption and to link a wider Sudano-Sahelian pastoral zone of cross-border trade. We aimed to describe the cow milk value chain and determine Sii/SBSEC prevalence with a cross-sectional study. Dairy production practices were described as non-compliant with basic hygiene standards. The system is influenced by secular sociocultural practices and environmental conditions affecting product properties. Phenotypic and molecular analyses identified SBSEC in 27/43 (62.8%) fermented and 26/67 (38.8%) unfermented milk samples. Stratified by collection stage, fermented milk at producer and vendor levels featured highest SBSEC prevalence of 71.4% and 63.6%, respectively. Sii with 62.8% and 38.8% as well as Streptococcus gallolyticus subsp. macedonicus with 7.0% and 7.5% were the predominant SBSEC species identified among fermented and unfermented milk samples, respectively. The population structure of Sii/SBSEC isolates seems to reflect evolving novel dairy-adapted, non-adapted and potentially pathogenic lineages. Northern Côte d'Ivoire was confirmed as area with high Sii presence in dairy products. The observed production practices and the high diversity of Sii/SBSEC supports in-depth investigations on Sii ecology niche, product safety and related technology in the dairy value chain potentially affecting large population groups across sub-Saharan Africa.

Risk factors for the carriage of Streptococcus infantarius subspecies infantarius isolated from African fermented dairy products.

Streptococcus infantarius subsp. infantarius (Sii) has been identified as predominant lactic acid bacteria in spontaneously fermented dairy products (FDPs) in sub-Saharan Africa including Côte d'Ivoire. However, Sii belongs to the Streptococcus bovis/Streptococcus equinus complex (SBSEC). Most SBSEC members are assumed to be involved as opportunistic pathogens in serious diseases in both humans and animals. A population-based cross-sectional survey, including 385 participants was conducted in Korhogo, northern Côte d'Ivoire, to identify risk factors for Sii fecal carriage, including consumption of local FDPs. A structured questionnaire was used to gather participant's socio-demographic and economic characteristics, their relation to livestock and dietary habits. In addition, fresh stool and milk samples were collected. The identification of Sii was done using a SBSEC-specific PCR assay targeting 16S rRNA and groEL genes. The overall prevalence of SBSEC and Sii carriage was 23.2% (confidence interval CI 95% = 18.9-27.5) and 12.0% (CI 95% = 8.4-15.5) for stool, respectively. Prevalence of Sii was significantly higher in consumers of artisanal butter compared with non-consumers (57.1% vs 10.1%, odds ratio OR: 11.9, 95% CI: 3.9-36.6), as well as in persons handling livestock (OR = 3.9; 95% CI = 1.6-9.3) and livestock primary products (OR = 5.7; 95% CI = 2.3-14.3). The closer contact with livestock was a risk factor for Sii fecal carriage. Sii strains were isolated from fresh and fermented milk products with a prevalence of 30.4% and 45.4%, respectively. Analysis of Sii population structure through the SBSEC multi locus sequence typing assay revealed a close relationship across human and dairy isolates, possibly linked to a Kenyan human isolate. All these outcomes underline the interest of in-depth investigations on the ecology, potential reservoirs and pathways of contamination by Sii at the human-animal-environment interface in comparison to yet to be collected data from Europe, Asia and the Americas to further elucidate the various roles of Sii.

Characterization of low-molecular-weight antiyeast metabolites produced by a food-protective Lactobacillus-Propionibacterium coculture.

We developed a pH-controlled batch fermentation process with separately immobilized cells of the protective coculture of Lactobacillus paracasei subsp. paracasei SM20 and Propionibacterium jensenii SM11 in supplemented whey permeate medium yielding cell-free supernatants with high antiyeast activity against Candida pulcherrima and Rhodotorula mucilaginosa. The antiyeast compounds were resistant to proteinase K and pronase E treatments and showed high heat resistance (121 degrees C for 15 min). Diafiltration (1,000-Da cutoff) revealed that the inhibitory metabolites have low molecular weights. Partial purification of active compounds was achieved by a microplate bioassay controlled procedure with solid-phase extraction (C18) followed by (i) gel filtration chromatography or (ii) semipreparative reverse-phase high-performance liquid chromatography (C18). In addition to propionic, acetic, and lactic acids, 2-pyrrolidone-5-carboxylic acid, 3-phenyllactic acid, hydroxyphenyllactic acid, and succinic acid were identified by chromatography and mass spectrometry. Accurate quantifications revealed only low concentrations (up to 7 mM) of 2-pyrrolidone-5-carboxylic acid, 3-phenyllactic acid, and hydroxyphenyllactic acid produced during fermentation in contrast to relatively high MICs (50 to more than 500 mM) determined at different pH values (4.0, 5.0, and 6.0). Succinic acid was present at higher concentrations (29 mM) in cell-free supernatants but with comparable high MICs (200 to more than 500 mM and pH 4.0, 5.0, and 6.0). Although none of these compounds was the main substance responsible per se for suppression of yeast growth, our study revealed a complex antiyeast mechanism with putative synergistic effects between several low-molecular-weight compounds.

The Road to Infection: Host-Microbe Interactions Defining the Pathogenicity of Streptococcus bovis/Streptococcus equinus Complex Members.

The Streptococcus bovis/Streptococcus equinus complex (SBSEC) comprises several species inhabiting the animal and human gastrointestinal tract (GIT). They match the pathobiont description, are potential zoonotic agents and technological organisms in fermented foods. SBSEC members are associated with multiple diseases in humans and animals including ruminal acidosis, infective endocarditis (IE) and colorectal cancer (CRC). Therefore, this review aims to re-evaluate adhesion and colonization abilities of SBSEC members of animal, human and food origin paired with genomic and functional host-microbe interaction data on their road from colonization to infection. SBSEC seem to be a marginal population during GIT symbiosis that can proliferate as opportunistic pathogens. Risk factors for human colonization are considered living in rural areas and animal-feces contact. Niche adaptation plays a pivotal role where Streptococcus gallolyticus subsp. gallolyticus (SGG) retained the ability to proliferate in various environments. Other SBSEC members have undergone genome reduction and niche-specific gene gain to yield important commensal, pathobiont and technological species. Selective colonization of CRC tissue is suggested for SGG, possibly related to increased adhesion to cancerous cell types featuring enhanced collagen IV accessibility. SGG can colonize, proliferate and may shape the tumor microenvironment to their benefit by tumor promotion upon initial neoplasia development. Bacteria cell surface structures including lipotheichoic acids, capsular polysaccharides and pilus loci (pil1, pil2, and pil3) govern adhesion. Only human blood-derived SGG contain complete pilus loci and other disease-associated surface proteins. Rumen or feces-derived SGG and other SBSEC members lack or harbor mutated pili. Pili also contribute to binding to fibrinogen upon invasion and translocation of cells from the GIT into the blood system, subsequent immune evasion, human contact system activation and collagen-I-binding on damaged heart valves. Only SGG carrying complete pilus loci seem to have highest IE potential in humans with significant links between SGG bacteremia/IE and underlying diseases including CRC. Other SBSEC host-microbe combinations might rely on currently unknown mechanisms. Comparative genome data of blood, commensal and food isolates are limited but required to elucidate the role of pili and other virulence factors, understand pathogenicity mechanisms, host specificity and estimate health risks for animals, humans and food alike.