Christoph Jans

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.

Genomics, evolution, and molecular epidemiology of the Streptococcus bovis/Streptococcus equinus complex (SBSEC)

The Streptococcus bovis/Streptococcus equinus complex (SBSEC) is a group of human and animal derived streptococci that are commensals (rumen and gastrointestinal tract), opportunistic pathogens or food fermentation associates. The classification of SBSEC has undergone massive changes and currently comprises 7 (sub)species grouped into four branches based on sequences identities: the Streptococcus gallolyticus, the Streptococcus equinus, the Streptococcus infantarius and the Streptococcus alactolyticus branch. In animals, SBSEC are causative agents for ruminal acidosis, potentially laminitis and infective endocarditis (IE). In humans, a strong association was established between bacteraemia, IE and colorectal cancer. Especially the SBSEC-species S. gallolyticus subsp. gallolyticus is an emerging pathogen for IE and prosthetic joint infections. S. gallolyticus subsp. pasteurianus and the S. infantarius branch are further associated with biliary and urinary tract infections. Knowledge on pathogenic mechanisms is so far limited to colonization factors such as pili and biofilm formation. Certain strain variants of S. gallolyticus subsp. macedonicus and S. infantarius subsp. infantarius are associated with traditional dairy and plant-based food fermentations and display traits suggesting safety. However, due to their close relationship to virulent strains, their use in food fermentation has to be critically assessed. Additionally, implementing accurate and up-to-date taxonomy is critical to enable appropriate treatment of patients and risk assessment of species and strains via recently developed multilocus sequence typing schemes to enable comparative global epidemiology. Comparative genomics revealed that SBSEC strains harbour genomics islands (GI) that seem acquired from other streptococci by horizontal gene transfer. In case of virulent strains these GI frequently encode putative virulence factors, in strains from food fermentation the GI encode functions that are pivotal for strain performance during fermentation. Comparative genomics is a powerful tool to identify acquired pathogenic functions, but there is still an urgent need for more physiological and epidemiological data to understand SBSEC-specific traits.

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

BackgroundStreptococcus 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.ResultsThe 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.ConclusionsThe 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.

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 β-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.

Adhesion Potential of Intestinal Microbes Predicted by Physico-Chemical Characterization Methods.

Bacterial adhesion to epithelial surfaces affects retention time in the human gastro-intestinal tract and therefore significantly contributes to interactions between bacteria and their hosts. Bacterial adhesion among other factors is strongly influenced by physico-chemical factors. The accurate quantification of these physico-chemical factors in adhesion is however limited by the available measuring techniques. We evaluated surface charge, interfacial rheology and tensiometry (interfacial tension) as novel approaches to quantify these interactions and evaluated their biological significance via an adhesion assay using intestinal epithelial surface molecules (IESM) for a set of model organisms present in the human gastrointestinal tract. Strain pairs of Lactobacillus plantarum WCFS1 with its sortase knockout mutant Lb. plantarum NZ7114 and Lb. rhamnosus GG with Lb. rhamnosus DSM 20021T were used with Enterococcus faecalis JH2-2 as control organism. Intra-species comparison revealed significantly higher abilities for Lb. plantarum WCSF1 and Lb. rhamnosus GG vs. Lb. plantarum NZ7114 and Lb. rhamnosus DSM 20021T to dynamically increase interfacial elasticity (10−2 vs. 10−3 Pa*m) and reduce interfacial tension (32 vs. 38 mN/m). This further correlated for Lb. plantarum WCSF1 and Lb. rhamnosus GG vs. Lb. plantarum NZ7114 and Lb. rhamnosus DSM 20021T with the decrease of relative hydrophobicity (80–85% vs. 57–63%), Zeta potential (-2.9 to -4.5 mV vs. -8.0 to -13.8 mV) and higher relative adhesion capacity to IESM (3.0–5.0 vs 1.5–2.2). Highest adhesion to the IESM collagen I and fibronectin was found for Lb. plantarum WCFS1 (5.0) and E. faecalis JH2-2 (4.2) whereas Lb. rhamnosus GG showed highest adhesion to type II mucus (3.8). Significantly reduced adhesion (2 fold) to the tested IESM was observed for Lb. plantarum NZ7114 and Lb. rhamnosus DSM 20021T corresponding with lower relative hydrophobicity, Zeta potential and abilities to modify interfacial elasticity and tension. Conclusively, the use of Zeta potential, interfacial elasticity and interfacial tension are proposed as suitable novel descriptive and predictive parameters to study the interactions of intestinal microbes with their hosts.

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.

African fermented dairy products – Overview of predominant technologically important microorganisms focusing on African Streptococcus infantarius variants and potential future applications for enhanced food safety and security

Milk is a major source of nutrients, but can also be a vehicle for zoonotic foodborne diseases, especially when raw milk is consumed. In Africa, poor processing and storage conditions contribute to contamination, outgrowth and transmission of pathogens, which lead to spoilage, reduced food safety and security. Fermentation helps mitigate the impact of poor handling and storage conditions by enhancing shelf life and food safety. Traditionally-fermented sour milk products are culturally accepted and widely distributed in Africa, and rely on product-specific microbiota responsible for aroma, flavor and texture. Knowledge of microbiota and predominant, technologically important microorganisms is critical in developing products with enhanced quality and safety, as well as sustainable interventions for these products, including Africa-specific starter culture development. This narrative review summarizes current knowledge of technologically-important microorganisms of African fermented dairy products (FDP) and raw milk, taking into consideration novel findings and taxonomy when re-analyzing data of 29 publications covering 25 products from 17 African countries. Technologically-important lactic acid bacteria such as Lactococcus lactis and Streptococcus infantarius subsp. infantarius (Sii), Lactobacillus spp. and yeasts predominated in raw milk and FDP across Africa. Re-analysis of data also suggests a much wider distribution of Sii and thus a potentially longer history of use than previously expected. Therefore, evaluating the role and safety of African Sii lineages is important when developing interventions and starter cultures for FDP in Africa to enhance food safety and food security. In-depth functional genomics, epidemiologic investigations and latest identification approaches coupled with stakeholder involvement will be required to evaluate the possibility of African Sii lineages as novel food-grade Streptococcus lineage.

Complete Genome Sequence of the Probiotic Bifidobacterium thermophilum Strain RBL67

ABSTRACT Bifidobacterium thermophilum RBL67, an isolate from infant feces, exhibits bacteriocin-like antimicrobial activity against Listeria spp. and Salmonella spp. and protects HT29-MTX cells against Salmonella infection. Here, the complete genome sequence of the probiotic B. thermophilum strain RBL67 is presented.

Bifidobacterium pseudolongum Strain PV8-2, Isolated from a Stool Sample of an Anemic Kenyan Infant.

ABSTRACT The complete genome sequence of Bifidobacterium pseudolongum PV8-2, isolated from feces of an anemic Kenyan infant, was determined using single-molecule real-time (SMRT) technology. The genome consists of a 2-Mbp chromosome and a 4-kb plasmid.

Complete and Assembled Genome Sequence of Bifidobacterium kashiwanohense PV20-2, Isolated from the Feces of an Anemic Kenyan Infant

ABSTRACT The complete genome sequence of Bifidobacterium kashiwanohense strain PV20-2, an infant feces isolate, was determined using single-molecule real-time sequencing (SMRT). Hierarchical genome assembly resulted in a completely assembled genome of 2,370,978 bp. The B. kashiwanohense PV20-2 genome is the first completely sequenced and assembled genome of the species.