Front. Microbiol. Frontiers in Microbiology Front. Microbiol. 1664-302X Frontiers Media S.A. 10.3389/fmicb.2026.1750102 Original Research Biodiversity of the antimicrobial potential of Weissella paramesenteroides strains isolated from dairy cattle Fischer Sebastian W.12* Conceptualization Data curation Formal analysis Funding acquisition Investigation Methodology Project administration Resources Supervision Validation Visualization Writing – original draft Writing – review & editing Euler Anna2 Data curation Investigation Writing – review & editing Bertels Leonie2 Investigation Writing – review & editing Mariani Corea Nadine2 Supervision Writing – review & editing Titgemeyer Fritz2 Conceptualization Funding acquisition Methodology Project administration Resources Supervision Validation Visualization Writing – original draft Writing – review & editing Institute for Hygiene and Public Health, University Clinics Bonn, Bonn, Germany Department of Food - Nutrition - Facilities, FH Muenster, Münster, Germany Correspondence: Sebastian W. Fischer, sebastian.fischer@fh-muenster.de 20 02 2026 2026 17 1750102 19 11 2025 20 12 2025 05 01 2026 Copyright © 2026 Fischer, Euler, Bertels, Mariani Corea and Titgemeyer. 2026 Fischer, Euler, Bertels, Mariani Corea and Titgemeyer https://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

Weissella species are lactic acid bacteria with a high potential for the fermentation of food items. They ferment fruits, vegetables, fish, and meat to deliver aromatic ingredients, produce antibacterial and antifungal compounds, and exert probiotic properties. Despite these attractive attributions, they are still poorly studied. We have isolated 40 strains of Weissella paramesenteroides from one biotope, the udder of milk cattle, to examine genomic plasticity, antibacterial efficacy, and organic acid and hydrogen peroxide formation. Each isolate was identified by DNA sequence comparisons of the 16S rRNA-encoding gene. Patterns of genomic DNA fragments from random amplification of polymorphic DNA showed that 38 isolates differed by more than 5%, thus representing subspecies. Nine isolates were selected for further characterization. They were able to inhibit all eight food-borne pathogens and surrogates used. Salmonella enterica and Pseudomonas aeruginosa were strongly inhibited, while inhibition of Listeria monocytogenes was weaker but still significant. However, the inhibition profiles varied considerably. Acid production was assessed for each isolate in the presence of indicator strains exhibiting different acidification patterns as well. The presence of S. enterica was associated with a drop in pH, whereas no notable acidification occurred when isolates were challenged with Klebsiella pneumoniae and Bacillus subtilis. To confirm the observed differences, we analyzed the pan-genomes of 10 genomes retrieved from the NCBI database. Each genome contains several 100 accessory genes and up to 6.5% unique genes, indicating a high genome fluidity and additional strain-specific metabolic capacity. We suggest that the antibacterial efficacy of W. paramesenteroides is based on a multi-barrier system, whereby strains have developed different genetic qualities for the expression of an individual set of barriers.

 antibacterial biodiversity food safety genome fluidity genome plasticity lactic acid bacteria Weissella The author(s) declared that financial support was received for this work and/or its publication. This study was financed by the Federal Ministry of Economic Affairs and Climate Action through a research grant of the “Zentrales Innovations programm Mittelstand (ZF4679802AJ2).” Further support was received by the Open Access Publication Fund of the University of Bonn. section-at-acceptance Food Microbiology Introduction

Weissella are Gram-positive, non-motile, facultative anaerobic microorganisms that grow heterofermentatively (Teixeira et al., 2021b; Fusco et al., 2023). They are closely related to Periweissellae, Furfurilactobacilli, and Leuconostocha (Qiao et al., 2023). The 26 currently described Weissella species belong to the order Lactobacillales, comprising 36 genera and over 400 species (Zheng et al., 2020; Bello and Gupta, 2025; Göker et al., 2025). Phylogenetic analyses have classified them into six species groups, each defined by a signature species: Weissella beninensis, Weissella kandleri, Weissella oryzae, Weissella halotolerans, Weissella confusa, and Weissella paramesenteroides (Fanelli et al., 2022). According to the PubMed database, members of the W. confusa and W. paramesenteroides groups, including W. cibaria, W. confusa, and W. paramesenteroides, have been predominantly investigated, with 440, 343, and 128 mentions in titles and abstracts, respectively. However, they are less well studied, compared to some well-characterized Lactobacillus species or closely related species such as Leuconostoc mesenteroides, which have several 1,000 entries in PubMed (Apostolakos et al., 2022).

Weissella species are ubiquitous. They are found among others in soil, tomatoes, bee pollen, papaya, the rhizosphere of olive trees, Chardonnay grapes, red pepper, desert plants, milk, dairy, kimchi, sourdough, fermented fish, fermented sausages, Mexican podzol, and especially in fermented beverages and fruits, and even in heroin (Fusco et al., 2015; Onur and Önlü, 2023). In humans, the oral cavity, breast milk, the gastrointestinal tract, the vagina, and feces are places where Weissella species occur (Walter et al., 2001; Martín et al., 2007a, 2007b; Albesharat et al., 2011; Kang et al., 2011; Teixeira et al., 2021a).

As gut-dwelling bacteria, they contribute to human health. This encompasses the degradation of dietary fibers, secretion of folic acid, anti-inflammatory effects, and immune system stimulation (Xiong et al., 2025). It has further been shown that W. confusa and W. cibaria produce exopolysaccharides and thus deliver prebiotic nutrients for beneficial gut-dwelling bacteria. Weissella species can tolerate low pH values, bile salts, and can adhere to the epithelial cells of the gut mucosa, three advantageous features to reside in the gastrointestinal tract (Pabari et al., 2020; Teixeira et al., 2021a; Onur and Önlü, 2023). Another characteristic is the inhibition of pathogens (Agirman et al., 2021). The mechanisms comprise the secretion of organic acids, nutrient and epithelial adhesion-site competition, hydrogen peroxide (H2O2) formation under oxygen stress, disruption of quorum-sensing communication of target bacteria, secretion of bacteriocins and small peptides, and so-called bacteriocin-like inhibitory substances (BLIS) (Papagianni and Papamichael, 2011; Silva et al., 2018; Fischer and Titgemeyer, 2023; Singh et al., 2024; Thuy et al., 2024; Rotta et al., 2025). Weissella species can inhibit foodborne pathogens like Bacillus cereus, Escherichia coli, Klebsiella pneumoniae, Listeria monocytogenes, Pseudomonas aeruginosa, Salmonella enterica, and Staphylococcus aureus, as well as various fungi, particularly by ion scavenging (Papagianni and Papamichael, 2011; Teixeira et al., 2021a; Wan et al., 2023; Thuy et al., 2024).

The abovementioned features, in combination with good growth capacities under standard laboratory conditions, indicate a favorable use of Weissella species in food items (Fessard and Remize, 2017). The frequent isolation from diverse biotopes points toward a rich, exploitable gene pool (Fusco et al., 2015).

Here, we report on an analysis of W. paramesenteroides strains isolated from foremilk and teat canal biofilms from dairy cows. We describe the genetic diversity and show differences in antibacterial efficacy and acid production against a range of foodborne pathogens and surrogates. To further explain the observed differences, we analyzed the genomic fluidity by examination of available completed W. paramesenteroides genomes retrieved from the NCBI genome database.

 Materials and methods Strain isolation and indicator strains

Samples were obtained from 18 farms situated in Münsterland, Germany. They were collected as follows. The udder of a cow was disinfected with an alcohol-soaked cloth before a biofilm was collected by introducing a swab for about 2 cm into the teat canal. Foremilk samples of 5–10 mL were withdrawn by milking. Samples were diluted in 0.9% sodium chloride solution and propagated as described previously (Mariani Corea et al., 2025). The isolated colonies were subjected to polymerase chain reactions (PCR) using primer 27f and 535R of the 16S rRNA gene (Fischer et al., 2025). The PCR products were subsequently sequenced by Microsynth Seqlab (Germany). Identification of species was achieved by using the Basic Local Alignment Tool (BLAST) provided by the National Center for Biotechnology Information (NCBI, United States) (Altschul et al., 1990). The isolates of W. paramesenteroides strains used in this study are listed in Table 1.

List of W. paramesenteroides isolates.

No. Strain Farm Animal husbandry, sample
1 05.3 92.2 VL-3 1 Conventional, biofilm
2 05.3162.2 HL-21 1 Conventional, biofilm
3 05.3162.2 HL-31 1 Conventional, biofilm
4 13.7 1A-4 13 Conventional grazing, biofilm
5 13.7 6A-9 13 Conventional grazing, biofilm
6 13.7 10A-2 13 Conventional grazing, biofilm
7 13.7 11A-5 13 Conventional grazing, biofilm
8 13.7 13A-1 13 Conventional grazing, biofilm
9 13.7 14A-1 13 Conventional grazing, biofilm
10 13.7 17–2 13 Conventional grazing, foremilk
11 13.7 19A-10 13 Conventional grazing, biofilm
12 13.7 21A-7 13 Conventional grazing, biofilm
13 13.7 22A-12 13 Conventional grazing, biofilm
14 13.7 22A-22 13 Conventional grazing, biofilm
15 13.7 22A-52 13 Conventional grazing, biofilm
16 13.7 22A-72 13 Conventional grazing, biofilm
17 18.5 1A-3 7 Conventional, biofilm
18 18.5 11–4 7 Conventional, foremilk
19 18.5 12A-10 7 Conventional, biofilm
20 18.5 19–3 7 Conventional, foremilk
21 20.4 7A-53 3 Organic, biofilm
22 20.4 7A-63 3 Organic, biofilm
23 22.4 S2-4 4 Collection tank
24 22.4 6A-2 4 Conventional, biofilm
25 22.4 7–3 4 Conventional, foremilk
26 22.4 7A-9 4 Conventional, biofilm
27 22.4 12A-3 4 Conventional, biofilm
28 26.3 5A-2 2 Conventional, biofilm
29 26.3 6A-5 2 Conventional, biofilm
30 27.4 5–4 4 Conventional, foremilk
31 27.4 6–1 4 Conventional, foremilk
32 27.4 8–1 4 Conventional, foremilk
33 27.8 11A-24 18 Conventional, biofilm
34 27.8 11A-54 18 Conventional, biofilm
35 27.8 11A-64 18 Conventional, biofilm
36 27.8 13A-4 18 Conventional, biofilm
37 31.5 4A-4 10 Conventional, biofilm
38 31.5 18–15 10 Conventional, foremilk
39 31.5 18–65 10 Conventional, foremilk
40 31.5 19–6 10 Conventional, foremilk

1, 2, 3, 4, 5The isolates were derived from the same quarter of the udder.

Eight bacterial species were employed as indicators and surrogates, representing foodborne pathogens and food spoilage organisms (Table 2). The strains were cultivated in Luria-Bertani (LB) Broth (Carl Roth, Germany) at 37 °C for 24 h. Brain Heart Infusion (BHI) Broth (BD, USA) was utilized for the cultivation of L. monocytogenes.

List of indicator strains.

Species Origin Comment
Bacillus subtilis Laboratory strain collection Surrogate food spoilage
Citrobacter koseri Laboratory strain collection Fish pathogen
Klebsiella pneumoniae Laboratory strain collection Opportunistic pathogen
Listeria monocytogenes Dsm 20,600 Foodborne pathogen
Pseudomonas aeruginosa Atcc 15442 Food spoilage and pathogen
Salmonella enterica* Lt-2 Foodborne pathogen
Staphylococcus haemolyticus Laboratory strain collection Surrogate food pathogen
Staphylococcus warneri Laboratory strain collection Surrogate food pathogen

*re-classified from originally S. typhimurium to S. enterica ssp. enterica ser. Typhimurium (Ames et al., 1975; Judicial Commission of the International Committee on Systematics of Prokaryotes, 2005).

 Random-amplified-polymorphic-DNA analysis (RAPD)

RAPD–PCR fingerprinting was carried out by applying the M13 minisatellite core primer (5′-GAGGGTGGCGGTTCT-3′) (Rossetti and Giraffa, 2005). PCRs were conducted as previously described (Fischer et al., 2025). The amplification products were separated by 2% agarose gel electrophoresis (AppliChem, Germany).

 Hydrogen peroxide production

Hydrogen peroxide production was conducted as described (Tomás et al., 2004). Colonies were streaked out from an overnight culture onto an mMRS agar plate supplemented with 1 mM 3,3′,5,5′-Tetramethylbenzidin (TMB) and 2 U/mL of type II horseradish peroxidase (Carl Roth, Germany). Agar plates were incubated anaerobically for 48 h at 30 °C before the lid of the Petri dish was removed for 10 min of aeriation. The formation of H2O2 became visible when the yellowish colonies turned black.

 Lawn-on-spot assays and acidification

Antimicrobial activities against foodborne pathogens and food spoilage organisms were evaluated with lawn-on-spot assays as described previously (Mariani Corea et al., 2025). W. paramesenteroides isolates were inoculated from a liquid overnight culture by positioning 2 μL into the center of an MRS agar plate supplemented with 0.5 g/L cysteine (Carl Roth, Germany) (mMRS) and incubated under anaerobic conditions at 30 °C for 48 h. A cell suspension of 0.2 mL containing 2.0×107 CFU/mL from a fresh overnight culture of the respective indicator strain was mixed with 10 mL of LB-soft agar (0.8%, Bacto Agar, BD, United States) or BHI-soft agar (0.8%) in the case of L. monocytogenes. The suspension was poured onto an mMRS plate. Assay plates were aerobically incubated in co-cultivation for 20–30 h at 37 °C. The clear halo (zone of inhibition), from which the colony diameter was subtracted, was measured in millimeters. Acid secretion of the W. paramesenteroides isolates was monitored with color-fixed indicator sticks (pH Strips; Carl Roth). In this way, it was possible to document changes in pH by comparing the pH in the inhibition zone with the pH of the indicator strain grown at the edge of the same agar plate. As a control, the pH was measured for all isolates and indicator strains solely cultivated on mMRS-soft agar plates.

 Data analyses and statistics

Data analyses comprised subspecies identification, antimicrobial activity in vivo, and the relationship between antimicrobial compounds and acid secretion. Therefore, the statistical and computing package R (v4.2.1) and Rstudio (v2023.03.0 + 386) with packages tidyverse (v2.0.0), readxl (v1.4.2), ggplot2 (v3.4.2), vegan (2.6–4), and scico (v1.5.0) were used (Crameri, 2018; Wickham et al., 2019; R Core Team, 2021; Oksanen et al., 2022; RStudio Team, 2022; Wickham and Bryan, 2022). Subspecies identification was based on the analysis of the data derived from RAPD-PCR DNA band patterns. The unweighted pair group method with arithmetic mean (UPGMA) was applied by using the hclust algorithm together with the Dice dissimilarity coefficient matrix from the R base package (Hilton et al., 1997; Ghazi et al., 2016). Phylogenetic trees were generated using a threshold of ≤5% dissimilarity to distinguish clonal isolates (Ghazi et al., 2016; Szaluś-Jordanow et al., 2018; Ruiz et al., 2014; Nielsen et al., 2014; Birch et al., 1996). Heatmaps showing antibacterial activities and acidification were generated on the basis of the average values from triplicates. Spearman correlation and scatterplot visualization performed on the raw replicate values assessed the contributions of antimicrobial compounds and acid secretion at a 95% confidence level. Phylogenetic trees, heatmaps, and scatterplots were visualized using ggplot2.

 Genome analyses

All available complete genomes of W. paramesenteroides together with the respective plasmids, were retrieved from the NCBI genome database: GCF_050613345.1, GCF_051903945.1, GCF_002386265.1, GCF_028994215.1, GCF_045005335.1, GCF_045006675.1, GCF_036327715.1, GCF_015689215.1, GCF_030168775.1, and GCA_052281005.1. Venn diagrams showing pan-genomes were computed with EDGAR 3.0 on the basis of the gene annotation list created with Bakta (Dieckmann et al., 2021; Schwengers et al., 2021). Singleton gene calculation was done with EDGAR 3.0 by running each of the 10 genomes against the remaining nine (Dieckmann et al., 2021). Genome screening of bacteriocin-encoding genes was performed using webserver BAGEL4 and BLASTP of the National Center for Biotechnology Information (Altschul et al., 1990; van Heel et al., 2018).

 Results Isolation of <italic>Weissella paramesenteroides</italic> strains and genetic biodiversity

A screening for lactic acid bacteria (LAB) from teat canal biofilms and foremilk samples from lactating cows across 18 dairy farms in Münsterland, Germany, yielded several 100 cultivable isolates. Species were assigned by BLASTN database alignments of the sequenced 16S rRNA gene fragments. Forty unequivocally identified isolates of W. paramesenteroides, which represented a prominent, well-cultivable group, were then selected for this study.

To determine whether W. paramesenteroides isolates differed genetically or represented clones, each strain was subjected to RAPD experiments. As shown in Figure 1A, differences in the gene fragment patterns were obvious between most isolates. The derived UPGMA dendrogram resolved multiple well-defined clusters alongside long-branch outliers (Figure 1D). Two pairs of isolates (27.4 6–1, 27.4 5–4, 13.7 22A-1, and 13.7 22A-2) co-clustered at 100% similarity, indicating clonal or near-clonal RAPD fingerprints (Figures 1B–D). The next highest grouping occurred near 75% similarity. Others resolved at the dendrogram periphery with markedly low relatedness to the remainder, reflecting more pronounced intraspecific divergence. Hence, 38 isolates represented subspecies as has been defined by a threshold of <95% (Figure 1D). Nine isolates were selected for further analyses.

RAPD fingerprints and UPGMA dendrogram. (A–C) Shown are 2% agarose gels with (A) the isolates that were used for further analyses: M, DNA Marker; 1, 05.3 162-HL-2; 2, 05.3 162-HL-3; 3, 13.7 17 -2; 4, 13.7 19A-10; 5, 20.4 7A-5; 6, 20.4 7A-6; 7, 22.4 7A-9; 8, 22.4 6A-2; 9, 22.4 S2-4. (B) Four W. paramesenteroides isolates were isolated from the same teat, of which 1 and 2 are identical: M, DNA Marker; 1, 13.7 22A-1; 2, 13.7 22 A-2; 3, 13.7 22A-5; and 13.7 22A-7. (C) Shown are the two identical isolates from the same farm isolated from different cows: M, DNA Marker; 1, 27.4 5–4; 2, 27.4 6–1. (D) UPGMA dendrogram depicting the relative relatedness (%) of the Weissella isolates. The red line shows the 95% threshold that was suggested to define subspecies (Nielsen et al., 2014). Data were obtained from duplicates.

Gel electrophoresis results and a dendrogram are displayed. The gels show DNA bands for multiple samples, marked M for the marker and lanes numbered 1-9. Bands vary in intensity and position. The dendrogram on the right represents the genetic similarity of isolates from different farms, with a similarity scale from 15% to 95%. Each isolate is color-coded to represent various farms labeled from 1 to 18, highlighting genetic diversity among them. Antimicrobial activity spectra

Antibacterial activities were assessed by lawn-on-spot assays challenging each Weissella isolate against a selection of Gram-negative and Gram-positive indicator strains representing foodborne pathogens and foodborne surrogates (Table 2). The diameters of the zones of inhibition varied between none and 4.9 cm. The differences in halo sizes are illustrated for six isolates against the Gram-negative C. koseri and the Gram-positive S. warneri (Figure 2). It can be seen that the isolates inhibited the indicators differently, and that the indicators also showed differences in halo size compared to the same isolate, as can be deduced for isolate 20.4 7A-6. A heatmap was calculated showing that all isolates were able to inhibit the indicators, with the exception of B. subtilis, which was weak but significantly inhibited by four isolates (Figure 3). The antimicrobial profiles were quite heterogeneous across all combinations. In general, the isolates performed best against the Gram-negatives P. aeruginosa and S. enterica, while K. pneumoniae was less sensitive. Several isolates, particularly 22.4 7A-9 and 05.3162.2 HL-2, displayed robust, broad-spectrum inhibition (Figure 3 and Table 3). In contrast, isolate 13.7 17–2 was the weakest isolate overall. In summary, the data presented in the heatmap reveal a broad but subspecies-specific antibacterial potential within the W. paramesenteroides collection of isolates. They will allow the systematic selection of the actual best choice against one, two, or any combination of indicators.

Zones of inhibition of six W. paramesenteroides isolates versus C. koseri and S. warneri. It shows the different sizes of the inhibition zones of the isolates against C. koseri and S. warneri and size differences by each isolate co-cultured with the respective indicator strain.

Twelve petri dish images show bacterial colonies reacting to different conditions. The top row features tests against C. koseri, with varying inhibition zones around a central colony. The bottom row shows tests against S. warneri, also with distinct inhibition zones. Each column contains two images, labeled with six isolates, in example “05.3 162.2-HL-2” or “22.4 7A-9.”

Heatmap of antibacterial activities. The color scale shows the zones of inhibition in cm. Experiments were carried out in triplicate.

Heatmap displaying the zone of inhibition for various samples against different bacterial strains. Rows represent samples with labels like “162.2 HL - 2,” and columns represent bacterial strains such as “Pseudomonas aeruginosa” and “Klebsiella pneumoniae.” Colors range from dark purple to light blue and orange, with a scale indicating inhibition zones diameters from zero to four centimeter. Acidification and hydrogen peroxide production

We also raised the question of whether the production of organic acids or H2O2 correlates with antibacterial activity. Acidification by the isolates was monitored in all lawn-on-spot assays by measuring the pH drop between the inhibition zone and the edge of the agar plate. The heatmap shown in Figure 4 displays to which extent the isolates could acidify the medium. Interestingly, this was variable and influenced by the presence of the respective indicator strain. While 22.4 7A-9 showed strong acid production against P. aeruginosa and S. enterica, the drop in pH was small against C. koseri and K. pneumoniae. The presence of S. enterica showed the highest medium acidification when exposed to the W. paramesenteroides strains, while other indicators caused the opposite (Figure 4). P. aeruginosa, however, gave a mixed picture. While isolate 22.4 7A-9 caused the highest acidification of all, co-culture with four other isolates resulted in mild acidification.

Heatmap of pH drop of each W. paramesenteroides isolate in co-culture with each indicator. Data represent the average of triplicates.

Heatmap showing pH drop across various bacterial strains in different samples. Rows represent Weissella isolates like 162.2 HL-2 and 22.4 S2-4, while columns denote pathogenic bacteria such as Pseudomonas aeruginosa and Listeria monocytogenes. Color intensity ranges from light peach (0.0) to dark blue (-1.5), indicating pH change severity.

To examine a relation between antimicrobial activity and acidification more rigorously, Pearson correlations were performed for each pair of strain and indicator. As shown in Figures 5A,B, significant negative correlations (cor − 0.48 to −0.61, p < 0.05) were observed for strains 05.3162.2 HL-2, 20–4 7A-5, 22.4 6A-2, 22.4 S2-4, indicating that acid production may cause an increase in the zone of inhibition (Figure 5A). For the remaining five isolates, it appeared that acid production played a minor role in causing antibacterial activity.

Pearson correlation analysis. Shown is the relation between pH drop and the sizes of zones of inhibition. (A) W. paramesenteroides isolates versus indicator strains and (B) indicator strains versus W. paramesenteroides isolates. cor, correlation coefficient; p-value (significance threshold <0.05).

Graphs displaying correlations between zone of inhibition size and pH drop for various bacterial strains. Each graph includes a blue trend line with gray confidence intervals and black data points. Panel A shows diverse strains, including "13.7 17 - 2" and "22.4 S2 - 4," with correlation coefficients and p-values. Panel B highlights bacteria such as Bacillus subtilis and Salmonella enterica, also with correlation data. The x-axis represents the zone of inhibition in centimeters, and the y-axis shows pH drop.

A second correlation analysis was conducted to calculate the behavior of the indicator strains regarding acid production (Figure 5B), and vice versa. It revealed that S. enterica (cor = −0.42, p = 0.028) was acid-driven, inhibited by the panel of isolates. That acidification did not significantly contribute to the antibacterial activity was found for K. pneumoniae (cor, 0.48; p, 0.011). The Pearson data on the remaining indicators showed no significant correlations.

All 40 isolates were tested for their ability to produce H2O2, with 18 testing positive (data not shown). Within this group, six of the nine isolates selected for lawn-on-spot assays synthesized the antibacterial compound under the conditions used (Table 3 and Figure 6).

Production of hydrogen peroxide. Experiments were carried out in triplicate.

Three petri dishes containing each three sections in which bacteria are streaked out to single colonies. Some of them form black colonies showing formation of hydrogen peroxide. Others that do not produce hydrogen peroxide stay whitish.

Ranking of antibacterial efficacy.

Strain Antibacterial efficacy ranking Sum (cm) Mean (cm) Antibacterial efficacy correlation to acid production Significant antibacterial correlation to acid production (*p < 0.05) H2O2 Best performer against indicator
22.4 7A-9 1 18.75 2.68 −0.41 0.061 − Bs, Ck, Pa, Se, Sw
162.2 HL-2 2 16.43 2.35 −0.48 0.017* + Ck, Kp, Sw
22.4 6A-2 3 15.88 2.28 −0.61 0.002* + Sh
20.4 7A-5 4 14.26 2.04 −0.58 0.003* + Lm
20.4 7A-6 5 14.10 2.02 −0.25 0.258 + /
162.2 HL-3 6 13.03 1.86 −0.07 0.768 + /
22.4 S2-4 7 12.44 1.78 −0.53 0.008* − /
13. 7 19A-10 8 12.39 1.77 −0.24 0.259 − /
13.7 17–2 9 11.24 1.60 −0.36 0.088 + /

Sum, added zones of inhibition; mean, average inhibition zone; Bs, B. subtilis; Ck, C. koseri; Kp, K. pneumoniae; Lm, L. monocytogenes; Pa, P. aeruginosa; Se, S. enterica; Sh, S. haemolyticus; Sw, S. warneri.

Table 3 shows the ranking of isolates according to their antibacterial efficacy. The 22.4 7A-9 strain performed best in five out of eight indicators. It appears that neither the production of organic acid nor H2O2 is a factor that contributes entirely to the antibacterial effect. In contrast, the second most effective isolate, 05.3162.2 HL-2, was found to produce both acid and H2O2. In summary, one-third of the isolates showed the formation of hydrogen peroxide in combination with significant acid production. This was not the case for the remaining six.

 Genome analysis

The above-described experimental data show that W. paramesenteroides strains exhibited pronounced genetic variations according to RAPD analysis. These go hand in hand with differences in antibacterial efficacy, acidification, and H2O2 production. To gain more information on the genomic plasticity, we analyzed the available genomes of W. paramesenteroides. Therefore, only completed genome assemblies from the NCBI genome database were included (Table 4). The key data show a variation in genome length from 1.947.910 to 2.149.463 base pairs and in annotated genes ranging from 1,936 to 2,184, indicating a decent genomic plasticity. The EDGAR web server platform was used for genomic comparisons. Since the server limits genome comparisons to five at a time due to complexity, we divided them into two groups (Figure 7). The Venn diagrams show that the two sets shared 1,311 and 1,504 core genes, respectively. Since the genomes have about 2,000 annotated genes, roughly 500 genes per genome can be considered as accessory. To find out the number of unique genes (singletons), each genome was compared to the remaining nine. As shown in Table 4, the number of singletons ranged from 14 to 132, with an average of 52 and a total of 524. This means an additional, unique metabolic capacity between 2% and 6%.

Key data of W. paramesenteroides genomes.

Strain Source Genome (Mbp) Plasmids Genes Singletons Accession
A47_1 Bombyx mori 2.080.060 1 2114 46 GCF_050613345.1
DSM 20288 Dairy 2.149.463 3 2147 77 GCF_051903945.1
FDAARGOS_414 Environment 1.947.910 0 1936 61 GCF_002386265.1
FL3 Fermented pepper 2.024.293 0 2069 46 GCF_028994215.1
JL_5 Moromi (soy sauce) 1.963.850 0 1952 14 GCF_045005335.1
LCW-28 Moromi (soy sauce) 1.990.966 2 1991 52 GCF_045006675.1
MbWp_142 Mulberry wine 2.007.410 1 2031 132 GCF_036327715.1
STCH_BD1 Ensiled sorghum 2.052.536 1 2072 36 GCF_015689215.1
W31 Idli batter 2.033.487 1 2039 27 GCF_030168775.1
WP12 Budu (fish sauce) 2.075.982 2 2,184 33 GCA_052281005.1

Pan-genomes of W. paramesenteroides. Ten completed genomes were divided into two sets for the shown genomic comparisons. The Venn diagrams illustrate the number of genes of the core genomes in the center of each group, surrounded by accessory genes between two to four genomes, and the number of singletons in the periphery. Strain numbering is as follows: 1, A47_1; 2, DSM 20288; 3, LCW-28; 4, STCH_BD1; 5, WP12; 6, MbWp_142; 7, W31; 8, FDAARGOS_414; 9, FL3; 10, JL_5.

Two complex Venn diagrams with overlapping colored regions, numbers, and intersections on a white background. Each diagram features different colored areas labeled with numbers. The left diagram uses numbers 1 to 5, and the right diagram uses numbers 6 to 10. The regions show numerical values indicating the quantities in each intersection zone.

A search for bacteriocin-encoding genes present in the 10 genomes was conducted with the BAGEL4 web server, which provides the most comprehensive database of bacteriocin-encoding genes. No bacteriocin genes were detected. In addition, the peptide sequence of the only so far reported bacteriocin Weissellin A from W. paramesenteroides DX (UniProtKB/Swiss-Prot: B3A0N4.1) was screened for homologous proteins using the NCBI BLASTP server. The 43 amino acids of Weissellin A showed 73% overall sequence identity to leucocin A/sakacin P family class II bacteriocins from L. mesenteroides and other LAB. No homologs were found within the 61 available genomes of W. paramesenteroides strains. This showed that known bacteriocins are not present in W. paramesenteroides.

 Discussion

Strains of W. paramesenteroides are frequently recovered from carbohydrate-rich niches such as raw milk, cheeses, fermented vegetables and cereals, and silages, consistent with their heterofermentative ecology (Fusco et al., 2015). The good growth characteristics of W. paramesenteroides strains, and the ubiquitous occurrence favors the use as probiotics, starters in food fermentation, and as protectives. This study aimed to investigate the antibacterial potential of W. paramesenteroides strains isolated from foremilk and teat canal biofilms. The data display a marked genetic plasticity, broad but varying antibacterial efficacies, differences in medium acidification depending on the confronted indicator strain, and H2O2 production in some isolated strains. Analysis of available genomes indicates that intraspecific genomic fluidity may underlie the observed phenotypic manifestations.

The data of RAPD analyses showed that 38 out of 40 isolates had less than 95% gene fragment pattern consistency and represented subspecies (Nielsen et al., 2014). Two pairs of isolates were clonal. The pair 27.4 5–4 and 27.4 6-1was isolated from foremilk samples of the same farm from cows number five and six. A third isolate, 27.4 8–1, from cow eight of the same farm was quite diverse. The second pair of clonal isolates, 13.7 22A-1 and 13.7 22A-2, was from the same teat canal biofilm of cow 22. However, we also isolated three subspecies, namely 27.8 11A-2, 27.8 11A-5, and 27.8 11A-6, from one teat canal of a cow from another farm. It also accounted for the isolates 20.4 7A-5 and 20.4 7A-6. It demonstrated that several subspecies can be present in one udder. We have recently characterized a set of 32 Pediococcus pentosaceus strains isolated from the same environment, in which data demonstrated that all isolates similarly represented subspecies (Fischer et al., 2025). Such genetic variations have been reported for Lactobacillus plantarum and for a set of more than a 1,000 lactobacilli isolated from raw cheese (Johansson et al., 1995; Rossetti and Giraffa, 2005). Three B. subtilis strains isolated from the sesame-flavored liquor Daqu also showed distinctly different amplification products in RAPD analysis experiments, indicating that similar genetic plasticity may be widespread among bacterial species (Wu et al., 2021).

All strains of W. paramesenteroides were able to inhibit the applied indicator organisms with the exception of B. subtilis. Strains 22.4 7A-9 and 05.3162.2 HL-2 were the best performers against five and three of eight indicators, respectively. In comparison to our previous study on P. pentosaceus, the findings corroborate that W. paramesenteroides and P. pentosaceus are able to defeat a broad range of target species (Fischer et al., 2025). While both species showed similar efficacies against some indicators, that is, P. aeruginosa or L. monocytogenes, W. paramesenteroides strains were clearly more efficient against C. koseri and S. warneri (Fischer et al., 2025). Pabari et al. have studied four other indicators with similar results, demonstrating broad antibacterial efficacies (Pabari et al., 2020). Nevertheless, W. paramesenteroides strains do not distinguish as they also inhibit many beneficial bacteria (Papagianni and Papamichael, 2011). This was shown by applying culture supernatants containing the bacteriocin Weissellin A (Papagianni and Papamichael, 2011).

When each isolate was exposed to each indicator strain, it became obvious that the acidification of the medium depended on the presence of the respective indicator strain. The highest drop in pH was detected when W. paramesenteroides and S. enterica were in co-culture, while almost none or even an increase in pH was found when K. pneumoniae or B. subtilis were applied. Among the W. paramesenteroides isolates, strains 05.3162.2 HL-2 and 22.4 7A-9 were the strongest acid producers. The responses of the indicator species suggest that they have different answers to acid stress (Guan and Liu, 2020; Zhou and Fey, 2020). A study of the transcriptome of B. subtilis cells that were stressed with the weak acids acetate and sorbate revealed that the ureABC operon, encoding urease, was up-regulated (Ter Beek et al., 2015). The enzyme is a major factor in coping with acid stress. It converts urea into two ammonia molecules that absorb protons, yielding ammonium (Ter Beek et al., 2015; Liu et al., 2020; Lin et al., 2022; Subramaniyan et al., 2023). The result is the buffering of acid. Ureases are also operative in the here used indicators K. pneumonia, P. aeruginosa, and S. warneri. Especially, K. pneumoniae strains are strong producers of urease, which is in good agreement with our data. Only one isolate caused medium acidification in co-culture with K. pneumoniae (Lin et al., 2022). The urease-negative indicators S. enterica, S. haemolyticus, and L. monocytogenes were less efficient against acidification. However, it should be noted that acid tolerance responses are multifaceted. Many genes have been identified, such as the synthesis of ammonia by the arginine deiminase pathway, amino acid decarboxylation systems, proton export by F1F0 ATPases, quorum sensing disruption or the involvement of global regulators, such as the carbon catabolite protein CcpA (Titgemeyer and Hillen, 2002; Ter Beek et al., 2015; Guan and Liu, 2020; Zhou and Fey, 2020; Roux et al., 2022; Rotta et al., 2025).

The production of H2O2 and thus the ability to exert oxidative stress was detected in about half of all isolates. The best performing isolate 27.4 7A-9 was not a producer of H2O2, indicating that other antibacterial mechanisms might be superior (Table 3). This raises the question of how the indicator strains might mask the full effect of hydrogen peroxide. The Gram-negative indicators used in this study possess various defense mechanisms to cope with reactive oxygen species. These are beside hydrogen peroxide, superoxide anions and hydroxyl radicals, which are able to primarily oxidize all RNA species, DNA, proteins, and cell membranes (Huang et al., 2013; Seixas et al., 2022). In K. pneumoniae, the catalases KatE, KatG, and peroxidases AhpC and GST neutralize H2O2 (Huang et al., 2013). They are triggered by the global regulator OxyR, which also stimulates the protective formation of biofilms and extracellular polysaccharides (EPS) (Hennequin and Forestier, 2009). This was shown by phenotype analysis of an oxyR deletion mutant, which is sensitive to H2O2 and unable to form biofilms (Hennequin and Forestier, 2009). For S. enterica, it has been shown that the catalases KatE and KatG are crucial for biofilm tolerance to H2O2, as their activity was reduced in the corresponding catalase mutant strains (Hahn et al., 2021). These catalases are also regulated by OxyR, which is itself activated by H2O2 (Pardo-Esté et al., 2018). Regarding C. koseri, defense mechanisms against oxidative stress have not yet been reported. We performed protein alignments with OxyR and KatG from K. pneumoniae and S. enterica, which show protein identities ranging from 83 to 97%, suggesting that C. koseri possesses very similar defense mechanisms. The OxyR regulon is also operative against oxygen stress in P. aeruginosa. Transcriptional analysis of the katA promoter and transcriptomics revealed that the expression of the katA gene, which encodes the major catalase, is mediated by OxyR in the presence of H2O2 (Palma et al., 2004; Heo et al., 2010). In B. subtilis, PerR is the main transcription factor involved in the expression of the catalase katA gene, the peroxidase-encoding ahpCF operon, and mgrA encoding a DNA-protecting protein (Bsat et al., 1996; Seixas et al., 2022). PerR binds as a metalloregulatory protein to ferrous iron (Fe2+) at a regulatory site. Increasing amounts of H2O2 oxidize the regulatory iron atom (Lee and Helmann, 2006). PerR-Fe3+ is released from its binding site, leading to derepression of katA and ahpCF (Seixas et al., 2022). In L. monocytogenes and in staphylococci, PerR regulates katA, ahpCF, and mgrA, as well (Horsburgh et al., 2001; Chen et al., 2006; Ruhland and Reniere, 2019; Cesinger et al., 2021).

The gene pool of W. paramesenteroides was described through genomic comparisons, revealing several hundred accessory genes and up to 6% singletons. The genome fluidity of W. paramesenteroides strains is comparable to that reported in our P. pentosaceus study (Fischer et al., 2025). A study of four genomes of W. confusa and another one, in which three complete and 39 draft genomes of W. paramesenteroides were analyzed, came to corroborating results (Yuan et al., 2021; Wan et al., 2023). Wan et al. (2023) reported the number of 1990 cloud genes that occurred in less than 15% of the W. paramesenteroides genomes.

The presence of bacteriocins from the genus Weissella remains rare (Leong et al., 2013; Fusco et al., 2015). Only one, Weissellin A, from W. paramesenteroides has been reported (Papagianni and Papamichael, 2011). It was able to inhibit many Gram-positive microorganisms but not S. enterica. Since the here examined strains were able to combat S. enterica, the mechanism must be independent of Weissellin A. Weissellin A showed high similarity to leucocin A/sakacin P family class II bacteriocins from L. mesenteroides and other LAB. It was, however, not detected in 61 W. paramesenteroides genomes according to BLASTP protein searches. Wan et al. reported that a pediocin-like bacteriocin is present in many W. paramesenteroides genomes (Wan et al., 2023). They concluded that the gene product may not have an antibacterial function, as strains encoding the pediocin-like gene did not inhibit Micrococcus luteus, which, however, is sensitive to Weissellin A (Wan et al., 2023). Thus, the currently available data suggest that W. paramesenteroides strains do not possess any known bacteriocins.

W. paramesenteroides strains have a repertoire of diverse mechanisms to combat competing bacteria. Our data suggest that the lowering of pH by organic acids is sufficient as long as the target bacterium is susceptible to acid stress. If not, other mechanisms become operative (Pabari et al., 2020; Singh et al., 2024; Thuy et al., 2024; Chaichana et al., 2025; Cui et al., 2025; Rotta et al., 2025). Although we did not sequence our isolates, the observed genome fluidity derived from RAPDs and public genomes of W. paramesenteroides strains points to the presence of strain-specific genes encoding antibacterial activity factors. To uncover these respective genes present in the pan-genome, further experiments will be necessary.

In conclusion, we support the suggestion of Ndagano et al. (2011) that the antibacterial activity is caused by different, synergistic acting mechanisms. It can be considered a multi-barrier system to meet diverse environmental conditions and exposure to diverse microorganisms.

 Data availability statement

The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.

 Ethics statement

Ethical approval was not required for the study involving animals in accordance with the local legislation and institutional requirements because the study falls under Directive 2010/63/EU in accordance with good veterinary practice [Art. 1(5)f, in conjunction with Art. 3(1)].

 Author contributions

SF: Conceptualization, Data curation, Formal analysis, Funding acquisition, Investigation, Methodology, Project administration, Resources, Supervision, Validation, Visualization, Writing – original draft, Writing – review & editing. AE: Data curation, Investigation, Writing – review & editing. LB: Investigation, Writing – review & editing. NM: Supervision, Writing – review & editing. FT: Conceptualization, Funding acquisition, Methodology, Project administration, Resources, Supervision, Validation, Visualization, Writing – original draft, Writing – review & editing.

 Acknowledgments

We would like to thank Till Sander-Titgemeyer for his professional support with graphic design.

 Conflict of interest

The author(s) declared that this work was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

 Generative AI statement

The author(s) declared that Generative AI was not used in the creation of this manuscript.

Any alternative text (alt text) provided alongside figures in this article has been generated by Frontiers with the support of artificial intelligence and reasonable efforts have been made to ensure accuracy, including review by the authors wherever possible. If you identify any issues, please contact us.

 Publisher’s note

All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.

 References Agirman G. Yu K. B. Hsiao E. Y. (2021). Signaling inflammation across the gut-brain axis. Science 374, 1087–1092. doi: 10.1126/science.abi6087, 34822299 Albesharat R. Ehrmann M. A. Korakli M. Yazaji S. Vogel R. F. (2011). Phenotypic and genotypic analyses of lactic acid bacteria in local fermented food, breast milk and faeces of mothers and their babies. Syst. Appl. Microbiol. 34, 148–155. doi: 10.1016/j.syapm.2010.12.001, 21300508 Altschul S. F. Gish W. Miller W. Myers E. W. Lipman D. J. (1990). Basic local alignment search tool. J. Mol. Biol. 215, 403–410. doi: 10.1016/S0022-2836(05)80360-2, 2231712 Ames B. N. McCann J. Yamasaki E. (1975). Methods for detecting carcinogens and mutagens with the salmonella/mammalian-microsome mutagenicity test. Mutat. Res. 31, 347–364. doi: 10.1016/0165-1161(75)90046-1, 768755 Apostolakos I. Paramithiotis S. Mataragas M. (2022). Functional and safety characterization of Weissella paramesenteroides strains isolated from dairy products through whole-genome sequencing and comparative genomics. Dairy 3, 799–813. doi: 10.3390/dairy3040055 Bello S. Gupta R. S. (2025). Genomic discovery of taxon-specific molecular markers for Lactobacillaceae genera. doi: 10.20944/preprints202506.2420.v2 Birch M. Denning D. W. Law D. (1996). Rapid genotyping of Escherichia coli O157 isolates by random amplification of polymorphic DNA. Eur. J. Clin. Microbiol. Infect. Dis. 15, 297–302. doi: 10.1007/BF01695661, 8781880 Bsat N. Chen L. Helmann J. D. (1996). Mutation of the Bacillus subtilis alkyl hydroperoxide reductase (ahpCF) operon reveals compensatory interactions among hydrogen peroxide stress genes. J. Bacteriol. 178, 6579–6586. doi: 10.1128/jb.178.22.6579-6586.1996, 8932315 Cesinger M. R. Schwardt N. H. Halsey C. R. Thomason M. K. Reniere M. L. (2021). Investigating the roles of Listeria monocytogenes peroxidases in growth and virulence. Microbiol Spectr 9, e00440–e00421. doi: 10.1128/Spectrum.00440-21, 34287055 Chaichana N. Boonsan J. Singkhamanan K. Wonglapsuwan M. Pomwised R. Yaikhan T. . (2025). Genomic and phenotypic characterization of Weissella paramesenteroides WP12 from fermented fish sauce (Budu) reveals promising strain for food fermentation and probiotic applications. LWT 215:117197. doi: 10.1016/j.lwt.2024.117197 Chen P. R. Bae T. Williams W. A. Duguid E. M. Rice P. A. Schneewind O. . (2006). An oxidation-sensing mechanism is used by the global regulator MgrA in Staphylococcus aureus. Nat. Chem. Biol. 2, 591–595. doi: 10.1038/nchembio820, 16980961 Crameri F. (2018). Geodynamic diagnostics, scientific visualisation and StagLab 3.0. Geosci. Model Dev. 11, 2541–2562. doi: 10.5194/gmd-11-2541-2018 Cui J. Xu Z. Yu Z. Zhang Q. Liu S. Du B. . (2025). High-alcohol-producing Klebsiella pneumoniae aggravates lung injury by affecting neutrophils and the airway epithelium. Cell Rep. Med. 6:101886. doi: 10.1016/j.xcrm.2024.101886, 39753141 Dieckmann M. A. Beyvers S. Nkouamedjo-Fankep R. C. Hanel P. H. G. Jelonek L. Blom J. . (2021). EDGAR3.0: comparative genomics and phylogenomics on a scalable infrastructure. Nucleic Acids Res. 49, W185–W192. doi: 10.1093/nar/gkab341, 33988716 Fanelli F. Montemurro M. Chieffi D. Cho G.-S. Franz C. M. A. P. Dell’Aquila A. . (2022). Novel insights into the phylogeny and biotechnological potential of Weissella species. Front. Microbiol. 13:914036. doi: 10.3389/fmicb.2022.914036 Fessard A. Remize F. (2017). Why are Weissella spp. not used as commercial starter cultures for food fermentation? Fermentation 3:38. doi: 10.3390/fermentation3030038 Fischer S. W. Mariani Corea N. Euler A. Bertels L. Titgemeyer F. (2025). Comparative analyses of Pediococcus pentosaceus strains isolated from Milk cattle reveal new insights for screening food-protective cultures. Microorganisms 13:2244. doi: 10.3390/microorganisms13102244, 41156704 Fischer S. W. Titgemeyer F. (2023). Protective cultures in food products: from science to market. Foods 12:1541. doi: 10.3390/foods12071541, 37048362 Fusco V. Chieffi D. Fanelli F. Montemurro M. Rizzello C. G. Franz C. M. A. P. (2023). The Weissella and Periweissella genera: up-to-date taxonomy, ecology, safety, biotechnological, and probiotic potential. Front. Microbiol. 14:1289937. doi: 10.3389/fmicb.2023.1289937, 38169702 Fusco V. Quero G. M. Cho G.-S. Kabisch J. Meske D. Neve H. . (2015). The genus Weissella: taxonomy, ecology and biotechnological potential. Front. Microbiol. 6:155. doi: 10.3389/fmicb.2015.00155, 25852652 Ghazi F. Kihal M. Altay N. Gürakan G. C. (2016). Comparison of RAPD-PCR and PFGE analysis for the typing of Streptococcus thermophilus strains isolated from traditional Turkish yogurts. Ann. Microbiol. 66, 1013–1026. doi: 10.1007/s13213-015-1185-z Göker M. Christensen H. Fingerle V. Kostovski M. Margos G. Moore E. R. B. . (2025). List of recommended names for bacteria of medical importance: report of the ad hoc committee on mitigating changes in prokaryotic nomenclature. Int. J. Syst. Evol. Microbiol. 75:006943. doi: 10.1099/ijsem.0.006943, 41129200 Guan N. Liu L. (2020). Microbial response to acid stress: mechanisms and applications. Appl. Microbiol. Biotechnol. 104, 51–65. doi: 10.1007/s00253-019-10226-1, 31773206 Hahn M. M. González J. F. Gunn J. S. (2021). Salmonella biofilms tolerate hydrogen peroxide by a combination of extracellular polymeric substance barrier function and catalase enzymes. Front. Cell. Infect. Microbiol. 11:683081. doi: 10.3389/fcimb.2021.683081, 34095002 Hennequin C. Forestier C. (2009). oxy0052, a LysR-type regulator involved in Klebsiella pneumoniae mucosal and abiotic colonization. Infect. Immun. 77, 5449–5457. doi: 10.1128/IAI.00837-09, 19786563 Heo Y.-J. Chung I.-Y. Cho W.-J. Lee B.-Y. Kim J.-H. Choi K.-H. . (2010). The major catalase gene (katA) of Pseudomonas aeruginosa PA14 is under both positive and negative control of the global Transactivator OxyR in response to hydrogen peroxide. J. Bacteriol. 192, 381–390. doi: 10.1128/JB.00980-09, 19933365 Hilton A. C. Mortiboy D. Banks J. G. Penn C. W. (1997). RAPD analysis of environmental, food and clinical isolates of Campylobacter spp. FEMS Immunol. Med. Microbiol. 18, 119–124. doi: 10.1111/j.1574-695X.1997.tb01036.x, 9223616 Horsburgh M. J. Clements M. O. Crossley H. Ingham E. Foster S. J. (2001). PerR controls oxidative stress resistance and Iron storage proteins and is required for virulence in Staphylococcus aureus. Infect. Immun. 69, 3744–3754. doi: 10.1128/IAI.69.6.3744-3754.2001, 11349039 Huang C.-J. Wang Z.-C. Huang H.-Y. Huang H.-D. Peng H.-L. (2013). YjcC, a c-di-GMP phosphodiesterase protein, regulates the oxidative stress response and virulence of Klebsiella pneumoniae CG43. PLoS One 8:e66740. doi: 10.1371/journal.pone.0066740, 23935824 Johansson M.-L. Quednau M. Molin G. Ahrné S. (1995). Randomly amplified polymorphic DNA (RAPD) for rapid typing of Lactobacillus plantarum strains. Lett. Appl. Microbiol. 21, 155–159. doi: 10.1111/j.1472-765X.1995.tb01030.x, 7576499 Judicial Commission of the International Committee on Systematics of Prokaryotes (2005). The type species of the genus Salmonella Lignieres 1900 is Salmonella enterica (ex Kauffmann and Edwards 1952) Le minor and Popoff 1987, with the type strain LT2T, and conservation of the epithet enterica in Salmonella enterica over all earlier epithets that may be applied to this species. Opinion 80. Int. J. Syst. Evol. Microbiol. 55, 519–520. doi: 10.1099/ijs.0.63579-0, 15653929 Kang M.-S. Lim H.-S. Kim S.-M. Lee H.-C. Oh J.-S. (2011). Effect of Weissella cibaria on Fusobacterium nucleatum -induced interleukin-6 and interleukin-8 production in KB cells. J. Bacteriol. Virol. 41:9. doi: 10.4167/jbv.2011.41.1.9 Lee J.-W. Helmann J. D. (2006). The PerR transcription factor senses H2O2 by metal-catalysed histidine oxidation. Nature 440, 363–367. doi: 10.1038/nature04537, 16541078 Leong K.-H. Chen Y.-S. Lin Y.-H. Pan S.-F. Yu B. Wu H.-C. . (2013). Weissellicin L, a novel bacteriocin from sian-sianzih -isolated Weissella hellenica 4-7. J. Appl. Microbiol. 115, 70–76. doi: 10.1111/jam.12218, 23594122 Lin W.-F. Hu R.-Y. Chang H.-Y. Lin F.-Y. Kuo C.-H. Su L.-H. . (2022). The role of urease in the acid stress response and fimbriae expression in Klebsiella pneumoniae CG43. J. Microbiol. Immunol. Infect. 55, 620–633. doi: 10.1016/j.jmii.2022.02.002, 35367140 Liu C. Zhao X. Xie H. Zhang X. Li K. Ma C. . (2020). Whole genome sequence and comparative genome analyses of multi-resistant Staphylococcus warneri GD01 isolated from a diseased pig in China. PLoS One 15:e0233363. doi: 10.1371/journal.pone.0233363, 32442199 Mariani Corea N. Titgemeyer F. Wachtarczyk J. Meyer F. Fischer S. W. (2025). Complete genome sequence of Pediococcus pentosaceus 13.7 2A-1 isolated from a Holstein Friesian dairy cattle. Microbiol. Res. Announc. 14, e00848–e00825. doi: 10.1128/mra.00848-25, 41036962 Martín R. Heilig H. G. H. J. Zoetendal E. G. Jiménez E. Fernández L. Smidt H. . (2007b). Cultivation-independent assessment of the bacterial diversity of breast milk among healthy women. Res. Microbiol. 158, 31–37. doi: 10.1016/j.resmic.2006.11.004, 17224259 Martín R. Heilig G. H. J. Zoetendal E. G. Smidt H. Rodríguez J. M. (2007a). Diversity of the Lactobacillus group in breast milk and vagina of healthy women and potential role in the colonization of the infant gut: diversity of the Lactobacillus group in breast milk. J. Appl. Microbiol. 103, 2638–2644. doi: 10.1111/j.1365-2672.2007.03497.x, 18045446 Ndagano D. Lamoureux T. Dortu C. Vandermoten S. Thonart P. (2011). Antifungal activity of 2 lactic acid Bacteria of the Weissella genus isolated from food. J. Food Sci. 76, M305–M311. doi: 10.1111/j.1750-3841.2011.02257.x, 21729073 Nielsen K. L. Godfrey P. A. Stegger M. Andersen P. S. Feldgarden M. Frimodt-Møller N. (2014). Selection of unique Escherichia coli clones by random amplified polymorphic DNA (RAPD): evaluation by whole genome sequencing. J. Microbiol. Methods 103, 101–103. doi: 10.1016/j.mimet.2014.05.018, 24912108 Oksanen J. Simpson G. L. Blanchet F. G. Kindt R. Legendre P. Minchin P. R. 2022 vegan: Community Ecology Package https://CRAN.R-project.org/package=vegan Onur M. Önlü H. (2023). Isolation, characterization of Weissella confusa and Lactococcus lactis from different milk sources and determination of probiotic features. Braz. J. Microbiol. 55, 663–679. doi: 10.1007/s42770-023-01208-7, 38158467 Pabari K. Pithva S. Kothari C. Purama R. K. Kondepudi K. K. Vyas B. R. M. . (2020). Evaluation of probiotic properties and prebiotic utilization potential of Weissella paramesenteroides isolated from fruits. Probiotics Antimicrob. Proteins 12, 1126–1138. doi: 10.1007/s12602-019-09630-w, 31942681 Palma M. DeLuca D. Worgall S. Quadri L. E. N. (2004). Transcriptome analysis of the response of Pseudomonas aeruginosa to hydrogen peroxide. J. Bacteriol. 186, 248–252. doi: 10.1128/JB.186.1.248-252.2004, 14679246 Papagianni M. Papamichael E. M. (2011). Purification, amino acid sequence and characterization of the class IIa bacteriocin weissellin a, produced by Weissella paramesenteroides DX. Bioresour. Technol. 102, 6730–6734. doi: 10.1016/j.biortech.2011.03.106, 21511463 Pardo-Esté C. Hidalgo A. A. Aguirre C. Briones A. C. Cabezas C. E. Castro-Severyn J. . (2018). The ArcAB two-component regulatory system promotes resistance to reactive oxygen species and systemic infection by Salmonella Typhimurium. PLoS One 13:e0203497. doi: 10.1371/journal.pone.0203497, 30180204 Qiao N. Bechtner J. Cnockaert M. Depoorter E. Díaz-Muñoz C. Vandamme P. . (2023). Comparative genomic analysis of Periweissella and the characterization of novel motile species. Appl. Environ. Microbiol. 89, e0103423–e0103423. doi: 10.1128/aem.01034-23, 37728921 R Core Team (2021) R: a language and environment for statistical computing. Available online at: https://www.R-project.org/ Rossetti L. Giraffa G. (2005). Rapid identification of dairy lactic acid bacteria by M13-generated, RAPD-PCR fingerprint databases. J. Microbiol. Methods 63, 135–144. doi: 10.1016/j.mimet.2005.03.001, 15893395 Rotta I. S. Rezende S. D. D. C. Perini H. F. Da Silva M. V. De Almeida F. A. Pinto U. M. . (2025). Exploring the potential of Weissella paramesenteroides UFTM 2.6.1 in disrupting quorum sensing and attenuating virulence in Listeria monocytogenes. Front. Microbiol. 16:1601203. doi: 10.3389/fmicb.2025.1601203, 40606152 Roux A.-E. Robert S. Bastat M. Rosinski-Chupin I. Rong V. Holbert S. . (2022). The role of regulator catabolite control protein a (CcpA) in Streptococcus agalactiae physiology and stress response. Microbiol Spectr 10, e02080–e02022. doi: 10.1128/spectrum.02080-22, 36264242 RStudio Team (2022) RStudio: integrated development environment for R. Available online at: http://www.rstudio.com/ Ruhland B. R. Reniere M. L. (2019). Sense and sensor ability: redox-responsive regulators in Listeria monocytogenes. Curr. Opin. Microbiol. 47, 20–25. doi: 10.1016/j.mib.2018.10.006, 30412828 Ruiz P. Seseña S. Palop M. L. (2014). A comparative study of different PCR-based DNA fingerprinting techniques for typing of lactic acid bacteria. Eur. Food Res. Technol. 239, 87–98. doi: 10.1007/s00217-014-2197-9 Schwengers O. Jelonek L. Dieckmann M. A. Beyvers S. Blom J. Goesmann A. (2021). Bakta: rapid and standardized annotation of bacterial genomes via alignment-free sequence identification: find out more about Bakta, the motivation, challenges and applications, here. Microb. Genomics 7:000685. doi: 10.1099/mgen.0.000685 Seixas A. F. Quendera A. P. Sousa J. P. Silva A. F. Q. Arraiano C. M. Andrade J. M. (2022). Bacterial response to oxidative stress and RNA oxidation. Front. Genet. 12:821535. doi: 10.3389/fgene.2021.821535, 35082839 Silva C. C. G. Silva S. P. M. Ribeiro S. C. (2018). Application of Bacteriocins and protective cultures in dairy food preservation. Front. Microbiol. 9:594. doi: 10.3389/fmicb.2018.00594, 29686652 Singh J. K. Devi P. B. Reddy G. B. Jaiswal A. K. Kavitake D. Shetty P. H. (2024). Biosynthesis, classification, properties, and applications of Weissella bacteriocins. Front. Microbiol. 15:1406904. doi: 10.3389/fmicb.2024.1406904, 38939182 Subramaniyan Y. Khan A. Fathima F. Rekha P. D. (2023). Differential expression of urease genes and ureolytic activity of uropathogenic Escherichia coli and Pseudomonas aeruginosa isolates in different nutritional conditions. Arch. Microbiol. 205:383. doi: 10.1007/s00203-023-03722-6, 37973630 Szaluś-Jordanow O. Krysztopa-Grzybowska K. Czopowicz M. Moroz A. Mickiewicz M. Lutyńska A. . (2018). MLST and RAPD molecular analysis of Staphylococcus aureus subsp. anaerobius isolated from goats in Poland. Arch. Microbiol. 200, 1407–1410. doi: 10.1007/s00203-018-1568-1, 30182255 Teixeira C. G. Fusieger A. Milião G. L. Martins E. Drider D. Nero L. A. . (2021a). Weissella: an emerging bacterium with promising health benefits. Probiotics Antimicrob. Proteins 13, 915–925. doi: 10.1007/s12602-021-09751-1, 33565028 Teixeira C. G. Silva R. R. D. Fusieger A. Martins E. Freitas R. D. Carvalho A. F. D. (2021b). O gênero Weissella na indústria de alimentos: Uma revisão. RSD 10:e8310514557. doi: 10.33448/rsd-v10i5.14557 Ter Beek A. Wijman J. G. E. Zakrzewska A. Orij R. Smits G. J. Brul S. (2015). Comparative physiological and transcriptional analysis of weak organic acid stress in Bacillus subtilis. Food Microbiol. 45, 71–82. doi: 10.1016/j.fm.2014.02.013, 25481064 Thuy T. T. D. Lu H.-F. Bregente C. J. B. Huang F.-C. A. Tu P.-C. Kao C.-Y. (2024). Characterization of the broad-spectrum antibacterial activity of bacteriocin-like inhibitory substance-producing probiotics isolated from fermented foods. BMC Microbiol. 24:85. doi: 10.1186/s12866-024-03245-0, 38468236 Titgemeyer F. Hillen W. (2002). Global control of sugar metabolism: a gram-positive solution. Antonie Van Leeuwenhoek 82, 59–71. doi: 10.1023/A:1020628909429, 12369205 Tomás M. S. J. Claudia Otero M. Ocaña V. Elena Nader-Macías M. (2004). “Production of antimicrobial substances by lactic acid Bacteria I: determination of hydrogen peroxide” in Methods in molecular biology. ed. Spencer J. F. T. (New Jersey: Humana Press), 337–346. van Heel A. J. de Jong A. Song C. Viel J. H. Kok J. Kuipers O. P. (2018). BAGEL4: a user-friendly web server to thoroughly mine RiPPs and bacteriocins. Nucleic Acids Res. 46, W278–W281. doi: 10.1093/nar/gky383, 29788290 Walter J. Hertel C. Tannock G. W. Lis C. M. Munro K. Hammes W. P. (2001). Detection of Lactobacillus, Pediococcus, Leuconostoc, and Weissella species in human feces by using group-specific PCR primers and denaturing gradient gel electrophoresis. Appl. Environ. Microbiol. 67, 2578–2585. doi: 10.1128/AEM.67.6.2578-2585.2001, 11375166 Wan X. Takala T. M. Huynh V. A. Ahonen S. L. Paulin L. Björkroth J. . (2023). Comparative genomics of 40 Weissella paramesenteroides strains. Front. Microbiol. 14:1128028. doi: 10.3389/fmicb.2023.1128028, 37065164 Wickham H. Averick M. Bryan J. Chang W. McGowan L. D. François R. . (2019). Welcome to the tidyverse. J. Open Source Softw. 4:1686. doi: 10.21105/joss.01686 Wickham H. Bryan J. (2022) Readxl: read excel files. Avaliable online at: https://CRAN.R-project.org/package=readxl Wu X. Jiang Q. Wang Z. Xu Y. Chen W. Sun J. . (2021). Diversity, enzyme production and antibacterial activity of Bacillus strains isolated from sesame-flavored liquor Daqu. Arch. Microbiol. 203, 5831–5839. doi: 10.1007/s00203-021-02552-8, 34491390 Xiong D. Liu M. Sun T. Ma Z. Sun Y. Yuan L. . (2025). Weissella paramesenteroides LAB14 mitigates the inflammation elicited by lipopolysaccharide and Salmonella through the suppression of the NF-κB and IRF pathways. Food Biosci. 68:106428. doi: 10.1016/j.fbio.2025.106428 Yuan S. Wang Y. Zhao F. Kang L. (2021). Complete genome sequence of Weissella confusa LM1 and comparative genomic analysis. Front. Microbiol. 12:749218. doi: 10.3389/fmicb.2021.749218, 34650545 Zheng J. Wittouck S. Salvetti E. Franz C. M. A. P. Harris H. M. B. Mattarelli P. . (2020). A taxonomic note on the genus Lactobacillus: description of 23 novel genera, emended description of the genus Lactobacillus Beijerinck 1901, and union of Lactobacillaceae and Leuconostocaceae. Int. J. Syst. Evol. Microbiol. 70, 2782–2858. doi: 10.1099/ijsem.0.004107, 32293557 Zhou C. Fey P. D. (2020). The acid response network of Staphylococcus aureus. Curr. Opin. Microbiol. 55, 67–73. doi: 10.1016/j.mib.2020.03.006, 32361405

Edited by: Vincenzina Fusco, National Research Council (CNR), Italy

Reviewed by: Malgorzata Ziarno, Warsaw University of Life Sciences, Poland

Nanzhen Qiao, University of Alberta, Canada