Gamboa, J. | bioRxiv Science

bioRxiv2026

Amphitrophic Listeria monocytogenes causes one-third of invasive listeriosis yet remains undetected by clonal complex-based risk classification

Conventional genomic risk classification of Listeria monocytogenes assigns clonal complexes to hypervirulent (CC1, CC2, CC4, CC6) or hypovirulent (CC9, CC121) categories based on population-level frequency ratios, leaving all remaining diversity in an undifferentiated "intermediate" category that carries no defined risk assessment. We analysed 436 genomes from confirmed invasive listeriosis across 19 countries using multi-dimensional genomic profiling of virulence and persistence determinants and demonstrate that this approach systematically misclassifies a major fraction of clinically relevant L. monocytogenes. Amphitrophic lineages -- carrying simultaneous genomic competence for clinical virulence (functional inlA, mean virulence score 52.7 +/- 6.6) and industrial persistence (SSI-1 in 94.1%, mean persistence score 66.8 +/- 11.6) -- constitute 31.0% of invasive disease, within 3.6 percentage points of the established hypervirulent category (34.6%). Of these 135 amphitrophic clinical isolates, 91.1% were classified as "intermediate" under conventional taxonomy. The five principal amphitrophic CCs (CC8, CC7, CC3, CC5, CC88) appear with indistinguishable dual-fitness genotypes in both clinical and food-chain datasets, establishing that the same organisms persist in processing facilities and cause invasive human disease. Decomposition of the species-level virulence-persistence trade-off (rho = -0.523) by trophic strategy reveals it to be a Simpsons paradox: no within-strategy correlation is significantly negative, and the only significant signal is a positive amphitrophic correlation (rho = +0.221, p = 0.010) indicating synergy rather than trade-off. Multi-dimensional profiling increases risk-stratified detection from 32.3% (conventional) to 65.6% of clinical isolates -- a 103% improvement. These findings demonstrate that clonal complex identity alone leaves one-third of clinically significant L. monocytogenes uncharacterised, and that effective One Health genomic surveillance requires simultaneous assessment of virulence and persistence at the isolate level.

microbiology↗

bioRxiv2026

Amphitrophic Listeria monocytogenes: multi-dimensional genomic profiling reveals a third ecological strategy that challenges the virulence-persistence trade-off paradigm

BackgroundThe virulence-persistence trade-off is considered a fundamental organizing principle of Listeria monocytogenes population biology: hypervirulent clonal complexes dominate clinical cases but are rarely found in processing environments, while hypovirulent lineages dominate industrial niches but are underrepresented in severe disease. However, whether this dichotomy operates as an absolute paradigm has not been quantitatively evaluated at the population scale. Here we develop a multi-dimensional genomic scoring approach that simultaneously quantifies virulence potential (V), environmental persistence capacity (P), clonal epidemiological context (C), and antimicrobial resistance (R) across 903 genomes from four independent datasets spanning five countries, and apply it to test the universality of the trade-off and to characterize the ecological strategies of L. monocytogenes at the population level. MethodsThe scoring approach integrates four components into a composite 0-100 score through empirically calibrated weights (V: 30%, P: 40%, C: 20%, R: 10%). Validation employed 903 L. monocytogenes genomes from four public BioProjects: longitudinal industrial surveillance in Norway (Fagerlund et al. 2022, n = 513, PRJNA689484), retail environments in the United States (Stasiewicz et al. 2015, n = 191, PRJNA245909), clinical-environmental context in China (Wang et al. 2021, n = 151, PRJNA759341), and meat processing in Poland (Kurpas et al. 2020, n = 48, PRJNA629756). ResultsThe composite score achieved excellent discriminatory performance for identifying persistent clones (AUC = 0.933; 95% CI: 0.910-0.954) with perfect specificity (1.000; zero false positives). The inverse V-P correlation was statistically significant across all four datasets (Spearman {rho} from -0.144 to -0.713; p < 0.01), providing the first cross-dataset quantitative confirmation of the trade-off. However, simultaneous evaluation of V-P profiles at the population scale revealed that the species does not conform to a binary dichotomy but rather exhibits three quantitatively distinguishable ecological strategies, for which we propose a functional trophic taxonomy: nosotrophic lineages (22.7%; V > 65, P < 35), specialized in the pathogenic niche; saprotrophic lineages (5.8%; V < 30, P > 45), with irreversible virulence attenuation and industrial specialization; and, as the central finding, amphitrophic lineages (39.1%; V [≥] 35, P [≥] 40), which simultaneously retain functional inlA and stress tolerance determinants (SSI-1) without detectable genomic sacrifice. The three strategies differed significantly (Kruskal-Wallis H = 138.7; p = 7.6 x 10-3{superscript 1}). The correspondence between trophic strategy and CC was predominant but not absolute, demonstrating that this phenotypic classification captures intra-CC functional heterogeneity inaccessible through conventional typing. Furthermore, comparison between genome-based and surveillance-informed classifications revealed that 60 hypervirulent isolates (CC1/CC14), genetically classified as nosotrophic, persisted for up to 8 years in industrial facilities despite lacking any recognized persistence markers -- indicating that their prolonged survival reflects environmental opportunity rather than intrinsic genomic adaptation. ConclusionsMulti-dimensional genomic profiling reveals that the virulence-persistence trade-off, while statistically robust, does not operate as an absolute paradigm. The amphitrophic strategy -- documented here for the first time as a quantitatively distinguishable category encompassing 39.1% of the analyzed population -- challenges the prevailing dichotomous model and identifies a previously unrecognized combined ecological niche. The ability to discriminate between genome-encoded persistence capacity and environmentally facilitated persistence provides a biological framework for understanding the ecological determinants of L. monocytogenes population dynamics in anthropogenic environments.

microbiology↗

bioRxiv2023

Novel microRNAs downregulated in breast cancer tumors bind to the 3'UTR of SNAIL, SLUG, ZEB1 and/or TWIST and decrease metastatic behavior in breast cancer cells

Metastasis, the leading cause of cancer-associated deaths, is promoted by transcription factors SNAIL, SLUG, ZEB1 and TWIST through the activation of epithelial-mesenchymal transition (EMT). MicroRNAs can suppress EMT, emerging as candidate molecular biomarkers and novel therapeutic targets. Herein, we evaluated microRNAs downregulated in breast cancer tissues expressing EMT transcription factors, to find new potential regulators of EMT. MiR-30a, miR-1271, miR-196a, miR-202, miR-210, miR-22, miR-331 and miR-34b were validated. Seven microRNAs downregulated luciferase activity through EMT transcription factors 3UTR, and all microRNAs decreased cell migration, invasion and/or proliferation. In MDA-MB-231 cells, miR-196a and miR-22 decreased endogenous ZEB1 levels, and miR-30a endogenous CCR7 levels. These results suggest that microRNAs studied are novel regulators of EMT through the control of SNAIL, SLUG, ZEB1 and TWIST. They also regulate the metastatic behavior of cancer cells, and may control the development of lymph node metastasis through the regulation of CCR7. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=111 SRC="FIGDIR/small/526978v1_ufig1.gif" ALT="Figure 1"> View larger version (24K): org.highwire.dtl.DTLVardef@cd3eecorg.highwire.dtl.DTLVardef@de10fborg.highwire.dtl.DTLVardef@12d09a7org.highwire.dtl.DTLVardef@1d4dc93_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗