In vivo monitoring of Lactiplantibacillus plantarum in the nasal and vaginal mucosa using infrared fluorescence.

Lactic acid bacteria (LAB) of the genus Lactiplantibacillus have been explored as potential mucosal vaccine vectors due to their ability to elicit an immune response against expressed foreign antigens and to their safety. However, tools for monitoring LAB distribution and persistence at the mucosal surfaces are needed. Here, we characterize Lactiplantibacillus plantarum bacteria expressing the infrared fluorescent protein IRFP713 for exploring their in vivo distribution in the mucosa and potential use as a mucosal vaccine vector. This bacterial species is commonly used as a vaginal probiotic and was recently found to have a niche in the human nose. Three different fluorescent L. plantarum strains were obtained using the nisin-inducible pNZRK-IRFP713 plasmid which contains the nisRK genes, showing stable and constitutive expression of IRFP713 in vitro. One of these strains was further monitored in BALB/c mice using near-infrared fluorescence, indicating successful colonization of the nasal and vaginal mucosae for up to 72 h. This study thus provides a tool for the in vivo spatiotemporal monitoring of lactiplantibacilli, allowing non-invasive bacterial detection in these mucosal sites. KEY POINTS: • Stable and constitutive expression of the IRFP713 protein was obtained in different L. plantarum strains. • IRFP713+ L. plantarum 3.12.1 was monitored in vivo using near-infrared fluorescence. • Residence times observed after intranasal and vaginal inoculation were 24-72 h.

A live auxotrophic vaccine confers mucosal immunity and protection against lethal pneumonia caused by Pseudomonas aeruginosa.

Pseudomonas aeruginosa is one of the leading causes of nosocomial pneumonia and its associated mortality. Moreover, extensively drug-resistant high-risk clones are globally widespread, presenting a major challenge to the healthcare systems. Despite this, no vaccine is available against this high-concerning pathogen. Here we tested immunogenicity and protective efficacy of an experimental live vaccine against P. aeruginosa pneumonia, consisting of an auxotrophic strain which lacks the key enzyme involved in D-glutamate biosynthesis, a structural component of the bacterial cell wall. As the amounts of free D-glutamate in vivo are trace substances in most cases, blockage of the cell wall synthesis occurs, compromising the growth of this strain, but not its immunogenic properties. Indeed, when delivered intranasally, this vaccine stimulated production of systemic and mucosal antibodies, induced effector memory, central memory and IL-17A-producing CD4+ T cells, and recruited neutrophils and mononuclear phagocytes into the airway mucosa. A significant improvement in mice survival after lung infection caused by ExoU-producing PAO1 and PA14 strains was observed. Nearly one third of the mice infected with the XDR high-risk clone ST235 were also protected. These findings highlight the potential of this vaccine for the control of acute pneumonia caused by this bacterial pathogen.

Methicillin-resistant Staphylococcus aureus in swine housed indoors in Galicia, Spain / Staphylococcus aureus resistente a la meticilina en cerdos estabulados en Galicia, España

INTRODUCTION: Livestock are known reservoirs of methicillin-resistant Staphylococcus aureus (MRSA) and this constitutes an important public health issue. The prevalence of nasal MRSA carriers in swine housed indoors in Galicia, Spain, was studied. METHODS: 197 samples from swine aged three, eight, 12, 16 and 24 weeks, and from adult pigs, were obtained from four farms. The cleaning procedures implemented to clean the barns and antimicrobial consumption were analyzed. Antimicrobial susceptibility and antimicrobial resistance genes were studied. PFGE, spa typing and MLST were used to classify the isolates. SCCmec, agr and pvl were analyzed. RESULTS: MRSA prevalence was 12.7%. Swine younger than 16 weeks had a higher colonization rate; 22.9% vs 3.5% (OR, 8.16; 95% CI, 2.47-29.79; p < 0.01). The only farm found to be MRSA-free used disinfectants as part of its cleaning procedure. All MRSA were tetracycline-resistant (identifying the tetK and tetM genes), 80% were resistant to erythromycin and clindamycin and 16% were only clindamycin-resistant. The ermC and vgaA genes were identified in these two phenotypes. A single genotype (PFGE type A) and ST398 - spa t011 (84%) and t1451 (16%) were identified. SCCmec type V and agrI were identified in all isolates, and all were pvl-negative. CONCLUSION: A correlation between swine age and MRSA colonization was observed. Appropriate cleaning procedures could have an impact on MRSA colonization in farming. Resistance to antibiotics used in human health was identified. Clinicians should be aware if their patients have come into contact with farm animals

INTRODUCCIÓN: Los animales de granja son reservorios de Staphylococcus aureus resistente a la meticilina (SARM), y constituyen un problema de salud pública. Se estudia la prevalencia de portadores nasales de SARM en cerdos estabulados en Galicia, España. MÉTODOS: En 4 explotaciones se obtuvieron 197 muestras de cerdos con edades en semanas de 3, 8, 12, 16, 24 y adultos. Se analizaron los métodos empleados para limpiar los establos y el consumo de antimicrobianos. Se estudió la resistencia a antimicrobianos, y los genes involucrados en esta. Los aislamientos fueron clasificados mediante PFGE, spa y MLST. Se analizaron SCCmec, agr y pvl. RESULTADOS: La prevalencia de SARM fue del 12,7%. Los cerdos de <16 semanas presentaron las frecuencias de colonización más elevadas 22,9 vs. 3,5% (OR: 8,16; IC 95%: 2,47-29,79; p < 0,01). En la única explotación libre de SARM se empleaban desinfectantes en la limpieza. Todos los SARM fueron resistentes a tetraciclina identificándose los genes tetK y tetM, el 80% fueron resistentes a eritromicina y clindamicina, y el 16% fueron únicamente resistentes a clindamicina. Se identificaron los genes ermC y vgaA en estos 2 fenotipos. Se identificó un único genotipo (PFGE-A) y ST398, siendo spa t011 (84%) y t1451 (16%). En todos los aislamientos se identificó SCCmec V y agrI, siendo estos pvl negativos. CONCLUSIONES: Se observó la asociación entre edad y colonización SARM. La limpieza adecuada podría modificar la colonización por SARM. Se detectaron resistencias a antibióticos empleados en humanos. Los médicos deberían conocer si los pacientes tienen contacto con animales de granja

Methicillin-resistant Staphylococcus aureus in swine housed indoors in Galicia, Spain.

INTRODUCTION: Livestock are known reservoirs of methicillin-resistant Staphylococcus aureus (MRSA) and this constitutes an important public health issue. The prevalence of nasal MRSA carriers in swine housed indoors in Galicia, Spain, was studied. METHODS: 197 samples from swine aged three, eight, 12, 16 and 24 weeks, and from adult pigs, were obtained from four farms. The cleaning procedures implemented to clean the barns and antimicrobial consumption were analyzed. Antimicrobial susceptibility and antimicrobial resistance genes were studied. PFGE, spa typing and MLST were used to classify the isolates. SCCmec, agr and pvl were analyzed. RESULTS: MRSA prevalence was 12.7%. Swine younger than 16 weeks had a higher colonization rate; 22.9% vs 3.5% (OR, 8.16; 95% CI, 2.47-29.79; p<0.01). The only farm found to be MRSA-free used disinfectants as part of its cleaning procedure. All MRSA were tetracycline-resistant (identifying the tetK and tetM genes), 80% were resistant to erythromycin and clindamycin and 16% were only clindamycin-resistant. The ermC and vgaA genes were identified in these two phenotypes. A single genotype (PFGE type A) and ST398 - spa t011 (84%) and t1451 (16%) were identified. SCCmec type V and agrI were identified in all isolates, and all were pvl-negative. CONCLUSION: A correlation between swine age and MRSA colonization was observed. Appropriate cleaning procedures could have an impact on MRSA colonization in farming. Resistance to antibiotics used in human health was identified. Clinicians should be aware if their patients have come into contact with farm animals.