Genome-wide analysis of mammary gland shows modulation of transcriptome landscape with alternative splice variants in Staphylococcus aureus mastitis in mice.

Epidemiological mapping shows Staphylococcus aureus to be the leading mastitis causing pathogen in India with diverse genetic lineages circulating in the dairy cattle population. We previously reported that endemic clonal strains of S. aureus isolated from subclinical mastitis lead to specific alteration of epigenetic modulators resulting in deviating immune response in intramammary infection mouse model. However, the extent of transcriptome modulation and associated alternative splicing in S. aureus mastitis is poorly understood. Hence, to gain a deeper insight of the extent of modulation of transcriptome landscape, we expanded the study here using high throughput, paired-end RNA sequencing analysis of the mouse mammary gland inoculated with three strains of S. aureus (SA1, SA2, and SA3) possessing specific genotype, virulence and enterotoxin traits. Overall, we detected 35,878 transcripts in S. aureus inoculated mammary gland, 23% more than those annotated in the reference genome. Expression of 20,756 transcripts was > 1 fragment per kilobase of transcript per million mapped fragments and 25.95% of multi-exonic genes were alternatively spliced. We noted Alternative Splicing (AS) events for > 100 immune-related genes. S. aureus infection quantitatively altered AS events in mice mammary gland. Collectively, the majority of differentially expressed significant genes clustered into immune-associated, cell adhesion and metabolic process categories. We observed AS events for 379 transcripts of genes putatively encoding several splicing associated proteins and transcription factors besides inflammatory mediators. The present analysis provides new insights into global transcriptome landscape and AS events in host-defense related genes in response to S. aureus intramammary infection, suggesting the need for studies focusing on multi-target and/or network therapeutics approach to combat mastitis.

Streptococcus uberis ST439 and ST475 induce differential inflammatory responses in a mouse intramammary infection model.

Streptococcus uberis causing mastitis is a growing challenge to the dairy industry. Molecular, epidemiological and population structure studies have revealed clonal diversity among the infecting strains. In this study, mouse intramammary infection model was used to uncover the host immune response to two epidemiologically important live strains of S. uberis (SU1and SU2) obtained from subclinical case of mastitis possessing specific and unique multi locus sequence types (ST), pulsed field gel electrophoresis (PFGE) pulsotypes and virulence profiles. Temporal (2h, 4h, 8h, 12h, 24h and 48h) expression of key inflammatory mediators (IL2, IL4, IL6, IL12, TNFα, IFNγ, GMCSF, TLR2, TLR4, TLR9, TLR11, TLR12, CD14, IL1ß, RANTES, Lactoferrin, and CXCl1) by reverse transcription and probe-based quantitative real-time PCR showed relative mRNA levels higher (p<0.05) in response to SU2 compared with SU1 with 24h PI serving as a critical point for the deviating behavior (SU1 versus SU2). Further employing the predicted biological processes under the influence of this pool of tested genes, the delineation of gene regulatory networks suggested SU1-favoring its persistence in the host environment; in contrast, SU2-which elevated gene expression indicating towards pathogen clearance or immune surveillance. This study suggested how these unique strains could manipulate the host immune response to influence the severity of mastitis; our results expand the available information on host pathogen interaction and provide a firm foundation needing further investigations to gain control over this pathogen.

In vivo antibacterial activity and pharmacological properties of the membrane-active glycopeptide antibiotic YV11455.

The membrane-active glycopeptide antibiotic YV11455 is a lipophilic cationic vancomycin analogue that demonstrates rapid and concentration-dependent killing of clinically relevant multidrug-resistant (MDR) Gram-positive bacteria in vitro. YV11455 was 2-fold and 54-270-fold more effective than vancomycin against clinical isolates of vancomycin-sensitive and vancomycin-resistant bacteria, respectively. In this study, the in vivo efficacy, pharmacodynamics, pharmacokinetics and acute toxicology of YV11455 were investigated. In vivo activity and pharmacodynamics were determined in the neutropenic mouse thigh infection model against meticillin-resistant Staphylococcus aureus (MRSA). YV11455 produced dose-dependent reductions in MRSA titres in thigh muscle. When administered intravenously, the 50% effective dose (ED(50)) for YV11455 against MRSA was found to be 3.3 mg/kg body weight, and titres were reduced by up to ca. 3log(10)CFU/g from pre-treatment values at a dosage of 12 mg/kg with single treatment. Single-dose pharmacokinetic studies demonstrated linear kinetics and a prolonged half-life, with an increase in drug exposure (area under the concentration-time curve) compared with vancomycin. The peak plasma concentration following an intravenous dose of 12 mg/kg was 543.5 µg/mL. Acute toxicology studies revealed that YV11455 did not cause any significant alterations in biochemical parameters or histological pictures related to major organs such as the liver and kidney at its pharmacodynamic endpoint (ED(3-log kill)). These findings collectively suggest that YV11455 could be used clinically for the treatment of infections caused by MDR Gram-positive bacteria.