Innate immune response to intramammary infection with Serratia marcescens and Streptococcus uberis

Streptococcus uberis and Serratia marcescens are Gram-positive and Gram-negative bacteria, respectively, that induce clinical mastitis. Once initial host barrier systems have been breached by these pathogens, the innate immune system provides the next level of defense against these infectious agents. The innate immune response is characterized by the induction of pro-inflammatory cytokines, as well as increases in other accessory proteins that facilitate host recognition and elimination of the pathogens. The objective of the current study was to characterize the innate immune response during clinical mastitis elicited by these two important, yet less well-studied, Gram-positive and Gram-negative organisms. The pro-inflammatory cytokine response and changes in the levels of the innate immune accessory recognition proteins, soluble CD14 (sCD14) and lipopolysaccharide (LPS)-binding protein (LBP), were studied. Decreased milk output, induction of a febrile response, and increased acute phase synthesis of LBP were all characteristic of the systemic response to intramammary infection with either organism. Infection with either bacteria similarly resulted in increased milk levels of IL-1$\beta$, IL-8, IL-10, IL-12, IFN-$\gamma$, TNF-$\alpha$, sCD14, LBP, and the complement component, C5a. However, the duration of and/or maximal changes in the increased levels of these inflammatory markers were significantly different for several of the inflammatory parameters assayed. In particular, S. uberis infection was characterized by the sustained elevation of higher milk levels of IL-1$\beta$, IL-10, IL-12, IFN-$\gamma$, and C5a, relative to S. marcescens infection. Together, these data demonstrate the variability of the innate immune response to two distinct mastitis pathogens

Topological organization of multichromosomal regions by the long intergenic noncoding RNA Firre

RNA, including long noncoding RNA (lncRNA), is known to be an abundant and important structural component of the nuclear matrix. However, the molecular identities, functional roles and localization dynamics of lncRNAs that influence nuclear architecture remain poorly understood. Here, we describe one lncRNA, Firre, that interacts with the nuclear-matrix factor hnRNPU through a 156-bp repeating sequence and localizes across an ~5-Mb domain on the X chromosome. We further observed Firre localization across five distinct trans-chromosomal loci, which reside in spatial proximity to the Firre genomic locus on the X chromosome. Both genetic deletion of the Firre locus and knockdown of hnRNPU resulted in loss of colocalization of these trans-chromosomal interacting loci. Thus, our data suggest a model in which lncRNAs such as Firre can interface with and modulate nuclear architecture across chromosomes

Expression of lymphocyte homing and adhesion molecules during intramammary infection of cows with Serratia marcescens or Streptococcus uberis: correlation with bacterial colonization and clinical signs

We wished to determine the expression of trafficking/adhesion molecules on the surface of lymphocytes isolated from infected mammary glands of cows challenged with either Serratia marcescens or Staphylococcus uberis. Healthy Holstein cows in mid lactation were infected by intramammary infusion with S. marcescens or S. uberis. Following infection, milk samples were collected at various time points. Body temperatures of the cows were taken, and milk was analyzed for colony forming units (CFU) of bacteria and somatic cell counts (SCC). Leukocytes were isolated from the milk and analyzed by flow cytometry. Percentages and types of lymphocytes were determined as well as expression of CD62L, CD11a, LPAM-1 and CD44 on these cells. We found that the percentage of lymphocytes expressing either CD62L or CD11a showed a marked increase 12 h post infection (PI) with S. marcescens that was not seen in cows infected with S. uberis. Conversely, the percentage of lymphocytes expressing CD44 increased in cows infected with S. uberis at 12 h PI, but the increase was not seen in cows infected with S. marcescens. Expression of LPAM-1 was low at all time points in both groups of cows. Body temperatures became elevated in both groups of cows, peaking at 24 h PI in S. marcescens-infected cows and dropping thereafter. In contrast, temperatures of S. uberis-infected cows continued to rise and were still elevated 96 h PI. CFU of bacteria isolated from mammary glands of S. marcescens-infected cows dropped precipitously 24 h PI but continued at high levels in S. uberis-infected cows. SCC began falling in S. marcescens-infected cows 48 h PI but continued to increase in S. uberis-infected cows. Thus, a greater percentage of lymphocytes in milk had a phenotype consistent with recruitment from the peripheral pool following infection with S. marcescens than was seen following infection with S. uberis. Concurrent with the increases seen in percentages of this lymphocyte phenotype, clinical signs lessened in the S. marcescens-infected cows. Published by Elsevier B.V

Evaluation of breed-dependent differences in the innate immune responses of Holstein and Jersey cows to Staphylococcus aureus intramammary infection

Mastitis is one of the most prevalent diseases of cattle. Various studies have reported breed-dependent differences in the risk for developing this disease. Among two major breeds, Jersey cows have been identified as having a lower prevalence of mastitis than Holstein cows. It is well established that the nature of the initial innate immune response to infection influences the ability of the host to clear harmful bacterial pathogens. Whether differences in the innate immune response to intramammary infections explain, in part, the differential prevalence of mastitis in Holstein and Jersey cows remains unknown. The objective of the current study was to evaluate several parameters of the innate immune response of Holstein and Jersey cows to intramammary infection with Staphylococcus aureus, a common mastitis-inducing pathogen. To control for non-breed related factors that could influence these parameters, all cows were of the same parity, in similar stages of milk production, housed and managed under identical conditions, and experimentally infected and sampled in parallel. The following parameters of the innate immune response were evaluated: acute phase protein synthesis of serum amyloid A and lipopolysaccharide-binding protein; total and differential circulating white blood cell counts; milk somatic cell counts; mammary vascular permeability; milk N-acetyl-beta-D-glucosaminidase (NAGase) activity; and production of the cytokines, interferon (IFN)-gamma, interleukin (IL)-12, tumour growth factor(TGF)-alpha, and TGF-beta 1. The temporal response of all of these parameters following infection was similar between Holstein and Jersey cows. Further, with the exception of changes in circulating neutrophils and NAGase activity, the overall magnitude of these parameters were also comparable. Together, these data demonstrate that the innate immune response of Holstein and Jersey cows to Staph. aureus intramammary infection remains highly conserved despite previously reported differences in mastitis prevalence, as well as genotypic and phenotypic traits, that exist between the two breeds

Characterization of Treponema phagedenis-Like Spirochetes Isolated from Papillomatous Digital Dermatitis Lesions in Dairy Cattle

Four spirochete strains were isolated from papillomatous digital dermatitis (PDD) lesions in Iowa dairy cattle and compared with two previously described spirochete strains isolated from dairy cattle in California. These six strains shared an identical 16S ribosomal DNA sequence that was 98% similar to Treponema phagedenis and 99% similar to the uncultivated PDD spirochete sequence DDLK-4. The whole-cell protein profiles resolved by sodium dodecyl sulfate-polyacrylamide gel electrophoresis of these six strains were similar. However, these strains showed differences in the antigenic diversity of lipopolysaccharide (LPS). Genetic diversity was also detected by pulsed-field gel electrophoresis of genomic DNA digests, revealing differences among five of the six strains. Serum immunoglobulin G antibodies from dairy cattle with active PDD lesions reacted with the LPS of all but one PDD spirochete strain. Likewise, peripheral blood mononuclear cells from cattle with active PDD lesions produced blastogenic responses to one of the two California isolates. Both antibody and lymphocyte blastogenic responses were reduced in convalescent dairy cattle, suggesting the immune response to these spirochetes has short duration. These results demonstrate genetic and antigenic diversity among T. phagedenis-like treponemes and provide further evidence for the involvement of these spirochetes in the pathogenesis of PDD