Bifidobacterium Infantis 35624 Protects Against Salmonella-Induced Reductions in Digestive Enzyme Activity in Mice by Attenuation of the Host Inflammatory Response

OBJECTIVES: Salmonella-induced damage to the small intestine may decrease the villi-associated enzyme activity, causing malabsorption of nutrients and diarrhea, and thus contribute to the symptoms of infection. The objective of this study was to determine the mechanism by which different doses and durations of Salmonella infection and lipopolysaccharide (LPS) affect brush border enzyme activity in the mouse, and to determine if the probiotic Bifidobacterium longum subspecies infantis 35624 could attenuate the intestinal damage. METHODS: BALB/c mice were challenged with Salmonella enterica serovar Typhimurium UK1 at various doses (10(2)-10(8) colony-forming unit (CFU)) and durations (10(6) CFU for 1-6 days). Mice were also treated with B. longum subsp. infantis 35624 for 2 weeks before and during a 6-day S. Typhimurium challenge (10(6) CFU), or before injection of LPS. The small intestine was assessed for morphological changes, mRNA expression of cytokines, and activity of the brush border enzymes sucrase-isomaltase, maltase, and alkaline phosphatase. RESULTS: S. Typhimurium infection significantly reduced the activity of all brush border enzymes in a dose- and time-dependent manner (P<0.05). This also occurred following injection of LPS. Pre-treatment with B. longum subsp. infantis 35624 prevented weight loss, protected brush border enzyme activity, reduced the small intestinal damage, and inhibited the increase in interleukin (IL)-10 and IL-8 expression due to Salmonella challenge. CONCLUSIONS: Salmonella infection reduces the small intestinal brush border enzyme activity in mice, with the level of reduction and associated weight loss increasing with dose and duration of infection. B. longum subsp. infantis 35624 treatment attenuated the effect of Salmonella infection on brush border enzyme activity and weight loss, which may be due to modulation of the host immune response

Life on Arginine for Mycoplasma hominis: Clues from Its Minimal Genome and Comparison with Other Human Urogenital Mycoplasmas

Mycoplasma hominis is an opportunistic human mycoplasma. Two other pathogenic human species, M. genitalium and Ureaplasma parvum, reside within the same natural niche as M. hominis: the urogenital tract. These three species have overlapping, but distinct, pathogenic roles. They have minimal genomes and, thus, reduced metabolic capabilities characterized by distinct energy-generating pathways. Analysis of the M. hominis PG21 genome sequence revealed that it is the second smallest genome among self-replicating free living organisms (665,445 bp, 537 coding sequences (CDSs)). Five clusters of genes were predicted to have undergone horizontal gene transfer (HGT) between M. hominis and the phylogenetically distant U. parvum species. We reconstructed M. hominis metabolic pathways from the predicted genes, with particular emphasis on energy-generating pathways. The Embden–Meyerhoff–Parnas pathway was incomplete, with a single enzyme absent. We identified the three proteins constituting the arginine dihydrolase pathway. This pathway was found essential to promote growth in vivo. The predicted presence of dimethylarginine dimethylaminohydrolase suggested that arginine catabolism is more complex than initially described. This enzyme may have been acquired by HGT from non-mollicute bacteria. Comparison of the three minimal mollicute genomes showed that 247 CDSs were common to all three genomes, whereas 220 CDSs were specific to M. hominis, 172 CDSs were specific to M. genitalium, and 280 CDSs were specific to U. parvum. Within these species-specific genes, two major sets of genes could be identified: one including genes involved in various energy-generating pathways, depending on the energy source used (glucose, urea, or arginine) and another involved in cytadherence and virulence. Therefore, a minimal mycoplasma cell, not including cytadherence and virulence-related genes, could be envisaged containing a core genome (247 genes), plus a set of genes required for providing energy. For M. hominis, this set would include 247+9 genes, resulting in a theoretical minimal genome of 256 genes

Effects of the TLR2 Agonists MALP-2 and Pam3Cys in Isolated Mouse Lungs

Background: Gram-positive and Gram-negative bacteria are main causes of pneumonia or acute lung injury. They are recognized by the innate immune system via toll-like receptor-2 (TLR2) or TLR4, respectively. Among all organs, the lungs have the highest expression of TLR2 receptors, but little is known about the pulmonary consequences of their activation. Here we studied the effects of the TLR2/6 agonist MALP-2, the TLR2/1 agonist Pam 3Cys and the TLR4 agonist lipopolysaccharide (LPS) on pro-inflammatory responses in isolated lungs. Methodology/Principal Findings: Isolated perfused mouse lungs were perfused for 60 min or 180 min with MALP-2 (25 ng/ mL), Pam3Cys (160 ng/mL) or LPS (1 mg/mL). We studied mediator release by enzyme linked immunosorbent assay (ELISA), the activation of mitogen activated protein kinase (MAPK) and AKT/protein kinase B by immunoblotting, and gene induction by quantitative polymerase chain reaction. All agonists activated the MAPK ERK1/2 and p38, but neither JNK or AKT kinase. The TLR ligands upregulated the inflammation related genes Tnf, Il1b, Il6, Il10, Il12, Ifng, Cxcl2 (MIP-2a) and Ptgs2. MALP-2 was more potent than Pam 3Cys in inducing Slpi, Cxcl10 (IP10) and Parg. Remarkable was the strong induction of Tnc by MALP2, which was not seen with Pam 3Cys or LPS. The growth factor related genes Areg and Hbegf were not affected. In addition, all three TLR agonists stimulated the release of IL-6, TNF, CXCL2 and CXCL10 protein from the lungs

Lipid-associated membrane proteins of Mycoplasma fermentans and M. penetrans activate human immunodeficiency virus long-terminal repeats through Toll-like receptors

Mycoplasmas are known to enhance human immunodeficiency virus (HIV) replication, and mycoplasma-derived lipid extracts have been reported to activate nuclear factor-κB (NF-κB) through Toll-like receptors (TLRs). In this study, we examined the involvement of TLRs in the activation of HIV long-terminal repeats (LTR) by mycoplasma and their active components responsible for the TLR activation. Lipid-associated membrane proteins (LAMPs) from two species of mycoplasma (Mycoplasma fermentans and M. penetrans) that are associated with acquired immune-deficiency syndrome (AIDS), were found to activate HIV LTRs in a human monocytic cell line, THP-1. NF-κB deletion from the LTR resulted in inhibition of the activation. The LTR activation by M. fermentans LAMPs was inhibited by a dominant negative (DN) construct of TLR1 and TLR6, whereas HIV LTR activation by M. penetrans LAMPs was inhibited by DN TLR1, but not by DN TLR6. These results indicate that the activation of HIV LTRs by M. fermentans and M. penetrans LAMPs is dependent on NF-κB, and that the activation of HIV LTR by M. fermentans LAMPs is mediated through TLR1, TLR2 and TLR6. In contrast, the LTR activation by M. penetrans LAMPs is carried out through TLR1 and TLR2, but not TLR6. Subsequently, the active component of M. penetrans and M. fermentans LAMPs was purified by reverse-phase high-performance liquid chromatography (HPLC). Interestingly, the purified lipoprotein of M. penetrans LAMPs (LPMp) was able to activate NF-κB through TLR1 and TLR2. On the other hand, the activation of NF-κB by purified lipoprotein of M. fermentans LAMPs (LPMf) was mediated through TLR2 and TLR6, but not TLR1

Triacylated lipoproteins derived from Mycoplasma pneumoniae activate nuclear factor-κB through toll-like receptors 1 and 2

Pathogenesis of Mycoplasma pneumoniae infection is considered to be in part attributed to excessive immune responses. Recently, a mycoplasma lipoprotein has been shown to induce nuclear factor κB (NF-κB) activation through toll-like receptor 1 (TLR1), TLR2 and TLR6. In this study, we examined the ability of lipoproteins from M. pneumoniae to activate NF-κB through TLR1- and TLR2-dependent, but TLR6-independent, pathways, and the active components responsible for the NF-κB activation through the TLR6-independent pathway were identified. The active lipoproteins were found to be MPN611 and MPN162 of M. pneumoniae (designated N-ALP1 and N-ALP2, respectively). Purified N-ALP1 and N-ALP2 from M. pneumoniae and triacylated partial synthetic lipopeptides of N-ALP1 and N-ALP2 augmented the levels of NF-κB induction through TLR1- and TLR2-dependent pathways, whereas diacylated partial synthetic lipopeptides of N-ALP1 and N-ALP2 activated NF-κB through TLR1-, TLR2- and TLR6-dependent pathways. These data suggest that N-ALP1 and N-ALP2 would be triacylated lipoproteins. The activity of N-ALP1 and N-ALP2 was decreased with a pretreatment of lipoprotein lipase, and partially decreased by protease treatment, indicating that the lipid moiety of N-ALP1 and N-ALP2 is critical for the NF-κB activation. Thus, triacylated lipoproteins derived from M. pneumoniae might activate NF-κB through TLR1 and TLR2, but not TLR6

The importance of B-cells and ecto-5′nucleotidase in Mycoplasma fermentans infection and the relevance to rheumatoid arthritis

The aim of this work was to discover if Mycoplasma fermentans, which is known to infect B cells, could be the cause of the raised ecto-5′-nucleotidase observed in the synovial fluid of rheumatoid arthritis patients. The ecto-5′-nucleotidase activity in the patients' serum has been shown to correlate with the erythrocyte sedimentation rate and DNA from the mycoplasma has been found in the synovial fluid. B lymphoblastoid cell lines were exposed to 16 strains of Mycoplasma fermentans and their ecto-5′-nucleotidase, CD73, was measured both biochemically and by mouse antibodies to human ecto 5′-nucleotidase using the fluorescence activated cell sorter. The type strain, PG 18, did not grow with the B cells. Some of the mycoplasma strains (9/15) increased the cellular ecto-5′-nucleotidase activity from twice to 17 fold, and usually showed 5′-nucleotidase activity themselves. At least one strain, M106, induced human 5′-nucleotidase on the normally 5′-nucleotidase negative Daudi and Raji Burkitt's lymphoma cell lines, and increased sevenfold the 5′-nucleotidase on the monocyte/macrophage cell line THP-1. Growing the cells in aged medium increased the level of mycoplasma infection. This mycoplasma-induced enzyme showed a conformational change and an increase in activity with a glycosylation change involving mannose groups. The other group of strains, mostly of respiratory or cell culture origin, usually did not have any 5′-nucleotidase of their own and decreased the B-cell enzyme activity by about half. Electron microscopy and flow cytometry showed that the strain M106 was filamentous and could be found inside the B-cells. The 5′-nucleotidase-inducing strains of M. fermentans may be important in the aetiology of rheumatoid arthritis

Kinases of two strains of Mycoplasma hyopneumoniae and a strain of Mycoplasma synoviae: an overview

Mycoplasma synoviae and Mycoplasma hyopneumoniae are wall-less eubacteria belonging to the class of Mollicutes. These prokaryotes have a reduced genome size and reduced biosynthetic machinery. They cause great losses in animal production. M. synoviae is responsible for an upper respiratory tract disease of chickens and turkeys. M. hyopneumoniae is the causative agent of enzootic pneumonia in pigs. The complete genomes of these organisms showed 17 ORFs encoding kinases in M. synoviae and 15 in each of the M. hyopneumoniae strain. Four kinase genes were restricted to the avian pathogen while three were specific to the pig pathogen when compared to each other. All deduced kinases found in the non pathogenic strain (J[ATCC25934]) were also found in the pathogenic M. hyopneumoniae strain. The enzymes were classified in nine families composing five fold groups