Natural-host animal models indicate functional interchangeability between the filamentous haemagglutinins of Bordetella pertussis and Bordetella bronchiseptica and reveal a role for the mature C-terminal domain, but not the RGD motif, during infection.

Bacteria of the Bordetella genus cause respiratory tract infections. Both broad host range (e.g. Bordetella bronchiseptica) and human-adapted (e.g. Bordetella pertussis) strains produce a surface-exposed and secreted protein called filamentous haemagglutinin (FHA) that functions in adherence and immunomodulation. Previous studies using B. pertussis and cultured mammalian cells identified several FHA domains with potential roles in host cell interactions, including an Arg-Gly-Asp (RGD) triplet that was reported to bind integrins on epithelial cells and monocytes to activate host signalling pathways. We show here that, in contrast to our previous report, the fhaB genes of B. pertussis and B. bronchiseptica are functionally interchangeable, at least with regard to the various in vitro and in vivo assays investigated. This result is significant because it indicates that information obtained studying FHA using B. bronchiseptica and natural-host animal models should apply to B. pertussis FHA as well. We also show that the C-terminus of mature FHA, which we name the MCD, mediates adherence to epithelial and macrophage-like cells and is required for colonization of the rat respiratory tract and modulation of the inflammatory response in mouse lungs. We could not, however, detect a role for the RGD in any of these processes.

Topical treatment with the vitamin D analogue calcipotriol enhances the upregulation of the antimicrobial protein hCAP18/LL-37 during wounding in human skin in vivo.

Cathelicidin antimicrobial protein, hCAP18, is the sole cathelin protein in human. Its C-terminal peptide, which is released enzymatically from the holoprotein, has broad antimicrobial activity but also has effects on eukaryotic cells. hCAP18 is present in leukocytes and is produced at epithelial interfaces as part of the innate immune system. In normal intact skin, there is low constitutive expression of hCAP18, which is rapidly upregulated upon injury. Accumulating evidence indicates that hCAP18/LL-37 may serve a key role in protecting the integrity of the epithelium and also actively promote re-epithelialization and tissue repair. Molecular mechanisms responsible for controlling hCAP18 gene expression in vivo are only partly understood. Vitamin D(3) and its analogue calcipotriol were recently found to directly induce transcription of the hCAP18 gene via functional vitamin D responsive elements in the hCAP18 gene promoter. Skin is the major source for vitamin D(3) in human, where its production is dependent on ultraviolet B (UVB) radiation. We have shown that exposure to UVB, sufficient to produce vitamin D(3), upregulates hCAP18 in human skin in vivo. In the present study, we demonstrate that the upregulation of hCAP18/LL-37 following acute skin injury is further enhanced, at both hCAP18 mRNA and protein levels, after topical treatment with the vitamin D(3) analogue calcipotriol. In chronic ulcers, calcipotriol treatment upregulated hCAP18 mRNA, whereas no consistent upregulation of hCAP18 protein was detected. Our results further support the role of vitamin D(3) as a key physiologic regulator of hCAP18/LL-37 in human skin.

Microarray evaluation of the Listeria monocytogenes infection and amoxicillin treatment in mice.

By using Affymetrix Mouse Genome Arrays and 20 biological replicates per experimental condition, the predictive value of liver and blood gene expression profiles previously identified was validated as predictive of Listeria monocytogenes infection severity (lethal and nonlethal infection). The ability of these genes to predict the outcome of antibiotic treatment was also assessed. Lethally infected BALB/c mice were treated with amoxicillin at 10 or 20 mg/kg; only the higher dose prevented death. The liver genes predicted that 70% of the animals treated at 10 mg/kg, but only 25% of the mice treated at 20 mg/kg, belonged to the lethal infection group, and this prediction was similar to the ultimate mortality outcome. These results confirm the value of microarrays as tools to predict host response to infection and efficacy of antibacterial therapy. These results might lead to applications that would help clinicians to adjust antibiotic dosages for efficient treatment but yet without toxicity.

Breastfeeding and health outcomes for the mother-infant dyad.

Worldwide, breastfeeding saves the lives of infants and reduces their disease burden. Breastfeeding also reduces the disease burden for mothers. This article examines who chooses to breastfeed and for how long in the United States. It also reviews the latest evidence about the consequences of breastfeeding for the health of the infant and mother. This review provides support for current national and international recommendations that support breastfeeding.

Modulation of the host interferon response and ISGylation pathway by B. pertussis filamentous hemagglutinin.

Bordetella pertussis filamentous hemagglutinin (FHA) is a surface-associated and secreted protein that serves as a crucial adherence factor, and displays immunomodulatory activity in human peripheral blood mononuclear cells (PBMCs). In order to appreciate more fully the role of secreted FHA in pathogenesis, we analyzed FHA-induced changes in genome-wide transcript abundance in human PBMCs. Among the 683 known unique genes with greater than 3-fold change in transcript abundance following FHA treatment, 125 (18.3%) were identified as interferon (IFN)-regulated. Among the latter group were genes encoding several members of the IFN type I response, as well as 3 key components of the ISGylation pathway. Using real-time RT-PCR, we confirmed FHA-associated increases in transcript abundance for the genes encoding ubiquitin-like protein, ISG15, and its specific protease USP18. Western-blot analysis demonstrated the presence of both, free ISG15 and several ISGylated conjugates in FHA-stimulated PBMC lysates, but not in unstimulated cells. Intracellular FACS analysis provided evidence that monocytes and a natural killer-enriched cell population were the primary producers of ISG15 in PBMCs after FHA stimulation. Our data reveal previously-unrecognized effects of B. pertussis FHA on host IFN and ISGylation responses, and suggest previously-unsuspected mechanisms by which FHA may alter the outcome of the host-pathogen interaction.

Interventions to increase the duration of breastfeeding in obese mothers: the Bassett Improving Breastfeeding Study.

OBJECTIVE: Maternal obesity is associated with poor breastfeeding outcomes, yet no intervention has been developed to improve them. To ascertain whether increased breastfeeding support or provision of a breast pump is a feasible, effective intervention to improve breastfeeding, we enrolled obese women who intended to breastfeed in two randomized trials. METHODS: In Bassett Improving Breastfeeding Study (BIBS) 1, 40 women received targeted breastfeeding support in the hospital and via telephone or usual care. Information regarding breastfeeding was collected via telephone for 7 days after delivery and at 30 and 90 days postpartum. In BIBS 2, 34 obese mothers received a manual or electric breast pump to use for 10-14 days or no pump; data collection was similar. RESULTS: In both experiments, randomization failed to distribute women of differing postpartum body mass index adequately among the treatment groups. When analyses were adjusted for this, there was no difference in BIBS 1 between targeted and usual care groups and in BIBS 2 among the treatment groups in the proportion of women still breastfeeding at the times studied. CONCLUSIONS: In future studies of obese women, stratified randomization may be necessary. Further development of interventions to help obese women achieve optimal breastfeeding outcomes is required.

Gene expression analysis reveals new possible mechanisms of vancomycin-induced nephrotoxicity and identifies gene markers candidates.

Vancomycin, one of few effective treatments against methicillin-resistant Staphylococcus aureus, is nephrotoxic. The goals of this study were to (1) gain insights into molecular mechanisms of nephrotoxicity at the genomic level, (2) evaluate gene markers of vancomycin-induced kidney injury, and (3) compare gene expression responses after iv and ip administration. Groups of six female BALB/c mice were treated with seven daily iv or ip doses of vancomycin (50, 200, and 400 mg/kg) or saline, and sacrificed on day 8. Clinical chemistry and histopathology demonstrated kidney injury at 400 mg/kg only. Hierarchical clustering analysis revealed that kidney gene expression profiles of all mice treated at 400 mg/kg clustered with those of mice administered 200 mg/kg iv. Transcriptional profiling might thus be more sensitive than current clinical markers for detecting kidney damage, though the profiles can differ with the route of administration. Analysis of transcripts whose expression was changed by at least twofold compared with vehicle saline after high iv and ip doses of vancomycin suggested the possibility of oxidative stress and mitochondrial damage in vancomycin-induced toxicity. In addition, our data showed changes in expression of several transcripts from the complement and inflammatory pathways. Such expression changes were confirmed by relative real-time reverse transcription-polymerase chain reaction. Finally, our results further substantiate the use of gene markers of kidney toxicity such as KIM-1/Havcr1, as indicators of renal injury.

Growth phase- and nutrient limitation-associated transcript abundance regulation in Bordetella pertussis.

To survive in a host environment, microbial pathogens must sense local conditions, including nutrient availability, and adjust their growth state and virulence functions accordingly. No comprehensive investigation of growth phase-related gene regulation in Bordetella pertussis has been reported previously. We characterized changes in genome-wide transcript abundance of B. pertussis as a function of growth phase and availability of glutamate, a key nutrient for this organism. Using a Bordetella DNA microarray, we discovered significant changes in transcript abundance for 861 array elements during the transition from log phase to stationary phase, including declining transcript levels of many virulence factor genes. The responses to glutamate depletion exhibited similarities to the responses induced by exit from log phase, including decreased virulence factor transcript levels. However, only 23% of array elements that showed at least a fourfold growth phase-associated difference in transcript abundance also exhibited glutamate depletion-associated changes, suggesting that nutrient limitation may be one of several interacting factors affecting gene regulation during stationary phase. Transcript abundance patterns of a Bvg+ phase-locked mutant revealed that the BvgAS two-component regulatory system is a key determinant of growth phase- and nutrient limitation-related transcriptional control. Several adhesin genes exhibited lower transcript abundance during stationary phase and under glutamate restriction conditions. The predicted bacterial phenotype was confirmed: adherence to bronchoepithelial cells decreased 3.3- and 4.4-fold at stationary phase and with glutamate deprivation, respectively. Growth phase and nutrient availability may serve as cues by which B. pertussis regulates virulence according to the stage of infection or the location within the human airway.