Strain-dependent proliferation in response to human gastric mucin and adhesion properties of Helicobacter pylori are not affected by co-isolated Lactobacillus sp

Background: Helicobacter pylori colonize the mucus layer that covers the gastric epithelium and can cause gastritis, ulcers, and gastric cancer. Recently, Lactobacillus sp. have also been found to reside in this niche permanently. This study compares adhesive properties and proliferation of co‐isolated lactobacilli and H. pylori in the presence of mucins and investigates possibilities for lactobacilli‐mediated inhibition of H. pylori. Materials and methods: Binding and proliferation of four H. pylori and four Lactobacillus strains, simultaneously isolated after residing in the stomachs of four patients for >4 years, to human gastric mucins were investigated using microtiter‐based methods. Results: The H. pylori strains co‐isolated with lactobacilli exhibited the same mucin binding properties as demonstrated for H. pylori strains previously. In contrast, no binding to mucins was detected with the Lactobacillus strains. Proliferation of mucin‐binding H. pylori strains was stimulated by the presence of mucins, whereas proliferation of non‐binding H. pylori and Lactobacillus strains was unaffected. Associative cultures of co‐isolated H. pylori and Lactobacillus strains showed no inhibition of H. pylori proliferation because of the presence of whole bacteria or supernatant of lactobacilli. Conclusions: The presence of lactobacilli in the stomach did not select for different mucin binding properties of H. pylori, and Lactobacillus sp. did neither compete for binding sites nor inhibit the growth of co‐isolated H. pylori. The effects of human gastric mucins on H. pylori proliferation vary between strains, and the host–bacteria interaction in the mucus niche thus depends on both the H. pylori strain and the microenvironment provided by the host mucins.     

An evaluation of the clinical assessments of under-five febrile children presenting to primary health facilities in rural Ghana

Background

The shift to test-based management of malaria represents an important departure from established practice under the Integrated Management of Childhood Illnesses (IMCI). The possibility of false results of tests for malaria and co-morbidity, however, make it important that guidelines in IMCI case assessment are still followed.

Methods and Findings

We conducted a cross-sectional observational study to evaluate current practices in IMCI-based assessment of febrile children in 10 health centres and 5 district hospitals, with follow up of a subset of children to determine day 7–10 post-treatment clinical outcome. Clinical consultation, examination and prescribing practices were recorded using a checklist by trained non-medical observers. The facility case management of 1,983 under-five years old febrile children was observed and 593 followed up at home on days 5–10. The mean number of tasks performed from the 11 tasks expected to be done by the IMCI guidelines was 6 (SD 1.6). More than 6 tasks were performed in only 35% of children and this varied substantially between health facilities (range 3–85%). All 11 tasks were performed in only 1% of children. The most commonly performed tasks were temperature measurement (91%) and weighing (88%). Respiratory rate was checked in only 4% of children presenting with cough or difficulty in breathing. The likelihood of performing “better than average number of tasks” (>6) was higher when the consultation was done by medical assistants than doctors (O.R. = 3.16, 1.02–9.20). The number of tasks performed during assessment did not, however, influence clinical outcome (O.R. = 1.02, 0.83–1.24).

Conclusion

Facility-tailored interventions are needed to improve adherence to IMCI guidelines incorporating test-based management of malaria. Studies are needed to re-evaluate the continued validity of tasks defined in IMCI case assessment guidelines.     

Acceptability of Rapid Diagnostic Test-Based Management of Malaria among Caregivers of Under-Five Children in Rural Ghana

Introduction

WHO now recommends test-based management of malaria (TBMM) across all age-groups. This implies artemisinin-based combination treatment (ACT) should be restricted to rapid diagnostic test (RDT)-positive cases. This is a departure from what caregivers in rural communities have been used to for many years.

Methods

We conducted a survey among caregivers living close to 32 health centres in six districts in rural Ghana and used logistic regression to explore factors likely to influence caregiver acceptability of RDT based case management and concern about the denial of ACT on account of negative RDT results. Focus group discussions were conducted to explain the quantitative findings and to elicit further factors.

Results

A total of 3047 caregivers were interviewed. Nearly all (98%) reported a preference for TBMM over presumptive treatment. Caregivers who preferred TBMM were less likely to be concerned about the denial of ACT to their test-negative children (O.R. 0.57, 95%C.I. 0.33–0.98). Compared with caregivers who had never secured national health insurance cover, caregivers who had valid (adjusted O.R. 1.30, 95% CI 1.07–1.61) or expired (adjusted O.R. 1.38, 95% CI 1.12–1.73) insurance cover were more likely to be concerned about the denial of ACT to their RDT-negative children. Major factors that promote TBMM acceptability include the perception that a blood test at health centre level represents improvement in the quality of care, leads to improvement in treatment outcomes, and offers opportunity for better communication between health workers and caregivers. Acceptability is also enhanced by engaging caregivers in the procedures of the test. Apprehensions about negative health worker attitude could however undermine acceptance.

Conclusion

Test (RDT)-based management of malaria in under-five children is likely to be acceptable to caregivers in rural Ghana. The quality of caregiver-health worker interaction needs to be improved if acceptability is to be sustained.     

A combination of behavioral and physiological indicators for assessing pig welfare on the farm

The purpose of this research was to identify pig welfare indicators that could help in recognizing stressful practices on farm. The study evaluated behavioral and physiological indicators (cortisol and negative acute phase proteins) in 2 groups of 20 female pigs 4 months old after a 48-hr transport. The first group (A) was transported at the end of May, the second (B) in June. Behavioral observations and blood collection occurred at arrival (D1) and 28 days later (D28). Compared with within-animal control samples obtained 28 days later, pigs of Group A had increased cortisol levels and decreased albumin concentrations after arrival. As demonstrated by lesion and behavior observations, the effect on cortisol and albumin was higher in Group B pigs after a tail-biting episode occurred. The study has reported no evidence of Retinol Binding Protein (RBP) in pigs. A method developed for swine RBP quantification found RBP strongly reduced in D28 samples of Group B, confirming it to be a negative protein in pigs. The suggested combination of physiological and behavioral indicators could provide useful information on the welfare state of an animal.     

Perceptions of an ideal point-of-care test for sexually transmitted infections--a qualitative study of focus group discussions with medical providers

Background

A point-of-care test (POCT) for sexually transmitted infections (STIs), which offers immediate diagnosis resulting in patients receiving diagnosis and treatment in a single visit, has the ability to address some of the STI control needs. However, needs assessment from STI experts and end users about currently available STI POCTs and their future new development has not been evaluated since World Health Organization Sexually Transmitted Diseases Diagnostics Initiative was formed over 15 years ago. Therefore, our objective was to explore the perceptions of the ideal types of STI POCT for use in health care settings.

Methodology/Principal Findings

A qualitative study, encompassing eight focus groups, was conducted from March 2008 through April 2009. Participants included 6 STD clinic directors, 63 clinicians, and 7 public health/laboratory/epidemiology professionals in the STI field. Discussion topics included currently available POCT, perceived barriers to using POCT in clinics, priority STI for the development of new POCT, and characteristics of the ideal POCT. All discussions were recorded and transcribed verbatim. Themes raised as barriers for current POCT included complexity, long time frames of the so-called “rapid” test, multiple time-driven steps, requiring laboratory technician, difficulty in reading result, interruption of workflow, unreliability, and invasiveness. Chlamydia trachomatis was identified as the priority organism for development of a new STI POCT. Themes indicated for the ideal POCT included rapid turnaround (up to 20 minutes), ease of use, non-invasive, accurate (preferred sensitivity and specificity in the range of high 90s), Clinical Laboratory Improvement Amendments (CLIA)-waived, user-friendly (for both patients and staff), compact, durable, and sturdy.

Conclusions/Significance

Focus group discussions with STI experts and professionals highlighted chlamydia as the top priority pathogen for POCT development, and identified the qualities of new POCT for STIs. Participants endorsed ease of use, rapid turnaround and high accuracy as essential characteristics of an ideal POCT.     

DegePrime,a Program for Degenerate Primer Design for Broad-Taxonomic-Range PCR in Microbial Ecology Studies

The taxonomic composition of a microbial community can be deduced by analyzing its rRNA gene content by, e.g., high-throughput DNA sequencing or DNA chips. Such methods typically are based on PCR amplification of rRNA gene sequences using broad-taxonomic-range PCR primers. In these analyses, the use of optimal primers is crucial for achieving an unbiased representation of community composition. Here, we present the computer program DegePrime that, for each position of a multiple sequence alignment, finds a degenerate oligomer of as high coverage as possible and outputs its coverage among taxonomic divisions. We show that our novel heuristic, which we call weighted randomized combination, performs better than previously described algorithms for solving the maximum coverage degenerate primer design problem. We previously used DegePrime to design a broad-taxonomic-range primer pair that targets the bacterial V3-V4 region (341F-805R) (D. P. Herlemann, M. Labrenz, K. Jurgens, S. Bertilsson, J. J. Waniek, and A. F. Andersson, ISME J. 5:1571–1579, 2011, http://dx.doi.org/10.1038/ismej.2011.41), and here we use the program to significantly increase the coverage of a primer pair (515F-806R) widely used for Illumina-based surveys of bacterial and archaeal diversity. By comparison with shotgun metagenomics, we show that the primers give an accurate representation of microbial diversity in natural samples.     

Human embryonic stem cell-derived mesenchymal progenitors—Potential in regenerative medicine

Tissue engineering and cell therapy require large-scale production of homogeneous populations of lineage-restricted progenitor cells that easily can be induced to differentiate into a specific tissue. We have developed straightforward protocols for the establishment of human embryonic stem (hES) cell-derived mesenchymal progenitor (hES-MP) cell lines. The reproducibility was proven by derivation of multiple hES-MP cell lines from 10 different hES cell lines. To illustrate clinical applicability, a xeno-free hES-MP cell line was also derived. None of the markers characteristic for undifferentiated hES cells were detected in the hES-MP cells. Instead, these cells were highly similar to mesenchymal stem cells with regard to morphology and expression of markers. The safety of hES-MP cells following transplantation was studied in severely combined immunodeficient (SCID) mice. The implanted hES-MP cells gave rise to homogeneous, well-differentiated tissues exclusively of mesenchymal origin and no teratoma formation was observed. These cells further have the potential to differentiate toward the osteogenic, adipogenic, and chondrogenic lineages in vitro. The possibility of easily and reproducibly generating highly expandable hES-MP cell lines from well-characterized hES cell lines with differentiation potential into several mesodermal tissues entails an enormous potential for the field of regenerative medicine.     

Nonlinear Strain Stiffening Is Not Sufficient to Explain How Far Cells Can?Feel on Fibrous Protein Gels

Recent observations suggest that cells on fibrous extracellular matrix materials sense mechanical signals over much larger distances than they do on linearly elastic synthetic materials. In this work, we systematically investigate the distance fibroblasts can sense a rigid boundary through fibrous gels by quantifying the spread areas of human lung fibroblasts and 3T3 fibroblasts cultured on sloped collagen and fibrin gels. The cell areas gradually decrease as gel thickness increases from 0 to 150 μm, with characteristic sensing distances of >65 μm below fibrin and collagen gels, and spreading affected on gels as thick as 150 μm. These results demonstrate that fibroblasts sense deeper into collagen and fibrin gels than they do into polyacrylamide gels, with the latter exhibiting characteristic sensing distances of <5 μm. We apply finite-element analysis to explore the role of strain stiffening, a characteristic mechanical property of collagen and fibrin that is not observed in polyacrylamide, in facilitating mechanosensing over long distances. Our analysis shows that the effective stiffness of both linear and nonlinear materials sharply increases once the thickness is reduced below 5 μm, with only a slight enhancement in sensitivity to depth for the nonlinear material at very low thickness and high applied traction. Multiscale simulations with a simplified geometry predict changes in fiber alignment deep into the gel and a large increase in effective stiffness with a decrease in substrate thickness that is not predicted by nonlinear elasticity. These results suggest that the observed cell-spreading response to gel thickness is not explained by the nonlinear strain-stiffening behavior of the material alone and is likely due to the fibrous nature of the proteins.