Using non-invasive neuroimaging to enhance the care, well-being and experimental outcomes of laboratory non-human primates (monkeys).

Over the past 10-20 years, neuroscience witnessed an explosion in the use of non-invasive imaging methods, particularly magnetic resonance imaging (MRI), to study brain structure and function. Simultaneously, with access to MRI in many research institutions, MRI has become an indispensable tool for researchers and veterinarians to guide improvements in surgical procedures and implants and thus, experimental as well as clinical outcomes, given that access to MRI also allows for improved diagnosis and monitoring for brain disease. As part of the PRIMEatE Data Exchange, we gathered expert scientists, veterinarians, and clinicians who treat humans, to provide an overview of the use of non-invasive imaging tools, primarily MRI, to enhance experimental and welfare outcomes for laboratory non-human primates engaged in neuroscientific experiments. We aimed to provide guidance for other researchers, scientists and veterinarians in the use of this powerful imaging technology as well as to foster a larger conversation and community of scientists and veterinarians with a shared goal of improving the well-being and experimental outcomes for laboratory animals.

Immunological consequences of strain variation within the Mycobacterium tuberculosis complex.

In 2015, there were an estimated 10.4 million new cases of tuberculosis (TB) globally, making it one of the leading causes of death due to an infectious disease. TB is caused by members of the Mycobacterium tuberculosis complex (MTBC), with human disease resulting from infection by M. tuberculosis sensu stricto and M. africanum. Recent progress in genotyping techniques, in particular the increasing availability of whole genome sequence data, has revealed previously under appreciated levels of genetic diversity within the MTBC. Several studies have shown that this genetic diversity may translate into differences in TB transmission, clinical manifestations of disease, and host immune responses. This suggests the existence of MTBC genotype-dependent host-pathogen interactions which may influence the outcome of infection and progression of disease. In this review, we highlight the studies demonstrating differences in innate and adaptive immunological outcomes consequent on MTBC genetic diversity, and discuss how these differences in immune response might influence the development of TB vaccines, diagnostics and new therapies.

Internalization of Vectorized Liposomes Loaded with Plasmid DNA in C6 Glioma Cells.

We studied internalization of vector nanocarriers loaded with plasmid DNA into C6 glioma cells. For improving selectivity of plasmid delivery, the liposomes were conjugated with monoclonal antibodies to VEGF and its receptor VEGFR2. Flow cytofluorometry and laser scanning confocal microscopy showed more intensive (more than 2-fold) internalization and accumulation of antibody-vectorized liposomes in C6 glioma cells in comparison with the control (liposomes conjugated with non-specific antibodies and non-vectorized liposomes). Using quantitative analysis of fluorescent signal, we showed that cationic immunoliposomes significantly more effective delivered pCop-Green-N plasmid DNA and ensured effective transfection of C6 glioma cells.

No added value of interferon-γ release to a prediction model for childhood tuberculosis.

The predictive value of a combination of clinical and radiological features with interferon-γ release assay (IGRA) for diagnosis of active tuberculosis (TB) disease among TB-exposed children is unknown.150 symptomatic HIV-negative children (aged 3 months to 14 years), prospectively recruited through active contact tracing, were included. Backward stepwise logistic regression and bootstrapping techniques were used for the development and internal validation of a clinical prediction model for active TB disease. Model discrimination and incremental value of a positive IGRA test were assessed by area under the receiver operating characteristic curve (AUC).35 (23%) children were diagnosed with active TB disease and started on treatment and 115 (77%) had other respiratory tract infections. A final parsimonious clinical model, comprising age <5 years (adjusted (a)OR 4.8, 95% CI 2.0-11.5) and lymphadenopathy on clinical examination (aOR 4.9, 95% CI 1.8-13.0) discriminated active TB disease from other disease with an AUC of 0.70 (95% CI 0.61-0.80). A positive IGRA result did not improve the discriminatory ability of the clinical model (c-statistic 0.72 versus 0.70; p=0.644).A clinical algorithm, including age <5 years and lymphadenopathy classified 70% of active TB disease among symptomatic TB-exposed children. IGRA does not add any discriminatory value to this prediction model.

The emerging threat of pre-extensively drug-resistant tuberculosis in West Africa: preparing for large-scale tuberculosis research and drug resistance surveillance.

BACKGROUND: Drug-resistant tuberculosis (TB) is a global public health problem. Adequate management requires baseline drug-resistance prevalence data. In West Africa, due to a poor laboratory infrastructure and inadequate capacity, such data are scarce. Therefore, the true extent of drug-resistant TB was hitherto undetermined. In 2008, a new research network, the West African Network of Excellence for Tuberculosis, AIDS and Malaria (WANETAM), was founded, comprising nine study sites from eight West African countries (Burkina Faso, The Gambia, Ghana, Guinea-Bissau, Mali, Nigeria, Senegal and Togo). The goal was to establish Good Clinical Laboratory Practice (GCLP) principles and build capacity in standardised smear microscopy and mycobacterial culture across partnering laboratories to generate the first comprehensive West African drug-resistance data. METHODS: Following GCLP and laboratory training sessions, TB isolates were collected at sentinel referral sites between 2009-2013 and tested for first- and second-line drug resistance. RESULTS: From the analysis of 974 isolates, an unexpectedly high prevalence of multi-drug-resistant (MDR) strains was found in new (6 %) and retreatment patients (35 %) across all sentinel sites, with the highest prevalence amongst retreatment patients in Bamako, Mali (59 %) and the two Nigerian sites in Ibadan and Lagos (39 % and 66 %). In Lagos, MDR is already spreading actively amongst 32 % of new patients. Pre-extensively drug-resistant (pre-XDR) isolates are present in all sites, with Ghana showing the highest proportion (35 % of MDR). In Ghana and Togo, pre-XDR isolates are circulating amongst new patients. CONCLUSIONS: West African drug-resistance prevalence poses a previously underestimated, yet serious public health threat, and our estimates obtained differ significantly from previous World Health Organisation (WHO) estimates. Therefore, our data are reshaping current concepts and are essential in informing WHO and public health strategists to implement urgently needed surveillance and control interventions in West Africa.

Position paper on olfactory dysfunction.

BACKGROUND: Olfactory dysfunction is an increasingly recognised condition, associated with reduced quality of life and major health outcomes such as neurodegeneration and death. However, translational research in this field is limited by heterogeneity in methodological approach, including definitions of impairment, improvement and appropriate assessment techniques. Accordingly, effective treatments for smell loss are limited. In an effort to encourage high quality and comparable work in this field, among others, we propose the following ideas and recommendations. Whilst the full set of recommendations are outlined in the main document, points include the following: - Patients with suspected olfactory loss should undergo a full examination of the head and neck, including rigid nasal endoscopy with small diameter endoscopes. - Subjective olfactory assessment should not be undertaken in isolation, given its poor reliability. - Psychophysical assessment tools used in clinical and research settings should include reliable and validated tests of odour threshold, and/or one of odour identification or discrimination. - Comprehensive chemosensory assessment should include gustatory screening. - Smell training can be helpful in patients with olfactory loss of several aetiologies. CONCLUSIONS: We hope the current manuscript will encourage clinicians and researchers to adopt a common language, and in so doing, increase the methodological quality, consistency and generalisability of work in this field.

Differences in T-cell responses between Mycobacterium tuberculosis and Mycobacterium africanum-infected patients.

In The Gambia, Mycobacterium tuberculosis (Mtb) and Mycobacterium africanum (Maf) are major causes of tuberculosis (TB). Maf is more likely to cause TB in immune suppressed individuals, implying differences in virulence. Despite this, few studies have assessed the underlying immunity to the two pathogens in human. In this study, we analyzed T-cell responses from 19 Maf- and 29 Mtb-infected HIV-negative patients before and after TB chemotherapy following overnight stimulation of whole blood with TB-specific antigens. Before treatment, percentages of early secreted antigenic target-6(ESAT-6)/culture filtrate protein-10(CFP-10) and purified protein derivative-specific single-TNF-α-producing CD4(+) and CD8(+) T cells were significantly higher while single-IL-2-producing T cells were significantly lower in Maf- compared with Mtb-infected patients. Purified protein derivative-specific polyfunctional CD4(+) T cells frequencies were significantly higher before than after treatment, but there was no difference between the groups at both time points. Furthermore, the proportion of CD3(+) CD11b(+) T cells was similar in both groups pretreatment, but was significantly lower with higher TNF-α, IL-2, and IFN-γ production in Mtb- compared with that of Maf-infected patients posttreatment. Our data provide evidence of differences in T-cell responses to two mycobacterial strains with differing virulence, providing some insight into TB pathogenesis with different Mtb strains that could be prospectively explored as biomarkers for TB protection or susceptibility.

Generation of recombinant extracellular fragment of vascular endothelial growth factor receptor 2 and specific monoclonal antibodies to this receptor.

cDNA extracellular Ig-like domains I-III of vascular endothelial growth factor receptor 2 (VEGFR2) was cloned in an expressing vector pET_32a. Western blotting showed immunochemical identity of recombinant VEGFR2I-III produced by prokaryotic expression system to the native receptor. BALB/c mice were immunized with VEGFR2I-III for obtaining specific antibodies to VEGFR2. Hybridomas producing monoclonal antibodies were selected by ELISA, Western blotting, and immunocytochemical assay. Thus, we obtained hybridoma producing monoclonal antibodies to VEGFR2 that selectively interact with both recombinant and native extracellular fragment of the receptor.

Host and pathogen factors that influence variability of Mycobacterium tuberculosis lipid body content in sputum from patients with tuberculosis: an observational study.

BACKGROUND: High proportions of Mycobacterium tuberculosis cells in sputum containing triacylglycerol-rich lipid bodies have been shown to be associated with treatment failure or relapse following antituberculous chemotherapy. Although lipid body determination is a potential biomarker for supporting clinical trial and treatment decisions, factors influencing variability in sputum frequencies of lipid body-positive (%LB+) M tuberculosis in patients are unknown. We aimed to test our hypothesis that exposure to host-generated NO and M tuberculosis strains are factors associated with differences in sputum %LB+. METHODS: In this observational study, we determined %LB+ frequencies before treatment by microscopy in patients with smear-positive tuberculosis from two separate prospective observational study settings (Gondar, Ethiopia, recruited between May 1, 2010, and April 30, 2011, and Fajara, The Gambia, who provided sputum samples before treatment between May 5, 2010, and Dec 22, 2011). In Ethiopia, fractional exhaled nitric oxide (FeNO) was measured as a biomarker of host NO, and M tuberculosis strain differences were determined by spoligotyping. Treatment response was assessed by percentage weight change after 7 months. In The Gambia, treatment responses were assessed as change in BMI and radiographic burden of disease after 6 months. Sputum M tuberculosis isolates were studied in vitro for their %LB+ and triacylglycerol synthase 1 (tgs1) mRNA responses to NO exposure. Propidium iodide staining was used as a measure of NO strain toxicity. Correlation between in vitro %LB+ frequencies following NO exposure and those of the same strain in sputum was examined with linear regression and Dunnett's multiple comparison test. FINDINGS: In Ethiopia, 73 patients who were smear positive for pulmonary tuberculosis were recruited (43 [59%] were male and 30 [41%] were female). Of these, the %LB+ in the sputum of 59 patients showed linear correlation with log10 FeNO (r2=0·28; p<0·0001) and an association with strain spoligotype was suggested. Seven M tuberculosis strains from The Gambia showed different dose-responses to NO in vitro, demonstrated by changing lipid body content, tgs1 transcription, and bacterial toxicity. In sputum %LB+ frequencies correlated with in vitro %LB+ responses to NO of the corresponding isolate. In a subset of 34 patients across both cohorts, higher sputum %LB+ frequencies before treatment were associated with weaker responses to treatment than lower sputum %LB+ frequencies. INTERPRETATION: M tuberculosis strain and exposure to host-generated NO are associated with sputum %LB+. Our results support the use of M tuberculosis strain-dependent sputum %LB+ as a predictive biomarker of treatment response. FUNDING: The Medical Research Council, the University of Leicester, and the University of Gondar.

Functional genetic variation in pe/ppe genes contributes to diversity in Mycobacterium tuberculosis lineages and potential interactions with the human host.

Introduction: Around 10% of the coding potential of Mycobacterium tuberculosisis constituted by two poorly understood gene families, the pe and ppe loci, thought to be involved in host-pathogen interactions. Their repetitive nature and high GC content have hindered sequence analysis, leading to exclusion from whole-genome studies. Understanding the genetic diversity of pe/ppe families is essential to facilitate their potential translation into tools for tuberculosis prevention and treatment. Methods: To investigate the genetic diversity of the 169 pe/ppe genes, we performed a sequence analysis across 73 long-read assemblies representing seven different lineages of M. tuberculosis and M. bovis BCG. Individual pe/ppe gene alignments were extracted and diversity and conservation across the different lineages studied. Results: The pe/ppe genes were classified into three groups based on the level of protein sequence conservation relative to H37Rv, finding that >50% were conserved, with indels in pe_pgrs and ppe_mptr sub-families being major drivers of structural variation. Gene rearrangements, such as duplications and gene fusions, were observed between pe and pe_pgrs genes. Inter-lineage diversity revealed lineage-specific SNPs and indels. Discussion: The high level of pe/ppe genes conservation, together with the lineage-specific findings, suggest their phylogenetic informativeness. However, structural variants and gene rearrangements differing from the reference were also identified, with potential implications for pathogenicity. Overall, improving our knowledge of these complex gene families may have insights into pathogenicity and inform the development of much-needed tools for tuberculosis control.

Cloning and expression of brain-specific anion transporter BSAT1 and isolation of monoclonal antibodies to its extracellular fragment.

Brain-specific anion transporter BSAT1 extracellular fragment (451-557) cDNA was cloned in a vector for prokaryotic expression, a producer E. coli strain was obtained, and recombinant extracellular fragment BSAT1(451-557)was purified and used for immunization of BALB/c mice. Splenic cells from mice with verified immune response were used for hybridoma generation. Several hybridoma clones producing monoclonal antibodies to BSAT1 extracellular fragment were selected. Antibody specificity was confirmed by ELISA, immunoblotting with recombinant BSAT1(451-557), and immunofluorescent BSAT1 assay on rat brain sections and cultured HEK293 cells. It was demonstrated that the obtained antibodies specifically bind native rat and human BSAT1 and can be used in both fundamental studies of structures forming the blood-brain barrier and development of targeted transport of diagnostic preparations and drugs across the blood-brain barrier.

Generation of monoclonal antibodies to recombinant vascular endothelial growth factor.

Female BALB/c mice were subcutaneously immunized with recombinant VEGF-164. After 3 immunization cycles, splenic B cells from immunized mouse were fused with immortalized myeloma culture SP2/0-Ag14 cells. Screening of hybrid cells producing anti-VEGF antibodies was performed by ELISA and immunocytochemical analysis on cultured C6 glioma cells. Subsequent cloning yielded hybridoma stably expressing monoclonal anti-VEGF antibodies recognizing recombinant and native VEGF.

Ligand-receptor assay for evaluation of functional activity of human recombinant VEGF and VEGFR-1 extracellular fragment.

cDNA encoding VEGF and Ig-like extracellular domains 2-4 of VEGFR-1 (sFlt-1(2-4)) were cloned into prokaryotic expression vectors pET32a and pQE60. Recombinant proteins were purified (metal affinity chromatography) and renatured. Chemiluminescent study for the interaction of recombinant VEGF and sFlt-1(2-4) showed that biotinylated VEGF specifically binds to the polystyrene-immobilized receptor extracellular fragment. Biotinylated recombinant sFlt-1 interacts with immobilized VEGF. Analysis of the interaction of immobilized recombinant VEGFR-1 and VEGF with C6 glioma cells labeled with CFDA-SE (vital fluorescent dye) showed that recombinant VEGFR-1 also binds to native membrane-associated VEGF. Recombinant VEGF was shown to bind to specific receptors expressed on the surface of C6 glioma cells. Functional activity of these proteins was confirmed by ligand-receptor assay for VEGF and VEGFR-1 (sFlt-1) and quantitative chemiluminescent detection.

Prototype-referenced shape encoding revealed by high-level aftereffects.

We used high-level configural aftereffects induced by adaptation to realistic faces to investigate visual representations underlying complex pattern perception. We found that exposure to an individual face for a few seconds generated a significant and precise bias in the subsequent perception of face identity. In the context of a computationally derived 'face space,' adaptation specifically shifted perception along a trajectory passing through the adapting and average faces, selectively facilitating recognition of a test face lying on this trajectory and impairing recognition of other faces. The results suggest that the encoding of faces and other complex patterns draws upon contrastive neural mechanisms that reference the central tendency of the stimulus category.

Position paper on olfactory dysfunction.

Background: Olfactory dysfunction is an increasingly recognised condition, associated with reduced quality of life and major health outcomes such as neurodegeneration and death. However, translational research in this field is limited by heterogeneity in methodological approach, including definitions of impairment, improvement and appropriate assessment techniques. Accordingly, effective treatments for smell loss are limited. In an effort to encourage high quality and comparable work in this field, among others, we propose the following ideas and recommendations. Whilst the full set of recommendations are outlined in the main document, points include the following: • Patients with suspected olfactory loss should undergo a full examination of the head and neck, including rigid nasal endoscopy with small diameter endoscopes. • Subjective olfactory assessment should not be undertaken in isolation, given its poor reliability. • Psychophysical assessment tools used in clinical and research settings should include reliable and validated tests of odour threshold, and/or one of odour identification or discrimination. • Comprehensive chemosensory assessment should include gustatory screening. • Smell training can be helpful in patients with olfactory loss of several aetiologies. Conclusions: We hope the current manuscript will encourage clinicians and researchers to adopt a common language, and in so doing, increase the methodological quality, consistency and generalisability of work in this field.

Comparison of TB-LAMP, GeneXpert MTB/RIF and culture for diagnosis of pulmonary tuberculosis in The Gambia.

BACKGROUND: Diagnosis of tuberculosis (TB) remains difficult, particularly in resource-limited settings. The development of nucleic acid-based tests for detection of Mycobacterium tuberculosis complex (MTBC) has significantly increased sensitivity compared to conventional smear microscopy and provides results within a matter of hours compared to weeks for the current gold-standard, liquid culture. METHODS: In this study we performed side-by-side comparison of mycobacterial detection assays on sputum samples from 285 subjects presenting with symptoms suggestive of TB in The Gambia and a cross-sectional cohort of 156 confirmed TB patients with a median of 2 months of treatment. A novel assay, Loop-Mediated Amplification test for TB (TB-LAMP), was compared to smear microscopy, MGIT culture and GeneXpert MTB/RIF for all samples. RESULTS: When culture was used as the reference standard, we found an overall sensitivity for TB-LAMP of 99% (95% CI: 94.5-99.8) and specificity of 94% (95% CI: 89.3-96.7). When latent class analysis was performed, TB-LAMP had 98.6% (95% CI: 95.9-100) sensitivity and 99% (95% CI: 98.2-100) specificity compared to 91.1% (95% CI: 86.1-96) sensitivity and 100% (95% CI: 98.2-100) specificity for MGIT culture. GeneXpert had the highest sensitivity 99.1% (95% CI: 97.1-100) but the lowest specificity 96% (95% CI: 92.6-98.3). Both TB-LAMP and GeneXpert showed high sensitivity and specificity regardless of age or strain of infection. CONCLUSION: Our findings show the diagnostic utility of both GeneXpert and TB-LAMP in The Gambia. Whilst TB-LAMP requires less infrastructure, it is unable to detect drug-resistant patterns and therefore would be most suitable as a screening test for new TB cases in peripheral health clinics.

Host Immune Responses Differ between M. africanum- and M. tuberculosis-Infected Patients following Standard Anti-tuberculosis Treatment.

Epidemiological differences exist between Mycobacterium africanum (Maf)- and Mycobacterium tuberculosis (Mtb)-infected patients, but to date, contributing host factors have not been characterised. We analysed clinical outcomes, as well as soluble markers and gene expression profiles in unstimulated, and ESAT6/CFP-10-, whole-Maf- and Mtb-stimulated blood samples of 26 Maf- and 49 Mtb-HIV-negative tuberculosis patients before, and after 2 and 6 months of anti-tuberculosis therapy. Before treatment, both groups had similar clinical parameters, but differed in few cytokines concentration and gene expression profiles. Following treatment the body mass index, skinfold thickness and chest X-ray scores showed greater improvement in the Mtb- compared to Maf-infected patients, after adjusting for age, sex and ethnicity (p = 0.02; 0.04 and 0.007, respectively). In addition, in unstimulated blood, IL-12p70, IL12A and TLR9 were significantly higher in Maf-infected patients, while IL-15, IL-8 and MIP-1α were higher in Mtb-infected patients. Overnight stimulation with ESAT-6/CFP-10 induced significantly higher levels of IFN-γ and TNF-α production, as well as gene expression of CCL4, IL1B and TLR4 in Mtb- compared to Maf-infected patients. Our study confirms differences in clinical features and immune genes expression and concentration of proteins associated with inflammatory processes between Mtb- and Maf-infected patients following anti-tuberculosis treatment These findings have public health implications for treatment regimens, and biomarkers for tuberculosis diagnosis and susceptibility.

Association of slow recovery of Mycobacterium africanum-infected patients posttreatment with high content of Persister-Like bacilli in pretreatment sputum.

OBJECTIVES/BACKGROUND: Mycobacterium africanum that causes 40% of tuberculosis (TB) in West Africa grows more slowly in culture and has similar transmission capacity compared with Mycobacterium tuberculosis, but M. africanum-exposed contacts progress more slowly to active disease. The presence of lipid body (LB) containing M. tuberculosis complex (MTBC) cells in sputum samples has been associated with mycobacterial transcriptomes indicating slow or no growth and persister-like antibiotic tolerance. Slow-growing bacilli have been found to display a persister-like phenotype with the accumulation of LBs and drug tolerance. Our previous study showed that the body mass index and lung damage resolution on chest X-ray were significantly improved slower in M. africanum-infected patients posttreatment than in M. tuberculosis-infected patients; however, the reason for this remains unclear. Therefore, we hypothesized that these differences could be either due to significant differences in drug resistance between the MTBC lineages or a difference in their content of persisters, as indicated by the percentage of LP-positive bacilli in sputum. METHODS: Sputum isolates collected before treatment from patients with TB were subjected to drug susceptibility testing using the BD BACTEC MGIT 960 SIRE kit. The percentage of acid-fast bacilli (AFB) and LB-positive bacilli in pretreatment sputum was determined by a dual staining procedure using Auramine O and LipidTOX Red neutral lipid stain, respectively, and fluorescence microscopy imaging. RESULTS: Out of the 77 isolates tested, 9 showed resistance to at least one drug and only 2 showed multidrug (rifampicin and isoniazid) resistance among M. tuberculosis-infected patients. The percentage of AFB-positive smears was similar between the two groups (p=0.821), whereas that of LP-positive bacilli was significantly higher (p=0.0059) in M. africanum-infected patients' sputa (n=24) than in M. tuberculosis-infected patients' sputa (n=36). In addition, the bacillary lengths were significantly higher in M. africanum-infected patients' sputa than in M. tuberculosis-infected patients' sputa (p=0.0007). A high frequency of LP-positive bacilli in pretreatment sputum was associated with a poor body mass index and lung damage on chest X-ray improvement following anti-TB treatment in both the groups (r2=0.022; p=0.017). CONCLUSION: The slow clinical recovery of M. africanum-infected patients compared with M. tuberculosis-infected patients posttreatment may be at least partially associated with the persistence of drug-tolerant "fat and lazy" bacilli.

Evaluation of poly(L-lactic acid) as a material for intravascular polymeric stents.

Poly(L-lactic acid) (PLLA) monofilaments were evaluated for use as intravascular polymeric stents. The PLLA monofilaments were extruded and drawn to different draw ratios. They were then subjected to different thermal treatments and their mechanical properties characterized. Stents constructed with similar monofilaments were tested under hydrostatic pressure, and the results correlated with the properties of the monofilaments. Stent collapse pressure was a decreasing function of stent diameter and filament draw ratio.

Gastric acid suppression does not promote clostridial diarrhoea in the elderly.

Gastric acid prevents bacterial colonization of the stomach and suppression of its secretion might predispose to Clostridium difficile (CD) diarrhoea. We retrospectively studied elderly patients admitted to medical wards of an acute hospital to determine whether the incidence of CD diarrhoea was greater among those previously treated with gastric acid suppressants. From records of stool CD toxin tests undertaken in 1995 and 1996, we found 126 cases with positive results, and selected 126 controls with negative results. Information about pre-morbid illness, predisposing factors for CD and medication received in the preceding 16 weeks was obtained from case-notes. A greater number of CD positive cases had received antibiotics such as Cefuroxime, ciprofloxacin or macrolides with or without metronidazole, were more severely disabled, required assistance for feeding, or had hypoalbuminaemia before the onset of diarrhoea. A greater number of controls had received lactulose, suggesting either that its laxative effect resembled CD infection prompting frequent stool tests, or that it offered protection against CD in this group. Both groups were similar for the use of proton-pump inhibitors or H2-receptor antagonists, suggesting that susceptible elderly patients are not more likely to develop CD diarrhoea after receiving gastric acid suppression therapy.