Cultural Recovery and Determination of Antimicrobial Susceptibility in Helicobacter pylori by Using Commercial Transport and Isolation Media

Abstract : Background: : Antimicrobial resistance of Helicobacter pylori is the main reason for eradication failure. We have studied the feasibility of a commercial transport medium for cultural recovery and subsequent drug susceptibility testing. Patients and Methods: : From March to December 2000, 79 consecutive gastric biopsies, positive in a rapid urease test, were transferred into a commercial transport medium and sent within 24 hours from the district hospital to the microbiological laboratory for culture and susceptibility testing. A commercial agar plate and an in-house Wilkins-Chalgren agar plate were used for culture. Susceptibility data were compared with data collected from 1992 to 2003 in the University Hospital of Zurich. Results: : Cultural recovery and susceptibility testing of H. pylori was successful in 55 of 79 patients. In 17 cases cultural recovery failed because of technical problems (n = 14), long transport time (n = 1) and unknown reason (n = 2). Failure of susceptibility testing (n = 7) was mainly due to fungal overgrowth. Resistance to metronidazole and clarithromycin was found in 15 (27%) and in 12 patients (22%), respectively; resistance to amoxicillin was not observed. Five patients (9%) showed resistance both to metronidazole and to clarithromycin. Eradication therapy failed in all patients with macrolide resistance. Resistance rates were higher in females than in males; 30% vs 12% for clarithromycin and 33% vs 20% for metronidazole. Resistance to metronidazole was significantly lower in Swiss patients (15%) than in non-Swiss patients (39%). Conclusion: : Antimicrobial resistance data can reliably be obtained by sending the biopsy specimen in a commercial transport medium to a microbiological laboratory. This is especially important after eradication failure. Resistance to metronidazole and clarithromycin is highly prevalent and more common in women and non-Swiss patient

Outcomes and safety of concomitant nevirapine and rifampicin treatment under programme conditions in Malawi.

SETTING: Thyolo District Hospital, rural Malawi. OBJECTIVES: To report on 1) clinical, immunological and virological outcomes and 2) safety among human immunodeficiency virus (HIV) infected patients with tuberculosis (TB) who received concurrent nevirapine (NVP) and rifampicin (RMP) based treatment. DESIGN: Retrospective cohort study. METHODS: Analysis of programme data, June-December 2007. RESULTS: Of a total of 156 HIV-infected TB patients who started NVP-based antiretroviral treatment, 136 (87%) completed TB treatment successfully, 16 (10%) died and 5 (4%) were transferred out. Mean body weight and CD4 gain (adults) were respectively 4.4 kg (95%CI 3.3-5.4) and 140 cells/mm(3) (95%CI 117-162). Seventy-four per cent of patients who completed TB treatment and had a viral load performed (n = 74) had undetectable levels (<50 copies/ml), while 17 (22%) had a viral load of 50-1000 copies/ml. Hepatotoxicity was present in 2 (1.3%) patients at baseline. Two patients developed Grade 2 and one developed Grade 3 alanine transaminase enzyme elevations during TB treatment (incidence rate per 10 years of follow-up 4.2, 95%CI 1.4-13.1). There were no reported deaths linked to hepatotoxicity. CONCLUSIONS: In a rural district in Malawi, concomitant NVP and RMP treatment is associated with good TB treatment outcomes and appears safe. Further follow-up of patients would be useful to ascertain the longer-term effects of this concurrent treatment

The GadX regulon affects virulence gene expression and adhesion of porcine enteropathogenic Escherichia coli in vitro

The ability of enteropathogenic Escherichia coli (EPEC) to express virulence factor genes and develop attaching and effacing (AE) lesions is inhibited in acidic environmental conditions. This inhibition is due to the activation of transcription factor GadX, which upregulates expression of glutamic acid decarboxylase (Gad). Gad, in turn, produces γ-aminobutyric acid (GABA), which was recently shown to have a beneficial effect on the jejunal epithelium in vitro due to increased mucin-1 levels. In the present study, we sought to test whether forced GadX activation/overexpression abolishes virulence associated features of EPEC and provokes increased GABA production. EPEC strains were isolated from diarrheic pigs and submitted to activation of GadX by acidification as well as gadX overexpression via an inducible expression vector plasmid. GABA concentrations in the growth medium, ability for adhesion to porcine intestinal epithelial cells (IPEC-J2) and virulence gene expression were determined. Growth in acidified media led to increased GABA levels, upregulated gadA/B expression and downregulated mRNA synthesis of the bacterial adhesin intimin. EPEC strains transformed with the gadX gene produced 2.1 to 3.4-fold higher GABA levels than empty-vector controls and completely lost their ability to adhere to IPEC-J2 cells and to induce actin accumulation. We conclude that intensified gadX activation can abolish the ability of EPEC to adhere to the intestinal epithelium by reducing the expression of major virulence genes

Randomized clinical trial to evaluate the effects of a prepartum cholecalciferol injection on postpartum serum calcium dynamics and health and performance in early-lactation multiparous dairy cows

The objectives of the present study were (1) to evaluate the effect of prepartum cholecalciferol treatment on serum Ca concentration during the first 10 d after calving and (2) to evaluate the effect of treatment on subsequent health and performance. Multiparous Holstein cows (n = 377) from one dairy farm were fed a negative dietary cation-anion difference diet (−31 mEq/kg of DM) for the last 21 d of gestation. On d 275, the animals were randomly assigned to a control or a treatment group. Cows in the control group were left untreated, and cows in the treatment group received an injection of 12 × 106 IU of cholecalciferol intramuscularly on the day of enrollment. If treated cows did not deliver the calf within 6 d, they were reinjected with 10 × 106 IU of cholecalciferol. Blood samples were drawn on 1, 2, 3, 5, 7, and 10 days in milk (DIM) and analyzed for serum Ca, P, and Mg concentrations. In a subsample of cows (50 control cows, 35 cows treated once with cholecalciferol, and 15 cows treated twice) serum haptoglobin, nonesterified fatty acids, β-hydroxybutyrate, and 25-hydroxycholecalciferol concentrations were analyzed on 1, 5, and 10 DIM. Binary data [retained placenta (RP), metritis] were analyzed using logistic regression models. Repeated measures ANOVA with first-order autoregressive covariance was performed to evaluate the treatment effect on milk yield over the first 10 test days after parturition, 25-hydroxycholecalciferol, serum Ca, P, Mg, β-hydroxybutyrate, nonesterified fatty acids, and haptoglobin concentrations. Cox proportional hazards were used to model the time to event outcomes (time to pregnancy within 200 d, culling until 300 DIM). After enrollment of 31.4% of cows and a preliminary analysis, adverse reactions became apparent, and the study was stopped. Cows treated with cholecalciferol had a greater risk of incurring RP and metritis. The adjusted mean incidences were 2.0%, 7.7%, and 4.0% for RP, and 21.6%, 39.3%, and 33.3% for metritis for control cows, cows treated once, and cows treated twice with cholecalciferol, respectively. Compared with control cows, cows injected once with 12 × 106 IU of cholecalciferol produced less energy-corrected milk on the first (−3.76 kg) and second (−2.75 kg) test days, respectively. Cows injected twice with cholecalciferol (12 × 106 IU of cholecalciferol and 10 × 106 IU 1 wk later) had a reduced milk yield only at first test day (−3.80 kg). Treatment with cholecalciferol led to a significant increase in 25-hydroxycholecalciferol on d 1, 5, and 10 after calving. Serum Ca and P concentrations were significantly increased in cows treated with cholecalciferol, but serum Mg concentrations were significantly reduced. Haptoglobin concentrations were significantly increased on 5 DIM in cows injected once with 12 × 106 IU of cholecalciferol. Although we observed no effect of treatment on culling until 300 DIM, time to pregnancy was delayed by 34 d in cows injected once with 12 × 106 IU of cholecalciferol. In the present study, injection with 12 × 106 IU of cholecalciferol had detrimental effects on health and milk production despite the beneficial effects on Ca homeostasis

Effects of broccoli extract and various essential oils on intestinal and faecal microflora and on xenobiotic enzymes and the antioxidant system of piglets

Objective: Since the ban of antibiotics as growth promoting feed additives in the EU in 2006 research in alternatives has gained importance. Phytogenic feed additives represent a heterogenous class of different plant derived substances that are discussed to improve the health of farm animals by direct and indirect antioxidant effects and by influencing microbial eubiosis in the gastrointestinal tract. Consequently our study aimed to investigate the influence of broccoli extract and the essential oils of tur- meric, oregano, thyme and rosemary, as selected individual additives, on intestinal and faecal microflora, on xenobiotic enzymes, and on the antioxidant system of piglets. Methods: 48 four weeks old male weaned piglets were assigned to 6 groups of 8. The piglets were housed individually in stainless steel pens with slatted floor. The control group (Con) was fed a diet without an additive for 4 weeks. The diet of group BE contained 0.15 g/kg sulforaphane in form of a broccoli extract. 535, 282, 373 and 476 mg/kg of the essential oils of turmeric (Cuo), oregano (Oo), thyme (To) and rosemary (Ro) were added to the diets of the remaining 4 groups to stan-dardise supplementation to 150 mg/kg of the oils’ key terpene compounds ar-turmerone, carvacrol, thymol and 1,8-cineole. The composition of bacterial microflora was examined by cultivating samples of jejeunal and colonic mucosa and of faeces under specific conditions. The mRNA expression of xenobiotic and antioxidant enzymes was determined by reversing transcrip- tase real time detection PCR (RT-PCR). Total antioxidant status was assayed using the Trolox Equivalent Antioxidant Capacity (TEAC), and lipid peroxidation was determined by measuring thiobarbioturic acid reactive substances (TBA- RS). Results: Compared to Con piglets all additives positively influenced weight gain and feed conversion in week 1. Over the whole trial period no significant differences in performance parameters existed between the experimental groups. Compared to group Con performance of Ro piglets was, however, slightly impaired. Com- pared to Con piglets Cuo, Oo and To increased the ratio of Lactobacilli:E. coli attached to the jejunal mucosa, whereas BE and Ro impaired this ratio slightly. In contrast in colonic mucosa Ro improved Lactobacilli:E. coli ratio. In faecal samples an improvement of Lactobacilli:E. coli ratio could be analysed for To and Ro. Ro was the only additive that reduced the incidence rate of piglets tested positive for enterotoxic E. coli (ETEC). All additives significantly increased jejunal TEAC and reduced TBA-RS. In the liver BE, Cuo, Oo and To increased TEAC in tendency and Ro significantly. Liver TBA-RS were slightly reduced by all additives compared to Con piglets. Whereas the influence of BE, To and Ro on jejunal TEAC mainly was derived from the induction of xenobiotic and antioxidant enzymes (indirect antioxidant effects), Cuo and Oo influenced TEAC by direct antioxidant effects. Discussion and Conclusions: Our results have shown: That within the labiatae oils Oo and To have the potential to improve performance slightly. That phytogenic substances have a small but not sig- nificant influence on intestinal microflora. That phytogenic feed additives up-regulate the anti- oxidant system of piglets either by direct or by indirect antioxidant effects and that they may thereby improve health status. That within the labiatae oils Oo has a high direct antioxidant potential whereas Ro potently induces xenobiotic and antioxidant enzymes. That broccoli extract is an attractive new phytogenic additive, improving antioxidant status by indirect antioxidant effects. That defined combinations of selected phytogenic substances may produce additive effects. That health promoting effects of phytogenic additives in the future should be studied systematically under the challenge with pathogenic microorganisms or food derived to-xins

a randomized clinical trial

The objective of the present study was to evaluate the effect of 2 dosages of prepartum cholecalciferol injection on blood minerals, vitamin D metabolites, and milk production. Cows entering their second or greater lactation (n = 158) were randomly assigned to a control group (CON) or one of 2 treatment groups receiving either 6 × 106 IU (6VitD) or 12 × 106 IU (12VitD) cholecalciferol intramuscularly on d 275 ± 1.2 (SD) of gestation. Concentrations of serum total Ca (tCa), phosphate, and Mg were determined on 1, 2, 3, 5, 7, and 10 d in milk (DIM). For a subsample of 30 cows entering the third lactation (n = 10/group), these samples were analyzed for cholecalciferol, 25-hydroxycholecalciferol (25-OHD3), and 24,25-dihydroxycholecalciferol (24,25-[OH]2D3). In these cows, we also determined 1,25-dihydroxycholecalciferol (1,25-[OH]2D3), the biologically most active metabolite, on 1, 2, 3, and 5 DIM. Repeated measures ANOVA was performed to evaluate the effect of different dosages of cholecalciferol on blood minerals, vitamin D metabolites, and milk yield over the first 5 test days after calving. Binary outcomes such as retained placenta and metritis were analyzed using a chi-squared test. Although the 12VitD treatment increased tCa concentrations on 1, 2, and 3 DIM compared with CON, administration of 6VitD increased tCa concentrations only on 1 DIM. Compared with CON cows and 6VitD cows, 12VitD cows had greater serum phosphate concentration during the first 10 DIM. Furthermore, 6VitD cows had greater serum phosphate concentrations compared with CON cows. On the contrary, 12VitD cows had lower serum Mg concentrations during the first 10 DIM compared with CON and 6VitD cows. Cholecalciferol was increased by the treatment and decreased quickly until 10 DIM. In respect to 25-OHD3, the 6VitD treatment resulted in a 4.1-fold increase in comparison to the CON group, while a 6.5-fold increase was observed in 12VitD animals. The vitamin D metabolite 24,25-(OH)2D3 increased linearly with 25-OHD3 serum levels, resulting in the highest concentrations in the 12VitD group. An increase of 1,25-(OH)2D3 until 3 DIM was observed in all cows. However, this rise was most pronounced in the CON group. The incidence of retained placenta was 1.9%, 11.5%, and 29.6%, and that of metritis was 11.5%, 15.4%, and 31.5% for CON, 6VitD, and 12VitD cows, respectively. Although none of the treated cows exerted clinical signs of hypocalcemia, one cow in CON incurred clinical hypocalcemia. Cows of the 12VitD group had a lower milk yield over the first 5 monthly test days compared with the control and 6VitD group (42.2 ± 0.5, 42.0, ± 0.5 and 40.7 ± 0.5 kg for control cows, 6VitD cows and 12VitD cows, respectively). Although no negative side effects were observed in 6VitD cows, we do not recommend the general application of 6 × 106 IU cholecalciferol before calving as positive effects on calcium homeostasis were marginal and restricted to the first DIM. The present findings confirm that the application of 12 × 106 IU cholecalciferol negatively affected milk production on this farm