Assessing Bias in Cognitive Testing for Older Adults with Sensory Impairment: An Analysis of Differential Item Functioning in the Baltimore Longitudinal Study on Aging (BLSA) and the Atherosclerosis Risk in Communities Neurocognitive Study (ARIC-NCS).

OBJECTIVES: Vision and hearing impairments affect 55% of people aged 60+ years and are associated with lower cognitive test performance; however, tests rely on vision, hearing, or both. We hypothesized that scores on tests that depend on vision or hearing are different among those with vision or hearing impairments, respectively, controlling for underlying cognition. METHODS: Leveraging cross-sectional data from the Baltimore Longitudinal Study of Aging (BLSA) and the Atherosclerosis Risk in Communities Neurocognitive Study (ARIC-NCS), we used item response theory to test for differential item functioning (DIF) by vision impairment (better eye presenting visual acuity worse than 20/40) and hearing impairment (better ear .5-4 kHz pure-tone average > 25 decibels). RESULTS: We identified DIF by vision impairment for tests whose administrations do not rely on vision [e.g., Delayed Word Recall both in ARIC-NCS: .50 logit difference between impaired and unimpaired (p = .04) and in BLSA: .62 logits (p = .02)] and DIF by hearing impairment for tests whose administrations do not rely on hearing [Digit Symbol Substitution test in BLSA: 1.25 logits (p = .001) and Incidental Learning test in ARIC-NCS: .35 logits (p = .001)]. However, no individuals had differences between unadjusted and DIF-adjusted measures of greater than the standard error of measurement. CONCLUSIONS: DIF by sensory impairment in cognitive tests was independent of administration characteristics, which could indicate that elevated cognitive load among persons with sensory impairment plays a larger role in test performance than previously acknowledged. While these results were unexpected, neither of these samples are nationally representative and each has unique selection factors; thus, replication is critical.

The effect of missing data and imputation on the detection of bias in cognitive testing using differential item functioning methods.

BACKGROUND: Item response theory (IRT) methods for addressing differential item functioning (DIF) can detect group differences in responses to individual items (e.g., bias). IRT and DIF-detection methods have been used increasingly often to identify bias in cognitive test performance by characteristics (DIF grouping variables) such as hearing impairment, race, and educational attainment. Previous analyses have not considered the effect of missing data on inferences, although levels of missing cognitive data can be substantial in epidemiologic studies. METHODS: We used data from Visit 6 (2016-2017) of the Atherosclerosis Risk in Communities Neurocognitive Study (N = 3,580) to explicate the effect of artificially imposed missing data patterns and imputation on DIF detection. RESULTS: When missing data was imposed among individuals in a specific DIF group but was unrelated to cognitive test performance, there was no systematic error. However, when missing data was related to cognitive test performance and DIF group membership, there was systematic error in DIF detection. Given this missing data pattern, the median DIF detection error associated with 10%, 30%, and 50% missingness was -0.03, -0.08, and -0.14 standard deviation (SD) units without imputation, but this decreased to -0.02, -0.04, and -0.08 SD units with multiple imputation. CONCLUSIONS: Incorrect inferences in DIF testing have downstream consequences for the use of cognitive tests in research. It is therefore crucial to consider the effect and reasons behind missing data when evaluating bias in cognitive testing.

The shifting picture of HIV treatment, comorbidity and substance use among US MSM living with HIV.

OBJECTIVES: People living with HIV (PLWH) have increased risk of chronic disease and poor mental health. We aimed to explore HIV disease indicators, comorbidity, and risk behavior of recent antiretroviral therapy (ART) initiators to inform current needs of PLWH. METHODS: Men who have sex with men (MSM) in the Multicenter AIDS Cohort Study (MACS) who initiated ART between 2010 and 2018 (recent initiators) were compared with age-, race- and geographic location-matched men who initiated ART during 2000-2009 (early initiators). Measures of HIV disease, behavior, comorbidity and mental health were collected prospectively every 6 months using standardized forms. RESULTS: Recent initiators had higher current CD4 (median CD4 451 vs. 307 cells/µL, P < 0.0001) and nadir CD4 (451 vs. 300 cells/µL, P < 0.0001) than earlier initiators. The proportion achieving viral suppression within a year of starting ART was significantly higher in recent compared with earlier initiators (92% vs. 74%, P < 0.0001). Median [interquartile range (IQR)] time from HIV diagnosis to ART initiation was 5.4 (1.7-23.1) months in recent initiators. Comorbidity prevalence was high in recent initiators, including obesity (24%), hypertension (25%) and kidney disease (15%). Substance use continues to be common, including cigarette use (40%), daily alcohol use (88%) and marijuana use (46%). CONCLUSIONS: Improvements in getting individuals onto ART at an early stage have led to substantially higher CD4 cell counts at initiation. However, the high burden of comorbidity, substance use and poor mental health affecting MSM living with HIV in the US underscore ongoing challenges and our need to adapt and coordinate care.

Kefir micro-organisms: their role in grain assembly and health properties of fermented milk.

Kefir is a homemade viscous and slightly effervescent beverage obtained by milk fermentation with kefir grains, which are built up by a complex community of lactic acid and acetic acid bacteria and yeasts confined in a matrix of proteins and polysaccharides. The present review summarizes the role of kefir micro-organisms in grain assembly and in the beneficial properties attributed to kefir. The use of both culture-dependent and independent methods has made possible to determine the micro-organisms that constitute this ecosystem. Kefir consumption has been associated with a wide range of functional and probiotic properties that could be attributed to the micro-organisms present in kefir and/or to the metabolites synthetized by them during milk fermentation. In this context, the role of micro-organisms in kefir health promoting properties is discussed with particular attention to the contribution of yeast as well as bioactive metabolites such as lactic and acetic acid, exopolysaccharides and bioactive peptides. Even though many advances on the knowledge of this ancient fermented milk have been made, further studies are necessary to elucidate the complex nature of the kefir ecosystem.

Association Between Depressive Symptom Patterns and Clinical Profiles Among Persons Living with HIV.

To describe patterns of depressive symptoms across 10-years by HIV status and to determine the associations between depressive symptom patterns, HIV status, and clinical profiles of persons living with HIV from the Multicenter AIDS Cohort Study (N = 980) and Women's Interagency HIV Study (N = 1744). Group-based trajectory models were used to identify depressive symptoms patterns between 2004 and 2013. Multinomial logistic regressions were conducted to determine associations of depression risk patterns. A 3-group model emerged among HIV-negative women (low: 58%; moderate: 31%; severe: 11%); 5-groups emerged among HIV-positive women (low: 28%; moderate: 31%; high: 25%; decreased: 7%; severe: 9%). A 4-group model emerged among HIV-negative (low: 52%; moderate: 15%; high: 23%; severe: 10%) and HIV-positive men (low: 34%; moderate: 34%; high: 22%; severe: 10%). HIV+ women had higher odds for moderate (adjusted odds ratio [AOR] 2.10, 95% CI 1.63-2.70) and severe (AOR 1.96, 95% CI 1.33-2.91) depression risk groups, compared to low depression risk. HIV+ men had higher odds for moderate depression risk (AOR 3.23, 95% CI 2.22-4.69), compared to low risk. The Framingham Risk Score, ART use, and unsuppressed viral load were associated with depressive symptom patterns. Clinicians should consider the impact that depressive symptoms may have on HIV prognosis and clinical indicators of comorbid illnesses.

Risk of End-Stage Renal Disease in HIV-Positive Potential Live Kidney Donors.

New federal regulations allow HIV-positive individuals to be live kidney donors; however, potential candidacy for donation is poorly understood given the increased risk of end-stage renal disease (ESRD) associated with HIV infection. To better understand this risk, we compared the incidence of ESRD among 41 968 HIV-positive participants of North America AIDS Cohort Collaboration on Research and Design followed for a median of 5 years with the incidence of ESRD among comparable HIV-negative participants of National Health and Nutrition Examination III followed for a median of 14 years. We used risk associations from multivariable Cox proportional hazards regression to derive cumulative incidence estimates for selected HIV-positive scenarios (no history of diabetes, hypertension, AIDS, or hepatitis C virus coinfection) and compared these estimates with those from similarly selected HIV-negative scenarios. For 40-year-old HIV-positive individuals with health characteristics that were similar to those of age-matched kidney donors, viral load <400 copies/mL, and CD4+ count ≥500 cells/µL, the 9-year cumulative incidence of ESRD was higher than that of their HIV-negative peers, yet still low: 2.5 versus 1.1 per 10 000 among white women, 3.0 versus 1.3 per 10 000 among white men, 13.2 versus 3.6 per 10 000 among black women, and 15.8 versus 4.4 per 10 000 among black men. HIV-positive individuals with no comorbidities and well-controlled disease may be considered low-risk kidney donor candidates.

18F-Fluorodeoxyglucose Positron Emission Tomography Parameters can Predict Long-Term Outcome Following Trimodality Treatment for Oesophageal Cancer.

AIMS: 18F-fluorodeoxyglucose positron emission tomography/computed tomography (18FDG-PET/CT) is routinely used for the pre-treatment staging of oesophageal or gastro-oesophageal junction cancers (EGEJC). The aim of this study was to identify objective 18FDG-PET/CT-derived parameters that can aid in predicting the patterns of recurrence and prognostication in patients with EGEJC. PATIENTS AND METHODS: EGEJC patients referred for consideration of preoperative chemoradiation therapy were identified and clinicopathological data were collected. 18FDG-PET/CT imaging data were reviewed and correlated with treatment outcomes. Maximum standardised uptake value (SUVmax), metabolic tumour volume (MTV) and total lesion glycolysis were assessed and association with recurrence-free survival (RFS), locoregional recurrence-free survival (LR-RFS), oesophageal cancer-specific survival (ECSS) and overall survival were evaluated using receiver operating characteristic curves, as well as Cox regression and Kaplan-Meier models. RESULTS: In total, 191 EGEJC patients completed trimodality treatment and 164 with 18FDG-PET/CT data were included in this analysis. At the time of analysis, 15 (9.1%), 70 (42.7%) and two (1.2%) patients were noted to have locoregional, distant and both locoregional and distant metastases, respectively. The median RFS was 30 months (9.6-50.4) and the 5-year RFS was 31.1%. The 5-year overall survival and ECSS were both noted to be 34.8%. Pre-treatment MTV25 > 28.5 cm3 (P = 0.029), MTV40 > 12.4 cm3 (P = 0.018) and MTV50 > 10.2 cm3 (P = 0.005) predicted for worse LR-RFS, ECSS and overall survival for MTV definition of voxels ≥25%, 40% and 50% of SUVmax. CONCLUSION: 18FDG-PET/CT parameters MTV and total lesion glycolysis are useful prognostic tools to predict for LR-RFS, ECSS and overall survival in EGEJC. MTV had the highest accuracy in predicting clinical outcomes. The volume cut-off points we identified for different MTV thresholds predicted outcomes with significant accuracy and may potentially be used for decision making in clinical practice.

Does Loosening the Inclusion Criteria of the CROSS Trial Impact Outcomes in the Curative-Intent Trimodality Treatment of Oesophageal and Gastroesophageal Cancer Patients?

AIM: To determine the efficacy of preoperative chemoradiotherapy as per the CROSS protocol for oesophageal/gastroesophageal junction cancer (OEGEJC), when expanded to patients outside of the inclusion/exclusion criteria defined in the original clinical trial. MATERIALS AND METHODS: Data were collected retrospectively on 229 OEGEJC patients referred for curative-intent preoperative chemoradiotherapy. Outcomes including pathological complete response (pCR), overall survival (OS), cancer-specific survival and recurrence-free survival (RFS) of patients who met CROSS inclusion criteria (MIC) versus those who failed to meet criteria (FMIC) were determined. RESULTS: In total, 42.8% of patients MIC, whereas 57.2% FMIC; 16.6% of patients did not complete definitive surgery. The MIC cohort had higher rates of pCR, when compared with the FMIC cohort (33.3% versus 20.6%, P = 0.039). The MIC cohort had a better RFS, cancer-specific survival and OS compared with the FMIC cohort (P = 0.006, P = 0.004 and P = 0.009, respectively). Age >75 years and pretreatment weight loss >10% were not associated with a poorer RFS (P = 0.541 and 0.458, respectively). Compared with stage I-III patients, stage IVa was associated with a poorer RFS (hazard ratio (HR) = 2.158; 95% confidence interval (CI) = 1.339-3.480, P = 0.001). Tumours >8 cm in length or >5 cm in width had a trend towards worse RFS (HR = 2.060; 95% CI = 0.993-4.274, P = 0.052). CONCLUSION: Our study showed that the robust requirements of the CROSS trial may limit treatment for patients with potentially curable OEGEJC and can be adapted to include patients with a good performance status who are older than 75 years or have >10% pretreatment weight loss. However, the inclusion of patients with celiac nodal metastases or tumours >8 cm in length or >5 cm in width may be associated with poor outcomes.

Exopolysaccharides produced by Lactobacillus and Bifidobacterium strains abrogate in vitro the cytotoxic effect of bacterial toxins on eukaryotic cells.

AIMS: To evaluate the capability of the exopolysaccharides (EPS) produced by lactobacilli and bifidobacteria from human and dairy origin to antagonize the cytotoxic effect of bacterial toxins. METHODS AND RESULTS: The cytotoxicity of Bacillus cereus extracellular factors on Caco-2 colonocytes in the presence/absence of the EPS was determined by measuring the integrity of the tissue monolayer and the damage to the cell membrane (extracellular lactate dehydrogenase activity). Additionally, the protective effect of EPS against the haemolytic activity of the streptolysin-O was evaluated on rabbit erythrocytes. The EPS produced by Bifidobacterium animalis ssp. lactis A1 and IPLA-R1, Bifidobacterium longum NB667 and Lactobacillus rhamnosus GG were able to counteract the toxic effect of bacterial toxins on the eukaryotic cells at 1mg ml(-1) EPS concentration. The EPS A1 was the most effective in counteracting the effect of B. cereus toxins on colonocytes, even at lower doses (0·5mg ml(-1) ), whereas EPS NB667 elicited the highest haemolysis reduction on erythrocytes. CONCLUSIONS: The production of EPS by lactobacilli and bifidobacteria could antagonize the toxicity of bacterial pathogens, this effect being EPS and biological marker dependent. SIGNIFICANCE AND IMPACT OF THE STUDY: This work allows gaining insight about the mechanisms that probiotics could exert to improve the host health.

Preservation of probiotic strains isolated from kefir by spray drying.

AIMS: This work aims to investigate the survival of Lactobacillus kefir CIDCA 8348, Lactobacillus plantarum CIDCA 83114 and Saccharomyces lipolytica CIDCA 812, all isolated from kefir, during spray drying and subsequent storage. METHODS AND RESULTS: Micro-organisms were grown in De Man, Rogosa, Sharpe (MRS) or yeast medium (YM) medium and harvested in the stationary phase of growth. The thermotolerance in skim milk (D and Z values), the survival of spray drying at different outlet air temperatures and subsequent storage in different conditions during 150 days were studied. The resistance to the heat treatments was higher in Lact. plantarum compared to Lact. kefir and S. lipolytica. The three micro-organisms studied varied considerably in their ability to survive to spray drying processes. Lactobacillus plantarum showed the highest survival rate for all the tested outlet air temperatures and also to the further storage in the dried state. The survival rates of Lact. kefir and S. lipolytica through drying and subsequent storage in the dried state decreased when the drying outlet air temperatures increased. CONCLUSIONS: Spray drying is a suitable method to preserve micro-organisms isolated from kefir grains. A high proportion of cells were still viable after 80 days of storage at refrigerated temperatures. SIGNIFICANCE AND IMPACT OF STUDY: It is the first report about spray-dried probiotic strains isolated from kefir grain and contributes to the knowledge about these micro-organisms for their future application in novel dehydrated products.

Urologic care and progression to end-stage kidney disease: a Chronic Kidney Disease in Children (CKiD) nested case-control study.

INTRODUCTION: Children with chronic kidney disease (CKD) risk progressing to end-stage kidney disease (ESKD). The majority of CKD causes in children are related to congenital anomalies of the kidney and urinary tract, which may be treated by urologic care. OBJECTIVE: To examine the association of ESKD with urologic care in children with CKD. STUDY DESIGN: This was a nested case-control study within the Chronic Kidney Disease in Children (CKiD) prospective cohort study that included children aged 1-16 years with non-glomerular causes of CKD. The primary exposure was prior urologic referral with or without surgical intervention. Incidence density sampling matched each case of ESKD to up to three controls on duration of time from CKD onset, sex, race, age at baseline visit, and history of low birth weight. Conditional logistic regression analysis was performed to estimate rate ratios (RRs) for the incidence of ESKD. RESULTS: Sixty-six cases of ESKD were matched to 153 controls. Median age at baseline study visit was 12 years; 67% were male, and 7% were black. Median follow-up time from CKD onset was 14.9 years. Seventy percent received urologic care, including 100% of obstructive uropathy and 96% of reflux nephropathy diagnoses. Cases had worse renal function at their baseline visit and were less likely to have received prior urologic care. After adjusting for income, education, and insurance status, urology referral with surgery was associated with 50% lower risk of ESKD (RR 0.50 [95% confidence interval [CI] 0.26-0.997), compared to no prior urologic care (Figure). After excluding obstructive uropathy and reflux nephropathy diagnoses, which were highly correlated with urologic surgery, the association was attenuated (RR 0.72, 95% CI 0.24-2.18). DISCUSSION: In this study, urologic care was commonly but not uniformly provided to children with non-glomerular causes of CKD. Underlying specific diagnoses play an important role in both the risk of ESKD and potential benefits of urologic surgery. CONCLUSION: Within the CKiD cohort, children with non-glomerular causes of CKD often received urologic care. Urology referral with surgery was associated with lower risk of ESKD compared to no prior urologic care but depended on specific underlying diagnoses.

Protective action of Lactobacillus kefir carrying S-layer protein against Salmonella enterica serovar Enteritidis.

Eight Lactobacillus kefir strains isolated from different kefir grains were tested for their ability to antagonize Salmonella enterica serovar Enteritidis (Salmonella enteritidis) interaction with epithelial cells. L. kefir surface properties such as autoaggregation and coaggregation with Salmonella and adhesion to Caco-2/TC-7 cells were evaluated. L. kefir strains showed significantly different adhesion capacities, six strains were able to autoaggregate and four strains coaggregated with Salmonella. Coincubation of Salmonella with coaggregating L. kefir strains significantly decreased its capacity to adhere to and to invade Caco-2/TC-7 cells. This was not observed with non coaggregating L. kefir strains. Spent culture supernatants of L. kefir contain significant amounts of S-layer proteins. Salmonella pretreated with spent culture supernatants (pH 4.5-4.7) from all tested L. kefir strains showed a significant decrease in association and invasion to Caco-2/TC-7 cells. Artificially acidified MRS containing lactic acid to a final concentration and pH equivalent to lactobacilli spent culture supernatants did not show any protective action. Pretreatment of this pathogen with spent culture supernatants reduced microvilli disorganization produced by Salmonella. In addition, Salmonella pretreated with S-layer proteins extracted from coaggregating and non coaggregating L. kefir strains were unable to invade Caco-2/TC-7 cells. After treatment, L. kefir S-layer protein was detected associated with Salmonella, suggesting a protective role of this protein on association and invasion.

Lactobacillus plantarum CIDCA 8327: An α-glucan producing-strain isolated from kefir grains.

Lactobacillus plantarum CIDCA 8327 is an exopolysaccharide (EPS)-producer strain isolated from kefir with promising properties for the development of functional foods. The aim of the present study was to characterize the structure of the EPS synthesized by this strain grown in skim milk or semidefined medium (SDM). Additionally, genes involved in EPS synthesis were detected by PCR. L. plantarum produces an EPS with a molecular weight of 104Da in both media. When grown in SDM produce an heteropolysaccharide composed mainly of glucose, glucosamine and rhamnose meanwhile the EPS produced in milk was composed exclusively of glucose indicating the influence of the sugar source. FTIR spectra of this EPS showed signals attributable to an α-glucan. Both by 1H NMR and methylation analysis it was possible to determine that this polysaccharide is a branched α-(1→4)-d-glucan composed of 80% linear α-(1→4)-d-glucopyranosyl units and 19% (1→4)-d-glucopyranosyl units substituted at O-3 by single α-d-glucopyranosil residues.

The impact of long-term moderate and heavy alcohol consumption on incident atherosclerosis among persons living with HIV.

BACKGROUND: Level of alcohol consumption is associated with differential risk of atherosclerosis, but little research has investigated this association among HIV+ persons. We evaluated the association between long-term alcohol use and incident atherosclerosis among HIV+ persons. METHODS: We utilized data from HIV+ participants of the Women's Interagency HIV Study (n=483) and the Multicenter AIDS Cohort Study (n=305) without history of cardiovascular disease. Atherosclerosis was assessed two times by B-mode carotid artery ultrasound imaging from 2004 to 2013. Presence of plaque was defined as focal carotid intima-media thickness over 1.5mm. Those with no plaque at baseline and plaque at follow-up were considered incident cases of atherosclerosis. Group-based trajectory models were used to categorize participants into 10-year drinking patterns representing heavy, moderate, or abstinent-low. Multivariable logistic regressions were conducted to assess the association of long-term moderate and heavy use on atherosclerosis, compared to abstinent-low. RESULTS: Heavy alcohol consumption was not statistically significantly associated with risk for incident atherosclerosis in women (AOR 1.10, CI 0.40-3.02) or men (AOR 1.31, CI 0.43-4.00), compared to abstinence-low. Moderate consumption was associated with 54% lower odds for incident disease in men (AOR 0.46, CI 0.21-1.00), but not in women (AOR 1.08, CI 0.58-2.00). In cohort-combined analyses, alcohol consumption was not statistically significantly association with incident atherosclerosis (moderate AOR 0.78, CI 0.48-1.27; heavy AOR 1.33, CI 0.66-2.69). CONCLUSION: Moderate alcohol consumption was associated with a significant protective effect on incident atherosclerosis in men only. No other levels of alcohol consumption significantly predicted atherosclerosis in men and women compared to abstinent-low.

Oral administration of kefiran exerts a bifidogenic effect on BALB/c mice intestinal microbiota.

The activity of kefiran, the exopolysaccharide present in kefir grains, was evaluated on intestinal bacterial populations in BALB/c mice. Animals were orally administered with kefiran and Eubacteria, lactobacilli and bifidobacteria populations were monitored in faeces of mice at days 0, 2, 7, 14 and 21. Profiles obtained by Denaturing Gradient Gel Electrophoresis (DGGE) with primers for Eubacteria were compared by principal component analysis and clearly defined clusters, correlating with the time of kefiran consumption, were obtained. Furthermore, profile analysis of PCR products amplified with specific oligonucleotides for bifidobacteria showed an increment in the number of DGGE bands in the groups administered with kefiran. Fluorescent In Situ Hybridisation (FISH) with specific probes for bifidobacteria showed an increment of this population in faeces, in accordance to DGGE results. The bifidobacteria population was also studied on distal colon content after 0, 2 and 7 days of kefiran administration. Analysis of PCR products by DGGE with Eubacteria primers showed an increment in the number and intensity of bands with high GC content of mice administered with kefiran. Sequencing of DGGE bands confirmed that bifidobacteria were one of the bacterial populations modified by kefiran administration. DGGE profiles of PCR amplicons obtained by using Bifidobacterium or Lactobacillus specific primers confirmed that kefiran administration enhances bifidobacteria, however no changes were observed in Lactobacillus populations. The results of the analysis of bifidobacteria populations assessed on different sampling sites in a murine model support the use of this exopolysaccharide as a bifidogenic functional ingredient.

Kefir-isolated bacteria and yeasts inhibit Shigella flexneri invasion and modulate pro-inflammatory response on intestinal epithelial cells.

The aim of this work was to evaluate the ability of a kefir-isolated microbial mixture containing three bacterial and two yeast strains (MM) to protect intestinal epithelial cells against Shigella flexneri invasion, as well as to analyse the effect on pro-inflammatory response elicited by this pathogen. A significant decrease in S. flexneri strain 72 invasion was observed on both HT-29 and Caco-2 cells pre-incubated with MM. Pre-incubation with the individual strains Saccharomyces cerevisiae CIDCA 8112 or Lactococcus lactis subsp. lactis CIDCA 8221 also reduced the internalisation of S. flexneri into HT-29 cells although in a lesser extent than MM. Interestingly, Lactobacillus plantarum CIDCA 83114 exerted a protective effect on the invasion of Caco-2 and HT-29 cells by S. flexneri. Regarding the pro-inflammatory response on HT-29 cells, S. flexneri infection induced a significant activation of the expression of interleukin 8 (IL-8), chemokine (C-C motif) ligand 20 (CCL20) and tumour necrosis factor alpha (TNF-α) encoding genes (P<0.05), whereas incubation of cells with MM did not induce the expression of any of the mediators assessed. Interestingly, pre-incubation of HT-29 monolayer with MM produced an inhibition of S. flexneri-induced IL-8, CCL20 and TNF-α mRNA expression. In order to gain insight on the effect of MM (or the individual strains) on this pro-inflammatory response, a series of experiments using a HT-29-NF-κB-hrGFP reporter system were performed. Pre-incubation of HT-29-NF-κB-hrGFP cells with MM significantly dampened Shigella-induced activation. Our results showed that the contribution of yeast strain Kluyveromyces marxianus CIDCA 8154 seems to be crucial in the observed effect. In conclusion, results presented in this study demonstrate that pre-treatment with a microbial mixture containing bacteria and yeasts isolated from kefir, resulted in inhibition of S. flexneri internalisation into human intestinal epithelial cells, along with the inhibition of the signalling via NF-κB that in turn led to the attenuation of the inflammatory response.

Selected Lactobacillus strains isolated from sugary and milk kefir reduce Salmonella infection of epithelial cells in vitro.

The isolation of potentially probiotic strains and the subsequent study of their properties are very important steps to gain insight in the health benefits ascribed to sugary and milk kefir. The aim of the present study was to characterise fifteen Lactobacillus strains isolated from these beverages by determining some surface properties and their ability to antagonise enterocyte cell damage after Salmonella infection in vitro. Lactobacillus surface properties were determined by hydrophobicity, autoaggregation, and coaggregation assays with Salmonella. In addition, lactobacilli adhesion to Caco-2/TC-7 cells and the effect on Salmonella invasion were evaluated. Finally, the disassembly of F-actin cytoskeleton on intestinal epithelial cells was assayed in vitro when Salmonella infection was performed in the presence of selected Lactobacillus strains. Ten out of the 15 strains showed a high adhesion capacity to Caco-2/TC-7 cells. Most of the strains were hydrophilic and non-autoaggregating. Strains isolated from sugary kefir were non-coaggregating with Salmonella, while strains Lactobacillus paracasei CIDCA 83120, 83121, 83123, 83124, 8339, 83102 isolated from milk kefir were able to coaggregate after 1 h. L. paracasei CIDCA 8339 and Lactobacillus kefiri CIDCA 83102 were able to diminish Salmonella invasion to the enterocytes. An antagonistic effect on cytoskeleton disruption elicited by the pathogen was also demonstrated. Our results suggest that both strains isolated from milk kefir could be considered as appropriate probiotic candidates.

Inhibitory power of kefir: the role of organic acids.

Milk and MRS broth fermented with kefir grains from different households were examined for inhibitory activity toward gram-negative and gram-positive strains. Fermented milk obtained with 10 g per 100 ml of inoculum (final pH 3.32 to 4.25) and MRS broth fermented with 1 and 10 g per 100 ml of inocula (final pH 4.18 to 5.25) had inhibitory power demonstrated by spot test and agar well diffusion assay. This inhibitory effect could be assigned to the undissociated form of lactic and acetic acid produced during the fermentation process. Kefir supernatants inhibited the growth of Escherichia coli 3 in nutrient broth at 37 degrees C for 24 h. However, supernatants of yogurt or milk artificially acidified with lactic and acetic acids allowed the growth of E. coli 3 in the same conditions. A bacteriostatic effect of milk fermented with kefir grains over E. coli 3 was also demonstrated.

Inhibitory activity of cheese whey fermented with kefir grains.

We investigated the chemical and microbiological compositions of three types of whey to be used for kefir fermentation as well as the inhibitory capacity of their subsequent fermentation products against 100 Salmonella sp. and 100 Escherichia coli pathogenic isolates. All the wheys after fermentation with 10% (wt/vol) kefir grains showed inhibition against all 200 isolates. The content of lactic acid bacteria in fermented whey ranged from 1.04 × 10(7) to 1.17 × 10(7) CFU/ml and the level of yeasts from 2.05 × 10(6) to 4.23 × 10(6) CFU/ml. The main changes in the chemical composition during fermentation were a decrease in lactose content by 41 to 48% along with a corresponding lactic acid production to a final level of 0.84 to 1.20% of the total reaction products. The MIC was a 30% dilution of the fermentation products for most of the isolates, while the MBC varied between 40 and 70%, depending on the isolate. The pathogenic isolates Salmonella enterica serovar Enteritidis 2713 and E. coli 2710 in the fermented whey lost their viability after 2 to 7 h of incubation. When pathogens were deliberately inoculated into whey before fermentation, the CFU were reduced by 2 log cycles for E. coli and 4 log cycles for Salmonella sp. after 24 h of incubation. The inhibition was mainly related to lactic acid production. This work demonstrated the possibility of using kefir grains to ferment an industrial by-product in order to obtain a natural acidic preparation with strong bacterial inhibitory properties that also contains potentially probiotic microorganisms.

Polysaccharide production by kefir grains during whey fermentation.

Fermentation of deproteinised whey with kefir grains CIDCA AGK1 was studied focusing on polysaccharide production from lactose. Kefir grains were able to acidify whey at different rates depending on the grain/whey ratio. During fermentation, kefir grains increased their weight and a water-soluble polysaccharide was released to the media. Exopolysaccharide concentration increased with fermentation time, reaching values of 57.2 and 103.4 mg/l after 5 days of fermentation in cultures with 10 and 100 g kefir grains/l, respectively. The polysaccharide fraction quantified after fermentation corresponded to the soluble fraction, because part of the polysaccharide became a component of the grain. Weight of kefir grains varied depending on the time of fermentation. Polysaccharide production was affected by temperature. Although the highest concentration of polysaccharide in the media was observed at 43 degrees C at both grain/whey ratios, the weight of the grains decreased in these conditions. In conclusion, kefir grains were able to acidify deproteinised whey, reducing lactose concentration, increasing their weight and producing a soluble polysaccharide.