Epp Sepp

The intestinal microflora in allergic Estonian and Swedish 2-year-old children.

The prevalence of allergic diseases seems to have increased particularly over the past 35–40 years. Furthermore, allergic disease is less common among children in the formerly socialist countries of central and Eastern Europe as compared with Western Europe. It has been suggested that a reduced microbial stimulation during infancy and early childhood would result in a slower postnatal maturation of the immune system and development of an optimal balance between TH1‐ and TH2‐like immunity.

Intestinal microbiota and immunoglobulin E responses in 5‐year‐old Estonian children

Background Over the last few decades, several studies from different parts of the world have indicated an increasing prevalence of allergic diseases. This has been related to environmental factors, like changes of microbial pressure. Our previous studies have demonstrated differences in the intestinal microbiota between allergic and non‐allergic children.

Development of intestinal microflora during the first month of life in Estonian and Swedish infants

Little is known about possible regional differences in the development of the intestinal microflora in infants. The aim of the present study was therefore to compare the development of the microflora in Estonian and Swedish infants during the first month of life. The intestinal microflora of one week old and one month old Estonian (n=20) and Swedish (n=20) infants was studied by quantitative culture of faecal samples. All newborns were delivered vaginally and fed with breast milk during the first month of life. At one week of age the counts of aerobes (coagulase negative staphylococci, enterococci, enterobacteria) were higher in Estonian than in Swedish newborns. The counts of lactobacilli and candida increased in the Estonian infants over the first month of life, while the counts of enterobacteria increased in Swedish infants. At one month of age, the Estonian infants were more frequently colonised with lactobacilli than the Swedish infants and the counts were higher. Our data indicate that previously described differences in intestinal microflora of Estonian and Swedish 1 to 2 year old children are present already at the first month of life.

Intestinal Lactobacillus sp. is associated with some cellular and metabolic characteristics of blood in elderly people.

The higher counts or particular groups (Firmicutes/Bacteroidetes) of intestinal microbiota are related to host metabolic reactions, supporting a balance of human ecosystem. We further explored whether intestinal lactobacilli were associated with some principal cellular and metabolic markers of blood in 38 healthy >65-year-old persons. The questionnaire, routine clinical and laboratory data of blood indices as much as the oxidized low-density lipoprotein (ox-LDL) and baseline diene conjugates in low-density lipoprotein (BDC-LDL) of blood sera were explored. The PCR-based intestinal Lactobacillus sp. composition and counts of cultivable lactobacilli (LAB) were tested. The facultative heterofermentative lactobacilli (Lactobacillus casei and Lactobacillus paracasei) were the most frequent (89 and 97%, respectively) species found, while Lactobacillus acidophilus, Lactobacillus plantarum and Lactobacillus reuteri were present in almost half of the elderly persons. The number of species simultaneously colonizing the individuals ranged from 1 to 7 (median 4). In elderly consuming probiotics the LAB counts were significantly higher than in these not consuming (median 7.8, range 4.2-10.8 vs. median 6.3, range 3.3-9.7 log cfu/g; p=0.005), adjusted (OR=1.71, CI95 1.04-2.82; p=0.035) for age and body mass index (BMI). The colonization by L. acidophilus was negatively related (r=-0.367, p=0.0275) to L. reuteri, staying significant after adjusting for age, sex and BMI (OR=0.16, CI95 0.04-0.73; p=0.018). However, the blood glucose concentration showed a tendency for a negative correlation for colonization with Lactobacillus fermentum (r=-0.309, p=0.062) adjusted for BMI (Adj. R(2)=0.181; p=0.013) but not for age and sex. The higher white blood cells (WBC) count was positively related (r=0.434, p=0.007) to presence of Lactobacillus reuteri adjusted for age, sex and BMI (Adj. R(2)=0.193, p=0.027). The lower values of ox-LDL were predicted by higher counts of cultivable lactobacilli adjusted by sex, age and BMI (r = -0.389, p = 0.016; Adj. R(2)=0.184 p=0.029). In conclusion, the pilot study of elderly persons shows that the intestinal lactobacilli are tightly associated with WBC count, blood glucose and content of ox-LDL which all serve as risk markers in pathogenesis of inflammation, metabolic syndrome and cardiovascular disease (CVD).

Persistence of Escherichia coli Clones and Phenotypic and Genotypic Antibiotic Resistance in Recurrent Urinary Tract Infections in Childhood

ABSTRACT We assessed the clonality of consecutive Escherichia coli isolates during the course of recurrent urinary tract infections (RUTI) in childhood in order to compare clonality with phenotypic antibiotic resistance patterns, the presence of integrons, and the presence of the sul1, sul2, and sul3 genes. Altogether, 78 urinary E. coli isolates from 27 children, who experienced recurrences during a 1-year follow-up after the first attack of acute pyelonephritis, were investigated. The MICs of sulfamethoxazole, trimethoprim-sulfamethoxazole (SXT), ampicillin, cefuroxime, cefotaxime, and gentamicin and the presence or absence of the intI gene for class 1 integrons and the sulfamethoxazole resistance-encoding genes sul1, sul2, and sul3 were determined. All E. coli strains were genotyped by pulsed-field gel electrophoresis. There were no significant differences in the prevalences of resistance to beta-lactams and SXT between initial and consecutive E. coli isolates (41 versus 45% and 41 versus 29%, respectively). However, the E. coli strains obtained after SXT administration more frequently carried two or more sul genes than the nonexposed strains (9/21 [43%] versus 11/57 [19%], respectively; P = 0.044). In 78% of the patients, the recurrence of unique clonal E. coli strains alone or combined with individual strains was detected. Phenotypic resistance and the occurrence of sul genes were more stable in clonal strains than in individual strains (odds ratios, 8.7 [95% confidence interval {95% CI}, 1.8 to 40.8] and 4.4 [95% CI, 1.1 to 17.7], respectively). Thus, in children with RUTIs, the majority of E. coli strains from consecutive episodes are unique persisting clones, with rare increases in the initially high antimicrobial resistance, the presence of sul genes, and the presence of integrons.

Risk factors associated with gut and nasopharyngeal colonization by common Gram-negative species and yeasts in neonatal intensive care units patients.

AIM To characterize dynamics of mucosal colonization of neonates by common aerobic Gram negative species and Candida spp. and to identify independent perinatal, neonatal, and environmental factors influencing the colonization process. STUDY DESIGN The nasopharyngeal (n=1145) and rectal (n=1242) swabs were collected on admission and thereafter twice a week in neonates with risk factors of early onset sepsis (n=276) admitted within the first 72 h of life. The association between colonization by different microbes and a total of 22 predefined risk factors was assessed using univariate and multiple logistic regression analyses. RESULTS Throughout the study about half of the patients had rectal (55.8%) or nasopharyngeal colonization (42.8%) with common Gram-negative microorganisms. Colonization dynamics and risk factors were in general similar for a given bacterial species in both mucosal sites; nonfermentative microbes more often found in nasopharyngeal swabs and Enterobacteriaceae in rectal swabs. All organisms except Escherichia coli were influenced by the duration of intensive care unit stay but other risk factors were species specific, perhaps reflecting their mode of acquisition. While colonization by E. coli and Candida albicans was associated with perinatal factors like term birth, vaginal delivery, and breast milk feeding; colonization by Klebsiella pneumoniae, Enteribacter cloacae, Acinetobacter spp. and non-albicans Candida spp. were mostly determined by hospital environment (treatment unit and period, artificial interventions and their duration) and gestation age ≤ 28 weeks. CONCLUSIONS The knowledge of risk factor profiles may permit the development of strategies to prevent heavy colonization and subsequent invasive disease in high risk infants.

Quantification of Clostridium difficile in Antibiotic-Associated-Diarrhea Patients

ABSTRACT Comparing culture- and non-culture-based methods for quantifying Clostridium difficile in antibiotic-associated-diarrhea patients, we found that the real-time PCR method correlated well with quantitative culture and was more sensitive. A positive association between the population levels of C. difficile and the presence of its toxins was found.

Colonization of infants and hospitalized patients with Clostridium difficile and lactobacilli.

Clostridium difficile is the most frequent agent of nosocomial diarrhea in industrialized countries. Approximately 10%–25% of hospitalized patients become colonized by C. difficile [1]. Among nonhospitalized adults, the rate of carriage of C. difficile in the intestinal tract varies in different countries, ranging from 2% in Sweden to 15% in Japan [2]. Asymptomatic carriage of C. difficile is more common among infants and young children, ranging from 15% to 63% [2]. It is generally agreed that the disruption of the indigenous intestinal microflora is an essential prerequisite for colonization by C. difficile. The presence of lactobacilli as a component of the indigenous intestinal microflora has traditionally been associated with resistance to colonization with intestinal pathogens [3, 4]. However, it is not clear if the counts of lactobacilli differ significantly between persons who are colonized by C. difficile and those who are not. The aim of our study was to compare the prevalence and counts of C. difficile and intestinal lactobacilli in Estonian and Swedish children and to compare the counts of lactobacilli in hospitalized Estonian patients in relation to the presence of C. difficile. We investigated the fecal microflora of 27 healthy Estonian infants and 29 healthy Swedish infants (all 1 year of age) and 34 consecutive patients (66 fecal samples) in the neurological intensive care unit of Tartu University Hospital (Tartu, Estonia). The patients had been hospitalized because of neurological surgery or brain trauma for periods ranging from 2 days to 5 months (median duration, 11 days) before sampling. These patients had been treated with up to nine different antimicrobial agents (median number, four; some had not received any agents). The study was done during two 2-month periods separated by a 10-month interval. To detect C. difficile in these hospitalized patients, the fecal samples were seeded after alcohol shock into cefoxitin-cycloserine-fructose agar and incubated anaerobically for 4 days. In addition, the counts of C. difficile and lactobacilli in all samples from the infants and in 20 randomized samples from the hospitalized patients that were collected in the second period were determined as described previously [5]. Ten (35%) of 29 Swedish infants and only one (4%) of 27 Estonian infants were colonized with C. difficile (P õ .05; figure 1A). Furthermore, counts of C. difficile in feces were significantly higher in Swedish infants than in Estonian infants (Põ .05; figure 1B). However, both the prevalence and counts of lactobacilli were significantly higher among the Estonian infants than among the Swedish infants (P õ .005). None of the 21 hospitalized patients was colonized by C. difficile during the first 2-month observation period, whereas seven of 13 patients were found to be colonized during the second period. The Figure 1. A, Frequency of the occurrence of Clostridium difficile counts of C. difficile varied from 4.3 log cfu/g to 8.1 log cfu/g and lactobacilli in Estonian (h) and Swedish ( ) infants. B, Ranges (median count, 7.0 log cfu/g). The counts of lactobacilli were and medians (j) of counts of C. difficile and lactobacilli in Estonian (h) and Swedish ( ) infants.

The occurrence of antimicrobial resistance and class 1 integrons among commensal Escherichia coli isolates from infants and elderly persons

BackgroundThe aim of our study was to compare the presence of the intI1 gene and its associations with the antibiotic resistance of commensal Escherichia coli strains in children with/without previous antibiotic treatments and elderly hospitalized/healthy individuals.MethodsOne-hundred-and-fifteen intestinal E. coli strains were analyzed: 30 strains from 10 antibiotic-naive infants; 27 from 9 antibiotic-treated outpatient infants; 30 from 9 healthy elderly volunteers; and 28 from 9 hospitalized elderly patients. The MIC values of ampicillin, cefuroxime, cefotaxime, gentamicin, ciprofloxacin, and sulfamethoxazole were measured by E-test and IntI1 was detected by PCR.ResultsOut of the 115 strains, 56 (49%) carried class 1 integron genes. Comparing persons without medical interventions, we found in antibiotic-naive children a significantly higher frequency of integron-bearing strains and MIC values than in healthy elderly persons (53% versus 17%; p < 0.01). Evaluating medical interventions, we found a higher resistance and frequency of integrons in strains from hospitalized elderly persons compared with non-hospitalized ones. Children treated with antibiotics had strains with higher MIC values (when compared with antibiotic-naive ones), but the integron-bearing in strains was similar. In most cases, the differences in resistance between the groups (integron-positive and negative strains separately) were higher than the differences between integron-positive and negative strains within the groups.ConclusionThe prevalence of integrons in commensal E. coli strains in persons without previous medical intervention depended on age. The resistance of integron-carrying and non-carrying strains is more dependent on influencing factors (hospitalization and antibiotic administration) in particular groups than merely the presence or absence of integrons.

Impact of empiric antibiotic regimen on bowel colonization in neonates with suspected early onset sepsis

The purpose of this study was to compare the impact of ampicillin and penicillin used for empiric treatment of early onset sepsis (EOS) on initial gut colonization by aerobic and facultative anaerobic microorganisms. A cluster-randomized, two-center, switch-over study was conducted in two paediatric intensive care units in Estonia and included 276 neonates. Rectal swabs were collected twice a week until discharge or day 60. Colonizing microbes were identified on species level and tested for ampicillin resistance (AR). The number of patients colonized with Gram negative microorganisms and Candida spp was similar in both treatment arms but ampicillin resulted in longer colonization duration (CD) of K. pneumonia (p = 0.012), AR Serratia spp (p = 0.012) and Candida spp (p = 0.02) and penicillin in that of AR Acinetobacter spp (p = 0.001). As for Gram positive microorganisms penicillin treatment was associated with a greater number of colonized patients and higher CD of Enterococcus spp and S. aureus but lower ones of S. haemolyticus and S. hominis. Influence of ampicillin and penicillin on initial gut colonization is somewhat different but these differences are of low clinical relevance and should not be a limiting step when choosing between these two antibiotics for the empiric treatment of EOS.