Objective To determine the effect of bovine lactoferrin on prevention of diarrhea in children. Study design We conducted a community-based randomized double-blind placebo controlled trial comparing supplementation with bovine lactoferrin versus placebo. Previously weaned children were enrolled at 12–18 months and followed for 6 months with daily home visits for data collection and supplement administration. Anthropometric measures were done monthly. Results 555 children were randomized: 277 to lactoferrin and 278 to placebo; 65 dropped out; 147,894 doses were administered (92% compliance). Overall there were 91,446 child-days of observation and 1,235 diarrhea episodes lasting 6,219 days. The main pathogens isolated during diarrheal episodes were norovirus (35.0%), enteropathogenic E. coli (11.4%), Campylobacter (10.6%), enteroaggregative E. coli (8.4%), enterotoxigenic E. coli (6.9%) and Shigella (6.6%). The diarrhea incidence was not different between groups: 5.4 vs. 5.2 episodes/child/year for lactoferrin and placebo, respectively (p=0.375). However, the diarrhea longitudinal prevalence was lower in the lactoferrin group (6.6% vs. 7.0%, p=0.017) as well as the median duration of episodes (4.8 vs. 5.3 days, p=0.046), proportion of episodes with moderate or severe dehydration (1.0% vs. 2.6%, p=0.045) and liquid stools load (95.0 vs. 98.6) liquid stools/child/year, p<0.001). There were no adverse events related to the intervention. Conclusions Although there was no decrease in diarrhea incidence, longitudinal prevalence and severity were decreased with lactoferrin.

The study was aimed to describe the serotype, mechanisms of antimicrobial resistance, and virulence determinants in Shigella spp. isolated from Peruvian children. Eighty three Shigella spp. were serogrouped and serotyped being established the antibiotic susceptibility. The presence of 12 virulence factors (VF) and integrase 1 and 2, along with commonly found antibiotic resistance genes was established by PCR. S. flexneri was the most relevant serogroup (55 isolates, 66%), with serotype 2a most frequently detected (27 of 55, 49%), followed by S. boydii and S. sonnei at 12 isolates each (14%) and S. dysenteriae (4 isolates, 5%). Fifty isolates (60%) were multi-drug resistant (MDR) including 100% of S. sonnei and 64% of S. flexneri. Resistance levels were high to trimethoprim-sulfamethoxazole (86%), tetracycline (74%), ampicillin (67%), and chloramphenicol (65%). Six isolates showed decreased azithromycin susceptibility. No isolate was resistant to nalidixic acid, ciprofloxacin, nitrofurantoin, or ceftriaxone. The most frequent resistance genes were sul2 (95%), tet(B) (92%), cat (80%), dfrA1 (47%), blaOXA-1 like (40%), with intl1 and intl2 detected in 51 and 52% of the isolates, respectively. Thirty-one different VF profiles were observed, being the ipaH (100%), sen (77%), virA and icsA (75%) genes the most frequently found. Differences in the prevalence of VF were observed between species with S. flexneri isolates, particularly serotype 2a, possessing high numbers of VF. In conclusion, this study highlights the high heterogeneity of Shigella VF and resistance genes, and prevalence of MDR organisms within this geographic region.

Infectious diarrhea can be classified based on its clinical presentation as noninflammatory or inflammatory disease. In developing countries, among inflammatory diarrhea cases, Shigella is the most common cause, followed by Campylobacter and Salmonella. Because the time frame in which treatment choices must be made is short and conventional stool cultures lack good sensitivity, there is a need for a rapid, sensitive, and inexpensive detection technique. The purpose of our study was to develop a multiplex real-time PCR procedure to simultaneously identify Campylobacter spp., Salmonella spp., and Shigella spp. Primers were designed to amplify the invA, ipaH, and 16S rRNA genes simultaneously in a single reaction to detect Salmonella, Shigella, and Campylobacter, respectively. Using this approach, we correctly identified 102 of 103 strains of the targeted enteropathogens and 34 of 34 other pathogens. The melting temperatures were 82.96 ؎ 0.05°C for invA, 85.56 ؎ 0.28°C for ipaH, and 89.21 ؎ 0.24°C for 16S rRNA. The limit of accurate quantification for the assay in stool samples was 10 4 CFU g ؊1 ; however, the limit of detection was 10 3 CFU g ؊1 . This assay is a simple, rapid, inexpensive, and reliable system for the practical detection of these three enteropathogens in clinical specimens. Infectious diarrhea is a global health problem that is still responsible for thousands of deaths worldwide, especially in children (1). It can be classified based on its clinical presentation as one of two syndromes-noninflammatory or inflammatory diarrhea (2). Among cases of inflammatory diarrhea, Shigella is the most common cause, followed by Campylobacter and Salmonella (3). These invasive organisms primarily target the lower bowel; they invade the intestinal mucosa to induce an acute inflammatory reaction and activate cytokines and inflammatory mediators (4).Because the time frame in which treatment choices must be made is short and the conventional stool cultures lack good sensitivity, there is a need for a rapid, sensitive, and inexpensive detection technique. We searched for DNA sequences that were highly conserved between the different species of each genera, and we selected the following genes as targets, invA (invasion A gene) for Salmonella spp., ipaH (invasion plasmid antigen H) for Shigella spp., and 16S rRNA for Campylobacter spp., to develop a fluorescence-based real-time PCR procedure to simultaneously identify these enteropathogens. In this method the post-PCR products are identified based on melting-point curve analysis. We have also standardized the technique to quantify these bacteria directly from stool samples. MATERIALS AND METHODSBacterial strains. A total of 147 enteropathogenic strains (Table 1) were analyzed, including clinical isolates representative of Salmonella spp., Shigella spp., and Campylobacter spp., as well as other enteropathogens. These clinical strains had previously been identified based on serology, biochemical assays, and real-time PCR for the diarrheagenic Escherichia coli (5). In add...

DEC are a major cause of diarrhea in Peruvian children. These pathogens are highly heterogeneous. Additional studies are required to determine the prevalence in rural areas of Peru and in severe diarrhea cases.