Measurements and predictions of thermophoretic soot deposition.

A thin laminar flow channel with a transverse temperature gradient was used to examine thermophoretic deposition of soot aerosol particles in experiments and modeled in Fire Dynamics Simulator (FDS) simulations. Conditions investigated included three flowrates, with nominal Reynolds number based on the hydraulic diameter of 55, 115 and 230, and two applied temperature gradients, nominally 10 °C/mm and 20 °C/mm, with repeats. Soot was generated from a propene diffusion flame. The burner exhaust was mixed with dilution air, and most large agglomerates greater than 1 µm aerodynamic diameter were removed prior to the channel inlet. The expected thermophoretic velocity of the aerosol was calculated from the applied temperature gradient. A calculated deposition velocity was determined from the mass of deposition, the channel inlet soot concentration, and the exposure time. Uniform soot deposition allowed targets to be used to measure the mass of deposition on the cold side of the channel. The mass of deposition was also determined by subtracting the mass of soot exiting the channel from the mass of soot entering the channel during the exposure time. The deposition velocities from these two methods generally agreed with the thermophoretic velocity and with each other. The deposition mass predicted by the FDS model also compared well with the experiments in most cases. The disagreements for the lowest flow rate cases are attributed to buoyant flow effects adding uncertainty to the actual temperature gradients present in the channel. (The opinions, findings, and conclusions expressed in this paper are the authors' and do not represent the views or policies of NIST or the United States Government.).

Randomized double-blind controlled trial of bovine lactoferrin for prevention of diarrhea in children.

OBJECTIVE: To determine the effect of bovine lactoferrin (bLF) on prevention of diarrhea in children. STUDY DESIGN: We conducted a community-based randomized double-blind placebo controlled trial comparing supplementation with bLF vs 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: Five hundred fifty-five children were randomized: 277 to bLF and 278 to placebo; 65 dropped out; 147 894 doses were administered (92% compliance). Overall there were 91 446 child-days of observation and 1235 diarrhea episodes lasting 6219 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 bLF and placebo, respectively (P = .375). However, the diarrhea longitudinal prevalence was lower in the bLF group vs placebo (6.6% vs 7.0%, P = .017), as well as the median duration of episodes (4.8 vs 5.3 days, P = .046), proportion of episodes with moderate or severe dehydration (1.0% vs 2.6%, P = .045), and liquid stools load (95.0 vs 98.6) liquid stools/child/year, P < .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 LF.

Clinical studies of lactoferrin in children.

Much has been learned in recent years about the mechanisms by which breastfeeding improves child health and survival. However, there has been little progress in using these insights to improve pediatric care. The aim of this study was to review all clinical studies of lactoferrin (LF) in children in an effort to determine which interventions may improve pediatric care or require further research. We conducted a systematic and critical review of published literature and found 19 clinical studies that have used human or bovine LF for different outcomes: iron metabolisms and anemia (6 studies), fecal flora (5 studies), enteric infections (3 studies), common pediatric illnesses (1 study), immunomodulation (3 studies), and neonatal sepsis (1 study). Although the efficacies have varied in each trial, the main finding of all published studies is the safety of the intervention. Protection against enteric infections and neonatal sepsis are the most likely biologically relevant activities of LF in children. Future studies on neonatal sepsis should answer critically important questions. If the data from these sepsis studies are proven to be correct, it will profoundly affect the treatment of low birth weight neonates and will aid in the reduction of child mortality worldwide.

Quantitative real-time polymerase chain reaction for enteropathogenic Escherichia coli: a tool for investigation of asymptomatic versus symptomatic infections.

BACKGROUND: Enteropathogenic Escherichia coli (EPEC) strains are pediatric pathogens commonly isolated from both healthy and sick children with diarrhea in areas of endemicity. The aim of this study was to compare the bacterial load of EPEC isolated from stool samples from children with and without diarrhea to determine whether bacterial load might be a useful tool for further study of this phenomenon. METHODS: EPEC was detected by polymerase chain reaction (PCR) of colonies isolated on MacConkey plates from 53 diarrheal and 90 healthy children aged <2 years. DNA was isolated from stool samples by cetyltrimethylammonium bromide extraction. To standardize quantification by quantitative real-time PCR (qRT-PCR), the correlation between fluorescence threshold cycle and copy number of the intimin gene of EPEC E2348/69 was determined. RESULTS: The detection limit of qRT-PCR was 5 bacteria/mg stool. The geometric mean load in diarrhea was 299 bacteria/mg (95% confidence interval [CI], 77-1164 bacteria/mg), compared with 29 bacteria/mg (95% CI, 10-87 bacteria/mg) in control subjects (P = .016). Bacterial load was significantly higher in children with diarrhea than in control subjects among children <12 months of age (178 vs 5 bacteria/mg; P = .006) and among children with EPEC as the sole pathogen (463 vs 24 bacteria/mg; P = .006). CONCLUSIONS: EPEC load measured by qRT-PCR is higher in diarrheal than in healthy children. qRT-PCR may be useful to study the relationship between disease and colonization in settings of endemicity.

Fecal leukocytes in children infected with diarrheagenic Escherichia coli.

The purpose of this study was to determine the presence and quantity of fecal leukocytes in children infected with diarrheagenic Escherichia coli and to compare these levels between diarrhea and control cases. We analyzed 1,474 stool samples from 935 diarrhea episodes and 539 from healthy controls of a cohort study of children younger than 2 years of age in Lima, Peru. Stools were analyzed for common enteric pathogens, and diarrheagenic E. coli isolates were studied by a multiplex real-time PCR. Stool smears were stained with methylene blue and read by a blinded observer to determine the number of polymorphonuclear leukocytes per high-power field (L/hpf). Fecal leukocytes at >10 L/hpf were present in 11.8% (110/935) of all diarrheal episodes versus 1.1% (6/539) in controls (P < 0.001). Among stool samples with diarrheagenic E. coli as the only pathogen isolated (excluding coinfection), fecal leukocytes at >10 L/hpf were present in 8.5% (18/212) of diarrhea versus 1.3% (2/157) of control samples (P < 0.01). Ninety-five percent of 99 diarrheagenic E. coli diarrhea samples were positive for fecal lactoferrin. Adjusting for the presence of blood in stools, age, sex, undernutrition, and breastfeeding, enterotoxigenic E. coli (ETEC) isolation as a single pathogen, excluding coinfections, was highly associated with the presence of fecal leukocytes (>10 L/hpf) with an odds ratio (OR) of 4.1 (95% confidence interval [CI], 1.08 to 15.51; P < 0.05). Although diarrheagenic E. coli was isolated with similar frequencies in diarrhea and control samples, clearly it was associated with a more inflammatory response during symptomatic infection; however, in general, these pathogens elicited a mild inflammatory response.

Effect of bovine lactoferrin in Salmonella ser. Typhimurium infection in mice.

Lactoferrin (LF) has in vitro antimicrobial activity against Gram-negative bacteria. Salmonella enterica subsp. enterica serovar Typhimurium causes systemic infection and acute diarrhea in humans, mainly in children younger than 2 years of age. The aim of the study was to determine the in vivo effect of bovine LF in Salmonella ser. Typhimurium infection in mice. 58 BALB/c mice were employed. Two hours before the infection with 300 microl of 10(7) CFU of Salmonella ser. Typhimurium, 29 mice received LF (2 mg) and 29 placebo (buffer). After the infection, the mice received LF (10 mg/ml) ad libitum or buffer, respectively, for 7 days. Mortality, weight and clinical signs (piloerection, hunched position and reduced movement) were monitored daily. The degree of inflammation and necrosis in the intestine, liver, spleen and brain were studied with a blinded observer. The mortality in the control group (8/29) was higher than in the LF group (1/29) (Kapplan Meier P < 0.05). From the third day post-infection the control group were significantly more symptomatic (P < 0.05). The blood culture for Salmonella spp. was positive for all mice studied in the control group (17/17), but positive in the LF group in only 6/17 animals (P < 0.05). In the LF group, the pathologic studies show less inflammation and focal necrosis in the four organs studied, with the greatest difference found in the intestine. Bovine LF protects against Salmonella ser. Typhimurium infection in mice, reducing the severity, mortality and the degree of inflammation of this infection.

Age-related susceptibility to infection with diarrheagenic Escherichia coli among infants from Periurban areas in Lima, Peru.

BACKGROUND: Diarrheagenic Escherichia coli strains are being recognized as important pediatric enteropathogens worldwide. However, it is unclear whether there are differences in age-related susceptibility to specific strains, especially among infants. METHODS: We conducted a passive surveillance cohort study of diarrhea that involved 1034 children aged 2-12 months in Lima, Peru. Control stool samples were collected from randomly selected children without diarrhea. All samples were analyzed for common enteric pathogens and for diarrheagenic E. coli with use of multiplex real-time polymerase chain reaction. RESULTS: The most frequently isolated pathogens in 1065 diarrheal episodes were diarrheagenic E. coli strains (31%), including enteroaggregative (15.1%) and enteropathogenic E. coli (7.6%). Diarrheagenic E. coli, Campylobacter species, and rotavirus were more frequently isolated from infants aged >or=6 months. Among older infants, diffusely adherent E. coli and enterotoxigenic E. coli were more frequently isolated from diarrheal samples than from control samples (P <.05). Children aged >or=6 months who were infected with enterotoxigenic E. coli had a 4.56-fold increased risk of diarrhea (95% confidence interval, 1.20-17.28), compared with younger children. Persistent diarrhea was more common in infants aged <6 months (13.5% vs 3.6%; P <.001). Among children with diarrheagenic E. coli-positive samples, coinfections with other pathogens were more common in children with diarrhea than in control children (40.1% vs 15.6%; P <.001). CONCLUSIONS: Diarrheagenic E. coli strains were more frequently isolated in samples from older infants. In this setting with high frequency of pathogen exposure and high frequency of breastfeeding, we hypothesize that the major age-related differences result from decreased exposure to milk-related protective factors and from increased exposure to contaminated food and water.

Development of a multiple gas analyzer using cavity ringdown spectroscopy for use in advanced fire detection.

A portable cavity ringdown spectroscopy (CRDS) apparatus was used to detect effluents from small test fires in the Fire Emulator/Detector Evaluator (FE/DE) and a small room in the Building Fire and Research Laboratory at the National Institute of Standards and Technology (NIST). The output from two lasers is combined to detect four combustion gases, CO, CO(2), HCN, and C(2)H(2), near simultaneously using CRDS. The goal of this work was to demonstrate the feasibility of using a CRDS sensor as a fire detector. Fire effluents were extracted from several test facilities and measurements of CO, CO(2), HCN, and C(2)H(2) were obtained every 25-30 s. In the FE/DE test, peak concentrations of the gases from smoldering paper were 420 parts in 10(6) (ppm) CO, 1600 ppm CO(2), 530 parts in 10(9) (ppb) HCN, and 440 ppb C(2)H(2). Peak gas concentrations from the small room were 270 ppm CO, 2100 ppm CO(2), and 310 ppb C(2)H(2).

Impact of lactoferrin supplementation on growth and prevalence of Giardia colonization in children.

We conducted a randomized, double-blind, placebo-controlled trial comparing supplementation with bovine lactoferrin versus placebo for the prevention of diarrhea in children. Comparison of overall diarrhea incidence and prevalence rates found no significant difference between the 2 groups. However, there was a lower prevalence of colonization with Giardia species and better growth among children in the lactoferrin group.

Detection of diarrheagenic Escherichia coli by use of melting-curve analysis and real-time multiplex PCR.

Diarrheagenic Escherichia coli strains are important causes of diarrhea in children from the developing world and are now being recognized as emerging enteropathogens in the developed world. Current methods of detection are too expensive and labor-intensive for routine detection of these organisms to be practical. We developed a real-time fluorescence-based multiplex PCR for the detection of all six of the currently recognized classes of diarrheagenic E. coli. The primers were designed to specifically amplify eight different virulence genes in the same reaction: aggR for enteroaggregative E. coli, stIa/stIb and lt for enterotoxigenic E. coli, eaeA for enteropathogenic E. coli and Shiga toxin-producing E. coli (STEC), stx(1) and stx(2) for STEC, ipaH for enteroinvasive E. coli, and daaD for diffusely adherent E. coli (DAEC). Eighty-nine of ninety diarrheagenic E. coli and 36/36 nonpathogenic E. coli strains were correctly identified using this approach (specificity, 1.00; sensitivity, 0.99). The single false negative was a DAEC strain. The total time between preparation of DNA from E. coli colonies on agar plates and completion of PCR and melting-curve analysis was less than 90 min. The cost of materials was low. Melting-point analysis of real-time multiplex PCR is a rapid, sensitive, specific, and inexpensive method for detection of diarrheagenic E. coli.

pic gene of enteroaggregative Escherichia coli and its association with diarrhea in Peruvian children.

Enteroaggregative Escherichia coli (EAEC) causes acute and persistent diarrhea among children, HIV-infected patients, and travelers to developing countries. We have searched for 18 genes-encoding virulence factors associated with aggregative adherence, dispersion, biofilm, toxins, serine protease autotransporters of Enterobacteriaceae (SPATEs) and siderophores, analyzed in 172 well-characterized EAEC strains (aggR(+)) isolated from stool samples of 97 children with diarrhea and 75 healthy controls from a passive surveillance diarrhea cohort study in Peru. Eighty-one different genetic profiles were identified, 37 were found only associated with diarrhea and 25 with control samples. The most frequent genetic profile was aggC(+)aatA(+)aap(+)shf(+)fyuA(+), present in 19 strains, including diarrhea and controls. The profile set1A(+)set1B(+)pic(+) was associated with diarrhea (P < 0.05). Of all genes evaluated, the most frequent were aatA (CVD 342) present in 159 strains (92.4%) and fyuA in 157 (91.3%). When EAEC strains were analyzed as a single pathogen (excluding co-infections), only pic was associated with diarrhea (P < 0.05) and with prolonged diarrhea (diarrhea ≥ 7 days) (P < 0.05). In summary, this is the first report on the prevalence of a large set of EAEC virulence genes and its association with diarrhea in Peruvian children. More studies are needed to elucidate the exact role of each virulence factor.

High Prevalence and Increased Severity of Norovirus Mixed Infections Among Children 12-24 Months of Age Living in the Suburban Areas of Lima, Peru.

In an active diarrhea surveillance study of children aged 12-24 months in Lima, Peru, norovirus was the most common pathogen identified. The percentage of mixed (bacterial and noroviral) infections was significantly higher among norovirus-positive samples (53%) than among norovirus-negative samples (12%). The combination of norovirus with the most common bacterial pathogens was associated with increased clinical severity over that of either single-pathogen norovirus or single-pathogen bacterial infections.

Rotavirus seasonal distribution and prevalence before and after the introduction of rotavirus vaccine in a peri-urban community of Lima, Peru.

We evaluated the monthly distribution of rotavirus diarrhea in a cohort of children 12-24 months of age followed as part of a diarrhea clinical trial in a peri-urban community of Lima. We observed a peak of rotavirus diarrhea in the winter months and a decrease in rotavirus prevalence after the introduction of the rotavirus vaccine in Peru.

Poliomyelitis-like syndrome in a child with West Nile virus infection.

West Nile virus (WNV) infections of the central nervous system are very uncommon in U.S. children. We report a child with a poliomyelitis-like presentation in which WNV was the only detected pathogen. WNV has not previously been associated with a poliomyelitis-like presentation in children.

The role of Shiga-toxin-producing Escherichia coli in hemorrhagic colitis and hemolytic uremic syndrome.

The Shiga-toxin-producing E. coli represent a major class of pathogens that have been defined over the last twenty years. They cause distinctive clinical manifestations such as afebrile bloody diarrhea with severe abdominal pain (hemorrhagic colitis) and microangiopathic hemolytic anemia with renal failure (hemolytic uremic syndrome). The most common Shiga-toxin-producing E. coli is serotype O157:H7, although at least one hundred different serotypes share the virulence traits and clinical manifestations with this organism. Understanding the pathophysicology, improving diagnostic tools, and developing a treatment strategy are important areas of ongoing investigations.

Management of children with infection-associated persistent diarrhea.

Diarrhea is the leading cause of death in children younger than 5 years of age; persistent diarrhea accounts for 30 to 50 percent of those deaths in developing countries. Malnutrition, immunosuppression, young age, and an increase in the preceding diarrhea burdens are risk factors for the development of persistent diarrhea. Although many viruses, bacteria, and parasites can produce persistent diarrhea, enteropathogenic Escherichia coli , enteroaggregative E. coli, Giardia, Cryptosporidium , and Cyclospora are the most important of these agents. With currently available tests, identifying a specific cause usually is difficult. Newer sensitive molecular tests must be used for studying the epidemiology of persistent diarrhea in children. Management includes rehydration, adequate diet, micronutrient supplementation, and antimicrobials. Persistent diarrhea seriously affects nutritional status, growth, and intellectual function. Meeting these challenges is profoundly important, particularly in developing countries.

Human milk protective mechanisms.

The survival of a nonimmune infant faced with a new pathogen depends in part on an array of specific and nonspecific human milk factors. Human milk protects via multiple redundant strategies. It targets shared virulence mechanisms to provide cross protection. By interfering with fundamental processes such as attachment to glycoconjugates, human milk anticipates new mutations and new pathogens that utilize carbohydrates as receptors. Furthermore, human milk IgA present appears to reflect long-term maternal immunologic memory. These overlapping protective strategies endow human milk with the unique ability to protect infants from organisms that mutate rapidly and are present in vast numbers.

Multiplex real-time PCR (MRT-PCR) for diarrheagenic.

Diarrheagenic Escherichia coli strains are important causes of diarrhea in children from the developing world and are now being recognized as emerging enteropathogens in the developed world. Current methods of detection are too expensive and labor-intensive for routine detection of these organisms to be practical. We developed a real-time fluorescence-based multiplex PCR for the detection of all six of the currently recognized classes of diarrheagenic E. coli. The primers were designed to specifically amplify eight different virulence genes in the same reaction: aggR for enteroaggregative E. coli (EAEC), stIa/stIb and lt for enterotoxigenic E. coli (ETEC), eaeA for enteropathogenic E. coli (EPEC), stx1 and stx2 for Shiga toxin-producing E. coli (STEC), ipaH for enteroinvasive E. coli (EIEC), and daaD for diffusely adherent E. coli (DAEC).

Detection of secretory immunoglobulin A in human colostrum as mucosal immune response against proteins of the type III secretion system of Salmonella, Shigella and enteropathogenic Escherichia coli.

BACKGROUND: Some enteropathogens use the type III secretion system to secrete proteins that allows them to interact with enterocytes and promote bacterial attachment or intracellular survival. These proteins are Salmonella invasion proteins (Sip), invasion plasmid antigens (Ipa) of Shigella and Escherichia coli secreted proteins (Esp) of enteropathogenic E. coli. There are no previous studies defining the presence of colostral sIgA against all these 3 major enteric pathogens. OBJECTIVE: To evaluate the presence of sIgA in colostrum against proteins of the type III secretion system of Salmonella, Shigella and enteropathogenic E. coli. METHODS: We collected 76 colostrum samples from puerperal women in Lima, Peru. These samples were reacted with type III secretion system proteins extracted from bacterial culture supernatants and evaluated by Western Blot. RESULTS: Antibodies were detected against Salmonella antigens SipA in 75 samples (99%), SipC in 62 (82%) and SipB in 31 (41%); against Shigella antigens IpaC in 70 (92%), IpaB in 68 (89%), IpaA in 66 (87%) and IpaD in 41 (54%); and against enteropathogenic E. coli EspC in 70 (92%), EspB-D in 65 (86%) and EspA in 41 (54%). Ten percent of samples had antibodies against all proteins evaluated and 42% against all except 1 protein. There was no sample negative to all these proteins. CONCLUSIONS: The extraordinarily high frequency of antibodies in colostrum of puerperal women detected in this study against these multiple enteric pathogens shows evidence of immunological memory and prior exposure to these pathogens, in addition to its possible protective role against infection.

Shiga toxin-producing Escherichia coli in Peruvian children with bloody diarrhea.

Shiga toxin-producing Escherichia coli (STEC) is not routinely sought in clinical laboratories in developing counties. Among 131 bloody diarrhea samples in Peruvian children <5 years of age, STEC was found in 9.2% and was associated with absence of fever, an observation that may increase suspicion of these pathogens. Because of the significant prevalence of STEC locally, proper diagnostics methods should be implemented in the region.