Effects of household concrete floors on maternal and child health - the CRADLE trial: a randomised controlled trial protocol.

Introduction: Early life soil-transmitted helminth infection and diarrhea are associated with growth faltering, anemia, impaired child development, and mortality. Exposure to fecally contaminated soil inside the home may be a key contributor to enteric infections, and a large fraction of rural homes in low-income countries have soil floors. The objective of this study is to measure the effect of installing concrete floors in homes with soil floors on child soil-transmitted helminth infection and other maternal and child health outcomes in rural Bangladesh. Methods and analysis: The Cement-based flooRs AnD chiLd hEalth (CRADLE) trial is an individually randomised trial in Sirajganj and Tangail districts, Bangladesh. Households with a pregnant woman, a soil floor, walls that are not made of mud will be eligible, and no plan to relocate for 3 years. We will randomise 800 households to intervention or control (1:1) within geographic blocks of 10 households to account for strong geographic clustering of enteric infection. Laboratory staff and data analysts will be blinded; participants will be unblinded. We will install concrete floors when the birth cohort is in utero and measure outcomes at child ages 3, 6, 12, 18, and 24 months. The primary outcome is prevalence of any soil-transmitted helminth infection (Ascaris lumbricoides, Necator americanus, or Trichuris trichiura) detected by qPCR at 6, 12, 18, or 24 months follow-up in the birth cohort. Secondary outcomes include household floor and child hand contamination with E. coli, extended-spectrum beta-lactamase producing E. coli, and soil-transmitted helminth DNA; child diarrhea, growth, and cognitive development; and maternal stress and depression. Ethics and dissemination: Study protocols have been approved by institutional review boards at Stanford University and the International Centre for Diarrheal Disease Research, Bangladesh (icddr,b). We will report findings on ClinicalTrials.gov, in peer-reviewed publications, and in stakeholder workshops in Bangladesh. Trial registration number: NCT05372068, pre-results.

Azithromycin for Bacterial Watery Diarrhea: A Reanalysis of the AntiBiotics for Children With Severe Diarrhea (ABCD) Trial Incorporating Molecular Diagnostics.

BACKGROUND: Bacterial pathogens cause substantial diarrhea morbidity and mortality among children living in endemic settings, yet antimicrobial treatment is only recommended for dysentery or suspected cholera. METHODS: AntiBiotics for Children with severe Diarrhea was a 7-country, placebo-controlled, double-blind efficacy trial of azithromycin in children 2-23 months of age with watery diarrhea accompanied by dehydration or malnutrition. We tested fecal samples for enteric pathogens utilizing quantitative polymerase chain reaction to identify likely and possible bacterial etiologies and employed pathogen-specific cutoffs based on genomic target quantity in previous case-control diarrhea etiology studies to identify likely and possible bacterial etiologies. RESULTS: Among 6692 children, the leading likely etiologies were rotavirus (21.1%), enterotoxigenic Escherichia coli encoding heat-stable toxin (13.3%), Shigella (12.6%), and Cryptosporidium (9.6%). More than one-quarter (1894 [28.3%]) had a likely and 1153 (17.3%) a possible bacterial etiology. Day 3 diarrhea was less common in those randomized to azithromycin versus placebo among children with a likely bacterial etiology (risk difference [RD]likely, -11.6 [95% confidence interval {CI}, -15.6 to -7.6]) and possible bacterial etiology (RDpossible, -8.7 [95% CI, -13.0 to -4.4]) but not in other children (RDunlikely, -0.3% [95% CI, -2.9% to 2.3%]). A similar association was observed for 90-day hospitalization or death (RDlikely, -3.1 [95% CI, -5.3 to -1.0]; RDpossible, -2.3 [95% CI, -4.5 to -.01]; RDunlikely, -0.6 [95% CI, -1.9 to .6]). The magnitude of risk differences was similar among specific likely bacterial etiologies, including Shigella. CONCLUSIONS: Acute watery diarrhea confirmed or presumed to be of bacterial etiology may benefit from azithromycin treatment. CLINICAL TRIALS REGISTRATION: NCT03130114.

Detection and absolute quantification of Lactiplantibacillus plantarum ATCC 202195 by quantitative real-time PCR.

IMPORTANCE: When administered for seven consecutive days shortly after birth, the probiotic bacterium Lactiplantibacillus plantarum ATCC 202195 plus fructooligosaccharide (FOS) was reported to reduce sepsis and lower respiratory tract infection events during early infancy in a randomized trial in India. Since probiotic effects are often strain specific, strain-level detection and quantification by routine molecular methods enables the monitoring of safety outcomes, such as probiotic-associated bacteremia, and allows for the quality of probiotic interventions to be assessed and monitored (i.e., verify strain identity and enumerate). Despite the potential clinical applications of L. plantarum ATCC 202195, an assay to detect and quantify this strain has not previously been described. Herein, we report the design of primer and probe sequences to detect L. plantarum ATCC 202195 and the development and optimization of a real-time PCR assay to detect and quantify the strain with high specificity and high sensitivity.

Genomic Heterogeneity of Cryptosporidium parvum Isolates From Children in Bangladesh: Implications for Parasite Biology and Human Infection.

Cryptosporidium species are a major cause of diarrhea and associated with growth failure. There is currently only limited knowledge of the parasite's genomic variability. We report a genomic analysis of Cryptosporidium parvum isolated from Bangladeshi infants and reanalysis of sequences from the United Kingdom. Human isolates from both locations shared 154 variants not present in the cattle-derived reference genome, suggesting host-specific adaptation of the parasite. Remarkably 34.6% of single-nucleotide polymorphisms unique to human isolates were nonsynonymous and 8.2% of these were in secreted proteins. Linkage disequilibrium decay indicated frequent recombination. The genetic diversity of C. parvum has potential implications for vaccine and therapeutic design. Clinical Trials Registration. NCT02764918.

Prevalence of Soil-Transmitted Helminths at Baseline and after Albendazole Treatment in the School-Age Children of Forcibly Displaced Myanmar Nationals in Bangladesh.

The forcibly displaced Myanmar nationals (FDMNs) known as Rohingya refugees are the largest group of stateless individuals globally. According to the emergencies humanitarian actors at the United Nations Office for the Coordination of Humanitarian Affairs, the worldwide refugee crisis involving FDMNs is intensifying at the fastest rate in history. Growing public health demands are being exacerbated by current difficulties in addressing poor access to health services, severe food shortages, and a lack of adequate housing. Infectious diseases constitute a major public health emergency in this vulnerable population. A study was carried out in FDMN children to investigate common soil-transmitted helminth (STH) infection at the time of enrollment and prospectively followed-up to 12 months after 2 doses albendazole treatment. At baseline, the prevalence of STH infection with at least one species was found to be 91.7% and 87.3% for Kato-Katz (KK) and quantitative polymerase chain reaction (qPCR) methods, respectively. Similarly, for follow-up children, the overall infection rate was 95.3% and 91.5%, respectively. Trichuris trichiura was the most predominant STH infection by both KK (baseline 87%, follow-up 89.1%) and qPCR (baseline 77.5%, follow-up 82.9%). The overall prevalence of stunting in the children was 37.8% at baseline and rose to 51.3% at 12 months. Alpha-1 antitrypsin (r = 0.13, P = 0.01) and myeloperoxidase (r = 0.12, P = 0.01) levels showed a positive correlation with Aascaris lumbricoides egg count per gram at baseline. An in-depth investigation is urgently needed to identify the underlying protective measures and the root cause of STH infections to improve the health of FDMN children.

Targeting hepatitis B vaccine escape using immunogenetics in Bangladeshi infants.

Hepatitis B virus (HBV) vaccine escape mutants (VEM) are increasingly described, threatening progress in control of this virus worldwide. Here we studied the relationship between host genetic variation, vaccine immunogenicity and viral sequences implicating VEM emergence. In a cohort of 1,096 Bangladeshi children, we identified human leukocyte antigen (HLA) variants associated with response vaccine antigens. Using an HLA imputation panel with 9,448 south Asian individuals DPB1*04:01 was associated with higher HBV antibody responses (p=4.5×10-30). The underlying mechanism is a result of higher affinity binding of HBV surface antigen epitopes to DPB1*04:01 dimers. This is likely a result of evolutionary pressure at the HBV surface antigen 'a-determinant' segment incurring VEM specific to HBV. Prioritizing pre-S isoform HBV vaccines may tackle the rise of HBV vaccine evasion.

Children develop Immunity to cryptosporidiosis in a high transmission intensity area.

Background: Cryptosporidium is one of the top causes of diarrhea in Bangladesh infants. Cryptosporidium infections lead to the production of antibody immune responses, which were associated with a decrease in parasite burden and decreased disease severity in subsequent infections. Methods: We conducted a longitudinal study of cryptosporidiosis from birth to five years of age in an urban slum of Dhaka Bangladesh. We then retrospectively tested the concentration of anti-Cryptosporidium Cp17 or Cp23 IgA in surveillance stool samples collected from 54 children during their first 3 years of life by enzyme-linked immunosorbent assay (ELISA). We also assessed the concentration of both IgA and IgG antibodies specific to Cryptosporidium Cp17 and Cp23 in the concentration of anti-Cryptosporidium Cp17 or Cp23 IgA and IgG antibodies in the children's plasma (1- 5 years). Results: The seroprevalence of both anti- Cp23 and Cp17 antibodies was high at ≤ one year of age and reflected the exposure of these children in this community to cryptosporidiosis. In Bangladesh, the prevalence of cryptosporidiosis is high during the rainy season (June to October) but decreases during the dry season. In younger infants' plasma anti-Cp17 and Cp23 IgG and anti-Cp17 IgA levels were markedly increased during the rainy season in line with the higher initial exposure to the parasite at this time. Both anti-Cp17, anti-Cp23 fecal IgA and the parasite burden declined during repeat infections. Conclusions: We found that anti-Cryptosporidium plasma and fecal antibody levels in children could contribute to the decrease in new infections in this study population.

Specific Cryptosporidium antigens associate with reinfection immunity and protection from cryptosporidiosis.

There is no vaccine to protect from cryptosporidiosis, a leading cause of diarrhea in infants in low- and middle-income countries. Here, we comprehensively identified parasite antigens associated with protection from reinfection. A Cryptosporidium protein microarray was constructed by in vitro transcription and translation of 1,761 C. parvum, C. hominis, or C. meleagridis antigens, including proteins with a signal peptide and/or a transmembrane domain. Plasma IgG and/or IgA from Bangladeshi children longitudinally followed for cryptosporidiosis from birth to 3 years of age allowed for identification of 233 seroreactive proteins. Seven of these were associated with protection from reinfection. These included Cp23, Cp17, Gp900, and 4 additional antigens - CpSMP1, CpMuc8, CpCorA and CpCCDC1. Infection in the first year of life, however, often resulted in no detectable antigen-specific antibody response, and antibody responses, when detected, were specific to the infecting parasite genotype and decayed in the months after infection. In conclusion, humoral immune responses against specific parasite antigens were associated with acquired immunity. While antibody decay over time and parasite genotype-specificity may limit natural immunity, this work serves as a foundation for antigen selection for vaccine design.

Prospective Cohort Study of Cryptosporidium Infection and Shedding in Infants and Their Households.

BACKGROUND: Cryptosporidium spp. are responsible for significant diarrheal morbidity and mortality in under-5 children. There is no vaccine; thus, a focus on prevention is paramount. Prior studies suggest that person-to-person spread may be an important pathway for transmission to young children. Here we describe a longitudinal cohort study of 100 families with infants to determine rates of cryptosporidiosis within households during the coronavirus disease 2019 (COVID-19) pandemic. METHODS: Families living in Mirpur, Bangladesh, with 1 infant aged 6-8 months were enrolled and followed with weekly illness survey and stool testing for Cryptosporidium for 8 months. RESULTS: From December 2020 to August 2021, 100 families were enrolled. Forty-four percent of index children and 35% of siblings had at least 1 Cryptosporidium infection. Shedding of Cryptosporidium occurred for a mean (standard deviation) of 19 (8.3) days in index infants, 16.1 (11.6) days in children 1-5 years, and 16.2 (12.8) days in adults. A longer duration of Cryptosporidium shedding was associated with growth faltering in infants. There was a spike in Cryptosporidium cases in May 2021, which coincided with a spike in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) cases in the region. CONCLUSIONS: In this intensive, longitudinal study of Cryptosporidium infection in families we found high rates of cryptosporidiosis in infants and children, and prolonged parasite shedding, especially among malnourished children. These data support that transmission within the household is an important route of exposure for young infants and that treatment of nondiarrheal infection to interrupt person-to-person transmission within the home may be essential for preventing cryptosporidiosis in infants.

Prospective cohort study of Cryptosporidium infection and shedding in infants and their households.

Background: Cryptosporidium spp are responsible for significant diarrheal morbidity and mortality in under-five children. There is no vaccine, thus a focus on prevention is paramount. Prior studies suggest that person-to-person spread may be an important pathway for transmission to young children. Here we describe a longitudinal cohort study of 100 families with infants to determine rates of cryptosporidiosis within households during the COVID-19 pandemic. Methods: Families living in Mirpur, Bangladesh with one infant age 6-8 months were enrolled and followed with weekly illness survey and stool testing for Cryptosporidium for 8 months. Results: From December 2020 to August 2021, 100 families were enrolled. Forty-four percent of index children, and 35% of siblings had at least one Cryptosporidium infection. Shedding of Cryptosporidium occurred for a mean of 19 days (sd 8.3 days) in index infants, 16.1 days (sd 11.6) in children 1-5 years, and 16.2 days (sd 12.8) in adults. A longer duration of Cryptosporidium shedding was associated with growth faltering in infants. There was a spike in Cryptosporidium cases in May 2021, which coincided with a spike in SARS-CoV-2 cases in the region. Conclusion: In this intensive, longitudinal study of Cryptosporidium infection in families we found high rates of cryptosporidiosis in infants and children, and prolonged parasite shedding, especially among malnourished children. These data support that transmission within the household is an important route of exposure for young infants, and that treatment of non-diarrheal infection to interrupt person-to-person transmission within the home may be essential for preventing cryptosporidiosis in infants. summary: Cryptosporidiosis is a leading cause of morbidity and mortality among children. We followed 100 families with infants living in Bangladesh and studied the incidence of Cryptosporidium infection. We found prolonged Cryptosporidium shedding in stool was common among infants and adults.

Small Intestine Bacterial Overgrowth in Bangladeshi Infants Is Associated With Growth Stunting in a Longitudinal Cohort.

INTRODUCTION: Small intestine bacterial overgrowth (SIBO) is common in children from low-income countries and has been cross-sectionally associated with growth stunting. We sought to determine whether SIBO was associated with poor growth and neurodevelopmental in a longitudinal analysis. METHODS: We measured SIBO by glucose hydrogen breath test (GHBT) at 18, 52, 78, and 104 weeks of life in a prospective longitudinal birth cohort of Bangladeshi children. Sociodemographic information and measures of enteric inflammation were analyzed as covariates. Diarrheal samples were tested for enteropathogens using polymerase chain reaction. Regression models were created using standardized mean GHBT area under the H2 curve (AUC) to determine associations with linear growth and cognitive, language, and motor scores on the Bayley-III Scales of Infant and Toddler Development at 2 years. We also investigated associations between GHBT AUC and enteropathogen exposure. RESULTS: A 1-ppm increase in standardized mean GHBT AUC was associated with a 0.01-SD decrease in length-for-age Z score (P = 0.03) and a 0.11-point decrease in Bayley language score (P = 0.05) at 2 years of age in adjusted analysis. Enteroaggregative Escherichia coli, Enteropathogenic Escherichia coli, Giardia, and Enterocytozoon bieneusi were associated with increased GHBT AUC, whereas Clostridium difficile, norovirus GI, sapovirus, rotavirus, and Cryptosporidium were associated with decreased GHBT AUC. None were consistent across all 4 time points. DISCUSSION: SIBO in the first 2 years of life is associated with growth stunting and decreased language ability in Bangladeshi infants and may represent a modifiable risk factor in poor growth and neurodevelopment in low-income countries.

MicroRNA Expression and Intestinal Permeability in Children Living in a Slum Area of Bangladesh.

Introduction: MicroRNAs (miRNAs) are small, non-coding RNAs that post-transcriptionally regulate gene expression. Changes in miRNA expression have been reported in a number of intestinal diseases, in both tissue samples and readily accessible specimens like stools. Pathogenic infections, diet, toxins, and other environmental factors are believed to influence miRNA expression. However, modulation of miRNAs in humans is yet to be thoroughly investigated. In this study, we examined the expression levels of two human miRNAs (miRNA-122 and miRNA-21) in stool samples of a group of Bangladeshi children who had an altered/increased intestinal permeability (IIP). Methods: Stool samples were collected from children with IIP (L:M > 0.09) and normal intestinal permeability (NIP) (L:M ≤ 0.09). Quantitative PCR was performed to quantify the levels of miRNA-122 and miR-21 in stools. Commercial ELISA kits were used to measure gut inflammatory markers Calprotectin and REG1B. Serum samples were tested using Human Bio-Plex Pro Assays to quantify IL-1ß, IL-2, IL-5, IL-10, IL-13, IFN-γ, and TNF-α. Total nucleic acid extracted from stool specimens were used to determine gut pathogens using TaqMan Array Card (TAC) system real-time polymerase chain reaction. Results: The expression levels of miRNA-122 (fold change 11.6; p < 0.001, 95% CI: 6.14-11.01) and miR-21 (fold change 10; p < 0.001, 95% CI: 5.05-10.78) in stool were upregulated in children with IIP than in children with normal intestinal permeability (NIP). Significant correlations were observed between stool levels of miR-122 and miR-21 and the inflammatory cytokines IL-1ß, IL-2, IFN-γ, and TNF-α (p < 0.05). Children with IIP were frequently infected with rotavirus, Campylobacter jejuni, Bacteroides fragilis, adenovirus, norovirus, astrovirus, and various Escherichia coli strains (ETEC_STh, ETEC_STp, EAEC_aaiC, EAEC_aatA) (p < 0.001). miR-122 significantly correlated with the fecal inflammatory biomarkers REG1B (p = 0.015) and Calprotectin (p = 0.030), however miR-21 did not show any correlation with these fecal biomarkers.

Nonsterile immunity to cryptosporidiosis in infants is associated with mucosal IgA against the sporozoite and protection from malnutrition.

We conducted a longitudinal study of cryptosporidiosis from birth to three years of age in an urban slum of Dhaka Bangladesh. Fecal DNA was extracted from monthly surveillance samples and diarrheal stool samples collected from 392 infants from birth to three years. A pan-Cryptosporidium qPCR assay was used to identify sub-clinical and symptomatic cryptosporidiosis. Anthropometric measurements were collected quarterly to assess child nutritional status. 31% (121/392) of children experienced a single and 57% (222/392) multiple infections with Cryptosporidium. Repeat infections had a lower burden of parasites in the stool (Cq slope = -1.85; p<0.0001) and were more likely to be sub-clinical (Chi square test for trend; p = 0.01). Repeat infections were associated with the development of growth faltering (Pearson correlation = -0.18; p = 0.0004). High levels of fecal IgA antibodies against the Cryptosporidium Cp23 sporozoite protein at one year of life were associated with a delay in reinfection and amelioration of growth faltering through three years of life (HAZ IgA high responders -1.323 ± 0.932 versus HAZ -1.731 ± 0.984 p = 0.0001). We concluded that nonsterile immunity to cryptosporidiosis in young children was associated with high levels of mucosal IgA anti-Cp23 and protection from diarrhea and growth faltering. Trial Registration: NCT02764918.

Megasphaera in the Stool Microbiota Is Negatively Associated With Diarrheal Cryptosporidiosis.

BACKGROUND: The protozoan parasites in the Cryptosporidium genus cause both acute diarrheal disease and subclinical (ie, nondiarrheal) disease. It is unclear if the microbiota can influence the manifestation of diarrhea during a Cryptosporidium infection. METHODS: To characterize the role of the gut microbiota in diarrheal cryptosporidiosis, the microbiome composition of both diarrheal and surveillance Cryptosporidium-positive fecal samples from 72 infants was evaluated using 16S ribosomal RNA gene sequencing. Additionally, the microbiome composition prior to infection was examined to test whether a preexisting microbiome profile could influence the Cryptosporidium infection phenotype. RESULTS: Fecal microbiome composition was associated with diarrheal symptoms at 2 timepoints. Megasphaera was significantly less abundant in diarrheal samples compared with subclinical samples at the time of Cryptosporidium detection (log2 [fold change] = -4.3; P = 10-10) and prior to infection (log2 [fold change] = -2.0; P = 10-4); this assigned sequence variant was detected in 8 children who had diarrhea and 30 children without diarrhea. Random forest classification also identified Megasphaera abundance in the pre- and postexposure microbiota as predictive of a subclinical infection. CONCLUSIONS: Microbiome composition broadly, and specifically low Megasphaera abundance, was associated with diarrheal symptoms prior to and at the time of Cryptosporidium detection. This observation suggests that the gut microenvironment may play a role in determining the severity of a Cryptosporidium infection. Clinical Trials Registration. NCT02764918.

Diarrheal Pathogens Associated With Growth and Neurodevelopment.

BACKGROUND: Diarrheal pathogens have been associated with linear growth deficits. The effect of diarrheal pathogens on growth is likely due to inflammation, which also adversely affects neurodevelopment. We hypothesized that diarrheagenic pathogens would be negatively associated with both growth and neurodevelopment. METHODS: We conducted a longitudinal birth cohort study of 250 children with diarrheal surveillance and measured pathogen burden in diarrheal samples using quantitative polymerase chain reaction. Pathogen attributable fraction estimates of diarrhea over the first 2 years of life, corrected for socioeconomic variables, were used to predict both growth and scores on the Bayley-III Scales of Infant and Toddler Development. RESULTS: One hundred eighty children were analyzed for growth and 162 for neurodevelopmental outcomes. Rotavirus, Campylobacter, and Shigella were the leading causes of diarrhea in year 1 while Shigella, Campylobacter, and heat-stable toxin-producing enterotoxigenic Escherichia coli were the leading causes in year 2. Norovirus was the only pathogen associated with length-for-age z score at 24 months and was positively associated (regression coefficient [RC], 0.42 [95% confidence interval {CI}, .04 to .80]). Norovirus (RC, 2.46 [95% CI, .05 to 4.87]) was also positively associated with cognitive scores while sapovirus (RC, -2.64 [95% CI, -4.80 to -.48]) and typical enteropathogenic E. coli (RC, -4.14 [95% CI, -8.02 to -.27]) were inversely associated. No pathogens were associated with language or motor scores. Significant maternal, socioeconomic, and perinatal predictors were identified for both growth and neurodevelopment. CONCLUSIONS: Maternal, prenatal, and socioeconomic factors were common predictors of growth and neurodevelopment. Only a limited number of diarrheal pathogens were associated with these outcomes.

Clinical Outcomes of Drug-resistant Shigellosis Treated With Azithromycin in Bangladesh.

BACKGROUND: Azithromycin is frequently used to treat shigellosis; however, clinical outcomes are uncertain. METHODS: We performed an observational cohort study in Bangladesh of patients with invasive diarrhea treated empirically with azithromycin. Susceptibility testing was performed by broth microdilution and disk diffusion post hoc on all Shigella isolates and clinical response was correlated with in vitro susceptibility. RESULTS: There were 149 Shigella culture-positive patients in the primary analysis. Infection with Shigella with decreased susceptibility to azithromycin was significantly associated with persistence of diarrhea at day 5 (31% vs 12%; relative risk [RR], 2.66; 95% confidence interval [CI], 1.34-5.28), culture positivity at day 5 or 6 (35% vs 5%; RR, 5.26; 95% CI, 1.84-14.85), and a higher rate of overnight hospitalization (58% vs 39%; RR, 1.49; 95% CI, 1.06-2.09). Shigella flexneri was more common than Shigella sonnei (58% vs 36%); however, S. sonnei constituted most of the isolates with decreased susceptibility to azithromycin (67%) and most of the multidrug-resistant strains (54%); thus, poor clinical outcomes were associated with S. sonnei. The current epidemiological cutoff for S. flexneri of ≥16 µg/mL to define decreased susceptibility to azithromycin was clinically predictive of poor outcome. Patients with S. sonnei and a low MIC (4 µg/mL) still had elevated rates of persistent diarrhea and culture positivity. CONCLUSIONS: This study documents worse clinical outcomes for S. flexneri with decreased susceptibility to azithromycin, as well as S. sonnei, and supports the utility of susceptibility testing and clinical breakpoints for azithromycin. S. sonnei is an emerging drug-resistant threat. CLINICAL TRIALS REGISTRATION: NCT03778125.

Delayed Time to Cryptosporidiosis in Bangladeshi Children is Associated with Greater Fecal IgA against Two Sporozoite-Expressed Antigens.

Cryptosporidiosis is common in early childhood, and both diarrheal and subclinical infections are associated with adverse developmental outcomes. Improved therapeutic medications may help reduce the burden of cryptosporidial diarrhea; however, an effective vaccine would be better able to prevent the detrimental impact of both diarrheal and subclinical disease. A more complete understanding of naturally occurring immunity may further inform strategies to develop an effective vaccine. In this prospective cohort study of Bangladeshi children, greater fecal IgA at 12 months, but not plasma IgG, directed against two sporozoite-expressed, immunodominant and vaccine candidate antigens was associated with delayed time to subsequent cryptosporidiosis to 3 years of life. These findings extend prior work and further support the role of mucosal antibody responses in naturally developing protective immunity to Cryptosporidium.

Pathogen flows from on-site sanitation systems in low-income urban neighborhoods, Dhaka: A quantitative environmental assessment.

BACKGROUND: Despite wide usage of on-site sanitation, there is limited field-based evidence on the removal or release of pathogens from septic tanks and other primary treatment systems, such as anaerobic baffled reactors (ABR). In two low-income areas in Dhaka, we conducted a cross-sectional study to explore pathogen loads discharged from commonly used on-site sanitation-systems and their transport in nearby drains and waterways. METHODS: We collected samples of drain water, drain sediment, canal water, and floodwater from April-October 2019. Sludge, supernatant, and effluent samples were also collected from septic tanks and ABRs. We investigated the presence and concentration of selected enteric pathogens (Shigella, Vibrio cholerae (V. cholerae), Salmonella Typhi (S. Typhi), Norovirus Genogroup-II (NoV-GII), and Giardia) and presence of Cryptosporidium in these samples using quantitative polymerase chain reaction (qPCR).The equivalent genome copies (EGC) of individual pathogens were estimated in each sample by interpolation of the mean Ct value to the corresponding standard curve and the dilution factor for each sample type. Absolute quantification was expressed as log10 EGC per 100 mL for the water samples and log10 EGC per gram for the sediment samples. RESULTS: Among all samples tested (N = 151), 89% were contaminated with Shigella, 68% with V. cholerae and NoV-GII, 32% with Giardia, 17% with S. Typhi and 6% with Cryptosporidium. A wide range of concentration of pathogens [range: mean log10 concentration of Giardia = 0.74 EGC/100 mL in drain ultrafiltration samples to mean log10 concentration of NoV-GII and Giardia = 7.11 EGC/100 mL in ABR sludge] was found in all environmental samples. The highest pathogen concentrations were detected in open drains [range: mean log10 concentration = 2.50-4.94 EGC/100 mL], septic tank effluent [range: mean log10 concentration = 3.32-4.65 EGC/100 mL], and ABR effluent [range: mean log10 concentration = 2.72-5.13 EGC/100 mL]. CONCLUSIONS: High concentrations of pathogens (particularly NoV-GII, V.cholerae and Shigella) were frequently detected in environmental samples from two low-income urban neighbourhoods of Dhaka city. The numerous environmental exposure pathways for children and adults make these findings of public health concern. These results should prompt rethinking of how to achieve safe sanitation solutions that protect public health in dense low-income areas. In particular, improved management and maintenance regimes, further treatment of liquid effluent from primary treatment processes, and appropriate application of onsite, decentralised and offsite sanitation systems given the local context.

Fecal MicroRNAs as Potential Biomarkers for Screening and Diagnosis of Intestinal Diseases.

MicroRNAs (miRNAs) are a class of conserved endogenous, small non-coding RNA molecules with a length of 18-25 nucleotides that regulate gene expression by RNA interference processes, including mRNA chopping, mRNA deadenylation, and translation inhibition. miRNAs maintain the physiological functions of the intestine and are instrumental in gut pathogenesis. miRNAs play an important role in intercellular communication and are present in all body fluids, including stools with different composition and concentrations. However, under diseased conditions, miRNAs are aberrantly expressed and act as negative regulators of gene expression. The stable and differentially expressed miRNAs in stool enables miRNAs to be used as potential biomarkers for screening of various intestinal diseases. In this review, we summarize the expressed miRNA profile in stool and highlight miRNAs as biomarkers with potential clinical and diagnostic applications, and we aim to address the prospects for recent advanced techniques for screening miRNA in diagnosis and prognosis of intestinal disorders.

Fecal Immunoglobulin A Against a Sporozoite Antigen at 12 Months Is Associated With Delayed Time to Subsequent Cryptosporidiosis in Urban Bangladesh: A Prospective Cohort Study.

In this prospective cohort study of Bangladeshi children, greater fecal immunoglobulin A, but not plasma immunoglobulin G, directed against the Cryptosporidium sporozoite-expressed antigen Cp23 at 12 months of age was associated with delayed time to subsequent cryptosporidiosis. This finding suggests a protective role for mucosal antibody-mediated immunity in naturally exposed children.