Gut Microbes, Enteropathy and Child Growth: The Role of the Microbiota in the Cycle of Diarrhea and Undernutrition in Peru

Background. The cycle between diarrhea and undernutrition continues to underlie a vast proportion of under-five mortality and is the primary driver of long-term disability among children living in lower and middle-income countries (LMIC). Interventions aimed at reducing childhood stunting have not achieved desired results, highlighting the need for novel research and strategies to target this problem. There is increasing evidence that the gut microbiota are implicated in growth acquisition and sustaining intestinal barrier integrity in a manner that impacts immunity to and consequences of disease; however, these relationships have not yet been examined in large-scale studies of children living in LMIC. Objective. To evaluate relationships between the gut microbial community, child growth, diarrhea and enteric infections (Campylobacter spp) in a birth cohort of 271 children aged 0-24 months in Iquitos, Peru. Methods. Analyses were conducted on children participating in the multi-site cohort study entitled ‘The Interactions of Malnutrition & Enteric Infections: Consequences for Child Health and Development (MAL-ED).’ Data were contributed over two years by mothers and children living in a peri-urban riverine community in Amazonian Peru. Regular home-visits were conducted to ascertain anthropometric indices, illness history, and dietary habits. Length-for-age (LAZ) and weight-for-length (WLZ) Z-scores below - 2 were used to classify stunting and wasting, respectively. Fecal specimens were collected during routine surveillance visits at monthly intervals (N=6004) and additionally during each maternal report of diarrheal symptoms (N=2436). Culture methods, immunoassays and amplification methods were employed according to a unified MAL-ED protocol to identify a panel of over 40 protozoa, bacteria and viruses of public health importance. Microbiota in fecal samples contributed at 6, 12, 18 and 24 months were analyzed by polymerase chain reactions using primers to identify variable regions of bacterial 16S ribosomal RNA genes at the Gordon Laboratory at Washington University. Members were binned into operational taxonomic units (OTU) sharing ≥97% nucleotide sequence identity, producing bacterial communities differentiated at the species level which were then used to generate metrics of maturity (microbiota-for-age Z score; MAZ), diversity (Shannon, Simpson indices) and richness (CHAO1, Faith’s Phylogenetic Diversity). Multivariable regression was used to detect and describe population-averaged associations between microbial metrics, growth acquisition, illness and infection with a generalized estimating equations approach to adjust for within-child correlations over time. Indicator species analysis (ISA) was employed to identify particular gut taxa whose presence and abundance was statistically indicative of phenotypes of interest. Results. Two-thirds of children (67%) were stunted and 9% of children experienced wasting before their 2nd birthday. Microbial diversity and richness increased significantly with age and weaning, and were suppressed by breastmilk exposure. In the first two years of life, we detected a suggestive relationship between microbial maturity and WLZ, but did not observe evidence of associations between microbial maturity, diversity or richness with LAZ in the full cohort. LAZ at birth was significantly associated with MAZ score throughout follow-up (β=0.10, p=0.012) and children born stunted had significantly lower gut microbial diversity and richness (Shannonβ=-0.19, CHAO1 =- 9.75; p-values <0.05) from birth to two years of age. In this subgroup, we additionally observed that children weaned before 24m of age experienced significantly pronounced deficits in microbial diversity and richness acquisition relative to those with continued breastfeeding. Nearly all children (96%) experienced diarrhea during follow-up. Odds of being severely stunted increased by 8% with each additional diarrheal episode throughout the first two years of life (OR=1.08; p<0.001). Cumulative diarrheal frequency, duration and severity were associated with significant reductions in microbial indices (p<0.05), and we observed evidence of enduring deficits beyond 1m after exposure. Children who were born stunted experienced greater insults to microbial diversity per diarrheal episode than those children who were not (Interaction terms: Shannon β =-0.04, p=0.037; Simpson β =-0.01, p=0.032). Time elapsed since last diarrheal episode was associated with recovery of Shannon (β =0.02, p=0.03) and phylogenetic diversity (β =0.11, p<0.01) and we detected evidence that this regeneration process was significantly slower among severely stunted children. Lower diversity and richness were associated with increased subsequent diarrheal incidence; a 1-unit increase in the Shannon and Simpson’s Diversity scales at 6m corresponding to a mean reduction of 1.3 and 3.4 diarrheal episodes from 6-24m of age, respectively. By two years of age, 251 (93%) of all children in the cohort had Campylobacter present in asymptomatic stools, and 221 (82%) experienced infection with diarrhea. Asymptomatic infection was associated with reduced LAZ concurrently and at 3, 6, and 9m thereafter (β=0.02, p<0.01 across all time points). Frequency of Campylobacter- positive diarrhea was associated with a concurrent reduction in -0.03 LAZ (p=0.002), independently from all-cause diarrhea. Asymptomatic Campylobacter infections were associated with changes to the gut microbial environment. Infection was associated with increased microbial diversity and richness metrics, and we identified 21 taxa indicative of being in the highest or lowest quartile of infection from birth to two years of age. Of these, seven indicator species showed suggestive evidence of a link with LAZ concurrently and 1m thereafter. Conclusions. This study provided evidence of associations between the gut microbial community, anthropometric indices, and enteric infections in a population of children experiencing the classical cycle of diarrhea and undernutrition. This is the first study to our knowledge to interrogate these pathways longitudinally in a large, representative sample of infants in LMIC. Our findings generate questions regarding the precise causal mechanisms underlying the observed associations, and should inform subsequent efforts to identify specific and actionable targets to interrupt pathways compounding childhood morbidity and mortality in LMIC

Impact of a malaria intervention package in schools on Plasmodium infection, anaemia and cognitive function in schoolchildren in Mali: a pragmatic cluster-randomised trial.

BACKGROUND: School-aged children are rarely targeted by malaria control programmes, yet the prevalence of Plasmodium infection in primary school children often exceeds that seen in younger children and could affect haemoglobin concentration and school performance. METHODS: A cluster-randomised trial was carried out in 80 primary schools in southern Mali to evaluate the impact of a school-based malaria intervention package. Intervention schools received two interventions sequentially: (1) teacher-led participatory malaria prevention education, combined with distribution of long-lasting insecticidal nets (LLINs), followed 7 months later at the end of the transmission season by (2) mass delivery of artesunate and sulfadoxine-pyrimethamine administered by teachers, termed intermittent parasite clearance in schools (IPCs). Control schools received LLINs as part of the national universal net distribution programme. The impact of the interventions on malaria and anaemia was evaluated over 20 months using cross-sectional surveys in a random subset of 38 schools(all classes), with a range of cognitive measures (sustained attention, visual search, numeracy, vocabulary and writing) assessed in a longitudinal cohort of children aged 9-12 years in all 80 schools. RESULTS: Delivery of a single round of IPCs was associated with dramatic reductions in malaria parasitaemia (OR 0.005, 95% CI 0.002 to 0.011, p<0.001) and gametocyte carriage (OR 0.02, 95% CI 0.00 to 0.17, p<0.001) in intervention compared with control schools. This effect was sustained for 6 months until the beginning of the next transmission season. IPCs was also associated with a significant decrease in anaemia (OR 0.56, 95% CI 0.40 to 0.78, p=0.001), and increase in sustained attention (difference +0.23, 95% CI 0.10 to 0.36, p<0.001). There was no evidence of impact on other cognitive measures. CONCLUSION: The combination of malaria prevention education, LLINs and IPCs can reduce anaemia and improve sustained attention of school children in areas of highly seasonal transmission. These findings highlight the impact of asymptomatic malaria infection on cognitive performance in schoolchildren and the benefit of IPCs in reducing this burden. Additionally, malaria control in schools can help diminish the infectious reservoir that sustains Plasmodium transmission

Gut microbiota features associated with Campylobacter burden and postnatal linear growth deficits in a Peruvian birth cohort

BACKGROUND: Campylobacter infection is associated with impaired growth of children, even in the absence of symptoms. To examine the underlying mechanisms, we evaluated associations between Campylobacter infection, linear growth, and fecal microbial community features in a prospective birth cohort of 271 children with a high burden of diarrhea and stunting in the Amazonian lowlands of Peru. METHODS: Campylobacter was identified using a broadly reactive, genus-specific enzyme-linked immunosorbent assay. 16S rRNA-based analyses were used to identify bacterial taxa in fecal samples at ages 6, 12, 18, and 24 months (N = 928). Associations between infection, growth, and gut microbial community composition were investigated using multiple linear regression adjusting for within-child correlations, age, and breastfeeding. Indicator species analyses identified taxa specifically associated with Campylobacter burden. RESULTS: Ninety-three percent (251) of children had Campylobacter present in asymptomatic fecal samples during the follow-up period. A 10% increase in the proportion of stools infected was associated with mean reductions of 0.02 length-for-age z scores (LAZ) at 3, 6, and 9 months thereafter (P \u3c .01). We identified 13 bacterial taxa indicative of cumulative Campylobacter burden and 14 taxa significantly associated with high or low burden of enteroaggregative Escherichia coli, norovirus, or Giardia. CONCLUSIONS: Campylobacter infection is common in this cohort and associated with changes in microbial community composition. These results support the notion that disruptions to the fecal microbiota may help explain the observed effects of asymptomatic infections on growth in early life

Diarrhea as a potential cause and consequence of reduced gut microbial diversity among undernourished children in Peru

BACKGROUND: Detrimental effects of diarrhea on child growth and survival are well documented, but details of the underlying mechanisms remain poorly understood. Recent evidence demonstrates that perturbations to normal development of the gut microbiota in early life may contribute to growth faltering and susceptibility to related childhood diseases. We assessed associations between diarrhea, gut microbiota configuration, and childhood growth in the Peruvian Amazon. METHODS: Growth, diarrhea incidence, illness, pathogen infection, and antibiotic exposure were assessed monthly in a birth cohort of 271 children aged 0-24 months. Gut bacterial diversity and abundances of specific bacterial taxa were quantified by sequencing 16S rRNA genes in fecal samples collected at 6, 12, 18, and 24 months. Linear and generalized linear models were used to determine whether diarrhea was associated with altered microbiota and, in turn, if features of the microbiota were associated with the subsequent risk of diarrhea. RESULTS: Diarrheal frequency, duration, and severity were negatively associated with bacterial diversity and richness (P \u3c .05). Children born stunted (length-for-age z-score [LAZ] ≤ -2) who were also severely stunted (LAZ ≤ -3) at the time of sampling exhibited the greatest degree of diarrhea-associated reductions in bacterial diversity and the slowest recovery of bacterial diversity after episodes of diarrhea. Increased bacterial diversity was predictive of reduced subsequent diarrhea from age 6 to 18 months. CONCLUSIONS: Persistent, severe growth faltering may reduce the gut microbiota\u27s resistance and resilience to diarrhea, leading to greater losses of diversity and longer recovery times. This phenotype, in turn, denotes an increased risk of future diarrheal disease and growth faltering

The scope of coverage under the Rotterdam Rules - Unimodal and multimodal aspects

U radu se proučavaju, raščlanjuju i interpretiraju sve relevantne odredbe o polju primjene Roterdamskih pravila. Analizira se njihovo opće polje primjene, govori se o ugovorima, prijevozima te subjektima prijevoznih poslova na koje se Roterdamska pravila primjenjuju. Proučavaju se i specifične odredbe o posebnim isključenjima. Tumači se u kojem opsegu Roterdamska pravila pokrivaju multimodalni prijevoz te se govori o multimodalnosti kao obilježju suvremenog prijevoza i unifikaciji multimodalnog prijevoza. Pomno se ulazi u sadržaj i doseg odredbi kojima se propisuju uvjeti koji moraju biti ispunjeni da bi odredbe druge međunarodne prijevozne konvencije imale prednost u odnosu na odredbe Roterdamskih pravila. Nastoji se ukazati i na razlike u pogledu njezinog polja primjene i polja primjene međunarodnih konvencija koje su na snazi, a kojima se uređuje prijevoz stvari morem. Odredbe o polju primjene iznimno su važne i zadiru u bit nove konvencije, njezinu koncepciju, temeljna načela i ciljeve novog pravnog uređenja koje Konvencija propisuje. Najvažnije novine polja primjene Roterdamskih pravila u odnosu na međunarodne konvencije koje su na snazi, a koje uređuju prijevoz stvari morem jesu proširenje polja primjene na prijevoze “od vrata do vrata” i novo uređenje prijevoznih isprava. Cilj rada je skrenuti pažnju na polje primjene kao važno pitanje koje utječe na recepciju Roterdamskih pravila i istaknuti ona rješenja koja se najviše kritiziraju i za koja se osnovano može pretpostaviti da predstavljaju zapreku za njihovu recepciju.This paper considers the application of provisions of the United Nations Convention on Contracts for the International Carriage of Goods Wholly or Partly by Sea (the Rotterdam Rules). The scope of coverage issues go to the heart of the new Convention. This article highlights the major topics relating to the scope of coverage. All aspects of this key issue are examined here: the general scope of application; criteria that must be satisfied for the Convention to be applied; types of transactions that the Convention might govern; application to certain parties. Some of these questions have been particularly controversial. Moreover, this paper addresses the circumstances under which the Rotterdam Rules give way to other conventions. The author discusses issues that may arise from the fact that the Convention will cover both the inward and the outward carriage. It further considers specific exclusions from the Convention. Differences between the Rotterdam Rules and the international legislation in force are pointed out. One of the most significant changes introduced by the Rotterdam Rules to the existing law is the expansion of its scope of coverage to include the door-to-door transport. The Convention covers both the inbound and the outbound international shipments to or from a Contracting State. The Convention applies to contracts in both the liner and the non-liner trades, but not to charterparties and other contracts for the use of a ship or of any space thereon. Volume contract, to which the Convention applies, may provide for greater or lesser obligations and liabilities than those imposed by the Rotterdam Rules

Sur l'origine des inhomogénéités de taille et de concentration des boucles de dislocation créées par irradiation électronique dans le CdTe

The chemical reaction rate theory has been developed in order to describe the dynamics of electron irradiation damage in CdTe. The surface effects have been taken into account through the diffusion of interstitials towards the surfaces. This model shows that, due to the surface effects, interstitial, vacancy and dislocation loop concentrations exhibit a gradient normal to the foil. For the same reason, the loop size is distributed around a mean value. The interstitial recombination rate on dislocation loops has been calculated and it has been shown that interstitial diffusion towards the loops is likely to occur. As a consequence, a density gradient is generated around the loop and modifies the growth dynamics.La théorie de la cinétique chimique est reprise et développée en écrivant la diffusion des interstitiels vers les surfaces et leur recombinaison sur celles-ci. Les résultats obtenus par cette modélisation montrent que les effets des surfaces sont à l'origine des gradients de concentration en interstitiels libres, lacunes et boucles de dislocation dans la lame ainsi que de la large distribution en taille de ces dernières. Après calcul de la vitesse de recombinaison des interstitiels sur les boucles de dislocation, il est montré qu'un flux diffusionnel d'interstitiels peut s'établir localement vers les boucles de dislocation et qu'un gradient en concentration de lacunes et interstitiels est ainsi engendré autour des défauts étendus, modifiant sensiblement leur dynamique de croissance

Simulation à l'échelle atomique de la formation des boucles de dislocation sous irradiation

Using the Monte carlo technique, we have developed a model for the Atomic Scale Simulation of the formation of dislocation loops in materials under irradiation. We assume that vacancy interstitial pairs are created by particle impact and diffuse through the solid. Three types of reaction are considered : vacancy interstitial recombination, interstitial association to form a nucleus for a new dislocation loop and incorporation of interstitials into already existing dislocation loops leading to their growth. We have determined the concentration of interstitials, vacancies and dislocation loops, together with the average radius of the latter. Our results are compared with those obtained by using the chemical rate theory and with experimental data on CdTe. Moreover, Atomic Scale Simulations lead to the spatial distribution of dislocation loops, in agreement with TEM experimental observations, and to indications about the distribution of vacancies around these loops. This kind of information is totally missing in the chemical rate theory.En utilisant une technique de Monte Carlo, nous avons développé un modèle de simulation à l'échelle atomique traitant de la formation des boucles de dislocation dans les matériaux sous irradiation. Les paires lacunes-interstitiels sont créées sous l'impact des particules incidentes et diffusent dans le matériau. Trois types de réaction sont supposées avoir lieu : la recombinaison lacune-interstitiel, l'association d'interstitiels pour former des germes de boucles de dislocation et l'incorporation d'interstitiels dans ces boucles conduisant à leur grossissement. Nous avons déterminé les concentrations des interstitiels, des lacunes et des boucles de dislocation, ainsi que la taille de ces dernières. Nous avons comparé ces résultats avec ceux de la théorie de la cinétique chimique et avec nos résultats expérimentaux sur le CdTe. En outre, la simulation à l'échelle atomique fournit les distributions spatiales des boucles de dislocation qui concordent avec les observations expérimentales en microscopie électronique en transmission, et des indications sur la répartition des lacunes autour de ces boucles. Ces dernières informations sont complètement absentes dans la théorie de la cinétique chimique

Disruptions to naloxone training among lay and occupational responders in Maryland during the emergence of COVID-19: Early impacts, recovery, and lessons learned

Background: Opioid overdose death rates increased during the COVID-19 pandemic. Disruptions in community-based naloxone trainings could have reduced the likelihood of overdose reversal and increased the chances of a fatal overdose. We investigated changes in the number of people trained in naloxone administration and distribution in Maryland before, during, and after COVID-related stay-at-home orders. Methods: Data on naloxone training are from the Maryland Department of Health. We used interrupted time series models to estimate changes in average monthly number of people trained: [1] pre-interruption (4/2019–3/2020), [2] 1-month post-interruption (4/2020–5/2020), and [3] 12-month post-interruption (4/2020–3/2021). Trainees were classified as lay (e.g., people who use drugs) or occupational (e.g., law enforcement officers and harm reduction workers) responders. Results: There were 101,332 trainees; 54.1% lay, 21.5% occupational, and 23.4% unknown responder status. We observed a decrease in the average monthly number of trainees in the pre-interruption period (-235, p<0.001), a larger decrease of 93.2% during the 1-month post-interruption (-846, p = 0.013), and an increase during the 12-month post-interruption (+217, p<0.001). There was a significant decrease among occupational responders 1-month post-interruption, and a significant increase among lay responders in the 12-month post-interruption period. Conclusions: Findings suggest a marked decrease in naloxone trainees immediately after stay-at-home order, followed by a moderate rebound in the 12-months after stay-at-home order. The decrease in occupational responders trained may have limited access to naloxone, but would likely have been offset by increases in number of lay responders trained. Strengthening lay and occupational responder connections could maintain naloxone distribution during public health crises