Soil stabilisation for DNA metabarcoding of plants and fungi. Implications for sampling at remote locations or via third-parties

Storage of soil samples prior to metagenomic analysis presents a problem. If field sites are remote or if samples are collected by third parties, transport to analytical laboratories may take several days or even weeks. The bulk of such samples and requirement for later homogenisation precludes the convenient use of a stabilisation buffer, so samples are usually cooled or frozen during transit. There has been limited testing of the most appropriate storage methods for later study of soil organisms by eDNA approaches. Here we tested a range of storage methods on two contrasting soils, comparing these methods to the control of freezing at -80 °C, followed by freeze-drying. To our knowledge, this is the first study to examine the effect of storage conditions on eukaryote DNA in soil, including both viable organisms (fungi) and DNA contained within dying/dead tissues (plants). For fungi, the best storage regimes (closest to the control) were storage at 4 °C (for up to 14 d) or active air-drying at room temperature. The worst treatments involved initial freezing, followed by thawing which led to significant later spoilage. The key spoilage organisms were identified as Metarhizium carneum and Mortierella spp., with a general increase in saprotrophic fungi and reduced abundances of mycorrhizal/biotrophic fungi. Plant data showed a similar pattern, but with greater variability in community structure, especially in the freeze-thaw treatments, probably due to stochastic variation in substrates for fungal decomposition, algal proliferation and some seed germination. In the absence of freeze drying facilities, samples should be shipped refrigerated, but not frozen if there is any risk of thawing

Effect of household-based drinking water chlorination on diarrhoea among children under five in Orissa, India: a double-blind randomised placebo-controlled trial.

BACKGROUND: Boiling, disinfecting, and filtering water within the home can improve the microbiological quality of drinking water among the hundreds of millions of people who rely on unsafe water supplies. However, the impact of these interventions on diarrhoea is unclear. Most studies using open trial designs have reported a protective effect on diarrhoea while blinded studies of household water treatment in low-income settings have found no such effect. However, none of those studies were powered to detect an impact among children under five and participants were followed-up over short periods of time. The aim of this study was to measure the effect of in-home water disinfection on diarrhoea among children under five. METHODS AND FINDINGS: We conducted a double-blind randomised controlled trial between November 2010 and December 2011. The study included 2,163 households and 2,986 children under five in rural and urban communities of Orissa, India. The intervention consisted of an intensive promotion campaign and free distribution of sodium dichloroisocyanurate (NaDCC) tablets during bi-monthly households visits. An independent evaluation team visited households monthly for one year to collect health data and water samples. The primary outcome was the longitudinal prevalence of diarrhoea (3-day point prevalence) among children aged under five. Weight-for-age was also measured at each visit to assess its potential as a proxy marker for diarrhoea. Adherence was monitored each month through caregiver's reports and the presence of residual free chlorine in the child's drinking water at the time of visit. On 20% of the total household visits, children's drinking water was assayed for thermotolerant coliforms (TTC), an indicator of faecal contamination. The primary analysis was on an intention-to-treat basis. Binomial regression with a log link function and robust standard errors was used to compare prevalence of diarrhoea between arms. We used generalised estimating equations to account for clustering at the household level. The impact of the intervention on weight-for-age z scores (WAZ) was analysed using random effect linear regression. Over the follow-up period, 84,391 child-days of observations were recorded, representing 88% of total possible child-days of observation. The longitudinal prevalence of diarrhoea among intervention children was 1.69% compared to 1.74% among controls. After adjusting for clustering within household, the prevalence ratio of the intervention to control was 0.95 (95% CI 0.79-1.13). The mean WAZ was similar among children of the intervention and control groups (-1.586 versus -1.589, respectively). Among intervention households, 51% reported their child's drinking water to be treated with the tablets at the time of visit, though only 32% of water samples tested positive for residual chlorine. Faecal contamination of drinking water was lower among intervention households than controls (geometric mean TTC count of 50 [95% CI 44-57] per 100 ml compared to 122 [95% CI 107-139] per 100 ml among controls [p<0.001] [n = 4,546]). CONCLUSIONS: Our study was designed to overcome the shortcomings of previous double-blinded trials of household water treatment in low-income settings. The sample size was larger, the follow-up period longer, both urban and rural populations were included, and adherence and water quality were monitored extensively over time. These results provide no evidence that the intervention was protective against diarrhoea. Low compliance and modest reduction in water contamination may have contributed to the lack of effect. However, our findings are consistent with other blinded studies of similar interventions and raise additional questions about the actual health impact of household water treatment under these conditions. TRIAL REGISTRATION: ClinicalTrials.govNCT01202383 Please see later in the article for the Editors' Summary

The time delay of the quadruple quasar RX J0911.4+0551

We present optical lightcurves of the gravitationally lensed components A (=A1+A2+A3) and B of the quadruple quasar RX J0911.4+0551 (z = 2.80). The observations were primarily obtained at the Nordic Optical Telescope between 1997 March and 2001 April and consist of 74 I-band data points for each component. The data allow the measurement of a time delay of 146 +- 8 days (2 sigma) between A and B, with B as the leading component. This value is significantly shorter than that predicted from simple models and indicates a very large external shear. Mass models including the main lens galaxy and the surrounding massive cluster of galaxies at z = 0.77, responsible for the external shear, yield H_0 = 71 +- 4 (random, 2 sigma) +- 8 (systematic) km/s/Mpc. The systematic model uncertainty is governed by the surface-mass density (convergence) at the location of the multiple images.Comment: 12 pages, 3 figures, ApJL, in press (June 20, 2002

Assessing Latrine Use in Rural India: A Cross-Sectional Study Comparing Reported Use and Passive Latrine Use Monitors.

Although large-scale programs, like India's Total Sanitation Campaign (TSC), have improved latrine coverage in rural settings, evidence suggests that actual use is suboptimal. However, the reliability of methods to assess latrine use is uncertain. We assessed the reliability of reported use, the standard method, by comparing survey-based responses against passive latrine use monitors (PLUMs) through a cross-sectional study among 292 households in 25 villages in rural Odisha, India, which recently received individual household latrines under the TSC. PLUMs were installed for 2 weeks and householders responded to surveys about their latrine use behavior. Reported use was compared with PLUM results using Bland-Altman (BA) plots and concordance statistics. Reported use was higher than corresponding PLUM-recorded events across the range of comparisons. The mean reported "usual" daily events per household (7.09, 95% confidence interval [CI] = 6.51, 7.68) was nearly twice that of the PLUM-recorded daily average (3.62, 95% CI = 3.29, 3.94). There was poor agreement between "usual" daily latrine use and the average daily PLUM-recorded events (ρc = 0.331, 95% CI = 0.242, 0.427). Moderate agreement (ρc = 0.598, 95% CI = 0.497, 0.683) was obtained when comparing daily reported use during the previous 48 hours with the average daily PLUM count. Reported latrine use, though already suggesting suboptimal adoption, likely exaggerates the actual level of uptake of latrines constructed under the program. Where reliance on self-reports is used, survey questions should focus on the 48 hours prior to the date of the survey rather than asking about "usual" latrine use behavior

Assessing patterns and determinants of latrine use in rural settings: A longitudinal study in Odisha, India.

INTRODUCTION: Monitoring of sanitation programs is often limited to sanitation access and coverage, with little emphasis on use of the facilities despite increasing evidence of widespread non-use. OBJECTIVES: We assessed patterns and determinants of individual latrine use over 12 months in a low- income rural study population that had recently received latrines as part of the Government of India's Total Sanitation Campaign (TSC) in coastal Puri district in Odisha, India. MATERIALS AND METHODS: We surveyed 1938 individuals (>3years) in 310 rural households with latrines from 25 villages over 12 months. Data collection rounds were timed to correspond with the seasons. The primary outcome was reported use by each member of the household over the prior 48h. We classified use into three categories-"never", "sometimes" and "always/usually". We also assessed consistency of use over six days across the three seasons (dry cold, dry hot, rainy). We explored the association between individual and household-level variables and latrine use in any given season and longitudinally using multinomial logistic regression. We also inquired about reasons for non-use. RESULTS: Overall, latrine use was poor and inconsistent. The average response probability at any given round of never use was 43.5% (95% CI=37.9, 49.1), sometimes use was 4.6% (95% CI=3.8, 5.5), and always/usual use was 51.9% (95% CI=46.2, 57.5). Only two-thirds of those who reported always/usually using a latrine in round one reported the same for all three rounds. Across all three rounds, the study population was about equally divided among those who reported never using the latrine (30.1%, 95% CI=23.0, 37.2), sometimes using the latrine (33.2%, 95% CI=28.3, 38.1) and always/usually using the latrine (36.8%, 95% CI=31.8, 41.8). The reported likelihood of always/usually versus never using the latrine was significantly greater in the dry cold season (OR=1.50, 95% CI=1.18, 1.89, p=0.001) and in the rainy season (OR=1.34, 95% CI=1.07, 1.69, p=0.012), than in the dry hot season. Across all three seasons, there was increased likelihood of always/usually and sometimes using the latrine versus never using it among females and where latrines had a door and roof. Older age groups, including those aged 41-59 years and 60+ years, and increase in household size were associated with a decreased likelihood of always/usually using the latrine versus never using it. The leading reason for non-use was a preference for open defecation. CONCLUSION: Results highlight the low and inconsistent use of subsidized latrines built under the TSC in rural Odisha. This study identifies individual and household levels factors that may be used to target behavior change campaigns to drive consistent use of sanitation facilities by all

Follow up study to assess the use and performance of household water filters in Zambia

Effective household water treatment has the potential to improve drinking water quality and prevent diarrhoeal disease if used correctly and consistently over time. We previously conducted a randomized, controlled trial of water filters among households in Zambia with children under two years of HIV-positive mothers. One year after completion of the trial, we conducted a follow up study to assess use and performance the household filters. Ninety percent of participating households met the criteria for current users, and 75% had stored water with lower levels of faecal contamination than source water. Although this study provides some encouraging evidence about the potential to maintain HWTS use and performance, further research is necessary to assess whether these results can be achieved over longer periods and with larger populations

Effects of particle stiffness on the extensional rheology of model rod-like nanoparticle suspensions

The linear and nonlinear rheological behavior of two rod-like particle suspensions as a function of concentration is studied using small amplitude oscillatory shear, steady shear and capillary breakup extensional rheometry. The rod-like suspensions are composed of fd virus and its mutant fdY21M, which are perfectly monodisperse, with a length on the order of 900 nm. The particles are semiflexible yet differ in their persistence length. The effect of stiffness on the rheological behavior in both, shear and extensional flow, is investigated experimentally. The linear viscoelastic shear data is compared in detail with theoretical predictions for worm-like chains. The extensional properties are compared to Batchelor\u27s theory, generalized for the shear thinning nature of the suspensions. Theoretical predictions agree well with the measured complex moduli at low concentrations as well as the nonlinear shear and elongational viscosities at high flow rates. The results in this work provide guidelines for enhancing the elongational viscosity based on purely frictional effects in the absence of strong normal forces which are characteristic for high molecular weight polymers