Multi-parallel qPCR provides increased sensitivity and diagnostic breadth for gastrointestinal parasites of humans: field-based inferences on the impact of mass deworming

BACKGROUND: Although chronic morbidity in humans from soil transmitted helminth (STH) infections can be reduced by anthelmintic treatment, inconsistent diagnostic tools make it difficult to reliably measure the impact of deworming programs and often miss light helminth infections. METHODS: Cryopreserved stool samples from 796 people (aged 2-81 years) in four villages in Bungoma County, western Kenya, were assessed using multi-parallel qPCR for 8 parasites and compared to point-of-contact assessments of the same stools by the 2-stool 2-slide Kato-Katz (KK) method. All subjects were treated with albendazole and all Ascaris lumbricoides expelled post-treatment were collected. Three months later, samples from 633 of these people were re-assessed by both qPCR and KK, re-treated with albendazole and the expelled worms collected. RESULTS: Baseline prevalence by qPCR (n = 796) was 17 % for A. lumbricoides, 18 % for Necator americanus, 41 % for Giardia lamblia and 15% for Entamoeba histolytica. The prevalence was <1% for Trichuris trichiura, Ancylostoma duodenale, Strongyloides stercoralis and Cryptosporidium parvum. The sensitivity of qPCR was 98% for A. lumbricoides and N. americanus, whereas KK sensitivity was 70% and 32%, respectively. Furthermore, qPCR detected infections with T. trichiura and S. stercoralis that were missed by KK, and infections with G. lamblia and E. histolytica that cannot be detected by KK. Infection intensities measured by qPCR and by KK were correlated for A. lumbricoides (r = 0.83, p < 0.0001) and N. americanus (r = 0.55, p < 0.0001). The number of A. lumbricoides worms expelled was correlated (p < 0.0001) with both the KK (r = 0.63) and qPCR intensity measurements (r = 0.60). CONCLUSIONS: KK may be an inadequate tool for stool-based surveillance in areas where hookworm or Strongyloides are common or where intensity of helminth infection is low after repeated rounds of chemotherapy. Because deworming programs need to distinguish between populations where parasitic infection is controlled and those where further treatment is required, multi-parallel qPCR (or similar high throughput molecular diagnostics) may provide new and important diagnostic information

Spectrochemical analysis of sycamore (Acer pseudoplatanus) leaves for environmental health monitoring

Terrestrial plants are ideal sentinels of environmental pollution, due to their sedentary nature, abundance and sensitivity to atmospheric changes. However, reliable and sensitive biomarkers of exposure have hitherto been difficult to characterise. Biospectroscopy offers a novel approach to the derivation of biomarkers in the form of discrete molecular alterations detectable within a biochemical fingerprint. We investigated the application of this approach for the identification of biomarkers for pollution exposure using the common sycamore (Acer pseudoplatanus) as a sentinel species. Attenuated total reflection Fourier-transform infrared (ATR-FTIR) spectroscopy was used to interrogate leaf tissue collected from three sites exposed to different levels of vehicle exhaust emissions. Following multivariate analysis of acquired spectra, significant biochemical alterations were detected between comparable leaves from different sites that may constitute putative biomarkers for pollution-induced stress. These included differences in carbohydrate and nucleic acid conformations, which may be indicative of sub-lethal exposure effects. We also observed several corresponding spectral alterations in both the leaves of A. pseudoplatanus exposed to ozone pollution under controlled environmental conditions and in leaves infected with the fungal pathogen Rhytisma acerinum, indicating that some stress-induced changes are conserved between different stress signatures. These similarities may be indicative of stress-induced reactive oxygen species (ROS) generation, although further work is needed to verify the precise identity of infrared biomarkers and to identify those that are specific to pollution exposure. Taken together, our data clearly demonstrate that biospectroscopy presents an effective toolkit for the utilisation of higher plants, such as A. pseudoplatanus, as sentinels of environmental pollution

Evaluation of a multi-level intervention to improve postpartum intrauterine device services in Rwanda [version 2; referees: 2 approved, 1 approved with reservations]

Background. The copper intrauterine device is one of the most safe, effective, and cost-effective methods for preventing unintended pregnancy. It can be used postpartum irrespective of breastfeeding to improve birth spacing and reduce unintended pregnancy and maternal-child mortality. However, this method remains highly underutilized. Methods. We developed a multi-level intervention to increase uptake of the postpartum intrauterine device (PPIUD, defined as insertion up to six weeks post-delivery) in Kigali, Rwanda. High-volume hospitals and health centers were selected for implementation of PPIUD counseling and service delivery. Formative work informed development of a PPIUD counseling flipchart to be delivered during antenatal care, labor and delivery, infant vaccination visits, or in the community. Two-day didactic counseling, insertion/removal, and follow-up trainings were provided to labor and delivery and family planning nurses followed by a mentored practicum certification process. Counseling data were collected in government clinic logbooks. Insertions and follow-up data were collected in logbooks created for the implementation. Data were collected by trained government clinic staff and abstracted/managed by study staff. Stakeholders were involved from intervention development through dissemination of results. Results. Two hospitals (and their two associated health centers) and two additional health centers were selected. In 6-months prior to our intervention, 7.7 PPIUDs/month were inserted on average at the selected facilities. From August 2017-July 2018, we trained 83 counselors and 39 providers to provide PPIUD services. N=9,020 women received one-on-one PPIUD counseling after expressing interest in family planning who later delivered at a selected health facility. Of those, n=2,575 had PPIUDs inserted (average of 214.6 insertions/month), a 29% uptake. Most PPIUDs (62%) were inserted within 10 minutes of delivery of the placenta. Conclusions. This successful, comprehensive intervention has the potential to make a significant impact on PPIUD uptake in Rwanda. The intervention is scalable and adaptable to other sub-Saharan African countries

Evaluation of a multi-level intervention to improve postpartum intrauterine device services in Rwanda [version 3; referees: 3 approved]

Background. The copper intrauterine device is one of the most safe, effective, and cost-effective methods for preventing unintended pregnancy. It can be used postpartum irrespective of breastfeeding to improve birth spacing and reduce unintended pregnancy and maternal-child mortality. However, this method remains highly underutilized. Methods. We developed a multi-level intervention to increase uptake of the postpartum intrauterine device (PPIUD, defined as insertion up to six weeks post-delivery) in Kigali, Rwanda. High-volume hospitals and health centers were selected for implementation of PPIUD counseling and service delivery. Formative work informed development of a PPIUD counseling flipchart to be delivered during antenatal care, labor and delivery, infant vaccination visits, or in the community. Two-day didactic counseling, insertion/removal, and follow-up trainings were provided to labor and delivery and family planning nurses followed by a mentored practicum certification process. Counseling data were collected in government clinic logbooks. Insertions and follow-up data were collected in logbooks created for the implementation. Data were collected by trained government clinic staff and abstracted/managed by study staff. Stakeholders were involved from intervention development through dissemination of results. Results. Two hospitals (and their two associated health centers) and two additional health centers were selected. In 6-months prior to our intervention, 7.7 PPIUDs/month were inserted on average at the selected facilities. From August 2017-July 2018, we trained 83 counselors and 39 providers to provide PPIUD services. N=9,020 women received one-on-one PPIUD counseling after expressing interest in family planning who later delivered at a selected health facility. Of those, n=2,575 had PPIUDs inserted (average of 214.6 insertions/month), a 29% uptake. Most PPIUDs (62%) were inserted within 10 minutes of delivery of the placenta. Conclusions. This successful, comprehensive intervention has the potential to make a significant impact on PPIUD uptake in Rwanda. The intervention is scalable and adaptable to other sub-Saharan African countries

Phenotypic Diversity and Altered Environmental Plasticity in Arabidopsis thaliana with Reduced Hsp90 Levels

The molecular chaperone HSP90 aids the maturation of a diverse but select set of metastable protein clients, many of which are key to a variety of signal transduction pathways. HSP90 function has been best investigated in animal and fungal systems, where inhibition of the chaperone has exceptionally diverse effects, ranging from reversing oncogenic transformation to preventing the acquisition of drug resistance. Inhibition of HSP90 in the model plant Arabidopsis thaliana uncovers novel morphologies dependent on normally cryptic genetic variation and increases stochastic variation inherent to developmental processes. The biochemical activity of HSP90 is strictly conserved between animals and plants. However, the substrates and pathways dependent on HSP90 in plants are poorly understood. Progress has been impeded by the necessity of reliance on light-sensitive HSP90 inhibitors due to redundancy in the A. thaliana HSP90 gene family. Here we present phenotypic and genome-wide expression analyses of A. thaliana with constitutively reduced HSP90 levels achieved by RNAi targeting. HSP90 reduction affects a variety of quantitative life-history traits, including flowering time and total seed set, increases morphological diversity, and decreases the developmental stability of repeated characters. Several morphologies are synergistically affected by HSP90 and growth temperature. Genome-wide expression analyses also suggest a central role for HSP90 in the genesis and maintenance of plastic responses. The expression results are substantiated by examination of the response of HSP90-reduced plants to attack by caterpillars of the generalist herbivore Trichoplusia ni. HSP90 reduction potentiates a more robust herbivore defense response. In sum, we propose that HSP90 exerts global effects on the environmental responsiveness of plants to many different stimuli. The comprehensive set of HSP90-reduced lines described here is a vital instrument to further examine the role of HSP90 as a central interface between organism, development, and environment

Predation and the Maintenance of Color Polymorphism in a Habitat Specialist Squamate

Multiple studies have addressed the mechanisms maintaining polymorphism within a population. However, several examples exist where species inhabiting diverse habitats exhibit local population-specific polymorphism. Numerous explanations have been proposed for the maintenance of geographic variation in color patterns. For example, spatial variation in patterns of selection or limited gene flow can cause entire populations to become fixed for a single morph, resulting in separate populations of the same species exhibiting separate and distinct color morphs. The mottled rock rattlesnake (Crotalus lepidus lepidus) is a montane species that exhibits among-population color polymorphism that correlates with substrate color. Habitat substrate in the eastern part of its range is composed primarily of light colored limestone and snakes have light dorsal coloration, whereas in the western region the substrate is primarily dark and snakes exhibit dark dorsal coloration. We hypothesized that predation on high contrast color and blotched patterns maintain these distinct color morphs. To test this we performed a predation experiment in the wild by deploying model snakes at 12 sites evenly distributed within each of the two regions where the different morphs are found. We employed a 2×2 factorial design that included two color and two blotched treatments. Our results showed that models contrasting with substrate coloration suffered significantly more avian attacks relative to models mimicking substrates. Predation attempts on blotched models were similar in each substrate type. These results support the hypothesis that color pattern is maintained by selective predation

Toxic marine microalgae and shellfish poisoning in the British isles: history, review of epidemiology, and future implications

The relationship between toxic marine microalgae species and climate change has become a high profile and well discussed topic in recent years, with research focusing on the possible future impacts of changing hydrological conditions on Harmful Algal Bloom (HAB) species around the world. However, there is very little literature concerning the epidemiology of these species on marine organisms and human health. Here, we examine the current state of toxic microalgae species around the UK, in two ways: first we describe the key toxic syndromes and gather together the disparate reported data on their epidemiology from UK records and monitoring procedures. Secondly, using NHS hospital admissions and GP records from Wales, we attempt to quantify the incidence of shellfish poisoning from an independent source. We show that within the UK, outbreaks of shellfish poisoning are rare but occurring on a yearly basis in different regions and affecting a diverse range of molluscan shellfish and other marine organisms. We also show that the abundance of a species does not necessarily correlate to the rate of toxic events. Based on routine hospital records, the numbers of shellfish poisonings in the UK are very low, but the identification of the toxin involved, or even a confirmation of a poisoning event is extremely difficult to diagnose. An effective shellfish monitoring system, which shuts down aquaculture sites when toxins exceed regularity limits, has clearly prevented serious impact to human health, and remains the only viable means of monitoring the potential threat to human health. However, the closure of these sites has an adverse economic impact, and the monitoring system does not include all toxic plankton. The possible geographic spreading of toxic microalgae species is therefore a concern, as warmer waters in the Atlantic could suit several species with southern biogeographical affinities enabling them to occupy the coastal regions of the UK, but which are not yet monitored or considered to be detrimental

Murid Herpesvirus-4 Exploits Dendritic Cells to Infect B Cells

Dendritic cells (DCs) play a central role in initiating immune responses. Some persistent viruses infect DCs and can disrupt their functions in vitro. However, these viruses remain strongly immunogenic in vivo. Thus what role DC infection plays in the pathogenesis of persistent infections is unclear. Here we show that a persistent, B cell-tropic gamma-herpesvirus, Murid Herpesvirus-4 (MuHV-4), infects DCs early after host entry, before it establishes a substantial infection of B cells. DC-specific virus marking by cre-lox recombination revealed that a significant fraction of the virus latent in B cells had passed through a DC, and a virus attenuated for replication in DCs was impaired in B cell colonization. In vitro MuHV-4 dramatically altered the DC cytoskeleton, suggesting that it manipulates DC migration and shape in order to spread. MuHV-4 therefore uses DCs to colonize B cells