Abstract.Hookworm infection affects 430 million people worldwide, causing iron deficiency, impaired cognitive development, and stunting in children. Because of the environmental conditions needed for the hookworm life-cycle, this parasite is endemic to resource-limited countries. Necator americanus was endemic in the southern United States before improvement of sewage disposal systems and eradication programs. With continued poverty, poor sanitation, and an environment suitable for the hookworm life-cycle in some regions of the southern United States, a current prevalence study using modern molecular diagnostics is warranted. Lowndes County, Alabama, was chosen as the study site given previous high hookworm burdens, degree of poverty, and use of open-sewage systems. Participants were interviewed, and stool, serum, and soil samples were tested for nine intestinal parasites using a multiparallel quantitative real-time PCR (qPCR) and enzyme-linked immunosorbent assays. We found that, among 24 households, 42.4% reported exposure to raw sewage within their home, and from 55 stool samples, 19 (34.5%) tested positive for N. americanus, four (7.3%) for Strongyloides stercoralis, and one (1.8%) for Entamoeba histolytica. Stool tested positive for N. americanus contained low levels of parasite DNA (geometric mean 0.0302 fg/μL). Soil studies detected one (2.9%) Cryptosporidium species, and Toxocara serology assay detected one (5.2%) positive in this population. Individuals living in this high-risk environment within the United States continue to have stool samples positive for N. americanus. Gastrointestinal parasites known to be endemic to developing countries are identifiable in American poverty regions, and areas with lower disease burden are more likely to be identified by using qPCR.

BackgroundIn resource-limited countries, stool microscopy is the diagnostic test of choice for intestinal parasites (soil-transmitted helminths and/or intestinal protozoa). However, sensitivity and specificity is low. Improved diagnosis of intestinal parasites is especially important for accurate measurements of prevalence and intensity of infections in endemic areas.MethodsThe study was carried out in Orán, Argentina. A total of 99 stool samples from a local surveillance campaign were analyzed by concentration microscopy and McMaster egg counting technique compared to the analysis by multi-parallel quantitative real-time polymerase chain reaction (qPCR). This study compared the performance of qPCR assay and stool microscopy for 8 common intestinal parasites that infect humans including the helminths Ascaris lumbricoides, Ancylostoma duodenale, Necator americanus, Strongyloides stercoralis, Trichuris trichiura, and the protozoa Giardia lamblia, Cryptosporidium parvum/hominis, and Entamoeba histolytica, and investigated the prevalence of polyparasitism in an endemic area.ResultsqPCR showed higher detection rates for all parasites as compared to stool microscopy except T. trichiura. Species-specific primers and probes were able to distinguish between A. duodenale (19.1 %) and N. americanus (36.4 %) infections. There were 48.6 % of subjects co-infected with both hookworms, and a significant increase in hookworm DNA for A. duodenale versus N. americanus (119.6 fg/μL: 0.63 fg/μL, P < 0.001) respectively. qPCR outperformed microscopy by the largest margin in G. lamblia infections (63.6 % versus 8.1 %, P < 0.05). Polyparasitism was detected more often by qPCR compared to microscopy (64.7 % versus 24.2 %, P < 0.05).ConclusionsMulti-parallel qPCR is a quantitative molecular diagnostic method for common intestinal parasites in an endemic area that has improved diagnostic accuracy compared to stool microscopy. This first time use of multi-parallel qPCR in Argentina has demonstrated the high prevalence of intestinal parasites in a peri-urban area. These results will contribute to more accurate epidemiological survey, refined treatment strategies on a public scale, and better health outcomes in endemic settings.

Background: Approximately 30% of children worldwide are infected with gastrointestinal parasites. Depending on the species, parasites can disrupt intestinal bacterial microbiota affecting essential vitamin biosynthesis.Methods: Stool samples were collected from 37 asymptomatic children from a previous cross-sectional Argentinian study. A multi-parallel real-time quantitative PCR was implemented for Ascaris lumbricoides, Ancylostoma duodenale, Necator americanus, Strongyloides stercoralis, Trichuris trichiura, Cryptosporidium spp., Entamoeba histolytica and Giardia duodenalis. In addition, whole-genome sequencing analysis was conducted for bacterial microbiota on all samples and analyzed using Livermore Metagenomic Analysis Toolkit and DIAMOND software. Separate analyses were carried out for uninfected, Giardia-only, Giardia + helminth co-infections, and helminth-only groups. Results:For Giardia-only infected children compared to uninfected children, DNA sequencing data showed a decrease in microbiota biodiversity that correlated with increasing Giardia burden and was statistically significant using Shannonʼs alpha diversity (Giardia-only > 1 fg/µl 2.346; non-infected group 3.253, P = 0.0317). An increase in diversity was observed for helminth-only infections with a decrease in diversity for Giardia + helminth co-infections (P = 0.00178). In Giardia-only infections, microbiome taxonomy changed from Firmicutes towards increasing proportions of Prevotella, with the degree of change related to the intensity of infection compared to uninfected (P = 0.0317). The abundance of Prevotella bacteria was decreased in the helminths-only group but increased for Giardia + helminth co-infections (P = 0.0262). Metagenomic analysis determined cobalamin synthesis was decreased in the Giardia > 1 fg/µl group compared to both the Giardia < 1 fg/µl and the uninfected group (P = 0.0369). Giardia + helminth group also had a decrease in cobalamin CbiM genes from helminth-only infections (P = 0.000754). Conclusion:The study results may provide evidence for an effect of parasitic infections enabling the permissive growth of anaerobic bacteria such as Prevotella, suggesting an altered capacity of vitamin B12 (cobalamin) biosynthesis and potential impact on growth and development in children .

Among 234 US youths with perinatal human immunodeficiency virus, 75% had antiretroviral resistance, substantially higher than that of the reference laboratory overall (36%-44%). Resistance to newer antiretrovirals and to all antiretrovirals in a class was uncommon. The only factor independently associated with future resistance was a higher peak viral load.

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