Worldwide absence of canonical benzimidazole resistance-associated mutations within β-tubulin genes from Ascaris

Background: The giant roundworm Ascaris is an intestinal nematode, causing ascariasis by infecting humans and pigs worldwide. Recent estimates suggest that Ascaris infects over half a billion people, with chronic infections leading to reduced growth and cognitive ability. Ascariasis affects innumerable pigs worldwide and is known to reduce production yields via decreased growth and condemnation of livers. The predominant anthelminthic drugs used to treat ascariasis are the benzimidazoles. Benzimidazoles interact with β-tubulins and block their function, and several benzimidazole resistance-associated mutations have been described in the β-tubulins of ruminant nematodes. Recent research on ascarids has shown that these canonical benzimidazole resistance-associated mutations are likely not present in the β-tubulins of Ascaris, Ascaridia or Parascaris, even in phenotypically resistant populations. Methods: To further determine the putative absence of key β-tubulin polymorphisms, we screened two β-tubulin isotypes of Ascaris, highly expressed in adult worms. Using adult and egg samples of Ascaris obtained from pigs and humans worldwide, we performed deep amplicon sequencing to look for canonical resistance-associated mutations in Ascaris β-tubulins. Subsequently, we examined these data in closer detail to study the population dynamics of Ascaris and genetic diversity within the two isotypes and tested whether genotypes appeared to partition across human and pig hosts. Results: In the 187 isolates, 69 genotypes were found, made up of eight haplotypes of β-tubulin isotype A and 20 haplotypes of isotype B. Single nucleotide polymorphisms were seen at 14 and 37 positions for β-tubulin isotype A and isotype B, respectively. No evidence of any canonical benzimidazole resistance-associated mutations was found in either human- or pig-derived Ascaris isolates. There was, however, a difference in the genetic diversity of each isotype and distribution of β-tubulin genotypes between human- and pig-derived Ascaris. Statistical tests of population differentiation show significant differences (p < 0.001) between pig- and human-derived worms; however, more diversity was seen between worms from different populations than worms from different hosts. Conclusions: Our work suggests an absence of canonical β-tubulin mutations within Ascaris, but alternative modes of anthelminthic resistance may emerge necessitating continued genetic scrutiny alongside monitoring of drug efficacy. Graphical Abstract

Toxocara (Nematoda: Ascaridida) and Other Soil-Transmitted Helminth Eggs Contaminating Soils in Selected Urban and Rural Areas in the Philippines

The extent of contamination of soils with soil transmitted helminthes (STH) eggs, particularly Toxocara, was determined in selected urban and rural towns of Laguna, Philippines. Soil samples were collected from public schools, house yards, and empty lots. Results revealed that, of the 1480 soil samples collected, 460 (31%) were positive for STH eggs. Toxocara sp. was the most prevalent (77%), followed by Ascaris sp. (11%), hookworms/strongyles/free-living nematodes (7%), and Trichuris sp. (5%). Some soil physicochemical parameters were also determined and associated with Toxocara eggs prevalence and density in soil. Results revealed that Toxocara sp. eggs were most prevalent in less acidic, relatively high temperature and high moisture soil conditions. They were also prevalent in sandy, silty, and loamy soil textures but less prevalent in clayey. No significant differences were found between depth 1 (0–5 cm) and depth 2 (6–10 cm). This study revealed that Toxocara sp. eggs are ubiquitous and the extent of contamination in soils from the selected towns of Laguna is relatively high. Hence, the data generated in this study can be used in promoting public awareness, particularly for pet owners and local health officials, for effective prevention and control of this parasitosis

Diversity and distribution of freshwater fish assemblages in Tayabas River, Quezon (Philippines)

Three stream sections (upstream, midstream, and downstream) of Tayabas River, Philippines were surveyed during the wet and dry seasons of 2010 to evaluate the poorly known status of freshwater fish assemblages. The study collected a total of 1,070 individuals comprising 15 species, 13 genera, and 8 families. The three most abundant groups were poeciliids (61.85%), gobiids (26.16%), and cichlid (5.51%). Shannon-Weiner’s diversity indices ranged from 1.270 to 2.171. Relatively high Shannon evenness indices (0.653–0.846) and low Simpson’s dominance values (0.142–0.322) were calculated implying a fairly equitable distribution of niche space for dominant and non-dominant fishes. Significant change on fish assemblage in longitudinal gradient was observed (p&lt;0.05), being the most diverse fish assemblage registered in the upstream. Species richness is mostly composed of native fish species (10 species) and mainly represented by stream gobiids (six species). The downstream, however, had the highest cumulative abundance, in which the larger proportion was from introduced species. Also, wet season had considerably more fish species and individuals relative to dry season (p&lt;0.05). This significant spatio-temporal differences in fish assemblage data were evaluated by multivariate analyses (p&lt;0.05). Canonical correspondence analysis identified the depth (seasonal water level fluctuations), vegetation growth, and dissolved oxygen concentrations (in order of importance) as the most influential environmental parameters affecting fish assemblage structure. Also, climatic stress (prolonged drought) and anthropogenically-induced habitat alteration could negatively affect the integrity of freshwater fishes within the river. The study suggests extensive management programs of the river for the protection of native fish species

Vectors and Spatial Patterns of Angiostrongylus cantonensis

In the Philippines, rats and snails abound in agricultural areas as pests and source of food for some of the local people which poses risks of parasite transmission to humans such as Angiostrongylus cantonensis. This study was conducted to determine the extent of A. cantonensis infection among rats and snails collected from rice-farming villages of Muñoz, Nueva Ecija. A total of 209 rats, 781 freshwater snails, and 120 terrestrial snails were collected for the study. Heart and lungs of rats and snail tissues were examined and subjected to artificial digestion for parasite collection. Adult worms from rats were identified using SSU rDNA gene. Seven nematode sequences obtained matched A. cantonensis. Results revealed that 31% of the rats examined were positive with A. cantonensis. Rattus norvegicus and R. tanezumi showed prevalence of 46% and 29%, respectively. Furthermore, only Pomacea canaliculata (2%) and Melanoides maculata (1%) were found to be positive for A. cantonensis among the snails collected. Analysis of host distribution showed overlapping habitats of rats and snails as well as residential and agricultural areas indicating risks to public health. This study presents a possible route of human infection for A. cantonensis through handling and consumption of P. canaliculata and M. maculata or crops contaminated by these snails

Parasite contamination of freshly harvested vegetables from selected organic and conventional farms in the Philippines

Vegetables are considered as an important part of a healthy diet. However, there have been reports showing contamination of vegetables with parasites. This study aimed to assess parasite contamination of freshly harvested vegetables from selected organic and conventional farms in the Philippines. A total of 252 freshly harvested vegetables were collected from 20 farms through systematic random sampling and were processed by means of sedimentation technique. Positive samples were subjected to molecular analysis for further identification of species. Results showed that 58 out of 252 (23.02%) vegetable samples were contaminated with parasites eggs/cysts/oocysts. The parasites found were Ancylostoma ceylanicum, Toxocara sp., Trichuris trichiura, Ascaris suum, Hymenolepis sp., unknown trematode egg, Isospora, Balantidium, Giardia intestinalis and Cryptosporidium. Ascaris suum had the highest contamination rate in organic and conventional farms at 13.09% and 8.33%, respectively. Cryptosporidium (≥800 oocysts/ kg) and Giardia intestinalis (≥240 cysts/kg) had the highest mean density in both farms. Also, lettuce showed the highest contamination rate among the sampled vegetables in both types of farms. Furthermore, results revealed that texture of vegetables, distance to the soil substrate, and farming practices could possibly contribute to the parasite contamination of vegetables in this study. These findings have important implications on public health that may aid regulatory agencies for prevention and control strategies for food safety