Ivermectin treatment of Loa loa hyper-microfilaraemic baboons (Papio anubis): Assessment of microfilarial loads, haematological and biochemical parameters and histopathological changes following treatment.

Individuals with high intensity of Loa loa are at risk of developing serious adverse events (SAEs) post treatment with ivermectin. These SAEs have remained unclear and a programmatic impediment to the advancement of community directed treatment with ivermectin. The pathogenesis of these SAEs following ivermectin has never been investigated experimentally. The Loa/baboon (Papio anubis) model can be used to investigate the pathogenesis of Loa-associated encephalopathy following ivermectin treatment in humans. 12 baboons with microfilarial loads > 8,000mf/mL of blood were randomised into four groups: Group 1 (control group receiving no drug), Group 2 receiving ivermectin (IVM) alone, Group 3 receiving ivermectin plus aspirin (IVM + ASA), and Group 4 receiving ivermectin plus prednisone (IVM + PSE). Blood samples collected before treatment and at Day 5, 7 or 10 post treatment, were analysed for parasitological, hematological and biochemical parameters using standard techniques. Clinical monitoring of animals for side effects took place every 6 hours post treatment until autopsy. At autopsy free fluids and a large number of standard organs were collected, examined and tissues fixed in 10% buffered formalin and processed for standard haematoxylin-eosin staining and specific immunocytochemical staining. Mf counts dropped significantly (p0.05). All animals became withdrawn 48 hours after IVM administration. All treated animals recorded clinical manifestations including rashes, itching, diarrhoea, conjunctival haemorrhages, lymph node enlargement, pinkish ears, swollen face and restlessness; one animal died 5 hours after IVM administration. Macroscopic changes in post-mortem tissues observed comprised haemorrhages in the brain, lungs, heart, which seen in all groups given ivermectin but not in the untreated animals. Microscopically, the major cellular changes seen, which were present in all the ivermectin treated animals included microfilariae in varying degrees of degeneration in small vessels. These were frequently associated with fibrin deposition, endothelial changes including damage to the integrity of the blood vessel and the presence of extravascular erythrocytes (haemorrhages). There was an increased presence of eosinophils and other chronic inflammatory types in certain tissues and organs, often in large numbers and associated with microfilarial destruction. Highly vascularized organs like the brain, heart, lungs and kidneys were observed to have more microfilariae in tissue sections. The number of mf seen in the brain and kidneys of animals administered IVM alone tripled that of control animals. Co-administration of IVM + PSE caused a greater increase in mf in the brain and kidneys while the reverse was noticed with the co-administration of IVM + ASA. The treatment of Loa hyper-microfilaraemic individuals with ivermectin produces a clinical spectrum that parallels that seen in Loa hyper-microfilaraemic humans treated with ivermectin. The utilization of this experimental model can contribute to the improved management of the adverse responses in humans

The Mitochondrial Genome of Baylisascaris procyonis

BACKGROUND: Baylisascaris procyonis (Nematoda: Ascaridida), an intestinal nematode of raccoons, is emerging as an important helminthic zoonosis due to serious or fatal larval migrans in animals and humans. Despite its significant veterinary and public health impact, the epidemiology, molecular ecology and population genetics of this parasite remain largely unexplored. Mitochondrial (mt) genomes can provide a foundation for investigations in these areas and assist in the diagnosis and control of B. procyonis. In this study, the first complete mt genome sequence of B. procyonis was determined using a polymerase chain reaction (PCR)-based primer-walking strategy. METHODOLOGY/PRINCIPAL FINDINGS: The circular mt genome (14781 bp) of B. procyonis contained 12 protein-coding, 22 transfer RNA and 2 ribosomal RNA genes congruent with other chromadorean nematodes. Interestingly, the B. procyonis mtDNA featured an extremely long AT-rich region (1375 bp) and a high number of intergenic spacers (17), making it unique compared with other secernentean nematodes characterized to date. Additionally, the entire genome displayed notable levels of AT skew and GC skew. Based on pairwise comparisons and sliding window analysis of mt genes among the available 11 Ascaridida mtDNAs, new primer pairs were designed to amplify specific short fragments of the genes cytb (548 bp fragment) and rrnL (200 bp fragment) in the B. procyonis mtDNA, and tested as possible alternatives to existing mt molecular beacons for Ascaridida. Finally, phylogenetic analysis of mtDNAs provided novel estimates of the interrelationships of Baylisasaris and Ascaridida. CONCLUSIONS/SIGNIFICANCE: The complete mt genome sequence of B. procyonis sequenced here should contribute to molecular diagnostic methods, epidemiological investigations and ecological studies of B. procyonis and other related ascaridoids. The information will be important in refining the phylogenetic relationships within the order Ascaridida and enriching the resource of markers for systematic, population genetic and evolutionary biological studies of parasitic nematodes of socio-economic importance

A murine macrofilaricide pre-clinical screening model for onchocerciasis and lymphatic filariasis

Background: New drugs effective against adult filariae (macrofilaricides) would accelerate the elimination of lymphatic filariasis and onchocerciasis. Anti-Onchocerca drug development is hampered by the lack of a facile model. We postulated that SCID mice could be developed as a fmacrofilaricide screening model. Methods: The filaricides: albendazole (ABZ), diethylcarbamazine (DEC), flubendazole (FBZ), ivermectin (IVM) and the anti-Wolbachia macrofilaricide, minocycline (MIN) were tested in Brugia malayi (Bm)-parasitized BALB/c SCID mice vs vehicle control (VC). Responses were compared to BALB/c wild type (WT). Onchocerca ochengi male worms or onchocercomata were surgically implanted into BALB/c SCID, CB.17 SCID, BALB/c WT mice or Meriones gerbils. Survival was evaluated at 7–15 days. BALB/c SCID were tested to evaluate the responsiveness of pre-clinical macrofilaricides FBZ and rifapentine (RIFAP) against male Onchocerca. Results: WT and SCID responded with >95% efficacy following ABZ or DEC treatments against Bm larvae (P < 0.0001). IVM was partially filaricidal against Bm larvae in WT and SCID (WT; 39.8%, P = 0.0356 and SCID; 56.7%, P = 0.026). SCID responded similarly to WT following IVM treatment of microfilaraemias (WT; 79%, P = 0.0194. SCID; 76%, P = 0.0473). FBZ induced a total macrofilaricidal response against adult Bm in WT and SCID (WT; P = 0.0067, SCID; P = 0.0071). MIN induced a >90% reduction in Bm Wolbachia burdens (P < 0.0001) and a blockade of microfilarial release (P = 0.0215) in SCID. Male Onchocerca survival was significantly higher in SCID vs WT mice, but not gerbils, after +15 days (60% vs 22% vs 39% P = 0.0475). Onchocercoma implants had engrafted into host tissues, with evidence of neovascularisation, after +7 days and yielded viable macro/microfilariae ex vivo. FBZ induced a macrofilaricidal effect in Onchocerca male implanted SCID at +5 weeks (FBZ; 1.67% vs VC; 43.81%, P = 0.0089). Wolbachia loads within male Onchocerca were reduced by 99% in implanted SCID receiving RIFAP for +2 weeks. Conclusions: We have developed a ‘pan-filarial’ small animal research model that is sufficiently robust, with adequate capacity and throughput, to screen existing and future pre-clinical candidate macrofilaricides. Pilot data suggests a murine onchocercoma xenograft model is achievable