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Larval migration in PERL chambers as an in vitro model for percutaneous infection stimulates feeding in the canine hookworm Ancylostoma caninum.

Franke D, Strube C, Epe C, Welz C, Schnieder T - Parasit Vectors (2011)

Bottom Line: Additionally, infective larvae of A. caninum were activated via serum-stimulation and feeding behaviour was analysed and compared between percutaneously migrated and serum-stimulated larvae.The medium beneath the skin had no effect on migration ratio, and no significant difference between the migration ratios through fresh and frozen/thawed skin was observed.The observed difference in time course of resumption of feeding indicates that percutaneously migrated larvae are not identical to serum-stimulated larvae, which are currently representing the model for early parasitic stages.

View Article: PubMed Central - HTML - PubMed

Affiliation: Institute for Parasitology, University of Veterinary Medicine Hannover, Buenteweg 17, 30559 Hannover, Germany.

ABSTRACT

Background: Ancylostoma caninum third-stage larvae are the non-feeding infective stage of this parasite and are able to infect potential hosts via different infection routes. Since percutaneous infection is one of the most important routes and skin penetration is the first step into parasitic life, an existing in vitro model for percutaneous migration was modified and evaluated. The main parameter used to evaluate migration was the migration ratio (migrated larvae as a percentage of total number of larvae recovered). Additionally, the skin lag was calculated, expressing the percentage of larvae remaining in the skin and therefore not being recovered. Since initiation of feeding is proposed to be an important step in the transition from free-living to parasitic A. caninum larvae, feeding assays were performed with in vitro percutaneously migrated larvae. Additionally, infective larvae of A. caninum were activated via serum-stimulation and feeding behaviour was analysed and compared between percutaneously migrated and serum-stimulated larvae.

Results: Maximum skin migration levels of infective larvae were observed at temperatures above 32°C when larvae were placed on the epidermal side of skin for more than 12 hours. The medium beneath the skin had no effect on migration ratio, and no significant difference between the migration ratios through fresh and frozen/thawed skin was observed. Maximum feeding levels of 93.2% were observed for percutaneously migrated larvae after 48 h incubation, whereas serum-stimulated larvae reached the maximum of 91.0% feeding larvae after 24 h.

Conclusions: The PERL chamber system was optimised and standardised as an in vitro model for percutaneous migration. The larvae recovered after percutaneous migration showed characteristic signs of activation similar to that of serum-stimulated larvae. The observed difference in time course of resumption of feeding indicates that percutaneously migrated larvae are not identical to serum-stimulated larvae, which are currently representing the model for early parasitic stages.

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Effect of incubation period on the migration behaviour of the L3. Each point represents the mean ± SD (n = 6). Larvae were incubated in PERL chambers for 1 h, 4 h, 8 h, 12 h, 16 h, and 24 h, respectively. A: Migration ratio over time. B: Total number of larvae recovered from the donor compartment (non-migrated larvae) and from the acceptor compartment (migrated larvae).
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Figure 4: Effect of incubation period on the migration behaviour of the L3. Each point represents the mean ± SD (n = 6). Larvae were incubated in PERL chambers for 1 h, 4 h, 8 h, 12 h, 16 h, and 24 h, respectively. A: Migration ratio over time. B: Total number of larvae recovered from the donor compartment (non-migrated larvae) and from the acceptor compartment (migrated larvae).

Mentions: The incubation time (Figure 4) during that larvae were allowed to migrate had a significant effect on the migration ratio (Kruskal-Wallis One-Way ANOVA, p < 0.001; H = 30.241 with 5 degrees of freedom) and skin lag (One-Way ANOVA, p < 0.001). After 1 hour of incubation most of the applied 300 larvae had invaded the skin, as the mean number of larvae recovered from the donor compartment was 42.33 (SD = 10.98), but the larvae could not be recovered from the acceptor compartment at that time point (Figure 4A). Therefore, the migration ratio was 0% for 1 h incubation time and increased over time (Figure 4B). The mean migration ratios were highest for incubation periods of 12 h and more (detailed results including mean skin lag are listed in Table 3). These migration ratios were not significantly different from each other (Dunn's test, p > 0.05) but from that after 1 h incubation (Dunn's test, p < 0.05). The mean skin lag also decreased from shorter to longer incubation periods. The skin lag was not statistically significant between the experiments with 1 h and 4 h incubation time and between the experiments with incubation periods of 12 h and longer (Holm-Sidak method, p > 0.01). All other comparisons detected significant differences (Holm-Sidak method, p < 0.01).


Larval migration in PERL chambers as an in vitro model for percutaneous infection stimulates feeding in the canine hookworm Ancylostoma caninum.

Franke D, Strube C, Epe C, Welz C, Schnieder T - Parasit Vectors (2011)

Effect of incubation period on the migration behaviour of the L3. Each point represents the mean ± SD (n = 6). Larvae were incubated in PERL chambers for 1 h, 4 h, 8 h, 12 h, 16 h, and 24 h, respectively. A: Migration ratio over time. B: Total number of larvae recovered from the donor compartment (non-migrated larvae) and from the acceptor compartment (migrated larvae).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3037914&req=5

Figure 4: Effect of incubation period on the migration behaviour of the L3. Each point represents the mean ± SD (n = 6). Larvae were incubated in PERL chambers for 1 h, 4 h, 8 h, 12 h, 16 h, and 24 h, respectively. A: Migration ratio over time. B: Total number of larvae recovered from the donor compartment (non-migrated larvae) and from the acceptor compartment (migrated larvae).
Mentions: The incubation time (Figure 4) during that larvae were allowed to migrate had a significant effect on the migration ratio (Kruskal-Wallis One-Way ANOVA, p < 0.001; H = 30.241 with 5 degrees of freedom) and skin lag (One-Way ANOVA, p < 0.001). After 1 hour of incubation most of the applied 300 larvae had invaded the skin, as the mean number of larvae recovered from the donor compartment was 42.33 (SD = 10.98), but the larvae could not be recovered from the acceptor compartment at that time point (Figure 4A). Therefore, the migration ratio was 0% for 1 h incubation time and increased over time (Figure 4B). The mean migration ratios were highest for incubation periods of 12 h and more (detailed results including mean skin lag are listed in Table 3). These migration ratios were not significantly different from each other (Dunn's test, p > 0.05) but from that after 1 h incubation (Dunn's test, p < 0.05). The mean skin lag also decreased from shorter to longer incubation periods. The skin lag was not statistically significant between the experiments with 1 h and 4 h incubation time and between the experiments with incubation periods of 12 h and longer (Holm-Sidak method, p > 0.01). All other comparisons detected significant differences (Holm-Sidak method, p < 0.01).

Bottom Line: Additionally, infective larvae of A. caninum were activated via serum-stimulation and feeding behaviour was analysed and compared between percutaneously migrated and serum-stimulated larvae.The medium beneath the skin had no effect on migration ratio, and no significant difference between the migration ratios through fresh and frozen/thawed skin was observed.The observed difference in time course of resumption of feeding indicates that percutaneously migrated larvae are not identical to serum-stimulated larvae, which are currently representing the model for early parasitic stages.

View Article: PubMed Central - HTML - PubMed

Affiliation: Institute for Parasitology, University of Veterinary Medicine Hannover, Buenteweg 17, 30559 Hannover, Germany.

ABSTRACT

Background: Ancylostoma caninum third-stage larvae are the non-feeding infective stage of this parasite and are able to infect potential hosts via different infection routes. Since percutaneous infection is one of the most important routes and skin penetration is the first step into parasitic life, an existing in vitro model for percutaneous migration was modified and evaluated. The main parameter used to evaluate migration was the migration ratio (migrated larvae as a percentage of total number of larvae recovered). Additionally, the skin lag was calculated, expressing the percentage of larvae remaining in the skin and therefore not being recovered. Since initiation of feeding is proposed to be an important step in the transition from free-living to parasitic A. caninum larvae, feeding assays were performed with in vitro percutaneously migrated larvae. Additionally, infective larvae of A. caninum were activated via serum-stimulation and feeding behaviour was analysed and compared between percutaneously migrated and serum-stimulated larvae.

Results: Maximum skin migration levels of infective larvae were observed at temperatures above 32°C when larvae were placed on the epidermal side of skin for more than 12 hours. The medium beneath the skin had no effect on migration ratio, and no significant difference between the migration ratios through fresh and frozen/thawed skin was observed. Maximum feeding levels of 93.2% were observed for percutaneously migrated larvae after 48 h incubation, whereas serum-stimulated larvae reached the maximum of 91.0% feeding larvae after 24 h.

Conclusions: The PERL chamber system was optimised and standardised as an in vitro model for percutaneous migration. The larvae recovered after percutaneous migration showed characteristic signs of activation similar to that of serum-stimulated larvae. The observed difference in time course of resumption of feeding indicates that percutaneously migrated larvae are not identical to serum-stimulated larvae, which are currently representing the model for early parasitic stages.

Show MeSH
Related in: MedlinePlus