Abstract
Nematode spicules vary in shape and size even between closely related species and, therefore, constitute key characters in nematode taxonomy for distinguishing between species. Spicules are seldom measured on fresh specimens, but rather at some time after extraction from culled hosts and after a period of preservation of the worms in chemical fixatives or by freezing. We carried out two experiments to assess the effects of freezing in Hanks' balanced salt solution, 70% or 80% ethanol and 10% formalin (both of the latter at room temperature and after storage at -80°C) on spicule length of Heligmosomoides bakeri at two time intervals after extraction from mice (Experiment 1, one and four weeks; Experiment 2, one and four months). In Experiment 1, no significant differences were detected, although there was some variation between treatments and over time. In Experiment 2, spicule length varied significantly between treatments and over time, the greatest shrinkage being in 80% ethanol and the least in 10% formalin. However, overall variation in spicule length was very limited, accounting for no more than 5.03% change in length over time and 4.95% between treatments at any of the periods of assessment. Therefore, while whole nematodes can shrivel and shrink in preservatives, making many measurements unreliable, our data indicated that spicule lengths are very little changed by preservation techniques over time, and so spicule length remains as a reliable taxonomic character.
Highlights
The characteristics of nematode spicules are used widely to distinguish between species, because they vary in shape and size between nematode genera and between congeners (MAFF, 1986; Kaufmann, 1996)
The main effect of freezing was not significant (F1, 195 = 0.03, P = 0.87), nor were any interactions incorporating freezing significant, the most marked shrinkage in mean length was in worms that had been frozen at −80°C in Hanks’ balanced salt solution (HBSS)
The largest discrepancy in mean lengths between treatments in month 1 was 4.32% and in month 4, 2.33%
Summary
The characteristics of nematode spicules are used widely to distinguish between species, because they vary in shape and size between nematode genera and between congeners (MAFF, 1986; Kaufmann, 1996). It is not usually possible to examine and measure worms immediately after their hosts have been culled, because hosts may have been trapped in remote locations, distant from laboratory facilities, and may have to be culled locally and preserved intact or after separation of body parts. Helminths are usually extracted at subsequent laboratory dissection of intestinal tracts or other body organs that have been preserved and stored for varying periods, sometimes for decades or even longer. Each method is known to have its own, distinct effects on tissues (Stoddard, 1989), the precise range of the possible effects on nematode spicules is not known. Knowledge about the extent of the influence of different preservatives on spicule length is crucial if measurements from different isolates are to be compared, for example, for taxonomic purposes
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