The in vitro development of the pentastomid Porocephalus
crotali from the infective, seventh (VII) instar, dissected from
the tissues of rat intermediate hosts, to the adult male (X) and female
(XI)
instars, normally resident in the lung of
rattlesnake definitive hosts, is described. The culture medium comprised
washed human blood cells, resuspended in bovine
serum (50[ratio ]50, v/v), with 20% minimum essential medium and
antibiotics. Two batches of approximately 100 pentastomids, maintained
at a
density of 2 worms/ml, were cultured in 500 ml bottles in an atmosphere
of 5% CO2 in air at
28°C. Culture bottles were rotated slowly on a Rollacell system and the
medium
was replenished every 2–3 days. Growth
was monitored at various intervals by weighing worms, measuring cast
cuticles with attached moulted hooks recovered
from the medium, and by dissecting worms for measurements of various organ
systems. Three moults separated the
infective and adult male instar, whereas 4 moults were necessary in the
case
of females. In both cultures natural mortality
was about 10% over a 160-day period, by which time 33–51% of
females and 62–70% of males had reached the
terminal instar. Some males attained full sexual maturity with seminal
vesicles loaded with viable sperm: such males were
indistinguishable from males recovered from naturally infected rattlesnakes.
However, females never achieved full size,
even after 200 days, and although copulation did not occur in vitro,
some females became patent.