Hess, R.

Poinar, G.O. Jr

Horizon / Pleins textes

Formation of crystals  within cells of two mermithid  nematodes 
George O. POINAR, Jr  and Roberta HESS 
Division of Entomology  and  Parasitologg,  University  of  California,  Berkeley,  California 94720, U S A .  
SUMMARY 
Disc-like platelets of supposed  proteinaceous  nature  composed of dense  subunits  approximately 8 nm‘in  diameter 
arranged  in a crystalline  pattern  were  observed  in  the  pseudocoelom of two  mermithid  nematodes, Gastromermis 
sp.  and Romanomermis culicivorax. In addition  morphologically  similar  subunits  were observed in the  nuclei  and 
cytoplasm of hypodermal  cord cells, It is  proposed  that  the  subunits  are  formed  intracellularly  and  pass  into  the 
pseudocoelom  where they  aggregate  into  large  platelets.  It  is  possible  that  these  crystals  represent  excretory  pro- 
ducts  stored  in  a  non-toxic  state, so that   they will not  weaken  the  relatively  small  invertebrate  host of these  internal 
parasites. 
RÉSUMÉ 
Formation de cristauz à l’intérieur des cellules de, deux Mermithides 
Les auteurs ont observé dans le pseudocoelome de deux nématodes Illermithides, Gasfro7ner1nis sp. e t  Romu- 
nomermis culicivoraz, des petites plaques ayant l’apparence de disques protéiques e t  composées de sous-unités 
denses  d’un  diamktre  approximatif  de 8 nrn présentant  un  arrangement  cristallin. 
Des éléments  cristallins  ont  été  aussi  observés  dans  les  noyaux  et le cytoplasme  des cellules des  cordes  hypoder- 
miques. Les sous-unités de ces petites plaques se formeraient à l’intérieur des cellules, puis passeraient dans le 
pseudocoelome où elles s’agrégeraient en formations plus importantes. Ces cristaux pourraient représenter des 
produits  d’excrétion  stockés sous forme  non  toxique,  donc  non  nocive  pour les insectes  h8tes  relativement  petits  de 
ces parasites internes que sont ces Mermithides. 
An earlier study  (Poinar, Leutenegger & 
Thomas,  1970) described disc-like platelets 
of supposed  proteinaceous nature from the 
pseudocoelom of free living post-parasitic  juve- 
niles of a mermithid nematode now considered 
in  the genus Gastromepnis. At  hat  ime, a 
survey showed that similar objects had been 
reported  in  various species of mermithid  nema- 
todes. Bugnion (1878) called them blood cor- 
puscles in Lirnrtornerrnis aquatilis (Duj.), Hag- 
meier (1912) referred t o  them as free-floating 
hyaline bodies in Para.merrnis aquafilis and 
other  mermithids  whereas Muller (1931) and 
Dollfus (1946) considered them as possible 
internal parasites. 
The  above  mentioned study showed the discs 
t o  consist of a highly  organized pattern of 
R e v u e  Nématol.  3 ( 1 )  : 107-113 (1980) 
parallel, dense lines. However, the origin anci 
formation of the discs were not investigated. 
Materials and methods 
The  platelets were examined in two species of 
mermithid  nematodes  in  the  present  study. 
One was the undescribed species of Gastro- 
rnermis  studied  by  Poinar, Leutenegger and 
Thomas  (1970) which parasitizes midges of 
the genus Cricotopus in fastflowing mountain 
streams in California. The second species was 
Rornanolnermis culicivorax Ross & Smith, a 
parasite of mosquito larvae originally collected 
in Louisiana and mass-reared by  Dr. J.J. Peter- 
sen. 
107 
G.O. Poinar Jr & R. Hess 
Fig. 1 (above) : Light microscope whole mount of Gastro~nermis sp. showing 
two adjacent. protein platelets floating in pseudocoelom (arrows). 
Fig. 2, (below) : Light mic,roscope whole mount of Romanomermis eulicivorax 
showing clusters of protein  platelets  accumulated  in  the pseudoc~oelom (arrows). 
Reurw Nématol. 3 ( 1 )  : 107-113 (1980) 
Fig. 3 (above) : Crystalline  lattice (C) in the nucleus of a hypodermal ce11 of Gastromermis 
sp.  Insert : higher  magnification of crystalline  subunits. 
Fig. 4 (below) : Crystalline  lattice (C) in the  nucleus of a hypodermal ce11 of Romanornermis 
culiciuorax. Insert : higher magnification of crystalline subunits. 
' Revue Nèmatol.  3 (1) : 107-113 (1980) 1 O9 
G.O. Poinar J r  dl. R. Hess 
Fig. 5 (above): Crystalline 1att.ice. (C) in t.he rytoplasm of a  hypodermal ce11 of Gastromermis Sp. 
Fig. 6 (below) : Crystalline lattice. ( C )  in the rytoplasm of a hypodermal cell of Romano- 
mermis culicivoran. 
Re 
Fig. 7 (above) : Protein  platelet  in  the  pseudocoelom of Gastromerrnis sp. Arrow indicates 
limiting b membrane o. Note elcctron dense outer boundary. Insert : portion of a protcin 
platelet in Gastrolnerrilis sp. showing structure similar to a crystalline lattice. 
Fig. 8 (below) : Protein  platelet in the pscudocoelom of H o n ~ a n o ~ n e r ~ n i s  culicivoras.  Insert 
shows structure similar to a crystalline lattice. Arrows indicate subunits in the pseu- 
dococlom. 
'vue Ne'nzatol. 3 (1)  : 107-113 (1980) 
G.O. Poinar J r  Le R. Hess 
For  ultrastructural  investigations,  adults  and 
postparasitic  juveniles of botth species and  para- 
sitic stages of R. crtlicivorax were prefixed for 
2 h in a 4 y0 solution of glutaraldehyde, buffered 
with  0.1 M phosphate  buffer,  then fixed  for 
1 h in 1 % osmium  tetrosicle.  After  dehydrat&ion 
in an alcohol series and  embedding  in  Araldite, 
sections cut with a Port.er-Blum MT-2 micro- 
tome were stained  with  urangl  acetde  and lead 
citrate and esamined with an RCA F3 and a 
Philips 300 electron microscope. 
Results 
In  both species of living  nemat,odes,  platNelets 
could be seen lloating  freely in t,he  pseudo- 
coelom or  aggregat.ed in clusters bet.meen the 
trophosome  and  muscle  layer  (Figs 1 and 2.). 
In  Gasfromermis sp., the  platelets  were  flat 
cireular dises and had an average diameter of 
23.2 pm (14.6-30.5) and an average  width of 
5.5 pm (3.7-7.3). They were  hyaline and not 
refringent  under  bright, field. They possessed an 
indist.inct limiting (( membrane 1) (Fig. 7) .  
In R. culicivorax the plat.elets were smaller, 
averaging 2 pm by 2.5 pm  in diameter. The 
structure of ' the p1atelet.s varied  from  aval 
(Fig. 2,) t.0 lobulate  (Fig. 8). They did not possess 
a limiting  membrane  or  interface  stmcture 
variation  (insert  Fig. 8). 
EIigh magnific,at,ion showecl that,  the  platelets 
were composed of dense subunits  approxima- 
tely 8 nm  in  diameter.  The su bu nit,^ were 
arrangecl in a cyst.alline pattern  with  an  average 
center t.o center  spacing of 23 nm  (inserts 
Fig. 7, 8). In  Gasfromernzis sp. the  subunik 
were  generally  aligned  within  one  or  two  planes, 
whereas  ïn R. c d i c i u o r n s  the  subunits were  more 
frequently aligned in many planes giving the 
platelet, a heterogeneous  appearanc,e  (Figs, 7, 8). 
In  both  nematodes, srnall units of c.rystalline 
1attic.e similar t o  those in t-he plat,elet.s of the 
pseudocoelonl  occ.urred within  the nuclei of 
t,he hypodermal cord cells. I n  Gasirornermis sp. 
these  units  often oc.c,urrecl adjac.ent, t o  extensive 
elwtron-dense nucleoli (Fig. 3). In  R. cul ic iuoraz,  
they were present in clear nuclei wit.h reduced 
chromatin and no apparent nucleoli (Fig. 4). 
- In ot_her--bs-o_dymal c&- 01- both  nzmatodes, 
crystalline  structures  similar to those  in  the 
platelets occurrecl in the  cytoplasm of the c.ells 
(Figs. 5, 6). 
When eut in cross section, the subunits of 
the  c.rystdline  arrays  found  in  the  nucleus, 
cytoplasm and pseudocoelom were round with 
a dense outer wall and less dense c.ent,er which 
gave  them a lzollow appearance  (inserts,  Figs  4, 
7, 8). Cross bridges  interconnecting tbe units 
could be seen (insert,, Fig. 8). Sec.tions c.ut in 
older  planes  showed t,he subunits  as dense 
parallel bands  (Figs 3, 4, 5, 6). The  angles 
between the subunit, array varied according to 
the  plane of seckion. The  insert of Fig. 8 shows 
units  similar t o  those. of the  platelet a t  the  
boundary of the pseudocoelom (arrows). 
Discussion 
The morphologic.al subunits composing the 
plat.elets of supposed  proteinaceous natnre  found 
in the pseudocoelo~n of Gasfromermis  sp. and 
R. culicivoran: are similar in structure t,o those 
crystals  found  in the nucleus and  cytoplasm of 
the hypodermal  cord cells. Similar subunits 
' were found at   the  edge of the  crystalline 1at.tic.e 
in the llypodermal ce11 cytoplasm and at, the 
boundaries of the plat,elet,s in  the pseudocoelom. 
This  suggests,  but does not  imply,  that,  the 
crystalline  subunits  are assembled within  the 
ce11 and sec.ret,ed into the pseudoc.oelom where 
they aggregate into platelets. 
The substxucture of the platlelets  fronl the 
two ne.mat.ode species is remarkahly  similar, 
heing  composed of subunits of the same size 
and  spacing  and  nmy  be of similar c.omposit.ion. 
blermithicl nematodes  ingeneral do not. 
possess a conqlete intestine wit.11 an external 
opening, nor do they possess  a funetional 
excretory system as do most other nematodes. 
Although the exaci chemic.al nature of t l m e  
plaLeletx is not yet lrnown they may possibly 
represent waste material stored in a non-toxic. 
form. This would ensure that  normal cellular 
metabolitles  would not  be releasecl in liquicl 
state  into  the  nematode  and in turn be  carried 
into  the inseck where  t,hey  could  interfere 
with normal  development of bath host, ancl 
rasite -pa---.------- -- - _ _ _ _  _ _ _  _ _  
R e v u e   N é m a t o l .  3 (1)  : 107-113 (1980) 112 
Cystals  within  cells of Merrnithids 
REFERENCES 
BUGNION, E. (1878). Note sur les  globules  sanguins  du 
Mermis  aquatil is  Duj.  suivie  de  quelques  remarques 
sur la structure anatomique de cette espèce. Actes 
Soc. Helvétique  Sei.  nat., 1876-1877 : 247-255. 
DOLLFUS, R. (1976). Parasites des Helminthes. Ency- 
clopédie biologique, vol. 27. Paris, P. Lechevalier, 
482 p. 
Accepté pour publication le 22 octolke 1979. 
HAGRIEIER, A. (1912). Beitrage zur Kenntnis  der 
Mermithiden. 1. Biologische  Notizen  und  syste- 
matische  Beschreibung  einiger  alter  und  neuer 
Arten. Zool. Jahrb.,  32 : 521-612. 
MULLER, G. W. (1931). Über Mermithiden. Z. Morph .  
Morph.  okol. Tiere,  24 : 82-147. 
POINAR, Jr. G. O., LEUTENEGGER,  R. & THOMAS, G. M. 
(1970). On the  occurrence of protein  platelets  in  the 
pseudocoelom of a nematode (Hydromermis  sp. : 
Mermithidae). Nematologica, 16 : 348-352. 
Revue Nhna to l .  3 (1) : 107-113 (1980) 113 


Formation of crystals within cells of two mermithid nematodes

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