Rev. Soc. Bras. Med. Trop. vol.38 número2

2 0 5

Revista da Sociedade Br asileir a de Medicina Tr opical 3 8 ( 2 ) :2 0 5 -2 0 6 , mar-abr, 2 0 0 5

In vitro

maintenance of

An giostron gylu s costaricen sis

does

not provide physiological conditions for egg laying

Manutenção in vitro

de Angio stro ngylus co sta ricensis

não constitui condição fisiológica para ovipostura

Márcia Bohrer Mentz

1

_{and Carlos Graeff-Teixeira}

2

ABSTRACT

Fi f te e n p a i rs ( m a le /f e m a le ) o f Angio str o ngylus c o star ic e nsis

we re k e p t in vitr o i n Wa ym o u th m e d i u m f o r th re e d a ys to

e va lu a te th e a m o u n t a n d d u ra ti o n o f e gg la yi n g. At 2 4 , 4 8 a n d 7 2 h o u rs, th e m e a n e gg c o u n ts we re 3 2 1 , 2 4 a n d 4 e ggs/

1 0 m i c ro li te rs, re sp e c ti ve ly. Mo st o f th e e ggs we re e li m i n a te d wi th i n th e f i rst 2 4 h o u rs, su gge sti n g th e y a re e x p e lle d

u n d e r n o n - p h ysi o lo gi c a l c o n d i ti o n s. Th e se re su lts i n d i c a te th a t in vitr o c o n d i ti o n s a re n o t a p p ro p ri a te f o r d ru g tri a ls o f

e gg- la yi n g i n h i b i to rs f o r tre a tm e n t o f a b d o m i n a l a n gi o stro n gylo si s.

Ke y-words

: Angio str o ngylus c o star ic e nsis. Ab d o m i n a l a n gi o stro n gyli a si s. Egg la yi n g.

RESUMO

Qu i n ze d u p la s ( m a c h o /f ê m e a ) d e Angio str o ngylus c o star ic e nsis f o ra m m a n ti d a s

in vitr o

, e m m e i o d e Wa ym o u th d u ra n te

3 d i a s, p a ra o b se rva ç ã o d a q u a n ti d a d e e d u ra ç ã o d a o vi p o si ç ã o . Mé d i a s d e 3 2 1 , 2 4 e 4 o vo s e m 1 0 m i c ro li tro s f o ra m

re gi stra d a s e m 2 4 , 4 8 e 7 2 h o ra s, re sp e c ti va m e n te . A m a i o ri a d o s o vo s f o i e li m i n a d a n a s p ri m e i ra s 2 4 h o ra s, su ge ri n d o

te re m si d o e x p u lso s e m c o n d i ç õ e s n ã o f i si o ló gi c a s. Este s re su lta d o s i n d i c a m q u e a s c o n d i ç õ e s in vitr o

n ã o sã o a d e q u a d a s

p a ra te ste s d e d ro ga s i n i b i d o ra s d a o vi p o si ç ã o , p a ra tra ta m e n to d a a n gi o stro n gi lí a se a b d o m i n a l.

Pal avr as-chave s

: Angio str o ngylus c o star ic e nsis. An gi o stro n gi lí a se a b d o m i n a l. Ovi p o si ç ã o .

1 . Se to r de Par asito lo gia do De par tame nto de Mic r o b io lo gia do Instituto de Ciê nc ias B ásic as da Saúde da Unive r sidade Fe de r al do Rio Gr ande do Sul, Po r to Ale gr e , RS, B r asil. 2 . Instituto de Pe sq uisas B io mé dic as da Po ntifíc ia Unive r sidade Cató lic a de Po r to Ale gr e , RS, B r asil.

Financ ial suppo r t: CAPES-PROB AL, 0 5 5 /9 7 AND PUC/RS

Addr e ss to: Dr a. Már c ia B o hr e r Me ntz. R. Fe r nande s Vie ir a 6 3 4 /8 0 1 , B air r o B o m Fim, 9 0 0 3 5 - 0 9 0 Po r to Ale gr e , RS. e - mail: mb me ntz@ uo l. c o m. b r

Re c e b ido par a pub lic aç ão e m 6 /2 /2 0 0 4 Ac e ito e m 1 6 /1 2 /2 0 0 4

COMUNICAÇÃO/COMMUNICATION

Angio stro ngylus co sta rice nsis Morera & Cespedes, 1 9 7 1 , is

a parasite nematode of the mesenteric arterial system in wild

rodents

_{. The intermediate hosts are terrestrial mollusks, mostly}

of the Veronicellidae family

_{. Abdominal angiostrongylosis may}

result from accidental human infection, leading to severe ischemic

and inflammatory intestinal lesions

_{. Sinc e the fir st r e po r t in}

Co sta Ric a

_{, many human c ase s have b e e n r e po r te d fr o m}

Mexic o

_{to B razil}

_{. B esides its medic al impo rtanc e,}

An gi o stro n gylu s c o sta ri c e n si s has b e e n se le c te d as a mo de l

to study the dise ase s c ause d b y tissue ne mato de s and the

e ffic ac y o f anthe lm intic s

_{. The lar ge sc ale pr o duc tio n o f}

lar vae and wo r ms, as we ll as the pr inc iple o f minimizing

l a b o r a to r y a n i m a l u s e j u s ti fi e s th e n e e d fo r

i n vi t r o

mainte nanc e o f par asite s. No ne o f the r e po r ts fr o m tr ials o f

i n vi tro

c ultivatio n o f A. c o sta ri c e n si s

we r e c o nc e r ne d with

the quantity and dur atio n o f e gg laying

_{. Ho we ve r,}

th e s e da ta a r e im po r ta n t to e va lua te th e us e fuln e s s o f a n

2 0 6

Me ntz MB e t al

After washing, a total of 1 5 male/female pairs were separated

and tr ansfe r r e d o nto ano the r plate , c o ntaining Waymo uth

medium ( GIBCO) plus antibiotic and amphotericin and incubated

at 3 7 ºC and 5 % CO

_{. The medium was c hanged at 1 -day intervals.}

Eggs were examined under a stereomic rosc ope and c ounted,

every 2 4 hours, for 7 2 hours. Most of the eggs were rec overed in

the first 2 4 hours, with a mean 3 2 1 eggs/female/1 0 mL. After 4 8

hours, the number of eggs dec reased dramatic ally, to 2 4 eggs/

female/1 0 mL. After 7 2 hours, only a few dark and deformed

eggs were rec overed ( Table 1 ) .

4 . Céspedes R, Salas J, Mekbe S, Troper L, Mülllner F, Morera P. Granulomas e nté r ic o s e linfátic o s c o m inte nsa e o sino filia tisular pr o duc ido s po r un strongilídeo ( strongylata) . Ac ta Médic a Costaric ensis 1 0 :2 3 5 -2 5 5 , 1 9 6 7 . 5 . Graeff-Teixeira C, Avila-Pires FD, Machado, RCC, Camillo-Coura L, Lenzi HL.

Identific aç ão de roedores silvestres c omo hospedeiros do Angio stro ngylus co sta ricensis no Sul do Brasil. Revista do Instituto de Medicina Tropical de São Paulo 3 2 : 1 4 7 -1 5 0 , 1 9 9 0 .

6 . Graeff-Teixeira C, Camillo-Coura L, Lenzi Hl. Clinic al and epidemiologic al aspec ts of abdominal angiostrongyliasis in Southern Brazil. Revista do Instituto de Medic ina Tropic al de São Paulo 3 3 : 3 7 3 -3 7 8 , 1 9 9 1 .

7 . Graeff-Teixeira C. Camillo-Coura L., Lenzi Hl. Histopathologic al c riteria for the dia gno s is o f a b do m ina l a ngio s tr o ngylia s is . Pa r a s ito lo gy Re s e a r c h 7 7 : 6 0 6 -6 1 1 , 1 9 9 1 b.

8 . Graeff-Teixeira C, Morera P. Método de digestão de moluscos em ácido clorídrico para isolamento de larvas de metastrongilídeo. Biociências3 :8 5 -8 9 , 1 9 9 5 . 9 . Gr aeff-Teixeir a C, Tho mé JW, Pinto SCC, Camillo -Co ur a L, Lenzi HL.

Phyllo ca ulis va rie ga tus an intermediate host of Angio stro ngylus co sta rice nsis

in South B razil. Memórias do Instituto Oswaldo Cruz 8 4 :6 5 -6 8 , 1 9 8 9 . 1 0 . Hata H. Essential amino ac ids and other essential c omponents for development

o f An gio stro n gylu s c o sta ric e n sis from third-stage larvae to young adults. Journal of Parasitology 8 0 : 5 1 8 -5 2 0 , 1 9 9 4 .

1 1 . Hata H. In vitro c ultivation of Angio stro ngylus c o sta ric e nsis eggs to first s ta ge la r va e in c he m ic a lly de fine d m e dium . I nte r na tio na l J o ur na l fo r Parasitology 2 6 : 2 8 1 -2 8 6 , 1 9 9 6 .

1 2 . Hata H, Kojima S. Angio stro ngylus co sta rice nsis: Culture of third-stage larvae to yo un g a dults in a De fin e d Me dium . Ex pe r im e n ta l Pa r a s ito lo gy 7 3 : 3 5 4 - 3 6 1 , 1 9 9 1 .

1 3 . Iabuki K, Montenegro MR. Apendic ite por Angio stro ngylus c o sta ric e nsis. Apresentaç ão de um c aso. Revista do Instituto de Medic ina Tropic al de São Paulo 2 1 : 3 3 -3 6 , 1 9 7 9 .

1 4 . Lobo-Sanahuja F, Loria-Cortes R, Gonzalez G. Angiostrongilosis abdominal. Aspec tos c línic os, tratamento y revisión de literatura. B oletin Médic o Hospital Infantil 4 4 : 4 -9 , 1 9 8 7 .

15. Loria-Cortes R, Lobo Sanahuja JF. Clinical abdominal angiostrongylosis. A study of 116 children with intestinal eosinophilic granuloma caused by Angiostrongylus costaricensis. American Journal of Tropical Medicine and Hygiene 29: 538-544, 1980. 1 6 . Mo r e r a P. I n ve s tiga c ió n de l h ué s p e d de fin itivo de An gi o s tr o n gylu s

c o sta ric e nsis ( Morera y Céspedes, 1 9 7 1 ) . B oletin Chileno de Parasitologia 2 5 : 1 3 3 -1 3 4 , 1 9 7 0 .

1 7 . Mo r e r a P. Ab do m in a l a n gio s tr o n gylia s is : a pr o b le m o f pub lic h e a lth . Parasitology Today 1 : 1 7 3 -1 7 5 , 1 9 8 5 .

1 8 . Morera P, Ash LR. Investigac ión del huésped intermediário de Angio stro ngylus c o sta ric e nsis ( Morera & Cespedes, 1 9 7 1 ) . B oletin Chileno de Parasitologia 2 5 :1 3 5 , 1 9 7 0 .

1 9 . Morera P, Céspedes R. Angiostrongylose abdominal. Una nueva parasitosis humana. Ac ta Médic a Costa Ric a 1 4 : 1 5 9 -1 7 3 , 1 9 7 1 .

2 0 . Terada M, Rodriguez O, Dharejo AM, Ishii AI , Sano M. Studies on c hemotherapy of parasitic helminths. ( XXVI) Comparative in vitro effects of various anthelmintics o n the m o tility o f An gi o stro n gylu s c o sta ri c e n si s and An gi o stro n gylu s ca nto ne nsis. Japanese Journal of Parasitology 3 5 : 3 6 5 -3 6 7 , 1 9 8 6 .

2 1 . Uga S, Ar ak i K, Matsumur a T. De po sitio n and de ve lo pme nt o f e ggs o f

n gi o stro n gylu s c a n to n e n si si n vi tro. Japane se Jo ur nal o f Par asito lo gy 3 : 2 5 7 -2 6 4 , 1 9 8 4 .

2 2 . Wallac e GD, Rosen L. Tec hniques for rec overing and identifying larvae of angiostrongylus cantonensis from molluscs. Malacologia 7 : 4 2 7 -4 3 8 , 1 9 6 9 . 2 3 . Zambrano PZ. Ileoc olitis pseudotumoral eosinofilic a de origem parasitário.

Revista Latinoameric ana de Patologia 1 2 : 4 3 -5 0 , 1 9 7 3 .

2 4 . Zilio tto A, Kunzle JE, Fe r nande s LAR, Pr ate s-Campo s JC, B r itto -Co sta R. Angiostrongilíase: apresentaç ão de um provável c aso. Revista do Instituto de Medic ina Tropic al de São Paulo 1 7 : 3 1 2 -3 1 8 , 1 9 7 5 .

Table 1 - Daily recovery of Angiostrongylus costaricensis eggs per m L from 15 pairs ( m ale/fem ale) of parasites m ain tain ed in Waym ou th m ediu m for 72 hou rs.

Pairs Number of eggs/1 0

µ

1 4 2 0 7 0 8 * *

2 5 1 0 9 0

3 1 4 2 1 0 0 2 * *

4 4 6 0 3 0 * 0

5 2 7 0 4 0

6 4 6 7 1 0 0

7 2 6 0 2 0

8 3 2 0 2 6 0

9 2 3 0 5 0

1 0 2 5 0 8 1 0

1 1 3 0 0 1 4 0

1 2 2 7 0 0 0

1 3 4 5 0 0 0

14 2 3 0 2 4 * * 8 * *

15 2 3 0 34 5 0 * *

* anembryonated * * dark and deformed

Chemic ally defined Waymouth medium, is reportedly the best

c ulture medium for the development of L3 into young adult stage,

with a 6 0 % suc c ess rate

_{. Otherwise, when mature female}

worms removed from the definitive host are kept in the same

c ulture medium, a very intense and extensive egg elimination

takes plac e, suggestive of a stress response.

I n c o n c lus io n , th e s e da ta in dic a te th a t th e

i n vi tro

c o nditio ns te ste d ar e no t a go o d e xpe r im e ntal m o de l fo r

te sting dr ug the r apy fo r ab do minal angio str o ngylo sis.

REFERENCES

1 . Agostini AA, Marc olan AM, Lisot JMC, Lisot JUF. Angiostrongilíase abdominal, estudo anátomo-patológic o de quatro c asos observados no Rio Gr ande do Sul, B r asil. Memó r ias do Instituto Oswaldo Cr uz 7 9 : 4 4 3 -4 4 5 , 1 9 8 4 . 2 . Ayala MAR. Angiostrongiloidíase abdominal: seis c asos observados no Paraná e