6tr ?Za

Adult identification in

SlOutlq4ql apparent

dependence

of polyteny

on food ^gouree

ln ^.olplghian

tubuLe and nurse ceLL

nuclei duri:lg

oogeneale.

Flnal

Report

l{lLliam S.

^Procunier

W.H.O. COIJABORATING CENru

f6r the studf ol

SIMULIIDAE & PHLEBOTOMINAB rn relation to

ONCHOCERCIASIS & LEISHMANIASIS

Department

of

Entomology

Briti.sh

l'luseun

(ttaturat Hletory),

Londou, Unlted K5rtgdom, SU| lBD

*^-rerary _adviser to the

^OCP/OMS Program, Ouagadougou Burkino Faso, l{ay ^1985.

,.sffir ^oT7

t4'

; ?Za

6tr

:.?".q',--;}

1.

This

study vas undertaken

to

determile

chromosone poJ.ytenizatioa and

to

^asoees tbo

parity

determinations..in

S.

damnosum

s.l.

the

opti.mum

tloe 36j

na.lplgblau

pbysiological

age

of the fllcs for

fhe

optinurn

time for

malpighlan tubul.e

"h"oro.ooe

polytenization

_{appeana to} occur

after a

bloodneal

for the

ti.me

period the

^oocarte eaters

aeiosls

up

to

rnd

iacluding the

karyosome stage

(I'igures IB-LD).

^The nucl,el (karyoeome) _becoaoa condensed and rernains

at this

etage

until fertillsed,

^uhereupon

the let

^and 8nd

mei.oti.c

divisions are

cornpletod and

the

spern fuses

nith the

functioaa-1 oocytr

to

form

a zygote,

The malpighian nuclej- appearr

to

break down

at the

onset

of vitellogenesis (laying

dowu

of yolk) ^(Figure IE).

Onset

of

meioete

varlee

anong

species and

especially

between aevann4$

anrl*forest llploe:

^{optimum tLne}

- forest I-4 hrs; savarurah

T-11 hrs.

Previous

to this

study

it

was suggestod

that

chromosome

polytenlzatlon

appeared

to

be dependent on food Bource (Procunier _{1984 and} Procunler & poat L985)

with the

degree

of

polyteny being stage

specific.

^I{ence

by the

tLme

adult flice

obtained

a

bLood meal moet ehould already be

at

stage

II

(Procunler ₁₉₈₄₎ polytcny.

Most

tissues

i.n which

high

metabollc

turnover is

required

duriag

ontogenetle

developnent appear

to

respoad

to the

demand

either

through

eelectlve anpllflcatlon of

genes and,/or

the

whoLe genone (Ashburner _198O).

In the

case

of the

whole gonome (chromosome conplenent)

thls

^{oay ta]<e} the form

of

^polyteny

or

endoduplicatloa (doubllng

of

cbromosome

number).

Siace ma-lpighian

tubules function

as an excretory organ which

j.lvolves

_water ba.Lance

(Wiggleswortb 1984)

great

demands would be

put

on

tbis

organ upon

acqulriag r

bl-oodrneal.

Further, the time to

synthesise DNA from

a

normal 2c value

to

2ol4o

value (tutt _polyteny)

_would

_also requlre a certain length of tine.

Consequently

the

observation that,

the flias are

already

at

stage

II

pollrEeay

is consistent ultb

tbe

tj.rne

constrainls put

on

the flles

syetem j-n

order to

^{oalce ready}

the

nucl"car oachiaery

for high

metabolic

turnover in

response

to the

bloodrneal.

Eovever,

littLe ig

^{known about ceIJ,}

^cycle in blackfliee ulth

regard to

DNA synthesj.s ti.me (except _{Bedo 1984}

-

^does

^not glve actual length of tlror ln his

^{paper) a.rrd} hov

long the celIs are in mitotic (melotic) divlslon.

s

G2 M

G1

DNA synthesis 'Gap

2

mltosts

^(meiosis)

Gap 1

) ({,

^o.;;

tq6L )

In

DrosophiLa (Kfng I97O)

the time required for

meiosis up

to

karyosone etagc

ts

27

hrs with the total time for

egg maturation

belag 4 days. In

noequitoea

this detailed

j-nformation

is not

avaiLable coucerning

aIL _the

atages

of

oogcacsla except

the half gravid (yolk ])

etage can occur 12

hre aftar a bloodneal.

Boncver,

i:r

mosquitoes readable chromosomes

are

frclm

the

nurse

ce1ls.

These organlaua provi.de some

clues

concerning

duration of

temporal events duri.:rg,^oogenesla.

ft

^J.s

important to establish that a particular

r,:t;rge can be used

consistantly

aa

a'.

physiological

_marker

of age. Cytologica]

observationsfrom a nunber

of differcnt species, !.

^yahenser

g. ryttpqu.].i,9.

^glggg4ggr

[.

^soubfense B (Procunier & Poet

1985),

S.

oyapockensy'roralmense anrl

3.

Iimbatum (SheHey, Moraes, Luna

Dtar

&

Procunier 1985) suggests

that a relative

tlrue

perlod exists for

coneistenbly

obtaiaing

readable malpighian

tubule

chromogomes. Thle concLusion

is firthcr

supported by

the

observation

that individaals

^{which show} dj-fferentia-L

folllcular

development

(Fig.

2C aad 2D) remaln

at

stage

I11 polytenizatioa

independent

of tine after a

^{bLoodmeal (a6}

long as

aome

follicles are at the right

^stage)

rhtle

those

individuals uhich

sbow gonotrophic concordance cannot be read

after

the ooclrte

enters vitellogenesis.

Tiaing of

onset

of meiosis, polfienlzation

and cluration

of

oogenesla La undcr hormonal

control.

^{These processes seem}

^t()

^{be dependeut on}

^the

production

of

R'ddt$"1 o',,1 Je'tata

llll

ecdysone and

its derivatives

and,/or

juvenile

hormone ⁽

, '.). _lcccrtly it

^{has been} shown

that

Juveni.Le ^hormone can induce

polyploldization la folllclo cerls

^(Lapoi

_nte et aI

^{I 9g5}

)

^{duri ng} vi tet logeriesi

! ^(r'igie

^{28 )}

6.

,.

The seguence

of

hormonally medlated evonts

during

oogenesis appear

to

^bc

a function of

^marry

factors -

^{eogo blood} motabolism, excretory

regulatioa tto.o

tr'ormation

of the peritrophic

merobrane occurs

very

rapidLy

after

acquirenent

of

^a

bloodnea1

in

many

blackfliee vith the laylag

doun

of

nenbrnnes

varyiag ln ttnc

Reirt ₊ LeLare'

and

type

⁽

1981).

^Formatlon

of the

^nembrane has been looked

at ulth rcgard

to understanding

factors Il-triting the traasolssion.of

onchocerca

(PUf[ppon

_X977 ancl

R"''<J*lqhcrt-

r 198f).

^However, formation

of the peritrophic

meobrane tray arso bc

a

x.l,nttlng

factor in

deterrniniag

the

onset

of

meiosis and srlbeeouent

vtteLlogerresLe. It ts interesting to note that

my

results for

^onset

of

meiosis ^(optinum

tlne for

chromosome

polytenization) correlate poeitively with the ti:ne

taken

for fornatlon of

membranes. Savannah species which have

a thick

membrana

require 6

Ura?

to lay

down

while forest

species

(again the idea of d,lscrealttr" periods for

^oarktng

developmental events) which have

a thin

^membrane

requlre only I tlr. ^(PnUlppo\

:lg??).

Further, differences in saturation

(oogenesis) tlmes between species ui].J.

certainly affect vector

dynanicswith respect

to transmission.

Paraelteg _havc evolved

a

^growth

time to L,

staSe which

correlate with

egg development

so

that gonotrophic concordance

will

ensure

paraeite survlval

because

it is infectlvc at the Jrd

bloodmea-l. Ti.me

for

oogenesis would provlde

a strong isolating

nrclranlco

for

parasi*'e

differentiation

which could aJlow

further selective

procesaea

to

account

for differences

between savpnnah and Fores.

:;;Ilr:;^tfiXi.:'itlii*',i"*u

be

that

species found

interfacing the

two

ecologlcal

zones would

exhibit ainilar

maturation tlmes and hence could

allow

developnent

of

botb rrstrainsrr

of paraeite.

A

series of

photomicrographs

depictlag _the

events

of

oogenesis

is

^gtvea

with

appropriate notes discussing

parlty

and gonotrophlc concordaace (Flguree

IA-LI

^and

2A-2G).

Photomicrographs

of tha adult

poLytene chromosome complenent

of 3

cpeclco

is also given. It

should be noted

that

lndividual"s whicb

give

mediocre prtparattoas

(marginal)

can be analyzed

with

appropriale photomicroscoplc

set-up

and hrvLng

preknowledge

of larval

banding aequences.

5

i-udividuals

of g.

sirbanum wcrc alao

positively identified;

however, photomicrographs wore

not

takon because alldee uere made

at

^Bamako.

Research Recorunendations

The f ol l owi ng stud'ies

are

envi saged:

l) To

determine

if malipighian tubule nuclei will give consistently

readablo chromosomes so as

to

be used

for routine adult identification

nov{

that the

optlmum

time for

chromosome

polytenization has been

detenfitlned

for 3 spccles ln !.

damnosum s. I .

2) To

determine

if a techno'logytransfer in adult ident'ification techniques'ls possible using

^DNA probesand malpigh'ian

tubule

chromosomes

from the

^same

fly.

Under

(l) study sites

and

protocol for adult identificat'ion as outlined'in

^the

previous advisorship (Procun'ier .l984) can be followed.

Under

12) as'in (l)

_except

_the fly

would

be

severed

in

^two

- with the

abdom{nal

part

being used

for

chromosome'identification and

the other half for

^DNA

analyrls.

Dr Post has 3 ^probes

now wh'ich

can differentiate 3 species pairs;

pS03

:

sancti ^paul i /soubrense

B,

^pS01

'l -

squamor,uT/yahense

affi

^pSQl

-

^si rbanug ⁽ Post ^'1985

-

^Penn

State). fhis

techn'ique

allows

populations

to

be separated by

quantitativa

means

based on the

amount

of

^{bi nd'i}

ng

⁽ radi oact'i vi ty

) the ^probe

^{shows when}

anneaiing

to the

^DNA

of a particular species.

However,

the

technique needs

to

be

verified

against

a

standard

to

ensure

the quantitat'ive difference don't overltp especially for siblings in sympathy. If quantitative difference hold

up

the

DNA

method would have tremendous pract'ica1

value not

on'ly

for identifying the

vector

but also to

^determine

relative vector potential (could

probably process 200-500 f1 i es/day) .

One

of the study sites

^should

chromosomes

preparations

were

approximate

time to look for

the

be at Tienfala, Mali

^because

^(l ) identifiable

obtai

ned

^f

rom there (S

^'indi vi dual s )

; ⁽²¹

^the

optimum.chromosome

pglytenization in S.

sirbanum

'i

s ^known;

⁽³

) 0r ^Post

^al

^{ready has a}

^probe

Consquently,

th'is population gives the

^greatest

'i

n

i denti f i cati on techni ^{ques .}

which can identify

S. s'irbanum.

prom'ise

for

a technology

transfer

A

second

s'ite

^coul

d ^be

Soubre where ^al though ^I

arval

^popu'l ati

ons of

_-S. ^yaheEe

and

sanctipauli are in allopatry the sites are close

enough

for the adults to be

considered

'in _sympatry.

_Again

_Dr

_{Post has} probes which can separate ^yahense

from s.anctipauli. Alternatively,

^some

other site

^where

populations occur ln

sympatry,

especially

damnosum

s.s, ^(Neither

one

of us has

looked

at

damnosum.)

Another

possibi'lity is to

^{go where}

S.

^s'irbanum

is zoophilic in its distribution.

3) A s'imilar study could

be undertaken

w'lth Dr

^Angela

Philips. I st'lll

^{do not}

know whether

they

have done

the critical analysis to show'if

^peak dlfferences between

2 populations (single adult) are additive - this is

^necessary

to

differentiate between intra and interspec'ific variatjon. i.e.

suppose the

folIowing

G.C.

profiIes

were obtained:

I adult

^{(sirbanum) 3mm}

I adult

^(yahense)

does A

+

B

give _a

peak height

4)

^Pari

ty

^determi

nat'ions

shoul

d ^be

^{f easi b'le}

(.l984).

_However,

an in-depth study

^{should be}

follicular relicts

are most

easily discernible'

6mm A

-/

^UM

4.5mm

A^^

using the

techniques

of

Procunler done

to verify the

stage

at

^whiqh

References

Ashburner, M.

1980.

Some aspects

of the structure

and

function of

^polytene.

chromosomes

of the Diptera.

^Symp.

^R.

Entomol. Soc. London.

I0.

Blackwell

Scientific Publicat'ions,

Oxford, 20pp.

Detinova,

T.S. 1962.

Age grouping methods

in

Diptera

of

^medical

importance.

t.lHO

monograph

series

No.

47.

Geneva, Swrueer'lBnd. 216pp.

Bedo, D.

G.

^I

982.

Patterns

of

polytene-chrornosome rep'l _i cati on

(Diptera:

_{Simul i idae)}

.

^Gentica

^59:

^{9-2.l .}

Cupp, E.I.l. & Colf

ins,

R.C.

.l979.

The gonotroph'ic

cycie

in Ann.J.

Trop.

Med. Hyg.

28:

422-426.

'ln

_{Simu I I um}

ornaflf$.

Simulium ochraceum.

and parous

rates of

anthropophi'lic transmission

of

^0nchocerca

Simul I um damnogum

Garms,

R. 1975.

0bservat'ions on

filarial ^jnfections

blackflies in

Guatemala,

with

reference

to

^the

volvulus. frop.

med.

Parasit. ?6:

^.I69-182.

King,

R.C. ^.l970. 0varian development

in Qrolophill lg&lfggasler.

Academlc Press

Inc.

^(London)

Ltd.,

Berkeley Squaare House,

London.

^2?7pp.

Lapointe, M.C. Koeppe,

J.K.

&

Nair, K.K.

^.l985.

'Fo'llicle _cel'l

polyp'loldy in

Leucophaea maderae:

regulation

by

juvenile

_hormone.

J. Insect. Physiol. 3l(2):

I 87-t 94.

Lewis,

D.J. 1957.

Aspects

of the structure, biology

and study

of

Ann.

trop.

Med.

Parasit. 5l:

340-358.

Madahar, D.P. 1967

"

Cytologica'l

studies

on

blackf'l'ies (Simuliidae: Diptera)

^Ph.D.

thesis, University of

Toronto, Toronto,

Ontario,

Canada.

-2-

Phil'ipp0n, B. 1977.

Etude de

la

transmission d'0ncherca

volvulus

^(Leuckart 1983)

( Nematoda,

Simuliidae) (oRsroM).

0nchocercidae)

par

Simulium damnosum Theobald, ^'1903 (Dlptera:

en

Afrique trop'icale. Trav.

Docum. Off

.

Rech.

Sc'i.

Tech.

53:

l-308

Procunier, l,l.S. 1984. In

Onchocerca/Simul'lum

relat'ionships in

regard

to

the transmi ss'ion

of

^human onchocerc'iasis

in

West

Ririca.

^l{HO/Fi 1 StlG document.

(

In

^{press ) .}

Procunier,

I.l.S.

.l975.

A

cytological

study

of

^two close-ly

related blacltfly

specles:

Cnephia dacotensis and Cnephia

or.nithophilia (Diptera: Simuliidae).

^Can

,

^,1.

Zool. 53:

I 622-1637.

Procunier,

W.S.

(D'i ptera ) ^:

and nurse

Rejd,

G.D.F. ^&

simu'lijds,

mi rgrat'ion

&

Post, R.J. ^'1985.

Adu'lt

identiflcation of

simulium damnosum

s.l.

apparent dependence on polyteny on

food

source

in

malpigh'ian tubule

cell

^nucle'i

during oogenesis.

Tropen. med.

Paraslt.

(subm.itted)

Lehane,

M.J. ^'1983. Peritrophic

membrane

formation'in three

temperate Simulium ornatum,

S.

equjmum and

S. lineatana, with

respect

to

the

of

onchocercal

microfilariae.

Ann.

Trop.

Med.

Parasit. 78(5):

527-539.

Riddiford,

L.M. & Truman, J.W.

1978.

Btodhemi$try

of rnsect

^hormones and Insect growth

regu'lations. In

Biochemistry

of Insects.

^Academic Press

Inc.

l'11

Fifth

Avenue, New

York,

New

York'10003.

Pp.308-355.

shell€y, A.J.,

Luna

Dias, A.P.A.,

Moraes, M.A.P. & Procunler,

l.l.S.1985.

The status

of

Simulium ^'---- oyapoekense _{-J -r} s.'l ./rora'imense and Simulium limbatum

of the

S.

amazonicum group as

vectors of

^human onchocerc'ias'is

ln

^lowland

tropical forest land

savannah areas

of Brazil. Bull. Ent,

Res. (submitted)

l,{igglesworth,

V.B. .l984. _Insect physiology. 8th edn.

^{Chapman &}

Hall,

733 Thlrd Avenue, New

York,

New York

.l00.l7. _lglpp.

0ogelesis

ln

^Simulium

Prev'ious

attempts to elucidate the events of

oogenes'ls

in blackflies has

boen

part'ia11y restricted by available

techniques

applied ln ^preparing

and

stainlng material

and

the type of

photomicroscopy

used'in

doEumenting

this process.

^(Lewls

1958,

Madahar

1967,

Garms

.l975,

Cupp and

Collins T$fgt. However.,'tillzatlon

of different fixation

and

staining

methods coupled

with the

use

of

interferenca microscopy has pnovided

the

means

for

understanding ovarian development more

fully.

Oogenesis

in adult Slnuliun

daranoaurn ^{; i,r.r}

i.I.

^aadf€'. roraltrcDse

Figure IA.

Magnification

XIOO.

NeuJ.y emorged

S.

yahense

adult:

a-geraarlunl

b -

^proxi.aal

follicJ.er c -

^distaJ.

folllcle.

Nota nuree

cells arc

uou

polyteni-zed (etage

I - ^arrov f).

Oocyte

is aot

dlsti-aguishable

at thia

stage.

rf

igure IB. fiagnification XIOO. 9. gSlgllgli tr.s.

^{Tuo bours}

after

bloodoeal.

Nurse

cells polytenized (stage II-III).

Oocyte

le apparent.

Nucleus

at

pa.chtrrtene stage

of

^meioels

(arrorl c).

figure lC. Magnificatioa XIOO. 9.

sa-nctipiruli

n.s. tlo

hours

after

bloodncal.

Precocious condensation

of

oocyte 'uucleua begindfifg

(arror. e).

Nurse celLa

are at

stage

III.

Fi.gure

LD.

Magnification

Xl@. E.

sanc-tipauli

n.s.

Four hours

aftar

blootlncel.

Oocyte shows condensed nuclear

material in the

form

of a

karyoooe

(arrou

_c)'.

Yolk

forrnation (arrow

e).

ViteJ"logeneeis appears

to

^be

starting.

Figure LE.

Magnification

XIOO. g.

EBBS!&gg}!

tr.s. Elght

hours

after

^bloodneal.

YoIk forrnation *.

^{Karyosome preeent}

^(arrow).

^Nurse

^cells

po).lrtenized

(stage III).

FolLicuLar epi-h,eL1a1 ceJ-ls appear

also to

be

partially

polytealzed

(arrov f).

Cel1 volume increasing.

figure LF.

M,agnification

Xj@.

_5..

eangtiq+uIi a.q Trelve

houre

after

bloodnoal.

{)

^yoJJx

formation.

^Karyosome

stl}I vlslble.

Nurse

ce}ls EtilL

stage

III

polyteny. r,ollicular

epitheU.un

partially

polytenized

(arrou g). CelI

volnoc increasing.

Fi.gure

lG. Magnification

X2OO.

S. ^yahense.

22 hours

after

^bloodmeal.

4/J

yo].k

forrnatlon.

^{Ce11 volu.ne}

lacreased.

Nurse ceJ.I etage

I.

Iuterfcrence microscopy shovs

yolk readily.

^Karyosome

^etiLL vialble.

Yolk doee not

cover oocyte.

Figr:re 18.

!{agnificati.on

X1@. g. Iglry.

^{24 hours}

^after

bLoodroeal.

Oocyte

sti11 vj.sibIe.

^{YoLk covers}

^nost of

egg.

Nurse cellspushed

to

one

end.

^{Karyosone appoara}

^to

^{be brokea doUn.}

FiSure

)-I. Magnification XI)o. E. rorairr.nse'

^{48 hours}

after bloodreal' Interference microscopy.

Chorion formatj.on around

egg.

^{Gernarluo and}

proximal,

foJ-licle

attached

outside

cborlon

(a, b).

Proximal

fol1icLe

appcara

to

be

at a sinil.ar

stage

as that

described

for Fig. IB.

^Nuroe ceLls aro

polytenized (arotr f).

Paroue females may

require less tj.ne to

nature tlJlce

proximal folLicle partial-ly

developed.

Figure IJ. Magnification X150. E.

Loraimonse. ^{48 hours}

after

bloodneal'.

Interference microscopy.

Chorlon present

uith

ce1ls eloughed

off

^(arrow

n).

Probably nurse

cells.

^Note

also

probable presencs

of vitellln

^membranc

(partiaJ.ly

_broken)

with yolk

underneath (amow

v).

w bc

B

Oogeuesis, Gonotrophlc Concordance and

parity

Figure

2A. Magnification XIOO. g. g!g,!gg.

Sour hours

aftar

bloodneaL.

Note

the

pre6ence

of 2 darkly

etalned heteropycnotlc ^bodj.ee

ln

each diataL

fo]-licle ^(arro,rs e)

i-naicati-ng

that ²

karyosbmea

are

present

per fol).lclc.

th.is is

^u:r-like

that

^seen

in

DrosophlLa, Toetse and mosquitoee, and hae

aot

^bcc!

(PrccuxiC." ^{lq? 5 )}

reported

before in blackfli""A ^a -

Sermariun,

b -

^proximal

follicle' c - distal ^foLlic1e.

Figure

28. {agnification X!@.

_,8. 933Siip39,ti

Ir.8. Zfi yolk fornatloa

sbowlng

polytenized foll,icular

epithella-L

nuclei

^(arrow

g)

covertng

the

oocyte.

Nurse

ce1ls stilL partially visible.

Fol.Ilc1e celLs

are

^probably

active in

synthesis

of viteLlin

(membrans).

Tlris

stage corrgsPonds

to

stage

III of Christopher for

mosquitoes and stage ]OA

of !ryIlilg.

Figure

2C. Magnification )€oo, 9. su*Sgli n.s.

^{T'relve hours}

Interference microscopy. Differential follicular

development

ig to

^presence

of rod

and

circular like

^{fun51 (arrow}

c). ^Fo1licles

of

development from uewly ^emerged

(arow p) to 4/)

^yu.Lk forrnation (arrow

5).

Figure

2D. Magnification

^X2OO.

l. S9!3:!9Eli !.s.

^{Same as}

^figure

^{2C excapt}

differential follicular

development probably under genic

control.

^Not

all' indj.viduals

^show gonotrophic concordance

(wilI affect vector

dynanics).

rUrtL fetor4 io tnqr.,3rr.rrrort

Parlty

Figure

2E. Magnificatioa

^X2OO.

9. IgIgE.

Four houre

after

^bloodmeal.

Relict

egg

(i)

and d.eveloping oocytes

(k).

after

bloodlncal.

probably duo '6hou _raJxgc

Figr-rre

2F.

Magnification

X5@. _,8.

eoubrense

of a stalk ^(pedicle - ^{arrou d)}

^{attached to}

a - gernariun, b -

^{proxinaS. foJ.licLe.}

B.

^Non

blood-fed.

Note prceencc

distal folIicIe ^(c);

One bour

after

blood.rneal. Probable

pro:rMhl folJ.icle, es-

dLstal

Figure 2G.

Magnification

fo].li.cular relict

^(d) ;

foLIicle.

X2OO.

S.

^yabense.

a-geraaf"luorb-

#.0

r.*

{i,

^I

r's l-

,f

"*$rh

I p

/

';,k**

b

Adult

Si-ouliun

sanctipaull

tr.s.

Magnification

XIOOO.

Diagnostic iaversions

i.nclude

I L

P+Q

,lrd II-!;!Q!.

At tbe

^Sassandra

R.

^(Soubre)

IT L 7 is polymorphic.

Tb.is

ladlvldual

shora

the II L-7 inverted

sequence

(the p.b. oits distaL

and

close to the

cnd

of

chromosome) nhereas

in the j:rdlvidual in the next flgure

shoue

fi

L-7 sequence as standard (tUe

p.b. sits

more

proxinal).

IS IL IIs IIt

urs

III

^L

short

arm

of

chronosome one

loag aro of

chronosome one

sbort

arm

of

chromosome one

long

a:m

of

chromosome one

short

arm

of

chromosome ono

long arn of

chromosome one

C

-

centromerer RB

- ring of ^Balblani,

^db

-

^{double bubbLe,}

pb

-

^para

^Balbiani,

^NO

- nucleolar

^organizer.

l-l

vf

frl

19/

l-i

^

o.o

J

FraF{

Fr

r/

/.x,

r i"{3

",ff

c _{hit ,t}

o,", 'L

_/t

tl _{zt rn}

-t H H

i,wE ^er\

--l*1rilr^'r-&:

J ''sl."q3-- Uf.

oro

H _{-o llcr\55\-}

= 2'

^L,

Adult

Sirnulium sanctipauLi

t1.8.

^ard,

E. 13!g

Maguification

^XIOOO.

A, C,

D show chromosome conplement

of S. sancttbault n.s.

which is*eharacterlzed

by II t -

^{4.6A being}

fixed.

^Note

II L-l

sequence

ie

standard (conpare

to other figure).

B

-

cbromosome

I of S.

Yahense.

yahense

is fixed for the

heavy Uana

(H) in the

centromere

region, unlike that for i. gg*g!i!3gE n.s.

^compare banding betneen A ^and

B.

Preknowledge

of diagnostic

^sequences

greatly facilitate ^analysis of

mediocre preparattone

especially in

areas

of

^{known cytotypeo'}

$na;=

. irl'ilb

Sl*

IL

-e',{^

"r(S

k *.b

ttu

NO

"61

lr"

',.;,.,:{i

-,. ,ii

,i

F

l

'il1l

._j il

:\ i,1

c

D

6r

t

lrw