FUNGI ASSOCIATED WITH BORRICHIA FRUTESCENS (ASTERACEAE): INSECT GALLS AND ENDOPHYTES

2006·4.5 MB·English

by  Diane Te Strake

WITH
FRUTESCENS
FUNGI ASSOCIATED BORRICHIA
AND
ENDOPHYTES
GALLS
(ASTERACEAE): INSECT
Amy
Haddock Keagy,and
Diane Te Peter D.Stiling
Strake,
Department of Biology
University of South Florida
Tompa, 33620-5200.
Florida U.S.A.
ABSTRACT
DC,
This report compares fungi found in galled and non-galled plants of Borrichia Jrutescens (L.)
&
Asteraceae. Fungi were observed in the plant galls of the midge, Asphondylia borhchiae Rossi
Strong. A variety of endophytic fungi were cultured from the apical meristems, stems and leaves of
galled and non-galled plants collected from several coastal sites in Florida. Fifteen percent of midges
examined in - 60) carried fungal spores of Alternaria sp.
RESUMEN
compara hongos cncontrados en plantas con sin agallas de Borrichia Jrutcscens
Esce articulo los y
&
(LJ DC. Asteraceae. Se observaron hongos en las agallas de Asphondylia horrichiac Rossi Strong.
Se cultivo una variedad de hogo cndof itico de los meristemos apicales, tallos y hojas de plantas con y
sin agallas colectadas en varios lugares costeros de Florida. El quince por ciento dc las agallas
examinadas (n = 60) tenian esporas de Altcrnayia sp.
INTRODUCTION
/
marsh communities Gulf
Asteraceae often found bordering the of the of
salt
Mexico and Atlantic Ocean. Insect galls on this host species are initiated in the
&
meristem by midge, Asphondylia horrichiae Rossi Strong (1990).
apical a
These galls are also impacted by four specific parasitoid wasps iGaleopsomyia
haemon Walker, Rileya cecidomyiae Ashmead, Tenuipetiolus teredon Walker,
and Torymusumhilicatus Gahan). Individuals from these parasites lay their eggs
inside the gall with A. horrichiae. The parasitoids devour the developing midges
and population horrichiae (Rossi 1992).
significantly affect levels of A. et al.
Several biological studies on this plant midge system elaborate on the lite his-
&
mcludmg
tory and ecological details parasitism rates (Rossi Strong 1990;
&
many
As
Stiling et al 1992; Stihng 1994; Rossi Stilmg 1998; Rossi et al. 1999). in
becomes
insect the tissue surrounding the developing larvae eventually
galls,
of chambers layered with fungal growth purportedly providing nutrition
full
knowledge
the developing larvae (Gagne 1989). Thus, of the fungi associ-
for
ated with both the galled and non-galled plants needed to gain insight into
is
host-parasite relationship.
this
r
Many known endophytes which predomi-
harbor microbial
plants are to
tern.
SIDA22(1):755-763.2006
756 BRILORG/SIDA
22(1)
nantly These organisms
arc fungi. live internally in plant tissue. Stone et
al.
endoph
(2000) suggest that these y tes secondarily invade insect galls. Batra and
Lichwardt grown
(1963) nidicate that "gall fungi are airborne, in a variety of
and
substrata are not species specific.^' Other nivestigators of nisect galls sus-
pect that spores of fungi are accidentally collected from leaves and
leaf litter
&
by the insects (Borkent Bissctt 1965), Haridass (1987) suggests that con-
fecal
means
tamniation another whereby
is possible of spore dispersal fungi are in-
troduced into the plant tissue. Gagne (1989) reports that midges feed by suck-
ing on the hyphae and states, "fungi are evidently obligatory in all galls...and
are food of those gall midges." Batra and Lichwardt (1963) alternatively indi-
cate that is difficult to determine whether midges actually feed on the hy-
it
phae
or the plant tissue degraded by the fungi.
we examined number
In this study a of midge galls of Asphondylia
from
borrichiae Borrich iafrutcsccnsirom several Florida the presence
sites for
or absence ol lungi. In addition plant apices, sterns, and leaf samples of galled
and
non-galled plants were cultured for fungi. Finally, a small population of
midges was
surveyed
for associated fungal spores.
MATERIALS AND METHODS
—
Sites for Collections, Borrichiafrutcscens and stem samples were
galls, leaf
m
from
collected the following locations from 1993-1995
Florida:
Site #1-Merritt Island National Wildlife Refuge, Titusville (Brevard
County).
Mayport
Site #2-Little Jetties Park, (Duval County).
Site #3-Ft. DeSoto State Park, Petersburg (Pinellas County)
St.
^4-Upper Tampa
Site Bay Park (Hillsborough County).
#5-Delnor-Wiggins
Site State Park, Naples (Collier County).
—
Culture Protocol. In May, 1995, five samples each of apex, stem, and leaf tissue
were collected from galled and non-galled plants from and
Sites All col-
1, 2, 3.
m
lected plant samples were placed individual,
sterile plastic bags, refrigerated,
and
returned to the laboratory within three days. Excised tissue from apices,
stem and both and
leaf tissue of galled non-galled plants tissues were rinsed in
running water lor 5 min, then each tissue type was placed separate
in flasks
mm
and 95%
rinsed with ethanol for 2 min. This was followed by 30 shake
a
20%
rinse in bleach solution. All tissues were held in flasks of sterile distilled
water (1- 2h) while tissues were prepared incubation. Three from each
for slices
m
tissue from each site were submerged Potato Dextrose Agar (PDA) and V-8
m
Agar
Juice (Difco, Detroit, Ml). These cultures were placed dark incubator
a
32^C PDA
seven from
at for days. Isolates successful cultures placed on
w^ere
slants and held at ambient temperature until sporulation after which identifi-
cations were made.
TE STRAKE ET FUNGI ASSOCIATED WITH BORRICHIA FRUTESCENS 757
AL.,
—
In Siiu Fungal Studies. In September, 1995, ten stems were collected ran-
domly from galled and non-galled plants of B.frutescens from four sites (# 1-
cm
Each stem was cut into segments. Using the protocol established by
4). 1.5
Hignight et (1993) stems were cleared and then all segments were micro-
al.
examined
scopically the presence of endophytes.
(or
—
Whole
Direct Observations of Insects. insects of AspJio?i-:Iy!]a horrichiae
were collected as they emerged from galls of Borrichiajrutesccns, (n - 60), Iva
imhricata Walter (n - 20), and IvaJrutesccnsL. (n = 5) and preserved in vials of
95% ethanol. All msect samples were examined for the presence or absence of
fungal spores on the surface using light microscopy.
AND
RESULTS DISCUSSION
chambers
Previous dissection of the Borrichiafrutescens gdlls revealed larval
Within white mycelium could be observed and fungal
these this
(pers.
obs.).
growth was arrested until the larvae became fully developed. Then the myce-
mat became and
denser, darker, quite distinct (Fig.
lial
1).
Midge galls observed in the Asteraceae, and fungi isolated Irom those galls
Aecidium
are listed in Table In addition, Farr et (1989) note the occurrence of
al.
1.
horrichiae, Puccinia mi and Cercosporidium sp. on stems and leaves of B.
rifica,
many
frutescens. Although plants have fungal endophytes (Clay et 1985; Clay
al.
known
were from
1990; Carroll 1988), prior to this they not B.frutescens.
and
Preliminary studies (1993-1994) including direct observations excis-
ing and culturing of a range of ages of gall tissue from B.frutescens from all five
two
sites consistently yielded a large variety of fungi including types of sterile
made
mycelium (data not shown). Similar observations have been in other stud-
ies of midge galls. Bissett and Borkent (1988) report that some midges from the
Lasiopterini and Asphondylidi inoculate galls with a variety of mitosporic fungi
and use them as a food source. Stone et (2000) observed that galls on Dou-
al.
(Pseudotsuga menziesiO support heavy fungal growth which, he sug-
glas-fir
may Wilson
be plant pathogens in addition to providing insect nutrition.
gests,
may
(1995) proposes that fungi in galls be saprobes or inquilines (organisms
maker
inhabiting insect galls not parasitizing the gall but otherwise utilizing
the gall tissue lor food).
and
To determine endophytic fungi could be observed microscopically
if
method Hignight preliminary study was conducted
using the of et al. (1993), a
in May 1995 using five steims each from galled, non-galled, flowering non-galled
from
and flowering galled plants of B.frutescens collected Site #3, Ft, Desoto.
100%
This mini-study revealed that endophytic fungi were present in of the
example
samples from plants with galls appearing similar to the seen in Figure
The remaining non-galled stems were heavily colonized with iungi. This
less
2.
quantify endophytic fungi
qualitative observation led to additional studies to
758 BRIIORG/SIDA
22(1)
Fig. ].Borrichia frutescens. Cross section through a gall showing the layer of the fungal mycelium.
We
involved in the stems of galled and non-galled plants. observed a consistent
occurrence of endophytic fungi in sampled stem segments from four diverse
all
Florida locations (Sites 1-4). In non-galled plants these occurrences range
all
64%
from 20 and
to in galled plants 30 to 64%. This suggests httle influence of
the role of the midge. But one might argue that the fungal endophytes
in galls
On
grow
out, invading other plant tissue. the other hand, depending on the time-
may
frame, endophytic fungi grow into the gall or exist in the pre-gall tissue
and mycof
contribute to the lora of the aging Either way, fungal endophytes
gall.
m
are extant Bjrutesccns.
and
In the apices, stems, leaves of both galled and non-galled plants from
and among
Sites 3 several fungal taxa occur (Table Included the
1, 2, 2). isolates
among
are several types of sterile mycelium. These were found all tissue types.
was among
Altcrnaria found Acrcmonium
sp. also all tissue types (Table strictum,
2).
759
TE STRAKE ET AL., FUNGI ASSOCIATED WITH BORRICHIA FRUTESCENS
Table Fungi reported In gall midge associations in Asteraceae.
1
.
Reference
Fungi
midge Host
Gall
Sderotium Weis 1982;Batra 1964
Asteromyio carbonifero Solidago canadensis asteris
Macrophoma Borkent&Bissett 1985
modesta mollis sp.
A. tumiflca,A. S.
carbonifero
A.
1964
Rhytisma Batra
Aster asteris
A.carbonifero sp.
1964
Batra
sempervirens bifrons
A.corbonifera R.
S.
1964
Batra
S.lanceolata R.solidaginis
A.carbonifera
Chaetoniium globosum Batra 963
Ratibida columnifera
A.rotibidoe
1
1963
Aureobasidium Batra
R.columnifera
A.ratibidae
pullulans
Plectosporium
R.columnifera
A.ratibidae
tabacinum
(asCephalosporium Batra 1963
ciferrii)
1963
Batra
A.carbonifera S.graminifolia Alternaria sp,
1963
Trichoderma Batra
simulans Helianthussp. viride
Bucculatrix
1963
Batra
simulans Heiianthus sp. Aspergillus sp.
B.
roseum 1963
Trichothecium Batra
Helianthus
simulans sp.
B.
1963
Batra
Heiianthus Penicillium spp.
simulans sp.
B.
J 963
Stemphylium Batra
Helianthus sp.
simulans sp.
B.
1963
simulans Heiianthus sp. Botrytis cinerea Batra
B.
more
irregular distribut^^^^
Bipolarfssp.,andVerticzniumlecaniidisplayed
from sampled. Other species of these genera have been
the
plant tissues sites
&
White
(Bacon 2000).
endophytic fungal hterature
reported in the
abdomen) midges
The mycangia pockets adjoining the of in
(small special
known be with fungal spores that are thought to be in-
wild
the are to filled
examined
newly emerged midges
in
serted with the eggs at oviposition. In the
were
no spores were seen in the mycangia. Spores of Alternaria sp.
this study,
15% midges emerging from Borrichiafrutescens
seen on the surfaces of of the
40%
and
= 10% the midges from Iva imhricata (n-20); of these
galls (n 60); of
/
/
Wh
seems that the diversity of fungi seen in these
fungal spores oviposition,
at
it
midge and host plant tissue are sourced by other means, possibly includ-
galls
some
ing of their parasitoids.
We endophy appear become denser in the plant
observed that fungal tes to
m
May preliminary study) and September revealing
between (seen the
tissues
m
were
hyphae and pigmentation The aging galls this study
increasing (Fig. 2).
became denser and darker with pigments
seen include fungal contents that
to
and both darkly pigmented, were consistently
Alternaria Bipolaris
(Fig. sp. sp.,
1).
760
BRIT.ORG/SIDA
22(1)
Table 2. Percent occurrence of fungi isolated from apical,bteni and leaf tissues isolated from Borrichia
and
frutescens: galled non-galled
plants.
Merritt Island Mayport
Desoto
Ft
non-gall
gall gall non-gall gall non-gall
mycelia
Sterile
Apex -
333
66.7 13.3 40
Stem
20
33,3 20
26.7
Leaf
6.7 40
.7 13.3 6.7
Alternaria
sp.
Apex
13.3
13,3
Stem
6.7
Leaf ?0
6. 33.3 6.7 13.3
Verticillium lecanii
(A.Zimmerm.) Viegas
Apex
Stem
6.7
Leaf
20
20
Bipolans
sp.
Apex
Stem
6.7
Leaf
6.
Acremonium Gams
strlctum W.
Apex
Stem
13 20
Leaf
Traaen
f7u/77/co/a gr/sea
Apex
Stem
6.7
Leaf
Monocillium indicum
S.B.Saksena
Apex
Stem
6,7
Leaf
Peniclllium
sp.
Apex
6
Stem
Leaf
Fusarium
sp
Apex
Stem
6.7
Leaf
STRAKE AL, FUNGI ASSOCIATED WITH BORRICHIA FRUTESCENS 761
TE ET
^
^
f
J
^
fe ^i
.:^
.r*^. .^ .^v"
Hyphae endophytes stem
hQ.I.Borrichiafrutescens. of in pith tissue.
Shaw
reviewed
from and non-galled plant (1992) several
isolated galled tissue.
pigmented
darkly
studies indicating that fungivores consistently favored litter
seems Altcrnaria and Bipolaris could play a
fungi. Interestingly, that sp. sp.
it
system.
nutritional role in this gall
study we establish the presence of entophytic fungi in insect galls.
In this
Endophytic fungi were cultured from tissues of galled and non-galled plants
These
Both these plant types carry several fungal taxa.
Borrichiafrutescens. of
of
common
observations are to all sites, but sample sizes are too limited to discern
exammed
on 15%
major Fungal spores were directly observed of
differences.
may
midges. This suggests that the midges play a role in fungal dispersal to gall
tissue as proposed by Batra and Lichtwardt (1963).
ACKNOWLEDGMENTS
comments and Hennen,
The authors greatly appreciate the suggestions of Joe
anonymous
and one
Erica Cline, reviewer.
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