Mycopathologia 143: 33–46, 1998.

© 1998 Kluwer Academic Publishers. Printed in the Netherlands.

33

Effect of various liquid culture media on morphology, growth, propagule

production, and pathogenic activity to Bemisia argentifolii of the
entomopathogenic Hyphomycete, Paecilomyces fumosoroseus

Claire Vidal1,2 , Jacques Fargues1 , Lawrence A. Lacey2,3 & Mark A. Jackson4

1 Unité
de Recherches en Lutte Biologique, Institut National de la Recherche Agronomique, Campus International
de Baillarguet, 34982 Montferrier-sur-Lez cedex, France; 2 European Biological Control Laboratory, USDA-ARS,
Parc Scientifique II Agropolis, 34397 Montpellier cedex 5, France; 3 Current address: Yakima Agricultural Re-
search Laboratory, USDA-ARS, 5230 Konnowac Pass Road, Wapato, WA 98951, U.S.A.; 4 National Center for
Agricultural Utilization Research, ARS-USDA, Peoria, IL 61604, U.S.A.

Received 10 April 1998; accepted in revised form 3 November 1998

Abstract
The entomopathogenic Hyphomycete Paecilomyces fumosoroseus was grown in five different liquid media, which
have been developed for mass production of Beauveria spp. or P. fumosoroseus. Production was followed for 96 h
by measuring both biomass and concentration of propagules. Maximal biomass was obtained with two media,
Jackson and Catroux media (40–60 mg ml−1 suspension produced after 42 h incubation), where the exponential
phase of growth began earlier than in the other media. While high concentrations of propagules (1.4–5.5 × 108
propagules ml−1 ) were produced in three media (Jackson, Paris, and Catroux media) after 48–72 h incubation,
production of propagules was lower in the two other media, containing maltose as carbon source (Goral and
Kondryatiev media) with 0.4–3.7 × 107 propagules ml−1 after 96 h incubation. P. fumosoroseus produced oblong
blastospores in the three most productive media and conidiospore-like (ovoid to subspherical) propagules in the two
other media. Infection potential of produced propagules was tested on the silverleaf whitefly (Bemisia argentifolii).
Whiteflies were sprayed as 2nd instars with P. fumosoroseus propagules produced in the five liquid media (1.9 ×
103 and 1.9 × 104 propagules cm−2 ). All the media produced propagules that were infectious for larvae. With the
lower dosage, mortality rates were significantly lower with propagules produced in one of the two least productive
media (57%) (in the Kondryatiev medium) compared to those obtained with the three most productive media
(>90%). However, when whiteflies were treated with the higher dosage, mortality rates (91–99%) between media
were not significantly different.
Key words: Bemisia argentifolii; liquid media; mass production; microbial control; Paecilomyces fumosoroseus;
propagule morphology

Introduction mosoroseus (Wize) Brown & Smith [2], which is one

The silverleaf whitefly, Bemisia argentifolii Bellows
& Perring (Homoptera: Aleyrodidae), has been among
the most serious insect pests in field and green-
house crops [1]. The rapid increase of resistance
to chemical pesticides in the whitefly and the nega-
tive impact of pesticides on its natural enemies have
prompted research into alternative methods of con-
trol. These include the use of biopesticides based on
the entomopathogenic Hyphomycete Paecilomyces fu-
of the most promising fungal species being studied for
control of B. argentifolii [2–5].
For successful use of a mycoinsecticide in bio-
control, it must be possible to produce high yields
of propagules which are pathogenic towards target in-
sects. Thus, quality as well as quantity of propagules
produced in liquid culture need to be examined with
equal attention, because both parameters can be af-
fected by medium composition and culture conditions.
34

When grown on solid media, Hyphomycetes pro- Table 1. Composition of the five media (g l−1 distilled water) of
shake-flask cultures in agitated liquid cultures of Paecilomyces
duce differentiated asexual spores, which are genet- fumosoroseus strain 92117.
ically stable [6]. In submerged culture, they usually
form single cells by schizolytic separation at septa or Ingredient Catroux Goral Jackson Kondryatiev Paris
by mechanical fragmentation of hyphae or can also be
KH2 PO4 6.8 2 2 2 0.36
produced from hyphae by yeast-like budding [7]. The
MgSO4 7H2 O 0.1 2 0.3 2 0.60
term ‘blastospore’ has been widely used in the litera-
Na2 HPO4 12H2 O – – – – 1.42
ture [8] to describe this cell, although other authors, FeSO4 7H2 O – – 0.05 – –
considering it functionally identical to short hyphal MnSO4 H2 O – – 0.0156 – –
cells, call it a ‘hyphal body’, in reference to its mode ZnSO4 7H2 O – – 0.014 – –
of formation [7]. Even though blastospores are usually CoCl2 6H2 O – – 0.0366 – –
the only type of propagule produced in submerged cul- KCl – – – – 1.00
ture, several studies reported the formation of conidia CaCO3 2 – – – –
[9–12]. Because of the lack of ultrastructural inves- CaCl2 – – 0.4 10 –
tigations, propagules formed in submerged culture KNO3 5 – – – –
which were microscopically identical to aerial coni- NaNO3 – 5 – 5 –
dia have been called ‘conidiospore-like’. Although NH4 NO3 – – – – 0.70
various entomopathogenic fungi are reported to pro- Casamino acids – – 13.2 – –
duce high concentrations of blastospores [13, 14], this Saccharose 30 – – – –
type of propagule is usually less resistant than aer- Glucose – – 80 – 10
ial conidia to conditions which are unfavorable, such Maltose – 20 – 20 –
as drying [15]. Production and pathogenic activity of Corn steep liquor 20 9 – – –
Yeast extract – – – – 5
blastospores have been largely studied in different Hy-
Vitamin mixturea – – 3.15 – –
phomycetes [7, 8, 16–22], including P. fumosoroseus
Chloramphenicol 0.50 0.50 0.50 0.50 0.50
[14, 23–25].
This paper describes the influence of nutrition in a Vitamin mixture: thiamin, riboflavin, pantothenate, niacin,

liquid cultures of P. fumosoroseus. This compara-
tive study was carried out with five different media,
consisting of three that have been patented as mass-
production media of Beauveria spp. [9, 17, 26], one
medium for liquid production of P. fumosoroseus [14],
and one reference medium [27]. We evaluated the ef-
fect of medium on both concentrations of propagules
and biomass and on the type of propagules produced
(i.e. blastospores or conidiospore-like), and their mor-
phological and pathogenic characteristics. Four of
the five media tested had not been studied for P.
fumosoroseus; thus this paper reports new elements
about the production of this fungal species in liquid
culture.
Materials and methods
Inoculum preparation and cultural conditions
The fungal isolate, P. fumosoroseus 92117 (Euro-
pean Biological Control Laboratory Fungal Collec-
tion, USDA, Montpellier, France), which had been
previously isolated from a cadaver of Bemisia tabaci
(Gennadius) in Multan, Pakistan in 1992, and selected
pyridoxamine, thiotic acid, 0.5 mg l−1 each; folic acid, biotin,
vitamin B12, 0.05 mg l−1 each.
for its pathogenic activity towards B. argentifolii [5]
was used. After host passaging through B. argen-
tifolii, conidia of the monosporal fungal strain were
harvested from the surface of cadavers and then plated
on ‘Paris’ nutrient agar (in g l−1 : 0.39 KH2 PO4 ; 1.42
Na2 HPO4 12H2 O; 0.60 MgSO4 7H2 O; 1.00 KCl; 0.70
NH4 NO3 ; 10 glucose; 5 yeast extract; 20 agar, supple-
mented with 0.5 chloramphenicol) [27] and incubated
for 14 d at 25 ◦ C.
After a second subculture onto Paris medium, a
stock suspension of the fungal isolate was made by
scraping conidia directly from the surface of 3-week-
old sporulating cultures. Spores were then suspended
in 10-ml sterile distilled water by shaking in 45-ml
flasks containing 5 to 6 dozen glass beads (3 mm
diam). No surfactant was added to the conidial sus-
pension, since 5 min of agitation at 700 oscillations per
min (10 cm vertical travel) on a mechanical shaker was
found to produce homogeneous suspensions of viable
single conidia. Conidial densities were determined
using a 10−2 dilution of stock and a hemacytometer.