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Letter
Chytrid Fungus in Europe
Trenton W.J. Garner,*
Susan Walker,*† Jaime Bosch,‡ Alex D. Hyatt,§ Andrew A. Cunningham,* and
Matthew C. Fisher†
*Zoological Society of London, London, United Kingdom; †Imperial College,
London, United Kingdom; ‡Museo Nacional de Ciencias Naturales, Madrid,
Spain; and §Commonwealth Scientific and Industrial Research Organisation,
Geelong, Victoria, Australia
Suggested
citation for this article
To the Editor: Amphibian species are declining at an alarming
rate on a global scale (1). One of the major reasons
for these declines is chytridiomycosis, caused by the chytridiomycete
fungus, Batrachochytrium dendrobatidis (1,2).
This pathogen of amphibians has recently emerged globally (2,3)
and has caused mass die-offs and extensive species declines on 4 continents
(1,3); knowledge of its distribution and effects on amphibian
populations remains poor. In Europe, little is known about B. dendrobatidis
distribution, which is disturbing when one considers that at least 3 European
amphibian species are undergoing chytrid-associated die-offs that will
likely lead to local extinction (4,5) (J. Bosch et al.,
unpub. data).
We screened 1,664 current and archived samples of wild amphibians collected
in Europe from 1994 to 2004 by researchers using amphibians as study organisms.
B. dendrobatidis infects the skin of adult amphibians and the mouthparts
of anuran larvae; samples included toe clippings and skin samples from
adults and mouthparts of tadpoles. Our sampling was opportunistic, including
both caudates and anurans. We screened all samples for chytrid fungus
with quantitative real-time polymerase chain reaction (PCR) of the ITS-1/5.8S
ribosomal DNA region of B. dendrobatidis (6),
including appropriate positive and negative controls. We confirmed real-time
PCR positives by amplifying a subset of these positives with a second
B. dendrobatidis–specific PCR with a nested reaction developed
from the ctsyn1 locus (3). To confirm that detection
with real-time PCR indicated a viable chytrid infection, when actual tissue
samples were available, we examined a generous subset using histologic
features for typical signals of pathogenic B. dendrobatidis infection.
Specifically, we found intracellular zoospore-carrying sporangia within
the stratum corneum and stratum granulosum of toe and skin samples. We
also compared real-time PCR amplification profiles of suspected positives
to those generated from samples from animals involved in chytrid-driven
die-offs and found these results to be comparable. Furthermore, attempts
to isolate the fungus from dead animals were successful when animals were
obtained in a suitable condition for this purpose (see below).
Our survey found B. dendrobatidis in amphibians in 5 European
countries, Spain, Portugal, Italy, Switzerland, and Great Britain. Previously,
chytrid infection has been reported in wild amphibians only in Spain,
Germany, and Italy (4,5,7,8). We detected chytrid fungus
in 20 of 28 amphibian species examined, representing 9 different genera,
5 anuran, and 4 caudate, in 6 families. We found signs of chytrid in archived
samples from as early as 1998. The number of infections per country we
found were Austria 0/24, Croatia 0/8, Czech Republic 0/18, Italy 2/101,
France 0/60, Germany 0/51, Greece 0/88, Portugal 1/25, Slovenia 0/29,
Spain 108/345, Sweden 0/197, Switzerland 63/252, and United Kingdom 2/466.
Infection prevalence was exceptionally high in Spain and Switzerland.
In Spain, ongoing chytridiomycosis-driven declines of midwife toads (Alytes
obstetricans) and salamanders (Salamandra salamandra) have
been documented since 1997 (4) and 1999 (5),
respectively, and confirmed with scanning electron microscopy, histologic
examination, and molecular detection methods (4,5). Common
toads have been suffering apparently minor chytrid-related die-offs in
Spain for several years, but mass die-offs were observed in 2004 (5)
(J. Bosch et al., unpub. data). No chytrid-related die-offs have been
reported in Switzerland. Furthermore, the infected animals from Switzerland
were all adults in good breeding condition, many of which reproduced successfully
in behavioral and ecologic experiments. Real-time PCR amplification profiles
for the Swiss samples were quantitatively equivalent to those generated
from samples of A. obstetricans collected during mass die-off events
in Spain; from these latter samples, we successfully isolated viable B.
dendrobatidis cultures from 2 geographically distinct areas. In Great
Britain, we found chytrid in 2 of 14 introduced North American bullfrogs
(Rana catesbeiana) caught in 2004 but did not find it in wild-captured
native species. Examination by microscope and electron microscope of 180
native British amphibians from 1992 to 1996 did not find chytrid infection
(A.A. Cunningham, unpub.data). The ability of the North American bullfrog
to act as a vector for chytrid range expansion has been hypothesized (9,10).
Our data may indicate that bullfrogs can fulfill this role in Great Britain
and other areas; we have found the molecular signal of chytrid infection
from introduced North American bullfrogs collected on 3 separate continents
(T.W.J. Garner et al., unpub. data).
This survey shows that B. dendrobatidis is widely and irregularly
distributed in Europe and infects a broad range of amphibian species.
Furthermore, because of the opportunistic nature of our sampling strategy,
our results certainly underestimate the overall prevalence of B. dendrobatidis
in Europe. These findings are surprising considering that chytrid-related
die-offs have been infrequently described in Europe. This may be because
B. dendrobatidis has only recently and rapidly expanded its range
into Europe (3), and the consequences are only now being
detected in wild amphibian populations; because the expression of chytridiomycosis
is environmentally limited (11); or because European
amphibians exhibit highly variable levels of resistance to chytrid infection.
Notwithstanding, our knowledge of the epidemiology of B. dendrobatidis
is insufficient to effectively manage wildlife and conduct disease abatement.
As data regarding the distribution of chytrid fungus accumulate and the
ecologic requirements for disease persistence and transmission are identified
(11), management of the pathogen can become more predictive.
Basic management practices, such as restricting transportation of potential
carriers and restricting pet trading and reintroduction projects, coupled
with field monitoring, must be improved to prevent further global emergence
of this pathogen. Our results also show that asymptomatic amphibians must
be included in any broad-scale epidemiologic screening for this emergent
pathogen.
Acknowledgments
We thank R. Jehle,
D. Schmeller, J.W. Arntzen, P. Lymberakis, B.R. Schmidt, B. Vincenz,
P.B. Pearman, K. Poboljšaj, E. Marzona, D. Seglie, H.-U. Reyer, C. Vorburger,
K. Grossenbacher, B. Schnüriger, V. Aguilar Sánchez, J. Foster, E. Ågren,
T. Mörner, I.U. Umo, and A.W. Sainsbury for providing tissue samples;
M. Perkins for providing technical assistance with the laboratory component
of this study; and Diverse Conservation Agencies of Spain for facilitating
permits for fieldwork in that country.
This study was supported
by an NERC standard grant (NER/A/S/2002/00832).
Partial funding
was provided to J. Bosch from a project supported by the Fundación BBVA.
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Suggested citation
for this article:
Garner TWJ, Walker
S, Bosch J, Hyatt AD, Cunningham AA, Fisher MC. Chytrid fungus in Europe
[letter]. Emerg Infect Dis [serial on the Internet]. 2005 Oct [date
cited]. Available from http://www.cdc.gov/ncidod/EID/vol11no10/05-0109.htm
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