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Dispatch
Weissella confusa
Infection in Primate (Cercopithecus mona)
Ana I. Vela,* Concepción Porrero,* Joaquín Goyache,* Ana Nieto,* Belen
Sánchez,* Víctor Briones,* Miguel Angel Moreno,* Lucas Domínguez,* and
José F. Fernández-Garayzábal*
*Universidad Complutense, Madrid, Spain
Suggested citation
for this article: Vela AI, Porrero C, Goyache J, Nieto A, Sánchez
B, Briones V, et al. Weissella confusa infection in primate (Cercopithecus
mona). Emerg Infect Dis [serial online] 2003 Oct [date cited].
Available from: URL: http://www.cdc.gov/ncidod/EID/vol9no10/02-0667.htm
We describe the
first systemic infection by Weissella confusa in a mona monkey
(Cercopithecus mona) on the basis of microbiologic, molecular
genetic, and histologic data. The same strain of W. confusa,
as determined by pulsed-field gel electrophoresis, was isolated in pure
culture from brain, liver, spleen, and intestine of this primate, illustrating
the clinical importance of the isolations.
Weissella microorganisms are gram-positive, catalase-negative
coccobacilli, which have been isolated from a wide variety of habitats
such as soil, fresh vegetables, fermented foods, or meat and meat products
(1,2). The genus Weissella is peculiar since it
currently includes 11 validated species, but only Weissella confusa
(basonym Lactobacillus confusus) and W. cibaria have been
isolated from human or animal clinical sources. W. cibaria has
been isolated from human bile and feces, the liver of a canary, and material
from an infected ear in a dog (1). W. confusa
has been isolated from feces of children with bacteremic infections (3)
and liver transplants (4), and from the peritoneal fluids
and abdominal walls of two patients (5). In animals,
W. confusa has been isolated from necropsy specimens from a dog
and from the ear of a dog with otitis (1). However, with
the exception of a thumb abscess caused by W. confusa in a healthy
49-year-old man (6), the clinical significance of all
other clinical isolates was not clearly established. This article describes
the first well-documented systemic infection caused by W. confusa
in a primate.
Case Report
A juvenile female mona monkey (Cercopithecus mona) was found dead
without clinical signs of disease in the previous 24 h. The animal had
no previous relevant medical history. The monkey was housed in a cage
with another monkey, which formed part of a primate bioacoustic research
unit. None of the other monkeys housed in the same reseach unit died or
exhibited any clinical sign. The dead monkey was sent to the hospital
of the veterinary school at the Complutense University in Madrid for necropsy.
Postmortem examination showed the existence of congestion, edema, and
petechial hemorrhages in most internal organs, especially marked in the
brain, liver, and spleen, which are typical lesions associated with systemic
infections. Samples from intestine, lung, liver, and brain were collected
under aseptic conditions for microbiologic studies. Tissue samples were
surface-plated on Columbia blood agar (bio-Mérieux España, s.a. Madrid)
and incubated aerobically and under anaerobic conditions for 48 h at 37°C.
Gram-positive, catalase-negative, facultative anaerobic coccobacilli were
isolated in pure culture from lung, liver, brain, and intestine. Biochemical
characterization was achieved by using the commercial Rapid ID32 Strep
version 2.0 system (bioMérieux España, s.a. Madrid) according to the manufacturer’s
instructions. The four isolates had identical biochemical profile (numerical
code 72007000000), being identified as Leuconostoc spp. Acid production
from ribose, L-arabinose, and galactose was also tested by using phenol
red base medium (Difco Laboratories, Detroit, MI), supplemented with 1%
(w/v) sugar, after 48 h of incubation at 37°C. Antimicrobial susceptibility
was tested by the microdilution method and haemophilus test medium with
lysed horse blood (7) with a commercially prepared dehydrated
panel (Sensititre, Trek Diagnostic Systems, East Grinstead UK) as previously
described (8). MICs (in mg/mL) were as follows: tetracycline,
<1; amoxicillin, <0.25; trimethoprim, 32; erythromycin,
<0.5; penicillin, <0.5; chloramphenicol, 8; ciprofloxacin,
<0.25; and vancomycin >128. Resistance of W. confusa
to vancomycin has been reported previously (4,6,7).
For histopathologic studies, tissues were fixed in 10% neutral-buffered
formalin, embedded in paraffin, cut in 4-mm sections, and stained with
hematoxylin and eosin. Histologic examination of the lungs, spleen, and
liver showed the existence of inflammatory infiltrates composed mainly
of neutrophils, and in lower proportion, of lymphocytes and macrophages
(Figure 1), suggesting the existence of an acute
septicemic process. Gram-positive coccobacilli emboli were observed in
some hepatic vessels, suggesting a hematogenous dissemination.
Identifying Weissella species by classic phenotypic methods can
be difficult (1,9). Comparing the 16S rRNA gene sequences
of bacterial species is a useful approach for the identifying unusual
clinical isolates or those which cannot be properly identified by conventional
phenotypic methods (10,11). The 16S
rRNA gene of each isolate was amplified by polymerase chain reaction (PCR)
and further sequenced to determine genotypic identity (12).
The determined sequences consisted of approximately 1,400 nucleotides
and were compared with the sequences of other gram-positive, catalase-negative
species available in the GenBank database, by using the BLAST program
(available from: URL: http://www.ncbi.nlm.nih.gov/BLAST).
The 16S rRNA gene analysis indicated that the four isolates were genotypically
identical, displaying the closest sequence similarity (99.9%) with W.
confusa (accession no. AB023241). Sequence similarity with W. cibaria
was 99.2%, which agrees with the high sequence similarity reported for
both species (1). Overall results of the phenotypic characterization
of the clinical isolates were consistent with those described for this
species (13). Like W. confusa, clinical isolates
were able to produce acid from ribose and galactose but not from L-arabinose,
one of the few biochemical tests that can differentiate this species and
W. cibaria (1). These results support the identification
of the clinical isolates as W. confusa. Weissella microorganisms
can be isolated as normal flora of the intestinal tract (l,14,15
). Thus, an extraintestinal origin of the systemic infection is most
likely.
W. confusa isolates were molecularly characterized by pulsed-field
gel electrophoresis (PFGE), according to the specifications of Vela et
al. (16) with the CHEF-DR III system (Bio-Rad Laboratories,
Hercules, CA). The restriction enzymes ApaI (Promega UK Ltd., Southhampton,
UK) and SmaI (MBI Fermentas Vilinius, Lithuania) were used according
to the manufacturer’s recommendations. These enzymes have been successfully
used for the molecular typing of microorganisms closely related to Weisella
(17). Gels were interpreted by standard criteria (18).
All W. confusa isolates displayed undistinguishable macrorestriction
patterns by PFGE with SmaI (data not shown) and ApaI (Figure
2) restriction enzymes, demonstrating that the systemic infection
was caused by a single strain of W. confusa.
Conclusions
Weissella are considered nonpathogenic microorganisms because
most of the strains isolated from clinical samples have been obtained
as mixed cultures without evidence of their clinical significance (1,7).
In this study, the same strain of W. confusa, as determined by
PFGE, was isolated in pure culture from the brain, liver, and spleen;
the isolations from these organs, together with the histopathologic data,
illustrate the clinical importance of the isolations. These results generate
further speculation about the potential of W. confusa as an opportunistic
pathogen. This is the first well-documented study in which, by combining
microbiologic, molecular genetic, and histologic, data, the clinical importance
of the isolation of W. confusa and its implication in an animal
infection is clearly established.
Dr. Fernández-Garayzábal
is a professor of microbiology at the Veterinary Faculty of the Complutense
University. His main research interests include the molecular detection
and epidemiology of bacterial pathogens of clinical relevance in veterinary
medicine.
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