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Letter
Enteropathogenic Klebsiella
pneumoniae HIV-Infected Adults, Africa
Phuong L. Nguyen Thi,* Simon Yassibanda, Awa Aidara, Chantal
Le Bouguénec,§ and Yves Germani*
*Institut Pasteur de Bangui, Bangui, Central African Republic; Hôpital
de l'Amitié, Bangui, Bangui, Central African Republic; Institut
Pasteur de Dakar, Dakar, Sénégal; and §Institut Pasteur, Paris, France
Suggested citation for this article: Nguyen Thi
PL, Yassibanda S, Aidara A, Le Bouguénec, C, Germani Y. Enteropathogenic
Klebsiella pneumoniae HIV-infected adults, Africa. Emerg Infect
Dis [serial online] 2003 Jan [date cited]. Available from: URL:
http://www.cdc.gov/ncidod/EID/vol9no1/02-0138.htm
To the Editor: Although Klebsiella pneumoniae lives as
a commensal in the intestine, this bacterium can occasionally cause diarrhea
in HIV-negative persons (1-4). Some of these diarrheagenic
strains encode thermostable or thermolabile toxins (2).
One group of researchers showed that a K. pneumoniae strain isolated
from bloody diarrhea can bind to HeLa cells and cytoskeletal proteins,
such as the actin that accumulates at the point of bacterium-host contact
(3). However, this isolate did not contain any of the
genes encoding virulence factors that have been ascribed to pathogenic
Escherichia coli strains and are responsible for bloody
diarrhea or dysenteric syndromes (3).
In Bangui, the rate of isolation of pure cultures of K. pneumoniae
from the stools of immunocompromised HIV-infected adults with chronic
diarrhea is increasing. This finding was observed during the routine biological
analyses performed in the Pasteur Institute Medical Laboratory and is
consistent of that made by Gassama et al. in Dakar (5).
The role of K. pneumoniae in HIV-infected adults is not well documented.
As no other known enteric pathogens were isolated from these samples,
we conducted a case-control study in Bangui in 19992001 to determine
the clinical significance of K. pneumoniae. The study population
included 31 adults hospitalized with chronic diarrhea and 31 matched controls.
(Because of civil unrest in Bangui due to military rebellions and the
difficulties involved in recruiting controls, the study was performed
on a small sample.) To be included in the study, the patients had to be
HIV positive, be >18 years of age, have provided a stool sample
containing K. pneumoniae on the day of recruitment, and have given
informed consent. Inclusion criteria were the same for controls except
that they did not have diarrhea on the day of recruitment or in the previous
month. Controls were family members or neighbors of the patients, matched
by age (within 5 years) and sex. Specimens from cases and controls were
collected over the same 1-month period. Known enteric pathogens were identified
by standard methods as described (6). Endoscopic examinations
were used to diagnose pseudomembranous colitis in patients with bloody
chronic diarrhea or watery chronic diarrhea. The median CD4+ cell count
was 122 cells/µL in the patients and 436 cells/µL in the controls.
AIDS-related symptoms were observed in all of the cases (Centers for Disease
Control and Prevention [CDC] stage C2 or C3) and none of the controls
(CDC stage A1). Of the 31 patients, 7 (22.6%) had bloody chronic diarrhea,
9 (29%) had watery chronic diarrhea, and 15 (48.4%) had mild chronic diarrhea.
Pseudomembranous colitis was diagnosed in nine patients (six with bloody
diarrhea, three with watery chronic diarrhea) who had been taking several
antibiotics, including ampicillin, for a long period (>1 month). Five
K. pneumoniae colonies were randomly picked up from each case sample
and examined. Control colonies were chosen if their appearance suggested
K. pneumoniae. The mean number of strains tested was 4.99 for patients
and 4.64 for controls (not significant, p=0.969). All of the enteric bacteria
isolated from the patients and grown on nonselective bromocresol purple
medium were K. pneumoniae, whereas an average of 10% to 20% of
the enteric bacteria isolated from the controls were K. pneumoniae.
We used assays typically used to identify the virulence factors of diarrheagenic
E. coli (7) to characterize the virulence properties
of the K. pneumoniae isolates, their genotypes, and their phenotypes
(their ability to bind to cultured HEp-2 cells and to promote cytoskeleton
modifications [fluorescent actin staining test], to invade epithelial
cells, to produce various enterotoxins and cytotoxins, and to induce fluid
accumulation in the intestines of newborn mice). The rabbit ligated ileal
loops test was performed when the genetic or phenotypic (on Vero or Y1
cells) tests were positive for toxins. All isolates from 27 patients (7,
9, and 11 with bloody, watery, and mild chronic diarrhea, respectively)
and two of the isolates from one control displayed an aggregative adherence
phenotype on HEp-2 cells. This phenotype appeared to be significantly
associated with chronic diarrhea (27/31 cases vs. 1/31 controls, Chi =40.7,
p<10-6). All HEp-2adherent K. pneumoniae isolated
from six of the patients produced toxins. The culture supernatants of
the HEp-2adherent K. pneumoniae strains isolated from four
of the patients with bloody chronic diarrhea and pseudomembranous colitis
had cytotoxic effects on Vero and Y1 cells, as characterized by the rounding
of cells after 24 h, followed by their detachment from the culture plate
and death after 72 h. These effects were not neutralized by rabbit antisera
raised against the Shiga toxin or the cholera toxin. The HEp-2adherent
K. pneumoniae strains isolated from two patients with watery chronic
diarrhea were enterotoxigenic in ligated rabbit ileal loops. Only the
HEp-2adherent K. pneumoniae strains isolated from one patient
with watery chronic diarrhea and pseudomembranous colitis (5 strains)
and from four patients with mild chronic diarrhea (20 strains) carried
sequences related to virulence genes from pathogenic enteroaggregative
E. coli. These isolates were all positive for the astA gene,
which encodes the EAST1 toxin, and for the genes that produce the AAF/I
fimbriae.
K. pneumoniae is normally resistant to beta-lactams. Multidrug-resistant
K. pneumoniae have been reported (1). All K.
pneumoniae isolates in this study were resistant to several antibiotics
including cotrimoxazole and ampicillin, which are largely used in Bangui
according to the recommendations of the World Health Organization (8).
In addition to the nonspecific measures used to correct and prevent fluid,
electrolyte, and nutritional imbalances, all persons with bloody and watery
chronic diarrhea (including those with pseudomembranous colitis) and 5
of the 15 patients with mild chronic diarrhea (10 were lost to follow-up)
were treated with ofloxacin (800 mg/day) or ceftriaxone (2 g/day), based
on the results of antimicrobial susceptibility testing. The state of all
patients with pseudomembranous colitis and mild chronic diarrhea, and
of five of the patients with watery chronic diarrhea (one patient died),
improved within 10 days of treatment.
In Dakar, during the study describing ordinary and opportunistic enteropathogens
associated with diarrhea in adults (5), stool samples
were collected from five HIV-infected adults with watery chronic diarrhea.
In all cases, heavy K. pneumoniae growth was observed on the primary
culture media, and no other known pathogens were recovered. These K.
pneumoniae strains were subjected to the same phenotypic and genotypic
tests as the strains isolated in Bangui. HEp-2adherent K. pneumoniae
was identified in four of these five samples. The condition of all the
patients rapidly improved after treatment with ofloxacin. In Bangui and
Dakar, repeated stool cultures were negative for K. pneumoniae
by the end of treatment, providing further evidence that these K.
pneumoniae were of etiologic importance, especially the HEp-2adherent
K. pneumoniae strains.
Only seven patients (four with mild, two with watery, and one with bloody
chronic diarrhea) had not taken antibiotics during the 2 weeks before
stool collection. The stool specimens from these seven patients yielded
pure primary cultures of HEp-2adherent K. pneumoniae and
no other bacterial enteric pathogens. None of these seven participants
was diagnosed with pseudomembranous colitis. The HEp-2adherent K.
pneumoniae strains isolated from the two participants with watery
chronic diarrhea induced the accumulation of fluid in ligated rabbit ileal
loops, and the HEp-2adherent strains isolated from three of the
participants with mild chronic diarrhea carried the astA gene,
which is associated with pathogenic EAEC. Among the five patients with
pseudomembranous colitis, all of whom had received antibiotics before
the onset of illness, we found that the four isolates from the patients
with bloody chronic diarrhea were cytopathogenic; the one isolate from
the patient with watery chronic diarrhea had the pathogenic marker for
enteroaggregative E. coli. These findings suggest that not only
is K. pneumoniae associated with chronic diarrhea in HIV-infected
persons but also that infection with particular HEp-2-adherent K. pneumoniae
subtypes may be associated with specific clinical illness.
Financial support
was provided by Agence Nationale de Recherche sur le SIDA (contract
1227) and Groupe d'Etude des Infections Diarrhéiques (ACIP, Réseau International
des Instituts Pasteur et Instituts Associés).
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