| dc.description.abstract | Burkholderia pseudomallei is a Gram-negative bacterium responsible for
causing melioidosis in a wide range of species. It may infect susceptible hosts
through cutaneous, ingestion, or inhalation routes. The disease has a moderate to
high case fatality rate, and the causative microorganism is categorized as Tier 1
Select Agent by the United States CDC. Concerns regarding melioidosis were
raised among nonhuman primate (NHP) researchers in Indonesia, following a fatal
melioidosis case in a cynomolgus macaque (Macaca fascicularis) in a primate
research center and fatal cases in orangutans. Those incidents were the driving force
to conduct surveillance in primate populations.
Serosurveillance was conducted on cynomolgus (Macaca fascicularis) and
pig-tailed macaques (Macaca nemestrina) in several facilities breeding facilities in
West Java and Banten, Indonesia: Jonggol, Darmaga, Lodaya, and Tinjil Island
facilities. The serosurveillance was conducted using a panel of enzyme-linked
immunosorbent assays targeting immunoglobulins G toward B. pseudomallei
proteins: purified lipopolysaccharide (LPS) and recombinant proteins of alkyl
hydroperoxide reductase subunit C (AhpC), hemolysin coregulated protein (Hcp1),
and putative outer membrane porin protein (OmpH). Demographic factor analysis
showed that species and breeding location were significant factors that were
associated with seropositivity. Cynomolgus macaques and Jonggol facility have
significantly association with seropositivity towards B. pseudomallei antigenic
proteins. Enclosures in Jonggol are designed with soil flooring that may act as the
reservoir of B. pseudomallei. Wet season and the observed grass-plucking behavior
in those macaques may have contributed to the exposure to B. pseudomallei in the
deeper layer of the soil. Pig-tailed macaques are predicted to be less-susceptible to
the bacteria, since they showed lower immune response even though being reared
in the same facility as some seropositive cynomolgus macaques. An evaluation of
the immunoassay panel showed that there were strong and positive correlations of
the recombinant protein assays (AhpC, Hcp1, and OmpH), whereas antibody level
in LPS assay was higher but showed no correlation to the results of other assays.
This may suggest that the development of antibody towards B. pseudomallei LPS
is quicker and stronger, since LPS is a strong T-cell independent antigen. It may
explain the reason that some animals had high level of antibody to LPS but not to
other antigens.
Even though the seropositivity ranges from 13-88% in several facilities, the
presence of immunoglobulin G against B. pseudomallei does not necessarily mean
that the macaques were currently having melioidosis, as the antibodies may reflect
asymptomatic infection or a resolved infection. Efforts to culture throat, oral,
wound, nasal, and rectal swabs of seropositive macaques resulted in no positive
culture for B. pseudomallei. Based on previous reports by other researchers,
macaques (including Macaca mulatta, Macaca fascicularis, and Macaca
nemestrina) have been considered as animal species with moderate to high
resistance towards melioidosis. Negative culture from seropositive animals in this
study may indicate that those animals have been exposed to B. pseudomallei, but
do not develop melioidosis, or the condition has been resolved. Environmental
sampling from soil in several breeding facilities also resulted in no culturable B.
pseudomallei, but the number of samples and area coverage was not adequate to
represent the very large area of the facilities.
In this study, B. pseudomallei was isolated from a fatal melioidosis case in a
Bornean orangutan (Pongo pygmaeus wurmbii). Diagnosis was confirmed through
culture of pulmonary abscess which was then followed by TTSS-1 real-time PCR
assay of the bacterial colonies. In order to provide information regarding pathogen
characteristics, morphology observation, multilocus sequence typing (MLST) and
YLF/BYFC PCR was conducted on B. pseudomallei isolate obtained from
pulmonary abscess. On Ashdown agar, the isolates resembled Type I and Type II
morphotypes of B. pseudomallei. The isolate showed positivity to YLF gene cluster,
which is almost exclusively found in isolates form Asia region. Sequence typing of
seven loci of B. pseudomallei housekeeping genes (ace, gltB, gmhD, lepA, lipA,
nark, and ndh) showed a combination of allele sequence type, that was further
identified as B. pseudomallei sequence type (ST) 54. That sequence type was
analyzed by E-BURST, a method that picture relatedness between similarity in
allele types. E-BURST was performed on multiple datasets; all sequence types in
Borneo or Kalimantan, all sequence types in Indonesia, all sequence types in
Southeast Asia, all sequence types found in animals, all sequence types found in the
environment, all sequence types found in humans, and all reported sequence types
worldwide. E-BURST analysis of this ST and other STs of B. pseudomallei found
in SE Asian countries showed that ST54 is clustered together with STs from
Malaysia, Thailand, and Cambodia. It is predicted that ST54 shares a common
ancestor with STs found in those countries. Information obtained in this study is the
first milestone in discovering melioidosis in nonhuman primates in Indonesia and
may serve as a stepping stone to discover the burden of the disease in other animal
species, by providing the benefits of using panel immunoassays and providing
information on local B. pseudomallei characteristics | id |
