laboratorymedicine> july 2003> number 7> volume 34

535

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Case 1
A 14-year-old boy presented at the
emergency room with sudden onset ab-
dominal pain and distention. One and a
half months prior to admission, the patient
had a history of bowel reduction secondary
to malrotation of small bowel. A concomi-
tant appendectomy was also performed.
Radiographic study revealed pneu-
moperitoneum. The patient underwent
exploratory laparotomy. Intraoperative
findings revealed gastric perforation and
necrotic stomach involving the fundus
and greater curvature. There was gross
contamination of the abdominal cavity by
food and blackening of the abdominal
viscera. The inferior pole of the spleen
had a small infarct. About 600 mL
serosanguinous abdominal fluid was aspi-
rated and submitted for culture and sensi-
tivity. The stomach was massively
dilated. The omentum along the greater
curvature was necrotic and friable.
The Patients Clinical Progression
Postoperative course was complicated
by acidosis, hypotension, coagulopathy,
and fever. Gentamicin and Metronidazole
were started and continued for 12 days.
Symptoms improved starting on the fifth
day and the patient was discharged well.
Sarcina ventriculi and coagulase-negative
staphylococci were recovered from the
abdominal fluid culture.
Pathologic Findings
The adipose and omental tissues
showed focal fat necrosis, hemorrhage, and
gram-positive cocci in packets of 8 or more
that are flattened in areas of contact with
adjacent cells. The stomach showed diffuse
acute hemorrhagic gastritis with perfora-
tion and ulceration of the greater curvature
and fundus of the stomach. [I1 and I2]
There was also ischemic necrosis
with ulceration and perforation of the
greater curvature of the stomach with an
incidental finding of gram-positive cocci
in packets of 8 or more similar to that
seen in the omentum. [I3 and I4]
Case 2
This is a 50-year-old Hispanic male
who complained of nausea and vomiting
for a year accompanied by significant
weight loss, coffee-ground emesis, and
melena. Significant past medical history
includes a gunshot wound to anterior
chest in 1979. Computed tomography
(CT) scan of the abdomen and pelvis
showed 2 lesions in the liver ranging in
size from 1.5 cm to 2.5 cm.
Esophagogastroduodenoscopy revealed
Grade II esophagitis, a 5-cm hiatal hernia,
and a duodenal lesion. The duodenal mass
Clinical progression
Pathological findings
Microbiology of Sarcina ventriculi
Isolation and identification
rounds [microbiology and virology]
A Case Report of Gastric Perforation
and Peritonitis Associated With Opportunistic
Infection by Sarcina ventriculi
Lucilene F. Tolentino, MD, Natash Kallichanda, MD, Brenda Javier, MD, Robert Yoshimori, PhD, Samuel W. French, MD
Harbor-UCLA Medical Center, Torrance, CA
DOI: 10.1309/CDFF04HE9FHDQPAN
[I1] Hemorrhagic necrosis on the gastric
mucosa (H&E stain, x400)
[I2] Sarcina ventriculi infection of the stomach
wall. (H&E stain, x1,000)
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536

biopsy showed chronic superficial gastritis

and ulcer crust with gram-positive cocci
in packets of 8 or more. [I5 and I6]
No cultures were performed on the
biopsy material. Esophageal biopsy of the
squamocolumnar junction showed
chronic superficial gastritis and
esophageal mucosa demonstrating repara-
tive changes.
Background
Sarcina ventriculi were first observed
by Goodsir (1842) in the stomach contents
of a patient with gastric problems resulting
in gas formation and vomiting. The organ-
ism was subsequently reported in similar
cases, but it was not isolated in pure cul-
ture from the human stomach until 1911
when Beijerinck showed that S. ventriculi
could be grown using strictly anaerobic
techniques; he had previously shown that
the same organism occur widely in the
soil.
1
Later, it was even isolated from air,
in addition to from the surface layer of
sand and soil (Smith, 1933). Smith found
that live cultures of S. ventriculi retained
their viability for only 2 or 3 days, and as
spores were not demonstrated, it was mys-
terious how the anaerobic bacteria could
survive in soil for years. A recent study
has shown that spores only form at very
high pH values (> 8.0) when cultured with
concomitant change in morphology.
2
Sarcina ventriculi have also been
cultivated from feces of healthy humans,
interestingly mainly from people living on
vegetarian diets.
3
As reported by the re-
searcher Synnove Vatn, S. ventriculi was
also considered as the main cause of the
changes of gastropathy (gas formation,
bleeding, and/or ulcers in the abomasums)
in lambs in conjunction with Clostridium
fallax and Clostridium sordelli. These in-
vestigations show a clear connection be-
tween gastropathy and Sarcina-like
bacteria, which are found in about 80% of
the examined lambs. These bacteria have
not been previously demonstrated or cul-
tured from the lambs and are hardly men-
tioned in the international literature.
3
Schemm and colleagues reported
Sarcina-like bacteria (possibly S. ventri-
culi) have been seen upon histopathologi-
cal examination of the abomasums of
calves that died of abomasal bloat in 80%
to 90% of cases. In calves with ruptured
abomasums, bacteria were observed on
the peritoneal surface. One of the abo-
masal content samples for which isolation
was attempted exhibited Sarcina-type
cells in the preenrichment medium. Se-
lective cultivation for S. ventriculi was
successful in 1 case. However, attempts
to isolate the cells in pure culture have
not succeeded. The researchers claimed
that the organisms ability to grow at a
low pH and produce large amounts of gas
suggested that it may be the cause of abo-
masal bloat.
4
Microbiology of Sarcina
ventriculi
Sarcina ventriculi are nearly spheri-
cal cells, 1.8 to 3 m in diameter, and
occur in packets of 8 or more. Generally,
the cells are flattened in areas of contact
with adjacent cells. They are gram-posi-
tive, non-motile, chemoorganotrophic
anaerobes, having exclusively fermenta-
tive metabolism. They are relatively
aerotolerant. Carbohydrates are
fermentable substances. The main prod-
ucts of glucose fermentation are ethanol,
acetic acid, carbon dioxide, and hydro-
gen. They utilize the Embden-Meyerhof
pathway for the fermentation of carbohy-
drates.
5
Two fermentation systems for
pyruvate metabolism are present: a yeast-
type decarboxylase that produces
acetaldehyde and carbon dioxide from
pyruvate and a clostridial-type
ferredoxin-dependent pyruvate-clastic
system that yields acetyl-phosphate, car-
bon dioxide, and hydrogen. Furthermore,
carbon dioxide and hydrogen are
produced from formate hydro-genlyase.
Phosphotransacetylase catalyzes the con-
version of acetyl-CoA to acetyl
phosphate which is metabolized to ac-
etate in a reaction catalyzed by acetyl
kinase. Furthermore, acetyl CoA is me-
tabolized to butyrate via a pathway simi-
lar to that present in saccharolytic
clostridia and involving butyryl
transferase and butyrate kinase.
6
[I3] Necrotic omentum showing S. ventriculi
(H&E stain, x100)
[I4] Spherical cells in packets of 8 or more (H&E
stain, x1,000).
[I5] H&E stain of chronic superficial gastritis
with ulcer crust and gram positive cocci in
packets of 8. (magnified 1,000x).
[I6] H&E stain of gram-positive cocci in packets
of 8 mixed with vegetable fibers and mucus on
top of an ulcer crust. (magnified 1,000x).
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537

The optimum temperature range for

growth is 30C to 37C which occurs at
pH 1 and 9.8. Growth of S. ventriculi
takes place in the human stomach as a
result of the development of certain
pathological conditions (eg, pyloric steno-
sis) that retard the flow of food to the
small intestine. Under these abnormal
circumstances, at the acid pH of the
stomach and in the presence of carbohy-
drates and other growth nutrients
contained in food, S. ventriculi thrives
and multiplies rapidly. It was previously
shown that Sarcina occur widely in soil
and could be ingested through soil parti-
cles present in vegetables.
6
Sarcina ventriculi Isolation and
Identification
1. Culture
a. Tissue submitted for culture was
ground in a Kendall precision
disposable tissue grinder.
b. Two blood agar plates (BAP),
MacConkey (Mac), colistin-
nalidixic acid (CNA), and
Schaedlers broth were inoculated.
c. A BAP was incubated anaerobically
for 2 days while the other BAP, Mac,
and CNA were incubated in a 5%
CO
2
incubator for 18 to 24 hours.
d. Schaedlers broth was incubated in
a non-CO
2
incubator for 5 days.
e. Growth was detected only anaerobi-
cally at 48 hours and using a stereo-
scope, colonies of different
morphologies were picked and
subcultured in duplicate onto BAP
and chocolate agar (CHOC).
f. The BAP was incubated anaerobi-
cally and the CHOC was incubated
aerobically in a CO
2
incubator for
48 hours.
g. After an incubation period of 48
hours, colonies growing on the BAP
and not on the CHOC were Gram
stained.
2. Identification (Virginia Polytechni
Institute, VPI Anaerobic Technique)
a. Gas Liquid Chromatography
(GLC), Hewlett-Packard
i. Only colonies of gram positive
cocci in cubes were isolated and
inoculated into prereduced peptone
yeast glucose or chopped meat
glucose broth (Anaerobic Systems
M, Morgan Hill, CA) and incubated
for 24 hours.
ii. Ether and chloroform extracts for
short-chain fatty acids and methy-
lated end products of fermentation
were performed according to the
protocol of VPI.
iii. GLC was performed and only a
major acetate peak was detected
from the ether extract. No signifi-
cant methylated products from the
chloroform extracts were detected.
b. Summation
i. Large gram-positive anaerobic
cocci (1.5 to 3 m) in packets of 8
is consistent with the genus
Sarcina.
ii. GLC with only acetate is consis-
tent with S. ventriculi.
iii. Sarcina ventriculi was the only
organism isolated which can with-
stand the low stomach acidity, pH 1,
for prolonged periods of time which
explains a pure culture of the
organism.
Discussion
One of the products of the
organisms fermentation system is
acetaldehyde. Acetaldehyde has been
shown to bind covalently to many cellular
and extracellular proteins.
7
One study
shows that both exogenous acetaldehyde
and that produced locally from ethanol
binds to gastric mucosa. This adduct for-
mation has been suggested to be a patho-
genetic factor behind ethanol-associated
gastric injury.
8
Acetaldehyde has proper-
ties that make it very suitable for poten-
tial nucleophilic attacks. Since many
nucleophilic groups are present in the
proteins, they are natural targets in vari-
ous tissues for reactive acetaldehyde. The
binding of acetaldehyde with proteins
results in the formation of 2 major types
of product, which have been classified as
unstable and stable acetaldehyde-protein
adducts. The unstable adducts may serve
as intermediates in stable adduct forma-
tion and can be stabilized by reducing
agents such as NADH, which is found in
large amounts in the liver during ethanol
metabolism.
The stable adducts appear to be the
most likely causes of toxic effects.
7
These
studies are the result of extensive studies
on alcohol-induced toxicities. Acute inges-
tion of alcohol has been shown to produce
inflammatory changes in the stomach and
duodenum of humans. The most promi-
nent findings included hemorrhagic ero-
sions, subepithelial blebs, and infiltration
of inflammatory cells into the lamina pro-
pria.
9
One of the first byproducts of alco-
hol metabolism is acetaldehyde.
Conclusion
Although the role of S. ventriculi in
the mechanism of mucosal injuries in
these cases are not clear, the local accumu-
lation of acetaldehyde formed from the
fermentation system for pyruvate metabo-
lism and indirectly, from ethanol, product
of glucose fermentation by the organism
could have induced the stomach and duo-
denal injuries similar to the mechanism of
acetaldehyde-induced mucosal injuries in
acute alcohol ingestion. Furthermore, the
tremendous CO
2
production from glucose
fermentation and pyruvate metabolism
suggests the mechanism of abdominal dis-
tention of the patient.
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J Med Microbiol. 1971;4:343-349.
2. Lowe SE, Zeikus J. Purification and
characterization of pyruvate decarboxylase
isolated from Sarcina ventriculi. Gen Microbiol.
1992;138:803-807.
3. Vatn S, Gunnes G, Nybo K, et al. Possible
involvement of Sarcina ventriculi in canine and
equine acute gastric dilatation. Acta Vet Scand.
2000:41.
4. DeBey BM, Blanchard PC, Durfee PT. Abomasal
bloat associated with Sarcina-like bacteria in goat
kids. J Am Vet Med Assoc. 1996;209:1468-1469.
5. Canale-Parola E. Biology of the sugar fermenting
sarcinae. Bacteriol Rev. 1970;34:82-97.
6. Canale-Parola E, Sneath NS, Mair ME, et al.
Genus Sarcina. Bergeys manual of systematic
bacteriology. Vol. 2. 1986;1100-1103.
7. Goward CR, Miller J, Nicholls DJ, et al.
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Ethanol-derived microbial production of
carcinogenic acetaldehyde in achlorhydric
atrophic gastritis. Scand J Gastroenterology.
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