J Periodontol • February 2014

Antimicrobial Resistance and Prevalence

of Resistance Genes of Obligate
Anaerobes Isolated From Periodontal
Abscesses
Yi Xie,* Jiazhen Chen,† Junlin He,* Xinyu Miao,† Meng Xu,* Xingwen Wu,* Beiyun Xu,* Liying Yu,*
and Wenhong Zhang†

Background: This study attempts to determine the anti-

microbial resistance profiles of obligate anaerobic bacteria
that were isolated from a periodontal abscess and to evalu-
ate the prevalence of resistance genes in these bacteria.
Methods: Forty-one periodontal abscess samples were
cultivated on selective and non-selective culture media to
isolate the oral anaerobes. Their antibiotic susceptibilities

P
eriodontal abscess has been de-
to clindamycin, doxycycline, amoxicillin, imipenem, cefra- fined as a suppurative lesion that is
dine, cefixime, roxithromycin, and metronidazole were de- associated with periodontal break-
termined using the agar dilution method, and polymerase down and localized pus in the gingival
chain reaction assays were performed to detect the pres- wall of the periodontal pocket.1 The vast
ence of the ermF, tetQ, nim, and cfxA drug resistance majority of dental abscesses with end-
genes. odontic or periodontal pocket origins
Results: A total of 60 different bacterial colonies was are polymicrobial anaerobic infections.2
isolated and identified. All of the isolates were sensitive These infections are predominantly at-
to imipenem. Of the strains, 6.7%, 13.3%, 16.7%, and 25% tributable to strict anaerobes, such as
were resistant to doxycycline, metronidazole, cefixime, and Gram-negative anaerobic bacteria, and
amoxicillin, respectively. The resistance rate for both clinda- partially facultative anaerobes, such as
mycin and roxithromycin was 31.7%. Approximately 60.7% the Streptococcus milleri group.3 How-
of the strains had the ermF gene, and 53.3% of the amoxicil- ever, in dental abscess, anaerobes out-
lin-resistant strains were found to have the cfxA gene. Two number aerobic and facultative bacteria
nim genes that were found in eight metronidazole-resistant in ratios ranging from 10:1 to 10,000:1,
strains were identified as nimB. with anaerobic Gram-negative bacilli
Conclusions: In the present study, the Prevotella species predominating.3 In periodontal abscesses,
are the most frequently isolated obligate anaerobes from Bacteroides forsythus, Fusobacterium
periodontal abscesses. The current results show their alarm- spp., Prevotella intermedia, Prevotella ni-
ingly high resistance rate against clindamycin and roxithro- grescens, and Porphyromonas gingivalis
mycin; thus, the use of these antibiotics is unacceptable for are the most prevalent species,4,5 and
the empirical therapy of periodontal abscesses. A brief these bacteria are more commonly iso-
prevalence of four resistance genes in the anaerobic bacte- lated from periodontal abscesses than
ria that were isolated was also demonstrated. J Periodontol endodontic abscesses. However, previous
2014;85:327-334. studies using swabs of purulent material
have demonstrated poor recovery of
KEY WORDS
strict anaerobes, which implicated strep-
Anti-infective agents; microbiology; periodontics. tococci or staphylococci as the causative
microorganisms of suppurative dental
* Department of Stomatology, Hua Shan Hospital, Fudan University, Yangpu, Shanghai, infection. This difference in recovery was
China.
† Department of Infectious Disease, Hua Shan Hospital, Fudan University. attributable to poor sampling techniques
and inadequate culture methods in these
investigations.6,7

doi: 10.1902/jop.2013.130081

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Drug Resistance of Obligate Anaerobes of Periodontal Abscess Volume 85 • Number 2

Antibiotics are often prescribed to limit the Microbial Samples

spread of the infection after the surgical drainage of The abscesses were drained after decontamination
pus from periodontal abscesses. However, reports of the mucosa. A sterile 10-mL inoculating loop‡ was
have indicated that abscess formation may be at- inserted into the deep area of the fistula for 20
tributable to the overgrowth of resistant pathogens.8 seconds. The loop was then immediately inoculated
Therefore, it is critical to know the susceptibility on two prereduced culture media using quadrate
profiles of clinically relevant oral pathogens (which section streak methods. Anaerobe basal agar§ me-
may vary considerably between patients in different dium was used as a non-selective culture medium
countries) for effective antibiotic treatment. and was supplemented with 5% sterile defibrinated
Amoxicillin, metronidazole, roxithromycin, and sheep blood.i The anaerobe basal sheep blood agar
clindamycin are commonly used to treat oral in- was supplemented with kanamycin¶ (100 mg/mL),
fections. Resistance against metronidazole was long and vancomycin# (75 mg/mL) was used as a selec-
considered to be rare in anaerobes. However, recent tive culture medium for the isolation of Gram-neg-
studies have shown that this resistance is no longer ative anaerobic bacteria. Both the selective and non-
uncommon.9 Metronidazole resistance in Bacter- selective culture media were immediately incubated
oides spp. appears to be encoded by the nimA in transparent bags** at 37C and were studied after
through nimG genes.9-11 The use of tetracycline has 2 to 4 days of growth. Each colony (usually two to six
decreased because of the high resistance rates colonies) with a distinct morphology (including the
observed in various microorganisms. The resistance colony size, pigmentation, transparency, viscidity,
in Gram-negative anaerobic bacteria, especially edge situation, apophysis situation, and hemolytic
in the genus Prevotella, was primarily attributable to features) was selected from the primary culture for
the presence of the tetQ gene,12,13 which encodes two or three repeated oxygen tolerance tests. The
the ribosomal protection protein.14 In oral anaer- isolates that only grew anaerobically were studied
obes, resistance to clindamycin–erythromycin has further. If several colonies from one specimen were
significantly increased during the past 20 years15 identified as the same species, only one colony was
and is usually encoded by the ermF gene (which used for additional study.
has the widest host range and was found in 10 of
Strain Species Identification
the examined anaerobic genera).16 Amoxicillin
Single anaerobic colonies were inoculated onto an-
continues to exhibit a high level of activity against
other anaerobe basal sheep blood agar plate for
the majority of oral anaerobes.17 However, cfxA
proliferation. Certain bacterium colonies from the
(class A/group 2e) has been characterized recently
same plate for the identification sequencing of 16S
in and cloned from oral amoxicillin-resistant Pre-
rRNA were sampled. Approximately 20 to 50 mg of
votella species.18
bacteria was harvested, suspended in a Tris–EDTA
In this study the aim is to investigate the an-
buffer, and pelleted for DNA extraction. The genomic
timicrobial resistance profiles of clinically cultured
DNA was extracted using a DNA extraction kit††
anaerobes from periodontal abscesses and their
according to the instructions of the manufacturer.
resistance to clindamycin, doxycycline, amoxicillin,
Approximately 1,400-bp 16S rDNA fragments were
imipenem, cefradine, roxithromycin, cefixime, and
amplified with the universal ribosomal 16S
metronidazole, which are commonly used in the
primers 8FLP (59-AGAGTTTGATCCTGGCTCAG-
treatment of periodontitis. The prevalence of four
39) and 1492RPL (59-GGTTACTTGTTACGACTT-39),
relevant resistance genes is also studied.
as described previously.19 The amplification products
MATERIALS AND METHODS were detected using electrophoresis in agarose gels
(1%) and sequenced using a DNA analyzer.‡‡ A se-
Participants
quence database of 16S ribosomal RNA sequences
Forty-one patients (21 males and 20 females; aged
(bacteria and archaea) from GenBank (using the
45 – 10 years [mean age: 47 years]) with peri-
BLASTN program through the National Center for
odontal abscesses were recruited at the Department
Biotechnology Information sever) was used to com-
of Stomatology of Hua Shan Hospital, Shanghai,
pare 1,400-bp 16S rDNA sequences.20 Identifica-
China, for this study from August 2011 to August
tion at the species level was defined as a 16S rDNA
2012. Informed written consent was obtained from
each patient, and the study was approved by the ‡ 10-mL Inoculating Loops, JET BIOFIL, Canada.
Ethics Committee of Hua Shan Hospital. The ex- § Anaerobe basal agar, Oxoid Limited, Basingstoke, Hampshire, U.K.
i Defibrinated sheep blood, Oxoid Limited.
clusion criteria were: 1) pregnancy; 2) consumption ¶ Kanamycin, Sigma-Aldrich, St. Louis, MO.
of systemic antimicrobials or anti-inflammatory # Vancomycin, Sigma-Aldrich.
** GENbag, bioMérieux, Marcy l’Etoile, France.
drugs in the previous 6 months; and 3) periodontal †† DNA extraction kit, TianGen Biotech, Shanghai, China.
therapy during the previous 6 months. ‡‡ 3730xl DNA Analyzer, Applied Biosystems, Foster City, CA.

328
J Periodontol • February 2014 Xie, Chen, He, et al.

Table 1.
Oligonucleotide Sequences and PCR Conditions Used to Detect Target Resistance Genes

Resistance Amplicon
Genes Oligonucleotide Sequence 59-39 Amplification Cycles Size (bp) Reference

cfxA/cfxA2 CGT AGT TTT GAG TAT AGC TTT 94C · 30 seconds, 35 cycles of 94C · 30 966 This study
seconds, 52C · 30 seconds, 72C · 75
seconds, and 72C · 5 minutes
GAT GTT GCC TAT ATA TGT C

tetQ TTA TAC TTC CTC Cgg CAT C 94C · 30 seconds, 35 cycles of 94C · 30 904 This study
seconds, 52C · 60 seconds, 72C ·
1.15 minutes, and 72C · 5 minutes
ATC ggT TCg AgA ATg TCCA
nim ATG TTC AGA GAA ATG GGG CGT 94C · 30 seconds, 35 cycles of 94C · 30 458 9
AAG seconds, 55C · 60 seconds, 72C · 10
minutes, and 72C · 5 minutes
GCT TCC TTG CCT GTC ATG TGC TC
ermF CGG GTC AGC ACT TTA CTA TTG 94C · 30 seconds, 35 cycles of 94C · 30 487 15
seconds, 50C · 30 seconds, 72C · 1
minutes, and 72C · 5 minutes
GGA CCT ACC TCA TAG AGA AG
bp = base pair.

sequence similarity of >99% to that of the prototype EUCAST, the breakpoints of ‡32 and ‡8 mg/mL for
strain sequence in GenBank; identification at the roxithromycin and cefixime, respectively, were used
genus level was defined as a 16S rDNA sequence in accordance with previous studies.25,26
similarity of >97% to that of the prototype strain
Detection of Drug Resistance Genes
sequence in GenBank.21
The ermF, tetQ, nim, and cfxA genes were amplified
Antimicrobial Susceptibility Testing using polymerase chain reaction (PCR) assays under
Antibiotic susceptibilities to clindamycin,§§ doxy- the conditions shown in Table 1 and with a total
cycline,ii amoxicillin,¶¶ cefradine,## cefixime,*** volume of 50 mL that was composed of 25 mL pre-
roxithromycin,††† metronidazole,‡‡‡ and imipenem§§§ mix,†††† 20 mM of each primer, and 5 to 100 ng
were determined using the agar dilution method22 bacteria genomic DNA. An ermF amplification was
with Brucella agariii plates that were supplemented performed in all strains because of its high per-
with 5 mg/mL hemin,¶¶¶ 1 mg/mL vitamin k1,### and centage of resistance to clindamycin, whereas tetQ,
5% defibrinated sheep blood. Two-day-old bacteria nim, and cfxA amplification were only performed in
(104 to 105 colony forming units) were inoculated the resistant strains because of their relatively low
onto the plates using multipoint inoculators.**** percentage of resistance to metronidazole, doxycy-
The minimal inhibitory concentration (MIC) was cline, and amoxicillin. The amplification products
defined as the lowest concentration of the antibiotic were sequenced using 3,730 and compared using
that repressed visible growth. Bacteroides fragilis the BLASTN program on the Nucleotide collection
ATCC 25285 served as the control strain. The (nr/nt) database in GenBank.
breakpoints were used in accordance with the rec-
ommendations of the Clinical and Laboratory §§ Clindamycin, National Institutes for Food and Drug Control, Beijing,
Standards Institute (CLSI):23 1) clindamycin (‡8 China.
ii
mg/mL); 2) imipenem (‡16 mg/mL); 3) metronida- ¶¶
Doxycycline, National Institutes for Food and Drug Control.
Amoxicillin, National Institutes for Food and Drug Control.
zole (‡32 mg/mL); 4) doxycycline (‡8 mg/mL); and ## Cefradine, National Institutes for Food and Drug Control.
5) amoxicillin (‡2 mg/mL). When CLSI antimicrobial *** Cefixime, National Institutes for Food and Drug Control.
††† Roxithromycin, National Institutes for Food and Drug Control.
breakpoints were not established, the breakpoints ‡‡‡ Metronidazole, National Institutes for Food and Drug Control.
§§§ Imipenem, Hangzhou MSD Pharmaceutical, Hangzhou, China.
of the European Committee on Antimicrobial iii Brucella agar, BD Difco, San Jose, CA.
Susceptibility Testing (EUCAST)24 were followed. ¶¶¶ Hemin, Sigma-Aldrich.
### Vitamin k1, Sigma-Aldrich.
Because the breakpoints of roxithromycin and ce- **** Multipoint inoculators, Denley Instruments, Billinghurst, Sussex, U.K.
fixime have not yet been determined by CLSI and †††† Premix Ex Taq, Takara, Kyoto, Japan.

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Drug Resistance of Obligate Anaerobes of Periodontal Abscess Volume 85 • Number 2

Table 2. The susceptibility assays were

repeated twice for all of the isolates.
Identification of Anaerobic Bacteria Isolated From
As shown in Table 3, the most ef-
Periodontal Abscesses by 16S rRNA fective antibiotic was imipenem, to
which all were sensitive. Low-to-in-
Genus of Strains/Total termediate resistance was found for
Microorganisms Strains Strains (percentage) other antimicrobial agents. In total,
Prevotella genus 42 42 of 60 (70%) four of 60 (6.7%), eight of 60
Black-pigmented Prevotella 29 29 of 60 (48.3%) (13.3%), 10 of 60 (16.7%), and 15 of
P. intermedia 12 — 60 (25%) of the strains were resistant
P. melaninogenica 7 — to doxycycline, metronidazole, ce-
P. nigrescens 3 — fixime, and amoxicillin, respec-
P. denticola 7 - tively. The least effective antibiotics
Non-black-pigmented Prevotella 13 13 of 60 (21.7%) in the present study are clindamy-
P dentalis 4 — cin and roxithromycin, which both
P. oris 3 —
had a 19 of 60 (31.7%) resistance
P. salivae 1 —
rate. Ten strains were resistant
Candidatus Prevotella conceptionensis 1 —
Prevotella sp. oral taxon 317 to both clindamycin and roxi-
Prevotella spp. 2 — thromycin. The resistances to
P. buccae 2 — amoxicillin and clindamycin were
high in the Prevotella species,
Veillonella genus 11 11 of 60 (18.3%) reaching 30.9% and 38.1%, re-
V. parvula/V. dispar 10 —
spectively, whereas the other
V. vegosae 1 —
anaerobic species had lower re-
Fusobacterium genus 2 2 of 60 (3.3%) sistances to these antibiotics (al-
F. canifelinum 1 — though not statistically significant
F. nucleatum 1 — because of the limited amount of
Actinomyces genus 1 1 of 60 (1.67%) strains) (Table 3).
A. odontolyticus 1 — The ermF gene was detected in
17 of the 60 strains, and 16 of these
Bacteroides 1 1 of 60 (1.67%) strains were phenotypically resistant
B. heparinolyticus 1 — to clindamycin and/or roxithromycin,
Shuttleworthia 1 1 of 60 (1.67%) whereas only one strain was sen-
S. satelle 1 — sitive to clindamycin and interme-
diately resistant to roxithromycin
Pentostrentococcus 1 1 of 60 (1.67%)
(MIC = 16 mg/mL). As shown in
P. stomatis 1 —
Table 4, in the amoxicillin-, metro-
Propionibacterium 1 1 of 60 (1.67%) nidazole-, and doxycycline- re-
P. acnes 1 — sistant strains, cfxA, nim, and tetQ
— = The proportion of species was not listed in this table. were detected in 53.3% (eight of
15), 25% (two of eight), and 50%
RESULTS (two of four) of the strains, respectively. Both of
A total of 60 different bacterial colonies were iso- the amplified nim genes had nucleotides that were
lated from the periodontal abscesses of 41 patients. 100% identical to nimB (National Center for Bio-
Identifications of these colonies are shown in Table technology Information gene identification no.
2; 42 isolates (70%) belonged to the Prevotella ge- 9496386); thus, they were identified as such.
nus, and 29 of 60 (48.3%) were black-pigmented
Prevotella, whereas 13 of 60 (21.7%) were non- DISCUSSION
black-pigmented Prevotella. Of the 60 strains, 11 In agreement with previous studies,1,3 Gram-nega-
(18.3%) belonged to the Veillonella genus. Other tive anaerobic rods are the most frequent isolates
genera, including Actinomyces, Peptostreptococcus, in periodontal abscesses, and the Prevotella spe-
Bacteroides, Shuttleworthia, and Propionibacterium, cies are those most frequently isolated in this
also had minor occurrences. Only three rods, Ac- research, suggesting their predominance in peri-
tinomyces odontolyticus, Shuttleworthia satelle, odontal abscess infection. It has been reported
and Propionibacterium acnes, were Gram positive, that non-black-pigmented Prevotella species (such
whereas the others were Gram negative. as P. oralis, P. buccae, and P. oris) are nearly as

330
J Periodontol • February 2014 Xie, Chen, He, et al.

Table 3.
MIC (mg/mL) Range and MIC50 and MIC90 Values of Prevotella and Non-Prevotella
Isolated From Periodontal Abscesses for Eight Antibiotics Tested in China

Resistant Strains/Total
Antibiotics Range (mg/mL) MIC50 MIC90 (resistance percentage)

Total (n = 60)
Cefradine <0.06 to >128 0.5 16 NA
Cefixime <0.06 to 32 0.125 8 10 of 60 (16.7%)
Amoxicillin <0.06 to >128 <0.06 32 15 of 60 (25%)
Clindamycin <0.06 to >128 <0.06 >128 19 of 60 (31.7%)
Metronidazole <0.06 to >128 0.5 >128 8 of 60 (13.3%)
Roxithromycin <0.06 to >128 2 >128 19 of 60 (31.7%)
Doxycycline <0.06 to >8 <0.06 1 4 of 60 (6.7%)
Imipenem <0.06 to 0.5 <0.06 <0.06 0 of 60 (0%)
Prevotella (n = 42)
Cefradine <0.06 to >128 0.5 32 NA
Cefixime <0.06 to 32 0.125 4 4 of 42 (9.5%)
Amoxicillin <0.06 to >128 <0.06 16 13 of 42 (30.9%)
Clindamycin <0.06 to >128 <0.06 >128 16 of 42 (38.1%)
Metronidazole <0.06 to >128 0.5 8 4 of 42 (9.5%)
Roxithromycin <0.06 to >128 2 64 11 of 42 (26.2%)
Doxycycline <0.06 to >8 <0.06 0.5 2 of 42 (4.8%)
Imipenem <0.06 to 0.5 <0.06 <0.06 0 (0%)
Non-Prevotella anaerobes (n = 18)
Cefradine <0.06 to 8 0.5 8 NA
Cefixime <0.06 to 16 1 16 6 of 18 (33.3%)
Amoxicillin <0.06 to 64 <0.06 8 2 of 18 (11.1%)
Clindamycin <0.06 to >128 <0.06 >128 3 of 18 (16.7%)
Metronidazole <0.06 to >128 0.5 128 3 of 18 (16.7%)
Roxithromycin <0.06 to >128 4 >128 6 of 18 (33.3%)
Doxycycline <0.06 to >8 <0.06 8 2 of 18 (11.1%)
Imipenem <0.06 to 0.5 <0.06 <0.06 0 of 60 (0%)
The following breakpoints were used in accordance with CLSI (2012):23 1) clindamycin (‡8 mg/mL); 2) imipenem (‡16 mg/mL); 3) metronidazole (‡32 mg/
24
mL); and 4) doxycycline (‡8 mg/mL). The following breakpoint was used according to EUCAST (2012): amoxicillin (‡2 mg/mL). The following breakpoints
were used according to references 25 and 26: 1) roxithromycin (‡32 mg/mL); and 2) cefixime (‡8 mg/mL).
MIC50 or MIC90 = MICs for 50% or 90% of the organisms, respectively; NA = Because no breakpoint was available for cefradine, resistance percentage was not
evaluated.

common as the black-pigmented species.1 How- rate renders it unacceptable for the empirical ther-
ever, in the research of the present authors, black- apy of severe infections that are predominantly
pigmented Prevotella (48.3%) had a higher ratio caused by Prevotella species. The detection rate for
than non-black-pigmented Prevotella (21.7%), and the ermF gene was 60.7% (13 of 19) in clindamycin-
this finding may be because of the higher virulence and/or roxithromycin-resistant Prevotella species,
of the black-pigmented Prevotella to cause in- which was slightly higher compared to the findings
fections.27 However, some of the non-Prevotella of Chung et al.,31 who found a 55.6% (20 of 36)
genera, such as Shuttleworthia and Propioni- detection rate in clinical anaerobic bacteria. Other
bacterium, that were isolated in this research have erm genes (ermB, ermC, ermQ, ermT, and erm35)
rarely been reported in periodontitis,28,29 and more have also been reported to encode macrolides and
information is needed to understand their clinical lincomycin resistance in anaerobes,15 which may
role in periodontal abscesses. explain the resistance of the non-tested ermF-neg-
In research by the present authors, clindamycin ative strains in this study.
was the least effective antibiotic against Prevotella, Tetracycline is one of the most widely used an-
which had 38.1% (16 of 42) resistant strains, and tibiotics worldwide, but its usage has decreased
this finding was in accordance with a previous because of increasing bacterial resistance to this
study30 in which 31% of the Prevotella species was drug. Doxycycline as a semisynthetic tetracycline
resistant to clindamycin. This alarming resistance has been shown to significantly reduce the anaerobic

331
Drug Resistance of Obligate Anaerobes of Periodontal Abscess Volume 85 • Number 2

Table 4.
Distribution of Resistance Genes of Resistant Strains Isolated From Periodontal Abscess

Positive Resistance Gene Strains/Phenotypic Resistant

Strains in the Genera (percentage)
Total Positive Resistance Gene Strains/
Resistance Genes Phenotypic Resistant Strains (percentage) Prevotella Veillonella Other Bacteria

nim 2 of 8 (25%) 2 of 2 (100%) — 0 of 6 (0%)

cfxA 8 of 15 (53.3%) 8 of 13 (62%) 0 of 2 (0%) —

ermF 17 of 28 (60.7%) 13 of 19 (68%) 3 of 7 (42%) 1 of 2 (50%)

tetQ 2 of 4 (50%) 2 of 4 (50%) — —
— = no phenotypic resistance strains in the genera.

population in plaque without increasing the pop- cfxA, although their ability to produce b-lactamases
ulation of resistant bacteria or allowing the bacteria remains untested.
to acquire antibiotic resistance.32 In the present Metronidazole is commonly used for the treatment
research, the rate of resistance to doxycycline is as of infections that are caused by anaerobic organ-
low as 6.7%, and some articles reported doxycy- isms. In the present research, it was found that eight
cline-resistant anaerobic bacteria that were isolated of the 60 anaerobic strains were resistant to met-
from periodontitis. Doxycycline could be recom- ronidazole and that four of these strains belonged to
mended as one of the first-line antimicrobial drugs Prevotella. Some studies have examined the distri-
against periodontal disease because of its high ac- bution of nim genes in Prevotella spp. Lubbe et al.39
tivity and low resistance. Tetracycline resistance has detected one nimA-positive strain among seven
always been associated with the presence of the tetQ metronidazole-resistant Prevotella species, whereas
gene, which has been found in 12 anaerobic gen- Katsandri et al.40 found one nimC-positive strain and
era.33,34 However, only two of four doxycycline- two nimE-positive isolates among 57 metronidazole-
resistant strains were detected to have tetQ, and resistant Prevotella species. In the present research,
other genes that promoted resistance are not iden- two Prevotella species strains (P. dentalis and P.
tified in this study. denticola) were detected to harbor nimB. Because
It is inspiring that all of the strains in the present five nim-negative strains with an MIC >128 mg/mL
study are susceptive to imipenem. However, imi- were recovered, other potential resistance mecha-
penem has an extremely broad spectrum of activity nisms may exist for these isolates, such as decreased
and is not recommended to use empirically in treating pyruvate:ferredoxin oxidoreductase activity, over-
light oral infections to prevent uncontrollable re- expression of efflux pumps, or alterations of the
sistance. rhamnose catabolism pathway.9,41
An amoxicillin–metronidazole combination has There are some limitations to the present study.
been used to treat periodontitis for the long term. In First, the anaerobic organisms should have been
the present study, 30.9% (13 of 42) of the Prevotella incubated for 7 to 14 days. However, after 4 days of
strains are resistant to amoxicillin; Kuriyama et al.35 incubation, the fast-growing colonies had increased
showed that amoxicillin resistance occurred in 34% in size and would have grown over some of the strict
of Prevotella isolates and that all of these resistant anaerobes. Therefore, the inoculations were limited
strains were found to produce b-lactamase. Reports to 4 days of incubation, and the development of the
indicated that the resistance rate for amoxicillin slow-growing anaerobes was likely inhibited. This
ranged from 9% to 54% in common isolates from may explain the high proportion of fast-growing
acute dental abscesses.36 Amoxicillin is advocated Prevotella and the loss of slow-growing anaerobes.
as a suitable first-line agent in these situations, Second, a limited amount of isolates from the primary
although the increased resistance of Prevotella plates based on typical morphology was chosen; thus,
strains to penicillins remains a matter of con- some anaerobes with similar phenotypes may have
cern.37 Prevotella species are known to produce been overlooked by the present study. Last, only the
b-lactamases that are encoded by cfxA,38 which antimicrobial resistance genes that had been fre-
may explain their resistance to amoxicillin. In the quently reported in Prevotella were detected; all other
present research, 61.5% (eight of 13) of amoxicillin- resistance genes that could encode for other anaer-
resistant Prevotella strains were found to harbor obes were not studied.

332
J Periodontol • February 2014 Xie, Chen, He, et al.

CONCLUSIONS microbiota associated with adult periodontitis. Oral

Microbiol Immunol 1996;11:282-288.
The use of macrolides and lincomycins should be
13. Roberts MC. Tetracycline resistance determinants:
monitored when treating periodontal abscesses be- Mechanisms of action, regulation of expression, ge-
cause of resistance of these anaerobic bacteria to netic mobility, and distribution. FEMS Microbiol Rev
these antibiotics. Their reduced susceptibility to 1996;19:1-24.
penicillins and metronidazole should also be noted 14. Roberts MC, Sutcliffe J, Courvalin P, Jensen LB,
Rood J, Seppala H. Nomenclature for macrolide and
and discussed. Some studies have addressed anti-
macrolide-lincosamide-streptogramin B resistance
microbial susceptibility profiles or detected resistance determinants. Antimicrob Agents Chemother 1999;43:
genes in the anaerobes that are isolated from peri- 2823-2830.
odontal abscesses. Careful monitoring of antimicro- 15. Roberts MC. Acquired tetracycline and/or macrolide-
bial resistance and the detection of these resistance lincosamides-streptogramin resistance in anaerobes.
Anaerobe 2003;9:63-69.
genes may verify the presence and spread of certain
16. Kuriyama T, Williams DW, Yanagisawa M, et al. Anti-
anaerobic strains, and these data could likely aid microbial susceptibility of 800 anaerobic isolates from
clinical practitioners in the empirical use of antibiotics patients with dentoalveolar infection to 13 oral antibi-
and could help identify the origin of the resistance. otics. Oral Microbiol Immunol 2007;22:285-288.
17. Madinier I, Fosse T, Giudicelli J, Labia R. Cloning and
ACKNOWLEDGMENTS biochemical characterization of a class A beta-lacta-
mase from Prevotella intermedia. Antimicrob Agents
This study was supported by the Postgraduate Inno- Chemother 2001;45:2386-2389.
vation Fund projects of Fudan University. Xie and Dr. 18. Handal T, Olsen I, Walker CB, Caugant DA. Detection
Chen contributed equally to this work. The authors and characterization of beta-lactamase genes in sub-
report no conflicts of interest related to this study. gingival bacteria from patients with refractory peri-
odontitis. FEMS Microbiol Lett 2005;242:319-324.
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