Keywords
flexor graft - anterior cruciate ligament reconstruction - infection - septic arthritis
Introduction
The arthroscopic reconstruction of the anterior cruciate ligament (ACL) is an efficient
procedure to regain knee stability after a lesion.[1]
[2] Infection post-ACL reconstruction is a rare complication, but potentially severe,
with an incidence ranging from 0.14% to 1.8%.[1]
[3]
[4]
[5]
[6]
[7] The development of infection is associated with a substantial morbidity (loss of
the graft used in the reconstruction, chondral degeneration, arthrofibrosis, and osteoarthrosis),
in addition to functional loss.[8]
[9] In an infection after ACL reconstruction, signs and symptoms, such as pain, joint
effusion, temperature increase, and movement restriction, are nonspecific and can
be acute or late alterations.[7] These alterations have low specificity, since they can be observed during the normal
postoperative period; in addition, when alone, they are not very helpful to confirm
an infection. Thus, a supplementary laboratorial evaluation is important for the diagnosis.[1]
[5]
[6]
[10]
Among the several reconstruction techniques, bone-patellar tendon-bone autografts
and hamstring tendons grafts are the most used for primary ACL reconstruction, and
their infection rates are similar.[11] Both require preparation at the auxiliary table before the final fixation. Hamstring
tendons are widely used nowadays because of the lower residual pain, of the lower
sensitivity change at the anterior region of the knee, of the higher tolerance to
extenuating activities, and of the lower rate of radiographical osteoarthritis.[12]
[13]
[14]
[15] There are two techniques for their preparation: the classical technique, with the
retraction of the hamstring tendons, and the other maintaining their tibial insertion.
Several authors defend that the preservation of this insertion makes the surgery more
biological, improves proprioception and blood supply, and reinforces the tibial portion
of the graft, considered its most fragile region.[16]
[17]
[18]
[19]
[20]
Therefore, the present study aims to evaluate the contamination rates of autografts
prepared with hamstring tendons with both techniques and to verify if the intraoperative
contamination is associated with the development of clinical infection in patients
submitted to ACL reconstruction. The secondary goal is to evaluate if the graft preparation
technique and the increased preparation time—between the harvesting and the definitive
implantation—can increase the risk of contamination.
Material and Methods
Between July 2016 and January 2017, prospective data from 124 patients submitted to
primary ACL reconstruction in our institution were collected. All of the patients
who underwent ACL reconstruction with hamstring grafts were enrolled. The exclusion
criteria included previous surgical procedures on the involved knee, the use of contralateral
hamstring tendon, another graft source, and noncompliance with the participation in
the study. The present study was approved by the Ethics Committee under the number
CAAE 48411115.0.0000.5127. All of the participants complied with the participation
and signed the informed consent form before the start of the study. No financial compensation
was offered to the patients in exchange for their participation in the present research.
Surgical Aspects of the Reconstruction of the Anterior Cruciate Ligament
Surgical Aspects of the Reconstruction of the Anterior Cruciate Ligament
During anesthetic induction, the patient was prepared with hair removal from the area
and received a prophylactic dose of cefazolin 2 g or of clindamycin 600 mg, in case
of allergy. The antibiotic therapy was maintained for 24 hours, administered every
8 hours. Skin antisepsis was performed with a soft brush and chlorhexidine soap followed
by the application of an alcoholic solution, always by the same assistant researcher.
After the field preparation, the tourniquet was triggered, and the surgical time count
started. The period between the start of the procedure up to the graft harvesting,
the period between the beginning of the surgery up to the fixation of the graft, the
total surgical time, and the graft preparation time were recorded. The preparation
time included the manipulation of the tendons and the standby time until the implantation,
while the final surgery time was set until the completion of the skin suture and the
tourniquet was turned off.
Three senior surgeons performed all of the reconstructions. Two techniques were used
for the harvesting and the preparation of the grafts during the reconstruction of
the ACL: the traditional technique with hamstring grafts harvest, tibial insertion
release, and free preparation in the auxiliary table (group 1); and the biological
technique, sparing this tibial insertion (group 2). The first technique was performed
by the surgeon de Lúcio Honório de Carvalho Júnior, and the second technique was performed
by the surgeons Eduardo Frois Temponi e Luiz Fernando Machado Soares.
Group 1–Classical Technique
Group 1–Classical Technique
The graft was harvested at the start of the procedure through a longitudinal incision
of between 2 and 3 cm along the distal insertion of the pes anserinus at the proximal
tibia. A 2-0 Vicryl suture (Ethicon Inc., Bridgewater, NJ, USA) was passed at the
most distal part of each tendon – gracile and semitendinosus tendons –, which were
disinserted with a blade and detached from the muscular belly with a closed harvester.
The two grafts were taken to the table by an assistant who removed the muscle part,
adjusted and prepared the borders and measured the tendons. At the same time, the
senior surgeon proceeded with the standard arthroscopy, the joint assessment, the
treatment of associated lesions, and with the preparation of the tunnels ([Fig. 1A]).
Fig. 1 Hamstring grafts preparation. A, group 1–classical technique with free hamstring
graft; B, group 2–technique with fixed hamstring graft.
Group 2–Technique with Tibial Insertion Preservation
Group 2–Technique with Tibial Insertion Preservation
Group 2 underwent the same access to expose the pes anserinus tendons. After their
individualization, the semitendinosus and gracile tendons were removed with an open
harvester. Next, the graft started to be prepared, maintaining its tibial insertion
to obtain a quadruple graft, kept between the fascia and the subcutaneous tissue;
all of the arthroscopy and fixation procedures were performed with the same routine
used in Group 1 ([Fig. 1B]).
Laboratorial Analysis
Two medium-sized fragments, measuring 4 × 4 × 2 mm, were obtained from the leftovers
of one end of the graft in order to not disturb its integrity. The first fragment
was obtained immediately after the preparation of the graft, and the second one was
obtained immediately before its passage for the final fixation; the elapsed time was
recorded. Both samples were immediately put in a sterile vial with 2 ml of saline
solution at 0.9% and sent to the laboratory at the end of the surgery. Each vial was
agitated for 1 minute in a vortex mixer; then, its liquid content was totally aspired
with a thick needle and transferred to a BacT/Alert FA blood culture vial (bioMérieux,
Marcy-l'Étoile, France) with brain-heart infusion (BHI) medium. This vial was incubated
at a BacT/Alert 3D microbial detection system (bioMérieux, Marcy-l'Étoile, France)
for up to 72 hours, and it was submitted to automated analysis every 10 minutes. If
the system accused positive growth, one sample of this material was incubated in a
Petri dish for qualitative evaluation.
The contents of another BHI vial were added to the initial culture vial and again
mixed with the graft fragment in the vortex mixer. Next, the first inoculation was
performed at a triple dish (blood agar, McConkey agar, and Mannitol salt agar). Both
the dish and the graft vial with BHI were incubated at 35.0 ± 2.0° C for 72 hours
in room atmosphere. The visual analysis was performed at 24 and 48 hours, and the
samples were inoculated in a new dish. After 72 hours with no dish or equipment growth,
the result was considered negative.
Data collection and Postoperative Period
Data collection and Postoperative Period
Anthropometric data, age, laterality, gender, comorbidities, presence of associated
joint lesions and 24-hour C-reactive protein levels were collected. No patient used
postoperative drain and the first dressing was kept closed for 48 hours. All of the
patients were discharged in 24 hours and reevaluated in scheduled visits at an outpatient
facility within 7, 14, 45, 90 and 180 days. The diagnosis of septic arthritis was
based on the clinical picture associated with the laboratorial evaluation with complete
blood count, C-reactive protein, erythrocyte sedimentation rate, and synovial fluid
analysis.
Statistical Analysis
The sample size was calculated prior to the study and the minimal number of 56 knees
was defined as required for the statistical treatment (28 in each group), considering
a significance level of 5% and a test power of 80%. Data was presented as mean and
standard deviations (SDs). The categorical data were compared by the chi-squares test
and by the Fischer exact test. After verifying the normality of the continuous variables
with the Kolmogorov-Smirnov test, the mean of the normal variables and the median
of the non-normal variables were calculated. Next, the existence of statistically
significant differences between the groups was determined with the Student t parametric
test for normal variables and with the Mann-Whitney nonparametric test for non-normal
variables. Significance was set at 0.05. The statistical analysis was performed with
IBM SPSS Statistics for Windows, Version 20.0 (IBM Corp., Armonk, NY, USA).
Results
A total of 110 patients were evaluated, with 14 (12.7%) patients lost at follow-up.
Anthropometric data and those related to the surgical procedure are listed in [Table 1]. Fifty-six (50.9%) patients did not present associated lesions, while 26 (23.6%)
presented with an isolated medial meniscal lesion, 18 (16.4%) presented with a lateral
meniscal lesion, 6 (11.0%) presented with lesions in both menisci, and 4 (3.6%) presented
with other lesion patterns. Comorbidities include hypertension, diabetes, hypothyroidism,
depression, asthma, and smoking, without differences between the groups ([Table 2]). The only significant difference between the groups was the harvesting time of
the graft. All of the cultures were negative, and the C-reactive protein levels did
not differ between the groups ([Table 3]).
Table 1
|
Variable
|
Category
|
Group 1
|
Group 2
|
Total
|
p-value
|
Test
|
|
Gender
|
Male
|
30
|
64
|
94
|
0.756
|
Chi-squared
|
|
Female
|
6
|
10
|
16
|
|
|
|
Total
|
36
|
74
|
110
|
|
|
|
Lesion side
|
Left
|
22
|
38
|
60
|
0.572
|
Fisher
|
|
Right
|
14
|
36
|
50
|
|
|
|
Total
|
36
|
74
|
110
|
|
|
|
Associated lesion
|
No lesion
|
20
|
36
|
56
|
0.446
|
Chi-squared
|
|
With lesion
|
16
|
38
|
54
|
|
|
|
Total
|
36
|
74
|
110
|
|
|
|
Septic arthritis
|
Negative
|
34
|
70
|
104
|
0.488
|
Chi-squared
|
|
(clinical infection)
|
Positive
|
0
|
2
|
2
|
|
|
|
Total
|
34
|
72
|
106
|
|
|
Table 2
|
Comorbidity
|
Group 1
|
Group 2
|
Total
|
p-value
|
Test
|
|
No comorbidities
|
Yes
|
24
|
46
|
24
|
ns
|
Chi-squared
|
|
No
|
12
|
28
|
12
|
|
|
|
Smoking
|
Yes
|
4
|
12
|
4
|
ns
|
Chi-squared
|
|
No
|
32
|
62
|
32
|
|
|
|
Hypertension
|
Yes
|
2
|
8
|
2
|
ns
|
Chi-squared
|
|
No
|
34
|
66
|
34
|
|
|
|
Asthma
|
Yes
|
0
|
2
|
0
|
ns
|
Chi-squared
|
|
No
|
36
|
72
|
36
|
|
|
|
Diabetes
|
Yes
|
2
|
2
|
2
|
ns
|
Chi-squared
|
|
No
|
34
|
72
|
34
|
|
|
|
Hypothyroidism
|
Yes
|
0
|
2
|
0
|
ns
|
Chi-squared
|
|
No
|
36
|
72
|
36
|
|
|
|
Depression
|
Yes
|
4
|
2
|
4
|
ns
|
Chi-squared
|
|
No
|
32
|
72
|
32
|
|
|
Table 3
|
Variable
|
Used technique
|
Average
|
Standard deviation
|
p-value
|
|
Graft harvesting time (min)
|
Group 1–classical technique
|
10.1
|
2.7
|
p < 0.05
|
|
Group 2–technique maintaining insertion
|
8.3
|
2.8
|
|
|
24-hour C-reactive protein (mg/dL)
|
Group 1–classical technique
|
8.6
|
5.0
|
ns
|
|
Group 2–technique maintaining insertion
|
13.5
|
11.0
|
|
|
Variable
|
Used technique
|
Median
|
Standard deviation
|
p
-value
|
|
Age (years old)
|
Group 1–classical technique
|
30.50
|
8.8
|
ns
|
|
Group 2–technique maintaining insertion
|
30.00
|
8.3
|
|
|
Weight (kg)
|
Group 1–classical technique
|
84.00
|
10.6
|
ns
|
|
Group 2–technique maintaining insertion
|
78.00
|
14.9
|
|
Height (cm)
|
Group 1–classical technique
|
173.00
|
10.6
|
ns
|
|
Group 2–technique maintaining insertion
|
176.00
|
6.9
|
|
Graft preparation time (minutes)
|
Group 1–classical technique
|
24.50
|
6.0
|
ns
|
|
Group 2–technique maintaining insertion
|
27.00
|
7.0
|
|
Total surgery time (minutes)
|
Group 1–classical technique
|
52.00
|
8.7
|
ns
|
|
Group 2–technique maintaining insertion
|
52.00
|
7.9
|
Two patients from group 2 presented septic arthritis at the postoperative period (1.8%),
in the acute phase. The first was submitted to antibiotic therapy and two more arthroscopies
for debridement. The good condition of the graft was demonstrated in both procedures,
allowing its maintenance. This patient had type 1 diabetes of difficult control, 24-hour
C-reactive protein level of 39 mg/dL, and negative graft cultures. The second case
was that of a healthy patient, also with negative cultures and preserved graft during
the debridement arthroscopy. In both patients, the first debridement culture showed
Staphylococcus epidermidis.
Discussion
The most important finding of the present study was the lack of statistical difference
between both groups regarding the degree of the contamination and the consequent clinical
infection, although 2 patients from group 2 had infections with negative perioperative
cultures. Another finding was that the graft preparation and fixation time did not
interfere with the contamination rates. Two cases of infection were described in group
2 (1.8%), both diagnosed at the acute phase and timely treated to avoid major damage
to the grafts. One case belonged to a patient with type 1 diabetes and problems with
blood sugar control at the immediate postoperative period, which can contribute to
infections; accordingly, some studies consider diabetes as an exclusion criterium.[11]
[21]
[22] We cannot prove that the source of the infection was the graft, since all its cultures
were negative. The contamination rate of the graft cultures from the present study
was 0%, different than those observed in similar trials ([Table 4]).[11]
[21]
[23]
[24]
[25] There is a great variation in the described methods of antisepsis, of culture, and
of incubation. Some authors used an iodine-based antiseptic,[21]
[25] others did not state which antiseptic was used,[11]
[22]
[23] some requested anaerobic cultures[11]
[23] and, in 1 trial, the incubation period was of up to 14 days.[24] It is known, for instance, that factors such as chlorhexidine use can influence
the number of positive cultures, since this agent is proven to reduce postoperative
infection indexes and skin colonization.[26]
[27]
[28]
Table 4
|
Author
|
Total N
|
Positive cultures/%
|
Organism (n)
|
Clinical infections/%
|
|
Hantes et al., 2008[11]
|
30
|
4/13%
|
Staphylococcus aureus (2)
|
0
|
|
Acinetobacter (1)
|
|
Staphylococcus epidermidis (1)
|
|
Gavriilidis et al., 2009[23]
|
89
|
9/10%
|
S. epidermidis (2)
|
0
|
|
Enterococcus (2)
|
|
Staphylococcus capitis (1)
|
|
Peptostreptococcus (1)
|
|
Corynebacterium (1)
|
|
Bacillus cereus (1)
|
|
Propionibacterium granulosum (1)
|
|
Plante et al., 2013[24]
|
30
|
7/23%
|
S. aureus (1)
|
0
|
|
Streptococcus viridians (1)
|
|
Corynebacterium (1)
|
|
Staphylococcus no aureus (1)
|
|
Lactobacillus (1)
|
|
Propionibacterium acnes (1)
|
|
Escherichia coli (1)
|
|
Nakayama et al., 2012[21]
|
50
|
1/2%
|
Bacillus sp. (1)
|
1/2%
|
|
Barbier et al., 2015[25]
|
25
|
3/12%
|
Staphylococcus hominis (1)
|
0
|
|
Staphylococcus capitis (1)
|
|
Candida parapsilosis (1)
|
|
Bradan et al., 2016[22]
|
60
|
10/16.7%
|
Staphylococcus epidermidis (4)
|
0
|
|
Staphylococcus aureus (2)
|
|
Acinetobacter (2)
|
|
Bacillus spp. (1)
|
|
Citrobacter spp. (1)
|
|
(present study)
|
110
|
0/0
|
No growth
|
2/1.8%
|
The microbiological analysis of the graft in the real use conditions of the trials,
with no contaminating or decontaminating intervention, shows positive cultures ranging
from 2 to 23% in the hamstring tendons.[11]
[21]
[23]
[24]
[25]
[29] Among these studies, only Nakayama et al.,[21] observed a case of positive graft culture and postoperative septic arthritis in
the same patient. However, the contaminating organism, methicillin-sensitive Staphylococcus aureus (MSSA), was different from the organism isolated from the infection, methicillin-resistant
S. aureus (MRSA) ([Table 4]). This lack of correlation between the clinical infection and the perioperative
microbiology can suggest that the contamination of the prepared graft with no complications
has a minor causative role in the current concept of ACL surgical reconstruction.
Regarding the supplementary evaluation, Hantes et al.,[11] found no differences in C-reactive protein levels from either contaminated or non-contaminated
grafts in the first 24 hours after the procedure. A similar situation was observed
by Graviilidis et al.,[23] who found an elevated C-reactive protein level at the 4th day, and normal values at the 20th day, but both with no differences between groups with either contaminated or non-contaminated
grafts, and by Bradan et al.,[22] who did not observe differences in the C-reactive protein levels between groups
with or without graft contamination at 7, 12 and 20 days after the surgery. These
trials, however, did not present clinical infection cases, in which a significant
increase in C-reactive protein levels can be noted.[1]
[5]
[6]
[10] Likewise, no significant variation was observed between 24-hour C-reactive protein
values in the present study.
Regarding the time variable, Hantes et al.[11] obtained an average time for the preparation of the hamstring graft of 19 (16–21)
minutes, and of 30 (28–43) minutes between the harvesting of the graft and its implantation.
These authors compared the bone-patellar tendon-bone graft preparation and implantation
times and found a significantly lower time for the patellar preparation, but with
no difference when comparing the implantation time. As such, they did not confirm
their hypothesis that a higher preparation time would be responsible for a higher
incidence of graft contamination of the hamstring tendon. Gavriilidis et al.[23] presented an average preparation time of 16 ( ± 2) minutes, and the average time
between the harvest of the hamstring graft and the implantation was 20 ( ± 2) minutes.
In addition, the sample from this study was not big enough to confirm the time and
higher risk of contamination hypothesis, but the authors state that there is a major
correlation in this association. Judd et al.[30] also affirmed that the time period between the harvest of the hamstring tendon and
the knee implantation is undoubtedly an important criterium for a possible contamination.
However, their study does not present numbers that confirm this hypothesis. The present
study compared two techniques for the harvest and the preparation of hamstring tendons,
and the only significant relationship was between the variable graft harvesting time
and the technique used (p = 0.038), indicating that individuals submitted to the classical technique (group
1; 10.1 minutes) presented a higher average graft harvest time compared with the ones
who underwent the technique with insertion preservation (group 2; 8.3 minutes). This
fact, however, was not related to a high graft exposure or to a higher risk of contamination.[23]
[30] There was no difference between both techniques regarding the time between the start
of the surgery until the fixation of the graft, the total surgery time, and the graft
preparation time. As such, the influence of the time variable cannot be considered
different regarding the possibility of contamination and/or infection.
Several limitations can be described in the present study. The adopted microbiological
protocol used for samples with no evidence or suspicion of anaerobic organisms prevented
the identification of certain pathogens typically found at the skin, as previously
reported.[23] The samples collected and stored in saline solution were kept in the surgical room
until the end of the procedure (∼ 30–40 minutes), which can be a factor of reduction
in the sensitivity of the test. The present study was conducted as a clinical activity,
applying actual procedures from the routine of the hospital, which prevented long
incubations or the use of culture medium and atmosphere for anaerobic organisms. Other
works evaluating cultures from “sterile” graft samples performed incubation until
5 to 14 days, while we did it for only 3 days, reducing the chance of identification
of fastidious organisms. Another limitation was the lack of laboratorial follow-up
during the postoperative period, which would increase costs and modify the clinical
routine. Despite the contamination rate of 0% in the present study, our findings reaffirm
important steps in the prevention of surgical infections, such as skin antisepsis,
antibiotic prophylaxis, that the surgery should be performed by a senior surgeon,
and meticulous graft protection, especially when the implantation is delayed. Through
a better understanding of the factors regarding graft contamination, prevention measures
for infectious diseases can be implemented. Moreover, further studies will be able
to confirm the actual contribution of factors such as surgical aggression, graft fixation
device, the number of assistants, drain use, operative wound care, associated diseases,
and preoperative sports activities.
Conclusion
Based on the obtained results, there was no association between graft contamination
and its preparation time or technique, neither between intraoperative contamination
and the development of clinical infection.
