Patients and bacteria strains
Patient clinical information was collected, and a total of 93 strains of non-repetitive S. maltophilia were isolated. Among them, 30 strains of S. maltophilia were isolated from the ICU, and the rest were from a range of hospital departments (Figure 1). The rest included 13 strains from neurosurgery, 9 strains from emergency internal medicine, 8 strains from cadre health care, 6 strains from cardiovascular surgery, 5 strains from hematology, 4 strains from liver surgery, 3 strains from oncology, 3 strains from nephrology, 2 strains from emergency medicine, 2 strains from neurology, 2 strains from cholangio-pancreatic surgery, 1 from thoracic surgery, 1 from gastrointestinal surgery, 1 from respiratory medicine, 1 from digestive medicine, 1 from general surgery, and 1 from urology.
Among the patients with S. maltophilia infections, 62 males (66.7%) and 31 females (33.3%) were included. A total of 73 patients were aged 60 years and above (78.5%). Before bacterial isolation, 35 patients (37.6%) were subjected to invasive examinations or treatments. Among all the S. maltophilia strains, there were 81 strains (87.1%) isolated from sputum, 7 strains (7.5%) from drains, 2 strains from pleural effusion, 1 strain from ascites, 1 strain from urine, and 1 strain from blood.
Of the 93 patients, 7 (7.5%) had no basic diseases but primarily had fractures and malnutrition, and 86 (92.5%) had one to five underlying diseases. Of these, 25 had malignant tumors, 24 had hypertension, 12 had coronary heart disease, 10 had renal insufficiency, 5 had leukemia, 14 had head trauma, 20 had chronic bronchitis or pneumonia, and 12 had liver injury. All of the patients had a history of antibiotic use (1-6 types), with an average of three antibiotics per person prior to isolation of S. maltophilia, of which 69 had used three or more antibiotics. The antibiotics primarily included cephalosporins (65/93), carbapenem (53/93), enzyme inhibitors (51/93), quinolones (35/93), glycopeptide (22/93), aminoglycosides (13/93), and tetracyclines (9/93) (Table 1).
Table 1. The clinical characteristics of adult and pediatric patients
|
Adult(n=93)
|
Demographics
|
|
Age(year, average, range)
|
66.3 (16-99)
|
Gender: male
|
62 (66.7%)
|
|
|
Baseline diseases, n(%)
|
|
Hypertension
|
24 (25.8)
|
Heart Disease
|
12 (12.9)
|
Malignancy
|
25 (26.9)
|
Pulmonary Disease
|
20 (21.5)
|
Liver Disease
|
12 (12.9)
|
Leukemia
|
5 (5.4)
|
Head trauma
|
14 (15.1)
|
|
|
Strain isolation n(%)
|
|
ICU
|
30 (32.3)
|
Sputum
|
81 (87.1)
|
|
|
Invasive operation n(%)
|
35 (37.6)
|
|
|
Previous antibiotics usage n(%)
|
|
The number of antibiotics ≥3
|
69 (74.2)
|
Cephalosporins
|
65 (69.9)
|
Carbapenems
|
53 (57.0)
|
Enzyme Inhibitors
|
51 (54.8)
|
Quinolones
|
35 (37.6)
|
Glycopeptides
|
22 (23.7)
|
Aminoglycosides
|
13 (14.0)
|
MLST analysis
The distribution of the clonal typing of S. maltophilia was relatively scattered. According to the different alleles, the strains were assigned to 61 sequence types. Among them, 45 types of the 60 strains were different from those published on the PubMLST database (recorded as STnew1-STnew45). The other 33 strains consisted of existing types in the database, of which a relatively larger number was ST23 (n=8). There were also strains of ST5 (n=3), ST15 (n=3), ST24 (n=3), ST3 (n=2), ST84 (n=2), ST89 (n=2), and ST99 (n=2) and ST4, ST8, ST13, ST36, ST77, ST98, ST102, and ST112. The eight S. maltophilia strains of ST23 were distributed in five different departments, and the 30 strains of S. maltophilia isolated from the ICU were classified into 24 sequence types. S. maltophilia strains of the exact same sequence types were not collected in other departments, indicating that there was no obvious clonal transmission of S. maltophilia infections in this study [1]. The detailed results are shown in Figure 2.
Virulence gene detection
The results of the virulence gene detection showed that the carriage rates of the four virulence genes were 79.6% (74/93) for Stmpr1, 91.4% (85/93) for Stmpr2, 94.6% (88/93) for Smf-1, and 52.7% (49/93) for Smlt3773. There were 31 strains of S. maltophilia that carried all four of the genes.
Analysis of drug resistance
The resistance rates of S. maltophilia to levofloxacin and TMP/SMX were 4.3% and 9.7%, respectively. All of S. maltophilia strains were sensitive to minocycline. Among these strains, one strain, numbered ji82, was resistant to both TMP/SMX and levofloxacin.
Biofilm forming ability
The average biofilm forming ability of S. maltophilia was OD492=0.54 ± 0.49 (0.044–2.34). The OD values of S. maltophilia isolated from the male and female patients were OD492 of 0.52 ± 0.51 and OD492 of 0.57 ± 0.47, respectively, and there was no significant difference between the two gender groups. There was no significant difference in the biofilm formation ability between people aged 60 and above and those under 60 years old, as shown in Figure 3. In addition, the drug resistance and biofilm forming ability of the strains were analyzed, and there was no obvious correlation between the drug-resistant phenotype and the biofilm forming ability, as shown in Table 2. The carrying rates of the three biofilm genes rmlA, spgM, and rpfF were 82.8% (77/93), 92.5% (86/93), and 64.5% (60/93), respectively. The point mutations of the spgM gene in the strains with strong biofilm forming abilities were relatively consistent and significantly different from those with weak biofilm forming abilities. The detailed sequencing results of some strains are shown in Figure 4 (the bases of the two strains with different biofilm forming abilities were selected as the representatives). However, the other two biofilm genes did not have obvious point mutations in the strains with different biofilm forming abilities.
Table 2. Drug-resistant rates and relationship between the drug resistance and biofilm formation
Antibiotics
|
Resistant rate
|
Pearson's correlation
|
levofloxacin
|
4.3%
|
0.02
|
sulfamethoxazole
|
9.7%
|
0.04
|
piperacillin / tazobactam
|
8.6%
|
0.1
|
cefoperazone / sulbactam
|
4.3%
|
0.08
|
minocycline
|
0%
|
NA
|
The carriage of the virulence genes
The carriage of the four virulence genes Stmpr1, Stmpr2, smf-1,and Smlt3773locus were 79.6%, 91.4%, 94.6%, and 52.7%, respectively.
Analysis of the risk factors in ICU patients infected with the S. maltophilia
By using a univariate analysis, it was concluded that the changes in lymphocytes, albumin, and the use of antibiotics were infection risk factors in the ICU patients (Table 3). After the multivariate analysis, the type of antibiotic use and lymphocyte count were found to be independent risk factors of infection with S. maltophilia, and the lymphocyte count was used as a common index for a routine blood examination (Table 4). These findings may be used as a new reference index for clinical sensitivity and control of S. maltophilia.
Table 3. Univariate analysis of risk factors of S. maltophilia infections in the ICU
Items
|
Patients (n=30)
|
Control (n=60)
|
P value
|
OR(95%CI)
|
male (sex)
|
23 (76.7%)
|
38 (63.3%)
|
0.263
|
0.565 (0.208-1.534)
|
Age (years)
|
64.8 ± 19.1
|
65.5 ± 16.9
|
0.873
|
|
leukocyte
|
11.5 ± 5.4
|
10.9 ± 4.1
|
0.777
|
|
neutrophil
|
9.4 ± 4.9
|
9.4 ± 3.9
|
0.767
|
|
lymphocyte
|
1.3 ± 0.9
|
0.9 ± 0.4
|
0.012
|
|
monocyte
|
0.7 ± 0.5
|
0.5 ± 0.4
|
0.536
|
|
albumin
|
30.6 ± 4.2
|
28.3 ± 5.7
|
0.033
|
|
globulin
|
29.0 ± 6.4
|
28.2 ± 7.0
|
0.286
|
|
prealbumin
|
129.0 ± 52.3
|
124.8 ± 49.9
|
1.000
|
|
surgeries
|
14 (46.7%)
|
27 (45.0%)
|
0.496
|
0.724 (0.286-1.835)
|
organ transplantation
|
5 (16.7%)
|
9 (15.0%)
|
0.987
|
0.990 (0.296-3.310)
|
malignant tumor
|
8 (26.7%)
|
15 (25.0%)
|
0.894
|
0.933 (0.337-2.585)
|
hypertension
|
7 (23.3%)
|
15 (25.0%)
|
0.923
|
0.949 (0.328-2.748)
|
diabetes
|
3 (10.0%)
|
9 (15.0%)
|
0.397
|
0.547 (0.135-2.213)
|
pulmonary infection
|
9 (30.0%)
|
16 (26.7%)
|
0.990
|
1.007 (0.374-2.712)
|
cardiopathy
|
4 (13.3%)
|
9 (15.0%)
|
0.841
|
0.875 (0.238-3.213)
|
liver injury
|
4 (13.3%)
|
7 (11.7%)
|
0.972
|
1.024 (0.272-3.856)
|
trachea intubation
|
12 (40.0%)
|
21 (35.0%)
|
0.941
|
1.036 (0.406-2.640)
|
chemotherapy
|
2 (6.7%)
|
3 (5.0%)
|
0.843
|
1.205 (0.189-7.681)
|
immunosuppressor
|
9 (30.0%)
|
17 (28.3%)
|
0.867
|
0.920 (0.343-2.464)
|
number of antibiotics
|
3.6 ± 1.2
|
3.0 ± 1.1
|
0.029
|
|
carbapenems
|
21 (70.0%)
|
40 (66.7%)
|
0.731
|
1.187 (0.445-3.167)
|
cephalosporins
|
20 (66.7%)
|
45 (75.0%)
|
0.604
|
0.771 (0.289-2.059)
|
quinolones
|
16 (53.3%)
|
30 (50.0%)
|
1.000
|
1.000 (0.396-2.523)
|
Table 4. Multivariate logistic regression analysis associated with S. maltophilia infections in the ICU
Risk factors
|
B value
|
Wals
|
P value
|
OR value
|
95% CI
|
lower limit
|
upper limit
|
lymphocyte
|
1.077
|
4.208
|
0.04
|
2.937
|
1.049
|
8.222
|
albumin
|
0.099
|
3.05
|
0.081
|
1.104
|
0.988
|
1.234
|
antibiotics
|
0.596
|
5.956
|
0.015
|
1.814
|
1.124
|
2.927
|
|
|
|
|
|
|
|
|