Sensitivity profile of blood culture isolates in a clinical analysis laboratory, Fortaleza, CE

Perfil de sensibilidade de isolados de hemocultura em um laboratório de análises clínicas, Fortaleza, CE

Perfil de sensibilidad de hemocultivos aislados en un laboratorio de análisis clínicos, Fortaleza, CE

Esta obra está bajo una Licencia Creative Commons Atribución 4.0 Internacional.

Bacteremia is associated with the presence of bacteria in the bloodstream, which may have a primary (direct entry into the bloodstream via needles, contaminated infusions, catheter, etc.) or secondary origin (from a primary focus of infection, with possible hematogenous or lymphatic spread), having great clinical importance due to the high morbidity and mortality when associated with sepsis¹, considering the increase in sepsis cases in Brazil². According to the Global Sepsis Alliance (GSA), sepsis can be caused by blood infections, urinary tract infections, lung infections, intestinal infections, and skin infections, being characterized by an inflammatory reaction that affects organs and systems from infections caused by viruses, bacteria, fungi and parasites³.

Given the increase in cases of bacteremia and sepsis, blood culture has emerged as the main laboratory methodology for identifying microorganisms, enabling the assessment of the sensitivity profile of clinical isolates^4,5. The main microorganisms present in bacteremia differ in terms of age. In children, the most reported cases are caused by Streptococcus pneumoniae, Neisseria meningitidis and Haemophilus influenzae⁵. In adult patients, the most frequent isolates are Staphylococcus aureus, Staphylococcus epidermidis, Enterococcus spp., S.pneumoniae, Escherichia coli, Pseudomonas aeruginosa, Klebsiella spp⁶. and Acinetobacter baumannii⁷.

The main blood culture isolates show considerable resistance to antibiotics of clinical importance⁷. Thus, it is possible to observe that antimicrobial resistance patterns have been altered and multidrug-resistant bacteria have increased alarmingly, resulting in serious infections with complicated treatments⁸. Antimicrobial resistance is a characteristic that can be intrinsic or acquired horizontally, through the acquisition of resistance genes, or through spontaneous mutations⁹. This resistance is an inevitable evolutionary process¹⁰, whose species with the highest frequency of antimicrobial resistance are S. aureus, Enterococcus faecium, E. faecalis, S. pneumoniae, E. coli, K. pneumoniae, P. aeruginosa and A. baumannii¹¹. Duval et al (2019) highlight the main multidrug-resistant microorganisms of global importance, such as carbapenem-resistant A. baumannii, carbapenem-resistant P. aeruginosa, and 3^rd generation carbapenem- cephalosporin-resistant Enterobacterales .K. pneumoniae, E. coli, Enterobacter spp., Serratia spp. and Proteus spp.)¹².

Identifying the pattern of antimicrobial resistance makes it possible to support empirical treatments⁴, since the release of blood cultures is a slow process, often unable to provide support and rapid therapeutic interventions. As a result, empirical treatment ends up being the only immediate solution, but empirical treatments end up being performed with broad-spectrum antibiotics, harming the patient due to drug toxicity and aiding in the selection of multidrug-resistant microorganisms⁶.

In Brazil, the main bacteria of clinical importance are non-fermenting bacteria, Extended Spectrum Beta-lactamase (ESBL) producing enterobacteria and methicillin-resistant S. aureus. Those related to cases of bacteremia are S. aureus, coagulase-negative Staphylococcus, Enterococcus spp, E. coli and Klebsiella spp⁷. Therefore, the present study aims to assess the main bacterial isolates in blood cultures and a possible change in their sensitivity profiles in a clinical analysis laboratory in Fortaleza, Ceará.

This is an epidemiological, descriptive, retrospective study with a quantitative approach of outsourced blood cultures from patients admitted to a tertiary hospital, a reference in Fortaleza, featuring 23 specialties, with 296 beds and eight operating rooms equipped with laminar flow.

This research was carried out taking into account the guarantee of the ethical and legal principles that govern research on human beings, recommended in Resolution 466/2012 of the Brazilian National Health Council (CNS), approved by the Research Ethics Committee, with CAAE (Certificadode Apresentação para Apreciação Ética - Certificate of Presentation for Ethical Consideration) 42401920.0.0000.5049.

Samples were collected from January 2019 to December 2020, and analyzed in a clinical analysis laboratory in Fortaleza, Ceará. Patients of both sexes without age restriction who were admitted to the hospital were included.

Patient data was collected in EpiCenter® system from the laboratory’s database, along with software linked to BD Phoenix™, MicroScan and AutoSCAN-4. The results were based on plotting performed in EpiCenter® software, dividing Gram-positive cocci (GPC) and Gram-negative bacilli (GNB) into groups, selecting the seven main microorganisms isolated from each group and performing the comparison through the elaboration of tables and graphs, demonstrating the total number of microorganisms isolated and analyzing the sensitivity profiles of the three main microorganisms of each group by drug tested. In addition to the aforementioned analyses, the analysis of the amounts of positive blood cultures and contaminants from 2019 and 2020 was carried out. The drugs used to compare the sensitivity profile of GNB and GPC were pre-established according to BrCast, version 9.0 of 2019, valid from 02/04/2019, assessing the minimum inhibitory concentration (MIC) of each drug to the respective identified microorganism¹³.

In the study, all samples of positive blood cultures and confirmed by the responsible laboratory were included. Through microbiological analysis, antimicrobial susceptibility tests were performed, following the standardization institution BrCast, classifying sensitive only the values referring to the usual doses.

The analysis of the sensitivity profile of the classes of antibiotics was performed through the arithmetic mean of the individual percentages of each drug against the species analyzed using Graphpad 7.0. The assessment of the change in antibiotic sensitivity profiles in 2019 and 2020 was compared using the chi-square test with Yates’ correction (α= 5%), using Graphpad 7.0, considering an increase or decrease in sensitivity if p-value is above 0.05.

During the research period, 840 microorganisms isolated from blood cultures of patients admitted to a tertiary hospital from January 2019 to December 2020 were identified.

In 2019, 64% (254/397) GNB and 36% (143/397) GPC were isolated from blood cultures. In 2020, 43.3% (192/443) GNB and 56.6% (251/443) GPC were isolated. The main isolates from the GNB group in 2019 were E. coli, 34.6% (88/254) K. pneumoniae, 29.1% (74/254), and P. aeruginosa, 18.5% (47/254). The main GPC isolates in 2019 were S. epidermidis, 39.8% (57/143), S. aureus, 24.4% (35/143), and S. haemolyticus, 12.6% (18/143).

In 2020, the main GNB isolates were E. coli, 31.2% (60/192), K. pneumoniae, 26.5% (51/192) and P. aeruginosa, 23% (44/192), while those of GPC were S. epidermidis, 51.4% (129/251), S. haemolyticus, 19.5% (49/251), and S. hominis, 13.5% (34/251), showing a change in prevalence between 2019 and 2020.

In 2019, the main GNB isolates showed the following sensitivity profiles: E. coli showed lower sensitivity to sulfonamides (53.85%), monobactams (58.82%) and penicillin (62.01%); K. pneumoniae showed lower sensitivity to phosphomycin (0%), to monobactams (58.82%) and to penicillin (34.09%); P. aeruginosa was less sensitive to phosphomycin (0%), glycylcycline (0%) and sulfonamides (0%). While GPC isolates showed the following sensitivity profile: S. epidermidis showed lower sensitivity to cephalosporins (7.78%), penicillin (15.05%) and quinolones (39.09%); S. aureus showed lower sensitivity to penicillin (41.03%), cephalosporins (72.92%) and quinolones (92.68%); and S. haemolyticus showed lower sensitivity to penicillin (16.67%), cephalosporins (33.33%) and aminoglycosides (37.50%) (Table 1).

In 2020, the main GNB isolates showed the following sensitivity profiles: E. coli showed lower sensitivity to monobactams (57.14%), penicillin (57.97%) and sulfonamides (59.38%); K. pneumoniae showed lower sensitivity to penicillin (46.43%), to monobactams (62.50%) and to sulfonamides (62.96%); P. aeruginosa showed lower sensitivity to phosphomycin (0%), sulfonamides (0%) and glycylcycline (6.25%). In GPC isolates, S. epidermidis showed a lower sensitivity to penicillin (17.70%), quinolones (38.07%) and sulfonamides (38.61%); S. aureus showed a lower sensitivity to penicillin (75.76%); and S. haemolyticus showed lower sensitivity to penicillin (8.14%), quinolones (25.74%) and aminoglycosides (32.43%). (Table1).

Comparative assessment of the change in sensitivity profile was observed for the main GNB and GPC isolates from blood cultures, from January 2019 to December 2020. As for GNB, E. coli isolates showed a decrease in their percentage of sensitivity to amikacin, amoxicillin-clavulanate, aztreonam, cefuroxime, ciprofloxacin, phosphomycin with G6P, gentamicin levofloxacin and tigecycline. However, no statistical difference was observed in susceptibility in the other antibiotics tested. K. pneumoniae isolates showed a percentage increase in the susceptibility of their isolates, comparing the two years, as 2019 isolates are less susceptible to the antibiotics tested. A change was observed in the susceptibility of K. pneumoniae to cefazolin (p=1.14), cefepime (p=0.39), ceftazidime (p=0.20), ceftriaxone (p=0.56), ertapenem (p=0.10), gentamicin (p=3.36), imipenem (p=2.34), meropenem (p=1.91), tigecycline (p=0.09), all representing an increase in sensitivity when compared to the years 2019 and 2020. P. aeruginosa isolates showed a decrease in the percentage of their sensitivity to amikacin and aztreonam (Table 2). A change in the susceptibility of P. aeruginosa to imipenem (P=3.54) was observed, representing an increase in sensitivity when compared to the years 2019 and 2020.

GPC isolates such as S. epidermidis demonstrated a percentage decrease in sensitivity to ciprofloxacin, clindamycin, erythromycin, levofloxacin, minocycline, oxacillin, rifampicin and trimethopim-sulfamethoxazole. A change in the susceptibility of S. epidermidis to minocycline was observed (p=0.01), representing a decrease in sensitivity when compared to the years 2019 and 2020. S. aureus isolates showed a decrease in sensitivity to clindamycin and erythromycin. An alteration was observed in the susceptibility of S. aureus to penicillin G (p=3.41), representing an increase in sensitivity when compared to the years 2019 and 2020. S. haemolyticus isolates showed a decrease in the percentage sensitivity to amoxicillin-clavulanate, ampicillin-sulbactam, ceftriaxone, ciprofloxacin, clindamycin, chloramphenicol, levofloxacin, rifampicin and trimethoprim-sulfamethoxazole. However, no statistical difference was observed in susceptibility in other antibiotics tested (Table 3).

Table 1
Sensitivity rate to the classes of antibiotics used for Gramnegative and Grampositive bacilli compared to the main isolates from blood cultures in 2019 and 2020

Table 2
Comparison of antimicrobial susceptibility profiles of the main Gramnegative bacilli GNB isolated from blood cultures between 2019 and 2020

Note: The main results of the minimum inhibitory concentration were cefazolin (≤ 4), cefepime (≤ 0.125), ceftazidime (≤ 0.5), gentamicin (≤ 2), imipenem (≤ 1), meropenem (≤ 0.125) and tigecycline (≤ 2), for K. pneumoniae, and imipenem (≤ 4), for P. aeruginosa.

Table 3
Comparison of antimicrobial susceptibility profiles of the main Gram-positive cocci (GPC) isolated from blood cultures, between 2019 and 2020

Note: the main results of the minimum inhibitory concentration were minocycline (≤ 0.25), for S. epidermidis, and penicillin G (≤ 0.125), for S.aureus.

The rate of positivity and contaminants in blood cultures is variable when comparing 2019 and 2020. In 2019, 4% (10/233) of the isolates were contaminants and 95% (223/233) were blood cultures confirmed as positive. In 2020, 16% (49/309) of the isolates were contaminants and 84% (260/309) were blood cultures confirmed as positive.

In a study carried out with six hospitals in Cascavel, Paraná, it was observed that the main microorganisms isolated in blood cultures were S. aureus (17.94%), S. epidermidis (16.26%), K. pneumoniae (14.52%), E. coli (8.97%), A. baumannii (8.81%) and P. aeruginosa (8.32%) ¹⁴.

In another study carried out at the Hospital das Clínicas of the UniversidadeFederal de Pernambuco (UFPE), Recife, it was identified that 57.64% of the blood culture isolates were Gram-positive bacteria, while 42.36% were Gram-negative bacteria. The main isolates were S. aureus (21.62%), coagulase negative Staphylococcus (20.54%), K. pneumoniae (9.72%), E. coli (7.56%) and A.baumannii (6.48%)¹⁵.

In a study carried out in a tertiary hospital in Minas Gerais, it was shown that 40.6% of those isolated were S. epidermidis, 17.2%, S. aureus, 7.8%, Enterobacter spp. and 6.3% of Pseudomonas spp.¹⁶. These studies are similar to what was found in the present study, in 2020, in which the main isolates were S. epidermidis (29.12%), E. coli (13.54%), K. pneumoniae (11, 51%), S. haemolyticus (11.06%), P. aeruginosa (9.93%) and S. hominis (7.67%).

It is possible to observe, considering the analyzed studies, that Gram-negative isolates showed resistance against ampicillin-sulbactam, ceftazidime, ciprofloxacin and levofloxacin. K. pneumoniae isolates are less sensitive to aztreonam, cefepime, ceftriaxone, ceftazidime, ciprofloxacin; E. coli showed a lower percentage of sensitivity to cefazolin and cefepime; and P. aeruginosa had a lower percentage of sensitivity to polymyxin b, ceftazidime and aztreonam¹⁵. Another study shows that the isolates belonging to the GPC group showed the following sensitivity pattern: 15.4% of S. epidermidis are sensitive to oxacillin; 92.3% are sensitive to vancomycin; 92.3 and 27.3% of S. aureus are sensitive to oxacillin; and 100% are sensitive to vancomycin¹⁶.

There is a divergence of data when we compare the assessment of this sensitivity profile of the main isolates, with only the data observed for the GPC group and, individually, for S. epidermidis and S. aureus agreeing. This divergence regarding the sensitivity profile may occur due to the difference in date between the works and due to the state of study of each work.

Some of the data cited may represent an analytical error in the laboratory or an error in sending the results to the software, as in the case of vancomycin-resistant S. aureus, isolated in 2019, and tigecycline-sensitive P. aeruginosa.

Clinical isolates from blood cultures showed a change in the sensitivity profile between 2019 and 2020. The sensitivity percentage assessment is changed according to the microorganism in question.

In GNB, when analyzing E. coli, it is possible to infer an increase in resistance against penicillin, monobactams, quinolones and phosphomycin, while P. aeruginosa shows an increase in resistance against monobactams, and K. pneumoniae showed a decrease in the resistance of their isolates, considering that all 2019 isolates are more resistant than those of 2020. While in GPC, S. epidermidis isolates show a considerable increase in their resistance against quinolones, macrolides, lincosamines and against a tetracycline (minocycline). S. aureus isolates showed an increase in sensitivity, given that the 2019 isolates were more resistant. In contrast, S. haemolyticus isolates showed increased resistance against penicillin, quinolones, lincosamines, amphenicols and macrolides.

The change in the sensitivity profile of the main clinical isolates from blood cultures is broad, taking into account that, for some species, this change resulted in an increase in the sensitivity of these isolates. In contrast, the species, with an increase in resistance observed, were against antibiotics of extreme clinical importance, resulting in the use of more potent broad-spectrum antibiotics.