Analysis On The Predominant Bacterial And Fungal Species In Blood Infection And Effect Of Verigene-DNA Chip For Rapid Bacterial Detection

ObjectiveTo investigate the species of bacteria and fungi in blood infection from patients in the Third Hospital of Hangzhou from2008to2014, and to determine the effect of Verigene-DNA chip for rapid detection of Gram-positive bacteria in blood infection.MethodsBy using BacT/ALERT3D blood sample culture system as well as the different culture models of single sample-single flask, single sample-double flasks or double samples-double flasks, the isolated cultures of bacteria and fungi in the peripheral blood samples from19789patients were obtained. The pure-cultured microbes were identified using VITEK2Compact system. The susceptibility of the bacterial and fungal isolates against different antibiotics was determined by both VITEK2Compact system and slip diffusion method. A slip diffusion-based confirmatory test recommend by CLSI was used to detect the β-lactamase activity of the bacterial isolates. The positive isolated culture rates with different blood sample culture models, predominant infected bacteria and fungi, and drug susceptibility of bacterial and fungal isolates were analyzed using statistic softwares. Furthermore, the Verigene-DNA chip was applied to identify the isolated98Gram-positive bacteria as well as detect the drug resistance genes in the isolates. The result of bacterial strain identification by the chip was compared to those by VITEK2Compact system while the result of drug resistance gene detection was checked by PCR.ResultsIn the19789peripheral blood samples, the positive rate of blood sample culture with the model of double samples-double flasks was significantly higher than that with the model of single sample-single flask or single sample-double flasks (p<0.01). In the1876bacterial isolates belonging to124species, the isolating rates of Staphylococcus hominis, Escherichia coli, Staphylococcus aureus, Staphylococcus epidermidis, Staphylococcus haemolyticus, Klebsiella pneumonia, Staphylococcus capitis, Enterococcus faecium, Acinetobacter baumannii and Enterococcus faecalis were11.46%-3.00%, respectively, and the isolating rates of other bacterial isolates belonging to114species were lower than2.3%. In the136fungal isolates belonging to18species, the isolating rates of Candida parapsilosis, Candida albicans, Candida glabrata and Candida guilliermondii were25.00%,21.32%,16.91%and11.76%, respectively, and the isolating rates of other fungal isolates belonging to14species were lower than8%. All the isolates of Staphylococcus species were susceptible to linezolid, vancomycin and tigecycline, but the detection rate (87.88%,116/132) of methicillin-resistant coagulase negative staphylococcus (MRCoNS) was higher than that (54.77%,109/199) of methicillin-resistant S. aureus (MRSA)(p<0.01). All the92isolates of E. faecium and56isolates of E. faecalis wre susceptible to tigecycline while the susceptible rates of the isolates of E. faecium and E. faecalis against linezolid were97.01%and95.56%, respectively. However, the detection rate of vancomycin-resistant enterococcus (VRE) in the isolates of E. faecium or E. faecalis was15.22%(14/92) or8.93%(5/56). In the208isolates of E. coli, the susceptible rates against imipenem, ertapenem, tigecycline, cefotetan, amikacin, piperacillin/tazobactam and nitrofurantoin were92.31%-99.52%, respectively, but1.03%and0.48%of the isolates were resistant to carbopenem antibiotics ertapenem and imipenem. In addition,54.31%of the E. coli isolates were detectable for β-lactamase activity. In the122iaolates of K. pneumonia, the susceptible rates against amikacin, cefotetan, tigecycline and SMZ-TMP were77.05%-79.51%, but32.71%and31.15%of the isolates were were resistant to carbopenem antibiotics ertapenem and imipenem. Moreover,28.21%of the K. pneumonia iaolates were detectable for β-lactamase activity. In the71isolates of A. baumannii, the susceptible rates against polymyxin B, tigecycline, amikacin and cefoperazone/sulbactam were60%-100%, respectively. All the29isolates of C. albicans were susceptible to fluconazol, itraconazole, clotrimazole,5-flucytosine, mycostatin and amphotericin B. The susceptible rates of34C. parapsilosis isolates against5-flucytosine and mycostatin are100%, but those against amphotericin B, itraconazole, fluconazol and clotrimazole were85.71%-97.06%, respectively. All the23C. glabrata were susceptible to5-flucytosine, amphotericin B and mycostatin, but the susceptible rates of the isolates against clotrimazole, itraconazole and fluconazol were56.52%-85.71%, respectively. The susceptible rates of16C. guilliermondii isolates against both5-flucytosine and amphotericin B were93.75%, but those of the isolates against both fluconazol and itraconazole were as low as6.25%. On the other hand, the total coincidence of identification results for98Gram-positive bacterial isolates belonging to16species using Verigene-DNA chip and VITEK2Compact system was91.84%(90/98), in which the coincidences for identification of isolates belonging to Staphylococcus, Enterococcus and Streptococcus species were91.67%(55/60),92.59%(25/27) and90.00%(9/10), respectively. Compared to the43h identification time in VITEK2Compact system, Verigene-DNA chip only took3.1h to complete the identification procedure. In the22S. aureus isolates and12S. epidermidis, Verigene-DNA chip confirmed that21S. aureus isolates and8S. epidermidis had methicillin resistance-associated mec A, gene, which completely coincident with PCR. In the9E. faecalis isolates and16E. faecium isolates, Verigene-DNA chip proved that one E. faecalis isolate and five E. faecium isolates were positive for vancomycin resistance-associated vanA gene, which also completely coincident with PCR. However, PCR but not Verigene-DNA chip revealed that two E. faecium had vanM gene.ConclusionThe culture model of double samples-double flasks can increase positive isolating culture rate of bacteria and fungi in blood samples. The predominant bacteria in blood infection are S. hominis, E. coli, S. aureus and S. epidermidis (>10%) while the predominant fungi in blood infection are C. parapsilosis, C. albicans, C. glabrata and C. guilliermondii (>10%). Although the differential susceptibility of bacterial and fungal isolates belonging to different species against different antibiotics is presented, the Gram-positive bacterial isolates have higher susceptible rates against linezolid and tigecycline, and the Gram-neagtive bacterial isolates have higher susceptible rates against tigecycline and amikacin as well as the fungal isolates have higher susceptible rates against5-flucytosine, amphotericin B and mycostatin. Since the Verigene-DNA chip has fast, accurate and convenient advantages as well as possesses the ability to the ability to simultaneously detect bacterial drug-resistant genes during identification of Gram-positive bacteria in blood infection compared to the VITEK2Compact system, this method is worthy for generalization and application in clinical laboratories.