ABSTRACT
@#Abstract: To report a case of Aspergillus salwaensis-induced spinal infection and its laboratory detection. The inflammatory granulation and necrotic tissue samples of a patient with spinal infection were collected from, the Affiliated Hospital of Chengde Medical College on June 17, 2020 for direct smear microscopy and culture, and the isolated strain was identified by microscopy by smear staining, matrix-assisted laser desorption ionization-time-of-flight mass spectrometry (MALDI-TOF-MS), molecular identification and in vitro antifungal susceptibility test. The patient was 62 years old female and presented with recurrent chest and back pain with no obvious cause. The initial diagnosis was spinal infection, after 7 days of treatment with levofloxacin, the effect was not good. Surgery was then performed remove the lesion via posterior thoracic debridement, and fungal hypha was observed under microscope in tissue specimens. The isolated strains had no typical structure, MALDI-TOF-MS was used for identification for many times, but there was no identification result. After 7 days of fluconazole treatment, the patient's condition improved, and her chest and back pain were alleviated compared to before surgery. The patient was discharged and followed up in the outpatient department, the fungus was later identified as Aspergillus salwaensis by sequence analysis of the internal transcribed spacer (ITS) gene sequencing, and the patient's antifungal medication was changed to voriconazole after with the attending physician. The patient consciously recovered well with no pain in the operative area and normal spinal activity at 1 year follow-up. The possibility of spinal fungal infection should be considered in patients with back pain without a clear cause and poor response to routine antibiotic treatment. Direct smear report of microscopic results are very important for guiding clinical antibiotic selection for rare filament fungi with atypical colony and microscopic morphology and unsuccessful MALDI-TOF-MS identification, molecular biological methods such as ITS sequence analysis can be helpful for early identification of the fungal species, improving identification speed.
ABSTRACT
【Objective】 To investigate the environmental pollution of blood collection and supply institutions by using matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) and evaluate its application value. 【Methods】 Colonies of air from blood donation sites, skin puncture sites of blood donors, platelet storage boxes, platelet collection equipment, object surfaces of related experimental consumables and cuff surfaces of staff after disinfection were collected, and typical colonies after cultivation were selected for microbial identification by microbial mass spectrometry and then compared with bacteria results detected in blood components from May 2017 to May 2021. 【Results】 Aseptic growth, the number of colonies ≤4.0 CFU/ dish, and the number of colonies > 4.0 CFU/dish accounted for 21.20%, 62.20% and 16.60%, respectively. The qualified rate from high to low was platelet storage box, bacteria settling in the air of blood donation room after disinfection, platelet collection equipment, skin puncture site of blood donors after disinfection, the surface of platelet consumables and the surface of medical staff's overalls. After disinfection, the blood donors' skin puncture sites were compared with other collection sites, and the t values were 2.0371, 1.508, 2.109, 1.961 and 1.778, respectively, with no significant difference (P>0.05). Thirty cases of bacterial contamination of blood components were detected from May 2017 to May 2021, among which the detection rate of apheresis platelets was the highest, and the t values were 1.731 and 2.272, relative to the contamination frequency of erythrocytes and plasma bacteria (P>0.05), while the t value was 2.875, relative to concentrated platelets, with significant difference (P<0.05). 【Conclusion】 Bacterial contamination of blood components mostly come from air bacteria settling, blood donors' arms and skin after disinfection, and surfaces of related equipment and materials. Therefore, it is of clinical significance to conduct strict disinfection of working sites, establish disinfection monitoring methods and formulate disinfection hygiene standards in blood stations.
ABSTRACT
【Objective】 To explore the application of matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) in the genotyping of difficult blood typing samples, and to provide evidence for clinical blood transfusion. 【Methods】 Three ambiguous blood group samples, submitted to Shanghai Blood Center by Shanghai regional hospitals, were studied, of which Sample1 included the proband and his parents. Serological methods were used to perform blood group typing, direct antibody test, unexpected antibody screening and identification test. Blood group genotyping was performed by using the MALDI-TOF MS detection systeme stablished in our laboratory. Sanger sequencing was used to confirm gene mutation sites, and serological or flow methods were used to verify specific samples′ phenotype. 【Results】 Serological results indicated the existence of antibodies against high frequency antigens in sample 1 (including proband and her mother), 2 and 3. The genotyping results of MALDI-TOF MS showed that the proband of sample 1 was Di(a+ b+ ), her father was Di(a-b+ ), her mother was Di(a+ b-), sample 2 was p, and sample 3 was Jr(a-). Sequencing results of three samples were consistent with mass spectrometry typing results. Serological results showed that sample 2 had a p phenotype. The flow cytometry results suggested that sample 3 had a Jr(a-) phenotype. 【Conclusion】 For the first time, we applied MALDI-TOF MS technology to blood type genotyping of ambiguous clinical samples in China. Compared with other genotyping methods such as PCR-SSP, MALDI-TOF MS has the advantages of rapid detection, high throughput and high specificity, which would contribute to identification of difficult blood typing samples in the future, as well as rare blood group screening.
ABSTRACT
Two types of Mycobacterium abscesses (Mab) were found in sputum from a patient with severe pneumonia in May 2020. One was Mycobacteriumabscessus with smooth morphology (Mab S), the other was Mycobacteriumabscessus with rough morphology (Mab R). Both of them were compared and drug susceptibility testing were performed to provide clinical scientific diagnosis and treatment. Morphology, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) and 16S rRNA were used to analyze Mab S and Mab R, phylogenetic evolution tree was constructed by gene sequence alignment for homology, proportion and broth drug test were used for in vitro drug sensitivity test. There were morphological differences between Mab S and Mab R. MALDI-TOF MS analysis showed that there were 223 protein peaks in Mab R and 147 protein peaks in Mab S. Mab S contained 1 397 bp and Mab R contained 1 402 bp as 16s rRNA gene sequencing revealed. Drug susceptibility testing showed that both of them were almost resistant to all antituberculosis drugs, but sensitive to most of antibiotics. Mab S and Mab R were not only different in manifestations, but also in protein and gene comparison. Both of them were generally resistant to antituberculosis drugs. Antibiotic combined therapy has been confirmed to be an effective treatment in clinic.
ABSTRACT
Objective To investigate the source of microbial contamination in the clean room of the workshop. Methods Microbiological sampling was carried out from the air, environment and personnel of the workshop. The samples were cultivated, the microorganisms were detected by MALDI-TOF-MS, and homology analysis was performed with the microbial identification system of the instrument. Results A total of 14 species and 41 strains of bacteria were detected. Nine strains of Staphylococcus epidermidis were selected for homology analysis, and the Staphylococcus epidermidis from personnel gloves and headgear had 94% homology. There was 83% homology among the staphylococcus epidermidis derived from the sedimentation bacteria, the ground environment and personnel, which was higher than the 71% of the standard strain. Conclusion The homology analysis demonstrates that the pollution in the clean room of the workshop mainly comes from personnel, and secondly comes from the environment outside the workshop. Enterprises need to strengthen management to prevent the occurrence of microbial contamination. MALDI-TOF-MS can be used for rapid detection of complex environmental bacteria and for homology analysis.
ABSTRACT
Fungal peritonitis is a rare but serious complication of peritoneal dialysis. This infection has been reported to be mostly caused by Candida species, and less frequently by a variety of other yeasts and moulds, such as Aspergillus, Penicillium, and Fusarium spp. are commonly isolated from soil, plants and environmental surfaces, and rarely from non-immunosuppressed subjects. In this report, author describe a case of infection caused by Fusarium solani in a 59-year-old man undergoing continuous ambulatory peritoneal dialysis. The fungus was recovered from cultures of peritoneal dialysate and the pathogen identification was carried out by mass spectrometry. The patient's outcome was favorable without complications after liposomal amphotericin B treatment along with peritoneal dialysis catheter removal.
ABSTRACT
To determine relative molecular weight of astragalus polysaccharides (APs), we used Shodex GS620 gel permeation chromatographic column and differential refraction detector (GPC-RI) with dextran as a reference. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) and GPC combined with multi-angle laser light scattering detection (GPC-MALLS) were also used．GPC-RI measure showed four peaks of APs, with the Mw of 1 380 000, 231 000, 18 000, and 480, respectively. Three main peaks were found by GPC-RI-MALLS with the Mw as 1 148 000, 180 000, and 43 000, respectively. Strong signals in 155 000 and 18 000 were detected by MALDI-TOF-MS, which also indicated the sugar moieties of the APs as hexoses. From our study, we found that the GPC-RI method with universal correction is most suitable for Mw determination of the APs. Nevertheless, the three methods should be combined and contrasted with each other to obtain accurate information in research of polysaccharides from Chinese medicine.
ABSTRACT
Objective@#To explore peptidomic changes of peptides in saliva and gingival crevicular fluid (GCF) before and after treatment of gingivitis.@*Methods@#From January 2017 to September 2017, seventeen participants at the age of 24-62 (6 males and 11 females) at Department of Preventive Dentistry, Peking University School and Hospital of Stomatology with gingivitis were recruited in the present study. Their clinical parameters were measured and recorded. Saliva and GCF samples were collected from each of the participants at the baseline and 7 days after ultrasonic supragingival scaling, respectively. Matrix-assisted laser desorption-ionization time-of-flight mass spectrometry (MALDI-TOF MS) was employed to detect the changes of peptidomic profiles, while ano-liquid chromatography-electrospray ionization-tandem mass spectrometry (nano-LC/ESI-MS/MS) was performed to identify the possible proteins from which the peptides might derive.@*Results@#Initially, four peptide peaks [mass-to-charge ratio (m/z) values: 1 030.6, 1 043.4, 1 053.4 and 1 064.6] were screened out exhibiting a decreasing trend after treatment (P<0.05). Besides, five peptide peaks from gingival crevicular fluid (P<0.05) exhibited differential expression, among which 1 055.5 and 1 168.3 demonstrating a decrease after treatment, while 3 363.7, 3 480.9 and 3 489.5 increased overtime. Certain positive correlations were detected between some peptides and clinical parameters. Principle component analysis using the above mentioned peptide peaks showed a distinct distribution before and after treatment and peptides from GCF showed a slightly better capacity to discriminate patients before and after treatment. The peptides with m/z values of 1 055.5 in GCF and 1 064.6 in saliva were identified to be segments of serum albumin and complement C3, respectively.@*Conclusions@#Several differentially expressed peptides were detected in saliva and GCF by MALDI-TOF MS, exhibiting the potentiality to act as biomarkers in gingivitis patients.
ABSTRACT
Objective To screen specific mass peaks (SMP) of carbapenem-resistant Klebsiella pneumoniae (CRKP) by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), and perform rapid homology analysis on CRKP based on SMP.Methods 76 strains of CRKP with multilocus sequence typing (MLST) were selected from a microbiology laboratory of a hospital, 52 strains were for establishment of method and 24 strains were for validation of method. Five different criteria for selecting SMP of CRKP were set, SMP of different ST-type were screened according to different criteria, CRKP was typed based on screened SMP, criterion with the highest coincidence rate of MLST results was determined as the best criterion. Twenty-four CRKP strains were typed according to the specific peaks screened through best criteria, accuracy of SMP typing method was verified, and was compared with principal component analysis (PCA) and main spectra profile (MSP) cluster analysis in mass spectrometer software.Results According to standard 2 (①signal-to-noise ratio [S/N]≥4, ②S/N ratio≥1.5, ③coefficient of variability [CV≤40％]), 45 SMP were selected from 52 strains of CRKP strains, 29 strains of CRKP were typed by SMP, with the highest coincidence rate (82.8％) with MLST, which was determined as the best criterion. Another 24 CRKP strains were typed according to SMP screened based on this criterion, and the coincidence rate with MLST was 83.3％. The coincidence rate between PCA cluster analysis and MLST was only 66.7％, consistency between MSP clustering analysis and MLST was poor, and it didn't conform to the grouping trend of ST typing.Conclusion MALDI-TOF MS can select SMP of CRKP of different ST, which can be used for rapid homology analysis on CRKP, provide basis for surveillance and control of outbreak of healthcare-associated infection.
ABSTRACT
Objective To investigate the relationship between the single nucleotide polymorphisms of SCN1A genes and the therapeutic effects of carbamazepine in Zhuang population with epilepsies. Methods We used Mass ARRAY-IPLEX and matrix assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF-MS) technology to detect the SCN1A gene rs4667869 and rs10497275 genotypes in peripheral blood of186 Zhuang individuals with epileptic (66 cases in effective group and 120 cases of ineffective group) who received the standardized treatment of carbamazepine in Baise Region. The reversed phase high-performance liquid chromatography was used to determine blood drug level of carbamazepine. The correlations between the genotypes, alleles and the carbamazepine efficacy of the two groups were evaluated, respectively. We also analyzed the difference of carbamazepine's blood concentration between different genotypes. Results Three genotypes of GG, GC and CC were detected in rs4667869 locus. There were 3 genotypes of GG, GA and AA found in rs 10497275 locus.The differences in the allele distribution (χ2 = 11.790, P = 0.001) and genotype distribution (χ2= 10.655, P =0.005) of the rs4667869 locus were statistically significant between the two groups (ineffective group vs. effective group). However, there was no significant difference in allele distribution (χ2 = 3.335, P= 0.068) and genotype (χ2= 3.046, P = 0.218) for rs 10497275 locus in these two groups. Compared with the GG + GC genotype, the CC genotype of rs4667869 locus significantly reduced the antiepileptic efficacy of carbamazepine (OR = 2.800, 95％CI : 1.495～5.244). W hile there were no significant differences in blood concentration of genotype CC (t=1.273, P = 0.083) comparing with genotypes GG + GC in rs4667869. No significant differences were found in blood concentration between genotype AA and genotypes GG + GA of rs 10497275 (t= 0.963, P = 0.064). Conclusions These results suggest that the single nucleotide polymorphisms of rs4667869 in SCN1A genes could be associated with the drug resistance of carbamazepine in Zhuang population with epilepsies.
ABSTRACT
ABSTRACT Introduction: The identification of anaerobic bacteria by conventional methods employed in clinical laboratories requires a lot of work and a long response time [turnaround time (TAT)]. Matrix assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) is an accurate, rapid and inexpensive technique with promising results for bacterial identification. Objective: To evaluate the MALDI-TOF mass spectrometry (VITEK-MS, bioMérieux, France) compared to the ANC card (VITEK 2, bioMérieux, France) for the identification of anaerobes, and also veriying the cost variation between both methodologies. Methods: 421 anaerobes were concomitantly identified by ANC (VITEK 2) and MALDI-TOF (VITEK MS). The conflicting results or those presenting low differentiation of the species were subjected to the 16S ribosomal ribonucleic acid (rRNA) sequencing. Results: Thirty-five strains were not identified by ANC (VITEK 2), and only one isolate was not identified by MALDI-TOF (VITEK MS). From the 386 anaerobes identified by the two methodologies, 97% agreement was observed on the identification of genus and species between the methodologies. Thirteen (3%) isolates were submitted to 16S sequencing. The agreement observed was 70% using ANC (VITEK 2) using 92% by MALDI-TOF (VITEK MS). Conclusion: Both methodologies showed an excellent performance for the identification of the strains tested with great differences in relation to cost-benefit. MALDI-TOF MS allowed 35 additional identifications and a saving of BRL$ 7,786 with the release of culture positive result five days ahead of the ANC (VITEK 2). TAT reduction may contribute to a successful clinical resolution.
RESUMO Introdução: A identificação das bactérias anaeróbias por métodos convencionais empregados nos laboratórios clínicos demanda muito trabalho e um longo tempo de resposta (TAT). A espectrometria de massa por ionização e dessorção a laser assistida por matriz (MALDI-TOF MS) é uma técnica precisa, rápida e barata, com resultados promissores para a identificação bacteriana. Objetivo: Avaliar a espectrometria de massas MALDI-TOF (VITEK MS, bioMérieux, France) diante do cartão ANC (VITEK 2, bioMérieux, France) para a identificação de anaeróbios, bem como verificar a variação de custos entre as metodologias. Métodos: Foram identificados 421 anaeróbios concomitantemente pelo ANC (VITEK 2) e pelo MALDI-TOF (VITEK MS). Os resultados discordantes ou que apresentaram baixa discriminação das espécies foram submetidos ao sequenciamento do 16S do ácido ribonucleico ribossonal (rRNA). Resultados: Trinta e cinco cepas não foram identificadas pelo ANC (VITEK 2), e somente um isolado ficou sem identificação pelo MALDI-TOF (VITEK MS). Dos 386 anaeróbios identificados pelas duas metodologias, a concordância na identificação de gênero e espécie foi observada em 97%. Treze (3%) isolados foram submetidos ao sequenciamento do 16S; a concordância observada foi de 70% com o ANC (VITEK 2) e 92% com MALDI-TOF (VITEK MS). Conclusão: Ambas as metodologias demonstraram ótimo desempenho para identificação das cepas testadas com grandes diferenças em relação ao custo-benefício. O MALDI-TOF MS permitiu 35 identificações adicionais e uma economia de R$ 7.786,00 com a liberação do resultado positivo da cultura cinco dias à frente do ANC (VITEK 2). A redução do TAT pode contribuir para uma resolução clínica bem-sucedida.
ABSTRACT
Objective To analyze the fingerprints of serum peptides in bloodstream infection induced by Candida albicans (C.albicans),with matrix assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS),and establish the corresponding diagnostic model Methods To establish ICR mice model of C.albicans bloodstream infection,and collect the serum samples which were purified by weak cation exchange beads.The serum peptide finger printing was recognized by MALDITOF MS,and BioExplorer software was used for analysis between infection group and normal control group.Furthermore,the peptide fingerprints were analyzed between C.albicans infection group,Escherichia coli (E.coli) infection group and Staphylococcus aureus (S.aureus) infection group.Results Comparison between C.albicans infection group and normal control group,there were 135 polypeptide peaks,of which 5 polypeptides up-regulated,6 down-regulated and 7 up-regulated first and down-regulated afterwards.The diagnostic model was established by combination of 6 peaks (i.e.m/z 1610.9,1742.3,2666.4,2778.0,3345.1 and 4528.8),which possessed an accurate rate of 80.0％ in diagnosis of C.albicans infection.It was found by comprehensive comparison between C.albicans,E.coli and S.aureus infection groups that there were 5 polypeptides expressed collectively,i.e.m/z1513.8,2910.8,3538.1,3884.9 and 5007.3.Conclusions MALDI-TOF MS can be used to distinguish the C.albicans infection and normal polypeptide peaks.The collective polypeptides expressed in the infection groups can be further used to diagnose the infection.Establishment of corresponding diagnostic models may be helpful for early diagnosis of fungal bloodstream infection and provide a basis for reasonable clinical medication.
ABSTRACT
Objective To establish Brucella peptide mass fingerprint database by MicroflexTMLT MALDI-TOF mass spectrometer ,and evaluate its application value in the identification of clinical isolates of Brucella. Methods In October 2013 ,a suspected Brucella strain was isolated in the microbiology laboratory of the hos-pital.It was confirmed as Brucella meltensis by VITEK ? 2 Compact automatic microbial analysis system and 16S rDNA gene sequencing.Brucella peptide mass fingerprint database was established with this strain ac-cording to the manufacturer's recommended method.Two suspected Brucella strains isolated in the laboratory in April 2015 and June 2016 were tested for Gram stain ,Kozlowski stain and biochemical reactions such as oxi-dase ,catalase and rapid urease activity.MicroflexTM LT mass spectrometer was used to collect the mass spec-trums of these two strains and three suspected Brucella strains isolated from other hospitals.When these spectrums were comparatively analyzed ,self-built database was added on the basis of the MALDI Biotyper da-tabase.The 16S rDNA gene sequencing was also used to identify the above-mentioned bacteria.Results Ac-cording to the recommended self-built database construction process ,Brucella peptide mass fingerprint data-base based on BRUxh1 was obtained successfully.The suspected Brucella strains isolated in April 2015 and June 2016 showed small Gram-negative coccobacillus under microscopic examination of stained smear ,and were positive for oxidase ,catalase ,urease activity (within 5 min).The identification results of mass spectrom- etry for the 5 suspected strains were all Brucella (mass spectrometry score :2.407 ,2.475 ,2.436 ,2.466 and 2.397).The 5 strains were confirmed as Brucella by 16S rDNA gene sequencing.Two strains among bacteria from other hospitals were falsely identified as Roseomonas gilardii by the VITEK?2 Compact system.Conclu-sion The combination of MALDI Biotyper database and self-built Brucella database can be used to compara-tively analyze the mass spectrometry of clinical isolates.Accurate identification of Brucella can be achieved by this means.The method which is fast and sensitive has high application value for rapid clinical diagnosis of Brucellosis.
ABSTRACT
The matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) is one of the first choice techniques for clinical microbiological identification owing to its fast,accuracy,low cost and easy to be operated.The accuracy of identification of MALDI-TOF MS relies on good instrument status,appropriate pre-trement,standardized operation and the rules of interpretation.Any errors in this process will directly influence the accuracy of identification.Therefore,strict internal quality control for the microbiological identification by MALDI-TOF MS is very important.Based on six years of research and clinical application experience,our laboratory establishes a relatively complete internal quality control system for the microbiological identification by MALDI-TOF MS,including all aspects of hardware,software and manual operation,the parameter settings,maintenance and calibration of the instrument,the preparation of strains and reagents,spectra acquisition and the interpretation and analysis of results,Which can make abnormal results be quickly traced and timely handled.
ABSTRACT
Objective To evaluate the value of matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF MS) in detecting carbapenemases-producing Enterobacteriaceae. Methods Sixty-one carbapenemase-producing Enterobacteriaceae (CPE) and 108 carbapenemase-none-producing Enterobacteriaceae(NCPE) strains that isolated in the Renmin Hospital of Wuhan University from February 2016 to February 2017 were used in this study. Twenty CPE strains and 20 NCPE strains were se-lected and used to establish the optimum reaction system of MALDI-TOF MS for the detection of carbapene-mase-producing Enterobacteriaceae strains. Results The optimal reaction system of MALDI-TOF MS was successfully established:equal volumes of imipenem solution (1 mg/ml) and bacterial suspension (2 Mc-Farland turbidity standard) were first mixed together and incubated at 37℃ for 20 min,and then the super-natant was detected by MALDI-TOF MS after centrifugating. It would be a carbapenemase-producing strain if the peak of (300±0.55) m/z in the mass spectrum of imipenem disappeared completely. MALDI-TOF MS was used to identify carbapenemase-producing Enterobacteriaceae strains among the 169 strains and the result was consistant with that by using carbapenemase gene detection. Conclusion MALDI-TOF MS could accu-rately and rapidly detect carbapenemases-producing Enterobacteriaceae. It could be used as a routine method to provide references for early anti-infective treatment and infection control in hospitals.
ABSTRACT
Se evaluaron 3 metodologías de extracción de proteínas para la identificación de hongos miceliales por MALDI-TOF MS en 44 aislados: la extracción con agua-ácido fórmico (E. agua), la extracción con zirconio-etanol-acetonitrilo-ácido fórmico (E. zirconio) y la recomendada por el proveedor del equipo (E. tubo). Se compararon 2 bases de datos: Bruker (BK) y BK + National Institutes of Health. Los resultados obtenidos utilizando dichas bases fueron los siguientes (en el orden citado): identificación correcta (IC) a nivel de género, 10 y 16 con E. agua; 27 y 32 con E. zirconio y 18 y 23 con E. tubo; IC a nivel de especie, 5 y 7 con E. agua; 17 y 20 con E. zirconio y 11 y 14 con E. tubo; identificaciones no confiables, 18 y 12 con E. agua y 9 y 4, tanto con E. zirconio como con E. tubo; ausencia de pico, 16 con E. agua, 8 con E. zirconio y 17 con E. tubo. La extracción con zirconio mostró el mejor rendimiento (p < 0,05).
In order to optimize the identification of molds with MALDI-TOF MS, three protein extraction-methodologies were evaluated against 44 isolates: water extraction (WE), zirconium extraction (ZE) and the provider's recommended method (PRM). Two data bases were compared, Bruker (BK) and Bruker + National Institutes of Health. Considering both databases, results were respectively as follows: correct identification (CI) at gender level, 10 and 16 by WE; 27 and 32 by ZE and 18 and 23 by PRM; CI at species level, 5 and 7 by WE; 17 and 20 by ZE and 11 and 14 by PRM; non-reliable identification, 18 and 12 by WE; 9 and 4 by ZE and by PRM. No peaks were observed in 16 by WE, 8 by ZE and 17 by PRM. ZE showed the best perfomance (p < 0.05).
Subject(s)
Proteins/analysis , Mycelium/classification , Fungi/classification , Mass Spectrometry/methods , Databases, Factual/statistics & numerical dataABSTRACT
Abstract Mastitis adversely affects milk production and in general cows do not regain their full production levels post recovery, leading to considerable economic losses. Moreover the percentage decrease in milk production depends on the specific pathogen that caused the infection and enterobacteria are responsible for this greater reduction. Phenotypic tests are among the currently available methods used worldwide to identify enterobacteria; however they tend to misdiagnose the species despite the multiple tests carried out. On the other hand The Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry (MALDI-TOF MS) technique has been attracting attention for its precise identification of several microorganisms at species level. In the current study, 183 enterobacteria were detected in milk (n = 47) and fecal samples (n = 94) from cows, and samples from water (n = 23) and milk lines (n = 19). All these samples were collected from a farm in Rio de Janeiro with the specific purpose of presenting the MALDI-TOF MS technique as an efficient methodology to identify Enterobacteriaceae from bovine environments. The MALDI-TOF MS technique results matched the biochemical test results in 92.9% (170/183) of the enterobacteria species and the gyrB sequencing confirmed 100% of the proteomic technique results. The amino acid decarboxylation test made the most misidentifications and Enterobacter spp. was the most misidentified genus (76.9%, 10/13). These results aim to clarify the current biochemical errors in enterobacteria identification, considering isolates from a bovine environment, and show the importance for more careful readings of phenotypic tests which are often used in veterinary microbiology laboratories.
Subject(s)
Animals , Female , Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization , Enterobacteriaceae/classification , Enterobacteriaceae/metabolism , Phenotype , Cattle , Sequence Analysis, DNA , DNA Gyrase/genetics , Proteomics/methods , Milk/microbiology , Enterobacteriaceae/isolation & purification , Genes, Bacterial , Mastitis, Bovine/diagnosis , Mastitis, Bovine/microbiologyABSTRACT
Objective The aim of this study was to evaluate the commercial SepsityperTM kit and serum separator tube coupled with matrix-assisted laser desorption/ionization time of flight (MALDI-TOF) mass spectrometry for direct identification of microorganisms in a blood culture system.Methods A total of 138 clinical blood samples from clinical laboratory in Renji hospital were tested with two methods respectively from April to June of 2016.Performance of the assays were compared against that of conventional bacterial culture as a reference.Results A total of 138 nonduplicate positive blood culture samples were collected,including 70 (53.03％) gram negative samples,57 (43.18％) gram positive samples,3 fungus samples,2 mixed samples,and 6 false positive samples which were excluded from further analysis.The accuracy rate of SepsityperTM kit and serum separator tube was 91.67％ and 84.09％ in rapid identification of pathogen from blood samples,83.33％ and 61.36％ in correct identification to species level.The accuracy rate of SepsityperTM kit and serum separator tube was 98.57％ and 95.71％ in identifying gram-negative bacteria,87.72％ and 78.59％ in identifying gram-positive bacteria,respectively.The turnaround time for identification of each sample was 40 min by the commercial SepsityperTM kit and 25 min by serum separator tube.Conclusions MALDI SepsityperTM kit has shown slightly higher accuracy rate in identification of pathogen from blood sample than serum separator tube,but the difference is not significant (91.67％ vs.84.09％,P＞0.05).Compared with MALDI SepsityperTM kit,serum separator tube is a rapid,easy,and cost-effective pretreatment method for direct identification of microorganisms from blood cultures using MALDI-TOF mass spectrometry.
ABSTRACT
Objective To study the serum peptide fingerprint using matrix-assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF MS) technology,and find the different peaks with potential significance and establish the diagnosis model of Staphylococcus aureus and Escherichia coli bloodstream infection.Methods To establish ICR mice model of S.aureus and E.coli bloodstream infections,and collect serum samples.The serum samples were purified by weak cation exchange beads,the serum peptide fingerprint was recognized by using MALDI-TOF MS and BioExplorer software between infections group and normal control group.Results Compared with the normal control group,6 peptides were up-regulated,7 peptides downregulated and 8 peptides up-regulated first and then down-regulated in S.aureus infection group;And 5 peptides down-regulated,4 peptides down-regulated first and then up-regulated,and 8 peptides up-regulated first and then down-regulated in E.coli infection group.Conclusion MALDI-TOF MS combined with BioExplorer software may be used as a tool to study the serum peptides of S.aureus and E.coli bloodstream infection,effectively find significant peptides for establishing a diagnosis model of these two bacterial infections,and has a certain value for the diagnosis of bacterial bloodstream infection.
ABSTRACT
Objective·To directly detect the bacteria in positive blood culture specimens by the separation gel tube combined with MicroflexTM matrixassisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS),perform direct antimicrobial susceptibility test based on the results of mass spectrometry,and preliminary explore the redesign of conventional positive blood culture specimen processing flow.Methods·449 positive-alarm blood culture specimens of monobacterial infections in clinical microbiology laboratory of Xirhua Hospital affiliated to Shanghai Jiao Tong University School of Medicine from September 1,2015 to August 31,2016 were collected and prepared according to the new positive blood culture specimens processing flow.The new redesigned processing flow included smear staining and microscopy,separation gel/mass spectrometry direct identification,bacteria film/mass spectrometry identification,pure colony/mass spectrometry identification,direct antimicrobial susceptibility test,and conventional antimicrobial susceptibility test,etc.According to the results of microscopic examination,identification test,and antimicrobial susceptibility test,level Ⅰ direct microscopic examination report,positive blood culture level Ⅱ (direct identification/bacteria film identification or direct antimicrobial susceptibility) report,and level Ⅲ final report were provided sequentially.Results·With the new redesigned processing flow,bacterial specieslevel identification reports for 82.2％,95.8％,and 100％ of positive blood samples could be obtain at 10 am and 17 pm on the current day and 10 am in the next morning,respectively,and be sent to clinical departments.Initial antimicrobial susceptibility reports for the bacteria that were considered as clinical significant pathogens by preliminary assessment could be provided in the next day of blood culture positive alarm with a higher coincidence rate as compared to the results of traditional antimicrobial susceptibility tests.Conclusion·The time from blood culture positive alarm to the provision of initial identification and antimicrobial susceptibility reports is shorter by 1-2 d for the redesigned processing flow than for the traditional one,which can provide fast and accurate etiologic diagnosis evidence for bloodstream infections for clinicians and is important for improving early diagnosis and treatment of clinical bloodstream infections.