scholarly journals Discovery of Species-unique Peptide Biomarkers of Bacterial Pathogens by Tandem Mass Spectrometry-based Proteotyping

2020 ◽  
Vol 19 (3) ◽  
pp. 518-528 ◽  
Author(s):  
Roger Karlsson ◽  
Annika Thorsell ◽  
Margarita Gomila ◽  
Francisco Salvà-Serra ◽  
Hedvig E. Jakobsson ◽  
...  

Mass spectrometry (MS) and proteomics offer comprehensive characterization and identification of microorganisms and discovery of protein biomarkers that are applicable for diagnostics of infectious diseases. The use of biomarkers for diagnostics is widely applied in the clinic and the use of peptide biomarkers is increasingly being investigated for applications in the clinical laboratory. Respiratory-tract infections are a predominant cause for medical treatment, although, clinical assessments and standard clinical laboratory protocols are time-consuming and often inadequate for reliable diagnoses. Novel methods, preferably applied directly to clinical samples, excluding cultivation steps, are needed to improve diagnostics of infectious diseases, provide adequate treatment and reduce the use of antibiotics and associated development of antibiotic resistance. This study applied nano-liquid chromatography (LC) coupled with tandem MS, with a bioinformatics pipeline and an in-house database of curated high-quality reference genome sequences to identify species-unique peptides as potential biomarkers for four bacterial pathogens commonly found in respiratory tract infections (RTIs): Staphylococcus aureus; Moraxella catarrhalis; Haemophilus influenzae and Streptococcus pneumoniae. The species-unique peptides were initially identified in pure cultures of bacterial reference strains, reflecting the genomic variation in the four species and, furthermore, in clinical respiratory tract samples, without prior cultivation, elucidating proteins expressed in clinical conditions of infection. For each of the four bacterial pathogens, the peptide biomarker candidates most predominantly found in clinical samples, are presented. Data are available via ProteomeXchange with identifier PXD014522. As proof-of-principle, the most promising species-unique peptides were applied in targeted tandem MS-analyses of clinical samples and their relevance for identifications of the pathogens, i.e. proteotyping, was validated, thus demonstrating their potential as peptide biomarker candidates for diagnostics of infectious diseases.

2021 ◽  
Vol 1 (S1) ◽  
pp. s31-s32
Author(s):  
Amanda Gusovsky ◽  
David Burgess ◽  
Donna Burgess ◽  
Emily Slade ◽  
Chris Delcher ◽  
...  

Background: A team of infectious diseases physicians, infectious diseases pharmacists, clinical laboratorians, and researchers collaborated to assess the management of lower respiratory tract infections (LRTIs). In 1 sample from our institution, 96.1% of pneumonia cases were prescribed antibiotics, compared to 85.0% in a comparison group. A collaborative effort led to the development of a protocol for procalcitonin (PCT)-guided antibiotic prescribing that was approved by several hospital committees, including the Antimicrobial Stewardship Committee and the Healthcare Pharmacy & Therapeutics Committee in December 2020. The aim of this analysis was to develop baseline information on PCT ordering and antibiotic prescribing patterns in LRTIs. Methods: We evaluated all adult inpatients (March–September 2019 and 2020) with a primary diagnosis of LRTI who received at least 1 antibiotic. Two cohorts were established to observe any potential differences in the 2 most recent years prior to adoption of the PCT protocol. Data (eg, demographics, specific diagnosis, length of stay, antimicrobial therapy and duration, PCT labs, etc) were obtained from the UK Center for Clinical and Translational Science, and the study was approved by the local IRB. The primary outcome of interest was antibiotic duration; secondary outcomes of interest were PCT orders, discharge antibiotic prescription, and inpatient length of stay. Results: In total, 432 patients (277 in 2019 and 155 in 2020) were included in this analysis. The average patient age was 61.2 years (SD, ±13.7); 47.7% were female; and 86.1% were white. Most patients were primarily diagnosed with pneumonia (58.8%), followed by COPD with complication (40.5%). In-hospital mortality was 3.5%. The minority of patients had any orders for PCT (29.2%); among them, most had only 1 PCT level measured (84.1%). The median length of hospital stay was 4 days (IQR, 2–6), and the median duration of antibiotic therapy was 4 days (IQR, 3–6). Conclusions: The utilization of PCT in LRTIs occurs in the minority of patient cases at our institution and mostly as a single measurement. The development and implementation of a PCT-guided therapy could help optimize antibiotic usage in patients with LRTIs.Funding: NoDisclosures: None


2021 ◽  
Vol 8 (4) ◽  
pp. 260-265
Author(s):  
Meryem Colak ◽  
Selin Yigit ◽  
Anil Tapisiz ◽  
Hager Muftah ◽  
Kenan Yuce ◽  
...  

Objective: Human Parainfluenza viruses (HPIVs) cause respiratory tract infections, and the second most common cause of acute respiratory illness-related hospitalizations after the respiratory syncytial virus in children <5 years of age. The aim of the study; determination of HPIVs positivity and common types in pediatric patients with respiratory tract infection; investigation the distribution of HPIV positivity by age groups, months and seasons, respectively. Material and Method: HPIV results of 1613 pediatric patients who were sent to the molecular virology laboratory from various pediatric clinics of Gazi Hospital between March 2016 and February 2021 (five years period) were investigated. Nucleic acid isolation was performed on the EZ1 Advanced (Qiagen, Germany) device using the EZ1 Virus Mini Kit by the manufacturer's protocol. Results: HPIV positivity was detected as 4.1% in clinical samples and, the most common HPIV type was found to be HPIV-3 (55%). The distribution of other HPIV types were; HPIV-2, HPIV-4 and HPIV-1 with 26%, 23% and 14%, respectively. HPIV-3 is the most common type in 2016, 2017, 2018 and 2019; however, HPIV-1 is the most common type in 2020. HPIVs co-infection was detected with other respiratory tract viruses in 51% of samples. The highest HPIV co-infection was detected in Rhinovirus. The highest HPIV positivity rate (45%) were determined in the 0-2 age group compared to other age groups (p<0.05). The highest positivity rate was in October in the autumn season (p<0.05), the lowest was in January and February in winter. The highest rate (8.1%) of HPIV positivity was found in 2016 and the lowest rate (0.7%) was in 2020. Conclusions: Since it is not possible to diagnose viral etiology of respiratory tract infections based on clinical findings, viral respiratory tract panel and Multiplex real-time PCR test are a fast and useful method in early diagnosis, treatment decision and prevention of unnecessary antibiotic use. HPIVs positivity is seen at higher rates in children aged 0-2 and in autumn months with seasonal differences.


2019 ◽  
Vol 10 (2) ◽  
pp. 14-19 ◽  
Author(s):  
Dharm Raj Bhatta ◽  
Deependra Hamal ◽  
Rajani Shrestha ◽  
Supram HS ◽  
Pushpanjali Joshi ◽  
...  

Background: Lower respiratory tract infections are one of the most common infections among the patients in Intensive Care Units (ICUs). Admission in ICUs and use of life supporting devices increase the risk of infection with multidrug resistant pathogens. Aims and Objectives: This study was aimed to determine the prevalence and antibiograms ofthe bacterial pathogens causing lower respiratory tract infectionsamong patients of ICUs. Materials and Methods: A total of 184 specimens from patients admitted in ICUswith lower respiratory tract infections were included in this study. Isolation, identification and antibiotic susceptibility testing of the isolates was performed by standard microbiological techniques. Carbapenamase detection was performed by modified Hodge test method.Detection of metallo beta lactamase (MBL) was tested by imipenem and imipenem/EDTA disc. Detection of Klebsiellapneumoniaecarbapenamase (KPC) was performed by imipenem and imipenem/phenyl boronic acid. Results: Out of 184 samples, 131 showed significant growth of bacterial pathogens. Acinetobacter species (42.6%), Staphylococcus aureus (16.9%) and Pseudomonasaeruginosa(13.9%)were the three most common isolates. Out of 22 imipenem resistant isolates of Acientobacter species, 9 were KPC producer, 4 were MBL producers and 3 isolates were positive for MBL and KPC both. Among the Acinetobacter species, 5.1% isolates were resistant to tigecycline and colistin. One isolate of Pseudomonas aeruginosa was positive for MBL. Conclusions:High prevalence of multidrug resistant bacteria in ICUs was recorded. Gram negative bacilli were predominantly associated with LRTI among ICU patients;Acinetobacterspecies being most common isolate. Detection of carbapenamase among the Acinetobacterand emergence of tigecycline resistancelimits the therapeutic options.Regular monitoring of such resistant isolates would be important for managing infection control in critical units.


2021 ◽  
Vol 11 ◽  
Author(s):  
Yanli Li ◽  
Rubén Fernández ◽  
Inma Durán ◽  
Rafael A. Molina-López ◽  
Laila Darwich

Pet animals are assumed to be potential reservoirs in transferring antimicrobial resistance (AMR) to humans due to the extensively applied broad-spectrum antimicrobial agents and their close contact with humans. In this study, microbiological data and antimicrobial susceptibility results of dog (n = 5,086) and cat (n = 789) clinical samples from a private Laboratory of Diagnosis in Barcelona were analyzed. Samples came from different counties of the Iberian Peninsula during 2016–2018. In dogs, clinical samples were most commonly from otitis, and in cats from wounds, respiratory tract infections and conjunctivitis. In both pet groups, Staphylococcus spp. (31% in dogs vs 30% in cats), Streptococcus spp. (19% vs 17%), Pseudomonas spp. (16% vs 10%), Escherichia coli (8% vs 5.6%), and Enterococcus spp. (5.5% vs 6.8%) were shown as the most predominant bacteria. However, higher frequencies of P. aeruginosa, P. canis, and S. pseudintermedius were found in dogs, while S. aureus and P. multocida were more prevalent in cats. The antimicrobial susceptibility testing demonstrated that Enterococcus spp. and Pseudomonas spp. presented the highest levels of AMR in both dogs and cats. Within the Enterobacteriaceae, E. coli showed low levels of AMR compared to Klebsiella, Proteus, or Enterobacter spp. Respiratory tract infections caused by K. pneumoniae presented higher AMR in cats. By contrast, Pasteurella isolates from the respiratory tract were highly sensitive to all the antimicrobials in cats and dogs. Data from this study could be used to guide empirical antimicrobial selection in companion animal veterinary practices in the Iberian Peninsula.


Author(s):  
Ahmed Atia ◽  
Najla Elyounsi ◽  
Ahmed Abired ◽  
Amal Wanis ◽  
Abdulsalam Ashour

1) Background: Respiratory tract infections (RTI) has been known to be a significant health concern for mortality and morbidity since many years. This study was aimed at determining the prevalence of bacterial pathogen causing upper respiratory tract (URTIs) and the susceptibility patterns to frequently used antibiotics among patients attending Abusetta hospital in Tripoli district; 2) Methods: A total of 1,110 throat swabs were collected between Jan, 2011 to December, 2014 and inoculated onto Blood agar, MacCkonkey agar and Chocolate agar then incubated at 37 oC for 24 hours. Bacterial pathogens were determined by bacteriological culture methods and antibiotic susceptibility of the isolates was identified following Clinical Laboratory Standard Institute guidelines (CLSI); 3) Results: Of the 1,110 respiratory samples tested, 71.1% (n = 789) of specimens were positive cultures with the dominant bacterial pathogens being streptococcus pneumonia 43.3% (n = 342), followed by pseudomonas aeruginosa 22.8% (n = 180), staphylococcus aurous 13.8% (n = 109), Escherichia Coli 6.9% (n = 55), Enterobacter spp 6.2% (n = 49), Citrobacter 4.5% (n = 36), and Klebsiella 2.2% (n = 18). Most isolates exhibited resistance against the commonly used antibiotics and to at least one antibiotic; and 4) Conclusions: The level of antibiotic resistance in this study is alarming and brings to light the timely and suitable diagnosis of the common bacteria causes of URTIs and proper antibiotic administration based on susceptibility test.


2021 ◽  
Vol 29 (3) ◽  
pp. 58-65
Author(s):  
O. L. Bororova ◽  
◽  
Y. O. Dziublyk ◽  
V.A. Iachnyk

MODERN METHODS OF ETIOLOGICAL DIAGNOSING OF ACUTE COMMUNITY-ACQUIRED LOWER RESPIRATORY TRACT INFECTIONS O. L. Bororova, Y. O. Dziublyk, V.A. Iachnyk Abstract The review presents the possibilities presented by various methods of etiological diagnostics used in pulmonology. The main method of diagnosing acute community-aquired lower respiratory tract infections is the microbiological approach which includes microscopy of patient’s material with Gram staining, cultures on nutrient media, isolation of culture, identification and determination of susceptibility of a microorganism to antibiotics. But unfortunately the etiologocal factor cannot be detected in about half of patients. Recently, the popularity of molecular methods of etiological diagnosis has grown. They are characterized by greater sensitivity to microbiological methods and allow to get results faster. Molecular diagnostic tests are divided into four categories depending on the mechanism based on them: immunoassay, hybridization methods, amplification and sequencing methods. Among the tests based on the principles of immunoassay, noteworthy are rapid tests, which are most consistent with the idea of an ideal diagnostic tool in the field of laboratory medicine. They are fast, simple, cheap, highly sensitive and highly specific. However, as the appearance of specific antibodies in the body takes some time, the results of tests based on immunoassay remain positive for several weeks after the delayed episode of acute community-acquired lower respiratory tract infection, so they have diagnostic value only in the presence of clinical manifestations of the disease. The genetic approach allows the detection of infectious agents in the early stages of the disease, when serological and immunological methods are ineffective. Tests based on nucleic acid amplification, including PCR, have also become increasingly common recently. These methods should be used for the diagnosis of atypical pathogens and respiratory viruses, because their cultivation in culture is difficult. Sequencing and mass spectrometry methods are being actively developed, but there are limitations that prevent their use in everyday clinical practice. So the combination of microbiological approach with molecular diagnostic methods is the most optimal for the identification of the causative agent of acute community-acquired lower respiratory tract infections and the use of targeted etiotropic treatment. Key words: acute community-acquired lower respiratory tract infections, etiological diagnosis, microbiological, serological, immunological, molecular genetic methods, ICA, PCR, sequencing, mass spectrometry. Ukr. Pulmonol. J. 2021;29(3):58–65


2021 ◽  
Vol 38 (4) ◽  
pp. 471-473
Author(s):  
Fahriye EKŞİ ◽  
Mehmet ERİNMEZ

Acinetobacter baumannii is an opportunistic pathogen which colonize inpatients and cause severe infections, septic shock and death. With emergence of multi-drug resistant gramnegative species and being effective in A. baumannii infections, colistin becomes a treatment option again. N-acetylcysteine (NAC), is a mucolitic agent which used commonly in lower respiratory tract infections especially patients who have cronic respiratory disorders like Chronic obstructive pulmonary disease, cystic fibrosis and bronchiectasis. In this study we aim to investigate the effect of NAC, which commonly added in lower respiratory tract infections tratment regime, on MIC values colistin used in A. baumanni tratment. Fifty A.baumannii isolates were included in the study. The isolates were identified by automated identification system. With broth microdilution method, we investigated and compared the MIC (minimum inhibitory concentration) values of colistin and NAC+Colistin combination. Colistin MIC50 value is 0.25 μg/mL and MİK90 value is 1 μg/mL, NAC+Colistin combination MIC50 value is 0.25 μg/mL and MİK90 value is 1 μg/mL. The screening for the effectiveness of clinical drugs may provide clinical strategy to improve treatment outcomes of A. baumannii and reduce hospitalization days.


2017 ◽  
Vol 4 (5) ◽  
pp. 1733
Author(s):  
Venkata Krishna Munagala ◽  
Ramisetty M. Uma Mahesh ◽  
Jithendra Kandati ◽  
Munilakshmi Ponugoti

Background: WHO estimated burden of respiratory tract infections in 2010, estimates four and half million deaths due to respiratory tract infections among children every year. In India, 1.2 million deaths have been reported among children due to RTI among 5.9 million deaths globally. Lower respiratory tract infections are most common causes of death than upper respiratory tract infections. Pneumonia and Bronchiolitis are most common types of LRTI in children. Pneumonia accounts for most of the deaths in children < 5 years of age. The present study was undertaken with an objective to know the various types of lower respiratory tract infections in children less than 12 years of age. The study also aims to know the various bacterial agents causing respiratory tract infections with their antibiotic susceptibility.Methods: Hospital based, prospective cross-sectional study was conducted for a period of one year and 375 children were enrolled. Demographic, clinical history and examination was done and signs and symptoms noted. All necessary investigations were performed and followed regularly for management and outcome.Results: Incidence of LRTI in the study was 9.76% with male preponderance (65.33%) and most common among children in 1-4 years age group. Ratio of males to females was 1.9:1. 73.6% of cases were in low socio-economic group, 35.2% were found with PEM-I grade and 18.13% had no immunization coverage. Cough and breathlessness were the major symptoms and respiratory distress and clubbing were major signs in the study. Bronchopneumonia was the commonest cause (38.7%) followed by bronchiolitis and Allergic bronchitis. 18.45 of cases had anemia and Leucocytosis was also present. Pulmonary infiltration was the major finding in the X-ray of chest. Streptococcus pneumoniae and Klebsiella pneumoniae were the common bacterial pathogens isolated.Conclusions: To conclude, our study clearly highlighted the various types of clinical presentations, risk factors and different types of LRTI in children <12 years of age. Understanding a clear knowledge of the etiology and bacterial pathogens clearly provides guidance for the physician in management and clinical outcome. 


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