Blood Culture Method Research Articles

A Comparative Study of Bacteriological Profile and Susceptibility Pattern by Rapid Direct Manual Identification and Antimicrobial Susceptibility Testing with Routine Blood Culture Method in Neonatal Septicemia.

A Comparative Study of Bacteriological Profile and Susceptibility Pattern by Rapid Direct Manual Identification and Antimicrobial Susceptibility Testing with Routine Blood Culture Method in Neonatal Septicemia.

Microbial cell-free DNA-sequencing as an addition to conventional diagnostics in neonatal sepsis.

Bloodstream infections remain a challenge for neonatologists, as traditional culture-based methods are time-consuming and rely on adequate blood volume. Next-generation sequencing (NGS) offers an alternative, as it can identify microbial cell-free DNA (mcfDNA) in a small blood sample, providing rapid pathogen detection. This study aimed to assess the diagnostic performance of DISQVER®-NGS compared to blood cultures in neonatal patients with suspected sepsis. In neonates with suspected sepsis, blood cultures and samples for NGS were prospectively collected. Patients were divided into four categories: 1) sepsis, blood culture positive, 2) clinical sepsis, culture negative, 3) suspected sepsis, 4) validation cohort. NGS detected bacterial, viral or fungal mcfDNA in 24 of 82 samples. Blood cultures were collected in 46 of 84 patients (15/46 positive). DISQVER® correctly identified pathogens in 9/15 patients with a positive blood culture, two with intrinsic resistance to their antibiotic regimen. In seven samples NGS reported the mcfDNA of bacteria that could have theoretically grown in culture but did not. NGS may enhance sensitivity in sepsis diagnostics by detecting mcfDNA in neonates with suspected sepsis. Interpreting NGS results requires correlation with clinical data, laboratory values, and routine microbiological tests for a comprehensive understanding of the patient's condition. Conventional blood culture methods have limitations in accuracy and turnaround time. The study aimed to investigate the diagnostic performance of the Next-Generation Sequencing method DISQVER® compared to traditional blood cultures in neonatal patients with suspected sepsis. Our findings suggest that NGS has the potential to augment the precision of conventional diagnostic techniques, can lead to improved detection of pathogens and targeted treatment approaches in neonatal sepsis. It is emphasized that further validation and integration with clinical and microbiological data are required to ensure optimal clinical utility.

Identification of sepsis-causing bacteria from whole blood without culture using primers with no cross-reactivity to human DNA

Sepsis is a major health concern globally, and identification of the causative organism usually takes several days. Furthermore, molecular amplification using whole blood from patients with sepsis remains challenging because of primer cross-reactivity with human DNA, which can delay appropriate clinical intervention. To address these concerns, we designed primers that could reduce cross-reactivity. By evaluating these primers against human DNA, we confirmed that the cross-reactivity observed with conventional primers was notably absent. In silico PCR further demonstrated the specificity and efficiency of the designed primers across 23 bacterial species that are often associated with sepsis. When tested using blood samples from sepsis patients, the designed primers showed moderate sensitivity and high specificity. Surprisingly, our method identified bacteria even in samples that were detected at other sites but tested negative using conventional blood culture methods. Although we identified some challenges, such as contamination with Acetobacter aceti due to the saponin pretreatment of samples, the developed method demonstrates remarkable potential for rapid identification of the causative organisms of sepsis and provides a new avenue for diagnosis in clinical practice.

Development of two multiplex PCR assays for rapid detection of eleven Gram-negative bacteria in children with septicemia

AimThis study aimed to develop a multiplex PCR assay for simultaneous detection of major Gram-negative etiologies of septicemia and evaluate its performance.MethodsMultiplex PCR (mPCR) assays were developed targeting 11 bacterial strains. Species-specific primers were confirmed using known clinical isolates and standard strains. Gradient PCR was performed on each primer against its target bacterial gene to determine its optimal amplification condition. The minimum detectable DNA concentration of the two assays was evaluated by adjusting bacterial DNA concentration to 100 ng/μL and, tenfold serially diluting it up to 10 pg/μL with DNAse-free water. The diagnostic accuracy of mPCR assays was established by subjecting the assays to 60 clinical blood samples.ResultsTwo mPCR assays were developed. Optimal primer annealing temperature of 55 °C was established and utilized in the final amplification conditions. The assays detected all targeted bacteria, with a 100 pg minimum detectable DNA concentration. Pathogens were not detected directly from whole blood, but after 4 h and 8 h of incubation, 41% (5/12) and 100% (12/12) of the bacteria were detected in culture fluids, respectively. The assays also identified Salmonella spp. and Klebsiella pneumoniae co-infections and extra pathogens (1 E. coli and 2 K. pneumoniae) compared with culture. The sensitivity and specificity of the mPCR were 100.0% (71.7–100.0) and 98.0% (90.7–99.0), respectively. The area under the ROC curve was 1.00 (1.00–1.00).ConclusionsThe mPCR assays demonstrated substantial potential as a rapid tool for septicemia diagnosis alongside the traditional blood culture method. Notably, it was able to identify additional isolates, detect co-infections, and efficiently detect low bacterial DNA loads with high sensitivity, implying its value in enhancing efficiency of diagnosis of septicemia.

Application of metagenomic next-generation sequencing technology in the etiological diagnosis of peritoneal dialysis-associated peritonitis.

Pathogens detected by metagenomic next-generation sequencing (mNGS) and the laboratory blood culture flask method were compared to understand the advantages and clinical significance of mNGS assays in the etiological diagnosis of peritoneal dialysis-associated peritonitis (PDAP). The study involved a total of 37 patients from the hospital's peritoneal dialysis centre, six of whom were patients with non-peritoneal dialysis-associated peritonitis. Peritoneal dialysis samples were collected from the 37 patients, who were divided into two groups. One group's samples were cultured using conventional blood culture flasks, and the other samples underwent pathogen testing using mNGS. The results showed that the positive rate of mNGS was 96.77%, while that of the blood culture flask method was 70.97% (p < 0.05). A total of 29 pathogens were detected by mNGS, namely 24 bacteria, one fungus, and four viruses. A total of 10 pathogens were detected using the bacterial blood culture method, namely nine bacteria and one fungus. The final judgment of the PDAP's causative pathogenic microorganism was made by combining the clinical condition, response to therapy, and the whole-genome sequencing findings. For mNGS, the sensitivity was 96.77%, the specificity was 83.33%, the positive predictive value was 96.77%, and the negative predictive value was 83.33%. For the blood culture flask method, the sensitivity was 70.97%, the specificity was 100%, the positive predictive value was 100%, and the negative predictive value was 0%. In conclusion, mNGS had a shorter detection time for diagnosing peritoneal dialysis-related peritonitis pathogens, with a higher positive rate than traditional bacterial cultures, providing significant advantages in diagnosing rare pathogens.

A novel Colorcult hemoculture in early diagnosis of blood stream infection: A boon in meagre resources

Blood stream infections (BSIs) account for major cause of morbidity and mortality. Conventional hemoculture methods are time consuming, with poor sensitivity and high risk of contamination. Automated blood culture systems are highly sensitive but are overpriced. Hence, it is not suitable for resource poor settings. In this background, this study was done to analyze the sensitivity of novel non automated chemical sensor-based blood culture and to compare the time to detection of both conventional and colorimetric method of blood culture. Blood specimens were collected from 50 study participants, admitted with suspected sepsis, inoculated into the colorcult &amp; conventional culture media and processed as per CLSI guidelines. Culture positive by both conventional and colorcult method was six (12%). Most of the study isolates were Gram negative bacteria (83.4%). The isolates were detected at the earliest within 96 hours by colorcult, whereas by conventional method only three (50 %) were detected within 96 hours and another three (50%) after seven days only. Owing to the escalation in BSIs associated morbidity and mortality, early diagnosis by a simple, cost effective, highly sensitive and specific method is an absolute necessity in resource poor settings. The performance of colorcult is superior to conventional method but cost effective than automated methods and can be used as an alternative method in the diagnosis of BSIs.

Analysis of Changes in Variation of Neutrophil and Monocyte Parameters, Including Volume, Conductivity and Scatter in Sepsis Patients and Healthy Controls: A Cross-sectional Study

Introduction: Sepsis continues to be a leading cause of mortality and prolonged hospitalisation. The conventional method of blood culture, while considered the gold standard, has limitations such as contamination and delayed reporting. The examination of peripheral smears has uncovered signs suggestive of septicaemia; however, these findings suffer from inter-observer variability and reliance on staining quality. Aim: To investigate the variation of neutrophil and monocyte parameters, including Volume, Conductivity, and Scatter (VCS), in sepsis compared to healthy controls. Materials and Methods: A cross-sectional analytical study was conducted at the Himalayan Institute of Medical Sciences, Swami Rama Himalayan University in Dehradun, Uttarakhand, over the course of one year, from January 2021 to December 2021, involving patients over 18 years of age categorised into sepsis group based on clinical suspicion, sepsis screen, blood culture, and Sequential Organ Failure Assessment (SOFA) score (n=117). A group of healthy controls was also included (n=140). Haematological investigations were performed using the DXH 800 Haematology Analyser (Beckman Coulter, CA, USA) with VCS Technology. Categorical variables were analysed using the Chi-square test, while non parametric data was compared using the Mann-Whitney U test. Results: The average age in the sepsis group was 50.17±13.17 years, while in the control group, it was 38.14±8.78 years. The results revealed higher White Blood Cell (WBC) counts (16.76±7.39)×103 / cumm in the sepsis group compared to healthy controls (6.68±1.42)×103 , absolute neutrophil counts (13.74±7.280)×103 in sepsis patients, and eosinopenia in the sepsis group (0.0114±0.0104)×103 compared to controls (0.23±0.116)×103 . Moreover, mean neutrophilic volumes (158.00±14.840) and monocytic volumes (182.58±18.64) were higher in the sepsis group, while they were lower in healthy controls, which were (149.52±5.23 and 171.17±6.28), respectively. Axial light loss for neutrophil and monocyte was 142.40±11.78 and 121.50±17.93, respectively, while it was lower in healthy controls showing a value of 135.51±7.63 and 119.45±8.25, respectively. Furthermore, mean neutrophilic and monocytic conductivity and scatter were decreased in sepsis. The observed higher WBC counts and absolute neutrophil counts in sepsis patients suggest a premature release of neutrophils from the bone marrow. The alterations in cell volume reflect an immune response. Additionally, the overall scatter of neutrophils and monocytes was reduced, accompanied by increased cellular transparency. Conclusion: The present study contributes valuable insights into the pathophysiological mechanisms underlying sepsis, emphasising the dynamic interplay between immune cells and their functional characteristics. Understanding these variations in cellular parameters could potentially aid in the development of more targeted diagnostic and therapeutic approaches for sepsis, ultimately improving patient outcomes. Further research is warranted to delve deeper into the specific mechanisms driving these observed changes and to explore their clinical implications in the context of sepsis management.

Detection of Salmonella species by Automated Blood Culture Method in Suspected Cases of Enteric Fever

Background: Enteric fever caused by Salmonella species is a systemic febrile illness that can be fatal since its characterization in 1880. Enteric fever may remain asymptomatic carrier and sometimes fatal that it can be difficult to diagnose based on symptoms alone because they are similar to those of other febrile disorders. In developing countries, enteric fever is common global health problem due to poor standard of personal hygiene, contaminated food, inadequate safe water and sanitation. Methods: This is a cross-sectional study that was conducted in the Microbiology Department of Rajshahi Medical College Hospital for one year from January 2021 to December 2021 to identify Salmonella species by Automated Blood Culture (Fastidious Antibiotic Neutralization method) followed by subculture in MacConkey’s agar, Xylose lysine deoxycholate agar, and Salmonella Shigella agar media in suspected enteric fever cases. This study included suspected cases of enteric fever as determined by the physician who fulfilled the inclusion and exclusion criteria. Results: Among 127 suspected cases of enteric fever 64(50.39%) culture positive bacteria isolated by automated blood culture (FAN) method and out of that Salmonella species 38 (59.37%) and others bacteria 26(40.63%) following subculture in MacConkey’s agar, Xylose lysine deoxycholate agar and Salmonella Shigella agar media. Regarding gender, the majorities (61.4%) of culture-positive patients were male, and the majority (65.35%) came from rural areas. Conclusion: Especially for patients who have received antibiotics, we propose that the automated blood culture approach followed by subculture could be employed as a novel diagnostic method for typhoid fever. TAJ 2023; 36: No-2: 71-76

Impact of quality improving interventions on blood culture contamination rate- A prospective observational study

Background: Blood cultures (BC) is the gold standard for diagnosis of bacteremia but false positive blood cultures due to specimen contamination can lead to increased usage of unnecessary antibiotics and have negative impact on antimicrobial stewardship program. Overgrowth of contaminants can also obscure growth of genuine pathogens and hamper accurate diagnosis. The aim of this study is to evaluate effectiveness of Quality improving (QI) interventions to reduce the blood culture contamination rate Material and Methods: Prospective observational study, conducted at a public sector hospital, for a total duration of 6 months. A taskforce developed and implemented QI interventions to reduce blood culture contamination, and evaluated effectiveness of these interventions by comparing contamination rates prior to and after implementation of the intervention. Results: The post-intervention average contamination rate was significantly lower with the mean contamination rate of 4.7 ± 1.5% as compared to baseline BC contamination rate which was 10.6 ± 1.8% (p=0.013) Conclusion: Quality improving (QI) interventions significantly reduced the blood culture contamination rate. Keywords: Quality improvement, Interventions, Blood culture, Contamination.

The impact of the BIOFIRE® Blood Culture Identification 2 Panel on antimicrobial treatment of children with suspected systemic inflammatory response syndrome and sepsis.

This study aimed to assess the therapeutic effects of implementing the BioFire® Blood Culture Identification 2 (BCID2) Panel (bioMérieux, Marcy l'Etoile, France) in the clinical practice of children with sepsis.This retrospective cross-sectional study included children from 15days of age to 18years old with sepsis and of whom the BCID2 Panel was studied from the positive blood culture. If the antimicrobial treatment was changed according to the results of BCID2 Panel, it was recorded and re-grouped as targeted antimicrobial therapy, de-escalation of the antimicrobial treatment and shifting to another antimicrobial drug if any antimicrobial resistance was detected. Seven-days and thirty-days mortality rate was recorded.Thirty-two patients with 36 septic episodes with positive BCID2 Panel results were included. The median age was 10months 15days (ranging from 15days to 16.5years). The mean difference between having positive results by the BCID2 Panel and conventional culture methods was 82.2 ± 45.4h (ranging from 12.3 to 207h). Effect of the BCID2 Panel on the antimicrobial treatment was detected in 69.4% of the episodes (n = 25). Glycopeptides were ceased at 6 patients, piperacillin/tazobactam was ceased at 6 patients, and cefotaxime was ceased at one patient and de-escalation was achieved in 13 episodes which formed the 36.1% of the initial antimicrobial treatment. Conclusion: The BCID2 Panel had an important impact on the patients care and optimization according to the principles of antimicrobial stewardship. The BCID2 Panel may be one of the key items for rapid and accurate diagnosis in children with sepsis. Blood culture is still the mainstay for especially detection of antimicrobial drug resistance, while BCID2 tests had several advantages such as speed and diagnostic accuracy as good as blood culture. What is Known: • The BioFire®Blood Culture Identification 2 (BCID2) Panel (bioMérieux, Marcy l'Etoile, France) is a well-validated assay that allows for the simultaneous identification of 43 nucleic acid targets associated with bloodstream infections within about 1h. What is New: • BCID2 Panel had a direct impact on the patient's treatment decision at the 69.4% of the sepsis episodes. • The clinicians received the microbiological results 82.2h earlier with BCID2 panel compared to blood culture methods with antimicrobial resistanceandde-escalation of the antimicrobial drugs was achieved at 13 episodes which formed the 36.1% of the initial treatment.

620. Development and Validation of a Novel qPCR Assay for Differentiation of Common Candida Pathogens including C. auris

Abstract Background Candida is a common asymptomatic fungal colonizer of humans. Of the 200+ total species in the genus Candida, only a few are known to cause infection. The most common are C. albicans, C. glabrata, C. parapsilosis, C. tropicalis, C. krusei, and C. auris. In the USA, candidemia is the fourth most common cause of bloodstream infections in hospitalized patients. Although antifungal medication can treat candidemia, mortality rates remain between 30 and 60%. Eurofins Viracor has developed a qPCR assay that will qualitatively detect and differentiate any of the 6 previously listed Candida species in whole blood. Speciation is critical for treatment due to variable drug susceptibility among different species, particularly C. auris due to its multi-drug resistance. Methods Multiple primer/probe mixes were designed around and within the ITS1, 5.8s, ITS2, and 28s ribosomal subunit regions in each of the six Candida target species. The 5.8s and 28s regions are heavily conserved, while the ITS regions are highly divergent and used to differentiate species. After optimization, the six primer/probe mixes were partially multiplexed resulting in four mixes covering all species. DNA was extracted from human blood spiked with viable Candida using the MagMAX™ DNA Multi-Sample Ultra 2.0 Kit, and eluates were amplified using the Applied Biosystems™ 7500 Fast platform. Results Each assay met all validation acceptance criteria and demonstrated excellent accuracy, precision, linearity and dynamic range, and sensitivity. 32 non-Candida fungal and bacterial pathogens that commonly cause bloodstream infections were used to assess specificity. All specificity samples were negative for all Candida targets. To assess accuracy, 75 of 75 Candida samples representing all six species across a range of concentrations were tested and correctly identified. 24 of 25 known-negative samples were negative, with one positive C. glabrata CT result observed just below the cutoff. Conclusion Traditional blood culture methods for Candida species identification can be too time consuming for a quickly progressing infection. The Candida qPCR assay described here not only detects an infection faster but provides accurate speciation allowing for rapid and targeted antifungal treatment options for the patient. Disclosures Clayton Thomas, BS in Biology, Eurofins Viracor: Employee James Grantham, n/a, Eurofins Viracor: Employee Emily Lute, n/a, Eurofins Viracor: Employee Jerod Davidson, n/a, Eurofins Viracor: Employee Scott Cowden, MS, MB ASCP(CM), Eurofins Viracor: Employee Jamie Nutt, n/a, Eurofins Viracor: Employee

Gender and Age Dependent Bacteriological Etiology of Community-Acquired Blood Stream Infection

Background: Bloodstream infection constitutes one of the most serious situations in infectious disease. Objective: The purpose of the presents study was to determine the gender and age dependent etiology of community- acquired urinary blood stream infection. Methodology: This was a retrospective analysis of blood samples from clinically suspected cases of blood stream infection. Samples were collected at Ad-din Women’s Medical College Hospital, Dhaka &amp; Rushmono Specialized Hospital, Dhaka from January 2018 to September 2018. All the samples were collected from inpatient’s and outpatient’s department of our hospital during the study period and processed in Microbiology laboratory. The BD BACTEC FX40 automated blood culture method was used to isolate bacterial pathogens and antimicrobial susceptibility test was performed by Kirby-Bauer disc diffusion method. Results: A total of 483 (16.1%) pathogens were isolated from 3018 bacteremia suspect patient blood specimens. Gram-negative cocci (58.4%) were predominant organisms recovered followed by Gram-positive bacilli (41.6%). Majority of BSI were caused by Gram negative bacteria predominantly Salmonella Typhi (31.1%). Salmonella species was found less prevalent in the children (55%) and more frequent in the age groups 16 to 30 years (85.0%) and more than 60 years (100.0%). Acinetobacter species was found less prevalent in age group 16 to 30 years (7.5%). Escherichia coli was found only few prevalent in the 1 to 15 (0.5%) and higher frequent in the age groups 16 to 30 years (2.5%). Salmonella species isolates appeared to be sensitive to ceftriaxone (91.3%), meropenem (90.7%) and cotrimoxazole (76%). Sensitivity rates of ciprofloxacin, gentamycin and levofloxacin were 87.3% for coagulase-negative Staphylococcus species (CoNS). Conclusion: Both patients’ age and gender are significant factors in determining bloodstream infection. Bangladesh Journal of Medical Microbiology, January 2023;17(1):3-9

CORRELATION OF C-REACTIVE PROTEIN AND BLOOD CULTURE IN NEONATAL SEPSIS AT INTEGRAL INSTITUTE OF MEDICAL SCIENCES AND RESEARCH, HOSPITAL LUCKNOW (UTTAR PRADESH)

Objective: Neonatal sepsis is a clinical condition of bacteremia characterized by systemic signs and symptoms of infection in children under 4 weeks of age. It involves systemic infections in newborns, including septicemia, meningitis, pneumonia, arthritis, osteomyelitis, and urinary tract infections. C-reactive protein (CRP) is an abnormal β-globulin produced by the liver during any inflammatory process. The gold standard method to diagnose neonatal sepsis is blood culture; however, it is time-consuming and requires a well-equipped laboratory and trained personnel. To study the correlation between CRP and blood culture in neonatal sepsis. Methods: The present study was a retrospective study conducted at the Integral Institute of Medical Sciences and Research, Hospital Lucknow. (UP) All the indoor neonates attending at the Integral Institute of Medical Science and Research Hospital who’s both parameters, CRP as well as blood culture were noted in the microbiology department register during my study period. In the blood culture, samples were collected aseptically and processed by either conventional or automated blood culture methods. CRP estimation was done by a latex agglutination card test. CRP was reported as positive if agglutination particles were seen. Results: CRP positivity rate: out of 235 samples, 72 (30.64%) were positive and 163 (69.36%) were negative. Blood culture positivity rate: out of 118 samples, 71 (60.17%) cases were culture positive and 47 (39.83%) were negative. After comparison of CRP samples with blood culture samples, 90 samples were tested for both blood culture and CRP, and the babies ages were under 4 weeks. Conclusion: So from our study, we are concluding that blood culture is the gold standard method for the diagnosis of neonatal sepsis, and although we can use CRP as the screening method, this test is not specific enough to be relied upon as the only indicator.

Deteksi Bakteri Salmonella sp. dengan Kultur Darah Pada Pasien Widal Positif di Laboratorium Klinik X

Until now, the gold standard test for diagnosing typhoid fever is a bacterial culture or also known as a gall culture examination. Blood cultures have the best sensitivity (40–60%) when performed in the first-early second week. Purpose: this study aims to identify the presence of Salmonella sp. in the blood of Widal positive patients. Method: this research is a descriptive study with a cross-sectional approach. This study used the blood culture method. The media used in this study were oxgall, SSA, KIA and IMViC media. The results showed that 20% of Salmonella typhi bacteria were found, 7% of other bacteria and 73% of bacteria did not grow on the media. The macroscopic characteristics of salmonella bacteria found were round colonies, clear in color, convex surface, flat colony edges, semi mucoid consistency, no reduction of tellurit. The microscopic characteristics are rod-shaped, spread arrangement, red in color and are Gram negative. Cultivation on IMViC media showed Indole -, MR +, VP -, Citrate -, KIA medium alkaline slope, acid base, H2S +, gas -. Conclusion: Salmonella thypi was found in the blood cultures of positive widal patients.

A new application of multiplex PCR combined with membrane biochip assay for rapid detection of 9 common pathogens in sepsis.

Rapid and accurate identification of specific sepsis pathogens is critical for patient treatment and disease control. This study aimed to establish a new application for the rapid identification of common pathogens in patients with suspected sepsis and evaluate its role in clinical application. A multiplex PCR assay was designed to simultaneously amplify specific conserved regions of nine common pathogenic microorganisms in sepsis, including Acinetobacter baumannii, Escherichia coli, Klebsiella pneumonia, Pseudomonas aeruginosa, Enterococcus faecalis, Staphylococcus aureus, Staphylococcus epidermidis, Streptococcus pneumonia, and Candida albicans. The PCR products were analyzed by a membrane biochip. The analytical sensitivity of the assay was determined at a range of 5-100 copies/reaction for each standard strain, and the detection range was 20-200 cfu/reaction in a series dilution of simulated clinical samples at different concentrations. Out of the 179 clinical samples, the positive rate for pathogens detected by the membrane biochip assay and blood culture method was 20.11% (36/179) and 18.44% (33/179), respectively. However, by comparing the positive rate of the nine common pathogens we detected, the membrane biochip assay tended to be more sensitive than the blood culture method (20.11% vs 15.64%). The clinical sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of the membrane biochip assay were 92.9%, 93.2%, 72.2% and 98.6%, respectively. Generally, this multiplex PCR combined membrane biochip assay can be used to detect major sepsis pathogens, and is useful for early initiation of effective antimicrobial treatment, and is feasible for sepsis pathogens identification in routine clinical practice.

A Comparative Study of Conventional and Automated Blood Culture System in Adult Patients

Introduction: Blood culture is considered as the gold standard for the diagnosis of bloodstream infection. Conventional blood culture system is less sensitive and takes longer duration for the detection of bloodstream infections whereas automated blood culture system is more sensitive and rapid in detecting causative organisms of bloodstream infections. This prospective study was undertaken to compare the automated blood culture system with the conventional blood culture system for the identification of microbial pathogens in bloodstream infections. Method: This prospective study was done in Department of Microbiology, Silchar Medical College &amp; Hospital, Silchar for a period of 7 months from November 2021 to May 2022. Blood samples from the patients were inoculated into BD BACTEC Plus Aerobic culture vials for automated blood culture system and Brain Heart Infusion broth for conventional blood culture system. Positive bottles flagged by the automated machine were isolated and identified by doing routine subcultures on Blood agar and MacConkey agar and necessary biochemical tests. The conventional blood culture bottles were processed as per standard protocols. Result: Out of 123 samples, 21(17.09%) showed culture positivity by automated blood culture method and 15(12.19%) showed culture positivity by conventional blood culture method. The most common isolate was Staphylococcus aureus followed by Escherichia coli in automated method whereas the most common isolate in conventional method was Staphylococcus aureus followed by Klebsiella pneumoniae. Conclusion: This study concluded that automated blood culture method is more sensitive than conventional method and detects the presence of microorganisms rapidly causing bloodstream infections. Keywords: Bloodstream infection, Automated blood culture, Conventional blood culture

Thromboembolic phenomenon to bilateral lower limbs is a catastrophic complication of blood culture-negative infective endocarditis

Background: Blood culture-negative infective endocarditis (BCNIE) refers to infective endocarditis (IE) with negative microorganism growth using usual blood culture method. It poses dilemma in terms of diagnosis, associated complication, and treatment. The study aims to highlight the occurrence of acute bilateral lower limb ischemia in BCNIE and its management approach. Case presentation: We present a BCNIE patient who developed acute bilateral lower limb ischemia, Rutherford stage 2b despite on course of antibiotic treatment. Patient underwent emergency bilateral open femoral embolectomy and right profundaplasty. A full six-weeks course of antibiotics and short-term anticoagulant successfully reverse the acute ischemic event. Conclusion: This case highlighted that despite the rarity of BCNIE causing limb ischemia, it can still occur due to significant risk factors. The early recognition of acute limb ischemia and prompt treatment are deemed important in saving the threatened limb from being amputated.

Combination of the insulator‐based dielectrophoresis and hydrodynamic methods for separating bacteria smaller than 3 μm in bloodstream infection: Numerical simulation approach

AbstractBloodstream infections have a high mortality rate with &gt;80,000 deaths per year in North America. The inability to detect pathogens quickly in the early stages of the infection causes high mortality. Such inability has led to a growing interest in developing a rapid, sensitive, and specific method for identifying these pathogens. The rapid detection of bloodstream infections requires the rapid and efficient separation of bacteria from the blood. But the problem is that the number of bacteria is much lower than other blood components. The blood culture step needs to be accomplished first for bacteria identification and antibiotic susceptibility testing. As the blood culture is time‐consuming, a method based on the insulator‐based has been presented that increases the number of bacteria by combining the blood culture method and increasing the concentration. In this model, the dielectrophoresis technique was utilized in a curved microchannel with a constriction for sorting three particle sizes including 9, 7–4 μm, as well as smaller than 3 μm. The results showed that the applied voltage and the channel dimensions affect separation efficiency. Suppose these values are properly selected (for example, a voltage of 110 V that was causing the maximum electric field of 200 V/cm). The proposed model can completely (100%) separate larger than 9 μm and smaller than 3 μm particles. The proposed model has simple geometry and is considered an appropriate technique for sorting all bacteria separation in bloodstream infection.

A diagnostic test of real-time PCR detection in the diagnosis of clinical bloodstream infection

Blood culture remains the standard for diagnosing bloodstream infections, but it is difficult to identify bacteria directly and timeliness. The real-time polymerase chain reaction (PCR) has the potential to fill this diagnostic gap. This study intends to explore the sensitivity and specificity of PCR in detecting bloodstream infection pathogens and to compare it with routine blood culture to explore its clinical application value. A total of 126 patients with bloodstream infections collected from various clinical departments of The First Hospital of Hebei Medical University. The patient's sample was divided into two parts. The one for multiplex PCR detection was performed using the Pathogeno Elite Multiplex PCR kit. Another blood culture was a fully automatic blood culture system from Autobio company. Among the 126 patients, a total of 17 pathogens were detected by PCR and blood culture both methods. PCR detected a total of 43 positive samples and 83 negative samples. Five samples were positive with blood culture, and 81 were negative. The negative predictive value of PCR was 0.98, with a sensitivity of 0.71 and a specificity of 0.68. A total of 38 specimens were positive for PCR but negative for blood culture, and 2 samples were positive for blood culture but negative for PCR. The top 5 pathogens with PCR detection were Epstein-Barr virus (27 cases), Human herpes virus 5 (9 cases), Klebsiella pneumoniae (5 cases), Staphylococcus (5 cases), and Stenotrophomonas maltophilia (4 cases). PCR detection can rapidly identify more pathogens and even multi-pathogen infections. Therefore, PCR testing may improve pathogen detection in patients with suspected bloodstream infections, enabling targeted treatment of patients.

Developing a multiplex PCR-based assay kit for bloodstream infection by analyzing genomic big data.

BackgroundIn recent years, the incidence of bloodstream infections (BSI) has increased, the composition of pathogenic bacteria has changed, and drug resistance among bacteria has gradually increased due to the widespread use of interventional techniques, broad‐spectrum antibacterial drugs, hormones, and immunosuppressive agents. Here, we have developed a multiplex PCR assay kit for the detection of pathogens (14 Gram‐negative bacteria, 15 Gram‐positive bacteria, and 4 fungi) in whole blood from patients with BSI using five‐color fluorescent multiplex PCR followed by capillary electrophoresis. Our assay exhibits a diagnosis of higher quality and an improved detection rate for common pathogens.MethodsA local genome DNA database of 33 pathogenic bacteria was constructed. Next, “Exhaustive” primer search of the full coding sequence of the reference genomes of these bacteria was performed. Panels with minimal interactions between primers and amplicons were selected by random sampling and testing by a recursive algorithm. Primers and Mg2+ concentrations and PCR reaction procedures were optimized to maximize the detection efficacy.ResultsThe LOD of the kit was determined as 100 copies/μl. Using clinical samples, results generated by this kit and regular blood culture method were found to be 95.08% consistent. Additionally, six pathogens which were unidentifiable by blood culture were successfully detected by this kit.ConclusionOur study provided a bioinformatics approach to the challenge of primer design in multiplex PCR, and combined with optimized wet lab practice, a multiplex PCR‐based assay kit for BSI with higher sensitivity and accuracy than blood culture was produced.