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.

Many methods are being tried for rapid and accurate identification of sepsis-causing microorganisms. We analyzed the performance of three different preparation methods [MBT Sepsityper IVD Kit (Bruker Daltonics GmbH, Germany), sodium dodecyl sulfate (SDS) lysis, and differential centrifugation with protein extraction (Centrifugation +PE)] and compared in standard and Sepsityper modules of the Bruker Biotyper MALDI-TOF MS for direct identification of bacteria from 240 positive blood culture bottles of BACTEC FX (Becton Dickinson, USA). By using the standard module, correct identification at species level (score ≥2) was done in 46.7% of the samples with SDS lysis, 44.2% with centrifugation +PE, and 25.4% with the Sepsityper kit. These ratios at the genus level (score range 1.70-1.99) were 34.6%, 31.3%, and 32.5%, respectively. With SDS lysis (195), more bacteria were identified correctly than centrifugation +PE (181) and the Sepsityper kit (139). A statistically significant difference was found between SDS and the Sepsityper kit and Centrifugation +PE and the Sepsityper kit (P < 0.001, both). By using the Sepsityper module, correct identification at species level (score ≥1.8) was determined in 74.2% of the samples with SDS lysis and centrifugation +PE each and 55% with the Sepsityper kit. These ratios at the genus level (score range 1.60-1.79) were 16.3%, 10%, and 19.2%, respectively. SDS lysis (217) had significantly higher identification rates than centrifugation +PE (202) and the Sepsityper kit (178) (P = 0.028 and P < 0.001). A statistically significant difference was also observed between centrifugation +PE and the Sepsityper kit (P < 0.001). Best performance was obtained with SDS lysis among the methods. Although better performance was achieved by using Sepsityper software module, risk of misidentification should not be ignored.
OBJECTIVE: Sepsis is a life-threatening condition, and rapid and accurate identification of the causative microorganisms from blood cultures is crucial for timely and effective treatment. Although there are many studies on direct identification from blood cultures with MALDI-TOF MS, further standardization is still needed. In our study, we analyzed the performance of three different preparation methods and compared by using two analysis modules of the Bruker Biotyper MALDI-TOF MS for direct identification of bacteria from numerous positive blood culture bottles. The literature reports a limited number of studies that compare different preparation methods for direct blood culture identification, processing a large number of blood samples concurrently and evaluating the same samples as in our study. Moreover, although SDS is used very frequently in medical laboratories, there are few studies on direct identification from blood culture bottles. In our study, the highest correct identification rate was observed with the SDS method.

Here, we report the complete genome sequences of Lactococcus petauri strains 473AN and 473GN, isolated from the blood culture of a Japanese patient with infective endocarditis. The complete genomes of 473AN and 473GN consist of single chromosomes of 2,065,772 and 2,094,461 bp, respectively.

UNASSIGNED: The diagnosis of tuberculosis (TB) disease and TB infection (TBI) remains a challenge, and there is a need for non-invasive and blood-based methods to differentiate TB from conditions mimicking TB (CMTB), TBI, and healthy controls (HC). We aimed to determine whether combination of cytokines and established biomarkers could discriminate between 1) TB and CMTB 2) TB and TBI 3) TBI and HC.
UNASSIGNED: We used hemoglobin, total white blood cell count, neutrophils, monocytes, C-reactive protein, and ten Meso Scale Discovery analyzed cytokines (interleukin (IL)-1β, IL-2, IL-4, IL-6, IL-8, IL-10, IL-12p70, IL-13, interferon (IFN)-ɣ, and tumor necrosis factor (TNF)-α) in TruCulture whole blood tubes stimulated by lipopolysaccharides (LPS), zymosan (ZYM), anti-CD3/28 (CD3), and unstimulated (Null) to develop three index tests able to differentiate TB from CMTB and TBI, and TBI from HC.
UNASSIGNED: In 52 persons with CMTB (n=9), TB (n=23), TBI (n=10), and HC (n=10), a combination of cytokines (LPS-IFN-ɣ, ZYM-IFN-ɣ, ZYM-TNF-α, ZYM-IL-1β, LPS-IL-4, and ZYM-IL-6) and neutrophil count could differentiate TB from CMTB with a sensitivity of 52.2% (95% CI: 30.9%-73.4%) and a specificity of 100 % (66.4%-100%). Null- IFN-ɣ, Null-IL-8, CD3-IL-6, CD3-IL-8, CD3-IL-13, and ZYM IL-1b discriminated TB from TBI with a sensitivity of 73.9% (56.5% - 91.3%) and a specificity of 100% (69.2-100). Cytokines and established biomarkers failed to differentiate TBI from HC with ≥ 98% specificity.
UNASSIGNED: Selected cytokines may serve as blood-based add-on tests to detect TB in a low-endemic setting, although these results need to be validated.

Clostridium perfringens (C. perfringens) is an anaerobic, spore-forming Gram-positive rod responsible for necrotizing gangrene, bacteremia in patients with cancer or gastrointestinal tract infection. C. perfringens virulence is due in large part to toxin production. In 2014, a new enterotoxin, BEC (binary enterotoxin of Clostridium perfringens) encoded by becA and becB genes, distinct from enterotoxin (CPE) encoded by the cpe gene, has been described. BEC-producing strains can be causative agents of acute gastroenteritis in humans. We present herein the case of a 64-year-old man who presented to the emergency department of Toulouse University Hospital with pneumonia and septic shock, without digestive symptoms. Blood cultures showed C. perfringens bacteremia and despite appropriate antibiotic treatment the patient passed away 7 h after admission. The characterization of the strain by whole genome sequencing revealed the presence of typical genes of C. perfringens: plc gene (alpha-toxin, phospholipase C) and pfoA (theta-toxin, perfringolysine). Surprisingly, this strain also harbored becA and becB genes encoding the recently described BEC toxin. Interestingly, alpha-toxin typing of our isolate and other published BEC isolates showed that they belonged to different PLC subtypes, confirming the high genetic diversity of these strains. To our knowledge, it is the first clinical case reporting bacteremia due to a BEC-producing C. perfringens isolate.

Management of suspected early-onset sepsis (EOS) is undergoing continuous evolution aiming to limit antibiotic overtreatment, yet current data on the level of overtreatment are only available for a select number of countries. This study aimed to determine antibiotic initiation and continuation rates for suspected EOS, along with the incidence of culture-proven EOS in The Netherlands. In this retrospective study from 2019 to 2021, data were collected from 15 Dutch hospitals, comprising 13 regional hospitals equipped with Level I-II facilities and 2 academic hospitals equipped with Level IV facilities. Data included birth rates, number of neonates started on antibiotics for suspected EOS, number of neonates that continued treatment beyond 48 h and number of neonates with culture-proven EOS. Additionally, blood culture results were documented. Data were analysed both collectively and separately for regional and academic hospitals. A total of 103,492 live-born neonates were included. In 4755 neonates (4.6%, 95% CI 4.5-4.7), antibiotic therapy was started for suspected EOS, and in 2399 neonates (2.3%, 95% CI 2.2-2.4), antibiotic treatment was continued beyond 48 h. Incidence of culture-proven EOS was 1.1 cases per 1000 live births (0.11%, 95% CI 0.09-0.14). Overall, for each culture-proven EOS case, 40.6 neonates were started on antibiotics and in 21.7 neonates therapy was continued. Large variations in treatment rates were observed across all hospitals, with the number of neonates initiated and continued on antibiotics per culture-proven EOS case varying from 4 to 90 and from 4 to 56, respectively. The high number of antibiotic prescriptions compared to the EOS incidence and wide variety in clinical practice among hospitals in The Netherlands underscore both the need and potential for a novel approach to the management of neonates with suspected EOS.

Bloodstream infections (BSI) caused by multidrug-resistant (MDR) bacteria, pose a major threat for patients, especially for those who are immunosuppressed. Rapid pathogen detection and characterization from positive blood cultures are crucial in the management of patients with BSI to enable an adequate and timely antimicrobial therapy. This study aimed to investigate the potential role of the Molecular Mouse system, a new CE IVD molecular test designed to rapidly detect the causative agents of bacteremia and their resistance determinants, in the management of the therapy in critically ill patients. Agreement between the results of the Molecular Mouse and the conventional routine method was also considered. Overall, 100 positive blood cultures were collected from septic critically ill patients from May 2023 to January 2024 and analyzed with Molecular Mouse and routine protocols. The new instrument consistently agreed with the routine protocols in the case of monomicrobial blood cultures, while some discrepancies were obtained in the polymicrobial samples. Antimicrobial resistance genes were detected in 35 samples, with vanA and CTX-M-1/9 groups being the most frequently detected targets. Therapy was adjusted in 42 critically ill patients confirming the importance of new rapid molecular tests in the management of positive blood cultures, to adjust empirical therapy and use new antibiotics accurately.

Discospondylitis is a well-recognized disease in dogs, but the relative prevalence of causal infectious agents and efficiency of relevant diagnostic tests are not well-established. Medical record review identified 117 dogs diagnosed with discospondylitis in our clinic over a 5-year period. In 32 dogs, discospondylitis was diagnosed as an incidental imaging finding; 24 of these dogs had concomitant neoplasia. A likely causal infection was identified in 45 of the remaining 85 dogs in which blood and urine cultures, serology for Brucella spp., and galactomannan fungal antigen testing were recommended. Ten dogs were diagnosed with Brucella canis, and ten were diagnosed with suspected fungal infection. Brucella suis serology was negative in all 35 dogs that were tested. Blood cultures were positive in 28 of 71 (39%) tested dogs, and urine culture was positive in 12 of 79 (15%). Cultures were positive from the lesion site of four of eight dogs that underwent surgery and one of the five dogs that underwent image-guided lesion sample collection. Subluxation secondary to discospondylitis was stabilized with metallic implants in four dogs. A similar proportion of known satisfactory treatment outcomes at last follow-up were recorded in dogs that had suspected fungal disease, other bacterial infections, or were Brucella-positive and in those dogs with imaging diagnosis only, although some individuals continued to receive anti-microbial agents or showed recurrent signs. These data support the value of blood culture in discospondylitis and suggest a relatively high prevalence of infection with Brucella spp. and suspected fungal infection.

We report discovery of a new bacterial genus and species of the family Pasteurellaceae by using phylogenetic and metabolic analysis. The bacterium, Emayella augustorita, was isolated from blood cultures of a patient in France diagnosed with an adenocarcinoma of the intestines and who was treated with a biliary prosthesis placement.

UNASSIGNED: Blood and urine are the most common culture testing for sepsis patients. This study aimed to compare clinical characteristics and outcomes of sepsis patients by blood and urine culture positivity and to identify factors associated with positive cultures.
UNASSIGNED: This retrospective study included patients aged ≥16 years with sepsis identified by the Sepsis-3 criteria presenting to the emergency department at four hospitals between 2017 and 2019 in Australia. Patient clinical outcomes were in-hospital mortality, intensive care unit (ICU) admission, hospital length of stay, and representation following discharge. Four culture groups were defined based on the positivity of blood cultures (BC) and urine cultures (UC) ordered within 24 h of triage.
UNASSIGNED: Of 4109 patient encounters with sepsis, 2730 (66%) were nonbacteremic, urine culture-negative (BC-UC-); 767 (19%) nonbacteremic, urine culture-positive (BC-UC+); 359 (9%) bacteremic, urine culture-negative (BC+UC-); and 253 (6%) bacteremic, urine culture-positive (BC+UC+). Compared with BC-UC- patients, BC+UC- patients had the highest risk of ICU admission (adjusted odds ratio [AOR] 95% CI: 1.60 [1.18-2.18]) while BC-UC+ patients had lowest risk (adjusted odds ratio [AOR]: 0.56 [0.41-0.76]). BC+UC- patients had the highest risk of 3-day representation (AOR: 1.51 [1.02-2.25]) and second longest hospital stay (adjusted relative risk 1.17 [1.03-1.34]). Antibiotic administration before sample collection for culture was associated with lower odds of positive blood or urine culture results (AOR: 0.38, p < 0.0001).
UNASSIGNED: Enhanced clinical care should be beneficial for nongenitourinary sepsis patients (BC+UC-) who had the highest comparative risk of adverse clinical outcomes. Every effort needs to be made to collect relevant culture samples before antibiotic administration, to follow up on culture results, and tailor treatment accordingly.