Laboratory automation, informatics, and artificial intelligence: current and future perspectives in clinical microbiology

Clinical diagnostic laboratories produce one product—information—and for this to be valuable, the information must be clinically relevant, accurate, and timely. Although diagnostic information can clearly improve patient outcomes and decrease healthcare costs, technological challenges and laboratory workflow practices affect the timeliness and clinical value of diagnostics. This article will examine how prioritizing laboratory practices in a patient-oriented approach can be used to optimize technology advances for improved patient care

In vitro antibacterial activity of ceftazidime/avibactam in combination against planktonic and biofilm carbapenemase-producing Klebsiella pneumoniae isolated from blood

Abstract Objectives The aim of this study was to report on in vitro tests of antibacterial activity of ceftazidime/avibactam in combination against planktonic or biofilm KPC carbapenemase-producing Klebsiella pneumoniae (KPC-Kp), the rate of KPC-Kp blood isolates in University of Perugia Hospital over a 5-year period, and their antimicrobial susceptibility patterns. Methods The antibacterial activity of ceftazidime/avibactam in combination with other antimicrobials was assessed against planktonic and biofilm bacteria by Etest and checkerboard assay. A retrospective review of laboratory data was performed to evaluate the rate of KPC-Kp from blood samples and their antimicrobial susceptibility patterns. Results Between 2014 and 2019, 130/4241 (3.1%) KPC-Kp were identified from blood cultures. Their rate increased from 2.3% in 2014–2015 to 4.5% over the last 3 years. Overall, 4.6% (6/130) of KPC-Kp isolates were susceptible to meropenem, 65.4% (85/130) to colistin, 65.1% (84/129) to tigecycline, 34.6% (45/130) to amikacin, 36.2% (42/116) to gentamicin, 40.2% (39/97) to fosfomycin and 91.5% (65/71) to ceftazidime/avibactam. Five of six ceftazidime/avibactam-resistant KPC-Kp were isolated from patients not treated with ceftazidime/avibactam. Synergism was detected both by Etest and checkerboard assay for the combination of ceftazidime/avibactam plus meropenem against planktonic isolates, whilst lower bactericidal activity was observed in biofilm KPC-Kp isolates. Conclusions Our in vitro data suggest that the combination of ceftazidime/avibactam plus meropenem has a synergistic antibacterial activity against planktonic bacteria, whilst a lower activity was detected against biofilm, suggesting worse clinical outcomes whenever biofilm infections are present. Further analyses are required to confirm these results before extending them to clinical practice

A synthetic peptide as a novel anticryptococcal agent.

Summary An engineered, killer decapeptide (KP) has been synthesized based on the sequence of a recombinant, single-chain anti-idiotypic antibody (KT-scFv) acting as a functional internal image of a yeast killer toxin. Killer decapeptide exerted a strong fungicidal activity against Candida albicans, which was attributed to peptide interaction with β-glucan. As this polysaccharide is also a critical component of the cryptococcal cell wall, we wondered whether KP was also active against Cryptococcus neoformans, a human pathogen of increasing medical importance. We found that KP was able to kill both capsular and acapsular C. neoformans cells in vitro. Furthermore, KP impaired the production of specific C. neoformans virulence factors including protease and urease activity and capsule formation, rendering the fungus more susceptible to natural effector cells. In vivo treatment with KP significantly reduced fungal burden in mice with cryptococcosis and, importantly, protected the majority of immunosuppressed animals from an otherwise lethal infection. Given the relevance of cryptococcosis in immunocompromised individuals and the inability of conventional drugs to completely resolve the infection, the results of the present study indicate KP as an ideal candidate for further studies on novel anticryptococcal agents

In vitro Antimicrobial Activity of Ampicillin-Ceftriaxone and Ampicillin-Ertapenem Combinations Against Clinical Isolates of Enterococcus faecalis with High Levels of Aminoglycoside Resistance

This paper reports on the in vitro antimicrobial activity of ampicillin-ceftriaxone and ampicillin-ertapenem combinations against five strains of E. faecalis with high-level aminoglycoside resistance recovered from blood of septicemic patients. Double disk diffusion test and time killing curves were used. A bacteriostatic synergistic effect between ampicillin and ceftriaxone was detected using the disk diffusion assay for three of the five enterococcal strains studied. With the same three isolates enhanced bactericidal activity was also observed using time killing experiments. Overall, for these three strains, after 24 hr of contact, a decrease ≥ 2 log10 from the initial bacterial inoculum was registered with most ampicillin-ceftriaxone combinations, reaching with some of them a colony reduction ≥ 3 log10. This bactericidal interaction was negatively influenced increasing the bacterial inoculum. In all five isolates neither a bacteriostatic nor a bactericidal cooperation was observed for ampicillin combined with 2 mg/l of ertapenem

Interleukin-4 Causes Susceptibility to Invasive Pulmonary Aspergillosis through Suppression of Protective Type I Responses

Aspergillus fumigatus, an opportunistic fungal pathogen, causes multiple allergic and non-allergic airway diseases. Invasive pulmonary aspergillosis (IPA) is a nonallergic, life-threatening disease of immunocompromised patients. In a murine model of IPA, interleukin (IL)—4-deficient (IL-4−/−) BALB/c mice were used to examine the role of IL-4 in lung pathology and immune responses. IL-4−/− mice were more resistant than wild-type mice to infection caused by multiple intranasal injections of viable A. fumigatus conidia. Resistance was associated with decreased lung inflammatory pathology, impaired T helper (Th)—2 responses (including lung eosinophilia), and an IL-12—dependent Th1 response. In contrast, development of host-detrimental antifungal Th2 cells occurred in IL-12−/− and interferon-γ−/− mice and in IL-4−/− mice when subjected to IL-12 neutralization. These results demonstrate that IL-4 renders mice susceptible to infection with A. fumigatus by inhibition of protective Th1 responses. IL-4 appears to have a distinct role in the pathogenesis of allergic and nonallergic lung diseases caused by the fungu

Evaluation of GeneXpert® system for detection of methicillin-resistant Staphyloccocus aureus in clinical samples

Infections caused by methicillin-resistant Staphyloccocus aureus strains (MRSA) have reached epidemic proportions globally, being the major cause of nosocomial infections. Rapid identification of MRSA in nasal swabs or in clinical samples is considered a useful strategy for control and treatment of these infections. GeneXpert system (Cepheid Europe,Vira-Solelch, Maurence-Scopont-France) can detect by real-time PCR in approximately one hour methicillin-resistant S. aureus or coagulase-negative staphylococci (CoNS) in clinical samples, in comparison with 24 hours for the culture or 48 hours for the antimicrobial susceptibility testing. In this study GeneXpert system was compared with traditional tests for MRSA detection in nasal swabs, bloodcultures and surgical wound swabs. Materials and methods. Eighteen nasal swabs, 23 blood-cultures and 13 surgical wound swabs were tested. The samples were cultured on blood-agar and mannitol-salt agar. Identification of isolates was carried out with traditional tests (Gram staining, catalase, coagulase) and automatic Phoenix system. Methicillin-susceptibility was evaluated according to 2010 CLSI guidelines. GeneXpert system was performed according to manufacturers instructions, by using the specific kits and methicillin-resistance was detected by amplification of the genic sequences spa, SCC e mecA. Results. The results showed a 100% accordance between GeneXpert system and traditional tests for detection of methicillin-resistant staphylococci. In particular, among 18 nasal swabs, no MRSA was detected, while 1 bloodculture (4.3%) and 4 surgical wound swabs (30.7%) were positive for MRSA. Conclusions. GeneXpert system allows a rapid detection of MRSA in clinical samples and shows the same sensitivity and specificity as traditional tests. Therefore, it represents a further effective diagnostic method for prevention and treatment of nosocomial infections due to methicillin-resistant staphylococci

Prevalence of cervical colonization by Ureaplasma parvum, Ureaplasma urealyticum, Mycoplasma hominis and Mycoplasma genitalium in childbearing age women by a commercially available multiplex real-time PCR: An Italian observational multicentre study

Background: Mycoplasmas are frequently isolated from the genital tract. New molecular PCR-based methods for the detection of mycoplasmas can better define the real epidemiology of these microorganisms. The aim of this study was to evaluate the prevalence of mycoplasmas in a population of childbearing age women by means of PCR. Methods: This 21-month multicentre observational study was conducted at four Italian clinical microbiology laboratories. Women reporting symptoms of vaginitis/cervicitis, or with history of infertility, pregnancy, miscarriage or preterm birth were included. Detection of Ureaplasma parvum, Ureaplasma urealyticum, Mycoplasma hominis, Mycoplasma genitalium was performed from cervical swabs by means of a commercially available multiplex real-time PCR. Results: a total of 1761 women fulfilled the inclusion criteria and were included in the study. The overall prevalence was: U. parvum 38.3%, U. urealyticum 9%, M. hominis 8.6% and M. genitalium 0.6%. The proportion of foreign patients positive for U. parvum was significantly higher compared to Italian patients (37% vs 30.1%, p = 0.007) and also for overall mycoplasma colonization (53.4% vs 45.8%, p = 0.011). The number of symptomatic patients positive for M. hominis was significantly higher than that of negative controls (2.9% vs 1%, p = 0.036). A significant positive trend in mycoplasma colonization was found in relation to the pregnancy week for U. urealyticum (p = 0.015), M. hominis (p = 0.044) and for overall mycoplasma colonization (p = 0.002). Conclusion: multiplex RT-PCR can be a valuable tool to evaluate the real epidemiology of cervical mycoplasma colonization. Keywords: Genital mycoplasmas, Mycoplasma genitalium, Mycoplasma hominis, Ureaplasma parvum, Ureaplasma urealyticum, Real-time PC