Clinical Evaluation of Nontuberculous Mycobacteria (NTM) Elite Agar, a New Medium for the Isolation of NTM: a Multicenter Study.

Nontuberculous mycobacteria (NTM) are gaining interest with the increased number of infected patients. NTM Elite agar is designed specifically for the isolation of NTM without the decontamination step. We assessed the clinical performance of this medium combined with Vitek mass spectrometry (MS) matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) technology for the isolation and identification of NTM in a prospective multicenter study, including 15 laboratories (24 hospitals). A total of 2,567 samples from patients with suspected NTM infection were analyzed (1,782 sputa, 434 bronchial aspirates, 200 bronchoalveolar lavage samples, 34 bronchial lavage samples, and 117 other samples). A total of 220 samples (8.6%) were positive with existing laboratory methods against 330 with NTM Elite agar (12.8%). Using the combination of both methods, 437 isolates of NTM were detected in 400 positive samples (15.6% of samples). In total, 140 samples of the standard procedures (SP) and 98 of the NTM Elite agar were contaminated. NTM Elite agar showed a higher performance for rapidly growing mycobacteria (RGM) species than SP (7% versus 3%, P < 0.001). A trend has been noted for the Mycobacterium avium complex (4% with SP versus 3% with NTM Elite agar, P = 0.06). The time to positivity was similar (P = 0.13) between groups. However, the time to positivity was significantly shorter for the RGM in subgroup analysis (7 days with NTM and 6 days with SP, P = 0.01). NTM Elite agar has been shown to be useful for the recovery of NTM species, especially for the RGM. Using NTM Elite agar + Vitek MS system in combination with SP increases the number of NTM isolated from clinical samples.

Microevolution, reinfection and highly complex genomic diversity in patients with sequential isolates of Mycobacterium abscessus.

Mycobacterium abscessus is an opportunistic, extensively drug-resistant non-tuberculous mycobacterium. Few genomic studies consider its diversity in persistent infections. Our aim was to characterize microevolution/reinfection events in persistent infections. Fifty-three sequential isolates from 14 patients were sequenced to determine SNV-based distances, assign resistance mutations and characterize plasmids. Genomic analysis revealed 12 persistent cases (0-13 differential SNVs), one reinfection (15,956 SNVs) and one very complex case (23 sequential isolates over 192 months), in which a first period of persistence (58 months) involving the same genotype 1 was followed by identification of a genotype 2 (76 SNVs) in 6 additional alternating isolates; additionally, ten transient genotypes (88-243 SNVs) were found. A macrolide resistance mutation was identified from the second isolate. Despite high diversity, the genotypes shared a common phylogenetic ancestor and some coexisted in the same specimens. Genomic analysis is required to access the true intra-patient complexity behind persistent infections involving M. abscessus.

Bacteriophage-Antibiotic Combination Therapy against Pseudomonas aeruginosa.

Phage therapy is an alternative therapy that is being used as the last resource against infections caused by multidrug-resistant bacteria after the failure of standard treatments. Pseudomonas aeruginosa can cause pneumonia, septicemia, urinary tract, and surgery site infections mainly in immunocompromised people, although it can cause infections in many different patient profiles. Cystic fibrosis patients are particularly vulnerable. In vitro and in vivo studies of phage therapy against P. aeruginosa include both bacteriophages alone and combined with antibiotics. However, the former is the most promising strategy utilized in clinical infections. This review summarizes the recent studies of phage-antibiotic combinations, highlighting the synergistic effects of in vitro and in vivo experiments and successful treatments in patients.

Rare Recombinant GI.5[P4] Norovirus That Caused a Large Foodborne Outbreak of Gastroenteritis in a Hotel in Spain in 2021.

Noroviruses are among the most important causes of acute gastroenteritis (AGE). In summer 2021, a large outbreak of norovirus infections affecting 163 patients, including 15 norovirus-confirmed food handlers, occurred in a hotel in Murcia in southeast Spain. A rare GI.5[P4] norovirus strain was identified as the cause of the outbreak. The epidemiological investigation determined that norovirus transmission might have been initiated through an infected food handler. The food safety inspection found that some symptomatic food handlers continued working during illness. Molecular investigation with whole-genome and ORF1 sequencing provided enhanced genetic discrimination over ORF2 sequencing alone and enabled differentiation of the GI.5[P4] strains into separate subclusters, suggesting different chains of transmission. These recombinant viruses have been identified circulating globally over the last 5 years, warranting further global surveillance. IMPORTANCE Due to the large genetic diversity of noroviruses, it is important to enhance the discriminatory power of typing techniques to differentiate strains when investigating outbreaks and elucidating transmission chains. This study highlights the importance of (i) using whole-genome sequencing to ensure genetic differentiation of GI noroviruses to track chains of transmission during outbreak investigations and (ii) the adherence of symptomatic food handlers to work exclusion rules and strict hand hygiene practices. To our knowledge, this study provides the first full-length genome sequences of GI.5[P4] strains apart from the prototype strain.

Subspecies Distribution and Antimicrobial Susceptibility Testing of Mycobacterium abscessus Clinical Isolates in Madrid, Spain: a Retrospective Multicenter Study.

Mycobacterium abscessus (MABS) is the most pathogenic and drug-resistant rapidly growing mycobacteria. However, studies on MABS epidemiology, especially those focusing on subspecies level, are scarce. We aimed to determine MABS subspecies distribution and its correlation with phenotypic and genotypic antibiotic profiles. A retrospective multicenter study of 96 clinical MABS isolates in Madrid between 2016 to 2021 was conducted. Identification at the subspecies level and resistance to macrolides and aminoglycosides were performed by the GenoType NTM-DR assay. The MICs of 11 antimicrobials tested against MABS isolates were determined using the broth microdilution method (RAPMYCOI Sensititer titration plates). Clinical isolates included 50 (52.1%) MABS subsp. abscessus; 33 (34.4%) MABS subsp. massiliense; and 13 (13.5%) MABS subsp. bolletii. The lowest resistance rates corresponded to amikacin (2.1%), linezolid (6.3%), cefoxitin (7.3%), and imipenem (14.6%), and the highest to doxycycline (100.0%), ciprofloxacin (89.6%), moxifloxacin (82.3%), cotrimoxazole (82.3%), tobramycin (81.3%), and clarithromycin (50.0% at day 14 of incubation). Regarding tigecycline, although there are no susceptibility breakpoints, all strains but one showed MICs ≤ 1 µg/mL. Four isolates harbored mutations at positions 2058/9 of the rrl gene, one strain harbored a mutation at position 1408 of the rrl gene, and 18/50 harbored the T28C substitution at erm(41) gene. Agreement of the GenoType results with clarithromycin and amikacin susceptibility testing was 99.0% (95/96). The rate of MABS isolates showed an upward trend during the study period, being M. abscessus subsp. abscessus the most frequently isolated subspecies. Amikacin, cefoxitin, linezolid, and imipenem showed great in vitro activity. The GenoType NTM-DR assay provides a reliable and complementary tool to broth microdilution for drug resistance detection. IMPORTANCE Infections caused by Mycobacterium abscessus (MABS) are increasingly being reported worldwide. Identifying MABS subspecies and assessing their phenotypic resistance profiles are crucial for optimal management and better patient outcomes. M. abscessus subspecies differ in erm(41) gene functionality, which is a critical determinant of macrolide resistance. Additionally, resistance profiles of MABS and the subspecies distribution can vary geographically, highlighting the importance of understanding local epidemiology and resistance patterns. This study provides valuable insights into the epidemiology and resistance patterns of MABS and its subspecies in Madrid. Elevated resistance rates were observed for several recommended antimicrobials, emphasizing the need for cautious drug use. Furthermore, we assessed the GenoType NTM-DR assay, which examines principal mutations in macrolides and aminoglycosides resistance-related genes. We observed a high level of agreement between the GenoType NTM-DR assay and the microdilution method, indicating its usefulness as an initial tool for early initiation of appropriate therapy.

Alternatives to Antibiotics against Mycobacterium abscessus.

Mycobacterium abscessus complex is extremely difficult to treat. Intrinsic and acquired bacterial resistance makes this species one of the most challenging pathogens and treatments last from months to years, associated with potential risky antibiotic toxicity and a high number of failures. Nonantibiotic antimicrobial agents against this microorganism have recently been studied so as to offer an alternative to current drugs. This review summarizes recent research on different strategies such as host modulation using stem cells, photodynamic therapy, antibiofilm therapy, phage therapy, nanoparticles, vaccines and antimicrobial peptides against M. abscessus both in vitro and in vivo.

Advances in Bacteriophage Therapy against Relevant MultiDrug-Resistant Pathogens.

The increase of multiresistance in bacteria and the shortage of new antibiotics in the market is becoming a major public health concern. The World Health Organization (WHO) has declared critical priority to develop new antimicrobials against three types of bacteria: carbapenem-resistant A. baumannii, carbapenem-resistant P. aeruginosa and carbapenem-resistant and ESBL-producing Enterobacteriaceae. Phage therapy is a promising alternative therapy with renewed research in Western countries. This field includes studies in vitro, in vivo, clinical trials and clinical cases of patients receiving phages as the last resource after failure of standard treatments due to multidrug resistance. Importantly, this alternative treatment has been shown to be more effective when administered in combination with antibiotics, including infections with biofilm formation. This review summarizes the most recent studies of this strategy in animal models, case reports and clinical trials to deal with infections caused by resistant A. baumannii, K. pneumoniae, E. coli, and P. aeruginosa strains, as well as discusses the main limitations of phage therapy.

Compassionate use of tocilizumab in severe SARS-CoV2 pneumonia.

INTRODUCTION: Tocilizumab (TCZ) is an interleukin-6 receptor antagonist, which has been used for the treatment of severe SARS-CoV-2 pneumonia (SSP), which aims to ameliorate the cytokine release syndrome (CRS) induced acute respiratory distress syndrome (ARDS). However, there are no consistent data about who might benefit most from it. METHODS: We administered TCZ on a compassionate-use basis to patients with SSP who were hospitalized (excluding intensive care and intubated cases) and who required oxygen support to have a saturation >93%. The primary endpoint was intubation or death after 24 h of its administration. Patients received at least one dose of 400 mg intravenous TCZ from March 8, 2020 to April 20, 2020. RESULTS: A total of 207 patients were studied and 186 analyzed. The mean age was 65 years and 68% were male patients. A coexisting condition was present in 68% of cases. Prognostic factors of death were older age, higher IL-6, d-dimer and high-sensitivity C-reactive protein (HSCRP), lower total lymphocytes, and severe disease that requires additional oxygen support. The primary endpoint (intubation or death) was significantly worst (37% vs 13%, p < 0·001) in those receiving the drug when the oxygen support was high (FiO2 >0.5%). CONCLUSIONS: TCZ is well tolerated in patients with SSP, but it has a limited effect on the evolution of cases with high oxygen support needs.