SARS-CoV-2 mutant spectra as variant of concern nurseries: endless variation?

Introduction: SARS-CoV-2 isolates of a given clade may contain low frequency genomes that encode amino acids or deletions which are typical of a different clade. Methods: Here we use high resolution ultra-deep sequencing to analyze SARS-CoV-2 mutant spectra. Results: In 6 out of 11 SARS-CoV-2 isolates from COVID-19 patients, the mutant spectrum of the spike (S)-coding region included two or more amino acids or deletions, that correspond to discordant viral clades. A similar observation is reported for laboratory populations of SARS-CoV-2 USA-WA1/2020, following a cell culture infection in the presence of remdesivir, ribavirin or their combinations. Moreover, some of the clade-discordant genome residues are found in the same haplotype within an amplicon. Discussion: We evaluate possible interpretations of these findings, and reviewed precedents for rapid selection of genomes with multiple mutations in RNA viruses. These considerations suggest that intra-host evolution may be sufficient to generate minority sequences which are closely related to sequences typical of other clades. The results provide a model for the origin of variants of concern during epidemic spread─in particular Omicron lineages─that does not require prolonged infection, involvement of immunocompromised individuals, or participation of intermediate, non-human hosts.

Synergism between remdesivir and ribavirin leads to SARS-CoV-2 extinction in cell culture.

BACKGROUND AND PURPOSE: There is a need for effective anti-COVID-19 treatments, mainly for individuals at risk of severe disease such as the elderly and the immunosuppressed. Drug repositioning has proved effective in identifying drugs that can find a new application for the control of coronavirus disease, in particular COVID-19. The purpose of the present study was to find synergistic antiviral combinations for COVID-19 based on lethal mutagenesis. EXPERIMENTAL APPROACH: The effect of combinations of remdesivir and ribavirin on the infectivity of SARS-CoV-2 in cell culture has been tested. Viral populations were monitored by ultra-deep sequencing, and the decrease of infectivity as a result of the treatment was measured. KEY RESULTS: Remdesivir and ribavirin exerted a synergistic inhibitory activity against SARS-CoV-2, quantified both by CompuSyn (Chou-Talalay method) and Synergy Finder (ZIP-score model). In serial passage experiments, virus extinction was readily achieved with remdesivir-ribavirin combinations at concentrations well below their cytotoxic 50 value, but not with the drugs used individually. Deep sequencing of treated viral populations showed that remdesivir, ribavirin, and their combinations evoked significant increases of the number of viral mutations and haplotypes, as well as modification of diversity indices that characterize viral quasi-species. CONCLUSION AND IMPLICATIONS: SARS-CoV-2 extinction can be achieved by synergistic combination treatments based on lethal mutagenesis. In addition, the results offer prospects of triple drug treatments for effective SARS-CoV-2 suppression.

Multicenter evaluation of BioFire JI panel related to improved microbiological diagnostics on acute osteoarticular infections.

Microbiological diagnosis of osteoarticular infections (OI) is crucial for a successful treatment. A prospective multicenter study including 262 synovial fluids with suspicion of acute OI was performed between July 2021 and October of 2022. BioFire Joint Infection Panel multiplex-PCR test was performed and results were compared with conventional cultures of synovial fluid specimens. In total, 136 microorganisms were detected, and fourteen samples were positive for more than one microorganism. In monomicrobial infections (n = 87) agreement with culture was 69%. In 26 samples, the multiplex PCR yield an additional positive result when culture result was negative. It helped in the detection of fastidious microorganisms as K. kingae and N. gonorrhoeae. This multiplex PCR has proven to be a useful technique that can be used for patients with high suspicion of acute OI in a rapid and automated manner.

Multicenter evaluation of the BIOFIRE Joint Infection Panel for the detection of bacteria, yeast, and AMR genes in synovial fluid samples.

The bioMérieux BIOFIRE Joint Infection (JI) Panel is a multiplex in vitro diagnostic test for the simultaneous and rapid (~1 h) detection of 39 potential pathogens and antimicrobial resistance (AMR) genes directly from synovial fluid (SF) samples. Thirty-one species or groups of microorganisms are included in the kit, as well as several AMR genes. This study, performed to evaluate the BIOFIRE JI Panel for regulatory clearance, provides data from a multicenter evaluation of 1,544 prospectively collected residual SF samples with performance compared to standard-of-care (SOC) culture for organisms or polymerase chain reaction (PCR) and sequencing for AMR genes. The BIOFIRE JI Panel demonstrated a sensitivity of 90.9% or greater for all but six organisms and a positive percent agreement (PPA) of 100% for all AMR genes. The BIOFIRE JI Panel demonstrated a specificity of 98.5% or greater for detection of all organisms and a negative percent agreement (NPA) of 95.7% or greater for all AMR genes. The BIOFIRE JI Panel provides an improvement over SOC culture, with a substantially shorter time to result for both organisms and AMR genes with excellent sensitivity/PPA and specificity/NPA, and is anticipated to provide timely and actionable diagnostic information for joint infections in a variety of clinical scenarios.

Atypical Mutational Spectrum of SARS-CoV-2 Replicating in the Presence of Ribavirin.

We report that ribavirin exerts an inhibitory and mutagenic activity on SARS-CoV-2-infecting Vero cells, with a therapeutic index higher than 10. Deep sequencing analysis of the mutant spectrum of SARS-CoV-2 replicating in the absence or presence of ribavirin indicated an increase in the number of mutations, but not in deletions, and modification of diversity indices, expected from a mutagenic activity. Notably, the major mutation types enhanced by replication in the presence of ribavirin were A→G and U→C transitions, a pattern which is opposite to the dominance of G→A and C→U transitions previously described for most RNA viruses. Implications of the inhibitory activity of ribavirin, and the atypical mutational bias produced on SARS-CoV-2, for the search for synergistic anti-COVID-19 lethal mutagen combinations are discussed.

Epidemiology and in vitro antimicrobial susceptibility of aerobic Actinomycetales in a clinical setting.

INTRODUCTION: The incidence of infections caused by aerobic actinomycetes is increasing. Recent changes in taxonomy and the variability in susceptibility patterns among species make necessary a proper identification and antibiotic susceptibility testing. MATERIAL AND METHODS: Fifty-three strains of aerobic actinomycetes were identified by MALDI-TOF MS using the VITEK MS Mycobacterium/Nocardia kit (bioMérieux, France) in a tertiary hospital in Spain during a six-year period. Antimicrobial susceptibility testing of the isolates was performed using the Sensititre Rapmycoi microdilution panel (Thermo Fisher Scientific, Massachusetts, USA). RESULTS: Forty strains of Nocardia spp. were identified in the study, being N. farcinica and N. cyriacigeorgica the most prevalent ones. All isolates were susceptible to linezolid and the resistance to amikacin was only observed in one isolate of Gordonia sputi. Resistance to cotrimoxazole was only found in five isolates. CONCLUSIONS: Routine identification and antimicrobial susceptibility testing of aerobic actinomycetes is advisable for an efficient identification of species and effective treatment.

Epidemiology and in vitro antimicrobial susceptibility of aerobic Actinomycetales in a clinical setting / Epidemiología y sensibilidad in vitro de especies de Actinomycetales aerobios en la rutina asistencial

Introduction: The incidence of infections caused by aerobic actinomycetes is increasing. Recent changes in taxonomy and the variability in susceptibility patterns among species make necessary a proper identification and antibiotic susceptibility testing. Material and methods: Fifty-three strains of aerobic actinomycetes were identified by MALDI-TOF MS using the VITEK MS Mycobacterium/Nocardia kit (bioMérieux, France) in a tertiary hospital in Spain during a six-year period. Antimicrobial susceptibility testing of the isolates was performed using the Sensititre Rapmycoi microdilution panel (Thermo Fisher Scientific, Massachusetts, USA). Results: Forty strains of Nocardia spp. were identified in the study, being N. farcinica and N. cyriacigeorgica the most prevalent ones. All isolates were susceptible to linezolid and the resistance to amikacin was only observed in one isolate of Gordonia sputi. Resistance to cotrimoxazole was only found in five isolates. Conclusions: Routine identification and antimicrobial susceptibility testing of aerobic actinomycetes is advisable for an efficient identification of species and effective treatment.(AU)

Introducción: La incidencia de infecciones por actinomicetos aerobios está aumentando. Los recientes cambios en la taxonomía y la variabilidad en la sensibilidad entre especies hacen necesaria una identificación y estudio de sensibilidad adecuados. Material y métodos: Se identificaron 53 cepas de actinomicetos aerobios mediante MALDI-TOF utilizando el kit VITEK-MS Mycobacterium/Nocardia (bioMérieux, Francia) en un hospital terciario español durante seis años. Los estudios de sensibilidad de los aislados se realizaron utilizando el panel de microdilución Sensititre Rapmycoi (Thermo Fisher Scientific, Massachusetts, EE. UU.). Resultados: Se identificaron 40 cepas de Nocardia spp., siendo Nocardia farcinica y Nocardia cyriacigeorgica las más prevalentes. Todos los aislados fueron sensibles a linezolid, y solo se detectó resistencia a amikacina en un aislado de Gordonia sputi. Solo se encontró resistencia al cotrimoxazol en cinco aislados. Conclusiones: Es aconsejable realizar la identificación de rutina y las pruebas de sensibilidad antimicrobiana de los actinomicetos aerobios para conseguir una identificación eficiente de las especies y un tratamiento eficaz.(AU)

Urinary Tract Infection in Pediatrics: Study of Uropathogens and Their Resistance in a Madrid Hospital

Introduction: Resistance to antibiotics is a growing problem with repercussions on the choice of first-line treatment in urinary tract infection (UTI) in childhood. Objectives: To know the current pattern of antibiotic susceptibility/resistance of the most frequent germs that cause UTI in our healthcare area. Secondary objective is to know the evolution of these patterns over time. Patients and Methods: A cross-sectional retrospective study of UTI episodes in a first-level hospital in two periods: 1st January 2008-31th December 2010 and 1st January 2017-31th December 2019 through a review of medical records, recording the following variables: Age, sex, fever, hospital admission, uropathy/bladder dysfunction, antibiotic prophylaxis. Results: First period: 174 UTI episodes (156 patients); Second period: 266 UTI episodes (218 patients). The most frequently isolated germ was E. coli, but in patients with uropathy or bladder dysfunction, the percentage of different germs is greater. A significant increase in resistance to amoxicillin/clavulanate (from 12.2 to 24%) is observed between both periods, it remains stable and in an acceptable range for gentamicin, cotrimoxazole and slightly increases to first-generation cephalosporins. In patients with uropathy/bladder dysfunction, resistance to all these antibiotics is significantly increased. Conclusions: The increased resistance of the most frequent uropathogens in the UTI of the pediatric population of our healthcare area to amoxicillin/clavulanate makes it unsuitable as empirical therapy. First-generation cephalosporins are an adequate alternative in patients without risk factors (AU)

Introducción: La resistencia a antibióticos es unproblema creciente con repercusión en la elección deltratamiento empírico en la infección del tracto urinario(ITU) en la infancia.Objetivos: Conocer cómo ha evolucionado la incidencia de uropatógenos causantes de ITU en poblaciónpediátrica y su patrón de resistencia antibiótica en el área11 de salud de la Comunidad de Madrid. Secundariamente,patrón de resistencia actual.Materiales y Métodos: Estudio de corte transversal retrospectivo de los episodios de ITU de un hospitalde Nivel I en dos periodos: 1 enero 2008 a 31 diciembre2010 y 1 enero 2017 a 31 diciembre 2019. Se recogió:Edad, sexo, fiebre, uropatía/disfunción vesical, profilaxisantibiótica, microorganismo aislado y su antibiograma. Serealizó estudio descriptivo de todas las variables recogidas ycomparación de proporciones independientes, utilizando laprueba chi cuadrado,tomando como variable de agrupación cada periodo. El análisis estadístico fue realizado con elprograma EPIDAT v 4.2.Resultados: Primer periodo: 174 episodios de ITU(156 pacientes); Segundo periodo: 266 episodios de ITU(218 pacientes). El germen más frecuentemente aisladofue E. coli, siendo mayor el aislamiento de gérmenes distintos en pacientes urópatas/disfunción vesical. Se observa un aumento significativo de la resistencia a amoxicilina/clavulánico, se mantiene estable y en rango aceptablepara gentamicina, cotrimoxazol y aumenta levemente a cefalosporinas de primera generación. En los pacientes conuropatía/disfunción vesical las resistencias a todos estos antibióticos se incrementansignificativamente.Conclusiones: El aumento de la resistencia de losuropatógenos más frecuentes en la ITU de la poblaciónpediátrica a amoxicilina/clavulánico del área sanitaria estudiada, lo hace no apto como terapia empírica. Las cefalosporinas de primera generación suponen una adecuadaalternativa en pacientes sin factores de riesgo (AU)

Microbiological Characterization of Cutibacterium acnes Strains Isolated from Prosthetic Joint Infections.

Aims: This study aimed to characterize 79 Cutibacterium acnes strains isolated from prosthetic joint infections (PJIs) originated from eight European hospitals. Methods: Isolates were phylotyped according to the single-locus sequence typing (SLST) scheme. We evaluated the ability of the biofilm formation of C. acnes strains isolated from PJIs and 84 isolates recovered from healthy skin. Antibiotic susceptibility testing of planktonic and biofilm cells of PJI isolates and skin isolates was performed. Results: Most of the isolates from PJIs belonged to the SLST class H/phylotype IB (34.2%), followed by class D/phylotype IA1 (21.5%), class A/phylotype IA1 (18.9%), and class K/phylotype II (13.9%). All tested isolates were biofilm producers; no difference in biofilm formation was observed between the healthy skin group and the PJI group of strains. Planktonic and sessile cells of C. acnes remained highly susceptible to a broad spectrum of antibiotics, including beta-lactams, clindamycin, fluoroquinolones, linezolid, rifampin, and vancomycin. The minimal inhibitory concentrations (MICs) for planktonic and biofilm states coincided in most cases. However, the minimal biofilm eradication concentration (MBEC) was high for all antimicrobial drugs tested (>32 mg/L), except for rifampin (2 mg/L). Conclusions: C. acnes strains isolated from healthy skin were able to produce biofilm to the same extent as isolates recovered from PJIs. All C. acnes strains in planktonic and sessile states were susceptible to most antibiotics commonly used for PJI treatment, although rifampin was the only antimicrobial agent able to eradicate C. acnes embedded in biofilm.

SARS-CoV-2 Mutant Spectra at Different Depth Levels Reveal an Overwhelming Abundance of Low Frequency Mutations.

Populations of RNA viruses are composed of complex and dynamic mixtures of variant genomes that are termed mutant spectra or mutant clouds. This applies also to SARS-CoV-2, and mutations that are detected at low frequency in an infected individual can be dominant (represented in the consensus sequence) in subsequent variants of interest or variants of concern. Here we briefly review the main conclusions of our work on mutant spectrum characterization of hepatitis C virus (HCV) and SARS-CoV-2 at the nucleotide and amino acid levels and address the following two new questions derived from previous results: (i) how is the SARS-CoV-2 mutant and deletion spectrum composition in diagnostic samples, when examined at progressively lower cut-off mutant frequency values in ultra-deep sequencing; (ii) how the frequency distribution of minority amino acid substitutions in SARS-CoV-2 compares with that of HCV sampled also from infected patients. The main conclusions are the following: (i) the number of different mutations found at low frequency in SARS-CoV-2 mutant spectra increases dramatically (50- to 100-fold) as the cut-off frequency for mutation detection is lowered from 0.5% to 0.1%, and (ii) that, contrary to HCV, SARS-CoV-2 mutant spectra exhibit a deficit of intermediate frequency amino acid substitutions. The possible origin and implications of mutant spectrum differences among RNA viruses are discussed.

SARS-CoV-2 Point Mutation and Deletion Spectra and Their Association with Different Disease Outcomes.

Mutant spectra of RNA viruses are important to understand viral pathogenesis and response to selective pressures. There is a need to characterize the complexity of mutant spectra in coronaviruses sampled from infected patients. In particular, the possible relationship between SARS-CoV-2 mutant spectrum complexity and disease associations has not been established. In the present study, we report an ultradeep sequencing (UDS) analysis of the mutant spectrum of amplicons from the nsp12 (polymerase)- and spike (S)-coding regions of 30 nasopharyngeal isolates (diagnostic samples) of SARS-CoV-2 of the first COVID-19 pandemic wave (Madrid, Spain, April 2020) classified according to the severity of ensuing COVID-19. Low-frequency mutations and deletions, counted relative to the consensus sequence of the corresponding isolate, were overwhelmingly abundant. We show that the average number of different point mutations, mutations per haplotype, and several diversity indices was significantly higher in SARS-CoV-2 isolated from patients who developed mild disease than in those associated with moderate or severe disease (exitus). No such bias was observed with RNA deletions. Location of amino acid substitutions in the three-dimensional structures of nsp12 (polymerase) and S suggest significant structural or functional effects. Thus, patients who develop mild symptoms may be a richer source of genetic variants of SARS-CoV-2 than patients with moderate or severe COVID-19. IMPORTANCE The study shows that mutant spectra of SARS-CoV-2 from diagnostic samples differ in point mutation abundance and complexity and that significantly larger values were observed in virus from patients who developed mild COVID-19 symptoms. Mutant spectrum complexity is not a uniform trait among isolates. The nature and location of low-frequency amino acid substitutions present in mutant spectra anticipate great potential for phenotypic diversification of SARS-CoV-2.

Vaccine breakthrough infections with SARS-CoV-2 Alpha mirror mutations in Delta Plus, Iota, and Omicron.

Replication of SARS-CoV-2 in the human population is defined by distributions of mutants that are present at different frequencies within the infected host and can be detected by ultra-deep sequencing techniques. In this study, we examined the SARS-CoV-2 mutant spectra of amplicons from the spike-coding (S-coding) region of 5 nasopharyngeal isolates derived from patients with vaccine breakthrough. Interestingly, all patients became infected with the Alpha variant, but amino acid substitutions that correspond to the Delta Plus, Iota, and Omicron variants were present in the mutant spectra of the resident virus. Deep sequencing analysis of SARS-CoV-2 from patients with vaccine breakthrough revealed a rich reservoir of mutant types and may also identify tolerated substitutions that can be represented in epidemiologically dominant variants.

Genomic Analysis of Cutibacterium acnes Strains Isolated from Prosthetic Joint Infections.

Cutibacterium acnes is a common cause of prosthetic joint infections (PJIs). The C. acnes population can be divided into six main phylotypes (IA1, IA2, IB, IC, II and III) that are associated with different clinical conditions and normal skin. A single-locus sequence typing (SLST) scheme can distinguish ten main SLST types: A-E (all IA1), F (IA2), G (IC), H (IB), K (II), L (III). We genome-sequenced and compared 16 strains of C. acnes isolated from healthy skin (n = 4) and PJIs (n = 12), including six PJI cases with a good outcome (four shoulder PJIs, one hip PJI, one knee PJI) and six with infection relapse (three shoulder PJIs, three hip PJIs). The sequenced strains belonged to four different phylotypes (IA1, IA2, IB and II) and seven different SLST types. All five type IB strains (all SLST type H1) were PJI isolates (three hip PJIs, two shoulder PJIs), and four of these caused infection relapse (three hip PJIs, one shoulder PJI). Isolates from PJI cases with a good outcome belonged to three different phylotypes (IA, IB, II). Interestingly, four strains (three strains from PJI cases with good outcome and one strain from healthy skin) contained a linear plasmid; these strains belonged to different SLST types (A1, C1, F4, H1) and were isolated in three different hospitals. This study suggests that type IB strains have the potential to cause infection relapse, in particular regarding hip PJIs. Moreover, our study revealed that strains belonging to the same SLST type can differ in their accessory genome in different geographic locations, indicative of microevolution.

Cutibacterium spp. isolated from orthopaedic implant-associated infection: A not-so-slowly growing organism.

INTRODUCTION: It has been reported that microbiological diagnosis of Cutibacterium spp. infection requires a prolonged incubation time (up to 14 days). We present our experience with regard to incubation time for detection of Cutibacterium spp. in orthopaedic samples over a 10-year period. METHODS: One hundred and nineteen samples were included in this retrospective study. Fifty-three were implants (having previously undergone sonication), 64 were periprosthetic tissue biopsies and two were synovial fluids. Atkins's criteria were used for interpreting the isolates. Quantification and number of days until a culture became positive for Cutibacterium spp. were evaluated. RESULTS: The median number of days to detection of a clinically significant isolate and a contaminant was 4 days. No clinically significant isolates grew after day eight. CONCLUSION: Most clinically significant isolates of Cutibacterium spp. are detected in the first 7 days of incubation, although a recommendation of prolonged incubation (up to 14 days) appears to be necessary for detecting other organisms.

A Janus-Faced Bacterium: Host-Beneficial and -Detrimental Roles of Cutibacterium acnes.

The bacterial species Cutibacterium acnes (formerly known as Propionibacterium acnes) is tightly associated with humans. It is the dominant bacterium in sebaceous regions of the human skin, where it preferentially colonizes the pilosebaceous unit. Multiple strains of C. acnes that belong to phylogenetically distinct types can co-exist. In this review we summarize and discuss the current knowledge of C. acnes regarding bacterial properties and traits that allow host colonization and play major roles in host-bacterium interactions and also regarding the host responses that C. acnes can trigger. These responses can have beneficial or detrimental consequences for the host. In the first part of the review, we highlight and critically review disease associations of C. acnes, in particular acne vulgaris, implant-associated infections and native infections. Here, we also analyse the current evidence for a direct or indirect role of a C. acnes-related dysbiosis in disease development or progression, i.e., reduced C. acnes strain diversity and/or the predominance of a certain phylotype. In the second part of the review, we highlight historical and recent findings demonstrating beneficial aspects of colonization by C. acnes such as colonization resistance, immune system interactions, and oxidant protection, and discuss the molecular mechanisms behind these effects. This new insight led to efforts in skin microbiota manipulation, such as the use of C. acnes strains as probiotic options to treat skin disorders.

Cutibacterium spp. isolated from orthopaedic implant-associated infection: A not-so-slowly growing organism / Cutibacterium spp. aislados de infecciones ortopédicas asociadas a implantes: Un microorganismo de crecimiento no tan lento

Introduction: It has been reported that microbiological diagnosis of Cutibacterium spp. infection requires a prolonged incubation time (up to 14 days). We present our experience with regard to incubation time for detection of Cutibacterium spp. in orthopaedic samples over a 10-year period. Methods: One hundred and nineteen samples were included in this retrospective study. Fifty-three were implants (having previously undergone sonication), 64 were periprosthetic tissue biopsies and two were synovial fluids. Atkins's criteria were used for interpreting the isolates. Quantification and number of days until a culture became positive for Cutibacterium spp. were evaluated. Results: The median number of days to detection of a clinically significant isolate and a contaminant was 4 days. No clinically significant isolates grew after day eight. Conclusion: Most clinically significant isolates of Cutibacterium spp. are detected in the first 7 days of incubation, although a recommendation of prolonged incubation (up to 14 days) appears to be necessary for detecting other organisms.(AU)

Introducción: Se ha reportado que el diagnóstico microbiológico de las infecciones por Cutibacterium spp. requiere un tiempo de incubación prolongado (hasta 14 días). Presentamos nuestra experiencia al respecto en muestras ortopédicas durante un período de 10 años. Métodos: Se incluyeron en este estudio retrospectivo 119 muestras de las que 53 fueron implantes (previa sonicación), 64 biopsias de tejido periprotésico y dos líquidos sinoviales. Para la interpretación se siguieron los criterios de Atkins. Se evaluó la cuantificación y el número de días hasta que el cultivo fue positivo para Cutibacterium spp. Resultados: La mediana del número de días para detectar un aislado clínicamente significativo y un contaminante fue de cuatro días. Ningún aislado clínicamente relevante creció después del día ocho. Conclusión: La mayoría de aislados clínicamente significativos de Cutibacterium spp. se detectan durante los siete primeros días de incubación, sin embargo, parece necesaria una incubación de hasta 14 días para la detección de otros microorganismos.(AU)

Epidemiology and in vitro antimicrobial susceptibility of aerobic Actinomycetales in a clinical setting.

INTRODUCTION: The incidence of infections caused by aerobic actinomycetes is increasing. Recent changes in taxonomy and the variability in susceptibility patterns among species make necessary a proper identification and antibiotic susceptibility testing. MATERIAL AND METHODS: Fifty-three strains of aerobic actinomycetes were identified by MALDI-TOF MS using the VITEK MS Mycobacterium/Nocardia kit (bioMérieux, France) in a tertiary hospital in Spain during a six-year period. Antimicrobial susceptibility testing of the isolates was performed using the Sensititre Rapmycoi microdilution panel (Thermo Fisher Scientific, Massachusetts, USA). RESULTS: Forty strains of Nocardia spp. were identified in the study, being N. farcinica and N. cyriacigeorgica the most prevalent ones. All isolates were susceptible to linezolid and the resistance to amikacin was only observed in one isolate of Gordonia sputi. Resistance to cotrimoxazole was only found in five isolates. CONCLUSIONS: Routine identification and antimicrobial susceptibility testing of aerobic actinomycetes is advisable for an efficient identification of species and effective treatment.