Clostridioides difficile is a significant cause of diarrhea in hospitals and the community. This bacterial pathogen is transmitted through the ingestion of endospores, which are challenging to eliminate due to intrinsic resistance to a variety of chemical disinfection agents. The well-characterized laboratory strain CD630 displays low virulence, has not caused outbreaks, and is highly susceptible to disinfectants. Nonetheless, a closely related strain termed NAPCR1 caused outbreaks in Costa Rica and later became endemic in many hospitals from this country. This strain causes disease through unusual mechanisms and is genotypically distinct from CD630. Consequently, its epidemic potential could be influenced by as yet unknown spore phenotypes, such as increased resistance to disinfectants. Objective: To determine whether the NAPCR1 strain is more resistant to a conventional and highly effective C. difficile sporicidal agent than strain CD630 and to identify potential explanatory mechanisms at the genomic level. Methods: We used an in vitro dilution-neutralization method to calculate the sporicidal activity of sodium dichloroisocyanurate (DCC) against purified spores from three subtypes of NAPCR1 isolates (LIBA-2945, LIBA-5761, and LIBA-6276), CD630, and a representative of the highly virulent and epidemic NAP1 strain (LIBA-5758). This phenotypic characterization was complemented with a genomics-steered search of polymorphisms in 15 spore- or sporulation-related genes. Results: Whereas DCC at a final concentration of 0.1 % (w/v) eradicated CD630 endospores with high efficacy (log10 reduction factor (LFR) ≥ 5), it only partially inactivated NAPCR1 (average LFR range: = 1.77-3.37) and NAP1 endospores (average LRF = 3.58). As hypothesized, the three NAPCR1 subtypes tested were more resistant to DCC than strain CD630 (ANOVA, P < 0.05), with LIBA-5761 showing the highest level of DCC resistance overall (ANOVA, P < 0.05). All three NAPCR1 isolates showed large deletions in bclA1. Besides, isolates LIBA-5761 and LIBA-6276 had deletions in bclA2. Conclusions: Our in vitro tests revealed a differential resistance of spores from the C. difficile NAPCR1 strain to DCC. They highlight the importance of continuously evaluating the efficacy of deployed disinfection agents against circulating strains and hint to a potential role of structural proteins from the exosporium in resistance to disinfectants in C. difficile. Introducción: Clostridioides difficile es una causa importante de diarrea a nivel hospitalario y comunitario. Esta bacteria se transmite por medio de la ingestión de endosporas, las cuales son difíciles de erradicar por su resistencia intrínseca a diferentes agentes químicos de desinfección. La cepa de referencia CD630 está bien caracterizada, es poco virulenta, no ha causado brotes, y es altamente susceptible a los desinfectantes. Además, pertenece al mismo clado MLST y es filogenéticamente muy cercana a la cepa NAPCR1. Sin embargo, solo la última ha causado brotes en Costa Rica y se ha convertido en una cepa endémica en varios hospitales locales. La cepa NAPCR1 causa enfermedad por mecanismos poco usuales y es genotípicamente diferente a la cepa CD630. Por lo tanto, su potencial epidémico podría estar influenciado por cambios fenotípicos en sus esporas, como una resistencia incrementada a los desinfectantes. Objetivo: Determinar si la cepa NAPCR1 presenta mayor resistencia que CD630 a un desinfectante de alta eficacia utilizado a nivel hospitalario y dilucidar posibles mecanismos a nivel genómico. Métodos: Se utilizó el método de dilución-neutralización para evaluar la actividad esporicida in vitro del dicloroisocianurato de sodio (DCC) contra esporas de 3 subtipos de la cepa NAPCR1 (LIBA-2945, LIBA-5761, y LIBA-6276), CD630 y un aislamiento representativo de la cepa epidémica e hipervirulenta NAP1 (LIBA-5758). Esta caracterización fenotípica fue complementada con una búsqueda genómica de polimorfismos en 15 genes relacionados con la estructura de la endospora o el proceso de esporulación. Resultados: El DCC a una concentración final de 0.1 % (p/v) erradicó las endosporas de la cepa CD630 con gran eficacia (factor de reducción logarítmica; FRL ≥ 5) y eliminó parcialmente las de las cepas NAPCR1 (FRL promedio = 1.77-3.64) y NAP1 (FRL promedio = 3.58). El perfil de susceptibilidad del aislamiento NAPCR1 LIBA-5761 fue único, ya que mostró un mayor nivel de resistencia hacia el DCC que los otros aislamientos NAPCR1 y la cepa NAP1 examinada (ANOVA, P < 0.05). Los tres aislamientos NAPCR1 mostraron deleciones en bclA1 y los aislamientos LIBA-5761 y LIBA-6276 tenían deleciones adicionales en bclA2. Conclusiones: Nuestros experimentos in vitro confirman la resistencia incrementada a los desinfectantes de la cepa NAPCR1 y una susceptibilidad diferencial en sus tres subtipos. Adicionalmente, señalan la importancia de evaluar continuamente la eficacia de los desinfectantes contra cepas circulantes y asignan un posible papel en la resistencia a los desinfectantes gracias a las proteínas del exosporio de C. difficile.

Carbapenem resistance is considered one of the greatest current threats to public health, particularly in the management of infections in clinical settings. Carbapenem resistance in bacteria is mainly due to mechanisms such as the production of carbapenemases (such as the imipenemase IMP, or other enzymes like VIM, NDM, and KPC), that can be detected by several laboratory tests, including immunochromatography and automated real-time PCR (qPCR). Methods: As part of local studies to monitor carbapenem-resistant bacteria in Costa Rica, two cases were initially identified with inconsistent IMP detection results. A possible gene drop-out in the automated qPCR test was suggested based on the negative result, contrasting with the positive result by immunochromatography and whole-genome sequencing. We hypothesized that molecular testing could be optimized through the development of tailored assays to improve the detection of IMP genes. Thus, using IMP gene sequences from the local isolates and regional sequences in databases, primers were redesigned to extend the detection of IMP alleles of regional relevance. Results: The tailored qPCR was applied to a local collection of 119 carbapenem-resistant isolates. The genomes of all 14 positive cases were sequenced, verifying the results of the custom qPCR, despite the negative results of the automated testing. Conclusions: Guided by whole-genome sequencing, it was possible to extend the molecular detection of IMP alleles circulating in Latin America using a tailored qPCR to overcome IMP gene drop-out and false-negative results in an automated qPCR.

Las enterobacterias son un grupo amplio y heterogéneo de bacilos Gram negativos que se aíslan de forma rutinaria en el laboratorio clínico y se asocian a una gran cantidad de cuadros clínicos. Aquellas resistentes a antibióticos de última línea, como a los carbapenémicos, representan un gran reto en los centros de salud. Ante la dificultad para tratar infecciones causadas por este tipo de bacterias, se ha retomado el uso de antimicrobianos clásicos como la colistina, la nitrofurantoína y la fosfomicina. El objetivo de este trabajo es detallar los principales mecanismos de resistencia para estos tres fármacos descritos en enterobacterias. Para ello, se efectuó una revisión bibliográfica de artículos científicos publicados entre los años 1999 y 2022, utilizando las bases de datos PubMed (NCBI), PLOS, Redalyc, Google Scholar y Science Direct. En este proceso, se usaron las palabras clave “Carbapenem-Resistant Enterobacteriaceae”, “colistin”, nitrofurantoin”, “fosfomycin”, “resistance” y “plasmids”. Se encontró que los mecanismos de resistencia son variados y abarcan fenómenos como modificación del sitio blanco, inactivación enzimática, impermeabilidad y eflujo. Además, los determinantes genéticos de resistencia se encuentran en cromosomas o en plásmidos. Conocer este tipo de información permite mejorar la vigilancia basada en el laboratorio, combatir el problema de resistencia a los antimicrobianos y optimizar el uso de estos antibióticos que forman parte del escaso arsenal para el tratamiento de ciertas infecciones causadas por microorganismos multidrogorresistentes.

Se presenta el caso de un paciente masculino de 70 años, hipertenso y extabaquista que fue hospitalizado por disnea, malestar general, fiebre y dolor abdominal. Tras el abordaje inicial con nifuroxazida y cefotaxime no presentó mejoría y, al realizar más estudios, fue diagnosticado con neumonía por L. pneumophila. Tras el diagnóstico y recibir terapia con levofloxacina, mostró mejoría y se recuperó de los síntomas iniciales, por lo que fue dado de alta. Se discuten las dificultades diagnósticas, de abordaje y la situación nacional para la detección de L. pneumophila en Costa Rica.

Background: Carbapenem-resistant Acinetobacter baumannii (CRAB) is a critical pathogen associated with severe hospital infections and high antimicrobial resistance. Despite the global significance of A. baumannii, there are limited data from Costa Rica regarding the resistance rate and genomic characteristics of CRAB. Methods: This study aimed to provide initial and exploratory epidemiological data on infections caused by A. baumannii and CRAB isolated in Costa Rica and to gain insights on the genome of selected strains, focusing on their resistance determinants and phylogenetic relationships. Results: Based on data from five main hospitals in Costa Rica, resistance rate to carbapenems was estimated at 9.8% to imipenem and 6.1% to meropenem. From 190 carbapenem-resistant clinical isolates available in a local collection, seven A. baumannii strains were identified, all showing resistance to carbapenems and carrying the bla_NDM-1 gene. Whole-genome sequencing of two strains yielded two distinct MLST profiles (Pasteur scheme: ST-150 for strain IPAT15 and ST-250 for IPAT72), as well as variations in the number and identity of plasmids, genomic islands, and other elements of the mobilome. Both isolates carried ten antimicrobial resistance genes, which are predicted to be harbored in plasmids for IPAT15, unlike the chromosomal determinants in IPAT72. A pangenome analysis of 878 genomes from a public database identified over 51,000 genes, with only 1338 (2.6%) forming the core genome. Phylogenetic analysis and assignation of international clones (ICs) showed predominance of IC2. Isolates from Costa Rica clustered near IC9 and shared some resistance determinants, but they were not directly assigned to an IC. Conclusions: Overall, this study provides exploratory insights regarding the occurrence of CRAB in Costa Rica using epidemiological and genomic data, with profiles that are comparable to other regions in Latin America and diverse genomic resistance determinants. While this study does not show the whole landscape of CRAB in Costa Rica, these data constitute an initial approach for improving clinical management and public health responses to CRAB infections, to ultimately improve outcomes for patients affected by this pathogen.