American crocodiles (Crocodylus acutus: Reptilia: Crocodilidae) visiting the facilities of a freshwater aquaculture of the Northern Pacific region, Costa Rica, carry tetracycline-resistant Escherichia coli.

Apex predators are exposed to antimicrobial compounds and resistant microbes, which accumulate at different trophic levels of the related ecosystems. The study aimed to characterize the presence and the antimicrobial resistance patterns of fecal Escherichia coli isolated from cloacal swab samples obtained from wild-living American crocodiles (Crocodylus acutus) (n = 53). Sampling was conducted within the distinctive context of a freshwater-intensive aquaculture farm in Costa Rica, where incoming crocodiles are temporarily held in captivity before release. Phenotypic antimicrobial susceptibility profiles were determined in all isolates, while resistant isolates were subjected to whole-genome sequencing and bioinformatics analyses. In total, 24 samples contained tetracycline-resistant E. coli (45.3%). Isolates carried either tet(A), tet(B), or tet(C) genes. Furthermore, genes conferring resistance to ß-lactams, aminoglycosides, fosfomycin, sulfonamides, phenicol, quinolones, trimethoprim, and colistin were detected in single isolates, with seven of them carrying these genes on plasmids. Genome sequencing further revealed that sequence types, prevalence of antibiotic resistance carriage, and antibiotic resistance profiles differed between the individuals liberated within the next 24 h after their capture in the ponds and those liberated from enclosures after longer abodes. The overall presence of tetracycline-resistant E. coli, coupled with potential interactions with various anthropogenic factors before arriving at the facilities, hinders clear conclusions on the sources of antimicrobial resistance for the studied individuals. These aspects hold significant implications for both the aquaculture farm's biosecurity and the planning of environmental monitoring programs using such specimens. Considering human-crocodile conflicts from the One Health perspective, the occurrence of antimicrobial resistance underscores the importance of systematical surveillance of antibiotic resistance development in American crocodiles.

Carriage of three plasmids in a single human clinical isolate of Clostridioides difficile.

A subset of clinical isolates of Clostridioides difficile contains one or more plasmids and these plasmids can harbor virulence and antimicrobial resistance determinants. Despite their potential importance, C. difficile plasmids remain poorly characterized. Here, we provide the complete genome sequence of a human clinical isolate that carries three high-copy number plasmids from three different plasmid families that are therefore compatible. For two of these, we identify a region capable of sustaining plasmid replication in C. difficile that is also compatible with the plasmid pCD630 that is found in many laboratory strains. Together, our data advance our understanding of C. difficile plasmid biology.

Plasmid carriage and the natural complexity of bacterial populations contributes to plasmid persistence / El transporte de plásmidos y la complejidad natural de las poblaciones bacterianas contribuyen a la persistencia del plásmido

Introduction: Plasmids carry and transport genes that assist their hosts to survive in many environments. Many studies have examined the conditions for plasmid persistence in bacterial populations. A limitation includes that a majority of the mathematical models for examining plasmid persistence only included bacteria from similar colonies. However, most bacterial cells inhabit complex communities where plasmids disseminate between varied bacterial host cells. Thus, there is a gap in knowledge concerning the persistence of plasmids in natural bacterial populations. To address a few of these gaps in knowledge, the present study attempted to examine the effects of plasmid carriage on intrinsic stages of bacterial populations in Bacillus subtilis co-cultures. Material and methods: B. subtilis cells were transformed with CRISPR-hCas-9 plasmid vectors where the natural phases of bacterial growth, biofilm production, and antibiotic resistance were examined in relation to plasmid carriage. These three natural phases were measured in relation to plasmid carriage through in vitro co-culture assays. Results: After calculating the CFU/mL, bacterial growth in the B. subtilis-Carrier with Escherichia coli (B. sub-C-E. coli) and Vibrio harveyi (B. sub-C-VH) co-cultures significantly decreased with a paired-t-test two-tailed P=0. The WT B. subtilis-V.H samples, the B. subtilis Carrier-V.H co-cultures, and the controls each scored a total of 40, 47, and 46 of crystal violet (CV) intensity of biofilm, respectively. Biofilm formation decreased after co-culturing E. coli with the B. subtilis-Carrier, yielding a P<0.001. The antibiotic resistance levels of the co-cultures increased by 3% for the B. sub-C-V.H samples while the B. sub-C-E. coli co-cultures decreased in antibiotic sensitivity by approximately 1.5%. Conclusions: Plasmid carriage contributes to plasmid persistence via altering the natural phases of bacterial populations (AU)

Introducción: Los plásmidos portan y transportan genes que ayudan a sus huéspedes a sobrevivir en muchos entornos. Muchos estudios han examinado las condiciones para la persistencia de plásmidos en poblaciones bacterianas. Una limitación incluye que la mayoría de los modelos matemáticos para examinar la persistencia de plásmidos solo incluyeron bacterias de colonias similares. Sin embargo, la mayoría de las células bacterianas habitan en comunidades complejas donde los plásmidos se diseminan entre diversas células huésped bacterianas. Por lo tanto, existe un vacío en el conocimiento sobre la persistencia de plásmidos en poblaciones bacterianas naturales. Para abordar algunas de estas lagunas en el conocimiento, el presente estudio intentó examinar los efectos del transporte de plásmidos en las etapas intrínsecas de las poblaciones bacterianas en cocultivos de Bacillus subtilis. Material y métodos: Células de B. subtilis se transformaron con vectores plasmídicos CRISPR-hCas-9 donde se examinaron las fases naturales de crecimiento bacteriano, producción de biopelículas y resistencia a los antibióticos en relación con el transporte del plásmido. Estas tres fases naturales se midieron en relación con el transporte de plásmidos a través de ensayos de cocultivo in vitro. Resultados: Después de calcular las UFC/mL, el crecimiento bacteriano en los cocultivos de B. subtilis-Carrier con Escherichia coli (B. sub-C-E. coli) y Vibrio harveyi (B. sub-C-VH) disminuyó significativamente con un -t-test de dos colas P=0. Las muestras WT B. subtilis-V.H, los cocultivos B. subtilis Carrier-V.H y los controles obtuvieron cada uno un total de 40, 47 y 46 de intensidad de biopelícula cristal violeta (CV), respectivamente. La formación de biopelículas disminuyó después de cocultivar E. coli con B. subtilis-Carrier, lo que arrojó un P<0,001 (AU)