Carbapenem-resistant Acinetobacter baumannii (CRAB): metabolic adaptation and transcriptional response to human urine (HU)

Abstract

Carbapenem-resistant Acinetobacter baumannii (CRAB) is a major human pathogen and a research priority for developing new antimicrobial agents. CRAB is a causative agent of a variety of infections in different body sites. One of the manifestations is catheter-associated urinary tract infection, which exposes the bacteria to the host's urine, creating a particular environment. Exposure of two CRAB clinical isolates, AB5075 and AMA40, to human urine (HU) resulted in the differential expression levels of 264 and 455 genes, respectively, of which 112 were common to both strains. Genes within this group play roles in metabolic pathways such as phenylacetic acid (PAA) catabolism, the Hut system, the tricarboxylic acid (TCA) cycle, and other processes like quorum sensing and biofilm formation. These results indicate that the presence of HU induces numerous adaptive changes in gene expression of the infecting bacteria. These modifications presumably help bacteria establish and thrive in the hostile conditions in the urinary tract. These analyses advance our understanding of CRAB's metabolic adaptations to human fluids, as well as expanding knowledge on bacterial responses to distinct human fluids containing different concentrations of human serum albumin (HSA).

Keywords: Acinetobacter baumannii; carbapenem-resistance; cefiderocol; human serum albumin; human urine.

Figure 1

Enriched Gene Ontology terms of the most representative differentially expressed genes. Gene ontology enrichment analysis of the DEGs of strains AB5075 (A) or AMA40 (B) incubated in the presence or in the absence of human urine. Data were obtaining using the Fisher’s test (FDR < 0.05) with the Blast2GO software. C) A. baumannii TCA-glyoxylate pathways in response to HU. Reactions and intermediates of the TCA and glyoxylate cycle are represented and based on BioCyc and MetaCyc. Green and red represents up and down-regulation, respectively. *, Indicates statically significant differential expressed genes (DEGs).

Figure 2

PAA degradation pathway is induced in CRAB by exposure to HU and affects neutrophile chemotaxis. Differential gene expression of genes in PAA and Phe pathways of A. baumannii AB5075 when exposed to HU in transcriptomic analyses (A) and by qRT-PCR (B). Differential gene expression of genes in PAA and Phe pathways of A. baumannii AMA40 when exposed to HU in transcriptomic analyses (C) and by qRT-PCR (D). Neutrophile chemotaxis assays (E) indicate a reduced neutrophile recruitment when both CRAB strains are exposed to HU. Statistical significance (P < 0.05) was determined by two-way ANOVA followed by Tukey’s multiple comparison test. Significance was indicated as follows: ***, P <0.001; ** P <0.01, and * P <0.05.

Figure 3

Differential expression of the genes involved on histidine catabolism in CRAB strains exposed to HU. (A) Heatmap outlining the differential expression of genes associated with histidine catabolism for AB5075 and AMA40 strains. (B) qRT-PCR results of the Hut system genes in AB5075 (A) and AMA40 (B). Statistical significance (P < 0.05) was determined by two-way ANOVA followed by Tukey’s multiple comparison test. Significance was indicated as follows: *** indicates p< 0.001; ** p<0.01, and * p<0.05.

Figure 4

Impact of HU on the expression of CRAB genes coding for K+ transport and benzoate and cysteine pathway functions. Benzoate pathway genes are upregulated in both strains, AB5075 and AMA 40, in transcriptomics studies (A) and corroborated by qRT-PCR (B). Expression of genes involved in the high-affinity potassium transporter are downregulated in both strains of A. baumannii studied (C and D). Cysteine metabolic pathway genes are also downregulated in the presence of HU for both strains AB5075 and AMA40 shown by transcriptomic analyses (E) and by qPCR (F). Statistical significance (P < 0.05) was determined by two-way ANOVA followed by Tukey’s multiple comparison test. Significance was indicated as follows: *** indicates P <0.001; ** P <0.01, and * P <0.05.