Prevalence of Type III Secretion System (T3SS) and Biofilm Development in Genetically Heterogeneous Clinical Isolates of Pseudomonas aeruginosa from Nigeria.

Pseudomonas aeruginosa infection in seriously ill patients is a major concern due to its ability to form biofilm and secrete effector toxins. There is little information on the prevalence of T3SS effector toxins and biofilm production in clinical isolates of P. aeruginosa from Nigeria. The goal of this study is to evaluate the prevalence of T3SS toxins and biofilm production among isolates from selected tertiary hospitals in Nigeria. This study examined 430 clinical isolates from our previous work, comprising 181 MDR (multidrug-resistant) and 249 non-MDR isolates. Biofilm production and type III secretion toxins were determined using colorimetric microtiter plate assay and polymerase chain reaction, respectively. Carbapenem-resistant isolates were typed using REP-PCR and BOX-PCR. Biofilm production was detected in 386/430 (89.8%) of the isolates. Out of 386 biofilm producers, 167 (43.3%) were multidrug-resistant isolates. PCR identified four T3SS virulence types among 430 isolates, including 78 (18.1%) exoU+/exoS- isolates, 343 (79.8%) exoU-/exoS + isolates, 5 (1.2%) exoU+/exoS + isolates, and 4 (0.9%) exoU-/exoS- isolates. Both REP- and BOX-PCR consist of eight clusters. On the REP-PCR dendrogram, ExoU+/ExoS- isolates majorly occupied cluster IV. Clusters IV, VII, and VIII consist of isolates from wounds on BOX-PCR dendrogram. There was a positive association between strong biofilm production and multidrug resistance in our P. aeruginosa isolates. This study identified multidrug-resistant, biofilm-producing P. aeruginosa strains that secrete cytotoxic effectors which are significant virulence factors in P. aeruginosa. This poses a severe risk to our healthcare system and highlights the importance of continuous surveillance to prevent infectious disease outbreaks.

Incidence and Molecular Characterization of Carbapenemase Genes in Association with Multidrug-Resistant Clinical Isolates of Pseudomonas aeruginosa from Tertiary Healthcare Facilities in Southwest Nigeria.

Pseudomonas aeruginosa, resistant to multiple antibacterial agents including carbapenems, is of great global public health concern. There is limited data available regarding incidence of Metallo-Beta Lactamase producing P. aeruginosa, their molecular basis of resistance in particular carbapenem resistance and any genetic relatedness among circulating clinical isolates in Southwest Nigeria. Four hundred and thirty P. aeruginosa isolates were collected from seven tertiary care hospitals (predominantly from wound, ear, and urinary tract infections) and verified by PCR targeting oprI and oprL. Antibiotic susceptibility using 16 selected antibiotics and MBL screening was performed. The integrons (class 1, 2 and 3) and carbapenemase genes- blaGES, blaNMC-A, blaBIC-1, blaSME, blaIMP, blaVIM, blaSPM, blaNDM, blaAIM, blaDIM, blaSIM, blaGIM, blaOXA-48, blaOXA-58 were detected by PCR and were sequenced. Quantitative real-time polymerase chain reaction was used to quantify expression levels of eight efflux pump genes, ampC cephalosporinase and outer membrane porin, oprD. The isolates were genotyped using Enterobacterial Repetitive Intergenic Consensus sequence Polymerase Chain Reaction (ERIC-PCR). Four hundred and thirty P. aeruginosa isolates were subjected to antibiotic susceptibility testing, revealing that 109 (25.4%) isolates were multidrug-resistant, 47 (10.9%) were extensively drug-resistant and 25 (5.8%) were pandrug-resistant. MBL was seen in 17.0% (73/430) isolates. MBL-encoding genes; blaVIM-5 and blaNDM-1 were detected in 86.3% (63/73) isolates, with blaVIM-5 and blaNDM-1 in 35.6% (26/73) and 38.4% (28/73), respectively, whereas co-occurrence of blaVIM-5 and blaNDM-1 was found in 12.3% (9/73). Forty-one (56.2%) carbapenem-resistant P. aeruginosa strains carried class 1 integrons, while co-occurrence of class 1 and 2 integrons was seen in 12.3%. qPCR results indicated that MexXY-OprM was highly expressed pump in 58.9%, ampC upregulated in 26.0%, while oprD porin was downregulated in 65.8% isolates. ERIC-PCR results suggest that carbapenem-resistant strains exhibit genetic heterogeneity. The high incidence of MBL-encoding genes and integrons in diversified clinical P. aeruginosa from southwestern Nigeria is of great concern. The co-occurrence of blaVIM-5 and blaNDM-1 as well as resistance in general manifesting a gradient based on genotypic variation suggests that there is a strong need for efficient surveillance programs and antibiotic stewardship.