engleski

Isolates of Acinetobacter pittii from municipal wastewater of the City of Zagreb, Croatia

Acinetobacter pittii is genetically closely related to A. baumannii, A. calcoaceticus, A. nosocomialis, genomic species “close to 13TU”, and genomic species “between 1 and 3”. Therefore, these six species of Acinetobacter spp. are grouped into A. calcoaceticus- baumannii complex. Human diseases have been attributed mainly to A. baumannii, and to a lesser extent to A. pittii. The latter is sensitive to antibiotics unlike A. baumannii which is usually multi-drug resistant. However, recent molecular studies revealed that A. pittii is often misidentified by routine laboratory methods. The prevalence of A. pittii in nature and its epidemiology are undefined to date. The objective of this study was to screen the municipal wastewater of Zagreb, the capital city of Croatia, for the presence of A. pittii. The sampling was done at the influent of the largest Croatian wastewater treatment plant of the City of Zagreb, where the municipal wastewater consisting of domestic, industrial, hospital and storm wastewaters are treated. In the composite 24h samples the physico-chemical and microbiological parameters were determined. The isolation of A. pittii was done on CHROMagar Acinetobacter supplemented with 15mg/L of cefsulodin after incubation at 42C/48h. The presumptive A. pittii colonies were firstly characterized to possess the following characteristics: Gram negative coccobacilli, negative oxidase, positive catalase reaction, no reaction on the Kligler Iron Agar. Further identification was carried out by using Vitek 2 system, and confirmed by MALDI-TOF MS and RNA polymerase β-subunit (rpoB) gene sequencing. Four isolates of A. pittii were obtained from municipal wastewater with high MALDI-TOF MS score values (2.156- 2.291). Isolates were sensitive to all 9 antibiotics tested by Vitek 2 system and E- test. One isolate showed MIC value of meropenem close to intermediate values (1.5mg/L) and twice as high MICs values of levofloxacin and colistin (0.5mg/L) than other three isolates. MALDI-TOF MS and PFGE dendrograms grouped three isolates with lower MICs in cluster different form one isolate with higher MICs values. Phylogenetic analysis has shown the close relatedness of the isolate with higher MICs values to the A. pittii strain CIP 70.15, with 100% sequence ID of the rpoB gene fragment. The isolate of A. pittii with higher MICs values was probably of clinical origin and entered the municipal wastewater trough hospital sewage. The finding of isolates with low MICs values suggests that A. pittii could also be an autochthonous species in water.

environment ; public health ; wastewater treatment plant

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