- Volume 67, Issue 9, 2018

Purpose. This study aimed to characterize A. baumannii strains isolated from patients in an intensive care unit (ICU) setting. Molecular techniques were used to study clonal relatedness and determine a fast, efficient and cost-effective way of detecting persistent clones.

Methodology. A. baumannii (n=17) were obtained in June and November 2015 from a single ICU setting in South India. DNA typing methods such as multilocus sequence typing (MLST), single-locus sequence-based typing (SBT) and DNA fingerprinting PCRs (M13, DAF4 and ERIC2) were employed to understand the association of clones. PCRs were performed for the antimicrobial resistance genes ISAba1-bla _OXA-51-like, ISAba1-bla _OXA-23-like, bla _NDM-1, bla _PER-7 and bla _TEM-1, and the virulence genes cpa 1, cpa2 and pkf.

Results. The MLST showed some degree of corroboration with the other DNA typing methods. The M13 PCR was found to give better results than the other fingerprinting methods. ST848 (CC92) was the dominant strain isolated in both June and November. All isolates were bla _OXA-51-like-positive, with 16 having ISAba1 upstream of the bla _OXA-51-like and bla _OXA-23-like genes. Genes such as bla _NDM-1 (23 %, n=4), bla _PER-7 (58.8 %, n=10), pkf (82 %, n=14), bla _TEM-1 (5.8 %, n=1), cpa1 (5.8 %, n=1) and cpa2 (5.8 %, n=1) were also detected.

Conclusion. M13 PCR can be used in routine environmental surveillance for the detection of persistent antibiotic resistant clones in an ICU setting because of its reliability and simplicity. Further studies based on greater sample size, conducted at the multi-centre level, can give us a better understanding of the reliability of the molecular methods that can be used for the detection of persistent clones in the hospital setting.

Purpose. From May 2015 to March 2016, an outbreak due to Listeria monocytogenes serotype 1/2a and clinical pulsotype never previously isolated in Europe occurred in central Italy, involving 24 confirmed clinical cases. The article provides a description of the outbreak and the investigation carried out by a multidisciplinary network.

Methodology. Epidemiological and microbiological surveillance was conducted to confirm the outbreak and to detect the food vehicle of infection. The origin and destination of the implicated food and its ingredients were investigated by tracing-back and -forward investigation.

Results. Next-generation sequencing confirmed the unique outbreak strain. On 4 January 2016, a L. monocytogenes strain with pulsotype indistinguishable from that isolated from clinical cases in the outbreak was detected in a sample of hog head cheese purchased from a retail supermarket by one of the patients. The hog head cheese was produced by a small meat processing plant in the Marche region, where microbiological investigation confirmed environmental and food contamination by the outbreak strain. Plant production was suspended and all contaminated batches of the hog head cheese were withdrawn from the market by 19 February by local health authority. We subsequently observed a sharp decline in clinical cases, the last being reported on 11 March 2016.

Conclusion. The key factor in the timely conclusion of this investigation was intersectoral collaboration among epidemiologists, microbiologists, veterinarians, statisticians and health and food safety authorities at national, regional and local levels.

Purpose. In this study, we aimed to investigate the genomic characteristics and evolution of pathogenicity islands of an enteropathogenic Escherichia coli (EPEC) strain, and to obtain a transcriptional profile of EPEC under different concentrations of ciprofloxacin using microarray analysis.

Methodology. The complete EPEC Deng genome was sequenced and compared to genomes of 12 previously sequenced E. coli strains. A 180 min time course experiment was performed in which the effect of ciprofloxacin on EPEC Deng growth was evaluated. Microarray profiling was used to study the effect of varying ciprofloxacin pressure on genome-wide transcriptional expression. Differential expression of the genes identified using microarray data was confirmed using real-time quantitative reverse transcriptase PCR (RTQ). Target gene-defective recombineering strains were created to investigate the influence of the grlA gene on ciprofloxacin susceptibility.

Results. Genomic comparisons revealed a close phylogenic relationship between EPEC Deng and E. coli strains O111_H_11128 and O26_H11_11368, with low genetic diversity among their type III secretion system genes and typically genetic variation in the map, tir, eae and espA genes of EPEC. It is noteworthy that 21 genes were down-regulated at all time points examined in the group exposed to 2 µg ml−1 of ciprofloxacin. A grlA-mutant derivative with increased susceptibility to ciprofloxacin was discovered.

Conclusions. The present findings provide an overview of the phylogenetic characteristics of EPEC Deng and its transcriptional response to ciprofloxacin, further suggesting that GrlA may play a clinically important role in EPEC responses to ciprofloxacin.

Purpose. Pasteurella multocida (P. multocida) is a principal pathogen of domestic animals and an opportunistic pathogen of humans. It is the causative agent of pneumonia and haemorrhagic septicaemia in cattle, sheep and goats, fowl cholera in chickens and progressive atrophic rhinitis in swine. In this study, we investigated the humoral and cellular immune responses and protective immunity conferred by an iron-inactivated vaccine with bacterial DNA (IIV+bDNA) as an adjuvant in mice.

Methodology. P. multocida was grown in BHI broth, inactivated with formalin and FeCl₃ and adjuvanted with alum and bDNA. Mice were immunized with two whole-cell inactivated vaccine doses 2 weeks apart. The animals were challenged 4 weeks after booster immunization. Immunogens (vaccines and bDNA) posed no safety problems when mice were injected subcutaneously (s/c) with these preparations. The serum antibody titres were tested by ELISA. At 28 days post immunization, cell-mediated immunity responses were determined. The responses were measured by assay of IL-6 and IL-12 in lymphocyte spleen culture supernatants.

Results. ELISA results showed that the levels of antibodies in iron inactivated with bDNA adjuvant groups were higher than in the formalin inactivated with alum adjuvant vaccine group. The protection rate of IIV+bDNA adjuvant vaccine was superior to that of the other vaccines and it protected 100 % of the challenge group mice. Following immunization, bDNA promoted increased production of interleukins compared to the control groups.

Conclusion. These studies indicate that bDNA is effective as an immune adjuvant, and along with stimulatory bDNA represent promising new humoral and cellular immune enhancers for vaccination applications. In addition, this vaccine is able to provide long-term protection against infection.