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The melioidosis bacterium, Burkholderia pseudomallei, is increasingly being recognised as a pathogen in patients with cystic fibrosis (CF). We have recently catalogued genome-wide variation of paired, isogenic B. pseudomallei isolates from seven Australasian CF cases, which were collected between 4 and 55 months apart. Here, we extend this investigation by documenting the transcriptomic changes in B. pseudomallei in five cases. Following growth in an artificial CF sputum medium, four of the five paired isolates exhibited significant differential gene expression (DE) that affected between 32 and 792 genes. The greatest number of DE events was observed between the strains from patient CF9, consistent with the hypermutator status of the latter strain, which is deficient in the DNA mismatch repair protein MutS. Two patient isolates harboured duplications that concomitantly increased expression of the β-lactamase-encoding gene penA, and a 35 kb deletion in another abolished expression of 29 genes. Convergent expression profiles in the chronically-adapted isolates identified two significantly downregulated and 17 significantly upregulated loci, including the resistance-nodulation-division (RND) efflux pump BpeEF–OprC, the quorum-sensing hhqABCDE operon, and a cyanide- and pyocyanin-insensitive cytochrome bd quinol oxidase. These convergent pathoadaptations lead to increased expression of pathways that may suppress competing bacterial and fungal pathogens, and that enhance survival in oxygen-restricted environments, the latter of which may render conventional antibiotics less effective in vivo. Treating chronically adapted B. pseudomallei infections with antibiotics designed to target anaerobic infections, such as the nitroimidazole class of antibiotics, may significantly improve pathogen eradication attempts by exploiting this Achilles heel.

Purpose. Antifungal resistance and several putative virulence factors have been associated with the pathogenicity of the Candida parapsilosis species complex. The objective of this study was to evaluate the antifungal susceptibility, the production of virulence factors and the pathogenicity of the C. parapsilosis complex.

Methodology. Overall, 49 isolates of C. parapsilosis sensu stricto, 19 C. orthopsilosis and nine C. metapsilosis were used. The planktonic and biofilm susceptibility to fluconazole, itraconazole, voriconazole, amphotericin B and caspofungin was assessed using a broth microdilution assay. Finally, the production of biofilm and hydrolytic enzymes and the fungal pathogenicity against Caenorhabditis elegans were investigated.

Results/Key findings. Overall, one C. orthopsilosis was resistant to caspofungin and susceptible-dose-dependent to itraconazole, the other two C. orthopsilosis were susceptible-dose-dependent to fluconazole and itraconazole, and one C. metapsilosis was susceptible-dose-dependent to azoles. A total of 67.5 % of the isolates were biofilm producers. Amphotericin B and caspofungin caused the greatest reduction in the metabolic activity and biomass of mature biofilms. Phospholipase and protease production was observed in 55.1 % of C. parapsilosis sensu stricto, 42.1 % of C. orthopsilosis and 33.3 % of C. metapsilosis isolates. Moreover, 57.9 % of C. orthopsilosis and 20.4 % of C. parapsilosis sensu stricto isolates were β-haemolytic, and all C. metapsilosis were α-haemolytic. Finally, the C. parapsilosis complex caused high mortality of C. elegans after 96 h of exposure.

Conclusion. These results reinforce the heterogeneity of these cryptic species for their antifungal susceptibility, virulence and pathogenic potential, emphasizing the relevance of monitoring these emerging pathogens.

Fluoroquinolone (FQ)-resistant Salmonella spp. were listed by the WHO in 2017 as priority pathogens for which new antibiotics were urgently needed. The overall global burden of Salmonella infections is high, but differs per region. Whereas typhoid fever is most prevalent in South and South-East Asia, non-typhoidal salmonellosis is prevalent across the globe and associated with a mild gastroenteritis. By contrast, invasive non-typhoidal Salmonella cause bloodstream infections associated with high mortality, particularly in sub-Saharan Africa. Most Salmonella strains from clinical sources are resistant to first-line antibiotics, with FQs now being the antibiotic of choice for treatment of invasive Salmonella infections. However, FQ resistance is increasingly being reported in Salmonella, and multiple molecular mechanisms are already described. Whole-genome sequencing (WGS) is becoming more frequently used to analyse bacterial genomes for antibiotic-resistance markers, and to understand the phylogeny of bacteria in relation to their antibiotic-resistance profiles. This mini-review provides an overview of FQ resistance in Salmonella, guided by WGS studies that demonstrate that WGS is a valuable tool for global surveillance.

This study characterizes KPC-2 producing Klebsiella pneumoniae belonging to ST101. Whole genome sequencing using the Ion Torrent PGM platform with 400 bp chemistry was performed. bla KPC-2 was found on an IncFIIK plasmid associated with ISKpn6 and ISKpn7 without Tn4401. This is the first report of KPC-2 K. pneumoniae from bacteremia in India. The isolate also coded for other resistance genes such as aadA1, aadA2, armA, aac(3)-Ild, aac(6′)-Ild for aminoglycoside; bla SHV-11 , bla TEM-1B , bla OXA-9, for β-lactams and aac(6′)-Ild, oqxA, oqxB, qnrB1 for fluoroquinolones. It belonged to the K17 capsular type. India is endemic to New Delhi metallo-β-lactamase and OXA48-like carbapenemases and K. pneumoniae carbapenemase (KPC) is seldom reported. With high rates of carbapenem resistance, emergence of KPC in India will challenge patient management. The isolate was susceptible to colistin. The patient had a fatal outcome.

The role of penetration limitation in Candida biofilm-associated antifungal resistance remains unclear. Most of the previous work has been done on Candida albicans, although non-albicans (NAC) species are also implicated in invasive candidiasis and the biofilm matrix has been shown to vary amongst different species. Only a few studies have evaluated clinical isolates. This study aimed to determine the relevance of penetration limitation in the antifungal resistance of biofilms formed by C. albicans and NAC clinical isolates, using an agar disk diffusion assay. The penetration of posaconazole and amphotericin B through the biofilms was significantly reduced. Fluconazole, voriconazole and caspofungin showed a superior penetration capacity in C. albicans, Candida tropicalis and Candida parapsilosis biofilms, but exhibited inter-species and strain/isolate variation. Candida krusei biofilms were the most resilient to antifungal permeation. All of the antifungal drugs failed to kill the biofilm cells, independent of penetration, suggesting that the other factors contribute markedly to the recalcitrance of the biofilms.

Bacteraemia caused by Escherichia coli is a growing problem with a significant mortality. The factors that influence the acquisition and outcome of these infections are not clear. Here, we have linked detailed genetic data from the whole-genome sequencing of 162 bacteraemic isolates collected in Scotland, UK, in 2013–2015, with clinical data in order to delineate bacterial and host factors that influence the acquisition in hospital or the community, outcome and antibiotic resistance. We identified four major sequence types (STs) in these isolates: ST131, ST69, ST73 and ST95. Nearly 50 % of the bacteraemic isolates had a urinary origin. ST69 was genetically distinct from the other STs, with significantly less sharing of accessory genes and with a distinct plasmid population. Virulence genes were widespread and diversely distributed between the dominant STs. ST131 was significantly associated with hospital-associated infections (HAIs), and ST69 with those from the community. However, there was no association of ST with outcome, although patients with HAI had a higher immediate mortality compared to those with community-associated infections (CAIs). Genome-wide association studies revealed genes involved in antibiotic persistence as significantly associated with HAIs and those encoding elements of a type VI secretion system with CAIs. Antibiotic resistance was common, and there were networks of correlated resistance genes and phenotypic antibiotic resistance. This study has revealed the complex interactions between the genotype of E. coli and its ability to cause bacteraemia, and some of the determinants influencing hospital or community acquisition. In part, these are shaped by antibiotic usage, but strain-specific factors are also important.

Purpose. To describe the antimicrobial resistance profile of Neisseria meningitidis isolates causing invasive disease in Brazil from 2009 to 2016.

Methodology. Among 3548 N. meningitidis isolates received, 2888 (81.4 %) were analysed for antimicrobial resistance using the broth microdilution technique, as recommended by the Clinical and Laboratory Standards Institute. Isolates were tested for ciprofloxacin, chloramphenicol, ceftriaxone, penicillin G, ampicillin and rifampin.

Results. All the isolates tested were susceptible to ceftriaxone, while 953 (33.0 %), 1307 (45.3 %) and 2 (0.07 %) isolates were penicillin G-, ampicillin- and rifampin-intermediate, respectively. Resistance to rifampin, ciprofloxacin and chloramphenicol was shown by three isolates (0.1 %), two isolates (0.07 %) and one (0.03 %) isolate, respectively. Although no isolates were resistant to penicillin G in the period of 2009–2016, our results show an upward trend in minimum inhibitory concentrations (MICs) for this drug as of 2010 (P<0.001). There was no significant difference between different gender and age groups of patients for reduced susceptibility to penicillin G. There was a higher frequency of isolates with reduced susceptibility to penicillin G in the South and Southeast regions (P<0.001). This reduced susceptibility was also associated with serotype 19 inside serogroup B (P<0.001).

Conclusion. Despite the decrease in susceptibility to penicillin G and ampicillin observed from 2010, the overall resistance of N. meningitidis isolates to the antimicrobials tested remained uncommon and sporadic, confirming their efficacy for chemoprophylaxis or treatment of invasive meningococcal disease (IMD) in Brazil. Continued surveillance of N. meningitidis antimicrobial susceptibility profiles is important in order to monitor variations in resistance either geographically, over time or in association with emergent clones.

The aim of this study was to characterize the presence of carbapenemase-encoding genes in distinct species of Acinetobacter spp. isolated from Brazilian hospitals. Five carbapenem-resistant Acinetobacter spp. isolates (two Acinetobacter pittii, two Acinetobacter bereziniae and one Acinetobacter junii) recovered from two distinct hospitals between 2000 and 2016 were included in this study. All of the isolates harboured bla IMP-1, which was inserted into In86, a class 1 integron. Pulsed field gel eletrophoresis analysis showed that both A. pittii were identical, while the two A. berezinae isolates were considered to be clonally related. In this study, we demonstrated that mobile elements carrying carbapenemase-encoding genes such as In86 may persist for a long period, allowing their mobilization from A. baumannii to other Acinetobacter spp. that are usually susceptible to multiple antimicrobials.

Although tuberculosis treatment is dependent on drug-susceptibility testing (DST) and molecular drug-resistance detection, treatment failure and relapse remain a challenge. This could be partially due to the emergence of antibiotic-tolerant dormant mycobacteria, where host lipids have been shown to play an important role. This study evaluated the susceptibility of Mycobacterium tuberculosis to two antibiotic combinations – rifampicin, moxifloxacin, amikacin and metronidazole (RIF-MXF-AMK-MTZ), and rifampicin, moxifloxacin, amikacin and pretomanid (RIF-MXF-AMK-PA) – in a lipid-rich dormancy model. Although their effectiveness in in vitro cultures with dextrose as a carbon source has been proved, we observed that none of the antibiotic mixtures were bactericidal in the presence of lipids. The presence of lipids may confer tolerance to M. tuberculosis against the mixture of antibiotics tested and such tolerance could be even higher during the dormant stages. The implementation of lipids in DST on clinical isolates could potentially lead to a better treatment strategy.

Acinetobacter baumannii is a common causative agent of hospital-acquired infections and a leading cause of infection in burns patients. Carbapenem-resistant A. baumannii is considered a major public-health threat and has been identified by the World Health Organization as the top priority organism requiring new antimicrobials. The most common mechanism for carbapenem resistance in A. baumannii is via horizontal acquisition of carbapenemase genes. In this study, we sampled 20 A. baumannii isolates from a patient with extensive burns, and characterized the evolution of carbapenem resistance over a 45 day period via Illumina and Oxford Nanopore sequencing. All isolates were multidrug resistant, carrying two genomic islands that harboured several antibiotic-resistance genes. Most isolates were genetically identical and represented a single founder genotype. We identified three novel non-synonymous substitutions associated with meropenem resistance: F136L and G288S in AdeB (part of the AdeABC efflux pump) associated with an increase in meropenem MIC to ≥8 µg ml−1; and A515V in FtsI (PBP3, a penicillin-binding protein) associated with a further increase in MIC to 32 µg ml−1. Structural modelling of AdeB and FtsI showed that these mutations affected their drug-binding sites and revealed mechanisms for meropenem resistance. Notably, one of the adeB mutations arose prior to meropenem therapy but following ciprofloxacin therapy, suggesting exposure to one drug whose resistance is mediated by the efflux pump can induce collateral resistance to other drugs to which the bacterium has not yet been exposed.