Publications by authors named "Seydina M Diene"

73 Publications

Historical, current, and emerging tools for identification and serotyping of Shigella.

Braz J Microbiol 2021 Sep 15. Epub 2021 Sep 15.

Laboratoire Microbiologie Santé et Environnement (LMSE), Doctoral School of Sciences and Technology, Faculty of Public Health, Lebanese University, Tripoli, Lebanon.

The Shigella genus includes serious foodborne disease etiologic agents, with 4 species and 54 serotypes. Identification at species and serotype levels is a crucial task in microbiological laboratories. Nevertheless, the genetic similarity between Shigella spp. and Escherichia coli challenges the correct identification and serotyping of Shigella spp., with subsequent negative repercussions on surveillance, epidemiological investigations, and selection of appropriate treatments. For this purpose, multiple techniques have been developed historically ranging from phenotype-based methods and single or multilocus molecular techniques to whole-genome sequencing (WGS). To facilitate the selection of the most relevant method, we herein provide a global overview of historical and emerging identification and serotyping techniques with a particular focus on the WGS-based approaches. This review highlights the excellent discriminatory power of WGS to more accurately elucidate the epidemiology of Shigella spp., disclose novel promising genomic targets for surveillance methods, and validate previous well-established methods.
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http://dx.doi.org/10.1007/s42770-021-00573-5DOI Listing
September 2021

First Genome Description of Isolate Bearing NDM-1 from Blood Culture.

Microorganisms 2021 Aug 17;9(8). Epub 2021 Aug 17.

Faculté de Pharmacie, Aix Marseille Université, IRD, APHM, MEPHI, IHU Méditerranée Infection, 19-21 Boulevard Jean Moulin, CEDEX 05, 13385 Marseille, France.

In this paper, we describe the first complete genome sequence of species, a clinical multidrug-resistant strain harboring the New Delhi Metallo-β-lactamase-1 (NDM-1) gene, isolated at the Kinshasa University Teaching Hospital, in Democratic Republic of the Congo. Whole genome sequencing of an imipenem-resistant clinical Gram-negative P8538 isolate was performed using MiSeq and Gridion, and then complete genome analysis, plasmid search, resistome analysis, and comparative genomics were performed. Genome assembly resulted in a circular chromosome sequence of 4,280,811-bp and 40.80% GC and a circular plasmid (pPV8538_NDM-1) of 151,684-bp and 51.93%GC, which was identified in an P8540 strain isolated in the same hospital. Interestingly, comparative genomic analysis revealed multiple sequences acquisition within the P8538 chromosome, including three complete prophages, a siderophore biosynthesis NRPS cluster, a Type VI secretion system (T6SS), a urease gene cluster, and a complete Type-I-F CRISPR-Cas3 system. Β-lactamase genes, including and , were found on the recombinant plasmid pPV8538_NDM-1, in addition to other antibiotic resistance genes such as , -, , , , --, , and . Genome comparison with species revealed 82.95% of average nucleotide identity (ANI), with species exhibiting 90.79% of proteome similarity. We report the first complete genome of species and for the first time the presence of the gene in this species. This work highlights the need to improve surveillance and clinical practices in DR Congo in order to reduce or prevent the spread of such resistance.
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http://dx.doi.org/10.3390/microorganisms9081751DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8398168PMC
August 2021

Extensive Comparative Genomic Analysis of and Reveals a Direct Association between the Absence of CRISPR-Cas Systems, the Presence of Anti-Endonuclease (ardA) and the Acquisition of Vancomycin Resistance in .

Microorganisms 2021 05 21;9(6). Epub 2021 May 21.

Aix Marseille University, IRD, APHM, MEPHI, IHU-Mediterranee Infection, 13005 Marseille, France.

Here, we performed a comparative genomic analysis of all available genomes of ( = 1591) and ( = 1981) and investigated the association between the presence or absence of CRISPR-Cas systems, endonuclease/anti-endonuclease systems and the acquisition of antimicrobial resistance, especially vancomycin resistance genes. Most of the analysed Enterococci were isolated from humans and less than 14% of them were from foods and animals. We analysed and detected CRISPR-Cas systems in 75.36% of genomes and only 4.89% of genomes with a significant difference (-value < 10). We found a negative correlation between the number of CRISPR-Cas systems and genome size (r = -0.397, -value < 10) and a positive correlation between the genome %GC content and the number of CRISPR-Cas systems (r = 0.215, -value < 10). Our findings showed that the presence of the anti-endonuclease gene may explain the decrease in the number of CRISPR-Cas systems in , known to deactivate the endonucleases' protective activities and enable the genome to be versatile in acquiring mobile genetic elements, including carriers of antimicrobial resistance genes, especially . Most importantly, we observed that there was a direct association between the absence of CRISPR-Cas, the presence of the anti-CRISPR gene and the acquisition of vancomycin resistance genes.
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http://dx.doi.org/10.3390/microorganisms9061118DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8224324PMC
May 2021

12/111phiA Prophage Domestication Is Associated with Autoaggregation and Increased Ability to Produce Biofilm in .

Microorganisms 2021 May 21;9(6). Epub 2021 May 21.

UMR INRAE 1282 Infectiologie et Santé Publique, Bactéries et Risque Materno-Foetal, Université de Tours, 37000 Tours, France.

CC17 carrying group-A prophages is increasingly responsible for neonatal infections. To investigate the impact of the genetic features of a group-A prophage, we first conducted an in silico analysis of the genome of 12/111phiA, a group-A prophage carried by a strain responsible for a bloodstream infection in a parturient. This revealed a Restriction Modification system, suggesting a prophage maintenance strategy and five ORFs of interest for the host and encoding a type II toxin antitoxin system RelB/YafQ, an endonuclease, an S-adenosylmethionine synthetase MetK, and an StrP-like adhesin. Using the WT strain cured from 12/111phiA and constructing deleted mutants for the ORFs of interest, and their complemented mutants, we demonstrated an impact of prophage features on growth characteristics, cell morphology and biofilm formation. Our findings argue in favor of 12/111phiA domestication by the host and a role of prophage features in cell autoaggregation, glycocalyx and biofilm formation. We suggest that lysogeny may promote GBS adaptation to the acid environment of the vagina, consequently colonizing and infecting neonates.
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http://dx.doi.org/10.3390/microorganisms9061112DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8223999PMC
May 2021

A metallo-β-lactamase enzyme for internal detoxification of the antibiotic thienamycin.

Sci Rep 2021 May 12;11(1):10062. Epub 2021 May 12.

IRD, APHM, MEPHI, IHU-Méditerranée Infection, Aix Marseille Univ, 19-21 Boulevard Jean Moulin, 13005, Marseille, France.

Thienamycin, the first representative of carbapenem antibiotics was discovered in the mid-1970s from soil microorganism, Streptomyces cattleya, during the race to discover inhibitors of bacterial peptidoglycan synthesis. Chemically modified into imipenem (N-formimidoyl thienamycin), now one of the most clinically important antibiotics, thienamycin is encoded by a thienamycin gene cluster composed of 22 genes (thnA to thnV) from S. cattleya NRRL 8057 genome. Interestingly, the role of all thn-genes has been experimentally demonstrated in the thienamycin biosynthesis, except thnS, despite its annotation as putative β-lactamase. Here, we expressed thnS gene and investigated its activities against various substrates. Our analyses revealed that ThnS belonged to the superfamily of metallo-β-lactamase fold proteins. Compared to known β-lactamases such as OXA-48 and NDM-1, ThnS exhibited a lower affinity and less efficiency toward penicillin G and cefotaxime, while imipenem is more actively hydrolysed. Moreover, like most MBL fold enzymes, additional enzymatic activities of ThnS were detected such as hydrolysis of ascorbic acid, single strand DNA, and ribosomal RNA. ThnS appears as a MBL enzyme with multiple activities including a specialised β-lactamase activity toward imipenem. Thus, like toxin/antitoxin systems, the role of thnS gene within the thienamycin gene cluster appears as an antidote against the produced thienamycin.
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http://dx.doi.org/10.1038/s41598-021-89600-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8115136PMC
May 2021

High frequency and diversity of Vancomycin-resistant Enterococci (VRE) in Algerian healthcare settings.

Infect Genet Evol 2021 08 30;92:104889. Epub 2021 Apr 30.

Aix-Marseille Univ., MEPHI, IRD, APHM, IHU-Méditerranée Infection, Marseille, France; IHU-Méditerranée Infection, Marseille, France. Electronic address:

The spread of vancomycin-resistant Enterococci (VRE) in Algerian hospital settings is poorly reported. Since the first report in 2006, few data have been available on the molecular mechanism of this resistance across the country. In this study, we investigate the frequency and antibiotic resistance mechanisms of Enterococci strains isolated from hospitalised patients in the Tlemcen university hospital. 191 Enterococcus spp. strains were collected from various clinical samples and were identified using MALDI-TOF-MS. The presence of van genes was investigated by standard PCR and sequencing. Results revealed that E. faecium and E. faecalis strains are the main pathogens identified in the study. Antibiotic susceptibility testing revealed that the resistance rate was high for the majority of antibiotic classes, including glycopeptides, and only linezolid was effective on all strains. Molecular analysis revealed that 52.2% of strains from intensive care unit (ICU) were positive for the vanA gene, including 44.44% E. faecium, 5.55% E. faecalis and 2.22% E. avium. 25.5% of these isolates co-harboured both the vanA and vanC genes, including E. gallinarum (n = 16) and E. faecium (n = 6). In surgical wards (SW) 29.70% of strains harboured the van genes, including 4.90% of E. faecalis harbouring the vanB gene, and of the rest of strains, (24.80%) harboured the vanC genes. Indeed, 9.90% E. gallinarum and 4.90% E. faecalis were positive for vanC1 and 9.90% of E. casseliflavus were positive for the vanC2/C3 gene. The glycopeptide resistance rate was higher among strains from the ICU and was mainly composed by E. faecium strains compared with surgical wards where resistant E. faecalis strains were predominant.
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http://dx.doi.org/10.1016/j.meegid.2021.104889DOI Listing
August 2021

Dissemination of Carbapenemases (OXA-48, NDM and VIM) Producing Isolated from the Mohamed VI University Hospital in Marrakech, Morocco.

Antibiotics (Basel) 2021 Apr 26;10(5). Epub 2021 Apr 26.

Aix Marseille Université, IRD, APHM, MEPHI, IHU-Mediterranée Infection, 13385 Marseille, France.

The emergence and spread of carbapenem-resistant (CRE) represent a major clinical problem and raise serious health concerns. The present study aimed to investigate and ascertain the occurrence of CRE among hospitalized patients of Mohamed VI University Hospital, Marrakech, Morocco. Biological samples were collected over a one-year period (2018). The bacterial isolates were identified by MALDI-TOF-MS. Antibiotic susceptibility testing was performed using disc diffusion and Etest. The modified Hodge test and combined disc diffusion test were used for phenotypic detection. CRE hydrolyzing enzyme encoding genes: OXA-48, KPC, IMP, VIM, and NDM were characterized by PCR and DNA sequencing. In total, 131 non-duplicate CRE clinical strains resistant to Ertapenem were isolated out of 1603 initial . was the most common species (59%), followed by (24%), (10%), (3%), (2%), (1%), and (1%). Of these, 56.49%, 21.37%, 15.27%, 3.38%, and 3.05% were collected from blood, urine, pus, catheters and respiratory samples, respectively. Approximately 85.5% (112/131) of the isolates were carbapenemase producers (40 OXA-48, 27 NDM, 38 OXA-48 + NDM and 7 VIM). All metallo-β-lactamases isolates were NDM-1 and VIM-1 producers. This is the first documentation of OXA-48 genes from and in Morocco.
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http://dx.doi.org/10.3390/antibiotics10050492DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8145435PMC
April 2021

Investigation of an XDR- ST2 outbreak in an intensive care unit of a Lebanese tertiary care hospital.

Future Microbiol 2020 10 20;15:1535-1542. Epub 2020 Nov 20.

Laboratoire Microbiologie Santé et Environnement (LMSE), Doctoral School of Science & Technology, Faculty of Public Health, Lebanese University, Tripoli, Lebanon.

We sought to investigate the genetic epidemiological relatedness of carbapenem-resistant (CRAB) strains of a suspected outbreak in a Lebanese tertiary care hospital to implement necessary infection prevention and control measures. Twenty-eight nonduplicate CRAB isolates detected among hospitalized patients between January 2016 and July 2017 were studied by real-time polymerase chain reaction (PCR), pulsed-field gel electrophoresis and multilocus sequence typing analyses. Twenty-seven isolates harbored , of which one also carried . The isolates distributed temporally in two presumably episodes were stratified by pulsed-field gel electrophoresis into many clusters. Although several clones have become endemic in the hospital, we have rapidly implemented appropriate infection prevention and control measures, achieving full eradication from August 2017 to November 2019. We have successfully investigated and controlled a polyclonal outbreak of OXA-23 producing ST2 CRAB.
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http://dx.doi.org/10.2217/fmb-2020-0079DOI Listing
October 2020

Dual RNase and β-lactamase Activity of a Single Enzyme Encoded in Archaea.

Life (Basel) 2020 Nov 14;10(11). Epub 2020 Nov 14.

MEPHI, IHU-Mediterranee Infection, Aix Marseille University, 19-21 Bd Jean Moulin, 13005 Marseille, France.

β-lactam antibiotics have a well-known activity which disturbs the bacterial cell wall biosynthesis and may be cleaved by β-lactamases. However, these drugs are not active on archaea microorganisms, which are naturally resistant because of the lack of β-lactam target in their cell wall. Here, we describe that annotation of genes as β-lactamases in Archaea on the basis of homologous genes is a remnant of identification of the original activities of this group of enzymes, which in fact have multiple functions, including nuclease, ribonuclease, β-lactamase, or glyoxalase, which may specialized over time. We expressed class B β-lactamase enzyme from Methanosarcina barkeri that digest penicillin G. Moreover, while weak glyoxalase activity was detected, a significant ribonuclease activity on bacterial and synthetic RNAs was demonstrated. The β-lactamase activity was inhibited by β-lactamase inhibitor (sulbactam), but its RNAse activity was not. This gene appears to have been transferred to the Flavobacteriaceae group especially the Elizabethkingia genus, in which the expressed gene shows a more specialized activity on thienamycin, but no glyoxalase activity. The expressed class C-like β-lactamase gene, from Methanosarcina sp., also shows hydrolysis activity on nitrocefin and is more closely related to DD-peptidase enzymes. Our findings highlight the need to redefine the nomenclature of β-lactamase enzymes and the specification of multipotent enzymes in different ways in Archaea and bacteria over time.
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http://dx.doi.org/10.3390/life10110280DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7697635PMC
November 2020

, a New Family of Fosfomycin Resistance Genes Identified in Bacterial Species Isolated from Human Microbiota.

Antimicrob Agents Chemother 2021 01 20;65(2). Epub 2021 Jan 20.

Aix Marseille University, IRD, APHM, MEPHI, IHU Méditerranée Infection, Marseille, France

Fosfomycin is a decades-old antibiotic, currently reused because of its activity against multidrug-resistant bacteria. Here, we used a combined approach to search for fosfomycin resistance determinants in 25 new bacterial species isolated from the human microbiota. Putative resistance genes were cloned into a susceptible strain. MIC values increased from 1 μg/ml to 1,024 μg/ml. Here, we report a new family of potential chromosomal fosfomycin resistance genes, named .
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http://dx.doi.org/10.1128/AAC.01712-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7848996PMC
January 2021

Culturing Ancient Bacteria Carrying Resistance Genes from Permafrost and Comparative Genomics with Modern Isolates.

Microorganisms 2020 Oct 3;8(10). Epub 2020 Oct 3.

Aix Marseille Université, IRD, AP-HM, MEPHI, 13005 Marseille, France.

Long considered to be a consequence of human antibiotics use by deduction, antibiotic resistance mechanisms appear to be in fact a much older phenomenon as antibiotic resistance genes have previously been detected from millions of year-old permafrost samples. As these specimens guarantee the viability of archaic bacteria, we herein propose to apply the culturomics approach to recover the bacterial content of a Siberian permafrost sample dated, using the in situ-produced cosmogenic nuclide chlorine36 (Cl), at 2.7 million years to study the dynamics of bacterial evolution in an evolutionary perspective. As a result, we cultured and sequenced the genomes of 28 ancient bacterial species including one new species. To perform genome comparison between permafrost strains and modern isolates we selected 7 of these species (i.e., and ). We observed a high level of variability in genomic content with a percentage of shared genes in the core genomes ranging from 21.23% to 55.59%. In addition, the Single Nucleotide Polymorphism (SNP) comparison between permafrost and modern strains for the same species did not allow a dating of ancient strains based on genomic content. There were no significant differences in antibiotic resistance profiles between modern and ancient isolates of each species. Acquired resistance to antibiotics was phenotypically detected in all gram-negative bacterial species recovered from permafrost, with a significant number of genes coding for antibiotic resistance detected. Taken together, these findings confirm previously obtained data that antibiotic resistance predates humanity as most of antimicrobial agents are natural weapons used in inter-microbial conflicts within the biosphere.
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http://dx.doi.org/10.3390/microorganisms8101522DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7600834PMC
October 2020

Promiscuous Enzyme Activity as a Driver of Allo and Iso Convergent Evolution, Lessons from the β-Lactamases.

Int J Mol Sci 2020 Aug 29;21(17). Epub 2020 Aug 29.

Aix-Marseille Univ IRD, APHM, MEPHI, IHU Méditerranée Infection, 19-21 Boulevard Jean Moulin, 13005 Marseille, France.

The probability of the evolution of a character depends on two factors: the probability of moving from one character state to another character state and the probability of the new character state fixation. The more the evolution of a character is probable, the more the convergent evolution will be witnessed, and consequently, convergent evolution could mean that the convergent character evolution results as a combination of these two factors. We investigated this phenomenon by studying the convergent evolution of biochemical functions. For the investigation we used the case of β-lactamases. β-lactamases hydrolyze β-lactams, which are antimicrobials able to block the DD-peptidases involved in bacterial cell wall synthesis. β-lactamase activity is present in two different superfamilies: the metallo-β-lactamase and the serine β-lactamase. The mechanism used to hydrolyze the β-lactam is different for the two superfamilies. We named this kind of evolution an allo-convergent evolution. We further showed that the β-lactamase activity evolved several times within each superfamily, a convergent evolution type that we named iso-convergent evolution. Both types of convergent evolution can be explained by the two evolutionary mechanisms discussed above. The probability of moving from one state to another is explained by the promiscuous β-lactamase activity present in the ancestral sequences of each superfamily, while the probability of fixation is explained in part by positive selection, as the organisms having β-lactamase activity allows them to resist organisms that secrete β-lactams. Indeed, an organism that has a mutation that increases the β-lactamase activity will be selected, as the organisms having this activity will have an advantage over the others.
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http://dx.doi.org/10.3390/ijms21176260DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7504333PMC
August 2020

First whole genome sequence of Paenalcaligenes suwonensis bearing bla Metallo-β-lactamase: A clinical isolate responsible for acute gastroenteritis.

Infect Genet Evol 2020 11 27;85:104513. Epub 2020 Aug 27.

MEPHI, IRD, APHM, IHU-Mediterranee Infection, Faculté de Pharmacie, Aix-Marseille University, Marseille, France. Electronic address:

Carbapenemase-producing Alcaligenes species has been described in only few studies, with none so far from the African continent. Here, we report the whole genome sequence of Peanalcaligenes suwonensis bearing bla metallo-β-lactamase and first detection of carbapenemase producing Alcaligenes faecalis isolated from patients attending tertiary healthcare facilities in Nigeria. The isolates were identified by MALDI-TOF Mass Spectrometry. Antibiotic susceptibility assay, modified Carba NP test and genomic investigation revealed that two isolates of Alcaligenes faecalis and an isolate of Paenalcaligenes suwonensis harboured bla gene. The genome sequence analysis of the P. suwonensis 191B isolate, responsible for acute gastroenteritis, reveal the presence of 18 antibiotic resistance genes coding for resistance to five different classes of antibiotics. Three of the genes (bla, bla and bla) codes for resistance to β-lactam antibiotics. To our best knowledge, we describe here the first genome sequence of P. suwonensis species and the first detection of class B carbapenemase bla in a clinical isolate of P. suwonensis species and Alcaligenes faecalis in Nigeria. The finding of this study is of concern, as lateral dissemination of the genes into clinically important Gram-negative pathogens is highly likely.
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http://dx.doi.org/10.1016/j.meegid.2020.104513DOI Listing
November 2020

Molecular Characterization of Multidrug-Resistant Escherichia coli Isolated from Milk of Dairy Cows with Clinical Mastitis in Algeria.

J Food Prot 2020 Dec;83(12):2173-2178

Aix-Marseille Université, UMR Microbes Evolution Phylogeny and Infections, Institut de Recherche pour le Développement, Assistance Publique-Hôpitaux de Marseille, Institut Hospitalo-Universitaire-Méditerranée Infection, Faculté de Pharmacie, 19-21 Boulevard Jean Moulin, 13005 Marseille, France.

The objective of this study was to investigate the occurrence of multidrug-resistant Escherichia coli in cows with clinical mastitis in 42 different dairy farms located in the Bordj Bou Arreridj region of Algeria. Milk samples were cultured on Columbia blood agar, and isolates were then identified by matrix-assisted laser desorption ionization-time of flight mass spectrometry. In total, 200 samples were screened and 52 E. coli strains confirmed as causative agents were obtained. The antimicrobial susceptibility testing was performed by disk diffusion method. Antibiotic resistance genes, including those conferring resistance to extended-spectrum β-lactamases (i.e., blaTEM, blaSHV, and blaCTX-M), tetracyclines (tetA, tetB, tetC, and tetJ), aminoglycosides [aph(3'), aac(3'), aac(6'), ant, aad, and armA], and quinolones (qnrA and qnrB) were amplified by standard PCR and sequenced when positive. Transferability of resistance genes has been investigated by conjugation experiments and multilocus sequence typing. The most frequently observed resistance was to amoxicillin (86.5%), followed by tetracycline (75%), amoxicillin-clavulanic acid (59.6%), trimethoprim-sulfamethoxazole (36.5%), doxycycline (13.5%), and ciprofloxacin (13.5%). Multidrug resistance was observed in 38.4% of isolates. Genotypic characterization showed that tetA (44.2%) and blaTEM-1 (30.7%) genes were the most prevalent. Screening for plasmid-mediated quinolone resistance genes demonstrated that seven isolates (13.5%) expressed qnrB and one isolate (1.9%) harbored qnrA. In addition, aminoglycoside resistance determinants including aadA1 and aac(3)-Id were detected in seven and two isolates, respectively. Moreover, blaTEM, tetA, tetB, qnrB, and aadA1 were successfully transferred horizontally to transconjugant strains. The multilocus sequence typing revealed the presence of three different sequence types (ST162, ST371, and ST 949).
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http://dx.doi.org/10.4315/JFP-20-198DOI Listing
December 2020

Bacterial infection during wars, conflicts and post-natural disasters in Asia and the Middle East: a narrative review.

Expert Rev Anti Infect Ther 2020 06 17;18(6):511-529. Epub 2020 Apr 17.

Faculté de Médecine et de Pharmacie, Aix Marseille Univ, IRD, APHM, MEPHI, IHU-Méditerranée Infection, Marseille, France.

: Bacterial infections resulting from wars and natural disasters represent a major public health problem. Over the past 50 years, Asia and the Middle East have suffered several wars. Moreover, East-Asian countries are considered the most natural disaster-prone countries in the world.: This review focuses on bacterial infection occurring during wars and after natural disasters, among refugees, wounded citizens and soldiers as well as the prevention and control measures that must be taken.: During wars, refugees and soldiers represent the two main sources of bacterial infections. Refugees coming from countries with a high prevalence of antimicrobial resistance can spread these pathogens to their final destination. In addition, these refugees living in inadequate shelters can contribute to the spread of bacterial infections. Moreover, some factors including the presence of fixed imported fragments; environmental contamination and nosocomial transmissions, play a key role in the dissemination of bacteria among soldiers. As for natural disasters, several factors are associated with increased bacterial transmissions such as the displacement of large numbers of people into over-crowded shelters, high exposure to disease vectors, lack of water and sanitation. Here, we carry out a systematic review of the bacterial infections that follow these two phenomena.
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http://dx.doi.org/10.1080/14787210.2020.1750952DOI Listing
June 2020

Massive analysis of 64,628 bacterial genomes to decipher water reservoir and origin of mobile colistin resistance genes: is there another role for these enzymes?

Sci Rep 2020 04 6;10(1):5970. Epub 2020 Apr 6.

Aix Marseille Univ., IRD, APHM, MEPHI, IHU-Mediterranee Infection, Marseille, France.

Since 2015, new worrying colistin resistance mechanism, mediated by mcr-1 gene has been reported worldwide along with eight newly described variants but their source(s) and reservoir(s) remain largely unexplored. Here, we conducted a massive bioinformatic analysis of bacterial genomes to investigate the reservoir and origin of mcr variants. We identified 13'658 MCR-1 homologous sequences in 494 bacterial genera. Moreover, analysis of 64'628 bacterial genomes (60 bacterial genera and 1'047 species) allows identifying a total of 6'651 significant positive hits (coverage >90% and similarity >50%) with the nine MCR variants from 39 bacterial genera and more than 1'050 species. A high number of MCR-1 was identified in Escherichia coli (n = 862). Interestingly, while almost all variants were identified in bacteria from different sources (i.e. human, animal, and environment), the last variant, MCR-9, was exclusively detected in bacteria from human. Although these variants could be identified in bacteria from human and animal sources, we found plenty MCR variants in unsuspected bacteria from environmental origin, especially from water sources. The ubiquitous presence of mcr variants in bacteria from water likely suggests another role in the biosphere of these enzymes as an unknown defense system against natural antimicrobial peptides and/or bacteriophage predation.
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http://dx.doi.org/10.1038/s41598-020-63167-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7136264PMC
April 2020

Inactivation of thymidine kinase as a cause of resistance to zidovudine in clinical isolates of Escherichia coli: a phenotypic and genomic study.

J Antimicrob Chemother 2020 06;75(6):1410-1414

Aix Marseille Univ, IRD, APHM, MEPHI, IHU-Méditerranée Infection, Marseille, France.

Objectives: The antiviral zidovudine has been recently identified as an active drug against resistant Enterobacteriaceae, but prevalence of resistance to this compound remains unknown. The aim was to estimate the prevalence of clinical Escherichia coli isolates resistant to zidovudine and to decipher the mechanism of zidovudine resistance.

Methods: We screened 537 isolates on zidovudine-containing agar plates and studied their thymidine kinase (tdk) gene sequences, the putative target involved in zidovudine resistance. Moreover, sequence analysis of 633 complete genomes of E. coli was performed to investigate mutation in the tdk gene. A comparative genomic analysis was done on an in vitro zidovudine-resistant mutant.

Results: After screening on our medium containing 2.7 mg/L (10 μM) zidovudine, nine strains had a zidovudine MIC >26.7 mg/L. The gene was absent in three isolates, inactivated by an IS (IS1X2 and ISApl1) in two isolates and mutated in four isolates. A genomic analysis of 633 E. coli genomes showed heterogeneity of the tdk gene sequence, with 27 different sequences. Among them, three genomes showed an inactivation of the gene (IS, stop codon and no tdk gene sequence). The in vitro mutant E. coli had 27 SNPs in eight genes of the core genome compared with the initial strain.

Conclusions: Our study reports zidovudine-resistant clinical isolates of E. coli, presumably related to tdk inactivation. Diversity of Tdk in bacterial genomes can be large. Other mechanisms need to be considered in zidovudine resistance. The use of zidovudine in antibiotic-resistant infections needs to be in combination and should be tested before clinical administration.
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http://dx.doi.org/10.1093/jac/dkaa057DOI Listing
June 2020

Intestinal carriage of colistin-resistant Enterobacteriaceae at Saint Georges Hospital in Lebanon.

J Glob Antimicrob Resist 2020 06 12;21:386-390. Epub 2019 Dec 12.

Aix Marseille University, IRD, APHM, MEPHI, IHU-Méditerranée Infection, Marseille, France, Faculté de Médecine et de Pharmacie, 19-21 Boulevard Jean Moulin, 13385 Marseille CEDEX 05, France. Electronic address:

Objectives: The increase in resistance to antibiotics has led to the revival of colistin as the last option for treatment, which automatically led to an increase of colistin-resistant, Gram-negative bacteria. In this study, we report the presence of clinical colistin-resistant Enterobacteriaceae isolated from a Lebanese hospital.

Methods: From 23 rectal swabs, eight colistin-resistant clinical strains (five Escherichia coli, two Enterobacter cloacae, and one Klebsiella pneumoniae) were isolated. Antibiotic susceptibility testing was performed using the disk diffusion method and Etest. The broth microdilution method was used to determine colistin susceptibility. Reverse transcription polymerase chain reaction (RT-PCR), standard PCR and sequencing were used to investigate genes encoding for extended-spectrum β-lactamases, carbapenemases and colistin resistance. Genotyping of these isolates was conducted by multilocus sequence typing (MLST).

Results: Results of antibiotic susceptibility testing revealed that all isolates were resistant to colistin. They had MICs for colistin that ranged from 8 to 32 mg/L. Real-time PCR results showed that five strains harboured bla and one strain harboured bla. Moreover, four strains were positive for bla, bla and bla, and K. pneumoniae harboured bla. Observed colistin resistance was linked to amino acid substitutions into protein sequences of pmrA/B, phoP/Q, and mgrB. Interestingly, we report here a mutation in the mgrB regulator and pmrA/B, phoP/Q in colistin-resistant E. cloacae and E. coli clinical isolates for the first time in Lebanon.

Conclusion: This study highlights the presence of colistin-resistant Gram-negative bacteria in a Lebanese hospital, which is worrisome. An urgent strategy needs to be adopted to avoid the spread of such bacteria.
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http://dx.doi.org/10.1016/j.jgar.2019.12.001DOI Listing
June 2020

Autochthonous case of mobile colistin resistance gene mcr-1 from a uropathogenic Escherichia coli isolate in Sétif Hospital, Algeria.

J Glob Antimicrob Resist 2019 12 9;19:356-357. Epub 2019 Nov 9.

Aix-Marseille Université, IRD, APHM, MEPHI, IHU Méditerranée Infection, 19-21 Boulevard Jean Moulin, 13385 Marseille Cedex 05, France; IHU Méditerranée Infection, Marseille, France. Electronic address:

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http://dx.doi.org/10.1016/j.jgar.2019.10.006DOI Listing
December 2019

Development of real-time PCR assay allowed describing the first clinical Klebsiella pneumoniae isolate harboring plasmid-mediated colistin resistance mcr-8 gene in Algeria.

J Glob Antimicrob Resist 2020 03 30;20:266-271. Epub 2019 Aug 30.

Aix-Marseille Univ., IRD, APHM, MEPHI, IHU-Mediterranee Infection, 19-21 boulevard Jean Moulin, 13005 Marseille, France; IHU-Mediterranee Infection, Marseille, France. Electronic address:

Objectives: We aimed to develop here a specific real-time PCR assay with TaqMan® probe to detect efficiently bacterial strains harboring the new plasmid mediated-colistin resistance mcr-8 gene.

Methods: Specific primers and probe for mcr-8 gene were designed from sequences alignment of all mcr genes variants. Specificity of the designed primers and probe were first checked par BlastN analysis and by in silico PCR. The analytical sensitivity and specificity tests were performed in vitro on a panel of 290 genomic DNA of Gram-negative bacteria and 250 metagenomic DNA from human stool samples. Whole genome sequencing (WGS) was performed here using MiSeq technology.

Results: Designed primers and probe were 100% specific tomcr-8 gene by BlastN and in silico PCR analysis. Real-time PCR screening of a collection of clinical isolates resulted to one positive Klebsiella pneumoniae isolate (KP95). WGS confirmed that this isolate harbored the mcr-8 gene and other resistance genes such as bla, bla β-lactamases. Our real-time PCR was highly sensitive on a 10-fold dilution serie from a calibrated inoculum at 10 CFU/mL with a limit of detection at 55 CFU/mL.

Conclusion: To the best of our knowledge, we propose here, the first real-time PCR assay targeting mcr-8 gene with high specificity and sensitivity, able to detect mcr-8 gene in less than 2 h from any DNA sample. This real-time PCR assay allowed the first description of a clinical K. pneumoniae strain harboring the mcr-8 gene in Algeria.
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http://dx.doi.org/10.1016/j.jgar.2019.08.018DOI Listing
March 2020

Human metallo-β-lactamase enzymes degrade penicillin.

Sci Rep 2019 08 21;9(1):12173. Epub 2019 Aug 21.

Aix Marseille Université, MEPHI, IHU-Méditerranée Infection, Marseille, France.

Nonribosomal peptides are assemblages, including antibiotics, of canonical amino acids and other molecules. β-lactam antibiotics act on bacterial cell walls and can be cleaved by β-lactamases. β-lactamase activity in humans has been neglected, even though eighteen enzymes have already been annotated such in human genome. Their hydrolysis activities on antibiotics have not been previously investigated. Here, we report that human cells were able to digest penicillin and this activity was inhibited by β-lactamase inhibitor, i.e. sulbactam. Penicillin degradation in human cells was microbiologically demonstrated on Pneumococcus. We expressed a MBLAC2 human β-lactamase, known as an exosome biogenesis enzyme. It cleaved penicillin and was inhibited by sulbactam. Finally, β-lactamases are widely distributed, archaic, and have wide spectrum, including digesting anticancer and β-lactams, that can be then used as nutriments. The evidence of the other MBLAC2 role as a bona fide β-lactamase allows for reassessment of β-lactams and β-lactamases role in humans.
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http://dx.doi.org/10.1038/s41598-019-48723-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6704141PMC
August 2019

Inactivation of Causes High Rhodomyrtone Resistance and Increased Pathogenicity in .

Front Microbiol 2019 28;10:1157. Epub 2019 May 28.

Microbial Genetics, Interfaculty Institute of Microbiology and Infection Medicine Tübingen (IMIT), University of Tübingen, Tübingen, Germany.

Rhodomyrtone (Rom) is an acylphloroglucinol antibiotic originally isolated from leaves of . Rom targets the bacterial membrane and is active against a wide range of Gram-positive bacteria but the exact mode of action remains obscure. Here we isolated and characterized a spontaneous Rom-resistant mutant from the model strain HG001 (Rom) to learn more about the resistance mechanism. We showed that Rom-resistance is based on a single point mutation in the coding region of [regulator of fatty acid (FA) resistance] that causes an amino acid change from Cys to Arg at position 116 in FarR, that affects FarR activity. Comparative transcriptome analysis revealed that mutated affects transcription of many genes in distinct pathways. FarR represses for example the expression of its own gene (), its flanking gene (effector of FA resistance), and other global regulators such as and . All these genes were consequently upregulated in the Rom clone. Particularly the upregulation of and leads to increased expression of virulence genes rendering the Rom clone more cytotoxic and more pathogenic in a mouse infection model. The Rom-resistance is largely due to the de-repression of . FarE is described as an efflux pump for linoleic and arachidonic acids. We observed an increased release of lipids in the Rom clone compared to its parental strain HG001. If is deleted in the Rom clone, or, if native is expressed in the Rom strain, the corresponding strains become hypersensitive to Rom. Overall, we show here that the high Rom-resistance is mediated by overexpression of in the Rom clone, that FarR is an important regulator, and that the point mutation in (Rom clone) makes the clone hyper-virulent.
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http://dx.doi.org/10.3389/fmicb.2019.01157DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6547885PMC
May 2019

Evaluation of different testing tools for the identification of non-gonococcal Neisseria spp. isolated from Lebanese male semen: a strong and significant association with infertility.

J Med Microbiol 2019 Jul 7;68(7):1012-1020. Epub 2019 Jun 7.

Aix-Marseille University, IRD, APHM, MEPHI, IHU Méditerranée infection, Faculté de Médecine et de Pharmacie, 19-21 boulevard Jean Moulin, 13385 Marseille CEDEX 05, France.

Purpose: The aim was to evaluate several microbiological tools for the identification of non-gonococcal Neisseria spp. isolated from semen samples from Lebanese men and to determine the putative link between the presence of Neisseria commensal species and infertility.

Methodology: Within a cross-sectional retrospective study design, the whole population included in this investigation was divided in 2 categories: 173 patients with symptoms of infertility and 139 patients with normal seminograms. Epidemiological and microbiological investigations were performed for 59 strains of Neisseria through several phenotypic and genotypic tools, including seminograms, an analytical profile index of Neisseria and Haemophilus (API-NH), matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), porA PCR, 16S rRNA and rplF gene sequencing, and antimicrobial susceptibility testing.

Results: The risk of Neisseria infection was twice as high in infertile patients compared to the control group [odds ratio (OR): 1.95, confidence interval (CI): 1.05-3.65, P =0.03]. Unreliable diagnosis of Neisseria urogenital infection has serious health and social consequences. Our findings showed that API-NH and 16S rRNA sequencing are poor tools to identify Neisseria at the species level. Therefore, reliable diagnosis of cases using MALDI-TOF MS and/or rplF sequencing is needed to provide critical treatment decisions and prevent antimicrobial resistance spreading in the community.

Conclusion: This work predicted a strong and significant association between the presence of Neisseria spp. in semen and male infertility among the Lebanese population. For a better understanding of this association, it is recommended that more genomic and large-scale epidemiological investigations are undertaken to reach definitive conclusions.
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http://dx.doi.org/10.1099/jmm.0.000990DOI Listing
July 2019

Temperate Prophages Increase Bacterial Adhesin Expression and Virulence in an Experimental Model of Endocarditis Due to From the CC398 Lineage.

Front Microbiol 2019 24;10:742. Epub 2019 Apr 24.

Genomic Research Laboratory, Service of Infectious Diseases, Medical University Center, Geneva University Hospitals, Geneva, Switzerland.

Until 2007, from clonal complex 398 (CC398) was exclusively associated with livestock species and companion animals. Recently, several studies described the emergence of CC398 as etiologies of severe infections in humans living in an animal-free environment. Recent sequencing efforts showed that the mobile genetic elements found in CC398 isolates were specific for each population and enabled differentiation of strains responsible for asymptomatic colonization from strains involved in bloodstream infections. We mobilized prophages from a human CC398 isolate and introduced them into two naïve ancestral isolates devoid of prophages that exclusively colonize animals. These lysogenized ancestral CC398 isolates acquired features related to virulence, such as an increased capacity to adhere to human extracellular matrix proteins and the ability to invade and survive within non-phagocytic cells. Pathogenicity of several clinical isolates from the CC398 lineage as well as ancestral and lysogenized ancestral counterparts was assessed in a model of infectious endocarditis in rats. Natural and artificial lysogens were not only more invasive than their prophage-free parent but also showed an increased capacity to multiply within aortic vegetations. This study identified prophages as mediators of bacterial virulence in a model of infectious endocarditis, probably through promotion of interaction with extracellular matrix components. Further studies are needed to identify mechanisms leading to promotion of intrinsic virulence.
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http://dx.doi.org/10.3389/fmicb.2019.00742DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6492496PMC
April 2019

Prevalence and characterization of Staphylococcus aureus in wastewater treatment plants by whole genomic sequencing.

Water Res 2019 Jul 15;158:193-202. Epub 2019 Apr 15.

Department of Biology, College of Arts and Sciences, University of Kentucky, USA.

Infections with Staphylococcus aureus are being spread through contact with the community environment, but the role of wastewater treatment plants in the transmission routes is not defined. This study investigated the prevalence, types, genetic elements, and potential for transmission of S. aureus by these engineered systems. Synchronized sampling events at two wastewater treatment plants were conducted with isolates of S. aureus obtained by a selective enrichment method using acriflavine that suppressed Staphylococcus epidermidis growth. DNA was extracted from a subset of the S. aureus isolates, checked by PCR to assure the absence of S. epidermidis, and sequenced to determine the multilocus sequence type, spa type, and carriage of the methicillin resistance and Panton-Valentine leukocidin genetic elements. Sequences were analyzed for single nucleotide polymorphism differences in pairwise comparison of isolates. There were two dominant S. aureus clonal complexes identified in the isolates, one commonly identified as hospital-related (CC5) and one community-related (CC8). Both types of isolates were found at both treatment facilities, even though only one facility had significant hospital sewage inputs. The presence of S. aureus persisted through treatment, with some isolates recovered from the final processes showing genetic diversity. The presence of the Panton-Valentine leukocidin genetic element was greater than the 1-5% expected from global reports. Our results suggest that treatment provides an opportunity for genetic shift, while the persistence and release of evolved strains of S. aureus may provide an environmentally relevant pathway to new hosts in the environment.
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http://dx.doi.org/10.1016/j.watres.2019.04.035DOI Listing
July 2019

How to discover new antibiotic resistance genes?

Expert Rev Mol Diagn 2019 04 21;19(4):349-362. Epub 2019 Mar 21.

a Microbes Evolution Phylogeny and Infections (MEPHI), IRD, APHM, IHU Méditerranée Infection, Faculté de Médecine et de Pharmacie , Aix-Marseille-Univ , Marseille , France.

Introduction: Antibiotic resistance (AR) is a worldwide concern and the description of AR have been discovered mainly because of their implications in human medicine. Since the recent burden of whole-genome sequencing of microorganisms, the number of new AR genes (ARGs) have dramatically increased over the last decade. Areas covered: In this review, we will describe the different methods that could be used to characterize new ARGs using classic or innovative methods. First, we will focus on the biochemical methods, then we will develop on molecular methods, next-generation sequencing and bioinformatics approaches. The use of various methods, including cloning, mutagenesis, transposon mutagenesis, functional genomics, whole genome sequencing, metagenomic and functional metagenomics will be reviewed here, outlining the advantages and drawbacks of each method. Bioinformatics softwares used for resistome analysis and protein modeling will be also described. Expert opinion: Biological experiments and bioinformatics analysis are complementary. Nowadays, the ARGs described only account for the tip of the iceberg of all existing resistance mechanisms. The multiplication of the ecosystems studied allows us to find a large reservoir of AR mechanisms. Furthermore, the adaptation ability of bacteria facing new antibiotics promises a constant discovery of new AR mechanisms.
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http://dx.doi.org/10.1080/14737159.2019.1592678DOI Listing
April 2019

An Integrative Database of β-Lactamase Enzymes: Sequences, Structures, Functions, and Phylogenetic Trees.

Antimicrob Agents Chemother 2019 05 25;63(5). Epub 2019 Apr 25.

Aix Marseille Université, IRD, APHM, MEPHI, IHU-Méditerranée Infection, Marseille, France

β-Lactamase enzymes have attracted substential medical attention from researchers and clinicians because of their clinical, ecological, and evolutionary interest. Here, we present a comprehensive online database of β-lactamase enzymes. The current database is manually curated and incorporates the primary amino acid sequences, closest structural information in an external structure database (the Protein Data Bank [PDB]) and the functional profiles and phylogenetic trees of the four molecular classes (A, B, C, and D) of β-lactamases. The functional profiles are presented according to the MICs and kinetic parameters that make them more useful for the investigators. Here, a total of 1,147 β-lactam resistance genes are analyzed and described in the database. The database is implemented in MySQL and the related website is developed with Zend Framework 2 on an Apache server, supporting all major web browsers. Users can easily retrieve and visualize biologically important information using a set of efficient queries from a graphical interface. This database is freely accessible at http://ifr48.timone.univ-mrs.fr/beta-lactamase/public/.
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http://dx.doi.org/10.1128/AAC.02319-18DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6496087PMC
May 2019

Investigation of multidrug-resistant ST2 Acinetobacter baumannii isolated from Saint George hospital in Lebanon.

BMC Microbiol 2019 02 2;19(1):29. Epub 2019 Feb 2.

Aix Marseille Univ, IRD, APHM, MEPHI, IHU-Méditerranée Infection, Faculté de Médecine et de Pharmacie, 19-21 boulevard Jean Moulin, 13385, Marseille, Cedex 05, France.

Background: Acinetobacter baumannii is an opportunistic pathogen causing various nosocomial infections. The spread of multidrug-resistant A. baumannii is a major public health problem. The aim of this study was to investigate the molecular epidemiology and the genetic support of multidrug-resistant A. baumannii isolates collected from Saint-Georges Hospital in Lebanon.

Methods: Between January and August 2016, 31 A. baumannii isolates were collected from sputum samples of patients infected with ventilator-associated pneumonia (VAP) and treated with colistin-carbapenem combination therapy. Antibiotic susceptibility testing was performed using the disk diffusion method. Carbapenemases, extended spectrum β-lactamases encoding genes and mcr-1/2 genes were investigated by RT-PCR and standard PCR. The epidemiological relatedness of the strains was studied using MLST analysis.

Results: Most of the isolates exhibited multidrug-resistant phenotypes. All the isolates were carbapenem-resistant and among them, 30 carried the class D carbapenemase bla gene while one isolate carried bla gene. MLST results revealed three sequence types, namely ST2, ST699, and ST627. Isolates having ST2 were the most prevalent clone (29/31, 93.5%).

Conclusions: This study shows a nosocomial spread of multidrug-resistant A. baumannii ST2 having bla gene in Saint-George in Lebanon. Monitoring and control measures need to be adopted to avoid the spread of A. baumannii to patients.
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http://dx.doi.org/10.1186/s12866-019-1401-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6359860PMC
February 2019

Detection of a new variant of OXA-23 carbapenemase in Acinetobacter radioresistens isolates from urban animals in Marseille, France.

J Glob Antimicrob Resist 2019 03 24;16:178-180. Epub 2019 Jan 24.

Aix-Marseille Université, IRD, APHM, MEPHI, IHU-Méditerranée Infection, 19-21 Bd. Jean Moulin, 13005 Marseille, France. Electronic address:

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http://dx.doi.org/10.1016/j.jgar.2019.01.021DOI Listing
March 2019

Colistin- and Carbapenem-Resistant Klebsiella pneumoniae Clinical_Isolates: Algeria.

Microb Drug Resist 2019 Mar 25;25(2):258-263. Epub 2018 Sep 25.

1 Aix Marseille University , IRD, APHM, MEPHI, IHU-Mediterranean Infection, Marseille, France .

This study investigates the molecular mechanisms of colistin and carbapenem resistance in Klebsiella pneumoniae ST101 strains. The three K. pneumoniae carried bla, bla, and bla genes and two coharbored bla. As for colistin resistance, the isolates had amino acid substitutions in PmrA/B and a truncated mgrB gene in one isolate.
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http://dx.doi.org/10.1089/mdr.2018.0147DOI Listing
March 2019
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