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This study utilized integrative bioinformatics' tools together with phenotypic assays to understand the whole-genome features of a carbapenem-resistant international clone II Acinetobacter baumannii AB073. Overall, we found the isolate to be resistant to seven antibiotic classes, penicillins, β -lactam/ β -lactamase inhibitor combinations, cephalosporins, carbapenems, aminoglycosides, fluoroquinolones, and folate pathway antagonists. These resistance phenotypes are related to various chromosomal-located antibiotic resistance determinants involved in different mechanisms such as reduced permeability, antibiotic target protection, antibiotic target alteration, antibiotic inactivation, and antibiotic efflux. IC2 A. baumannii AB073 could not transfer antibiotic resistance by conjugation experiments. Likewise, mobilome analysis found that AB073 did not carry genetic determinants involving horizontal gene transfer. Moreover, this isolate also carried multiple genes associated with the ability of iron uptake, biofilm formation, immune invasion, virulence regulations, and serum resistance. In addition, the genomic epidemiological study showed that AB073-like strains were successful pathogens widespread in various geographic locations and clinical sources. In conclusion, the comprehensive analysis demonstrated that AB073 contained multiple genomic determinants which were important characteristics to classify this isolate as a successful international clone II obtained from Thailand., Competing Interests: The authors declare that they have no conflicts of interest., (Copyright © 2024 Rattiya Cheewapat et al.)

Acinetobacter baumannii is an important opportunistic pathogen, usually associated with immunocompromised individuals with a prolonged period of stay in a hospital. Currently, the incidence of multi-drug resistant A. baumannii (MDR-AB) and extensively drug-resistant A. baumannii (XDR-AB) is increasing rapidly in Thailand, mirroring the trend worldwide. Novel therapeutic approaches for the treatment of A. baumannii infection using bacteriophages are being evaluated, and the use of phage-derived peptides is being tested as alternative approach to fighting infection. In this study, we isolated and determined the biological features of a lytic A. baumannii phage called vB_AbaAut_ChT04 (vChT04). The vChT04 phage was classified as a member of the family Autographiviridae of the class Caudoviricetes. It showed a short latent period (10 min) and a large burst size (280 PFU cell -1 ), and it was able to infect 52 out of 150 clinical MDR-AB strains tested (34.67%). Most of the phage-sensitive strains were A. baumannii strains that had been isolated during the same year that the phage was isolated. The phage showed activity across a broad pH (pH 5.0-8.0) and temperature (4-37°C) range. Whole-genome analysis revealed that the vChT04 genome comprises 41,158 bp with a 39.3% GC content and contains 48 open reading frames (ORFs), 28 of which were assigned putative functions based on homology to previously identified phage genes. Comparative genomic analysis demonstrated that vChT04 had the highest similarity to phage vB_AbaP_WU2001, which was isolated in the southern part of Thailand. An endolysin gene found in the vChT04 genome was used to synthesize an antimicrobial peptide (designated as PLysChT04) and its antimicrobial activity was evaluated using minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) assays. The MIC and MBC values of peptide PLysChT04 against MDR-AB and XDR-AB were 312.5-625 µg/mL, and it was able to inhibit both phage-susceptible and phage-resistant isolates collected over different time periods. PLysChT04 showed good efficacy in killing drug-resistant A. baumannii and other bacterial strains, and it is a promising candidate for development as an alternative therapeutic agent targeting A. baumannii infections., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Austria, part of Springer Nature.)

Phage lytic enzymes are promising antimicrobial agents. In this study, an endolysin derived from vB_AbaM_PhT2 (vPhT2), was identified. This endolysin represented the conserved lysozyme domain. Recombinant endolysin (lysAB- vT2) and hydrophobic fusion endolysin (lysAB-vT2-fusion) were expressed and purified. Both endolysins showed lytic activity against bacterial crude cell wall of Gram-negative bacteria. The MIC of lysAB-vT2-fusion was 2 mg/ml corresponding to 100 µM, while the MIC of lysAB-vT2 was more than 10 mg/ml (400 µM). Combination of lysAB-vT2-fusion with colistin, polymyxin B or copper was synergistic against A. baumannii (FICI value as 0.25). Antibacterial activity of lysAB-vT2-fusion plus colistin at the fractional inhibitory concentrations (FICs) revealed that it can inhibit Escherichia coli, Klebsiella pneumoniae and various strains of extremely drug-resistant A. baumannii (XDRAB) and phage resistant A. baumannii. The lysAB- vT2-fusion still retained its antibacterial activity after incubating the enzyme at 4, 20, 40 and 60 °C for 30 min. The lysAB-vT2-fusion could inhibit the mature biofilm, and incubation of lysAB-vT2-fusion with T24 human cells infected with A. baumannii led to a partial reduction of LDH release from T24 cells. In summary, our study highlights the antimicrobial ability of engineered lysAB-vT2-fusion endolysin, which can be applied for the control of A. baumannii infection., (© 2023. Crown.)

Background: Acinetobacter baumannii ( A. baumannii ) is an important cause of nosocomial infection, especially in intensive care units (ICUs). It has the propensity to tolerate various environments and multiple classes of antibiotics. Our study aimed to characterize the comparative genomes of A. baumannii from hospital environments and clinical isolates., Methods: Clinical and environmental A. baumannii isolates were collected from a university hospital. Antibiotic susceptibility testing was performed, antibiotic resistance genes (ARGs) were characterized, and repetitive element palindromic-PCR (rep-PCR) typing was performed. Eight representative A. baumannii isolated from environmental and clinical samples from the same wards were selected for whole-genome sequencing (WGS) using the Illumina platform., Results: A total of 106 A. baumannii isolates were obtained from 312 hospital environmental samples. A high percentage of samples with A. baumannii colonization were detected from AMBU bags (77.9%), followed by bedrails (66.7%) and suction tubes (66.7%). We found that 93.4% of the environmental isolates were multidrug-resistant A. baumannii (MDRAB), and 44.7% were extremely drug-resistant A. baumannii (XDRAB). bla OXA-23 bla NDM, and bla OXA-58 were present in 80.2%, 78.3%, and 0.9% of all isolates, respectively. Sixty-one A. baumannii isolates were collected from patient specimens in the same ward. Among all A. baumannii clinical isolates, MDRAB and XDRAB accounted for 82% and 55.7%, respectively. The most dominant ARGs identified was bla OXA-23 (80.3%), followed by bla NDM (55.7%). The genetic diversity of all isolates using rep-PCR could be divided into 33 genotypes. The genome size of eight A. baumannii ranged from 3.78-4.01 Mb. We found six of eight strains to be bla NDM-5 -harboring A. baumannii . Mobile genetic elements (MGEs), such as integron1 ( intl 1), located upstream of bla NDM-5 were observed. The phylogenomic relationship of the core and pan genomes as well as the single nucleotide polymorphism (SNP) count matrix revealed the genetic similarity of A. baumannii environmental and clinical strains obtained from the same ward., Conclusion: This study confirmed that A. baumannii colonized in hospital environments were the main reservoir of nosocomial infection and provides critical information to guide the control of A. baumannii infection., Competing Interests: The authors declare that they have no competing interests., (© 2023 Kitti et al.)

Laccases are multicopper oxidase family enzymes that can oxidize various substrates. In this study, we isolated laccase-producing Acinetobacter spp. from the environment, and one isolate of laccase-producing Acinetobacter baumannii, designated NI-65, was identified. The NI-65 strain exhibited constitutive production of extracellular laccase in a crude extract using 2,6-dimethoxyphenol as a substrate when supplemented with 2 mM CuSO 4 . Whole-genome sequencing of the NI-65 strain revealed a genome size of 3.6 Mb with 3,471 protein-coding sequences. The phylogenetic analysis showed high similarity to the genome of A. baumannii NCIMB8209. Three laccase proteins, PcoA and CopA, that belong to bacterial CopA superfamilies, and LAC-AB, that belongs to the I-bacterial bilirubin oxidase superfamily, were identified. These proteins were encoded by three laccase-coding genes (pcoA, copA, and lac-AB). The lac-AB gene showed a sequence similar to that of polyphenol oxidase (PPO). Gene clusters encoding the catabolized compounds involved in the utilization of plant substances and secondary metabolite biosynthesis gene clusters encoding antimicrobial compounds were identified. This is the first report of whole-genome sequencing of laccase-producing A. baumannii, and the data from this study help to elucidate the genome of A. baumannii to facilitate its application in synthetic biology for enzyme production., (© 2022. The Author(s).)

Acinetobacter baumannii is a major cause of nosocomial infection, and the incidence of extensively drug-resistant A. baumannii (XDRAB) infections has dramatically increased worldwide. In this study, we aimed to explore the complete genome sequence of XDRAB 329, ST1166/98 (Oxford/Pasteur), which is an outbreak clone from a hospital in Thailand. Whole-genome sequencing (WGS) was performed using short-read Illumina and long-read PacBio sequencing, and a conjugation assay of its plasmid was performed. The complete genome sequence of A. baumannii AB329 revealed a circular chromosome 3,948,038 bp in length with 39% GC content. Antibiotic resistance genes (ARGs), including beta-lactam resistance ( bla OXA-51 , bla ADC-25 , bla OXA-23 , bla TEM-1D) , aminoglycoside resistance ( aph(3') -Ia, aph (3″)-Ib, aph (6)-Id, arm A), tetracycline resistance ( tet (B), tet (R)), macrolide resistance ( mph (E), msr (E)), and efflux pumps, were found. Mobile genetic elements (MGEs) analysis of A. baumannii AB329 revealed two plasmids (pAB329a and pAB329b), three prophages, 19 genomic islands (GIs), and 33 insertion sequences (ISs). pAB329a is a small circular plasmid of 8,731 bp, and pAB329b is a megaplasmid of 82,120 bp. aph (3')-VIa was detected in pAB329b, and a major facilitator superfamily (MFS) transporter was detected in the prophage . Acinetobacter baumannii resistance island 4 (AbaR4) harboring tetracycline and aminoglycoside resistance was detected in the genome of A. baumannii AB329. pAB329b, which belongs to Rep-type GR6 (plasmid lineage LN_1), is a conjugative plasmid with the ability to transfer an aminoglycoside resistance gene to sodium azide-resistant A. baumannii. This study provides insights into the features of the MGEs of XDRAB , which are the main reservoir and source of dissemination of ARGs., Competing Interests: The authors declare there are no competing interests., (©2022 Khongfak et al.)

Escherichia coli, a bacterium that causes severe foodborne diseases, is transmitted to humans primarily through the consumption of contaminated foods. These foodborne pathogens are causing a public health problem that requires alternative control approaches, such as bacteriophage (phage) biocontrol. In this study, we characterized vB_EcoM_Tw01 (vTw01) isolated from sewage and vB_EcoM_Tcm05 (vTcm05) isolated from chicken meat. Both vTw01 and vTcm05 were assigned to the family Myoviridae based on their morphology, with the former exhibiting a narrow host range with low minimum inhibitory multiplicity of infection (miMOI), and the latter exhibiting a broad host range with high miMOI. The latent periods of these phages were 20 and 30 min for vTw01 and vTcm05, while the burst sizes were ∼140 and ∼300 PFU/cell, respectively. They were relatively stable over a wide range of pH values and temperatures. The bioinformatics analysis of the genomic sequence suggests that vTw01 and vTcm05 have double-stranded DNA with genome sizes of 170,107 bp and 149,059 bp, respectively. Bacteriophage encoded enzymes, such as tail-lysozyme, spanin Rz, holin, cell wall hydrolase (CWH), and endolysin, were identified in the genome of both phages. In conclusion, this study investigated the morphological, physiological, and genomic features of two E. coli phages isolated from different sources. It was confirmed that these phages and their enzymes can serve as potential candidates for phage biocontrol., (Copyright © 2022. Published by Elsevier B.V.)

Extensively drug-resistant Acinetobacter baumannii (XDR-AB) is a major threat to public health worldwide. A retrospective study for 27 XDR-AB isolates from four tertiary hospitals in Thailand was conducted. Beta-lactamase and virulence genes were characterized by PCR. The bla ADC , bla OXA-51 , and bla OXA-23 were detected in all isolates, whereas bla PER-1 and bla NDM-1 genes were present in 7.4% and 3.7% of isolates. All isolates had virulence genes, including genes in iron acquisition system, biofilm formation and secretion systems. The plasmids in XDR-AB belonged to GR2 (100%), GR6 (40.7%), and GR1 (7.4%). Multilocus sequence typing sequence types (STs) were further investigated. The data demonstrated that XDR-AB isolates had nine STs: ST195 ( n  = 4), ST208 ( n  = 4), ST368 ( n  = 1), ST451 ( n  = 5), ST457 ( n  = 2), ST1947 ( n  = 1), ST1166 ( n  = 7), including two novel STs namely ST1682 ( n  = 2) and ST1684 ( n  = 1). We observed that the majority ST1166 (25.9%) was associated with the prevalence of GR2 and GR6 plasmids and tr a U virulence gene. Genome-based single nucleotide polymorphism phylogenetic analysis of the isolates with two novel ST types indicated that the two isolates belonged to the international clone II (IC2) within the same cluster. In conclusion, our data showed the dissemination of XDR-AB isolates harbored virulence genes and antibiotic resistance genes among four hospitals in Thailand. The results highlighted the difficulty posed for the empirical treatment of the patients with the A. baumannii infection.

In this study, we examined the association between antimicrobial resistance, CRISPR/Cas systems and virulence with phage susceptibility in Acinetobacter baumannii and investigated draft genomes of phage susceptible multidrug resistant A. baumannii strains from Thailand. We investigated 230 A. baumannii strains using 17 lytic A. baumannii phages and the phage susceptibility was 46.5% (107/230). Phage susceptibility was also associated with resistance to numerous antibiotics (p-value < 0.05). We also found association between biofilm formation and the presence of ompA gene among phage susceptible A. baumannii strains (p-value < 0.05). A. baumannii isolates carrying cas5 or combinations of two or three other cas genes, showed a significant increase in phage resistance. Whole-genome sequences of seven phage susceptible A. baumannii isolates revealed that six groups of antibiotic resistance genes were carried by all seven phage susceptible A. baumannii. All strains carried biofilm associated genes and two strains harbored complete prophages, acquired copper tolerance genes, and CRISPR-associated (cas) genes. In conclusion, our data exhibits an association between virulence determinants and biofilm formation among phage susceptible A. baumannii strains. These data help to understand the bacterial co-evolution with phages.

The multi-drug resistance of the opportunistic pathogen Acinetobacter baumannii is of growing concern, with many clinical isolates proving to be resistant to last resort as well as front line antibiotic treatments. The use of bacteriophages is an attractive alternative to controlling and treating this emerging nosocomial pathogen. In this study, we have investigated bacteriophages collected from hospital wastewater in Thailand and we have explored their activity against clinical isolates of A. baumannii . Bacteriophage vB&#95;AbaM&#95;PhT2 showed 28% host range against 150 multidrug resistant (MDR) isolates and whole genome sequencing did not detect any known virulence factors or antibiotic resistance genes. Purified vB&#95;AbaM&#95;PhT2 samples had endotoxin levels below those recommended for preclinical trials and were not shown to be directly cytotoxic to human cell lines in vitro. The treatment of human brain and bladder cell lines grown in the presence of A. baumannii with this bacteriophage released significantly less lactate dehydrogenase compared to samples with no bacteriophage treatment, indicating that vB&#95;AbaM&#95;PhT2 can protect from A. baumannii induced cellular damage. Our results have also indicated that there is synergy between this bacteriophage and the end line antibiotic colistin. We therefore propose bacteriophage vB&#95;AbaM&#95;PhT2 as a good candidate for future research and for its potential development into a surface antimicrobial for use in hospitals.

The emergence and dissemination of colistin resistance among Gram-negative bacteria is a global problem. We initiated a surveillance of colistin-resistant and -susceptible Escherichia coli in raw meats from chicken in Nepal. A total of 180 meat samples were collected; from these, 60 E. coli strains were isolated (33.33%), of which 16 (26.66%) were colistin-resistant and harboured the mcr-1 gene. All isolates were characterised by antibiotic susceptibility testing, the presence of antibiotic resistance genes, phylogenetic analysis and plasmid replicon typing. Most of the colistin-resistant E. coli had the antibiotic resistant pattern CIP/CN/SXT/TE (43.75%). Coexistence of tet, qnr, sul and dfr genes was detected in both colistin-resistant and -susceptible E. coli. Most colistin-resistant E. coli strains belonged to phylogroup C, whereas 10% of isolates belonged to phylogroup D. Inc FIB was the dominant plasmid Inc type in the isolates. Dissemination of antibiotic-resistant E. coli in raw meats is a public health concern in Nepal and requires further investigation to ascertain the sources of contamination., (© FEMS 2019.)

Dissemination of mcr-1 encoding colistin resistance in Gram-negative bacteria has created critical situation in poultry, livestock farming, and public health. In Nepal, for the first time, we initiated surveillance of colistin-resistant Escherichia coli in broilers from seven different chicken farms. A total of 324 cloacal swabs were collected and 118 E. coli were isolated, of which 27 (22.8%) were colistin resistance all harboring mcr-1 gene, but lacking IS ApI1 . Colistin-resistant isolates were characterized by antibiotic susceptibility testing, detecting antibiotic resistance genes, phylogenetic analysis, and plasmid replicon typing. These isolates belonged to the phylo-group A (70.37%) and phylo-group D (29.63%). In addition, most isolates (>80%) were resistant to ciprofloxacin, tetracycline, and sulfamethoxazole-trimethoprim. As much as 3 of the 27 mcr-1 encoding isolates were confirmed as extended-spectrum β-lactamase (ESBL) producer, all 3 isolates carrying bla CTX-M gene. We performed the conjugation experiment to check transferability of mcr-1 , tet , and bla CTX-M genes, and only two donors were found to have transferred resistance to ticarcillin. The transfer of colistin and tetracycline resistance was not detected, which suggests the chromosomal location of mcr-1 and tet genes. The prevalence of Inc K/B and Inc I1 was 96.3% and 81.48%, respectively. This study shows the co-existence of mcr-1 with tet , sul , qnr , dfr , and bla CTX-M genes and dissemination of these resistant isolates in Nepalese chicken farms, which may pose huge threat to the livestock, especially chickens, and public health in Nepal.

Conjugation is a type of horizontal gene transfer (HGT) that serves as the primary mechanism responsible for accelerating the spread of antibiotic resistance genes in Gram-negative bacteria. The present study aimed to elucidate the mechanisms underlying the conjugation-mediated gene transfer from the extensively drug-resistant Acinetobacter baumannii (XDR-AB) and New Delhi Metallo-beta-lactamase-1-producing Acinetobacter baumannii (NDM-AB) to environmental isolates of Acinetobacter spp. Conjugation experiments demonstrated that resistance to ticarcillin and kanamycin could be transferred from four donors to two sodium azide-resistant A. baumannii strains, namely, NU013R and NU015R. No transconjugants were detected on Mueller-Hinton Agar (MHA) plates containing tetracycline. Plasmids obtained from donors as well as successful transconjugants were characterized by PCR-based replicon typing and S1-nuclease pulsed-field gel electrophoresis (S1-PFGE). Detection of antibiotic resistance genes and integrase genes (int) was performed using PCR. Results revealed that the donor AB364 strain can transfer the blaOXA-23 and blaPER-1 genes to both recipients in association with int1. A 240-kb plasmid was successfully transferred from the donor AB364 to recipients. In addition, the aphA6 and blaPER-1 genes were co-transferred with the int1 gene from the donor strains AB352 and AB405. The transfer of a 220-kb plasmid from the donors to recipient was detected. The GR6 plasmid containing the kanamycin resistance gene (aphA6) was successfully transferred from the donor strain AB140 to both recipient strains. However, the blaNDM-1 and tet(B) genes were not detected in all transconjugants. Our study is the first to demonstrate successful in vitro conjugation, which indicated that XDR-AB contained combination mechanisms of the co-transfer of antimicrobial resistance elements with integron cassettes or with the plasmid group GR6. Thus, conjugation could be responsible for the emergence of new types of antibiotic-resistant strains., Competing Interests: The authors have declared that no competing interests exist.

Acinetobacter baumannii is a major threat to public health due to the emergence and dissemination of antibiotic-resistant strains. The purpose of this study was to determine the molecular epidemiology of antibiotic-resistant A. baumannii isolates collected from four tertiary hospitals in Thailand during the period November 2013-February 2015. We screened 339 A. baumannii, nonrepetitive clinical isolates to determine drug susceptibility. Among all isolates, we found that 7.9% was nondrug-resistant A. baumannii (NR-AB). Carbapenem-resistant A. baumannii (CR-AB) strains accounted for 84.9% of the total isolates, with extensively drug-resistant A. baumannii (XDR-AB) accounting for 7.9% of the total isolates. We further investigated class D carbapenemase genes using multiplex-PCR amplification and class B metallo-β-lactamase genes, including bla IMP , bla VIM , and bla NDM-1 genes, using PCR and sequencing methods. We found that 300 (88.5%) isolates carried acquired class D carbapenemase genes, including bla OXA-23 (82.6%), bla OXA-24 (0.3%), and bla OXA-58 (6.5%). The genes bla IMP and bla VIM were not detected in any isolates. The bla NDM-1 was detected in 31 isolates from two hospitals (9.1%). All of the bla NDM-1 -positive A. baumannii (NDM-AB) had ISAba125 sequences upstream of the bla NDM-1 gene. A coexistence of three resistance genes-bla OXA-23 -bla OXA-58 -bla NDM-1 -was found in one isolate. A repetitive element palindromic-PCR (REP-PCR) revealed that all A. baumannii isolates were genetically diverse and could be divided into 33 genotypes. Only three genotypes were found to be predominant in all hospitals. Data from our study indicate the widespread emergence of multiple resistance determinants in A. baumannii isolates in Thailand, suggesting the need for more stringent infection control measures.

Methicillin-resistant coagulase negative staphylococci (MR-CoNS) are the major cause of infectious diseases because of their potential ability to form biofilm and colonize the community or hospital environments. This study was designed to investigate the biofilm producing ability, and the presence of mecA, icaAD, bap and fnbA genes in MR-CoNS isolates. The MR-CoNS used in this study were isolated from various samples of community environment and five wards of hospital environments, using mannitol salt agar (MSA) supplemented with 4 μg/ml of oxacillin. The specie level of Staphylococcus haemolyticus, Staphylococcus epidermidis, Staphylococcus hominis and Staphylococcus warneri was identified by specific primers of groESL (S. haemolyticus), rdr (S. epidermidis) and nuc (S. hominis and S. warneri). The remainder isolates were identified by tuf gene sequencing. Biofilm production was determined using Congo red agar (CRA) and Microtiter plate (MTP) assay. The mecA and biofilm associated genes (icaAD, fnbA and bap) were detected using PCR method. From the 558 samples from community and hospital environments, 292 MR-CoNS were isolated (41 from community environments, and 251 from hospital environments). S. haemolyticus (41.1%) and S. epidermidis (30.1%) were the predominant species in this study. Biofilm production was detected in 265 (90.7%) isolates by CRA, and 260 (88.6%) isolates were detected by MTP assay. The staphylococci isolates derived from hospital environments were more associated with biofilm production than the community-derived isolates. Overall, the icaAD and bap genes were detected in 74 (29.5%) and 14 (5.6%) of all isolates from hospital environments. When tested by MTP, the icaAD gene from hospital environment isolates was associated with biofilm biomass. No association was found between bap gene and biofilm formation. The MR-CoNS isolates obtained from community environments did not harbor the icaAD and bap genes. Conversely, fnbA gene presented in MR-CoNS isolated from both community and hospital environments. The high prevalence of biofilm producing MR-CoNS strains demonstrated in this study indicates the persisting ability in environments, and is useful in developing prevention strategies countering the spread of MR-CoNS.

The present study was conducted to isolate and characterize the molecular epidemiology of the methicillin-resistant staphylococci in the general university environment, where all five locations; the library, restrooms, canteens, computer rooms and outdoor surfaces, are in common use by a large population of students. We used Mannitol Salt Agar (MSA) supplemented with 4 μg/ml of oxacillin to screen the methicillin-resistant staphylococci. The species level was identified by PCR of rdr (Staphylococcus epidermidis), groESL (Staphylococcus haemolyticus) and nuc (Staphylococcus aureus and Staphylococcus warneri) genes and DNA sequencing of tuf and dnaJ genes. The susceptibility patterns of the isolates were determined using the disk diffusion method. Antibiotic and disinfectant resistance genes, together with SCCmec types, were detected by the PCR method. The methicillin resistant-staphylococci were isolated from 41 of 200 samples (20.5%), and all of them were found to be methicillin-resistant coagulase negative staphylococci (MR-CoNS). The library had the highest percentage of contamination, with 43.3% of the samples found to be contaminated. All isolates belonged to 6 different species including S. haemolyticus, S. epidermidis, S. warneri, S. cohnii, S. saprophyticus and S. hominis. The antimicrobial resistance rates were highest against penicillin (100%), then cefoxitin (73.1%), erythromycin (73.1%) and oxacillin (68.3%). Altogether, the isolates were approximately 61.0% multidrug resistant (MDR), with the S. epidermidis isolates being the most multidrug resistant. The prevalence of the qacA/B gene was detected in 63.4% of the isolates, and SCCmec could be typed in 43.9% (18/41) of the isolates. The type range was: II (n = 1), IVd (n = 1), I (n = 2), V (n = 6), IVa (n = 8) and untypeable (n = 23). This result indicates that these university environments are contaminated with methicillin-resistant coagulase negative staphylococci that carry various SCCmec types and high rate of disinfectant resistance genes. [Int Microbiol 20(2):65-73 (2017)]., (Copyright© by the Spanish Society for Microbiology and Institute for Catalan Studies.)

Background: Molecular analysis of carbapenem-resistant genes in Acinetobacter baumannii , an emerging pathogen, is less commonly reported from Nepal. In this study we determined the antibiotic susceptibility profile and genetic mechanism of carbapenem resistance in clinical isolates of A. baumannii., Methods: A. baumannii were isolated from various clinical specimens and identified based on Gram staining, biochemical tests, and PCR amplification of organism specific 16S rRNA and bla OXA-51 genes. The antibiotic susceptibility testing was performed using disc diffusion and E-test method. Multiplex PCR assays were used to detect the following β-lactamase genes: four class D carbapenem hydrolyzing oxacillinases ( bla OXA-51 , bla OXA-23 , bla OXA-24 and bla OXA-58 ). Uniplex PCRs were used to detect three class B metallo-β-lactamases genes ( bla IMP , bla VIM and bla NDM-1 ), class C cephalosporin resistance genes ( bla ADC ), aminoglycoside resistance gene ( aphA6 ), and IS Aba1 of all isolates. Insertion sequence IS Aba125 among NDM-1 positive strains was detected. Clonal relatedness of all isolates were analyzed using repetitive sequence-based PCR (rep-PCR)., Results: Of total 44 analyzed isolates, 97.7% ( n  = 43) were carbapenem-resistant A. baumannii (CR-AB) and 97.7% ( n  = 43) were multidrug resistant A. baumannii (MDR-AB). One isolate was detected to be extremely drug resistant A. baumannii (XDR-AB). All the isolates were fully susceptible to colistin (MICs < 2 μg/ml). The bla OXA-23 gene was detected in all isolates, while bla NDM-1 was detected in 6 isolates (13.6%). Insertion sequence, IS Aba1 was detected in all of bla OXA-23 positive isolates. IS Aba125 was detected in all bla NDM-1 positive strains. The bla ADC and aphA6 genes were detected in 90.1 and 40.1%, respectively. The rep-PCR of all isolates represented 7 different genotypes., Conclusion: We found high prevalence of CR-AB and MDR-AB with bla OXA-23 gene in a tertiary care hospital in Nepal. Systemic network surveillance should be established for monitoring and controlling the spread of these resistant strains.

Acinetobacter baumannii is an opportunistic bacterial pathogen that is the major cause of hospital-acquired infections. It has been shown that A. baumannii with high biofilm formation increases the risk of acquiring infection. In this study, the prevalence of virulence genes involved in biofilm formation was determined in 225 A. baumannii clinical isolates from three hospitals in Thailand. Most of the isolates were multidrug-resistant A. baumannii strains (86.2%). Among all isolates, 76.9% (173/225) showed biofilm formation ability. The association between biofilm forming ability and gentamicin resistance was found. The presence of virulence genes, epsA, bap, ompA, bfmS and blaPER-1 genes, was investigated by PCR. The prevalence of ompA, bfmS, bap, blaPER-1 and epsA genes among the isolated strains was 84.4%, 84%, 48%, 30.2%, respectively. Biofilm formation related genes, ompA and bap were associated with multidrug-resistant A. baumannii strains. The result of this study revealed that a high prevalence of biofilm-forming phenotypes among A. baumannii strains obtained from different hospitals. Effective strategies to prevent infection due to A. baumannii that produce biofilms are therefore needed. [Int Microbiol 19(2):121-129 (2016)]., (Copyright© by the Spanish Society for Microbiology and Institute for Catalan Studies.)