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A recent occurrence of carbapenemase-producing Acinetobacter ursingii was reported in the Netherlands and comprised three unrelated strains carrying the blaIMP-4 and blaOXA-58 encoding genes. The objective was to investigate a putative common source of the carbapenemase resistance genes and plasmids in these A. ursingii strains.

Objectives The aim of this study was to identify Acinetobacter spp. strains from paediatric patients, to determine their genetic relationship, to detect antibiotic resistance genes and to evaluate the role of efflux pumps in antibiotic resistance. Methods A total of 54 non-duplicate, non-consecutive Acinetobacter spp. isolates were collected from paediatric patients. Their genetic relationship, antibiotic resistance profile, efflux pump activity, antibiotic resistance genes and plasmid profile were determined. Results The isolates were identified as 24 Acinetobacter haemolyticus, 24 Acinetobacter calcoaceticus–baumannii (Acb) complex and 1 strain each of Acinetobacter junii, Acinetobacter radioresistens, Acinetobacter indicus, Acinetobacter lwoffii, Acinetobacter ursingii and Acinetobacter venetianus. The 24 A. haemolyticus were considered genetically unrelated. One strain was resistant to carbapenems, two to cephalosporins, two to ciprofloxacin and sixteen to aminoglycosides. The antibiotic resistance genes blaOXA-214 (29%), blaOXA-215 (4%), blaOXA-264 (8%), blaOXA-265 (29%), blaNDM-1 (4%), aac(6ʹ)-Ig (38%) and the novel variants blaOXA-575 (13%), blaTEM-229 (75%), aac(6ʹ)-Iga (4%), aac(6ʹ)-Igb (13%) and aac(6ʹ)-Igc (42%) were detected. Among 24 Acb complex, 5 were multidrug-resistant, carbapenem-resistant strains carrying blaOXA-51 and blaOXA-23; they were genetically related and had the same plasmid profile. Other species were susceptible. In some strains of A. haemolyticus and Acb complex, the role of RND efflux pumps was evidenced by a decrease in the MICs for cefotaxime, amikacin and ciprofloxacin in the presence of an efflux pump inhibitor. Conclusions This study identified isolates of A. haemolyticus carrying new β-lactamase variants and shows for the first time the contribution of efflux pumps to antibiotic resistance in this species.

BACKGROUND AND AIM Nowadays, anti-inflammation treatment is a promising approach for preventing tumorigenesis, and human microflora is closely related to inflammation. This study aimed to investigate the gastric cardiac microbiome and identify inflammation-related microorganisms for gastric cardiac inflammation. METHODS We performed 16S rRNA sequencing on a total of 11 healthy individuals and 89 individuals with different degree of gastric cardiac inflammation. Immunohistochemistry was used for verifying candidate bacteria. Phylogenetic reconstruction of unobserved states (picrust) was used for predicting the pathways involved by cardiac microflora. RESULTS The resident phyla in normal were Proteobacteria, Firmicutes, Bacteroides, and Actinobacteria, and the dominant genus in normal were Halomonas, shewanella, and Comamonas. In the progression of gastric cardiac inflammation, the diversity of cardiac microflora did not change (P > 0.05). However, the composition structure of cardiac microflora varied between healthy and inflamed tissues (P

Purpose The study aims to explore the characteristic microorganisms of the common tongue coatings in patients with gastric cancer (GC). Methods A total of 115 GC patients were assigned to four groups: White-thin coating (W-thin) group, White-thick coating (W-thick) group, Yellow-thin coating (Y-thin) group and Yellow-thick coating (Y-thick) group. Thirty-five healthy volunteers with White-thin coating were recruit as controls. High-throughput sequencing was used to describe the microbial community of the tongue coatings based on 16S rRNA and 18S rRNA genes. Multi-factors statistical analysis was carried out to present the microbial biomarkers of the tongue coating in GC patients. Results At bacterial phylum level, Saccharibacteria had higher relative abundance in W-thick group than W-thin group, Proteobacteria was more abundant in W-thin group than Y-thick group and less abundant in Y-thick group than Y-thin group. At fungal genus level, Guehomyces and Aspergillus presented to be significantly different among the common tongue coatings. Forteen significantly increased taxa were sorted out as the microbial biomarkers of common tongue coatings by LEfSe and ROC analysis. At species level, bacterial Capnocytophaga leadbetteri and fungal Ampelomyces_sp_IRAN_1 may be the potential biomarkers of W-thin coating, four bacterial species (Megasphaera micronuciformis, Selenomonas sputigena ATCC 35185, Acinetobacter ursingii, Prevotella maculosa) may be the potential biomarkers of W-thick coating. In general, the white coatings held more complex commensal relationship than the yellow coatings. Conclusion The common tongue coating owned characteristic microorganisms and special commensal relationship in the GC patients.

Background: Acinetobacter is widely distributed in nature as saprophytes. Recently, they have emerged as a nosocomial pathogendue to its ability for survival in the hospital environment on a wide range of dry and moist surface. They cause pneumonia, urinarytract infection (UTI), and surgical site infection (SSI) where drain tips are inserted, endocarditis, meningitis, peritonitis, andbacteremia. Antibiotic susceptibility pattern of Acinetobacter may vary geographically. Due to multidrug resistance patterns ofAcinetobacter, it is imperative to know the institutional prevalent susceptibility profiles.Aims and Objectives: This study was conducted to isolate Acinetobacter species from various clinical samples, to determine theantibiotic susceptibility pattern and to carry out the epidemiological investigation of the isolates.Materials and Methods: The study was conducted in a tertiary care hospital, over a period of 2 years. After identification, thespeciations of Acinetobacter isolates were done by biochemical tests and by VITEK 2. Antibiotic susceptibility was determined bydisc diffusion method. Extended-spectrum β-lactamases (ESBLs) and metallo-β-lactamases (MBLs) production were detectedby the combined disc diffusion test. An epidemiological study of Acinetobacter was carried out.Results: Of 5096 infected samples, 505 (9.9%) were non-fermenting Gram-negative bacilli, among which 170 (33.8%) werefound to be Acinetobacter. The highest numbers of isolates were Acinetobacter baumannii, followed by Acinetobacter lwoffii,Acinetobacter radioresistance, Acinetobacter calcoaceticus, Acinetobacter Haemolyticus, and Acinetobacter ursingii. Highestincidences of susceptibility were to imipenem (60%), chloramphenicol, and gentamicin. ESBL and MBL productions were detectedin 23% and 17%, respectively.Conclusion: A high level of antibiotic resistance was observed in this study and maximum isolation rate was in SSI. Most ofthe patients had high-risk factors such as prolonged hospitalization, indwelling catheters, and orthopedics implants in situ orother catheterization and diabetes. The analysis of susceptibility pattern will be useful in understanding the epidemiology of thisorganism in our hospital setup.

The potential of indigenously isolated bacteria from the Estância Velha stream to reduce Cr(VI) was evaluated and also the chromium contamination over the past ten years was verified in one of the most important industrial centers of Brazil, the “Brazilian Capital of Tanneries,” Estância Velha municipality in the Rio Grande do Sul State, South Brazil. Samples were collected from the Estância Velha stream at the source (P1), as well as at upstream (P2) and downstream (P3) of the most demographically area. The bacterial strains reduced between 52.5 and 61.6% of 250 mg L–1 Cr(VI) in 48 h. The genus Acinetobacter was the most abundant and could efficiently reduce 500 mg L–1 of Cr(VI); for example, P2.8 and P2.9 strains of Acinetobacter ursingii reduced 21.3 and 24.5% of 500 mg L–1 of Cr(VI), respectively, after 48 h. Moreover, an analysis of Cr levels in the stream sediment reported up to 3594 mg. L–1 of total Cr and up to 138 mg. L–1 of Cr(VI) in 2009. Acinetobacter strains were identified as the most abundant and efficient in reducing Cr(VI), makes them an ideal candidate for cleaning environments contaminated with tannery effluents, an approach that is more cost-effective than the traditional methods.

Background/purpose: Acinetobacter is an aerobic, gram-negative coccobacillus, which causes nosocomial infections including bacteremia. Recent development of molecular techniques has made classification of the Acinetobacter genomospecies possible, but there are still only a few studies comparing clinical features of the subspecies. We investigated bacteremia caused by Acinetobacter, isolated subspecies, and compared clinical features for each group. Methods: A retrospective analysis of Acinetobacter bacteremia cases was made in a 900-bed hospital in Japan. In addition to conventional procedures, subspecies identification based on rpoB sequence was made, and comparison of clinical characteristics between each subspecies were analyzed. Results: We collected 35 cases (Acinetobacter baumannii 14, A. nosocomialis 12, Acinetobacter ursingii 6, and A. seifertii 3). All of the A. seifertii bacteremia cases were blood stream infection occurring in cerebrovascular disease patients, showing particularly higher incidence of shock (100%) and high Pitt bacteremia score (PBS) (6.33 ± 2.52) in comparison to A. baumannii (43% and 2.86 ± 2.25, respectively). Sequential Organ Failure Assessment (SOFA) score and the PBS were slightly higher in A. nosocomialis in comparison to A. baumannii, and the 7 day mortality rate was higher in A. nosocomialis (25%) than in A. baumannii (7%), though this difference was not found to be significant. Conclusions: A.seifertii, the recently defined novel species, showed distinctive clinical features of bacteremia. And, in contrast to previous studies, the severity of A. nosocomialis infection was not lower than that of A. baumannii, which might suggest the influence of local epidemiology. Further characterization of these subspecies should be continued. Keywords: Acinetobacter seifertii, Acinetobacter nosocomialis, Acinetobacter baumanii, Bacteremia, rpoB, Genomospecies

The aim of this study was to investigate the phenotypic and genotypic antimicrobial resistance, integrons, and transferability of resistance markers in 243 aerobic bacteria recovered from pork at slaughter in the People's Republic of China. The organisms belonged to 22 genera of gram-negative bacteria (92.2%) and gram-positive bacteria (7.8%). High levels of resistance were detected to tetracycline, trimethoprim-sulfamethoxazole, and ampicillin (36.2 to 54.3%), and lower levels were detected to nitrofurantoin, cefotaxime, gentamicin, ciprofloxacin, and chloramphenicol (7.8 to 29.2%). Across species, genes conferring antimicrobial resistance were observed with the following frequencies: blaTEM, 40.7%; blaCMY-2, 15.2%; blaCTX-M, 11.5%; sul2, 27.2%; sul1, 14.4%; tet(A), 5.4%; tet(L), 5.4%; tet(M), 5.0%; tet(E), 3.7%; tet(C), 3.3%; tet(S), 2.5%; and tet(K), 0.8%. Various antimicrobial resistance genes were found in new carriers: blaTEM in Lactococcus garvieae, Myroides odoratimimus, Aeromonas hydrophila, Staphylococcus sciuri, Raoultella terrigena, Macrococcus caseolyticus, Acinetobacter ursingii, Sphingobacterium sp., and Oceanobacillus sp.; blaCMY-2 in Lactococcus lactis, Klebsiella oxytoca, Serratia marcescens, Acinetobacter baumannii, and Myroides phaeus; tet(L) in M. caseolyticus; sul1 in Vibrio cincinnatiensis; sul2 in Acinetobacter bereziniae, Acinetobacter johnsonii, and V. cincinnatiensis; and the class 1 integron and gene cassette aadA2 in V. cincinnatiensis. Approximately 6.6% of isolates contained class 1 integrons, and one isolate harbored class 2 integrons. Plasmid associated intI1 and androgen receptor- encoding genes were transferred into Escherichia coli J53 and E. coli DH5α by conjugation and transformation experiments, respectively. Our study highlights the importance of aerobic bacteria from pork as reservoirs for antimicrobial resistance genes and mobile genetic elements that can readily be transferred intra- and interspecies.

Acinetobacter spp. are opportunistic pathogens being A. baumannii the most frequently identified in nosocomial settings. A. ursingii was mainly described as causing bacteremia and outbreaks in neonatal intensive care units. Ten A. ursingii isolates were recovered from rectal swab screening for carbapenemase-producing bacteria between June 2013 and December 2015 from a children hospital in Argentina. All ten isolates were metallo-β-lactamase-producing, nine were positive for blaIMP-1 and one for blaNDM-1. IMP-positive isolates were also positive for blaOXA-58 gene. All isolates were susceptible to ciprofloxacin, colistin and minocycline, and nine were susceptible to ampicillin-sulbactam and gentamicin. Two A. ursingii displayed high level of resistance to aztreonam associated with blaCTX-M-15 in one isolate, and blaVEB-1 in the other. Eight SmaI-PFGE patterns were recognized. We evaluated the usefulness of Acinetobacter MLST-Pasteur scheme, to analyse A. ursingii isolates, however the rpoB gene was not amplified. A new set of primers were designed for specific amplification and sequencing, allowing the analysis of rpoB gene for this species. New alleles and the sequence types 748, 749, 750, 751, 993, 1186, 1187, and 1189 were included at the Acinetobacter MLST-Pasteur database. Those isolates showing related PFGE patterns were assigned to the same ST. To the best of our knowledge, this is the first report of MBL-producing A. ursingii in Argentina. The inclusion of A. ursingii species to the Acinetobacter MLST-Pasteur scheme allows deeper molecular characterization and a better understanding about the epidemiology of this germen. Fil: Faccone, Diego Francisco. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Dirección Nacional de Institutos de Investigación. Administración Nacional de Laboratorios e Institutos de Salud. Instituto Nacional de Enfermedades Infecciosas. Área de Antimicrobianos; Argentina Fil: Martino, Florencia. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Dirección Nacional de Institutos de Investigación. Administración Nacional de Laboratorios e Institutos de Salud. Instituto Nacional de Enfermedades Infecciosas. Área de Antimicrobianos; Argentina Fil: Pasteran, Fernando. Dirección Nacional de Institutos de Investigación. Administración Nacional de Laboratorios e Institutos de Salud. Instituto Nacional de Enfermedades Infecciosas. Área de Antimicrobianos; Argentina Fil: Albornoz, Ezequiel Pablo. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Dirección Nacional de Institutos de Investigación. Administración Nacional de Laboratorios e Institutos de Salud. Instituto Nacional de Enfermedades Infecciosas. Área de Antimicrobianos; Argentina Fil: Biondi, Estefania. Gobierno de la Ciudad de Buenos Aires. Hospital General de Niños "Ricardo Gutiérrez"; Argentina Fil: Vazquez, Miryam. Gobierno de la Ciudad de Buenos Aires. Hospital General de Niños "Ricardo Gutiérrez"; Argentina Fil: Rapoport, Melina. Dirección Nacional de Institutos de Investigación. Administración Nacional de Laboratorios e Institutos de Salud. Instituto Nacional de Enfermedades Infecciosas. Área de Antimicrobianos; Argentina Fil: Rodrigo, Veronica. Gobierno de la Ciudad de Buenos Aires. Hospital General de Niños "Ricardo Gutiérrez"; Argentina Fil: de Belder, Denise Gisele. Dirección Nacional de Institutos de Investigación. Administración Nacional de Laboratorios e Institutos de Salud. Instituto Nacional de Enfermedades Infecciosas. Área de Antimicrobianos; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Gómez, Sonia Alejandra. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Dirección Nacional de Institutos de Investigación. Administración Nacional de Laboratorios e Institutos de Salud. Instituto Nacional de Enfermedades Infecciosas. Área de Antimicrobianos; Argentina Fil: Corso, Alejandra. Dirección Nacional de Institutos de Investigación. Administración Nacional de Laboratorios e Institutos de Salud. Instituto Nacional de Enfermedades Infecciosas. Área de Antimicrobianos; Argentina

Acinetobacter ursingii M3 and Asticcacaulis excentricus M6 are plant growth-inhibiting bacteria that reduce the yield of the duckweed Lemna minor. We report here the complete genome sequences of A. ursingii M3 and A. excentricus M6, sequenced using the PacBio RS II platform.

In this paper, we report the data from whole-genome shotgun sequencing of an Acinetobacter ursingii isolate from healthy human skin of the forearm. The bacterial genome includes 3,473 genes and carries beta-lactamase resistance genes as well as resistance genes for several heavy metals.

International audience; INTRODUCTION: Surgical site infection (SSI) following orthopedic surgery can have a substantial impact on patients and families. The rate remains high, ranging from 0.5% to 8.5% in pediatric spine surgery. It is common to allow children to bring a teddy bear (or similar toy) to the surgical ward to help reduce the stress of surgery. We hypothesize that despite their known benefits for children, teddies would increase the bacterial load in the surgical room.METHODS: A blinded descriptive study was conducted from June 2015 to September 2016. The study included children entering the hospital through the emergency ward for a traumatic cause requiring surgery. Patients admitted for infectious problems and those who had been hospitalized less than 6 months before the inclusion date were excluded. A picture of the teddy was taken and stored in a blind fashion. The AFNOR (Association française de normalisation) standardized rules for bacteriological surface control and the ISO/DIS 14698 protocol were strictly followed. Two independent observers performed blind bacteriologic analyses of the teddy bears with bacteria identification and colony counts. Photos of the teddy bears were then analyzed by two blinded, independent observers: one doctor and one parent from outside the hospital. Cleanliness and fluffiness of the toy was evaluated using a numeric scale.RESULTS: Bacteria were identified on 100% of the 53 teddies included. The mean number of bacteria was 182.5±49.8 CFU/25 cm2. Eight teddies (15.1%) tested positive for potential pathogenic bacteria (two staphylococcus aureus, one acinetobacter ursingii, four acinetobacter baumannii, one pseudomonas stutzeri). Three teddies (5.7%) tested positive for fungi. The median cleanliness score was 2 (interquartile range (IQR)=1) if rated by the doctor and 2 (IQR=1) if rated by the parent. No statistical difference was found between these two values in the global teddy bear population. We found no any statistical link between the number of CFUs and the cleanliness scores given by the doctor. The median fluffiness score given by the parent was 2 (IQR=1). Looking at the correlative CFUs, we found a statistically significant difference between each stage of fluffiness with a higher stage showing higher CFU (P

Acinetobacter species can be important opportunistic pathogens in humans, especially in healthcare settings. We report here the first isolation of Acinetobacter ursingii from an animal species; it was isolated from a canine urinary tract infection, and phenotypic identification proved unreliable. Keywords: Acinetobacter ursingii, canine, molecular diagnostics, urinary tract infection, veterinary microbiology

The authors report a case of a premature male newborn admitted to the neonatal intensive care unit after an emergent caesarean due to maternal pre-eclampsia and foetal bradycardia at 32 weeks of gestational age and birth weight of 1440 g. There were no infection risk factors reported. On day 3 his clinical condition deteriorated, with tachycardia and subfebrile temperature and C-reactive protein at 1.25 mg/dL. Empirical antibiotics (flucloxacillin and gentamicin) were started, with no clinical improvement and C-reactive protein increasing to a maximum of 19 mg/dL (upper normal level of 1 mg/dL) after 3 days. Blood cultures from the third to the eighth day of life were positive for Acinetobacter ursingii. Targeted therapy was administered for 14 days with clinical and laboratorial improvement and he was discharged on the 28th day of life without any known sequelae. A. ursingii is emerging as an infectious agent of late-onset sepsis in immunosuppressed neonates.

Acinetobacter ursingii is an aerobic, gram-negative, opportunistic microorganism which is rarely isolated among Acinetobacter species. We present two immunocompetent infants who developed bacteremia due to A.ursingii. The first patient is a two -month- old boy who had been hospitalized in pediatric surgery unit for suspected tracheo-esophageal fistula because of recurrent aspiration pneumonia unresponsive to antibiotic therapy. The second patient is a fourteen -month- old boy with prolonged vomiting and diarrhea. A. ursingii was isolated from their blood cultures. They were successfully treated with ampicillin-sulbactam. Although A.ursingii has recently been isolated from a clinical specimen; reports of infection with A.ursingii in children are rare. A.ursingii should be kept in mind as an opportunistic microorganism in children.Pan African Medical Journal 2016; 23

Error-prone and error-free DNA damage repair responses that are induced in most bacteria after exposure to various chemicals, antibiotics or radiation sources were surveyed across the genus Acinetobacter. The error-prone SOS mutagenesis response occurs when DNA damage induces a cell’s umuDC- or dinP-encoded error-prone polymerases. The model strain Acinetobacter baylyi ADP1 possesses an unusual, regulatory umuD allele (umuDAb) with an extended 5′ region and only incomplete fragments of umuC. Diverse Acinetobacter species were investigated for the presence of umuDC and their ability to conduct UV-induced mutagenesis. Unlike ADP1, most Acinetobacter strains possessed multiple umuDC loci containing either umuDAb or a umuD allele resembling that of Escherichia coli. The nearly omnipresent umuDAb allele was the ancestral umuD in Acinetobacter, with horizontal gene transfer accounting for over half of the umuDC operons. Despite multiple umuD(Ab)C operons in many strains, only three species conducted UV-induced mutagenesis: Acinetobacter baumannii, Acinetobacter ursingii and Acinetobacter beijerinckii. The type of umuDC locus or mutagenesis phenotype a strain possessed was not correlated with its error-free response of survival after UV exposure, but similar diversity was apparent. The survival of 30 Acinetobacter strains after UV treatment ranged over five orders of magnitude, with the Acinetobacter calcoaceticus–A. baumannii (Acb) complex and haemolytic strains having lower survival than non-Acb or non-haemolytic strains. These observations demonstrate that a genus can possess a range of DNA damage response mechanisms, and suggest that DNA damage-induced mutation could be an important part of the evolution of the emerging pathogens A. baumannii and A. ursingii.

The prevalence of the currently known Acinetobacter species and related trends of antimicrobial resistance in a Dutch university hospital were studied. Between 1999 and 2006, Acinetobacter isolates from clinical samples were collected prospectively. Isolates were analyzed by amplified fragment length polymorphism fingerprinting. For species identification, a profile similarity cutoff level of 50% was used, and for strain identification, a cutoff level of 90% was used. Susceptibility for antimicrobial agents was tested by disk diffusion by following the CLSI guideline. The incidences of Acinetobacter isolates ranged from 1.7 to 3.7 per 10,000 patients per year, without a trend of increase, during the study years. Twenty different species were distinguished. Acinetobacter baumannii (27%) and Acinetobacter genomic species (gen. sp.) 3 (26%) were the most prevalent. Other species seen relatively frequently were Acinetobacter lwoffii (11%), Acinetobacter ursingii (4%), Acinetobacter johnsonii (4%), and Acinetobacter junii (3%). One large cluster of A. baumannii , involving 31 patients, and 16 smaller clusters of various species, involving in total 39 patients, with at most 5 patients in 1 cluster, occurred. Overall, 37% of the A. baumannii isolates were fully susceptible to the tested antibiotics. There was a borderline significant ( P = 0.059) trend of decreasing susceptibility. A. baumannii was the Acinetobacter species causing the largest burden of multiple-antibiotic resistance and transmissions in the hospital.

Acinetobacter species other than Acinetobacter baumannii have rarely been reported to be associated with nosocomial outbreaks of bloodstream infections. Within a period of 1 week, seven Acinetobacter -like isolates were recovered from peripheral blood and catheter specimens of five patients at a neonatal intensive care unit (NICU) in a tertiary hospital in Turkey. All five patients had placement of central venous catheters and had received total parenteral nutrition before the onset of bacteremia. Two of the five patients died. Medical devices, tap water, aerators, water samples, various surfaces, intravenous fluids, and the hands of health care workers in the NICU were sampled and were culture negative for the bacterium. All seven of the isolates had identical biochemical reactions, antimicrobial susceptibility results, and pulsed-field gel electrophoresis patterns, indicating a clonal nosocomial outbreak. A panel of standard biochemical reaction profiles and three phenotypic commercial identification systems failed to identify these isolates. Phenotypically, the isolate differed from Acinetobacter ursingii by its hemolysis on sheep blood agar and its negative citrate utilization. Sequences of the full 16S rRNA gene, which contained at least three different gene copies with polymorphic sequences between nucleotide positions 70 and 206, were determined from the first recovered isolate. The complete 1,529- to 1,531-bp 16S rRNA gene sequences and partial 801-bp rpoB gene sequences had similarities of 99.5% and 97.2%, respectively, to an A. ursingii isolate. The DNA-DNA similarities of the strain against the type strain of A. ursingii were 64.7 and 68.7%, which were lower than the recommended threshold value of 70% for the definition of bacterial species. These data indicate that a novel Acinetobacter organism caused the nosocomial outbreak of bacteremia in the NICU unit. We propose the designation of Acinetobacter septicus sp. nov. for these isolates, with isolate AK001 as the type strain.

We report a community-acquired bloodstream infection with Acinteobacter ursingii in an HIV-negative woman who injected drugs. The infection was successfully treated with meropenem. Species identification was performed by using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Improved identification of Acinetobacter spp. by using this method will help identify clinical effects of this underdiagnosed pathogen.

The genus Acinetobacter is amongst the most common causes of nosocomial bacterial infections. Even though Acinetobacter baumannii is the most frequent identified species causing a wide range of infections in humans, other species such as Acinetobacter pittii, Acinetobacter nosocomialis, Acinetobacter haemolyticus, Acinetobacter johnsonii, Acinetobacter lwoffii and Acinetobacter ursingii are sporadically observed as nosocomial pathogens. The four species that show similar phenotypes were grouped together as “Acinetobacter calcoaceticus – A. baumannii (ACB) complex”, i. e. A. baumannii, A. nosocomialis, A. pittii and less clinically important A. calcoaceticus. Owing to the high complexity within Acinetobacter genus, it is difficult to differentiate Acinetobacter into species level and these organisms are frequently misidentified. Acinetobacter spp. exhibit a great propensity to acquire antimicrobial resistance determinants and rapidly develop multidrug-resistant (MDR) phenotypes especially resistance to carbapenems, the last resort of antimicrobials to treat Acinetobacter infections. A high prevalence and endemic situations of carbapenem-resistant A. baumannii (CRAB) have been observed in multiple geographical areas such as Asia-Pacific and South America where predominant A. baumannii clonal lineages were noted. The international clone (IC) 2 is recognised as the most successful clone of A. baumannii causing outbreaks and persisting in hospital environments worldwide, particularly in Asia. Despite the increased amount of research on Acinetobacter epidemiology, potential virulence and evolution, little is known about the factors that may have contributed to the success of this well-known clone, IC2. The broad objectives of my PhD are to 1) develop a rapid method to assist in species identification of Acinetobacter non-baumannii, 2) determine the molecular epidemiology of A. baumannii, primarily from Thailand and 3) describe the genome of A. baumannii IC2 isolates from Thailand and compare these genome data with A. baumannii IC2 isolates from Japan, Malaysia and Singapore. Firstly, a multiplex PCR was developed to detect intrinsic oxacillinases encoding genes (blaOXAs), which assisted in rapid identification of multiple Acinetobacter species including A. lwoffii/Acinetobacter schindleri, A. johnsonii, A. calcoaceticus, A. haemolyticus and Acinetobacter bereziniae. Additionally, 30 novel blaOXAs variants were identified in this study. The investigation of Acinetobacter spp. collected worldwide revealed that acquired-type blaOXAs disseminated globally in Acinetobacter spp. as opposed to A. baumannii causing carbapenem resistance particularly in A. pittii. Carbapenem resistance was also observed in A. pittii from Australia and Thailand. Through the genomes of the two A. pittii strains ST119 and a novel ST655, several classes of antimicrobial resistance genes including a novel blaOXA-421, blaOXA-23, blaOXA-96, blaOXA-10, blaIMP-4, blaVEB-7, blaCRAB-2, floR, cmlA1, aar-2 and dfrA10 were identified. This emphasises the importance of A. pittii as an impending multidrug resistance pathogen in this region. Secondly, to explore an endemic situation of multidrug-resistant Acinetobacter spp., 300 non-repetitive ACB complex isolates, mainly A. baumannii from the largest tertiary hospital in Thailand were characterised for their molecular epidemiology and antimicrobial resistance mechanisms. Of these, 270 isolates were carbapenem-resistant and 92.2% resistant to amikacin. A. baumannii IC2 was the dominant clone of A. baumannii (80%) and blaOXA-23-like was detected in most of CRAB isolates. The genomes of 13 representative isolates of A. baumannii (n=11), A. nosocomialis (n=1) and A. pittii (n=1) were analysed via whole genome sequencing. Antimicrobial resistance island, AbaR4-type containing Tn2006, was found in all CRAB isolates. armA was the only 16S rRNA methylase gene found that caused resistance to amikacin (located within Tn1548) and in close proximity to macrolide resistance genes (mphE and msrE). csu locus, bap, bfmRS and pga locus associated with biofilm formation were found in all IC2 isolates in addition to the typical set of antimicrobial resistance genes; blaOXA-23, blaOXA-66, blaADC, strA, strB and tetB in this clone. Therefore, the presence of arrays of antimicrobial resistance genes and biofilm-related loci may contribute to the spread and persistence of A. baumannii IC2 in this hospital. Lastly, comparative genome analysis was performed on 21 representative Acinetobacter spp. isolates from Thailand and other countries, i.e. Japan, Malaysia and Singapore. The range of the CRAB genome size was 3.9 - 4.1 Mb with GC content approximately 39%. Interestingly, the susceptible A. baumannii genome size was 3.7-3.8 Mb. The size difference was due to the absence of resistance genes and regions for bacterial competition and biofilm formation in susceptible isolates. The genetic contexts of key antimicrobial resistance genes and resistance islands were investigated. Our study confirmed that the antimicrobial resistance genes and other genomic features of IC2 isolates were homogenous. The diversity was mainly found in the composition of the antimicrobial resistance genes of IC2 isolates and locus for capsule synthesis (K locus). The genome and composition of antimicrobial resistance genes between IC2 isolates from Thailand and all IC2 from other countries were closely related. Two IC2 carbapenem-susceptible isolates from Japan harboured less antimicrobial resistance genes (blaOXA-66, strA, strB, sul2 and tetB). The locus for the outer core of lipid A-core moiety (OC locus) was generally conserved - OCL1 was a common type within IC2. Such variations observed of the K locus may impact in the difficulties to generate human immune response to A. baumannii. Lastly, one key feature of IC2 was the integrity of all chromosomal regions for type VI secretion system (T6SS) and biofilm formation (csu). In contrast, these regions in non-IC2 isolates were disrupted. This may indicate the incapability of non-IC2 isolates to persist in the hospital environment. The study of various aspects of carbapenem-resistant Acinetobacter spp. in our region has revealed specific insights into this pathogen locally and globally. These include the species identification, molecular epidemiology and the genome of Acinetobacter spp. Genomic analysis has described all necessary attributes of the superiority of IC2 to spread further under antimicrobial pressure and harsh environments. Further research is required to understand greater detail of these many unique findings.

Many Gram-negative bacteria use N -acyl- l -homoserine lactones (AHLs) as quorum-sensing signal molecules. We have reported that Acinetobacter strains isolated from activated sludge have AHL-degrading activity. In this study, we cloned the amiE gene as an AHL-degradative gene from the genomic library of Acinetobacter sp. strain Ooi24. High-performance liquid chromatography analysis revealed that AmiE functions as an AHL acylase, which hydrolyzes the amide bond of AHL. AmiE showed a high level of degrading activity against AHLs with long acyl chains but no activity against AHLs with acyl chains shorter than eight carbons. AmiE showed homology with a member of the amidases (EC 3.5.1.4) but not with any known AHL acylase enzymes. An amino acid sequence of AmiE from Ooi24 showed greater than 99% identities with uncharacterized proteins from Acinetobacter ursingii CIP 107286 and Acinetobacter sp. strain CIP 102129, but it was not found in the draft or complete genome sequences of other Acinetobacter strains. The presence of transposase-like genes around the amiE genes of these three Acinetobacter strains suggests that amiE is transferred by a putative transposon. Furthermore, the expression of AmiE in Pseudomonas aeruginosa PAO1 reduced AHL accumulation and elastase activity, which were regulated by AHL-mediated quorum sensing.

Neonatal infections may be caused by various microorganisms, but as far as we are aware, Acinetobacter ursingii has not yet been reported in connection with nosocomial infections of premature infants. During 2 months, 3 premature babies were treated with nosocomial infection caused by A. ursingii at the same ward, and on the basis of molecular typing results the same strain was responsible for all of these cases. Traditional biochemical methods and automatic identification systems failed to identify this bacterium on the species level, and only 16S rDNA sequencing gave acceptable species identifications. The isolated strains proved to be susceptible to all of the tested antimicrobials, including ampicillin/sulbactam, doxycyclin, netilmicin, ciprofloxacin, piperacillin/tazobactam, ceftazidime, imipenem, meropenem, trimethoprim/sulfametoxazole, gentamicin, tobramycin, amikacin, and levofloxacin according to the CLSI standard. In spite of the environmental screening, the source of the infection could not be clarified. One of 3 neonates died, the others recovered and were discharged home after several months of hospitalization.

Objectives To assess the contamination risk with CF-relevant pathogens of the Tobi Podhaler dry powder inhaler (DPI) device compared to established nebulizers and to obtain "real-world" data from nebulizers of patients with chronic colonization of the lung with Gram-negative bacteria in the patient's usual inhalation setting, to learn more about patient's cleaning and inhalation habits. Methods Microbiological samples from the mouthpiece and medication reservoir were collected using an eSwab® from a device that was prepared in the patient's personal way before next inhalation. Results Microbiological analysis showed that the samples of 8/26 dry powder devices were sterile while the other 16 samples were contaminated with non-pathogenic bacteria of the human respiratory flora. Only 1/20 nebulizer devices were sterile. In 4 devices Staphylococcus aureus was detected. The other nebulizers were contaminated with a variety of bacterial species belonging to the human respiratory tract flora and bacteria typically found in humid sources (e.g. Pseudomonas fluorescens, Acinetobacter ursingii, Mycobacterium fortuitum ), indicating the potential transmission risk of CF-relevant species such as P. aeruginosa . Conclusion Tobi Podhaler is used 7 days twice daily. For cleaning it is supposed to be wiped with a dry and clean cloth. The cleaning procedure of a nebulizer is more time consuming and challenging. Under "real-world" conditions we found that the typical cleaning procedure of a nebulizer is failing to sterilize the device completely. Using a DPI with almost zero cleaning effort seems to be a satisfactory alternative regarding the hygienic requirements of an inhalation device.

The species belonging to the Acinetobacter genus are currently reported as opportunistic pathogens in hospitalized patients with underlying predispositions. However, except for the Acinetobacter calcoaceticus-Acinetobacter baumannii complex, the identification of other species is frequently unreliable, especially for Acinetobacter ursingii and Acinetobacter schindleri , newly described in 2001. Thus, the clinical significance, phenotypic features, and antimicrobial susceptibilities of these two misidentified species remain unclear. Of 456 Acinetobacter sp. clinical strains isolated from 2002 to 2005 in Henri Mondor Hospital, 15 isolates (10 A. ursingii and 5 A. schindleri isolates) were studied. They were characterized using a phenotypic approach (API 20 NE and VITEK 2 systems), 16S rRNA gene sequencing, and susceptibility to antimicrobial agents with evaluation of impact in clinical relevance. The two corresponding type strains were also included for comparison. All isolates were identified to the species level using molecular tools, whereas the phenotypic methods remained unreliable due to the absence of these two species in the manufacturers' databases. However, the API 20 NE system appeared to be a reasonably reliable phenotypic alternative for the identification of A. ursingii when the numerical code 0000071 was found. Conversely, no discriminative phenotypic alternative existed for A. schindleri isolates. Concerning antimicrobial susceptibility, A. ursingii strains appeared to be more resistant to antibiotics than A. schindleri strains, which could imply therapeutic consequences. Finally, the prevalence of infections caused by A. ursingii and A. schindleri (representing 9.7% and 4.8% of non- A. calcoaceticus-A. baumannii complex strains, respectively) seems to be underestimated.

The taxonomic status of two recently described phenetically distinctive groups within the genus Acinetobacter, designated phenon 1 and phenon 2, was investigated further. The study collection included 51 strains, mainly of clinical origin, from different European countries with properties of either phenon 1 (29 strains) or phenon 2 (22 strains). DNA-DNA hybridization studies and DNA polymorphism analysis by AFLP revealed that these phenons represented two new genomic species. Furthermore, 16S rRNA gene sequence analysis of three representatives of each phenon showed that they formed two distinct lineages within the genus Acinetobacter. The two phenons could be distinguished from each other and from all hitherto-described Acinetobacter (genomic) species by specific phenotypic features and amplified rDNA restriction analysis patterns. The names Acinetobacter ursingii sp. nov. (type strain LUH 3792T = NIPH 137T = LMG 19575T = CNCTC 6735T) and Acinetobacter schindleri sp. nov. (type strain LUH 5832T = NIPH 1034T = LMG 19576T = CNCTC 6736T) are proposed for phenon 1 and phenon 2, respectively. Clinical and epidemiological data indicate that A. ursingii has the capacity to cause bloodstream infections in hospitalized patients.

We describe what we believe to be the first case of biliary sepsis caused by Acinetobacter ursingii. The patient was a healthy woman with no comorbidities who presented with choledocholithiasis and cholangitis. The performance of an endoscopic cholangiopancreatography was the trigger for A. ursingii bacteraemia. This report highlights the inadequacies of conventional phenotypic tests usually available in clinical microbiology laboratories for the identification of Acinetobacter species.

In their recent paper, Kilic et al. (2) reported on a hospital outbreak of bloodstream infections caused by a non-glucose-acidifying Acinetobacter strain. The authors showed that this strain (AK001) was phenotypically most similar to Acinetobacter ursingii but had two properties atypical of this species: i.e., the ability to hemolyse sheep blood and the inability to grow on citrate. Additional taxonomic methods to clarify the taxonomic status of AK001 included 16S rRNA and rpoB gene sequence analysis and DNA-DNA reassociation. Although the results of the sequence analyses supported the close relationship of the strain to A. ursingii, the value of DNA-DNA reassociation between AK001 and the A. ursingii type strain was found to be 64.7 to 68.7%, which is slightly less than 70%, the recommended threshold for species delineation (6). The authors concluded that the strain represents a new species, for which the name “Acinetobacter septicus” was proposed. As, in our opinion, the data provided by Kilic et al. (2) do not convincingly substantiate the proposal of a novel species, we analyzed strain AK001 using methods that had previously been used to resolve Acinetobacter taxonomy (3). AK001 was obtained by the Culture Collection, University of Goteborg (CCUG) from Y.-W. Tang and was designated CCUG 56015. The strain was shown to have properties consistent with the original description of A. ursingii, except for the inability to utilize citrate (4). It is of note, however, that there are other citrate-negative A. ursingii strains in our current collection, which indicates the intraspecies variation of this property. Importantly, the hemolytic activity of AK001 reported by Kilic et al. (2) could not be confirmed, although sheep blood agar plates obtained from different suppliers were used at different cultivation temperatures and with several control hemolytic Acinetobacter strains. In the study by Kilic et al. (2), the 16S rRNA gene sequences of AK001 were 99.5% similar to that of the A. ursingii type strain, which is the value that can be found in strains within one species (4). To obtain more comprehensive information on the rpoB-based position of AK001 within the genus Acinetobacter, we determined the rpoB sequence of AK001 and compared it to that of eight A. ursingii strains and all hitherto-described species. The similarity values between AK001 (GenBank no. {"type":"entrez-nucleotide","attrs":{"text":"EU742582","term_id":"190688713","term_text":"EU742582"}}EU742582) and the A. ursingii strains ({"type":"entrez-nucleotide","attrs":{"text":"EU477105","term_id":"526150767","term_text":"EU477105"}}EU477105 and {"type":"entrez-nucleotide-range","attrs":{"text":"EU742575 to EU742581","start_term":"EU742575","end_term":"EU742581","start_term_id":"190688699","end_term_id":"190688711"}}EU742575 to EU742581) ranged from 96.9% to 98.0%, which partially overlaps with the range of intraspecies values found for Acinetobacter beijerinckii (3). In contrast, the rpoB similarity values between AK001 and the other species were between 78.8% and 84.5%. The proposal of “A. septicus” was based mainly on the results of DNA-DNA reassociation. However, DNA-DNA reassociation values close to the proposed value of 70% have to be treated with caution as incongruent results for the same strains can be obtained in different laboratories and/or by different methods, which may have taxonomic implications (5). Concerning also other limitations of DNA-DNA reassociation (1), no conclusions based on these intermediate values should be made without further evidence of genetic isolation between strains. To address this problem, the relatedness of AK001 to hitherto-described species was investigated by AFLP (Keygene NV) analysis, a whole-genome fingerprinting method. Previous studies using this method have shown that strains of the same (genomic) species cluster together at a ≥50% similarity level (4). AFLP analysis showed that AK001 and 25 reference A. ursingii strains were linked in a distinct cluster at a 59% similarity level, while AK001 was separated from other species at 34% similarity, which strongly supports the assignment of AK001 to A. ursingii. Based on the above arguments, strain AK2001 is considered as belonging to A. ursingii and not representing a novel species.

Acinetobacter ursingii has not been reported in infectious processes apart from its recent description as a new species. A bacteremia caused by A. ursingii in a patient with a pulmonary adenocarcinoma confirms that this microorganism is an opportunistic human pathogen. The isolate was susceptible to imipenem, aminoglycosides, rifampin, and fluoroquinolones.

Acinetobacter species can be important opportunistic pathogens in humans, especially in healthcare settings. We report here the first isolation of Acinetobacter ursingii from an animal species; it was isolated from a canine urinary tract infection, and phenotypic identification proved unreliable.