Proteus Mirabilis Isolates Research Articles

Prevalence of ESBL/AmpC genes and specific clones among the third-generation cephalosporin-resistant Enterobacteriaceae from canine and feline clinical specimens in Japan

In recent years, besides the widespread occurrence of extended-spectrum β-lactamase (ESBL)- and/or plasmid-mediated AmpC (pAmpC)-producing Enterobacteriaceae in both healthcare and community settings of humans, the third-generation cephalosporin (3GC)-resistant microbes have also been reported from companion animals worldwide. Here, we characterized ESBL- and/or pAmpC-producing Enterobacteriaceae clinical isolates from companion animals. Among the 487 clinical isolates mainly from urine of dogs and cats between May and September 2016, 104 non-repetitive isolates were resistant to the 3GC, and they consisted of 81 of 381 (21.3%) Escherichia coli, 21 of 50 (42.0%) Klebsiella pneumoniae, and 2 of 56 (3.6%) Proteus mirabilis isolates. In the 81 E. coli, the predominant bla genes were blaCTX-M-27 and blaCMY-2 (n = 15 each), followed by blaCTX-M-15 (n = 14), blaCTX-M-14 (n = 10), and blaCTX-M-55 (n = 5). In 21 K. pneumoniae, 10 bla gene types including blaCTX-M-15 (n = 4), blaCTX-M-2 (n = 4), and blaCTX-M-14 (n = 3) were found. The blaCTX-M-2 was identified in 2 P. mirabilis. Twenty-four of the 42 E. coli belonging to phylogroup B2 were O25b-ST131 clone, mostly associated with uropathogenic E. coli pathotype, and 22 isolates of this clone were identified as specific H30R subclone. High prevalence of the blaCTX-M-27-harboring isolates were noted among the H30R/non-Rx lineage (13/19, 68.4%) (p < 0.05). The genetic environment of blaCTX-M-27 of most isolates of this lineage was identical to that of human isolates, but unique flanking genetic structures were also identified. Newly emerging virulent lineage B2-non-O25b-ST1193 was also confirmed in 5 isolates. The fosA3 and/or armA genes were detected in E. coli and K. pneumoniae isolates. These data suggest that companion animals serve as a potential reservoir of antimicrobial resistant E. coli and K. pneumoniae. This also has considerable veterinary importance, since urinary tract infections are an important disease causing therapeutic challenges worldwide.

Detection of ESBL genes from ciprofloxacin resistant Gram negative bacteria isolated from urinary tract infections (UTIs)

Urinary tract infections (UTIs) are the most disregarded diseases in both developing and developed countries and accountable for one fourth of the health care related infections. Even though the women and children are the most susceptible reservoir, the infections are at a significant amount in men of all ages. In UTI, the multidrug resistant bacteria (MDRB) is an emerging burden and now represents a daily challenge for the management of antimicrobial therapy in healthcare settings. A total of 100 uropathogens of Gram negative bacteria (GNB) were used in the current study to detect the ciprofloxacin resistant effect due to extended spectrum beta lactameses production (ESBLs). The phenotypic identification of ESBLs producing uropathogens from UTIs were detected by primary ESBL identification test (PMIT) and double disc combination method (DDCM). In addition, the ESBL production was further confirmed by MIC stripe method. Further, the prevalence of TEM, SHV, OXA and CTX-M type genes of isolated GNB and ciprofloxacin resistant strains was detected by multiplex PCR method. Among the 100 uropathogens, 84% was found to produce ESBLs. Out of 84, 60 strains were identified as ciprofloxacin resistant by Hexa discs and they developed resistance against all antibiotics. The PMIT and DDCM proved the result including Escherichia coli (26), 21 isolates of Proteusmirabilis, 17 of Pseudomonas aeruginosa, 14 of Klebsiellapneumoniae and 6 of Acinetobacter sp. In particular, the high number of CTX-M and TEM genes were frequently detected from collected uropathogens and all the TEM, SHV, OXA, and CTX-M genes were identified from ciprofloxacin resistant strains only. Due to the increase of multiple ESBL genes in uropathogens, sustained supervision for using favorable antibiotics and the decreasing the infection is essential.

Antimicrobial Properties, Phytochemical Composition, and Phenotypic Resistance Pattern of Selected Enteropathogenic Microorganism on Ageratum conyzoides (Goat Weed) Leaf Extract

The aim of this study is to determine the phytochemical composition, antimicrobial properties and phenotypic resistance pattern of selected enteropathogenic microorganism on Ageratum conyzoides leaf extract. Enteropathogenic isolates were collected from Adekunle Ajasin University Health Centre, Akungba Akoko, Ondo State and were identified using Microbact™ 24E Identification kit. Antibiotic sensitivity testing was carried out against the enteropathogenic organism using Kirby-Bauer disc diffusion method, imipenem was active against Proteus mirabilis, Enterobacter agglomerans, Enterobacter gerogoviae, with diameter zone of inhibition of 41 mm, 26 mm and 30 mm. ofloxacin were active against Proteus mirabilis, Enterobacter agglomerans, Enterobacter gergoviae, with diameter zone of inhibition of 21, 31 and 21 mm. All enteropathogenic organisms shows high resistant rate against oxacilin, vancomycin, amoxicillin, ceftazidime, piperacillin, cefoxitin, tazobactam. Antimicrobial screening of the leaf extract of Ageratum conyzoides revealed high bioactivity against Serratia marcescens, Proteus vulgaris, Enterobacter agglomerans and Proteus mirabilis isolates with diameter zone of inhibition ranging from 15 mm to 22 mm at 100 mg/ml. Using broth dilution method, the minimum inhibitory concentrations (MIC) of the Ageratum conyzoides (Goatweed) extract range from 25 mg/ml and 100 mg/ml. The phytochemical screening results shows that Ageratum conyzoides extract contained alkaloid, pholobotannins, Cardiac glycoside, phenol, tannins, saponin, and flavonoids. It can be deduced from this research work that Ageratum conyzoides has better antimicrobial properties and it contains very important phytochemical constituents which can be a magic bullet to deal with the menace of antimicrobial resistance enteropathogenic microorganism.

Antimicrobial Resistance Patterns of Proteus mirabilis isolates from Urinary tract, burn wound and Diabetic foot Infections

Proteus mirabilis is a common etiologic agent of urinary tract, burn wound and diabetic foot infections. Resistance to Proteus mirabilis is also common and represents a challenge to antibiotic therapy. This study aimed to investigate the antibiotic resistance of Proteus mirabilis isolated from three sources; urinary tract infections, burn wound infections in addition to diabetic foot infections. Forty-five clinical isolates of Proteus mirabilis (15 from each source) were used in this study. Complete resistance was found with each of ampicillin and tetracycline. High resistance was exhibited with cefepime. The resistance was intermediate against ceftazidime, cefotaxime, sulfamethoxazole-trimethoprim, amoxicillin-clavulinic, chloramphenicol, cefoperazone, aztreonam and ampicillin-sulbactam. Low resistance was found with piperacillin. These low resistance rates were also shown against tested aminoglycosides and fluoroquinolones. Very little resistance was found to imipenem, while no resistance was exhibited against piperacillin- tazobactam. The resistance pattern showed variation among different sources. Generally, burn wound isolates showed the highest resistance rates followed by diabetic foot isolates, while urinary tract isolates were the least resistant. High resistance was found with cefepime only in isolates from urinary tract infections and no diabetic foot isolate was highly resistant to any of the tested antibiotics. However, such resistance was observed with amoxicillin-clavulinic acid, cefepime, ceftazidime, cefotaxime and sulphamethoxazole-trimethoprim in burn wound isolates. Multidrug resistance (MDR) was also found with varying rates in isolates from different sources. MDR was more common in burn wound isolates than in diabetic foot isolates or urinary tract isolates. This study suggests that there is a variation in antibiotic resistance of Proteus mirabilis among different sources and alarms against high resistance especially in burn wound isolates that requires a strict policy in antibiotic dispensing to minimize such tesistance.

Detection of Virulence Factors of Local Isolates of Proteus mirabilis

Detection of Virulence Factors of Local Isolates of Proteus mirabilis

The Antibacterial Properties of Aqueous and Alcoholic Extracts of Punica granatum Seeds on Infectious Diarrhea Produced by Bacteria

Diarrhea is one of the gastrointestinal diseases that some reasons associated with an inappropriate of antibiotics treatment and, increased antibiotic resistance, has led to problems in the health system which vary in different societies. Control and resolution of this problem is necessary and the acquisition of a new group of antimicrobial compounds has been mentioned. So, this study aimed to determine the antibacterial activity of aqueous and alcoholic extracts of pomegranate seeds ( Punica granatum ) on common gastrointestinal producing bacteria. The antibacterial property of aqueous and alcoholic extracts of Punica granatum seeds against 100 bacteria was considered. Fifty and three isolates of E. coli , 19 isolates of Shigella sonneii , 8 isolates of Shigella flexneri , 5 isolates of Shigella dysentery , 7 strains of Proteus vulgaris , 5 isolates of Proteus mirabilis and 3 isolates of Citrobacter fraundii gathered from patients with diarrhea during July 2015-August 2016. Minimum inhibitory concentration (MIC) of various concentrations (mg/ml) of the extracts was used as an index of antimicrobial activity. The minimum inhibitory concentration of aqueous extract vs. ethanolic based on mg / ml were following: E. coli (75 vs 37.5), Shigella sonnei (37.5 vs., 18.75), Shigella flexneri (18.75 vs., 9.37), Shigella dysentery (18.75 vs., 9.37), Proteus vulgaris (18.75 vs., 9.37), Proteus mirabilis (9.37 vs., 9.37) and Citrobacter fraundii (150 vs., 75), respectively. The aqueous and alcoholic extracts had inhibitory effects on the studied bacteria, which affects the concentration of the substances used. Maybe, these inhibitory effects against each bacterium depend on effective substances and the type of the solvent properties. It is suggested to extract new antimicrobial compounds by chromatography to gain new antimicrobial agents.

Uropathogens and Antibiotic Resistance Among Nursing Home Residents - National Healthcare Safety Network (NHSN)

BackgroundKnowledge of urinary tract infection (UTI) pathogen and susceptibility patterns is necessary to inform antibiotic prescribing and monitor resistance. We describe bacterial pathogens and UTI antibiotic resistance patterns among residents in nursing homes (NHs) reporting to the National Healthcare Safety Network (NHSN) long-term care facility (LTCF) component.MethodsAll UTI events from January 1, 2013 to December 31, 2016 were included; up to three organisms per UTI event may be reported. Pathogen susceptibility results for selected antibiotics are reported as: Susceptible (S), Intermediate (I), Resistant (R), or Not tested (N). For this analysis, resistance was defined as I or R. We described pathogens and summarized antibiotic resistance only when ≥100 isolates of a bacterial species had susceptibility test results for a particular antibiotic reported to NHSN.ResultsIn 166 NHs located in 37 states, a total of 4,054 pathogens were reported for 2,827 residents. Six organisms accounted for 81% of all UTI events (n = 3,599) (Table). A large proportion of Escherichia coli isolates, which accounted for 41% of uropathogens, were resistant to trimethoprim-sulfamethoxazole (35%) and levofloxacin (50%). Among Proteus mirabilis isolates, 53% were resistant to levofloxacin (Figure). Methicillin resistance was 74% among Staphylococcus aureus, and vancomycin resistance among Enterococcus spp. was 18%.ConclusionThis is the first summary of UTI pathogens and susceptibility data from U.S. nursing homes reporting to a national surveillance system. Resistance to antibiotics commonly used to treat UTIs was high. Tracking and preventing resistance for key pathogens is a CDC priority and NHSN reporting by NHs provides a crucial opportunity to track antibiotic resistance, highlighting the importance of enrolling more NHs into NHSN.Table: Top Nursing Home Uropathogens - NHSN, January 2013–December 2016 Pathogen N % Escherichia coli 1,64841 Proteus mirabilis 52413 Enterococcus spp. 44111 Klebsiella pneumoniae 41210 Pseudomonas aeruginosa 2566 Staphylococcus aureus 1223Note: Enterococcus spp. includes E. faecium and E. fecalis.Disclosures All authors: No reported disclosures.

Identification and Susceptibility Testing of Non-lactose Fermenting Gram-Negative Bacilli in Urinary Tract Infection

Background: Urinary tract infections are one of the most common bacterial infections in humans both in community and hospital settings. The etiology of UTIs and the antimicrobial susceptibility of urinary pathogens have been changing over the years. Objectives: This work aimed to compare the accuracies of API 20E, Vitek1 system with GNI card with conventional reference biochemical tests for the identification of non-lactose fermenting Gram-negative bacilli in patients with urinary tract infection. Methodology: This study was conducted on 320 patients attending outpatient urology clinic. Physical, chemical, and microscopic examinations were done first to urine samples. Then, urine samples with pus cells <10 per high power field (HPF) or with bacterial cells or yeast were chosen for culture. These samples were 142 out 320 urine samples. These urine samples were isolated on nutrient, blood & MacConkey's agar media. Results: Standard bacteriological identification was done for all isolates (187 isolates) by colony morphology and microscopic examination. Only non-lactose fermenting Gram-negative bacilli (60 isolates) were further examined. Other growths were excluded. Motility test, biochemical reactions by conventional methods, API 20E, and vitek1 system with GNI card were done on non-lactose fermenting Gram-negative bacilli. Antibiotic susceptibility test by reference broth microdilution MIC procedure and Vitek1 system with GNI-108 card were done on non-lactose fermenting Gram-negative bacilli. One hundred thirty-five strains isolated from 94 hospitalized catheterized patients and fifty-two strains isolated from 48 outpatients sixty isolates (32.1%) were non-lactose fermenting Gram-negative bacilli. Out of them (thirty-one (16.5%) Pseudomonas aeruginosa, twenty (10.7%) Proteus mirabilis, six (3.2%) Proteus vulgaris, and three (1.6%) Providencia stuartii). After completion of supplemental tests, there was no difference between API 20E and vitek1 system in their ability to correctly identify Proteus mirabilis, Proteus vulgaris, and Providencia stuartii to the species level with there was difference between identifying isolates from Pseudomonas aeruginosa with vitek1 system. Conclusion: API 20E needs more time than vitek1 for identification of Proteus mirabilis, Proteus vulgaris, Providencia stuartii, and Pseudomonas aeruginosa, the most effective antibiotic against Pseudomonas aeruginosa isolates was ticarcillin/clavulant (77.4%), for Proteus mirabilis isolates were aztreonam and imipenem (each with 100%), for Proteus vulgaris isolates was imipenem (100%), and for Providencia stuartii isolates were amikacin, aztreonam, ciprofloxacin, imipenem, and TMP/SMX (100%).

Antimicrobial susceptibility profile in urinary pathogens causing community-acquired infections in diabetic patients in Colombia

Urinary tract infection is the most common pathology in diabetic patients, and an important determinant of morbidity and mortality among them. The increasing resistance of uropathogens acquired in the community to commonly used antibiotics is alarming. To identify the profile of antibiotic susceptibility of uropathogens responsible for communityacquired infections among diabetic patients in hospitals in Colombia. We conducted a descriptive study in a subgroup of diabetic patients in the framework of a larger study in adults with urinary tract infection acquired in the community. Over one year, we collected Escherichia coli, Klebsiella spp. and Proteus mirabilis isolates from nine hospitals in Colombia. Their susceptibility profile was determined using microbiological and molecular methods to establish the presence of extended-spectrum AmpC betalactamases and KPC carbapenemases. We collected 68 isolates (58 E. coli, nine Klebsiella spp. and one Proteus mirabilis). Four (6.9%) of the E. coli isolates expressed extended spectrum betalactamases, two (3.4%) of them belonged to the phylogenetic group B2 and to ST131 clone and expressed the TEM-1 and CTM-X-15 betalactamases. The AmpC phenotype was found in four (6.9%) of the E. coli isolates, three of which produced TEM-1 and CMY-2 betalactamases. One K. pneumoniae isolate expressed the KPC-3 carbapenemase. The presence of extended spectrum betalactamases and carbapenemases in uropathogens responsible for community-acquired infection was confirmed in diabetic patients.

Phenotypic detection of extended –spectrum betalactamase production in Proteus. mirabilis isolation from Patients with Significant Bacteriuria in Najaf provina

250 mid-stream urine samples were collected from patients suspected of urinary tract infection (UTI) who were attended to Three hospitals (Medical Al-Sader City, Al-Hakeem, and Al-Zahra Maternity and Children) in Najaf provina during the period of October 2011 to January 2012 and screened for the presence of Proteus mirablis .The growth of ≥105 colony forming units/ml was considered as significant bacteriuria.&#x0D; A total of 198 (79.2%) significant bacteriuria were detected. The study showed higher incidence of UTI in females 129 (65.2%)than males 69 (34.8% )However , 52 (26.3%) isolates of Proteus mirabilis , 146 (73.7%) isolates other species pathogens. The 52 isolates of Proteus mirabilis were tested for their antibiotic resistance against 11 antibiotics by the Kirby-Bauer disk diffusion test. All the isolates were found to be resistant to at least 5 antibiotics to which they were subjected. Therefore, all these isolates were considered to be multidrug resistant. The majority 39 (75.0%) isolates of Proteus mirabilis were able to produce β-lactamase enzymes with rapid iodometric method. All isolates of Proteus mirabilis were tested for their ability to produce Extended-spectrum β-lactamase . The result showed that 18 (34.6%) isolates were able to produce ESBLs.

Genetic characteristic of class 1 integrons in proteus mirabilis isolates from urine samples.

Background: Proteus mirabilis is an opportunistic pathogen, commonly associated with complicated urinary tract infections (UTIs). UTIs caused by multidrug-resistant Proteus mirabilis have increased worldwide. Multidrug-resistance of Gram-negative enteric bacteria is usually associated with class 1 integrons.Purposes: To investigate the prevalence and characterize gene cassettes of class 1 integrons in multidrug-resistant P. mirabilis Methods: From 2006 to 2008, 314 P. mirabilis isolates from urine were collected from a regional teaching hospital. Antimicrobial resistance of the isolates was determined by disk diffusion methods. The phenotypic confirmatory test of extended-spectrum β-lactamase (ESBL) production was performed as described in the Clinical and Laboratory Standards Institute (CLSI) guideline. The genetic organization of the class 1 integron cassettes was investigated by PCR, cloning, and sequencing of the regions surrounding these genes.Results: Seventy-nine (25%, 79/314) P. mirabilis isolates were ESBL-producing and most ESBL-producing P. mirabilis were positive for blaCTX-M. Class 1 integrons were presented in 76 isolates (24.2%, 76/314), and were more frequently found in ESBL-positive (55/79, 70%) than ESBL-negative (21/235, 8.9%) P. mirabilis isolates. The most prevalence of the cassettes encoded resistance genes were aminoglycoside (aac(6’)-Ib, aacA7, aadAl, aadA2, and aadAla), trimethoprim (dfrAl and dfrA12) and chloramphenicol (catB3 and cmlA6). The most prevalent cassette of dfr12-orfF-aadA2 was found in 49 isolates. The cassette array aadB-catB3-oxa10-aadA1 was first found in P. mirabilis. The enterobacterial repetitive intergenic consensus (ERIC)-PCR fingerprinting patterns were detected in these 76 integron positive P. mirabilis isolates and belonged to 8 profiles.Conclusion: This study investigated the prevalence and characterized gene cassettes of class 1 integrons in MDR P. mirabilis isolates from urine samples. The frequency of gene cassettes in P. mirabilis were partially by clonal spread of the carriers and the results could provide information for effective antimicrobial therapy and infection control.

First report in Africa of two clinical isolates of Proteus mirabilis carrying Salmonella genomic island (SGI1) variants, SGI1-PmABB and SGI1-W

Two Proteus mirabilis strains, designated PmTAN59 and PmKAF126, were isolated from two different Egyptian cities in 2014 and 2015, respectively. PmTAN59 was isolated from a sputum swab from a pneumonia patient in Tanta University Teaching Hospital. PmKAF126 was isolated from a patient with a diabetic foot infection in a hospital in the city of Kafr El-Sheikh. The two isolates were identified with bacterial small ribosomal RNA (16S rRNA) gene amplification and sequencing and tested for antimicrobial sensitivity with a Kirby–Bauer disk diffusion assay. The two strains were resistant to amoxicillin/clavulante, ampicillin, cefotaxime, cefoxitin, ceftriaxone, chloramphenicol, ciprofloxacin, colistin, gentamicin, kanamycin, nalidixic acid, spectinomycin, streptomycin, sulfamethoxazole/trimethoprime, and tetracycline, but sensitive to aztreonam, imipenem, and meropenem. Molecular characterization was used to map the entire backbone, including the multiple antibiotic resistance (MDR) region, of Salmonella genomic island 1 (SGI1). Both isolates carried a structure similar to SGI1, with two different MDR regions corresponding to SGI1-PmABB in PmTAN59 and SGI1-W in PmKAF126. SGI1-PmABB carried an integron of ~1.5kb with a two-gene cassette, aacCA5–aadA7, which confers resistance to gentamicin, streptomycin, and spectinomycin, whereas SGI1-W carried an integron of ~1.9kb containing aadA2–lnuF, which confers resistance to spectinomycin, streptomycin, and lincosamides. PmKAF126 carried the entire SGI1 sequence, however PmTAN59 carried a SGI1 structure with a deletion in the region from ORF S005 to ORF S009 and accompanied by insertion of IS1359 (1258bp). Furthermore, PmTAN59 carried class 2 integron of ~2.2kb containing dfrA1-sat2-aadA1. An ERIC-PCR analysis detected no clonal relationship between the two strains. Molecular screening for other antimicrobial resistance genes and a plasmid analysis indicated that PmTAN59 carried an IncFIB plasmid type. This strain also carried blaTEM-1 and the plasmid-mediated quinolone-resistance gene qnrA1. However, PmKAF126 carried no plasmids and no resistance gene other than that contained in the MDR region of SGI1 and floR gene conferring resistance to florfenicol. To the best of our knowledge, this is the first report of an SGI1-positive P. mirabilis strain in Egypt or on the entire African continent.

Chromosomal location of the fosA3 and blaCTX-M genes in Proteus mirabilis and clonal spread of Escherichia coli ST117 carrying fosA3-positive IncHI2/ST3 or F2:A-:B- plasmids in a chicken farm

The aim of this study was to investigate the spread and location of the fosA3 gene among Enterobacteriaceae from diseased broiler chickens. Twenty-nine Escherichia coli and seven Proteus mirabilis isolates recovered from one chicken farm were screened for the presence of plasmid-mediated fosfomycin resistance genes by PCR. The clonal relatedness of fosA3-positive isolates, the transferability and location of fosA3, and the genetic context of the fosA3 gene were determined. Seven P. mirabilis isolates with three different pulsed-field gel electrophoresis (PFGE) patterns and five E. coli isolates belonging to sequence type 117 (ST117) and phylogenetic group D were positive for fosA3 and all carried the blaCTX-M gene. In E. coli, the genetic structures IS26–ISEcp1–blaCTX-M-65–IS26–fosA3–1758 bp–IS26 and IS26–ISEcp1–blaCTX-M-3–blaTEM-1–IS26–fosA3–1758 bp–IS26 were present on transferable IncHI2/ST3 and F2:A-:B- plasmids, respectively. However, fosA3 was located on the chromosome of the seven P. mirabilis isolates. IS26–ISEcp1-blaCTX-M-65–IS26–fosA3–1758 bp–IS26 and IS26–blaCTX-M-14–611 bp–fosA3–1222 bp–IS26 were detected in three and four P. mirabilis isolates, respectively. Minicircles that contained both fosA3 and blaCTX-M-65 were shared between E. coli and P. mirabilis. This is the first report of the fosA3 gene integrated into the chromosome of P. mirabilis isolates with the blaCTX-M gene. The emergence and clonal spread of avian pathogenic E. coli ST117 with the feature of multidrug resistance and high virulence are a serious problem.

Dissemination of blaOXA-58 in Proteus mirabilis isolates from Germany.

Characterization of Proteus mirabilis isolates harbouring bla OXA-58 with emphasis on the genetic environment of this resistance determinant. Strains of P. mirabilis ( n = 37) isolated from different patients were tested for the presence of bla OXA-58 . The genetic context of bla OXA-58 was determined by WGS of two strains and Sanger sequencing. Clonality of the strains was assessed by PFGE. Susceptibility testing was performed by microdilution according to EUCAST. Four strains isolated in different geographical regions of Germany were positive for bla OXA-58 , and WGS showed that this resistance gene was harboured on a plasmid. Sanger sequencing confirmed the presence of two nearly identical plasmids, 6219 and 6208 bp in size, in all four strains. Upstream of bla OXA-58 an IS Aba 3-like transposase gene was located. The P. mirabilis strains were not clonally related according to PFGE. MICs of meropenem for three of the strains were only just above the EUCAST breakpoint and the Carba NP test was positive for only two of the strains. To our knowledge, this is the first description of bla OXA-58 in the species P. mirabilis . The resistance gene is harboured by almost identical plasmids in strains not clonally related and from different geographical regions. Apart from an IS Aba 3-like transposase gene upstream of bla OXA-58 the genetic context is different from bla OXA-58 harboured on plasmids in the genus Acinetobacter . With MICs of meropenem well below the EUCAST breakpoint or only just above it and equivocal or false negative results from the Carba NP test, bla OXA-58 can be easily overlooked in P. mirabilis .

Hatchery workers' IgG antibody profiles to airborne bacteria.

Occupational exposure to high concentrations of airborne bacteria in poultry production is related to an increased risk of respiratory disorders. However, etiology and in particular microorganisms' potential role in pathogenesis still needs to be elucidated. Thus, detection of specific antibodies against occupational microbial antigens may lead to identification of potentially harmful species. For the purpose of IgG titer determination, indirect immunofluorescence on various bacterial isolates from duck hatchery air was combined with image-based quantification of fluorescence intensity. Moreover, in addition to established assays with pure bacterial cultures, a new approach utilized complex bioaerosol samples for detection of anti-microbial antibodies in human sera by determination of percentages of antibody-bound cells in different serum dilutions. Mean titers in sera from hatchery workers and a non-exposed control group did not display significant differences for most tested isolates and application of comprehensive cluster analysis to entire titer data revealed no structure reflecting workers and controls group. Furthermore, determination of immunoreactivity to the complete microbial community in workplace air displayed similar proportions of antibody-bound cells in both groups. Although no general differences in immunoreaction patterns were observed, mean titers to a Proteus mirabilis isolate and to 3 of 4 distinct Acinetobacter baumannii isolates were higher in the group of hatchery workers than in the reference group indicating a potential applicability as exposure markers. We conclude, despite long term bioaerosol exposure, hatchery workers' IgG antibody profiles to tested antigens did not differ substantially from those of the control group. However, increased workers' titers to A. baumannii and clinical relevance of this species should lead to further investigations regarding potential involvement in pathogenesis of occupational respiratory disorders.

Regional Spread of CTX-M-2-Producing Proteus mirabilis with the Identical Genetic Structure in Japan.

In this study, we analyzed the molecular epidemiology of extended-spectrum β-lactamase (ESBL)-producing Proteus mirabilis isolates collected from the central region of Japan. Between 2005 and 2012, 820 clinical P. mirabilis isolates were obtained from ten acute care hospitals in Japan. We characterized ESBL confirmatory test-positive isolates by sequencing the ESBL genes and their flanking regions, detecting plasmid replicons, and performing pulsed-field gel electrophoresis (PFGE). Ninety-six isolates (12%) were positive according to the ESBL confirmatory test; all these isolates possessed blaCTX-M-2 with the same flanking structure of upstream ΔISEcp1 and a downstream region identical to downstream blaKLUA-1. IncT was the prevalent, and only, replicon found in 63 isolates. PFGE analysis detected eight clusters with more than one isolate, among which three included 56 isolates and six included isolates from multiple hospitals. CTX-M-2-producing P. mirabilis with an identical genetic structure flanking blaCTX-M-2 is dominant in this Japanese region, and there is evidence for the clonal spread of isolates.

Characterization of CTX-M-140, a Variant of CTX-M-14 Extended-Spectrum β-Lactamase with Decreased Cephalosporin Hydrolytic Activity, from Cephalosporin-Resistant Proteus mirabilis.

CTX-M-140, a novel CTX-M-type extended-spectrum β-lactamase (ESBL), was identified in cephalosporin-resistant clinical isolates of Proteus mirabilis CTX-M-140 contained an alanine-to-threonine substitution at position 109 compared to its putative progenitor, CTX-M-14. When it was expressed in an Escherichia coli isogenic background, CTX-M-140 conferred 4- to 32-fold lower MICs of cephalosporins than those with CTX-M-14, indicating that the phenotype was attributable to this single substitution. For four mutants of CTX-M-14 that were constructed by site-directed mutagenesis (A109E, A109D, A109K, and A109R mutants), MICs of cephalosporins were similar to those for the E. coli host strain, which suggested that the alanine at position 109 was essential for cephalosporin hydrolysis. The kinetic properties of native CTX-M-14 and CTX-M-140 were consistent with the MICs for the E. coli clones. Compared with that of CTX-M-14, a lower hydrolytic activity against cephalosporins was observed for CTX-M-140. blaCTX-M-140 is located on the chromosome as determined by I-CeuI pulsed-field gel electrophoresis (I-CeuI-PFGE) and Southern hybridization. The genetic environment surrounding blaCTX-M-140 is identical to the sequence found in different plasmids with blaCTX-M-9-group genes among the Enterobacteriaceae Genome sequencing and analysis showed that P. mirabilis strains with blaCTX-M-140 have a genome size of ∼4 Mbp, with a GC content of 38.7% and 23 putative antibiotic resistance genes. Our results indicate that alanine at position 109 is critical for the hydrolytic activity of CTX-M-14 against oxyimino-cephalosporins.

Dissemination of a Multidrug-Resistant VIM-1- and CMY-99-ProducingProteus mirabilisClone in Bulgaria

The aim of this study was to analyze the beta-lactamases and the molecular epidemiology of 19 clinically significant isolates of Proteus mirabilis with decreased susceptibility to imipenem, which have been collected from seven hospitals, located in different Bulgarian towns (Sofia, Varna, and Pleven). The isolates were obtained from blood, urine, tracheal and wound specimens. One additional isolate from hospital environment was included. Susceptibility testing, conjugation experiments, and plasmid replicon typing were carried out. Beta-lactamases were characterized by isoelectric focusing, PCR, and sequencing. Clonal relatedness was investigated by RAPD and PFGE. Integron mapping was performed by PCR and sequencing. All isolates showed a multidrug-resistance profile, but remained susceptible to piperacillin/tazobactam, cefepime, meropenem, and fosfomycin. They produced identical beta-lactamases, namely: TEM-1, VIM-1, and CMY-99. PCR mapping revealed that the blaVIM-1 gene was part of a class 1 integron that additionally included the aac(6')-I, dhfrA1, and ant(3″)-Ia genes. In addition, 17 of the isolates carried the armA gene. Conjugation experiments and plasmid replicon typing were unsuccessful. The isolates were clonally related according to RAPD and PFGE typing. This study reveals the nationwide distribution of a multidrug-resistant P. mirabilis clone producing VIM-1 and CMY-99 along with the presence of different aminoglycoside resistance mechanisms.

Emergence of multidrug-resistant Proteus mirabilis in along-term care facility in Croatia.

An increased frequency of Proteus mirabilis isolates resistant to expanded-spectrum cephalosporins was observed recently in along-term care facility in Zagreb (Godan). The aim of this study was the molecular characterization of resistance mechanisms to new cephalosporins in P.mirabilis isolates from this nursing home. Thirty-eight isolates collected from 2013-2015 showing reduced susceptibility to ceftazidime were investigated. Antibiotic susceptibilities were determined by broth microdilution method. Inhibitor-based tests were performed to detect extended-spectrum (ESBLs) and AmpC β-lactamases. AmpC β-lactamases were characterized by polymerase chain reaction (PCR) followed by sequencing of bla ampC genes. Quinolone resistance determinants (qnr genes) were characterized by PCR. Genotyping of the isolates was performed by repetitive element sequence (rep)-PCR and pulsed-field gel electrophoresis (PFGE). Presence of an AmpC β-lactamase was confirmed in all isolates by combined-disk test with phenylboronic acid. All isolates were resistant to amoxicillin alone and combined with clavulanate, cefotaxime, ceftriaxone, cefoxitin, and ciprofloxacin; but susceptible to cefepime, imipenem, and meropenem. PCR followed by sequencing using primers targeting bla ampc genes revealed CMY-16 β-lactamase in all but one strain. Bla cmy-16 was carried by anon-conjugative plasmid which did not belong to any known plasmid-based replicon typing (PBRT) group. Rep-PCR identified one large clone consisting of 15isolates, three pairs or related isolates, one triplet, and four singletons. PFGE confirmed the clonality of the isolates. This is the first report of multidrug resistant P.mirabilis in anursing home in Croatia. Cephalosporin resistance was due to plasmid-mediated AmpC β-lactamase CMY-16.

Antimicrobial Resistance and &amp;lt;i&amp;gt;β&amp;lt;/i&amp;gt;-Lactamase Production among Hospital Dumpsite Isolates

Metallo-β-Lactamases (MBLs) and Extended Spectrum β-Lactamses (ESBLs) have emerged world-wide as a significant source of β-lactam resistance. The emergence of MBLs and ESBLs encoded on plasmids among Gram-negative pathogens in hospital dumpsites was investigated. Soils of different government and private hospitals were collected and processed following standard bacteriological techniques. Antimicrobial susceptibility testing was carried out by the disk-diffusion technique using Ceftazidime (30 μg), Cefuroxime (30 μg), Cefotaxime (30 μg), Cefixime (5 μg), Trimethprim-sulfamethoxazole (25 μg), Gentamycin (100 μg) Amoxicillin-Clavunalate (30 μg), Ciprofloxacin (5 μg), Ofloxacin (5 μg), Nitrofurantoin (300 μg) and Imipenem (10 μg). The role of plasmids in resistance was evaluated by subjecting isolates to curing using Sodium Dodecyl Sulfate (SDS). ESBLs production by Double-Disk Synergy Test (DDST) was carried out. Isolates resistant to Imipenem were subjected to a confirmatory test using Modified Hodge’s test and to MBLs production by DDST. Eighty-two Gram-negative isolates comprising of 32 (39.02%) Escherichia coli, 20 (24.39%) Serratia marcescens, 14 (17.07%) Klebsiella pneumonia, 10 (12.28%) Proteus mirabilis and 6 (7.32%) Enterobacter aerogenes were obtained. Susceptibility results revealed a 100% resistance of all isolates to Ceftazidime, Cefuroxime, Cefixime, Amoxycillin-clavulanate and Cefotaxime. A total of 66 (80.48%) isolates harboured plasmids out of which 26 (31.71%) isolates were ESBL producers. MBLs production was observed in 8 (25.00%) E. coli, 2 (2.41%) Klebsiella pneumonia and 2 (2.41%) Proteus mirabilis isolates. All MBLs producing isolates were ESBLs producers. The finding of highly resistant isolates producing ESBLs and MBLs in a hospital environment is quite disturbing and should be addressed urgently.