Search

Introduction Citrobacter bacteria are gram-negative anaerobic bacilli commonly found in water, soil, food, and the intestinal tracts of animals and humans. Patients at highest risk of these nosocomial infections include neonates and adults who are ≥65 years of age, debilitated, or immunocompromised. Citrobacter braakii ( C. braakii), specifically, has been reported to cause sepsis in immunocompromised patients. Herein, we describe a case of an allogeneic stem cell transplant (SCT) adult patient with C. braakii bloodstream infection. Case Report We report our experience managing a central line-associated bloodstream infection (CLABSI) due to C. braakii in an allogeneic SCT patient. Management and Outcomes: Our patient was initially managed with cefepime. The central venous catheter (CVC) was removed. Blood cultures cleared 24 hours after antibiotic initiation. Therapy was transitioned to oral levofloxacin once susceptibilities resulted. Discussion The course of this patient highlights the important relationship between an unusual pathogen, C. braakii, in an immunocompromised allogeneic SCT patient. In our case, the source of the bacteremia was most likely a CLABSI given the positive catheter tip cultures. Although this report describes the successful utilization of cefepime and levofloxacin in the treatment of C. braakii infection, caution should be exercised when choosing empiric antimicrobial therapy as AmpC resistance. This clinical scenario can aid health care providers in making informed treatment decisions when faced with patients diagnosed with this relatively uncommon pathogen. Further reports should be published to determine C. braakii bacteremia management in hematopoietic stem cell transplant patients.

A likely symbiotic association between tephritid fruit flies and gut bacteria has been recognized since the beginning of the last century. However, direct evidence for a link between gut bacteria and fruit fly fitness is still limited or absent for many species. Similar to other tephritids, the gut of Bactrocera minax (Enderlein) (Diptera: Tephritidae) is known to contain bacteria throughout the life stage, but what, if any, impact these bacteria have on B. minax fitness is entirely unknown. In order to elucidate the effects of bacteria on the fitness of B. minax, resident bacteria were isolated from the adult gut using culture-dependent techniques. Adult fly diets were subsequently supplemented with three bacterial isolates (Klebsiella pneumonia, Citrobacter braakii and Pantoea dispersa), or bacteria were removed from flies by antibiotics treatment: untreated adults provided a control. Adult fitness parameters (male and female longevity, female fecundity, male copulation number) were measured for the two treatments and one control group. Results were complex depending on the fitness parameter measured and the bacterial species. Compared to the controls, antibiotic treated B. minax had significantly decreased fecundity, but male and female longevity was increased. When flies were fed diets supplemented with any of the three bacterial isolates, female fecundity was significantly enhanced. However, only Citrobacter braakii significantly increased male mating frequency than control males. The results show that gut bacteria directly influence fitness of B. minax, but impacts are dependent on the bacterial species and the fitness parameters measured.

Citrobacter braakii and Citrobacter freundii are Gram-negative opportunistic pathogens associated with many infectious diseases, including septicemia, in humans and animals. Here, we report the draft genome sequences of seven C. braakii strains and one C. freundii strain isolated from Canadian wastewater treatment facilities.

Presence of extended spectrum beta-lactamases (ESBL) in bacteria is a growing health concern of global significance. The local, regional, national, and international epidemiological studies for extended spectrum beta-lactamases-producing Enterobacteriaceae and their encoding genes in foods are still incomplete. The objective of this study was to determine the occurrence of extended spectrum beta-lactamases-producing Enterobacteriaceae and the characteristics of their encoding genes from a total of 250 samples of various foods of animal-origin (100 raw chicken meat, 100 raw cow milk, and 50 raw cow milk cheese) sold in Turkey. Overall, 55 isolates were positive as extended spectrum beta-lactamases-producing Enterobacteriaceae. The most prevalent extended spectrum beta-lactamases-producing strain were identified as Escherichia coli (80%), followed by Enterobacter cloacae (9.1%), Citrobacter braakii (5.5%), Klebsiella pneumoniae (3.6%), and Citrobacter werkmanii (1.8%) by Vitek® MS. The simultaneous production of extended spectrum beta-lactamases and AmpC was detected in five isolates (9.1%) in E. coli (80%) and E. cloacae (20%). The frequency rates of blaTEM, blaCTX-M, and blaSHV were 96.4%, 53.7%, and 34.5%, respectively. The co-existence of bla -genes was observed in 82% of extended spectrum beta-lactamases producers with a distribution of blaTEM & blaCTX-M (52.7%), blaTEM & blaSHV (20%), blaTEM & blaCTX-M & blaSHV (12.7%), and blaSHV & blaCTX-M (1.8%). The most prevalent variant of blaCTX-M clusters was defined as blaCTX-M-1 (97.2%), followed by blaCTX-M-8 (2.8%). In summary, the analysed foods were found to be posing a health risk for Turkish consumers due to contamination by Enterobacteriaceae with a diversity of extended spectrum beta-lactamases encoding genes.

PGPB enhance plant growth through nitrogen fixation, IAA production, phosphorus and pottasium solubilization. PGPB as biofertilizer provide an alternative for chemical fertilizer to reduce environmental damage. PGPB in consortium could be more effective instead of single strain inoculant. The aim of this study is to investigate compatibility, design soil bacteria consortium isolated from rubber-Canna intercropping plantation and evaluate the consortium’s potential as biofertilizer. Functional roles of PGPB was tested such as nitrogen fixation, potassium and phosphate solubilization, and IAA production. Detection of pathogenic bacteria was tested by Blood Agar method. Compatibility test was performed by cross streak method. Bacteria were identified using sequencing of 16srRNA gene and verified using BLAST to database of NCBI. The results showed that among 19 bacterial isolates, all showed the nitrogen-fixing activity, G5 had the highest phosphorus and potassium solubilization index, whereas Citrobacter braakii strain 167 produced the highest IAA concentration. Compatibility analysis showed that Citrobacter freundii strain LMG 3246, Citrobacter braakii strain DSM 17596 and G5 are compatible as a bacteria consortium and may developed as biofertilizer. This study found a possible new and beneficial biofertilizer formulation to enhance plant growth and to reduce the application of chemical fertilizer.

The aim of the present study was to isolate several bacterial consortia from a soil sample and to establish if they could colonize zirconium-tin alloy, such as Zircaloy-4. Two bacterial consortia containing aerobic heterotrophic bacteria and anaerobic sulfate-reducing bacteria were isolated from a soil sample. The aerobic heterotrophic bacteria exhibited a higher capability to utilize different sole carbon sources, as compared with anaerobic sulfate-reducing bacteria. Based on a morphological, biochemical, and molecular analysis, bacterial isolates were identified as Pseudomonas putida IBBHA1, Pseudomonas aeruginosa IBBHA2, Achromobacter spanius IBBHA3, Citrobacter freundii IBBSR1, Citrobacter youngae IBBSR2, and Citrobacter braakii IBBSR3. Isolated bacterial consortia which possess distinct DNA fingerprints were able to form biofilms and colonize the surface of zirconium-tin alloy coupons, although the colonization of coupons by the aerobic heterotrophic bacteria or anaerobic sulfate-reducing bacteria alone was lower compared with that observed when the coupon was immersed in a mixture of both bacterial consortia. Coupons immersed in these bacterial consortia revealed changes in the surface characteristics, which can facilitate or accelerate zirconium-tin alloy corrosion. The accumulation of corrosion products on coupons surface was less significant when the coupons were immersed solely in aerobic heterotrophic bacteria or anaerobic sulfate-reducing bacteria, compared with that observed when the coupon was immersed in a mixture of both bacterial consortia.

Domestic ruminants are regarded as the major reservoir of Shiga toxin-producing Escherichia coli (STEC) closely related to human infection. A total of 363 ovine carcasses were swabbed in an Algiers city slaughterhouse for research on STEC. First of all, screening of the STECs was carried out by a multiplex PCR searching for the genes coding for the virulence factors stx1 , stx2 and eae. This step was followed by STEC isolation and serotyping. The presence of stx+ /stx+ eae+ genes was shown in 116 sheep carcasses (31·95%). From the 116 positive samples, 20 bacterial strains (17·24%) were isolated. Nineteen strains belonged to the species E. coli (STEC), and 1 belonged to Citrobacter braakii (eae+ stx1 + ). During this study, the presence of potentially pathogenic STEC for humans on the surface of sheep carcasses was confirmed. Corrective measures should be considered at the slaughterhouse level to avoid outbreaks of STEC in Algeria. SIGNIFICANCE AND IMPACT OF THE STUDY: PCR screening revealed the significant presence of the genetic markers of Shiga toxin-producing Escherichia coli (STEC) (stx+ /stx+ eae+ ) on the surfaces of sheep carcasses. Citrobacter braakii (stx1 + eae+ ) was isolated for the first time in this study. The risk of foodborne diseases due to STEC must be taken into account in Algeria. To prevent the emergence of epidemic outbreaks among children and older by people, preventive measures should be taken.

International audience; The spread of carbapenemase-producing Enterobacteriaceae (CPE) is a significant problem for healthcare worldwide. The prevalence of carbapenem-resistant Enterobacteriaceae (CPE) in water environments in Algeria are unknown. The aim of this study was to screen for the presence of CPE isolates in the Soummam River in Bejaia, Algeria. Isolates of Enterobacteriaceae recovered from twelve samples of river water and showing reduced susceptibility to carbapenems were included in this study. The isolates were identified by matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS). Isolates were subjected to antimicrobial susceptibility testing and the modified Carba NP test. Carbapenemase and extendedspectrum b-lactamase (ESBL) determinants were studied by PCR amplification and sequencing. The clonal relatedness between isolates was studied by Multilocus Sequence Typing (MLST) method. A total of 20 carbapenem-resistant Enterobacteriaceae strains were included in this study, identified as Escherichia coli (n = 12), Klebsiella pneumoniae (n = 3), Raoultella ornithinolytica (n = 3), Citrobacter freundii (n = 1) and Citrobacter braakii (n = 1). Carbapenemase genes identified in this study included blaOXA-48, observed in 17 isolates (9 E. coli, 3 K. pneumoniae, 3 R. ornithinolytica, 1 C. freundii and 1 C. braakii), and blaOXA-244, a variant of blaOXA-48, was found in three E. coli isolates. MLST showed that 12 E. coli strains belonged to six different sequence types (ST559, ST38, ST212, ST3541, 1972 and ST2142), and we identified three different STs in K. pneumoniae isolates, including ST133, ST2055, and a new sequence type: ST2192. This study showed the presence of OXA-48-like-producing Enterobacteriaceae in water environments and highlighted the potential role of aquatic environments as reservoirs of clinically relevant antimicrobial-resistant bacteria, with the potential to spread throughout the community. (C) 2017 Elsevier Ltd. All rights reserved.

Non-Human Primates are one of the most displayed exhibits in zoological facilities in Nigeria. They share phylogenetic similarities with humans and a high probability of pathogen transfer exists between humans and these species during zoo visit. The resident microbial flora plays a vital part in animal health. A total of 36 swab (18 oral and 18 rectal) samples were collected from anaesthetised apparently healthy NHPs of the family Cercopithecidae (sub family Cercopithecinae) from two urban wildlife facilities. Standard bacteriological procedures were used in the isolation and identification of the bacteria. Antimicrobial susceptibility testing was carried out using disc diffusion method according to Bauer-Kirby technique. All the samples yielded at least one bacterial organism. The oral swabs had 69.7% Gram positive and 30.3% Gram negative organisms while the rectal samples had 38.8% and 61.2% of Gram positive and negative organism respectively. The bacteria included; Bacillus spp., Staphylococcus saprophyticus, Micrococcus luteus, Proteus spp, Enterococcus faecalis, Escherichia coli, Klebsiella oxytoca, Citrobacter braakii, Serratia odorifera and Aeromonas hydrophila. . Antimicrobial susceptibility testing revealed high susceptibility of the isolates to ciprofloxacin (73.2%), sparfloxacin (70.7%), ofloxacin and pefloxacin (67.1%) and gentamycin (53.7%) while a high resistance was recorded against augmentin, amoxicillin, streptomycin and septrin. Some of the bacteria showed multidrug resistance to many of the antimicrobials. Isolation of cultivable bacteria with multidrug resistance from the oral and rectal orifices of NHPs strengthens the need for holistic sanitary and management regulations in zoological facilities housing these animals bearing in mind the probable routes of transmission of identified organisms.

The intestinal tract is a huge reservoir of Enterobacteriaceae, some of which are opportunist pathogens. Several genera of these bacteria harbour intrinsic antibiotic resistance genes, such as ampC genes in species of Citrobacter, Enterobacter or Escherichia genera. In this work, beta-lactamases and other resistance mechanisms have been characterized in Enterobacteriaceae isolates recovered from healthy human faecal samples, focusing on the ampC beta-lactamase genes. Fifty human faecal samples were obtained, and 70 Enterobacteriaceae bacteria were isolated: 44 Escherichia coli, 4 Citrobacter braakii, 9 Citrobacter freundii, 8 Enterobacter cloacae, 1 Proteus mirabilis, 1 Proteus vulgaris, 1 Klebsiella oxytoca, 1 Serratia sp. and 1 Cronobacter sp. A high percentage of resistance to ampicillin was detected (57%), observing the AmpC phenotype in 22 isolates (31%) and the ESBL phenotype in 3 isolates. AmpC molecular characterization showed high diversity into bla CMY and bla ACT genes from Citrobacter and Enterobacter species, respectively, and the pulsed-field gel electrophoresis (PFGE) analysis demonstrated low clonality among them. The prevalence of people colonized by strains carrying plasmid-mediated ampC genes obtained in this study was 2%. The unique plasmid-mediated bla AmpC identified in this study was the bla CMY-2 gene, detected in an E. coli isolate ascribed to the sequence type ST405 which belonged to phylogenetic group D. The hybridization and conjugation experiments demonstrated that the ISEcp1-bla CMY-2-blc structure was carried by a ~78-kb self-transferable IncK plasmid. This study shows a high polymorphism among beta-lactamase genes in Enterobacteriaceae from healthy people microbiota. Extensive AmpC-carrier studies would provide important information and could allow the anticipation of future global health problems.

Among five American Type Culture Collection (ATCC) Citrobacter strains, qnrB60 in Citrobacter freundii ATCC 6879, an isolate from the preantibiotic era, and qnrB61 in Citrobacter braakii ATCC 51113 T , a type strain belonging to the C. freundii complex, were identified. Meanwhile, a truncated qnrB -like pseudogene was identified in C. freundii ATCC 8090 T and ATCC 43864. No qnrB -like sequence was found in Citrobacter koseri ATCC 27028 T . These findings underscore the close relationship between this species and qnrB .

Objectives: There is little information about carbapenemase-producing (CP) Citrobacter spp. We studied the molecular epidemiology and microbiological features of CP Citrobacter spp. isolates collected in Spain (2013-15). Methods: In total, 119 isolates suspected of being CP by the EUCAST screening cut-off values were analysed. Carbapenemases and ESBLs were characterized using PCR and sequencing. The genetic relationship among Citrobacter freundii isolates was studied by PFGE. Results: Of the 119 isolates, 63 (52.9%) produced carbapenemases, of which 37 (58.7%) produced VIM-1, 20 (31.7%) produced OXA-48, 12 (19%) produced KPC-2, 2 (3.2%) produced NDM-1 and 1 (1.6%) produced VIM-2; 9 C. freundii isolates co-produced VIM-1 plus OXA-48. Fourteen isolates (22.2%) also carried ESBLs: 8 CTX-M-9 plus SHV-12, 2 CTX-M-9, 2 SHV-12 and 2 CTX-M-15. Fifty-seven isolates (90.5%) were C. freundii, 4 (6.3%) were Citrobacter koseri, 1 (1.6%) was Citrobacter amalonaticus and 1 (1.6%) was Citrobacter braakii. By EUCAST breakpoints, eight (12.7%) of the CP isolates were susceptible to the four carbapenems tested. In the 53 CP C. freundii analysed by PFGE, a total of 44 different band patterns were observed. Four PFGE clusters were identified: cluster 1 included eight isolates co-producing VIM-1 and OXA-48; blaVIM-1 was carried in a class 1 integron (intI-blaVIM-1-aacA4-dfrB1-aadA1-catB2-qacEΔ1/sul1) and blaOXA-48 was carried in a Tn1999.2 transposon. Conclusions: We observed the clonal and polyclonal spread of CP Citrobacter spp. across several Spanish geographical areas. Four species of Citrobacter spp. produced up to five carbapenemase types, including co-production of VIM-1 plus OXA-48. Some CP Citrobacter spp. isolates were susceptible to the four carbapenems tested, a finding with potential clinical implications. This work was supported by Plan Nacional de I + D+i 2013–2016 and Instituto de Salud Carlos III, Subdirección General de Redes y Centros de Investigación Cooperativa, Ministerio de Economía, Industria y Competitividad, Spanish Network for Research in Infectious Diseases (REIPI RD16CIII/0004/0002) - co-financed by European Development Regional Fund (ERDF) ‘A way to achieve Europe’, Operative Program Intelligent Growth 2014–2020. This work was also supported by a grant from the Instituto de Salud Carlos III (grant number MPY 1135/16), by the Antibiotic Resistance Surveillance Program of the Centro Nacional de Microbiología (Instituto de Salud Carlos III, Ministerio de Economía y Competitividad) of Spain and by a grant from the Agencia Española de Medicamentos y Productos Sanitarios (grant number MEG 1281/14). Sí

Bacterial growth and histamine formation in mackerel during storage at 0, 4, 15 and 25C were monitored. Histamine production and bacterial multiplication were accelerated with increase of the storage temperature. The highest histamine level of 2,080 mg/kg was detected after 2 days of storage at 25C. Selective media were used for isolation of histamine-producing bacteria (HPB) and the histamine production was confirmed by high-performance liquid chromatography analysis. A total number of 305 strains were isolated from spoiled mackerel and 32 of them were identified as HPB. Most of the HPB were isolated from violet red bile glucose agar. Morganella morganii, Citrobacter braakii, Hafnia alvei and Raoultella planticola were confirmed as high histamine-producing strains, whereas other microorganisms were found to possess weak histamine-producing ability. Practical Applications In order to effectively inhibit the activity of HPB, it is necessary to have a good understanding of bacterial species contributing to histamine production in mackerel during storage. Therefore, we studied the variation of histamine content and the growth of bacteria related to histamine production in mackerel during storage. Results of this study could be useful to assure the safety of mackerel products by controlling histamine content.

Continuous European Union programmes with specified methods for enumeration of Escherichia coli in bivalves for human consumption are currently running. The objective of this research was to examine the species accuracy of the five times three tube Most Probable Number (MPN) EU reference method used for detection of E. coli in marine bivalves. Among 549 samples of bivalves harvested from Norwegian localities during 2014 and 2015, a total number of 200 bacterial isolates were prepared from randomly selected culture-positive bivalves. These presumptive E. coli isolates were characterized biochemically by the Analytical Profile Index (API) 20E, as well as by Matrix Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry (MALDI-TOF MS). The majority of isolates (90%) were identified as E. coli, by both API 20E and MALDI-TOF MS. Ten isolates (5%) were identified as Klebsiella pneumoniae, while one isolate was identified as K. oxytoca by both methods, whereas three isolates were identified as Acinetobacter baumannii, Citrobacter braakii, and Enterobacter cloacae, respectively. The identification of the remaining six isolates were not in compliance between the two methods.

Histidine, lysine, ornithine and tyrosine decarboxylase activities were tested in 79 strains of Enterobacteriaceae (41 of Hafnia alvei, 17 of Serratia liquefaciens, 5 of Enterobacter cloacae, 4 of Citrobacter braakii, 2 of Proteus vulgaris, 2 of Proteus mirabilis, 2 of Providencia stuartii, 2 of Klebsiella terrigena, 1 of Rahnella aquatilis, 1 of Salmonella arizonae, 1 of Citrobacter youngae and 1 of Escherichia coli) isolated from Botillo, a Spanish traditional sausage. In general, the strains were positive for all four activities, with the exception of two strains of H. alvei and the E. coli strain, which did not display histidine decarboxylase activity. The strains of P. mirabilis and P. stuartii did not exhibit any of the four activities tested. Accumulation of putrescine and cadaverine was studied throughout growth of the 75 strains that displayed ornithine and lysine decarboxylase activities. Biogenic amines were produced particularly in the exponential phase, with maximum accumulation occurring after between 12 to 72 h, depending on the biogenic amine and microbial species considered. Maximum accumulation of putrescine varied greatly between species and within the same species, and ranged from 18 mg/l in the R. aquatilis strain to 7325 mg/l in a H. alvei strain. Maximum accumulation of cadaverine varied less than that of putrescine, and ranged from 30 mg/l in the R. aquatilis strain to 1935 mg/l in a S. liquefaciens strain.

Rhizosphere microorganisms possessing phytase activity are considered important for rendering phytate-phosphorus (P) available to plants. In the present study, the Citrobacter braakii phytase gene (appA) was overexpressed in rhizobacteria possessing plant growth promoting (PGP) traits, for increasing their potential as bioinoculants. AppA was cloned under the lac promoter in the broadhost-range expression vector pBBR1MCS-2. Transformation of the recombinant construct pCBappA resulted in high constitutive phytase activity in all of the eight rhizobacterial strains belonging to genera Pantoea, Citrobacter, Enterobacter, Pseudomonas (two strains), Rhizobium (two strains), and Ensifer that were studied. Transgenic rhizobacterial strains were found to display varying levels of phytase activity, ranging from 10-folds to 538-folds higher than the corresponding control strains. The transgenic derivative of Pseudomonas fluorescens CHA0, a well-characterized plant growth promoting rhizobacterium, showed the highest expression of phytase (~8 U/mg) activity in crude extracts. Although all transformants showed high phytase activity, rhizobacteria having the ability to secrete organic acid showed significantly higher release of P from Caphytate in buffered minimal media. AppA overexpressing rhizobacteria showed increased P content, and dry weight (shoot) or shoot/ root ratio of mung bean (Vigna radiata) plants, to different extents, when grown in semisolid agar (SSA) medium containing Na-phytate or Ca-phytate as the P sources. This is the first report of the overexpression of phytase in rhizobacterial strains and its exploitation for plant growth enhancement.

Aims: To determine whether American alligators (Alligator mississippiensis) are an unrecognized poikilothermic source of faecal coliform and/or potential pathogenic bacteria in South Carolina’s coastal waters. Methods and Results: Bacteria from the cloaca of American alligators, as well as bacteria from surface water samples from their aquatic habitat, were isolated and identified. The predominant enteric bacteria identified from alligator samples using biochemical tests included Aeromonas hydrophila, Citrobacter braakii, Edwardsiella tarda, Escherichia coli, Enterobacter cloacae, Plesiomonas shigelloides and putative Salmonella, and these were similar to bacteria isolated from the surface waters in which the alligators inhabited. Based on most-probable-number enumeration estimates from captive alligator faeces, faecal coliform bacteria numbered 8·0 × 109 g−1 (wet weight) of alligator faecal material, a much higher concentration than many other documented endothermic animal sources. Conclusions: A prevalence of enteric bacteria, both faecal coliforms and potential pathogens, was observed in American alligators. The high faecal coliform bacterial density of alligator faeces may suggest that alligators are a potential source of bacterial contamination in South Carolina coastal waters. Significance and Impact of the Study: These findings help to increase our understanding of faecal coliform and potential pathogenic bacteria from poikilothermic reptilian sources, as there is the potential for these sources to raise bacterial water quality levels above regulatory thresholds.

The efficacy of the agents was evaluated on the basis of minimum inhibitory concentration (MIC) and time-kill curve analysis in Pseudomonas aeruginosa, Staphylococcus aureus, Acinetobacter baumannii, Mycobacterium smegmatis, and Citrobacter braakii; MIC was found to be 0.5, 4.0, 0.015625, 0.0078125, and 0.0625 mg/L in FDC of ceftazidime + sulbactam, respectively, which is lower than ceftazidime and sulbactam individually. Time-kill curve analysis demonstrated maximum killing of bacteria after 4 h. A fixed dose combination of ceftazidime + sulbactam was found to have stronger antibacterial properties than ceftazidime and sulbactam alone at in vitro analysis.

We examine the use of multiwavelength ultraviolet (UV) resonance–Raman signatures to identify the effects of growth phase and growth medium on gram-positive and gram-negative bacteria. Escherichia coli(E. coli), Citrobacter koseri(C. koseri), Citrobacter braakii (C. braakii), and Bacillus cereus (B. cereus)were grown to logarithmic and stationary phases in nutrient broth and brain heart infusion broth. Resonance Raman spectra of bacteria were obtained at multiple wavelengths between 220 and 260 nm; a range that encompasses the resonance frequencies of cellular constituents. We find that spectra of the same bacterial species exhibit differences due to both growth condition and growth phase, but the larger differences reflect changes due to growth phase. The differences in the Raman spectra correlate with genetic differences among the species. Using a Pearson correlation based algorithm, we achieve successful identification of these bacteria in 83% of the cases.

Citrobacter braakii is a Gram-negative bacterium belonging to the Enterobacteriaceae family. Here, we report 5.2- and 5.0-Mb genome assemblies for C. braakii strains GTA-CB01 and GTA-CB04, respectively.

Microbial phytases catalyze dephosphorylation of phytic acid, thereby potentially releasing chelated iron and improving human iron absorption from cereal-based diets. For this catalysis to take place in vivo, the phytase must be robust to low pH and proteolysis in the gastric ventricle. This study compares the robustness of five different microbial phytases, evaluating thermal stability, activity retention, and extent of dephosphorylation of phytic acid in a simulated low-pH/pepsin gastric environment and examines secondary protein structural changes at low pH via circular dichroism. The Peniophora lycii phytase was found to be the most thermostable, but the least robust enzyme in gastric conditions, whereas the Aspergillus niger and Escherichia coli phytases proved to be most resistant to gastric conditions. The phytase from Citrobacter braakii showed intermediate robustness. The extent of loss of secondary structure at low pH correlated positively with the extent of activity loss at low pH.

In this study, we investigated the effects of enzyme-producing probiotic bacteria isolated from the gastrointestinal tract of rainbow trout on the growth performance, feed conversion ratio, and digestive enzyme activity, of fry (Oncorhynchus mykiss). Three isolates (G8/2013, T7/2013 and U5/2013) of candidate bacteria elicited the highest protease, lipase, and amylase activities, respectively. Isolates were identified as Aeromonas sp., Bacillus sp. and Citrobacter braakii by morphological, physiological, biochemical characterizations as well as 16S rRNA gene sequence analysis. The fry basal diet was supplemented with probiotics at varying concentrations; G8 group, Aeromonas sp. 1.72 x 108 CFU/g; U5 group, Bacillus sp. 3.01 x 108 CFU/g; T7 group, C. braakii 2.96 x 108 CFU/g and a mixed group (same bacterial concentrations), and control group (no bacteria). The rainbow trout fry were fed ad libitum in triplicate treatments with supplemented and non-supplemented probiotic diets for 70-days. The total bacterial count in the intestine was significantly higher in the mixed group (30th and 50th days) and U5 group (50th and 70th days) compared with the control group. However, there was no significant difference in weight gain, specific growth rate (SGR), feed conversion ratio (FCR), nutrient digestibility, or digestive enzyme activity among the groups.a

To further identify the origins of plasmid-mediated cephalosporinases that are currently spreading worldwide, the chromosomal beta-lactamase genes of Citrobacter braakii, Citrobacter murliniae, Citrobacter werkmanii reference strains and of Escherichia fergusonii and Enterobacter cancerogenus clinical isolates were cloned and expressed into Escherichia coli and sequenced. These beta-lactamases had all a single pI value8 and conferred a typical AmpC-type resistance pattern in E. coli recombinant strains. The cloned inserts obtained from genomic DNAs of each strain encoded Ambler class C beta-lactamases. The AmpC-type enzymes of C. murliniae, C. braakii and C. werkmanii shared 99%, 96% and 95% amino acid sequence identity, respectively, with chromosomal AmpC beta-lactamases from Citrobacter freundii. The AmpC-type enzyme of E. cancerogenus shared 85% amino acid sequence identity with the chromosomal AmpC beta-lactamase of Enterobacter cloacae OUDhyp and the AmpC-type enzyme of E. fergusonii shared 96% amino acid sequence identity with that of E. coli K12. The ampC genes, except for E. fergusonii, were associated with genes homologous to regulatory ampR genes of other chromosomal class C beta-lactamases that explain inducibility of beta-lactamase expression in these strains. This work provides further evidence of the molecular heterogeneity of class C beta-lactamases.

The growth of Listeria monocytogenes during the pathogen specific enrichment of food samples can be limited by the presence of additional microorganisms that are resistant to the selective conditions being applied. If growth is severely limited and minimum post-enrichment threshold levels are not met then the presence of L. monocytogenes may go undetected. Several food products were screened for non-pathogenic commensal or spoilage microorganisms that are capable of growth under the conditions commonly used by regulatory testing laboratories to select for Listeria species. The effect of these potential competitor microorganisms on the ability to detect L. monocytogenes by several common molecular screening assays was then determined. Eight species of bacteria were isolated from foods that demonstrated the ability to grow in buffered Listeria enrichment broth under selective conditions. Growth of these competitor microorganisms during the enrichment incubation resulted in a decrease ranging from 1 to 4 logs in the 48 h population of L. monocytogenes. Three strains of L. monocytogenes representing serotypes 1/2a, 1/2b, and 4b were included in this study but no one serotype appeared to be most or least sensitive to the presence of competitor microorganisms. One additional strain of L. monocytogenes was identified as displaying minimal growth during the enrichment period in the presence of the Citrobacter braakii with the final population only reaching approximately 2.6 log CFU/ml after 48 h which was a 2 log increase over the initial population. This particular strain was subsequently shown to be difficult to detect following enrichment by an automated immunofluorescence assay and an antibody-based lateral flow device assay. In some enrichments, this strain was also difficult to detect by real-time PCR.

We will examine the use of multi-wavelength UV resonance-Raman signatures to identify the effects of growth phase on different types of bacteria. Gram positive and gram-negative species, Escherichia coli, Bacillus cereus, Citrobacter koseri and Citrobacter braakii were grown to logarithmic and stationary phases in different culture media. Raman spectra of bacteria were obtained by sequential illumination of samples between 220 and 260 nm; a range which encompasses the resonance frequencies of cellular components. In addition to the information contained in the single spectrum, this two-dimensional signature contains information reflecting variations in resonance cross sections with illumination wavelength. Results of our algorithms in identifying the differences between these germs are discussed. Preliminary results indicate that growth affects the Raman signature, but not to an extent that would negate identification of the species.

A new biological inspired method to produce nanopalladium is the precipitation of Pd on a bacterium, i.e., bio-Pd. This bio-Pd can be applied as catalyst in dehalogenation reactions. However, large amounts of hydrogen are required as electron donor in these reactions resulting in considerable costs. This study demonstrates how bacteria, cultivated under fermentative conditions, can be used to reductively precipitate bio-Pd catalysts and generate the electron donor hydrogen. In this way, one could avoid the costs coupled to hydrogen supply. The catalytic activities of Pd(0) nanoparticles produced by different strains of bacteria (bio-Pd) cultivated under fermentative conditions were compared in terms of their ability to dehalogenate the recalcitrant aqueous pollutants diatrizoate and trichloroethylene. While all of the fermentative bio-Pd preparations followed first order kinetics in the dehalogenation of diatrizoate, the catalytic activity differed systematically according to hydrogen production and starting Pd(II) concentration in solution. Batch reactors with nanoparticles formed by Citrobacter braakii showed the highest diatrizoate dehalogenation activity with first order constants of 0.45 ± 0.02 h−1 and 5.58 ± 0.6 h−1 in batches with initial concentrations of 10 and 50 mg L−1 Pd, respectively. Nanoparticles on C. braakii, used in a membrane bioreactor treating influent containing 20 mg L−1 diatrizoate, were capable of dehalogenating 22 mg diatrizoate mg−1 Pd over a period of 19 days before bio-Pd catalytic activity was exhausted. This study demonstrates the possibility to use the combination of Pd(II), a carbon source and bacteria under fermentative conditions for the abatement of environmental halogenated contaminants.

This study aims to establish the isolation frequency of species of the family Enterobacteriaceae in fecal samples of South American common sea lion (Otaria flavescens). Thirty fecal samples were collected from the resting ground of an urban sea lions colony in Valdivia, southern Chile. The bacteria species and their isolation frequencies were Edwarsiella tarda (73%), Escherichia coli (70%), Hafnia alvei (33%), Morganella morganii (7%), Proteus mirabilis (7%), Klebsiella pnuemoniae subsp. pneumoniae (3%), Serratia rubidea (3%), Providencia rustigianii (3%) and Citrobacter braakii (3%).

The phytase gene appAS was isolated from Shigella sp. CD2 genomic library. The 3.8 kb DNA fragment contained 1299 bp open reading frame encoding 432 amino acid protein (AppAS) with 22 amino acid signal peptide at N-terminal and three sites of N-glycosylation. AppAS contained the active site RHGXRXP and HDTN sequence motifs, which are conserved among histidine acid phosphatases. It showed maximum identity with phytase AppA of Escherichia coli and Citrobacter braakii. The appAS was expressed in Pichia pastoris and E. coli to produce recombinant phytase rAppAP and rAppAE, respectively. Purified glycosylated rAppAP and nonglycosylated rAppAE had specific activity of 967 and 2982 U mg(-1), respectively. Both had pH optima of 5.5 and temperature optima of 60°C. Compared with rAppAE, rAppAP was 13 and 17% less active at pH 3.5 and 7.5 and 11 and 18% less active at temperature 37 and 50°C, respectively; however, it was more active at higher incubation temperatures. Thermotolerance of rAppAP was 33% greater at 60°C and 24% greater at 70°C, when compared with rAppAE. Both the recombinant enzymes showed high specificity to phytate and resistance to trypsin. To our knowledge, this is the first report on cloning and expression of phytase from Shigella sp.

Arylsulfatase activity was detected in a bacterial strain, Citrobacter braakii 69-b, isolated from soil by enrichment cultivation using porcine gastric mucin. The production of arylsulfatase was derepressed markedly in a synthetic medium by the addition of tyramine. The purified enzyme hydrolyzed 4-nitrophenyl sulfate, 4-nitrocatechol sulfate, and 3-indoxyl sulfate, and was classified as type I arylsulfatase.

The use of molecular tools for early and rapid detection of gram-negative histamine-producing bacteria is important for preventing the accumulation of histamine in fish products. To date, no molecular detection or identification system for gram-negative histamine-producing bacteria has been developed. A molecular method that allows the rapid detection of gram-negative histamine producers by PCR and simultaneous differentiation by single-strand conformation polymorphism (SSCP) analysis using the amplification product of the histidine decarboxylase genes ( hdc ) was developed. A collection of 37 strains of histamine-producing bacteria (8 reference strains from culture collections and 29 isolates from fish) and 470 strains of non-histamine-producing bacteria isolated from fish were tested. Histamine production of bacteria was determined by paper chromatography and confirmed by high-performance liquid chromatography. Among 37 strains of histamine-producing bacteria, all histidine-decarboxylating gram-negative bacteria produced a PCR product, except for a strain of Citrobacter braakii . In contrast, none of the non-histamine-producing strains (470 strains) produced an amplification product. Specificity of the amplification was further confirmed by sequencing the 0.7-kbp amplification product. A phylogenetic tree of the isolates constructed using newly determined sequences of partial hdc was similar to the phylogenetic tree generated from 16S ribosomal DNA sequences. Histamine accumulation occurred when PCR amplification of hdc was positive in all of fish samples tested and the presence of powerful histamine producers was confirmed by subsequent SSCP identification. The potential application of the PCR-SSCP method as a rapid monitoring tool is discussed.

Tributyltin compound (TBT) released into the aquatic environment is generally degraded by bacteria in water and sediment. The isolation of TBT-degrading bacteria from TBT polluted sediment leads to the indication of specific potential TBT degraders. Two new strains of bacteria designated as B2 and B3 were successfully isolated using glycerol medium containing tributyltin chloride (TBTC) at 130 μM from contaminated sediment collected from Bowling Basin in Glasgow. The observed degradation after 14 days of the microcosm from the sediment and the isolated bacteria were investigated at an initial concentration of 1 μM TBTC. It was found that TBT was degraded by the bacterial strains B2 and B3 at 8.3 and 16.9 %, respectively. The results indicate that B2 and B3 are effective as TBT degraders. EC50 of B2 and B3 in water were 88.73 and 112.53 μM TBTC, which were significantly higher than the concentration of TBT measured at the basin, suggesting a low effect of TBT on the growth and activity of bacteria. After identification using API 20E and 16S sequencing, the bacterial isolate strain B2 is Citrobacter braakii and B3 is Enterobacter cloacae. Therefore, this study has discovered two species of high resistance TBT degrader which have never been previously studied or isolated based upon TBT degradation ability.

Bacteria that thrive in extreme conditions and the bacteriophages that infect them are sources of valuable enzymes resistant to denaturation at high temperatures. Many of these heat-stable proteins are useful for biotechnological applications, nevertheless, none have been utilized as antibacterial agents. Here, we demonstrate the bactericidal potential of Ts2631 endolysin from the extremophilic bacteriophage vB_Tsc2631, which infects Thermus scotoductus, against the alarming multidrug-resistant clinical strains of Acinetobacter baumannii, Pseudomonas aeruginosa and pathogens from the Enterobacteriaceae family. A 2&ndash, 3.7 log reduction in the bacterial load was observed in antibacterial tests against A. baumannii and P. aeruginosa after 1.5 h. The Ts2631 activity was further enhanced by ethylenediaminetetraacetic acid (EDTA), a metal ion chelator (4.2 log reduction in carbapenem-resistant A. baumannii) and, to a lesser extent, by malic acid and citric acid (2.9 and 3.3 log reductions, respectively). The EDTA/Ts2631 combination reduced all pathogens of the Enterobacteriaceae family, particularly multidrug-resistant Citrobacter braakii, to levels below the detection limit (&gt, 6 log), these results indicate that Ts2631 endolysin could be useful to combat Gram-negative pathogens. The investigation of A. baumannii cells treated with Ts2631 endolysin variants under transmission electron and fluorescence microscopy demonstrates that the intrinsic antibacterial activity of Ts2631 endolysin is dependent on the presence of its N-terminal tail.