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Klebsiella spp. are important opportunistic pathogens commonly defined as environmental clinical mastitis agents. Despite Klebsiella mastitis being clinically impairing in cows and costly to the industry, only a few studies describe Klebsiella isolated from mastitis cases. The aim of this work was to characterize species of Klebsiella involved in clinical mastitis cases in Canada. Klebsiella isolated from clinical mastitis cases (n = 53) were identified to the species level using a biochemical test panel and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. The rpoB gene sequence was used as the gold standard method and identified Klebsiella pneumoniae (n = 40), Klebsiella oxytoca (n = 9), Raoultella ornithinolytica (n = 2), and Raoultella planticola (n = 2). Raoultella , a genus closely related to Klebsiella, was also accurately identified using mass spectrometry but not via biochemical testing. Using the disc diffusion technique, 31 (58%) isolates were found to be susceptible to all antimicrobials tested (n = 18). The remaining 22 (42%) isolates were resistant to 1 or more of the following antimicrobials: kanamycin (2%), streptomycin (38%), spectinomycin (13%), sulfisoxazole (13%), and tetracycline (19%). The following antimicrobial resistance genes were identified: tetA , tetB , sul1 , strA/strB , and aadA. Random amplified polymorphic DNA revealed the majority of our isolates as unrelated and having different patterns, indicating environmental contamination as the primary source of infection. All isolates were shown to be biofilm producers. In conclusion, although antimicrobial resistance was low for both Klebsiella and Raoultella species, genetically related Klebsiella spp. isolates appeared to be more resistant. [ABSTRACT FROM AUTHOR]

Abstract The removal of diclofenac, which is a widely used non-steroidal anti-inflammatory drug and a dangerous environmental contaminant of surface water, was studied. Its complete degradation by Raoultella sp. KDF8 to primary metabolic products was described with individual metabolites identified by mass spectrometry. The Raoultella strain KDF8 was cultivated with unlabelled and 13C6-labelled diclofenac. A total of 32 compounds were detected as intermediates in diclofenac metabolism by high resolution tandem mass spectrometry. Furthermore, proteomic analysis showed that the presence of diclofenac resulted in the upregulation of 24 enzymes associated with the benzoate, catechol, protocatechuate and ketoadipate pathways. The presence of genes encoding specific enzymes, such as catechol 1,2-dioxygenase, protocatechuate 3,4-dioxygenase and quercetin 2,3-dioxygenase, has been verified by sequence analysis of PCR products. The description of diclofenac metabolism by strain KDF8 provides a detailed view of the degradation of xenobiotic aromatic compounds by microorganisms in general. Highlights • The bacterium Raoultella sp. KDF8 catabolized diclofenac. • 13C-labelled and unlabelled DCF was used to identify biodegradation products (BPs). • 32 BPs were identified by a high resolution tandem electrospray mass spectrometry. • Proteomics revealed upregulation of 24 enzymes of aromatic and ketoadipate pathways. • Key metabolic enzymes were confirmed by the sequence analysis of PCR products. [ABSTRACT FROM AUTHOR]

Raoultella electrica , a Gram-negative, non-spore-forming, rod-shaped facultative anaerobe, was identified during a regular investigation of bacterial contamination in table eggs in the winter season. A total of 165 hen's eggs were collected in the winter season from 15 different areas of the city of Jaipur, India. Gram-negative Enterobacteriales were isolated on selective and differential media by the conventional plate method and were further identified by several biochemical tests and 16S rRNA gene sequencing. Commonly prescribed antibiotics for enteric infection were used for antibiotic susceptibility testing. For isolated microorganisms, different resistance patterns were found against the different antibiotics used (p < 0.01). The multiple antibiotic resistance index of bacterial isolates ranged from 0.10 to 0.60. R. electrica strain 1GB/NBRC 109676/KCTC 32430 was isolated for the first time from commercial chicken's eggs. [ABSTRACT FROM AUTHOR]

The genus Raoultella was excised from Klebsiella in 2001, but difficulties in its identification may have led to an underestimation of its incidence and uncertainty on its pathogenic role. Recently, clinical reports involving Raoultella have increased, probably through the introduction of mass-spectrometry in clinical microbiology laboratories and the development of accurate molecular techniques. We performed a retrospective analysis using our blood culture collection (2011–14) to identify Raoultella isolates that could have been erroneously reported as Klebsiella . PCR and gene sequencing of highly specific chromosomal class A β-lactamase genes was established as the reference method, and compared with 16S rRNA and rpoβ sequencing, as well as matrix-assisted laser desorption/ionization time-of-flight mass spectroscopy (MALDI-TOF MS), MicroScan Walkaway system and API20E biochemical identification. MALDI-TOF and rpoβ correctly identified all Raoultella isolates, whereas 16S rRNA provided inconclusive results, and MicroScan and API20E failed to detect this genus. The analysis of the clinical characteristics of all Raoultella bacteraemia cases reported in the literature supports the role of Raoultella as an opportunistic pathogen that causes biliary tract infections in elderly patients who suffer from some kind of malignancy or have undergone an invasive procedure. Two salient conclusions are that Raoultella shows tropism for the biliary tract and so its identification could help clinicians to suspect underlying biliary tract disease when bacteraemia occurs. Concomitantly, as most phenotypic identification systems are not optimized for the identification of Raoultella , the use of MALDI-TOF or additional phenotypic tests is recommended for the reliable identification of this genus. [ABSTRACT FROM AUTHOR]

Pathogens from nonpoint sources are the leading cause of water quality impairments in US surface waters. This study assessed the capacity of basidiomycetous fungal mycelium on cellulosic substrates to remove Escherichia coli from synthetic stormwater under unsaturated vertical-flow conditions. The mycelium of Stropharia rugoso-annulata was tested in mycofiltration columns consisting of 18.6 L containers with mycelium grown on either wood chips or a mixture of wood chips and straw. S. rugoso-annulata mycofiltration columns were loaded with water spiked with 600–900 cfu/100 mL of E. coli at low (0.5 L/min; 0.57 m/d) and high (2.2 L/min; 2.5 m/d) hydraulic loading. Influent and effluent were monitored for thermotolerant coliform and E. coli using the Coliscan membrane filter chromogenic method. Alder wood chips infused with S. rugoso-annulata mycelium yielded a removal rate of around 20% relative to control filters. Wood chip and straw media appeared less effective with substantial net export of bacteria from both mycelium-infused and un-inoculated control media. The un-inoculated control media used in this study commonly exported high concentrations of thermotolerant coliform bacteria. On wood chip-based media, the presence of actively growing mycelium reduced the thermotolerant coliform exports by >90% relative to the control media. The study highlights the limitations of using thermotolerant coliform to assess pathogen removal in cellulose rich ecotechnologies like mycofiltration. [ABSTRACT FROM AUTHOR]

Abstract: Although Raoultella planticola and Raoultella ornithinolytica were described more than 20 years ago, identifying them remains difficult. The reliability of the chromosomal bla gene for this identification was evaluated in comparison with that of the 16S rDNA and rpoB genes in 35 Raoultella strains from different origins. Of the 26 strains previously identified as R. planticola by biochemical tests alone or in association with molecular methods, 21 harboured a bla gene with 99.8% identity with the bla gene of two reference R. ornithinolytica strains (bla ORN gene) and 5 harboured a bla gene with 99.2% identity with the bla gene of two reference R. planticola strains (bla PLA gene). The 9 isolates previously identified as R. ornithinolytica harboured a bla ORN gene. The bla gene-based identification was confirmed by 16S rDNA and rpoB sequencing. The 21 isolates newly identified as R. ornithinolytica had a test negative for ornithine decarboxylase (ODC). Molecular experiments suggested one copy of ODC-encoding gene in both ODC-negative R. ornithinolytica and R. planticola strains and two copies in ODC-positive R. orninthinolytica strains. Analysis of the 35 bla genes allowed us (i) to confirm an identity of only 94% between the bla genes of the two Raoultella species while this identity was > 98% for rpoB and > 99% for 16S rDNA genes and (ii) to develop and successfully apply a bla PCR RFLP assay for Raoultella spp. identification. Overall, this study allowed us to discover ODC-negative R. ornithinolytica and to provide a reliable Raoultella identification method widely available as not requiring sequencing equipment. [Copyright &y& Elsevier]

Abstract: Ninety-nine clinical isolates previously identified as Klebsiella oxytoca were evaluated using matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS). Eight isolates were identified as Raoultella spp., being 5 Raoultella spp. and 3 K. oxytoca, by 16S rRNA sequencing. These isolates were correctly identified by applying the 10% differential rule for the MALDI-TOF MS score values. This approach might be useful to discriminate Raoultella species from K. oxytoca. [Copyright &y& Elsevier]