Rapid identification of pathogenic bacteria using Raman spectroscopy and deep learning.

Raman optical spectroscopy promises label-free bacterial detection, identification, and antibiotic susceptibility testing in a single step. However, achieving clinically relevant speeds and accuracies remains challenging due to weak Raman signal from bacterial cells and numerous bacterial species and phenotypes. Here we generate an extensive dataset of bacterial Raman spectra and apply deep learning approaches to accurately identify 30 common bacterial pathogens. Even on low signal-to-noise spectra, we achieve average isolate-level accuracies exceeding 82% and antibiotic treatment identification accuracies of 97.0±0.3%. We also show that this approach distinguishes between methicillin-resistant and -susceptible isolates of Staphylococcus aureus (MRSA and MSSA) with 89±0.1% accuracy. We validate our results on clinical isolates from 50 patients. Using just 10 bacterial spectra from each patient isolate, we achieve treatment identification accuracies of 99.7%. Our approach has potential for culture-free pathogen identification and antibiotic susceptibility testing, and could be readily extended for diagnostics on blood, urine, and sputum.

Bacterial communities associated with anthracnose symptomatic and asymptomatic leaves of guarana, an endogenous tropical crop, and their pathogen antagonistic effects.

Plants are colonized by diverse microorganisms that can substantially impact their health and growth. Understanding bacterial diversity and the relationships between bacteria and phytopathogens may be key to finding effective biocontrol agents. We evaluated the bacterial community associated with anthracnose symptomatic and asymptomatic leaves of guarana, a typical tropical crop. Bacterial communities were assessed through culture-independent techniques based on extensive 16S rRNA sequencing, and cultured bacterial strains were evaluated for their ability to inhibit the growth of Colletotrichum sp. as well as for enzyme and siderophore production. The culture-independent method revealed that Proteobacteria was the most abundant phylum, but many sequences were unclassified. The emergence of anthracnose disease did not significantly affect the bacterial community, but the abundance of the genera Acinetobacter, Pseudomonas and Klebsiella were significantly higher in the symptomatic leaves. In vitro growth of Colletotrichum sp. was inhibited by 11.38% of the cultured bacterial strains, and bacteria with the highest inhibition rates were isolated from symptomatic leaves, while asymptomatic leaves hosted significantly more bacteria that produced amylase and polygalacturonase. The bacterial isolate Bacillus sp. EpD2-5 demonstrated the highest inhibition rate against Colletotrichum sp., whereas the isolates EpD2-12 and FD5-12 from the same genus also had high inhibition rates. These isolates were also able to produce several hydrolytic enzymes and siderophores, indicating that they may be good candidates for the biocontrol of anthracnose. Our work demonstrated the importance of using a polyphasic approach to study microbial communities from plant diseases, and future work should focus on elucidating the roles of culture-independent bacterial communities in guarana anthracnose disease.

Klebsiella sp. strain C2A isolated from olive oil mill waste is able to tolerate and degrade tannic acid in very high concentrations.

Four bacterial strains capable of growing in the presence of tannic acid as sole carbon and energy source were isolated from olive mill waste mixtures. 16S rRNA gene sequencing assigned them to the genus Klebsiella. The most efficient strain, Klebsiella sp. strain C2A, was able to degrade 3.5 g L(-1) tannic acid within 35 h with synthesizing gallic acid as main product. The capability of Klebsiella sp. strain C2A to produce tannase was evidenced at high concentrations of tannic acid up to 50 g L(-1) . The bacteria adapted to the toxicity of tannic acids by an increase in the membrane lipid fatty acids degree of saturation, especially in the presence of concentrations higher than 20 g L(-1) . The highly tolerant and adaptable bacterial strain characterized in this study could be used in bioremediation processes of wastes rich in polyphenols such as those derived from olive mills, winery or tanneries.

Screening and optimization of indole-3-acetic acid production and phosphate solubilization from rhizobacteria aimed at improving plant growth.

A total of 216 bacterial strains were isolated from rice rhizospheric soils in Northern Thailand. The bacterial strains were initially tested for solubilization of inorganic phosphate, indole acetic acid (IAA) production, selected strains were then tested for optimized conditions for IAA production and whether these caused stimulatory effects on bean and maize seedling growth. It was found that all strains had solubilized inorganic phosphate (P), but only 18.05% produced IAA. The best IAA producer was identified by biochemical testing and 16S rDNA sequence analysis as Klebsiella SN 1.1. In addition to being the best IAA producer, this strain was a high P-solubilizer and produced the highest amount of IAA (291.97 ± 0.19 ppm) in culture media supplemented with L-tryptophan. The maximum production of IAA was achieved after 9 days of incubation. The culture requirements were optimized for maximum IAA production. The tested of IAA production by selected isolates was studied in a medium with 0, 0.1, 0.2, 0.5, 0.7, and 0.9% (v/v) L-tryptophan. Low levels (12.6 ppm) of IAA production was recorded without tryptophan addition. Production of IAA in Klebsiella SN 1.1 increased with an increase to 0.2% (v/v) tryptophan concentration. The production of IAA was further confirmed by extraction of crude IAA from this isolate and subsequent Thin Layer Chromatography (TLC) analysis. A specific spot from the extracted IAA production was found to correspond with a standard spot of IAA with the same R (f) value. The Klebsiella strain SN 1.1 also demonstrated stimulatory effects on bean seedlings in vivo.

Clinical implications of extended spectrum beta-lactamase (ESBL) producing Klebsiella species and Escherichia coli on cefepime effectiveness.

OBJECTIVE: To determine the affect of ESBL production among Klebsiella species and Escherichia coli on cefepime effectiveness. METHODS: This was a retrospective, case-controlled study comparing the clinical and microbiologic responses of patients receiving cefepime for ESBL producing Klebsiella species or E. coli from a non-urine source with matched controls receiving cefepime for non-ESBL strains. Cases with ESBLs were included if they received monotherapy and were clinically evaluable. Non-ESBL controls were matched in a 2:1 ratio based on age, infection site, intensive care unit (ICU) stay, pathogen species and date of hospitalization. RESULTS: Ten patients receiving cefepime for ESBLs were matched to 20 controls. Most patients received cefepime 1g q12h. Patients receiving cefepime for an ESBL infection were 9.7 (95% CI: 1.4-68.8) and 28.5 (95% CI: 2.6-306.6) times as likely to have an unsuccessful clinical and microbiological response compared with those with a non-ESBL infection. The presence of an ESBL did not have a statistically significant effect on all cause or infection-related mortality. CONCLUSION: These data indicate that ESBL production among non-urinary Klebsiella species and E. coli negatively affected cefepime effectiveness. Further studies are required to evaluate if higher doses of cefepime may improve responses in ESBLs that are initially susceptible.

Antibiotic resistance and plasmid profiles of Escherichia coli and Klebsiella isolated from in-patients receiving prolonged antibiotic therapy.

Distribution by serogroup, phage type, colicin production, colicin type, sensitivity to antibiotics and plasmid characteristics of 74 Escherichia coli and 11 Klebsiella strains isolated from hospitalized patients receiving prolonged antibiotic therapy indicated that the infections were not associated with the hospital environment. Resistance was tested to 26 antibiotics, some of them being not generally used in therapy; 30 strains were resistant to 4 to 17 antibiotics. There was a significant difference in the antibiotic resistance of strains derived from patients with urinary-tract infections (UTI) and with leukaemia (LP). As compared to the UTI group, among E. coli strains in the LP group the frequency of multiple resistance was significantly higher, the MIC values were higher and R-plasmids were more frequent. Out of 30 multiple resistant E. coli strains 27 were R-plasmid carriers. Three different kinds of plasmid profile were shown in more than one strain (2 out of 10 UTI strains and 3 and 2 out of 10 LP strains). The rest of the isolates differed in plasmid profile from these and from one another; the presence of "epidemic plasmid" was not demonstrated. Plasmid epidemiological examinations may forecast the efficacy of an antibiotic or of a group of antibiotics.

Augmentation of antibody responses of mice to inhaled protein antigens by simultaneously inhaled bacterial lipopolysaccharides.

Serum antibody responses of mice to repeatedly inhaled protein antigens such as bovine serum albumin and ovalbumin, plus or minus bacterial lipopolysaccharide (LPS) in the form of an aerosol were studied. Results showed that the levels of responses to inhaled protein antigens varied, depending on the mouse strain-antigen combination and that LPS inhaled simultaneously with the antigens definitely augmented the responses which were not otherwise very high. LPS extracted from Klebsiella O3 (LPS-K) but not LPS from Escherichia coli O55 (LPS-E), which was inhaled at the time of initial inhalation of antigen, significantly intensified the priming for the secondary antibody response to the antigen subsequently inhaled. Both LPS-K and LPS-E, however, definitely acted to augment the response when they were inhaled repeatedly together with the antigen. Oral administration of antigen or antigen plus LPS-K did not induce any detectable antibody response in our experiment, ruling out the possibility that the antigen and LPS stimulated the immune system via alimentary canal rather than via lung. Tissue distribution of the radioactivity soon after inhalation of 131I-labeled antigen and decay speed of the radioactivity were not significantly changed by LPS-K inhaled simultaneously. This suggested that the augmentation of responses was not mediated by the action of LPS to modulate the air-blood barrier against the entry of antigen via lung. All the results prove for the first time that inhaled LPS displays a definite adjuvant action on antibody responses to inhaled antigens.

An investigation into the properties of klebsiella strains isolated from ankylosing spondylitis patients.

Thirty-nine strains of klebsiella isolated from ankylosing spondylitis patients were examined by the methods of Cowan & Steel (1974), those described by Edmondson et al. (1980) and by capsular typing. No significant difference was detected by any of these methods between these strains and those examined by other workers from non-ankylosing spondylitis patients and other environments.

[Utilization of organic compounds by "Klebsiella" (author's transl)]. / Utilisation de substrats carbonés par Klebsiella.

A total of 209 Klebsiella strains were investigated for their ability to grow on minimal mineral media supplemented by different organic compounds. The "respiratory" strains of Klebsiella described by Cowan as K. pneumoniae (sensu stricto), K. edwardsii and K. atlantae, assimilate substrates at a low rate. They are in this respect closely related to the dystrophic species K. ozaenae and K. rhinoscleromatis, although they taxonomically fit into the more ubiquitous species K. pneumoniae. These findings might result in a reconsideration of the taxonomic status of these strains.

A comparison of the properties of Klebsiella strains isolated from different sources.

The ability to form gas in lactose bile-salt broth at 44.5 degrees C (the "faecal coliform" or FC test), growth in nutrient broth at 10 degrees C, indole production and pectin liquefaction were studied in 480 strains of Klebsiella isolated from human and animal infections, from various sites in the hospital environment and hospital food, and from river water and flowers. A positive FC response was correlated inversely with the ability to grow at 10 degrees C. Most strains of human and animal clinical origin were FC positive, whereas strains from water and flowers were mainly FC negative. The frequency of a positive FC response in strains from the hospital environment fell between these two extremes. The production of indole and liquefaction of pectin by klebsiellas was correlated directly with the ability to grow at 10 degrees C and a negative FC response. Nearly all of the strains could be allocated to one of four groups on the basis of these tests. The capsular serotype, bacteriocine-inhibition patterns and antibiotic sensitivities of the strains were examined. No correlation was evident between the first two properties and klebsiellas from any particular source. Strains of clinical origin were more often resistant to five or more antibiotics than were strains from the hospital environment, which in turn showed a greater frequency of antibiotic resistance than did strains from river water and flowers.

Significance of low-temperature growth associated with the fecal coliform response, indole production, and pectin liquefaction in Klebsiella.

In the genus Klebsiella, the growth respnse in nutient broth at 10 degrees C correlates inversely with the operational definition of a fecal coliform and not merely with the ability to grow at 44.5 degrees C. Of the fecal coliform-positive Klebsiella, 97% did not grow at 10 degrees C after 72 h of incubation. Conversely, 97% of the fecal coliform-negative isolates grew at 10 degrees C. The amount of growth at 10 degrees C varied among the fecal coliform-negative isolates and was found to correlate with indole production and pectin liquefaction. Low-temperature growth associated with specific biochemical tests can be used to differentiate several groups in the genus Klebsiella. Three main groups were discerned. Group I consists of indole-negative, pectin-nonliquefying, fecal coliform-positive isolates that do not grow at 10 degrees C. Group II isolates are differentiated from group I by a fecal-coliform-negative response and growth at 10 degrees C. Group III are indole-positive, pectin-liquefying, fecal coliform-negative isolates that grow at 10 degrees C. In our culture collection, isolates of group I are most frequently of human/animal clinical origins, whereas isolates of groups II and III are predominantly derived from the environment.