Deep Profiling of the Proteome Dynamics of Pseudomonas aeruginosa Reference Strain PAO1 under Different Growth Conditions.

As one of the most common bacterial pathogens causing nosocomial infections, Pseudomonas aeruginosa is highly adaptable to survive under various conditions. Here, we profiled the abundance dynamics of 3489 proteins across different growth stages in the P. aeruginosa reference strain PAO1 using data-independent acquisition-based quantitative proteomics. The proteins differentially expressed during the planktonic growth exhibit several distinct patterns of expression profiles and are relevant to various biological processes, highlighting the continuous adaptation of the PAO1 proteome during the transition from the acceleration phase to the stationary phase. By contrasting the protein expressions in a biofilm to planktonic cells, the known roles of T6SS, phenazine biosynthesis, quorum sensing, and c-di-GMP signaling in the biofilm formation process were confirmed. Additionally, we also discovered several new functional proteins that may play roles in the biofilm formation process. Lastly, we demonstrated the general concordance of protein expressions within operons across various growth states, which permits the study of coexpression protein units, and reversely, the study of regulatory components in the operon structure. Taken together, we present a high-quality and valuable resource on the proteomic dynamics of the P. aeruginosa reference strain PAO1, with the potential of advancing our understanding of the overall physiology of Pseudomonas bacteria.

Emerging Trends and Hot Topics in Cardiopulmonary Resuscitation Research: A Bibliometric Analysis from 2010 to 2019.

BACKGROUND Cardiopulmonary resuscitation (CPR) is a topic of great scientific and clinical interest that has received much attention in the past decade. Our study aimed to predict the trends in CPR research activities and evaluate hot topics via bibliometric means, quantitatively and qualitatively. MATERIAL AND METHODS All data were collected from a search of the Web of Science Core Collection on May 12, 2020. Retrieved information was investigated with bibliometric analysis by CiteSpace and VOSviewer software and the Online Analysis Platform of Literature Metrology to analyze and predict the trends and hotspots in this field. RESULTS Our search returned a total of 9563 articles and reviews on CPR published from 2010 through 2019. The number of original research studies on CPR has been increasing annually. The journal Resuscitation published the greatest number of manuscripts involved CPR, and the leading country and institution with regard to contributions on CPR were the United States and the University of Pennsylvania. Keyword co-occurrence/co-citation-cluster analysis showed that the most popular terms associated with CPR occurred in the manner of cluster labels, such as therapeutic hypothermia and treatment recommendation, among others. In addition, palliative care, sepsis, extracorporeal membrane oxygenation, and brain injury were identified as new foci through burst detection analysis. CONCLUSIONS Our study showed that the scientific research focus on CPR is switching from traditional therapeutic treatments to a public health practice, with in-depth understanding and development of CPR-related techniques expanding over the past decade. These results demonstrate trends in the CPR research and detected the possible neo-foci for ensuing research.

Practical Fixed-Time Bipartite Synchronization of Uncertain Coupled Neural Networks Subject to Deception Attacks via Dual-Channel Event-Triggered Control.

This article investigates the practical fixed-time synchronization of uncertain coupled neural networks via dual-channel event-triggered control. Contrary to some previous studies, the bipartite synchronization of signed graphs representing cooperative and antagonistic interactions is studied. The communication channel is introduced into deception attacks, which are described by Bernoulli's stochastic variables. Based on the concept of two channels, event-triggered mechanisms are designed for sensor-to-controller and controller-to-actuator channels to reduce communication consumption and controller update consumption as much as possible. Lyapunov and comparison theories are used to derive synchronization criteria and explicit expression of settling time. An example of Chua's circuit system is presented to demonstrate the feasibility of the obtained theoretical results.

Embedded EEG Feature Selection for Multi-Dimension Emotion Recognition via Local and Global Label Relevance.

Due to the problem of a small amount of EEG samples and relatively high dimensionality of electroencephalogram (EEG) features, feature selection plays an essential role in EEG-based emotion recognition. However, current EEG-based emotion recognition studies utilize a problem transformation approach to transform multi-dimension emotional labels into single-dimension labels, and then implement commonly used single-label feature selection methods to search feature subsets, which ignores the relations between different emotional dimensions. To tackle the problem, we propose an efficient EEG feature selection method for multi-dimension emotion recognition (EFSMDER) via local and global label relevance. First, to capture the local label correlations, EFSMDER implements orthogonal regression to map the original EEG feature space into a low-dimension space. Then, it employs the global label correlations in the original multi-dimension emotional label space to effectively construct the label information in the low-dimension space. With the aid of local and global relevance information, EFSMDER can conduct representational EEG feature subset selection. Three EEG emotional databases with multi-dimension emotional labels were used for performance comparison between EFSMDER and fourteen state-of-the-art methods, and the EFSMDER method achieves the best multi-dimension classification accuracies of 86.43, 84.80, and 97.86 percent on the DREAMER, DEAP, and HDED datasets, respectively.

A Multi-Source Transfer Joint Matching Method for Inter-Subject Motor Imagery Decoding.

Individual differences among different subjects pose a great challenge to motor imagery (MI) decoding. Multi-source transfer learning (MSTL) is one of the most promising ways to reduce individual differences, which can utilize rich information and align the data distribution among different subjects. However, most MSTL methods in MI-BCI combine all data in the source subjects into a single mixed domain, which will ignore the effect of important samples and the large differences in multiple source subjects. To address these issues, we introduce transfer joint matching and improve it to multi-source transfer joint matching (MSTJM) and weighted MSTJM (wMSTJM). Different from previous MSTL methods in MI, our methods align the data distribution for each pair of subjects, and then integrate the results by decision fusion. Besides that, we design an inter-subject MI decoding framework to verify the effectiveness of these two MSTL algorithms. It mainly consists of three modules: covariance matrix centroid alignment in the Riemannian space, source selection in the Euclidean space after tangent space mapping to reduce negative transfer and computation overhead, and further distribution alignment by MSTJM or wMSTJM. The superiority of this framework is verified on two common public MI datasets from BCI competition IV. The average classification accuracy of the MSTJM and wMSTJ methods outperformed other state-of-the-art methods by at least 4.24% and 2.62% respectively. It's promising to advance the practical applications of MI-BCI.

The phosphate-induced small RNA EsrL promotes E. coli virulence, biofilm formation, and intestinal colonization.

Escherichia coli are part of the normal intestinal microbiome, but some enterohemorrhagic E. coli (EHEC) and enteropathogenic E. coli (EPEC) strains can cause potentially life-threatening gastroenteritis. Virulence factors underlying the ability of EHEC and EPEC to cause disease include those encoded in the locus of the enterocyte effacement (LEE) pathogenicity island. Here, we demonstrated that EsrL, a small RNA present in many E. coli strains, promoted pathogenicity, adhesion, and biofilm formation in EHEC and EPEC. PhoB, the response regulator of the two-component system that controls cellular responses to phosphate, directly repressed esrL expression under low-phosphate conditions. A phosphate-rich environment, similar to that of the human intestine, relieved PhoB-mediated repression of esrL. EsrL interacted with and stabilized the LEE-encoded regulator (ler) transcript, which encodes a transcription factor for LEE genes, leading to increased bacterial adhesion to cultured cells and colonization of the rabbit colon. EsrL also bound to and stabilized the fimC transcript, which encodes a chaperone that is required for the assembly of type 1 pili, resulting in enhanced cell adhesion in pathogenic E. coli and enhanced biofilm formation in pathogenic and nonpathogenic E. coli. Our findings demonstrate that EsrL stimulates the expression of virulence genes in both EHEC and EPEC under phosphate-rich conditions, thus promoting the pathogenicity of EHEC and EPEC in the nutrient-rich gut environment.

Protection Against Post-resuscitation Acute Kidney Injury by N-Acetylcysteine via Activation of the Nrf2/HO-1 Pathway.

Background and Objective: Acute kidney injury (AKI), the common complication after cardiopulmonary resuscitation (CPR), seriously affects the prognosis of cardiac arrest (CA) patients. However, there are limited studies on post-resuscitation AKI. In addition, it has been demonstrated that N-acetylcysteine (N-AC) as an ROS scavenger, has multiorgan-protective effects on systemic and regional ischaemia-reperfusion injuries. However, no studies have reported its protective effects against post-resuscitation AKI and potential mechanisms. This study aimed to clarify the protective effects of N-AC on post-resuscitation AKI and investigate whether its potential mechanism was mediated by activating Nrf-2/HO-1 pathway in the kidney. Methods: We established cardiac arrest models in rats. All animals were divided into four groups: the sham, control, N-AC, and ZnPP groups. Animals in each group except for the ZnPP group were assigned into two subgroups based on the survival time: 6 and 48 h. The rats in the control, N-AC, and ZnPP groups underwent induction of ventricular fibrillation (VF), 8 min untreated VF and cardiopulmonary resuscitation. Renal function indicators, were detected using commercial kits. Renal pathologic changes were assessed by haematoxylin-eosin (HE) staining. Oxidative stress and inflammatory responses were measured using the corresponding indicators. Apoptosis was evaluated using terminal uridine nick-end labeling (TUNEL) staining, and expression of proteins associated with apoptosis and the Nrf-2/HO-1 pathway was measured by western blotting. Results: N-AC inhibited post-resuscitation AKI. We observed that N-AC reduced the levels of biomarkers of renal function derangement; improved renal pathological changes; and suppressed apoptosis, oxidative stress, and inflammatory response. Additionally, the production of ROS in the kidneys markedly decreased by N-AC. More importantly, compared with the control group, N-AC further upregulated the expression of nuclear Nrf2 and endogenous HO-1 in N-AC group. However, N-AC-determined protective effects on post-resuscitation AKI were markedly reversed after pretreatment of the HO-1 inhibitor zinc protoporphyrin (ZnPP). Conclusions: N-AC alleviated renal dysfunction and prolonged survival in animal models of CA. N-AC partially exerts beneficial renal protection via activation of the Nrf-2/HO-1 pathway. Altogether, all these findings indicated that N-AC as a common clinical agent, may have the potentially clinical utility to improve patients the outcomes in cardiac arrest.

Mapping the research trends and hot topics of ventricular arrhythmia: A bibliometric analysis from 2001 to 2020.

Background: Studies of ventricular arrhythmia (VA) have drawn much scholarly attention over the past two decades. Our study aimed to assess the current situation and detect the changing research trends of VA quantitatively and qualitatively. Materials and methods: All the information used in our statistical and bibliometric analysis were collected and summarized from papers retrieved from the Web of Science Core Collection (WoSCC) database on December 22, 2021 using certain criteria. Visual analytics were realized using CiteSpace, VOSviewer, the bibliometrix R package, and the bibliometric online analysis platform. Results: A total of 6,897 papers (6,711 original articles, 182 proceedings papers, three book chapters, and one data paper) were published in 796 journals that concentrated on the research areas of cardiovascular and critical care medicine. The most productive country and influential institution was the USA and the Mayo Clinic, respectively. Heart Rhythm (551 articles and 8,342 local citations) published the most manuscripts. The keyword co-occurrence and co-citation network of references analyses revealed that the most popular terms were ventricular tachycardia, ventricular fibrillation, catheter ablation, implantable cardioverter defibrillator (ICD), and sudden cardiac death (SCD). Further, the burst detection analysis demonstrated that topics strongly associated with clinical prognosis, such as meta-analysis, long-term outcomes, and impact, were new concerns. Conclusion: Our study offers a comprehensive picture of VA research and provides profound insights into the current research status. Moreover, we show that new topics within the VA research field have focused more on prognosis and evidence-based clinical guidelines.

EFFECTS OF SHENFU INJECTION ON SUBLINGUAL MICROCIRCULATION IN SEPTIC SHOCK PATIENTS: A RANDOMIZED CONTROLLED TRIAL.

ABSTRACT: Background and Objective: The optimization of macrocirculatory hemodynamics is recommended by current sepsis guidelines. However, microcirculatory dysfunction is considered the cause of severe sepsis. In the present study, we designed to verify whether the application of Shenfu injection (SFI) restores microcirculation, thereby improving tissue perfusion and inhibiting organ dysfunction, resulting in improved outcomes. Design: We conducted a prospective, single-center, randomized, double-blind, placebo-controlled clinical trial. Intervention: Patients were randomly assigned to group receiving SFI (n = 20) or placebo (n = 20) for 5 days. We administered SFI or glucose injection for 5 days and blinded the investigators and clinical staff by applying light-proof infusion equipment that concealed therapy allocation. Measurements and Results: We measured the systemic dynamics and lactate levels, biomarkers of endothelial dysfunction, and inflammatory cytokines in the plasma. The parameters of sublingual microcirculation were assessed using side-stream dark-field imaging. Sequential Organ Failure Assessment (SOFA) score, Acute Physiology and Chronic Health Evaluation (APACHE) score, total dose, and duration of vasopressor use, emergency intensive care unit (EICU) stay, and 28-day mortality were evaluated. After treatment with SFI, the disturbance of the sublingual microcirculation was considerably alleviated, as indicated by the significant increase in total vessel density, perfused vessel density, and microvascular flow index. Moreover, the plasma biomarker levels of endothelial dysfunction, including Ang-2, Syn-1, and ET-1, were reversed after SFI treatment. Importantly, the SFI group had a more favorable prognosis than the control group in terms of the APACHE-II score, SOFA score, duration of vasopressor administration, and length of EICU stay. However, the difference in mortality at day 28 was not statistically different between the SFI (15%, 3/20) and placebo (25%, 5/20) groups ( P = 0.693). Conclusions : Shenfu injection provided apparent effects in improving sublingual microcirculatory perfusion in patients with septic shock, and this protection may be related with the inhibition of endothelial dysfunction and vasodilatory effects.

The AhR ligand phthiocol and vitamin K analogs as Pseudomonas aeruginosa quorum sensing inhibitors.

The aryl hydrocarbon receptor (AhR) protein senses microbial-secreted metabolites to trigger the host's innate immune system. The Pseudomonas quinolone signal (PQS) and Mycobacterium tuberculosis (MTb) metabolite phthiocol (Pht) are both ligands of AhR with similar chemical structures. As PQS is an essential quorum-sensing molecule that regulates a wide range of virulence factors in Pseudomonas aeruginosa, we hypothesized that Pht and its analogs are potential P. aeruginosa quorum-sensing inhibitors (QSIs) with immune-modulating functions. In this study, we demonstrated that Pht was able to inhibit the P. aeruginosa pqs QS system and reduce both biofilm formation and the production of pyocyanin. Molecular docking analysis suggested that Pht competes with PQS at the binding site of its receptor, PqsR. An electrophoretic mobility shift assay confirmed the Pht-PqsR interaction and showed that Pht attenuated PqsR from binding to the pqsA promoter. Proteomic analysis showed that synthesis of the key pqs QS proteins decreased upon the addition of Pht to the bacterial cultures. Furthermore, Pht analogs vitamins K1 (Phylloquinone), K2 (Menaquinones), and K3 (Menadione) were also showed to inhibit the P. aeruginosa pqs QS system while able to activate the AhR signaling pathways. Our study suggests that the AhR ligands Pht and its vitamin K analogs are promising QSIs for the alternative treatment of P. aeruginosa infections.

Biosynthesis of UDP-2-acetamido-4-formamido-2,4,6-trideoxy-hexose by WekG, WekE, WekF, and WekD: Enzymes in the Wek pathway responsible for O-antigen Assembly in Escherichia coli O119.

Considering the importance of bacterial glycoconjugates on virulence and host mimicry, there is a need to better understand the biosynthetic pathways of these unusual sugars to identify critical targets involved in bacterial pathogenesis. In this report, we describe the cloning, overexpression, purification, and biochemical characterization of the four central enzymes in the biosynthesis pathway for UDP-2-acetamido-4-formamido-2,4,6-trideoxy-hexose, WekG, WekE, WekF, and WekD. Product peaks from enzyme-substrate reactions were detected by using a combination of capillary electrophoresis (CE) and electrospray ionization-mass spectrometry (ESI-MS). Putative enzyme assignments were provided by protein sequence analysis. Combined with the mass spectrometric characterization of pathway intermediates, we propose a biosynthetic pathway for UDP-2-acetamido-4-formamido-2,4,6-trideoxy-hexose. This process involves C-4, C-6 dehydration, C-4 amination, and formylation. CID-ESI-MSn result confirmed that the final product is a 4 formamido derivative too rather than the 3 formamido derivatives as reported earlier.

Levosimendan Ameliorates Post-resuscitation Acute Intestinal Microcirculation Dysfunction Partly Independent of its Effects on Systemic Circulation: A Pilot Study on Cardiac Arrest in a Rat Model.

BACKGROUND: Cardiac arrest (CA) is recognized as a life-threatening disease; however, the initial resuscitation success rate has increased due to advances in clinical treatment. Levosimendan has shown potential benefits in CA patients. However, its exact function on intestinal and systemic circulation in CA or post-cardiac arrest syndrome (PCAS) remained unclear. This study preliminarily investigated the link between dynamic changes in intestine and systemic hemodynamics post-resuscitation after levosimendan administration. METHODS: Twenty-five rats were randomized into three groups: sham control group (n = 5), levosimendan group (n = 10), and vehicle group (n = 10). Intestinal microcirculation was observed using a sidestream dark-field imaging device at baseline and each hour of the return of spontaneous circulation (≤6 h). Systemic hemodynamics, serum indicators of cardiac injury, and tissue perfusion/metabolism were measured by echo-cardiography, a biological signal acquisition system, and an enzyme-linked immunosorbent assay, respectively. RESULTS: Myocardial injury and global and intestinal perfusion/metabolism were significantly improved by levosimendan treatment. There was no statistically significant difference in the mean arterial pressure values between the vehicle and levosimendan groups (P > 0.05). The intestinal and systemic circulation measurements showed poor correlation (Pearson r-value of variable combinations in the levosimendan group was much less than 0.75; P < 0.01, levosimendan vs. vehicle group). CONCLUSIONS: Levosimendan significantly reduced the cardiac injury and corrected the metabolic status in an experimental rat model of ventricular fibrillation induced CA and cardiopulmonary resuscitation. Levosimendan may ameliorate PCAS-induced intestinal microcirculation dysfunction, partly independent of its effects on macrocirculation.

In silico species identification and serotyping for Cronobacter isolates by use of whole-genome sequencing data.

Cronobacter spp. are foodborne pathogens that can cause severe infections in neonates through contaminated powdered infant formula. Accurate and rapid pathogen identification and serotyping are crucial to limit the detrimental effects of bacterial infections, and to prevent outbreaks and sporadic infections. Conventional serotyping is tedious, laborious, and time-consuming; however, with whole-genome sequencing (WGS) becoming faster and cheaper, WGS has vast potential in routine typing and surveillance. Hence, in this study, we developed a publicly available tool, CroTrait (CronobacterTraits), for in silico species identification and O serotyping of Cronobacter isolates based on WGS data. CroTrait showed excellent performance in species identification and O serotyping when 810 genomes with known species identities and 276 genomes with known O serotype were tested. Moreover, CroTrait allows rapid prediction of new potential O serotypes. We identified 11 novel potential O serotypes of Cronobacter using CroTrait. Therefore, CroTrait is a convenient and promising tool for species identification and O serotyping of Cronobacter isolates.

A Novel Small RNA Promotes Motility and Virulence of Enterohemorrhagic Escherichia coli O157:H7 in Response to Ammonium.

Enterohemorrhagic Escherichia coli serotype O157:H7 (O157) is a critical, foodborne, human intestinal pathogen that causes severe acute hemorrhagic diarrhea, abdominal cramping, and even death. Small RNAs (sRNAs) are noncoding regulatory molecules that sense environmental changes and trigger various virulence-related signaling pathways; however, few such sRNAs have been identified in O157. Here, we report a novel sRNA, EsrF that senses high ammonium concentrations in the colon and enhances O157 pathogenicity by promoting bacterial motility and adhesion to host cells. Specifically, EsrF was found to directly interact with the 5' untranslated regions of the flagellar biosynthetic gene, flhB, mRNA and increase its abundance, thereby upregulating expression of essential flagellar genes, including flhD, flhC, fliA, and fliC, leading to elevated O157 motility and virulence. Meanwhile, an infant rabbit model of O157 infection showed that deletion of esrF and flhB significantly attenuates O157 pathogenicity. Furthermore, NtrC-the response regulator of the NtrC/B two-component system-was found to exert direct, negative regulation of esrF expression. Meanwhile, high ammonium concentrations in the colon release the inhibitory effect of NtrC on esrF, thereby enhancing its expression and subsequently promoting bacterial colonization in the host colon. Our work reveals a novel, sRNA-centered, virulence-related signaling pathway in O157 that senses high ammonium concentrations. These findings provide novel insights for future research on O157 pathogenesis and targeted treatment strategies.IMPORTANCE The process by which bacteria sense environmental cues to regulate their virulence is complex. Several studies have focused on regulating the expression of the locus of enterocyte effacement pathogenicity island in the typical gut pathogenic bacterium, O157. However, few investigations have addressed the regulation of other virulence factors in response to intestinal signals. In this study, we report our discovery of a novel O157 sRNA, EsrF, and demonstrate that it contributed to bacterial motility and virulence in vitro and in vivo through the regulation of bacterial flagellar synthesis. Furthermore, we show that high ammonium concentrations in the colon induced esrF expression to promote bacterial virulence by releasing the repression of esrF by NtrC. This study highlights the importance of sRNA in regulating the motility and pathogenicity of O157.

Attachment of Enterohemorrhagic Escherichia coli to Host Cells Reduces O Antigen Chain Length at the Infection Site That Promotes Infection.

Many enteropathogenic bacteria express a needle-like type III secretion system (T3SS) that translocates effectors into host cells promoting infection. O antigen (OAg) constitutes the outer layer of Gram-negative bacteria protecting bacteria from host immune responses. Shigella constitutively shortens the OAg molecule in its three-dimensional conformation by glucosylation, leading to enhanced T3SS function. However, whether and how other enteropathogenic bacteria shorten the OAg molecule that probably facilitates infection remain unknown. For the first time, we report a smart mechanism by which enterohemorrhagic Escherichia coli specifically reduces the size of the OAg molecule at the infection site upon sensing mechanical signals of intestinal epithelial cell attachment via the membrane protein YgjI. YgjI represses expression of the OAg chain length regulator gene fepE via the global regulator H-NS, leading to shortened OAg chains and injection of more T3SS effectors into host cells. However, bacteria express long-chain OAg in the intestinal lumen benefiting their survival. Animal experiments show that blocking this regulatory pathway significantly attenuates bacterial virulence. This finding enhances our understanding of interactions between the surfaces of bacterial and host cells and the way this interaction enhances bacterial pathogenesis. IMPORTANCE Little is known about the regulation of cell wall structure of enteropathogenic bacteria within the host. Here, we report that enterohemorrhagic Escherichia coli regulates its cell wall structure during the infection process, which balances its survival in the intestinal lumen and infection of intestinal epithelial cells. In the intestinal lumen, bacteria express long-chain OAg, which is located in the outer part of the cell wall, leading to enhanced resistance to antimicrobial peptides. However, upon epithelial cell attachment, bacteria sense this mechanical signal via a membrane protein and reduce the OAg chain length, resulting in enhanced injection into epithelial cells of T3SS effectors that mediate host cell infection. Similar regulation mechanisms of cell wall structure in response to host cell attachment may be widespread in pathogenic bacteria and closely related with bacterial pathogenesis.

The K90 capsular polysaccharide produced by Acinetobacter baumannii LUH5553 contains di-N-acetylpseudaminic acid and is structurally related to the K7 polysaccharide from A. baumannii LUH5533.

Acinetobacter baumannii isolate LUH5553 carries the KL90 capsule gene cluster, which includes genes for three glycosyltransferases (Gtrs) and the ItrA3 initiating transferase, as well as a set of genes for synthesis of a higher sugar, 5,7-diacetamido-3,5,7,9-tetradeoxy-l-glycero-l-manno-non-2-ulosonic (di-N-acetylpseudaminic) acid (Pse5Ac7Ac). The K90 capsular polysaccharide (CPS) has a tetrasaccharide repeat (K90 unit), which begins with d-GlcpNAc and contains Pse5Ac7Ac. The higher sugar was cleaved by mild acid hydrolysis of the CPS, and structures of the initial and modified polysaccharides were established by 1D and 2D 1H and 13C NMR spectroscopy. K90 contains α-d-Galp-(1 → 6)-d-GlcpNAc and α-d-GlcpNAc-(1 → 3)-d-GlcpNAc fragments, and formation of these glycosidic linkages is catalysed respectively by Gtr14 and Gtr15. The gtr14 and gtr15 genes occur in several A. baumannii KL gene clusters, including KL5 and KL7 that carry itrA2 rather than itrA3. As ItrA2 introduces d-GalpNAc rather than d-GlcpNAc as the first monosaccharide, Gtr15 can transfer d-GlcpNAc to either of these amino sugars, suggesting that this enzyme has relaxed specificity. Consequently, the third, novel glycosyltransferase, Gtr163, forms the ß-(2 → 3) linkage between Pse5Ac7Ac and d-Galp. Wzy polymerases encoded by KL90 and KL7 are 54% identical and form the same linkage between the K units to give branched polysaccharides with the same main chain but different disaccharide side chains, ß-Pse5Ac7Ac-(2 → 3)-d-Galp in K90 and α-Leg5Ac7Ac-(2 → 6)-d-Galp in K7.