MEET: A Multi-Band EEG Transformer for Brain States Decoding.

OBJECTIVE: Electroencephalography (EEG) is among the most widely used and inexpensive neuroimaging techniques. Compared to the CNN or RNN based models, Transformer can better capture the temporal information in EEG signals and focus more on global features of the brain's functional activities. Importantly, according to the multiscale nature of EEG signals, it is crucial to consider the multi-band concept into the design of EEG Transformer architecture. METHODS: We propose a novel Multi-band EEG Transformer (MEET) to represent and analyze the multiscale temporal time series of human brain EEG signals. MEET mainly includes three parts: 1) transform the EEG signals into multi-band images, and preserve the 3D spatial information between electrodes; 2) design a Band Attention Block to compute the attention maps of the stacked multi-band images and infer the fused feature maps; 3) apply the Temporal Self-Attention and Spatial Self-Attention modules to extract the spatiotemporal features for the characterization and differentiation of multi-frame dynamic brain states. RESULTS: The experimental results show that: 1) MEET outperforms state-of-the-art methods on multiple open EEG datasets (SEED, SEED-IV, WM) for brain states classification; 2) MEET demonstrates that 5-bands fusion is the best integration strategy; and 3) MEET identifies interpretable brain attention regions. SIGNIFICANCE: MEET is an interpretable and universal model based on the multiband-multiscale characteristics of EEG. CONCLUSION: The innovative combination of band attention and temporal/spatial self-attention mechanisms in MEET achieves promising data-driven learning of the temporal dependencies and spatial relationships of EEG signals across the entire brain in a holistic and comprehensive fashion.

MM-GLCM-CNN: A multi-scale and multi-level based GLCM-CNN for polyp classification.

Distinguishing malignant from benign lesions has significant clinical impacts on both early detection and optimal management of those early detections. Convolutional neural network (CNN) has shown great potential in medical imaging applications due to its powerful feature learning capability. However, it is very challenging to obtain pathological ground truth, addition to collected in vivo medical images, to construct objective training labels for feature learning, leading to the difficulty of performing lesion diagnosis. This is contrary to the requirement that CNN algorithms need a large number of datasets for the training. To explore the ability to learn features from small pathologically-proven datasets for differentiation of malignant from benign polyps, we propose a Multi-scale and Multi-level based Gray-level Co-occurrence Matrix CNN (MM-GLCM-CNN). Specifically, instead of inputting the lesions' medical images, the GLCM, which characterizes the lesion heterogeneity in terms of image texture characteristics, is fed into the MM-GLCN-CNN model for the training. This aims to improve feature extraction by introducing multi-scale and multi-level analysis into the construction of lesion texture characteristic descriptors (LTCDs). To learn and fuse multiple sets of LTCDs from small datasets for lesion diagnosis, we further propose an adaptive multi-input CNN learning framework. Furthermore, an Adaptive Weight Network is used to highlight important information and suppress redundant information after the fusion of the LTCDs. We evaluated the performance of MM-GLCM-CNN by the area under the receiver operating characteristic curve (AUC) merit on small private lesion datasets of colon polyps. The AUC score reaches 93.99% with a gain of 1.49% over current state-of-the-art lesion classification methods on the same dataset. This gain indicates the importance of incorporating lesion characteristic heterogeneity for the prediction of lesion malignancy using small pathologically-proven datasets.

Jian-Pi-Yi-Shen formula restores iron metabolism from dysregulation in anemic rats with adenine-induced nephropathy.

ETHNOPHARMACOLOGICAL RELEVANCE: Jian-Pi-Yi-Shen (JPYS) is a herbal decoction being used to relieve the symptoms of chronic kidney disease (CKD) and its complications, including anemia, for over twenty years. Nonetheless, it is unclear how JPYS influences renal anemia and iron metabolism. AIM OF THE STUDY: An analysis of network pharmacology, chemical profiling, and in vivo experiments was conducted to identify the impact of JPYS on JAK2-STAT3 pathway and iron utilization in renal anemia and CKD. MATERIALS AND METHODS: The chemical properties of JPYS and its exposed ingredients were detected in vivo. And based on the aforesaid chemical compounds, the potential targets and signaling pathways of JPYS for renal anemia treatment were predicted by network pharmacology. Afterward, an adenine-feeding animal model of CKD-related anemia was developed to verify the mechanism by which JPYS modulates iron recycling to treat renal anemia. Renal injury was estimated by serum creatinine (Scr), blood urea nitrogen (BUN), histopathological examinations and fibrosis degree. Western blot, enzyme-linked immunosorbent assay (ELISA), quantitative real-time polymerase chain reaction (qPCR) and immunohistochemistry approaches were utilized to assess the levels of JAK2, STAT3 and iron metabolism-related factors. RESULTS: There were 164 active ingredients identified in JPYS, including prototypes and metabolites in vivo were identified in JPYS, and 21 core targets were found through network pharmacology based on topological characteristics. Combined with the core targets and pathway enrichment analysis, the majority of the candidate targets were associated with the JAK2-STAT3 signaling pathways. Experimental results indicated that JPYS treatment significantly decreased the expression of BUN and Scr, restored renal pathological damage, down-regulated fibrosis degree, and improved hematological parameters such as red blood cell, hemoglobin and hematocrit in CKD rats. Furthermore, JPYS significantly restored iron metabolism from dysregulation by increasing the levels of iron and ferritin in the serum, inhibiting the production of hepcidin in liver and serum, and regulating transferrin receptor 1 in bone marrow. Meanwhile, the expression of JAK2 and STAT3 was suppressed by JPYS treatment. CONCLUSIONS: Based on these results, JPYS reduces hepcidin levels by inhibiting the activation of JAK2-STAT3 signaling, thereby protecting against iron deficiency anemia.

Effectiveness of early glucocorticoids in myasthenia gravis: a retrospective cohort study.

Purpose: This study aimed to clarify the effect of early glucocorticoid (GC) application on achieving minimal manifestation (MM) status or better in the treatment of myasthenia gravis (MG) in the early clinical phase. Methods: A retrospective analysis was performed using data from 336 patients with MG who received GC therapy from January 2015 to September 2022 in the Zhengzhou University Henan Institute of Medical and Pharmaceutical Sciences Myasthenia Gravis Biobank (ZMB). Patients were divided into two groups: the early mono-GC group (treated with GC within 6 months of MG onset) and the delayed mono-GC group. Results: Kaplan-Meier analysis showed that the early mono-GC group achieved MM status earlier and more frequently than the delayed mono-GC group (log-rank test, p = 0.0082; hazard ratio [HR], 1.66; p = 0.011). The early mono-GC group had a lower maintenance oral GC dose than the delayed mono-GC group. In multivariate Cox regression analysis, early mono-GC (HR, 1.50; p = 0.043), early-onset MG (EOMG) (HR, 1.74; p = 0.034), and ocular MG (OMG) (HR, 1.90; p = 0.007) were associated with MM status or better. In conclusion, early mono-GC, EOMG, and OMG were positive predictors of treatment goals. In EOMG, OMG, and acetylcholine receptor antibody-positive MG (AChR-MG) subgroups, the maintenance oral GC doses in the early mono-GC group were significantly lower than the doses in the delayed mono-GC group (p < 0.05). Conclusion: Early intervention with GC led to better long-term outcomes and reduced the necessary maintenance dose of oral GC for patients with MG. EOMG and OMG were positive predictors of MM status or better with mono-GC.

Differentiate preterm and term infant brains and characterize the corresponding biomarkers via DICCCOL-based multi-modality graph neural networks.

Preterm birth is a worldwide problem that affects infants throughout their lives significantly. Therefore, differentiating brain disorders, and further identifying and characterizing the corresponding biomarkers are key issues to investigate the effects of preterm birth, which facilitates the interventions for neuroprotection and improves outcomes of prematurity. Until now, many efforts have been made to study the effects of preterm birth; however, most of the studies merely focus on either functional or structural perspective. In addition, an effective framework not only jointly studies the brain function and structure at a group-level, but also retains the individual differences among the subjects. In this study, a novel dense individualized and common connectivity-based cortical landmarks (DICCCOL)-based multi-modality graph neural networks (DM-GNN) framework is proposed to differentiate preterm and term infant brains and characterize the corresponding biomarkers. This framework adopts the DICCCOL system as the initialized graph node of GNN for each subject, utilizing both functional and structural profiles and effectively retaining the individual differences. To be specific, functional magnetic resonance imaging (fMRI) of the brain provides the features for the graph nodes, and brain fiber connectivity is utilized as the structural representation of the graph edges. Self-attention graph pooling (SAGPOOL)-based GNN is then applied to jointly study the function and structure of the brain and identify the biomarkers. Our results successfully demonstrate that the proposed framework can effectively differentiate the preterm and term infant brains. Furthermore, the self-attention-based mechanism can accurately calculate the attention score and recognize the most significant biomarkers. In this study, not only 87.6% classification accuracy is observed for the developing Human Connectome Project (dHCP) dataset, but also distinguishing features are explored and extracted. Our study provides a novel and uniform framework to differentiate brain disorders and characterize the corresponding biomarkers.

GLIM criteria using NRS-2002 and MUST as the first step adequately diagnose the malnutrition in Crohn's disease inpatients: A retrospective study.

Objective: The Global Leader Initiative on Malnutrition (GLIM) criteria have been recommended for malnutrition diagnosis recently, for which the first step is malnutrition risk screening with any validated tool. This study aims to investigate the incidence of nutritional risk and malnutrition in Crohn's disease inpatients and compare the suitability of Nutritional Risk Screening 2002 (NRS-2002) and Malnutrition Universal Screening Tool (MUST) as the first-step screening tool for GLIM criteria. Methods: We retrospectively analyzed the clinical data of Crohn's disease inpatients in our hospital from August 2016 to December 2019. NRS-2002 and MUST were used for nutritional screening at the time of admission. GLIM and Patient Generated-Subjective Global Assessment (PG-SGA) were used for malnutrition assessment, respectively. Patients without nutritional risk screened by NRS-2002 but with malnutrition risk screened by MUST were especially screened out. The appendicular skeletal muscle mass index (ASMI), fat-free mass index (FFMI), body fat percent (BFP), and body cell mass (BCM) were measured by the Biospace Inbody S10 composition analyzer. Results: A total of 146 Crohn's disease patients were enrolled, of which 62.3 and 89.7% had nutritional or malnutrition risk according to NRS-2002 and MUST, respectively. The prevalence of malnutrition assessed by GLIM was 59.6% (87 cases) and 82.2% (120 cases) when NRS-2002 and MUST were used as the first step of GLIM respectively. Meanwhile, 99 patients (67.8%) had malnutrition when assessed by PG-SGA. There were 41 patients who were not at nutritional risk according to NRS-2002 but were at malnutrition risk determined by MUST. At last, 33 patients were GLIM-defined, and 16 patients were PG-SGA-defined malnutrition among the 41 patients. Conclusion: The nutritional risk or malnutrition is common in Crohn's disease inpatients. It is recommended to use a variety of nutritional assessment tools for Crohn's disease inpatients. MUST can be used as a good supplement for the patients with a score of NRS-2002 lower than 3 in order to decrease the miss rate of GLIM-defined malnutrition.

Relationships Among Pre-Pregnancy BMI, Gestational, and Postpartum Oral Glucose Tolerance Results in Women With Gestational Diabetes Mellitus.

Background and Aims: To investigate the relationship among maternal demographic and clinical characteristics, gestational and postpartum oral glucose tolerance test (ppOGTT) results in patients with gestational diabetes mellitus (GDM). Methods: Patients with gestational diabetes mellitus from January 1, 2016, to August 31, 2019, were enrolled. General characteristics, dietary energy intake, pre-gestational body mass index (BMI), gestational oral glucose tolerance test (gOGTT), and 42 days ppOGTT results of all participants were collected. The relationships among maternal clinical characteristics, fasting glucose of gOGTT (gOGTT-FPG), 1 h postprandial glucose of gOGTT (gOGTT-1h PG), 2 h postprandial glucose of gOGTT (gOGTT-2h PG), and maternal postpartum glucose outcomes were evaluated. Results: A total of 156 patients with GDM were included in this study. Among them, 73.7% had inadequate daily total energy intake, an insufficient ratio of carbohydrates and protein, and an excessive fat ratio. Most of the patients (81.4%) were normal in their ppOGTT examination. Less than 20% of the patients (16.7%) were in the pre-diabetic situation, and 3 patients (1.9%) had diabetes. Pre-pregnancy BMI of patients with GDM was a risk factor for increased gOGTT-FPG levels. Those who were overweight before pregnancy had a greater risk for a higher gOGTT-FPG compared to those who had normal pre-pregnancy BMI (P = 0.021, odds ratio [OR] = 4.583). Abnormal gOGTT-2hPG was a risk factor for abnormal ppOGTT (P = 0.04). Those who had an elevated gOGTT-2hPG (≧8.5 mmol/L) had a 2.426 times higher risk for abnormal ppOGTT than those who had normal gOGTT-2hPG (<8.5 mmol/L) results. Conclusion: For women who are overweight before pregnancy, it is better to control their BMI to normal before getting pregnant. Women who had abnormal gOGTT-2h PG should pay more attention to the ppOGTT results.

Role of Muscle Mass and Nutritional Assessment Tools in Evaluating the Nutritional Status of Patients With Locally Advanced Nasopharyngeal Carcinoma.

Objective: This study was to explore the role and necessity of muscle mass [fat-free mass index (FFMI) and appendicular skeletal muscle index (ASMI) measured by bioelectrical impedance analysis (BIA)] in nutritional status evaluation of patients with locally advanced (III, IVa) nasopharyngeal carcinoma (NPC). Methods: One hundred and thirty locally advanced NPC patients were recruited. Their nutritional status was assessed by albumin (ALB), body mass index (BMI), Nutritional Risk Screening 2002 (NRS 2002), Patient generated-Subjective Global Assessment (PG-SGA), and muscle mass. Consistency test and McNemar test were used to evaluate the consistency of muscle mass with ALB, BMI, NRS 2002, and PG-SGA, and correlation analysis was performed on muscle mass and PG-SGA or BMI. Results: 61/130 (46.9%) of the patients had nutritional risks according to NRS 2002, 68/130 (53.1%) of the patients had malnutrition according to PG-SGA assessment. FFMI and ASMI could determine the loss of muscle mass that cannot be detected by albumin (30.2 and 65.6%), BMI (28.0 and 35.3%), NRS 2002 (26.1 and 25.0%), and PG-SGA (18.6 and 55.6%). McNemar test showed that the malnutrition results assessed by FFMI and BMI were inconsistent (P <0.001), but further Pearson correlation analysis showed that BMI was positively correlated with FFMI (rs = 0.300, P = 0.001). Conclusion: The commonly used nutritional assessment scale/parameters cannot identify the muscle mass loss in patients with locally advanced NPC. Analysis of human body composition is important for nutritional assessment in patients with locally advanced NPC.

Identification of RT-PCR-Negative Asymptomatic COVID-19 Patients via Serological Testing.

Asymptomatic individuals with coronavirus disease (COVID-19) have been identified via nucleic acid testing for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2); however, the epidemiologic characteristics and viral shedding pattern of asymptomatic patients remain largely unknown. In this study, serological testing was applied when identifying nine asymptomatic cases of COVID-19 who showed persistent negative RT-PCR test results for SARS-CoV-2 nucleic acid and no symptoms of COVID-19. Two asymptomatic cases were presumed to be index patients who had cleared the virus when their close contacts developed symptoms of COVID-19. Three of the asymptomatic cases were local individuals who spontaneously recovered before their presumed index patients developed symptoms of COVID-19. This report presents the epidemiologic and clinical characteristics of asymptomatic individuals with SARS-CoV-2 infection that were undetected on RT-PCR tests in previous epidemiologic investigations probably due to the transient viral shedding duration.

Role of Human Body Composition Analysis and Malnutrition Risk Questionnaire in the Assessment of Nutritional Status of Patients With Initially Diagnosed Crohn's Disease.

Objective: This study was carried out to investigate the role and necessity of human body composition analysis in assessing the nutritional status of initially diagnosed Crohn's disease (CD) patients. Methods: A total of 47 initially diagnosed CD patients were recruited. The skeletal muscle mass index (SMI), fat-free mass index (FFMI), body fat mass, body fat percent, visceral fat area (VFA), and body cell mass were determined with the Biospace Inbody S10 composition analyzer. Results: In 47 patients with initially diagnosed CD, SMI could determine the muscular mass reduction that could not be determined by the body mass index (BMI) (35.3%), albumin (ALB) (65.6%), nutrition risk screening (NRS)2002 (25.0%), and Patient-Generated Subjective Global Assessment (PG-SGA) (55.6%). FFMI could determine the malnutrition that could not be determined by the BMI (58.8%), albumin (90.6%), NRS2002 (50.0%), and PG-SGA (55.6%). VFA in the fistulizing CD patients was significantly higher than in the stricturing and non-fistulizing, non-stricturing patients (P < 0.05). SMI and BMI had the same performance (P = 1.000) and general consistence (Kappa = 0.487, P = 0.001) in the assessment of malnutrition; SMI and ALB had different performance (P < 0.001) and inconsistence was noted (Kappa = 0.069, P = 0.489) in the assessment of malnutrition; the results of the nutrition assessment were different between SMI and NRS2002 (P = 0.002), and inconsistence was observed (Kappa = 0.190, P = 0.071). SMI and PG-SGA had the same performance in the assessment of nutrition (P = 0.143), but there was inconsistence (Kappa = 0.099, P = 0.464). FFMI and BMI had general consistence in the assessment of malnutrition (Kappa = 0.472, P < 0.001), but the positive rate determined by FFMI (85.1%) was markedly higher than that by BMI (63.8%) (P = 0.002). FFMI and ALB had different performance in the assessment of malnutrition (P < 0.001) and there was inconsistence (Kappa = -0.008, P = 0.877). FFMI and NRS2002 had the same performance in the assessment of malnutrition (P = 0.453), but the consistence was poor (Kappa = 0.286, P = 0.039). The results determined by SMI and PG-SGA were consistent (P = 0.727), but the consistence was poor (Kappa = 0.399, P = 0.006). Conclusion: Human body composition analysis can identify the patients with muscular mass reduction that cannot be identified by commonly used nutrition assessment scales/parameters. Thus, it is helpful for the assessment of disease severity and also important for the nutrition assessment in CD patients.

Identification of the first case of SFTSV infection in the Hunan Province of China and epidemiological surveillance in the locality.

This study reports the etiological identification, clinical diagnosis, and the results of the local epidemiological surveillance of the first case of severe fever with thrombocytopenia syndrome virus (SFTSV) infection in 2014 in Hunan Province, China. The infected patient was isolated and closely monitored. The virus is a member of the Bunyaviridae sandfly family and is characterized by real-time PCR, electron microscopy, immunofluorescence, and whole-genome sequencing. We also detected IgG and IgM antibodies against SFTSV among the local human population and domestic animals in a serological surveillance. Prevalence of SFTSV-specific antibodies was monitored in the local population for two years after the identification of the first SFTS case. Approximately 5% (4/77) of the people who had direct contact with the patient were seropositive, which is significantly higher than the seropositivity of the general local population [1.57% (44/2800), P < 0.05]. Furthermore, the percentage of the general population who were seropositive was higher in 2015 than in 2014 (χ2 = 7.481, P = 0.006). The epidemiological investigation found that the SFTSV is epidemic in goats, cattle, and chickens in Hunan Province. The risk of infection of domestic animals can be minimized by feeding in pens rather than allowing foraging.

Polymerase chain reaction-restriction fragment length polymorphism analysis of the genes involved in the biosynthesis of the lipopolysaccharide of Pasteurella multocida.

The lipopolysaccharide, also known as the somatic antigen or O-antigen, is an important virulence factor of Pasteurella multocida. In the current study, the genes involved in the biosynthesis of the outer core region of the lipopolysaccharide, which were obtained from somatic type reference strains and field strains of P. multocida, were subjected to polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis. The PCR-RFLP analysis classified 11 out of the 16 serotypes into 5 PCR-RFLP types (I-V). Types I and V contain strains belong to serotypes 1 and 13, respectively. The rest of the PCR-RFLP types contain strains belong to certain groups of serotypes. Typing of 38 field strains from poultry using PCR-RFLP analysis and the gel diffusion precipitation test showed consistent results. These results indicate that the PCR-RFLP analysis can be a useful tool for rapid somatic typing of some strains of P. multocida.

Recombinant proteins containing the hypervariable region of the haemagglutinin protect chickens against challenge with Avibacterium paragallinarum.

The haemagglutinin (HA) protein plays a key role in the immunogenicity and pathogenicity of Avibacterium paragallinarum, but the domain organization and antigenicity exhibited by different domains of this protein remain unknown. This study reports the presence of a hypervariable region in the HA proteins of strains of serovars A and C of A. paragallinarum. This hypervariable region is located approximately at residues 1100-1600 of the HA protein. The sequence identity found in this hypervariable region was only 18.1%, whereas those upstream and downstream of this region were 83.8 and 97.8%, respectively. Western blot analyses using antisera against the whole-cell antigens of A. paragallinarum showed that the hypervariable region was more antigenic than other regions of the HA protein. Moreover, the antigenicity of the hypervariable region was serovar-specific. Chickens immunized with recombinant proteins that contained the hypervariable region were protected (83-100% protection rate) against challenge infection with A. paragallinarum of the homologous serovar. These results suggest that recombinant proteins containing the hypervariable region may be useful antigens for use in the development of a vaccine against A. paragallinarum.

Analysis of the biosynthesis genes and chemical components of the capsule of Avibacterium paragallinarum.

The aim of this study was to investigate biosynthesis genes and chemical components of the capsule of Avibacterium paragallinarum. The sequence of a 10-kb region containing the capsule biosynthetic locus of Av. paragallinarum was determined. Two reference strains, i.e., 221 (serovar A) and H18 (serovar C), together with four Taiwanese field strains (all serovar C) were sequenced. The results showed that there are two genotypes (I and II) of the capsule biosynthetic locus in Av. paragallinarum, and the capsule genotype is independent of the serovar. The capsule biosynthetic loci of genotypes I and II consisted of six and five genes, respectively. The genotype I genes encoded proteins that are most similar to proteins from Pasteurella multocida capsule types A and F while the genotype II genes encoded proteins most similar to proteins from P. multocida capsule type D and Escherichia coli K5. The results suggested that genotype I strains contain hyaluronan or chondroitin in the capsule wall while genotype II contain heparosan. Enzymatic digestion of the capsule materials extracted from Av. paragallinarum showed that genotype I strains contained chondroitin while genotype II strains contained heparosan in the capsule. This is the first report on the existence of different genotypes of capsule biosynthesis genes in Av. paragallinarum and the presence of chondroitin and heparosan as chemical components of the capsule of Av. paragallinarum.

Development of blocking ELISA for detection of antibodies against avian influenza virus of the H7 subtype.

BACKGROUND AND PURPOSE: The conventional method used for subtyping of antibodies against avian influenza viruses is hemagglutination inhibition (HI) test. However, the HI test is laborious and requires preparation of antigen from viable viruses that might be hazardous. The aim of this study was to develop a blocking enzyme-linked immunosorbent assay (B-ELISA) for detection of antibody of avian influenza of the H7 subtype. The B-ELISA is fast and avoids the need to culture whole viruses. METHODS: The B-ELISA was based on the reaction between a monoclonal antibody and a recombinant hemagglutinin protein purified from Escherichia coli. The specificity of the B-ELISA was determined by testing H7-negative field sera and the sensitivity of the B-ELISA was determined by testing sera collected from experimentally immunized chickens. RESULTS: The specificity of the B-ELISA was found to be 97.7% when compared with the HI test. The sensitivity was found to vary with the HI titer of sera. A sensitivity of 100% was achieved when test sera had HI titers >or=2(7). The sensitivity dropped to 33% and 20% when test sera had HI titers of 2(6) and 2(5), respectively. Nearly all test sera with HI titers

Protective immunity conferred by recombinant Pasteurella multocida lipoprotein E (PlpE).

The genes encoding Pasteurella multocida lipoprotein E (PlpE) and lipoprotein B (PlpB) were cloned from P. multocida strain X-73 (serotype A:1) and expressed in Escherichia coli. The protective immunity conferred by recombinant PlpE (r-PlpE) and PlpB (r-PlpB) on mice and chickens was evaluated. The results showed that mice immunized with 10microg of purified r-PlpE were protected (80-100% survival rate) against challenge infection with 10 or 20 LD(50) of P. multocida strains X-73 (serotype A:1), P-1059 (serotype A:3) and P-1662 (serotype A:4). In contrast, mice immunized with r-PlpB were not protected. Chickens immunized with 100microg of purified r-PlpE were protected (63-100% survival rate) against lethal challenge infection with strains X-73 and P-1662, whereas those immunized with r-PlpB were not. Sequence analyses showed that PlpE from different strains of P. multocida exhibited 90.8-100% sequence identity to each other, suggesting that PlpE might serve as a cross-protective antigen. This is the first report of a recombinant P. multocida antigen that confers cross protection on animals.

Cloning of the gene and characterization of the enzymatic properties of the monomeric alkaline phosphatase (PhoX) from Pasteurella multocida strain X-73.

We have identified a new phoX gene encoding the monomeric alkaline phosphatase from Pasteurella multocida X-73. This gene was not found in the published genome sequence of Pasteurella multocida pm70. Characterization of the recombinant PhoX of Pasteurella multocida X-73 showed that it is a monomeric enzyme, activated by Ca(2+) and possibly secreted by the Tat pathway. These features distinguish phosphatases of the PhoX family from those of the PhoA family. All proteins of the PhoX family were found to contain a conserved motif that shares significant sequence homology with the calcium-binding site of a phosphotriesterase known as diisopropylfluorophosphatase. Site-directed mutagenesis revealed that D527 of PhoX might be the ligand bound to the catalytic calcium. This is the first report on identification of homologous sequences between PhoX and the phosphotriesterase and on the potential calcium-binding site of PhoX.

Molecular characterization of plasmids with antimicrobial resistant genes in avian isolates of Pasteurella multocida.

The complete nucleotide sequences of two plasmids from avian isolates of Pasteurella multocida that caused outbreaks of fowl cholera in Taiwan were determined. The entire sequences of the two plasmids, designated as pJR1 and pJR2, were 6792 bp and 5252 bp. Sequence analysis showed that the plasmid pJR1 contained six major genes: the first gene (sulII) encoded a type II sulfonamide resistant dihydropteroate synthase, the second gene (tetG) encoded a tetracycline resistance protein, the third gene (catB2) encoded a chloramphenicol acetyltransferase, the fourth gene (rep) encoded a replication protein, and the fifth and sixth genes (mbeCy and deltambeAy) encoded proteins involved in the mobilization of plasmid. The plasmid pJR2 contained five major genes: the first gene (deltaintI1) encoded a truncated form of a type I integrase, the second gene (aadA1) encoded an aminoglycoside adenylyltransferase that confers resistance to streptomycin and spectinomycin, the third gene (blaP1) encoded a beta-lactamase that confers resistance to ampicillin and carbenicillin, and the fourth and fifth genes might encode proteins involved in the plasmid replication or segregation. Sequence comparisons showed that the antibiotic resistance genes found in pJR1 and pJR2 exhibited a high degree of sequence homology to the corresponding genes found in a great variety of gram-negative bacteria, including Escherichia coli, Salmonella enterica Typhimurium DT104, Psedomonas spp., P. multocida, Mannheimia spp., and Actinobacills pleuropneumoniae, which suggests that these resistance genes were disseminated in these bacteria. Although sulII and tetG genes were found previously in P. multocida or Mannheimia spp., this is the first report on the presence of catB2, aadA1, and blaP1 genes in bacteria of the family Pasturellaceae. Moreover, the aadA1 and blaP1 genes found in pJR2 were organized into an integron structure, which is a site-specific recombination system capable of capturing and mobilizing antibiotic resistance genes. This is also the first report on the presence of an integron in bacteria of the family Pasteurellaceae. The presence of a P. multocida integron might facilitate the spreading of antibiotic resistance genes between P. multocida and other gram-negative bacteria.