Preparation,identification and comparison of murine and rabbit polyclonal antibodies against matrix protein of rabies virus strain CVS-11 / 中国生物制品学杂志

@#Objective To prepare murine and rabbit polyclonal antibodies against rabies virus(RV) matrix(M) protein and compare their reactivity.Methods The prokaryotic expression vector pET-28a-M was constructed by using the cDNA of cells infected with RV CVS-11 strain as template,then transformed into E.coli BL21(DE3),and the induced by IPTG to express M protein.After nickel column affinity chromatography and dialysis renaturation,female BALB/c mice and New Zealand white rabbits were immunized with the M protein,and the whole blood was taken to separate the serum.The titers of the murine and rabbit polyclonal antibodies were detected by ELISA,and the reactivity was measured by Western blot,indirect immunofluorescence assay(IFA) and immunoprecipitation(IP).Results The plasmid pET-28a-M was constructed correctly as identified by sequencing.The titers of murine and rabbit polyclonal antibodies were 1:100 and 1:256 000respectively,and the polyclonal antibodies had reactivity with different RV strains.Conclusion The murine and rabbit polyclonal antibodies against M protein were successfully prepared,which provides important biological tools for exploring the interaction between M protein and host protein as well as studying the pathogenesis of RV.

Analysis of drug resistance gene variation of influenza A（H1N1） pdm09 virus in Huzhou City，Zhejiang Province / 上海预防医学

ObjectiveTo investigate the mutation and genetic evolution of drug resistance gene of A（H1N1） pdm09 influenza pandemic strain in 2023 in Huzhou City， Zhejiang Province. MethodsRespiratory tract specimens from 2 influenza monitoring hospitals were collected forA（H1N1） pdm09 influenza virus nucleic acid detection. Positive specimens were inoculated with MDCK cells for influenza virus isolation and sequencing. DNA Star 7.1 software and Mega 4.0 software were used to analyze the neuraminidase （NA） enzyme active site and the amino acid sites related to drug resistance in M2 protein. ResultsNucleotide homology and amino acid homology of NA between the isolated and the vaccine strains were 98.87%‒99.22% and 98.94%‒99.36%， respectively. The nucleotide homology range of M gene was 99.07% to 99.85%， and the amino acid homology range was 99.02%‒99.94%. The isolates and vaccine strains belong to the evolutionary clades of 6B.1A.5a.2a. The amino acids at the key sites of the enzyme activity center of NA were still highly conserved， and the 9 key amino acid sites related to NA inhibitor resistance did not change， but some mutations occurred at the non-enzyme active sites in some popular strains. The 5 amino acid sites related to drug resistance of M2 protein were not replaced， but the 31st amino acid sites changed from serine to asparagine. ConclusionThe A（H1N1） pdm09 pandemic strain in Huzhou in 2023 has high homology with the 2023‒2024 vaccine strain recommended by WHO. All endemic strains are resistant to amantadines.

Effects of histidine 74 in rabies virus matrix protein on mitochondrial localization / 中国人兽共患病学报

This study investigated the effect of histidine(H)at position 74 on the localization of full-length viral matrix protein(M)in mitochondria.A eukaryotic expression vector expressing the N-terminus or C-terminus of wild-type M protein or its histidine 74 mutant protein(H→P,H74P)fused with V5(tag protein)-APEX2(modified ascorbate peroxidase)or A-PEX2-V5 was constructed and transfected into 293T cells.Fluorescence staining and laser confocal microscopy were used to an-alyze the effect of histidine 74 mutation on the localization of full-length M protein in mitochondria.Immunoblotting showed that the eukaryotic expression vector expressed the fusion protein normally.Fluorescence staining indicated that the H→P mu-tation at position 74 did not affect the ability of the full-length viral M protein to localize to mitochondria.Our findings regard-ing the effect of histidine 74 on the mitochondrial localization of full-length virus M protein provide a foundation for future study of the biological function of M protein and the replication mechanism of rabies virus.

Application of the emm cluster typing system in related studies of Group A Streptococcus / 中华实用儿科临床杂志

Group A Streptococcus (GAS) is one of the most important pathogens leading to children deaths all over the world.Bacterial typing is the commonest approach to analyzing the epidemiological characteristics of pathogenic microorganisms.The emm typing is frequently adopted to study GAS.The emm cluster typing is a recently proposed typing method based on the amino acid sequence homology of M proteins and the ability to bind to the host serum protein.This typing system has been extensively used for epidemiological investigation, strain selection and vaccine deve-lopment in foreign countries.However, it has not been applied in China yet. emm typing is based on a small variable region of emm genes, while the emm cluster typing system defines GAS types according to nearly intact sequences of emm genes.Besides, the emm cluster grouping system is acquired directly by emm typing comparison, so it is simple and feasible.Furthermore, the emm cluster typing can provide more information regarding the functional and structural properties of M proteins in different emm types of GAS.In this review, the methods, principles and applications of the emm cluster typing system in GAS research were summarized, in order to promote its application in China.

A case of the mutation of the poems syndrome with M protein negative in ascites / 中国综合临床

POEMS syndrome is a rare clinical disease associated with plasma cell diseases.Classic pentalogy includes: multiple peripheral neuropathy, organ enlargement, endocrine disorders, M-proteinemia and skin lesions.Due to its rarity, multiple system involvement and high clinical heterogeneity, the missed diagnosis rate and misdiagnosis rate are high.This paper reports a case of M protein negative POEMS syndrome with ascites as the prominent manifestation.

Clinical and electrophysiological profiles in early recognition of polyneuropathy, organomegaly, endocrinopathy, M-protein, and skin changes syndrome / 中华医学杂志(英文版)

Background@#The detection of polyneuropathy, organomegaly, endocrinopathy, M-protein, and skin changes (POEMS) syndrome at early stage is challenging for neurologists. Since polyneuropathy could be the first manifestation, it could be misdiagnosed as chronic inflammatory demyelinating polyneuropathy (CIDP). The present study aimed to determine the clinical and electrophysiological features of POEMS syndrome to distinguish from CIDP.@*Methods@#The data of a group of patients with POEMS (n = 17) and patients with CIDP (n = 17) in Zhongshan Hospital Fudan University from January 2015 to September 2017 were analyzed in this retrospective study. The clinical features, neurological symptoms, and electrophysiological findings were compared between the two groups.@*Results@#Clinically, patients with POEMS demonstrated significantly more neuropathic pain in the lower extremities than patients with CIDP (58.8% vs. 11.8%, P = 0.01). Multisystem features like edema, skin change, organomegaly, and thrombocytosis were also pointed towards the diagnosis of POEMS syndrome. Electrophysiologically, terminal latency index (TLI) was significantly higher in patients with POEMS than that in patients with CIDP (median nerve: 0.39 [0.17–0.52] vs. 0.30 (0.07–0.69), Z = –2.413, P = 0.016; ulnar nerve: 0.55 [0.23–0.78] vs. 0.42 [0.12–0.70], Z = –2.034, P = 0.042). Patients with POEMS demonstrated a higher frequency of absent compound muscle action potential of the tibial nerve (52.9% vs. 17.6%, P = 0.031), less conduction block (ulnar nerve: 0 vs. 35.3%, P = 0.018), and less temporal dispersion (median nerve: 17.6% vs. 58.8%, P = 0.032) than CIDP group. The combination of positive serum monoclonal protein and high TLI (if either one or both were present) discriminated POEMS from CIDP with a sensitivity of 94.1% and 47.1% and specificity of 76.5% and 100.0%, respectively.@*Conclusions@#POEMS syndrome could be distinguished from CIDP through typical clinical and electrophysiological characteristics in practice. The combination of serum monoclonal protein and high TLI might raise the sensitivity of detecting POEMS syndrome.

Clinical features, treatment and follow-ups of Sj(o)gren's syndrome-associated monoclonal gammopathy / 中华风湿病学杂志

Objective To investigate the clinical characteristics,immunological features,treatment and follow-ups of Sj(o)gren's syndrome-associated monoclonal gammopathy (SS-MG).Methods A retro-spective,case-control study was conducted for 18 cases diagnosed with SS-MG and 36 age-and sex-matched non-MG-SS patients from Janurary 2010 to Janurary 2017 in Peking University People's Hospital.The clinical and laboratory features,treatment and follow-ups were recorded and compared.Comparisons between groups were made using t test for normally distributed numerical data,Mann-Whitney U test for non-normally distributed numerical data,and Pearson Chi-square,continuity correction or Fisher's exact tests for categorical data.Results SS patients,when complicated with MG,had significantly increased level of TP [(78± 11) g/L,(71±10) g/L,t=-2.382,P=0.021] and erythrocyte sedimentation rate (ESR) [52.5(45.3) mm/1 h,33.0(42.5) mm/1 h,Z=-2.179,P=0.029],higher prevalence of urine NAG positivity [75％(9/12),28％(7/25),x2=7.298,P=0.007],hypoglobulinemia [33％(6/18),3(1/36),x2=7.407,P=0.006] and thrombotic events [17％(3/18),0％(0/36),P=0.033],and less previous exposure to glucocorticoid [22％(4/18),64％(23/36),x2=8.333,P=0.004],compared to the control group.Primary SS patients complicated with MG had significantly higher ESSDAI [26.0(25.0),12.0 (9.0),Z=-2.724,P=0.006] and Clin EULAR Sj(o)gren's syndrome disease activity index (ESSDAI) [24.0(25.0),10.5 (10.0),Z=-2.523,P=0.011].Among the 18 patients,2 were diagnosed with multiple myeloma,1 was diagnosed with non-Hodgkin lymphoma,and the left were diagnosed as MG of undetermined significance (MGUS).Ten patients were followed up,among whom 2 patients with MGUS experienced increased levels of M protein,newly developed genetic abnormalities,and renal involvement.Conclusion SS patients may be complicated with MG.MGUS is the most common form.However,malignant hematologic disorders are revealed as well.In SS patients with high serum TP and ESR,hypoglobulinemia,tubulointerstitial kidney involvement and unexplained thrombotic events,especially in those with high disease activity,so MG should be an alert for further work-ups,monitoring and treatment.

The Inductive Effect of Purified Streptococcal M-protein on Immunity against Streptococcus pyogenes in Rabbit Model.

Aims: The study aimed to know the effects of the purified M-protein on immune system to produce protection against Streptococcus pyogenes in rabbits. Study Design: Case-control study. Place and Duration of Study: In this study, collection samples and bacterial identification were carried out in two hospitals; Child Protection Hospital and Central Child Hospital in Baghdad city, and experimental work was done in Department of Medical Microbiology, College of Medicine-Babylon university, Iraq. The study was done during the period between January to July 2014. Methodology: A total of 260 samples were collected from tonsillitis and pharyngitis cases. Three main parts involved in this study: the first part is bacterial diagnosis based on relied diagnostic procedures. The second part is detection of serogroup of GAS and antistreptolysin O (ASO) antibodies by using latex agglutination test, and the third part is experimental study conducted on the protective immune response against the group A streptococci using rabbit model. M-protein was purified by using Ion exchange chromatography. The rabbit models were immunized with purified M protein according to standard method. The immune response generated against the M protein was checked in an rabbit population. Results: From a total of 260 samples of tonsillitis and pharyngitis cases among children, only 8 (3.07%) isolates were identified as Streptococcus pyogenes. High amount of M-protein was detected in two isolates by indirect bacterial test. The concentration of purified M-protein ranged from 20-24.68 µg/ml. The purified M protein has important role in an induction of the immune response in experimental model. It leads to increased phagocytosis, stimulation of T-cell, and high level of antibody in serum of an immunized rabbits. Conclusion: The purified streptococcal M protein has strong antigenicity, and it has important role in an induction the strong protective immune response in experimental rabbit model. It may be used in future studies as vaccine against streptococcal infection among humans.

Accelerating the development of a group A Streptococcus vaccine: an urgent public health need

Group A Streptococcus (GAS) infections cause substantial worldwide morbidity and mortality, mostly associated with suppurative complications such as pharyngitis, impetigo, and non-suppurative immune syndromes such as acute rheumatic fever, rheumatic heart disease, and acute post-streptococcal glomerulonephritis. Deaths occur mostly in children, adolescents, and young adults in particular pregnant women in low- and middle-income countries. GAS strains are highly variable, and a GAS vaccine would need to overcome the issue of multiple strains. Several approaches have been used multivalent vaccines using N-terminal polypeptides of different M protein; conserved M protein vaccines with antigens from the conserved C-repeat portion of the M protein; incorporation selected T- and B-cell epitopes from the C-repeat region in a synthetic polypeptide or shorter single minimal B-cell epitopes from this same region; and non-M protein approaches utilizing highly conserved motives of streptococcal C5a peptidase, GAS carbohydrate and streptococcal fibronectin-binding proteins. A GAS vaccine represents urgent need for this neglected disease and should therefore deserve the greatest attention of international organizations, donors, and vaccine manufacturers.

Progression of Extramedullary Plasmacytoma in a Multiple Myeloma Patient with No Increment in Serum M Protein Level / 대한내과학회지

A 76 year-old female who was diagnosed with multiple myeloma (IgG, lambda) had received bortezomib, melphalan and prednisolone as first-line treatment. After completing six cycles of chemotherapy, her serum monoclonal protein level decreased from 7.28 g/dL to 0.65 g/dL, indicating a partial response. However, at the next scheduled visit she complained of slowly progressing dyspnea. On chest X-ray, newly developed pleural effusion was found, and rapidly progressing extramedullary plasmacytoma was detected in the anterior mediastinum on chest computerized tomography. However, there was no change in her serum monoclonal protein level. In Korea, extramedullary involvement is encountered in 5% of patients with multiple myeloma. However, evaluation of treatment response using solely the serum monoclonal protein level may not accurately reflect disease status in these patients.

Observation and Analysis of the M Protein in Immunofixation Electrophoresis / 国际检验医学杂志

Objective To analysis the varying degrees of the M protein staining after immunofixation electrophoresis(IFE)and study its applications in clinical diagnosis.Methods 196 cases of clinical serum samples were tested by using IFE,we analyzed the positive electrophoretic bands of M protein and performed statistical analysis by using SPSS17.0.The M proteins were analyzed ret-rospectively.Results 103 patients were diagnosed with monoclonal gammopathy in 196 patients with positive M protein bands,in-cluding 96 cases of multiple myeloma(MM)and 7 cases of other monoclonal gammopathy;93 patients were non-monoclonal gam-mopathy.By analyzing the M band staining in different clinical groups,we found that M bands were mainly with dense and thick staining in monoclonal immunoglobulin group,the dense staining rate of MM was 90.6%,and the difference between MM and the other monoclonal gammopathy was not significant(P >0.05).In contrast,M bands were in light and narrow staining in non-mono-clonal immunoglobulin group,the rate of which was 25.8%,the difference between non-monoclonal immunoglobulin group and monoclonal immunoglobulin group was statistically significant(P <0.01).The proportion of allelic band in MM,other monoclonal gammopathy,non-monoclonal gammopathy were 39.6%,28.6% and 2.2% respectively,the differences were statistically significant (P <0.01).Conclusion The M band,accompanied by allelic band in IFE staining,is helpful in the diagnosis of monoclonal gam-mopathy,especially MM.The appearance of M protein provides early warning of monoclonal gammopathy.

Emm Typing of Invasive Infections Caused by Streptococcus dysgalactiae / 대한내과학회지

Recently, invasive infections with the human pathogen Streptococcus dysgalactiae subspeciesequisimilis (SDSE) have increased around the globe. Typing of the emm gene of SDSE, which encodes a virulence factor (M protein), has provided important information. Here, we report two cases of invasive SDSE infection that presented with endocarditis and bacteremia, and their emm gene types.

Emm Typing of Invasive Infections Caused by Streptococcus dysgalactiae / 대한내과학회지

Recently, invasive infections with the human pathogen Streptococcus dysgalactiae subspeciesequisimilis (SDSE) have increased around the globe. Typing of the emm gene of SDSE, which encodes a virulence factor (M protein), has provided important information. Here, we report two cases of invasive SDSE infection that presented with endocarditis and bacteremia, and their emm gene types.

Celulitis por Streptococcus equi: reporte de caso y revisión de la literatura / Streptococcus equi Cellulitis: Case Report and Literature Review

El Streptococcus equi es un coco gram positivo, perteneciente al grupo C de Lancefield, causa una enfermedad de gran relevancia en caballos, la gurma o adenitis equina (1-2); en humanos, estas infecciones son poco frecuentes, siendo más frecuentes las infecciones de piel y tejidos blandos, faringitis, neumonía, síndrome tóxico similar al shock y endocarditis. Cuando la infección está asociada a bacteriemia, la mortalidad reportada es del 25%.(3) Presentamos el caso de un hombre de 44 años que ingresa al servicio de urgencias de la Clínica universidad de la Sabana con un cuadro clínico de celulitis en mano derecha por Streptococcus equi .

Streptococcus equi is a gram-positive cocci, from group C of Lance 􀃀 eld. It causes an important disease in horses, strangles or equine adenitis (1-2). In humans, these infections are rare, and skin and soft tissue infections, pharyngitis, pneumonia, toxic shock-like syndrome and endocarditis are more frequently observed. When the infection is associated with bacteremia, the reported mortality is near 25% (3). We report the case of a 44-year old man who was admitted to the emergency department of the University of Sabana Clinic with cellulitis due to Streptococcus equi in his right hand.

Phylogenetic characterization of genes encoding for glycoprotein 5 and membrane protein of PRRSV isolate HH08

A porcine reproductive and respiratory syndrome virus (PRRSV) was obtained from clinic samples. Genes 5 and 6 encoding for the viral glycoprotein 5 and a membrane protein of the PRRSV designated as HH08 were amplified by reverse transcription-PCR. These sequences were compared with reference sequences derived from different geographical locations. The results indicated that the virus belongs to the North American type rather than European. Comparative analyses of the genetic diversity between the PRRSV isolate HH08 and other Chinese as well as foreign reference strains of PRRSV were discussed based on the sequence comparison and the topology of phylogenetic trees constructed in this study.

Clinical analysis of 639 patients with M-proteinemia / 中国实用内科杂志

Objective To understand the typing and clinical significance of M-protein,to raise the understanding and diagnostic level through analysis of 639 patients with M-protein.Methods To analysis age,sex and disease of M-protein positive patients,charateristic and content difference of multiple myeloma(MM)and monoclonal gammopathy of undeterminal significance(MGUS).Results In 639 patients with M-proteinemia,there were 409 patients with IgG-type(64%),80 IgA-type(12.5%),79 IgM-type(12.4%),4 IgD-type(0.6%),9 ? light chain(1.4%),18 ? light chain(2.8%),27 double clone(4.2%),oligoclone(2.0%).There were 115 MM(18.0%),13 Waldenstrm's macroglobulinemia(2.0%),1 primary amyloidosis of kidney,19 non-Hodgkin's lymphoma(NHL)(3%),5 chronic lymphocytic leukemia(CLL)(0.8%),473 MGUS(74.0%).Conclusion M-proteinemia is a clinical phonomenon,mainly in MM and lymphocytic proliferative disease.There is an improving tendency in MGUS detectable rate.

Serotyping of Group A Streptococci Isolated from Healthy School Children and Patients with Pharyngotonsillitis / 감염

BACKGROUND: To evaluate serological typing of T(epidemiologic marker) and M protein(major virulence antigen) is important to understand pathogenesis and epidemiology of streptococcal infection. The purpose of this study is to find out whether there were major difference in distribution of serotypes isolated from healthy school children and patients with pharyngotonsillitis, and to characterize the geographical differences in distribution of the serotypes. METHOD: Twenty-three strains of group A streptococci were isolated from healthy school children in two different areas(Dongdaemun-Ku and Kangsuh-Ku) in Seoul in April and July 1996. 23 strains came from patients living in Dongjak-Ku with pharyngotonsillitis in April 1996. All isolated were serotyped by T agglutination, M precipitation and opacity factor at the WHO Collaborative Center for Reference and Research on Streptococci, University of Minnesota, Minneapolis. RESULTS: 89.1% of the strains were typable by T agglutination, 56.5% by M precipitation, and 52.2% were positive in opacity factor. T types 1, 25, 4, and 12 accounted for 65.2% of patients with pharyngotonsillitis, T types 12, and 25 accounted for 71.5% of healthy children in Dongdaemun-Ku, and T types 28, 6, and 3 accounted for 62.6% of healthy children in Kangsuh-Ku. T types 1, 25, 28, 12, 4 and M types 1, 75, 28, 4, 12 were typed in decreasing order. CONCLUSION: We characterized the differences in serotypes of group A streptocpcci between healthy children and patients. The periodic and seasonal serotyping analysis is important in monitoring and understanding of the epidemiologic patterns of group A streptococci.

A Case of Multiple Myeloma with Biclonal (IgG-K and IgA-K) M-proteins / 대한임상병리학회지

Two M-protein peaks in serum protein electrophoresis are rarely present in patients with plasma cell discrasia. We describe a 72-year-old male patient with multiple myeloma secreting biclonal M-proteins, which were confirmed by immunofixation. Immunoelectrophoresis has some difficulties to dectect M components when a very small amount of M-protein develops an equivocally abnormal precipitation arc. In this case, serum protein electrophoresis revealed two M peaks, one in beta and the other in gamma globulin region. An immunoelectrophoresis revealed unequivocally abnormal precipitation arcs in IgA and K light chain regions, but the arc in IgG region was equivocal. We performed an immunofixation and confirmed biclonal gammopathy, IgA-K and IgG-K types. This result supports the view that immunofixation is an useful confirmatory test when immunoelectrophoresis results are equivocal.