Isolation and Phenotypic Characterization of Lytic Bacteriophage with Activity against Pseudomonas aeruginosa

Aims: To perform the isolation and phenotypic characterization of bacteriophage with lytic activity against Pseudomonas aeruginosa. To demonstrate that this type of viral agent can be isolated from the environment and used for the biocontrol of resistant bacterial types, such as Pseudomonas aeruginosa. Study Design: This study was an experimental study. Place and Duration of Study: The study was conducted at, Bacteriology and Mycology Laboratory in the Veterinary Hospital at the School of Agricultural Sciences, Innovation and Business of the University of Passo Fundo (ESAN/UPF) and Center for Diagnosis and Research in Animal Health of the University of Passo Fundo (CDSA/UPF), between April 2022 and June 2022. Methodology: Samples of untreated water were inoculated with the host bacterium strain Pseudomonas aeruginosa ATCC 27853 in an enriched media After the incubation period in, a phage filtrate was obtained by centrifugation followed by filtration. We verified the presence of bacteriophages using spot test and we carried out its purification by the method of sterile toothpick plate transfer on bacterial overlay semi-solid agar. Amplification was performed using an SM buffer elution procedure to produce a stock of viral material. Through assays in Petri dishes with bacterial overlay, we performed titration and phenotypic characterization regarding the lysis spectrum and efficiency of phage infection in the host. Results: We managed to isolate a morphologically characterized lytic bacteriophage with approximately 1 mm of diameter, high clarity in the inhibition area, the presence of halo and well-demarcated edges. The bacteriophage, named as Pseudomonas aeruginosa Phage UPF_PaBP1, demonstrated the infection capacity of the target bacteria in all tested dilutions and a stock preparation with a titre of 6.5 x 10? PFU/ml was obtained for future use. Conclusion: The isolated phage showed strong lytic activity against the bacterial host, a finding that nourishes our expectations regarding the use of this phage as a biocontrol agent and phage therapy.

Characterization of a Coliphage AS1 isolated from sewage effluent in Pakistan / Caracterização de um colifago AS1 isolado de efluente de esgoto no Paquistão

The emergence of multi-drug resistant (MDR) bacterial strains, which are posing a global health threat has developed the interest of scientists to use bacteriophages instead of conventional antibiotics therapy. In light of an increased interest in the use of phage as a bacterial control agent, the study aimed to isolate and characterize lytic phages from sewage effluent. During the current study, bacteriophage AS1 was isolated from sewage effluent against E.coli S2. The lytic activity of phageAS1 was limited to E.coli S2 strain showing monovalent behavior. The calculated phage titer was 3.5×109 pfu/ml. PhageAS1 was stable at a wide range of pH and temperature. The maximum stability was recorded at 37ºC and pH 7.0, while showing its normal lytic activity at temperature 60ºC and from pH 5.0 to 11.0 respectively. At temperature 70ºC, phage activity was somewhat reduced whereas, further increase in temperature and decrease or increase in pH completely inactivated the phage. From the current study, it was concluded that waste water is a best source for finding bacteriophages against multi-drug resistant bacterial strains and can be used as bacterial control agent.

O surgimento de cepas bacterianas multirresistentes (MDR), que representam uma ameaça global à saúde, desenvolveu o interesse dos cientistas em usar bacteriófagos em vez da terapia convencional com antibióticos. Diante do crescente interesse no uso de fago como agente de controle bacteriano, o estudo visou isolar e caracterizar fagos líticos de efluente de esgoto. Durante o estudo atual, o bacteriófago AS1 foi isolado de efluente de esgoto contra E. coli S2. A atividade lítica de phageAS1 foi limitada à cepa E. coli S2, apresentando comportamento monovalente. O título de fago calculado foi de 3,5 x 109 ufp/ml. PhageAS1 foi estável em uma ampla faixa de pH e temperatura. A estabilidade máxima foi registrada a 37ºC e pH 7,0, enquanto mostrou atividade lítica normal em temperatura de 60ºC e pH 5,0 a 11,0, respectivamente. Na temperatura de 70ºC, a atividade do fago foi um pouco reduzida, enquanto o aumento adicional da temperatura e a diminuição ou aumento do pH inativaram completamente o fago. Com base no estudo atual, concluiu-se que a água residual é a melhor fonte para encontrar bacteriófagos contra cepas bacterianas multirresistentes e pode ser usada como agente de controle bacteriano.

Characterization of a Coliphage AS1 isolated from sewage effluent in Pakistan

Abstract The emergence of multi-drug resistant (MDR) bacterial strains, which are posing a global health threat has developed the interest of scientists to use bacteriophages instead of conventional antibiotics therapy. In light of an increased interest in the use of phage as a bacterial control agent, the study aimed to isolate and characterize lytic phages from sewage effluent. During the current study, bacteriophage AS1 was isolated from sewage effluent against E.coli S2. The lytic activity of phageAS1 was limited to E.coli S2 strain showing monovalent behavior. The calculated phage titer was 3.5×109 pfu/ml. PhageAS1 was stable at a wide range of pH and temperature. The maximum stability was recorded at 37ºC and pH 7.0, while showing its normal lytic activity at temperature 60ºC and from pH 5.0 to11.0 respectively. At temperature 70ºC, phage activity was somewhat reduced whereas, further increase in temperature and decrease or increase in pH completely inactivated the phage. From the current study, it was concluded that waste water is a best source for finding bacteriophages against multi-drug resistant bacterial strains and can be used as bacterial control agent.

Resumo O surgimento de cepas bacterianas multirresistentes (MDR), que representam uma ameaça global à saúde, desenvolveu o interesse dos cientistas em usar bacteriófagos em vez da terapia convencional com antibióticos. Diante do crescente interesse no uso de fago como agente de controle bacteriano, o estudo visou isolar e caracterizar fagos líticos de efluente de esgoto. Durante o estudo atual, o bacteriófago AS1 foi isolado de efluente de esgoto contra E. coli S2. A atividade lítica de phageAS1 foi limitada à cepa E. coli S2, apresentando comportamento monovalente. O título de fago calculado foi de 3,5 x 109 ufp/ml. PhageAS1 foi estável em uma ampla faixa de pH e temperatura. A estabilidade máxima foi registrada a 37ºC e pH 7,0, enquanto mostrou atividade lítica normal em temperatura de 60ºC e pH 5,0 a 11,0, respectivamente. Na temperatura de 70ºC, a atividade do fago foi um pouco reduzida, enquanto o aumento adicional da temperatura e a diminuição ou aumento do pH inativaram completamente o fago. Com base no estudo atual, concluiu-se que a água residual é a melhor fonte para encontrar bacteriófagos contra cepas bacterianas multirresistentes e pode ser usada como agente de controle bacteriano.

Characterization of a Coliphage AS1 isolated from sewage effluent in Pakistan / Caracterização de um colifago AS1 isolado de efluente de esgoto no Paquistão

The emergence of multi-drug resistant (MDR) bacterial strains, which are posing a global health threat has developed the interest of scientists to use bacteriophages instead of conventional antibiotics therapy. In light of an increased interest in the use of phage as a bacterial control agent, the study aimed to isolate and characterize lytic phages from sewage effluent. During the current study, bacteriophage AS1 was isolated from sewage effluent against E.coli S2. The lytic activity of phageAS1 was limited to E.coli S2 strain showing monovalent behavior. The calculated phage titer was 3.5×109 pfu/ml. PhageAS1 was stable at a wide range of pH and temperature. The maximum stability was recorded at 37ºC and pH 7.0, while showing its normal lytic activity at temperature 60ºC and from pH 5.0 to11.0 respectively. At temperature 70ºC, phage activity was somewhat reduced whereas, further increase in temperature and decrease or increase in pH completely inactivated the phage. From the current study, it was concluded that waste water is a best source for finding bacteriophages against multi-drug resistant bacterial strains and can be used as bacterial control agent.

O surgimento de cepas bacterianas multirresistentes (MDR), que representam uma ameaça global à saúde, desenvolveu o interesse dos cientistas em usar bacteriófagos em vez da terapia convencional com antibióticos. Diante do crescente interesse no uso de fago como agente de controle bacteriano, o estudo visou isolar e caracterizar fagos líticos de efluente de esgoto. Durante o estudo atual, o bacteriófago AS1 foi isolado de efluente de esgoto contra E. coli S2. A atividade lítica de phageAS1 foi limitada à cepa E. coli S2, apresentando comportamento monovalente. O título de fago calculado foi de 3,5 x 109 ufp/ml. PhageAS1 foi estável em uma ampla faixa de pH e temperatura. A estabilidade máxima foi registrada a 37ºC e pH 7,0, enquanto mostrou atividade lítica normal em temperatura de 60ºC e pH 5,0 a 11,0, respectivamente. Na temperatura de 70ºC, a atividade do fago foi um pouco reduzida, enquanto o aumento adicional da temperatura e a diminuição ou aumento do pH inativaram completamente o fago. Com base no estudo atual, concluiu-se que a água residual é a melhor fonte para encontrar bacteriófagos contra cepas bacterianas multirresistentes e pode ser usada como agente de controle bacteriano.

Reducing Salmonella enterica serovar Enteritidis contamination in food: lytic bacteriophages in a homemade mayonnaise-like matrix / Reducción de la contaminación por Salmonella enterica serovar Enteritidis en alimentos: bacteriófagos líticos en una matriz similar a la mayonesa casera / Redução da contaminação por Salmonella enterica serovar Enteritidis em alimentos: bacteriófagos líticos em uma matriz caseira semelhante à maionese

Abstract Background: Salmonella enterica serovar Enteritidis (SE) is one of the major causes of food-borne disease worldwide, mainly associated with the consumption of poultry products, such as eggs. Several control methods have been implemented in the egg production process, but they have not effectively reduced the outbreaks. Therefore, the use of bacteriophages for the biocontrol of food-borne pathogens is gaining increasing acceptance. Objective: To evaluate a bacteriophage cocktail's effectiveness in reducing SE counts in an experimentally contaminated mayonnaise-like matrix. Methods: Homemade mayonnaise was contaminated with SE (103 CFU/ml) with equal volume to a matrix (1:1) treated with a bacteriophage cocktail (five phages, MOI 105), and stored at 21 °C for 24 and 72 h. Bacterial counts were performed to evaluate the bio-controlling activity of the cocktail and compared with a contaminated but not treated group. Results: Significant reductions (up to 3.75 log10 CFU/ml) were observed in the bacteriophage-treated groups (p<0.0001). Conclusions: These results demonstrate the effectiveness of bacteriophages as biocontrol agents for Salmonella Enteritidis in a raw-egg-derivative foodstuff. Further studies are needed to prove the reduction in an undiluted homemade mayonnaise.

Resumen Antecedentes: La Salmonella enterica, serovar Enteritidis (SE), es una de las principales causas de enfermedades transmitidas por alimentos en todo el mundo, asociadas principalmente al consumo de productos avícolas tales como los huevos. Diferentes métodos de control se han ensayado en el proceso de producción de huevos, pero no han sido capaces de reducir eficazmente los brotes de salmonelosis en las personas. Por esta razón, el uso de bacteriófagos para el control biológico de patógenos transmitidos por los alimentos está ganando cada vez más aceptación. Objetivo: Evaluar la eficacia de un cóctel de bacteriófagos para reducir los recuentos de SE en una matriz similar a la de mayonesa contaminada experimentalmente. Método: La mayonesa casera fue contaminada con SE (103 UFC/ml) en igual volumen que la matriz (1:1), tratada con una mezcla de bacteriófagos (cinco fagos, MOI 105), y almacenada a 21 °C por 24 y 72 h. Se realizaron recuentos bacterianos para evaluar la actividad biocontroladora de la mezcla y compararlos con un grupo contaminado, pero no tratado. Resultados: Se observaron reducciones significativas (hasta 3,75 log10 CFU/ml) en los grupos tratados con bacteriófagos (p<0,0001). Conclusiones: Estos resultados demuestran la efectividad del uso de bacteriófagos como agentes de biocontrol de Salmonella Enteritidis en un alimento crudo derivado del huevo. Sin embargo, se necesita realizar más estudios para comprobar la reducción en mayonesa casera no diluida.

Resumo Antecedentes: Salmonella enterica serovar Enteritidis (SE) é uma das principais causas de doenças transmitidas por alimentos em todo o mundo, principalmente associada ao consumo de produtos derivados de aves, como ovos. Diferentes métodos de controle foram implementados no processo de produção de ovos, mas não foram capazes de reduzir efetivamente os surtos nas pessoas. Por esse motivo, o uso de bacteriófagos para o controle biológico de patógenos de origem alimentar está ganhando crescente aceitação. Objetivo: Avaliar a eficácia de um coquetel de bacteriófagos na redução da contagem de SE em uma matriz experimentalmente semelhante a maionese contaminada. Método: A maionese caseira foi contaminada com SE (103 UFC/ml) no mesmo volume da matriz (1:1), tratada com uma mistura de bacteriófagos (cinco fagos, MOI 105) e armazenada a 21 °C por 24 e 72 h. As contagens bacterianas foram realizadas para avaliar a atividade de biocontrole da mistura e comparadas com um grupo contaminado, mas não tratado. Resultados: Reduções significativas (até 3,75 log10 UFC/ ml) foram observadas nos grupos tratados com bacteriófagos (p<0,0001). Conclusões: Esses resultados demonstram a eficácia do uso de bacteriófagos como agentes de biocontrole de Salmonella Enteritidis em alimentos crus derivados de ovos, mas são necessários mais estudos para verificar a redução da maionese caseira não diluída.

Advances in the release mechanisms of bluetongue virus / 生物工程学报

Bluetongue virus (BTV) causes Bluetongue (BT) of ruminants vectored by culicoides midges. It is also a classic model for studying the release mechanism of non-enveloped virus. This review begins with the infection and assembly of BTV, then summarizes the advances of different ways of releasing BTV. This includes BTV-induced autophagy and the release as extracellular vesicles via multivesicular bodies, BTV-induced apoptosis and the lytic release, as well as different pathways of release through budding via plasma membrane. The regulatory mechanisms of NS3 which is a key non-structural protein during the release of BTV are also discussed, providing a basis for further understanding the molecular mechanisms underpinning the infection, proliferation and release of BTV.

Antagonic and plant growth-promoting effects of bacteria isolated from mine tailings at El Fraile, Mexico / Efecto antagónico y promotor del crecimiento vegetal de bacterias aisladas de losjales mineros de El Fraile, México

Abstract Mine tailings contain high concentrations of heavy metals such as As, Pb, Cu, Mn, andFe, which are detrimental to the health of humans and the environment. In tailings at the ElFraile mine in Guerrero, Mexico, some plant species are apparently tolerant of heavy metals andcan be found growing in the tailings. These plants could be associating with heavy metal-tolerantbacteria that promote plant growth and improve biomass production, and these bacteria couldbe a useful alternative for bacteria-assisted phytoremediation. The objective of this study wasto isolate bacteria detected in the mine tailings at El Fraile-Taxco, focusing on those in the soilfrom the rhizosphere, the inner tissue of the root, leachate, and water, which have the poten-tial to promote plant growth. The ability of the isolated bacteria to promote plant growth wasevaluated in vitro. Of the 151 morphotypes isolated, 51% fix nitrogen, 12% dissolve phosphates,and 12%, 39.7%, and 48.3% produce indole acetic acid, gibberellins, and siderophores, respec-tively. In addition, 66.7% were observed to produce lytic enzymes, such as proteases, celluloses,lipases, esterases, and amylases, which exhibited activity against Fusarium, Aspergillus, andColletotrichum. The use of 16S rRNA analysis led to the identification of the bacterial generaChryseobacterium, Bacillus, Pseudomonas, Mycobacterium, Staphylococcus, Curtobacterium,Enterobacter, Agrobacterium, Ochrobactrum, Serratia, Stenotrophomonas, and Acinetobac-ter. The bacteria isolated from the rhizosphere exhibited the greatest ability to fix nitrogenand produced indole acetic acid, gibberellins, siderophore, and lytic enzymes. In addition, theisolates collected from the soil samples demonstrated ability to solubilize phosphate.

Resumen Los jales mineros contienen una alta concentración de metales pesados como As, Pb, Cu, Mn y Fe. Estas altas concentraciones de metales son perjudiciales para la salud humana y el medio ambiente. En los jales mineros de El Fraile, México, es posible detectar especies de plantas tolerantes a los metales pesados; estas plantas podrían estar asociadas con bacterias capaces de promover su crecimiento, además de poseer actividad antagonista contra hongos. El objetivo de este estudio fue aislar de diferentes microambientes (suelo rizosférico, tejido de raíz, lixiviado y agua) del área del jale El Fraile bacterias con potencial de promover el crecimiento vegetal y actividad antagonista contra hongos fitopatógenos. Estudios in vitro demostraron que el 51% de los morfotipos aislados (151 en total) fijan nitrógeno y el 12% disuelven fosfatos. Asimismo, el 12, 39,7 y 48,3% producen ácido indolacético, giberelinas y sideróforos, respectivamente. Por otro lado, se observó que el 66,7% producía enzimas líticas como proteasas, celulasas, lipasas, esterasas y amilasas, además de exhibir actividad antagonista contra Fusarium, Aspergillus y Colletotrichum. Mediante análisis del gen 16S ARNr, se identificó a estas bacterias como pertenecientes a los géneros Chryseobacterium, Bacillus, Pseudomonas, Mycobacterium, Staphylococcus, Curtobacterium, Enterobacter, Agrobacterium, Ochrobac-trum, Serratia, Stenotrophomonas y Acinetobacter. Las bacterias de la rizosfera exhibieron la mayor capacidad para fijar nitrógeno y produjeron ácido indolacético, giberelinas, sideróforos y enzimas líticas. Además, se detectó que las cepas aisladas de suelo rizosférico eran las que tenían la capacidad de solubilizar fosfatos.

Knee and the farm animals: An unusual presentation of Brucella melitensis masquerading as tuberculosis

In many developed countries, brucellosis has been successfully eradicated. However, brucellosis, with its myriad presentations, continues to be a clinical and diagnostic challenge in primarily agrarian countries such as India. We present a case of a rare manifestation of brucellosis i.e., septic arthritis of the knee joint associated with a lytic lesion of the proximal tibia. The patient belonged to a Brucella endemic country, and clinical features were of chronic reactive knee arthritis with synovial hypertrophy and effusion. Advanced diagnostic methods played a pivotal role in excluding the diagnosis of tuberculosis, and thus unnecessary administration of antitubercular therapy and initiating focused narrowed anti-Brucella management, achieving the goal of antimicrobial stewardship also.

Biological characteristics and lytic effectiveness of phages isolated from domestic wastewater against indigenous Salmonella spp

Aim: This study was carried out to isolate and study the effectiveness of lytic phage from domestic wastewater to reduce the population of Salmonella spp. in patients suffering from diarrhea and to characterize biological phages. Methodology: The lytic phages from several domestic wastewater were identified using a transmission electron microscope to know morphological phages. After identifying the molecular weight protein by sodium dodecyl sulphate-polyacrylamide gel electrophoresis, to know the effectiveness, the lytic phages were infected to Salmonella spp. from diarrheal disease patients and non-pathogenic Escherichia coli. Phage stability on thermal, pH, and buffer was then analyzed to determine the biological characteristics. Results: Three lytic phages (F-SB1, F-SB2, and F-SB3), successfully isolated from domestic wastewater, showed an icosahedral head with a short or long tail as their morphological characteristic. These phages were morphologically similar to the phages of family Siphoviridae, Myoviridae and Podoviridae. The three isolated lytic phages were stable at 27 °C to 37 °C, pH 4-7 in sodium magnesium buffer and effectively decreased the population of Salmonella spp., however could not lyse E. coli. Interpretation: All the isolated lytic phages in this study can contribute as cocktail phages in decreasing the population Salmonella spp

Single-Staged Transoral Intra-Lesional Excision of Aneurysmal Bone Cyst of Axis (C2) Vertebra and Temporary Stabilization of Upper Cervical Spine in an Adolescent: A Case Report

@#Aneurysmal bone cyst (ABC) infrequently occurs within the upper cervical vertebrae. Various therapeutic options have been reported in the literature. We would like to share our experience in managing a case of a 16-year-old girl diagnosed with ABC at the body of axis (C2) vertebra. Serious attention had to be given on the stability of the cervical spine following tumour resection, which can be affected by the mode of treatment chosen. Instability can have a detrimental effect on the cervical spine, in which case may necessitate further surgery. We performed a single-staged intra-lesional curettage via a transoral approach and temporary non-fusion posterior stabilization of C1 lateral mass screw and C2 pedicle screw. The implants were removed after six months once ossification of C2 has taken place to regain full motion of the neck. There was no evidence of recurrence or instability of the cervical spine three years following surgery.

Infective endocarditis due to Streptococci and Enterococci: A 3-year retrospective study

Introduction: Infective endocarditis (IE) is an infection of the heart valves with an aggregation of bacteria in a fibrin plaque called vegetation. Aims and Objectives: This is a retrospective study of all infective endocarditis cases due to alpha haemolytic streptococci and enterococci. Methods: All cases of infective endocarditis cases due to alpha haemolytic streptococci and enterococci in a period of three years from 1st January 2010 to 31st December 2012 were included. Isolation of the same organism from more than one set of blood cultures was taken as a confirmed case of infective endocarditis. Clinical and serological parameters were recorded using a proforma. Results: Native valve endocarditis was more common with only five prosthetic valves being involved. Out of 89 clinically suspected cases of IE in the three years from Jan 2010 to Dec 2012, for which blood was sent for culture, 63(70.78%) samples were positive by culture. Of these, 42/63(66.66%) were due to alpha-lytic Streptococci, enterococci and rare gram positive cocci. The rare ones included Enterococcus gallinarum, abiotropha defective, Vagococcus fluvialis and Nutritionally Variant Streptococci(NVS). High level Aminoglycoside resistance(HLAR) was also encountered. The varied and important features of these isolates are discussed. Complications and treatment are described. Conclusion: From a clinical microbiology point of view, the major challenge faced by the microbiologist in diagnosis of IE is proper aseptic collection of sample before starting antibiotics with a need for multiple samples to detect and also to prove the causative organism. Sensitivity reporting can be a difficult task in the context of NVS, HLAR and gram positives that are slow growing. Congestive failure and embolisation occurs even when the antibiotic treatment is successful.When patients go in for complications, it is very rarely due to wrong antibiotics.

Progress in studies on mechanism of immune evasion of Epstein-Barr virus / 中华实验和临床病毒学杂志

Epstein-Bar virus (EBV) infection is widespread within the human population with over 95% of adults being infected. In response to primary EBV infection, the host mounts an antiviral immune response comprising both innate and adaptive immune system. In healthy populations, the immune system can control EBV infection to a large extent. However, the virus cannot be cleared. Instead, EBV establishes a persistent latent infection in B lymphocytes characterized by limited viral gene expression. To establish a persistent infection efficiently, EBV have evolved a number of strategies to avoid immune elimination. In this review, we focus on the immune evasion mechanisms of EBV encoded immune-evasion proteins, microRNA and host exosome pathway.

Mutation studies of the gene encoding YuiC, a stationary phase survival protein in Bacillus subtilis

Aims@#YuiC is a stationary phase survival (Sps) protein from the Firmicute Bacillus subtilis that possesses muralytic activity to cleave bacterial cell-wall peptidoglycan. It has a small lytic transglycosylase (MltA) fold analogous to the resuscitation promoting factors (Rpfs) of Actinobacteria which have a hybrid of a mini lysozyme and soluble lytic transglycosylase (Slt35/70) fold. The present study aimed at identifying key residues of YuiC/Sps that are catalytically active and studying the effect of B. subtilis cell growth upon sps/yuiC deletion. @*Methodology and results@#Four forms of mutated yuiC were created through Site-directed, Ligase-Independent Mutagenesis Polymerase Chain Reaction (SLIM PCR) that include the substitutions of D129A, D151A, D162A and K102A. These individual mutated yuiC genes were cloned and expressed in the Escherichia coli BL21 (DE3) expression system and subsequently purified to homogeneity using affinity, cation exchange and size exclusion chromatography. The D129A variant was shown to be insoluble, indicating its role in maintaining the right protein folding of YuiC. The remaining three variants resulted in soluble proteins but were inactive on zymograms indicating that they may be responsible for catalysis. B. subtilis cells harbouring individual sps genes (yuiC, yabE, yocH and yorM) knocked out showed stationary phase defects and altered colony morphologies compared to the wild type. @*Conclusion, significance and impact of study@#This study has identified the key residues involved in catalysis of YuiC, which are the D151, D162 and K102. These are conserved in Sps domains. The catalytic mechanism of YuiC is similar to the mechanism reported for Neisseria gonorrhoeae MltA. sps/yuiC knock outs have implied that each sps/yuiC has a significant role on B. subtilis late growth stage. The B. subtilis YuiC/Sps model has given an insight into Sps functions in the final growth stage of the Firmicutes, which members include etiologic agents of anthrax, botulism and listeriosis. Inhibition of Sps protein may inactivate pathogen replication and facilitate entrance into a non-contagious dormant sporulation stage.

JMT-1: a novel, spherical lytic halotolerant phage isolated from Yuncheng saline lake

ABSTRACT This work described a novel halotolerant phage, JMT-1, with a spherical morphology. JMT-1, which was isolated from a hypersaline lake, could produce clear plaques on Chromohalobacter sp. LY7-3. The purified virions are spherical, have no visible tail, and are about 3050 nm in diameter. JMT-1 has a wide host range, and this study showed that the phage can infect at least five halophilic bacteria. The proteins of JMT-1 were analyzed using sodium dodecyl sulfate polyacrylamide gel electrophoresis, and six proteins were detected. Results show that JMT-1 is a bacteriophage with a linear double-stranded DNA. Meanwhile, the genome is approximately 23 kb in length and is sensitive to the restriction endonucleases Bam I, EcoR I, Hind III and Kpa I. JMT-1 has a high titer, approaching 1.5 × 109 pfu/mL after dilution to 106 pfu/mL. The phage is also sensitive to chloroform but not to temperature, pH, and lowered salt concentration. JMT-1 is a spherical lytic halotolerant phage with a wide host range and has the tolerance to specific extreme environments. These data could provide references for studying phage resources in extreme environments and would also provide the useful methods for isolation and identification of other valuable phage in the salt lake environment.

JMT-1: a novel, spherical lytic halotolerant phage isolated from Yuncheng saline lake

ABSTRACT This work described a novel halotolerant phage, JMT-1, with a spherical morphology. JMT-1, which was isolated from a hypersaline lake, could produce clear plaques on Chromohalobacter sp. LY7-3. The purified virions are spherical, have no visible tail, and are about 30-50 nm in diameter. JMT-1 has a wide host range, and this study showed that the phage can infect at least five halophilic bacteria. The proteins of JMT-1 were analyzed using sodium dodecyl sulfate polyacrylamide gel electrophoresis, and six proteins were detected. Results show that JMT-1 is a bacteriophage with a linear double-stranded DNA. Meanwhile, the genome is approximately 23 kb in length and is sensitive to the restriction endonucleases Bam I, EcoR I, Hind III and Kpa I. JMT-1 has a high titer, approaching 1.5 × 109 pfu/mL after dilution to 10−6 pfu/mL. The phage is also sensitive to chloroform but not to temperature, pH, and lowered salt concentration. JMT-1 is a spherical lytic halotolerant phage with a wide host range and has the tolerance to specific extreme environments. These data could provide references for studying phage resources in extreme environments and would also provide the useful methods for isolation and identification of other valuable phage in the salt lake environment.

Progress in lytic polysaccharide monooxygenase / 生物工程学报

Lignocellulose is the most abundant renewable biomass resource. Enzymatic breakdown of lignocellulose into oligosaccharides or monosaccharides is the key to exploit lignocellulosic biomass. However, traditional glycoside hydrolases are insufficient to degrade lignocellulose. The emergence of lytic polysaccharide monooxygenase, a novel enzyme for lignocellulose degradation, has enriched the deconstruction schema and accelerated the enzymatic conversion of polysaccharides, by introducing new chain breaks that allow hydrolases to initiate further degradation. Here, we review the discovery, classification and catalytic mechanism of the enzyme, as well as the methods for assaying its activity. The prospect for its application in feed additive, functional food and biofuel development is further discussed.

Peptidoglycan recycling and bacterial resistance to β-lactams / 中国药学杂志

To introduce peptidoglycan recycling and the β-lactams resistance mechanisms of bacteria, so that some help would be supplied to corresponding scientific workers and university teachers. By searching literatures, combined with our own studies, the bactericidal mechanisms of β-lactams and the resistance mechanisms of bacteria to β-lactams were summarized. The bactericidal activity of β-lactams is resulted from the inhibition of cell wall biosynthesis through combination with penicillin binding proteins such as transpeptidase destruction of peptidoglycan balances between biosynthesis and hydrolysis. The drug resistance of bacteria is resulted from the induction of β-lactamase, expression of out-pumping proteins, increase of outmembrane permeability, and modification of antibiotic target proteins. The proteins related to peptidoglycan recycling, such as transpeptidase and glycosyltransferase, would be potential targets for screening new β-lactams. The proteins related to β-lactams resistance, such as β-lactamase, would be potential targets for screening adjuvant drugs of β-lactams.

Effects of cyclin-dependent kinase 9 ( CDK9) on lytic replication of Kaposi′s sarcoma-associated her-pesvirus ( KSHV) / 中华微生物学和免疫学杂志

Objective To investigate the role of cyclin-dependent kinase 9 ( CDK9) in regulating lytic replication of Kaposi′s sarcoma-associated herpesvirus ( KSHV) .Methods Percentages of red fluores-cent protein (RFP) positive cells were counted after treating and stimulating iSLK .219 cells with FIT-039, a CDK9 specific inhibitor , and doxycycline ( Dox ) , respectively , or transfecting and stimulating them with CDK9 siRNA (si-CDK9) and Dox, respectively.Quantitative real-time PCR was used to detect the expres-sion of KSHV genes at mRNA level in TREx-K-Rta BCBL-1 cells treated with FIT-039 or transfected with si-CKD9 in the presence of Dox.Chromatin immunoprecipitation (ChIP) assay was used to examine the enrich-ment of RNA polymerase Ⅱ( RNA PolⅡ) and phosphorylated CTD-S2 of RNA PolⅡon the PAN promot-er in TREx-K-Rta BCBL-1 cells treated with FIT-039.Results Both FIT-039 intervention and CDK9 knockdown dramatically deceased the percentage of RFP-positive iSLK.219 cells and suppressed the expres-sion of ORF50, PAN and K2 in TREx-K-Rta BCBL-1 cells at mRNA level.Furthermore, FIT-039 signifi-cantly inhibited the enrichment of RNA Pol Ⅱand phosphorylated CTD-S2 of RNA Pol Ⅱon the PAN pro-moter in TREx-K-Rta BCBL-1 cells.Conclusion CDK9 is involved in regulating the lytic replication of KSHV through promoting the phosphorylation of CTD-S2 of RNA Pol Ⅱ.

Different characteristics between mycobacteriophage Chy1 and D29, which were classified as cluster A2 mycobacteriophages.

Purpose: The aim of this study was to isolate a novel mycobacteriophage and then explore its anti‑tuberculosis (TB) potential. Materials and Methods: Phage was isolated from enriched soil sample. A total of 36 mycobacterial strains obtained from clinical specimens were subjected to investigate the host range of phage by the spot lysis assay. Biological characteristics were investigated through growth curve, host range and phage antimicrobial activity in vitro. Then, genome sequencing and further analysis were accomplished by using an ABI3730XL DNA sequencer and comparative genome, respectively. Results: A lytic mycobacteriophage (Chy1) was isolated and the plaque morphology was similar to D29. The genome of Chy1 was estimated to be about 47,198 base pair (bp) and strong similarity (97.4% identity) to D29, especially, the Chy1 gene 7 encoding holin which is considered as a clock controlling growth cycle of the corresponding phage, was identical (100% identity) to phage D29 gene 11, thus classifying Chy1 as a member of the cluster A2 family. However, to our surprise, Chy1 can infect a narrower range of host‑mycobacterial strains than that of D29. The latent period of Chy1 was quite longer compared to D29. Moreover, Chy1 has a weaker ability to lyse Mycobacterium smegmatis compared to D29. Conclusions: The sequence of Chy1 showed 97.4% homology with the genome sequence of D29, but there was a large difference in their biological characteristics. Overall, the results of this investigation indicate that Chy1 is not an ideal candidate for developing mycobacteriophage‑based anti‑TB therapies but for future researches to investigate the reason why biological characteristics of Chy1 and D29 were remarkably different.

Determination of lytic enzyme activities of indigenous Trichoderma isolates from Pakistan

Abstract This study investigated lytic enzyme activities in three indigenous Trichoderma strains namely, Trichoderma asperellum, Trichoderma harzianum and Trichoderma sp. Native Trichoderma strains and a virulent strain of Rhizoctonia solani isolated from infected bean plants were also included in the study. Enzyme activities were determined by measuring sugar reduction by dinitrosalicylic acid (DNS) method using suitable substrates. The antagonists were cultured in minimal salt medium with the following modifications: medium A (1 g of glucose), medium B (0.5 g of glucose + 0.5 g of deactivated R. solani mycelia), medium C (1.0 g of deactivated respective antagonist mycelium) and medium D (1 g of deactivated R. solani mycelia). T asperellum showed presence of higher amounts of chitinases, β-1, 3-glucanases and xylanases in extracellular protein extracts from medium D as compared to medium A. While, the higher activities of glucosidases and endoglucanses were shown in medium D extracts by T. harzianum. β-glucosidase activities were lower compared with other enzymes; however, activities of the extracts of medium D were significantly different. T. asperellum exhibited maximum inhibition (97.7%). On the other hand, Trichoderma sp. did not show any effect on mycelia growth of R. solani on crude extract.