In vitro lytic efficiency of Staphylococcus aureus bacteriophages in bacteria from bovine mastitis: a meta-analysis / Eficiência lítica in vitro de bacteriófagos de Staphylococcus aureus em bactérias de mastites bovina: uma meta-análise

ABSTRACT: Bacteriophages have been investigated as alternative to the treatment of bacterial infections, including bovine mastitis, in production animals. In this meta-analysis, we evaluated in vitro efficiency of phages of Staphylococcus aureus against S. aureus, which is involved in the etiology of bovine mastitis. Seventeen studies were included and the bacterial lytic activity was extracted using proportion analysis. The lytic efficiency of phages was obtained in this meta-analysis using a random-effects model [significant difference (P<0.05)]. Forest plots were used to graphically represent the efficiency of phages on bacterial isolates. Most phages (e.g., CS1, DW2, ΦSA011, ΦSA012, ΦSA022, ΦSA023, ΦSA024, ΦSA025, ΦSA037, ΦSA038, ΦSA039, ΦSA041, ΦSA042, ΦSA043, ΦSA044, MSA6, Ufv-aur2 to Ufv-aur11, SAH-1, SPW, vB_SauM_JS25, SaPh1 to SaPh6, SA, SANF, SA2, ΦSA012, ΦSA039, phi11, phiIPLA88, phiIPLA35, phiIPLA-RODI, phiIPLA-C1C, SAJK-IND, vBSP-A1, vBSP-A2, STA1.ST29, EB1.ST11, EB1.ST27, Remus, and ISP) were efficiently lytics or infected most S. aureus isolates, demonstrating 80% (P<0.05) lytic efficiency. The phages SA, SANF and SA2, also demonstrated lytic activity or infected the non-Staphylococcus aureus and Macrococcus caseolyticus isolates. In this meta-analysis, we compared and demonstrated the in vitro efficiency and host range of S. aureus phages. Additionally, the phages represent an alternative to be researched to treat bovine mastitis in dairy cattle caused by the prevalent microorganism, S. aureus.

RESUMO: Os bacteriófagos têm sido investigados como alternativa ao tratamento de infecções bacterianas em animais de produção, incluindo a mastite bovina. Nesta meta-análise, avaliamos a eficiência in vitro de fagos de Staphylococcus aureus contra S. aureus envolvidas na etiologia da mastite bovina. Dezessete estudos foram incluídos e a atividade lítica bacteriana foi extraída usando análise de proporção. A eficiência lítica dos fagos foi obtida nesta meta-análise, usando um modelo de efeitos aleatórios (diferença significativa (P <0,05)). Os gráficos de Forest plots foram usados para representar graficamente a eficiência dos fagos em isolados bacterianos. Os fagos avaliados, na sua grande maioria, (por exemplo, CS1, DW2, ΦSA011, ΦSA012, ΦSA022, ΦSA023, ΦSA024, ΦSA025, ΦSA037, ΦSA038, ΦSA039, ΦSA041, ΦSA042, ΦSA043, ΦSAf e U0f, ua04 SPW, vB_SauM_JS25, SaPh1 a SaPh6, SA, SANF, SA2, ΦSA012, ΦSA039, phi11, phiIPLA88, phiIPLA35, phiIPLA-RODI, phiIPLA-C1C, SAJK-IND, vBSP-A1, vBSP-A2, STA1.ST29, EB1.ST11, EB1.ST27, Remus, e ISP) foram eficientemente líticos ou infectaram a maioria dos isolados de S. aureus, demonstrando 80% (P < 0,05) de eficiência lítica. Os fagos SA, SANF e SA2 também demonstraram atividade lítica ou infectaram os isolados Staphylococcus não-aureus e Macrococcus caseolyticus. Nesta meta-análise, comparamos e demonstramos a eficiência in vitro e gama de hospedeiros de fagos de S. aureus. Adicionalmente, os fagos representam uma alternativa a ser pesquisada para o tratamento da mastite bovina em gado leiteiro causada pelo microrganismo prevalente, ou seja S. aureus.

Effects of phage therapy on sepsis / 中华烧伤杂志

Phages can fight against sepsis through directly lysing the bacteria and influence the patients′ self-response to the pathogens through the immunomodulation effects in a coordinated way. Under the situation of the rising antimicrobial resistance, phage has attracted wide attention of researchers at home and abroad. Along with the development of researches and clinical related trials, we believe phage therapy in sepsis treatment can be expected soon in the future.

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

Encapsulation of specific Salmonella Enteritidis phage f3αSE on alginate-spheres as a method for protection and dosification

Background: Bacteriophages have been proposed as an alternative to control pathogenic bacteria resistant to antibiotics. However, they are not extensively used due to different factors such as vulnerability under environmental conditions and the lack of efficient administration methods. A potential solution is the encapsulation of bacteriophages in hydrogel polymers to increase their viability and as a controlled release method. This work describes the use of alginate-Ca+2 matrixes as mechanisms for protection and dosification of the phage f3αSE which has been successfully used to prevent infections produced by Salmonella Enteritidis. Results: The viability of the pure phage is reduced in near 100% after 1-h incubation at pH 2 or 3. However, the encapsulated phage remains active in 80, 6% at pH 3, while no differences were observed at pH 2, 4 or 7. Exposition of f3αSE to different T° showed that the viability of this phage decreased with increased T° to near 15% at 60°C, while the encapsulated phage remains with 50% viability at same temperature. Finally, the encapsulation of phages showed to extend their presence for 100 h in the medium compared to non-encapsulated phages in a water flow system, which simulate automatic birdbath used in poultry industry, maintaining the phage concentration between 102 and 104 PFU/mL during 250 h. Conclusions: Encapsulation in alginate-Ca+2 spheres can be a good alternative to extend viability of phages and can be used as a phage method dosification method in water flow systems.

Expression and relationship of PRL-3,tumor associated macrophags and lymphangiogenesis in papillary thyroid carcinoma / 实用医学杂志

Objective To investigate the relationship of PRL-3,tumor associated macrophages and lym-phangiogenesis in papillary thyroid carcinoma. Methods SP immunohistochemistry was used to study the levels of PRL-3,CD68,and D2-40 in papillary thyroid carcinoma and thyroid adenoma.Results The positive expression rates of PRL-3,CD-68 and D2-40 were higher in papillary thyroid carcinoma than those in thyroid adenoma (P < 0.01). High PRL-3,CD-68 or D2-40 was associated with lymphatic metastasis in patients with papillary thyroid carcinoma(P<0.01).Conclusion The expression levels of PRL-3,CD-68 and D2-40 are positively cor-related in papillary thyroid carcinoma,and they are related to invasion and lymphangiogenesis of papillary thyroid carcinoma.

The induction, purification, whole genome sequencing and comparative genome analysis of a novel staphylococcus haemolyticus bacteriophage IME-SA4 / 中国医师杂志

Objective To induce a novel temperate bacteriophage from staphylococcus haemolyticus strain SA98,observe the morphology and size,complete the whole genome sequencing,analyse the structure of genome and evolutionary relationship.Methods The mitomycin C was used to induce the temperate phage from staphylococcushaemolyticus strain SA98,the induced phage was observed by transmission electron microscopy after be concentrated and purified.The genome DNA was extracted and high through sequenced.The feature of whole genome and evolutionary relationship was analyzed.Results A temperate phage IME-SA4 was successfully induced from staphylococcus haemolyticus strain SA98.Transmission electron microscopy analysis indicated that IME-SA4 had an isometric head and a non-contractile long tail.The whole genome of IME-SA4 was long as 41 843 bp,and the whole genome blast result indicated IME-SA4 shared only 13％ homology with most related strain phiRS7.Conclusions A novel staphylococcus haemolyticus temperate phage with low homology with other staphylococcusphages was successfully induced from staphylococcus haemolyticus strain SA98.The research of its morphology,whole genome sequencing and comparative genome analysis make the foundation for further study of staphylococcus phages' properties and practical application.

Los enzibióticos como alternativa terapéutica contra las enfermedades bacterianas / The enzybiotics as therapeutic alternative against bacterial diseases

El constante desarrollo de las enfermedades infecciosas, conjuntamente con la aparición de la resistencia microbiana a los antibióticos, ha originado que nuevamente se piense en los fagos como opción terapéutica. De hecho, existe una importante aportación bibliográfica sobre los bacteriófagos y su utilidad para eliminar los procesos infecciosos, lo que ha justificado el continuar investigando acerca del posible uso de estos y de sus productos génicos, como esperanzadora alternativa a los tratamientos con antimicrobianos disponibles en la actualidad. Por ello, en este artículo se ofrece información sobre estos microorganismos, en específico sobre los enzibióticos, y se propone que sean considerados en el combate contra las infecciones bacterianas.

The constant development of the infectious diseases, together with the emergence of the microbial resistance to the antibiotics, has originated that again it is thought on the phages as therapeutic option. In fact, an important literature contribution exists about the bacteriophages and their use to eliminate infectious processes, what has justified the continuity in investigating about the possible use of them and of their genic products, as a promising alternative for treatments with antimicrobials currently available. That is why, information on these microorganisms is offered in this work, specifically on the enzibiotics, and is it intended them to be considered in the bacterial infections control.

Differentially expressed genes of HepG2 cells treated with gecko polypeptide mixture / 中国药理学与毒理学杂志

OBJECTIVE In order to investigate the possible anti-tumor molecular mechanisms of gecko polypeptide mixture (GPM). METHODS RNA-seq technology was used to identify the differen?tially expressed genes of human hepatocellular carcinoma (HCC) HepG2 cells treated with or without GPM. The HepG2 cells were treated with different concentration of GPM (0, 0.1, 0.2, 0.3, 0.4 mg·mL-1) for 6 h, 12 h and 24 h, respectively. MTT assay was used to detect the viability of HepG2 cells. DAPI fluorescence staining was performed to observe nucleus morphological changes of HepG2 cells. Western blot analysis was applied to observe the expression of apoptosis-related proteins in HepG2 cells. RESULTS The results showed that GPM could induce HepG2 cells apoptosis and influence HepG2 cells proliferation in a dose-dependent manner. We applied many analysis methods, including differen?tially expressed genes analysis, Gene Ontology (GO) enrichment analysis, KEGG pathway enrichment analysis, protein- protein interaction network analysis to screen out possible molecular mechanisms. ER-nucleus signaling pathway, cellular response to stress and apoptotic processes were identified the potential anti-cancer molecular biological process of GPM. GPM may also induce apoptosis in HepG2 cells via endoplasmic reticulum stress pathway. The mechanism is closely related to ERs, which might be beneficial for clinical therapy of HCC. CONCLUSION GPM can inhibit cells proliferation and induce apoptosis in HepG2 cells. The gene expression profile of GPM in HepG2 cells was obtained. The present study revealed the potential anti-tumor mechanism of GPM.

Isolation and characterization of bacteriophage against extended-spectrumβ-lactamase-producing Escherichia coli strains / 上海交通大学学报（医学版）

Objective · To isolate phages which can fight against extended-spectrum β-lactamase (ESBLs)-producing Escherichia coli (E. coli), and provide basic research for establishment of E. coli phage library and treatment of bacterial infection. Methods · Samples collected from sewage were co-cultured with 93 ESBLs-producing E. coli strains. A phage named JDEC001 was isolated by double agar overlay plaque assay. The biological characteristics, complete genome sequence and comparative genome analyses of JDEC001 were studied respectively. Results · JDEC001 belongs to the lytic phage as a member of the Caudovirales order, Podoviridae family. It has high activity at pH from 5 to 11 and with temperature from 0 to 39 ℃ .Whole-genome sequencing of JDEC001 demonstrated double-stranded DNA genome of 38745 bp with GC content of 49.93%, which encoded 46 open reading frames. The comparative genomics also showed that there was no virulent genes or antibiotic resistant genes in its genome. Conclusion · The phage JDEC001 against ESBLs-producing E. coli was isolated and purified, with good stability in a broad range of pH and temperature.

Inhibitory effects of the Chlamydiaphage phiCPG1 capsid protein Vp1 on Chlamydia psittaci strain guinea pig inclusion conjunctivitis and Chlamydia trachomatis serovar E / 中华皮肤科杂志

Objective To evaluate inhibitory effects of the Chlamydiaphage phiCPG1 capsid protein Vp1 on Chlamydia psittaci strain guinea pig inclusion conjunctivitis (GPIC) and Chlamydia trachomatis (Ct) serovar E,and to provide new ideas for the treatment of Ct infection.Methods The Chlamydiaphage phiCPG1 capsid protein Vp1 was expressed in Escherichia coli BL21 transfected with the recombinant plasmid Vp1-pET30a (+),identified by Western blot analysis and purified by using dialysis bags.Bicinchonininc acid (BCA) assay was performed to determine the concentration of Vp1 protein.GPIC and Ct serovar E strains were both classified into 4 groups to be firstly incubated with Vp1 protein (Vp1 group),Tris-glycine solution (Tris group),S protein (S group) or Dulbecco's Modified Eagle Medium (DMEM,DMEM group) at room temperature for 3 hours,then were used to infect Hela cells followed by 72-hour (GPIC) or 48-hour (Ct serovar E) culture with the presence of Vp 1 protein (Vp 1 group),Tris-glycine solution (Tris group),S protein (S group) or DMEM (DMEM group).Subsequently,immunofluorescence staining was conducted to observe and count chlamydial inclusions.Results The number of GPIC inclusions was significantly different between the 4 groups after 72-hour culture (F=476.632,P＜ 0.05),and lower in the Vp1 group (5.0 ± 1.5) than in the Tris group (24 ± 1.2,P＜ 0.05),S group (25 ± 1.7,P＜ 0.05) and DMEM group (25 ± 1.5,P＜ 0.05),but insignificantly different between the latter 3 groups (P ＞ 0.05).Compared with the DMEM group,the Vp1 group showed a significant decrease of 80.2％ ± 3.99％ and 77.2％ ± 1.79％ in the number of GPIC and Ct serovar E inclusions respectively,with no significant difference in the inhibitory effect of Vp1 on GPIC versus Ct serovar E (t =2.057,P ＞ 0.05).Conclusion The phiCPG1 capsid protein Vp1 can obviously inhibit GPIC and Ct serovar E infections to a similar degree.

Influences of chrysophanol on immune function of lead poisoning mice / 中国药理学通报

Aim To investigate the protective effects of chrysophanol( Chry) on immune function of lead poi-soning mice. Methods The lead poisoning model was established by peritoneally injecting mice with 7 mg · kg-1 lead acetate every day for 8 days. After Chry was ip injected for 14 days,spleen and thymus index, the white blood cell count, T, B lymphocyte proliferation, phagocytic function of macrophages, natural killer cell ( NK cell) activity were detected. The concentrations of IFN-γ,IL-2,IL-4,IL-10 in the lead poisoning mice ser-um were determined by enzyme-linked immunosorbent assay ( ELISA) . Results Intraperitoneal injection of 7 mg · kg-1 lead acetate for 8 consecutive days could cause an immunity decline in lead poisoning mice, Chry could significantly improve the immunity of lead poisoning mice. Compared with model group, Chry sig-nificantly improved growth rate, viscera index and white blood cell count of lead poisoning mice at differ-ent extent ( P0. 05 ) , only IL-10 was significantly increased in Chry ( 0 . 1 mg · kg-1 ) treatment group ( P<0 . 05 ) . Conclusion Chry can significantly improve the im-mune function of lead poisoning mice.

Liquid based formulations of bacteriophages for the management of waterborne bacterial pathogens in water microcosms.

Water resources are contaminated by life-threatening multidrug resistant pathogenic bacteria. Unfortunately, these pathogenic bacteria do not respond to the traditional water purification methods. Therefore, there is a need of environmentally friendly strategies to overcome the problems associated with the antimicrobial resistant bacterial pathogens. In the present study, highly potent lytic phages against multidrug-resistant Salmonella enterica serovar Paratyphi B, Pseudomonas aeruginosa and Klebsiella pneumoniae were isolated from the Pavana river water. They belonged to the Podoviridae and Siphoviridae families. These phages were purified and enriched in the laboratory. Monovalent formulations of φSPB, BVPaP-3 and KPP phages were prepared in three different liquids viz., phage broth, saline and distilled water. The phages were stable for almost 8-10 months in the phage broth at 4 °C. The stability of the phages in saline and distilled water was 5-6 months at 4 °C. All of the phages were stable only for 4-6 months in the phage broth at 30 °C. The monovalent phage formulation of φSPB was applied at MOI < 1, as disinfectant against an exponential and stationary phase cells of Salmonella enterica serovar Paratyphi B in various water microcosms. The results indicated that there was almost 80 % reduction in the log phase cells of Salmonella serovar Paratyphi B in 24 h. In stationary phase cells, the reduction was comparatively less within same period. At the same time, there was concomitant increase in the phage population by 80% in all the microcosms indicating that φSPB phage is highly potent in killing pathogen in water. Results strongly support that the formulation of φSPB in the phage broth in monovalent form could be used as an effective biological disinfectant for preventing transmission of water- borne bacterial pathogens, including antimicrobial resistant ones.

Characterization and lytic activity of Pseudomonas fluorescens phages from sewage

Pseudomonas fluorescens phages from sewage were tested against P. fluorescens isolates of soil and sewage. The phages were characterized as to host range, morphology, structural proteins and genome fingerprint. Of the seven phages isolated, one was found to be abundant in sewage (5.9×10(7) pfu/mL), having broad host range, and distinct protein and DNA profile when compared to the other six phages. DNA restriction and protein profiles of the phages and their morphology indicate the diversity in the sewage environment. None of the isolates from the rhizosphere regions of various cultivated soils were susceptible to phages isolated from sewage.

Detection of Chlamydia trachomatis phage Vp1 gene in clinical swab specimens as well as anti-Vp1 antibodies in serum specimens / 中华皮肤科杂志

Objective To detect Chlamydia trachomatis phage Vp1 gene in clinical swab specimens and anti-Vp1 antibodies in serum specimens.MethodsCervical and urethral swab as well as serum specimens were collected from attendees to the sexually transmitted disease(STD) clinic in the Tianjin Institute of STD,during March 2008 to March 2011.PCR was conducted to detect chlamydial phage Vp1 gene in swab samples,enzyme linked immunosorbent assay(ELISA) and Western blot to detect anti-Vp1 antibody in sera.The swab specimens positive for Vp1 gene were subjected to cell culture followed by the detection of Vp1 protein with an immunofluorescence-based method.ResultsTotally,36 out of 1542 swab specimens turned out to be positive for Vp1 gene,and 23 out of 453 serum specimens for anti-Vp1 antibody.No positive results were obtained in the Vp1 gene-positive swab specimens by cell culture and immunofluorescence-based assay.ConclusionThe Vp1 gene of Chlamydial trachomatis phage and anti-Vp1 antibody are successfully detected from clinical swab and serum specimens respectively.

loxP-cre system based in vivo recombination / 肿瘤研究与临床

Phage display technology is improved in recent years.LoxP-cre system based in vivo recombination is the most important one. From double recombination system to single recombination system,from large scale sample sequencing to visual judgement, in vivo recombination makes it feasible to construct large natural antibody library, and it is favorable to get high affinity antibody. This review introduces the important improvement of in vivo recombination system.

Cloning, expression and identification of phage's capsid Vp3 protein of guinea pig inclusion conjunctivitis chlamydia / 中华传染病杂志

Objective To get phagc's capsid Vp3 gene and protein of guinea pig inclusion conjunctivitis (GPIC) chlamydia. Methods The genome DNA was extracted from the φCPG1 phage.The full sequence of Vp3 gene was amplified by polymerase chain reaction (PCR) from the above genome DNA. The Vp3 gene was digested by restriction endonuclease and then inserted into prokaryotic plasmid vector pET30a (+). The recombinant plasmid was transformed into E. coil BL21, and was identified by restriction endonuclease, PCR and sequencing. The E. coil BL21 with expected recombinant plasmid was induced and the expressed recombinant Vp3 protein was identified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blot, then purified by agarose gel. Results The recombinant gene was sequenced and proved to be 447 bp which was consistent with the φCPG1 Vp3 gene sequence in GenBank. A 25 000 capsid protein was expressed and confirmed by SDS-PAGE and Western blot. The purified protein was obtained. Conclusion The capsid Vp3 protein of φCPG1 is successfully expressed and purified, which is helpful for the further study on its mechanism and clinical applications.

Characterization of a Vibrio parahaemolyticus Phage Isolated from Marine

A novel bacteriophage, designated as VPP97, that infects the strains of Vibiro parahaemolyticus (hallophilic, Gram-negative bacterium) isolated most commonly from marine environments, has been discovered, and several of its properties have been determined. The plaques were clear and sized 0.6-1.0 mm in diameter. The virion forms a single band on 70% sucrose gradient and p1.50 CsC1 gradient by sucrose gradient centrifugation and CsCI gradient centrifugation respectively. It has a hexagonal head and a relatively long tail, as shown by electron microscopy. Vibrio alginolyticus, Vibrio fluvialis and Vibrio furnissii were also sensitive to this phage It was almost totally inactivated at 70 degree C and at pH below 5 or over 10. The nucleic acid of VPP97 is composed of DNA. The VPP97 had 9 specific structural proteins sized between 21.5 kDa and 97.4 kDa on SDS-PAGE. When V. parahaemolyticus cultures were treated with either phage VPP97 or one of the several antibiotics for 2 hours, the viable number of V. parahaemolyticus treated with the phage VPP97 is lower than that treated with chloramphenicol, erythromycin or penicillin, but not lower than that treated with tetracycline. Mice that have responded to the phage treatment revealed the lower numbers of V. parahaemolyticus in small intestine and less damage on small intestine compared to the untreated mice. Therefore, we suggest that the phage treatment appears effective to the infection by V. parahaemolyticus.

Construction of phage random eight-peptide library / 中国病理生理杂志

AIM: To construct random eight-peptide library for the study on atherosclerosis and restenosis. METHODS and RESULTS: Random oligodeoxynucleotides encoded eight peptides were synthesized and amplified by polymerase chain reaction(PCR). The product was cloned into phage surface display vector fUSE5 in Sfi I site and electroporated into competent MC1061. The library was identified through PCR, hybridization, DNA sequencing and affinity biopanning of streptavidin. Because the upstream primer is complementary to part vector clone site sequences and part exogenous gene sequences, and the other one complementary to pIII gene of vector, thus only clones inserted exogenous gene could be amplified easily. Additionally we used the probe oligodeoxynucleotide complementary to vector clone site sequences to identify clones which were not inserted exogeneous genes. Furthermore, two hybridizing positive clones were sequenced. Their sequences are consistent with two oligodeoxynucleotide probe sequences. As a result, 2.1?108 special clones were obtained. Affinity biopanning proved that the libraries could be amplified steadily. CONCLUSION: The eight-peptide library is reliable.

Construction of a packaged strain XL-pⅢ and preparation of defective helper phage / 解放军医学杂志

Objective To construct a packaged bacteria strain and, with it, to prepare gene III-defective helper phage. Methods Whole gene III and chloramphenicol-resistant gene were amplified and joined to form a fragment flanking 36bp homologous sequence of hisBCD using overlapping extending PCR, and it was then integrated into chromosome of E. coli XL1-Blue to replace the hisBCD gene using Red recombination system. The recombinant strain was identified with chloramphenicol-resistant screening, PCR and histidine-phenotype analysis, and named as XL-pⅢ. The gene III-deleted genome of phage was constructed with PCR and transfected into the recombinant strain XL-pⅢ to prepare the defective helper phage. It was quantified by infecting E. coli XL1-Blue and formed colonies were counted in kanamycin plate. Results The recombinant strain could grow in chloramphenicol-resistant plate, but couldn't grow in M9 medium without histidine, implying that it had lost its histidine phenotype. The aimed gene fragment could be amplified as designed. The constructed recombinant was named as XL-pⅢ. The prepared defective helper phage could infect E. coli XL1-Blue only once and the CFU was 3.7?108. Conclusions The packaged strain XL-pⅢ is successfully constructed and used to prepare the gene III-deleted helper phage. It is expected to be used in SIP system.