Journal of Microbiology

Volume 50(3); June 2012

Research Support, Non-U.S. Gov'ts

Seasonal Changes in Nitrogen-Cycle Gene Abundances and in Bacterial Communities in Acidic Forest Soils: Jaejoon Jung , Jinki Yeom , Jiwon Han , Jisun Kim , Woojun Park; J. Microbiol. 2012;50(3):365-373. Published online June 30, 2012; DOI: https://doi.org/10.1007/s12275-012-1465-2

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46 Citations

Abstract: The abundance of genes related to the nitrogen biogeochemical cycle and the microbial community in forest soils (bacteria, archaea, fungi) were quantitatively analyzed via real-time PCR using 11 sets of specific primers amplifying nifH, bacterial amoA, archaeal amoA, narG, nirS, nirK, norB, nosZ, bacterial 16S rRNA gene, archaeal 16S rRNA gene, and the ITS sequence of fungi. Soils were sampled from Bukhan Mountain from September of 2010 to July of 2011 (7 times). Bacteria were the predominant microbial community in all samples. However, the abundance of archaeal amoA was greater than bacterial amoA throughout the year. The abundances of nifH, nirS, nirK, and norB genes changed in a similar pattern, while narG and nosZ appeared in sensitive to the environmental changes. Clone libraries of bacterial 16S rRNA genes were constructed from summer and winter soil samples and these revealed that Acidobacteria was the most predominant phylum in acidic forest soil environments in both samples. Although a specific correlation of environmental factor and gene abundance was not verified by principle component analysis, our data suggested that the combination of biological, physical, and chemical characteristics of forest soils created distinct conditions favoring the nitrogen biogeochemical cycle and that bacterial communities in undisturbed acidic forest soils were quite stable during seasonal change.

Antioxidant Capacity of Novel Pigments from an Antarctic Bacterium: Daniela N. Correa-Llantén , Maximiliano J. Amenábar , Jenny M. Blamey; J. Microbiol. 2012;50(3):374-379. Published online June 30, 2012; DOI: https://doi.org/10.1007/s12275-012-2029-1

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51 Citations

Abstract: In Antarctica microorganisms are exposed to several conditions that trigger the generation of reactive oxygen species, such as high UV radiation. Under these conditions they must have an important antioxidant defense system in order to prevent oxidative damage. One of these defenses are pigments which are part of the non-enzymatic antioxidant mechanisms. In this work we focused on the antioxidant capacity of pigments from an Antarctic microorganism belonging to Pedobacter genus. This microorganism produces different types of pigments which belong to the carotenoids group. The antioxidant capacity of a mix of pigments was analyzed by three different methods: 1,1-diphenyl-2-picrylhydrazyl, ROS detection and oxygen electrode. The results obtained from these approaches indicate that the mix of pigments has a strong antioxidant capacity. The oxidative damage induced by UVB exposure to liposomes was also analyzed. Intercalated pigments within the liposomes improved its resistance to lipid peroxidation. Based on the analysis carried out along this research we conclude that the antioxidant properties of the mix of pigments protect this bacterium against oxidative damage. These properties make this mix of pigments a powerful antioxidant mixture with potential biotechnological applications.

Biological Control and Plant Growth Promoting Capacity of Rhizobacteria on Pepper under Greenhouse and Field Conditions: Mi-Seon Hahm , Marilyn Sumayo , Ye-Ji Hwang , Seon-Ae Jeon , Sung-Jin Park , Jai Youl Lee , Joon-Hyung Ahn , Byung-Soo Kim , Choong-Min Ryu , Sa-Youl Ghim; J. Microbiol. 2012;50(3):380-385. Published online June 30, 2012; DOI: https://doi.org/10.1007/s12275-012-1477-y

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60 Citations

Abstract: Plant growth promoting rhizobacteria Ochrobactrum lupini KUDC1013 and Novosphingobium pentaromativorans KUDC1065 isolated from Dokdo Island, S. Korea are capable of eliciting induced systemic resistance (ISR) in pepper against bacterial spot disease. The present study aimed to determine whether plant growth-promoting rhizobacteria (PGPR) strains including strain KUDC1013, strain KUDC1065, and Paenibacillus polymyxa E681 either singly or in combinations were evaluated to have the capacity for potential biological control and plant growth promotion effect in the field trials. Under greenhouse conditions, the induced systemic resistance (ISR) effect of treatment with strains KUDC1013 and KUDC1065 differed according to pepper growth stages. Drenching of 3-week-old pepper seedlings with the KUDC-1013 strain significantly reduced the disease symptoms. In contrast, treatment with the KUDC1065 strain significantly protected 5-week-old pepper seedlings. Under field conditions, peppers treated with PGPR mixtures containing E681 and KUDC1013, either in a two-way combination, were showed greater effect on plant growth than those treated with an individual treatment. Collectively, the application of mixtures of PGPR strains on pepper might be considered as a potential biological control under greenhouse and field conditions.

Zygomycota Associated with Traditional Meju, a Fermented Soybean Starting Material for Soy Sauce and Soybean Paste: Seung-Beom Hong , Dae-Ho Kim , Mina Lee , Seong-Yeol Baek , Soon-wo Kwon , Jos Houbraken , Robert A. Samson; J. Microbiol. 2012;50(3):386-393. Published online June 30, 2012; DOI: https://doi.org/10.1007/s12275-012-1437-6

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37 Citations

Abstract: Various zygomycota species were detected during a study of the mycobiota of meju, a brick of dried fermented soybeans, used in Korean cuisine. Two hundred and sixty-eight strains were isolated from 98 finished meju products collected in various regions of Korea from 2009 to 2011, and 96 strains were isolated from in-process meju on various farms from 2010 to 2011. The isolated zygomycota were identified using phenotypic characteristics combined with DNA sequences of the internal transcribed spacer regions of ribosomal DNA and the D1/D2 nuclear ribosomal large subunit. Of 364 zygomycota strains, 108 were identified as Mucor circinelloides, 96 as M. racemosus, 60 as Lichtheimia ramosa, 22 as Rhizopus stolonifer, 16 as Lichtheimia corymbifera, and the other 62 strains comprised 10 other species. The psychrotrophic species, Mucor circinelloides and M. racemosus were predominantly present during low temperature fermentation (LTF) and the thermotolerant species Lichtheimia ramosa and Rhizomucor species were predominant during high temperature fermentation (HTF). The results suggest that temperature has a large influence on the zygomycota composition during the fermentation process of meju.

Paenibacillus xylaniclasticus sp. nov., a Xylanolytic-Cellulolytic Bacterium Isolated from Sludge in an Anaerobic Digester: Chakrit Tachaapaikoon , Somboon Tanasupawat , Patthra Pason , Somphit Sornyotha , Rattiya Waeonukul , Khin Lay Kyu , Khanok Ratanakhanokchai; J. Microbiol. 2012;50(3):394-400. Published online June 30, 2012; DOI: https://doi.org/10.1007/s12275-012-1480-3

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2 Citations

Abstract: A mesophilic, facultative, anaerobic, xylanolytic-cellulolytic bacterium, TW1T, was isolated from sludge in an anaerobic digester fed with pineapple waste. Cells stained Gram-positive, were spore-forming, and had the morphology of straight to slightly curved rods. Growth was observed in the temperature range of 30 to 50°C (optimum 37°C) and the pH range of 6.0 to 7.5 (optimum pH 7.0) under aerobic and anaerobic conditions. The strain contained meso-diaminopimelic acid in the cell-wall peptidoglycan. The predominant isoprenoid quinone was menaquinone with seven isoprene units (MK-7). Anteiso-C15:0, iso-C16:0, anteiso-C17:0, and C16:0 were the predominant cellular fatty acids. The G+C content of the DNA was 49.5 mol%. A phylogenetic analysis based on 16S rRNA showed that strain TW1T belonged within the genus Paenibacillus and was closely related to Paenibacillus cellulosilyticus LMG 22232T, P. curdlanolyticus KCTC 3759T, and P. kobensis KCTC 3761T with 97.7, 97.5, and 97.3% sequence similarity, respectively. The DNA-DNA hybridization values between the isolate and type strains of P. cellulosilyticus LMG 22232T, P. curdlanolyticus KCTC 3759T, and P. kobensis KCTC 3761T were found to be 18.6, 18.3, and 18.0%, respectively. The protein and xylanase patterns of strain TW1T were quite different from those of the type strains of closely related Paenibacillus species. On the basis of DNA-DNA relatedness and phenotypic analyses, phylogenetic data and the enzymatic pattern presented in this study, strain TW1T should be classified as a novel species of the genus Paenibacillus, for which the name Paenibacillus xylaniclasticus sp. nov. is proposed. The type strain is TW1T (=NBRC 106381T =KCTC 13719T =TISTR 1914T).

Tularemia Progression Accompanied with Oxidative Stress and Antioxidant Alteration in Spleen and Liver of BALB/c Mice: Miroslav Pohanka , Oto Pavlis , Branislav Ruttkay-Nedecky , Jiri Sochor , Jakub Sobotka , Jiri Pikula , Vojtech Adam , Rene Kizek; J. Microbiol. 2012;50(3):401-408. Published online June 30, 2012; DOI: https://doi.org/10.1007/s12275-012-1621-8

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10 Citations

Abstract: Francisella tularensis is the causative agent of tularemia. It is an intracellular pathogen with the ability to survive within phagosomes and induce pyroptotic cell death. In this study, we attempted to prove whether oxidative imbalance plays a significant role in tularemia pathogenesis. In our experimental model, we subcutaneously infected female BALB/c mice (dose 105 CFU of F. tularensis LVS). Liver, spleen, and blood were collected from mice at regular intervals from days 1–15 after infection. The bacterial burden was assessed by a cultivation test. The burden was unchanging from the 2nd to 6th day after infection. The bacterial burden corresponded to the plasmatic level of IFN-γ, IL-6, and liver malondialdehyde. After the phase of acute bacteraemia and the innate immunity reaction, the levels of reduced glutathione and total low molecular weight antioxidants decreased significantly and the activity of caspase-3 increased in the liver. The level of reduced glutathione decreased to 25% of the original level, and the total level of low molecular weight antioxidants was less than 50% of the initial amount. The demonstrated effects of tularemia-induced pathology had a more extensive impact on the liver than on the spleen.

Journal Article

Disruption of SCO5461 Gene Coding for a Mono-ADP-Ribosyltransferase Enzyme Produces a Conditional Pleiotropic Phenotype Affecting Morphological Differentiation and Antibiotic Production in Streptomyces coelicolor: Krisztina Szirák , Judit Keser&# , Sándor Biró , Iván Schmelczer , György Barabás , András Penyige; J. Microbiol. 2012;50(3):409-418. Published online June 30, 2012; DOI: https://doi.org/10.1007/s12275-012-1440-y

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Abstract: The SCO5461 gene of Streptomyces coelicolor A3(2) codes for an ADP-ribosyltransferase enzyme that is predicted to be a transmembrane protein with an extracellular catalytic domain. PCR-targeted disruption of the gene resulted in a mutant that differentiated normally on complex SFM medium; however, morphological differentiation in minimal medium was significantly delayed and this phenotype was even more pronounced on osmotically enhanced minimal medium. The mutant did not sporulate when it was grown on R5 medium, however the normal morphological differentiation was restored when the strain was cultivated beside the wild-type S. coelicolor M145 strain. Comparison of the pattern of ADP-ribosylated proteins showed a difference between the mutant and the wild type, fewer modified proteins were present in the cellular crude extract of the mutant strain. These results support our previous suggestions that protein ADP-ribosylation is involved in the regulation of differentiation and antibiotic production and secretion in Streptomyces.

Research Support, Non-U.S. Gov't

Evaluation of the Cell Growth of Mycobacteria Using Mycobacterium smegmatis mc2 155 as a Representative Species: Jorge A. Gonzalez-y-Merchand , Ruben Zaragoza-Contreras , Rosalina Guadarrama-Medina , Addy C. Helguera-Repetto , Sandra Rivera-Gutierrez , Jorge F. Cerna-Cortes , Leopoldo Santos-Argumedo , Robert A. Cox; J. Microbiol. 2012;50(3):419-425. Published online June 30, 2012; DOI: https://doi.org/10.1007/s12275-012-1556-0

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7 Citations

Abstract: The study of the in vitro cell growth of mycobacteria still remains a fastidious, difficult, and time-consuming procedure. In addition, assessing mycobacterial growth in the laboratory is often complicated by cell aggregation and slow growth-rate. We now report that the use of a stainless steel spring in the culture led to an absence of large cell clumps, to a decrease of dead cells in the exponential phase and to growth of a more homogeneous population of large cells. We also report that flow cytometry is a rapid, simple and reliable approach to monitor mycobacterial cell growth and viability. Here, we monitored Mycobacterium smegmatis cellular growth by optical density, dry cell mass, and colony forming units; in addition, viability, cell size and granularity profiles were analyzed by flow cytometry, and cell morphology by electron microscopy. Cultures monitored by flow cytometry may lead to a better understanding of the physiology of mycobacteria. Moreover, this methodology may aid in characterizing the cell growth of other fastidious species of microorganisms.

Journal Article

Regulatory Role of cAMP Receptor Protein over Escherichia coli Fumarase Genes: Yu-Pei Chen , Hsiao-Hsien Lin , Chi-Dung Yang , Shin-Hong Huang , Ching-Ping Tseng; J. Microbiol. 2012;50(3):426-433. Published online June 30, 2012; DOI: https://doi.org/10.1007/s12275-012-1542-6

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8 Citations

Abstract: Escherichia coli expresses three fumarase genes, namely, fumA, fumB, and fumC. In the present study, catabolite repression was observed in the fumA-lacZ and fumC-lacZ fusion strains, but not in the fumB-lacZ fusion strain. The Crp-binding sites in fumA and fumC were identified using an electrophoretic mobility shift assay and footprint analysis. However, the electrophoretic mobility shift assay did not detect band shifts in fumB. Fnr and ArcA serve as transcription regulators of fumarase gene expression. In relation to this, different mutants, including Δcya, Δcrp, Δfnr, and ΔarcA, were used to explore the regulatory role of Crp over fumA and fumC. The results show that Crp is an activator of fumA and fumC gene expression under various oxygen conditions and growth rates. ArcA was identified as the dominant repressor, with the major repression occurring at 0–4% oxygen. In addition, Fnr was confirmed as a repressor of fumC for the first time. This study elucidates the effects of Crp on fumarase gene expression.

Research Support, Non-U.S. Gov't

Extracellular Stress and Lipopolysaccharide Modulate Acinetobacter baumannii Surface-Associated Motility: Christin N. McQueary , Benjamin C. Kirkup , Yuanzheng Si , Miriam Barlow , Luis A. Actis , David W. Craft , Daniel V. Zurawski; J. Microbiol. 2012;50(3):434-443. Published online June 30, 2012; DOI: https://doi.org/10.1007/s12275-012-1555-1

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65 Citations

Abstract: Acinetobacter baumannii is a nosocomial bacterial pathogen, and infections attributed to this species are further complicated by a remarkable ability to acquire antimicrobial resistance genes and to survive in a desiccated state. While the antibiotic resistance and biofilm formation of A. baumannii is well-documented, less is known about the virulence attributes of this organism. Recent studies reported A. baumannii strains display a motility phenotype, which appears to be partially dependent upon Type IV pili, autoinducer molecules, and the response to blue light. In this study, we wanted to determine the prevalence of this trait in genetically diverse clinical isolates, and any additional required factors, and environmental cues that regulate motility. When strains are subjected to a wide array of stress conditions, A. baumannii motility is significantly reduced. In contrast, when extracellular iron is provided or salinity is reduced, motility is significantly enhanced. We further investigated whether the genes required for the production of lipopolysaccharide (lpsB) and K1 capsule (epsA/ptk) are required for motility as demonstrated in other Gram-negative bacteria. Transposon mutagenesis resulted in reduced motility by the insertion derivatives of each of these genes. The presence of the parental allele provided in trans, in the insertion mutant background, could only restore motility in the lpsB mutant. The production of core LPS directly contributes to the motility phenotype, while capsular polysaccharide may have an indirect effect. Further, the data suggest motility is regulated by extracellular conditions, indicating that A. baumannii is actively sensing the environment and responding accordingly.

Journal Article

Identification and Methicillin Resistance of Coagulase-Negative Staphylococci Isolated from Nasal Cavity of Healthy Horses: Jolanta Karakulska , Karol Fijałkowski , Paweł Nawrotek , Anna Pobucewicz , Filip Poszumski , Danuta Czernomysy-Furowicz; J. Microbiol. 2012;50(3):444-451. Published online June 30, 2012; DOI: https://doi.org/10.1007/s12275-012-1550-6

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31 Citations

Abstract: The aim of this study was an analysis of the staphylococcal flora of the nasal cavity of 42 healthy horses from 4 farms, along with species identification of CoNS isolates and determination of resistance to 18 antimicrobial agents, particularly phenotypic and genotypic methicillin resistance. From the 81 swabs, 87 staphylococci were isolated. All isolates possessed the gap gene but the coa gene was not detected in any of these isolates. Using PCR-RFLP of the gap gene, 82.8% of CoNS were identified: S. equorum (14.9%), S. warneri (14.9%), S. sciuri (12.6%), S. vitulinus (12.6%), S. xylosus (11.5% ), S. felis (5.7%), S. haemolyticus (3.4%), S. simulans(3.4%), S. capitis (1.1%), S. chromogenes (1.1%), and S. cohnii subsp. urealyticus (1.1%). To our knowledge, this was the first isolation of S. felis from a horse. The species identity of the remaining Staphylococcus spp. isolates (17.2%) could not be determined from the gap gene PCR-RFLP analysis and 16S rRNA gene sequencing data. Based on 16S-23S intergenic transcribed spacer PCR, 11 different ITS-PCR profiles were identified for the 87 analyzed isolates. Results of API Staph were consistent with molecular identification of 17 (19.5%) isolates. Resistance was detected to only 1 or 2 of the 18 antimicrobial agents tested in the 17.2% CoNS isolates, including 6.9% MRCoNS. The mecA gene was detected in each of the 5 (5.7%) phenotypically cefoxitin-resistant isolates and in 12 (13.8%) isolates susceptible to cefoxitin. In total, from 12 horses (28.6%), 17 (19.5%) MRCoNS were isolated. The highest percentage of MRCoNS was noted among S. sciuri isolates (100%).

Research Support, Non-U.S. Gov'ts

Identification of a New Bacillus licheniformis Strain Producing a Bacteriocin-Like Substance: Yaoqi Guo , Zhanqiao Yu , Jianhua Xie , Rijun Zhang; J. Microbiol. 2012;50(3):452-458. Published online June 30, 2012; DOI: https://doi.org/10.1007/s12275-012-2051-3

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20 Citations

Abstract: The emergence of antibiotic resistance has spurred a great number of studies for development of new antimicrobials in the past decade. The purpose of this study was to screen environmental samples for Bacillus strains producing potent antimicrobial agents. A new strain, which showed strong antimicrobial activity against Staphylococcus aureus and Salmonella enterica ser. Pullorum, was isolated from soil and designated as B116. This new isolate was identified as Bacillus licheniformis by morphological, biochemical and genetic analyses. The production of bacteriocin-like substance (BLS) started at early exponential phase and achieved highest level at early stationary phase. The BLS was precipitated by ammonium sulfate and its molecular mass was determined as ~4 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Culture supernatant of the new isolate exhibited antimicrobial activity against both Gram-positive and Gram-negative bacteria, including Bacillus cereus, Staphylococcus aureus, Listeria monocytogenes, Micrococcus luteus, Escherichia coli, and Salmonella spp. The BLS was resistant to heat, acid and alkaline treatment. Activity of the BLS was totally lost after digestion by pronase and partially lost after digestion by papain and lipase. The new isolate and relevant BLS are potentially useful in food and feed applications.

Molecular Screening of Streptomyces Isolates for Antifungal Activity and Family 19 Chitinase Enzymes: Youssuf Gherbawy , Hesham Elhariry , Abdulla Altalhi , Bahig El-Deeb , Ghada Khiralla; J. Microbiol. 2012;50(3):459-468. Published online June 30, 2012; DOI: https://doi.org/10.1007/s12275-012-2095-4

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33 Citations

Abstract: Thirty soil-isolates of Streptomyces were analyzed to determine their antagonism against plant-pathogenic fungi including Fusarium oxysporum, Pythium aristosporum, Colletotrichum gossypii, and Rhizoctonia solani. Seven isolates showed antifungal activity against one or more strain of the tested fungi. Based on the 16S rDNA sequence analysis, these isolates were identified as Streptomyces tendae (YH3), S. griseus (YH8), S. variabilis (YH21), S. endus (YH24), S. violaceusniger (YH27A), S. endus (YH27B), and S. griseus (YH27C). The identity percentages ranged from 98 to 100%. Although some isolates belonged to the same species, there were many differences in their cultural and morphological characteristics. Six isolates out of seven showed chitinase activity according to a chitinolytic activity test and on colloidal chitin agar plates. Based on the conserved regions among the family 19 chitinase genes of Streptomyces sp. two primers were used for detection of the chitinase (chiC) gene in the six isolates. A DNA fragment of 1.4 kb was observed only for the isolates YH8, YH27A, and YH27C. In conclusion, six Streptomyces strains with potential chitinolytic activity were identified from the local environment in Taif City, Saudi Arabia. Of these isolates, three belong to family 19 chitinases. To our knowledge, this is the first reported presence of a chiC gene in S. violaceusniger YH27A.

Identification and Characterization of Host Factors Interacting with Bombyx mori Nucleopolyhedrovirus ORF8: WonKyung Kang , Susumu Katsuma , Noriko Matsuda-Imai , Masaaki Kurihara , Toyoshi Yoshiga , Toru Shimada , Shogo Matsumoto; J. Microbiol. 2012;50(3):469-477. Published online June 30, 2012; DOI: https://doi.org/10.1007/s12275-012-2010-z

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4 Citations

Abstract: The orf8 gene (Bm8) in Bombyx mori nucleopolyhedrovirus (BmNPV) is one of 17 genes unique to group I NPVs and is expressed as an early gene. We have reported that Bm8 may play an important role during viral infection and that Bm8 protein co-localized with IE1 to specific nuclear foci throughout infection. It was also demonstrated that both IE1 and BmNPV hr facilitate this localization of Bm8. To investigate further, host proteins interacting with Bm8 were screened using a yeast two-hybrid system. We identified 6 host clones as Bm8-interacting partners from three cDNA libraries derived from BmN cells or B. mori larvae. Further assays showed that the N-terminal region of Bm8 is important for the interaction with most host clones and that two of the clones can associate with IE1. Cloning and sequencing of full-length cDNAs revealed that most of the clones potentially encode either membrane-bound proteins or secreted proteins. Quantitative RT-PCR analysis revealed that some of these host genes were slightly induced during the early stage of infection in BmN cells, and that the expression of all genes was markedly reduced during the late stage of infection. Generation of mutant BmNPVs over-expressing these host genes also identified a gene that potentially functions as a negative factor during BmNPV infection. These features of Bm8-interacting host proteins strongly support that Bm8 is a multifunctional protein involved in multiple signaling pathways in host cells.

Evaluation of the Efficacy of a Pre-pandemic H5N1 Vaccine (MG1109) in Mouse and Ferret Models: Min-Suk Song , Ho-Jin Moon , Hyeok-il Kwon , Philippe Noriel Q. Pascua , Jun Han Lee , Yun Hee Baek , Kyu-Jin Woo , Juhee Choi , Sangho Lee , Hyunseung Yoo , In gyeong Oh , Yeup Yoon , Jong-Bok Rho , Moon-Hee Sung , Seung-Pyo Hong , Chul-Joong Kim , Young Ki Choi; J. Microbiol. 2012;50(3):487-488.

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Abstract: The threat of a highly pathogenic avian influenza (HPAI) H5N1 virus causing the next pandemic remains a major concern. In this study, we evaluated the immunogenicity and efficacy of an inactivated whole-virus H5N1 pre-pandemic vaccine (MG1109) formulated by Green Cross Co., Ltd containing the hemagglutinin (HA) and neuraminidase (NA) genes of the clade 1 A/Vietnam/1194/04 virus in the backbone of A/Puerto Rico/8/34 (RgVietNam/04xPR8/34). Administration of the MG1109 vaccine (2-doses) in mice and ferrets elicited high HI and SN titers in a dose-dependent manner against the homologous (RgVietNam/04xPR8/34) and various heterologous H5N1 strains, (RgKor/W149/06xPR8/34, RgCambodia/04xPR8/34, RgGuangxi/05xPR8/34), including a heterosubtypic H5N2 (A/Aquatic bird/orea/W81/05) virus. However, efficient cross-reactivity was not observed against heterosubtypic H9N2 (A/Ck/Korea/H0802/08) and H1N1 (PR/8/34) viruses. Mice immunized with 1.9 μg HA/dose of MG1109 were completely protected from lethal challenge with heterologous wild-type HPAI H5N1 A/EM/Korea/W149/06 (clade 2.2) and mouse-adapted H5N2 viruses. Furthermore, ferrets administered at least 3.8 μg HA/dose efficiently suppressed virus growth in the upper respiratory tract and lungs. Vaccinated mice and ferrets also demonstrated attenuation of clinical disease signs and limited virus spread to other organs. Thus, this vaccine provided immunogenic responses in mouse and ferret models even against challenge with heterologous HPAI H5N1 and H5N2 viruses. Since the specific strain of HPAI H5N1 virus that would potentially cause the next outbreak is unknown, pre-pandemic vaccine preparation that could provide crossprotection against various H5 strains could be a useful approach in the selection of promising candidate vaccines in the future.

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