Restriction Digest Profiles Research Articles

Molecular investigation of two novel bacteriophages of a facultative methylotroph, Raoultella ornithinolytica: first report of Raoultella phages.

Bacteria of the genus Raoultella are known to inhabit aquatic environments and can be found in medical samples. The pathogenicity of Raoultella ornithinolytica isolates in human has become increasingly important, and several cases of infections have been reported recently. However, there are no reports of isolation of bacteriophages infecting this bacterium. In this study, two novel phages (ISF3 and ISF6) of a methylotrophic Raoultella strain were isolated from sewage. To characterize the isolated phages, morphological features, protein profiles, restriction digestion patterns, and partial genome sequences were studied. Despite morphological differences, electron microscopy revealed that both phages had an icosahedral capsid connected to a contractile tail, suggesting that ISF3 and ISF6 both belong to the family Myoviridae. Partial nucleotide sequences of the ISF3 genome showed 99% to 100% identity to DNA of Klebsiella pneumonia phages KP15, KP27 and BMBT1; however, the restriction digestion profiles of ISF3 genome digested by EcoRI and EcoRV differed from those of Klebsiella phages KP15 and KP27. A partial sequence alignment showed that ISF6 can be classified as a member of a new viral genus due to its significant differences from other previously identified phages. To the best of our knowledge, this is the first report of the isolation and characterization of the specific Raoultella phages that have potential to be used as new pharmaceuticals against R. ornithinolytica.

National collection of type cultures: the bacteriophage and plasmid collections and repositories

The National Collection of Type Cultures (NCTC) is the world’s oldest bacterial collection that was specifically established to provide strains globally to support scientific research. In addition to the general catalogue NCTC has a fully curated bacteriophage and plasmid archive that to date has not recently been made available to the wider scientific community. The NCTC bacteriophage collection consists of over 100 bacteriophage and their corresponding bacterial hosts which were originally deposited primarily for their value in bacterial typing. The collection consists of bacteriophage from the following hosts: Streptococcus agalactiae, Staphylococcus aureus, Escherichia coli and Campylobacter. The NCTC Datta plasmid collection is a curated collection of over 500 unique plasmids which were originally curated to examine the biology of plasmid transfer between and within bacterial strains. The NCTC bacteriophage collection is currently being fully characterised using a range of modern methods including genomic sequencing and electron microscopy. The plasmid collection is also being characterised using genomic sequencing and restriction digest profiles. It is intended that once characterised and rebanked both the plasmid and bacteriophage collections will be made available in 2019 to scientists to support research and development. The NCTC bacteriophage and plasmid collections will both be dynamic, representing an active repositories into which microbiologists can deposit phages and plasmids to support accessibility and reproducibility in science.

Bacteriophages with potential to inactivate Salmonella Typhimurium: Use of single phage suspensions and phage cocktails

The aim of this study was to compare the dynamics of three previously isolated bacteriophages (or phages) individually (phSE-1, phSE-2 and phSE-5) or combined in cocktails of two or three phages (phSE-1/phSE-2, phSE-1/phSE-5, phSE-2/phSE-5 and phSE-1/phSE-2/phSE-5) to control Salmonella enterica serovar Typhimurium (Salmonella Typhimurium) in order to evaluate their potential application during depuration. Phages were assigned to the family Siphoviridae and revealed identical restriction digest profiles, although they showed a different phage adsorption, host range, burst size, explosion time and survival in seawater. The three phages were effective against S. Typhimurium (reduction of ∼2.0 log CFU/mL after 4h treatment). The use of cocktails was not significantly more effective than the use of single phages. A big fraction of the remained bacteria are phage-resistant mutants (frequency of phage-resistant mutants 9.19×10−5–5.11×10−4) but phage- resistant bacterial mutants was lower for the cocktail phages than for the single phage suspensions and the phage phSE-1 presented the highest rate of resistance and phage phSE-5 the lowest one. The spectral changes of S. Typhimurium resistant and phage-sensitive cells were compared and revealed relevant differences for peaks associated to amide I (1620cm−1) and amide II (1515cm−1) from proteins and from carbohydrates and phosphates region (1080–1000cm−1). Despite the similar efficiency of individual phages, the development of lower resistance indicates that phage cocktails might be the most promising choice to be used during the bivalve depuration to control the transmission of salmonellosis.

Gender Specific Differences in RNA Polymerase III Transcription.

RNA polymerase (pol) III transcribes a variety of untranslated RNAs responsible for regulating cellular growth and is deregulated in a variety of cancers. In this study, we examined gender differences in RNA pol III transcription in vitro and in vivo. Expression levels of U6 snRNA, tMet, and known modulators of RNA pol III transcription were assayed in male and female derived adenocarcinoma (AC) lung cancer cell lines and male and female C57BL/6J mice using real time quantitative PCR. Methylation status of the U6 snRNA promoter was determined for lung and liver tissue isolated from male and female C57BL/6J mice by digesting genomic DNA with methylation sensitive restriction enzymes and digestion profiles were analyzed by qPCR using primers spanning the U6 promoter. Here, we demonstrate that RNA pol III transcription is differentially regulated by EGCG in male and female derived AC lung cancer cell lines. Basal RNA pol III transcript levels are significantly different in male and female derived AC lung cancer cell lines. These data prompted an investigation of gender specific differences in RNA pol III transcription in vivo in lung and liver tissue. Herein, we report that U6 snRNA RNA pol III transcription is significantly stimulated in the liver tissue of male C57BL/6J mice. Further, the increase in U6 transcription correlates with a significant inhibition in the expression of p53, a negative regulator of RNA pol III transcription, and demethylation of the U6 promoter in the liver tissue of male C57BL/6J mice. To the best of our knowledge, this is the first study demonstrating gender specific differences in RNA pol III transcription both in vivo and in vitro and further highlights the need to include both male and female cell lines and animals in experimental design.

Catfish Species Identification Using Lab-On-Chip PCR-RFLP

Lab-on-a-chip based Polymerase Chain Reaction-Restriction Fragment Length Polymorphism (PCR-RFLP) technology was developed for the identification of seven catfish species including an Ictalurid hybrid. RFLP profiles of mitochondrial cytochrome b fragments digested by three different restriction endonucleases (DdeI, HaeIII, and NlaIII) were constructed from morphologically verified catfish samples on an Agilent 2100 Bioanalyzer (Agilent Technologies, Inc., Santa Clara, CA, USA). High sensitivity and ease of handling through end-point microchip-based capillary electrophoresis increased resolution and accuracy of DNA fragment sizing. RFLP profiles derived from a combination of all three enzymes produced consistent species-specific identification profiles. Observed restriction fragment patterns for Clarias batrachus and Ictalurus punctatus were similar, but both of these species could be consistently differentiated using a single band of the HaeIII restriction site. Assay advantages due to shorter assay times, assay ease, and minimum usage of harmful solvents and chemicals when compared to traditional DNA barcoding were counterbalanced by the need to develop and optimize specific restriction digest profiles for all potential species of interest. With further development, this method may be utilized in testing of catfish products to ensure the enforcement of seafood labeling regulations.

Polymorphisms in Exon 2 of MHC Class II DRB3 Gene of 10 Domestic Goats in Southwest China

Polymorphism of the second exon of the caprine leukocyte antigen-DRB3 gene (CLA-DRB3*02) was investigated in this study. The 285 bp PCR product of 258 individuals from 10 domestic goat breeds in Southwest China was digested with restriction endonucleases PstI and HaeIII and then genotyped. Three alleles and 4 restriction digestion profiles were distinguished by digestion of the PCR fragment by PstI, and 8 alleles and 13 genotypes by HaeIII. For HaeIII restriction enzyme sites, the Chi-square (X(2)) test showed that all goat breeds in this study did not fit with the Hardy-Weinberg equilibrium (p<0.01 or p<0.05). The highly polymorphic nature of CLA-DRB3*02 was demonstrated and the ranges of gene heterozygosity (He) and polymorphism information content (PIC) were 0.36-0.63 and 0.32-0.55, respectively. Clustering analysis showed that the 10 goat breeds clustered into two groups and Dazu Black goat had a close genetic relationship with Chengdu Grey, Jintang Black and Nanjiang Yellow goats.

First Report of a "Candidatus Phytoplasma australiense"-Related Strain in Lucerne (Medicago sativa) in Australia.

Australian lucerne yellows (ALuY), a phytoplasma-associated disease, is a major problem in Australia that causes the pasture seed industry millions of dollars in losses annually (3). Samples were collected from lucerne (Medicago sativa L.) plants exhibiting symptoms indicative of ALuY (4) in a seed lucerne paddock (cv CW 5558) at Griffith, southwestern New South Wales (NSW), Australia, in November 2005 and again in January 2006. Samples were kept at 4°C and processed within 36 h of collection. Total DNA was extracted from approximately 0.3 g of leaf midribs and petioles of each plant sample and used as template in a nested PCR assay with phytoplasma universal primer pairs P1/P7 and fU5/m23sr. PCR products resulting from the first amplification were diluted (1:30) with sterile distilled water (SDW) before reamplification with fU5/m23sr. DNA of Australian tomato big bud (TBB) phytoplasma and SDW were used as positive and negative assay controls, respectively. Ten of fifteen plant samples collected in November tested positive for phytoplasma DNA. Restriction digestion profiles of nested PCR amplicons with HpaII endonuclease were the same for all symptomatic plants but differed from the control. Phytoplasma identity was determined by sequencing two nested PCR products that yielded identical sequences. One was deposited in the GenBank database (Accession No. DQ786394). BLAST analysis of the latter sequence revealed a >99.6% similarity with "Candidatus Phytoplasma australiense" (L76865) and related strains papaya dieback (Y10095), phormium yellow leaf (U43570), strawberry green petal (AJ243044), and strawberry lethal yellows (AJ243045). Direct PCR with primers FP 5'-GCATGTCGCGGTGAATAC-3' and RY 5'-TGAGCTATAGGCCCTTAATC-3' designed to specifically amplify DNA of "Ca. P. australiense" detected the phytoplasma in 8 of 40 samples collected in January. Whether this phytoplasma is the etiological agent solely responsible for ALuY is currently under investigation. "Ca. P. asteris" and stolbur group (16SrXII) phytoplasmas have been reported in lucerne in the United States (2) and Italy (1), respectively. Within the stolbur group 16SrXII, "Ca. P. australiense" and stolbur phytoplasma are regarded as separate species and both are distinct from "Ca. P. asteris", a group 16SrI strain. To our knowledge, this is the first report of a "Ca. P. australiense" related strain in lucerne.

Allelic variations in exon 2 of caprine MHC class II DRB3 gene in Chinese indigenous goats

Allelic variations in the second exon of caprine leukocyte antigen-DRB3 gene (CLA-DRB3*02) were investigated. A total of 459 animals from 12 Chinese indigenous goat populations were genotyped. Six alleles and 18 restriction digestion profiles were distinguished by digestion of PCR amplification product of CLA-DRB3*02 with HaeIII. The results suggested that there were significant difference ( P < 0.000) in both allele frequencies and genotype frequencies of exon 2 restriction fragment length polymorphisms (RFLPs) among the breeds. The new allele CLA-DRB3*0206 was identified only in three goat populations from the three-gorge reservoir, and the breed-specific allele CLA-DRB3*0205 was merely detected in the three populations of Tibetan goat. The results also indicated that the potentially different ecological factors, such as climate, disease, topography, pasture conditions and pathogens might be account for the significantly different distribution of alleles and genotypes among the goat populations. The nucleotide and predicted amino acid sequence alignment of CLA-DRB3*02 alleles provided evidence for considerable variation among the six alleles. Phylogenetic analysis showed that the six alleles of CLA-DRB3*02 were closely related. They clustered together and were separated from the corresponding sequences of sheep and cattle, which indicated the trans-species evolution model of CLA-DRB3*02 was not present in the Chinese goat populations studied.

New variations in intron 4 of growth hormone gene in Chinese native chickens.

Polymorphism in intron 4 of chicken growth hormone (cGH) gene was studied in 20 Chinese native chicken populations and broiler or layer populations. A total of eight restriction digestion profiles were identified in intron 4 and confirmed by sequencing. Among 20 populations, there were distinctively different allele numbers and frequencies of intron 4 restriction fragment length polymorphisms (RFLPs) between Chinese native chickens and broilers or layers. Two new alleles, allele D and allele E, were identified in Taihe Silkies. Allele D was also identified in other Chinese native breeds and a 50 bp fragment deletion was identified in allele E.

Chloroplast DNA diversity in the wild shrub Cytisus scoparius L. (Leguminosae)

Cytoplasmic DNA (chloroplast and mitochondrial DNA) of 40 Cytisus scoparius individuals collected from four deciduous forests across Europe were studied using the PCR-RFLP technique. Twenty-one pairs of universal primers in combination with four restriction enzymes were screened for cpDNA and mtDNA amplification and restriction digestion profiles of the amplified fragments. In all, 10 primer-restriction enzyme combinations showed polymorphisms. Fourteen cpDNA haplotypes were found, of which 10 are unique to a population. All the populations are polymorphic in their haplotype composition. Intraspecific cpDNA diversity detected in this species will be useful for population genetic studies. High cpDNA diversity detected in the study must be dealt with cautiously when using chloroplast genomes for phylogenetic studies involving this species.

Human monoclonal autoantibodies to B-cell epitopes outside the thyroid peroxidase autoantibody immunodominant region.

Human autoantibodies to thyroid peroxidase (TPO) interact with a restricted or immunodominant region (IDR) on intact TPO. However, a smaller proportion of polyclonal serum TPO autoantibodies bind outside this region. To isolate monoclonal nonimmunodominant region (non-IDR) TPO autoantibodies, we screened a thyroid-derived immunoglobulin gene phage display library while "epitope masking" the TPO IDR with four human TPO monoclonal autoantibodies that define the IDR. Among 31 non-IDR autoantibodies obtained (expressed as Fab), 8 representatives were analyzed further based on their restriction digestion profiles. All are encoded by almost identical H chains (VH3 family), with extremely long D regions, paired with three different types of light chains. In contrast, IDR TPO Fab from the same patient utilize seven different heavy chains (VH1 and VH5 families) paired nonpromiscuously with different light chains. Use of VH5 genes has not been reported previously for TPO autoantibodies. Both non-IDR and IDR Fab bind specifically to TPO and not to other proteins. The non-IDR Fab affinities for TPO are moderately high (Kd 1-2 x 10(-9) M), somewhat lower than those for most IDR Fab (Kd 1-4 x 10(-10) M). The epitopes of the three types of non-IDR Fab overlap with each other, indicating a major role for their heavy chain in TPO binding. Most importantly, the epitopes of non-IDR Fab are recognized by patients' serum autoantibodies. In summary, we provide the first insight into the immunoglobulin genes, affinities and epitopes of human monoclonal autoantibodies that bind outside the TPO-immunodominant region.

Analysis of restriction profiles of mitochondrial DNA from Sporothrix schenckii.

Mitochondrial DNA (mtDNA) diversity was analyzed in 42 clinical isolates of Sporothrix schenckii from Mexico (n = 29), Guatemala (n = 4) and Colombia (n = 9). Based on HaeIII restriction digestion profiles, the isolates were classified into eight types. In addition to 24 mtDNA types previously reported in another study, 6 new types were found in this study. Most of the strains belong to type 14 and type 30, the former restricted to Mexico, whereas the latter was distributed in Mexico, Guatemala and Colombia. The new types (25-30) were identified in Mexico, Guatemala and Colombia. Restriction-fragment length polymorphism in mtDNA of S. schenckii revealed high levels of genetic variation attributable to differences in restriction sites as well as in mtDNA size. Based on genetic distances S. schenckii types were clustered into two main groups.

Distribution and molecular characterization of the root endophyte Phialocephala fortinii along an environmental gradient in the boreal forest of Alberta

Phialocephala fortinii is a common root endophytic fungus with a wide geographic distribution and little, if any, host specificity. Little is known about its habitat specificity, although there is evidence to suggest that high water tables may restrict the occurrence of P. fortinii in wetlands. We tested this hypothesis by determining the distribution of P. fortinii along a sand dune – wetland complex. Isolates of P. fortinii, identified on the basis of cultural and morphological characteristics, were obtained from the roots of vascular plants across the moisture gradient. Three ‘cultural groups’ were recognized among these isolates. Thirty-three of these isolates were compared among themselves and to strains of known identity using PCR/RFLP analysis of the ITS region and a portion of the 28S subunit of rDNA. The restriction digest profiles of all isolates were identical to those of P. fortinii for 4 restriction enzymes. DNA sequences, from a subset of these strains, showed a low percent sequence divergence confirming the reliability of the RFLP data. The same analyses were done with two strains of Leptodontidium orchidicola a culturally similar root endophyte, to ensure that this taxon was not among the transect isolates. DNA data showed a clear difference between P. fortinii and L. orchidicola but did not discriminate among cultural groups. Thus, P. fortinii showed no habitat specificity and occurred in both xeric and hydric sites. RFLP profiles and ITS sequences showed little variation among isolates of P. fortinii and among the isolates of L. orchidicola.

Monophyly of endosymbiont containing trypanosomatids: phylogeny versus taxonomy.

To obtain additional information on the phylogenetic relationships within the family Trypanosomatidae (order Kinetoplastida), we have sequenced the small subunit ribosomal RNA genes from the endosymbiont containing species Herpetomonas roitmani TCC080, Herpetomonas sp. TCC263, Crithidia oncopelti ATCC 12982 and a partial large subunit rRNA gene from H. roitmani. The small subunit sequences in the two isolates of Herpetomonas are very similar but not identical, and so are their restriction digest profiles of kinetoplast DNA. The size of minicircles in both isolates is 4.2 kilobases. The inferred ribosomal RNA phylogenetic trees shows the genera Herpetomonas and Crithidia as polyphyletic. Endosymbiont-bearing herpetomonads cluster with the endosymbiont-bearing crithidias and a blastocrithidia to form a monophyletic clade, whereas the endosymbiont-free members of these genera are found elsewhere in the tree. These data support the hypothesis of a monophyletic origin of endosymbiosis in trypanosomatid evolution and also suggest that a taxonomic revision is needed in order to better describe the natural affinities in this family.

Specific Amplification and Restriction Polymorphisms of the Cyanobacterial rRNA Operon Spacer Region

Summary Forty-two strains of cyanobacteria commonly associated with toxic bloom events and representing 10 cyanobacterial genera were examined by RFLP analysis of the PCR amplified 16S-23S rRNA gene internal transcribed spacer (ITS). A total of 97 different DNA profiles were generated by the application of 8 restriction endonucleases to digest the PCR products. The length of the PCR products obtained for strains assigned to the same genus seemed to be a useful taxonomic character and could probably be used for rapid identification. Nine characteristic amplification products delineated the 10 genera studied. Intrageneric strain differentiation was provided by restriction digest profiles which, when combined for each strain, resulted in 27 distinct genotypes. Specific amplification of cyanobacterial strains from mixed populations and environmental samples containing algae and heterotrophic bacteria was possible due to the use of a cyanobacteria specific 16S rRNA gene-directed PCR primer. The genetic relatedness observed between the taxa studied coincided with the taxonomic identification of the studied strains, particularly within the genera Anabaena and Microcystis.

Reversible fluconazole resistance in Candida albicans: a potential in vitro model.

To study the development and potential mechanisms of antifungal resistance in relation to antifungal exposure, reversible fluconazole resistance was examined in vitro. Candida albicans ATCC 36082 blastospores were passed in liquid yeast nitrogen base medium containing either 4, 8, 16, or 128 micrograms of fluconazole per ml, and susceptibility testing was performed after each passage. High-level fluconazole resistance (50% inhibitory concentration, > 256 micrograms/ml) developed in the isolates after serial passage in medium containing 8, 16, or 128 micrograms of fluconazole per ml, but not in isolates passed in 4 micrograms of fluconazole per ml. Reduced susceptibility was noted within four to seven passages, which was equivalent to 14 to 19 days of exposure to the drug. However, all isolates returned to the susceptible phenotype after 8 to 15 passages in medium lacking the drug; thus, fluconazole resistance was reversible in vitro. In vivo, organisms retained the resistant phenotype after a single passage in the rabbit model of infective endocarditis. Restriction digest profiles and karyotypic analysis of the parent strain and selected fluconazole-resistant and -susceptible isolates from each group were identical. Investigations into the molecular mechanisms of this reversible resistance failed to reveal increased accumulation of mRNA for 14 alpha-demethylase, the target enzyme for fluconazole, or for the candidal multidrug transporters CDR1 and BENr. This process of continuous in vitro exposure to antifungal drug may be useful as a model for studying the effects of different antifungal agents and dosing regimens on the development of resistance and for defining the mechanism(s) of reversible resistance.

Presence of multiple amylases encoded by independent genes in Cellulomonas sp. NCIM 2353

Four genomic fragments encoding independent amylases from Cellulomonas sp. NCIM 2353 were cloned in Escherischia coli DH10B, using plasmid Bluescript. The enzyme encoded by these clones could be induced in the presence of starch, without IPTG. On zymogram amylase activity staining, two of these clones were seen to produce independent enzymes, corresponding to the Cellulomonas amylases. The restriction digestion profiles confirmed that these four DNA fragments are situated at different locations on the Cellulomonas genome. Some of the clones also harbored xylanase activity.

Allozyme and mitochondrial DNA separation of Pacific northern bluefin tuna, Thunnus thynnus orientalis (Temminck and Schlegel), from southern bluefin tuna, Thunnus maccoyii (Castelnau)

Northern and southern bluefin tunas are morphologically similar and can be misidentified, posing problems for fishery management and marketing. Allozyme variation and restriction-site variation in mitochondrial DNA (mtDNA) were used to distinguish between the two species. A survey of 36 allozyme loci active in white muscle and liver tissue showed that the genetic identity between the species was high (Nei's I = 0.907). One diagnostic locus (sAH*) and two nearly diagnostic loci (ADA* and GDA*) were found, and four loci showed highly significant allele frequency differences (FH*, GPI-A*, PGDH* and sSOD*). A survey of the mtDNA genome, using 15 restriction enzymes and southern blotting, revealed five restriction enzymes that gave species-diagnostic restriction digest profiles (Ban I, Bcl I, Dra I, Pvu II, Xba I) and a further three enzymes (Pst I, Barn HI and Nco I) with large haplotype frequency differences. Mitochondrial DNA analysis provided more reliable discrimination of specimens than did allozyme analysis, although the more rapid allozyme identification will be accurate for most specimens. The two biochemical genetic methods were then used to identify Australian-caught fish of uncertain identity. Six of 12 tuna originally considered to be northern bluefin tuna were confirmed as northern bluefin and six were identified as southern bluefin. The presence of northern bluefin tuna as far south as south-western Tasmania was confirmed.

Multiply resistant Klebsiella pneumoniae strains from two Chicago hospitals: identification of the extended-spectrum TEM-12 and TEM-10 ceftazidime-hydrolyzing beta-lactamases in a single isolate

Ceftazidime-resistant Klebsiella pneumoniae strains began to appear when ceftazidime usage was increased in two unrelated Chicago hospitals. These strains produced a beta-lactamase with an isoelectric point of 5.6 (RP-5.6) and strong hydrolyzing activity against ceftazidime. Two different restriction digest profiles were associated with the ceftazidime resistance plasmids. A second beta-lactamase with a pI of 5.2 (RP-5.2) was coproduced in two representative strains. The second beta-lactamase hydrolyzed ceftazidime, cefotaxime, and aztreonam with relative hydrolysis rates of < 8% of that observed for benzylpenicillin. Both enzymes were inhibited by clavulanic acid and tazobactam. Nucleotide sequencing of the genes coding for RP-5.2 and RP-5.6 revealed sequences identical to those of the TEM-12 and TEM-10 beta-lactamase genes, respectively. Both genes were derived from a TEM-1 sequence related to that of the gene encoded on the Tn2 transposon. Single point mutations are required to progress from TEM-1 to TEM-12 and from TEM-12 to TEM-10. Extracts from broths grown from single cell isolates of the strain producing TEM-12 and TEM-10 were shown to contain both enzymes. Transconjugants producing either the TEM-12 or the TEM-10 beta-lactamase were obtained. A significant finding was that both enzymes were encoded by plasmids with identical restriction digest patterns. These studies show that mutations leading to extended-spectrum beta-lactamases can occur sequentially in the same organism, with the genes encoding both enzymes maintained stably.

Biological and biophysical characteristics of phages isolated from Clostridium botulinum type C and D strains, and physicochemical properties of the phage DNAs.

Toxin-converting phages CE beta and d-16 phi and non-converting phages CE gamma and d-1', isolated from toxigenic strains C-468 and D-CB16 of Clostridium botulinum types C and D, respectively, were characterized biologically and biophysically. DNAs isolated from these four phages were studied physicochemically. Phages CE beta, d-16 phi, CE gamma and d-1' were adsorbed to their susceptible cells at the constant rates of 1.1 x 10(-8), 3.3 x 10(-8), 1.4 x 10(-8) and 1.4 x 10(-8) ml/min, and grew after latent periods of 35, 45, 35 and 35 min at the burst sizes of 38, 85, 35 and 35, respectively. Converting phages were more susceptible than non-converting phages to physical and chemical treatments such as temperature, pH, time, UV-irradiation and organic solvents. GC% of phage DNAs were determined by melting temperature, buoyant density and HPLC analysis to be 26, 26, 29 and 29% for CE beta, d-16 phi, CE gamma and d-1' phage DNAs, respectively. The restriction digestion profiles of phage DNAs with seven endonucleases were compared by agarose gel electrophoresis, revealing that the two converting phage DNAs were similar in regard to five enzyme digestion profiles, but different in the other two enzymatic profiles. Non-converting phage DNAs produced the same restriction digestion profiles with all seven endonucleases. The molecular sizes determined from the size of restriction enzyme digestion fragments were about 110 kb for converting phage CE beta and d-16 phi DNAs and 65 kb for non-converting phage CE gamma and d-1' DNAs. Dot blot hybridization experiments revealed that DNA homology between the converting phages CE beta and d-16 phi was 50-75%, while that between non-converting phages CE gamma and d-1' was about 100%. Converting phage and non-converting phage DNAs did not hybridize at all.