Smaller Burst Size Research Articles

Characterization of a novel genus of jumbo phages and their application in wastewater treatment

Phages widely exist in numerous environments from wastewater to deep ocean, representing a huge virus diversity, yet remain poorly characterized. Among them, jumbo phages are of particular interests due to their large genome (>200kb) and unusual biology. To date, only six strains of jumbo phages infecting Klebsiella pneumoniae have been described. Here, we report the isolation and characterization of two jumbo phages from hospital wastewater representing the sixth genus: φKp5130 and φKp9438. Both phages showed lytic activity against broad range of clinical antibiotic-resistant K. pneumoniae strains and distinct physiology including long latent period, small burst size, and high resistance to thermal and pH stress. The treatment of sewage water with the phages cocktail resulted in dramatic decline in K. pneumoniae population. Overall, this study provides detailed molecular and genomics characterization of two novel jumbo phages, expands viral diversity, and provides novel candidate phages to facilitate environmental wastewater treatment.

Isolation and characterization of a novel phage belonging to a new genus against Vibrio parahaemolyticus

BackgroundVibrio parahaemolyticus is a major foodborne pathogen that contaminates aquatic products and causes great economic losses to aquaculture. Because of the emergence of multidrug-resistant V. parahaemolyticus strains, bacteriophages are considered promising agents for their biocontrol as an alternative or supplement to antibiotics. In this study, a lytic vibriophage, vB_VpaM_R16F (R16F), infecting V. parahaemolyticus 1.1997T was isolated, characterized and evaluated for its biocontrol potential.MethodsA vibriophage R16F was isolated from sewage from a seafood market with the double-layer agar method. R16F was studied by transmission electron microscopy, host range, sensitivity of phage particles to chloroform, one-step growth curve and lytic activity. The phage genome was sequenced and in-depth characterized, including phylogenetic and taxonomic analysis.ResultsR16F belongs to the myovirus morphotype and infects V. parahaemolyticus, but not nine other Vibrio spp. As characterized by determining its host range, one-step growth curve, and lytic activity, phage R16F was found to highly effective in lysing host cells with a short latent period (< 10 min) and a small burst size (13 plaque-forming units). R16F has a linear double-stranded DNA with genome size 139,011 bp and a G + C content of 35.21%. Phylogenetic and intergenomic nucleotide sequence similarity analysis revealed that R16F is distinct from currently known vibriophages and belongs to a novel genus. Several genes (e.g., encoding ultraviolet damage endonuclease and endolysin) that may enhance environmental competitiveness were found in the genome of R16F, while no antibiotic resistance- or virulence factor-related gene was detected.ConclusionsIn consideration of its biological and genetic properties, this newly discovered phage R16F belongs to a novel genus and may be a potential alternate biocontrol agent.

When Plaquing Is Not Possible: Computational Methods for Detecting Induced Phages.

High-throughput sequencing of microbial communities has uncovered a large, diverse population of phages. Frequently, phages found are integrated into their bacterial host genome. Distinguishing between phages in their integrated (lysogenic) and unintegrated (lytic) stage can provide insight into how phages shape bacterial communities. Here we present the Prophage Induction Estimator (PIE) to identify induced phages in genomic and metagenomic sequences. PIE takes raw sequencing reads and phage sequence predictions, performs read quality control, read assembly, and calculation of phage and non-phage sequence abundance and completeness. The distribution of abundances for non-phage sequences is used to predict induced phages with statistical confidence. In silico tests were conducted to benchmark this tool finding that PIE can detect induction events as well as phages with a relatively small burst size (10×). We then examined isolate genome sequencing data as well as a mock community and urinary metagenome data sets and found instances of induced phages in all three data sets. The flexibility of this software enables users to easily include phage predictions from their preferred tool of choice or phage sequences of interest. Thus, genomic and metagenomic sequencing now not only provides a means for discovering and identifying phage sequences but also the detection of induced prophages.

Quorum sensing signal disrupts viral infection dynamics in the coccolithophore Emiliania huxleyi

Viruses that infect phytoplankton are abundant in all regions of the global ocean. Despite their ubiquity, little is understood regarding how biotic interactions can alter virus infection success as well as the fate of phytoplankton hosts. In previous work, the bacterially derived compound 2-heptyl-4-quinolone (HHQ) has been shown to protect the cosmopolitan coccolithophore Emiliania huxleyi from virus-induced mortality. The present study explores the potential mechanisms through which protection is conferred. Using a suite of transmission electron microscopy and physiological diagnostic staining techniques, we show that when E. huxleyi is exposed to HHQ, viruses can gain entry into cells but viral replication and release is inhibited. These findings are supported by a smaller burst size, as well as lower infectious and total virus production when the host is treated with nanomolar concentrations of HHQ. Additionally, diagnostic staining results indicate that programmed cell death markers commonly associated with viral infection are not activated when infected E. huxleyi are exposed to HHQ. Together, these results suggest that the ability of HHQ to inhibit infectious viral production protects the alga not from getting infected, but from cell lysis. This work identifies a new mechanistic role of bacterial quorum sensing molecules in mediating viral infections in marine microbial systems.

Characteristics of a novel temperate bacteriophage against Staphylococcus arlettae (vB_SarS_BM31).

Staphylococcus arlettae is a rarely reported coagulase-negative staphylococcus (CoNS) isolated from infected humans and livestock. Observing phage-bacteria interaction could improve the understanding of bacterial pathogenetic mechanisms, providing foundational evidence for phage therapy or phage detection. Herein, we aimed to characterise and annotate a novel bacteriophage, vB_SarS_BM31 (BM31), specific to S. arlettae. This bacteriophage was isolated from a milk sample associated with bovine mastitis and collected in the Sichuan Province, China. The BM31 genome comprised a linear double-stranded DNA of 42,271 base pair in length with a G + C content of 34.59%. A total of 65 open reading frames (ORFs) were assembled from phage DNA, of which 29 were functionally annotated. These functional genes were divided into four modules: the structural, DNA packing and replication, lysis, and lysogeny modules. Holin (ORF25), lysin (ORF26), and integrase (ORF28) were located closely in the entire BM31 genome and were important for lyse or lysogeny cycle of BM31. The phage was identified as a temperate phage according to whole genome analysis and life cycle assay, with basic biological characteristics such as small burst size, short latency period, and narrow host range, consistent with the characteristics of the family Siphoviridae, subcluster B14 of the Staphylococcus bacteriophage. The present isolation and characterisation of BM31 contributes to the Staphylococcus bacteriophage database and provides a theoretical foundation for its potential applications. To the best of our knowledge, BM31 is the only shared and completely reported phage against S. arlettae in the entire public database.

A Novel Alteromonas Phage Lineage with a Broad Host Range and Small Burst Size.

ABSTRACTAlteromonas is an opportunistic marine bacterium that persists in the global ocean and has important ecological significance. However, current knowledge about the diversity and ecology of alterophages (phages that infect Alteromonas) is lacking. Here, three similar phages infecting Alteromonas macleodii ATCC 27126T were isolated and physiologically characterized. Transmission electron microscopy revealed Siphoviridae morphology, with an oblate icosahedral head and a long noncontractile tail. Notably, these members displayed a small burst size (15–19 plaque-forming units/cell) yet an extensively broad host spectrum when tested on 175 Alteromonas strains. Such unique infection kinetics are potentially associated with discrepancies in codon usage bias from the host tRNA inventory. Phylogenetic analysis indicated that the three phages are closely evolutionarily related; they clustered at the species level and represent a novel genus. Three auxiliary metabolic genes with roles in nucleotide metabolism and putative biofilm dispersal were found in these phage genomes, which revealed important biogeochemical significance of these alterophages in marine ecosystems. Our isolation and characterization of these novel phages expand the current understanding of alterophage diversity, evolution, and phage–host interactions.IMPORTANCE The marine bacterium Alteromonas is prevalent in the global ocean with crucial ecological significance; however, little is known about the diversity and evolution of its bacteriophages that profoundly affect the bacterial communities. Our study characterized a novel genus of three newly isolated Alteromonas phages that exhibited a distinct infection strategy of broad host spectrum and small burst size. This strategy is likely a consequence of the viral trade-off between virulence and lysis profiles during phage–host coevolution, and our work provides new insight into viral evolution and infection strategies.

Phenotypic and genotypic characterization of the new Bacillus cereus phage SWEP1.

A new Bacillus cereus phage, SWEP1, was isolated from black soil. The host lysis activity of phage SWEP1 has a relatively short latent time (20min) and a small burst size of 83 PFU. The genome of SWEP1 consists of 162,461bp with 37.77% G+C content. The phage encodes 278 predicted proteins, 103 of which were assigned functionally. No tRNA genes were found. Comparative genomics analysis indicated that SWEP1 is related to Bacillus phage B4 (86.91% identity, 90% query coverage). Phenotypic and genotypic characterization suggested that SWEP1 is a new member of a new species in the genus Bequatrovirus, family Herelleviridae.

Homburgvirus LP-018 Has a Unique Ability to Infect Phage-Resistant Listeria monocytogenes.

Listeria phage LP-018 is the only phage from a diverse collection of 120 phages able to form plaques on a phage-resistant Listeria monocytogenes strain lacking rhamnose in its cell wall teichoic acids. The aim of this study was to characterize phage LP-018 and to identify what types of mutations can confer resistance to LP-018. Whole genome sequencing and transmission electron microscopy revealed LP-018 to be a member of the Homburgvirus genus. One-step-growth curve analysis of LP-018 revealed an eclipse period of ~60–90 min and a burst size of ~2 PFU per infected cell. Despite slow growth and small burst size, LP-018 can inhibit the growth of Listeria monocytogenes at a high multiplicity of infection. Ten distinct LP-018-resistant mutants were isolated from infected Listeria monocytogenes 10403S and characterized by whole genome sequencing. In each mutant, a single mutation was identified in either the LMRG_00278 or LMRG_01613 encoding genes. Interesting, LP-018 was able to bind to a representative phage-resistant mutant with a mutation in each gene, suggesting these mutations confer resistance through a mechanism independent of adsorption inhibition. Despite forming plaques on the rhamnose deficient 10403S mutant, LP-018 showed reduced binding efficiency, and we did not observe inhibition of the strain under the conditions tested. Two mutants of LP-018 were also isolated and characterized, one with a single SNP in a gene encoding a BppU domain protein that likely alters its host range. LP-018 is shown to be a unique Listeria phage that, with additional evaluation, may be useful in biocontrol applications that aim to reduce the emergence of phage resistance.

A68 Characterization and comparative genomic analysis of two Bacillus megaterium lytic bacteriophages

Next-generation sequencing technologies provide unique possibilities for the comprehensive assessment of the environmental diversity of bacteriophages. Many Bacillus bacteriophages have been isolated, but very few Bacillus megaterium bacteriophages have been characterized. Here, we describe biological characteristics and whole-genome sequences and their annotations for two new isolates of the B. megaterium bacteriophages (BM5 and BM10), isolated from Egyptian soil samples and representing two different groups according to their host range and amplified fragment length polymorphism profiles. Both phages have been displaying different thermal inactivation points (82 and 78 °C) and pH tolerance range (5–9.2 and 5–8.4 pH) while having the same longevity in vitro (192 h). Electron microscopy observation has proved that both phages belonged to the Myoviridiae family. Furthermore, growth analyses indicated that phages BM5 and BM10 have a shorter latent period (20 and 25 min) and smaller burst size (103 and 117 PFU) than is typical for Bacillus phages. The genome sizes of phages BM5 and BM10 were 165,031 bp and 165,213 bp, respectively, with a modular organization. Bioinformatic analyses of BM5 and BM10 genomes enabled assignments of putative functions to 97 and 65 putative ORFs, respectively. Comparative analysis of BM5 and BM10 genome structures with other B. megaterium bacteriophages revealed relatively high levels of sequence and organizational identity. Both genomic comparisons and phylogenetic analyses support the conclusion that the sequenced phages (BM5 and BM10) belong to different sub-clusters (L5 and L7) within L cluster and display different lifestyles (lysogenic and lytic). Sequenced phages encode proteins associated with Bacillus pathogenesis. BM5 does not contain any tRNA sequences, while BM10 genome codes for 17 tRNAs.

Characterization and comparative genomic analysis of virulent and temperate Bacillus megaterium bacteriophages.

Next-Generation Sequencing (NGS) technologies provide unique possibilities for the comprehensive assessment of the environmental diversity of bacteriophages. Several Bacillus bacteriophages have been isolated, but very few Bacillus megaterium bacteriophages have been characterized. In this study, we describe the biological characteristics, whole genome sequences, and annotations for two new isolates of the B. megaterium bacteriophages (BM5 and BM10), which were isolated from Egyptian soil samples. Growth analyses indicated that the phages BM5 and BM10 have a shorter latent period (25 and 30 min, respectively) and a smaller burst size (103 and 117 PFU, respectively), in comparison to what is typical for Bacillus phages. The genome sizes of the phages BM5 and BM10 were 165,031 bp and 165,213 bp, respectively, with modular organization. Bioinformatic analyses of these genomes enabled the assignment of putative functions to 97 and 65 putative ORFs, respectively. Comparative analysis of the BM5 and BM10 genome structures, in conjunction with other B. megaterium bacteriophages, revealed relatively high levels of sequence and organizational identity. Both genomic comparisons and phylogenetic analyses support the conclusion that the sequenced phages (BM5 and BM10) belong to different sub-clusters (L5 and L7, respectively), within the L-cluster, and display different lifestyles (lysogenic and lytic, respectively). Moreover, sequenced phages encode proteins associated with Bacillus pathogenesis. In addition, BM5 does not contain any tRNA sequences, whereas BM10 genome codes for 17 tRNAs.

Characterization, sequencing and comparative genomic analysis of vB_AbaM-IME-AB2, a novel lytic bacteriophage that infects multidrug-resistant Acinetobacter baumannii clinical isolates.

BackgroundWith the use of broad-spectrum antibiotics, immunosuppressive drugs, and glucocorticoids, multidrug-resistant Acinetobacter baumannii (MDR-AB) has become a major nosocomial pathogen species. The recent renaissance of bacteriophage therapy may provide new treatment strategies for combatting drug-resistant bacterial infections. In this study, we isolated a lytic bacteriophage vB_AbaM-IME-AB2 has a short latent period and a small burst size, which clear its host’s suspension quickly, was selected for characterization and a complete genomic comparative study.ResultsThe isolated bacteriophage vB_AbaM-IME-AB2 has an icosahedral head and displays morphology resembling Myoviridae family. Gel separation assays showed that the phage particle contains at least nine protein bands with molecular weights ranging 15–100 kDa. vB_AbaM-IME-AB2 could adsorb its host cells in 9 min with an adsorption rate more than 99% and showed a short latent period (20 min) and a small burst size (62 pfu/cell). It could form clear plaques in the double-layer assay and clear its host’s suspension in just 4 hours. Whole genome of vB_AbaM-IME-AB2 was sequenced and annotated and the results showed that its genome is a double-stranded DNA molecule consisting of 43,665 nucleotides. The genome has a G + C content of 37.5% and 82 putative coding sequences (CDSs). We compared the characteristics and complete genome sequence of all known Acinetobacter baumannii bacteriophages. There are only three that have been sequenced Acinetobacter baumannii phages AB1, AP22, and phiAC-1, which have a relatively high similarity and own a coverage of 65%, 50%, 8% respectively when compared with our phage vB_AbaM-IME-AB2. A nucleotide alignment of the four Acinetobacter baumannii phages showed that some CDSs are similar, with no significant rearrangements observed. Yet some sections of these strains of phage are nonhomologous.ConclusionvB_AbaM-IME-AB2 was a novel and unique A. baumannii bacteriophage. These findings suggest a common ancestry and microbial diversity and evolution. A clear understanding of its characteristics and genes is conducive to the treatment of multidrug-resistant A. baumannii in the future.

Isolation and characterization of a virulent bacteriophage SPW specific for Staphylococcus aureus isolated from bovine mastitis of lactating dairy cattle.

Mastitis in dairy cattle continues to be an economically important disease. However, control is complicated by a high prevalence of resistance to antibiotics. Phage therapy, therefore, is considered as an alternative way of controlling bacterial infections and contaminations. In this study, we have described isolation and characterization of a highly virulent phage SPW from wastewater of dairy farm, which possesses a strong lytic capability against mastitis-associated Staphylococcus aureus, the most important pathogen in bovine clinical and subclinical mastitis. The phage SPW produced large, round and clear plaques on bacterial culture plates. TEM showed phage SPW has an icosahedral head 62.5 nm in diameter and long tail of 106 nm, head and tail were held together by a connector of 18 ± 1.5 nm long and can be classified as a member of the Myoviridae family. Restriction analysis indicated that phage SPW was a dsDNA virus with an approximate genome size of 65-69 kb. One-step growth kinetics showed a short latency period of about 10-15 min and a rise period of 50 min and a relatively small burst size was 44 ± 3 phages particles/infected cell. Moreover, adsorption rates were not influenced by calcium ions and phage SPW was relatively stable in a wide range of temperature and pH values, and resistant to chloroform and isopropanol. The optimal multiplicity of infection (MOI) was 0.01. When phage SPW was used to infect five other clinically isolated pathogenic isolates, it showed relatively wide spectrum host range. Phage SPW was capable of eliciting efficient lysis of S. aureus, revealing it potentially as an effective approach to prophylaxis or treatment of S. aureus-associated mastitis in dairy cows.

Isolation and Characterization of a Novel Bacteriophage φ4D Lytic Against Enterococcus faecalis Strains

In recent years, Enterococcus faecalis has emerged as an important opportunistic nosocomial pathogen capable of causing dangerous infections. Therefore, there is an urgent need to develop novel antibacterial agents to control this pathogen. Bacteriophages have very effective bactericidal activity and several advantages over other antimicrobial agents and so far, no serious or irreversible side effects of phage therapy have been described. The objective of this study was to characterize a novel virulent bacteriophage φ4D isolated from sewage. Electron microscopy revealed its resemblance to Myoviridae, with an isometric head (74 ± 4 nm) and a long contractile tail (164 ± 4 nm). The φ4D phage genome was tested using pulsed-field gel electrophoresis and estimated to be 145 ± 2 kb. It exhibited short latent period (25 min) and a relatively small burst size (36 PFU/cell). Tests were conducted on the host range, multiplicities of infection (MOI), thermal stability, digestion of DNA by restriction enzymes, and proteomic analyses of this phage. The isolated phage was capable of infecting a wide spectrum of enterococcal strains. The results of these investigations indicate that φ4D is similar to other Myoviridae bacteriophages (for example φEF24C), which have been successfully used in phagotherapy.

Characterization of a bacteriophage, isolated from a cow with mastitis, that is lytic against Staphylococcus aureus strains

Methicillin-resistant strains of Staphylococcus aureus (MRSA) are now the most commonly reported antibiotic-resistant bacterium in clinical settings. Therefore, there is an urgent need to develop novel antibacterial agents to control this pathogen. Bacteriophage therapy is a potential alternative treatment for MRSA infections. The objective of this study was characterization of a novel virulent bacteriophage (MSA6) isolated from a cow with mastitis. Electron microscopy showed its resemblance to members of the family Myoviridae, with an isometric head (66 nm) and a long contractile tail (173 nm). The genome of phage MSA6 was tested by pulsed-field gel electrophoresis and estimated to be about 143 kb. It exhibited rapid adsorption (>82% in 5 min), a short latent period (15 min) and a relatively small burst size (23 PFU/cell). Isolated phage was capable of infecting a wide spectrum of staphylococcal strains of both human and bovine origin. The results of this investigation indicate that MSA6 is similar to other bacteriophages belonging to the family Myoviridae (Twort, K, G1, 812) that have been successfully used in bacteriophage therapy.

Large Pseudomonas phages isolated from barley rhizosphere

Abstract Five bacteriophages infecting common fluorescent pseudomonads (Pseudomonas fluorescens and Pseudomonas putida) were isolated from barley rhizosphere soil. Morphological and molecular characteristics of the phages are described together with selected phage-host interactions. All phages belonged to the Myoviridae family with isometrical heads on contractile tails; 4 of them were unusually large and had complex protein and DNA profiles. The large phages had estimated genome sizes of 200 kb or more. Restriction enzyme analyses and DNA-DNA hybridizations showed that all isolates represented different phage species. None of the isolates were observed to establish lysogeny with the main host strain, P. putida MM1. The large phages multiplied slowly on their hosts, producing very small plaques; one-step growth experiments with one of the large phages (Psp 4) hence demonstrated a long latent period (2.5 h) and a very small burst size (10 particles). One of the large phages (Psp 3) was abundant in the rhizosphere (approx. 104 pfu g−1 soil) and had a particularly broad host range which extended to both fluorescent (Pseudomonas aeruginosa, P. fluorescens, P. putida and Pseudomonas chlororaphis) and non-fluorescent (Pseudomonas stutzeri) Pseudomonas spp. occurring in soil. The ecological importance of the large Pseudomonas phages must be further studied, but their slow multiplication rates suggested a possible mechanism of balanced phage-host co-existence in the rhizosphere.

Adaptive Assembly on Delay Fairness in Optical Burst Switched Networks

This paper focuses on burst assembly at edge node in optical burst switching networks, which can be implemented to provide the delay QoS guarantee for the whole network. Based on traffic prediction and delay fairness of different classes, a Prediction and delay QoS Burst Assembly (PQBA) mechanism is proposed to reduce the assembly delay and improve the delay fairness especially for high priority service at edge node. We compare the delay, burst size, burst utilization and length error rate of all service classes under different offered load. Simulation studies are carried out for PQBA and the results show that high priority service with relative small burst size achieves a low assembly delay and high burst utilization. Consequently, PQBA realizes the QoS-based delay fairness and service class differentiation at edge node.

Characterization of six bacteriophages of serratia liquefaciens CP6 isolated from the sugar beet phytosphere

Six phages (PhiCP6-1 to PhiCP6-6) that are commonly found in the phytosphere of sugar beet (Beta vulgaris var. Amethyst) were investigated, and their relative impacts on their host (Serratia liquefaciens CP6) were compared. There were fundamental differences between the two most abundant predators of CP6 (PhiCP6-1 and PhiCP6-4). Like PhiCP6-2 and PhiCP6-5, PhiCP6-1 belonged to the family Siphoviridae, while PhiCP6-4 exhibited the morphology of the family Podoviridae. The other phages were members of the family Myoviridae. DNA-DNA cross-hybridization revealed that PhiCP6-1 and PhiCP6-4 had little common DNA, although all of the other phages exhibited some genetic similarity. Like PhiCP6-2, PhiCP6-3, and PhiCP6-5, PhiCP6-1 was capable of forming a lysogenic association with its host, while PhiCP6-4 and PhiCP6-6 appeared to be entirely virulent. Single-step growth curve experiments revealed that PhiCP6-4 had a much shorter latent period and a smaller burst size than PhiCP6-1. Also, PhiCP6-1 could transduce a number of host chromosomal markers with transfer frequencies of 2.9 x 10(-9) to 3.9 x 10(-7), whereas PhiCP6-4 could not transduce S. liquefaciens CP6 genes. When viewed in the context of the strikingly different temporal niches of these phages, our data provide an insight into how bacteriophage interactions with their hosts might reflect the natural ecology of bacteriophages. Our data also illustrate how the potential for gene transfer changes over time in an environment that supports several different phages.

Large Pseudomonas phages isolated from barley rhizosphere

Five bacteriophages infecting common fluorescent pseudomonads ( Pseudomonas fluorescens and Pseudomonas putida) were isolated from barley rhizosphere soil. Morphological and molecular characteristics of the phages are described together with selected phage-host interactions. All phages belonged to the Myoviridae family with isometrical heads on contractile tails; 4 of them were unusually large and had complex protein and DNA profiles. The large phages had estimated genome sizes of 200 kb or more. Restriction enzyme analyses and DNA-DNA hybridizations showed that all isolates represented different phage species. None of the isolates were observed to establish lysogeny with the main host strain, P. putida MM1. The large phages multiplied slowly on their hosts, producing very small plaques; one-step growth experiments with one of the large phages (Psp 4) hence demonstrated a long latent period (2.5 h) and a very small burst size (10 particles). One of the large phages (Psp 3) was abundant in the rhizosphere (approx. 10 4 pfu g −1 soil) and had a particularly broad host range which extended to both fluorescent ( Pseudomonas aeruginosa, P. fluorescens, P. putida and Pseudomonas chlororaphis) and non-fluorescent (Pseudomonas stutzeri) Pseudomonas spp. occurring in soil. The ecological importance of the large Pseudomonas phages must be further studied, but their slow multiplication rates suggested a possible mechanism of balanced phage-host co-existence in the rhizosphere.

T1 pip: A mutant which affects packaging initiation and processive packaging of T1 DNA

The pip mutation of phage T1 is located between the tar (gene 2.5) and am6 (gene) mutations in the region of the T1 genome which codes for early functions. The tar and pip mutations are additive in increasing the efficiency of transduction by T1 When T1 carries the pip mutation the initiation of DNA packaging by the phage at the non-T1, esp-λ site is more efficient than when the phage is pip +; the small average burst size of 8 to 10 by T1 pip suggests that pip causes a reduction in the efficiency with which TI utilizes pac, the normal packaging initiation site of the phage. The presence of the BglII-D fragment (cut at one end at pac and the other by BglII) after digestion of Tlpip DNA by BglII shows that Tlpip continues to initiate DNA packaging at pac. The increased molarity of BglllD coupled with the absence of the BglII-C fragment (which contains DNA on both sides of pac and can only be cut from processively packaged genomes) shows that T1 pip packages only genomes which are initiated at pac and is defective in processive packaging.

Replication of the algal virus PBCV-1 in UV-irradiated Chlorella.

The large dsDNA algal virus, PBCV-1, replicates in UV-irradiated Chlorella, albeit more slowly and with a smaller burst size than in untreated Chlorella. Irradiated cells are unable to form colonies, and endogenous RNA and DNA synthesis are reduced to background levels. Thus a fully function host nucleus is not required for PBCV-1 replication.