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1.  Ecological Genomics of Marine Picocyanobacteria†  
Summary: Marine picocyanobacteria of the genera Prochlorococcus and Synechococcus numerically dominate the picophytoplankton of the world ocean, making a key contribution to global primary production. Prochlorococcus was isolated around 20 years ago and is probably the most abundant photosynthetic organism on Earth. The genus comprises specific ecotypes which are phylogenetically distinct and differ markedly in their photophysiology, allowing growth over a broad range of light and nutrient conditions within the 45°N to 40°S latitudinal belt that they occupy. Synechococcus and Prochlorococcus are closely related, together forming a discrete picophytoplankton clade, but are distinguishable by their possession of dissimilar light-harvesting apparatuses and differences in cell size and elemental composition. Synechococcus strains have a ubiquitous oceanic distribution compared to that of Prochlorococcus strains and are characterized by phylogenetically discrete lineages with a wide range of pigmentation. In this review, we put our current knowledge of marine picocyanobacterial genomics into an environmental context and present previously unpublished genomic information arising from extensive genomic comparisons in order to provide insights into the adaptations of these marine microbes to their environment and how they are reflected at the genomic level.
doi:10.1128/MMBR.00035-08
PMCID: PMC2698417  PMID: 19487728
3.  Effects of Some Disinfectants on African Swine Fever Virus 
Applied Microbiology  1973;25(1):115-122.
Ten commercially available disinfectants were tested at high pH in 2% sodium hydroxide and low pH in 2% acetic acid as inactivants for African swine fever (ASF) in a protein-rich blood-spleen homogenate. As assayed in leukocyte cultures, sodium hydroxide and acetic acid, sodium meta silicate and Roccal did not inactivate ASF virus in 1 hr at 22 to 25 C. Some viricidal activity as assayed in leukocyte cultures was found with Weladol, Triton X-100 Amphyl, pHisoHex, sodium dodecyl sulfate, LpH, Environ, Environ D, and One-Stroke Environ. Of these, the last four appeared to be most promising. When assayed in pigs, only One-Stroke Environ (1/E) was viricidal. Concentrations of 1.0, 0.75, and 0.5 were effective, but, at 0.25%, virus was not inactivated. The minimal time to inactivate ASF virus by 1% 1/E is 60 min. A room contaminated with ASF virus was made safe for pigs after 1 hr by spraying with 1% 1/E. The most active component of 1/E is o-phenylphenol. Although another component of 1/E, i.e., o-benzyl-p-chlorophenol, also has some activity, the mixture of the active components of 1/E is most effective against ASF virus. One of the soluble antigens associated with ASF virus is destroyed by 1/E.
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PMCID: PMC380746  PMID: 4631432
4.  Diel Expression of Cell Cycle-Related Genes in Synchronized Cultures of Prochlorococcus sp. Strain PCC 9511 
Journal of Bacteriology  2001;183(3):915-920.
The cell cycle of the chlorophyll b-possessing marine cyanobacterium Prochlorococcus is highly synchronized under natural conditions. To understand the underlying molecular mechanisms we cloned and sequenced dnaA and ftsZ, two key cell cycle-associated genes, and studied their expression. An axenic culture of Prochlorococcus sp. strain PCC 9511 was grown in a turbidostat with a 12 h–12 h light-dark cycle for 2 weeks. During the light periods, a dynamic light regimen was used in order to simulate the natural conditions found in the upper layers of the world's oceans. This treatment resulted in strong cell cycle synchronization that was monitored by flow cytometry. The steady-state mRNA levels of dnaA and ftsZ were monitored at 4-h intervals during four consecutive division cycles. Both genes exhibited clear diel expression patterns with mRNA maxima during the replication (S) phase. Western blot experiments indicated that the peak of FtsZ concentration occurred at night, i.e., at the time of cell division. Thus, the transcript accumulation of genes involved in replication and division is coordinated in Prochlorococcus sp. strain PCC 9511 and might be crucial for determining the timing of DNA replication and cell division.
doi:10.1128/JB.183.3.915-920.2001
PMCID: PMC94958  PMID: 11208789
5.  Comparative analysis of splicing of the complete set of chloroplast group II introns in three higher plant mutants. 
Nucleic Acids Research  1999;27(19):3866-3874.
The barley mutant albostrians and the maize mutants crs1 and crs2 are defective in the splicing of various plastid group II introns. By analysing tRNA precursors and several mRNAs not previously examined, the investigation of in vivo splicing defects in these mutants has been completed. The albostrians mutation causes the loss of plastid ribosomes resulting secondarily in a disruption of splicing of all subgroup IIA introns in the chloroplast. Thus MatK, the only putative chloroplast intron-specific maturase of higher plants, might have evolved to function in splicing of multiple introns. We show that in the case of tRNA-Ala(UGC)the first step of splicing is affected, as suggested by the absence of lariat molecules. Thus the plastid-encoded splicing factor lacking in albostrians must participate in the formation of the catalytically active structure. In contrast, a mutation in the nuclear gene crs1 prevents splicing of only one intron but causes specific additional effects as precursor transcripts for tRNA-Ile(GAU), tRNA-Ala(UGC), tRNA-Lys(UUU)and tRNA-Val(UAC), but not tRNA-Gly(UCC), have significantly enhanced steady-state levels in this mutant. Our data provide evidence for a variety of splicing factors and pathways in the chloroplast, some encoded by nuclear and some by chloroplast genes, and possibly for a dual function of some of these factors.
PMCID: PMC148650  PMID: 10481026
6.  Prochlorococcus, a Marine Photosynthetic Prokaryote of Global Significance 
The minute photosynthetic prokaryote Prochlorococcus, which was discovered about 10 years ago, has proven exceptional from several standpoints. Its tiny size (0.5 to 0.7 μm in diameter) makes it the smallest known photosynthetic organism. Its ubiquity within the 40°S to 40°N latitudinal band of oceans and its occurrence at high density from the surface down to depths of 200 m make it presumably the most abundant photosynthetic organism on Earth. Prochlorococcus typically divides once a day in the subsurface layer of oligotrophic areas, where it dominates the photosynthetic biomass. It also possesses a remarkable pigment complement which includes divinyl derivatives of chlorophyll a (Chl a) and Chl b, the so-called Chl a2 and Chl b2, and, in some strains, small amounts of a new type of phycoerythrin. Phylogenetically, Prochlorococcus has also proven fascinating. Recent studies suggest that it evolved from an ancestral cyanobacterium by reducing its cell and genome sizes and by recruiting a protein originally synthesized under conditions of iron depletion to build a reduced antenna system as a replacement for large phycobilisomes. Environmental constraints clearly played a predominant role in Prochlorococcus evolution. Its tiny size is an advantage for its adaptation to nutrient-deprived environments. Furthermore, genetically distinct ecotypes, with different antenna systems and ecophysiological characteristics, are present at depth and in surface waters. This vertical species variation has allowed Prochlorococcus to adapt to the natural light gradient occurring in the upper layer of oceans. The present review critically assesses the basic knowledge acquired about Prochlorococcus both in the ocean and in the laboratory.
PMCID: PMC98958  PMID: 10066832
7.  Precise branch point mapping and quantification of splicing intermediates. 
Nucleic Acids Research  1997;25(10):2030-2031.
Lariat intermediates of a group II intron were investigated via RT-PCR. Several reverse transcriptases appeared capable of reading through a branched nucleotide. A new method has been established that yields precise information about the location of the branch point within an intron. As an extension of our approach, antisense transcripts of the previously cloned PCR products were successfully used in RNase Protection Assays, providing a tool for quantification of splicing intermediates. Application of the method presented to other self-splicing introns as well as introns in nuclear pre-mRNAs is envisaged.
PMCID: PMC146694  PMID: 9115373
8.  New method of antibody detection by indirect immunoperoxidase plaque staining for serodiagnosis of African swine fever. 
Journal of Clinical Microbiology  1982;16(4):650-655.
An indirect immunoperoxidase plaque-staining method was developed for detecting antibody to African swine fever virus infection. In both sensitivity and specificity, the test was comparable to indirect immunofluorescence. Because it has all of the desirable features of the indirect immunofluorescence test and may also be readily used for testing large numbers of sera, the indirect immunoperoxidase plaque-staining method can be used as a single and final serodiagnostic test in a large-scale survey of African swine fever. The indirect immunoperoxidase plaque-staining method should be applicable to other viruses that can be adapted to and grown in cell cultures.
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PMCID: PMC272439  PMID: 6185528
9.  Residual viruses in pork products. 
Partly cooked canned hams and dried pepperoni and salami sausages were prepared from the carcasses of pigs infected with African swine fever virus and pigs infected with hog cholera virus. Virus was not recovered from the partly cooked canned hams; however, virus was recovered in the hams before heating in both instances. Both African swine fever virus and hog cholera virus were recovered from the dried salami and pepperoni sausages, but not after the required curing period.
PMCID: PMC242793  PMID: 564162
10.  Immunofluorescence Plaque Assay for African Swine Fever Virus 
Suitably diluted cell culture adapted African swine fever virus preparations were inoculated on VERO cell monolayers and grown on coverslips. Gum tragacanth was used as an overlay. After three days incubation at 37°C the infected cultures were fixed with acetone and stained with fluorescent antibody conjugate. Fluorescing plaques consisted of 20-30 infected cells.
Three statistical criteria for a quantitatively reliable assay were met: the Poisson distribution for plaque counts, linearity of the relationship between the concentration of virus and the plaque count and reproducibility of replicate titrations. The method is suitable for counts up to at least 70 plaques per 5 cm2 coverslip and computed titers are reproducible within 0.16 log units with a total of 300 plaques enumerated.
PMCID: PMC1319849  PMID: 4279763
11.  African Swine Fever: Application of Immunoelectroosmophoresis for the Detection of Antibody 
Thirty-three pigs in three groups of nineteen, ten, and four pigs were infected with three different African swine fever (ASF) virus isolates, respectively. All virus isolates were attenuated to varying degrees by passaging in cell cultures, and they retained sufficiently low virulence to produce subacute and chronic infections in pigs. Sera collected at various intervals were tested for antibody activity by the immunoelectroosmophoresis, agar gel diffusion precipitin, and complement-fixation tests using a modified Kolmer technique. Results clearly indicated that the immunoelectroosmophoresis test is a rapid (30 minute) and accurate method with extreme sensitivity and superior to the complement-fixation and agar gel diffusion precipitin tests in detecting antibody against ASF virus. Possible use of this method in detecting ASF virus infection is suggested.
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PMCID: PMC1319686  PMID: 4114980
12.  A Plaque Assay for Duck Plague Virus 
A plaque assay for duck plague virus was developed for a chicken embryo-adapted virus and a duck lethal virus and used to determine the identity of these viruses. Using the plaque inhibition neutralization test, duck plague virus was differentiated from Newcastle disease, fowl plague, and duck hepatitis viruses. The plaque morphology is described.
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PMCID: PMC1319285  PMID: 15846902

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