Regulation, replication, and integration functions of the Vibrio cholerae CTXφ are encoded by region RS2

CTXφ is a filamentous phage that encodes cholera toxin, one of the principal virulence factors of Vibrio cholerae . CTXφ is unusual among filamentous phages because it can either replicate as a plasmid or integrate into the V. cholerae chromosome at a specific site. The CTXφ genome has two regions, the 'core' and RS2. Integrated CTXφ is frequently flanked by an element known as RS1 which is related to RS2. The nucleotide sequences of RS2 and RS1 were determined. These related elements contain three nearly identical open reading frames (ORFs), which in RS2 were designated rstR , rstA2 and rstB2 . RS1 contains an additional ORF designated rstC . Functional analyses indicate that rstA2 is required for CTXφ replication and rstB2 is required for CTXφ integration. The amino terminus of RstR is similar to the amino termini of other phage-encoded repressors, and RstR represses the expression of rstA2 . Although genes with related functions are clustered in the genome of CTXφ in a way similar to those for other filamentous phages, the CTXφ RS2-encoded gene products mediating replication, integration and repression appear to be novel.     

Long-Term Drug Treatment of Obesity in a Private Practice Setting

ATKINSON, RICHARD L, ROY C BLANK, DONALD SCHUMACHER, NIKHIL V DHURANDHAR, DOUGLAS L RITCH. Long-term drug treatment of obesity in a private practice setting. This study evaluated the long-term efficacy and safety of the combination of phentermine and fenfluramine for the treatment of obesity in a private practice setting. A total of 1388 consecutive, qualified patients presenting to a private general internal medicine practice in Charlotte, NC, were enrolled with eligibility criteria including: age 18 years to 60 years, 20% over “desirable” bodyweight or body mass index <27, no serious medical or psychiatric disease, and no contraindications to drug therapy. Patients were instructed in diet, exercise, and behavior modification techniques and received phentermine (15 mg/day to 30 mg/day) and fenfluramine (20 mg/day to 60 mg/day) continuously for over 3 years. Average duration of treatment was 15. 9 months, and average weight loss at the last visit was 11. 6 kg, or 11. 7% of initial bodyweight. For patients completing 1 year of drug treatment, mean weight loss was 16. 5 kg, or 16% of initial weight. Weight loss persisted for 2 years, but partial regain was seen at 3 years. The dropout rates were 18% at 6 months, 39% at 1 year, 68% at 2 years, and 78% at 3 years. At 1 year, blood pressure of hypertensive patients fell from 151/95 mm Hg to 127/78 mm Hg, and serum cholesterol and triglycerides of hyperlipidemic patients fell by 0. 750 mmol/L (29 mg/dL) and 0. 937 mmol/L (83 mg/dL), respectively. Adverse events were modest. We conclude that, in a private practice setting, long-term treatment of obesity with the combination of phentermine, fenfluramine, and a weight maintenance program is generally safe and effective. More research is needed to determine efficacy and safety for longer than 3 years.     

Surgical Treatment of Constipation

A 30-year retrospective review of 544,354 Seattle area hospital admissions yielded 25 patients who underwent surgical therapy for the relief of intractable idiopathic constipation. All patients were refractory to conventional medical treatment consisting of the daily use of laxatives, cathartics, emollients or enemas. Long-term follow-up was available for 13 of the 25 patients. All 13 patients had clinical improvement as a result of the operation. This confirms results reported by other authors. Subtotal colectomy and left hemicolectomy are the procedures generally favored.     

Selected aspects of collared peccary (Dicotyles tajacu) reproductive biology in a captive texas herd

Collared peccaries (Dicotyles tajacu) from 32 litters were born in captivity during the period from September, 1981 through July, 1983. Incidence of singleton, twin, and triplet litters was 28%, 66%, and 6%, respectively. Mean (± SE) weight of young from singleton, twin, and triplet litters was 751 (± 35) gm, 698 (± 17) gm, and 612 (± 74) gm, respectively. Sex ratio among all litters was 56:44 (M:F). Neonatal mortality amounted to 9% of the young born. Mean duration of estrus was 2.60 days and mean length of gestation was 145.6 days. Evidence indicated that multiple copulations increased ovulation rate and the chance for conception.     

Cytochemical and electrophoretic studies of haemoglobin synthesis in the fat body of a midge, Chironomus thummi

The fat body of developing mid- and late fourth instar larvae of a midge, Chironomus thummi, has been investigated by means of the benzidine reaction for the localization of haemoglobin within cells. In the subepidermal fat body the reaction deposits of the haemoglobin pseudo-peroxidase activity appear predominantly in the intracisternal cavities of ER and the Golgi, and later, in the pharate pupal stage, in small dense granules (0.5–1 μm in. diameter).All the major protein bands of fat body extracts, which are resolved in electrophoresis, give the benzidine reaction and show incorporation of ¹⁴C-amino levulinic acids, in this case a specific marker for haemoglobin synthesis. In addition, labelled proteins show identical electrophoretic mobility as the haemoglobins of the haemolymph, suggesting that haemoglobins are synthesized in the fat body. Two types of fat body cells seem to differ with respect to their rôle in haemoglobin metabolism.     

Selenocystine-resistant mutants of Histoplasma capsulatum

Cysteine metabolism has been thought to be important to the phenomenon of dimorphism inHistoplasma capsulatum. We sought mutants with genetic blocks in the metabolism of cysteine by selection of colonies resistant to the toxic analogue, selenocystine. The 22 resistant strains thus obtained were all deficient in uptake of cystine from the surrounding medium but were normally able to convert from mycelium to yeast and back again. Furthermore, they had normal quantities of NADH-dependent cystine reductase when this enzyme was measured. We conclude that mutants defective in cystine uptake can be readily obtained by selection of colonies resistant to selenocystine, and that a lesion in cystine-uptake does not appear to affect the phenomenon of dimorphism in this organism.Preliminary reports of this work were presented at the Second International Congress of Mycology, Tampa, 1977 and at the first International Conference on Histoplasmosis, Atlanta, 1978.     

Oviposition in Triatoma protracta: Role of mating and relationship to egg growth

Yolk deposition begins in the terminal oocytes of virgins of Triatoma protracta a few days after adult eclosion, and while a few eggs may be matured before vitellogenesis ceases, none are laid. Mating at day four stimulates egg maturation and oviposition in fed and unfed females, egg-laying beginning as early as nine days after eclosion. If mating is delayed until day 16, by which time vitellogenesis normally has ceased, stored eggs are laid within two to four days and yolk deposition is resumed. Removal of the brain prevents oviposition, as does the severance of the ventral nerve cord. Thus, an intact central nervous system is required for egg-laying. Also, since neither operation inhibits egg maturation or ovulation, it appears that the latter reproductive responses to mating are independent of oviposition.     

Protection of cryopreserved Onchocerca microfilariae (nematoda) from dilution shock by the use of serum

This paper describes the use of newborn calf serum during the cooling and warming/dilution phases of the cryopreservation of Onchocerca gutturosa microfilariae using an interrupted slow cool to ?196 °C in the presence of 5% (v/v) methanol. Serum proved detrimental at concentrations above 20% (v/v) in the cooling medium unless it was also present in high concentrations, 60% (v/v) in the warming/ dilution medium.Damage to the organisms occurred predominantly during the thawing/dilution phase of cryopreservation and not the cooling phase and could be reduced greatly by the presence of high serum concentrations when thawing. This indicates that the major protective action of serum is that of reducing dilution shock—shock produced by a rapid influx of water and/or the effects of high solute concentrations established during cooling.     

Antibiotics and the search for new principles

Originally presented as an Invited Lecture at the 1990 Society for Industrial Microbiology Annual Meeting in Orlando, Florida.     

Nucleotide sequence of the genome of the filamentous bacteriophage I2-2: Module evolution of the filamentous phage genome

Summary The nucleotide sequence of the circular single-stranded genome of the filamentous Escherichia coli phage I2-2 has been determined and compared with those of the filamentous E. coli phages Ff(M13, fl, or fd) and IKe. The I2-2 DNA sequence comprises 6744 nucleotides; 139 nucleotides less than that of the N- and I2-plasmid-specific phage IKe, and 337 (336) nucleotides more than that of the F-plasmid-specific phage Ff. Nucleotide sequence comparisons have indicated that I2-2, IKe, and Ff have a similar genetic organization, and that the genomes of I2-2 and IKe are evolutionarily more closely related than those of I2-2 and Ff. The studies have further demonstrated that the I2-2 genome is a composite replicon, composed of only two-thirds of the ancestral genome of IKe. Only a contiguous I2-2 DNA sequence of 4615 nucleotides encompassing not only the coat protein and phage assembly genes, but also the signal required for efficient phage morphogenesis, was found to be significantly homologous to sequences in the genomes of IKe and Ff. No homology was observed between the consecutive DNA sequence that contains the origins for viral and complementary strand replication and the replication genes. Although other explanations cannot be ruled out, our data strongly suggest that the ancestor filamentous phage genome of phages I2-2 and IKe has exchanged its replication module during evolution with that of another replicon, e.g., a plasmid that also replicates via the so-called rolling circle mechanism. Offprint requests to: R.N.H. Konings