Comparative study of various methods for determining Vibrio cholerae toxigenicity

Testing of 138 Vibrio cholerae strains for gene determinants responsible for the production of cholera enterotoxin by the polymerase chain reaction (PCR) and gene probing using molecular CT-probe showed good correlation of the results of different methods and correlation of these data with studies of V. cholerae strain virulence in vivo and in hemolytic activity test. The advantages of PCR in rapid assessment of the toxigenicity and epidemic significance of V. cholerae strains are demonstrated.     

Comparative study of two culture conservation methods in medical mycology

The efficiency of two fungal conservation methods was compared: Suspension in sterile distilled water and subcultures on potato dextrose agar (PDA) slants at 4 °C. One hundred and eleven strains corresponding to 84 different-species of microorganisms studied in medical mycology were evaluated. The efficiency of each method was estimated by the survival percentage and the preservation of the morphological features of each strain within a seven-year period. From the 111 strains, 79 (71.2%) were preserved viable in water, compared to 86 (77.5%) strains preserved by subculture on PDA slants. Concerning morphological features 75 of the 79 water viable strains (94.9%) conserved their morphology. In contrast, only 60 of the 86 strains (69.8%) conserved their typical morphology by the PDA subculture method. The water conservation method offers important benefits over serial subculture such as: Minimal pleomorphism, simple, rapid and requiring few materials. Thus, the water conservation method is recommended for laboratories where specialized conservation equipment is not available.     

Polymer-cushioned bilayers. I. A structural study of various preparation methods using neutron reflectometry.

This neutron reflectometry study evaluates the structures resulting from different methods of preparing polymer-cushioned lipid bilayers. Four different techniques to deposit a dimyristoylphosphatidylcholine (DMPC) bilayer onto a polyethylenimine (PEI)-coated quartz substrate were examined: 1) vesicle adsorption onto a previously dried polymer layer; 2) vesicle adsorption onto a bare substrate, followed by polymer adsorption; and 3, 4) Langmuir-Blodgett vertical deposition of a lipid monolayer spread over a polymer-containing subphase to form a polymer-supported lipid monolayer, followed by formation of the outer lipid monolayer by either 3) horizontal deposition of the lipid monolayer or 4) vesicle adsorption. We show that the initial conditions of the polymer layer are a critical factor for the successful formation of our desired structure, i.e., a continuous bilayer atop a hydrated PEI layer. Our desired structure was found for all methods investigated except the horizontal deposition. The interaction forces between these polymer-supported bilayers are investigated in a separate paper (Wong, J. Y., C. K. Park, M. Seitz, and J. Israelachvili. 1999. Biophys. J. 77:1458-1468), which indicate that the presence of the polymer cushion significantly alters the interaction potential. These polymer-supported bilayers could serve as model systems for the study of transmembrane proteins under conditions more closely mimicking real cellular membrane environments.     

Comparative anatomical studies of the vomeronasal complex and the rostral palate of various mammals. I

The anatomy of the vomeronasal complex and, in connection with this, the structures of the rostral palate were studied in different species of mammals, namely members of the order Marsupialia, Scandentia, Insectivora, Primates, Rodentia, and Lagomorpha. The following results were obtained: The organs of Jacobson of all forms studied are well-developed. The organ of Jacobson is situated at the base of the nasal septum and opens rostrally, always closely connected to the nasopalatine duct. Even in rodents, lagomorphs and Solenodon, where the openings of the organs are positioned rostral to the ductus, both systems are nevertheless connected by means of special furrows. Accordingly the organs of Jacobson are functionally much more closely related to the oral cavity than to the nasal cavity, which they actually belong to. This can be emphasized by the peculiar structures of the rostral palate inclosing the papilla palatina and with it the oral openings of the nasopalatine ducts. In all species studied, the anterior part of the upper jaw presents a very interesting situation because the median furrow of the rhinarium communicates directly or indirectly with the sulcus papillae palatinae, thus forming a very distinct system of grooves which preserves a connection between the nasopalatine ducts and the preoral surroundings. In rodents, lagomorphs, and Solenodon, we find in this part of the palate a special situation because of their unusually arranged incisors, which are not separated by a diastema. However, also in these cases, there are distinct connecting passages between the papilla palatina and the extraoral surroundings. The conditions found in Ratufa bicolor and in early stages of the rat demonstrate that the extraordinary topography of the rostral palate in rodents is a secondary formation by means of ontogeny and phylogeny. Cebus apella, a platyrrhine simian, shows already a clear reduction of palatal structures compared to those found in prosimians. In Setifer setosus and Echinops telfairi, we find the papilla palatina and with it the oral openings of the nasopalatine ducts overgrown by a bipartite caudal branch of the rhinarium. The neonate Setifer allows us to reconstruct the mechanism of this overgrowing procedure. We find a similar situation in Erinaceus, where the papilla palatina remains uncovered, however. Because of contradictory bibliographical data, some elements of the vomeronasal complex in mammals needed to be carefully analysed in regard to structure and nomenclature: in many species the paraseptal cartilage bifurcates rostrally into a dorsal and a ventral branch.(ABSTRACT TRUNCATED AT 400 WORDS)     

Amphibian cells in culture. I. Nutritional studies.

Nutritional requirements of amphibian cells in culture were studied for the purpose of modifying a minimal medium in which frog cells could proliferate and which could be used for obtaining drug-resistant and auxotrophic variants. The serum, purine, CO2, and amino acid requirements for ICR 2A (a Rana pipiens haploid cell strain) have been investigated employing two different media: L-15, a nonbicarbonate, amino acid-buffered medium and Eagle's MEM, a bicarbonate-buffered medium. In this paper we present evidence to support the following conclusions: (1) With L-15 as the base medium, 10% fetal calf serum (FCS) supports optimal cell growth during exponential phase. Calf serum, whole, dialyzed, or heat-inactivated, cannot substitute for FCS and, in fact, is inhibitory. (2) Purines are required by ICR 2A cells only if grown in a nonbicarbonate-buffered medium, since the cells under these conditions cannot produce enough endogenous CO2 to support de novo purine synthesis. (3) In addition to the amino acids considered essential for mammalian cells in culture, ICR 2A cells depend upon exogenous asparagine. Glutamine and/or aspartic acid cannot replace the asparagine requirement. However, ICR 2A cells do utilized exogenous glutamine as an oxidative substrate.     

ICMSF methods studies. VIII. Comparative study for the enumeration of Clostridium perfringens in foods.

Four methods were compared in an international comparative study for the enumeration of Clostridium perfringens: the SFP (Shahidi-Ferguson perfringens) agar (A), TSC (tryptose-sulfite-cycloserin) agar (B), SC (sulfite-cycloserine) agar (C), and neomycin blood agar (D) methods. The confirmed C. perfringens counts were slightly lower for D than for A-C. The percentages of presumptive colonies confirmed as C. perfringens were essentially the same in each method. The relative numbers of nonspecific colonies were the lowest in C, followed by B, D, and A. The methods were also compared for simplicity and for aspects associated with the recognition and selection of presumptive colonies.     

ICMSF methods studies. XII. Comparative study for the enumeration of Clostridium perfringens in feces.

As the second phase of an international comparative study for the enumeration of Clostridium perfringens, four methods were compared for "total" and spore counts of C. perfringens in fecal specimens: the SFP (Shahidi-Ferguson perfringens) agar (A), TSC (tryptose-sulfite-cycloserine) agar (B), SC (sulfite-cycloserine) agar (C), and neomycin blood agar (D) methods. In both the total and spore count procedures, the confirmed C. perfringens counts in method D were lower than in methods A, B, and C. Little differences among methods were found in the percentages of presumptive colonies confirmed as C. perfringens. The nonspecific counts in methods A and D were generally greater than in B and C, but nonspecific microorganisms did not interfere in the enumeration of C. perfringens spores by any of the four methods. In overall performance, methods B and C were superior to A and D. The mean C. perfringens spore count was only 0.17 log lower than the mean total count. Spore counts alone are, therefore, adequate in investigations of C. perfringens outbreaks.     

Organ culture of suckling rat intestine: Comparative study of various hormones on brush border enzymes

Summary Jejunal mucosa of 6 d-old rats were cultured for 24 and 48 h in the presence of thyroxine, insulin, pentagastrin, glucagon, epidermal growth factor (EGF) or dibutyryl-A-3:5-MP cyclic with or without dexamethasone (DX). The enzymes were assayed on the purified brush borders. The various agents added alone to the basic culture medium had no effect with the exception of DX on the levels of enzyme activities. Dexamethasone alone induced sucrase, stimulated maltase, and protected other brush border enzyme activities (aminopeptidase, lactase, and alkaline phosphatase). When added to DX-supplemented medium, only the following factors modified the levels of enzymatic activities observed with DX alone. Insulin (10⁻⁶ M) increased maltase, alkaline phosphatase, and lactase activity to a greater extent than DX at 24 h culture, the effect being maintained at 48 h on alkaline phosphatase only. At 48 h culture, both EGF (10⁻⁸ M) and dbcAMP (10⁻³ M) decreased DX-induced sucrase activity. The latter agent also depressed DX-stimulated aminopeptidase activity. This work was supported by the Institut National de la Santé et de la Recherche Médicale, Centre National de la Recherche Scientifique, and a grant 79.7.1243 from the Délégation Générale a la Recherche Scientifique et Technique. P. M. S. is a recipient of a grant from Fondation de la Recherche Médicale (France).