Toxigenic Aspergillus flavus and aflatoxins in Sri Lankan medicinal plant material

The fungal flora of 6 Asian medicinal plants, Aerva lanata (Linn.) Juss. Alyssicarpus vaginalis D.C., Tribulus terrestris Linn. Adhatoda vasica Nees., Centella asciatica (L.) Urb., Cardiospermum halicacabum Linn. was determined. After surface disinfection Aspergillus spp. were most frequently observed. Aspergillus flavus, isolated from Alyssicarpus vaginalis and Aerva lanata produced aflatoxins in culture. Aflatoxin B₁ was also detected in a sample of Aerra lanata at a level of 0.5 g/g. Plant material destined for medicinal use should be stored carefully prior to its use to prevent growth of naturally occurring toxigenic mold fungi.     

Two kinds of calcium channels in canine atrial cells. Differences in kinetics, selectivity, and pharmacology

Currents through Ca channels were recorded in single canine atrial cells using whole-cell recording with patch pipettes. Two components of Ca channel current could be distinguished. One ("Ifast") was present only if cells were held at negative potentials, was most prominent for relatively small depolarizations, and inactivated within tens of milliseconds. The other ("Islow"), corresponding to the Ca current previously reported in single cardiac cells, persisted even at relatively positive holding potentials, required stronger depolarizations for maximal current, and inactivated much more slowly. Both currents were unaffected by tetrodotoxin and both were reduced by Co. Ifast had the same size and kinetics when Ca was exchanged for Ba, while Islow was bigger and slower with Ba as the charge carrier. In isotonic BaCl2, fluctuation analysis showed that Ifast had a smaller single channel current than Islow. Islow was much more sensitive to block by nitrendipine than was Ifast; also, Islow, but not Ifast, was increased by the dihydropyridine drug BAY K8644. Isoproterenol produced large increases in Islow but had no effect on Ifast.     

Characterization of virulent and avirulent A/chicken/Pennsylvania/83 influenza A viruses: potential role of defective interfering RNAs in nature.

In April 1983, an influenza virus of low virulence appeared in chickens in Pennsylvania. Subsequently, in October 1983, the virus became virulent and caused high mortality in poultry. The causative agent has been identified as an influenza virus of the H5N2 serotype. The hemagglutinin is antigenically closely related to tern/South Africa/61 (H5N3) and the neuraminidase is similar to that from human H2N2 strains (e.g., A/Japan/305/57) and from some avian influenza virus strains (e.g., A/turkey/Mass/66 [H6N2]). Comparison of the genome RNAs of chicken/Penn with other influenza virus isolates by RNA-RNA hybridization indicated that all of the genes of this virus were closely related to those of various other influenza virus isolates from wild birds. Chickens infected with the virulent strain shed high concentrations of virus in their feces (10(7) 50% egg infective dose per g), and the virus was isolated from the albumin and yolk of eggs layed just before death. Virus was also isolated from house flies in chicken houses. Serological and virological studies showed that humans are not susceptible to infection with the virus, but can serve as short-term mechanical carriers. Analysis of the RNA of the viruses isolated in April and October by gel migration and RNA-RNA hybridization suggested that these strains were very closely related. Oligonucleotide mapping of the individual genes of virulent and avirulent strains showed a limited number of changes in the genome RNAs, but no consistent differences between the virulent and avirulent strains that could be correlated with pathogenicity were found. Polyacrylamide gel analysis of the early (avirulent) isolates demonstrated the presence of low-molecular-weight RNA bands which is indicative of defective-interfering particles. These RNAs were not present in the virulent isolates. Experimental infection of chickens with mixtures of the avirulent and virulent strains demonstrated that the avirulent virus interferes with the pathogenicity of the virulent virus. The results suggest that the original avirulent virus was probably derived from influenza viruses from wild birds and that the virulent strain was derived from the avirulent strain by selective adaptation rather than by recombination or the introduction of a new virus into the population. This adaptation may have involved the loss of defective RNAs, as well as mutations, and thus provides a possible model for a role of defective-interfering particles in nature.     

Are seals frequently infected with avian influenza viruses?

Influenza A virus isolates of the H4N5 subtype (which has previously been detected only in birds) were recovered from harbor seals dying of viral pneumonia on the New England coast from June 1982 through March 1983. When these isolates were compared with other mammalian and avian viruses in serological assays and RNA-RNA competitive hybridization, it was found that the seal viruses were most closely related antigenically and genetically to recent avian virus strains and were readily distinguishable from mammalian viruses, including H7N7 isolates recovered from seals in 1980. Unlike any previous isolates from mammals, these recent seal viruses replicate in the intestinal tracts of ducks, a characteristic of avian viruses. The association of avian viruses with influenza outbreaks in seals suggests that transmission of avian viruses to seals is occurring in nature. Potentially, this may be an example of the adaptation of avian viruses to mammals, which would represent an intermediate step in the evolution of new mammalian strains.     

Site recognition by protein-primed T cells shows a non-specific peptide size requirement beyond the essential residues of the site. Demonstration by defining an immunodominant T site in myoglobin.

In previous studies, six T sites within myoglobin (Mb) were localized. To define precisely the boundaries of the T sites, a new approach is introduced and applied here to the T site residing within residues 107-120 of Mb. Two sets of peptides were synthesized. One set represents a stepwise elongation by one-residue increments of the Mb sequence. The other set represents an identical stepwise addition of one-residue increments of the Mb sequence, but which were extended by additional unrelated (nonsense) residues to a uniform size of 14 residues. The longer peptides (nonsense-extended) usually gave higher proliferative responses than did their shorter counterparts having the same Mb region. Thus a minimum peptide size is required for optimal T-cell stimulation. The T site subtends, in three high-responder mouse strains, residues 109-119 or 110-120, depending on strain, and, in three low-responder strains, maps to residues 108-120. Thus, in this case, the T site coincides with the site of B-cell recognition and resides in a small discrete surface region of the protein chain.     

A comparison of the effect of three fungicides on growth and roridin E production by Myrothecium roridum

Three fungicides, chlorothalonil, dichloran and mancozeb were studied to determine the effects on growth and production of roridin E by Myrothecium roridum in vitro. With increasing concentrations of fungicides, both growth and production of roridin E were inhibited. Of the three fungicides, chlorothalonil was much more effective in the inhibition of growth and production of roridin E, than dichloran and mancozeb.     

Suppression of the responsiveness of lymphocytes from cancer patients triggered by co-culture with autologous tumor-derived cells

The question as to whether or not cancer patients have "tumor antigen"-induced suppressor T cells is of considerable interest and importance. As an approach to that question, the effect of addition of autologous irradiated tumor-derived cells (TDC) on the mixed lymphocyte response (MLR) of patients' lymphocytes (Ly) and of healthy donor Ly was tested. The rationale for these experiments was based on the fact that circulating antigen-responsive blood lymphocytes can be reactivated in vitro by exposure to the appropriate antigen. Thus, if there are circulating tumor "antigen"-reactive suppressor Ly, exposure to TDC as a source of the antigen should reactivate those cells. Reactivation of suppressor cells might result in diminished responsiveness to other stimuli such as alloantigens in the mixed leukocyte culture. We found that the addition of TDC to Ly cultures produced four distinct patterns of reaction. In 26 of the 74 different patient-tumor assays, the addition of autologous TDC to the patient cultures inhibited MLR, but the addition of the same TDC to cultures of Ly from healthy donors had no effect or increased their responsiveness (Specific Suppression). In 21 cases, the addition of autologous TDC to the patient cultures suppressed the MLR and the addition of the same TDC to control cultures suppressed the response of some but not all the healthy donors (Selective Suppression). In four cases, the addition of TDC to the cultures suppressed the MLR of the patients and all of the control donors (Nonspecific Suppression). In 23 cases, the addition of autologous TDC resulted in no suppression of the patient MLR or of any of the simultaneously tested normal donors (No Suppression). When TDC of patients with noninvasive bladder cancer were added to their own Ly cultures, only four of 11 produced specific or selective suppression compared to 11 of 12 when TDC came from patients with superficially invasive cancer. These data provide indirect evidence to support the hypothesis that human tumors induce circulating suppressor cells that may be reactivated in vitro by co-culture with TDC.     

Selective Utilization of Pyrimidine Deoxyribonucleosides for Deoxyribonucleic Acid Synthesis in Pneumococcus

When pyrimidine deoxyribonucleosides are supplied to growing cultures of Diplococcus pneumoniae, they are selectively used for incorporation into deoxyribonucleic acid (DNA). Differently labeled molecules of deoxyuridine, thymidine, and deoxycytidine were used to study the precursor pathways of this organism. Each of these preformed pyrimidine deoxynucleosides is incorporated intact (i.e., without cleavage of the glycosidic bond) and is predominantly recoverable as DNA thymidine. During the utilization of deoxycytidine and deoxyuridine by pneumococci, large proportions of the available precursor are converted to free thymidine, which is secreted back into the growth medium. The biochemical pathways for selective incorporation into DNA and the regulation of concentrations of intracellular thymidine compounds by excretion of free thymidine are discussed.     

Discrete conductance fluctuations in lipid bilayer protein membranes

Discrete fluctuations in conductance of lipid bilayer membranes may be observed during the initial stages of membrane interaction with EIM ("excitability inducing material"), during destruction of the EIM conductance by proteolysis, and during the potential-dependent transitions between low and high conductance states in the "excitable" membranes. The discrete conductance steps observed during the initial reaction of EIM with the lipid membranes are remarkably uniform, even in membranes of widely varying lipid composition. They range only from 2 to 6 x 10_-10 ohm_-1 and average 4 x 10_-10 ohm_-1. Steps found during destruction of the EIM conductance by proteolysis are somewhat smaller. The transition between high conductance and low conductance states may involve steps as small as 0.5 x 10_-10 ohm_-1. These phenomena are consistent with the formation of a stable protein bridge across the lipid membrane to provide a polar channel for the transport of cations. T6he uniform conductance fluctuations observed during the formation of these macromolecular channels may indicate that the ions in a conductive channel, in its open state, are largely protected from the influence of the polar groups of the membrane lipids. Potential-dependent changes in conductance may be due to configurational or positional changes in the protein channel. Differences in lipid-lipid and lipid-macromolecule interactions may account for the variations in switching kinetics in various membrane systems.     

Changes in the titer of the female-predominant storage protein (81K) during larval and pupal development of the waxmoth,galleria mellonella

The levels of an 81K storage protein in the waxmoth, Galleria mellonella, were monitored during the course of development using rocket immunoelectrophoresis. During the fifth and sixth larval stadia, 81K protein levels increased during feeding and growth but sharply declined at each larval molt. During the fifth and sixth stadia hemolymph levels of the 81K protein increased to about 1 and 2.5 mg/ml, respectively, with no discernible differences between levels in males and females. Neither the fat body nor the remainder of the carcass contained the 81K protein, indicating that the accumulation of this protein during the intermolt period was exclusively in the hemolymph and redistribution of the 81K protein into other tissues does not occur at the final two larval molts. During the seventh (final) larval stadium the absolute quantities of the 81K protein increased from 23 μg per insect to over 1,600 μg in females and to 300 μg in males. The hemolymph concentration of the 81K protein reached 28 mg/ml in females and 6 mg/ml in males with only low levels found in the remaining tissues. Shortly after pupal apolysis, marked by eyespot retraction, the fat body in both sexes rapidly and quantitatively sequestered the 81K protein from the hemolymph. The 81K protein in the hemolymph of both males and females rapidly dropped to nearly zero concentration by pupation. The 81K storage protein remained localized in the fat body cells after uptake occurred, even though the fat body cells disaggregate and reaggregate during metamorphosis. During pharate adult development the 81K storage protein disappeared from the fat body without entering the hemolymph. At adult eclosion 81K was virtually absent from the tissues of both males and females.