- Volume 61, Issue 1, 1970

Summary: A simple and rapid colorimetric test has been developed for the detection of pyrrolidonpl peptidase (PLP) activity in bacteria. Of 2354 strains belonging to various groups of enteric bacteria tested, 451 were PLP-positive. These included Citrobacter, 223 out of 226 strains, Klebsiella (178/188), Enterobacter (49/52), and one strain of Serratia.

Pyrrolidonyl peptidase was not detected in 1146 Shigella strains, 354 Salmonella strains, 333 Escherichia strains, 21 Arizona strains, nor in 33 strains of Proteus, Providencia and Hafnia.

In addition, 56 cultures of various Gram-negative and Gram-positive bacteria were tested for PLP. Enzyme activity was found in strains belonging to the genera Bacillus, Streptococcus, Staphylococcus, Micrococcus, Sarcina, Neisseria and Pseudomonas. The possibility of employing the PLP test for differentiation within the family Enterobacteriaceae is discussed.

SUMMARY:Phenol-water extracted lipopolysaccharides (LPS) from Bacteroides fragilis nctc 9343 did not contain heptose or 2-keto-3-deoxy-octonate. The sugar components identified were glucosamine, galactosamine, glucose, galactose, fucose, rhamnose and traces of mannose. An unidentified amino compound was found. The preparations were free of nucleic acids and protein constituted only a minor fraction of the LPS. The LPS preparations had endotoxic effect when tested in rabbits, but the endotoxic potency was low.

SUMMARY: Breakdown of DNA to acid-soluble fragments induced in strains of Escherichia coli b by nalidixic acid (10 μg./ml.) differed in linear rate according to the ability of the strain to repair DNA lesions. Strains which were exr (bs-2) and exr, uvr (bs-i) exhibited excessive DNA breakdown following nalidixic add treatment. The excision-defective uvr strains, bs-8, bs-12 and wwp-2 her, also degraded their DNA to acid-soluble fragments at a rate which initially was greater than that of the parental strain (E. coli b). This degradation was unaffected by the rad mutation. Removal of nalidixic acid from the system considerably lowered the rate of DNA breakdown. The responses to nalidixic acid and other agents potentiating DNA breakdown are compared.

SUMMARY: Nitrogen fixation has been obained with strains of Desulfovibrio vulgaris and D. gigas, organisms hitherto believed to be incapable of using molecular nitrogen. Fixation has been demonstrated by increases in total nitrogen and by uptake of 15N2. Fixation of N2 may be widespread in this genus of the sulphate-reducing bacteria.

SUMMARY: Chloramphenicol (1 mg. /ml.) increased the mean doubling time of Polytomella caeca from 3·6 to 5 hr and gave 62 % inhibition of the final yield of organisms on an acetate + mineral salts + thiamine medium. The rates of acetate oxidation and tetrazolium reduction by whole organisms decreased progressively when grown in the presence of increasing chloramphenicol concentrations; no ultrastructural abnormalities of the mitochondria of such organisms were seen in electron micrographs. Succinoxidase and NADH oxidase activities of isolated mitochondria were decreased by 43 % and 72 %, respectively, by growth of organisms with 1 mg. chlor-amphenicol/ml. The content of cytochromes (a + a3 ) was decreased by 40 %, and cytochrome oxidase activity was about 50 % that of normal mitochondria. No alteration in the activities of rotenone-sensitive NADH-cytochrome c oxidoreductase or succinate-cytochrome c oxidoreductase was detected, using mammalian cytochrome c as electron acceptor. However, the rate of reduction of electron-transport components during the aerobic-anaerobic transition (with succinate as substrate) suggested that the supply of electrons to the respiratory chain was rate-limiting and might be more important in leading to respiratory deficiency than the decreased cytochrome oxidase activity. No major cytochrome dislocation was detected, suggesting that the respiratory chain was itself intact.

SUMMARY: The enzymic properties of 46 strains of β - lactamase-producing enteric bacteria were examined. Eight distinct types of β - lactamase were detected among these strains when substrate profile, sensitivity to p-chloromercuri- benzoate (pCMB) and to cloxacillin inhibition, reaction with antiserum and charge properties are used as parameters of enzyme type. The types of enzyme detected ranged from molecules with a predominantly cephalo- sporinase profile to those where penicillins were hydrolysed much more rapidly than cephalosporins. The majority of isolates synthesized an enzyme that was almost equally active against penicillins and cephalosporins. To date only three of the eight types of enzyme have been shown to be transferable by conjugation.

Non-restricting (r−) mutants of two different host specificities carried by resistance transfer factors have been isolated. As previously found with other host specificities, the non-restricting mutants were of two phenotypes: those that retained the ability to modify DNA, and those which had lost the ability to modify DNA. These mutants were tested for complementation with wild-type host specificities carried either on the Escherichia coli chromosome, on resistance transfer factors, or on the phage Pi. No complementation was observed and possible explanations for this finding are considered.

Summary: This paper deals with taxonomic problems caused by the occurrence in nature of forms of Mallomonas species which are considered to be stages in development. They are called ‘imperfect’ forms; that is, forms that lack features that have been included in the diagnosis. These individuals might be described as separate species except when intermediate forms are noticed which link them with the perfect form of diagnosed species. The fine details of their scales are the same as in the perfect form.

SUMMARY: A new species of the torquata group of the genus Mallomonas is described by light and electron microscopes. A list is given of those members of the torquata group which have been published with illustrations obtained with the use of light and electron microscopes. A few of the torquata group which have been published only with light microscope illustrations are listed and discussed. It is considered that no clear decision about their nomenclature can be reached without an electron micrograph of the scales.

Summary: Resting Aspergillus flavus synthesized more aflatoxin on a medium with glucose than they did with any other substrate tested. d-Xylose and ethanol fostered the formation of kojic acid but not aflatoxin. Yields of kojic acid and aflatoxin responded differently to alteration in temperature, pH, surface-volume ratio of the culture medium, and the presence of various chemicals in the medium. [14C] Acetate as a substrate led to strongly labelled aflatoxin being formed. The simultaneous addition of unlabelled kojic acid did not lower the relative isotope content of the synthesized toxin. On the other hand, addition of unlabelled acetate to medium with [14C]kojic acid did reduce the relative isotope content of the toxin synthesized. It is concluded that the synthesis of kojic acid and aflatoxin follow separate pathways, and that kojic acid is not an intermediate in aflatoxin synthesis by resting cells of A. flavus.

SUMMARY: Sporidesmin production by 37 recently isolated strains and one heavily sporing laboratory isolate of Pithomyces chartarum was assayed by toxicity in vitro to tissue culture cells and related to sporulation and growth. Under standard cultural conditions strains varied greatly in their ability to produce sporidesmin and spores. Heavily sporing cultures produced most sporidesmin and the level of sporidesmin production by a strain could be changed by manipulation of cultures in ways which also stimulated or depressed sporulation. Ultraviolet radiation increased sporulation and sporidesmin production in 33 strains. Shaken cultures, in which growth was good but sporulation suppressed, produced no sporidesmin. The close association of sporidesmin production with sporulation supports the reliability of spore counts as an index of pasture toxicity. Most and probably all strains of P. chartarum are potentially able to produce moderate to high levels of sporidesmin. Ultraviolet radiation may stimulate production of sporidesmin by the growing fungus but destroys it in aqueous solutions leached from the senescent mycelium.

SUMMARY:The mode of addition of the steroid substrate, 16α-hydroxycortexolone 16,17-acetonide, was shown to influence markedly the rate of enzymic conversion of the steroid as well as the form of growth of the fungal organism, in a mixed culture of Arthrobacter simplex (1-dehydrogenator) and Curvularia lunata (11β-hydroxylator). The effects observed were apparently related to the size and solubility of the steroid particles added in suspension or precipitated in the medium by addition of the steroid in any of several water-miscible non-aqueous solvents. The best rates of steroid conversion were observed when the mould grew in elongated pellets rather than in filamentous form.

SUMMARY: Methionine-requiring mutants of Saccharomyces cerevisiae produce large amounts of hydrogen sulphide from sulphate, sulphite or thiosulphate when grown in the presence of suboptimum concentrations of methionine. O-Acetylhomoserine and homocysteine act like methionine with a methionine-requiring mutant which can use them for growth. Wild-type strains of S. cerevisiae and S. carlsbergensis also form large amounts of hydrogen sulphide from inorganic sulphur sources when the yeast is deficient in either pantothenate or vitamin B6. This excess sulphide production is inhibited by methionine or its immediate precursors, suggesting that both vitamins are required for methionine biosynthesis. O-Acetylhomoserine is a normal precursor of homocysteine and methionine in S. cerevisiae and S. carlsbergensis. The effect of pantothenate on sulphide production by these yeasts is probably due to its involvement in the formation of O-acetylhomoserine.

Summary: Fifty-two strains, comprising six Rhizobium species, were examined for their esterase patterns using electrophoresis on cellulose acetate. Esterase activity was detected in five Rhizobium species. The sixth species, R.japonicum, was characterized by the absence of esterase activity in all but one of the strains examined. Rhizobium trifolii and R. leguminosarum strains showed similarities in their esterase profiles. Rhizobium meliloti strains formed a group distinct from these on the basis of their esterase patterns. Rhizobium lotus sp. and R. phaseoli also exhibited esterase activity.

Heat denaturation and metal inhibition studies suggest that the esterase activity is truly enzymic. The inability of the bacterial esterase to react with a synthetic peptide suggests that residual esterase activity associated with certain proteolytic enzymes is not involved. Heat tests revealed differences in the sensitivity of the multiple forms of esterases in rhizobia to inactivation.

SUMMARY: Vitamin requirements for growth in a casein hydrolysate medium were determined for 27 strains of Aerococcus viridans from diverse sources. Pantothenic acid, nicotinic acid and biotin were either absolutely required by, or markedly stimulatory to, all strains; none required thiamin, riboflavin, pyridoxine, folic acid or folinic acid. Tween 80 replaced the biotin requirement of most strains. Amino acid requirements were not sharply defined and varied from strain to strain. As the amino acid composition of the medium was simplified the amount of growth was decreased and most strains would not grow when biotin was replaced by Tween 80. A single purine base was required: either guanine or xanthine alone satisfied this requirement for each of the strains tested; adenine was a suitable alternative source for some strains. Exogenous pyrimidine was not required.

SUMMARY: It has been established that, in pneumococcus, donor sites on transforming deoxyribonucleic acid (DNA) can be divided into at least two types, known as low-efficiency (LE) sites and high-efficiency (HE) sites, by comparing the number of transformants of a particular character with that obtained for a standard reference gene ( Ephrussi- Taylor, Sicard & Kamen, 1965 ; Lacks, 1966 ). The reference gene used by Ephrussi- Taylor and her colleagues was str-41, whilst Lacks used the sulf-d gene. Ephrussi- Taylor & Gray (1966) proposed that LE markers are integrated into both chains of the recipient DNA by an excision and repair mechanism, whilst HE markers have to wait for a further division before both the recipient strands become altered. Evidence that the LE markers are transmitted into both strands whilst HE markers are transmitted to only one strand has been put forward by Ephrussi-Taylor (1966) , Gray & Ephrussi-Taylor (1967) and Louarn & Sicard (1968) . Louarn & Sicard (1969) have taken advantage of this difference between the integration of the markers to show that either of the recipient strands of DNA can be transformed by the HE marker. They calculated the ratio R = (D.N)/(A.B), where D = number of doubles transformed for genes A and B, N = the total colony-forming units (c.f.u.) present, and A and B the number of transformants for genes A and B respectively. By comparing the ratios obtained for pairs of markers having the composition HE-HE, LE-LE, or HE-LE, Louarn and Sicard showed that the results were consistent with either strand being transformed by an HE marker. They also discussed the effect of replication events on the value of R, particularly with respect to the location on the chromosome of such events in relation to the markers. The results reported below support these conclusions.

When Euglena gracilis is grown in the presence of some antibiotics (Provasoli, Hutner & Schatz, 1948 ; Ebringer, 1962, 1966 ; Celmer & Ebringer, 1967 ), organisms with no chloroplasts are soon produced. These aplastidic organisms can then be isolated and maintained as permanently colourless races. The mechanism of the bleaching action is still in dispute. We have suggested an interaction with plastid DNA or some hereditary apparatus responsible for chloroplast replication (Ebringer, Mego & Jurasek, 1969 ). To test this hypothesis, we have examined some antibacterial agents which inhibit DNA synthesis in bacteria or in other organisms. Nalidixic acid inhibits specifically the synthesis of DNA in bacteria (Goss, Deitz & Cook, 1965 a, b; Cook et al. 1966 ) and exerts a mutagenic effect upon proliferating bacteria (Cook, Goss & Deitz, 1966 ). Some other agents which affect DNA synthesis in bacteria also exert a bleaching action on Euglena gracilis (to be published), so the question arises whether nalidixic acid is able to produce permanent colourless races of this organism.