South African Strains Research Articles

The effect of temperature on the development of wilting and on progeny tuber infection of potatoes inoculated with South African strains of biovar 2 and 3 ofPseudomonas solanacearum

Potato plants, after inoculation with a South African strain of biovar 2 or a strain of biovar 3 ofPseudomonas solanacearum were grown in growth chambers at temperatures ranging from 14/16°C (dark/light) up to 25/30°C. When grown at 14/16°C or higher, plants inoculated with the biovar 2 strain developed wilting symptoms and internal symptoms on progeny tubers and the disease was transmitted by the progeny tubers. When the plants were inoculated with the biovar 3 strain, wilting symptoms developed from 18/20°C, internal symptoms on progeny tubers from 20/22°C and the disease was transmitted by progeny tubers from plants grown at 16/18°C or higher.

Characterization of Pseudomonas syringae pv. tomato by Numerical Analysis of Phenotypic Features and Total Soluble Proteins11)

Abstract The homogeneity of South African and other strains of Pseudomonas syringae pv. tomato was determined by numerical analysis of differential phenotypic features and computer‐assisted comparison of gel electropherograms of soluble proteins. Two closely‐related groups, that could only be separated by their reaction in litmus milk, were distinguished in a phenotypic dendrogram. Three non‐South African reference strains fell outside these groups, indicating the possible existence of more groups, worldwide. No clear‐cut correlation was found between clustering of strains and their geographical origin. Forty‐seven strains examined had similar protein patterns, reflecting their close genetic resemblance. All strains were pathogenic on tomatoplants.

Characteristics of South African Strains of Pseudomonas solanacearum

Characteristics of South African Strains of Pseudomonas solanacearum

Comparative immunochemical and biological analysis of African and South American yellow fever viruses

Four geographic variants (topotypes) of yellow fever (YF) virus from Africa and South America previously determined by RNA oligonucleotide mapping were analyzed for their structural, antigenic and virulence characteristics. The electrophoretic migration mobility and carbohydrate content of the envelope protein E characterized YF virus strains of South America. The NS 1 protein of South American and Central African YF virus strains was not precipitated by anti-NS 1 monoclonal antibodies (MAB) that precipitated NS 1 of West African strains. No distinction could be made among the YF virus strains on the basis of the neutralizing capacity of available MAB. South American, but not African YF virus strains were virulent for 8-day-old mice upon intraperitoneal inoculation. The results permitted characterization of topotypes of Central Africa and South America but failed to differentiate YF strains from West Africa.

Preparation and use of monoclonal antibodies for identifying Crimean-Congo hemorrhagic fever virus.

Seven monoclonal antibodies were prepared against a South African strain of Crimean-Congo hemorrhagic fever (CCHF) virus and were found to be directed against viral nucleocapsid protein. Five of the monoclonal antibodies reacted to high titer in indirect immunofluorescence (IF) tests and enzyme-linked immunosorbent assays (ELISA) with 22 strains of CCHF virus and failed to cross-react with the closest antigenic relative of CCHF, Hazara virus, or with 4 other nairoviruses which need to be distinguished from CCHF virus in Africa. These antibodies, used in the IF technique, readily detected antigens induced by all strains of CCHF virus included in the study in cell culture monolayers and mouse brain tissue, which represent the systems commonly used for isolation of CCHF virus. The IF technique with monoclonal antibodies constitutes a rapid and specific means of identifying newly isolated strains of CCHF virus.

Penicillin-binding proteins of penicillin-susceptible and -resistant pneumococci: immunological relatedness of altered proteins and changes in peptides carrying the beta-lactam binding site.

There are several major differences between the penicillin-binding proteins (PBPs) of highly penicillin-resistant and -susceptible strains of pneumococci. The highest-molecular-size PBP 1a (98 kilodaltons [kDa]) of susceptible pneumococci is not detectable in resistant bacteria. Instead, resistant strains contain a PBP of smaller size: 92 and 94 kDa in South African strains 8249 and A95210, respectively, and 96 kDa in New Guinea strain 2955 (S. Zighelboim and A. Tomasz, Antimicrob. Agents Chemother. 17:434-442, 1980). Using antibodies prepared against PBP 1a of penicillin-susceptible pneumococci, we demonstrated that these anomalous-sized proteins in the resistant strains are immunologically related to PBP 1a of penicillin-susceptible bacteria. A second difference between the PBP patterns of strain 8249 and the susceptible pneumococci is that the 78-kDa PBP 2b is not detectable by the radioactive penicillin binding assay in the resistant strain. Using antibodies prepared against PBP 2b of susceptible cells, we demonstrated the presence of PBP 2b in membrane preparations from strain 8249 cells. Thus, the poor detection of this PBP appears to be related to its greatly decreased affinity for the antibiotic molecule. We also compared the patterns of penicillin-labeled peptides derived from PBPs of resistant and susceptible cells during partial proteolysis by V8 protease. Several changes were observable in small peptides carrying the beta-lactam binding site generated from the high Mr (PBP 1a-related) binding proteins. In contrast, no differences in the pattern of penicillin-labeled peptides were seen when the pattern of PBP 2a of susceptible pneumococci was compared with the peptide pattern of PBP 2a from resistant strains. One of the resistant isolates (strain 2955) also had a PBP 3 with a higher-than-normal molecular weight. This protein gave strong positive reaction with antibodies against PBP 3 of susceptible cells. Examination of the pattern of penicilloyl peptides generated from the susceptible and resistant PBP 3s during partial proteolysis revealed only differences which seem to reside distant from the beta-lactam binding site.

Alterations in kinetic properties of penicillin-binding proteins of penicillin-resistant Streptococcus pneumoniae.

Earlier studies have shown that the highly penicillin-resistant South African Strains of pneumococci contain altered penicillin-binding proteins (PBPs) (S. Zighelboim and A. Tomasz, Antimicrob. Agents Chemother. 17:434-442, 1980). We now describe a detailed quantitative characterization of the reaction of radioactively labeled penicillin with the PBPs of the penicillin-susceptible and penicillin-resistant pneumococci and several intermediate-resistance-level genetic transformants as well. The altered binding of the antibiotic by the PBPs of resistant cells appears to be due to a combination of two factors: lower drug affinity and change in the cellular amounts of PBPs. No alteration in the rates of deacylation of the penicilloyl-PBPs of the resistant cells was detected.

Penicillin tolerance in multiply drug-resistant natural isolates of Streptococcus pneumoniae.

Five of six multiply drug-resistant clinical isolates of Streptococcus pneumoniae from South Africa demonstrated penicillin tolerance. In contrast to the common wild-type strains of pneumococci, treatment of the tolerant strains with penicillin above the minimum inhibitory concentration did not induce cell wall degradation, lysis, or leakage of intracellular components, and the rate of loss of viability was reduced compared with that of nontolerant strains. While these South African strains contained lower specific activity of autolytic enzyme than did nontolerant strains, the residual autolytic activity (15%-26% of the nontolerant wild type) was much more than that found in lysis-defective laboratory mutants of pneumococci (less than or equal to 1%); the rate of penicillin-induced lysis did not correlate with the specific activity of residual autolysin. Also, in contrast to the complete lysis resistance of lysis-defective mutants to all lytic agents, the tolerant South African strains were resistant primarily to lysis by beta-lactam antibiotics but could still be lysed by other cell wall inhibitors (e.g., cycloserine) and detergents. The penicillin resistance and penicillin tolerance traits could be separated by genetic transformation. We suggest that the drug-specific tolerance of the South African pneumococcal strains is related to some alteration in the control of autolysin activity.

Genetic and molecular studies of plasmids coding for colonization factor antigen I and heat-stable enterotoxin in several escherichia coli serotypes.

Plasmids coding for colonization factor antigen I (CFA/I) and heat-stable enterotoxin (ST) were identified in 10 strains of human enterotoxigenic Escherichia coli. The strains, which belonged to serogroups O63, O114, O128, and O153, were isolated in Bangladesh, Latin America, Spain, and South Africa. Two strains produced heat-labile enterotoxin in addition to ST. CFA/I-ST plasmids were mobilized from two O128 strains into E. coli K-12 with the R factor R1-19K-. Like the prototype CFA/I-ST plasmid NTP113, mobilized previously from an E. coli O78 strain into K-12, these two plasmids were non-autotransferring. All 10 CFA/I-ST plasmids were incompatible with NTP113 and had molecular weights ranging from 59 X 10(6) to 72 X 10(6). The molecular properties of seven of these plasmids were compared with those of six CFA/I-ST plasmids previously mobilized from O78 strains from Ethiopia, South Africa, and Bangladesh and with those of one plasmid coding for CFA/I, ST and heat-labile enterotoxin from a South African strain of serogroup O63. Digestion with the restriction endonuclease HindIII showed that several plasmids had very similar fragment patterns and two were identical. Generally, a larger proportion of HindIII fragments were of common size in digests of plasmids identified in strains from related geographical areas, regardless of serogroup. However, all except one plasmid shared five or six HindIII fragments of the same size, one of which had been shown previously to be involved in CFA/I production. There was at least 90% DNA homology between CFA/I-ST plasmids with a molecular weight of about 58 X 10(6) from O78 strains from different sources. Most of the DNA sequences of these plasmids were present in a larger CFA/I-ST plasmid (72 X 10(6) from an O128 strain. The results of genetic and molecular studies suggest that CFA/I and ST production is determined by very similar plasmids in different serogroups of human enterotoxigenic E. coli from several sources.

Multiple antibiotic resistance in South African strains of Streptococcus pneumoniae: mechanism of resistance to beta-lactam antibiotics.

Multiply antibiotic-resistant strains of Streptococcus pneumoniae appeared in South Africa in 1977. In these organisms resistance to chloramphenicol is caused by an inducible, drug-inactivating enzyme; however, the basis for resistance to other antibiotics is unknown. Pneumococci with increased resistance to beta-lactam antibiotics do not produce beta-lactamases, a finding indicating the presence of intrinsic resistance to these drugs. One approach to the understanding of the mechanism of this resistance was to study the pattern of penicillin-binding proteins (PBPs) in the South African pneumococci. With the use of highly radioactive penicillin to label PBPs in vivo, it was found that the South African pneumococci have a PBP pattern that differs from that of the sensitive laboratory strain R6 in several respects. Differences include (1) a lack of PBPs la and lb; (2) the presence of a new, faster moving protein (lower molecular weight), named PBP lc; (3) an apparent decrease in the affinity of PBP 2a for [3H]penicillin; and (4) a lack of PBP 2b. Taking advantage of the fact that penicillin resistance is a property acquired in a stepwise process, a series of isogenic and progressively more resistant transformants was constructed. DNA from the resistant South African strain 8249 of S. pneumoniae was used as the donor in a series of transformations for which the recipients were either strain R6 or transformants of organisms with lower levels of resistance. In vivo labeling of the PBPs of these transformants revealed a gradual shift from a pattern similar to that of the sensitive strain (in the transformants of lower resistance) to a pattern resembling that of the highly resistant donor strain (in the transformants of higher resistance) as the level of penicillin resistance increased.

Penicillin susceptibility of Streptococcus pneumoniae in 1978. Screening for resistance by disk testing.

Recently, strains of Streptococcus pneumoniae with greatly increased resistance to penicillin (minimal inhibitory concentrations or MICs, 1--8 microgram/ml) were recovered in cultures of blood from patients in South Africa and from one in Minnesota who had serious pneumococcal infections. The authors undertook a study to determine whether these resistant strains have become prevalent in their locale. Between January and July 1978, the laboratories of hospitals serving the greater Madison, Wisconsin, area (population 200,000) contributed 243 pneumococcal isolates for susceptibility testing by an agar dilution technic. Strains with greatly increased resistance (MICs greater than or equal to 1.0 microgram/ml) were not found; only six strains (2.4%) had relative resistance to penicillin (MICs .125--.50 microgram/ml), a range of susceptibility that has been associated with inconsistent clinical responses to treatment with penicillin. Overall, the susceptibility patterns of these 243 isolates are similar to those reported from other centers in North America over the past 30 years. Routine susceptibility testing of pneumococci by hospital laboratories in our area does not appear to be necessary now, but laboratories are advised to screen blood and spinal fluid isolates by a disk-diffusion method. Studies of these 243 isolates and seven South African multiply-resistant strains using a modified Kirby-Bauer technic, showed that a zone of inhibition less than 35 mm around a 10-unit penicillin disk, or better, less than 17 mm around a 1-microgram oxacillin disk, correlates strongly (P less than .001) with resistance to penicillin (MIC greater than or equal to .1 microgram/ml).

Hybrids between Schistosoma haematobium and S. mattheei and their identification by isoelectric focusing of enzymes

Some biological features of F 1 hybrids between South African strains of Schistosoma haematobium and S. mattheei are described and compared with those of both parental species. The distinctive patterns of the G6PD and PGM isoenzymes, resolved by isoelectric focusing, of both species and of the hybrid are defined and the results of enzyme analyses of parasites isolated from human infections in the Transvaal are reported. These show that hybridization does occur naturally in man and that the shape of the eggs produced is not necessarily a guide to the genetic constitution of the enclosed larvae. The experimentally produced F 1 hybrids exhibit heterosis in their increased infectivity to both snails and hamsters, in their more rapid growth and earlier maturation and in the increased daily egg production per female worm when compared with both of the parental species. The possible practical implications of this are discussed.

Antimicrobial susceptibility testing of pneumococci: determination of Kirby-Bauer breakpoints for penicillin G, erythromycin, clindamycin, tetracycline, chloramphenicol, and rifampin.

Antimicrobial susceptibility testing of pneumococci is now essential to monitor for the presence of resistance to agents such as the penicillins, macrolides, lincomycins, chloramphenicol, and tetracycline. In this study, clinical isolates of a selection of resistant South African strains were tested for antimicrobial susceptibility by minimal inhibitory concentration (MIC) determination and by a modified Kirby-Bauer disk diffusion technique, using Mueller-Hinton medium supplemented with 5% horse blood. Disk diffusion breakpoints were determined for penicillin G, erythromycin, clindamycin, tetracycline, chloramphenicol, and rifampin. Reliable results were obtained on disk diffusion for all these agents except for penicillin G. With 6-mug penicillin G disks, zones of strains with intermediate penicillin susceptibility overlapped those of sensitive and resistant strains. With 5-mug methicillin disks, clearer separation of strains based on susceptibility to penicillin G occurred. Strains with zones of <35 mm around penicillin G disks and <25 mm around methicillin disks should have penicillin G MICs determined to confirm their resistance to penicillin G. In view of the potential for pneumococci to be resistant to the agents used in this study, antimicrobial susceptibility of all clinically significant isolates should be determined.

Interconversion of Type C and D Strains of Clostridium botulinum by Specific Bacteriophages

These studies show that Clostridium botulinum types C and D cultures can be cured of their prophages and converted to either type C or D depending on the specific phage used. Strains of types C and D were cured of their prophages and simultaneously ceased to produce their dominant toxins designated as C(1) and D, respectively. Cured nontoxigenic cultures derived from type C strain 162 were sensitive to the phages from the toxigenic type C strain 162 and type D strain South African. When cured nontoxigenic cultures derived from strain 162 were infected with the tox(+) phages from the 162 strain of type C and the South African strain of type D, they then produced toxin neutralized by types C and D antisera, respectively. Cured nontoxigenic cultures isolated from the type D South African strain were only sensitive to the parent phage, and, when reinfected with the tox(+) phage, they produced toxin neutralized by type D antiserum. Type C strain 153 and type D strain 1873, when cured of their respective prophages, also ceased to produce toxins C(1) and D, but, unlike strain 162 and the South African strain, they continued to produce a toxin designated as C(2). When the cured cultures from strains 153 and 1873 were infected with the tox(+) phage from type D strain 1873, the cultures simultaneously produced toxin that was neutralized by type D antiserum. When these cured cultures were infected with the tox(+) phage from type C strain 153, the cultures produced toxin that was neutralized by type C antiserum. These studies with the four strains of C. botulinum confirm that the toxigenicity of types C and D strains requires the continued participation of tox(+) phages. Evidence is presented that types C and D cultures may arise from a common nontoxigenic strain.

Interconversion of type C and D strains of Clostridium botulinum by specific bacteriophages.

These studies show that Clostridium botulinum types C and D cultures can be cured of their prophages and converted to either type C or D depending on the specific phage used. Strains of types C and D were cured of their prophages and simultaneously ceased to produce their dominant toxins designated as C(1) and D, respectively. Cured nontoxigenic cultures derived from type C strain 162 were sensitive to the phages from the toxigenic type C strain 162 and type D strain South African. When cured nontoxigenic cultures derived from strain 162 were infected with the tox(+) phages from the 162 strain of type C and the South African strain of type D, they then produced toxin neutralized by types C and D antisera, respectively. Cured nontoxigenic cultures isolated from the type D South African strain were only sensitive to the parent phage, and, when reinfected with the tox(+) phage, they produced toxin neutralized by type D antiserum. Type C strain 153 and type D strain 1873, when cured of their respective prophages, also ceased to produce toxins C(1) and D, but, unlike strain 162 and the South African strain, they continued to produce a toxin designated as C(2). When the cured cultures from strains 153 and 1873 were infected with the tox(+) phage from type D strain 1873, the cultures simultaneously produced toxin that was neutralized by type D antiserum. When these cured cultures were infected with the tox(+) phage from type C strain 153, the cultures produced toxin that was neutralized by type C antiserum. These studies with the four strains of C. botulinum confirm that the toxigenicity of types C and D strains requires the continued participation of tox(+) phages. Evidence is presented that types C and D cultures may arise from a common nontoxigenic strain.

Studies on the effects of bacteria on experimental schistosome infections in animals.

The initial discovery of the schistosomicidal effects of Klebsiella and E. coli on mice infected with S. mansoni is described. This antischistosomal effect was confined to Gram-negative organisms. The bacteria found to possess this property belonged to the genera: Klebsiella, Escherichia, Salmonella, Arizona, Enterobacter, Serratia and less frequently to the genera Providencia, Achromobacter and Flavobacterium. Of the 24 Gram-positive and 9 other Gram-negative organisms examined none had a lethal effect on the schistosomes. The activity of Klebsiella E.15 and E.10 against strains of S. mansoni from Egypt, Liberia and St. Lucia in mice and hamsters was similar. Klebsiella E.15 was also effective against an Egyptian strain of S. mansoni in monkeys (Erythrocebus patas) and against a South African strain of S. mattheei in mice and hamsters. Activity is confined to mature schistosomes. Immature flukes of 3 weeks or less were unaffected. The rate of activity, the infection and death of the schistosome and the dose response relationship, i.e. the number of organisms given, the route of administration and the resulting kill of schistosomes are described. The precise mode of action on the schistosome is unknown. Living bacteria are essential to produce this effect. Dead bacteria of an active strain and extra-bacterial products were inactive. The effect of antibiotics on the bacterial infection of schistosomes and the interaction between the bacteria and the host are discussed. A freeze-dried preparation of Klebsiella E.10 when ampouled and kept under vacuum at 4°C for 30 months retained its viability and schistosomicidal activity. Some of the problems involved in contemplating the use of bacteria to control schistosomiasis are discussed. The possible inter-relationship between concomittant salmonellosis and schistosomiasis is briefly discussed.

Cyclic development and longevity of Theileria parva in the tick Rhipicephalus appendiculatus

Full details of the technical methods evolved in the study of Theileria parva in the tick Rhipicephalus appendiculatus are described. Stages of T. parva discovered in intestinal contents, gut epithelial cells, and the cells of the salivary glands are fully described. Only the stages of the parasite found in the salivary glands and ducts could definitely be said to be a part of the normal life cycle of T. parva. Multiplication occurred in this site by a process of multiple fission. No sexual stages were discovered. Salivary gland inclusions of T. lawrencei (South African strain) were similar to those of T. parva. The many sources of error that have confused earlier workers are fully discussed. Studies on the longevity of T. parva in its tick vector confirmed the fact that the contained parasites die before the tick. Ticks become noninfective within 34–40 weeks after molting but are capable of feeding up to 15 months. It was found that ticks infected with T. parva could transmit the parasite to cattle 24 hours after being placed on the host.

POLYGENES IN EYE PIGMENT PRODUCTION IN DROSOPHILA SPECIES

During previous investigations on eye pigments in Drosophila species (Nolte, 1958), it was found that wild type South African strains of D. melanogaster and American strains of D. pseudoobscura possessed significantly different amounts of the red and the brown eye pigments, and it was postulated that two series of polygenes were segregating in these populations, and affecting the total amounts of the two pigments, respectively. In order to study this problem further and to test the effect of segregation of these genes, a series of investigations was instituted on outcrossing and inbreeding.

Eye Pigment Relationships in Three Species Groups of Drosophila

The system of pigment cells in the com-pound eyes of Drosophila melanogaster has been described by Nolte (1950), Gersh (1952) and Clayton (1952). In these cells the pigment granules are laid down and they carry deposits of two pigment components, a red and a brown. These pigments can be extracted differentially (Ephrussi and Herold, 1944; Nolte, 1952); the red pigment is extracted in an acidified ethyl alcohol (AEA) and the brown pigment in acidified methyl alcohol (AMA). Quantitative estimations of the two pigments have been made by means of spectrophotometric analysis of the two extracts, and a comparison has been made between the wild type and a large number of mutant strains (Nolte, 1952-1955). In a later study (Nolte, 1958b) the degree of dominance of wild type alleles of various mutant eye-color genes was investigated, and quantitative data were included for two South African strains of D. melanogaster; these were found to differ greatly from the data for the Canton-S strain which had mainly been used as a basis for the quantitative comparison of mutant strains of this species. This fact led to a consideration of the status of this quantitative characteristic in the various species and strains of Drosophila in South Africa. Since it then appeared that these pigment measurements might be useful characters for the analysis of natural populations, and could possibly be of interest in the tracing of phylogenies, the investigation was extended to include foreign species and strains. Since it is possible that the melanogaster and willistoni species groups are directly derived from the obscura group (Sturtevant, 1940), the investigation was extended to include species and mutant strains of these three groups of the subgenus Sophophora.

A note on the heat resistance of a South African strain of Clostridium botulinum type B

AbstractThe z and D values for a South African strain of Clostridium botulinum, type B, have been determined by a method described. The results indicate that the South African strain at least equals in heat resistance the maximum resistance for Clostridium botulinum assumed in process calculations for canned foods.