Development of selective media for Bacillus sphaericus and Bacillus thuringiensis var. israelensis

Summary The resistance of 8 strains of Bacillus sphaericus and of 2 strains of Bacillus thuringiensis var. israelensis (B.t.i.) to various antibiotics and antibiotic combinations were tested. All B. sphaericus strains were resistant to streptomycin, lincomycin and bacitracin, and six strains were resistant to combinations of these antibiotics. This antibiotic resistance could be utilised to establish selective media to identify and follow the fate of B. sphaericus and of B.t.i. in the field.     

Incidence of resistance to chloramphenicol and tetracyclines among 13502Salmonella strains isolated in 1961

Summary The rate of resistance to tetracycline and chloramphenicol amongSalmonella strains isolated in the Netherlands in 1961 was found to be 3.96%, the corresponding figures for 1958/1959 and 1961 being 2.08 and 1.29 respectively. In this country the total number ofSalmonella types found to develop resistance to either tetracycline or chloramphenicol now amounts to 38. Almost 77% of all resistant strains isolated in 1961 were found among the human pathogenS.typhimurium. The relative frequency of resistance in this organism was 8.18%, as compared with 2.50% in 1958/1959 and 1.80% in 1960. In 1961 some cross infections caused by tetracycline resistant strains ofS.typhimurium were observed in man and on one occasion also in a herd of calves. A similar outbreak due to a tetracycline resistant strain ofS.bovis morbificans was seen in a hospital. As almost 87% of all antibiotic resistant strains found in 1961 originated from human patients, the resistance must be largely attributed to the therapeutic use of the drugs in question.     

Growth of dimorphic human pathogenicfungi on media containing cycloheximide and chloramphenicol

Summary Studies of 24 strains ofBlastomyces dermatitidis confirmed previously published results that the yeast-phase of this fungus is more sensitive than the mycelial-phase to cycloheximide and chloramphenicol.Studies of 5 strains each ofHistoplasma capsulatum, Paracoccidioides brasiliensis andSporotrichum schenckii show that that these species also have a similar yeast-phase mycelial -phase sensitivity differential in regard to these antibiotics.A cycloheximide resistant strain ofB. dermatitidis was developed from a sensitive strain.The experimental results support the general practice of using 0.5 mg/ml cycloheximide and 0.05 mg/ml chloramphenicol in media for the isolation of the four fungi at 25° C. The results indicate, however, that some strains would not be recovered at 37° C with similar concentrations of these antibiotics.It is recommended that a concentration of not more than 0.2 mg/ml chloramphenicol should be used to preserve sputum which is subsequently to be cultured forB. dermatitidis, Histoplasma capsulatum, Paracoccidioides brasiliensis orS. schenckii.     

Antibiotic resistance of Shigella in different regions of the USSR]

Shigella were most sensitive to polymyxin ceporin, ampicillin, neomycin and furazolidone and resistant to chloramphenicol, tetracycline and streptomycin. Shigella resistant simultaneously to two or three drugs mainly to tetracycline + chloramphenicol, tetracycline + streptomycin and tetracycline + chloramphenicol + streptomycin were most frequent. The frequency of the Shigella strains carrying R-plasmids increased from 28 per cent in 1969--1970 to 72.6 per cent in 1977. The Shigella strains isolated during the dysentery outbreak in 1973--1977 carried the R-factor controlling resistance to tetracycline + chloramphenicol, tetracycline + chloramphenicol + streptomycin, tetracycline + chloramphenicol + streptomycin + neomycin. Interaction between separate biochemical types, colicinogenicity and drug resistance classes was found in the Shigella isolates. The data on the effect of antibiotic (tetracyclines) intensive use in stock-raising defining wide spread of the R-plasmids controlling resistance to these drugs were obtained.     

Study on Staphylococcus aureus Strain HPC-250 for Associated Antibacterial Property

We isolated a Staphylococcus aureus strain HPC-250 producing antibacterial agent against Paenibacillus strain HPC-251. Both strains were isolated from the same environmental niche. The bacteria were identified using the partial sequencing of their TA-cloned 16S rDNA. Spectrum of the antibacterial agent was also tested against routine observed bacteria with drinking water contamination such as Escherichia coli, Salmonella, Pseudomonas, and Vibrio and these were found to be sensitive. Bacteria like Acinetobacter and Burkholderia were found to be resistant. The differential antibacterial activity of the HPC-250 was observed for the genus Bacillus where B. subtilis remained resistant although B. sphaericus was sensitive.     

Conjugal Transfer of a Toxin-Coding Megaplasmid from Bacillus thuringiensis subsp. israelensis to Mosquitocidal Strains of Bacillus sphaericus

Both Bacillus sphaericus and Bacillus thuringiensis subsp. israelensis produce mosquitocidal toxins during sporulation and are extensively used in the field for control of mosquito populations. All the known toxins of the latter organism are known to be encoded on a large plasmid, pBtoxis. In an attempt to combine the best properties of the two bacteria, an erythromycin resistance-marked pBtoxis plasmid was transferred to B. sphaericus by a mating technique. The resulting transconjugant bacteria were significantly more toxic to Aedes aegypti mosquitoes and were able to overcome resistance to B. sphaericus in a resistant colony of Culex quinquefasciatus, apparently due to the production of Cry11A but not Cry4A or Cry4B. The stability of the plasmid in the B. sphaericus host was moderate during vegetative growth, but segregational instability was observed, which led to substantial rates of plasmid loss during sporulation.     

Survival of orally administered isolated intestinalLactobacillus acidophilus in different parts of gastrointestinal tract of mice

A strain ofLactobacillus acidophilus (Strain HF) was isolated from human faeces. A chloramphenicol resistant strain (HFCm) and a strain (HFCmSm) restant to both chloramphenicol and streptomycin were developed from the isolated strain (HF). All the three strains showed similarin vitro susceptibility against host defence factors like gastric acid, bile salts and volatile as well as non-volatile fatty acids.In vivo tests were done by feeding these strains to mice. When the resistant strains were orally administered along with the antibiotic(s) they were stable up to 72 h     

Selective Medium for Mosquito-Pathogenic Strains of Bacillus sphaericus

A selective medium (BATS), which contains arginine as the sole carbon and nitrogen source and which also contains streptomycin, allowed growth of 18 mosquito-pathogenic strains of Bacillus sphaericus. It inhibited the growth of 68% of the nonpathogenic B. sphaericus strains tested as well as other Bacillus spp. and aquatic bacteria.     

Action of Lincomycin on Staphylococci

On a solid medium, 0.1 to 1 μg/ml of lincomycin hydrochloride had a bacteriostatic effect upon 95 of 100 strains of staphylococci. Using cellophane transfers, we observed a bactericidal effect upon 54 of these strains after 3 to 14 hr of contact with 1 μg/ml. Five staphylococcal strains resistant to 100 μg/ml of lincomycin were also resistant to penicillin G, streptomycin, erythromycin, tetracycline, chloramphenicol (three strains), and rovamycin (three strains). Other staphylococcal strains resistant to methicillin, ampicillin, tetracycline, streptomycin, chloramphenicol, and erythromycin were sensitive to lincomycin.     

Various Levels of Cross-Resistance to Bacillus sphaericus Strains in Culex pipiens (Diptera: Culicidae) Colonies Resistant to B. sphaericus Strain 2362

We studied the cross-resistance to three highly toxic Bacillus sphaericus strains, IAB-59 (serotype H6), IAB-881 (serotype H3), and IAB-872 (serotype H48), of four colonies of the Culex pipiens complex resistant to B. sphaericus 2362 and 1593, both of which are serotype H5a5b strains. Two field-selected highly resistant colonies originating from India (KOCHI, 17,000-fold resistance) and France (SPHAE, 23,000-fold resistance) and a highly resistant laboratory-selected colony from California (GeoR, 36,000-fold resistance) showed strong cross-resistance to strains IAB-881 and IAB-872 but significantly weaker cross-resistance to IAB-59 (3- to 43-fold resistance). In contrast, a laboratory-selected California colony with low-level resistance (JRMM-R, 5-fold resistance) displayed similar levels of resistance (5- to 10-fold) to all of the B. sphaericus strains tested. Thus, among the mosquitocidal strains of B. sphaericus we identified a strain, IAB-59, which was toxic to several Culex colonies that were highly resistant to commercial strains 2362 and 1593. Our analysis also indicated that strain IAB-59 may possess other larvicidal factors. These results could have important implications for the development of resistance management strategies for area-wide mosquito control programs based on the use of B. sphaericus preparations.     

Sensitivity of 56 strains of Streptococcus group B to 12 antibiotics: its determination by dilution and diffusion in agar

Sensitivity of 56 streptococcal strains of group B to 12 antibiotics was studied with the method of dilution in a special solid medium for cultivation of streptococci and with the method of agar diffusion in the same medium. All the strains were found to be sensitive to chloramphenicol, ristomycin and erythromycin. The predominating majority of the strains were sensitive to beta-lactam antibiotics and lincomycin. All the strains were resistant to aminoglycoside antibiotics, such as streptomycin and gentamicin. More than 85 per cent of the strains were resistant to tetracycline. Strains with multiple resistance to 2-7 antibiotics were detected. Satisfactory correlation between the two methods was observed. It was shown to be clinically advisable to determine the sensitivity of streptococci of group B to beta-lactam antibiotics, erythromycin and lincomycin.     

Multiple drug resistance inAerobacter aerogenes

Multiple drug resistance resulting from continued growth in various drugs in turn followed by serial subculture in the simultaneous presence of all the drugs has been studied. In this way strains ofA. aerogenes resistant, in considerable measure, to 3, 4 and 5 drugs have been obtained. Although the final state was always a compromise between the various new properties it was often a good and very effective compromise and for this reason kinetic properties of the cells such as growth rate had to be intensively followed. Tests for presence or absence of growth after a fixed interval of time would not have sufficed. At the 5-drug stage a good compromise was obtained when the agents used were streptomycin, sulphanilamide, chloramphenicol, ampicillin and 5-aminoacridine. When tretamine (triethylene melamine) replaced 5-aminoacridine as the fifth drug, the rate of growth in the various drugs both singly and in admixture was not as good.Continued subculture in sulphanilamide medium led to a condition of hypersensitivity to chloramphenicol. Ampicillin-resistant strains were more inhibited by 2-phenylethanol than the ampicillin-sensitive parent organism. Strains resistant to chloramphenicol were also resistant to tretamine and vice versa. Such cross-resistance did not occur with any of the other drugs at the concentrations used.     

Occurrence of heterogeneous R-plasmids during two concurrent epidemics due to multidrug-resistantSalmonella oranienburg

During two different epidemics that started in August–September, 1980, 140 and 6 multidrug-resistant strains ofSalmonella oranienburg were isolated from a Children's Hospital in New Delhi (epidemic 1) and H.N. Das Hospital in Bombay (epidemic 2) respectively. Out of 140, 123 strains showed high levels of resistance to ampicillin, kanamycin, streptomycin, sulfamethoxazole, tetracycline, and trimethoprim. All six strains of epidemic 2 and two strains of epidemic 1 were also found resistant to chloramphenicol in addition to the above antibiotics. The genetic characterization of conjugative R-plasmids harbored by these strains revealed that, while strains of epidemic 1 carry a fi^–, 96 megadalton, an unclassified plasmid, the strains isolated during epidemic 2 contain a fi^–, 62 megadalton plasmid of incompatibility group I₁. All the six strains of epidemic 2 were found to produce bacteriocin of Col Ib group. Plasmid transfer studies revealed that the genes for antibiotic resistance and bacteriocin production were coded on a single plasmid of 62 megadalton. The data show the significance of detailed genetic analysis while dealing with the clonal outbreaks due to same resistant serotypes.     

Isolation of antibiotic resistance bacterial strains from Eastern Siberia permafrost sediments

A collection of bacterial antibiotic resistance strains isolated from arctic permafrost subsoil sediments of various age and genesis was created. The collection included approximately 100 strains of Gram-positive (Firmicutes, Arthrobacter) and Gram-negative bacteria (Bacteroidetes, γ-Proteobacteria, and α-Proteobacteria) resistant to aminoglycoside antibiotics (gentamicin, kanamycin, and streptomycin), chloramphenicol and tetracycline. Antibiotic resistance spectra were shown to differ in Gram-positive and Gram-negative bacteria. Multidrug resistance strains were found for the first time in ancient bacteria. In studies of the molecular nature of determinants for streptomycin resistance, determinants of the two types were detected: strA-strB genes coding for aminoglycoside phosphotransferases and genes aadA encoding aminoglycoside adenylyltransferases. These genes proved to be highly homologous to those of contemporary bacteria.     

Leishmania tarentolae: streptomycin and chloramphenicol resistance of promastigotes.

Stable mutant strains of Leishmania tarentolae promastigotes resistant to chloramphenicol (CAP) were isolated by replica-plating techniques. In addition, cell lines stress-adapted to streptomycin and to high culture temperature (33 C) were obtained. Drug resistance was influenced by temperature, culture media, and plating technique. Inhibition of amino acid incorporation into protein occurs in CAP-resistant cells when exposed to 600 μg CAP/ml but this inhibition was 50–80% lower than that found in wild type sensitive cells. The primary site of CAP action appears to be inhibition of protein synthesis. CAP also adversely affected proline oxidation.     

Incidence of resistance to tetracycline,chloramphenicol and ampicillin amongSalmonella species isolated in The Netherlands in 1965 and 1966

In 1965 and 1966 respectively 9628 and 10467Salmonella strains were screened for resistance to tetracycline and chloramphenicol. In 1965 the major half of the strains were also tested for ampicillin resistance and in 1966 all strains passed this test. In 1965 20.9% and in 1966 21.7% were found resistant, mainly to tetracycline. The incidence of resistance to chloramphenicol remained low (less than 1%). By far the majority of the resistant strains belonged toS. typhi murium andS. panama. The rate of resistance in the former organism remained nearly equal to that observed in 1963 – 1964, whereas inS. panama it has diminished.Ampicillin-resistant strains were found in as many as 23 different types ofSalmonella. In 8 types in which ampicillin resistance was found, only animal strains were concerned. These findings may represent cases of primary ampicillin resistance. In the human strains ofSalmonella the incidence of ampicillin resistance increased from 11.6% in 1965 to 15.3% in 1966.     

A Strain of Bacillus sphaericus Causes Slower Development of Resistance in Culex quinquefasciatus

Two field-collected Culex quinquefasciatus colonies were subjected to selection pressure by three strains of Bacillus sphaericus, C3-41, 2362, and IAB59, under laboratory conditions. After 13 and 18 generations of exposure to high concentrations of C3-41 and IAB59, a field-collected low-level-resistant colony developed >144,000- and 46.3-fold resistance to strains C3-41 and IAB59, respectively. A field-collected susceptible colony was selected with 2362 and IAB59 for 46 and 12 generations and attained >162,000- and 5.7-fold resistance to the two agents, respectively. The pattern of resistance evolution in mosquitoes depended on continuous selection pressure, and the stronger the selection pressure, the more quickly resistance developed. The resistant colonies obtained after selection with B. sphaericus C3-41 and 2362 showed very high levels of cross-resistance to B. sphaericus 2362 and C3-41, respectively, but they displayed only low-level cross-resistance to IAB59. On the other hand, the IAB59-selected colonies had high cross-resistance to both strains C3-41 and 2362. Additionally, the slower evolution of resistance against strain IAB59 may be explained by the presence of another larvicidal factor. This is in agreement with the nontoxicity of the cloned and purified binary toxin (Bin1) of IAB59 for 2362-resistant larvae. We also verified that all the B. sphaericus-selected colonies showed no cross-resistance to Bacillus thuringiensis subsp. israelensis, suggesting that it would be a promising alternative in managing resistance to B. sphaericus in C. quinquefasciatus larvae.     

Phenotypic suppression by streptomycin of amber mutants in the ribonucleic Acid bacteriophage coat protein cistron

After mutagenesis with nitrosoguanidine or ultraviolet light, 298 streptomycin high-resistant and 98 streptomycin high-dependent mutants were isolated from HfrC Su⁻. They were tested for their ability to phenotypically suppress five different amber ribonucleic acid (RNA) bacteriophage mutants in the presence of streptomycin. The phage mutants are all in the coat protein, which is 129 amino acids long; the uracil-adenine-guanine codons were at the following positions: sus3 and amB2, 6; amB11, 50; amB21, 54; sus11, 70. Only sus3 and amB2 could be phenotypically suppressed by streptomycin; this was clearly demonstrated in nine mutant strains, seven str-HR and two str-HD. The suppression was always dependent upon added streptomycin and was dose-dependent in all cases. None of the mutants showed measurable suppression in absence of the drug. Among revertants to streptomycin independence from streptomycin-dependent strains that could show phenotypic suppression, most of those that were still resistant to streptomycin (10 μg or more) retained the capacity to show phenotypic suppression; whereas among those revertants sensitive to 10 μg of streptomycin or less, none retained the capacity. Eight different amber polar mutants (strong and weak) in gene 34 of phage T4 were also tested for pleiotypic suppression by streptomycin in all the streptomycin-resistant and -dependent strains isolated. No suppression was found in any of the 396 strains tested.     

Induced variations in microorganisms

Genetic transfer of markers has been found to occur through a sexual cycle between intact cells in a mixed culture of two different genotypes. When rS (radiosensitive and streptomycin resistant) and Rs (radioresistant and streptomycin sensitive) cells ofRhizobium trifolii were brought into varying periods of cellular contact, the cell mixture yielded RS recombinants. The recombination frequency was higher in young mating cultures than in aged ones. Except for the acquisition of the R and S markers, the recombinants did not differ appreciably from the parental strains in other characters.