Skin Commensal Staphylococci May Act as Reservoir for Fusidic Acid Resistance Genes

We analyzed the occurrence and mechanisms of fusidic acid resistance present in staphylococci isolated from 59 healthy volunteers. The fingers of the volunteers were screened for the presence of staphylococci, and the collected isolates were tested for resistance to fusidic acid. A total of 34 fusidic acid resistant staphylococcal strains (all were coagulase-negative) were isolated from 22 individuals (22/59, 37.3%). Examination of the resistance genes revealed that acquired fusB or fusC was present in Staphylococcus epidermidis, Staphylococcus capitis subsp. urealyticus, Staphylococcus hominis subsp. hominis, Staphylococcus warneri and Staphylococcus haemolyticus. Resistance islands (RIs) carrying fusB were found in S. epidermidis and S. capitis subsp. urealyticus, while staphylococcal chromosome cassette (SCC)-related structures harboring fusC were found in S. hominis subsp. hominis. Genotypic analysis of S. epidermidis and S. hominis subsp. hominis indicated that the fus elements were disseminated in diverse genetic strain backgrounds. The fusC elements in S. hominis subsp. hominis strains were highly homologous to SCCfusC in the epidemic sequence type (ST) 239/SCCmecIII methicillin-resistant S. aureus (MRSA) or the pseudo SCCmec in ST779 MRSA. The presence of acquired fusidic acid resistance genes and their genetic environment in commensal staphylococci suggested that the skin commensal staphylococci may act as reservoir for fusidic acid resistance genes.     

Identification of Staphylococci Isolated from Clinical Material

A total of 350 staphylococci isolated from various clinical sources were examined for bound and free coagulase, fermentation of mannitol, and deoxyribonuclease. The economical coagulase-mannitol-agar method of Esber and Faulcomer was found to be suitable for the detection of free coagulase and mannitol fermentation. A significant number of coagulase- and mannitol-negative staphylococci proved to be deoxyribonuclease-positive.     

Characterization of Staphylococci Isolated from Raw Milk

To evaluate the pathogenicity of staphylococci from bovine raw milk, the general characteristics of 775 strains isolated from 798 samples of milk were studied. The coagulase test was performed by use of rabbit plasma. Chromogenesis, mannitol fermentation, and gelatin liquefaction were investigated on Chapman's Medium 110, after 48 hr of incubation. Production of β-hemolysin, which has been considered indicative of pathogenic staphylococci of animal origin, was determined by streaking different strains on sheep blood-agar plates in the presence of a strain of Lancefield group B streptococci. Plates were incubated at 37 C for 24 hr, and strong hemolysis was produced in the zone of interaction of β-hemolysin and some substance liberated by streptococcus (CAMP test). Of 404 strains found to be coagulase-positive, 95.8% exhibited a deep-orange pigment, 76.5% produced β-hemolysin, 91.8% fermented mannitol, and 75% liquefield gelatin. Of 371 strains which gave a negative coagulase test, about 16% fermented mannitol and liquefied gelatin; none of these strains produced β-hemolysin. When results are grouped according to pigmentation and coagulase production, β-hemolysin seems to be developed by pathogenic strains of Staphylococcus aureus only. If suitability of these tests for investigation of pathogenicity is compared, production of β-hemolysin appears to be the most useful one, since no “false positive” results were found. The use of the CAMP test as a simple and rapid technique to determine production of β-hemolysin by pathogenic strains of animal staphylococci during routine bacteriological work is suggested.     

High Prevalence of Multidrug-Tolerant Bacteria and Associated Antimicrobial Resistance Genes Isolated from Ornamental Fish and Their Carriage Water

Background

Antimicrobials are used to directly control bacterial infections in pet (ornamental) fish and are routinely added to the water these fish are shipped in to suppress the growth of potential pathogens during transport.

Methodology/Principal Findings

To assess the potential effects of this sustained selection pressure, 127 Aeromonas spp. isolated from warm and cold water ornamental fish species were screened for tolerance to 34 antimicrobials. Representative isolates were also examined for the presence of 54 resistance genes by a combination of miniaturized microarray and conventional PCR. Forty-seven of 94 Aeromonas spp. isolates recovered from tropical ornamental fish and their carriage water were tolerant to ≥15 antibiotics, representing seven or more different classes of antimicrobial. The quinolone and fluoroquinolone resistance gene, qnrS2, was detected at high frequency (37% tested recent isolates were positive by PCR). Class 1 integrons, IncA/C broad host range plasmids and a range of other antibiotic resistance genes, including floR, bla _TEM−1, tet(A), tet(D), tet(E), qacE2, sul1, and a number of different dihydrofolate reductase and aminoglycoside transferase coding genes were also detected in carriage water samples and bacterial isolates.

Conclusions

These data suggest that ornamental fish and their carriage water act as a reservoir for both multi-resistant bacteria and resistance genes.     

The Vitamin Requirements of Staphylococci Isolated from Human Skin

The vitamin requirements of 46 strains representing nine species of Staphylococcus isolated from human skin together with nine authentic reference strains of these species were determined using a chemically defined medium. Strains of Staphylococcus saprophyticus and Staphylococcus cohnii were isolated on selective media. All the strains investigated required nicotinic acid and thiamine for growth. Biotin was essential or stimulatory for all coagulase negative strains except one strain of Staphylococcus capitis. Oleic acid substituted for biotin in all cases except with one strain of Staphylococcus haemolyticus. No species pattern of biotin requirement or of the ability of oleic acid to substitute for biotin was apparent. Five out of six strains of Staph. cohnii required pantothenic acid.     

Permeability of Isolated Infected Cells from Soybean Nodules

Infected cells from the central zone of mature soybean noduleswere isolated by incubating nodule slices with hydrolytic enzymes,and their permeability to various organic compounds measured.The cells were effectively impermeable to sucrose, fructoseand glucose, readily permeable to malate and succinate and partiallypermeable to glutamate and glutamine. Malate uptake showed saturationkinetics and was competitively inhibited by succinate. Malateuptake was also inhibited by a protonophore and the respiratorypoison antimycin, as well as by butylmalonate, cyanocinnamicacid and other substrate analogues, but not by phthalonate.We conclude that there is a dicarboxylate carrier on the plasmamembraneof soybean infected cells, which catalyses the uptake of malateand which depends on mitochondrial ATP for maximum rates.     

Trypanosoma theileri Associated with T-lymphocytes Isolated from a Latently Infected Cow

. Trypanosoma theileri infection, latent in a mature Hereford cow, could not be demonstrated in routine blood smears or cultures. Throughout the 2-year period an intravenous injection of dexamethasone consistently produced parasitaemia which was detectable in peripheral blood mononuclear cell (PBMC) cultures. Fractionation techniques such as plastic adherence and Sephadex-G10 fractionation, designed to deplete monocytes and enrich T-lymphocytes, increased trypanosome-positive cultures from 25 to 100%. Removal of B-lymphocytes from Sephadex, non-adherent (SE-NA) cells did not reduce the percentage of positive cultures. Light and transmission electron microscopy of SE-NA PBMC cultured for 36 or 45 h revealed numerous trypanosomes and widespread T-lym-phocyte destruction. No trypanosomes were observed in 12-h cultures. A close association, either extra- or intracellular, of a parasitic stage of T. theileri with T-lymphocytcs is inferred.     

Diterpene Constituents of Euphorbia exigua L. and Multidrug Resistance Reversing Activity of the Isolated Diterpenes

Phytochemical investigation of the MeOH extract obtained from the aerial parts of the annual weed Euphorbia exigua L. resulted in the isolation of two novel ( 1, 2 ) and one known ( 3 ) jatrophane diterpenes. Their structures were established by extensive 1D‐ and 2D‐NMR spectroscopy and HR‐ESI‐MS. The isolated compounds were evaluated for multidrug resistance (MDR) reversing activity on human MDR gene‐transfected L5178 mouse lymphoma cells; and all three compounds were found to modulate the intracellular drug accumulation.     

The Ocean as a Global Reservoir of Antibiotic Resistance Genes

Recent studies of natural environments have revealed vast genetic reservoirs of antibiotic resistance (AR) genes. Soil bacteria and human pathogens share AR genes, and AR genes have been discovered in a variety of habitats. However, there is little knowledge about the presence and diversity of AR genes in marine environments and which organisms host AR genes. To address this, we identified the diversity of genes conferring resistance to ampicillin, tetracycline, nitrofurantoin, and sulfadimethoxine in diverse marine environments using functional metagenomics (the cloning and screening of random DNA fragments). Marine environments were host to a diversity of AR-conferring genes. Antibiotic-resistant clones were found at all sites, with 28% of the genes identified as known AR genes (encoding beta-lactamases, bicyclomycin resistance pumps, etc.). However, the majority of AR genes were not previously classified as such but had products similar to proteins such as transport pumps, oxidoreductases, and hydrolases. Furthermore, 44% of the genes conferring antibiotic resistance were found in abundant marine taxa (e.g., Pelagibacter, Prochlorococcus, and Vibrio). Therefore, we uncovered a previously unknown diversity of genes that conferred an AR phenotype among marine environments, which makes the ocean a global reservoir of both clinically relevant and potentially novel AR genes.     

Thermal Resistance of Salmonellae and Staphylococci in Foods

The heat-resistant Salmonella senftenberg 775W and two strains of Staphylococcus aureus were tested at temperatures up to 68.3 C (71.1 C for S. senftenberg) in four different media. From the survival data, decimal reduction times (D values) were calculated for each set of conditions, and decimal reduction time curves were constructed for each bacterial strain in each medium. Slopes of decimal reduction time curves (Z(D)) ranged from 4.52 to 6.38 C with a single exception. There was no statistical heterogeneity among the remaining values. Results were in close agreement with published results of similar studies conducted at somewhat lower temperatures and support the practice of using a slope value (Z(D)) of 5.56 C for establishing time-temperature relationships for food processing. It is recommended that such a decimal reduction time curve not be extrapolated to temperatures more than 5.56 C higher than those actually tested.