The prevalence and cotrimoxazole susceptibility of Streptococcus pneumoniae isolated from sputum of 100 HIV-positive patients attending the Nigeria Institute of Medical Research clinic was investigated using standard microbiological methods. Eleven of the sputum specimens grew Streptococcus pneumoniae. Antimicrobial susceptibility test showed that all the isolates were sensitive to amoxicillin, augmentin, erythromycin and chloramphenicol but were resistant to cotrimoxazole. Continuous surveillance of S pneumoniae in sputum samples of HIV-positive subjects in this environment is necessary in order to regulate treatment regimen, considering that cotrimoxazole is the drug recommended by WHO for respiratory infections in HIV patients.
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Unhygienic poultry feedstuffs can lead to nutrient losses and detrimental effect on poultry production and public health. In the present study, mycobiota and colony-forming units per gram in ingredients and finish poultry feed was evaluated with special reference to potentially mycotoxigenic fungi.
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Avian infectious bronchitis virus (IBV) defective RNAs (D-RNAs) have been used for the expression of heterologous genes in a helper-virus-dependent expression system. The heterologous genes were expressed under the control of an IBV transcription-associated sequence (TAS) derived from gene 5 of IBV Beaudette. However, coronavirus D-RNA expression vectors display an inherent instability following serial passage with helper virus, resulting in the eventual loss of the heterologous genes. The use of the picornavirus encephalomyocarditis virus (EMCV) internal ribosome entry site (IRES) sequence to initiate gene translation was investigated as an alternative method to the coronavirus-mediated TAS-controlled heterologous gene expression system. IBV D-RNAs containing the chloramphenicol acetyltransferase (CAT) reporter gene, under EMCV IRES control, were assessed for IRES-mediated CAT protein translation. CAT protein was detected from T7-derived IBV D-RNA transcripts in a cell-free protein synthesis system and in situ in avian chick kidney (CK) cells following T7-derived D-RNA synthesis from a recombinant fowlpox virus expressing the bacteriophage T7 DNA-dependent RNA polymerase. However, CAT protein was not detected in CK cells from IRES-containing IBV D-RNAs, in which the IRES-CAT construct was inserted at two different positions within the D-RNA, in the presence of helper IBV. Northern blot analysis demonstrated that the IRES-containing D-RNAs were not rescued on serial passage with helper virus, indicating that the EMCV IRES sequence had a detrimental effect on IBV D-RNA rescue.
The aim of this study was to investigate the phenotypic and genotypic antibiotic resistance profiles of pseudomonads isolated from surfaces of a goat and lamb slaughterhouse, which were representative of areas that are possible sources of meat contamination. Mesophilic (85 isolates) and psychrotrophic (37 isolates) pseudomonads identified at the species level generally were resistant to sulfamethoxazole, erythromycin, amoxicillin, ampicillin, chloramphenicol, trimethoprim, rifampin, and ceftazidime (especially mesophiles), as well as colistin and tetracycline (especially psychrotrophes). However, they generally were sensitive to ciprofloxacin, gentamicin, imipenem, and kanamycin regardless of species identity. Worryingly, in the present study, we found multidrug resistance (MDR) to up to 13 antibiotics, which was related to intrinsic and acquired resistance mechanisms. Furthermore, a link between various antimicrobial resistance genes was shown for beta-lactams and tetracycline, trimethoprim, and sulfonamides. The distribution and resistome-based analysis of MDR pseudomonads in different slaughterhouse zones indicated that the main sources of the identical or related pseudomonad strains were the animals (feet and wool) and the slaughterhouse environment, being disseminated from the beginning, or entrance environment, to the environment of the finished meat products. Those facts must be taken into consideration to avoid cross-contamination with the subsequent flow of mobile resistance determinants throughout all slaughterhouse zones and then to humans and the environment by the application of adequate practices of hygiene and disinfection measures, including those for animal wool and feet and also the entrance environment.
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This study demonstrates the outbreak of typhoid fever occurs through asymptomatic carrier. In addition, this study also reveals the occurrence of considerable drug resistant among the S. typhi isolates.
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Bacteriological analyses of seawater from three main beaches in Fortaleza, Brazil were performed during 1997. Thirty-six samples per beach were collected for a total of 108 samples. For Meireles Beach, 44% of the samples had MPN total coliforms values of at least 1100 or over 2400/100 ml, followed by Formosa and Diários beaches showing lower counts. For fecal coliforms the highest numbers were demonstrated for Formosa, followed by Meireles and Diários beaches in this descending order: 13.0%, 11.1% and 8.3%, respectively. Escherichia coli strains were identified in 76.8% of the 108 samples. Among 295 strains of E. coli, 21 belonged to serogroups O25, O26, O91, O112, O119, O158 and O164. Strains from serogroup O26 were tested using PCR, ELISA and Vero cells to detect Verotoxins VT1 and VT2 and all strains were negative. No LT and ST, as determined by ELISA and suckling mice assays, were detected among the 295 strains. All strains of E. coli were sensitive to ampicillin, cephalothin, gentamicin, tetracycline, sulfametox-trimethoprim, chloramphenicol and ciprofloxacin. Although the E. coli strains were not toxigenic, their presence in high numbers could be of public health significance.
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This study suggests that it is possible to induce thermotolerance in biocontrol yeasts such as C. sake. However, this does not improve survival of cells exposed to spray-drying sufficiently to consider this a suitable formulation method for this biocontrol agent. HSPs, sugars and sugar polyols were not directly responsible for induced thermotolerance in yeast cells.
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This study is to present in-depth evaluations of in vitro antimicrobial activities of four natural coumarins 5-geranyloxy-7-methoxycoumarin (Gm, 1), (5,7-dimethoxy-8-prenyloxycoumarin (artanin, Ar, 2)), isopimpinellin (Is, 3) and phellopterin (Ph, 4) from Zanthoxylum nitidum (Roxb.) DC. (Rutaceae) extracts, focusing on their potential restoration the activity of conventional antibacterial agents against clinical MRSA strains.
Nonribosomal peptide synthetases (NRPSs) are large, multidomain enzymes that biosynthesize medically important natural products. We report the crystal structure of the free-standing NRPS condensation (C) domain VibH, which catalyzes amide bond formation in the synthesis of vibriobactin, a Vibrio cholerae siderophore. Despite low sequence identity, NRPS condensation enzymes are structurally related to chloramphenicol acetyltransferase (CAT) and dihydrolipoamide acyltransferases. However, although the latter enzymes are homotrimers, VibH is a monomeric pseudodimer. The VibH structure is representative of both NRPS condensation and epimerization domains, as well as the condensation-variant cyclization domains, which are all expected to be monomers. Surprisingly, despite favorable positioning in the active site, a universally conserved histidine important in CAT and in other C domains is not critical for general base catalysis in VibH.
Outbreak 1 peaked in February 1997; 31 patients were confirmed by culture as having Salmonella Typhimurium var Copenhagen infection, isolates of which showed indistinguishable pulsed-field gel electrophoresis (PFGE) patterns. The outbreak strain was phage type DT104 with the 5-drug resistance pattern. Sixteen cases and 25 controls were enrolled in the case-control study; 15 of 16 Salmonella Typhimurium var Copenhagen cases compared with 14 of 24 matched controls reported eating unpasteurized Mexican-style cheese, (matched odds ratio, 7.9; 95% confidence interval, 1.1-354.9). Enhanced surveillance uncovered outbreak 2, which peaked in April 1997 and was caused by a non-Copenhagen variant of Salmonella Typhimurium. During outbreak 2, Salmonella Typhimurium was isolated from 79 persons who ate fresh Mexican-style cheese from street vendors and from cheese samples and raw milk. The PFGE pattern of the milk isolate matched 1 of the 3 patterns recovered from patients; all strains were phage type DT104b with the 5-drug resistance pattern.
A 58-year-old woman presented to eye emergency with a chronic conjunctivitis which was diagnosed by laboratory microbiological testing to be due to the environmental pathogen Raoultella planticola. The organism was sensitive to Chloramphenicol and the patient made a rapid recovery on these drops. This is the first report of this organism infecting the eye.
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The aim of the study was to test the hypothesis that the efflux pump inhibitors (EPIs) 1-(1-naphthylmethyl)-piperazine (NMP) and phenyl-arginine-beta-naphthylamide (PAbetaN) can inhibit the Vibrio cholerae resistance-nodulation-division (RND) family efflux systems, and thereby render V. cholerae susceptible to antimicrobial agents and inhibit the production of the virulence factors cholera toxin (CT) and the toxin coregulated pilus (TCP).
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Resurgence of serogroup A and recent increase in ST-11 among serogroup W135 isolates were worrying when considered with the epidemic wave of serogroup A meningitis, which affected neighboring countries and the serogroup W135 epidemic in Niger in 2009-2010.
FEJ was clearly confirmed to promote the recovery of bone marrow hemopoietic function in a myelosuppressed mouse model, which may be attributed to (i) improving bone marrow hematopoietic microenvironment; (ii) facilitating the cell proliferation and preventing BMNC from apoptosis; (iii) stimulating the expressions of IL-1β, IL-3, IL-6, SCF and GM-CSF and inhibiting the expression of TGF-β.
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When challenged by the dietary soybean cysteine protease inhibitor scN, the cowpea bruchid (Callosobruchus maculatus) adapts to the inhibitory effects by readjusting the transcriptome of its digestive system, including the specific activation of a cathepsin B-like cysteine protease CmCatB. To understand the transcriptional regulation of CmCatB, we cloned a portion of its promoter and demonstrated its activity in Drosophila cells using a chloramphenicol acetyltransferase reporter system. EMSAs detected differential DNA-binding activity between nuclear extracts of scN-adapted and -unadapted midguts. Two tandem chicken ovalbumin upstream promoter (COUP) elements were identified in the CmCatB promoter that specifically interacted with a protein factor unique to nuclear extracts of unadapted insect guts, where CmCatB expression was repressed. Seven-up (Svp) is a COUP-TF-related transcription factor that interacted with the COUP responsive element. Polyclonal anti-(mosquito Svp) serum abolished the specific DNA-binding activity in cowpea bruchid midgut extracts, suggesting that the protein factor is an Svp homolog. Subsequent cloning of a cowpea bruchid Svp (CmSvp) indicated that it shares a high degree of amino acid sequence similarity with COUP-TF/Svp orphan nuclear receptor family members from varied species. The protein was more abundant in scN-unadapted insect guts than scN-adapted guts, consistent with the observed DNA-binding activity. Furthermore, CmCatB expression was repressed when CmSvp was transiently expressed in Drosophila cells, most likely through COUP binding. These findings indicate that CmSvp may contribute to insect counter-defense, in part by inhibiting CmCatB expression under normal growth conditions, but releasing the inhibition when insects are challenged by dietary protease inhibitors.
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Islet-specific glucose-6-phosphatase catalytic subunit-related protein (IGRP) is selectively expressed in islet beta cells and is a major autoantigen in a mouse model of type I diabetes. The analysis of IGRP-chloramphenicol acetyltransferase (CAT) fusion gene expression through transient transfection of islet-derived betaTC-3 cells revealed that a promoter region, located between -273 and -254, is essential for high IGRP-CAT fusion gene expression. The sequence of this promoter region does not match that for any known islet-enriched transcription factor. However, data derived from gel retardation assays, a modified ligation-mediated polymerase chain reaction in situ footprinting technique and a SDS-polyacrylamide separation/renaturation procedure led to the hypothesis that this protein might be Pax-6, a conclusion that was confirmed by gel supershift assays. Additional experiments revealed a second non-consensus Pax-6 binding site in the -306/-274 IGRP promoter region. Pax-6 binding to these elements is unusual in that it appears to require both its homeo and paired domains. Interestingly, loss of Pax-6 binding to the -273/ -246 element is compensated by Pax-6 binding to the -306/-274 element and vice versa. Gel retardation assays revealed that another islet-enriched transcription factor, namely Pdx-1, binds four non-consensus elements in the IGRP promoter. However, mutation of these elements has little effect on IGRP fusion gene expression. Although chromatin immunoprecipitation assays show that both Pax-6 and Pdx-1 bind to the IGRP promoter within intact cells, in contrast to the critical role of these factors in beta cell-specific insulin gene expression, IGRP gene transcription appears to require Pax-6 but not Pdx-1.
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Thirty promoter-carrying fragments were isolated. Three C. glutamicum clones harboring pAKC6 with promoter fragments displayed chloramphenicol acetyltransferase (CAT) activity of more than 24 U/mg. The fragment F57 led to the highest CAT activity of 32.50 U/mg, even more than that produced by the promoter Ptrc, 26.33 U/mg.
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Blood samples were cultured for bacterial isolates and identified by standard procedures. Susceptibility testing was performed according to the National Committee for Clinical Laboratory Standards. Plasmid DNA extraction was performed using the alkaline lysis method with Escherichia coli v517 used as the standard. Conjugation and transformation experiments were performed using standard methods. For the latter, E. coli K12 HB 101 (ara-14, galK2, hsd 520, lacyl, leu, mtl-1, Pro A2, rec A13, rps L20, sup E44, thii xyl-5) was used as the recipient and plasmid PBR 322 was used as the positive control.
Surfactant protein A (SP-A) is a member of the collectin family of innate host defense molecules expressed primarily in respiratory epithelial cells of the lung. SP-A concentrations are influenced by both cell-specific and ubiquitous nuclear proteins that regulate SP-A gene transcription in a cell-selective and temporally regulated manner. In this work, a consensus GATA-binding site (GBS) was identified at positions -69 to -64 of the mouse SP-A gene. The transcriptional activity of wild-type SP-A reporter constructs in HeLa cells was increased 5-10-fold when cotransfected with a GATA-6 expression plasmid. Deletion of the GBS completely blocked transactivation by GATA-6. Transfection of a construct expressing GATA-6-engrailed fusion protein inhibited basal expression of the SP-A/chloramphenicol acetyltransferase construct in MLE-15 cells. Nuclear extract proteins from MLE-15 cells bound to the GBS in the mouse SP-A gene, and a supershifted band was detected with a GATA-6-specific antibody. Transactivation of the wild-type SP-A constructs by GATA-6 increased transcriptional activity 7-10-fold, whereas thyroid transcription factor-1 (TTF-1) increased the activity of these constructs 12-18-fold. The effects of cotransactivating with both GATA-6 and TTF-1 expression constructs were additive. However, mutation of the TTF-1-binding sites alone or in combination decreased GATA-6 transactivation. Likewise, mutation of the GBS blocked TTF-1 activation of the SP-A promoter. In situ hybridization demonstrated GATA-6 mRNA in the peripheral epithelial cells of fetal mouse lung, consistent with the sites of SP-A expression. GATA-6 is expressed in respiratory epithelial cells and binds to a cis-acting element in the SP-A gene promoter, activating the transcriptional activity of the gene.
Mathematical models of the transfer of charged macromolecules have been constructed on the basis of the classical equations of electromigration diffusion of Helmholtz-Smolukhovskii, Goldman, and Goldman-Hodgkin-Katz. It was shown that ion transfer in placental (mimicking lipid-protein barriers) and muscle barriers occurs by different mechanisms. In placental barriers, the electromigration diffusion occurs along lipid-protein channels formed due to the conformational deformation of phospholipid and protein molecules with the coefficients of diffusion D = (2.6-3.6) x 10(-8) cm2/s. The transfer in muscle barriers is due to the migration across charged interfibrillar channels with the negative diffusion activation energy, which is explained by changes in the structure of muscle fibers and expenditures of thermal energy for the extrusion of Cl- from channel walls with the diffusion coefficient D = (6.0-10.0) x 10(-6) cm2/s.
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Studies were carried out to investigate the incidence of multiple antibiotic resistance among E .coli (total 152) isolated from poultry in Karachi to eight commonly used antibiotics: ampicillin (A), chloramphenicol (C), gentamycin (G), anamycin (K), neomycin (N), polymyxin B (P), streptomycin (S) and tetracycline (T) at the levels of 50 microg/ml, 100 microg/ml and 500 microg/ml. Tables of the results are given, showing the number of resistant strains of different patterns of antibiotic resistance at different levels. A comparison of antibiotic resistance to different number of antibiotics and the frequency of resistance to individual antibiotic at different levels is also reported. The highest frequency of resistance was against tetracycline whereas the lowest frequency of resistance was against gentamycin. Thirty R plasmids were isolated from the resistant strains and will be reported elsewhere.
This observational study was conducted at Dr. Essa's Laboratory over a period of 12 months ending in March 2012. Two hundred samples taken from conjunctiva of patients with conjunctivitis were cultured on routine medium and the antibiograms of bacterial isolates were determined by Kirby- Bauer disc diffusion method.
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While plasmids are very commonly associated with the majority of the lactic acid bacteria, they are only very rarely associated with Lactobacillus delbrueckii, with only four characterized to date. In this study, the complete sequence of a native plasmid, pDOJ1, from a strain of Lactobacillus delbrueckii subsp. bulgaricus was determined. It consisted of a circular DNA molecule of 6,220 bp with a G+C content of 44.6% and a characteristic ori and encoded six open reading frames (ORFs), of which functions could be predicted for three-a mobilization (Mob) protein, a transposase, and a fused primase-helicase replication protein. Comparative analysis of pDOJ1 and the other available L. delbrueckii plasmids (pLBB1, pJBL2, pN42, and pLL1212) revealed a very similar organization and amino acid identities between 85 and 98% for the putative proteins of all six predicted ORFs from pDOJ1, reflecting a common origin for L. delbrueckii plasmids. Analysis of the fused primase-helicase replication gene found a similar fused organization only in the theta replicating group B plasmids from Streptococcus thermophilus. This observation and the ability of the replicon to function in S. thermophilus support the idea that the origin of plasmids in L. delbrueckii was likely from S. thermophilus. This may reflect the close association of these two species in dairy fermentations, particularly yogurt production. As no vector based on plasmid replicons from L. delbrueckii has previously been constructed, an Escherichia coli-L. delbrueckii shuttle cloning vector, pDOJ4, was constructed from pDOJ1, the p15A ori, the chloramphenicol resistance gene of pCI372, and the lacZ polylinker from pUC18. This cloning vector was successfully introduced into E. coli, L. delbrueckii subsp. bulgaricus, S. thermophilus, and Lactococcus lactis. This shuttle cloning vector provides a new tool for molecular analysis of Lactobacillus delbrueckii and other lactic acid bacteria.
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Cultural characters, biochemical tests, antibiotic sensitivity test (disc diffusion), agarose gel electrophoresis, and conjugation protocols were done. Thirty five stool samples were collected from the suspected food handlers for the study.
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The aim of this study is to characterize the epidemiological features of typhoid fever, categorized as class 1 notifiable disease in Korea and to analyze the recent change of antimicrobial resistance of Salmonella enterica serotype Typhi isolated nationwide. We retrospectively analyzed the 1,692 culture-proven cases from 1992 to 2000, using the data of the Korean National Institute of Health. The overall incidence of culture-proven typhoid fever was 0.41 per 100,000 population. It occurred all over the country, but the southeastern part of Korean peninsula had the higher incidence rate than other areas. There were several outbreaks suspected, of which two outbreaks were confirmed. The resistance rate against chloramphenicol showed mild increase, but the ampicillin, trimethoprim/sulfamethoxazole, kanamycin, or nalidixic acid resistance remained at the similar levels for the past 9 yr. There were 21 (1.3%) multidrug-resistant (MDR) strains isolated since 1992, and the number of those has increased. Two strains resistant to ciprofloxacin were first identified in Korea.