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In this paper it is shown a process to demarcate areas with analogous wind conditions. For this purpose a dispersion graph between wind directions will be traced for all stations placed in the studied zone. These distributions will be compared among themselves using the hierarchical clustering algorithm. This information will be used to build a matrix, letting us work with all relations simultaneously. By permutation of elements in this matrix it is possible to group relationed stations.

In this paper several architectures of neural network multilayer artificial are used to forecast mean daily wind speed at a target station. For this purpose, meteorological variables of reference stations are chosen as exogenous inputs. The mean daily wind speed and direction of 88 measuring stations, from 2005 to 2008 were used. From these stations we will identify zones with similar wind patterns to a target station, using a method based on clustering techniques. Data of target station were acquired from a unit located in Southern Andalusia (Penaflor, Sevilla), with a soft orography (10 minutes between measurements). The network inputs are basically historical values of the predicted variable as well as a number of support variables. Different feed-forward models and Radial Basis networks have been elected with the aim of carrying out the treatment of the data. These models are compared to the persistence model to select which prediction models are better than persistence model in a short-time prediction.

The susceptibility phenotypes of 964 clinical isolates of Mycobacterium tuberculosis were studied over a 7-year period in Seville, Spain. Thirty-eight (3.9%) strains were rifampin (RMP) resistant, 79 (8.2%) were isoniazid (INH) resistant and 22 (2.3%) were resistant to at least both antimicrobials (multidrug-resistant, MDR). We studied the mechanisms of resistance to these drugs in 94 resistant clinical isolates of M. tuberculosis using three molecular methods: 1) PCR-single strand conformation polymorphism (SSCP) analysis, 2) RFLP analysis using B1/B2 primers, and 3) sequence analysis. Five different mutations were detected in the rpoB gene: Ser531--Leu (72.3%), His526--Asp (12.8%), Asn518--Ser (2.1%), Gln513--Leu (2.1%) and a nine-nucleotide deletion (2.1%). In the case of resistance to INH, four different mutations in the katG gene were detected, Ser315--Thr (58.0%), Ser315--Leu (2.9%), partial deletion (5.8%) and Ile304--Val (1.4%), while in the inhA regulatory region the only mutation was the nucleotide substitution C209T (4.3%). No mutation was found in the ahpC promoter.

Characterization and frequency of the rpoB gene mutations associated with rifampin resistance in Mycobacterium tuberculosis clinical isolates in Sevilla.Characterization of rpoB mutations in 21 rifampicin-resistant strains of M. tuberculosis isolated during a three-year period (1994-1996) by three different molecular methods: a nonradioactive Single-strand conformation polymorphism (SSCP) analysis, DNA sequence analysis and a commercial method the line probe assay InnoLiPA.Five distinct rpoB mutations were identified. Ser531--Leu mutation was detected in 14 strains (66.7%), H526--Asp in 3 strains (14.3%), Ans512--Ser in 1 strain (4.8%), Glu513--Leu in 1 strain (4.8%). A nine nucleotide deletion (codon 510-513) was found in one strain (4.8%) while in the remaining resistant strain (4.8%) no mutation was detected.The frequency of the different mutations found in the rpoB gene, associated with rifampicin resistance in Mycobacterium tuberculosis clinical isolates in Seville, are similar to those previously reported. However, two new mutations has been detected: a nine nucleotide deletion (codon 510-513), and the Asn512--Ser point mutation. The characterization of the mutations in the rpoB gene could serve as epidemiological marker for the rifampicin resistant clinical isolates of M. tuberculosis.

The paper summarizes at a subspecific level 11 clinical strains of K. pneumoniae. The objective of the work was to determine in a simple and effective way differences between different strains of the mentioned taxon. Biochemical characteristics, antibiogram and part of the plasmid spectrum were used for assessment of inter-species differences between different strains and at the same time their use as simple markers of epidemiological analyses is presented.

The authors describe a false outbreak of tuberculosis by contamination in sample processing.The longitudinal polymorphisms of restriction fragments (RFLPs) of 6 strains of Mycobacterium tuberculosis isolated in different patients over a three week period and which were apparently implicated in an outbreak of tuberculosis were analyzed.Four of the strains studied presented identical restriction pattern and the remaining two presented totally different patterns. Following study of the clinical histories and the epidemiologic relationships three cases of tuberculosis were confirmed. The other three strains isolated corresponded to contamination during the sampling process.In a possible outbreak of six cases of tuberculosis, molecular techniques have allowed identification of three true cases of tuberculosis and have demonstrated contamination during the sampling process in three other cases. The latter could not have been shown with the clinical and phenotypical data of the strains.

This study evaluates a four-hour fluorescent DNA hybridization assay using both known bacterial isolates and clinical specimens. A biotinylated oligonucleotide probe from a sequence of the plasmid-encoded gene cppB was used. Hybrids were detected by addition of a streptavidin-alkaline phosphatase conjugate, followed by incubation for 30 min in a fluorescent substrate for alkaline phosphatase. The level of detection of the fluorescent assay was 0.1 pg of cryptic plasmid DNA or 200 cfu of the plasmid-containing strain NG 34/85 of Neisseria gonorrhoeae. A total of 119 reference strains of Neisseria gonorrhoeae and other related bacteria were tested for reactivity with the probe. All Neisseria gonorrhoeae strains, including eight plasmid-free strains, hybridized with the probe. Fluorescence ratios were 2.67 for plasmid-free strains and 3.85 for plasmid-containing strains. Of the heterologous microorganisms tested, only one of six strains of Neisseria cinerea gave a fluorescence ratio above the 2.0 cut-off value for positivity with the probe at a cell density of 1 x 10(4) cfu. The probe was also evaluated using clinical specimens from 100 patients attending a clinic for sexually transmitted diseases. The sensitivity of the assay was 100% while the specificity was 97.5%. Positive and negative predictive values were 91.2% and 100%, respectively. The fluorescent DNA hybridization assay for the detection of Neisseria gonorrhoeae described here thus appears to be a highly specific and sensitive assay.

With arbitrary primer PCR technique it is possible to obtain amplification lane patterns easily and with good reproducibility from the genomic DNA of bacteria studied. There is also no need for prior information regarding the DNA sequence.This method implies two cycles of amplification with low stringency, followed by a PCR of high stringency.Using the above mentioned technique, we were able to show that Campylobacter spp from clinical samples could be separated as well as different strains from the same species.According to our results as well as the ones from different authors applied to other microorganisms, we can assume that the method could be used in any bacterial species for epidemiologic studies purposes.

A one-chain DNA probe, that complements ribosomal RNA of Chlamydia trachomatis was used as a detection method for this microorganism on clinical samples. We compare the method with the cell culture one.A total of 175 samples (cervix swabs) from women seen at the STD center of the Facultad de Medicina de Sevilla were examined by both diagnostic techniques. When the results were different, a third method (ELISA) was also used.Using serial dilutions of a C. trachomatis cell culture as reference pattern, we determine the minimum number of inclusion forming units needed in order to be detected by the probe was 1000. Of all 175 samples, in 24 (14%) cell culture was positive for C. trachomatis, and 26 were positive using the DNA probe test. Sensitivity and specificity for this test were 93% and 95%, respectively.We believe that the DNA probe test was similar to the cell culture test as screening test in Chlamydia trachomatis infections diagnosis, specially among high risk populations.

A fluorometric procedure for the detection of DNA-DNA hybrids is described. The procedure involved the detection of probe-bound alkaline phosphatase with the fluorescent substrate ATTOPHOS. This substrate is converted to ATTOFLUOR by alkaline phosphatase and fluoresces strongly at 550 nm when excited with a wavelength of 440 nm. DNA hybridization assays were performed both with dilutions of purified target plasmid DNA (pSE9 or PBR322) and whole bacterial cells. Streptavidin-alkaline phosphatase conjugates were added to react with bound probe. Fluorometric assays, as well as colorimetric assays, using 5-bromo-4-chloro-3-indolylphosphate + nitroblue tetrazolium for alkaline phosphatase activity were performed. The fluorescence of the substrate was measured at time intervals, and the slope of the regression line calculated. A slope four times greater than that of background was considered positive. One hundred femtograms or 2.2 x 10(4) molecules of homologous DNA were detected with the fluorescent assay as compared with 10,000 femtograms or 2.2 x 10(6) molecules of homologous DNA with the colorimetric assay. Similar results were obtained with whole cells. Approximately 1 x 10(3) homologous cells were detected fluorometrically and 1 x 10(5) cells were detected colorimetrically. Based on these results, we conclude that, in our hands, the DNA hybridization assay described here using ATTOPHOS as the substrate for alkaline phosphatase is a very sensitive assay for the detection of DNA-DNA hybrids.

The antibiotics resistance pattern, the plasmid profile, the auxotypes and serotypes of 116 Neisseria gonorrhoeae clinical isolates obtained in one year were examined. The incidence of penicillinase producing (PPNG) strains was 12% (14 strains). The most frequent plasmid pattern was the combination of 4.5, 2.6 and 24.5 MDa plasmids. The conjugative plasmid of 24.5 MDa showed a high prevalence (32% of the total strains), and almost all the PPNG strains harboured this plasmid. The strains with the 4.5 MDa plasmid belonged to the auxotypes Pro-, Zero and Pro-Hyx-Ura-, whereas that with the 3.2 MDa plasmid was of auxotype Pro-Hyx-His-. The serotypes Aedih/Arst (WI serogroup) and Bak/Bropt, Back/Bropyt and Bak/Bropyt (WII/III serogroup) were predominant.

The diagnosis of congenital syphilis is difficult since it depends mainly on the results of serological tests. The results of five serological tests (three specific and two non-specific) in nine neonates with congenital syphilis are compared with those obtained in three with passively acquired antibodies. It appeared that the serological diagnosis of congenital syphilis must be based on the finding of specific neonatal antibodies in cord serum, which give positive results to the fluorescent treponemal antibody absorption test for immunoglobulin M, together with high titres of total IgM and negative results to latex tests. The non-specific tests are useful for confirming the efficacy of treatment. The mean number of cases of congenital syphilis in Seville is 0.81/1000 live births.

A comparative study of a DNA probe for the detection of Neisseria gonorrhoeae with Gram stain and modified Thayer-Martin medium was performed. The probe was the 2.6 megadaltons (Mda) cryptic plasmid of N. gonorrhoeae, labeled with a nonradioactive system combining sulphonation of DNA with the antibody detection of sulphonated groups (Organics Chemiprobe). Overall 101 samples were evaluated: 39 urethral exudates, 3 rectal exudates, and 59 cervical exudates, from 42 males and 54 females in whom gonococcal infection was suspected. Simultaneously, Gram stain and culture of the several exudates were carried out. The in vitro sensitivity of the method was evaluated with different dilutions of N. gonorrhoeae, from 1 to 1000 cfu/ml, and a minimum of 50 cfu/ml were detected. In the detection of N. gonorrhoeae in clinical samples, there was a 100% sensitivity and a specificity of 83% (males) and 79% (females), with positive predictive value of 97% in males and 53% in females. The negative predictive value was 100% both for males and females.

The ecology of R plasmids was studied in the intestinal flora of 19 patients with salmonellosis without antibiotic treatment. The plasmids found in the Salmonella strains and the accompanying non-pathogenic Enterobacteriaceae were characterized in each patient. We determined the transferability by conjugation, the fi character and the incompatibility group and did enzyme restriction analysis of these plasmids. The results obtained showed that S. typhimurium is the species of this genus with the highest incidence of R plasmids, and Escherichia coli among the non-pathogenic Enterobacteriaceae. The plasmids found in Salmonella are different from the plasmids found in the other Enterobacteriaceae in fi character (50% fi+ in Salmonella and 5% in the other Enterobacteriaceae) and incompatibility group (33% belong to the FII group in Salmonella plasmids and none on the other Enterobacteriaceae), thereby expressing a different origin.

S. typhi strains harboring R plasmids are a common finding in Mexico City hospitals. The predominant R plasmids share the following properties: they belong to the same incompatibility group (H1)’ have a molecular weight of 135 Mdal, carry a temperature sensitive transfer system, and code for the resistance to Cm, Sm, Su, Hg and Tc. The general organization of the resistance genes resembles that of RlOO, since homogenic derivatives which have lost the r-determinants, Tn1O or all the resistance genes can be isolated in vitro by several methods. Furthermore, naturally occurring a-plasmids deleted for the r-determinants or Tn1O have been found, although at low frequency. Complementation experiments indicated that the ts transfer system of the mexican plasmids is not related to that of Flac, Rl-19 or Col Ibdrd.

The in vitro activity of the new semisynthetic ureidopenicillins azlocillin (AZ) and mezlocillin (MZ), and of the new cephalosporin cefsulodin (CEF) were determined against 50 carbenicillin-sensitive (CARs) and 50 carbenicillin-resistant (CARr) P. aeruginosa clinical isolates. In the CARs group the most active antibiotics are AZ and CEF with MICs between 0.5 and 8 microgram/ml. Ticarcillin (TIC) and MZ showed more activity than CAR with MICs from 2 to 32 mu/ml. In this group there is a predominance of pyocine-type groups 1 and 3. In the CARr group, AZ is the most active antibiotic at low concentrations. At 64 microgram/ml of CEF, 72% of strains are inhibited as compared to 70% with AZ, 62% with MZ and 50% with TIC. In this group there is predominance of non-typeable strains. Interaction with mecillinam (MEC) and these antibiotics was studied on three different culture media (MH, NIH and DST). There were few cases of synergism with MEC and TIC combinations, mostly on DST medium. No appreciable synergism was found with other combinations of antibiotics.

The effects of subinhibitory concentrations (sub-MICs) of ampicillin on R plasmid transfer in Escherichia coli were studied. Each donor strain culture was separated into two parts; one was mixed with a recipient strain culture for mating, the other was treated with the sub-MIC of ampicillin and then mixed with the recipient strain culture. In both cases the R plasmid transfer frequency was determined at 30, 60, 90 and 120 min of mating. Results showed that there exists a general decrease in the transfer frequencies under sub-MIC treatment (two plasmids did not transfer at all). The proportion of aggregates and the number of cells that compose them were not affected by the sub-MIC of ampicillin. Our study supports the idea that the changes induced in E. coli by sub-MICs of ampicillin did not affect the function of the surface structures responsible for aggregation but did affect the proteins implicated in DNA transfer, situated on the cell surface.