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PubMed1983

Etiological diagnosis of bacterial pneumonia by gram stain and quantitative culture of expectorates. Leukocytes or alveolar macrophages as indicators of sample representativity.

Sputum samples from 151 patients admitted to Roslagstull Hospital for Infectious Diseases, Stockholm, from Sept. 1978 through May 1979 with acute community-acquired lower respiratory tract disease and roentgenological evidence of acute pneumonia were examined by direct microscopy of gram-stained smears and semiquantitative culture. It was carefully noted if the specimen was collected before or after initiation of antibiotic therapy. For an estimate of the suitability of the samples for bacteriological examination 2 criteria were applied: (i) presence of alveolar macrophages, and (ii) purulence, i.e. a ratio leukocytes/squamous epithelial cells of greater than 5. The latter was found to be a good indicator of sample suitability, while the presence of macrophages was not. Of the 266 samples examined 76% were deemed purulent. Potentially pathogenic bacteria in numbers of greater than or equal to 10(5) colony forming units/ml were found in 67% of the purulent sputum samples obtained before antibiotic therapy but in only 36% if such treatment had already been started. Pneumococci were isolated from 52% of pre-treatment samples but from only 8% after treatment. H. influenzae was found as often in post-treatment samples (17%) as in pre-treatment ones (15%) and enteric gram-negative rods twice as often in post-treatment samples (11 vs. 6%). The use of gram-stained smears was a valuable aid in the interpretation of the culture results and the results could be made available to the clinician within minutes after receipt of the specimen. The results were in agreement with those of the cultures for about 75% of the purulent samples.

Acute Disease↗
PubMed2002

Field validation of a milk line sampling device for monitoring milk quality and udder health.

The objective of this study was to investigate the ability of a milk line sampling device to obtain a representative sample by comparing SCC and bacterial culture results between milk line and bulk tank samples for milk harvested from the same group of cows at the same milking. A total of 42 paired milk line and bulk tank samples were collected at separate milking events from 21 different herds. Concordance correlation coefficients showed a high level of agreement between the two sample types, with values ranging between 0.74 and 0.99 for all parameters and bacterial species measured. ANOVA showed that SCC and bacterial culture results for Streptococcus agalactiae, Staphylococcus aureus, Streptococcus non-agalactiae, Coliforms, and coagulase-negative staphylococci were neither numerically or statistically different between milk line and bulk tank samples. KAPPA analysis showed that overall agreement beyond chance between milk line and bulk tank samples in determining whether a herd was positive or negative for either Strep. agalactiae or Staph. aureus were 100 and 75%, respectively. While further research is needed to fully assess the utility of this tool for the purpose of bacterial culture, the results of this study suggest that the strategy of milk line sampling is a very promising monitoring tool. This sampling strategy should provide producers with inexpensive and timely information that will help to improve programs for monitoring milk quality and udder health in commercial dairy herds.

Analysis of Variance↗
PubMed2002

Field validation of a milk-line sampling device for monitoring milk component data.

The objective of this study was to investigate the ability of a milk-line sampling device to obtain a representative sample under field conditions by comparing the milk component composition between milk-line and bulk-tank samples, for milk harvested from the same group of cows at the same milking. A total of 42 paired milk-line and bulk-tank samples were collected at separate milking events from 21 different dairy herds. Samples were analyzed for milk fat (%), true protein (%), and milk urea nitrogen (mg/dl). Concordance correlation coefficients showed a very high level of agreement between the two sample types, with values ranging between 0.82 and 0.95 for the components measured. ANOVA showed that milk component data derived from milk-line samples were neither statistically nor numerically different from milk component data derived from bulk-tank samples. The strategy of monitoring milk component data through milk line sampling should provide dairy producers with inexpensive, timely, and accurate information that will help to improve ongoing nutritional monitoring programs for individual groups of cows within the dairy herd.

Animal Nutritional Physiological Phenomena↗
PubMed2004

Effects of storage time and thawing methods on the recovery of Mycoplasma species in milk samples from cows with intramammary infections.

This study was executed to determine the effects of storage and thawing on the viability of Mycoplasma spp. in milk from cows with intramammary infections. The trial was designed using a control sample and seven handling regimens subjected to two methods of thawing. There was a significant treatment effect on the recovery of colony-forming units in milk samples when comparing the control sample with handling regimens 1 through 7. There was a continuous decline in log (10) mean number of cfu/mL recovered. Mean concentrations were 6.29, 4.64, 3.69, 3.01, 1.86, 4.41, 4.13, and 3.18 for control and handling regimens 1 to 7, respectively. To determine the best thawing method, handling regimen 1 through 7 samples were thawed using two methods. On average, more mycoplasma were recovered from milk samples thawed at ambient temperature (4.04 cfu/mL) than milk samples thawed in a 37 degrees C water bath (3.76 cfu/mL). A final comparison was made between individual treatments. With the exception of the handling regimen 5 to 6 pair-wise comparison, all pair-wise comparisons between handling regimens were significantly different. The results of this study indicate that storage and thawing of milk samples is harmful to mycoplasma organisms. Fresh samples should be used to improve detection of Mycoplasma spp. from milk of infected cattle. If frozen samples are used, then length of storage time should be minimized, and thawing milk at ambient temperature will improve recovery of mycoplasma as opposed to thawing in a 37 degrees C water bath.

Animals↗
PubMed1987

A study of the stability of Record of Performance milk samples for infrared milk analysis.

The effect of switching individual cow Record of Performance milk samples from on-farm Babcock analytical system to centralized infrared analysis was investigated. The effects of lipolytic activity on the signals of the fat (5.73/5.58 and 3.48/3.56 microns) and protein (6.46/6.68 microns) wavelengths of filter instruments showed that lipolysis decreased the infrared fat signal and concurrently increased the protein signal. The chain length (3.48/3.56 microns) signal was unaffected by lipolysis due to the concordant movement of the reference wavelength baseline with the changes in the sample wavelength signal. Storage trials (3 d) of preserved milk samples indicated no direct relation between fat signal depression and chemically determined free fatty acid levels. Microbial growth and agitation also contributed to changes observed in samples studied under controlled conditions. Based on concurrent infrared, Babcock, and Mojonnier analyses of approximately 900 samples on d 1 and 3, there was a consistent but marginal drop in the carbonyl (5.73/5.56 microns) fat signal and an increase in the amide (6.46/6.68 microns) protein signal, but these changes were of limited significance. A study of 3-d and older samples indicated that significant levels of acid were produced, indicative of microbial growth. A comparison of Babcock results carried out on-farm, Mojonnier analyses in the laboratory, and their corresponding infrared results did not show statistically significant differences. Other than the inherent variability present in the individual cow samples, there were minor quality changes occurring under the present sample handling system due to a combination of lipolysis, microbial growth, and acid production. Refrigeration of Record of Performance samples during transport and at the central laboratory is recommended to minimize these changes.

Animals↗
PubMed2005

Comparison of lethal versus non-lethal sample sources for the detection of infectious salmon anemia virus (ISAV).

The emergence of infectious salmon anemia virus (ISAV) in Canada and the USA has led to the establishment of ISAV surveillance programs for cultured Atlantic salmon (Salmo salar L.) and wild fish species, including Atlantic salmon. Current testing procedures for ISAV consist of viral culture, reverse-transcription polymerase chain reaction (RT-PCR) and indirect fluorescent antibody testing (IFAT), and require lethal sampling. As the focus of this study, blood was evaluated as a possible non-lethal sample source for ISAV diagnostic screening by viral culture and RT-PCR. Tissue samples (consisting of kidney/spleen for viral culture or kidney only for RT-PCR), blood and, to a lesser extent, mucus were tested from Atlantic salmon survivors of laboratory ISAV infection trials and moribund fish from marine salmon grow-out facilities participating in a USDA-sponsored surveillance program. The trial fish represented a potential carrier population, while the surveillance fish were composed of moribund individuals from ISA clinical sites. Sample sources and diagnostic techniques were compared. Blood compared well to tissue samples for viral culture and produced a greater number of positives than did kidney samples for ISAV detection by RT-PCR. RT-PCR using both kidney and blood samples was determined to be a more sensitive assay than viral isolation. Mucus did not perform well in either assay compared to the other sample sources. Blood appears to be a reliable non-lethal sample source for the detection of ISAV by viral culture and RT-PCR in both moribund and asymptomatic fish.

Animals↗
PubMed2000

Detection of DNA of nocturnally periodic Brugia malayi in night and day blood samples by a polymerase chain reaction-ELISA-based method using an internal control DNA.

An internal control was used in a polymerase chain reaction (PCR)-ELISA-based technique to detect the Hha I repeat of the filarial parasite Brugia malayi. A single microfilaria added to 200 microl of blood was reliably detected. The assay was evaluated on field samples from persons living in an area endemic for Anopheles-transmitted, nocturnally periodic B. malayi in central Sulawesi, Indonesia. Examination of night blood of 138 individuals for the presence of microfilariae by filtration revealed 44 microfilaria carriers. All microfilaria carriers were also positive in the PCR-ELISA and, in addition, 14 more samples were proven to contain parasite DNA. The sensitivity of both methods was compared on night and on day blood samples collected from 113 persons. Whereas 37 microfilaria carriers were identified by filtration of night blood, no microfilariae were observed in the corresponding day blood samples. The PCR-ELISA result was positive in all 37 night blood samples of microfilaria carriers and in an additional 13 night blood samples without microfilariae. Parasite DNA was detected in 31 day blood samples of microfilaria carriers and in 3 day blood samples of amicrofilaremic persons. Assuming a sensitivity of the PCR-ELISA on night blood of 100%, the sensitivity of night blood filtration is 74% and that of the PCR-ELISA on day blood is 68%. These data suggest that the described PCR-ELISA method is capable of detecting infections with nocturnally periodic B. malayi in day blood samples. Therefore, this method may facilitate both the identification of endemic areas and the monitoring of control programs.

Adult↗
PubMed1998

Some remarks on the design of three-class sampling plans.

Among the variety of sampling plans for the evaluation of bacterial counts the attributive three-class sampling plan has widely gained acceptance because of its simple application and its robust functionality. However, the performance characteristics of three-class sampling plans depend on lot heterogeneity and the distance between the microbiological limits m and M, which are the maximum level of target organisms under conditions of good manufacturing practice (GMP) and the level of target organisms that is considered as unacceptable or defective. The probability of lot rejection due to a single sample result above M increases with increasing lot heterogeneity and/or with decreasing distance between these limits. Especially for investigations on nonpathogenic microorganisms it is questionable whether a lot still meeting GMP conditions should be rejected solely because a single sample result lies above M. Taking the often used three-class sampling plan (n = 5; cm = 2, cm = 0) as an example, it is demonstrated how insight into the relationship between sample variability and an appropriate distance between m and M can be gained. These calculations are based on the assumptions that logarithmically transformed bacterial counts follow a normal distribution and that an indifference lot with a contamination level at m is to be evaluated. For this kind of lot the probability of acceptance or rejection is equally 0.5 according to the two-class sampling plan. Introducing a limit for the additional risk of rejection of an indifference lot with acceptable heterogeneity when the three-class sampling plan is applied, a criterion for choosing the distance between m and M is developed.

Bacteriological Techniques↗
PubMed1999

Broiler skin sampling for optimum recovery of Salmonella spp.

The objective of this research was to determine the appropriate sample size (1, 5, or 10 g) and location (neck, breast, or vent) from which to sample processed poultry skin for Salmonella spp. Postkill, prescald broiler carcasses were used to help ensure that Salmonella spp. would be found. Mean Salmonella spp. counts from skin samples of 1 g (2.91 log10 CFU/g) were significantly lower (P < 0.05) than skin samples of 5 and 10 g: 3.52 log10 CFU/g and 3.42 log10 CFU/g, respectively. Mean Salmonella spp. counts from breast (3.62 log10 CFU/g) or neck (3.40 log10 CFU/g) skin samples were significantly higher (P < 0.05) than counts from vent skin samples (2.84 log10 CFU/g). Neck skin is the preferred sampling location because it contained a representative number of Salmonella spp., it had a slightly higher incidence of Salmonella spp. than vent skin, and removal of neck skin for microbiological testing did not decrease the quality grade of the bird, as would the removal of breast skin. Research results will increase the accuracy and precision of the microbiological analytical procedures for processed poultry by providing guidelines for the amount and location of skin to be sampled, as well as the preparatory procedures involved to release the Salmonella spp. from the skin samples.

Animals↗
PubMed2001

Microbiological sampling of meat cuts and manufacturing beef by excision or swabbing.

Groups of 25 beef or pork loin primal cuts or of pieces of stored or not stored manufacturing beef were sampled by excision and by swabbing with cotton wool, sponge, and gauze. Total aerobic counts, coliforms, and Escherichia coli from each sample were enumerated. Values for the mean log10, log10 mean, and/or the log10 total numbers recovered were calculated for each set of 25 bacterial counts. Those statistics indicated that, for product sampled without storage, swabbing with cotton wool or sponge recovered about 30%, and swabbing with gauze recovered about 10% of the bacteria recovered by excision sampling; but that for product sampled after storage, swabbing with cotton wool or sponge recovered about 50% and swabbing with gauze recovered about 15% of the bacteria recovered by excision sampling. However, the incidences of samples positive for coliforms and E. coli were less for stored than for nonstored product with all methods of sampling. The findings indicate that the conditions of meat surfaces, the handling of product, and the state of the microflora might all affect the numbers of bacteria recovered by any sampling technique. Thus, the relationship between the numbers recovered by excision or any selected swabbing technique may differ for different types of noncomminuted, raw meat product.

Animals↗
PubMed2002

Detection of Escherichia coli O157:H7 in 10- and 25-gram ground beef samples with an evanescent-wave biosensor with silica and polystyrene waveguides.

A portable evanescent-wave fiber-optic biosensor was used to detect Escherichia coli O157:H7 in seeded 10- and 25-g ground beef samples. The biosensor works by launching light from a 635-nm laser diode into specially designed optical fiber probes, generating an evanescent field that extends approximately 1,000 nm from the fiber surface. Fluorescent molecules within the evanescent field are excited, and a portion of their emission recouples into the fiber probe. The return path emission is transported by an optical fiber to a photodiode within the biosensor that detects and quantifies the fluorescent signal. A sandwich immunoassay was performed on the fiber probes with cyanine 5 dye-labeled polyclonal anti-E. coli O157:H7 antibodies for generation of the specific fluorescent signal. Biotin-streptavidin interactions were used to attach polyclonal anti-E. coli O157:H7 antibodies to the surface of the fiber probe. A centrifugation method was developed to obtain samples suitable for biosensor analysis from 10- and 25-g ground beef samples. The assay was shown to be sensitive and repeatable. One hundred percent correct identification of positive samples was demonstrated at 9.0 x 10(3) CFU/g for 25-g ground beef samples with silica waveguides and at 5.2 x 10(2) CFU/g for 10-g ground beef samples with polystyrene waveguides. The reaction was highly specific. No false positives were observed for 10-g ground beef samples not spiked with the pathogen. In addition, when samples were spiked with high concentrations of a variety of non-E. coli O157:H7 organisms, no false positives were observed. The method was rapid, with results being obtained within 25 min of sample processing.

Animals↗
PubMed1999

Prospective validation of single plasma sample 99mTc-ethylenedicysteine clearance in adults.

UNLABELLED: 99mTc-L,L-ethylene, L, dicysteine (EC) clearance shows strong correlation with orthoiodohippurate clearance, and it is possible to estimate effective renal plasma flow from 99mTc-EC clearance. In routine clinical studies, it is practical to use the one or two plasma sample method instead of multiple plasma samples for clearance determination. A single-sample technique was developed for 99mTc-EC, and a regression formula was generated. A prospective study tested the validity of this regression formula. METHODS: The study population was composed of 26 patients with a wide range of renal function. Multiple plasma sample 99mTc-EC clearances were calculated from all patients using the open two-compartment model. Single plasma sample clearances were also determined from the 54-min plasma sample using the regression formula published previously. RESULTS: The multiple-sample plasma clearance of 99mTc-EC ranged from 46 to 668 mL/min with a mean of 300.76 +/- 150.73 mL/min. The clearances obtained from the 54-min plasma sample ranged from 49 to 699 mL/min, with a mean of 297.39 +/- 152.23 mL/min. There was an excellent correlation between the clearances obtained by the two techniques (r = 0.99, slope = 0.9911). The standard error of estimation was found to be 25.9 mL/min. CONCLUSION: This study suggests that 99mTc-EC clearance can be estimated from 54-min plasma samples with an acceptable error of estimation for most routine clinical studies.

Adolescent↗
PubMed1997

[Study on the best sampling time and the time distribution of atmospheric bacterial particle].

The changes of atmospheric bacterial particle concentrations in three areas--Beijing, Tianjin and Shenyang, were investigated in different seasons and period of one day. The samplers used in this study were MF-45 and HTK-201 air microbial samplers. The results showed that the concentrations of atmospheric bacterial particle in Beijing and Tianjin are higher in spring than those in other seasons. They are 2053/m3 and 2556/m3 respectively in spring, 995/m3 and 1064/m3 respectively in summer. The concentrations of atmospheric bacterial particle in Shenyang is higher in autumn (10108/m3) and lower in winter (1294/m3). The changes of atmospheric bacterial particle concentrations have two peak values during a day. The two high peak values are at 6:00 to 7:00 and 18:00, while the two low values are at 11:00 to 13:00 and 1:00 to 2:00. We sampled the atmospheric bacterial particles at 4 different combinations of sampling time in the three cities. They were 12, 8, 6 and 4 times a day. The sampling time combination was analyzed by mathematical statistics according to the results. The best sampling time was decided by the statistics results and time distribution character of atmospheric bacterial particle concentration in different seasons and period of one day. The concentration of atmospheric bacterial particle should be detected in the middle month of the four seasons: January, April, July and October. Sampling time at 7:00, 10:00, 13:00, 16:00, 19:00, 22:00, 1:00 and 4:00 could be selected for 8 times sampling a day. The sampling time sequence should be at 5:00, 11:00, 17:00 and 23:00 for 4 times sampling a day. The four times sampling at daytime in spring and fall should be at 6:00, 9:00, 12:00 and 19:00; 5:00, 9:00, 13:00 and 17:00 in summer; 7:00, 10:00, 13:00 and 16:00 in winter.

Air Microbiology↗
PubMed1999

Reliability of an intravenous intermittent access port (saline lock) for obtaining blood samples for coagulation studies.

BACKGROUND: Patients admitted to rule out myocardial infarction often receive a heparin infusion that requires measurement of partial thromboplastin time every 6 to 8 hours until coagulation values stabilize. If the patient's acuity at admission does not necessitate placement of arterial or central catheters, insertion of an intermittent intravenous access port (saline lock) would decrease the need for frequent venipuncture. Many studies on obtaining blood samples via arterial and central catheters have been reported, but no reports have described the method of using a saline lock or the amount of blood discarded during that procedure. OBJECTIVE: To evaluate the efficacy of inserting a saline lock specifically for obtaining blood samples and to determine the amount of blood that must be discarded in order to obtain prothrombin and partial thromboplastin times that match the values from blood samples obtained via venipuncture. METHODS: Coagulation values determined with 2 consecutive blood samples obtained via the saline lock after the first 0.5 mL of blood withdrawn was discarded were compared with measurements determined with a blood sample obtained via venipuncture. RESULTS: Prothrombin and partial thromboplastin times were not significantly different among the 3 blood samples. CONCLUSION: The untoward effects of frequent venipunctures to obtain blood samples for measurement of prothrombin and partial thromboplastin times can be lessened by using an 18-gauge saline lock to obtain blood samples and discarding the first 0.5 mL of blood withdrawn before the samples for coagulation studies are collected.

Adult↗
PubMed1999

Increased detection of circulating tumor cells in the blood of colorectal carcinoma patients using two reverse transcription-PCR assays and multiple blood samples.

The objectives of this study were to assess whether the use of two reverse transcription-PCR (RT-PCR) cDNA assays and multiple blood sampling increased circulating tumor cell detection in colorectal cancer patients. Systemic blood was sampled three times at 1-min intervals in 100 colorectal cancer patients (50 primary tumors only and 50 liver metastases), and in 70 control patients without known cancer. After removal of the erythrocytes, samples were subjected to separate RT-PCR reactions using specific primers for carcinoembryonic antigen (CEA) and cytokeratin 20 (CK20). Statistical analysis was performed by the two-sample binomial test and the one-sided McNemar test. There were significant increases in circulating tumor cell positivity when CEA and CK20 assays were used together as compared with either CEA or CK20 assay used alone. There were also significant increases in circulating tumor cell positivity for either CEA or CK20 assay used alone when the results from two blood samples were compared with the results from one sample. Circulating colorectal cancer cell positivity rose from 48% (CEA) and 34% (CK20) with one assay of one sample to 74% when both assays of three samples were used to identify circulating tumor cells. Three non-cancer control patients (4.3%) were positive for either CEA (two patients) or CK20 (one patient). Tumor cells were identified more frequently in the circulation of colorectal cancer patients than had been suggested previously. RT-PCR-based studies of the clinical significance of circulating cancer cells in colorectal cancer should involve multiple blood samples with identification of multiple tumor-related cDNA products.

Aged↗
PubMed1999

Serum eosinophil cationic protein determination in asthmatic children-effect of different collecting tubes used for blood sampling.

We compared the effect of using 2 different serum collecting tubes, serum separation tubes (SST, with clot activator and gel barrier) and conventional glass tubes (with no additives), on circulating levels of eosinophil cationic protein (ECP) in asthmatic children and controls. The serum ECP values obtained from both SST and glass tubes were significantly higher in asthmatic children than in corresponding controls. ECP values were higher in serum samples using SST than in those using glass tubes (P<0.01), while no difference was found between the two in controls. ECP levels correlated with peripheral eosinophil counts, for SST samples and glass tube samples alike. The difference in ECP levels between these two tubes also correlated with circulating eosinophil counts (r = 0.62, P = 0.004) After 18-hour storage at room temperature, the ECP values increased significantly in samples obtained from asthmatic children. No difference in ECP values between SST samples and glass tube samples was found for 18 hour samples. Thus, ECP levels obtained from SST samples and glass tube samples, though reliable, should not be directly compared, especially in asthmatic children with eosinophilia.

Asthma↗
PubMed2000

[Percutaneous umbilical blood sampling in pregnant women with thrombocytopenia. Apropos of 35 cases].

OBJECTIVE: To determine the interest of percutaneous umbilical blood sampling in maternal thrombocytopenia. PATIENTS AND METHOD: We performed a retrospective study of 36 percutaneous umbilical samples in pregnant women with thrombocytopenia. We included patients with a platelet count less than 100,000/mm(3) or a previous diagnosis of idiopathic thrombocytopenic purpura. Patients were divided into two groups. Group A consisted of women whose thrombocytopenia was caused by idiopathic thrombocytopenic purpura. The other patients were included in the group B. Percutaneous umbilical blood samples were performed by direct ultrasound guidance under sterile conditions, after 37 weeks gestation. Cesarean section was indicated if the fetal platelet count was < or =50 000/mm(3), in other cases obstetric indications were followed. RESULTS: Percutaneous umbilical blood samples were performed at a mean age of 38.5 gestational weeks. Fetal blood sampling was successful in 34 of the 36 cases. There was no fetal complication. In group A, two fetuses had low cord-blood platelet counts (20,000 and 98,000 per cubic millimeter). There was no fetal thrombopenia in group B. Three cesarean sections were performed, in one case because of severe fetal thrombocytopenia and in 2 cases because of an unsuccessful percutaneous umbilical blood sampling (in both, neonatal platelet count was normal). There was no significant complication in infants. There was a significant correlation between fetal and neonatal platelet count. CONCLUSION: There is no maternal predictive factor for fetal thrombocytopenia. Percutaneous umbilical blood sampling is feasible and has a good diagnostic value. The overall risk of fetal loss is low. But there is no indication of percutaneous umbilical blood sampling in the management of thrombocytopenia in pregnancy because severity of thrombopenia is not directly related to the severity of maternal disease and there are no series large enough to assess the association between delivery method and intracranial hemorrhage in thrombocytopenic infants born of mothers with idiopathic thrombocytopenic purpura. More subjectively than on scientific arguments a percutaneous umbilical blood sampling could be performed for patients with idiopathic thrombocytopenia purpura and a past history of neonatal intracranial hemorrhage. A cesarean section could be done if the platelet count is less than 100 000/mm(3), no intracranial hemorrhage has been described above this level.

Cordocentesis↗
PubMed2000

[Evaluation of different clinical samples and cell lines in the isolation of enterovirus in pediatric patients].

BACKGROUND: To evaluate prospectively the efficacy of different clinical samples and cellular lines in the isolation of Enteroviruses from pediatric patients. METHODS: In the period july-1997 to july-1999 we analyze the samples of 102 pediatric patients (< 2 years old) with an unknown febrile etiology syndrome. After the process of decontamination the samples were inoculated in the MRC-5, Hep-2 and Vero cell lines by the shell-vial assay. After 2-3 days of incubation the monolayers were stained by a monoclonal antibody against VP1; then the strains were identified as Poliovirus, ECHO-virus or Coxsacke by specific antibodies. RESULTS: 96 clinical samples were studied (45 pharyngeal swabs, 28 feces, 13 cerebrospinal fluid, 5 bloods, 4 urines and one bronchoalveolar lavage). Enteroviruses were isolated in 48 patients (47%), corresponding to 60 clinical samples (62.5%). The enteroviruses were isolated in 75.5% of pharyngeal swabs, 71.4% of feces, 30.7% of cerebrospinal fluid, one blood (20%) and the bronchoalveolar lavage. 28 patients presented simultaneously both pharyngeal and rectal swabs; in this group the use of both samples permitted the isolation of Enteroviruses in 26 cases (92.8%). Of 60 Enteroviruses, 59 (98.3%) were isolated in the MRC-5 cell line, 23 (38.3%) in the Hep-2, and 14 (23.3%) in the Vero; the MRC-5 demonstrated a significant difference in the isolation of Enteroviruses compared with the other cell lines. The Enteroviruses were identified as 30 (50%) ECHO-virus, 15 (25%) as Poliovirus vaccine, 12 (20%) could not to be identified and in 3 cases (5%) no growth of virus was detected in the culture. The MRC-5 cell line was significatively superior to the other in the isolation of the different Enteroviruses. CONCLUSIONS: To obtain the maximum diagnostic efficacy in front a patient with a possible infection by Enteroviruses, is necessary to use different clinical samples, specially the more related to the infection focci (cerebrospinal fluid or blood). The MRC-5 cell line has been the most efficace in the isolation of Enteroviruses independently of the type of clinical sample or the viral genus. The shell-vial assay is a good method for the isolation and identification of Enteroviruses isolated from clinical samples.

Animals↗