Hybrid & El Tor variant biotypes of Vibrio cholerae O1 in Thailand.

BACKGROUND & OBJECTIVES: El Tor Vibrio cholerae O1 carrying ctxB C trait, so-called El Tor variant that causes more severe symptoms than the prototype El Tor strain, first detected in Bangladesh was later shown to have emerged in India in 1992. Subsequently, similar V. cholerae strains were isolated in other countries in Asia and Africa. Thus, it was of interest to investigate the characteristics of V. cholerae O1 strains isolated chronologically (from 1986 to 2009) in Thailand. METHODS: A total of 330 V. cholerae O1 Thailand strains from hospitalized patients with cholera isolated during 1986 to 2009 were subjected to conventional biotyping i.e., susceptibility to polymyxin B, chicken erythrocyte agglutination (CCA) and Voges-Proskauer (VP) test. The presence of ctxA, ctxB, zot, ace, toxR, tcpA C , tcpA E, hlyA C and hlyA E were examined by PCR. Mismatch amplification mutation assay (MAMA) - and conventional- PCRs were used for differentiating ctxB and rstR alleles. RESULTS: All 330 strains carried the El Tor virulence gene signature. Among these, 266 strains were typical El Tor (resistant to 50 units of polymyxin B and positive for CCA and VP test) while 64 had mixed classical and El Tor phenotypes (hybrid biotype). Combined MAMA-PCR and the conventional biotyping methods revealed that 36 strains of 1986-1992 were either typical El Tor, hybrid, El Tor variant or unclassified biotype. The hybrid strains were present during 1986-2004. El Tor variant strains were found in 1992, the same year when the typical El Tor strains disappeared. All 294 strains of 1993-2009 carried ctxBC ; 237 were El Tor variant and 57 were hybrid. INTERPRETATION & CONCLUSIONS: In Thailand, hybrid V. cholerae O1 (mixed biotypes), was found since 1986. Circulating strains, however, are predominantly El Tor variant (El Tor biotype with ctxB C).

Detection of Opisthorchis viverrini antigens in stools using specific monoclonal antibody.

Detection of Opisthorchis viverrini antigens in stools using specific monoclonal antibody. International Journal for Parasitology 22: 527-531. A sandwich enzyme-linked immunosorbent assay (ELISA) was developed for detecting Opisthorchis viverrini antigen in faecal extracts of four groups of individuals. These were 24 patients with O. viverrini infection only (group 1), 31 patients with O. viverrini and other parasitic infections (group 2), 141 patients with other parasitic infections (group 3) and 21 normal, parasite-free individuals (group 4). The first antibody used in the ELISA was polyclonal immunoglobulin G prepared from the serum of a rabbit previously immunized with crude extract of O. viverrini. The second antibody was monoclonal antibody specific to an antigen located in the worm tegument and muscular tissue. Sensitivity of the assay was 31% while specificity was 100%. Considerations for improving the sensitivity are discussed.

Monoclonal antibody to a diagnostic Mr 24,000 antigen of Gnathostoma spinigerum.

Crude water extract (CA) was prepared from the advanced third-stage larvae of Gnathostoma spinigerum collected from livers of naturally infected eels. The extract was partially purified by chromatofocussing column chromatography and the fraction which contained specific antigen of G. spinigerum which was an Mr 24,000 glycoprotein was used to immunize five Balb/c mice for preparing immune splenocytes. Spleen cells were collected from one mouse which showed high serum titre by indirect enzyme-linked immunosorbent assay and contained specific antibody to the Mr 24,000 antigen as checked by Western blot analysis. The spleen cells were fused with myeloma Sp2/0 cells at a ratio of 10 spleen cells per one myeloma cell using polyethylene glycol 3350 as a fusogen. Thirteen out of 174 growing polyclones (7.5%) produced antibodies to the partially purified CA fraction. Among them, two polyclones produced antibody directed to the Mr 24,000 protein. These two polyclones were subjected to monocloning by limiting dilution and a monoclone GN6/24 which produced monoclonal antibody to the specific Mr 24,000 protein of G. spinigerum was obtained.

Specific monoclonal antibodies to Opisthorchis viverrini.

A Balb/c mouse was immunized with a crude soluble antigen of Opisthorchis viverrini adult worms (OVAA) over a period of 7 months. Spleen cells from the immune mouse were fused with Sp2/0 myeloma cells. Among the 264 tissue culture wells containing the fused cells, cells of 96 wells (36%) produced antibodies to the immunizing agent. Antibodies produced by cells in several wells reacted with antigens from other species of parasite. Cells of 17 wells produced antibodies specific only to OVAA, thus cells from three representative wells were cloned by limiting dilution. Hybrids obtained produced antibodies which could be classified according to their tissue specificities into three groups. The first group of antibodies reacted strongly to the worm integument and weakly with the muscles while those belonging to the second group reacted only to muscles of the worms. The monoclonal antibodies of the third group gave a positive reaction to both muscles and tegument.

Field applications of an immunoradiometric assay for the detection of Plasmodium falciparum antigen in a population in a malaria-endemic area in Thailand.

Two surveys were made in people living in a malaria-endemic area in West Thailand in October 1985 (a transmission season) and in May 1986 (a nontransmission season) to detect Plasmodium falciparum antigen using the immunoradiometric assay (IRMA). In the first survey involving 101 people, the IRMA-positive rate was 56.4% and then significantly declined to 16.5% during the second survey involving 79 people of the same group. The parasitological-positive rates were likewise decreased from 11.9% to 1.3% (P = 0.015) during these two seasons. IRMA-positive rates were significantly higher than the corresponding parasitological-positive rates (P less than 0.0001 and 0.002 for the first and the second surveys, respectively). The geometric mean IRMA binding activity of samples collected in the first survey (1,726 cpm) was significantly higher than those collected during the second survey (920 cpm, P = 0.001). Regression analysis showed that IRMA activities were linearly correlated with the parasite counts by microscopic examination (r = 0.629, P = 0.022). IRMA was specific for P. falciparum since all 30 healthy controls and 6 of 7 vivax malaria cases were negative.

Factors contributing to the development of cerebral malaria. I. Humoral immune responses.

Humoral immune responses to malaria were studied in 100 patients with cerebral malaria of whom 53 had added complications, 108 patients with acute malaria, and 100 blood donors. The methods employed were indirect hemagglutination (IHA), indirect fluorescent antibody (IFA), enzyme-linked immunosorbent assay (ELISA), and parasite growth inhibition (PGI) tests. Patients with cerebral malaria, especially those with complications, had histories of fewer attacks of malaria in the previous 5 years than did those with acute malaria, suggesting that the cerebral malaria patients were less immune. The combined cerebral malaria group (complicated and uncomplicated) did not show defective humoral immune responses, since the initial seronegative rate and the mean initial IHA and IFA antibody titers were not significantly different from those of acute malaria patients and the mean initial ELISA titer was even higher than that of the acute malaria group. Reduced humoral responses were found only in complicated cerebral malaria patients, as their mean initial IHA titer was lower and their IHA seronegative rate was higher than those in acute malaria patients and in the uncomplicated cerebral malaria group. The combined cerebral malaria group had greater PGI activity than that of acute malaria patients, but this increased activity was entirely due to the higher results obtained in the complicated cerebral malaria group. The increased PGI activity returned to normal after recovery. An IgG preparation from seven of eight of these sera failed to exert the growth inhibition effect. Factors other than IgG were therefore responsible for the inhibition of parasite growth.

Genotypes and phenotypes of Shiga toxin producing-Escherichia coli isolated from healthy cattle in Thailand.

Shiga toxin producing-Escherichia coli (STEC) has not yet been identified as an important aetiologic agent of human disease in Thailand. To evaluate the potential for STEC to contribute to human disease in Thailand, 139 fecal samples were collected from healthy cattle from five different provinces and analysed by genotypic and phenotypic methods for STEC. Of 139 samples, 27 (19.4%) were positive for stx1 and/or stx2 by multiplex polymerase chain reaction, or for O157 lipopolysaccharide (LPS) by immunoassay. Isolates positive for stx and/or O157 were subdivided into 49 strains that varied in the presence of the virulence determinants stx1+/stx2+ (22 strains), stx2+ (22 strains), stx1+ (4 strains), and O157 LPS (1 strain). Within these 49 distinguishable strains, other virulence determinants varied as follows: hlyA+ (77.6%), eae+ and tir+ (4.1%), and katP+ (6.12%). The most predominant profile (22 isolates) was stx1+/stx2+, eae-, tir-, etpD-, hlyA+, katP-. For further characterization of the isolated strains by two molecular typing assays, plasmid profiles and ERIC PCR were performed. The results suggest that the genetic and phenotypic profiles of STEC associated with human disease are not prevalent at this time in cattle in Thailand.

Differential diagnosis of schistosomiasis mekongi and trichinellosis in human.

An indirect (plate) ELISA and, a more convenient version, a dot-blot (membrane) ELISA have been developed using haemocyanin of a mollusk, Megathura crenulata, i.e. keyhole limpet haemocyanin (KLH) and purified, specific antigen of Trichinella spiralis (APTsAg) obtained from a monoclonal antibody-affinity column chromatography, for differential diagnosis of schistosomiasis mekongi and trichinellosis. Serum samples of patients with parasitologically confirmed trichinellosis were reactive to both antigens in both versions of ELISA while sera of patients with schistosomiasis mekongi were positive only to the KLH. Both ELISA were negative when used to test sera of normal controls and patients with gnathostomiasis, paragonimiasis and opisthorchiasis.

Antibodies against Vibrio cholerae lipopolysaccharide, cell-bound haemagglutinin and toxin in the intestinal fluid during convalescence.

Specific antibodies to V. cholerae lipopolysaccharide (LPS), cell-bound haemagglutinin (CHA) and toxin (CT) in the intestinal lavages of healthy Thais and Thai cholera patients during the convalescence period were determined by enzyme-linked immunosorbent assay. Only IgM and IgA specific antibodies were detectable in the specimens. All of the persons who were just recovered from cholera had IgA anti-CT and IgA anti-LPS and 82.4% had IgA anti-CHA. The IgA anti-CT, anti-LPS and anti-CHA were detected also in the gut fluids of 70.6%, 94.1% and 88.2%, respectively, of the healthy controls. The mean levels of the IgA antibodies of all specificities between the two groups of individuals were not different. However, the IgM anti-CT and IgM anti-LPS of the cholera patients increased during the convalescence period. The levels, therefore, were significantly higher than those of the controls. The ratios of IgA anti-CT: IgM anti-CT and IgA anti-LPS: IgM anti-LPS among the patients were 2.93:1 and 2.02:1, respectively while those of the controls were 10:1 and 34:1, respectively. IgA antibodies predominated in the lavages of both groups of the individuals.

Simple serological tests for detecting classical heat labile enterotoxin (LT-I) of Escherichia coli.

Diarrhoea caused by enterotoxigenic Escherichia coli (ETEC) remains a problem in Southeast Asia. At present, no routine laboratories as yet are available for ETEC detection. In this study, attempts were made to produce reagents for use in simple serological tests for detecting LT. The serological methods were the Biken, the staphylococcal coagglutination and the reverse passive hemagglutination tests. For the Biken test, medium was prepared locally by mixing constituents as described previously by Honda et al., (1981). Anti-CT-B subunit was prepared by immunizing a rabbit with commercial CT-B subunits (Sigma). Other chemical reagents e.g. colistin, lincomycin etc. were obtained from the local supplies. Using the locally made reagents to detect LT from 100 WHO reference strains of E. coli by the Biken test, it was found that the test had 100%, 92%, 96%, 100% and 92.5% of specificity, sensitivity, accuracy, positive predictive value and negative predictive value, respectively. Protein A rich Staphylococcus aureus from the stock culture of the Department of Microbiology and Immunology, Faculty of Tropical Medicine, Mahidol University were grown in suitable medium i.e. blood agar containing lincomycin (BA-Lin). Suitable amount of the rabbit anti CT-B subunit (0.1 ml) was used to sensitize each ml of the formalinized, heat-fixed bacteria. The sensitized bacteria were used for detecting LT in the lysates of the 100 E. coli reference strains. The lysates were prepared by growing the E. coli strains on BA-Lin medium for 8 hours, then a loopful of each strain was inoculated into colistin solution (20,000 unit/ml).(ABSTRACT TRUNCATED AT 250 WORDS)

Oral vaccine against cholera prepared from Vibrio cholerae antigen(s).

Albino rats aged 7-8 weeks old purchased from the National Laboratory Animal Centre, Salaya, Nakhon Pathom, were found to be a good animal model for the study on immunogenicity of V. cholerae antigens. Seventy-two rats were fasted for 15 hours before feeding each one with 1 ml of 5% NaHCO3 to reduce gastric acidity prior to immunization. They were divided into 9 groups of 8 rats and immunized orally with 2 ml, each, of the V. cholerae antigens dissolved or suspended in Cassamino acid as follows: group 1 (control): Cassamino acid (Ca) alone; group 2 (control): 2.5% formalinized sheep red blood cells (F-SRBC); group 3: 1,000 micrograms of lipopolysaccharide (LPS); group 4: 100 micrograms of procholeragenoid (P); group 5: 80 haemagglutinating units of cell-bound haemagglutinin (CHA) adsorbed onto the surface of F-SRBC (CH-SRBC); group 6: 500 micrograms of LPS + 50 micrograms of P; group 7: CH-SRBC + 50 micrograms of P; group 8: combined vaccine formula 1 consisted of 500 micrograms of LPS, CH-SRBC and 50 micrograms of P and group 9: combined vaccine formula 2 consisted of 1,000 micrograms of LPS, CH-SRBC and 100 micrograms of P. The immunization was repeated once more 14 days later. Five days, thereafter, the rats were killed and their jejuni were removed for cryostat sectioning. Antibody producing cells against LPS (anti-LPS cells), P (anti-CT cells) and CHA (anti-CHA cells) in the intestinal lamina propria were enumerated by double antibody sandwich method of immunofluorescence using pure LPS, cholera toxin (CT) and pure CHA as the antigens in the assay, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Comparison of various serological test results using antigens from different strains of Plasmodium falciparum.

Suitability of different strains of Plasmodium falciparum grown continuously in vitro was compared using the indirect haemagglutination (IHA) the indirect immunofluorescent (IFA) tests and ELISA. In the tests employing soluble antigens (IHA and ELISA), there was a significant higher mean log titer of the same sera tested against different strains. Ranking of the strains in term of sensitivity for the detection of malaria antibody in people in the endemic area were G-112 = SO = CC greater than SU greater than PS in the IHA test and G-112 = SO greater than CC greater than SU greater than PS in the ELISA. The difference in the mean log titers appear to relate neither to the geographical location not the isoenzyme markers tested. There was also an apparent correlation between the results of the IHA and the IFA test but not between these two tests and ELISA.

Cross-sectional seroepidemiological survey of malaria in endemic areas with different activities of malaria control.

A single cross-sectional seroepidemiological survey of malaria antibody was conducted in 1982 in Klang District, Rayong Province in three villages under different phases of malaria control activity to determine whether a single survey could be used to delineate malaria endemicity in Thailand and to compare the usefulness of ELISA and the indirect haemagglutination test (IHA) in the assessment of malaria endemicity. Village 11 was a control area with high infection rate with an annual slide positive rate of 16.3% in 1981. Village 6 was also a control area but was in the late attack phase in which residual insecticide spraying has been ceased since 1976. Village 7 was a consolidation area. Finger-tipped blood was collected from 189, 191 and 132 individuals from villages 11, 6 and 7 respectively, and the plasma tested for anti-P. falciparum antibody with ELISA and IHA. With ELISA, it was shown that the seropositive rate in population of village 11 (84.6%) was significantly higher than those of other two villages (48.9% in village 6 and 28.8% in village 7). After age stratification, it was shown that the differences were observed in every age group except in the greater than or equal to 45 year age group of village 6. With IHA, a significantly higher seropositive rates in population of village 11 was evident when they were compared with the corresponding age groups of 6-14, 15-29 and 30-44 years in village 7, and the age group of less than or equal to 5 year in village 6.(ABSTRACT TRUNCATED AT 250 WORDS)

Human monoclonal anti-Plasmodium falciparum antibodies produced by stable EBV-transformed lymphocytes from patients with falciparum malaria.

Peripheral blood lymphocytes (PBL) from 10 persons living in a malaria endemic area and 18 patients recovered from falciparum malaria were studied, nine of whom were admitted to the Hospital for Tropical Diseases and the remaining nine patients were from Trad District Hospital. PBL were divided into two portions, one of which was transformed directly by EBV in the presence of cyclosporin A to eliminate T cell suppression and the other was pre-incubated before transformation with the extract of ultrasonically disrupted, schizont-enriched P. falciparum parasites from in vitro culture. The products of transformed cells were tested for antibodies against blood stages and sporozoites and cells from positive wells were cloned and propagated. With antigen pre-stimulation, cells from 212 of 317 wells (64.5%) were transformed, and this level of transformation was not significantly different from that in the absence of antigen stimulation in which 193 of 311 wells (62.5%) showed transformation (p greater than 0.05). In contrast, 85 of 212 (40.2%) clones from antigen prestimulated wells secreted antibodies whereas 18 of 193 (9.3%) wells without prior antigen stimulation did (p less than 0.0001). Only 44 of 103 antibody-positive clones were subjected to further analysis, of which 42 had activities against blood stages and two against sporozoites. Based on indirect immunofluorescent reactivities, our anti-blood stage monoclonal antibodies (MABs) were conformed to group I (21 clones), III (11 clones) and V (5 clones) and group VI (5 clones).(ABSTRACT TRUNCATED AT 250 WORDS)

Antibodies against circumsporozoite proteins of Plasmodium falciparum induced by natural infection.

Sera from 10 individuals who lived in a malaria endemic area, 10 patients with acute uncomplicated falciparum malaria and 10 patients with cerebral malaria and hyperimmune mouse serum were tested for their reactivities against Plasmodium falciparum sporozoite antigens by Western blot analysis using 125I-labeled staphylococcal protein A as the detecting reagent. These sera were shown by indirect immunofluorescence and/or circumsporozoite precipitation test to have antibodies reacting against the parasites. It was found that all serum antibodies from the three groups of individuals and the mouse serum reacted in a similar pattern with circumsporozoite (CS) proteins of P. falciparum. Ten sera from normal individuals were negative in all reactions. Monoclonal antibody (MAB) specific against CS proteins of the parasites showed that the proteins exhibited as four different molecular weight (MW) polypeptides, i.e., 67,000, 65,000, 60,000, and 58,000 daltons. These CS proteins of P. falciparum were found to be species and stage specific. Radioimmunoprecipitation using 35S-methionine-labeled parasites and sera of individuals from the various categories or MABs gave a similar result. Another protein antigen of P. falciparum sporozoites had a MW of 80,000 daltons. This antigen was not species specific, probably not membrane associated and was present in a minute quantity in the parasite's extract.

Vibriocidal antibody and antibodies to Vibrio cholerae lipopolysaccharide, cell-bound haemagglutinin and toxin in Thai population.

Vibriocidal antibody and antibodies to Vibrio cholerae lipopolysaccharide (anti-LPS), cell-bound haemagglutinin (anti-CHA) and toxin (anti-CT) were determined in Thai individuals of various age groups who lived in areas with high (H) and low (L) cholera endemicity. The enzyme-linked immunosorbent assay (ELISA) was performed to detect levels of class specific anti-LPS, anti-CHA and anti-CT. It was found that Thai individuals acquired the vibriocidal antibody early in life. Fifty percent of individuals aged 5 to 15 years old had detectable titre while more than 80% of adults had titres ranged from 1:5 to 1:125 or higher. Thai adults who lived in area with high cholera endemicity had significantly higher vibriocidal antibody levels than their counterparts who lived in area with low cholera endemicity. Lipopolysaccharide was not the only antigen responsible for stimulating the vibriocidal antibody production. Adult levels of all classes of anti-CHA from L were higher than those of H while the anti-LPS in the forms of total immunoglobulins, IgG and IgA were similar but IgM of L was higher than that of H. The levels of all classes of anti-CT from H seemed to increase with age except at the school age (5 years to 15 years old) when there were marked decreases of all antibody classes. Total immunoglobulin and IgM anti-CT at adult age of H and L were not different, although IgG anti-CT of L was higher than that of H and IgA anti-CT of H was higher than that of L.

Localization of Vibrio cholerae O1 in the intestinal tissue.

Colonization of V. cholerae O1 in vivo is known to be a non-invasive type which the vibrios are confined only to the intestinal tissues. The pathway by which the vibrio antigens reach the lymphoid cells and subsequently give rise to the immune responses is not entirely clear. Thus, experiments were performed in experimental rats by inoculating live V. cholerae O1 into the ligated ileal loops. The fate of the vibrios in the intestinal tissues was then studied by transmission electron microscopy at different times after the inoculation. It was concluded that live V. cholerae O1 were initially taken up by the M cells which overlay Peyer's patches and which subsequently delivered the intact vibrios to phagocytic cells in the Peyer's patches. These phagocytic cells processed (digested) the vibrios while the lymphocytes and plasma cells infiltrated around them. During the late period of infection (12-15 hours after inoculation of the vibrios), vibrios were also found passing through the loose intercellular spaces between the absorptive epithelial cells into the underlying intestinal tissues.

Immunodiagnosis of trichinellosis: efficacy of somatic antigen in early detection of human trichinellosis.

Crude antigens prepared from the infective stage larvae of Trichinella spiralis were used for antibody detection by indirect ELISA and Western blotting in serum samples taken from trichinellosis patients and from normal, parasite-free controls. The serum specimens were collected from acute ill, symptomatic patients on the first day of treatment (Day 0), and then two months (M2) and 4 months (M4) later. The sensitivities of the indirect ELISA and Western blotting on Day 0 were 81% and 92%, respectively. Both tests were 100% sensitive for M2 and M4 serum samples. Every serum sample from the parasite-free controls tested negative by both immunological assays, indicating 100% specificity. Crude somatic antigens can therefore be used for the early detection of human trichinellosis (acute trichinellosis).

Monoclonal antibody-based dot-blot ELISA for the detection of Salmonella in foods.

Monoclonal antibody (MAb) produced to polysaccharides in the LPS molecule of salmonellae was used in a dot-blot ELISA for detecting Salmonella in 873 food samples, ie 100 fresh chicken, 261 frozen chicken, 78 pork, 84 beef, 100 hen eggs, 100 duck eggs, 50 sea-mussels, 50 shrimps and 50 squids in comparison with the conventional culture method. Salmonella culture from foods involved the following steps: pre-enrichment, enrichment in selective medium, isolation on selective and indicator media, followed by biochemical and serological identification of appropriate colonies, respectively. The whole culture procedure took 5 days. Food samples from the selective enrichment medium were also subjected to the MAb-based dot-blot ELISA. The whole procedure of dot-blot ELISA took less than 2 hours. Among 873 food samples, salmonellae could be recovered from 7.4% of the samples by the bacterial isolation method (16% of fresh chicken, 8.8% of frozen chicken, 24.4% of pork, 3.6% of beef and 2% each of hen eggs and duck eggs, respectively). Salmonella derby were predominant among pork samples while S.paratyphi B biovar java predominated in chicken. The MAb-based dot-blot ELISA were positive in 19.5% of the food samples, i.e. 30% of fresh chicken, 27.6% of frozen chicken, 34.6% of pork, 21.4% of beef, 20% of shrimp, 16% of sea-mussels, 2% of hen eggs and 4% of duck eggs. The sensitivity and specificity of the MAb-based dot-blot ELISA compared to the bacterial culture method were 81.5% and 85%, respectively. The discrepancy of the data between the culture method and the dot-blot ELISA might be due to the fact that the culture method could detect only living cells at numbers that gave at least one isolated colony on the selective/differential plate while the dot-blot ELISA detects any form of Salmonella antigen. The monoclonal antibody-based dot-blot ELISA offers several advantages over the conventional bacterial culture method when it is used for screening of Salmonella contamination in foods, especially export foods. These include rapidity, cost-effectiveness and simplicity (the dot-blot ELISA does not need highly trained personnel or equipment, in contrast to the culture method). The test can be performed in field conditions and the result can be read visually. It also offers multisample analysis at one time which renders more samples of food for screening possible, thus false negative results are fewer which, in turn, assures the quality of the export food in a cost-saving, short time frame.

Detection of Salmonella contamination in food samples by dot-ELISA, DNA amplification and bacterial culture.

A dot-blot enzyme-linked immunosorbent assay (dot-ELISA) employing a genus Salmonella specific monoclonal antibody (MAb) was used for detection of the bacteria in food samples in comparison with the conventional culture method and the DNA amplification. Among the 200 chicken and pork samples (100 each) tested, 9% and 33%, 7% and 20% and 7 and 23% were positive for salmonellae by the dot-ELISA, the culture method and the DNA amplification, respectively. Statistical analyses revealed that the sensitivity, specificity, efficacy, and positive and negative predictive values of the detection of Salmonella in the food samples by dot-ELISA compared with the culture method were 93.33%, 91.76%, 92%, 66.66% and 98.73%, respectively. Comparison of the DNA amplification and the culture method revealed the sensitivity, specificity, efficacy, and positive and negative predictive values of 100%, 91.58%, 92%, 65.21% and 100%, respectively. The dot-ELISA and the DNA amplification results were in a better agreement when the two assays were compared. The sensitivity, specificity, efficacy, positive and negative predictive values of the dot-ELISA compared to the DNA amplification were 91.3%, 100%, 98%, 100% and 97.5%, respectively. From this study, the dot-ELISA is rapid, simple, sensitive, specific at low cost with limited amount of infectious waste to be disposed and offers another advantage in that it detects only the smooth LPS of Salmonella which implies the possible presence of the virulent organisms.