Aptamer functionalized MoS2-rGO nanocomposite based biosensor for the detection of Vi antigen.

We report a novel aptamer functionalized MoS2-rGO based electrochemical method for Vi polysaccharide antigen mediated detection of enteric fever. Herein, highly selective anti-Vi aptamers were screened from a pool of oligonucleotides using a microtitre based SELEX approach and characterized for its specificity and stability. The MoS2-rGO nanocomposite was synthesized using a liquid assisted exfoliation by taking optimum ratio of MoS2 and rGO. The nanocomposite presented synergistic effect owing to easy biomolecular functionalization and enhanced conductivity. The screened anti-Vi aptamers were embedded on the MoS2-rGO nanocomposite via thiol linkage to give a stable biointerface. The developed aptasensor was characterized and further evaluated for its performance with different concentrations of Vi antigen using ferrocene labeled boronic acid as an electroactive probe. The aptasensor responded linearly in the range between 0.1 ng mL-1 to 1000 ng mL-1with a detection limit of 100 pg mL-1, and did not show any cross-reactivity with other bacterial polysaccharides indicating high specificity. The applicability of the developed aptasensor was further validated in urine and sera specimens spiked with Vi antigen.

Detection of Salmonella typhi by nested polymerase chain reaction in blood, urine, and stool samples.

A nested polymerase chain reaction (PCR) specific for Salmonella enterica serovar Typhi was used for the detection of the pathogen in blood, urine, and stool samples from 131 patients with clinical suspicion of typhoid fever. The sensitivity of blood culture, the PCRs with blood, urine, and feces, and the Widal test were 61.8%, 84.5%, 69.3%, 46.9%, and 39.0%, respectively. The sensitivity of the PCRs with blood (P < 0.001) and urine (P = 0.01) were significantly higher, and the sensitivity of the PCR with feces (P > 0.05) was similar to that of blood culture. Combined, the PCRs on urine and feces showed positive results for 16 (70%) of 23 typhoid patients with negative results with blood culture and PCR with blood. These results show that the PCR with blood is a sensitive method for the diagnosis of typhoid fever, and that the PCRs with urine and feces could be useful complementary tests.

Sexually acquired Salmonella Typhi urinary tract infection.

We report a case of isolated urinary Salmonella enterica serotype Typhi in an HIV-positive man who has sex with men. He was clinically well and blood and stool cultures were negative, indicating that this may have been a sexually acquired urinary tract infection.

Bacteria isolated from blood, stool and urine of typhoid patients in a developing country.

Blood, stool and urine samples were collected from 100 patients diagnosed as having typhoid in 5 hospitals in Akwa Ibom State and analyzed for the presence of Salmonella species and other bacteria. Of the 100 blood samples screened, 55 (55%) were positive with the Widal test and 39 (39%) were positive on blood culture. Thirteen (14.1%) out of 92 urine samples were positive for bacterial growth, while 22 (26.8%) of the stool cultures were positive out of the 82 samples screened. Those within the age range 11-20 years old were infected most frequently (33%), followed by the age range 21-30 (19%) and 41-50 (18%) years old. Those in the age range of 0-2 years old (4%) were least infected. Female subjects were more infected than males. The commonest organisms isolated from the blood samples were Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa, Klebsiella pneumoniae, Proteus vulgaris, Streptococcus faecalis, Salmonella paratyphi and Salmonella typhi. S. aureus, S. epidermidis, E. coli, K. aerogenes, S. faecalis, Proteus mirabilis and P. aeruginosa were isolated from urine, while those isolated from stool were S. aureus, E. coli, S. typhi, S. paratyphi, Shigella sp, K. pneumoniae, P. vulgaris, P. aeruginosa and Vibrio cholerae 01. The isolates were sensitive to peflacine, ceftazidine, ciprofloxacin, ceftriaxone, cefotaxime and chloramphenicol. These antibiotics are recommended as the drugs of choice in therapy. The results suggest the existence of symptomless carriers of enteric fever bacilli in the state. This is worrisome, since some of the S. typhi isolates exhibited multiple resistance to commonly used antibiotics.

Production of monoclonal antibodies to Vi polysaccharide antigen of Salmonella typhi.

Twenty-four Vi antigen-specific monoclonal antibodies were produced in this study. The MAbs were found to be highly specific to Vi possessing bacteria. Selected MAbs were used in a direct agglutination assay for rapid identification of S. typhi in primary bacterial culture and also used to develop an assay to detect Vi antigen in clinical specimens. The result showed that they could not detect the antigen in urine and serum from acute patients even they could detect as low as 0.02 micrograms/ml of Vi antigen added in normal urine. The study has shown that these MAbs are very useful for rapid identification of S. typhi in primary bacterial culture and they can replace polyclonal anti-Vi antibodies which have been used routinely in bacteriological laboratories.

Duodenal string test in typhoid fever.

Twenty five children between 4-12 years of age hospitalized with a clinical diagnosis of enteric fever were studied for evaluating the practicality and sensitivity of duodenal string-capsule culture (DSCC) and compared with conventional cultures from blood (BC), urine (UC) and stool (SC). Duodenal string capsule (DSCC) was successfully inserted in 18 patients (72%). Insertion of DSCC failed in 7 patients (28%) and all of them were below 6 years of age. Salmonella typhi was isolated from DSCC and/or BC in 13 cases (72.2%). DSCC was positive in 11 out of 13 confirmed cases of typhoid fever (84.6%). BC was positive in 8 cases (61.5%). DSCC was successful in isolating the organism in about 30% more cases than BC. Duodenal string test was a simple, non-invasive and a reliable test which when used in combination with BC could identify almost all cases of enteric fever irrespective of duration of fever and prior use of antibiotics.

[The demonstration of specific Salmonella typhi antigens by the latex agglutination method]. / Indikatsiia spetsificheskikh antigenov Salmonella typhi metodom lateksnoi aggliutinatsii.

The evaluation of the diagnostic importance of the method of latex agglutination with antibody diagnostica for the detection S.typhi O and Vi antigens in biosubstrates obtained from patients is presented. The clinical approbation of the method has made it possible to characterize its diagnostic specificity (whose indices varied from 95.8 to 100%), sensitivity and significant diagnostic effectiveness (67.7-91.3%). The method has been recommended for the early diagnosis of typhoid fever.

Enzyme-linked immunosorbent assay for detection of Salmonella typhi Vi antigen in urine from typhoid patients.

Because typhoid fever continues to be a major cause of illness in many developing countries, there is a clear need for a sensitive and specific test that will permit rapid laboratory diagnosis of the disease. An enzyme-linked immunosorbent assay (ELISA) has recently been developed and tested, both in the laboratory and in a clinical situation, for its ability to detect Vi antigen in urine. The ELISA was capable of detecting as little as 1 ng of purified Vi antigen per ml in urine, compared with 100 ng/ml detectable by a previously tested coagglutination method. It could also detect antigen in urine diluted as much as 1:1,024 in normal urine. In tests of urine specimens from six stool culture-positive persons in a small typhoid outbreak in the United States, the ELISA detected antigen in specimens from four of the six patients. The ELISA also proved to be specific, giving no false-positive results for specimens from 50 persons who did not have typhoid fever. The apparent high sensitivity and specificity of this ELISA make it a promising test for rapid diagnosis of typhoid fever.

Detection of urinary Vi antigen as a diagnostic test for typhoid fever.

Since Vi antigen is limited primarily to Salmonella typhi, it has been thought that detection of the antigen may be a useful method for diagnosing acute typhoid fever. The slide coagglutination method and enzyme-linked immunosorbent assay have recently been suggested as ways to detect small quantities of Vi antigen in urine. In Santiago, Chile, we compared the results of these two methods in patients with acute typhoid fever, paratyphoid fever, and other febrile illnesses and in afebrile control subjects. Using a cut-off value that maximally separated typhoid patients from controls, the enzyme-linked immunosorbent assay was positive in 62.4% of 141 patients with culture-proven typhoid infections and in 13.2% of 159 afebrile control subjects. The enzyme-linked immunosorbent assay was false positive in 64.7% of 34 culture-proven paratyphoid A or B patients and 47.1% of 21 patients with other nontyphoidal febrile illnesses. The coagglutination test was positive in 34% of typhoid patients, 14% of afebrile control subjects, and 46% of febrile control subjects. We conclude that these tests when performed with the Vi antibodies employed in this study are of little value for the diagnosis of typhoid fever in this setting.

Randomized prospective study comparing two dosage regimens of ciprofloxacin for the treatment of typhoid fever.

Sixty-two patients with blood culture-proven typhoid fever were randomly assigned to receive either 500 or 750 mg of ciprofloxacin orally, twice daily for 7 days or for two days following defervescence, whichever was greater. Thirty-four and 28 patients received 500 mg and 750 mg respectively. Strains of Salmonella typhi resistant to ampicillin, chloramphenicol and co-trimoxazole were isolated from the blood of 27 patients (43.5%). No resistance to ciprofloxacin was encountered. Both regimens were equally effective; fever subsided in mean times of 4.9 +/- 1.7 days in the 500 mg group and 5.2 +/- 2.2 days in the 750 mg group (P = 0.54). All patients were cured, although one patient in the 750 mg group experienced a presumed relapse two months following completion of therapy. Ciprofloxacin administered for 7-10 days was adequate treatment for 57 of the 62 patients (92%); only five patients required therapy for more than 10 days. Patients with pretreatment symptoms of > or = 10 days duration defervesced in a mean of 5.7 +/- 2.3 days compared with 4.5 +/- 1.3 days (P = 0.01) for those with symptoms of shorter duration. We conclude that 500 mg of ciprofloxacin taken orally twice daily is adequate treatment for typhoid fever.