Paraplegia caused by invasive spinal aspergillosis.
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Adult↗
Biomedical subjects
Amita Gupta
Publications and source records attributed to Amita Gupta.
The traveling farm wife.
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Acute Kidney Injury↗
Same-sex behavior and high rates of HIV among men attending sexually transmitted infection clinics in Pune, India (1993-2002).
OBJECTIVES: To determine HIV/sexually transmitted infection (STI) prevalence, trends, and risk behaviors of men who have sex with men (MSM) and compare these with those of non-MSM attending STI clinics in Pune, India over a 10-year period. DESIGN: Cross-sectional. METHODS: From 1993 through 2002, men attending 3 STI clinics in Pune underwent HIV/STI screening. Demographic, risk behavior, clinical, and laboratory data were collected using standardized questionnaires and laboratory procedures. RESULTS: Of 10,785 men screened, 708 (6.6%) were MSM. Among these 708 MSM, 189 (31.7%) had 10 or more lifetime partners, 253 (35.7%) were married, 163 (23.1%) had sex with a hijra (eunuch), and 87 (13.3%) had exchanged money for sex. A total of 134 (18.9%) were HIV-positive, 149 (21.5%) had genital ulcer disease (GUD), 37 (5.8%) had syphilis, and 29 (4.3%) had gonorrhea (GC). Over the decade, neither HIV nor GC prevalence changed among MSM (P = 0.7), but syphilis and GUD decreased significantly (P < 0.0001). Compared with non-MSM, MSM were more likely to initiate sexual activity at age <16 years, to have >10 lifetime partners, to have sex with a hijra, and to use condoms regularly, but they did not differ significantly in HIV prevalence and had a lower prevalence of GC, GUD, and syphilis. Independent factors associated with HIV among MSM were employment (adjusted odds ratio [AOR] = 3.08; P = 0.02), history of GUD (AOR = 1.86; P = 0.003), and syphilis (AOR = 2.09; P = 0.05). CONCLUSIONS: Same-sex and high-risk sexual behaviors are prevalent among men attending STI clinics in India. Although syphilis and GUD rates decreased, HIV prevalence remained high during the decade, highlighting the importance of additional targeted efforts to reduce HIV risk among all men, including MSM, in India.
Adult↗
Bifunctional recombinant fusion proteins for rapid detection of antibodies to both HIV-1 and HIV-2 in whole blood.
BACKGROUND: Availability of accurate diagnostic tests has been helpful in curtailing the spread of HIV infection. Among these, simple, point of care, inexpensive tests which require only a drop of blood from finger-prick and give reliable results within minutes are a must for expansion of testing services and for reaching mobile and marginalized populations. Such tests will not only be a boon for the infrastructure-starved developing and underdeveloped countries but will also be extremely useful in developed countries where post-testing compliance is a major problem. Our laboratory has been involved in developing reagents for heamagglutination-based rapid detection of antibodies to HIV in whole blood using recombinant molecules specific for either HIV-1 or HIV-2. Since it is not required of a screening test to differentially detect HIV and HIV-2, it would useful to create a single molecule capable of simultaneous detection of both HIV-1 and HIV-2 in a drop of blood. RESULTS: The present paper describes designing, high-level expression and large-scale purification of new molecules comprising recombinant anti-RBC Fab fused to immunodominant regions of envelope sequences from both gp41 of HIV-1 and gp36 of HIV-2. These immunodominant regions of HIV envelope contain cysteine residues, which make disulfide bond and can interfere with the assembly of light chain and heavy chain fragment to make Fab molecule in vitro. To circumvent this problem, a series of fusion proteins having different combinations of native and mutant envelope sequences were constructed, purified and evaluated for their efficacy in detecting antibodies to HIV-1 and HIV-2. A chimeric molecule comprising native envelope sequence of gp41 of HIV-1 and modified envelope sequence of gp36 of HIV-2 gave good production yield and also detected both HIV-1 and HIV-2 samples with high sensitivity and specificity. CONCLUSION: The new bifunctional antibody fusion protein identified in this study detects both HIV-1 and HIV-2 infected samples efficiently and can be used in place of molecules that detect only HIV-1 or HIV-2. This will make reagent production more economical as only one molecule has to be produced in place of two molecules. Also, it will simplify the testing procedure allowing detection of both HIV-1 and HIV-2 infections in a single drop of blood.
Antibodies, Bispecific↗
Human papillomavirus DNA in urine samples of women with or without cervical cancer and their male partners compared with simultaneously collected cervical/penile smear or biopsy specimens.
Infection of specific types of high-risk human papillomaviruses (HPVs) causes cervical cancer in women. Conventional test for genital HPV infection requires collection of scraped cervical cells or biopsy specimens, which involves invasive procedures. Utility of non-invasive urine sampling for detection of HPV in women and their male sexual partners is controversial. The validation of this urine-based HPV DNA test is of immense value not only in screening large population and children but also for HPV vaccine monitoring in adolescents. We examined the frequency of high risk HPV types 16 and 18 in simultaneously collected urine samples and cervical scrapes or biopsy specimens from women with cervical cancer and their single lifetime male sexual partners in order to validate the utility of urine sampling as a reliable non-invasive method for detection of genital HPV infection. Thirty women with invasive cervical cancer and their husbands along with 30 age-matched normal healthy women including their husbands were recruited for the study. Cervical biopsies/scrapes from women subjects and penile scrapes from their husbands and urine samples from all of them were collected before taking biopsy or scrapes. HPV-L1 consensus primer as well as high-risk HPV (HPV 16 and 18) type-specific oligo-primers were used for PCR detection of HPV DNA. The total frequency of HPV in women with cervical cancer was found to be 83% (25/30) while it was only 67% (20/30) in their male partners but there was virtually no difference in results between urine and scrape or tissue biopsy either in women or their male partners. Although healthy women and their husbands showed similar frequency of HPV infection both in urine and scrape samples, there was a significant difference (p=0.05) in the prevalence of high risk HPV type 16 in women with cervical cancer (70%) and their male partners (30%). Similar was the trend between control women and their male partners. The results also showed a very high prevalence of HPV type 16 among Indian women with cervical cancer while its frequency was significantly low in their single lifetime male partners. The case by case matching of HPV positivity and negativity between urine and cervical/penile scrapes or biopsies obtained from women and their male partners demonstrated that the non-invasive urine sampling can be reliably used for screening genital HPV infection in both men and women.
Adult↗
High HIV prevalence among a high-risk subgroup of women attending sexually transmitted infection clinics in Pune, India.
OBJECTIVE: To investigate changes over a decade in prevalence and correlates of HIV among high-risk women attending sexually transmitted infection (STI) clinics in Pune, India, who deny a history of commercial sex work (CSW). DESIGN: Cross-sectional. METHODS: From 1993 to 2002, 2376 women attending 3 STI clinics in Pune were offered HIV screening. Women who denied CSW were included (n = 1020). RESULTS: Of 1020 women, 21% were HIV infected. The annual HIV prevalence increased from 14% in 1993 to 29% in 2001-2002 (P < 0.001). The change in HIV prevalence over time was paralleled by changes in clinic visitor characteristics; in later periods, women were older, more often employed, less likely to be currently married, and more likely to report condom use. In multivariate analysis, factors independently associated with HIV were calendar period (adjusted odds ratio [AOR], 1.9 for 1997-1999 vs. 1993-1996; 95% CI, 1.2-3.0; AOR, 2.3 for 2000-2002 vs. 1993-1996; 95% CI, 1.5-3.6), lack of formal education (AOR, 2.0; 95% CI, 1.4-2.9), having been widowed (AOR, 3.1; 95% CI, 1.6-6.1), current employment (AOR, 1.8; 95% CI, 1.2-2.6), and genital ulcer disease on examination (AOR, 1.8; 95% CI, 1.2-2.7). CONCLUSIONS: Women attending STI clinics in India who deny a history of CSW represent a small, hidden subgroup, likely put at risk for HIV because of high-risk behavior of their male partners, generally their husbands. Educational and awareness efforts that have targeted other subgroups in India (men and CSWs) should also focus on these hard-to-reach women. Risk reduction in this subgroup of Indian women would also be expected to reduce perinatal infections in India.
Adolescent↗
Images in clinical medicine. Eye worm.
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Adult↗
Necrotizing enterocolitis associated with clostridium perfringens type A in previously healthy north american adults.
BACKGROUND: Necrotizing enteritis associated with Clostridium perfringens type C ("pigbel") is a well-known syndrome in severely protein-deprived populations in the Pacific. It is exceedingly rare in the developed world. C perfringens type A is a common cause of acute gastroenteritis and, in a handful of infections, has been reported in association with a syndrome resembling necrotizing enteritis. STUDY DESIGN: This study includes a case series and literature review. Charts and autopsy reports from four patients with adult necrotizing enterocolitis (ANEC) were reviewed. C perfringens isolates were subtyped by mouse bioassay and pulsed-field gel electrophoresis. Fixed tissue specimens were tested with an anticlostridial antibody using an immunohistochemical assay. RESULTS: Between 2000 and 2003, ANEC developed in four previously healthy men; three died. The small bowel was affected in three patients and the colon in two patients. Portal or mesenteric vein thrombosis occurred in three patients. C perfringens type A was isolated from three patients and immunohistochemical assay demonstrated clostridial antigens limited to affected areas of the intestine of all four. The nonculture positive patient had a strong epidemiologic link to one of the others, and a compatible clinical course. C perfringens of the same pulsed-field gel electrophoresis-defined molecular subtyped was isolated from stool samples of one patient, his wife, and food from a restaurant they patronized. CONCLUSIONS: ANEC associated with C perfringens type A infection occurred in four North American adults. Culture for C perfringens type A should be performed in cases of ANEC. Alternative tests such as immunohistochemical assay were diagnostically useful. Additional research might uncover virulence factors, host factors, and the burden of disease in the population.
Adult↗
Expression and purification of recombinant 38-kDa and Mtb81 antigens of Mycobacterium tuberculosis for application in serodiagnosis.
Availability of genome sequence of Mycobacterium tuberculosis has accelerated identification of antigens for serodiagnosis of tuberculosis and a number of new antigens are being tested in various combinations to produce cocktails with high sensitivity and specificity. For producing a highly specific diagnostic test, it is important that the recombinant antigens be highly pure, free of host protein, and correctly folded so that they bind only to specific antibodies. Also, for commercial viability they need to be produced in high yields. We have cloned, expressed, and purified a number of mycobacterial antigens in Escherichia coli. This paper describes, expression and purification of two important mycobacterial proteins with serodiagnostic potential, namely, 38-kDa and Mtb81 antigens, in monomeric form. The protocol involves using a T7 promoter based expression vector under conditions of regulated and slow expression followed by three-step column chromatography procedure to obtain highly purified proteins. The yields of the two proteins were several folds higher than previously reported. The purified proteins were useful in detecting antibodies in sera of tuberculosis patients (smear positive, smear negative, and extra-pulmonary categories).
Antigens, Bacterial↗
Expression and purification of recombinant antigens of Mycobacterium tuberculosis for application in serodiagnosis.
Accurate diagnosis is essential for the treatment, prevention, and control of tuberculosis. Poor specificity of the tuberculin skin test in BCG-vaccinated populations and constraints to implementation of PCR and CMI-based diagnostic assays in developing countries warrant development of easy-to perform robust serological tests. Due to great heterogeneity in humoral response in TB patients, it will be necessary to include several antigens in any diagnostic assay to achieve useful levels of sensitivity and specificity. This needs production of recombinants, soluble versions of mycobacterial antigens in high yields. We have cloned, expressed, and purified a number of mycobacterial proteins in Escherichia coli. This paper describes the expression and purification of four promising sero-reactive proteins namely, ESAT6, CFP10, MTC28, and 14-kDa antigen of Mycobacterium tuberculosis. The protocol involves regulated and slow expression of proteins by using a T7 promoter-based expression vector for obtaining soluble protein followed by a three-step column chromatography procedure employing media with high binding capacity and flow characteristics. The yields of these proteins obtained were several folds higher than previously reported. The purified proteins were useful in detecting antibodies in sera of TB patients (smear positive, smear negative, and extra-pulmonary categories) and in combination with other immunodominant antigens will be useful in increasing the sensitivity to detect M. tuberculosis specific antibodies.
Antibodies, Bacterial↗
Limited evolution in the HIV type 1 pol region among acute seroconverters in Pune, India.
India has the second largest burden of HIV-1-infected persons worldwide. Access to antiretroviral drugs in India is increasing. We analyzed HIV-1 protease and reverse transcriptase sequences in 12 acute seroconverters from Pune, India, and evaluated HIV-1 evolution in these individuals over time. HIV-1 genotyping was performed with the ViroSeq HIV-1 Genotyping System. Baseline samples, collected between 1999 and 2001, had viral loads from 3,523 to 8,556,280 copies/ml. All subjects had subtype C HIV-1. None of the samples had primary drug resistance mutations. The sequence identity between baseline and 1-year samples ranged from 99.7% to 99.9%, and between baseline and 2-year samples ranged from 99.4% to 100%. Most of the nucleotide changes were silent (synonymous). Amino acid substitutions were rare, and varied from subject to subject. In this cohort, drug resistance was not observed and evolution in the pol region was very limited during the first 2 years of infection.
Acute Disease↗
Antimicrobial-resistant Shigella sonnei: limited antimicrobial treatment options for children and challenges of interpreting in vitro azithromycin susceptibility.
BACKGROUND: Antimicrobial-resistant Shigella sonnei is a growing problem in the United States and poses treatment challenges particularly among children. Azithromycin is recommended as an alternative oral agent for shigellosis. METHODS: All isolates of Shigella submitted to Johns Hopkins clinical laboratory during the outbreak year (2002) were compared with a historical comparison group (1996-2000). Isolates were considered multiresistant if they were resistant to ampicillin and trimethoprim-sulfamethoxazole (TS). Selected outbreak and reference isolates were tested for azithromycin susceptibility by E-test, disk diffusion and broth dilution methods. RESULTS: Between 1996-2000, among the 111 isolates submitted, 63% were from pediatric patients; 63% of isolates were resistant to ampicillin and 12% to TS. In 2002, among the 205 isolates submitted, 82% were from pediatric patients; 91% isolates were resistant to ampicillin and 67% to TS. The proportion of multiresistant isolates increased from 6% in 1996 to 65% in 2002 (P < 0.05). Azithromycin susceptibility by E-test and disk diffusion demonstrated 2 zones of inhibition for S. sonnei. Interpretation using the inner zone resulted in higher MICs (minimal inhibitory concentration) compared with the outer zones by E-test (P < 0.0001) and disk diffusion (P < 0.0001). CONCLUSIONS: With increasing interest in using azithromycin for shigellosis, clinical laboratories should be aware of the interpretation difficulty caused by the dual-zone phenomenon seen with E-test and disk diffusion methods for S. sonnei.
Adolescent↗
Laboratory-confirmed shigellosis in the United States, 1989-2002: epidemiologic trends and patterns.
During 1989-2002, a total of 208,368 laboratory-confirmed Shigella infections were reported to the Centers for Disease Control and Prevention. Shigella sonnei accounted for 71.7%, Shigella flexneri accounted for 18.4%, Shigella boydii accounted for 1.6%, and Shigella dysenteriae accounted for 0.7% of infections; for 7.6%, no serogroup was reported. National incidence rates ranged from 7.6 cases per 100,000 persons in 1993 to 3.7 cases per 100,000 persons in 1999. Incidence rates for S. boydii, S. dysenteriae, and S. flexneri decreased over the 14-year period by 81%, 83%, and 64%, respectively; S. sonnei rates only decreased by 8%. The highest rates were reported from western states (10.0 cases per 100,000 persons) and among children 1-4 years of age (20.6 cases per 100,000 persons). The female-male S. sonnei incidence rate ratio among 20-39-year-old adults decreased from 2.3 during 1989-1999 to 1.4 during 2000-2002. Approximately 1% of isolates were from extraenteric sources; 0.25% were from blood. S. sonnei remains an important cause of diarrhea in the United States. Prevention efforts that target high-risk groups are needed.
Dysentery, Bacillary↗
GB virus type C: a virus in search of a disease or a role in HIV therapy?
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CD4 Lymphocyte Count↗
Evaluation of rapid diagnostic tests for typhoid fever.
Laboratory diagnosis of typhoid fever requires isolation and identification of Salmonella enterica serotype Typhi. In many areas where this disease is endemic, laboratory capability is limited. Recent advances in molecular immunology have led to the identification of sensitive and specific markers for typhoid fever and technology to manufacture practical and inexpensive kits for their rapid detection. We evaluated three commercial kits for serologic diagnosis of typhoid fever. Patients presenting with > or = 4 days of fever were enrolled at two hospitals in Southern Vietnam. Cases were patients with serotype Typhi isolated from blood samples, and controls were patients with other laboratory-confirmed illnesses. Serotype Typhi isolates were confirmed and tested for antimicrobial susceptibility at the Pasteur Institute in Ho Chi Minh City. The Widal test was run at the hospitals and the Pasteur Institute. Sera were shipped frozen to the Centers for Disease Control and Prevention and tested by using Multi-Test Dip-S-Ticks, TyphiDot, and TUBEX to detect immunoglobulin G (IgG), IgG and IgM, and IgM, respectively. Package insert protocol instructions were followed. We enrolled 59 patients and 21 controls. The sensitivity and specificity findings were as follows: 89 and 53% for Multi-Test Dip-S-Ticks, 79 and 89% for TyphiDot, 78 and 89% for TUBEX, and 64 and 76% for Widal testing in hospitals and 61% and 100% for Widal testing at the Pasteur Institute. For all assays, the sensitivity was highest in the second week of illness. The Widal test was insensitive and displayed interoperator variability. Two rapid kits, TyphiDot and TUBEX, demonstrated promising results.
Adolescent↗
Antimicrobial resistance among Campylobacter strains, United States, 1997-2001.
We summarize antimicrobial resistance surveillance data in human and chicken isolates of Campylobacter. Isolates were from a sentinel county study from 1989 through 1990 and from nine state health departments participating in National Antimicrobial Resistance Monitoring System for enteric bacteria (NARMS) from 1997 through 2001. None of the 297 C. jejuni or C. coli isolates tested from 1989 through 1990 was ciprofloxacin-resistant. From 1997 through 2001, a total of 1,553 human Campylobacter isolates were characterized: 1,471 (95%) were C. jejuni, 63 (4%) were C. coli, and 19 (1%) were other Campylobacter species. The prevalence of ciprofloxacin-resistant Campylobacter was 13% (28 of 217) in 1997 and 19% (75 of 384) in 2001; erythromycin resistance was 2% (4 of 217) in 1997 and 2% (8 of 384) in 2001. Ciprofloxacin-resistant Campylobacter was isolated from 10% of 180 chicken products purchased from grocery stores in three states in 1999. Ciprofloxacin resistance has emerged among Campylobacter since 1990 and has increased in prevalence since 1997.
Animals↗
Escherichia coli O157 cluster evaluation.
We investigated a multistate cluster of Escherichia coli O157:H7 isolates; pulsed-field gel electrophoresis subtyping, using a single enzyme, suggested an epidemiologic association. An investigation and additional subtyping, however, did not support the association. Confirmating E. coli O157 clusters with two or more restriction endonucleases is necessary before public health resources are allocated to follow-up investigations.
Adolescent↗
High-density functional display of proteins on bacteriophage lambda.
We designed a bacteriophage lambda system to display peptides and proteins fused at the C terminus of the head protein gpD of phage lambda. DNA encoding the foreign peptide/protein was first inserted at the 3' end of a DNA segment encoding gpD under the control of the lac promoter in a plasmid vector (donor plasmid), which also carried lox P(wt) and lox P(511) recombination sequences. Cre-expressing cells were transformed with this plasmid and subsequently infected with a recipient lambda phage that carried a stuffer DNA segment flanked by lox P(wt) and lox P(511) sites. Recombination occurred in vivo at the lox sites and Amp(r) cointegrates were formed. The cointegrates produced recombinant phage that displayed foreign protein fused at the C terminus of gpD. The system was optimised by cloning DNA encoding different length fragments of HIV-1 p24 (amino acid residues 1-72, 1-156 and 1-231) and the display was compared with that obtained with M13 phage. The display on lambda phage was at least 100-fold higher than on M13 phage for all the fragments with no degradation of displayed products. The high-density display on lambda phage was superior to that on M13 phage and resulted in selective enrichment of epitope-bearing clones from gene-fragment libraries. Single-chain antibodies were displayed in functional form on phage lambda, strongly suggesting that correct disulphide bond formation takes place during display. This lambda phage display system, which avoids direct cloning into lambda DNA and in vitro packaging, achieved cloning efficiencies comparable to those obtained with any plasmid system. The high-density display of foreign proteins on bacteriophage lambda should be extremely useful in studying low-affinity protein-protein interactions more efficiently compared to the M13 phage-based system.
Bacteriophage M13↗