Improved serological diagnosis of rubella.

The use of an enzyme immunoassay-immunoglobulin G antibody test instead of the hemagglutination inhibition test as a primary test for the serological diagnosis of current infection, with complement fixation as an alternate test for use when enzyme immunoassay results are high and stationary, improved the serological diagnosis of rubella.     

Evaluation of a commercial rubella IgM assay for use on oral fluid samples for diagnosis and surveillance of congenital rubella syndrome and postnatal rubella.

BACKGROUND: Clinical diagnosis (surveillance) of rubella is unreliable and laboratory confirmation is essential. Detection of virus specific IgM in serum is the most commonly used method. However, the use of serum necessitates the drawing of blood, either through venipuncture or finger/heel prick, which can be difficult in young babies. Oral fluid samples have proved useful as an alternative, less invasive sample for virus specific IgM detection however until recently no commercial rubella IgM tests were available, restricting the usefulness of this approach. OBJECTIVES: To evaluate the performance of the Microimmune Rubella IgM capture EIA using oral fluid samples from outbreaks as well as in cases of suspected congenital rubella syndrome (CRS). STUDY DESIGN: Paired serum and oral fluids were collected from cases during a rubella outbreak in three provinces in Turkey. Matched serum and oral fluid samples were collected from children with suspected CRS in an active surveillance programme at the Aravind Eye Hospital in South India. Serum samples were collected as part of the measles surveillance programme in Ethiopia. RESULTS: On serum samples the sensitivity and specificity of the Microimmune Rubella IgM capture EIA compared to Behring Enzygnost rubella IgM test was 96.9% (62/64; 95% CI 94.2-100%) and 100% (53/53; 95% CI 93.2-100%). On oral fluids compared to matched Behring results on serum the sensitivity was 95.5% (42/44; 95% CI 84.5-99.4%). The sensitivity and specificity of Microimmune Rubella IgM capture EIA on oral fluids from suspected CRS cases compared to serum results using Behring Enzygnost IgM assay was 100% (95% CI 84.5-100%) and 100% (95% CI 95.8-100.0%) respectively. CONCLUSION: Microimmune Rubella IgM capture EIA has adequate performance for diagnosis and surveillance of rubella in outbreak using either serum or oral fluid specimens.     

Assessment of the diagnostic value of RT-PCR on amniotic fluid for prenatal diagnosis of congenital rubella infection

AIM OF THE STUDY: To assess the diagnostic value of RT-PCR on amniotic fluid (AF) for prenatal diagnosis of congenital rubella infection. MATERIAL AND METHODS: RT-PCR on AF was compared to specific IgM antibody detection in foetuses and/or newborns in 45 pregnant women with confirmed primary infection. RESULTS: specificity of RT-PCR was 100% and sensitivity ranged between 83 and 95%. CONCLUSION: RT PCR may be considered as a valuable tool for prenatal diagnosis of foetal rubella infection.     

Detection of low-avidity immunoglobulin G in oral fluid samples: new approach for rubella diagnosis and surveillance

Low-avidity rubella immunoglobulin G (IgG) was detected in oral fluid samples from 30 of 32 rubella IgM-positive patients (sensitivity, 94%) and from 4 of 34 IgM-negative patients (specificity, 88%). Measuring IgG avidity in oral fluid samples could improve the reliability of rubella surveillance when the incidence of the disease and the positive predictive value of IgM tests are low.     

Enzyme-linked immunosorbent assay for the diagnosis of recent rubella infection.

This paper presents further evidence of the sensitivity and reproducibility of ELISA for the detection of rubella antibody. This test can also be extended to include the diagnosis of recent rubella infections once the index of significance has been computed based on the estimated ELISA ratio of absorbance between the acute and convalescent serum samples. There is a greater than 90% agreement when compared to the results obtained by the haemagglutination-inhibition test. It should be further evaluated for rubella using purified rubella antigen to increase sensitivity and reproducibility.     

A lymphocyte transformation assay for the diagnosis of congenital rubella.

A rubella-specific lymphocyte transformation assay, using cryopreserved mononuclear cells, has been developed and used to evaluate specific responses among 21 children with congenitally acquired rubella (CAR), 25 healthy control children and 10 children with sensorineural deafness of unknown aetiology. Although all 21 children with CAR were seropositive, 12 (57.1%) failed to respond to rubella antigen in the transformation assay. Negative in vitro lymphocyte transformation responses were detected significantly more frequently among congenitally infected children below 3 years of age. Thirteen of the 25 (52%) control children were seropositive; only one of these seropositive children (7.6%) gave a negative transformation response. A negative rubella-specific lymphocyte transformation response in a seropositive child, particularly when aged 3 years or younger, is therefore suggestive of CAR. Four of the 10 children with deafness of unknown aetiology were rubella seropositive but gave negative responses in the transformation assay, suggesting that these children had CAR. Our assay may provide a very useful test for retrospective diagnosis of CAR, particularly in children under the age of 3.     

Evaluation of sensitivity of TaqMan RT-PCR for rubella virus detection in clinical specimens

BackgroundAn easy and reliable assay for detection of the rubella virus is required to strengthen rubella surveillance. Although a TaqMan RT-PCR assay for detection of the rubella virus has been established in Japan, its utility for diagnostic purposes has not been tested.ObjectivesTo allow introduction of the TaqMan RT-PCR into the rubella surveillance system in Japan, the sensitivity of the assay was determined using representative strains for all genotypes and clinical specimens.Study designThe detection limits of the method for individual genotypes were examined using viral RNA extracted from 13 representative strains. The assay was also tested at 10 prefectural laboratories in Japan, designated as local reference laboratories for measles and rubella, to allow nationwide application of the assay.ResultsThe detection limits and amplification efficiencies of the assay were similar among all the representative strains of the 13 genotypes. The TaqMan RT-PCR could detect approximately 90% of throat swab and urine samples taken up to 5 days of illness. These samples were determined positive by a highly sensitive nested RT-PCR.ConclusionsThe TaqMan RT-PCR could detect at least 10 pfu of rubella virus. Although the sensitivity was somewhat lower than that of the conventional nested RT-PCR, the TaqMan RT-PCR could be more practical to routine tests for rubella laboratory diagnosis and detection in view of the rapid response and reducing risks of contamination.     

Improved method for cholera diagnosis.

The use of the oxidase test is proposed to discover sparse colonies of cholera vibrios on a plate of Pril nutrient agar seeded directly with the suspected stool sample or after enrichment. This method also enables the detection of other oxidase-positive potential pathogens, such as non-agglutinating vibrios, Aeromonas hydrophila, Pseudomonas spp., and other nonfermentative organisms.     

Use of ribotyping in epidemiological surveillance of nosocomial outbreaks.

Over the past few years, genotypic methods based on the study of bacterial DNA polymorphism have shown high discriminatory power for strain differentiation and superiority over most phenotypic methods commonly available in the clinical microbiology laboratory. Some of the methods used, however, required either a high level of technology and sophisticated equipment (e.g., pulsed-field gel electrophoresis) or species-specific reagents of restricted availability (randomly cloned DNA probes or gene-specific probes). Because ribotyping uses a universal probe (rRNA) and is a rather simple technology, particularly since the advent of nonradioactive labelling systems, it has been widely used for strain differentiation of most bacterial species involved in nosocomial outbreaks. In vitro and in vivo stability of the markers studied has been demonstrated. Although there may be limitation to this approach, ribotyping was found to be highly discriminative, particularly for typing members of the family Enterobacteriaceae, Pseudomonas cepacia, and Xanthomonas maltophilia. In many cases, it has improved the understanding of the mechanism of nosocomial acquisition of organisms by allowing a distinction between endogenous and exogenous infections. Among exogenous infections, it has distinguished between individual and epidemic strains, thus differentiating cross-infection from independent acquisition.     

Molecular genetic methods for diagnosis and characterisation of Chlamydia trachomatis and Neisseria gonorrhoeae: impact on epidemiological surveillance and interventions

One of the mainstays in the prevention of Chlamydia trachomatis and Neisseria gonorrhoeae infections is the availability of laboratory diagnostics with high sensitivity and specificity. Assays for diagnosis of C. trachomatis include cell culture and nucleic acid amplification tests (NAATs). The major target sequences for C. trachomatis diagnosis by NAATs are located at the cryptic plasmid and the major target used for characterisation is the omp1 gene. The gold standard for diagnosis of N. gonorrhoeae is culture. However, numerous NAATs for identification of N. gonorrhoeae and a number of molecular genetic methods for characterisation of N. gonorrhoeae have been developed. Probably no routine laboratory can attain as high sensitivity by culturing C. trachomatis or N. gonorrhoeae as by using NAATs. For that reason NAATs can be recommended for diagnosing C. trachomatis, but not as the only diagnostic assay for N. gonorrhoeae, due to lack of antibiotic susceptibility testing and specificity problems, most pronounced for pharyngeal and rectal samples. Genotyping of C. trachomatis or N. gonorrhoeae provides additional information for contact tracing. It is recommended for N. gonorrhoeae, at least in low prevalence geographic areas, but cannot today be recommended for C. trachomatis. This is due to the low genetic variability and hence the limited benefits for partner notification. However, genotyping of C. trachomatis may play an important role under special circumstances.     

Improved strategies for diagnosis and treatment of osteoporosis.

OBJECTIVE: Osteoporosis is a silent epidemic that afflicts millions of postmenopausal women around the world. Osteoporosis places an enormous economic burden on society, including significant morbidity and mortality. Because of the increasing numbers of patients with osteoporosis, primary care physicians have become the front line for the diagnosis and treatment of this condition. Thus, the primary care provider should be able to diagnose osteoporosis in both asymptomatic and symptomatic women, perform a thorough workup to exclude secondary causes of osteoporosis, and optimally prevent and treat osteoporosis using the various forms of pharmacologic and nonpharmacologic therapies. DESIGN: Review of current literature and articles dealing with pathophysiology, diagnosis, and treatment of osteoporosis. RESULTS: Diagnostic and therapeutic modalities can reduce the incidence of fractures. CONCLUSIONS: The diagnosis and treatment of osteoporosis has greatly improved but needs further efforts to prevent the disease.     

DNA amplification methods for diagnosis and epidemiological investigations of avian mycoplasmosis.

This review describes some applications of DNA amplification methods for diagnosis or epidemiological investigations of avian mycoplasmosis. Tests for direct detection of pathogenic mycoplasmas have been developed. Moreover, most avian mycoplasma species can be differentiated, according to their unique restriction fragment length polymorphism (RFLP) patterns generated after digestion of PCR products with different restriction enzymes. In order to characterize isolates below the species level, PCR-based subtyping methods have been introduced. One of them, arbitrarily primed-PCR, results in strain-specific arrays of DNA fragments that can distinguish even closely related strains of a given species. This method was successfully used to investigate the molecular epidemiology of vaccine strains and of Mycoplasma gallisepticum conjunctivitis in songbirds. Major issues in the development of DNA-amplification tests concern the selection of the appropriate target, specimen collection, DNA preparation and detection of amplification reaction inhibitors. Careful consideration to the design and work flow of the facility are necessary to avoid false-positive results.     

Improved diagnosis on a daily basis of enterovirus meningitis using a one-step real-time RT-PCR assay

The detection of the enterovirus genome in cerebrospinal fluid (CSF) by PCR techniques has proved to be more sensitive than traditional cell culture for the diagnosis of enterovirus meningitis. However, PCR assays are time consuming and labor intensive, particularly if separate hybridization steps are used to confirm the specificity of positive findings. The aim of this study was to develop a one-step real-time RT-PCR assay with LightCycler (LC) technology that was sensitive, rapid, and easy to perform in routine practice. The enterovirus detection limit was determined by testing 10-fold limiting dilution series of cell culture stocks with the echovirus 25 (E-25) prototype strain and with the third European Union Quality Control Concerted Action (EU-QCCA) enterovirus proficiency panel. A total of 100 CSF specimens were investigated in a comparative study. With the E-25 strain, the detection limit of the real-time assay was 286 TCID50/ml (50% tissue culture infective dose). When samples of the EU-QCCA panel were tested, our assay gave identical results (detection limit down to 3.6 TCID50/ml) to those of the reference laboratory, which used one-step RT-PCR assay. When CSF specimens were tested, there was a correlation between the real-time assay and the conventional in-house assay in 96 of 100 CSFs tested. This one-step real-time assay allows rapid enterovirus detection in CSF since results are obtained in 3 hr as against 36 hr with the "in-house" RT-PCR assay. This new assay is now being used in routine practice, and allows diagnosis on a daily basis.     

Improved diagnosis of reactivated toxoplasmosis.

AIMS: To identify antigens detected by western blotting in primary Toxoplasma gondii infection and determine their role in diagnosis of reactivated toxoplasmosis. METHODS: Twenty three immunocompromised patients were tested by IgG western blotting. Patients were grouped retrospectively. Group 1 comprised 15 human immunodeficiency (HIV)/AIDS patients and included: group 1A (six patients with clinical and/or serological evidence of reactivation), group 1B (five patients with clinical evidence only), and group 1C (four asymptomatic patients). Group 2 comprised eight non-HIV/AIDS immunocompromised patients with clinical and/or serological evidence of reactivation. Immunocompetent patients (n = 23) with primary toxoplasmosis were a control group used to determine the progression of the antigens detected. RESULTS: In primary toxoplasmosis, antibodies against 6, 20, 22, 23, 25, 28, 29, and 36 kDa antigens predominated. Detection of four or more of the 6, 20, 22, 23, 25, and 36 kDa antigens was considered to be western blot positive. In two group 1A patients, western blotting indicated past infection. During reactivation, this reverted to being western blot positive. Three other group 1A patients were western blot positive. In three of five group 1B patients, western blot positive results improved serological diagnosis of reactivated toxoplasmosis (p < 0.05). In two of five group 1B patients and all four group 1C patients, western blot indicated past infection. In group 2, two of eight patients reverted from a pattern of past infection to western blot positive. Five other patients from group 2 were western blot positive. CONCLUSIONS: Detection of some low molecular weight antigens is diagnostic of reactivated toxoplasmosis. These antigens can be detected even with normal dye test titres and their detection improves the diagnosis of reactivated toxoplasmosis. They might be the result of the release of bradyzoites from ruptured tissue cysts.     

Introduction of software tools for epidemiological surveillance in infection control in Colombia

Introduction:

Healthcare-Associated Infections (HAI) are a challenge for patient safety in the hospitals. Infection control committees (ICC) should follow CDC definitions when monitoring HAI. The handmade method of epidemiological surveillance (ES) may affect the sensitivity and specificity of the monitoring system, while electronic surveillance can improve the performance, quality and traceability of recorded information.

Objective:

To assess the implementation of a strategy for electronic surveillance of HAI, Bacterial Resistance and Antimicrobial Consumption by the ICC of 23 high-complexity clinics and hospitals in Colombia, during the period 2012-2013.

Methods:

An observational study evaluating the introduction of electronic tools in the ICC was performed; we evaluated the structure and operation of the ICC, the degree of incorporation of the software HAI Solutions and the adherence to record the required information.

Results:

Thirty-eight percent of hospitals (8/23) had active surveillance strategies with standard criteria of the CDC, and 87% of institutions adhered to the module of identification of cases using the HAI Solutions software. In contrast, compliance with the diligence of the risk factors for device-associated HAIs was 33%.

Conclusions:

The introduction of ES could achieve greater adherence to a model of active surveillance, standardized and prospective, helping to improve the validity and quality of the recorded information.     

Molecular surveillance of rubella viruses in Taiwan from 2005 to 2011

Rubella has been listed as a mandatory notifiable disease in Taiwan since 1988. Because of high coverage rates with an effective vaccine, rubella cases have decreased dramatically in Taiwan since 1994. However, rubella outbreaks still occur due to imported transmission. Five large clusters were detected in Taiwan from 2007 to 2011. In 2007, one cluster was caused by rubella genotype 1E viruses that were imported from Vietnam, whereas another cluster was caused by genotype 2B viruses and was untraceable. In 2008, two clusters were caused by different lineages of genotype 1E viruses that were imported from Malaysia. In 2009, a cluster that was caused by genotype 2B viruses was associated with imported cases from Vietnam. The rubella viruses from 124 confirmed cases from 2005 to 2011 were characterized, and the data revealed that these viruses were distributed in the following four genotypes: 1E (n = 56), 1h (n = 1), 1j (n = 4), and 2B (n = 63). Of these viruses, 93 (75%) were associated with imported cases, and 43 of 56 genotype 1E viruses were associated with imported cases from China, Vietnam, Malaysia, and Indonesia. One genotype 1h virus was imported from Belarus, and three of four genotype 1j viruses were imported from the Philippines. Of 63 rubella genotype 2B viruses, 46 were imported from Vietnam, Thailand, Malaysia, China, Germany, and South Africa. Molecular surveillance allows for the differentiation of circulating rubella viruses and can be used to investigate transmission pathways, which are important to identify the interruption of endemic virus transmission. J. Med. Virol. 85:745–753, 2013. © 2013 Wiley Periodicals, Inc.