Identification of Streptococcus gallolyticus subsp. macedonicus as the etiological agent in a case of culture-negative multivalve infective endocarditis by 16S rDNA PCR analysis of resected valvular tissue.

Today, PCR using broad-range primers is being used increasingly to detect pathogens from resected heart valves. Herein is described the first case of multivalve infective endocarditis where 16S rDNA PCR was used to detect a single pathogen from two affected valves in a 61-year-old man. Triple heart valve replacement was required despite six weeks of appropriate antimicrobial therapy. The organism was confirmed as Streptococcus gallolyticus subsp. macedonicus, a member of the 'S. equinus/S. bovis' complex. To date, only one report has been made of human infection due to this organism. This may be due to the limited resolution of the routine diagnostic methods used and/or as a consequence of the complex nomenclature associated with this group of organisms.

Human papilloma virus genotype distribution and risk factor analysis amongst reproductive-age women in urban Gambia.

PURPOSE: Cervical cancer is the most frequently diagnosed female cancer in The Gambia, representing approximately 30 % of cases. In 2014, the quadrivalent human papilloma virus (HPV) vaccine was introduced, which offers protection against HPV genotypes 6, 11, 16 and 18. To evaluate the potential effectiveness of this vaccine, genotype distribution and risk factor analysis were assessed. METHODOLOGY: Endocervical samples (n=232) were collected from women aged 20-49 years residing in urban Gambia. A questionnaire was administered to capture socio-demographic and cervical cancer risk factors. HPV detection and genotyping was performed by PCR amplification of the L1 major capsid gene and analysis of sequenced PCR products.Results/Key findings. The prevalence of HPV was 12 % (28/232), and the high-risk (HR) genotype HPV 52 (5/28) was the most prevalent genotype. HR-HPV sequences had high identity (≥90 %) to isolates which originated from America, Europe and Asia but not from Africa. Half (14/28) of participants were co-infected with Ureaplasma urealyticum/parvum, which increases the risk of progression to cervical cancer. Female genital mutilation and the use of hormone contraception for >5 years were identified as potential risk factors for HPV infection. Ethnicity-associated differences were also noted; participants of the Fula ethnic group had a higher prevalence of HR-HPV infection (31.3 %) compared to the Mandinka (18.8 %) and Wollof (12.5 %) groups. CONCLUSION: These data may have a significant public health impact as the HPV quadrivalent vaccine may be of limited value if the circulating non-HPV 16/18 HR-genotypes are responsible for cytological abnormalities of the cervix.

A consensus on fungal polymerase chain reaction diagnosis?: a United Kingdom-Ireland evaluation of polymerase chain reaction methods for detection of systemic fungal infections.

The limitations of classical diagnostic methods for invasive fungal infections (IFIs) have led to the development of molecular techniques to aid in the detection of IFIs. Despite good published performance, interlaboratory reproduction of these assays is variable, and no consensus has been reached for an optimal method. This publication describes the first multicenter study of polymerase chain reaction methods, for the detection of Aspergillus and Candida species, currently used in the UK and Ireland by distribution and analysis of multiple specimen control panels. All three Candida methods were comparable, achieving a satisfactory level of detection (10 cfu), and the method of preference was dependent on the requirements of the particular laboratory. The results for the five Aspergillus assays were more variable, but two methods (2Asp and 4Asp) were superior (10(1) conidia). Formally, the overall performances of the two Aspergillus assays were comparable (kappa statistic = 0.77). However, on the Roche LightCycler, there was a clear sample-type effect that greatly reduced the detection limit of the 4Asp method when testing whole blood samples. Therefore, the preferred Aspergillus method relied on the amplification platform available to the user. This study represents the initial process to achieve a consensus method for the diagnosis of IFIs.

A two-tube combined TaqMan/SYBR Green assay to identify mycobacteria and detect single global lineage-defining polymorphisms in Mycobacterium tuberculosis.

We have developed a novel real-time PCR assay to identify and perform preliminary genotyping of mycobacteria in a manner tailored to our local service. Within a single thermocycler run, mycobacterial 16S rDNA and the Mycobacterium tuberculosis global lineage-defining RD750 polymorphism are targeted in separate reaction tubes, each of which includes both TaqMan and SYBR Green chemistries. The results of this 16S-RD assay differentiate M. tuberculosis complex (MTBC) from nontuberculous mycobacteria (NTM) and recognize whether or not MTBC isolates belong to the East African-Indian lineage, the single most frequently isolated global MTBC lineage in our service. If required, NTM amplicons may be sequenced to provide more specific identities. We report the technical performance of this assay on 88 mycobacteria-positive cultures and discuss its use in the initial management of mycobacterial infections. The 16S-RD assay correctly identified all 70 MTBC-positive cultures and 17 NTM-positive cultures while contemporaneously recognizing 26 MTBC isolates as within and 44 outside the East African-Indian lineage. In artificial samples, the combined assay also showed limited potential to detect mixed mycobacterial infections (MTBC/NTM) and tuberculosis infections involving more than one global MTBC lineage. The approach we have established can be readily tailored to targets of particular value for any mycobacterial diagnostic service, thereby optimizing the value of the results for local clinical and public health management of mycobacterial infections.

Identification of Neisseria gonorrhoeae as the causative agent in a case of culture-negative dermatitis-arthritis syndrome using real-time PCR.

Sexually transmitted infections (STIs) are an increasingly common and important cause of a fever in a returning traveler. Systemic complications of STIs, human immunodeficiency virus seroconversion illness, and secondary syphilis are diagnoses that can easily be missed. We present a case of culture-negative disseminated gonococcal infection presenting with fever, malaise, polyarthralgia, arthritis, and a rash that developed following orogenital contact and was diagnosed using real-time polymerase chain reaction. This technology has major potential to improve the speed and sensitivity of diagnosis and consequent management of patients with this syndrome.