Implementation of pooled saliva tests for universal screening of cCMV infection.

Congenital cytomegalovirus (cCMV) is the most common intrauterine infection, leading to neurodevelopmental disabilities. Universal newborn infant screening of cCMV has been increasingly advocated. In the absence of a high-throughput screening test, which can identify all infected newborn infants, the development of an accurate and efficient testing strategy has remained an ongoing challenge. Here we assessed the implementation of pooled saliva polymerase chain reaction (PCR) tests for universal screening of cCMV, in two hospitals of Jerusalem from April 2022 through April 2023. During the 13-month study period, 15,805 infants (93.6% of all live newborn infants) were screened for cCMV using the pooled approach that has since become our routine screening method. The empirical efficiency of the pooling was six (number of tested newborn infants per test), thereby sparing 83% of the saliva tests. Only a minor 3.05 PCR cycle loss of sensitivity was observed for the pooled testing, in accordance with the theoretical prediction for an eight-sample pool. cCMV was identified in 54 newborn infants, with a birth prevalence of 3.4 per 1,000; 55.6% of infants identified with cCMV were asymptomatic at birth and would not have been otherwise targeted for screening. The study demonstrates the wide feasibility and benefits of pooled saliva testing as an efficient, cost-sparing and sensitive approach for universal screening of cCMV.

Saliva Real-Time Polymerase Chain Reaction for Targeted Screening of Congenital Cytomegalovirus Infection.

BACKGROUND: Saliva real-time polymerase chain reaction (PCR) was shown to be sensitive and specific for the detection of congenital cytomegalovirus (cCMV) in universal screening studies. In the current study, we assessed the performance of saliva real-time PCR in newborns undergoing targeted cCMV screening. METHODS: Saliva real-time PCR results were prospectively correlated with reference-standard urine detection in newborns undergoing targeted cCMV screening over a 3-year period, in successive validation (concurrent testing of all saliva and urine specimens) and routine-screening (confirmatory urine testing of positive saliva results) implementation phases. RESULTS: The sensitivity, specificity, and positive and negative predictive values of saliva real-time PCR were 98.3% (95% confidence interval, 90.8%-99.9%), 91.5% (89.3%-93.3%), 45.6% (36.7%-54.7%), and 99.9% (99.2%-99.9%), respectively, in 856 concurrently tested newborns. True-positive saliva real-time PCR detection (defined in relation to urine detection) was associated with earlier saliva sampling (P = .002) and a higher saliva viral load (P < .001). We further identified a saliva viral load cutoff value that reliably distinguished between true-positive and false-positive saliva results. CONCLUSIONS: In newborns undergoing targeted screening for cCMV, saliva real-time PCR is highly sensitive yet has a low positive predictive value, necessitating confirmatory testing. Early sampling and application of a validated viral load cutoff could improve the assay performance and support its large-scale implementation in this growing clinical setting.

HPV genotype distribution among women with normal and abnormal cervical cytology presenting in a tertiary gynecology referral Clinic in Ethiopia.

BACKGROUND: Cervical cancer is the second most prevalent cancer among women of child-bearing age in Ethiopia. The aim of this study was to determine human papilloma virus (HPV) genotype distribution among HIV-negative women with normal and abnormal cervical cytology results. METHODS: We investigated a consecutive of 233 HIV-negative women between December 2008 and March 2009 presenting in a Tertiary Gynecology Referral Clinic in Ethiopia. Screening was done by Pap cytology and HPV detection and genotyping method was nested PCR (direct amplification with MY09/MY11 primers, followed by nested amplification with GP5/GP6 primers) and sequencing of the nested products. Sequencing of the non-purified nested PCR products was performed following re-amplification with Big dye terminator, using the GP6 primer. RESULTS: Of the 233 study participants, 92 (39.5%) had abnormal cytology. All women with abnormal cervical cytology had positive HPV DNA compared to only 48.9% of those presenting with normal cytology. Of these women, the frequency of high-risk (HR)-HPV was 83.2% and its prevalence in women with abnormal cervical cytology was significantly higher than those with normal cytology (92.4% vs. 71%, p < 0.0001). The most frequent genotypes identified were HPV16 (44.1%), followed by HPV35 and HPV45 (each 6.2%), HPV31 (4.4%), HPV56 (3.7%), HPV18 and HPV59 (each 3.1%), HPV58 (2.5%) and HPV39 (1.9%). While the most common HR-HPV infections among women with normal cytology were HPV16 (20.3%), followed by HPV35 (8.7%), HPV56 and HPV58 (each 5.8%), HPV18, HPV31 and HPV39 (each 4.4%), HPV45 (2.9%) and HPV59 and HPV68 (each 1.5%), the most common HR-HPV infections in women with abnormal cytology included HPV16 (62%), followed by HPV45 (8.7%), HPV 31, HPV35 and HPV59 (each 4.4%), and HPV18, HPV52 and HPV56 (each 2.2%). We also noted low prevalence of multiple HPV infections in women with normal or abnormal cytology. Multivariable logistic analysis showed that residing in rural area (OR 3.24, 95% CI: 1.13-9.30), being multipara (OR 7.35, 95% CI: 1.78-30.38) and having abnormal cervical cytology results (OR 6.75, 95% CI: 1.78-25.57) were all independently associated with HPV16 genotype. CONCLUSIONS: Our study revealed a significant risk of infection with HR-HPV, in particular with HPV16 genotype, in women attending a referral center in Ethiopian women presenting with or without abnormal cervical cytology. Moreover, Pap smear cytology missed a significant proportion of women compared to those who were identified by PCR for HR-HPV infections. In addition, the PCR method we used was not suitable for sensitive detection of co-existent multiple infections. Data from the present study indicate that currently available HPV vaccines could prevent nearly 67% of all cervical cancer cases in women in Ethiopia.

Efficacy of pulsed dye laser treatment for common warts is not influenced by the causative HPV type: a prospective study.

Verruca vulgaris (VV) is a prevalent skin condition caused by various subtypes of human papilloma virus (HPV). The most common causes of non-genital lesions are HPV types 2 and 4, and to a lesser extent types 1, 3, 26, 29, and 57. Although numerous therapeutic modalities exist, none is universally effective or without adverse events (AE). Pulsed dye laser (PDL) is a favorable option due to its observed efficacy and relatively low AE rate. However, it is not known which verrucae are most likely to respond to PDL, or whether the causative viral subtype influences this response. The objective of this prospective blinded study was to assess whether the HPV subtype was predictive of response to PDL. For that matter, 26 verrucae from 26 immunocompetent patients were biopsied prior to treatment by PDL. HPV coding sequences were isolated and genotyped using PCR analysis. Patients were treated by PDL (595 nm wavelength, 5 mm spot size, 1.5 ms pulse duration, 12 J/cm2 fluence) once a month for up to 6 months, and clinical response was assessed. Binary logistic regression analysis and linear logistic regression analysis were used in order to evaluate statistical significance. Different types of HPV were identified in 22 of 26 tissue samples. Response to treatment did not correlate with HPV type, age, or gender. As no association between HPV type and response to PDL therapy could be established, it is therefore equally effective for all HPV types and remains a favorable treatment option for all VV.

Diagnosis of Cytomegalovirus Infection during Exacerbation of Ulcerative Colitis.

BACKGROUND/AIMS: The role of cytomegalovirus (CMV) reactivation during exacerbations of ulcerative colitis (UC) is yet a matter of debate, and assessment of CMV infection in UC patients remains an ongoing challenge. We aimed to identify associated parameters and compare detection methods for CMV infection during UC exacerbation. METHODS: Clinical, pathological and virological parameters were retrospectively analyzed in all patients hospitalized in our institution for UC exacerbation between January 2009 and April 2015, who underwent full evaluation for CMV infection in colonic tissue by histopathology, immunohistochemistry (IHC) and CMV-PCR. RESULTS: Of 28 patients who underwent full examination for tissue CMV-infection, 13 (46.4%) were found to be positive for CMV. Tissue CMV-PCR was more sensitive for the detection of CMV infection than histopathology and IHC. CMV-positive patients had a statistically higher frequency of recent steroid treatment and fever, with higher mean partial Mayo scores and lower mean albumin levels. There were no significant differences between CMV-positive and CMV-negative patients in terms of age, severity of colitis and disease duration. In a multivariable model, only recent steroid treatment and fever were independently associated with colonic CMV infection. CONCLUSIONS: This study provides a clinical model to detect the presence of CMV infection in patients hospitalized with UC exacerbation, which could direct proper investigation and facilitate timely empirical therapy.

High rate of cytomegalovirus drug resistance among patients receiving preemptive antiviral treatment after haploidentical stem cell transplantation.

We examined the rate, clinical impact, and risk factors of cytomegalovirus (CMV) drug resistance in 561 patients who underwent 616 hematopoietic stem cell transplantations (HSCTs) over 5 years. Drug resistance was exclusively identified in haploidentical (haplo)-HSCT recipients receiving preemptive therapy, among whom the rate was 14.5%. Resistance appeared after prolonged treatment (median, 70 days), was associated with higher preceding viral load (P < .001), and was the strongest predictor for disease by multivariate analysis. The high rate of drug resistance as interlinked with severe disease in haplo-HSCT recipients suggests the potential advantage of prophylactic over preemptive treatment in high-risk patients and highlights the need for better-tolerable anti-CMV drugs.

Human cytomegalovirus kinetics following institution of artesunate after hematopoietic stem cell transplantation.

The anti-malaria drug artesunate has been shown to be an effective inhibitor of cytomegalovirus (CMV) in vitro, in an experimental animal model, and in a recent single-case clinical use. In this first case-series of 6 stem cell transplant recipients who received preemptive artesunate treatment for CMV infection, we have examined the viral kinetics following institution of artesunate, and employed first-phase viral kinetics studies to calculate its antiviral effectiveness. Two patients demonstrated a rapid 0.8-2.1 log viral load decline by 7 days, with a viral decay half-live of 0.9-1.9 days. Four patients demonstrated a continued yet stalled viral growth slope during treatment. No adverse events were noted in treatment courses of up to 28 days. Overall, a divergent antiviral efficacy was revealed, ranging from 43% to 90%, which appeared to be primarily dependent on the virus baseline growth dynamics. Further dose escalation studies are needed to examine the role of artesunate in the treatment of CMV infection in the transplantation setting.

Artesunate as a potent antiviral agent in a patient with late drug-resistant cytomegalovirus infection after hematopoietic stem cell transplantation.

This is the first report of treatment of cytomegalovirus infection with artesunate, for a stem cell transplant recipient with a newly identified foscarnet-resistant and ganciclovir-resistant DNA polymerase L776M mutation. Artesunate treatment resulted in a 1.7-2.1-log reduction in viral load by treatment day 7, with a viral half-life of 0.9-1.9 days, indicating a highly effective block in viral replication.