Impact of transition from open bay to single room design neonatal intensive care unit on multidrug-resistant organism colonization rates.

BACKGROUND: The influence of the neonatal intensive care unit (NICU) design on the acquisition of multidrug-resistant organisms (MDROs) has not been well-documented. AIM: To examine the effect of single room unit (SRU) versus open bay unit (OBU) design on the incidence of colonization with MDROs and third-generation cephalosporin-resistant bacteria (3G-CRB) in infants admitted to the NICU. METHODS: Retrospective cohort study, including all infants admitted to the NICU of a tertiary care academic hospital two years prior to and two years following the transition from OBU to SRU in May 2017. Weekly cultures of throat and rectum were collected to screen for MDRO carriership. Incidence of colonization (percentage of all infants and incidence density per 1000 patient-days) with MDROs and 3G-CRB were compared between OBU and SRU periods. FINDINGS: Incidence analysis of 1293 NICU infants, identified 3.2% MDRO carriers (2.5% OBU, 4.0% SRU, not significant), including 2.3% extended-spectrum ß-lactamase-producing Enterobacterales carriers, and 18.6% 3G-CRB carriers (17% OBU, 20% SRU, not significant). No differences were found in MDRO incidence density per 1000 patient-days between infants admitted to OBU (1.56) compared to SRU infants (2.63). CONCLUSION: Transition in NICU design from open bay to SRUs was not associated with a reduction in colonization rates with MDROs or 3G-CRB in our hospital. Further research on preventing the acquisition and spread of resistant bacteria at high-risk departments such as the NICU, as well as optimal ward design, are needed.

Persistence of human papillomavirus DNA in benign and (pre)malignant skin lesions from renal transplant recipients.

An extremely diverse group of human papillomavirus (HPV) types consisting of epidermodysplasia verruciformis (EV)-associated HPV types and other cutaneous HPV types (e.g., HPV types 2 and 3) is associated with nonmelanoma cancers and benign lesions of the skin. The frequent presence of multiple HPV types in single skin biopsy specimens of renal transplant recipients prompted us to develop PCR techniques for the detection of distinct (sub)groups of genotypically related cutaneous HPV types, i.e., three subgroups of EV-associated HPV types and two groups (A2 and A4) of other cutaneous HPV types. This approach generally allowed a reliable identification of HPV genotypes by direct sequencing of the PCR products, despite the frequent occurrence of multiple infections. The targeted spectrum of HPV types comprises 66 cutaneous HPV types including 21 putative novel HPV types. We also detected 17 putative novel HPV subtypes. We demonstrated that the skin of nearly all renal transplant recipients who developed various benign and (pre)malignant skin lesions was persistently infected with one or more EV-associated HPV types and/or HPV types belonging to groups A2 and A4. The frequency and distribution of EV-associated HPV and HPV types belonging to groups A2 and A4 were similar in biopsy specimens from hyperkeratotic papillomas (77.5%), squamous cell carcinomas (77. 8%), and actinic keratoses (67.9%) but appeared to be lower in specimens of basal cell carcinomas (35.7%), benign lesions (38.5%), and clinically normal skin (32.3%). These findings suggest that renal transplant recipients are prone to persistent cutaneous HPV infection. Our data do not support the existence of high-risk cutaneous HPV types.

The prevalence of human papillomavirus DNA in benign keratotic skin lesions of renal transplant recipients with and without a history of skin cancer is equally high: a clinical study to assess risk factors for keratotic skin lesions and skin cancer.

UNLABELLED: DNA of the epidermodysplasia-verruciformis associated subgroup of HPV (EV-HPV) is frequently detected in biopsies of premalignant lesions and nonmelanoma skin cancers of renal transplant recipients. The prevalence of EV-HPVs, however, has never been systematically studied in benign keratotic skin lesions of patients with or without a history of skin cancer. This study included 42 renal transplant recipients with and 36 without a history of skin cancer. A total of 176 skin biopsies were tested for the presence of EV-HPV DNA, using a nested polymerase chain reaction (PCR). METHOD: EV-HPV typing was done by comparison of the sequence of the amplified PCR products with the sequence of all known EV-HPVs. The natural history of the development of keratotic skin lesions was studied. The number of keratotic skin lesions rapidly increased after transplantation. This increase was most pronounced in patients who developed skin cancer. The prevalence of EV-HPV DNA in benign keratotic skin lesions was equally high in patients with and without a history of skin cancer, i.e., 55 and 53% in the two groups, respectively. A large variety of EV-HPV types was found, but of these none were predominantly present in either patient groups. A higher prevalence of EV-HPV DNA was found in benign skin lesions from sun-exposed sites, but only in patients with a history of skin cancer. The association between the number of keratotic skin lesions and the development of skin cancer strongly supports the hypothesis that EV-HPVs play a role in cutaneous oncogenesis. The equally high prevalence of EV-HPV infection in patients with and without a history of skin cancer, however, may indicate that besides EV-HPV infection, other factors, such as sun exposure may also be important.

Detection of human papillomavirus DNA in plucked hairs from renal transplant recipients and healthy volunteers.

We have previously detected a group of human papillomaviruses originally found in skin lesions of epidermodysplasia verruciformis (EV) patients in skin cancers from renal transplant recipients and from non-immunosuppressed patients. The reservoir of EV-HPVs is still unknown. In the current study we investigated whether EV-HPV DNA can be detected in plucked hairs from renal transplant recipients and healthy volunteers. Hairs were plucked from eyebrows, scalp, arms, and/or legs and DNA was subsequently isolated. To detect EV-HPV, we used nested PCR with degenerate primers located in the HPV L1 open reading frame. HPV DNA was detected in hairs from one or more sites in all 26 renal transplant recipients tested. Forty-five of 49 samples (92%) from these 26 patients were positive. The HPV type was successfully determined by sequencing in 38 samples, and all types belonged to the EV-HPVs. In ten of 22 healthy volunteers (45%), EV-HPV DNA was also detected in hairs from one or more sites. Twenty of 38 samples (53%) were positive, of which 17 samples were typed as EV-HPV types. These findings indicate that EV-HPV is subclinically present in the skin of the general population. Immunosuppression may lead to activation of the virus, explaining the finding that the apparent prevalence of EV-HPV in plucked hairs from renal transplant patients is higher than in those from the volunteers. If a dose-response situation exists for the carcinogenic potential of HPV infection, this finding may be relevant to the increased risk of skin cancer in this group of patients.

p53 mutations implicate sunlight in post-transplant skin cancer irrespective of human papillomavirus status.

Mutations in p53 were detected in 11/23 (48%) of non melanoma skin cancers in renal allograft recipients and in 5/8 (63%) of sporadic tumours from immune competent patients. 9/12 (75%) of mutations in transplant patients and all 5 mutations in non transplant tumours were consistent with damage caused by ultraviolet (u.v.) irradiation. DNA sequences, predominantly of the epidermodysplasia verruciformis (EV) subgroup, were detected in 9/23 (39%) of transplant tumours and in 2/8 (25%) of eight non-transplant tumours. There was no relationship between HPV status and p53 mutation, HPV DNA being present in 5/16 (31%) of tumours with p53 mutation and 6/15 (40%) of tumours lacking p53 mutation. These data are consistent with an important role for sunlight in the development of post-transplant skin cancer, and with limited functional data suggesting that E6 proteins of the cutaneous and EV-related papillomaviruses do not target p53 for ubiquitin-mediated degradation.

High frequency of detection of epidermodysplasia verruciformis-associated human papillomavirus DNA in biopsies from malignant and premalignant skin lesions from renal transplant recipients.

Based on immunologic and epidemiologic data, it is plausible that skin cancer in renal transplant recipients is associated with human papillomaviruses (HPV). At present, conflicting evidence exists concerning the presence of HPV DNA in these cancers. We recently described a nested polymerase chain reaction method that enables the detection of all previously isolated epidermodysplasia verruciformis (EV)-associated HPVs. We now describe the detection of EV-associated HPV DNA in 49 (80%) of 61 biopsies from squamous cell carcinomas, in four (50%) of eight basal cell carcinomas, in 14 (93%) of 15 actinic keratoses, in two (40%) of five cases of Bowen's disease, and in four (57%) of seven keratoacanthomas. HPV DNA typing revealed that all detected HPV types belonged to the EV-associated HPV types. A wide spectrum of EV-associated HPVs was found, including six putative new HPV types. In a high percentage of the lesions more than one HPV type was detected. We often found the same HPV types in different skin biopsies from both malignant and premalignant lesions from the same patient. The high frequency of detection of EV-associated HPV types in biopsies from malignant and premalignant lesions is in agreement with the hypothesis that EV-associated HPVs are involved in the pathogenesis of skin cancer in renal transplant recipients.

Multifocal vulvar intraepithelial neoplasia grade III and multicentric lower genital tract neoplasia is associated with transcriptionally active human papillomavirus.

BACKGROUND: The incidence of vulvar intraepithelial neoplasia Grade III (VIN III) is increasing and is diagnosed at a younger age than previously. VIN III is often multifocal and frequently coexists with multicentric dysplastic lesions in the cervix and vagina. Warty-type VIN III more often has been found to contain human papillomavirus (HPV) DNA than basaloid-type VIN III: The authors performed HPV DNA polymerase chain reaction (PCR) analysis in 48 VIN III biopsies and reverse transcriptase (RT)-PCR in 8 HPV-16 DNA-positive multifocal VIN III biopsies to detect E6/E7 transcripts. METHODS: Human papillomavirus DNA detection and histologic analysis were performed on alternating slides of paraffin embedded biopsies. Polymerase chain reaction was performed with consensus primers, and HPV typing was performed by direct sequencing. Total RNA was isolated from frozen biopsies by centrifuging a guanidinium thiocyanate (GTC) lysate through a cesium chloride (CsCl) cushion. The RT reaction was performed using a 3' primer, located just downstream of the E7 stop codon, and the PCR reaction was performed using the same 3' primer and a 5' primer located just downstream of the E6 start codon. RESULTS: The mean age of the 48 patients was 37.7 years. Eighty-one percent had multifocal VIN III: Sixty-six percent had multicentric neoplasia. Forty-six percent of the biopsies were warty-type, 17% basaloid-type, 35% mixed-type and 2% differentiated-type. Ninety-two percent were HPV-positive and 83% contained HPV-16 DNA. Human papillomavirus DNA was more often present in multifocal VIN III lesions than in unifocal VIN III lesions and also more often in VIN III lesions coexisting with other dysplastic multicentric lesions than in unicentric VIN III lesions. Warty-type VIN III more often contained koilocytes than basaloid-type VIN III: A correlation between different morphologic forms of VIN III and the presence of HPV DNA was not found. Both types of VIN III often coexist in one lesion. In all the RT-PCRs, a 593-base-pair fragment was detected, corresponding to the expected length of the major E6*-E7 mRNA. CONCLUSIONS: The observed high prevalence of transcriptionally active HPV DNA associated with multifocal and multicentric dysplasia suggests a role of HPV in the pathogenesis of these lesions. A positive correlation between different morphologic forms of VIN III and the presence of HPV DNA was not found.

Nested PCR approach for detection and typing of epidermodysplasia verruciformis-associated human papillomavirus types in cutaneous cancers from renal transplant recipients.

The epidermodysplasia verruciformis (EV)-associated human papillomaviruses (HPVs) constitute a group of HPV genotypes isolated mostly from the cutaneous lesions of patients with the genetic disorder of EV. Broad-spectrum detection of EV HPVs in cutaneous lesions of non-EV patients was previously difficult because no EV HPV consensus PCR was available. We describe a nested PCR that enables the detection of all known EV HPV types at relatively low-copy-number levels. The deduced sequences of a 92-amino-acid stretch of the L1 open reading frames of all types are shown for convenient typing. The technique proved very valuable in viral studies of skin cancers from renal transplant recipients. A high prevalence (81%) of EV HPV types was found in skin cancer biopsies. A wide spectrum of EV HPV types that differed from HPV-5 and -8 was found to be involved. The technique also proved useful in detecting potentially novel EV HPV types in skin cancers. The relationship of these new types to known HPV types is demonstrated by phylogenetic tree analysis.

Detection of epidermodysplasia verruciformis-like human papillomavirus types in malignant and premalignant skin lesions of renal transplant recipients.

To evaluate the putative role of human papillomaviruses (HPV) in the development of skin cancer in renal transplant recipients, a series of skin biopsies from premalignant and malignant skin lesions was analysed using the polymerase chain reaction. Four different consensus primer pairs were used. HPV DNA was detected in five of 24 cases of squamous cell carcinoma, in one of three cases of Bowen's disease, in none of four basal cell carcinomas, in two of seven cases of actinic keratosis and in one of five cases of keratoacanthoma. Typing by direct sequencing of the amplified HPV DNA was possible in seven of nine cases, and revealed epidermodysplasia verruciformis (EV)-associated HPV types, or HPV types related to EV-associated types. Hence, HPV DNA could be detected in a significant proportion of (pre)malignant skin tumours in renal transplant recipients. The finding that some of the detected HPV types were as yet uncharacterized EV-related types, suggests that HPV DNA could be present in a higher percentage of lesions, and might be detected with refinement of the techniques.

Detection of cutaneous and genital HPV types in clinical samples by PCR using consensus primers.

Two sets of consensus PCR primers consisting of a common 3' primer CP-I and two 5'-primers, CP-IIG (primer set A) and CP-IIS (primer set B), in the E1 open reading frame of the human papillomavirus (HPV) genome are presented. These two primer sets enabled the detection of a 188 base pair (bp) fragment of HPV 1, 2, 3, 4, 5, 6b, 7, 8, 9, 10a, 11, 12, 14a, 16, 17, 18, 19, 20, 21, 22, 24, 25, 31, 33, 36, 37, 38, 39 and 46. HPV types 15, 23, 49 and 50 were poorly amplified and HPV type 41 was not amplified. The method is suitable for the detection of HPV DNA sequences in clinical samples of both cervical and cutaneous lesions.