Contamination of environmental surfaces by genital human papillomaviruses

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  1. S Straussane,
  2. P Sastry1,
  3. C Sonnexii,
  4. Due south Edwardsthree,
  5. J Grey1
  1. 1Clinical Microbiology and Public Health Laboratory, Addenbrooke's Hospital, Cambridge, United kingdom of great britain and northern ireland
  2. 2Department of Genitourinary Medicine, Addenbrooke'southward Hospital, Cambridge, UK
  3. 3Department of Genitourinary Medicine, West Suffolk Hospital, Bury St Edmunds, UK
  1. Correspondence to:
 Dr Jim Gray, Clinical Microbiology and Public Health Laboratory, Addenbrooke's Hospital, Hills Road, Cambridge CB2 2QW, Uk: jg2{at}mole.bio.cam.air-conditioning.uk

Abstruse

Objective: To investigate contamination of environmental surfaces with human papillomaviruses (HPV) DNA in ii genitourinary medicine (Glue) clinics and in an on-site staff leisure and fitness center.

Methods: Samples were collected from the treatment rooms and patients' toilets of 2 GUM clinics situated at ii hospital sites and from the toilets of the staff leisure and fitness centre on one of the sites. Samples were tested for the presence of HPV Dna by nested polymerase concatenation reaction (PCR), and HPV amplicons were typed by reverse line hybridisation using HPV type specific oligonucleotide probes complementary to 35 HPV types. All samples were as well tested for β globin Deoxyribonucleic acid by PCR in order to assess their quality.

Results: HPV Deoxyribonucleic acid was found to be present at more 50% of the sites sampled in i of the Glue clinics, but was absent in the 2d, and also from the staff leisure and fitness heart. All HPV Dna detected was constitute to be cell associated. The most commonly establish HPV types were 6, 11, and 16, respectively. HPV infected cells were institute to be localised mainly to surfaces used predominantly past medical staff.

Conclusions: This report has identified contamination of the environment of a Mucilage clinic. Possible sources for the contamination of the clinic toilets were from genital sites via easily to the environment. Within the treatment rooms the near probable route of HPV DNA contamination of the environment was via the doctor's gloved easily.

  • human papillomaviruses
  • environmental contamination
  • polymerase chain reaction

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  • human papillomaviruses
  • ecology contamination
  • polymerase concatenation reaction

Genital human papillomaviruses (HPV) infections are spread predominantly through sexual intercourse, although other routes of transmission have been postulated,1, two and papillomaviruses may stay infectious within cells for up to 7 days, even afterward desiccation.3 HPVs infect squamous epithelial cells, and desquamated cells are regularly shed from the skin and the mucous membranes. Virus particles contained within these cells may exist transmitted betwixt sexual partners and cause productive infection in the recipient. HPV infections are the well-nigh common sexually transmitted viral infections in the United Kingdom. To date there are more than than 100 unlike HPV types distinguished on the basis of Deoxyribonucleic acid sequence homology.iv The replication cycle of HPV is heavily dependent on that of the host jail cell. This means that early stages of replication occur in basal cells, while the afterwards stages of virus replication are only seen in differentiated epithelial cells. Clinically, HPVs have been subdivided into cutaneous and mucosal HPVs according to their site of infection. Infection with HPVs may event in asymptomatic infection or the development of clinically apparent lesions such as warts—that is, a localised proliferation of epithelial cells, on hands, feet, or within the genital tract. Patients with genital warts commonly nowadays to genitourinary medicine (Mucilage) departments, and in 1999 at that place were 125 000 attendances in the United kingdom for this reason (KC 60 returns). Genital warts are predominately acquired by HPV types half-dozen and 11five although coinfection with other HPV types, in item type 16,6 is well documented.

There are a diverseness of therapeutic options for genital warts including cryotherapy and chemical treatments. Cryotherapy is a method used commonly for the removal of genital warts which may be applied by spraying liquid nitrogen onto the lesions, or by directly applying a probe cooled past liquid nitrogen. Chemical treatment with podophyllotoxin, which relies on local tissue devastation, may be self practical past the patient every bit a home treatment.

Although genital HPV types have been detected in non-genital epithelium and on non-epithelial surfaces,7– 9 the clinical significance of these findings is uncertain.

The objective of this study was to determine whether HPV DNA, and which types, were nowadays on environmental surfaces in two Gum clinics and a hospital staff leisure and fitness heart, and to place procedures associated with the risks of high levels of contamination.

MATERIALS AND METHODS

Environmental samples were collected from the treatment rooms and the male person and female person patients' toilets of Glue clinics A and B on six occasions and one occasion, respectively. The diverse sites sampled are listed in table one. Environmental samples were also collected from male person and female toilets at a hospital based staff leisure and fettle centre on i occasion. Samples were usually obtained during the working twenty-four hours, although the sampling at clinic A on the fifth and sixth occasion was performed at 8.30 am and 4.30 pm of the same day.

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Tabular array 1

Detection of HPV in ecology samples nerveless from dispensary A. (β Globin DNA could be detected in all HPV positive samples)

Samples were collected using cotton tipped swabs soaked in 0.1M phosphate buffered saline (PBS) pH 7.2. All environmental surfaces were swabbed firmly in order to pick upward any cells that may have been present and gloves were changed between each sample. Samples were assigned a number on the inflow at the laboratory in club to mask the source of each individual sample during testing. Simply afterwards the testing of all samples was performed were laboratory numbers correlated with the sample source.

Serial dilutions of CaSki cells, containing 600 copies HPV-16 DNA per jail cell (starting concentration 4 × 106 cells/ml) and RNase-free water were included as positive and negative controls, respectively. Nucleic acids were extracted from all samples and controls using the guanidinium isothiocyanate/silica procedure.10

All samples were tested for the presence of β globin Dna using a existent fourth dimension PCR (LightCycler; Roche Diagnostics, United kingdom), with the GH20/PC04 primersxi earlier they were tested for HPV Deoxyribonucleic acid, using a single tube nested real time PCR,12 in order to determine the sample quality. The presence of β globin DNA was determined by melting the PCR product in order to measure out its melting temperatures (Tm). The Tm values of the reaction products were compared to that of a positive control. A minimum of xv copies of β globin DNA could be detected using this method.

HPV Dna was detected using the LightCycler, with the MY09/MY1113 and GP5+/GP6+ primersxiv in a single tube nested PCR.12 The presence of HPV Deoxyribonucleic acid was determined by melting the PCR product in order to measure its Tm. A minimum of 10 HPV Deoxyribonucleic acid input copies could be detected through the use of the unmarried tube nested PCR.

Ii microlitres, of HPV DNA positive, nested PCR amplicons were used to repeat the 2d circular of the PCR with the GP5+ fluorophore labelled primer15 and the GP6+ primer in club to generate products for HPV typing. These PCRs were carried out on a conventional block based thermal cycler (Perkin Elmer, GenAmp 2400) as described previously.16

Reverse line hybridisation was performed every bit described by Jordens et al.fifteen Probes complementary to sequences of the L1 region of HPV types 2, 6, 11, xvi, xviii, 31, 33, 35, 39, 41, 42, 43, 45, 50, 51, 52, 53, 54, 56, 58, 59, 61, 62, 66, 67, seventy, 72, Han 831, and CP8304 were used.

RESULTS

HPV Dna was detected in environmental samples collected from Glue clinic A on more than one occasion. HPV DNA was constitute mostly associated with surfaces of treatment areas and equipment used for the assessment and treatment of patients, although HPV DNA was besides found in both the male and female person patients' toilets (table 1). β Globin Dna was detected in all HPV DNA positive samples, and was non detected in any of the HPV negative samples collected from GUM clinic A. HPV DNA was not detected in any of the samples collected from GUM clinic B or from the staff leisure and fettle centre (table 2). β Globin Deoxyribonucleic acid was non detected in whatever of the samples from the staff leisure and fitness eye and was found in only three samples from GUM clinic B (tabular array 2).

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Table two

Sites sampled in clinic B and the staff leisure and fitness middle and the detection of β globin and HPV Deoxyribonucleic acid

Tabular array three shows the accumulation of HPV types throughout GUM clinic A during a single day. At 8.30 am HPV DNA was detected only in samples from the female and male toilets, and on the surface of the cryotherapy guns. In samples collected at 4.xxx pm, HPV DNA was detected throughout the dispensary, toilets, and on the equipment used for the cess and handling of patients (Table 3).

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Tabular array 3

Accumulation of HPV DNA in GUM clinic A during one twenty-four hours. (β Globin Deoxyribonucleic acid could be detected in all HPV positive samples)

A total of 19 HPV types, all associated with genital HPV infection, were detected on ecology surfaces. HPV types 6, sixteen, and 11 were the most commonly institute types, bookkeeping for 28.6%, 24.ii%, and ix.8%, respectively, of all HPV types detected in the environment. These represent to the distribution and prevalence of HPV types identified from genital swabs of Glue clinic patients on different occasions (unpublished data).

Word

The aim of this study was to investigate the possibility of environmental contamination with genital HPV types in "high take chances" and "depression gamble" environments.

HPV infected cells were shown to be present on many of the surfaces within the handling rooms in Mucilage clinic A. The spectrum of HPV types detected was similar to that constitute in the attendees of this GUM clinic (unpublished data).

The failure to blazon 4 of the HPV amplicons may have been due to the absenteeism of a specific probe, complementary to sequences in the PCR amplicon. The probes used were specific for fewer than 50% of known HPV types, although they were called to match the spectrum of mainly genital HPV types detected in the population of the Britain.

All identified HPV types detected in the handling rooms were mucosal HPV types, ruling out the possibility of contamination from cutaneous wart virus. HPV DNA detected on the bed control panel, lite switch, and the colposcope handle were much more likely to indicate contamination from the medico'due south gloved easily afterwards he/she had examined the patient. Contamination of the examination bed and of the patients' toilets is more likely to accept arisen from the patients themselves, as these are areas with which medical and nursing staff take niggling or no contact. The toilets sampled were used exclusively by patients, and all HPV types detected were of mucosal origin indicating transfer from genital sites to the environment via hands or by direct contact. HPV DNA was detected at a higher frequency in the male toilets and then in the female person toilets of GUM clinic A, and only ii HPV types were detected in the female toilets compared to v HPV types in the male toilets, once again suggesting the transfer of HPV DNA was from the ballocks via the hands onto the environmental surfaces. Hand carriage of genital HPV Dna has been shown in a previous study, where patients with genital HPV infections were found to carry the aforementioned HPV type on their easily.9

The cleaning regimes employed in the clinics and the space bachelor for treating patients were factors associated with the presence or absence of contamination. Gum clinic A had no formal cleaning procedures in place, whereas Gum clinic B had established very rigorous and strict cleaning procedures. The patients' toilets of clinic A were cleaned in one case a twenty-four hour period past the contract cleaning staff. Treatment rooms, as well as equipment, were only cleaned at the end of each week, with the exception of the exam bed, which was cleaned at the end of each day. GUM clinic A houses two pocket-sized treatment rooms and one male and female toilet adjacent to the waiting area. GUM clinic B was very spacious, with 6 new treatment rooms, each with their ain adjacent patient toilet.

In Glue clinic A equipment used, such as cryotherapy guns, was frequently moved between treatment rooms and was never cleaned. In Glue clinic B, private cryotherapy spray guns were used less often, owing to greater utilize of self handling, and were allocated to each room and after use were wiped down with Azowipe (70% isopropyl alcohol BP, Vernon-Carus Ltd, Great britain). Interestingly, eight out of the 14 HPV types detected on the cryoguns of dispensary A were HPV types which were never detected on whatever ecology surfaces within the clinic. All these viii HPV types (43, 45, 51, 58, 59, 67, 70, Han 831) are associated with cervical abnormalities or neoplasia rather then clinically obvious lesions.

The number of clinic attendees in Gum clinic A was higher than that of clinic B, resulting in the throughput for each individual room being higher for Glue clinic A than GUM dispensary B. Out of a total of approximately 100 patients attending Gum clinic A per day up to xx had genital warts. However, in Glue clinic B a total of nineteen patients, of which only iii patients had warts and four patients attended for colposcopy, were seen on the day of sampling. On the day before sampling a total of 25 patients, of whom three had genital warts, were seen in GUM clinic B.

Samples obtained before and subsequently working hours in GUM clinic A showed an absenteeism of HPV and β globin Deoxyribonucleic acid before patients attention. This suggests degradation of cellular and HPV Dna which had contaminated surfaces on the previous working solar day.

The detection of HPV DNA does not necessarily indicate the presence of feasible virus. The lack of in vitro civilization procedures for HPV now does not let the viability of the virus to be determined. The correlation betwixt the detection of HPV Deoxyribonucleic acid and β globin DNA would suggest that only cell associated viral Deoxyribonucleic acid was recovered. Bovine papillomavirus type 1 has been shown to survive desiccation in cell extracts for upwards to 7 days.three

The quantity of virus nowadays on ecology surfaces was not adamant. However, the single tube real time nested PCR enables the detection of a minimum of 10 input copies of HPV Dna; therefore, fifty-fifty a very minor number of HPV Deoxyribonucleic acid present could have been recovered from the sites sampled in clinic A. It is possible that even if viable virus was nowadays this may be insufficient to infect an epithelial site past manus transfer. Given that the possibility of transmission via ecology surfaces is neither confirmed nor disproved, precautions should be taken to prevent the transfer of infected cells via human being "vectors." Regular cleaning of the dispensary surroundings and equipment would reduce the potential for infected cells to contaminate environmental surfaces. It is likely that the more rigorous cleaning regime of Glue clinic B, together with fewer patient attendances was the reason for the absence of detectable β globin and HPV Deoxyribonucleic acid. Gum clinic A has implemented a more than rigorous cleaning policy since we undertook this study, and nosotros promise to re-examine the effect of environmental viral contamination in the nigh future.

CONTRIBUTORS

The master author, SS, with the co-writer, PS, collected the samples, and performed the PCR and the contrary hybridisation on the environmental samples; CS supervised the sample collection in GUM clinic A and was co-author; SE supervised the sample collection in GUM clinic B and was co-author; JG supervised the projection and was senior author.

Funding was provided past the Public Health Laboratory Service for whom the Cambridge laboratory acts as the National Man Papillomavirus Reference Laboratory.

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    Sexually Transmitted Infections 2002; 78 80-lxxx Published Online First: 01 April 2002. doi: 10.1136/sti.78.ii.fourscore-a