Effects of antimicrobial therapy on sperm-mucus interaction

Sperm-mucus interaction under in-vitro or in-vivo conditionsmight be affected by microorganisms colonizing the reproductivetract. In order to study the influence of antimicrobial therapy,an extensive microbial screening was performed including Chlamydiatrachomatis, Mycoplasma hominis, Ureaplasma urealyticum, Neisseriagonorrhoeae, a broad spectrum of potentially pathogenic aerobicand anaerobic bacteria, Trichomonas vaginalis, herpes simplexvirus and yeasts. One-hundred-and-six couples with a mean durationof infertility of 5.5 years (range 1–12 years) and withisolation of potentially pathogenic microorganisms in semensamples and/or cervical swabs were submitted to a prospectivepilot study. None of the patients displayed signs or symptomsof infection in the lower genital tract. Before and after specifictherapy, based on antimicrobial susceptibility testing, spermanalyses and in-vitro sperm penetration meter tests (SPMT) (Kremer)were performed. SPMT was evaluated with cervical mucus of patients'wives, collected after a standardized oral treatment with oestrogensand, additionally, in a crossed manner with cervical mucus andspermatozoa of fertile donors. The success of antimicrobialtherapy was controlled by repeating the same mkrobial screeningand was 96%. However, there was a marked change in the mkrobialpattern. A comparison of the results of sperm analyses beforeand after treatment revealed neither significant differencesfor sperm volume, sperm count, propulsive motility, morphology,vitality, pH, fructose concentration or number of round cells,nor was there a significant influence on the cervical indexand the number of leukocytes in cervical mucus. In the SPMTpenetration distance, sperm density, quality and duration ofmotility were not significantly influenced by antimicrobialmedication, but were improved in the group of patients withvery poor SPMT results before treatment. The overall pregnancyrate was low (17% after 6 months, 21% after 12 months). Theresults suggest that microbial colonization is of minor importancefor sperm-mucus interaction and that the benefits of antimicrobialtherapy in asymptomatic couples are limited.     

Chlamydial serology in 1303 asymptomatic subfertile couples

The clinical significance of antichlamydial antibodies (Chlam Ab) was determined in a total of 1303 subfertile couples consulting for infertility investigation and treatment. Median age of the women was 30 (range 22-44) years and of the men 33 (range 21-53) years. The median duration of infertility was 4 (range 1-21) years. All patients were asymptomatic for genital tract infection. A comprehensive infertility investigation included examination of the endocrine, cervical, and tubal factor, and semen analysis, antisperm antibody (ASA) testing, sperm-mucus interaction testing in vitro using a standardized protocol, and post-coital testing (PCT). Screening for Chlam IgG Ab was performed in serum of both partners, obtained at the same time. Simultaneous microbial cultures in genital secretions of both partners included a broad spectrum of potentially pathogenic bacteria. Elevated titres of Chlam IgG Ab as seromarker for previous infection were found in 20.8% of all women, and in 12.6% of men. Chlam Ab were significantly more frequent in partners of seropositive patients (in 51.8% of women with a Chlam Ab positive partner, compared to 15.8% of the other women). Microbial screening outcome was not significantly related to results of chlamydial serology in both partners. In women, elevated titres of Chlam Ab were significantly associated with a tubal factor, but were not related to reduced quality of the endocervical mucus (CM), including the in-vitro penetrability of the CM (using partners' or donors' spermatozoa). In males, Chlam Ab were not significantly related to the outcome of semen analysis, including screening for ASA (IgG and/or IgA) in semen, and several parameters of sperm functional capacity. After exclusion of couples with tubal disease, subsequent male fertility did not significantly differ in males with or without Chlam Ab. The results suggest that during basic infertility investigation, positive chlamydial serology as an easy screening procedure indicates a higher risk for a tubal infertility factor. However, in asymptomatic patients, Chlam IgG Ab in serum are not associated with a cervical factor or with the male factor, using several determinants for evaluation of semen quality including subsequent fertilizing capacity.      

Circulating antisperm antibodies and fertility prognosis: a prospective study

In a prospective study performed on 235 couples with long standinginfertility (median 5 years), circulating antisperm antibodies(ASA) were determined with both a standard tray-agglutinationtest (TAT) and a radloimmunoassay (RIA). Serum levels were correlatedwith results of sperm analysis (SA), postcoital testing (PCT)and in-vitro sperm-cervical mucus penetration testing (SCMIPT),performed with cervical mucus and spermatozoa of patients andin parallel with material of fertile donors. A simultaneousmicrobial screening included Chiamydia trachomatis, mycoplasmas,Herpes simplex virus, gonococci, other potentially pathogenicaerobic and anaerobic bacteria and yeasts. The pregnancy rate12 months after determination of ASA in the serum samples was27% (64/235). Between ASA-positive and -negative male and/orfemale patients (TAT as well as RIA), no significant differencewas found for any of the tested variables of SA, PCT, crossedSCMPT and colonization of genital secretions with microorganisms.Patients who later achieved pregnancy and those who did notdid not differ with regard to serum antibody status (TAT aswell as RIA). The findings suggest that the clinical significanceof circulating ASA determined with the currently available methodsis low. ASA in serum samples of infertile female andlor malepatients do not influence the fertility prognosis.     

Cytomegalovirus infection and viral shedding in the genital tract of infertile couples

The prevalence of cytomegalovirus (CMV) infection and viral shedding in infertile couples in Taiwan and its role in infertility were studied. Two hundred fifty couples were enrolled in this study. Anti-CMV IgG was measured in serum samples from these infertile couples. Viral shedding examined by DNA hybridization was detected in semen from the husband and cervical mucus from the wife. Anti-CMV IgG was detected in 249 (99.6%) of the 250 male serum samples and in 247 (98.9%) of the 250 female serum samples. Viral shedding was detectable in 83 (33.5%) of 248 semen samples and 83 (33.7%) of 246 cervical mucus samples by dot-blot DNA hybridization assay. Semen quality was not apparently affected by the existence of viral shedding. The co-shedding rate in semen and acervical mucus was high (15.9%). It is concluded that the seroprevalence and genital tract viral shedding were relatively high in infertile couples in Taiwan. Viral shedding did not affect the semen quality. Nevertheless, screening of donor semen is recommended. © 1995Wiley-Liss, Inc.     

Seminal antibodies to human 60kd heat shock protein (HSP 60) in male partners of subfertile couples

BACKGROUND: Heat shock proteins (HSP) are essential mammalian and bacterial stress proteins. At the cellular level, they act as chaperones, have important regulatory functions, and are considered to be an essential factor for reproduction. Scarce information exists on the role of sensitization to HSP and the potential role in the aetiology of male infertility. METHODS: The potential association of immunoglobulin (Ig)A antibodies (Ab) to the human 60 kDa heat shock protein (HSP 60) with several parameters of subclinical male genital tract infection/inflammation and with semen quality and sperm fertilizing capacity was analysed in a prospective study. IgA Ab to human HSP 60 were determined in seminal plasma of 202 randomly chosen male partners of subfertile couples with a median duration of infertility of 4 years (range 1-15 years), who were asymptomatic for genital tract infection. After medical history and clinical examination, a comprehensive evaluation of semen quality, in aliquots of the same ejaculates used for HSP Ab determination, included: sperm analysis; local antisperm antibody (ASA) screening; standardized sperm-cervical mucus (CM) penetration testing; immunocytochemical round cell differentiation to determine seminal leukocyte counts; evaluation of complement fraction C(3) and of some pro-inflammatory cytokines; and microbial screening. Subsequent fertility was recorded after 6 months. RESULTS: The presence of HSP 60 IgA Ab in seminal fluid (total positive 6.9%) was significantly associated with leukocytospermia, the presence of C(3), and also with high interleukin (IL) levels in seminal plasma. HSP 60 Ab were not related to the bacterial colonization of ejaculates. There was no association of seminal IgA Ab to human HSP 60 with semen quality, determined with microscopical semen analysis, nor with local IgG- or IgA-class ASA. There was no relationship with sperm intrinsic motility and duration of motility in the sperm CM-penetration test, nor with sperm fertilizing capacity. CONCLUSIONS: The combined presence of IgA Ab to human 60 kDa HSP, leukocytes and other established infection/inflammation markers in semen might suggest a potential role of the immune response to heat shock proteins (HSP) in cases of silent male genital tract infection, but the results do not indicate a marked relationship of HSP 60 Ab in seminal fluid with standard parameters of semen quality.     

Influence of microbial colonization on sperm-mucus interaction in vivo and in vitro*

Two-hundred-and-thirty-three asymptomatic couples with a meanduration of infertility of 5 years were submitted to postcoitaltesting (PT) and to sperm penetration meter test (SPMT) andsimultaneous microbial screening. Cervical swabs and semen specimenswere collected for culture of Mycophma homink, Ureaphma urealyticum,Chlamydia tmhomufis, Neisseria gonorrhoeae, other potentiallypathogenic and commensal aerobic and anaerobic bacteria, herpessimplex virus, vaginal swabs for Trichomonas vaginalis and yeasts.Results of microbial screening were analysed with regard tosperm penetration ability into wives' cervical mucus in vivoand in vitro, but no marked influence was revealed for mostmicmrganisms. Samples of only one of the 233 couples provedto be completely sterile. The findings suggest that in asymptomaticpatients microbial colonization is of minor importance for sperm-mucusinteraction.     

Human papillomavirus infection among sexual partners attending a Sexually Transmitted Disease Clinic in Rio de Janeiro, Brazil

Cervical cancer is a major source of illness and death among women worldwide and genital infection with oncogenic human papillomavirus (HPV) its principal cause. There is evidence of the influence of the male factor in the development of cervical neoplasia. Nevertheless, the pathogenic processes of HPV in men are still poorly understood. It has been observed that different HPV types can be found among couples. The objective of the present study was to investigate HPV infections in female patients (n = 60 females/group) as well as in their sexual partners and to identify the concordance of HPV genotypes among them. By using the polymerase chain reaction, we detected a 95% prevalence of HPV DNA in women with cervical intraepithelial neoplasia (CIN) compared to 18.3% in women with normal cervical epithelium, with a statistically significant difference (P < 0.001). The HPV DNA prevalence was 50% in male partners of women with CIN and 16.6% in partners of healthy women. In the control group (healthy women), only 9 couples were simultaneously infected with HPV, and only 22.2% of them had the same virus type, showing a weak agreement rate (kappa index = 0.2). Finally, we observed that HPV DNA was present in both partners in 30 couples if the women had CIN, and among them, 53.3% shared the same HPV type, showing moderate agreement, with a kappa index of 0.5. This finding supports the idea of circulation and recirculation of HPV among couples, perpetuating HPV in the sexually active population, rather than true recurrences of latent infections.     

Anaerobic bacteria commonly colonize the lower airways of intubated ICU patients

Objectives  To investigate respiratory tract colonization by aerobic and anaerobic bacteria in mechanically ventilated patients.
Methods  Bacterial colonization of the stomach and the respiratory tract was qualitatively and quantitatively analyzed over time in 41 consecutive mechanically ventilated patients in a Swedish intensive care unit (ICU), with special emphasis on elucidation of the role of anaerobic bacteria in the lower respiratory tract. Samples were taken from the oropharynx, gastric juice, subglottic space and trachea within 24 h (median 14 h) of intubation, and then every third day until day 18 and every fifth day until day 33.
Results  The patients were often heavily colonized with microorganisms not considered to belong to a healthy normal oropharyngeal and gastric flora on admission to the ICU. A majority harbored enterococci, coagulase-negative staphylococci and Candida spp. in at least one site on day 1. Anaerobic bacteria, mainly peptostreptococci and Prevotella spp., were isolated from subglottic and/or tracheal secretions in 59% of the patients. Different routes of tracheal colonization for different groups of microorganisms were found. Primary or concomitant colonization of the oropharynx with staphylococci, enterococci, enterobacteria and Candida was often seen, while Pseudomonas spp., other non-fermenting Gram-negative rods and several anaerobic species often primarily colonized the trachea, indicating exogenous or direct gastrointestinal routes of colonization.
Conclusions  Mechanically ventilated patients were heavily colonized in their lower airways by potential pathogenic microorganisms, including a high load of anaerobic bacteria. Different routes of colonization were shown for different species.     

Sexually transmitted diseases and Chlamydia trachomatis in women consulting for contraception.

To study the frequency of genital infections in women consulting their family doctor for contraception, 248 women (median age 23 years) were examined for a range of genital microorganisms. The prevalence of clue cells, Candida albicans and Trichomonas vaginalis were 21.0%, 12.9% and 2.0%, respectively. Neisseria gonorrhoeae was isolated in only one case, whereas Chlamydia trachomatis was found in 6.3% of women. A specific clinical picture for an infection with C. trachomatis in women was not seen. Given the prevalence of over 5% for C. trachomatis and the absence of typical signs and symptoms in infected women, screening for this organism is recommended in women requesting an intrauterine contraceptive device, to prevent complications such as pelvic inflammatory disease and their sequelae.     

Isolation of Mycoplasma pneumoniae from the human urogenital tract.

Mycoplasma pneumoniae is a common etiologic agent of lower respiratory tract infections in humans. However, it has been reported previously that the organism has occasionally been isolated from sites other than the oropharynx and respiratory tract. We report the isolation of 24 strains of M. pneumoniae from urogenital specimens obtained from 22 female patients. Most isolates were of cervical origin from patients attending several local gynecological clinics over a 2-year period. Strains were also isolated from the urethra of one of three healthy male sexual partners of female patients positive for the organism. Single serum specimens obtained from three female patients and three different male sexual partners showed antibody levels suggestive of either recent respiratory infection or genital tract colonization with M. pneumoniae. Although there is no apparent definitive explanation for the localized outbreak of the organism at these unusual sites, the possible transfer through sexual and/or orogenital contact remains the most likely mode of transmission. The occurrence of an organism with obvious pathogenicity for human epithelial tissue in the urogenital tract suggests such transfer could play a role in genital tract infection.     

Evaluation of Immunoglobulin A1 (IgA1) Protease and IgA1 Protease-Inhibitory Activity in Human Female Genital Infection with Neisseria gonorrhoeae

Immunoglobulin A1 (IgA1) protease, an enzyme that selectively cleaves human IgA1, may be a virulence factor for pathogenic organisms such as Neisseria gonorrhoeae. Host protection from the effects of IgA1 protease includes antibody-mediated inhibition of IgA1 protease activity, and it is believed that the relative balance between IgA1 protease and inhibitory antibodies contributes to the pathogenesis of disease caused by IgA1 protease-producing organisms. We have examined the levels of these two opposing factors in genital tract secretions and sera from women with uncomplicated infection with N. gonorrhoeae. When IgA1 in cervical mucus was examined by Western blotting, no evidence of cleavage fragments characteristic of IgA1 protease activity was seen in gonococcus-infected or control patients. Cleavage fragments typical of IgA1 protease were detected, however, after the addition of exogenous IgA1 protease to cervical mucus. Degraded IgA1 was detected in some vaginal wash samples, but the fragment pattern was not typical of IgA1 protease activity. All N. gonorrhoeae isolates from the infected patients produced IgA1 protease in vitro. All but two serum samples and 16 of 65 cervical mucus samples displayed inhibitory activity against gonococcal IgA1 protease, but there was no significant difference in the level of inhibitory activity between gonococcus-infected and noninfected patients in either cervical mucus or serum. There was no difference in the levels of IgA1 protease-inhibitory activity in serum or cervical mucus collected from patients at recruitment and 2 weeks later. These results suggest that cleavage of IgA1 by gonococcal IgA1 protease within the lumen of the female lower genital tract is unlikely to be a significant factor in the pathogenesis of infections by N. gonorrhoeae.     

Chronic asymptomatic genital tract infection and semen quality.

Chronic asymptomatic genital tract infection has been postulated to cause infertility in man. This study demonstrates the correlation of positive seminal fluid bacteriological cultures with abnormal parameters of seminal fluid analysis in the male partners of 100 uninvestigated couples presenting with infertility. Gram negative bacteria were cultured only in patients with semen abnormalities while Gram positive bacteria were cultured from both normal and abnormal semen specimens.     

The microbiological and immunological characteristics of disbiotic disturbances of the biotopes of the mucose membranes of the respiratory and urogenital tracts

The study consisted in comparison of the microecology and the parameters of the humoral immunity of the biotopes of the respiratory and urogenital tracts under normal conditions and in inflammatory processes caused by bacterial agents. The study revealed significant changes in the microbiocenosis of the respiratory and urogenital tracts, caused by decline of indigenous microflora and increase of the content of conditionally pathogenic microorganisms. These changes depended on the degree of the infectious process severity. Measurement of the humoral immunity parameters revealed a significant correlation between the content of conditionally pathogenic microorganisms and the levels of M and A immunoglobulins, as well as secretory IgA and free secretory component (sc) in the vaginal secretions of patients with non-specific inflammatory diseases of the genital tract, as well as G and A immunoglobulins, secretory IgA, and sc in the saliva of patients with chronic inflammatory diseases of the respiratory tract.     

Variations of the Amnionless gene in recurrent spontaneous abortions

Recurrent spontaneous abortions (RSA) are estimated to affect 0.5-1% of couples trying to have a child. The causes of RSA are unknown in the majority of cases. This study aimed to determine whether homozygous mutations in the AMN gene in a fetus cause spontaneous abortions in humans, as they are known to cause spontaneous abortions in mice. The study was conducted by screening 40 couples and 5 women with three or more unexplained spontaneous abortions for heterozygous mutations in the AMN gene using denaturing high-performance liquid chromatography. Altogether, 3 exonic and 11 intronic sequence variations were found. There were no significant differences in the frequencies of the variations between the patients and a control group. One of the exonic variations was non-synonymous, and three of the variations may affect gene splicing. None of the putative phenotype-affecting variations were found in both partners in any couple. These results indicate that RSA in the couples studied cannot be explained by homozygous AMN mutations in the fetus. However, two couples had different, potentially deleterious variations in both partners. If these variations have a phenotypic effect, the RSA experienced by these couples may be caused by mutations in the AMN gene. In addition, birthplaces of the patients' ancestors revealed some clustering, suggesting that some patients may carry a founder mutation in another gene which may contribute to RSA.     

Pregnancy: Seminal white blood cells and recurrent abortion

The incidence of leukocytospermia (>1 million white bloodcells/ml semen) and concentrations of white blood cell populationsin semen from men whose partners experience unexplained recurrentspontaneous abortion by conventional testing criteria are describedand compared to those of men whose partners have normal reproductivehistories. Comparable seminal granulocyte and total white bloodcell concentrations between groups suggest that couples withunexplained recurrent abortion do not have an abnormally highincidence of subclinical genital tract infections for whichleukocytospermia could serve as a marker. However, a significantlyhigher prevalence of elevated CD4⁺ and CD8⁺ T lymphocyte concentrationsin semen from men whose partners have cellular immunity to spermantigens (i.e. produce embryotoxic factors in response to spermantigen stimulation) warrants further investigation.     

Seminal tract infections: impact on male fertility and treatment options

Bacterial and viral infections of the genital tract may be important aetiological factors for male infertility. Infectious processes may lead to deterioration of spermatogenesis, impairment of sperm function and/or obstruction of the seminal tract. Detection of bacteria in semen does not necessarily signify infection since bacteriospermia may represent contamination, colonization or infection. Reported prevalence of Ureaplasma urealyticum in human semen varies from 10 to 40%. Enterobacteria can even be found in up to 90% of semen samples depending on the sensitivity of detection methods used. Chlamydia trachomatis is the most frequent sexually transmitted bacterial organism in industrialized countries. It is suggested that its main influence is due to sexual transmission resulting in tubal disease and subsequent infertility in the female partner rather than a direct influence on male reproductive functions. The effect of leukocytospermia on male fertility is controversial. This is probably due to different detection methods, different populations studied and to the fact that leukocyte subtypes in semen may have different functions. In addition to potentially negative effects, leukocytes may even have protective effects on spermatozoa. Only recently have amplification methods been established to detect viruses in semen with high sensitivity and specificity. It is unclear if these infections significantly contribute to male infertility.     

Determination of sperm antibodies in human genital secretions by an ELISA technique

Sera and genital secretions of 178 infertile men and 40 infertilewomen were evaluated for antibodies against spermatozoa by anenzymelinked immunosorbent assay (ELISA). It was shown thatcirculating sperm antibodies are not usually transudated intogenital secretions and, on the other hand, that local antibodiesfound in cervical mucus or seminal plasma are not detectablein serum. Furthermore it was demonstrated that the solubilizationof cervical mucus by bromelain almost completely removes antibodyactivities detectable by ELISA, whereas sonkation of mucus doesnot affect the immunoglobulins. Sonication should thus be appliedfor liquefaction of cervical mucus in order to assess its antibodyproperties.     

Chlamydia trachomatis in subfertile women undergoing uterine instrumentation: an alternative to direct microbial testing or prophylactic antibiotic treatment

Chlamydia trachomatis is the major cause of tubal occlusion, and is also associated with IVF failure and spontaneous abortion. These infections are asymptomatic in most individuals and can persist in the genital tract for long periods of time in a form resistant to immune destruction. A significant percentage of couples seeking treatment for infertility might, therefore, harbour C. trachomatis in their genital tract. An unresolved question is what to do about this possible chlamydial persistence. Cervical, endometrial and semen samples can be tested for C. trachomatis and only positive individuals treated. Alternatively, all couples undergoing infertility treatment can receive prophylactic antibiotics. We advocate a third option, to screen and treat only individuals who are positive for systemic and/or local anti-chlamydial antibody production. Detection of species-specific C. trachomatis antibodies in peripheral blood will determine which individuals have been exposed to this organism and who, therefore, may be at risk for harbouring persistent forms. Identification of IgA antibodies in genital tract secretions may be an even better indicator of the presence of C. trachomatis in the genital tract. Circulating antibodies to the chlamydial 60kDa heat shock protein (hsp60) is a specific indicator of tubal occlusion and, furthermore, correlates with the continued presence of this micro-organism in the genital tract of non-human primates. Screening for both cervical IgA antibodies to C. trachomatis and serum IgG anti-chlamydial hsp60 appears to provide the best indication as to which women may be harbouring C. trachomatis.     

Cytokine expression pattern in the genital tract of Chlamydia trachomatis positive infertile women - implication for T-cell responses

Human genital infection caused by Chlamydia trachomatis is thought to be immunologically mediated, resulting in local recruitment of lymphocyte subsets and inducing the production of cytokines. Little information is available about the role of lymphocyte recruitment and the regulation of cytokine production in the genital tract of C. trachomatis positive infertile women. We have evaluated the recruitment of lymphocyte subsets in the genital tract and production of Th1/Th2 cytokines in cervical secretions and laparoscopic specimens from the fallopian tubes of C. trachomatis positive infertile women (n = 17) and compared them with controls, viz. C. trachomatis negative infertile women (n = 20) using ELISA and flow cytometry. None of these patients were found to be infected either with Candida sps., bacterial vaginosis, Trichomonas vaginalis, Neisseria gonorrhoeae, Mycoplasma hominis or Ureaplasma urealyticum in the cervix. Flow cytometric analysis of cervical secretions in Chlamydia positive women revealed recruitment of both CD4 and CD8 lymphocytes to the genital tract was up-regulated and a variation in the production rates of different cytokines in cervical secretions and fallopian tube was observed. We found that the immune responses in cervical secretions were of Th0 type, since all the analysed cytokines, viz. IFN-gamma, TNF-alpha, IL-10 and IL-12 were up-regulated. As, both CD4 and CD8 cells contribute to the production of IFN-gamma and IL-10, these results suggest that along with CD4 cells, CD8 lymphocytes also may be important for local regulation of Th1/Th2 responses in the genital tract during C. trachomatis infection.     

The case for routine HIV screening before IVF treatment: a survey of UK IVF centre policies

The case for routine human immunodeficiency virus (HIV) screening of all couples seeking assisted reproductive treatment is so strong that it should be made obligatory for all couples entering IVF programmes to be given information about HIV transmission, and offered testing. In August 1999, questionnaires regarding routine HIV screening of couples seeking IVF treatment were sent to the medical directors of the 74 licensed assisted conception units in the UK. Of the 45 (60.8%) centres who responded, 19 (42.2%) routinely screen both partners for HIV antibodies, 25 (55.5%) do not screen and one centre selectively screens high-risk patients. There was no significant difference in the proportion of centres that routinely carried out screening with regards to the unit size: six out of 13 (46.2%) small units compared with 13/32 (40.6%) large units. In all, 17 centres (37.8%) rated HIV screening as essential, nine (20%) as desirable, 11 (24.4%) as not required, while eight (17. 8%) centres did not comment. Of the 19 centres that have a routine screening policy, 18 have management protocols in the event that the test is positive. Of these 18 centres, 12 adhere rigidly to the protocol, while five centres adhere to the protocol with few exceptions and the remaining one uses its protocol for guidance only. The main reasons for not employing routine HIV screening were: the lack of cost effectiveness, low prevalence of HIV infection in their population, necessity for and cost of counselling, uncertainty about the need for screening and potential delay to start of treatment.