Managing taste and odour metabolite production in drinking water reservoirs: The importance of ammonium as a key nutrient trigger.

Taste and odour (T&O) compounds (most commonly 2-MIB and Geosmin) in drinking water are becoming an increasingly global problem for water management. Here, the trigger(s) for 2-MIB and Geosmin production were investigated in Plas Uchaf reservoir (North Wales, UK) with detailed water sample analysis between 2015 and 2016. Historical abstraction data from this reservoir and 4 reservoirs in Somerset (England, UK) were compared statistically using Self-Organising Map (SOM) analysis. In-reservoir measurements (2015-2016) revealed an 85% reduction in ammonium from the primary external loading source led to lower 2-MIB and Geosmin concentrations, with peak concentrations of 2-MIB declining from 60 to 21  ng l-1 and Geosmin declining from 140 to 18  ng l-1. No other measured water chemistry parameter showed a significant difference between years. The SOM results support the in-reservoir findings, revealing 2-MIB and Geosmin to be associated with high ammonium relative to nitrate for all 5 reservoirs. We conclude that ammonium is key for stimulating cyanobacterial productivity and production of T&O compounds. Whilst it is well understood that adequate availability of phosphorus is required for rapid growth in cyanobacteria, and hence should still be considered in management decisions, we suggest that monitoring sources and concentrations of ammonium is key for managing T&O outbreaks in drinking water reservoirs.

Modelling treated waste disposal in Port Phillip Bay and Bass Strait biogeochemical and physical removal.

Large cities, such as Melbourne, generate substantial quantities of sewage, which, after treatment, must be disposed. Melbourne's sewage is disposed via two routes, that treated at the Western Treatment Plant (WTP) is discharged in enclosed Port Phillip Bay, while the Boags Rock outfall empties into the exposed Bass Strait. In Port Phillip Bay biogeochemical processes control the fate of waste, while in the Bass Strait physical mixing rapidly disperses the waste. These different processes require very different ecosystem models. Port Phillip Bay requires detailed modelling of water-column and in-sediment processes, in particular detailed models of recycling processes, and also modelling of benthos-water-column interactions. Interaction of these components gives the model a nonlinear response to change in load. The Bass Strait ecosystem model is simple with no modelling of the sediment and limited modelling of water-column recycling. This model's behaviour is largely controlled by the physical environment.

Characterisation of a sperm coating auto-antigen reacting with antisperm antibodies of infertile males using monoclonal antibodies.

OBJECTIVE: In a previous study a number of sperm-specific antigens were identified which reacted with antisperm antibodies from both infertile and vasovasostomised males. To investigate the localisation and distribution of these antigens and their role in male fertility, monoclonal antibodies were raised against them; immunoblotting techniques were used to select only those antibodies which competed with human antisperm antibodies for these human auto-antigens. DESIGN: One antibody, NW21, reacted with an 18 kDa auto-antigen present on epididymal sperm but absent from testicular sperm. Immunohistochemical studies showed that the antigen is produced in small basal cells between the columnar epithelium of the corpus epididymis, passes up into the tubule and then coats sperm passing along the epididymis. Sperm stored in the cauda epididymis and ductus deferens stain strongly for this sperm coating glycoprotein. CONCLUSIONS: The localisation of this antigen supports the suggestion that auto-immune infertility may represent a response to epididymal rather than testicular sperm. Monoclonal antibodies raised to unique and immunologically accessible sperm coating proteins, produced in the epididymis rather than in the testis, would seem to present an excellent theoretical solution to male contraception.

Unilateral testicular obstruction: orchidectomy or reconstruction?

Unilateral testicular obstruction can cause infertility in man by reducing sperm output, or by stimulating antisperm antibody production. Amongst 125 subfertile men with unilateral blocks, the causes were defined as post-infective (n = 72), post-traumatic (n = 41) or congenital (n = 12) by surgical exploration. Of these, 50 had severe oligozoospermia or azoospermia, and another 30 had oligozoospermia; 67 had significantly raised serum antisperm antibody titres. The outflow tracts were reconstructed in 87 men, while in 24 the obstructed testis was removed and replaced with a prosthesis. Follow-up information is available in 104 men. Reconstruction produced significant improvement in sperm concentration in the ejaculate but little change in antisperm antibody titres. Orchidectomy lowered seminal plasma antibody titres and had no effect on sperm output. Orchidectomy plus prednisolone was most effective in terms of pregnancies produced in the female partners. Testicular biopsies showed that spermatogenesis was normal on the side of the block in 90% of cases, whereas it was impaired in the contralateral testis in 40% of men. Although sperm output can be improved by reconstruction, the stimulus to antibody production is more reliably eliminated by orchidectomy. Sperm concentration, antisperm antibody titres and bilateral testicular biopsy results should be taken into account before making a final therapeutic recommendation in men with unilateral testicular obstruction.

The effects on in-vitro fertilization of autoantibodies to spermatozoa in subfertile men.

Thirty-six infertile couples underwent treatment by in-vitro fertilization. In 16 couples (group 1) the male partner was positive for antisperm antibodies measured by direct mixed antiglobulin reaction, direct immunobead test, and serum and/or seminal plasma tray agglutination test. In 20 couples (group 2) the men had no such antibodies. Men with poor sperm motility were excluded. The female partners had no antisperm antibodies, and in the controls (group 2) infertility was due to a known female factor. The fertilization rate in couples without antisperm antibodies (group 2) was 72.7% compared to 50.5% when the men had antibodies. However, the pregnancy rate per embryo transfer was not significantly different in the two groups (46.1% in group 1, 33.3% in group 2). This indicates that antisperm antibodies in the male interfere with sperm--egg fusion and subsequent fertilization but once fertilization has occurred, the pregnancy rate remains the same.

Comparison of mixed antiglobulin reaction and direct immunobead test for detection of sperm-bound antibodies in subfertile males.

OBJECTIVE: To compare two methods of detection of surface bound antibodies on spermatozoa from subfertile males. DESIGN: Prospective comparison of direct mixed antiglobulin reaction (MAR) for immunoglobulin (Ig)G with direct immunobead test (IgG and IgA) applied to spermatozoa from male partners of infertile couples. Circulating unbound antibody measured by tray agglutination test in serum and seminal plasma in a representative proportion. SETTING: Seminology laboratory. PATIENTS: One hundred nine male partners of infertile couples. RESULTS: Highly significant correlation between direct MAR (IgG) and direct immunobed test (IgG) and between both of these tests and serum unbound antibody measured by tray agglutination test. Highly significant correlation between direct immunobead test (IgA) and seminal plasma unbound antibody measured by tray agglutination test, but no correlation with MAR (IgG). CONCLUSIONS: Mixed antiglobulin reaction (IgG) is a cheap, quick, and sensitive screening test, but immunobead test (IgA) provides useful additional information on class of antibody on spermatozoa that may be clinically more important.

The concentrations of transferrin, beta 2-microglobulin, and albumin in seminal plasma in relation to sperm count.

A number of studies have shown a relationship between transferrin levels in seminal plasma and sperm count. In the present study, the levels of transferrin, beta 2-microglobulin and albumin were measured in 171 samples of seminal plasma (50 subjects with normal sperm count; 32 with azoospermia; 49 with oligozoospermia; 10 with polyzoospermia; and 31 vasectomized). All three proteins were related to sperm count. It is concluded that there is a general relationship between seminal plasma proteins and sperm numbers and that some products of the sperm themselves must control the entry of plasma proteins into seminal plasma.

Testicular obstruction: clinicopathological studies.

Genital tract reconstruction has been attempted in subfertile men with obstructive azoospermia (370 patients) or unilateral testicular obstruction (80 patients), and in vasectomised men undergoing reversal for the first (130 patients) or subsequent (32 patients) time. Histopathological changes in the obstructed testes and epididymes, and immunological responses to the sequestered spermatozoa have been studied to gain insight into possible causes of failure of surgical treatment. The results of surgery have been assessed by follow-up sperm counts and occurrence of pregnancies in the female partners. The best results were obtained with vasectomy reversal (patency 90%, pregnancy 45%), even after failed previous attempts (patency 87%, pregnancy 37%). Epididymovasostomy gave good results with postinfective caudal blocks (patency 52%, pregnancy 38%), while postinfective vasal blocks were better corrected by total anatomical reconstruction (patency 73%, pregnancy 27%) than by transvasovasostomy (patency 9%, no pregnancies). Poor results were obtained with capital blocks (patency 12%, pregnancy 3%), in which substantial lipid accumulation was demonstrated in the ductuli efferentes; three-quarters of these patients had sinusitis, bronchitis or bronchiectasis (Young's syndrome). There is circumstantial evidence to suggest that this syndrome may be a late complication of mercury intoxication in childhood. After successful reconstruction, fertility was relatively reduced in those men who had antibodies to spermatozoa, particularly amongst the postinfective cases. Similarly, impaired fertility was found in men with unilateral testicular obstruction and antibodies to spermatozoa. Mononuclear cell infiltration of seminiferous tubules and rete testis was noted occasionally, supporting a diagnosis of autoimmune orchitis; although rare, this was an important observation as the sperm output became normal with adjuvant prednisolone therapy.

Influence of antisperm antibodies on human sperm function.

Antisperm antibodies have been found in about 8% of men with infertility and in 60 to 80% of patients following vasectomy. In order to investigate the way antibodies influence sperm function we studied serum and seminal plasma from patients with infertility (n = 61) or undergoing vasovasostomy (n = 25). These antisera were characterised to determine their TAT titre, the nature of the target antigens and their capacity to interfere directly with fertilisation. The results indicate that antibodies from both groups of patients exhibit a capacity to stimulate or suppress sperm/oocyte fusion. The proportion of samples showing stimulatory activity was higher (50%) in the vasovasostomised population than in patients with infertility (21%). The remainder of the antisera suppressed sperm/oocyte fusion. There was no correlation between the titre of antisperm antibodies and their capacity to influence sperm function, indicating that it is the nature of the target antigens which is of significance rather than the antibody concentration. Western blot analysis indicated that these antisera targeted a group of sperm surface antigens with molecular weights of 30kD (35,45,66,90 and 115kD). Monoclonal antibodies are now being prepared in order to determine which of these specific components are involved in the suppression of sperm function.

Characterization of sperm antigens reacting with human antisperm antibodies.

Antibodies reacting with human spermatozoa have been detected by various immunological techniques in the sera of subfertile men. Different patterns of sperm agglutination are observed with different sera, either head-to-head, tail-to-tail, or tail-tip-to-tail-tip. Differences have been detected between the clinically relevant antibodies in spontaneously infertile males and the less important antibodies in males who have undergone reversal of vasectomy. It has been suggested that the variations in agglutination patterns are due either to different classes of antibody or to binding of antibody to different antigens. In the present study immunoblotting techniques were used to characterize the reactivity of solubilized sperm proteins with serum samples exhibiting different modes of sperm agglutination. This involved the electrophoretic transfer of proteins from SDS gels to nitrocellulose sheets followed by overlay with serum antibody. Using these techniques we have attempted to characterize the antigens of spermatozoa which react with sera from both spontaneously infertile and vasovasostomized men. The results showed that although antisperm antibodies bind to discrete and sperm-associated antigens, there is no substantial difference between the antigenic patterns observed with antibodies producing different types of sperm agglutination. Neither the antigens detected, nor the intensity of reaction showed significant differences although there was a tendency for head-to-head agglutinating antibodies to react more strongly with the higher molecular weight antigens. Moreover, although with sequential serum samples the patterns of agglutination may change, the antigenic pattern remains unchanged.

Effect of cyclosporin A in male autoimmune infertility.

Nine infertile men (mean duration of infertility, 8.3 years) with antisperm antibodies were treated with the immunosuppressive drug cyclosporin A for 6 months with a dose of 5 to 10 mg/kg/day. Seminal plasma and serum antisperm antibodies fell in three subjects on treatment, and sperm count and motility increased substantially in one. Three successful pregnancies occurred in the study group: one on treatment, one in the first cycle of artificial insemination with the husband's sperm after treatment (twin infants), and one 3 months after cessation of treatment (twin infants). Successful conceptions with cyclosporin were unrelated to falls in antisperm antibody titer.

Cyclic prednisolone therapy for male infertility associated with autoantibodies to spermatozoa.

Seventy-six subfertile men with significant titers of antisperm antibodies were treated with a new corticosteroid regimen, consisting of prednisolone, 40 mg daily, rising to 80 mg daily if antibody titers did not fall, given from days 1 to 10 of the partner's menstrual cycle, for up to nine cycles. Twenty-five (33%) of the partners became pregnant during a treatment cycle, more than twice the expected incidence without treatment. No serious complications occurred, although one half of the patients had transient minor side effects. This regimen appears to be encouraging and suitable for further assessment in a prospective controlled trial.

The clinical relevance of classes of immunoglobulins on spermatozoa from infertile and vasovasostomized males.

Following reversal of vasectomy, conceptions occur even when antisperm antibodies are present in the seminal plasma, but this is most unusual in men with similar titers of such antibodies who are spontaneously infertile. To clarify the differences between antisperm antibodies occurring in infertile men and those associated with vasectomy reversal, we have studied 23 spontaneously infertile men and 22 men who underwent vasectomy reversal, all of whom had antisperm antibodies detected in seminal plasma by the same tray agglutination test. The class of antibody on spermatozoa was defined by a double-antibody technique using diluted rabbit anti-human IgG, IgM, or IgA or secretory component, followed, after washing, by 125I-labeled donkey anti-rabbit Ig. The results have shown that similar amounts of IgG and IgM were present on the spermatozoa, but infertile men had significantly more IgA and especially more secretory component than men who underwent vasectomy reversal. This was associated with significantly greater impairment of penetration of cervical mucus in the former group. It appears that the type of antibody on the spermatozoa may vary according to the stimulus for its production.

PIP: To clarify the differences between antisperm antibodies occurring in spontaneously infertile men and in men who have undergone reversal of vasectomy, classes of immunoglobulins were compared in 23 men from the former group and 22 men from the latter group. Antisperm antibodies were detected in seminal plasma by the same tray agglutination test in all subjects. The class of antibody on spermatozoa was defined by a double-antibody technique that used diluted rabbit antihuman IgG, IgA, or secretory component, followed, after washing, by 125I-labled donkey antirabbit Ig. Although similar amounts of IgG amd IgM were present on the spermatozoa in both groups, infertile men had significantly more IgA and especially more secretory component than men who underwent vasectomy reversal. This was associated with significantly greater impairment of penetration of cervical mucus in the former group. 8 of the 20 vasovasectomized men with antisperm antibodies in their seminal plasma produced pregnancies, whereas no pregnancies have been observed in 63 spontaneously infertile untreated men with seminal plasma antibodies. Spermatozoa from 86% of vasovasectomized men penetrated the cervical mucus, while spermatozoa from only 27% of spontaneously infertile men were able to do so. It is concluded that the type of antibody on the spermatozoa may vary according to the stimulus for its production.

Exploratory scrototomy in 168 azoospermic males.

In 168 azoospermic males with normal or only slightly raised serum FSH levels, serum antisperm antibodies were measured, and the site of obstruction or the nature of the failure of spermatogenesis was defined by exploratory scrototomy with inspection of epididymes, vasography and testicular biopsy. When possible, surgical reconstruction was done by side-to-side epididymovasostomy, with vasovasostomy when necessary using 6/0 Prolene and no splints. Acquired blocks of cauda epididymis (34 cases) and vas (23 cases) were significantly more commonly associated with serum antisperm antibodies than congenital bilateral absence of vasa (29 cases) or blocks at the caput epididymis (48 cases), most of which were associated with sinusitis, bronchitis or bronchiectasis (Young's syndrome). Many of the former patients came from abroad, whereas most of the latter came from the British Isles. Sperm counts of 10 million per ml or more were produced by 23 (45%) of 51 adequately followed up patients with acquired blocks following surgical reconstruction, and 11 pregnancies (21.5%) were produced. Amongst those with spermatozoa in the ejaculate after surgery, serum antisperm antibodies were found significantly less often in those whose wives became pregnant compared with those who failed to produce pregnancies. It is concluded that failure of surgical treatment in some of these cases may have an immunological basis. No success was achieved with other groups.

The effects of sperm antibodies on fertility after vasectomy reversal.

Antisperm antibodies have been measured in serum and in seminal plasma in 130 males before and after vasectomy reversal, and the occurrence of pregnancy has been analyzed in those partners who were trying to produce a pregnancy. All patients have been followed for at least one year. Sperm-agglutinating antibodies were found in the serum of 79% of patients; seminal plasma antibodies were present in only 9.5% before reversal, and this rose to 29.5% afterwards. Overall, pregnancies occurred in the partners of 44.6% of those men who were trying to produce children. Production of pregnancy was significantly less likely when the preoperative serum antisperm antibody titer was 512 or more, but no decrease in fertility was seen with titers below this. Similar numbers of pregnancies were produced by patients with or without seminal plasma antibodies in titers of up to 16; there are too few patients with titers above this level to permit further analysis. A randomized controlled trial of perioperative steroids showed that they produced no benefit. It appears that the antisperm antibodies associated with vasectomy reversal may differ fundamentally from those occurring in naturally subfertile males.

The diagnosis of unilateral testicular obstruction in subfertile males.

Thirty-two subfertile males with sperms in the ejaculate and unilateral testicular obstruction are reported: the diagnosis was established by exploration of scrotum in 26, clinically in 2, 3 had had previous partially successful epididymovasostomies, and 1 had had an epispadias repair. The past medical history gave relevant information in 27 (84%), and useful findings were made on clinical examination in a further 3 cases. Fifteen patients had sperm counts over 20 million per ml, and 15 were less than 10 million per ml. Twenty-six (81%) had serum antisperm antibodies detected by tray agglutination test (TAT), 21 (81%) of whom had evidence of head-to-head (HH) agglutinins in pure or mixed form. Comparison with 162 vasectomised males and 160 naturally infertile males with antisperm antibodies showed that 55% of the former and 24% of the latter had HH agglutinins on TAT, differences that were highly significant on statistical analysis. Evidence of obstruction was found in 14 (37%) of 38 naturally infertile males with antisperm antibodies and HH or mixed agglutination, but only in 12 (10%) of 122 with TT agglutinins: this difference was also highly significant. Clinical history, physical examination and serum antisperm antibodies, especially if HH agglutinins are present, can suggest the possibility of unilateral testicular obstruction, but confirmation of the diagnosis requires exploration of scrotum.

Reversal of vasectomy: the effects of sperm antibodies on subsequent fertility.

Antisperm antibodies were measured in serum and seminal plasma in 130 males before and after vasectomy reversal and the occurrence of pregnancy was analysed in the partners of 77 who were followed for more than one year. Sperm-agglutinating antibodies were found in the serum of 79% of patients; seminal plasma antibodies were present in only 9.5% before reversal and this rose to 26% afterwards. Pregnancies occurred in the partners of 53% of those men who were trying to produce children. A pregnancy was significantly less likely when the pre-operative serum antisperm antibody titre was 512 or more, but no decrease in fertility was seen with titres below this. Several pregnancies were produced by patients with seminal plasma antibodies, but numbers and follow-up are too small to permit detailed analysis. A randomised controlled trial of per-operative steroids showed that they produced no benefit. The antisperm antibodies associated with vasectomy reversal appear to differ fundamentally from those occurring in naturally subfertile males.

The results of intermittent high dose steroid therapy for male infertility due to antisperm antibodies.

Forty-five males who had been infertile for 2 to 10 (average 5.3) years, with significantly positive antisperm antibody tests (serum spermagglutination titers more than 32) and demonstrably improved sperm penetration of cervical mucus, were treated with repeated 7-day courses of methylprednisolone (MP) 32 mg three times a day from days 21 to 28 (in a few cases days 1 to 7) of their wives' menstrual cycles. Antisperm antibody tests were repeated after treatment and related to the occurrence of pregnancy in the wife. Fourteen wives (31%) became pregnant in a cycle following treatment of the husband. The production of pregnancy was always associated with a marked drop in sperm immobilizing titer and usually with disappearance of antibodies from seminal plasma; both of these effects were sometimes observed without a significant change in serum spermagglutination titers. Three patients (6%) had transient severe side effects, and 14 (26%) have had mild side effects, out of 54 patients treated so far.