Mutants of human rhinovirus type 2 (HRV-2) resistant to and dependent on the antirhinoviral compound chalcone Ro 09-0410 were selected in cell culture under clean laboratory conditions. A total of 42 volunteers were challenged with either the drug-resistant mutant ,ug of amphotericin B (Fungizone) per ml, 1 mg of neomycin per ml, 50 IU of penicillin G per ml, and 100 ,ug of streptomycin per ml.The methods used for growing and determining the infectivity of human rhinovirus type 2 (HRV-2) in tissue culture were described elsewhere (3).Drug. Chalcone Ro 09-0410 (4'-ethoxy-2'-hydroxy-4,6'-dimethoxy-chalcone) is a potent inhibitor of rhinovirus uncoating (14) and was kindly supplied by Ian Lennox-Smith, Roche Research Laboratory, WeLyn Garden City, United Kingdom. It was stored as a stock solution (10 mg/ml) in dimethyl sulfoxide and diluted in maintenance medium shortly before use.Cytotoxicity and estimation of MIC. The cytotoxic concentration and MIC of Ro 09-0410 were estimated as described previously (2).Neutralization test. Neutralization tests were done as described previously (3). Briefly, doubling dilutions of serum were mixed with equal volumes of HRV-2 (100 50%o tissue culture infective doses per ml) and incubated at room temperature for 1 h. Cells from freshly dispersed sheets of Ohio HeLa cells (3.0 x 105 ml) were added to the neutralization mixture. A virus control (no serum), serum control (no virus), and cell control (no virus or serum) were included in each test. Plates were incubated at 33°C and read microscopically for cytopathic effect when virus controls showed 100%o cytopathic effect.Development of HRV-2 mutants. Human diploid MRC-5 cell monolayers were infected with 0.2 ml of the HRV-2 parental strain (10486 50% tissue culture infective doses per ml) under clean laboratory conditions in the absence of chalcone Ro 09-0410 and in the presence of serial doubling dilutions of the drug starting with a concentration of 0.128 ,ug/ml (Fig. 1). When the cells showed cytopathic effect, the supernatant was collected and 0.2 ml was used to infect new MRC-5 monolayers.

The long arm of the Y chromosome contains nonoverlapping regions termed azoospermia factor (AZF) with great influence on male fertility. Microdeletions at these regions minimise the males' ability to father offsprings. In this preliminary study, we attempted to screen the presence or absence of twenty Y chromosome's sequence-tagged sites (STS) associated with fertility in infertile and Down syndrome (DS) males. Genomic DNA from 35 fertile, 74 infertile and 22 karyotyped DS males was extracted and amplified in multiplex polymerase chain reaction (PCR) containing 20 primer pairs that amplify Y-specific STS that cover functional regions associated with AZF and spermatogenesis-related genes. Our results indicated the integrity of the Y chromosome at the 20 fertility markers for both the fertile and Down syndrome males. However, the results of the infertile males showed the presence of microdeletions at these Y-specific STS. Three samples showed Y chromosome microdeletion when blood and seminal fluid genomic DNA were assayed, while two samples showed microdeletion only when seminal fluid genomic DNA was assayed. The current study demonstrated that the molecular genetic aspect of infertility should be given proper attention when dealing with infertility cases. Furthermore, our results indicate the importance of genetic counselling in managing infertility cases.

We have isolated a human rhinovirus type-2 (HRV-2) mutant that is resistant to the antiviral agent chalcone Ro 09-0410 (4′-ethoxy-2′-hydroxy-4, 6′-dimethoxychalcone). This Ro 09-0410-resistant HRV-2 mutant (SR2-0410) exhibited altered biological properties when compared with the parental wild-type (wt) HRV-2. It was unstable when exposed to acid and heat in the presence of the drug, was incapable of producing plaques and produced an early cytopathic effect (CPE) at high temperatures (35°C and 37°C). Furthermore, compared with the parental wild-type, it showed a reduced ability to be neutralized by an anti-HRV-2 polyclonal serum and monoclonal antibodies. This SR2-0410 mutant demonstrated cross-resistance to other synthetic anti-rhinovirus compounds, which are also thought to bind to the viral capsid protein (VPI), such as 4′,6-dichloroflavan, 3-methoxy-6-[4-(3-methylphenyl)-I-piperazinyl] pyridazine (R 61837) and WIN 51711 (5-[7-[4-(4,5-dihydro-2-oxazolyl) phenoxyl] heptyl]-3-methyl-isoxazole). Furthermore, it was also resistant to various antiviral combinations synergistic against HRV-2. However, it was still sensitive to enviroxime [2-amino-I-(isopropyl sulphonyl)-6-benzimidazole phenyl ketone oxime], which has a different mode of action and is thought to interfere with viral RNA synthesis.