Phenol ammonium sulphate basic fuchsin staining of sputum in pot for the detection of acid-fast bacilli.

BACKGROUND & OBJECTIVES: Improper practices of making direct smears of sputum for detection of acid-fast bacilli (AFB) and of disposing sputum cups are hazardous. The present study was undertaken with the objective to stain sputum samples in their containers by 'phenol (10%) ammonium sulphate (4%) basic fuchsin (2%) solution' and to decolourize and counterstain their smears for detection of AFB- (henceforth called pot method) and to compare the smear results of pot method with the standard Ziehl-Neelsen (ZN) method. METHODS: A total of 575 selected sputum samples from pulmonary tuberculosis patients were stained by the standard ZN and pot methods and the proportions of AFB positive smears were compared. RESULTS: Of the 575 samples, 126 were AFB positive for both the staining methods and the difference was not statistically significant. Pot method missed 9 ZN positive smears (8 scanty and one 1+) and ZN method missed 9 pot positive smears (9 scanty) and the difference was not significant. High grade smears (3+) were seen more in pot method (42) than in ZN method (25) and the difference was significant. INTERPRETATION & CONCLUSION: Our findings showed that pot method was comparable to standard ZN method and had many advantages. Pot method can be explored further for the detection of AFB in sputum samples obtained from pulmonary tuberculosis suspects.

Application of lot sampling of sputum AFB smears for the assessment of microscopy centres.

SETTING: Designated microscopy centres (DMC) and additional microscopy centres (AMC) performing sputum acid-fast bacilli (AFB) microscopy, the District TB Centre (DTC) and a reference laboratory (RL). OBJECTIVES: To ascertain the feasibility of adopting lot sampling of AFB smears and to assess the performance of MCs employing Senior Tuberculosis Laboratory Supervisors (STLS) with no knowledge about the principles of quality assurance of AFB microscopy and RL-based laboratory technicians with training on quality assurance for blinded checking of AFB smears. METHODS: Slides from MCs were transported to the DTC and the RL; 20 smears per month per MC were selected systematically; 1547 slides from DMCs and 726 from AMCs were checked, respectively, by STLSs at the DTC and by RL laboratory technicians. Discrepancies were resolved by referee. RESULTS: The discrepancy between MC laboratory technicians and STLSs at the DTC was 4.7%, compared to 1% at the RL. The STLSs and RL-based laboratory technicians had 70 and 2 errors, respectively. CONCLUSIONS: Lot sampling of AFB smears is feasible under field conditions. Assessment of MCs was more valid with RL-based technicians trained in principles of quality assurance of sputum AFB microscopy than with STLSs with no such training and working in the field.

Storage of heat-fixed unstained sputum AFB smears for panel testing in a tuberculosis unit in South India.

To evaluate the suitability for panel testing of heat-fixed unstained sputum AFB smears stored for up to 10 months, panels of slides were prepared at the national laboratory and stored under ambient conditions. Every month, three slides were utilised for panel testing in each of 12 microscopy centres; 70 smears were checked in a blinded fashion after 10 months. Reading errors occurred in 15/360 slides used in panel testing and in 4/70 slides used in blinded checking. The quality and grading of heat-fixed unstained smears were unaffected for up to 10 months and were found suitable for panel testing.

Lot quality assurance sampling of sputum acid-fast bacillus smears for assessing sputum smear microscopy centers.

Assessment of 12 microscopy centers in a tuberculosis unit by blinded checking of eight sputum smears selected by using a lot quality assurance sampling (LQAS) method and by unblinded checking of all positive and five negative slides, among the slides examined in a month in a microscopy centre, revealed that the LQAS method can be implemented in the field to monitor the performance of acid-fast bacillus microscopy centers in national tuberculosis control programs.

Blinded rechecking of sputum smears for acid-fast bacilli to ensure the quality and usefulness of restaining smears to assess false-positive errors.

SETTING: Microscopy Centres, Velliyur Tuberculosis Unit, Tiruvallur District, Tamil Nadu, India. PROCEDURES: Twelve microscopy centre laboratory technicians examined 41978 direct sputum smears for acid-fast bacilli. Senior Tuberculosis Laboratory Supervisors (STLS) checked all positive smears (4696) and 10% of negative smears (4776) in an unblinded fashion as per Revised National Tuberculosis Control Programme (RNTCP) guidelines. Ten per cent of the positive and negative slides and another 10% of unchecked negative slides were selected systematically for blinded rereading at the Tuberculosis Research Centre (TRC); 422 slides were reread without and with restaining. RESULTS: Unblinded checking by STLS of the smears read by the laboratory technicians yielded 95 to 100% agreement. Blinded rereading at the TRC revealed that false-negative errors were greater among the laboratory technicians (2-7%) than the STLS (0-3%). Restaining and blinded rereading of slides reduced false-positive errors from 27% to 7%. CONCLUSIONS: Blinded rereading at the reference centre facilitates assessment of laboratory technicians and STLS. Restaining before rereading the smears was found to be useful for precise estimation of false-positive errors.

Cobalt-catalyzed one-pot three-component coupling route to beta-acetamido carbonyl compounds: a general synthetic protocol for gamma-lactams.

An efficient improved procedure for the synthesis of beta-acetamido carbonyl compounds is developed by a cobalt(II) chloride-catalyzed three-component coupling protocol. The procedure is also amenable to the synthesis of gamma-lactams by a three-component coupling reaction with use of 2-carbomethoxybenzaldehyde. The beta-acetamido carbonyl compounds derived from 2-carbomethoxybenzaldehyde are useful intermediates as they can be transformed to the corresponding gamma-lactams on treatment with base.

Reverse turn induced pi-facial selectivity during polyaniline-supported cobalt(II) salen catalyzed aerobic epoxidation of N-cinnamoyl L-proline derived peptides.

A novel chemo- and diastereoselective aerobic epoxidation of the N-cinnamoyl peptides catalyzed by polyaniline-supported cobalt(II) salen (PASCOS) is described. The N-cinnamoyl proline derived peptides 1 show a high pi-facial selectivity during these epoxidations. The origin of this diastereoselectivity in 1 has been attributed to (i) the propensity of the N-cinnamoyl proline amide to exist predominantly as trans rotamer in CDCl3, DMSO-d6, and CH3CN medium and (ii) existence of these peptides as organized structures (gamma- and beta-turns) due to the presence of intramolecular hydrogen bonds. An extensive solution NMR and MD simulation study on 1d and 1f indicates that the origin of the high pi-facial selectivity is due to the well-defined gamma- and beta-turns which result in the hindrance of one face of the cinnamoyl double bond in the transition state of the epoxidation reaction.

Synthesis of small cyclic peptides via reverse turn induced ring closing metathesis of tripeptides.

A reverse turn induced (gamma/beta-turn) cyclization of tripeptides 1 can be performed in a ring closing metathesis reaction with Grubbs' catalyst to the corresponding cyclic peptides 2. These cyclic peptides may be useful probes as a conformationally constrained mimic of the bioactive conformation of structurally related HIV protease inhibitors.

Washing of new microscopic glass slides in dichromate solution does not influence sputum AFB smear results.

SETTING: Microscopy centres in Tiruvallur District, Tamil Nadu, India, implementing DOTS. OBJECTIVE: To know whether washing new glass slides in dichromate solution is essential for effective sputum acid-fast bacilli microscopy. METHODS: Two direct smears were prepared from each of 1750 sputum samples. One was made on dichromate solution-cleaned new glass slides and the other was made on unwashed new glass slides. The smears were blinded and examined. RESULTS: Of the 1750 specimens, 13.5% and 13.08% were positive for AFB using washed and unwashed slides, respectively (P = 0.12). The concordance between these two (including one grade above and one below) was 98.7%. CONCLUSION: Washing of new glass slides in dichromate solution is not essential for AFB microscopy.

Nonconventional carbon additions to azomethines. aryl amination/indoline synthesis by direct aryl radical addition to azomethine nitrogen.

[reaction: see text]. The generality of a new method for aryl amination has been defined. Ketimines derived from o-bromophenethylamine cyclize to the N-substituted indoline when treated with nBu3SnH and a radical initiator. The pH-neutral conditions tolerate base- and acid-sensitive functionality. The observed regioselectivity is nonconventional for addition reactions involving carbon radicals and carbon-heteroatom pi-bonds.