Spatio-temporal dynamics and distributional trend analysis of African swine fever outbreaks (2020-2021) in North-East India.

African swine fever (ASF) is a highly contagious, notifiable, and fatal hemorrhagic viral disease affecting domestic and wild pigs. The disease was reported for the first time in India during 2020, resulted in serious outbreaks and economic loss in North-Eastern (NE) parts, since 47% of the Indian pig population is distributed in the NE region. The present study focused on analyzing the spatial autocorrelation, spatio-temporal patterns, and directional trend of the disease in NE India during 2020-2021. The ASF outbreak data (2020-2021) were collected from the offices of the Department of Animal Husbandry and Veterinary Services in seven NE states of India to identify the potential clusters, spatio-temporal aggregation, temporal distribution, disease spread, density maps, and risk zones. Between 2020 and 2021, a total of 321 ASF outbreaks were recorded, resulting in 59,377 deaths. The spatial pattern analysis of the outbreak data (2020-2021) revealed that ASF outbreaks were clustered in 2020 (z score = 2.20, p < .01) and 2021 (z score = 4.89, p < .01). Spatial autocorrelation and Moran's I value (0.05-0.06 in 2020 and 2021) revealed the spatial clustering and spatial relationship between the outbreaks. The hotspot analysis identified districts of Arunachal Pradesh, Assam and districts of Mizoram, Tripura as significant hotspots in 2020 and 2021, respectively. The spatial-scan statistics with a purely spatial and purely temporal analysis revealed six and one significant clusters, respectively. Retrospective unadjusted, temporal, and spatially adjusted space-time analysis detected five, five, and two statistically significant (p < .01) clusters, respectively. The directional trend analysis identified the direction of disease distribution as northeast-southwest (2020) and north-south (2021), indicate the possibility of ASF introduction to India from China. The high-risk zones and spatio-temporal pattern of ASF outbreaks identified in the present study can be used as a guide for deploying proper prevention, optimizing resource allocation and disease control measures in NE Indian states.

Development and evaluation of a novel polymerase spiral reaction based testing technique for same-day visual detection of Campylobacter coli in pork.

The developed polymerase spiral reaction-based technique specifically amplified the ceuE gene of C. coli and involved a three-step centrifugation method for DNA extraction. PSR, real-time and end-point PCR were able to detect 62 fg, 620 fg and 6.2 pg C. coli DNA/tube, respectively. PSR detection limits for artificially contaminated pork samples without enrichment, with 12 h enrichment and after 24 h enrichment were 1000 CFU/g, 100 CFU/g, and 10 CFU/g samples, respectively which were ten times better than real-time PCR. The detection performance of PSR (with 12 h enrichment) was also compared to culture (ISO10272-1:2017) method using 75 naturally-contaminated samples, which revealed the sensitivity, specificity, PPV, NPV and accuracy of 100% (95%CI, 73.2%-100%), 98.4% (95%CI, 90%-99.9%), 93.3% (95%CI, 66%-99.6%), 100% (95%CI, 92.5%-100%) and 98.7% (95%CI, 92.8%-99.9%), respectively. The advantage and novelty of this assay are its equipment-free nature, dye-based interpretation by the naked eye, and the requirement of one enzyme and one primer pair. This assay could be a better alternative to other molecular methods and may help in reducing the possible troubles (e.g., gastroenteritis, hospitalization, or death) of belated detection of C. coli in food products. This is the primary report applying the PSR for C. coli detection.

Cross-sectional study on rotavirus A (RVA) infection and assessment of risk factors in pre- and post-weaning piglets in India.

Rotavirus A (RVA) infections are known to retard the piglets' growth and minimize the profit to the pig farming community. Between August 2014 and July 2017, in a cross-sectional study, we surveyed 13 organized pig farms located in the eight states of India representing northern, north-eastern and southern regions, to identify the risk factors associated with RVA infection in pre- and post-weaning piglets. Faecal samples (n = 411) comprising of non-diarrhoeic (n = 320) and diarrhoeic (n = 91) were collected and screened for RVA infection using VP6 gene-based RT-PCR. RVA positivity of 52.5% (168/320) in non-diarrhoeic and 59.3% (54/91) in diarrhoeic piglets was noticed. Further, 53.3% (120/225) and 54.8% (102/186) of the samples from pre- and post-weaned samples were positive for RVA, respectively. To note, no statistically significant association was noticed between RVA infection, health and weaning status. Additionally, a questionnaire-based survey was conducted to identify the risk factors for RVA infections in piglets. The analysis revealed that good ventilation (OR 0.2, 95% CI 0.15-0.39), use of deep well water (OR 0.2, 95% CI 0.13-0.43) and feeding of commercial feed (OR 0.3, 95% CI 0.18-0.41) were associated with reduced risk of RVA infection compared with poor ventilation, use of shallow well water and feeding of own milled feed, respectively. Contrarily, mixed farms (OR 2.1, 95% CI 1.26-3.37), use of heater or cooler (OR 5.9, 95% CI 3.74-9.30), sheds in different elevation (OR 2.5, 95% CI 1.20-5.01) and weekly and occasional use of disinfectant for surface cleaning (OR 1.8, 95% CI 1.12-2.96) were associated with higher RVA infection. Mitigating the risk factors might help in better health management of piglets and increase the economic return to pig farming community in the country.

Risk factor analysis, antimicrobial resistance and pathotyping of Escherichia coli associated with pre- and post-weaning piglet diarrhoea in organised farms, India.

A cross-sectional study was conducted from 2014 to 2017 in 13 organised pig farms located in eight states of India (Northern, North-Eastern and Southern regions) to identify the risk factors, pathotype and antimicrobial resistance of Escherichia coli associated with pre- and post-weaning piglet diarrhoea. The data collected through questionnaire survey were used to identify the risk factors by univariable analysis, in which weaning status, season, altitude, ventilation in the shed, use of heater/cooler for temperature control in the sheds, feed type, water source, and use of disinfectant, were the potential risk factors. In logistic regression model, weaning and source of water were the significant risk factors. The piglet diarrhoea prevalence was almost similar across the regions. Of the 909 faecal samples collected (North - 310, North-East - 194 and South - 405) for isolation of E. coli, pathotyping and antibiotic screening, 531 E. coli were isolated in MacConkey agar added with cefotaxime, where 345 isolates were extended spectrum ß-lactamase (ESBL) producers and were positive for blaCTX-M-1 (n = 147), bla TEM (n = 151), qnrA (n = 98), qnrB (n = 116), qnrS (n = 53), tetA (n = 46), tetB (n = 48) and sul1 (n = 54) genes. Multiple antibiotic resistance (MAR) index revealed that 14 (2.64%) isolates had MAR index of 1. On the virulence screening of E. coli, 174 isolates harboured alone or combination of Stx1, Stx2, eaeA, hlyA genes. The isolates from diarrhoeic and post-weaning samples harboured higher number of virulence genes than non-diarrhoeic and pre-weaning. Alleviating the risk factors might reduce the piglet diarrhoea cases. The presence of multidrug-resistant and ESBL-producing pathogenic E. coli in piglets appears a public health concern.

Epidemiology of carbapenem-resistant Escherichia coli and first report of blaVIM carbapenemases gene in calves from India.

A cross-sectional study on six dairy farms was conducted to ascertain the occurrence of carbapenem-resistant Escherichia coli in calves. Two-hundred and seventy-nine isolates of E. coli were recovered from 90 faecal samples from apparently healthy (45) and diarrhoeal (45) calves. The isolates were screened for phenotypic susceptibility to carbapenems and production of metallo ß-lactamase, as well as five carbapenemase resistance genes by PCR, and overexpression of efflux pumps. Eighty-one isolates (29.03%) were resistant to at least one of three carbapenem antibiotics [meropenem (23.30%), imipenem (2.15%) and ertapenem (1.43%)], and one isolate was positive for the blaVIM gene which was located on an Incl1 plasmid of a novel sequence type (ST 297) by multilocus sequence typing. The majority (83.95%) of isolates had an active efflux pump. Calves housed on concrete floors were approximately seven times more likely to acquire meropenem-resistant isolates than those housed on earthen floors (95% CI 1.27-41.54). In India, carbapenem drugs are not used in food animal treatment, hence carbapenem-resistant strains in calves possibly originate from the natural environment or human contact and is of public health importance. To our knowledge, this is the first report of blaVIM carbapenemases gene in calves from India.

Spatial molecular epidemiology of carbapenem-resistant and New Delhi metallo beta-lactamase (blaNDM)-producing Escherichia coli in the piglets of organized farms in India.

AIM: A cross-sectional study was conducted in 10 government-organized pig farms between 2014 and 2016 representing seven states of India to understand the epidemiology of carbapenem resistance in the Escherichia coli. METHODS AND RESULTS: In this study, fecal sample (n = 673) from non-diarrheic (n = 501) and diarrheic (n = 172) piglets were processed for isolation of carbapenem resistant E. coli. Of 673, E. coli isolate (n = 112) was genotyped for confirming the carbapenem resistance and associated virulence factors. Of the 112 isolates, 23 were phenotypically resistant to carbapenem and 8 were carrying the New Delhi metallo beta-lactamase (blaNDM) gene. The carbapenem-resistant isolates also produced extended spectrum beta-lactamases and were multidrug resistant. The PCR-based pathotyping revealed the presence of stx1, stx2, eae and hlyA genes. The enterobacterial repetitive intergenic consensus PCR dendrogram analysis of the isolates yielded three distinct clusters. The statistical analysis revealed no association between carriages of carbapenem-resistant E. coli in different breed of piglets however, location, sex, health status of piglets and age showed significant difference. The spatial analysis with SaTScan helped in identification of carbapenem-resistant clusters. CONCLUSIONS: The presence of carbapenem resistant E. coli isolates with virulence genes in the piglet poses a potential public health risk through possible access and spread via the food chain and environment. Efflux pump may also play an important role in carbapenem resistance in piglet E. coli isolates. Furthermore, identification of risk factors in relation to spatial clusters will help in designing preventive strategies for reducing the risk of spread of carbapenem resistant bacteria. SIGNIFICANCE AND IMPACT OF THE STUDY: 1. Piglets harbor carbapenem resistant E. coli and have great public health significance. 2. Apart from carbapenemase, efflux pump is also important for carbapenem resistance. 3. This is the first report of blaNDM in the piglets from India.

Burden and predictors of hypertension in India: results of SEEK (Screening and Early Evaluation of Kidney Disease) study.

BACKGROUND: Hypertension (HTN) is one of the major causes of cardiovascular morbidity and mortality. The objective of the study was to investigate the burden and predictors of HTN in India. METHODS: 6120 subjects participated in the Screening and Early Evaluation of Kidney disease (SEEK), a community-based screening program in 53 camps in 13 representative geographic locations in India. Of these, 5929 had recorded blood pressure (BP) measurements. Potential predictors of HTN were collected using a structured questionnaire for SEEK study. RESULTS: HTN was observed in 43.5% of our cohort. After adjusting for center variation (p < 0.0001), predictors of a higher prevalence of HTN were older age ≥ 40 years (p < 0.0001), BMI of ≥ 23 Kg/M2 (p < 0.0004), larger waist circumference (p < 0.0001), working in sedentary occupation (p < 0.0001), having diabetes mellitus (p < 0.0001), having proteinuria (p < 0.0016), and increased serum creatinine (p < 0.0001). High school/some college education (p = 0.0016), versus less than 9th grade education, was related with lower prevalence of HTN. Of note, proteinuria and CKD were observed in 19% and 23.5% of HTN subjects. About half (54%) of the hypertensive subjects were aware of their hypertension status. CONCLUSIONS: HTN was common in this cohort from India. Older age, BMI ≥ 23 Kg/M2, waist circumference, sedentary occupation, education less, diabetes mellitus, presence of proteinuria, and raised serum creatinine were significant predictors of hypertension. Our data suggest that HTN is a major public health problem in India with low awareness, and requires aggressive community-based screening and education to improve health.

Expression of Orseolia oryzae nucleoside diphosphate kinase (OoNDPK) is enhanced in rice gall midge feeding on susceptible rice hosts and its over-expression leads to salt tolerance in Escherichia coli.

The Asian rice gall midge, Orseolia oryzae, is a major dipteran pest of rice, with many known biotypes. The present investigation was initiated to understand the molecular mechanisms of infestation for developing novel integrated pest management strategies. We isolated and characterized a gene, nucleoside diphosphate kinase (OoNDPK), from the rice gall midge, encoding a protein with 169 amino acid residues and with a secretory signal sequence - an observation that assumes significance as salivary gland secretions have been implicated to play a major role in insect-plant interactions. Furthermore, up-regulation (> 18 folds) of OoNDPK was observed in the salivary glands of maggots feeding on susceptible host in contrast to those feeding on resistant host. Phylogenetic analysis revealed similarity of OoNDPK with its dipteran orthologues. 3DLigandSite analysis, of the predicted OoNDPK and its orthologues, revealed phenylalanine and tyrosine residues to be specifically present in NDPK proteins from the plant feeders. Results suggest secretion of OoNDPK into the host plant and its probable involvement in gall midge-rice interaction. Using the coleoptile cell elongation assay, we demonstrated that the recombinant OoNDPK is capable of causing elongation of rice coleoptile cells. Additionally, heterologous expression of OoNDPK in Escherichia coli increased the tolerance of these cells to salt (NaCl; up to 1 mM), hinting at the involvement of this gene in abiotic stress response as well.

Pathotyping and antimicrobial susceptibility testing of Escherichia coli isolates from neonatal calves.

Neonatal calf mortality is a major concern to livestock sector worldwide. Neonatal calf diarrhoea (NCD), an acute severe condition causes morbidity and mortality in calves. Amongst various pathogens involved in NCD, E. coli is considered as one of the major causes. The study was targeted to characterize E. coli isolates from neonatal calves for diarrhoeagenic Escherichia coli (DEC) types (pathotyping), antimicrobial resistance (AMR) profiling and to correlate with epidemiological parameters. From neonates, a total of 113 faecal samples were collected, out of that 308, lactose fermenting colonies were confirmed as E. coli. Pathotypable isolates (12.3%) were represented by STEC (6.1%), EPEC (2.9%), ETEC (1.9%), EAEC (0.9%) and EHEC (0.3%). Occurrence of STEC was more in non-diarrhoeic calves, whereas ETEC was observed more in diarrhoeic calves. EPEC occurrence was observed in both diarrhoeic and non-diarrhoeic calves. Fishers extract test showed no significant association for occurrence of DEC types to type of dairies, health status, species, breed, age and sex of neonatal calves. Two hundred and eighty isolates were tested for antimicrobial susceptibility. The isolates showed maximum resistance towards ampicillin (55.4%) followed by tetracycline (54.3%), while minimum resistance was observed towards meropenem (2.5%). Multidrug resistant E. coli isolates were found to be 139 (49.6%), and Extended-spectrum beta-lactamase (ESBL) producers were 120 (42.9%). DEC pathotypes like STEC, ETEC, EHEC and EAEC that are also multidrug resistant present in neonatal calves have zoonotic potential and hence are of public health significance.

Compatible interaction with its rice host leads to enhanced expression of the gamma subunit of oligosaccharyl transferase in the Asian rice gall midge, Orseolia oryzae.

The Asian rice gall midge, Orseolia oryzae, is a fast evolving, damaging pest of rice. Understanding the underlying molecular mechanism of interaction between the gall midge and rice will help in devising strategies to control and manage the pest. The present study aims to identify rice-responsive genes in the gall midge that aid pest survival. The abundance of transcripts coding for enzymes related to glycosylation, in a cDNA library prepared from maggots of the rice gall midge feeding on susceptible hosts, indicated their probable involvement in the gall midge-rice interaction. Hence, a full-length transcript for a gamma subunit of the oligosaccharyl transferase gene (OoOST) from the gall midge was cloned and characterized. It has 72% similarity to its orthologue cloned from Aedes aegypti. Tissue-specific analysis of the expression of OoOST revealed an increase (> sevenfold) in the transcripts of the gene in the salivary glands of maggots in susceptible plants when compared with the transcript level in the salivary glands of maggots feeding on resistant hosts. Using quantitative PCR, performed on different developmental stages of the maggots in two susceptible and two resistant hosts, we observed similar expression patterns (i.e. overexpression in the compatible interaction). These results indicate the involvement of OoOST in maggot survival and establishment in the susceptible host. In order to identify polymorphism in the gene, OoOST was cloned from three gall midge biotypes GMB1, GMB4 and GMB4M.

Multi-drug resistant (MDR), extended spectrum beta-lactamase (ESBL) producing and carbapenem resistant Escherichia coli in rescued Sloth bears (Melursus ursinus), India.

The study reports the multi-drug resistant (MDR), extended spectrum beta-lactamase (ESBL) producing and carbapenem resistant Escherichia coli (CRE) isolated from rescued sloth bear (Melursus ursinus), India. Non-duplicate faecal samples from 21 adult rescued sloth bears were collected at once during 2015-2016 and processed for isolation of E. coli and antibacterial susceptibility pattern. From 21 samples, 45 E. coli were isolated and on phenotypic screening, 23 were MDR, 17 were ESBL producers, and five were carbapenem-resistant (CR). Three E. coli isolates (6.67%, 3/45) showed no resistance, however 42 isolates (93.33%, 42/45) exhibited resistant to at least one antibiotics. The MDR isolates carried beta-lactamase, chloramphenicol, aminoglycosides, tetracycline, fluroquinolone, and sulphadimidine resistance genes. All the phenotypic ESBL producing isolates harbored blaCTX-M genes. On genotypic screening, three CRE (60.0%, 3/5) were positive for blaNDM carbapenemase gene and efflux pump-mediated carbapenem resistance was detected in two CRE isolates (40.0%, 2/5) which were negative for carbapenemase genes. The CRE isolates (n = 5) also co-harbored AMR genes like blaTEM-1, blaAmpC, qnrA, qnrB, qnrS, tetA, tetB and sulI. Virulence screening of the resistant isolates detected the presence of Stx1(n = 1), Stx2 (n = 3), eaeA (n = 4) and hlyA (n = 3) genes. Plasmid incompatibility (Inc) typing revealed that two isolates harboured blaNDM-5 gene on Incl1 and one isolate on IncF plasmid. Apart from the NDM gene, the plasmids also carried tetracycline, beta-lactamase and quinolone resistance genes. The plasmid multilocus sequence typing (pMLST) of the E. coli Incl1 plasmid showed the Sequence Type (ST) 297. This appears to be the first report of MDR, ESBL producing and blaNDM-5 genes on Incl1 and IncF plasmids from rescued sloth bear.

Faecal carriage of extended spectrum beta-lactamase (ESBL) and New Delhi metallo beta-lactamase(NDM) producing Escherichia coli between piglets and pig farmworkers.

A cross-sectional study on five organized pig farms was conducted to assess the faecal carriage of ESBL and blaNDM carbapenemase-producing E. coli in piglets and pig farmworkers. Faecal samples from piglets (n = 155) and pig farmworkers (n = 21) were processed for isolation and characterization of E. coli. A total of 124 E. coli isolates from piglets and 21 E. coli isolates pig farmworkers were recovered and screening for ESBL production showed that 44.4 % (55/124) of the isolates from piglets and 42.9 % (9/21) of the isolates from farmworkers were ESBL positive. The ESBL positive isolates from piglets and farmworkers harbored blaCTX-M and also co-harbored other beta-lactams, sulphonamide, quinolone and tetracycline resistance genes. Diarrhoeic (50%, 49/98) and crossbred piglets (52.7%, 39/74) harbored a significantly higher number of ESBL producing isolates than non-diarrhoeic (23.1 %, 6/26) and purebred piglets (32%, 16/50) (p < 0.05). Piglets and pig farmworkers harbored nine and two carbapenem-resistant isolates, respectively. Interestingly, two isolates from piglets and one isolate from farmworkers harbored the blaNDM gene. The blaNDM positive E. coli isolated from piglets and farmworkers of the same farm revealed similar antibacterial resistance patterns, resistant genes, sequence (ST-167) and plasmid type (IncX3). In India, carbapenems are not used in food animal treatment, hence carbapenem resistant E. coli in piglets possibly originated from the human contact or common environment and is of public health importance.

Bluetongue in India: a systematic review and meta-analysis with emphasis on diagnosis and seroprevalence.

Bluetongue (BT) is an infectious viral disease which affects a wide range of ruminants and was first reported in India in 1964. In view of the absence of comprehensive information on the BT status in India, this study presents the seroprevalence on BT in farm animals of India based-on a systematic review and meta-analysis. A systematic review was conducted to identify the published articles (2001-2018) reporting the seroprevalence of BT in sheep, goats, cattle, buffalo, camels, and Mithun (Bos frontalis) from India. From 409 research articles, 71 fulfilled the inclusion criteria and meta-analysis for proportions was carried out targeting the eligible studies. From these, 144 strata level data were extracted with a sample size of 14048 sheep, 14696 goats, 5218 cattle, 2653 buffaloes, 2062 camels, and 222 Mithun. Overall, the analyses showed that the BT seroprevalence of 43% (95% CI: 38-49%) in goats, 39% (95% CI: 33-46%) in sheep, 38% (95% CI: 25-45%) in cattle, 34% (95% CI: 20-51%) in buffaloes, 16% (95% CI: 10-22%) in camels, and 66% (95% CI: 17-95%) in Mithun. Furthermore, the meta-regression analysis suggested that serological tests, geographical region, and sample size were the prime moderators. Meta-analytic study indicates the BT seropositivity in 25.35 million sheep (95% CI: 21.5-29.9), 58 million goats (95% CI: 51.3-66.2), 66.8 million cattle (95% CI: 47.7-86), 37.0 million buffaloes (95% CI: 21.7-55.4), 0.06 million camels (95% CI: 0.04-0.09), and 0.19 million Mithun (95% CI: 0.05-0.28). The findings highlight the variation of BT seropositivity in different geographical regions of India.

Molecular characterisation of blaOXA-48 carbapenemase-, extended-spectrum ß-lactamase- and Shiga toxin-producing Escherichia coli isolated from farm piglets in India.

OBJECTIVES: The aim of this study was to characterise carbapenemase-, extended-spectrum ß-lactamase (ESBL)- and Shiga toxin-producing Escherichia coli isolated from farm piglets in India. METHODS: Faecal samples (n=741) from 10 organised pig farms, including non-diarrhoeic (n=546) and diarrhoeic (n=195) piglets, were processed for isolation of carbapenem-resistant and ESBL-producing E. coli. RESULTS: A total of 27 and 243 isolates were phenotypically confirmed as carbapenem-resistant and ESBL-producers, respectively. The meropenem minimum inhibitory concentration (MIC) of carbapenem-resistant isolates ranged from 8-128µg/mL. On genotypic screening of the 27 carbapenem-resistant isolates, 3 isolates were positive for the blaOXA-48 carbapenemase gene; no other carbapenemase genes were detected. The 243 ESBL-producing isolates were positive for blaCTX-M-1 (n=135), qnrA (n=92), qnrB (n=112), qnrS (n=49), tetA (n=42), tetB (n=45) and sul1 (n=43). The Shiga toxin virulence markers stx1 and stx2 were detected in 41 and 38 of the 243 phenotypic ESBL-producing isolates, respectively. Multilocus sequence typing (MLST) of blaOXA-48-positive E. coli isolates showed ST10- and ST5053-like sequence types. CONCLUSION: This is the first report on the presence of blaOXA-48-carrying E. coli in piglets in India, which pose a potential risk to public health.

Induction of mammary tumors in aging rats by 7,12-dimethylbenz[a]anthracene: role of DNA synthesis during carcinogenesis.

Two routes of administration were used to test the susceptibility of the mammary gland of the rat to 7,12-dimethylbenz[a]anthracene (DMBA) carcinogenesis in relation to age of the tissue. In one series of experiments, 60-, 70-, 90-, 120-, 150-, and 200-day-old female noninbred Sprague-Dawley rats were given DMBA iv. In parallel experiments, rats of the same ages as those above were given DMBA by local application. Mammary tumors developed in 80-90% of the 60- and 70-day-old rats and in 40% of the 90-day-old rats. Rats 120 days old and older were completely "refractory" to DMBA. In contrast, all rats, irrespective of their ages, developed tumors when DMBA was applied locally. The DNA synthesis labeling index (LI) of mammary glands of rats in two series of experiments was examined by [3H]thymidine incorporation and autoradiography at 24, 48, and 72 hours after DMBA treatment. DMBA, given iv significantly inhibited DNA synthesis in mammary glands, but DMBA applied locally significantly increased the LI of the mammary glands. Thus the LI of the mammary glands in 60-day-old rats increased from 8% (control, untreated) to 16% (rats receiving local applications of DMBA), whereas the LI of the mammary glands in 150-day-old rats increased from 1.15% (control, untreated) to 8% (rats receiving local application of DMBA).

Comparative chromosome study of normal mammary tissue, carcinogen-induced mammary tumors, and hyperplastic alveolar nodules in the rat: Brief communication.

The chromosome constituents of 101 normal mammary gland cells, 122 hyperplastic alveolar nodule (HAN) cells, and 360 cells from eleven 7,12-dimethylbenz[a]anthracene (DMBA)-induced mammary tumors in the WF rat were examined. Both mammary gland and HAN had normal diploid karyotype patterns, whereas mammary tumors showed frequent (approximately 30%) abnormalities, e.g., aneuploidy, pseudodiploidy, polyploidy, and structural aberrations such as breakage and translocation. These results suggested that chromosome constituents of HAN cells are essentially the same as those of the normal mammary epithelial cells.

Estrogenic properties of 3,9-dihydroxy-7,12-dimethylbenz[a]anthracene in rats.

Previously, the 3,9-dihydroxy derivative of benz[a]-anthracene was shown to be weakly estrogenic. The availability of the related diol of the mammary carcinogen dimethylbenz[a]-anthracene, i.e., 3,9-dihydroxy-7,12-dimethylbenz[a]anthracene (3,9-diOHDMBA), prompted a similar study of its estrogenic properties. The competitive binding studies of 3,9-diOHDMBA with 17beta-estradiol in the uterine cytosol of immature SD rats gave a Ka of 1.7 x 10(8) M-1. 17beta-Estradiol (10(-9) M) binding to the 8S binding protein was inhibited by 3,9-diOHDMBA at concentrations similar to those of nafoxidine HCl (1 x 10(-5) M). Bioassay demonstrated that the diol possesses 1/4,464 the activity of 17beta-estradiol.

Estrogenic properties of 3,9-dihydroxybenz[a]anthracene, a potential metabolite of benz[a]anthracene.

The estrogen receptor binding properties of 3,9-dihydroxybenz[a]anthracene (3,9-diOHBA) were determined in uterine cytosol of immature Sprague-Dawley rats by competitive binding experiments with [3H]estradiol and sucrose density centrifugation. 3,9-DiOHBA inhibited estradiol binding to the 8S binding protein at a concentration (1.2 X 10(-5) M) approximately equal to that of nafoxidine-HCl (3 X 10(-5) M) required to inhibit estradiol-specific binding, and bioassay for estrogenic activity substantiated this finding.

Enhancement of mammary tumorigenesis by dietary selenium deficiency in rats with a high polyunsaturated fat intake.

The effect of selenium depletion on mammary tumorigenesis following dimethylbenz[a]anthracene administration was examined in female Sprague-Dawley rats that were fed different levels and types of fats. Four basal diets deficient in selenium were used: (a) 1% corn oil; (b) 5% corn oil; (c) 25% corn oil; and (d) a high saturated fat diet containing 1% corn oil and 24% hydrogenated coconut oil. The comparable selenium-adequate diets were obtained by adding 0.1 ppm of selenium to each of the basal diets. In animals that received an adequate supplement of selenium, an increase in fat intake was accompanied by an increased tumor incidence when corn oil was used in the diets. A high saturated fat ration, on the other hand, was much less effective in this respect. Only in those rats that were maintained on a high polyunsaturated fat diet (25% corn oil) did selenium depletion result in a further increase in tumor incidence and tumor yield. Such an augmentation was not observed in animals given either a 1 or a 5% corn oil ration or a diet rich in saturated fat. Regardless of selenium status, almost all of the tumors found were adenocarcinomas. An enhancement of tumorigenesis as a result of selenium deficiency in rats fed the 1% corn oil regimen was detected provided a high dose of dimethylbenz[a]anthracene was used, suggesting that alterations in dimethylbenz[a]anthracene metabolism might be involved under this condition. The antioxidant property of selenium is discussed as a possible mechanism by which selenium protects against tumorigenesis, especially in animals with a high polyunsaturated fat intake.

Effect of estrogen and progesterone on cellular replication of human breast tumors.

A total of 54 infiltrating carcinomas of the breast were studied for estrogen receptor concentration in the tumor cytosol and thymidine-labeling indexes. The results showed that there was no significant difference in the level of thymidine-labeling index between 22 primary and 32 metastatic breast cancers. No significant association between the levels of thymidine-labeling index and the presence or absence of estrogen receptors was observed. The effect of "physiological" doses of estrogen and progesterone on cell proliferative activity was studied by the level of thymidine-labeling indexes in the tumor cells in 10 patients with multiple skin and s.c. metastases. Tumor biopsies were performed for labeling indexes both before and after hormonal treatment. The results showed that physiological doses of estrogen and progesterone induced a significant rise in thymidine-labeling index within 3 days after hormonal treatment in seven of the 10 tumors. Of the seven tumors that showed a rise in thymidine-labeling index, three were estrogen receptor positive and four were estrogen receptor negative. Of the three nonresponsive tumors, one was estrogen receptor positive and two were estrogen receptor negative. The study suggests that estrogen and progesterone can induce cell replication in both estrogen receptor-positive and -negative tumors.