Development of reverse transcription loop-mediated isothermal amplification (RT-LAMP) for rapid detection of viruses infecting patchouli (Pogostemon cablin).

Patchouli (Pogostemon cablin), a highly valued medicinal plant, suffers significant economic losses following infection with Broad bean wilt virus 2 (BBWV-2) and Peanut stripe virus (PStV). In this study, a field-based isothermal technique called reverse transcription loop-mediated isothermal amplification (RT-LAMP) was established for an early and specific detection of BBWV-2 and PStV. The oligo primers were designed to target the coat protein genes of PStV and BBWV-2. The reaction conditions, such as temperature and time duration, were optimized to 65 °C for 60 min. The LAMP amplicons positive for PStV and BBWV-2 revealed characteristic ladder-type bands following agarose gel electrophoresis. Further, a colorimetric assay using a metal ion-based indicator (Hydroxy-naphthol blue, HNB) was conducted to visualize the amplified products with the naked eye, thus facilitating accessibility to field practices. The assay developed in this study was found to be virus specific, and was 100 times more sensitive than RT-PCR. Thus, the RT-LAMP assay established in this study is quick, reliable, and cost-effective for the accurate identification of BBWV-2 and PStV. It will facilitate the screening of patchouli planting materials.  Further, it may reduce the risk of virus spread and could be helpful in phytosanitary programs.

Effect of Piper cubeba L. fruit on ethylene glycol and ammonium chloride induced urolithiasis in male Sprague Dawley rats.

BACKGROUND: To investigate the antilithiatic effect of hydroalcoholic extract of Kabab Chini (Piper cubeba L.) fruit in male Sprague Dawley rats. METHODS: Rats were divided into six groups of six each. Group I received regular rat food and drinking water ad libitum. Groups II to VI were administered with ethylene glycol (EG) 0.75% (V/V) and ammonium chloride (AC) 1% (W/V) in drinking water for 7 days to induce urolithiasis. From 8th day Group I received 1 mL of 5% gum acacia. Group IV was treated with Cystone; V and VI groups with the hydro-alcoholic extract of Piper cubeba L. Treatment was continued for further 14 days, thereafter animals sacrificed. While Group II animals were sacrificed just after 7 days treatment with EG and AC. Group III was left untreated until 14 days and sacrificed on 22nd day. Crystalluria was analyzed on 8th and 22nd day while, urinary calcium, phosphorus, creatinine, sodium and magnesium on 22nd day. Biochemistry and histopathological studies of kidney were also carried out. RESULTS: Test groups showed significant reduction (p < 0.001) of crystals in urine. Serum creatinine and urea (p < 0.01) were also decreased significantly. Urine analysis showed significant increase in magnesium while calcium, sodium, chloride and phosphorus significantly decreased along with histopathological improvement in kidney tissue in treated groups. CONCLUSION: From the above results it can be concluded that hydroalcoholic extract of P. cubeba L. fruit has significant inhibitory effect in calcium oxalate urolithiasis.

Antilithiatic effect of Peucedanum grande C. B. Clarke in chemically induced urolithiasis in rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Duqu (Peucedanum grande C. B. Clarke) has been used by Unani physicians since ancient times in retention of urine, renal and bladder calculi, nephritis and other associated disorders in different dosage forms. AIM OF THE STUDY: The aim of the study was to evaluate the antiurolithiatic activity of Peucedanum grande C. B. Clarke in experimental model. MATERIALS AND METHODS: The experiment was carried out in male Sprague Dawley rats divided into 5 groups of 8 animals each. Animals of negative control received 1ml of 5% Gum acacia throughout the study. Remaining four groups received Ethylene glycol 0.75% and Ammonium chloride 1% by adding in their drinking water for first seven days to induce urolithiasis. From 8th day, positive control received 1ml of 5% Gum acacia, standard control received Cystone in the dose of 750mg/kg while test groups A and B were treated with hydroalcoholic extract of test drug in the dose of 56mg/kg and 97mg/kg respectively up to 21 days, thereafter the animals were sacrificed. Number of CaOx crystals in urine, levels of serum calcium, phosphorus, creatinine, urea, urinary calcium and sodium were observed. Kidney homogenate analysis and histopathology were also carried out. RESULTS: Test drug reduced number of CaOx crystals in urine (p<0.001); levels of serum calcium, phosphorus, creatinine, (p<0.001) urea, (p<0.05); urinary calcium (p<0.001) and sodium decreased significantly in standard and test groups. The urine volume increased significantly (p<0.05, 0.01) in both the test groups. Histopathology of kidney showed no CaOx crystal deposition in both the test groups. CONCLUSION: On the basis of above findings, it can be concluded that the test drug possesses significant antiurolithiatic activity.

Effect of combination of Withania somnifera Dunal and Tribulus terrestris Linn on letrozole induced polycystic ovarian syndrome in rats.

BACKGROUND: To investigate the effect of the combination of hydroalcoholic extract of Withania somnifera (WS) and Tribulus terrestris (TT) on letrozole induced polycystic ovarian syndrome (PCOS) in rat. METHODS: Twenty four female Wistar rats of regular estrus cycle were divided into four groups of six animals each. The negative control group received 1 mL of 0.5% carboxy methyl cellulose. The animals of the other groups were treated with letrozole (1 mg/kg) for 21 days for induction of PCOS. The animals of the positive control group were sacrificed on the 22nd day. In the test and standard groups, the treatment was started from the 22nd day and continued for a further 28 days. The test group was treated with hydroalcoholic extract of the combination of WS and TT (198 mg/kg) and the standard group with clomiphene citrate (1 mg/kg). Throughout the study, vaginal smears were collected daily from each animal for the determination of different phases of the estrus cycle. After completion of the treatment schedule all the animals of each group were sacrificed; analysis of hormones, total cholesterol, blood glucose, ovarian and uterine weight, and histopathological study of the ovary were carried out. RESULTS: The combination of the test drugs showed significant effects in normalizing the estrus cycle after being altered by letrozole. In the positive control group follicle-stimulating hormone level was decreased although luteinizing hormone, estradiole, and testosterone levels were increased (p < 0.05), however, after treatment the reverse effect was observed in the level of these hormones. Significant reduction in serum total cholesterol was also observed (p < 0.05). The test drugs decreased ovarian weight, and uterine weight was also returned to normalcy. Histopathology of the ovary showed almost normal ovary. CONCLUSION: The above findings indicate the effectiveness of the combination of hydroalcoholic extract of WS and TT against letrozole induced polycystic ovarian syndrome in rat. This validates the usefulness of combination in PCOS and other related disorders as mentioned by Unani physicians.