First Report of Ratoon Stunt of Sugarcane Caused by Leifsonia xyli subsp. xyli in Pakistan.

Sugarcane (Saccharum hybrids), the second largest cash crop of Pakistan, is planted on 1.029 million ha with an annual production of 50 million tons. During a survey of the sugarcane crop in Faisalabad, Sargodha, and the Dera Ghazi Khan Division of the Punjab Province of Pakistan from 2007 to 2010, symptoms consistent with ratoon stunting, including stunted growth and reddening of the vascular bundles at the nodal regions (1), was observed on sugarcane cvs. CP77-400, SPF-241, CP72-2086, and NCo-310. CP72-2086 and NCo-310 showed severely stunted growth in both crop cycles. A chemical test was performed for detecting ratoon stunt from the field. Longitudinal sections of mature nodes were treated with a combination of hydrogen peroxide and hydrochloric acid. Healthy canes developed a blue-green color in the parenchymatous tissue around the fibrovascular bundles, diseased cane did not. This field test illustrated that as much as 25% of the plants were infected by ratoon stunt in the survey area. Aerobic bacteria were isolated from a stunted sample (NCo-310) on modified sugarcane medium (17 g of cornmeal agar, 8 g of peptone from soy meal, 1 g of K2HPO4, 1 g of KH2PO4, 0.2 g of MgSO4·7H2O, 0.5 g of glucose, 1 g of cysteinefree base, 2 g of bovine serum albumin, and 15 mg of bovine hemin chloride) and incubated for 3 to 4 weeks at 28°C. Light, off-white, round, and raised growth bacterial colonies (1.5 to 4.5 × 0.2 to 0.35 µm). Isolates were positive for the gram and catalase reactions and negative for oxidase, aesculin hydrolysis, urease production, and motility. The pathogen was identified as Leifsonia xyli subsp. xyli (formerly Clavibacter xyli subsp. xyli) based on its morphological characteristics (2). A direct antigen coating-ELISA was developed with antiserum raised against L. xyli subsp. xyli at the National Institute for Biotechnology and Genetic Engineering, Faisalabad, Pakistan. Infected or suspected to be infected plants of different cultivars were used for an ELISA test. Results showed that sugarcane cvs. NCo-310 (Log 1.342 CFU/ml) and CP72-2086 (Log 0.118 CFU/ml) had higher L. xyli subsp. xyli titres than the other cultivars tested (SPF-213 [Log 0.071CFU/ml], CPF-237 [Log 0.077CFU/ml], HSF-240 [Log 0.069 CFU/ml], NSG-555 [Log 0.060 CFU/ml], SPSG-26 [Log 0.076 CFU/ml], SPSG-79 [Log 0.074 CFU/ml], SPF-238 [Log 0.057 CFU/ml], and CP77-400 [Log 0.063 CFU/ml]). Cv. SPF-241 (Log 0.107 CFU/ml) was weakly positive for ratoon stunt (4). Axillary buds of sugarcane were injected via a sterile hypodermic syringe with an 18-gauge needle to deliver a bacterial suspension of 109 cells/ml (3). Inoculated sugarcane plants were examined at intervals over 9 months for the development of symptoms and the presence of bacteria. Cultivars were evaluated on the basis of average number of colonized vascular bundles. SPF-213, CPF-237, HSF-240, NSG-555, SPSG-26, SPSG-79, SPF-238, and CP77-400 were resistant; SPF-241 showed moderate resistance and CP72-2086 and NCo-310 were highly susceptible to ratoon stunt. The pathogen was reisolated from the inoculated plants and identified as L. xyli subsp. xyli by bacteriological tests and its serological reaction. To our knowledge, this is the first report of ratoon stunt of sugarcane in Punjab Province of Pakistan. References: (1) M. J. Davis et al. Science 210:1365, 1980. (2) L. I. Evtushenko et al. Int. J. Syst. Evol. Microbiol. 50:371, 2000. (3) M. P. Nayiager et al. Phytopathol. Z. 99:273, 1980. (4) G.-P. Rao and G.-P. Singh. Sugar Tech. 2:35, 2000.

Clinical administration of microneedles: skin puncture, pain and sensation.

Injections using hypodermic needles cause pain, discomfort, localised trauma and apprehension. Additionally, careful use and disposal of needles is required to avoid transmission of blood-borne pathogens. As an alternative, microneedles can facilitate drug delivery without significantly impacting on pain receptors or blood vessels that reside beneath the skin outer layers. In this study we aim to determine the pain and sensory response to the application of wet-etch silicon microneedles, when used in such a way as to reliably penetrate skin, and provide a preliminary indication of how skin responds to microneedle injury with time. Twelve subjects received single-blinded insertions of a 25-G hypodermic needle and two microneedle arrays (36 needles of 180 and 280 mum height). The optimal method for microneedle application was determined in a pilot study. Pain intensity was scored using a visual analogue scale (VAS) and sensory perception determined using an adapted McGill Pain Questionnaire Short Form. Skin penetration was determined by external staining and measurement of trans-epidermal water loss (TEWL). Mean VAS scores, verbal descriptions and questionnaire responses showed that the 180 and 280 mum microneedles caused significantly less pain and discomforting sensation in participants than the hypodermic needle. Methylene blue staining and TEWL analysis confirmed that microchannels were formed in the skin following microneedle application. Evidence of microchannel repair and resealing was apparent at 8-24 h post-application. In summary, this study shows that pyramidal wet-etch microneedles can penetrate human skin with minimal pain and sensory discomfort, creating transient pathways for potential drug, vaccine and DNA delivery.

A study on diarrheal diseases in children under five years of age.

A cross sectional house-to-house survey on diarrheal disease in children less than five-years-of-age was conducted in the Eastern Province of Saudi Arabia. The main objective was to determine diarrheal disease morbidity and its relationship with different variable factors and management practices during the episode of diarrhea. The survey population included 1,105 households with 2,005 children under five-years-of-age selected from the entire Eastern Province. The prevalence observed was 7.6% during a two-week period giving an annual incidence of two episodes per child per year. Diarrheal morbidity was less prevalent among with age, water storage, and feeding patterns. Diarrhea morbidity was less prevalent among children receiving breast milk. The majority of cases with diarrhea were treated in health facilities where 27% were given drugs only. A solution containing sugar and salt in addition to home fluids were given to 46.4% while 23% received oral rehydration solution (ORS).