INTERNATIONAL JOURNAL OF AGRICULTURE & BIOLOGY ISSN Print: 1560–8530; ISSN Online: 1814–9596 20–1387/2021/25–3–567–574 DOI: 10.17957/IJAB/15.1702 http://www.fspublishers.org Full Length Article To cite this paper: Sajid SM, A Yousaf, H Irshad, MA Zafar, SU Rehman (2021). Effect of various stabilizers on viability of lyophilized Pasteurella multocida B:3,4 for use as hemorrhagic septicemia vaccine. Intl J Agric Biol 25:567‒574 Effect of Various Stabilizers on Viability of Lyophilized Pasteurella multocida B:3,4 for use as Hemorrhagic Septicemia Vaccine Sajid Mahmood Sajid 1 , Arfan Yousaf 1 , Hamid Irshad 2* , Muhammad Arif Zafar 1 and Saif ur Rehman 1 1 Faculty of Veterinary and Animal Sciences, PMAS-Arid Agriculture University, Rawalpindi 2 Animal Health Program, Animal Sciences Institute, National Agricultural Research Center, Islamabad * For correspondence: hamidirshad@hotmail.com Received 29 September 2020; Accepted 21 November 2020; Published 25 January 2021 Abstract Hemorrhagic septicemia (HS) is a devastating disease of cattle and buffaloes. The live aerosol vaccine is the best option to control HS. However, stability and viability of live vaccine is an issue. The present study was conducted to investigate the effect of three extraneous stabilizers trehalose, skimmed milk and lactalbumin on the viability of the live vaccine strain Pasteurella multocida B:3,4. The viability of the strain was evaluated using various concentrations (5, 10, 15 and 20%) of these three stabilizers. Moreover, viability of P. multocida B:3,4 was also determined at four different storage temperatures (- 20, 4, 25 and 37°C). The duration of lyophilization cycle was also standardized for highest survival of cells. The data showed that trehalose and lactalbumin ensued percentage of viability as 91.89±0.08 and 80.38±2.57 respectively. Skimmed milk as stabilizer did not prove to defend cells during lyophiliztion and subsequent storage and exhibited cell viability approximately 0.47±0.009%. The study indicated that most effective stabilizer for lyophiliztion of P. multocida B:3,4 was trehalose at 15% concentration and was most suitable temperature for storage of lyophilized P. multocida B:3,4. © 2021 Friends Science Publishers Keywords: Stabilizers; Pasteurella multocida; Viability; Lyophilization; Trehalose; Skimmed Milk; Lactalbumin Introduction Hemorrhagic septicemia is a short course, lethal and septicemic disease of buffaloes and cattle caused by a Gram negative, non-motile, coccobacillus bacterium, the Pasteurella multocida (Boyce et al. 2000). HS is a main epizootic disease in cattle and buffaloes in several countries with high morbidity and mortality (El-Jakee et al. 2016). In Asia, HS has caused severe economic losses (Benkirane and De Alwis 2002; Abba et al. 2017). Treatment of HS with antibiotics is prolonged, expensive and ineffective because of emerging resistance of antibiotics against P. multocida. The use of antibiotics might cause toxicity to human consumers. Acute nature of HS is another hindrance in treatment (Ahmad et al. 2014). Different vaccine types including oil adjuvanted, multiple emulsion and alum precipitated vaccines are used to control this fatal disease. Killed vaccines have been extensively used against HS in the world (Shivachandra et al. 2011). Nonetheless, these injectable vaccines are difficult to administer as restraining the animals has been found to be very difficult (Sarah 2007). Moreover, oil adjuvant vaccine is fairly disliked by farmers because it possesses dense viscosity (Sarah et al. 2006; De Alwis 1992). The application of live vaccine that can be given intra-nasally is a substitute method to save the animals from HS. Live vaccines have played a pivotal part from the start of immunology (Detmer and Glenting 2006). Yet, there has been failure to develop vaccine having greater viable inhabitants of live microorganism for protection from disease. Thus, the initiation of an effective and systematic formulation approach is critical to increase survival rate, storage stability and bacterial cell activity in live vaccine. Freeze-drying or lyophilization is a convenient technique to conserve bacteria and viruses by dehydrating cellular fluid (Das et al. 2018). The purpose of lyophilization is to convert bacteria in a stable form which can be stored for a longer period of time. Moreover, it is mainly required to attain high viability of cells (Winters and Winn 2010). At present, freeze drying is a frequently used method for this purpose. It has been reported that many strains of bacteria have shown better survival rates after lyophilization for long storage duration (Peiren et al. 2015). This biophysical process is relatively comfortable and generally used for cultural collection of microorganisms. The discovery also narrates the way for preparing a stabilized product, vaccine (Morgan et al. 2006). However, this method might render the microorganisms to extra stressful operational steps therefore