Alopecia induced by salmonellosis in guinea pigs

DESPITE extensive research on the functional, genetic and morphological aspects, the cycling and morphogenesis of the hair follicle is poorly understood due to the effects of multiple genetic and environmental factors (Stenn and others 1996, Panteleyev and others 1998, 1999, 2000, Paus 1998). Although all the malfunctions associated with hair follicles appear to be under the control of the hairless (hr) gene, how the gene is activated, and alopecia precipitated, is still not well understood. Environmental factors such as deficiencies of certain vitamins or minerals, or the acquisition of some infections, have been shown to have a definite association with hair loss (Radostits and others 2003). Bovine viral diarrhoea in cattle, Border disease in sheep, and dermatitis and mycosis in most mammalian species have long been recognised among the infectious causes of alopecia, but bacterial diseases have rarely been shown to be a cause of hair loss (Radostits and others 2003). Alopecia has often been reported in descriptions of chronic cases of typhoid and paratyphoid fever in human beings and in experimental salmonellosis in laboratory rodents (Sharma and Singh 1997, Singh and others 1997, Singh and Verma 2002). This short communication describes a study of the role of chronic salmonellosis in hair loss in guinea pigs. Chronic salmonellosis was induced (Singh and others 2002) in guinea pigs, under the approval of the institute’s animal ethics committee, with four nalidixic acid-resistant isogenic strains of wild origin (wild type S-787, aroA mutant S-28, htrA mutant S-29 and aroA-htrA mutant S-30) available at the National Salmonella Centre (Singh and others 2000), and an ampicillin-resistant reference strain (E-156) of Salmonella enterica serovar Abortusequi. A total of 120 female guinea pigs, aged 60 to 75 days, procured from the pathogen-free laboratory animal facility of the institute, were divided into 12 groups (A to L) of 10; all were maintained on a balanced ration recommended by the laboratory animal section of the institute, with an ample supply of water and green vegetables, on hygienic galvanised iron trays (2 m x 1·2 m x 0·3 m) padded with sterilised wheat straw.Vitamin C (ascorbic acid; Hi-Media) was supplemented at the rate of 1 g/kg of feed for the animals in group E. For the inoculation with salmonellae, the animals in groups A to F were inoculated intramuscularly in the thigh muscle, while the animals in groups G to J and L were fed orally with 0·1 ml of trypticase soya broth (TSB; Difco B & D India) cultures of different strains, grown overnight, with predetermined doses (Table 1). For setting the doses of inocula, the number of colony-forming units (cfu) was determined by the spread agar method (Singh and others 2002), by plating, in triplicate, 100 μl aliquots of 10-fold dilutions of eight-hour cultures grown in TSB at 37°C. The plates were incubated at 37°C for 24 hours before the colonies were counted. Suspected colonies were tested by the plate agglutination test (Edwards and Ewing 1986) using antisera to somatic factor 4 and flagellar factor x (Difco B & D India). For counting and plating the cultures of strains S-787, S-28, S-29 and S-30, plates of Hektoen enteric (HE) agar (Difco B & D India) supplemented with 30 μg/ml nalidixic acid (Difco B & D India) were used. For counting the strain E-156 colonies, plates of HE agar supplemented with 30 μg/ml ampicillin (Difco B & D India) were used. The animals in group K were inoculated with 0·1 ml of 0·15M phosphate-buffered saline intramuscularly in the thigh, as a negative control. The animals’ trays were cleaned and the straw padding was replaced twice weekly, and approximately 25 g of faecal pellets were collected from the trays for re-isolation of Salmonella species (Edwards and Ewing 1986). Rectal swabs were also collected on every seventh day from individual animals, until the 91st day after infection, for reisolation of salmonellae (Edwards and Ewing 1986). The guinea pigs were examined for signs of hair loss daily from the day of inoculation until the end of the experiment; however, quantitative recording for hair loss was carried out every seventh day. The degree of alopecia was scored 0 if no loss of hair was apparent; 1 when hair was depleted on the back, near the hindquarters; 2 when hair depletion was frank on the back and abdomen; and 3 when more than three-quarters of the guinea pig’s body was affected (Fig 1). The association between different strains of Salmonella and alopecia was determined by a normal Z test and phi coefficient (Fleiss 1981). The onset and degree of depilation and the number of affected animals varied with the route of inoculation and the strain of Salmonella (Table 1). Loss of hair started as early as 27 and 24 days after infection, respectively, in the animals in groups A and F, which had respectively been inoculated intramuscularly with wild-type strain S-787 and reference strain